CHAPTER XII.

THE ORGANS OF EXCRETION.

The earliest stages in the development of the excretory system have already been described in a previous chapter[335] of this memoir, and up to the present time no investigator, with the exception of Dr Alex. Schultz[336], has gone over the same ground. Dr Schultz' descriptions are somewhat brief, but differ from my own mainly in stating that the segmental duct arises from an involution instead of as a solid knob. This discrepancy is, I believe, due to Dr Schultz drawing his conclusions as to the development of the segmental duct from its appearance at a comparatively late stage. He appears to have been unacquainted with my earlier descriptions.

Footnote 335: Chapter VI. p. 345, _et seq._

Footnote 336: _Archiv f. Micr. Anat_. Bd. XI.

The adult anatomy and later stages in the development of the excretory organs form the subject of the present chapter, and stand in marked contrast to the earlier stages in that they have been dealt with in a magnificent monograph[337] by Professor Semper, whose investigations have converted this previously almost unknown field of vertebrate embryology into one of the most fully explored parts of the whole subject. Reference is frequently made to this monograph in the succeeding pages, but my references, numerous as they are, give no adequate idea of the completeness and thoroughness of Professor Semper's investigations. In Professor Semper's monograph are embodied the results of a considerable number of preliminary papers published by him in his _Arbeiten_ and in the _Centralblatt_. The excretory organs of Elasmobranchii have also formed the subject of some investigations by Dr Meyer[338] and by myself[339]. Their older literature is fully given by Professor Semper. In addition to the above-cited works, there is one other paper by Dr Spengel[340] on the Urinogenital System of Amphibians, to which reference will frequently be made in the sequel, and which, though only indirectly connected with the subject of this chapter, deserves special mention both on account of the accuracy of the investigations of which it forms the record, and of the novel light which it throws on many of the problems of the constitution of the urinogenital system of Vertebrates.

Footnote 337: "Urogenital System d. Plagiostomen," Semper, _Arbeiten_, Vol. II.

Footnote 338: _Sitzungsberichte d. Naturfor. Ges. Leipzig_, 1875. No. 2.

Footnote 339: "Preliminary account of the development of Elasmobranch Fishes," _Quarterly Journal of Microscopical Science_, 1874. "Origin and History of the Urinogenital Organs of Vertebrates," _Journal of Anat. and Physiol._ Vol. X.

Footnote 340: _Arbeiten_, Semper, Vol. III.

_Excretory organs and genital ducts in the adult._

The kidneys of Scyllium canicula are paired bodies in contact along the median line. They are situated on the dorsal wall of the abdominal cavity, and extend from close to the diaphragm to a point a short way behind the anus. Externally, each appears as a single gland, but by the arrangement of its ducts may be divided into two distinct parts, an anterior and a posterior. The former will be spoken of as the Wolffian body, and the latter as the kidney, from their respective homology with the glands so named in higher Vertebrates. The grounds for these determinations have already been fully dealt with both by Semper[341] and by myself.

Footnote 341: Though Professor Semper has come to the same conclusion as myself with respect to these homologies, yet he calls the Wolffian body Leydig's gland after its distinguished discoverer, and its duct Leydig's duct.

Externally both the Wolffian body and the kidney are more or less clearly divided into segments, and though the breadth of both glands as viewed from the ventral surface is fairly uniform, yet the hinder part of the kidney is very much thicker and bulkier than the anterior part and than the whole of the Wolffian body. In both sexes the Wolffian body is rather longer than the kidney proper. Thus in a male example, 33 centimetres long, the two glands together measured 8-1/4 centimetres and the kidney proper only 3-1/2. In the male the Wolffian bodies extend somewhat further forwards than in the female. Leaving the finer details of the glands for subsequent treatment, I pass at once to their ducts. These differ slightly in the two sexes, so that it will be more convenient to take the male and female separately.

A partly diagrammatic representation of the kidney and Wolffian body of the male is given on Pl. 20, fig. 1. The secretion of the Wolffian body is carried off by a duct, _the Wolffian duct (w.d.)_, which lies on the ventral surface of the gland, and receives a separate ductule from each segment (Pl. 20, fig. 5). The main function of the Wolffian duct in the male is, however, that of a vas deferens. The testicular products are brought to it through the coils of the anterior segments of the Wolffian body by a number of vasa efferentia, the arrangement of which is treated of on pp. 487, 488. The section of the Wolffian duct which overlies the Wolffian body is much contorted, and in adult individuals at the generative period enormously so. The duct often presents one or two contortions beyond the hind end of the Wolffian body, but in the normal condition takes a straight course from this point to the unpaired urinogenital cloaca, into which it falls independently of its fellow of the opposite side. It receives no feeders from the kidney proper.

The excretion of the kidney proper is carried off not by a single duct, but by a series of more or less independent ducts, which, in accordance with Prof. Semper's nomenclature, will be spoken of as _ureters_. These are very minute, and their investigation requires some care. I have reason, from my examinations of this and other species of Elasmobranchii, to believe that they are, moreover, subject to considerable variations, and the following description applies to a definite individual. Nine or possibly ten distinct ureters, whose arrangement is diagrammatically represented in fig. 1, Pl. 20, were present on each side. It will be noticed that, whereas the five hindermost are distinct till close to their openings into the urinogenital cloaca, the four anterior ones appear to unite at once into a single duct, but are probably only bound up in a common sheath. The ureters fall into the common urinogenital cloaca, immediately behind the opening of the Wolffian duct (so far as could be determined), by four apertures on each side. In a section made through the part of the wall of the cloaca containing the openings of the ureters of both sides, there were present on the left side (where the section passed nearer to the surface than on the right) four small openings posteriorly, viz. the openings of the ureters and one larger one anteriorly, viz. the opening of the Wolffian duct. On the other side of the section where the level was rather deeper, there were five distinct ducts cut through, one of which was almost on the point of dividing into two. This second section proves that, in this instance at least, the two ureters did not unite till just before opening into the urinogenital cloaca. The same section also appeared to shew that one of the ureters fell not into the cloaca but into the Wolffian duct.

As stated above both the Wolffian duct and the ureters fall into an unpaired urinogenital cloaca. This cloaca communicates at one end with the general cloaca by a single aperture situated at the point of a somewhat conspicuous papilla, just behind the anus (Pl. 20, fig. 1, _o_), and on the other it opens freely into a pair of bladders, situated in close contact with each other, on the ventral side of the kidney (Pl. 20, fig. 1, _sb_). To these bladders Professor Semper has given the name _uterus masculinus_, from having supposed them to correspond with the lower part of the oviducts of the female. This homology he now admits to be erroneous, and it will accordingly be better to drop the name uterus masculinus, for which may be substituted _seminal bladder_--a name which suits their function, since they are usually filled with semen at the generation season. The seminal bladders communicate with the urinogenital cloaca by wide openings, and it is on the borders of these openings that the mouths of the Wolffian duct and ureters must be looked for. My embryological investigations, though they have not been specially directed to this point, seem to shew that the seminal bladders do not arise during embryonic life, and are still absent in very young individuals. It seems probable that both the bladders and the urinogenital cloaca are products of the lower extremities of the Wolffian duct. The only other duct requiring any notice in the male is the rudimentary oviduct. As was first shewn by Semper, rudiments of the upper extremities of the oviducts, with their abdominal openings, are to be found in the male in the same position as in the female, on the front surface of the liver.

In the female the same ducts are present as in the male, viz. the Wolffian duct and the ureters. The part of the Wolffian duct which receives the secretion of the Wolffian body is not contorted, but is otherwise similar to the homologous part of the Wolffian duct in the male. The Wolffian ducts of the two sides fall independently into an unpaired urinal cloaca, but their lower ends, instead of remaining simple as in the male, become dilated into urinary bladders. Vide Pl. 20, fig. 2. There were nine ureters in the example dissected, whose arrangement did not differ greatly from that in the male--the hinder ones remaining distinct from each other, but a certain amount of fusion, the extent of which could not be quite certainly ascertained, taking place between the anterior ones. The arrangement of the openings of these ducts is not quite the same as in the male. A somewhat magnified representation of it is given in Pl. 20, fig. 3, _o.u._ The two Wolffian ducts meet at so acute an angle that their hindermost extremities are only separated by a septum. In the region of this septum on the inner walls of the two Wolffian ducts were situated the openings of the ureters, of which there were five on each side arranged linearly. In a second example, also adult, I found four distinct openings on each side similarly arranged to those in the specimen described. Professor Semper states that all the ureters in the female unite into a _single duct_ before opening into the Wolffian duct. It will certainly surprise me to find such great variations in different individuals of this species as is implied by the discrepancy between Professor Semper's description and my own.

The main difference between the ureters in the male and female consists in their falling into the urinogenital cloaca in the former and into the Wolffian duct in the latter. Since, however, the urinogenital cloaca is a derivative of the Wolffian duct, this difference between the two sexes is not a very important one. The urinary cloaca opens, in the female, into the general cloaca by a median papilla of somewhat smaller dimensions than the corresponding papilla in the male. Seminal bladders are absent in the female, though possibly represented by the bladder-like dilatations of the Wolffian duct. The oviducts, whose anatomy is too well known to need description, open independently into the general cloaca.

Since the publication of Professor Semper's researches on the urinogenital system of Elasmobranch fishes, it has been well known that, in most adult Elasmobranchii, there are present a series of funnel-shaped openings, leading from the perivisceral cavity, by the intermediation of a short canal, into the glandular tubuli of the kidney. These openings are called by Professor Semper, _Segmentaltrichter_, and by Dr Spengel, in his valuable work on the urogenital system of Amphibia, _Nephrostomen_. In the present work the openings will be spoken of as segmental openings, and the tubes connected with them as segmental tubes. Of these openings there are a considerable number in the adults of both sexes of Scy. canicula, situated along the inner border of each kidney. The majority of them belong to the Wolffian body, though absent in the extreme anterior part of this. In very young examples a few certainly belong to the region of the kidney proper. Where present, there is one for each segment[342]. It is not easy to make certain of their exact number. In one male I counted thirteen. In the female it is more difficult than in the male to make this out with certainty, but in one young example, which had left the egg but a short time, there appeared to be at least fourteen present. According to Semper there are thirteen funnels in both sexes--a number which fairly well agrees with my own results. In the male, rudiments of segmental tubes are present in all the anterior segments of the Wolffian body behind the vasa efferentia, but it is not till about the tenth segment that the first complete one is present. In the female a somewhat smaller number of the anterior segments, six or seven, are without segmental tubes, or only possess them in a rudimentary condition.

Footnote 342: The term segment will be more accurately defined below.

A typical segment of the Wolffian body or kidney, in the sense in which this term has been used above, consists of a number of factors, each of which will be considered in detail with reference to its variations. On Pl. 20, fig. 5, is represented a portion of the Wolffian body with three complete segments and part of a fourth. If one of these be selected, it will be seen to commence with (1) a segmental opening, somewhat oval in form (_st.o_) and leading directly into (2) a narrow tube, the segmental tube, which takes a more or less oblique course backwards, and, passing superficially to the Wolffian duct (_w.d_), opens into (3) a Malpighian body (_p.mg_) at the anterior extremity of an isolated coil of glandular tubuli. This coil forms the fourth section of each segment, and starts from the Malpighian body. It consists of a considerable number of rather definite convolutions, and after uniting with tubuli from one or two (according to size of the segment) accessory Malpighian bodies (_a.mg_), smaller than the one into which the segmental tube falls, eventually opens by a (5) narrowish tube into the Wolffian duct at the posterior end of the segment. Each segment is completely isolated (except for certain rudimentary structures to be alluded to shortly) from the adjoining ones, _and never has more than one segmental tube and one communication with the Wolffian duct_.

The number and general arrangement of the segmental tubes have already been spoken of. Their openings into the body-cavity are, in Scyllium, very small, much more so than in the majority of Elasmobranchii. The general appearance of a segmental tube and its opening is somewhat that of a spoon, in which the handle represents the segmental tube, and the bowl the segmental opening. Usually amongst Elasmobranchii the openings and tubes are ciliated, but I have not determined whether this is the case in Scy. canicula, and Semper does not speak definitely on this point. From the segmental openings proceed the segmental tubes, which in the front segments have nearly a transverse direction, but in the posterior ones are directed more and more obliquely backwards. This statement applies to both sexes, but the obliquity is greater in the female than in the male.

As has been said, each segmental tube normally opens into a Malpighian body, from which again there proceeds the tubulus, the convolutions of which form the main mass of each segment. This feature can be easily seen in the case of the Malpighian bodies of the anterior part of the Wolffian gland in young examples, and sometimes fairly well in old ones, of either sex[343]. There is generally in each segment a second Malpighian body, which forms the commencement of a tubulus joining that from the primary Malpighian body, and, where the segments are larger, there are three, and possibly in the hinder segments of the Wolffian gland and segments of the kidney proper, more than three Malpighian bodies.

Footnote 343: My observations on this subject completely disprove, if it is necessary to do so after Professor Semper's investigations, the statement of Dr Meyer, that segmental tubes in Scyllium open into lymph organs.

The accessory Malpighian bodies, or at any rate one of them, appear to have curious relations to the segmental tubes. The necks of some of the anterior segmental tubes (Pl. 20, fig. 5) close to their openings into the primary Malpighian bodies are provided with a small knob of cells which points towards the preceding segment and is usually connected with it by a fibrous band. This knob is most conspicuous in the male, and in very young animals or almost ripe embryos. In several instances in a ripe male embryo it appeared to me to have a lumen, and to be continued directly forwards into the accessory Malpighian body of the preceding segment. One such case is figured in the middle segment on Pl. 20, fig. 5. In this embryo segmental tubes were present in the segments immediately succeeding those connected with the vasa efferentia, and at the same time these segments contained ordinary and accessory Malpighian bodies. The segmental tubes of these segments were not, however, connected with the Malpighian body of their proper segment, but instead, turned forwards and entered the segment in front of that to which they properly belonged. I failed to trace them quite definitely to the accessory Malpighian body of the preceding segment, but, in one instance at least, there appeared to me to be present a fibrous connection, which is shewn in the figure already referred to, Pl. 20, fig. 5, _r.st_. In any case it can hardly be doubted that this peculiarity of the foremost segmental tubes is related to what would seem to be the normal arrangement in the next few succeeding segments, where each segmental tube is connected with a Malpighian body in its own segment, and more or less distinctly with an accessory Malpighian body in the preceding segment.

In the male the anterior segmental tubes, which even in the embryo exhibit signs of atrophy, become in the adult completely aborted (as has been already shewn by Semper), and remain as irregular tubes closed at both ends, which for the most part do not extend beyond the Wolffian duct (Pl. 20, fig. 4, _r.st_). In the adult, the first two or three segments with these aborted tubes contain only accessory Malpighian bodies; the remaining segments, with aborted segmental tubes, both secondary and primary Malpighian bodies. In neither case are the Malpighian bodies connected with the aborted tubes.

The Malpighian bodies in Scyllium present no special peculiarities. The outer layer of their capsule is for the most part formed of flattened cells; but, between the opening of the segmental tube and the efferent tubulus of the kidney, their cells become columnar. Vide Pl. 20, fig. 5. The convoluted tubuli continuous with them are, I believe, ciliated in their proximal section, but I have not made careful investigations with reference to their finer structure. Each segment is connected with the Wolffian duct by a single tube at the hinder end of the segment. In the kidney proper, these tubes become greatly prolonged, and form the ureters.

It has already been stated that the semen is carried by vasa efferentia from the testes to the anterior segments of the Wolffian body, and thence through the coils of the Wolffian body to the Wolffian duct. The nature of the vasa will be discussed in the embryological section of this chapter: I shall here confine myself to a simple description of their anatomical relations. The consideration of their connections naturally falls under three heads: (1) the vasa efferentia passing from the testes to the Wolffian body, (2) the mode in which these are connected with the Wolffian body, and (3) with the testis.

In Pl. 20, fig. 4, drawn for me from nature by my friend Mr Haddon, are shewn the vasa efferentia and their junctions both with the testes and the kidney. This figure illustrates better than any description the anatomy of the various parts. Behind there are two simple vasa efferentia (_v.e._) and in front a complicated network of vasa, which might be regarded as formed of either two or four main vessels. It will be shewn in the sequel that it is really formed of four distinct vessels. Professor Semper states that there is but a single vas efferens in Scyllium canicula, a statement which appears to me unquestionably erroneous. All the vasa efferentia fall into a _longitudinal duct (l.c)_, which is connected in succession with the several segments of the Wolffian body (one for each vas efferens) which appertain to the testis. The hind end of the longitudinal duct is simple, and ends blindly close to its junction with the last vas efferens; but in front, where the vasa efferentia are complicated, the longitudinal duct also has a complicated constitution, and forms a network rather than a simple tube. It typically sends off a duct to join the coils of the Wolffian body between each pair of vasa efferentia, and is usually swollen where this duct parts from it. A duct similar to this has been described by Semper as _Nierenrandcanal_ in several Elasmobranchii, but its existence is expressly denied in the case of Scyllium! It is usually found in Amphibia, as we know from Bidder and Spengel's researches. Spengel calls it _Längscanal des Hoden_; the vessels from it into the kidney he calls _vasa efferentia_, and the vessels to it, which I speak of as vasa efferentia, he calls _Quercanale_.

The exact mode of junction of the separate vasa efferentia with the testis is difficult to make out on account of the opacity of the basal portion of the testis. My figure shews that there is a network of tubes (formed of four main tubes connected by transverse branches) which is a continuation of the anterior vasa efferentia, and joined by the two posterior ones. These tubes receive the tubuli coming from the testicular ampullæ. The whole network may be called, with Semper, the _testicular network_. While its general relations are represented in my figure, the opacity of the testes was too great to allow of all the details being with certainty filled in.

The kidneys of Scyllium stellare, as might be expected, closely resemble those of Scy. canicula. The ducts of the kidney proper, have, in the former species, a larger number of distinct openings into the urinogenital cloaca. In two male examples I counted seven distinct ureters, though it is not impossible that there may have been one or two more present. In one of my examples the ureters had seven distinct openings into the cloaca, in the other five openings. In a female I counted eleven ureters opening into the Wolffian duct by seven distinct openings. In the remaining parts of the excretory organs the two species of Scyllium resemble each other very closely.

As may be gathered from Prof. Semper's monograph, the excretory organs of Scyllium canicula are fairly typical for Elasmobranchii generally. The division into kidney and Wolffian body is universal. The segmental openings may be more numerous and larger, _e.g._ Acanthias and Squatina, or absent in the adult, _e.g._ Mustelus and Raja. Bladder-like swellings of the Wolffian duct in the female appear to be exceptional, and seminal bladders are not always present. The variations in the ureters and their openings are considerable, and in some cases all the ureters are stated to fall into a single duct, which may be spoken of as the ureter _par excellence_[344], with the same relations to the kidneys as the Wolffian duct bears to the Wolffian body. In some cases Malpighian corpuscles are completely absent in the Wolffian body, _e.g._ Raja.

Footnote 344: I feel considerable hesitation in accepting Semper's descriptions of the ureters and their openings. It has been shewn above that for Scyllium his statements are probably inaccurate, and in other instances, _e.g._ Raja, I cannot bring my dissections to harmonise with his descriptions.

The vasa efferentia of the testes in Scyllium are very typical, but there are some forms in which they are more numerous as well as others in which they are less so. Perhaps the vasa efferentia are seen in their most typical form in Centrina as described and figured (Pl. XXI) by Professor Semper, or in Squatina vulgaris, as I find it, and have represented it on Pl. 20, fig. 8. From my figure, representing the anterior part of the Wolffian body of a nearly ripe embryo, it will be seen that there are five vasa efferentia (_v.e_) connected on the one hand with a longitudinal canal at the base of the testes (_n.t_) and on the other with a longitudinal canal in the Wolffian body. Connected with the second longitudinal canal are four Malpighian bodies, three of them stalked and one sessile; from which again proceed tubes forming the commencements of the coils of the anterior segments of the Wolffian body. These Malpighian bodies are clearly my primary Malpighian bodies, but there are in Squatina, even in the generative segments, secondary Malpighian bodies. What Semper has described for Centrina and one or two other genera, closely correspond with what is present in Squatina.

_Development of the Segmental Tubes._

On p. 345, _et seq._ an account was given of the first formation of the segmental tubes and the segmental duct, and the history of these bodies was carried on till nearly the period at which it is taken up in the exhaustive Memoir of Professor Semper. Though the succeeding narration traverses to a great extent the same ground as Semper's Memoir, yet many points are treated somewhat differently, and others are dealt with which do not find a place in the latter. In the majority of instances, attention is called to points on which my results either agree with, or are opposed to, those of Professor Semper.

From previous statements it has been rendered clear that _at first_ the excretory organs of Elasmobranchii exhibit no division into Wolffian body or kidney proper. Since this distinction is merely a question of the ducts, and does not concern the glandular tubuli, no allusion is made to its appearance in the present section, which deals only with the glandular part of the kidneys and not with their ducts.

Up to the close of stage K the urinogenital organs consist of a segmental duct opening in front into the body-cavity, and terminating blindly behind in close contact with the cloaca, and of a series of segmental tubes, each opening into the body-cavity on the inner side of the segmental duct, but ending blindly at their opposite extremities. It is with these latter that we have at present to deal. They are from the first directed obliquely backwards, and coil close round the inner and dorsal sides of the segmental duct. Where they are in contact (close to their openings into the body-cavity) with the segmental duct, the lumen of the latter diminishes and so comes to exhibit regular alternations of size. This is shewn in Pl. 12, fig. 18, _s.d_. At the points where the segmental duct has a larger lumen, it eventually unites with the segmental tubes.

The segmental tubes rapidly undergo a series of changes, the character of which may be investigated, either by piecing together transverse sections, or more easily from longitudinal and vertical sections. They acquire a Lambda-shaped form with an anterior limb opening into the body-cavity and posterior limb, resting on a dilated portion of the segmental duct. The next important change which they undergo consists in a junction being effected between their posterior limbs and the segmental duct. In the anterior part of the body these junctions appear before the commencement of stage L. A segmental tube at this stage is shewn in longitudinal section on Pl. 21, fig. 7_a_, and in transverse section on Pl. 18, fig. 2. In the former the actual openings into the body-cavity are not visible. In the transverse section only one limb of the Lambda is met with on either side of the section; the limb opening into the body-cavity is seen on the left side, and that opening into the segmental duct on the right side. This becomes quite intelligible from a comparison with the longitudinal section, which demonstrates that it is clearly not possible to see more than a single limb of the Lambda in any transverse section.

After the formation of their junctions with the segmental duct, other changes soon take place in the segmental tubes. By the close of stage L four distinct divisions may be noticed in each tube. Firstly, there is the opening into the body-cavity, with a somewhat narrow stalk, to which the name segmental tube will be strictly confined in the future, while the whole products of the original segmental tube will be spoken of as a segment of the kidney. This narrow stalk opens into a vesicle (Pl. 18, fig. 2, and 21, fig. 6), which forms the second division. From the vesicle proceeds a narrower section forming the third division, which during stage L remains very short, though in later stages it grows with great rapidity. It leads into the fourth division, which constitutes the posterior limb of the Lambda, and has the form of a dilated tube with a narrow opening into the segmental duct.

The subsequent changes of each segment do not for the most part call for much attention. They consist mainly in the elongation of the third division, and its conversion into a coiled tubulus, which then constitutes the main mass of each segment of the kidney. There are, however, two points of some interest, viz. (1) the formation of the Malpighian bodies, and (2) the establishment of the connection between each segmental tube and the tubulus of the preceding segment which was alluded to in the description on p. 486. The development of the Malpighian body is intimately linked with that of the secondary connection between two segments. They are both products of the metamorphosis of the vesicle which forms the termination of the segmental tube proper.

At about stage O this vesicle grows out in two directions (Pl. 21, fig. 10), viz. towards the segment in front (_p.x_) and posteriorly into the segment of which it properly forms a part (_mg_). That portion which grows backward remains continuous with the third division of its proper segment, and becomes converted into a Malpighian body. It assumes (Pl. 21, figs. 6 and 10) a hemispherical form, while near one edge of it is the opening from a segmental tube, and near the other the opening leading into a tubulus of the kidney. The two-walled hemisphere soon grows into a nearly closed sphere, with a central cavity into which projects a vascular tuft. For this tuft the thickened inner wall of cells forms a lining, and at the same time the outer wall becomes thinner, and formed of flattened cells, except in the interval between the openings of the segmental tube and kidney tubulus, where its cells remain columnar.

The above account of the formation of the Malpighian bodies agrees very well with the description which Pye[345] has given of the formation of these bodies in the embryonic Mammalian kidney. My statements also agree with those of Semper, in attributing the formation of the Malpighian body to a metamorphosis of part of the vesicle at the end of the segmental tube. Semper does not however enter into full details on this subject.

Footnote 345: _Journal of Anatomy and Physiology_, Vol. IX.

The elucidation of the history of the second outgrowth from the original vesicle towards the preceding segment is fraught with considerable difficulties, which might no doubt be overcome by a patient investigation of ample material, but which I have not succeeded in fully accomplishing.

The points which I believe myself to have determined are illustrated by fig. 10, Pl. 21, a longitudinal vertical section through a portion of the kidney between stages O and P. In this figure parts of three segments of the kidney are represented. In the hindermost of the three--the one to the right--there is a complete segmental tube (_s.t_) which opens at its upper extremity into an irregular vesicle, prolonged _behind_ into a body which is obviously a developing Malpighian body, _m.g_, and in _front_ into a wide tube cut obliquely in the section and ending apparently blindly (_p.x_). In the preceding segment there is also a segmental tube (_s.t_) whose opening into the body-cavity passes out of the plane of the section, but which is again connected with a vesicle dilating behind into a Malpighian body (_m.g_) and in front into the irregular tube (_p.x_), as in the succeeding segment, _but this tube is now connected_ (and this could be still more completely seen in the segment in front of this) _with a vesicle which opens into the thick-walled collecting tube (fourth division) of the preceding segment_ close to the opening of the latter into the Wolffian duct. The fact that the anterior prolongation of the vesicle ends blindly in the hinder-most segment is due of course to its terminal part passing out of the plane of the section. _Thus we have established between stages O and P a connection between each segmental tube and the collecting tube of the segment in front of that to which it properly belongs; and it further appears that in consequence of this each segment of the kidney contains two distinct coils of tubuli which only unite close to their common opening into the Wolffian duct!_

This remarkable connection is not without morphological interest, but I am unfortunately only able to give in a fragmentary manner its further history. During the greater part of embryonic life a large amount of interstitial tissue is present in the embryonic kidneys, and renders them too opaque to be advantageously studied as a whole; and I have also, so far, failed to prepare longitudinal sections suitable for the study of this connection. It thus results that the next stage I have satisfactorily investigated is that of a nearly ripe embryo already spoken of in connection with the adult, and represented on Pl. 20, fig. 5. This figure shews that each segmental tube, while distinctly connected with the Malpighian body of its own segment, also sends out a branch towards the secondary Malpighian body of the preceding segment. This branch in most cases appeared to be rudimentary, and in the adult is certainly not represented by more than a fibrous band, but I fancy that I have been able to trace it (though not with the distinctness I could desire) in surface views of the embryonic kidney of stage Q. _The condition of the Wolffian body represented on Pl. 20, fig. 5 renders it probable that the accessory Malpighian body in each segment is developed in connection with the anterior growth from the original vesicle at the end of the segmental tube of the succeeding segment._ How the third or fourth accessory Malpighian bodies, when present, take their origin I have not made out. It is, however, fairly certain that they form the commencement of two additional coils which unite, like the coil connected with the first accessory Malpighian body, with the collecting tube of the primitive coil close to its opening into the Wolffian duct or ureter.

The connection above described between two successive kidney segments appears to have escaped Professor Semper's notice, though I fancy that the peculiar vesicle he describes, _loc. cit._ p. 303, as connected with the end of each segmental tube, is in some way related to it. It seems possible that the secondary connection between the segmental tube and the preceding segment may explain a peculiar observation of Dr Spengel[346] on the kidney of the tailless Amphibians. He finds that, in this group, the segmental tubes do not open into Malpighian bodies, but into the fourth division of the kidney tube. Is it not just possible that in this case the primitive attachment of the segmental tubes may have become lost, and a secondary attachment, equivalent to that above described, though without the development of a secondary Malpighian body, have been developed? In my embryos the secondary coil of the segmental tubes opens, as in the Anura, into the fourth section of a kidney tubulus.

Footnote 346: _Loc. cit._ pp. 85-89.

_Development of the Müllerian and Wolffian ducts._

The formation of the Müllerian and Wolffian ducts out of the original segmental duct has been dealt with in a masterly manner by Professor Semper, but though I give my entire assent to his general conclusions, yet there are a few points on which I differ from him. These are for the most part of a secondary importance; but they have a certain bearing on the homology between the Müllerian duct of higher Vertebrates and that of Elasmobranchii. The following account refers to Scy. canicula, but so far as my observations go, the changes in Scy. stellare are nearly identical in character.

I propose treating the development of these ducts in the two sexes separately, and begin with the female.

Shortly before stage N a horizontal split arises in the segmental duct[347], commencing some little distance from its anterior extremity, and extending backwards. This split divides the duct into a dorsal section and a ventral one. The dorsal section forms the Wolffian duct, and receives the openings of the segmental tubes, and the ventral one forms the Müllerian duct or oviduct, and is continuous with the unsplit anterior part of the primitive segmental duct, which opens into the body-cavity. The nature of the splitting may be gathered from the woodcut, fig. 6, p. 511, where _x_ represents the line along which the segmental duct is divided. The splitting of the primitive duct extends slowly backwards, and thus there is for a considerable period a single duct behind, which bifurcates in front. A series of transverse sections through the point of bifurcation always exhibits the following features. Anteriorly two separate ducts are present, next two ducts in close juxtaposition, and immediately behind this a single duct. A series of sections through the junction of two ducts is represented on Plate 21, figs. 1A, 1B, 1C, 1D.

Footnote 347: For the development of the segmental duct, vide p. 345, _et seq._

In my youngest example, in which the splitting had commenced, there were two separate ducts for only 14 sections, and in a slightly older one for about 18. In the second of these embryos the part of the segmental duct anterior to the front end of the Wolffian duct, which is converted directly into the oviduct, extended through 48 sections. In the space included in these 48 sections at least five, and I believe six, segmental tubes with openings into the body-cavity were present. These segmental tubes did not however unite with the oviduct, or at best, but one or two rudimentary junctions were visible, and the evidence of my earlier embryos appears to shew that the segmental tubes in front of the Wolffian duct never become in the female united with the segmental duct. The anterior end of the Wolffian duct is very much smaller than the oviduct adjoining it, and as the reverse holds good in the male, an easy method is afforded of distinguishing the two sexes even at the earliest period of the formation of the Wolffian duct.

Hitherto merely the general features of the development of the oviduct and Wolffian duct have been alluded to, but a careful inspection of any good series of sections, shewing the junction of these two ducts, brings to light some features worth noticing in the formation of the oviduct. It might have been anticipated that, where the two ducts unite behind as the segmental duct, their lumens would have nearly the same diameter, but normally this appears to be far from the case.

To illustrate the formation of the oviduct I have represented a series of sections through a junction in an embryo in which the splitting into two ducts had only just commenced (Pl. 21, fig. 1), but I have found that the features of this series of sections are exactly reproduced in other series in which the splitting has extended as far back as the end of the small intestine. In the series represented (Pl. 21) 1A is the foremost section, and 1D the hindermost. In 1A the oviduct (_od_) is as large or slightly larger than the Wolffian duct (_w.d_), and in the section in front of this (which I have not represented) was considerably the larger of the two ducts. In 1B the oviduct has become markedly smaller, but there is no indication of its lumen becoming united with that of the Wolffian duct--the two ducts, though in contact, are distinctly separate. In 1C the walls of the two ducts have fused, and the oviduct appears merely as a ridge on the under surface of the Wolffian duct, and its lumen, though extremely minute, _shews no sign of becoming one with that of the Wolffian duct_. Finally, in 1D the oviduct can merely be recognised as a thickening on the under side of the segmental duct, as we must now call the single duct, but a slight bulging downwards of the lumen of the segmental duct appears to indicate that the lumens of the two ducts may perhaps have actually united. But of this I could not be by any means certain, and it seems quite possible that the lumen of the oviduct never does open into that of the segmental duct.

The above series of sections goes far to prove that the posterior part of the oviduct is developed as a nearly solid ridge split off from the under side of the segmental duct, into which at the utmost a very small portion of the lumen of the latter is continued. One instance has however occurred amongst my sections which probably indicates that the lumen of the segmental duct may sometimes, in the course of the formation of the oviduct and Wolffian duct, become divided into two parts, of which that for the oviduct, though considerably smaller than that for the Wolffian duct, is not so markedly so as in normal cases (Pl. 21, fig. 2).

Professor Semper states that the lumen of the part of the oviduct split off from the hindermost end of the segmental duct becomes continuously smaller, till at last close to the cloaca it is split off as a solid rod of cells without a lumen, and thus it comes about that the oviduct, when formed, ends blindly, and does not open into the cloaca till the period of sexual maturity. My own sections do not include a series shewing the formation of a terminal part of the oviduct, but Semper's statements accord precisely with what might probably take place if my account of the earlier stages in the development of the oviduct is correct. The presence of a hymen in young female Elasmobranchii was first made known by Putmann and Garman[348], and subsequently discovered independently by Semper[349].

Footnote 348: "On the Male and Female Organs of Sharks and Skates, with special reference to the use of the claspers," _Proceed. American Association for Advancement of Science_, 1874.

Footnote 349: _Loc. cit._

The Wolffian duct appears to receive its first segmental tube at its anterior extremity.

In the male the changes of the original segmental duct have a somewhat different character to those in the female, although there is a fundamental agreement between the two sexes. As in the female, a horizontal split makes its appearance a short way behind the front end of the segmental duct, and divides this into a dorsal Wolffian duct and a ventral Müllerian duct, the latter continuous with the anterior section of the segmental duct, which carries the abdominal opening. The differences in development between the two sexes are, in spite of a general similarity, very obvious. In the first place, the ventral portion split off from the segmental duct, instead of being as in the female larger in front than the Wolffian duct, is very much smaller; while behind it does not form a continuous duct, but in some parts a lumen is present, and in others again absent (Pl. 21, fig. 6). It does not even form an unbroken cord, but is divided in disconnected portions. Those parts with a lumen do not appear to open into the Wolffian duct.

The process of splitting extends gradually backwards, so that there is a much longer rudimentary Müllerian duct by stage O than by stage N. By stage P the posterior portions of the Müllerian ducts have vanished. The anterior parts remain, as has been already stated, till adult life. A second difference between the male and female depends on the fact that, in the male, the splitting of the segmental duct into Müllerian duct and Wolffian duct never extends beyond the hinder extremity of the small intestine. A third and rather important point of difference consists in the splitting commencing far nearer the front end of the segmental duct in the male than in the female. In the female it was shewn that about 48 sections intervened between the front end of the segmental duct and the point where this became split, and that this region included five or six segmental tubes. In the male the homologous space only occupies _about 7 to 12 sections, and does not contain the rudiment of more than a single segmental tube_. Although my sections have not an absolutely uniform thickness, yet the above figures suffice to shew in a conclusive manner that the splitting of the segmental duct commences far further forwards in the male than in the female. This difference accounts for two facts which were mentioned in connection with the excretory organs of the adult, viz. (1) the greater length of the Wolffian body in the male than in the female, and (2) the fact that although a nearly similar number of segmental tubes persist in the adults of both sexes, yet that in the male there are five or six more segments in front of the first fully developed segmental opening than in the female.

The above description of the formation of the Müllerian duct in the male agrees very closely with that of Professor Semper for Acanthias. For Scyllium however he denies, as it appears to me erroneously, the existence of the posterior rudimentary parts of the Müllerian duct. He further asserts that the portions of the Müllerian duct with a lumen open into the Wolffian duct. The most important difference, however, between Professor Semper's and my own description consists in his having failed to note that the splitting of the segmental duct commences much further forwards in the male than in the female.

I have attempted to shew that the oviduct in the female, with the exception of the front extremity, is formed as a nearly solid cord split off from the ventral surface of the segmental duct, and not by a simple splitting of the segmental duct into two equal parts. If I am right on this point, it appears to me far easier to understand the relationship between the oviduct or Müllerian duct of Elasmobranchii and the Müllerian duct of Birds, than if Professor Semper's account of the development of the oviduct is the correct one. Both Professor Semper and myself have stated our belief in the homology of the ducts in the two cases, but we have treated their relationship in a very different way. Professor Semper[350] finds himself compelled to reject, on theoretical grounds, the testimony of recent observers on the development of the Müllerian duct in Birds, and to assert that it is formed out of the Wolffian duct, or, according to my nomenclature, 'the segmental duct.' In my account[351], the ordinary statements with reference to the development of the Müllerian duct in Birds are accepted; but it is suggested that the independent development of the Müllerian duct may be explained by the function of this duct in the adult having, as it were, more and more impressed itself upon the embryonic development, till finally all connection, even during embryonic life, between the oviduct and the segmental duct (Wolffian duct) became lost.

Footnote 350: _Loc. cit._ pp. 412, 413.

Footnote 351: "The Urinogenital Organs of Vertebrates," _Journal of Anatomy and Physiology_, Vol. X. p. 47. [This edition, p. 164.]

Since finding what a small portion of the segmental duct became converted into the Müllerian duct in Elasmobranchii, I have reexamined the development of the Müllerian duct in the Fowl, in the hope of finding that its posterior part might develop nearly in the same manner as in Elasmobranchii, at the expense of a thickening of cells on the outer surface of the Wolffian duct. I have satisfied myself, in conjunction with Mr Sedgwick, that this is not the case, and that the general account is in the main true; but at the same time we have obtained evidence which tends to shew that the cells which form the Müllerian duct are in part derived from the walls of the Wolffian duct. We propose giving a full account of our observations on this point, so that I refrain from mentioning further details here. It may however be well to point out that, apart from observations on the actual development of the Müllerian duct in the Bird, the fact of its abdominal opening being situated some way behind the front end of the Wolffian duct, is of itself a sufficient proof that it cannot be the metamorphosed front extremity of the Wolffian (= segmental) duct, in the same way that the abdominal opening of the Müllerian duct is the front extremity of the segmental duct in Elasmobranchii.

Although the evidence I can produce in the case of the Fowl of a direct participation of the Wolffian duct in the formation of the Müllerian is not of an absolutely conclusive kind, yet I am inclined to think that the complete independence of the two ducts, if eventually established as a fact, would not of itself be sufficient (as Semper is inclined to think) to disprove the identity of the Müllerian duct in Birds and Elasmobranchii.

We have, no doubt, almost no knowledge of the magnitude of the changes which can take place in the mode of development of the same organ in different types, yet this would have to be placed at a very low figure indeed in order to exclude the possibility of a change from the mode of development of the Müllerian duct in Elasmobranchii to that in Birds. We have, it appears to me, in the smallness of the portion of the segmental duct which goes to form the Müllerian duct in Elasmobranchii, evidence that a change has already appeared in this group in the direction of a development of the Müllerian duct independent of the segmental duct, and therefore of the Wolffian duct; and it has been in view of this consideration, that I have devoted so much attention to the apparently unimportant point of how much of the segmental duct was concerned in the formation of the Müllerian duct. An analogous change, in a somewhat different direction, would seem to be taking place in the development of the rudimentary Müllerian duct in the male Elasmobranchii.

It is, perhaps, just worth pointing out, that the blindness of the oviduct of female Elasmobranchii, and its mode of development from an imperfect splitting of the segmental duct, may probably be brought into connection with the blindness of the extremity of the Müllerian duct or oviduct which so often occurs in both sexes of Sturgeons (Accipenser).

I may, perhaps, at this point, be permitted to say a few words about my original account of the development of the Wolffian duct This account was incorrect, and based upon a false interpretation of an imperfect series of sections, and I took the opportunity, in a general account of the urinogenital system of Vertebrates, to point out my mistake[352]. Professor Semper has, however, subsequently done me the honour to discuss, at considerable length, my original errors, and to attempt to explain them. Since it appears to me improbable that the continuation of such a discussion can be of much general interest, it will suffice to say now, that both Professor Semper's and my own original statements on the development of the Wolffian duct were erroneous; but that both of us have now recognised our mistakes; and that the first morphologically correct account of the development was given by him.

Footnote 352: _Journal of Anatomy and Physiology_, Vol X. 1875. [This edition, No. VII.]

* * * * *

With reference to the formation of the urinal cloaca there is not much to say. The originally widely separated openings of the two Wolffian ducts gradually approximate in both sexes. By stage O (Pl. 19, fig. 1_b_) they are in close contact, and the lower ends of the two ducts actually coalesce at a somewhat later period, and open by a single aperture into the common cloaca. The papilla on which this is situated begins to make its appearance considerably before the actual fusion of the lower extremities of the two ducts.

_Formation of Wolffian Body and Kidney proper._

Between stages L and M the hindermost ten or eleven segments of the primitive undivided excretory organ commence to undergo changes which result in their separation from the anterior segments as a distinct gland, which was spoken of in the description of the adult as the kidney proper, while the unaltered preceding segments of the kidney were spoken of as the Wolffian body.

It will be remembered that each segment of the embryonic kidney consists of four divisions, the last or fourth of which opens into the Wolffian duct. The changes which take place in the hindermost ten or eleven segments, and cause them to become distinguished as the kidney proper, concern alone the fourth division of each segment, which becomes prolonged backwards, and its opening into the Wolffian duct proportionately shifted. These changes affect the foremost segments of the kidney much more than the hindermost, so that the fourth division in the foremost segments becomes very much longer than in the hindermost, and at last all the prolongations of the kidney segments come to open nearly on the same level, close to the cloacal termination of the Wolffian duct (Pl. 21, fig. 8). The prolongations of the fourth division of the kidney-segments have already (p. 481) been spoken of in the description of the adult as ureters, and this name will be employed for them in the present section.

The exact manner in which the changes, that have been briefly related, take place is rather curious, and very difficult to unravel without the aid of longitudinal sections. First of all, the junction between each segment of the kidney and the Wolffian duct becomes so elongated as to occupy the whole interval between the junctions of the two neighbouring segments. The original opening of each tube into the Wolffian duct is situated at the anterior end of this elongated attachment, the remaining part of the attachment being formed solely of a ridge of cells on the dorsal side of the Wolffian duct. The general character of this growth will be understood by comparing figs. 7_a_ and 7_b_, Pl. 21--two longitudinal vertical sections through part of the kidneys. Fig. 7 _a_ shews the normal junction of a segmental tube with the Wolffian duct in the Wolffian body, while in figure 7_b_ (_r.u_) is shewn the modified junction in the region of the kidney proper in the same embryo. The latter of these figures (fig. 7_b_) appears to me to prove that the elongation of the attachments between the segmental tubes and Wolffian duct takes place _entirely at the expense of the former_. Owing to the length of this attachment, every transverse section through the kidney proper at this stage either presents a solid ridge of cells closely adhering to the dorsal side of the Wolffian duct, or else passes through one of the openings into the Wolffian duct.

During stage M the original openings of the segmental tubes into the Wolffian duct appear to me to become obliterated, and at the same time the lumen of each ureter is prolonged into the ridge of cells on the dorsal wall of the duct.

Both of these changes are illustrated in my figures. The fact of the obliteration of the original opening into the Wolffian duct is shewn in longitudinal section in Pl. 21, fig. 9, _u_, but more conclusively in the series of transverse sections represented on Pl. 21, figs. 3A, 3B, 3C. In the hindermost of these (3C) is seen the solid terminal point of a ureter, while the same ureter possesses a lumen in the two previous sections, but exhibits no signs of opening into the Wolffian duct. Sections may however be met with which appear to shew that in some instances the ureters still continue to open into the Wolffian duct, but these I find to be rare and inconclusive, and am inclined to regard them as abnormalities. The prolongation of the lumen of the ureters takes place in a somewhat peculiar fashion. The lumen is not, as might be expected, _completely_ circumscribed by the wall of the ureter, but only _dorsally and to the sides_. Ventrally it is closed in by the dorsal wall of the Wolffian duct. In other words, each ureter is at first an incomplete tube. This peculiarity is clearly shewn in the middle figure of the series on Pl. 21, fig. 3B.

During stages M and N the ureters elongate considerably, and, since the foremost ones grow the most rapidly, they soon come to overlap those behind. As each ureter grows in length it remains an incomplete tube, and its lumen, though proportionately prolonged, continues to present the same general relations as at first. It is circumscribed by its proper walls only dorsally and laterally; its floor being formed in the case of the front ureter by the Wolffian duct, and in the case of each succeeding ureter by the dorsal wall of the ureter in front. This is most easily seen in longitudinal sections, and is represented on Pl. 21, fig. 9, or on a larger scale in fig. 9A. In the latter figure it is especially clear that while the wall on the dorsal side of the lumen of each ureter is continuous with the dorsal wall of the tubulus of its own segment, the wall on the ventral side is continuous with the dorsal wall of the ureter of the preceding segment. This feature in the ureters explains the appearance of transverse sections in which the ureters are not separate from each other, but form together a kind of ridge on the dorsal side of the Wolffian duct, in which there are a series of perforations representing the separate lumens of the ureters (Pl. 21, fig. 4). The peculiarities in the appearance of the dorsal wall of the Wolffian duct in fig. 9A, and the difference between the cells composing it and those of the ventral wall, become intelligible on comparing this figure with the representation of transverse section in figs. 3B and 3C, and especially in fig. 4. Most of the ureters continue to end blindly at the close of stage N, and appear to have solid posterior terminations like that of the Müllerian duct in Birds.

By stage O all the ureters have become prolonged up to the cloacal end of the Wolffian duct, so that the anterior one has a length equal to that of the whole kidney proper. For the most part they acquire independent openings into the end section of the Wolffian duct, though some of them unite together before reaching this. The general appearance of the hindermost of them between stages N and O is shewn in longitudinal and vertical section in Pl. 21, fig. 8, _u_.

They next commence to develop into complete and independent tubes by their side walls growing inwards and meeting below so as to completely enclose their lumen. This is seen already to have occurred in most of the posterior ureters in Pl. 21, fig. 8.

Before stage P the ureters cease to be united into a continuous ridge, and each becomes separated from its neighbours by a layer of indifferent tissue: by this stage, in fact, the ureters have practically attained very nearly their adult condition. The general features of a typical section through them are shewn on Pl. 21, fig. 5. The figure represents the section of a female embryo, not far from the cloaca. Below is the oviduct (_od_). Above this again is the Wolffian duct (_w.d_), and still dorsal to this are four ureters (_u_). In female embryos more than four ureters are not usually to be seen in a single section. This is probably owing to the persistence, in some instances, of the intimate connection between the ureters found at an earlier stage of development, and results in a single ureter coming to serve as the collecting duct for several segments. A section through a male embryo of stage P would mainly differ from that through a female in the absence of the oviduct, and in the presence of probably six[353], instead of four, ureters.

Footnote 353: This at least holds good for one of my embryos at this stage, which is labelled Scy. canicula, but which may possibly be Scy. stellare.

The exact amount of fusion which takes place between the ureters, and the exact number of the ureters, cannot easily be determined from sections, but the study of sections is chiefly of value in shewing the general nature of the changes which take place in the process of attaining the adult condition.

It may be noticed, as a consequence of the above account, that the formation of the ureters takes place by a growth of the original segmental tubes, and not by a splitting off of parts of the wall of the Wolffian duct.

The formation of ureters in Scyllium, which has been only very cursorily alluded to by Professor Semper, appears to differ very considerably from that in Acanthias as narrated by him.

_The Vasa Efferentia._

A comparison of the results of Professor Semper on Elasmobranchii, and Dr Spengel on Amphibians, suggests several interesting questions with reference to the development of the vasa efferentia, and the longitudinal canal of the Wolffian body.

Professor Semper was the first to describe the adult anatomy and development of vasa efferentia in Elasmobranchii, and the following extracts will fully illustrate his views with reference to them.

"In[354] dem frühesten Stadium finden sich wie früher angegeben ungefahr 34 Trichter in der Leibeshöhle, von diesen gehen die 27 hintersten in die persistirenden Segmentaltrichter über, von denen 4 beim erwachsenen Thiere auf dem Mesorchium stehen. Die übrigen 7 schliessen sich vollständig ab zu den erwähnten länglichen und später mannigfach auswachsenden varicösen Trichterblasen; von diesen sind es wiederum 3-4 welche untereinander in der Längsrichtung verwachsen und dadurch den in der Basis der Hodenfalte verlaufenden Centralcanal des Hodens bilden. Ehe aber diese Verwachsung zu einem mehr oder minder geschlängelten Centralcanal vollständig wird, hat sich einmal das Lumen der Trichterblasen fast vollständig geschlossen und ausserdem von ihnen aus durch Verwachsung und Knospung die erste Anlage des rete vasculosum Halleri gebildet (Taf. XX. Figs. 1, 2_c_). Es erstreckt sich nämlich mehr oder minder weit in die Genitalfalte hinein ein unregelmässiges von kleinen Zellen begränztes Canalnetz welches zweifellos mit dem noch nicht ganz vollständigen Centralcanale des Hodens (Taf. XX. Fig. 2_c_) in Verbindung steht. Von diesem letzteren aus gehen in regelmässigen Abständen die Segmentalgänge (Taf. XX. Fig. 2 _sg._) gegen die Niere hin; da sie meist stark geneigt oder selbst geschlängelt (bei 6{ctm} langen Embryonen) gegen die Niere zu verlaufen, wo sie sich an die primären _Malpighi_'schen Körperchen und deren Bildungsblasen ansetzen, so kann ein verticaler Querschnitt auch nie einen solchen nun zum vas efferens gewordenen Segmentalgang seiner ganzen Länge nach treffen. Gegen die Trichterfurche zu aber steht namentlich am hinteren Theile der Genitalfalte der Centralcanal häufig noch durch einen kurzen Zellstrang mit dem Keimepithel der Trichterfurche in Verbindung; mitunter findet sich hier sogar noch eine kleine Höhlung, Rest des ursprünglich hier vorhandenen weiten Trichters" (Taf. XX. Fig. 3_c_).

Footnote 354: _Loc. cit._ p. 364.

And again: "Dieser[355] Gegensatz in der Umbildung der Segmentalgänge an der Hodenbasis scheint nun mit einem anderen Hand in Hand zu gehen. Es bildet sich nämlich am Innenrande der Niere durch Sprossung und Verwachsung der Segmentalgänge vor ihrer Insertion an das primäre _Malpighi_'sche Körperchen ein Canal beim Männchen aus, den ich als _Nierenrandcanal_ oben bezeichnet habe. Ich habe denselben bei Acanthias Centrina (Taf. XXI. Fig. 13) und Mustelus (Taf. XV. Fig. 8) gefunden. Bei Centrina ist er ziemlich lang und vereinigt mindestens 7 Segmentalgänge, aber von diesen letzteren stehen nur 5 mit dem Hodennetz in Verbindung. Dort nun wo diese letzteren sich an den Nierenrandcanal ansetzen (Taf. XXI. Fig. 13 sg.1-sg.5) findet sich jedesmal ein typisch ausgebildetes _Malpighi_'sches Körperchen, mit dem aber nun nicht mehr wie ursprünglich nur 2 Canäle verbunden sind (Taf. XXI. Fig. 14) sondern 3. Einer dieser letzteren ist derjenige Ast des Nierenrandcanals welcher die Verbindung mit dem nächst folgenden Segmentalgang zu besorgen hat. An den Stellen aber wo sich an den Nierenrandcanal die hinteren blind gegen den Hoden hin endenden Segmentalgänge ansetzen fehlen diese _Malpighi_'schen Körperchen (Taf. XXI. Fig. 13 _sg_7) vollständig. Auch bei Mustelus (Taf. XV. Figs. 8, 10) findet genau dasselbe Verhältniss statt; da aber hier nur 2 (oder 3) Segmentalgänge zu vasa efferentia umgewandelt werden, so stehen hier am kurzen Randcanal der Niere auch nur 2 oder 3 _Malpighi_'sche Körperchen. Diese aber sind typisch ausgebildet" (Taf. XV. Fig. 10).

Footnote 355: _Loc. cit._ p. 395.

From these two extracts it is clear that Semper regards both the vasa efferentia, and central canal of the testis network, as well as the longitudinal canal of the Wolffian body, as products of the anterior segmental tubes.

The appearance of these various parts in the fully grown embryos or adults of such genera as Acanthias and Squatina strongly favours this view, but Semper appears to have worked out the development of these structures somewhat partially and by means of sections, a method not, in Scyllium at least, very suitable for this particular investigation. I myself at first unhesitatingly accepted Semper's views, and it was not till after the study of the paper of Dr Spengel on the Amphibian kidney that I came to have my doubts as to their accuracy. The arrangement of the parts in most Amphibians is strikingly similar to that in Elasmobranchii. From the testis come transverse canals corresponding with my vasa efferentia; these fall into a longitudinal canal of the kidneys, from which again, as in Squatina (Pl. 20, fig. 8), Mustelus and Centrina, canals (the vasa efferentia of Spengel) pass off to Malpighian bodies. So far there is no difficulty, but Dr Spengel has made the extremely important discovery, that in young Amphibians each Malpighian body in the region of the generative ducts, in addition to receiving the vasa efferentia, is connected with a fully developed segmental tube opening into the body-cavity. In Amphibians, therefore, it is improbable that the vasa efferentia are products of the open extremities of the segmental tubes, considering that these latter are found in their unaltered condition at the same time as the vasa efferentia. When it is borne in mind how strikingly similar in most respects is the arrangement of the testicular ducts in Amphibia and Elasmobranchii, it will not easily be credited that they develop in entirely different methods. Since then we find in Amphibians fully developed segmental tubes in the same segments as the vasa efferentia, it is difficult to believe that in Elasmobranchii the same vasa efferentia have been developed out of the segmental tubes by the obliteration of their openings.

I set myself to the solution of the origin of the vasa efferentia by means of surface views, after the parts had been made transparent in creosote, but I have met with great difficulties, and so far my researches have only been partially successful. From what I have been able to see of Squatina and Acanthias, I am inclined to think that the embryos of either of these genera would form far more suitable objects for this research than Scyllium. I have had a few embryos of Squatina which were unfortunately too old for my purpose.

Very early the vasa efferentia are fully formed, and their arrangement in an embryo eight centimetres long is shewn in Pl. 20, fig. 6, _v.e_. It is there seen that there are six if not seven vasa efferentia connected with a longitudinal canal along the base of the testes (Semper's central canal of the testis), and passing down like the segmental tubes to spaces between the successive segments of the Wolffian body. They were probably connected by a longitudinal canal in the Wolffian body, but this could not be clearly seen. In the segment immediately behind the last vas efferens was a fully developed segmental tube. This embryo clearly throws no light on the question at issue except that on the whole it supports Semper's views. I further failed to make out anything from an examination of still younger embryos.

In a somewhat older embryo there was connected with the anterior vas efferens a peculiar structure represented on Pl. 20, fig. 7, _r.st_? which strangely resembled the opening of an ordinary segmental tube, but as I could not find it in the younger embryo, this suggestion as to its nature, is, at the best, extremely hazardous. If, however, this body really is the remnant of a segmental opening, it would be reasonable to conclude that the vasa efferentia are buds from the segmental tubes as opposed to their openings; a mode of origin which is not incompatible with the discoveries of Dr Spengel. I have noticed a remnant, somewhat similar to that in the Scyllium embryo, close to the hindermost vas efferens in an embryo Squatina (Pl. 20, fig. 8, _r.st_?).

With reference to the development of the longitudinal canal of the Wolffian body, I am without observations, but it appears to me to be probably a further development of the outgrowths of the vesicles of each segmental tube, which were described in connection with the development of the segmental tubes, p. 492. Were an anterior outgrowth of one vesicle to meet and coalesce with the posterior outgrowth of the preceding vesicle, a longitudinal canal such as actually exists would be the result. The central canal of the base of the testes and the network connected with it in the adult (Pl. 20, fig. 4), appear to be derivatives of the vasa efferentia.

I am thus compelled to leave open the question of the real nature of the vasa efferentia, but am inclined to regard them as outgrowths from the anterior segmental tubes, though not from their open terminations.

* * * * *

My views upon the homologies of the various parts of the urinogenital system, the development of which has been described in the present chapter, have already been expressed in a paper on Urinogenital organs of Vertebrates[356]. Although Kölliker's[357] discovery of the segmental tubes in Aves, and the researches of Spengel[358], Gasser[359], Ewart[360] and others, have rendered necessary a few corrections in my facts, I still adhere in their entirety to the views expressed in that paper, and feel it unnecessary to repeat them in this place. I conclude the chapter with a résumé of the development of the urinogenital organs in Elasmobranchii from their first appearance to their permanent condition.

Footnote 356: _Journal of Anatomy and Physiology_, Vol. X. [This edition, No. VII.]

Footnote 357: _Entwicklungsgeschichte des Menschen u. der höheren Thiere._

Footnote 358: _Loc. cit._

Footnote 359: _Beiträge zur Entwicklungsg. d. Allantois d. Müller'schen Gänge u. d. Afters._

Footnote 360: "Abdominal Pores and Urogenital Sinus of Lamprey," _Journal of Anatomy and Physiology_, Vol. X. p. 488.

* * * * *

_Résumé._--The first trace of the urinary system makes its appearance as a knob springing from the intermediate cell-mass opposite the fifth protovertebra (woodcut, fig. 5A, _p.d_). This knob is the rudiment of the abdominal opening of the segmental duct, and from it there grows backwards to the level of the anus a solid column of cells, which constitutes the rudiment of the segmental duct itself (woodcut, fig. 5B, _p.d_). The knob projects towards the epiblast, and the column connected with it lies between the mesoblast and epiblast. The knob and column do not long remain solid, but the former acquires an opening into the body-cavity continuous with a lumen, which makes its appearance in the latter.

While the lumen is gradually pushing its way backwards along the solid rudiment of the segmental duct, the first traces of the segmental tubes, or proper excretory organs, make their appearance in the form of solid outgrowths of the intermediate cell-mass, which soon become hollow and open into the body-cavity. Their blind ends curl obliquely backwards round the inner and dorsal side of the segmental duct. One segmental tube makes its appearance for each protovertebra, commencing with that immediately behind the abdominal opening of the segmental duct, the last tube being situated a short way behind the anus. Soon after their formation the blind ends of the segmental tubes open into the segmental duct, and each of them becomes divided into four parts. These are (woodcut 7) (1) a section carrying the abdominal opening or segmental tube proper, (2) a dilated vesicle into which this opens, (3) a coiled tubulus proceeding from (2) and terminating in (4), a wider portion opening into the segmental duct. At the same time, or shortly before this, each segmental duct unites with and opens into one of the horns of the cloaca, and also retires from its primitive position between the epiblast and mesoblast, and assumes a position close to the epithelium lining the body-cavity. The general features of the excretory organs at this period are diagrammatically represented on the woodcut, fig. 6. In this fig. _p.d_ is the segmental duct and _o_ its abdominal opening. _s.t_ points to the segmental tubes, the finer details of whose structure are not represented in the diagram. The kidneys thus form at this period an unbroken gland composed of a series of isolated coiled tubes, one extremity of each of which opens into the body-cavity, and the other into the segmental duct, which forms the only duct of the kidney, and communicates at one end with the body-cavity, and at the other with the cloaca.

The next important change concerns the segmental duct, which becomes longitudinally split into two complete ducts in the female, and one complete duct and parts of a second in the male. The manner in which this takes place is diagrammatically represented in woodcut 6 by the clear line _x_, and in transverse section in woodcut 7. The resulting ducts are the (1) Wolffian duct dorsally, which remains continuous with the excretory tubules of the kidney, and ventrally (2) the oviduct or Müllerian duct in the female, and the rudiments of this duct in the male. In the female the formation of these ducts takes place by a nearly solid rod of cells, being gradually split off from the ventral side of all but the foremost part of the original segmental duct, with the short undivided anterior part of which duct it is continuous in front. Into it a very small portion of the lumen of the original segmental duct is perhaps continued (Pl. 21, fig. 1A, etc.). The remainder of the segmental duct (after the loss of its anterior section and the part split off from its ventral side) forms the Wolffian duct. The process of formation of the ducts in the male chiefly differs from that in the female in the fact of the anterior undivided part of the segmental duct, which forms the front end of the Müllerian duct, being shorter, and in the column of cells with which it is continuous being from the first incomplete.

The tubuli of the primitive excretory organ undergo further important changes. The vesicle at the termination of each segmental tube grows forwards towards the preceding tubulus, and joins the fourth section of it close to the opening into the Wolffian duct (Pl. 21, fig. 10). The remainder of the vesicle becomes converted into a Malpighian body. By the first of these changes a connection is established between the successive segments of the kidney, and though this connection is certainly lost (or only represented by fibrous bands) in the anterior part of the excretory organs in the adult, and very probably in the hinder part, yet it seems most probable that traces of it are to be found in the presence of the secondary Malpighian bodies of the majority of segments, which are most likely developed from it.

Up to this time there has been no distinction between the anterior and posterior tubuli of the primitive excretory organ which alike open into the Wolffian duct. The terminal division of the tubuli of a considerable number of the hindermost of these (ten or eleven in Scyllium canicula), either in some species elongate, overlap, and eventually open by apertures (not usually so numerous as the separate tubes), on nearly the same level, into the hindermost section of the Wolffian duct in the female, or into the urinogenital cloaca, formed by the coalesced terminal parts of the Wolffian ducts, in the male; or in other species become modified in such a manner as to pour their secretion into a single duct on each side, which opens in a position corresponding with the numerous ducts of the other type (woodcut, fig. 8). It seems that both in Amphibians and Elasmobranchii the type with a single duct, or approximations to it, are more often found in the females than in the males. The subject requires however to be more worked out in Elasmobranchii[361]. In both groups the modified posterior kidney-segments are probably equivalent to the permanent kidney of the amniotic Vertebrates, and for this reason the numerous ducts of the first group or single duct of the second were spoken of as ureters. The anterior tubuli of the primitive excretory organ retain their early relation to the Wolffian duct, and form the Wolffian body.

Footnote 361: The reverse of the above rule is the case with Raja, in the male of which a closer approximation to the single-duct type is found than in the female.

The originally separate terminal extremities of the Wolffian ducts always coalesce, and form a urinal cloaca, opening by a single aperture situated at the extremity of a median papilla behind the anus. Some of the abdominal openings of the segmental tubes in Scyllium, or in other cases all the openings, become obliterated.

In the male the anterior segmental tubes undergo remarkable modifications. There appear to grow from the first three or four or more of them (though the point is still somewhat obscure) branches, which pass to the base of the testis and there unite into a longitudinal canal, form a network, and receive the secretion of the testicular ampullæ (woodcut 9, _nt_). These ducts, the vasa efferentia, carry the semen to the Wolffian body, but before opening into the tubuli of this they unite into the _longitudinal canal of the Wolffian body (l.c)_, from which pass off ducts equal in number to the vasa efferentia, each of which normally ends in a Malpighian body. From the Malpighian body so connected start the convoluted tubuli of what may be called the generative segments of the Wolffian body along which the semen is conveyed to the Wolffian duct (_v.d_). The Wolffian duct itself becomes much contorted and acts as vas deferens.

In the woodcuts, figs. 8 and 9, are diagrammatically represented the chief constituents of the adult urinogenital organs in the two sexes. In the adult female, fig. 8, there are present the following parts:

(1) The oviduct or Müllerian duct (_m.d_) split off from the segmental duct of the kidneys. Each oviduct opens at its anterior extremity into the body-cavity, and behind the two oviducts have independent communications with the general cloaca.

(2) The Wolffian ducts (_w.d_), the other product of the segmental ducts of the kidneys. They end in front by becoming continuous with the tubulus of the anterior segment of the Wolffian body on each side, and unite behind to open by a common papilla into the cloaca. The Wolffian duct receives the secretion of the anterior part of the primitive kidney which forms the Wolffian body.

(3) The ureter (_d_) which carries off the secretion of the kidney proper. It is represented in my diagram in its most rare and differentiated condition as a single duct.

(4) The glandular tubuli (_s.t_), some of which retain their original openings into the body-cavity, and others are without them. They are divided into two groups, an anterior forming the Wolffian body, which pour their secretion into the Wolffian duct, and a posterior group forming the kidney proper, which are connected with the ureter.

In the male the following parts are present (woodcut 9):

(1) The Müllerian duct (_md_), consisting of a small rudiment attached to the liver representing the foremost end of the oviduct of the female.

(2) The Wolffian duct (_w.d_) which precisely corresponds to the Wolffian duct of the female, but, in addition to functioning as the duct of the Wolffian body, also acts as a vas deferens (_vd_). In the adult male its foremost part has a very tortuous course.

(3) The ureter (_d_), which has the same fundamental constitution as in the female.

(4) The segmental tubes (_st_). The posterior of these have the same arrangement in both sexes, but in the male modifications take place in connection with the anterior ones to fit them to act as transporters of the testicular products.

Connected with the anterior ones there are present (1) the vasa efferentia (VE), united on the one hand with (2) the central canal in the base of the testis (_nt_), and on the other with the longitudinal canal of the Wolffian body (_l.c_). From the latter are seen passing off the successive tubuli of the anterior segments of the Wolffian body in connection with which Malpighian bodies are typically present, though not represented in my diagram.

_Postscript._

It was my original intention to have given an account of the development of the generative organs. In the course, however, of my work a number of novel and unexpected points turned up, which have considerably protracted my investigations, and it has appeared to me better no longer to delay the appearance of this monograph, but to publish elsewhere my results on the generative organs. In chapter VI. p. 349 _et seq._ the early stages of the generative organs are described, but in contemplation of the completion of the account no allusion was made to their literature, and more especially to Professor Semper's important contributions. I may perhaps say that I have been able to confirm the most important result to which he and other anatomists have nearly simultaneously arrived with respect to Vertebrates, viz. _that the primitive ova give rise to both the male and female generative products_.

EXPLANATION OF PLATES 20 AND 21.

COMPLETE LIST OF REFERENCE LETTERS.

_amg._ Accessory Malpighian body. _cav._ Cardinal vein. _ge._ Germinal epithelium. _k._ True kidney. _l.c._ Longitudinal canal of the Wolffian body connected with vasa efferentia. _mg._ Malpighian body. _nt._ Network and central canal at the base of the testis. _o._ External aperture of urinal cloaca. _od._ Oviduct or Müllerian duct of the female. _od´._ Müllerian duct of the male. _ou._ Openings of ureters in Wolffian duct in the female (fig. 3). _pmg._ Primary Malpighian body. _px._ Growth from vesicle at the end of a segmental tube to join the collecting tube of the preceding segment. _rst._ Rudimentary segmental tube. _ru._ Ureter commencing to be formed. _sb._ Seminal bladder. _sd._ Segmental duct. _st._ Segmental tube. _sto._ Opening of segmental tube into body-cavity. _sur._ Suprarenal body. _t._ Testis. _u._ Ureters. _ve._ Vas efferens. _wb._ Wolffian body. _wd._ Wolffian duct.

PLATE 20.

Fig. 1. Diagrammatic representation of excretory organs on one side of a male Scyllium canicula, natural size.

Fig. 2. Diagrammatic representation of the kidney proper on one side of a female Scyllium canicula, natural size, shewing the ducts of the kidney and the dilated portion of the Wolffian duct.

Fig. 3. Opening of the ureters into the Wolffian duct of a female Scyllium canicula. The figure represents the Wolffian ducts (_wd_) with ventral portion removed so as to expose their inner surface, and shews the junction of the two W. ducts to form the common urinal cloaca, the single external opening of this (_o_), and openings of ureters into one Wolffian duct (_ou_).

Fig. 4. Anterior extremity of Wolffian body of a young male Scyllium canicula shewing the vasa efferentia and their connection with the kidneys and the testis. The vasa efferentia and longitudinal canal are coloured to render them distinct. They are intended to be continuous with the uncoloured coils of the Wolffian body, though this connection has not been very successfully rendered by the artist.

Fig. 5. Part of the Wolffian body of a nearly ripe male embryo of Scyllium canicula as a transparent object. Zeiss a a, ocul. 3. The figure shews two segmental tubes opening into the body-cavity and connected with a primary Malpighian body, and also, by a fibrous connection, with a secondary Malpighian body of the preceding segment. It also shews one segmental tube (_rst_) imperfectly connected with the accessory Malpighian body of the preceding segment of the kidney. The coils of the kidney are represented somewhat diagrammatically.

Fig. 6. Vasa efferentia of a male embryo of Scyllium canicula eight centimetres in length. Zeiss a a, ocul. 2.

There are seen to be at the least six and possibly seven distinct vasa going to as many segments of the Wolffian body and connected with a longitudinal canal in the base of the testis. They were probably also connected with a longitudinal canal in the Wolffian body, but this could not be clearly made out.

Fig. 7. The anterior four vasa efferentia of a nearly ripe embryo. Connected with the foremost one is seen a body which looks like the remnant of a segmental tube and its opening (_rst?_).

Fig. 8. Testis and anterior part of Wolffian body of an embryo of Squatina vulgaris.

The figure is intended to illustrate the arrangement of the vasa efferentia. There are five of these connected with a longitudinal canal in the base of the testis, and with another longitudinal canal in the Wolffian body. From the second longitudinal canal there pass off four ducts to as many Malpighian bodies. Through the Malpighian bodies these ducts are continuous with the several coils of the Wolffian body, and so eventually with the Wolffian duct. Close to the hindermost vas efferens is seen a body which resembles a rudimentary segmental tube (_rst?_).

PLATE 21.

Figs. 1A, 1B, 1C, 1D. Four sections from a female Scyllium canicula of a stage between M and N through the part where the segmental duct becomes split into Wolffian duct and oviduct. Zeiss B, ocul. 2. 1A is the foremost section.

The sections shew that the oviduct arises as a thickening on the under surface of the segmental duct into which at the utmost a very narrow prolongation of the lumen of the segmental duct is carried. The small size of the lumen of the Wolffian duct in the foremost section is due to the section passing through nearly its anterior blind extremity.

Fig. 2. Section close to the junction of the Wolffian duct and oviduct in a female embryo of Scyllium canicula belonging to stage N. Zeiss B, ocul. 2.

The section represented shews that in some instances the formation of the oviduct and Wolffian duct is accompanied by a division of the lumen of the segmental duct into two not very unequal parts.

Figs. 3A, 3B, 3C. Three sections illustrating the formation of a ureter in a female embryo belonging to stage N. Zeiss B, ocul. 2.

3A is the foremost section.

The figures shew that the lumen of the developing ureter is enclosed in front by an independent wall (fig. 3A), but that further back the lumen is partly shut in by the subjacent Wolffian duct, while behind no lumen is present, but the ureter ends as a solid knob of cells without an opening into the Wolffian duct.

Fig. 4. Section through the ureters of the same embryo as fig. 3, but nearer the cloaca. Zeiss B, ocul. 2.

The figure shews the appearance of a transverse section through the wall of cells above the Wolffian duct formed by the overlapping ureters, the lumens of which appear as perforations in it. It should be compared with fig. 9A, which represents a longitudinal section through a similar wall of cells.

Fig. 5. Section through the ureters, the Wolffian duct and the oviduct of a female embryo of Scy. canicula belonging to stage P. Zeiss B, ocul. 2.

Fig. 6. Section of part of the Wolffian body of a male embryo of Scyllium canicula belonging to stage O. Zeiss B, ocul. 2.

The section illustrates (1) the formation of a Malpighian body (_mg_) from the dilatation at the end of a segmental tube, (2) the appearance of the rudiment of the Müllerian duct in the male (_od´_).

Figs. 7_a_, 7_b_. Two longitudinal and vertical sections through part of the kidney of an embryo between stages L and M. Zeiss B, ocul. 2.

7_a_ illustrates the parts of a single segment of the Wolffian body at this stage, vide p. 491. The segmental tube and opening are not in the plane of the section, but the dilated vesicle is shewn into which the segmental tube opens.

7_b_ is taken from the region of the kidney proper. To the right is seen the opening of a segmental tube into the body-cavity, and in the segment to the left the commencing formation of a ureter, vide p. 502.

Fig. 8. Longitudinal and vertical section through the posterior part of the kidney proper of an embryo of Scyllium canicula at a stage between N and O. Zeiss A, ocul. 2.

The section shews the nearly completed ureters, developing Malpighian bodies, &c.

Fig. 9. Longitudinal and vertical section through the anterior part of the kidney proper of the same embryo as fig. 8. Zeiss A, ocul. 2.

The figure illustrates the mode of growth of the developing ureters.

9A. More highly magnified portion of the same section as fig. 9.

Compare with transverse section fig. 4.

Fig. 10. Longitudinal and vertical section through part of the Wolffian body of an embryo of Scyllium canicula at a stage between O and P.

The section contains two examples of the budding out of the vesicle of a segmental tube to form a Malpighian body in its own segment and to unite with the tubulus of the preceding segment close to its opening into the Wolffian duct.

XI. ON THE PHENOMENA ACCOMPANYING THE MATURATION AND IMPREGNATION OF THE OVUM[362].

Footnote 362: From the _Quarterly Journal of Microscopical Science_, April, 1878.

The brilliant discoveries of Strasburger and Auerbach have caused the attention of a large number of biologists to be turned to the phenomena accompanying the division of nuclei and the maturation and impregnation of the ovum. The results of the recent investigations on the first of these points formed the subject of an article by Mr Priestley in the sixteenth volume of this Journal, and the object of the present article is to give some account of what has so far been made out with reference to the second of them. The matters to be treated of naturally fall under two heads: (1) the changes attending the ripening of the ovum, _which are independent of impregnation_; (2) the changes which are directly due to impregnation.

Every ovum as it approaches maturity is found to be composed (Fig. 1) of (1) a protoplasmic body or vitellus usually containing yolk-spherules in suspension; (2) of a germinal vesicle or nucleus, containing (3) one or more germinal spots or nucleoli. It is with the germinal vesicle and its contents that we are especially concerned. This body at its full development has a more or less spherical shape, and is enveloped by a distinct membrane. Its contents are for the most part fluid, but may be more or less granular. Their most characteristic component is, however, a protoplasmic network which stretches from the germinal spot to the investing membrane, but is especially concentrated round the former (Fig. 1). The germinal spot forms a nearly homogeneous body, with frequently one or more vacuoles. It occupies an often excentric position within the germinal vesicle, and is usually rendered very conspicuous by its high refrangibility. In many instances it has been shewn to be capable of amoeboid movements (Auerbach, and Os. Hertwig), and is moreover more solid and more strongly tinged by colouring reagents than the remaining constituents of the germinal vesicle. These peculiarities have caused the matter of which it is composed to be distinguished by Auerbach and Hertwig as nuclear substance.

In many instances there is only one germinal spot, or one main spot, and two or three accessory smaller spots. In other cases, _e.g._ Osseous Fish, there are a large number of nearly equal germinal spots. The eggs which have been most investigated with reference to the changes of germinal vesicle are those with a single germinal spot, and it is with these that I shall have more especially to deal in the sequel.

The germinal vesicle occupies in the first instance a central position in the ovum, but at maturity is almost always found in close proximity to the surface. Its change of position in a large number of instances is accomplished during the growth of the ovum in the ovary, but in other cases does not take place till the ovum has been laid.

The questions which many investigators have recently set themselves to answer are the two following:--(1) What becomes of the germinal vesicle when the ovum is ready to be impregnated? (2) Is any part of it present in the ovum at the commencement of segmentation? According to their answers to these questions the older embryologists roughly fall into two groups: (1) By one set the germinal vesicle is stated to completely disappear and not to be genetically connected with the subsequent nuclei of the embryo. (2) According to the other set it remains in the ovum and by successive divisions forms the parent nucleus of all the nuclei in the body of the embryo. Though the second of these views has been supported by several very distinguished names the first view was without doubt the one most generally entertained, and Haeckel (though from his own observations he was originally a supporter of the second view) has even enunciated the theory that there exists an anuclear stage, after the disappearance of the germinal vesicle, which he regards as an embryonic repetition of the monad condition of the Protozoa.

While the supporters of the first view agree as to the disappearance of the germinal vesicle they differ considerably as to the manner of this occurrence. Some are of opinion that the vesicle simply vanishes, its contents being absorbed in the ovum; others that it is ejected from the ovum and appears as the _polar cell_ or _body_, or _Richtungskörper_ of the Germans--a small body which is often found situated in the space between the ovum and its membrane, and derives its name from retaining a constant position in relation to the ovum, and thus serving as a guide in determining the similar parts of the embryo through the different stages. The researches of Oellacher (15)[363] in this direction deserve special mention, as having in a sense formed the foundation of the modern views upon this subject. By a series of careful observations upon the egg of the trout and subsequently of the bird, he demonstrated that the germinal vesicle of the ovum, while still in the ovary, underwent partial degeneration and eventually became ejected. His observations were made to a great extent by means of sections, and the general accuracy of his results is fairly certain, but the nature of the eggs he worked on, as well as other causes, prevented his obtaining so deep an insight into the phenomena accompanying the ejection of the germinal vesicle as has since been possible. Lovén, Flemming (6), and others have been led by their investigations to adopt views similar in the main to Oellacher's. As a rule, however, it is held by believers in the disappearance of the germinal vesicle that it becomes simply absorbed, and many very accurate accounts, so far as they go, have been given of the gradual atrophy of the germinal vesicle. The description of Kleinenberg (14) for Hydra, and Götte for Bombinator, may perhaps be selected as especially complete in this respect; in both instances the germinal vesicle commences to atrophy at a relatively early period.

Footnote 363: The numbers appended to authors' names refer to the list of publications at the end of the paper.

Coming to the more modern period the researches of five workers, viz. Bütschli, E. van Beneden, Fol, Hertwig, and Strasburger have especially thrown light upon this difficult subject. It is now hardly open to doubt that while part of the germinal vesicle is concerned in the formation of the polar cell or cells, when such are present, and is therefore ejected from the ovum, part also remains in the ovum and forms a nuclear body which will be spoken of as the _female pronucleus_, the fate of which is recorded in the second part of this paper. The researches of Bütschli and van Beneden have been especially instrumental in demonstrating the relation between the polar bodies and the germinal vesicle, and those of Hertwig and Fol, in shewing that part of the germinal vesicle remained in the ovum. It must not, however, be supposed that the results of these authors are fully substantiated, or that all the questions connected with these phenomena are settled. The statements we have are in many points opposed and contradictory, and there is much that is still very obscure.

In the sequel an account is first given of the researches of the above-named authors, followed by a statement of those results which appear to me the most probable.

The researches of van Beneden (3 and 4) were made on the ovum of the rabbit and of Asterias, and from his observations on both these widely separated forms he has been led to conclude that the germinal vesicle is either ejected or absorbed, but that it has in no case a genetic connection with the first segmentation sphere. He gives the following description of the changes in the rabbit's ovum. The germinal vesicle is enclosed by a membrane, and contains one main germinal spot, and a few accessory ones, together with a granular material which he calls _nucleoplasma_, which affects, as is usual in nuclei, a reticular arrangement. The remaining space in the vesicle is filled by a clear fluid. As the ovum approaches maturity the germinal vesicle assumes an excentric position, and fuses with the peripheral layer of the egg to constitute the _cicatricular lens_. The germinal spot next travels to the surface of the cicatricular lens and forms the _nuclear disc_: at the same time the membrane of the germinal vesicle vanishes though it probably unites with the nuclear disc. The nucleoplasma then collects into a definite mass and forms the nucleoplasmic body. Finally the nuclear disc assumes an ellipsoidal form and becomes the nuclear body. Nothing is now left of the original germinal vesicle but the nuclear body and the nucleoplasmic body both still situated within the ovum. In the next stage no trace of the germinal vesicle can be detected in the ovum, but outside it, close to the point where the modified remnants of the vesicle were previously situated, there is present a polar body which is composed of two parts, one of which stains deeply and resembles the nuclear body, and the other does not stain but is similar to the nucleoplasmic body. Van Beneden concludes that the polar bodies are the two ejected products of the germinal vesicle. In the case of Asterias, van Beneden has not observed the mode of formation of the polar bodies, and mainly gives an account of the atrophy of the germinal vesicle, but adds very little to what was already known to us from Kleinenberg's (14) earlier observations. He describes with precision the breaking up of the germinal spot into fragments and its eventual disappearance.

Though there are reasons for doubting the accuracy of all the above details on the ovum of the rabbit, nevertheless, the observations of van Beneden taken as a whole afford strong grounds for concluding that the formation of the polar cells is connected with the disappearance, partial or otherwise, of the germinal vesicle. A very similar account of the apparent disappearance of the germinal vesicle is given by Greeff (19) who states that the apparent disappearance of the germinal spot precedes that of the vesicle.

The observations of Bütschli are of still greater importance in this direction. He has studied with a view to elucidating the fate of the germinal vesicle, the eggs of Nephelis, Lymnæus, Cucullanus, and other Nematodes; and Rotifers. In all of these, with the exception of Rotifers, he finds polar bodies, and in this respect his observations are of value as tending to shew the widespread existence of these structures. Negative results with reference to the presence of the polar bodies have, it may be remarked, only a very secondary value. Bütschli has made the very important discovery that in perfectly ripe eggs of Nephelis, Lymnæus and Cucullanus and allied genera a _spindle_, similar to that of ordinary nuclei in the act of division, appears close to the surface of the egg. This spindle he regards as the metamorphosed germinal vesicle, and has demonstrated that it takes part in the formation of the polar cells. He states that the whole spindle is ejected from the egg, and that after swelling up and forming a somewhat spherical mass it divides into three parts.

In the Nematodes generally, Bütschli has been unable to find the spindle modification of the germinal vesicle, but he states that the germinal vesicle undergoes degeneration, its outline becoming indistinct and the germinal spot vanishing. The position of the germinal vesicle continues to be marked by a clear space which gradually approaches the surface of the egg. When it is in contact with the surface a small spherical body, the remnant of the germinal vesicle, comes into view, and eventually becomes ejected. The clear space subsequently disappears. This description of Bütschli resembles in some respects that given by van Beneden of the changes in the rabbit's ovum, and not impossibly refers to a nearly identical series of phenomena. The discovery by Bütschli of the spindle and its relation to the polar body has been of very great value.

The publications of van Beneden, and more especially those of Bütschli, taken by themselves lead to the conclusion that the whole germinal vesicle is either ejected or absorbed. Nearly simultaneously with their publications there appeared, however, a paper by Oscar Hertwig (11) on the eggs of one of the common sea urchins (_Toxopneustes lividus_), in which he attempted to shew that part of the germinal vesicle, at any rate, was concerned in the formation of the first segmentation nucleus. He believed (though he has himself now recognised that he was in error on the point) that no polar cell was formed in Toxopneustes, and that the whole germinal vesicle was absorbed, with the exception of the germinal spot which remained in the egg as the female pronucleus.

The following is the summary which he gives of his results, pp. 357-8.

"At the time when the egg is mature the germinal vesicle undergoes a retrogressive metamorphosis and becomes carried towards the surface of the egg by the contraction of the protoplasm. Its membrane becomes dissolved and its contents disintegrated and finally absorbed by the yolk. The germinal spot appears, however, to remain unaltered and to continue in the yolk and to become the permanent nucleus of the ripe ovum capable of impregnation."

After the publication of Bütschli's monograph, O. Hertwig (12) continued his researches on the ova of Leeches (_Hæmopis_ and _Nephelis_), and not only added very largely to our knowledge of the history of the germinal vesicle, but was able to make a very important rectification in Bütschli's conclusions. The following is a summary of his results:--The germinal vesicle, as in other cases, undergoes a form of degeneration, though retaining its central position; and the germinal spot breaks up into fragments. The stages in which this occurs are followed by one when, on a superficial examination, the ovum appears to be absolutely without a nucleus; but there can be demonstrated by means of reagents in the position previously occupied by the germinal vesicle a spindle nucleus with the usual suns at its poles, which Hertwig believes to be a product of the fragments of the germinal spot. This spindle travels towards the periphery of the ovum and then forms the spindle observed by Bütschli. At the point where one of the apices of the spindle lies close to the surface a small protuberance arises which is destined to form the first polar cell. As the protuberance becomes more prominent one half of the spindle passes into it. The spindle then divides in the normal manner for nuclei, one half remaining in the protuberance, the other in the ovum, and finally the protuberance becomes a rounded body united to the egg by a narrow stalk. It is clear that if, as there is every reason to think, the above description is correct, the polar cell is formed by a simple process of cell-division and not, as Bütschli believed, by the forcible ejection of the spindle.

The portion of the spindle in the polar cell becomes a mass of granules, and that in the ovum becomes converted without the occurrence of the usual nuclear stage into a fresh spindle. A second polar cell is formed in the same manner as the first one, and the first one subsequently divides into two. The portion of the spindle which remains in the egg after the formation of the second polar cell reconstitutes itself into a nucleus--the female pronucleus--and travelling towards the centre of the egg undergoes a fate which will be spoken of in the second part of this paper.

The most obscure part of Hertwig's work is that which concerns the formation of the spindle on the atrophy of the germinal vesicle, and his latest paper, though it gives further details on this head, does not appear to me to clear up the mystery. Though Hertwig demonstrates clearly enough that this spindle is a product of the metamorphoses of the germinal vesicle, he does not appear to prove the thesis which he maintains, that it is the metamorphosed germinal spot.

Fol, to whom we are indebted in his paper on the development of Geryonia (7) for the best of the earlier descriptions of the phenomena which attend the maturation of the egg, and later for valuable contributions somewhat similar to those of Bütschli with reference to the development of the Pteropod egg (8), has recently given us a very interesting account of what takes place in the ripe egg of _Asterias glacialis_ (9). In reference to the formation of the polar cells, his results accord closely with those of Hertwig, but he differs considerably from this author with reference to the preceding changes in the germinal vesicle. He believes that the germinal spot atrophies more or less completely, but that in any case its constituents remain behind in the egg, though he will not definitely assert that it takes no share in the formation of the spindle at the expense of which both the polar cells and the female pronucleus are formed. The spindle with its terminal suns arises, according to him, from the contents of the germinal vesicle, loses its spindle character, travels to the surface, and reacquiring a spindle character is concerned in the formation of the polar cells in the way described by Hertwig.

Giard (10) gives a somewhat different account of the behaviour of the germinal vesicle in _Psammechinus miliaris_. At maturity the contents of the germinal vesicle and spot mix together and form an amoeboid mass, which, assuming a spindle form, divides into two parts, one of which travels towards the centre of the egg and forms the female pronucleus, the other remains at the surface and gives origin to two polar cells, both of which are formed after the egg is laid. What Giard regards as the female pronucleus is perhaps the lower of the two bodies which take the place of the original germinal vesicle as described by Fol. Vide the account of Fol's observations on p. 531.

Strasburger, from observations on _Phallusia_, accepts in the main Hertwig's conclusion with reference to the formation of the polar bodies, but does not share Hertwig's view that either the polar bodies or female pronucleus are formed at the expense of the germinal spot alone. He has further shewn that the so-called canal-cell of conifers is formed in the same manner as the polar cells, and states his belief that an equivalent of the polar cells is widely distributed in the vegetable subkingdom.

This sketch of the results of recent researches will, it is hoped, suffice to bring into prominence the more important steps by which the problems of this department of embryology have been solved. The present aspects of the question may perhaps be most conveniently displayed by following the history of a single ovum. For this purpose the eggs of _Asterias glacialis_, which have recently formed the subject of a series of beautiful researches by Fol (9), may conveniently be selected.

The ripe ovum (Fig. 2), when detached from the ovary, is formed of a granular vitellus without a vitelline membrane, but enveloped in a mucilaginous coat. It contains an excentrically situated germinal vesicle and germinal spot. In the former is present the usual protoplasmic reticulum. As soon as the ovum reaches the sea water the germinal vesicle commences to undergo a peculiar metamorphosis. It exhibits frequent changes of form, its membrane becomes gradually absorbed and its outline indented and indistinct, and finally its contents become to a certain extent confounded with the vitellus (Fig. 3).

The germinal spot at the same time loses its clearness of outline and gradually disappears from view.

At a slightly later stage in the place of the original germinal vesicle there may be observed in the fresh ovum two clear spaces (fig. 4), one ovoid and nearer the surface, and the second more irregular in form and situated rather deeper in the vitellus. By treatment with reagents the first clear space is found to be formed of a spindle with two terminal suns on the lower side of which is a somewhat irregular body (Fig. 5). The second clear space by the same treatment is shewn to contain a round body. Fol concludes that the spindle is formed out of part of the germinal vesicle and not of the germinal spot, while he sees in the round body present in the lower of the two clear spaces the metamorphosed germinal spot. He will not, however, assert that no fragment of the germinal spot enters into the formation of the spindle. It may be observed that Fol is here obliged to fill up (so far at least as his present preliminary account enables me to determine) a lacuna in his observations in a hypothetical manner, and O. Hertwig's (13) most recent observations on the ovum of the same or an allied species of Asterias tend to throw some doubt upon Fol's interpretations.

The following is Hertwig's account of the changes in the germinal vesicle. A quarter of an hour after the egg is laid the protoplasm on the side of the germinal vesicle towards the surface of the egg develops a prominence which presses inwards the wall of the vesicle. At the same time the germinal spot develops a large vacuole, in the interior of which is a body consisting of nuclear substance, and formed of a firmer and more refractive material than the remainder of the germinal spot. In the above-mentioned prominence towards the germinal vesicle, first one sun is formed by radial striæ of protoplasm, and then a second makes its appearance, while in the living ovum the germinal spot appears to have vanished, the outline of the germinal vesicle to have become indistinct, and its contents to have mingled with the surrounding protoplasm. Treatment with reagents demonstrates that in the process of disappearance of the germinal spot the nuclear mass in the vacuole forms a rod-like body, the free end of which is situated between the two suns which occupy the prominence of the germinal vesicle. At a slightly later period granules may be seen at the end of the rod and finally the rod itself vanishes. After these changes there may be demonstrated by the aid of reagents a spindle between the two suns, which Hertwig believes to grow in size as the last remnants of the germinal spot gradually vanish, and he maintains, as before mentioned, that the spindle is formed at the expense of the germinal spot. Without following Hertwig so far as this[364] it may be permitted to suggest that his observations tend to shew that the body noticed by Fol in the median line, on the inner side of his spindle, is in reality a remnant of the germinal spot and not, as Fol supposes, part of the germinal vesicle. Considering how conflicting is the evidence before us it seems necessary to leave open for the present the question as to what parts of the germinal vesicle are concerned in forming the first spindle.

Footnote 364: Hertwig's full account of his observations, with figures, in the 4th vol. of the _Morphologische Jahrbuch_, has appeared since the above was written. The figures given strongly support Hertwig's views.

The spindle, however it be formed, has up to this time been situated with its axis parallel to the surface of the egg, but not long after the stage last described a spindle is found with one end projecting into a protoplasmic prominence which makes its appearance on the surface of the egg (Fig. 6). Hertwig believes that the spindle simply travels towards the surface, and while doing so changes the direction of its axis. Fol finds, however, that this is not the case, but that between the two conditions of the spindle an intermediate one is found in which a spindle can no longer be seen in the egg, but its place is taken by a compact rounded body. He has not been able to arrive at a conclusion as to what meaning is to be attached to this occurrence. In any case the spindle which projects into the prominence on the surface of the egg divides it into two parts, one in the prominence and one in the egg (Fig. 7). The prominence itself with the enclosed portion of the spindle becomes partially constricted off from the egg as the first polar body (Fig. 8). The part of the spindle which remains in the egg becomes directly converted into a second spindle by the elongation of its fibres without passing through a typical nuclear condition. A second polar cell next becomes formed in the same manner as the first (Fig. 9), and the portion of the spindle remaining in the egg becomes converted into two or three clear vesicles (Fig. 10) which soon unite to form a single nucleus, the female pronucleus (Fig. 11). The two polar cells appear to be situated between two membranes, the outer of which is very delicate and only distinct where it covers the polar cells, while the inner one is thicker and becomes, after impregnation, more distinct and then forms what Fol speaks of as the vitelline membrane. It is clear, as Hertwig has pointed out, that the polar bodies originate by a regular cell division and have the value of cells.

_General conclusions._

Considering how few ova have been adequately investigated with reference to the behaviour of the germinal vesicle any general conclusions which may at present be formed are to be regarded as provisional, and I trust that this will be borne in mind by the reader in perusing the following paragraphs.

There is abundant evidence that at the time of maturation of the egg the germinal vesicle undergoes peculiar changes, which are, in part at least, of a retrogressive character. These changes may begin considerably before the egg has reached the period of maturity, or may not take place till after it has been laid. They consist in appearance of irregularity and obscurity in the outline of the germinal vesicle, the absorption of its membrane, the partial absorption of its contents in the yolk, and the breaking up and disappearance of the germinal spot. The exact fate of the single germinal spot, or the numerous spots where they are present, is still obscure; and the observations of Oellacher on the trout, and to a certain extent my own on the skate, tend to shew that the membrane of the germinal vesicle may in some cases be ejected from the egg, but this conclusion cannot be accepted without further confirmation.

The retrogressive metamorphosis of the germinal vesicle is followed in a large number of instances by the conversion of what remains into a striated spindle similar in character to a nucleus previous to division. This spindle travels to the surface and undergoes division to form the polar cell or cells in the manner above described. The part which remains in the egg forms eventually the female pronucleus.

The germinal vesicle has up to the present time only been observed to undergo the above series of changes in a certain number of instances, which, however, include examples from several divisions of the Coelenterata, the Echinodermata, and the Mollusca, and also some of the Vermes (Nematodes, Hirudinea, Sagitta). It is very possible, not to say probable, that it is universal in the animal kingdom, but the present state of our knowledge does not justify us in saying so. It may be that in the case of the rabbit, and many Nematodes as described by van Beneden and by Bütschli, we have instances of a different mode of formation of the polar cells.

The case of Amphibians, as described by Bambeke (2) and Hertwig (12) cannot so far be brought into conformity with our type, though observations are so difficult to make with such opaque eggs that not much reliance can be placed upon the existing statements. In both of these types of possible exceptions it is fairly clear that, whatever may be the case with reference to the formation of the polar cells, part of the germinal vesicle remains behind as the female pronucleus.

There are a large number of types, including the whole of the Rotifera[365] and Arthropoda, with a few doubtful exceptions, in which the polar cells cannot as yet be said to have been satisfactorily observed.

Footnote 365: Flemming (6) finds that, in the summer and probably parthenogenetic eggs of _Lacinularia socialis_, the germinal vesicle approaches the surface and becomes invisible, and that subsequently a slight indentation in the outline of the egg marks the point of its disappearance. In the hollow of the indentation Flemming believes a polar cell to be situated, though he has not definitely seen one.

Whatever may be the eventual result of more extended investigation, it is clear that the formation of polar cells according to our type is a very constant occurrence. Its importance is also very greatly increased by the discovery by Strasburger of the existence of an analogous process amongst plants. Two questions about it obviously present themselves for solution: (1) What are the conditions of its occurrence with reference to impregnation? (2) What meaning has it in the development of the ovum or the embryo?

The answer to the first of these questions is not difficult to find. The formation of the polar bodies is independent of impregnation, and is the final act of the normal growth of the ovum. In a few types the polar cells are formed while the ovum is still in the ovary, as, for instance, in some species of Echini, Hydra, &c., but, according to our present knowledge, far more usually after the ovum has been laid. In some of the instances the budding off of the polar cells precedes, and in others follows impregnation; but there is no evidence to shew that in the later cases the process is influenced by the contact with the male element. In Asterias, as has been shewn by O. Hertwig, the formation of the polar cells may indifferently either precede or follow impregnation--a fact which affords a clear demonstration of the independence of the two occurrences.

To the second of the two questions it does not unfortunately seem possible at present to give an answer which can be regarded as satisfactory.

The retrogressive changes in the membrane of the germinal vesicle which usher in the formation of the polar bodies may very probably be viewed as a prelude to a renewed activity of the contents of the vesicle; and are perhaps rendered the more necessary from the thickness of the membrane which results from a protracted period of passive growth. This suggestion does not, however, help us to explain the formation of polar cells by a process identical with cell division. The ejection of part of the germinal vesicle in the formation of the polar cells may probably be paralleled by the ejection of part or the whole of the original nucleus which, if we may trust the beautiful researches of Bütschli, takes place during conjugation in Infusoria as a preliminary to the formation of a fresh nucleus. This comparison is due to Bütschli, and according to it the formation of the polar bodies would have to be regarded as assisting, in some as yet unknown way, the process of regeneration of the germinal vesicle. Views analogous to this are held by Strasburger and Hertwig, who regard the formation of the polar bodies in the light of a process of excretion or removal of useless material. Such hypotheses do not unfortunately carry us very far.

I would suggest that in the formation of the polar cells part of the constituents of the germinal vesicle which are requisite for its functions as a complete and independent nucleus are removed to make room for the supply of the necessary parts to it again by the spermatic nucleus (vide p. 541). More light on this, as on other points, may probably be thrown by further investigations on parthenogenesis and the presence or absence of a polar cell in eggs which develop parthenogenetically. Curiously enough the two groups in which parthenogenesis most frequently occurs in the ordinary course of development (_Arthropoda_ and _Rotifera_) are also those in which polar cells, with the possible exception mentioned above, of the parthenogenetic eggs of Lacenularia, are stated to be absent. This curious coincidence, should it be confirmed, may perhaps be explained on the hypothesis, I have just suggested, viz. _that a more or less essential part of the nucleus is removed in the formation of the polar cells; so that in cases, e.g. Arthropoda and Rotifera, where polar cells are not formed, and an essential part of the nucleus not therefore removed, parthenogenesis can much more easily occur than when polar cells are formed_.

That the part removed in the formation of the polar cells is not absolutely essential, seems at first sight to follow from the fact of parthenogenesis being possible in instances where impregnation is the normal occurrence. The genuineness of all the observations on this head is too long a subject to enter into here[366], but after admitting, as we probably must, that there are genuine cases of parthenogenesis, it cannot be taken for granted without more extended observation that the occurrence of development in these rare instances may not be due to the polar cells not having been formed as usual, and that when the polar cells are formed the development without impregnation is less possible.

Footnote 366: The instances quoted by Siebold from Hensen and Oellacher are not quite satisfactory. In Hensen's case impregnation would have been possible if we can suppose the spermatozoa to be capable of passing into the body-cavity through the open end of the uninjured oviduct; and though Oellacher's instances are more valuable, yet sufficient care seems hardly to have been taken, especially when it is not certain for what length of time spermatozoa may be able to live in the oviduct. For Oellacher's precautions, vide _Zeit. für wiss. Zool._ Bd. XXII. p. 202.

The remarkable observations of Professor Greeff (19) on the parthenogenetic development of the eggs of _Asterias rubens_ tell, however, very strongly against this explanation. Greeff has found that under normal circumstances the eggs of this species of starfish will develop without impregnation in simple sea water. The development is quite regular and normal though much slower than in the case of impregnated eggs. It is not definitely stated that polar cells are formed, but there can be no doubt that this is implied. Professor Greeff's account is so precise and circumstantial that it is not easy to believe that any error can have crept in; but neither Hertwig nor Fol have been able to repeat his experiments, and we may be permitted to wait for further confirmation before absolutely accepting them.

It is possible that the removal of part of the protoplasm of the egg in the formation of the polar cells may be a secondary process due to an attractive influence of the nucleus on the cell protoplasm, such as is ordinarily observed in cell division.

_Impregnation of the Ovum._

A far greater amount of certainty appears to me to have been attained as to the effects of impregnation than as to the changes of the germinal vesicle which precede this, and there appears, moreover, to be a greater uniformity in the series of resulting phenomena. For convenience I propose to reverse the order hitherto adopted and to reserve the history of the literature and my discussion of disputed points till after my general account. Fol's paper on _Asterias glacialis_, is again my source of information. The part of the germinal vesicle which remains in the egg, after the formation of the second polar cell, becomes converted into a number of small vesicles (Fig. 10), which aggregate themselves into a single clear nucleus which gradually travels toward the centre of the egg and around which as a centre the protoplasm becomes radiately striated (Fig. 11). This nucleus is known as the _female pronucleus_[367]. In _Asterias glacialis_ the most favourable period for fecundation is about an hour after the formation of the female pronucleus. If at this time the spermatozoa are allowed to come in contact with the egg, their heads soon become enveloped in the investing mucilaginous coat. A prominence, pointing towards the nearest spermatozoon, now arises from the superficial layer of protoplasm of the egg and grows till it comes in contact with the spermatozoon (Figs. 12 and 13), Under normal circumstances the spermatozoon, which meets the prominence, is the only one concerned in the fertilisation, and it makes its way into the egg by passing through the prominence. The tail of the spermatozoa, no longer motile, remains visible for some time after the head has bored its way in, but its place is soon taken by a pale conical body which is, however, probably in part a product of the metamorphosis of the tail itself (Fig. 14). This body vanishes in its turn.

Footnote 367: According to Hertwig's most recent statement a nucleolus is present in this nucleus.

At the moment of contact between the spermatozoon and the egg the outermost layer of the protoplasm of the latter raises itself as distinct membrane, which separates from the egg and prevents the entrance of any more spermatozoa. At the point where the spermatozoon entered a crater-like opening is left in the membrane (Fig. 14).

The head of the spermatozoon when in the egg forms a nucleus for which the name _male pronucleus_ may be conveniently adopted. It grows in size by absorbing, it is said, material from the ovum, though this may be doubted, and around it is formed a clear space free from yolk-spherules. Shortly after its formation the protoplasm in its neighbourhood assumes a radiate arrangement (Fig. 15). At whatever point of the egg the spermatozoon may have entered, it gradually travels towards the female pronucleus. This latter, around which the protoplasm no longer has a radial arrangement, remains motionless till it comes in contact with the rays of the male pronucleus, after which its condition of repose is exchanged for one of activity, and it rapidly approaches the male pronucleus, and eventually fuses with it (Fig. 16).

The product of this fusion forms the first segmentation nucleus (Fig. 17), which soon, however, divides into the two nuclei of the two first segmentation spheres. While the two pronuclei are approaching one another the protoplasm of the egg exhibits amoeboid movements.

Of the earlier observations on this subject there need perhaps only be cited one of E. van Beneden, on the rabbit's ovum, shewing the presence of two nuclei before the commencement of segmentation. Bütschli was the earliest to state from observations on _Rhabditis dolichura_ that the first segmentation nucleus arose from the fusion of two nuclei, and this was subsequently shewn with greater detail for _Ascaris nigrovenosa_, by Auerbach (1). Neither of these authors gave at first the correct interpretation of their results. At a later period Bütschli (5) arrived at the conclusion that in a large number of instances (_Lymnæus_, _Nephelis_, _Cucullanus_, &c.), the nucleus in question was formed by the fusion of two or more nuclei, and Strasburger at first made a similar statement for _Phallusia_, though he has since withdrawn it. Though Bütschli's statements depend, as it seems, upon a false interpretation of appearances, he nevertheless arrived at a correct view with reference to what occurs in impregnation. Van Beneden (3) described in the rabbit the formation of the original segmentation nucleus from two nuclei, one peripheral and the other central, and he gave it as his hypothetical view that the peripheral nucleus was derived from the spermatic element. It was reserved for Oscar Hertwig (11) to describe in _Echinus lividus_ the entrance of a spermatozoon into the egg and the formation from it of the male pronucleus.

Though there is a general agreement between the most recent observers, Hertwig, Fol, Selenka, Strasburger, &c., as to the main facts connected with the entrance of one spermatozoon into the egg, the formation of the male pronucleus, and its fusion with the female pronucleus, there still exist differences of detail in the different descriptions which partly, no doubt, depend upon the difficulties of observation, but partly also upon the observations not having all been made upon the same species. Hertwig does not enter into details with reference to the actual entrance of the spermatozoon into the egg, but in his latest paper points out that considerable differences may be observed in occurrences which succeed impregnation, according to the relative period at which this takes place. When, in Asterias, the impregnation is effected about an hour after the egg is laid and previously to the formation of the polar cells, the male pronucleus appears at first to exert but little influence on the protoplasm, but after the formation of the second polar cell, the radial striæ around it become very marked, and the pronucleus rapidly grows in size. When it finally unites with the female pronucleus it is equal in size to the latter. In the case when the impregnation is deferred for four hours the male pronucleus never becomes so large as the female pronucleus. With reference to the effect of the time at which impregnation takes place, Asterias would seem to serve as a type. Thus in _Hirudinea_, _Mollusca_, and _Nematodes_ impregnation normally takes place before the formation of the polar bodies is completed, and the male pronucleus is accordingly as large as the female. In _Echinus_, on the other hand, where the polar bodies are formed in the ovary, the male pronucleus is always small.

Selenka, who has investigated the formation of the male pronucleus in _Toxopneustes variegatus_, differs in certain points from Fol. He finds that usually, though not always, a single spermatozoon enters the egg, and that though the entrance may be effected at any part of the surface, it generally occurs at the point marked by a small prominence where the polar cell was formed. The spermatozoon first makes its way through the mucous envelope of the egg, within which it swims about, and then bores with its head into the polar prominence. The head of the spermatozoon on entering the egg becomes enveloped by the superficial protoplasm, and travels inward with its envelope, while the tail remains outside. As Fol has described, a delicate membrane becomes formed shortly after the entrance of the spermatozoon. The head continues to make its way by means of rapid oscillations, till it has traversed about one eighth of the diameter of the egg, and then suddenly becomes still. The tail in the meantime vanishes, while the neck swells up and forms the male pronucleus. The junction of the male and female pronucleus is described by Fol and Selenka in nearly the same manner.

Giard gives an account of impregnation which is not easily brought into harmony with that of the other investigators. His observations were made on _Psammechinus miliaris_. At one point is situated a polar body and usually at the pole opposite to it a corresponding prominence. The spermatozoa on gaining access to the egg attach themselves to it and give it a rotatory movement, but according to Giard none of them penetrate the vitelline membrane which, though formed at an earlier period, now retires from the surface of the egg.

Giard believes that the prominence opposite the polar cells serves for the entrance of the spermatic material, which probably passes in by a process of diffusion. Thus, though he regards the male pronucleus as a product of impregnation, he does not believe it to be the head of a spermatozoon.

Both Hertwig and Fol have made observations on the result of the entrance into the egg of several spermatozoa. Fol finds that when the impregnation has been too long delayed the vitelline membrane is formed with comparative slowness and several spermatozoa are thus enabled to penetrate. Each spermatozoon forms a separate pronucleus with a surrounding sun; and several male pronuclei usually fuse with the female pronucleus. Each male pronucleus appears to exercise a repulsive influence on other male pronuclei, but to be attracted by the female pronucleus. When there are several male pronuclei the segmentation is irregular and the resulting larva a monstrosity. These statements of Fol and Hertwig are at first sight in contradiction with the more recent results of Selenka. In _Toxopneustes variegatus_ Selenka finds that though impregnation is usually effected by a single spermatozoon yet that several may be concerned in the act. The development continues, however, to be normal if three or even four spermatozoa enter the egg almost simultaneously. Under such circumstances each spermatozoon forms a separate pronucleus and sun.

It may be noticed that, while the observations of Fol and Hertwig were admittedly made upon eggs in which the impregnation was delayed till they no longer displayed their pristine activity, Selenka's were made upon quite fresh eggs; and it seems not impossible that the pathological symptoms in the embryos reared by the two former authors may have been due to the imperfection of the egg and not to the entrance of more than one spermatozoon. This, of course, is merely a suggestion which requires to be tested by fresh observations. We have not as yet a sufficient body of observations to enable us to decide whether impregnation is usually effected by a single spermatozoon, though in spite of certain conflicting evidence the balance would seem to incline towards the side of a single spermatozoon[368].

Footnote 368: The recent researches of Calberla on the impregnation of the ovum of _Petromyzon Planeri_ support this conclusion.

The discovery of Hertwig as to the formation of the male pronucleus throws a flood of light upon impregnation.

The act of impregnation is seen essentially to consist in the fusion of a male and female nucleus; not only does this appear in the actual fusion of the two pronuclei, but it is brought into still greater prominence by the fact that the female pronucleus is a product of the nucleus of a primitive ovum, and the male pronucleus is the metamorphosed _head_ of the spermatozoon which is itself developed from the nucleus of a spermatic cell[369]. The spermatic cells originate from cells (in the case of Vertebrates at least) identical with the primitive ova, so that the fusion which takes place is the fusion of morphologically similar parts in the two sexes.

Footnote 369: This seems the most probable view with reference to the nature of the head of the spermatozoon, though the point is not perhaps yet definitely decided.

It must not, however, be forgotten, as Strasburger has pointed out, that part of the protoplasm of the generative cells of the two sexes also fuse, viz. the tail of the spermatozoon with the protoplasm of the egg. But there is no evidence that the former is of importance for the act of impregnation. The fact that impregnation mainly consists in the union of two nuclei gives an importance to the nucleus which would probably not have been accorded to it on other grounds.

Hertwig's discovery is in no way opposed to Mr Darwin's theory of pangenesis and other similar theories, but does not afford any definite proof of their accuracy, nor does it in the meantime supply any explanation of the origin of two sexes or of the reasons for an embryo becoming male or female.

_Summary._

In what may probably be regarded as a normal case the following series of events accompanies the maturation and impregnation of an egg:--

(1) Transportation of the germinal vesicle to the surface of the egg.

(2) Absorption of the membrane of the germinal vesicle and metamorphosis of the germinal spot.

(3) Assumption of a spindle character by the remains of germinal vesicle, these remains being probably largely formed from the germinal spot.

(4) Entrance of one end of the spindle into a protoplasmic prominence at the surface of the egg.

(5) Division of the spindle into two halves, one remaining in the egg, the other in the prominence. The prominence becomes at the same time nearly constricted off from the egg as a polar cell.

(6) Formation of a second polar cell in same manner as first, part of the spindle still remaining in the egg.

(7) Conversion of the part of the spindle remaining in the egg after the formation of the second polar cell into a nucleus--the female pronucleus.

(8) Transportation of the female pronucleus towards the centre of the egg.

(9) Entrance of one spermatozoon into the egg.

(10) Conversion of the head of the spermatozoon into a nucleus--the male pronucleus.

(11) Appearance of radial striæ round the male pronucleus which gradually travels towards the female pronucleus.

(12) Fusion of male and female pronuclei to form the first segmentation nucleus.

_List of important recent Publications on the Maturation and Impregnation of the Ovum._

1. Auerbach. _Organologische Studien_, Heft 2.

2. Bambeke. "Recherches s. Embryologie des Batraciens." _Bull. de l'Acad. royale de Belgique_, 2me sér., t. LXI. 1876.

3. E. Van Beneden. "La Maturation de l'OEuf des Mammifères." _Bull. de l'Acad. royale de Belgique_, 2me sér., t. XL, no. 12, 1875.

4. E. Van Beneden. "Contributions à l'Histoire de la Vésicule Germinative, &c." _Bull. de l'Acad. royale de Belgique_, 2me sér., t. XLI, no. 1, 1876.

5. Bütschli. _Eizelle, Zelltheilung, und Conjugation der Infusorien._

6. Flemming. "Studien in d. Entwicklungsgeschichte der Najaden." _Sitz. d. k. Akad. Wien_, B. LXXI. 1875.

7. Fol. "Die erste Entwicklung des Geryonideneies." _Jenaische Zeitschrift_, Vol. VII.

8. Fol. "Sur le Développement des Pteropodes." _Archives de Zoologie Expérimentale et Générale_, Vols. IV and V.

9. Fol. "Sur le Commencement de l'Hénogénie." _Archives des Sciences Physiques et Naturelles_. Genève, 1877.

10. Giard. _Note sur les premiers phénomènes du développement de l'Oursin._ 1877.

11. Hertwig, Oscar. "Beit. z. Kenntniss d. Bildung, &c., d. thier. Eies." _Morphologisches Jahrbuch_, Bd. I.

12. Hertwig, Oscar. Ibid. _Morphologisches Jahrbuch_, Bd. III, Heft. 1.

13. Hertwig, Oscar. "Weitere Beiträge, &c." _Morphologisches Jahrbuch_, Bd. III, Heft 3.

14. Kleinenberg. _Hydra_. Leipzig, 1872.

15. Oellacher, J. "Beiträge zur Geschichte des Keimbläschens im Wirbelthiereie." _Archiv f. micr. Anat._, Bd. VIII.

16. Selenka. _Befruchtung u. Theilung des Eies von Toxopneustes variegatus_ (Vorläufige Mittheilung). Erlangen, 1877.

17. Strasburger. _Ueber Zellbildung u. Zelltheilung._ Jena, 1876.

18. Strasburger. _Ueber Befruchtung u. Zelltheilung._ Jena, 1878.

19. R. Greeff. "Ueb. d. Bau u. d. Entwicklung d. Echinodermen." _Sitzun. der Gesellschaft z. Beförderung d. gesammten Naturwiss. z. Marburg_, No. 5. 1876.

_Postscript_.--Two important memoirs have appeared since this paper was in type. One of these by Hertwig, _Morphologisches Jahrbuch_, Bd. IV, contains a full account with illustrations of what was briefly narrated in his previous paper (13); the other by Calberla, "Der Befruchtungsvorgang beim Ei von _Petromyzon Planeri_," _Zeit. für wiss. Zool._, Bd. XXX, shews that the superficial layer of the egg is formed by a coating of protoplasm free from yolk-spheres, which at one part is continued inwards as a column, and contains the germinal vesicle. The surface of this column is in contact with a micropyle in the egg-membrane. Impregnation is effected by the entrance of the head of a single spermatozoon (the tail remaining outside) through the micropyle, and then along the column of clear protoplasm to the female pronucleus.

XII. ON THE STRUCTURE AND DEVELOPMENT OF THE VERTEBRATE OVARY[370].

Footnote 370: From the _Quarterly Journal of Microscopical Science_, Vol. 18, 1878.

(With Plates 24, 25, 26.)

The present paper records observations on the ovaries of but two types, viz., Mammalia and Elasmobranchii. The main points dealt with are three:--1. The relation of the germinal epithelium to the stroma. 2. The connection between _primitive ova_ in Waldeyer's sense and the permanent ova. 3. The homologies of the egg membranes.

The second of these points seems to call for special attention after Semper's discovery that the primitive ova ought really to be regarded as _primitive sexual cells_, in that they give rise to the generative elements of both sexes.

THE DEVELOPMENT OF THE ELASMOBRANCH OVARY.

The development of the Elasmobranch ovary has recently formed the subject of three investigations. The earliest of them, by H. Ludwig, is contained in his important work, on the 'Formation of the Ovum in the Animal Kingdom[371].' Ludwig arrives at the conclusion that the ovum and the follicular epithelium are both derived from the germinal epithelium, and enters into some detail as to their formation. Schultz[372], without apparently being acquainted with Ludwig's observations, has come to very similar results for Torpedo.

Footnote 371: _Arbeiten a. d. zool.-zoot. Institut Würzburg_, Bd. I.

Footnote 372: _Archiv f. micr. Anat._ Vol. XI.

Semper[373], in his elaborate memoir on the urogenital system of Elasmobranchii, has added very greatly to our knowledge on this subject. In a general way he confirms Ludwig's statements, though he shews that the formation of the ova is somewhat more complicated than Ludwig had imagined. He more especially lays stress on the existence of nests of ova (Ureiernester), derived from the division of a single primitive ovum, and of certain peculiarly modified nuclei, which he compares to spindle nuclei in the act of division.

Footnote 373: _Arbeiten a. d. zool.-zoot. Institut Würzburg_, Bd. II.

My own results agree with those of previous investigators, in attributing to the germinal epithelium the origin both of the follicular epithelium and ova, but include a number of points which I believe to be new, and, perhaps, of some little interest; they differ, moreover, in many important particulars, both as to the structure and development of the ovary, from the accounts of my predecessors.

The history of the female generative organs may conveniently be treated under two heads, viz. (1) the history of the ovarian ridge itself, and (2) the history of the ova situated in it. I propose dealing in the first place with the ovarian ridge.

_The Ovarian ridge in Scyllium._--At the stage spoken of in my monograph on Elasmobranch Fishes as stage L, the ovarian ridge has a very small development, and its maximum height is about 0.1 mm. It exhibits in section a somewhat rounded form, and is slightly constricted along the line of attachment. It presents two surfaces, which are respectively outer and inner, and is formed of a layer of somewhat thickened germinal epithelium separated by a basement membrane from a central core of stroma. The epithelium is far thicker on the outer surface than on the inner, and the primitive ova are entirely confined to the former. The cells of the germinal epithelium are irregularly scattered around the primitive ova, and have not the definite arrangement usually characteristic of epithelial cells. Each of them has a large nucleus, with a deeply staining small nucleolus, and a very scanty protoplasm. In stage N the ovarian ridge has a pointed edge and narrower attachment than in stage L. Its greatest height is about 0.17 mm. There is more stroma, and the basement membrane is more distinct than before; in other respects no changes worth recording have taken place. By stage P a distinction is observable between the right and left ovarian ridges; the right one has, in fact, grown more rapidly than the left, and the difference in size between the two ridges becomes more and more conspicuous during the succeeding stages, till the left one ceases to grow any larger, though it remains for a great part of life as a small rudiment.

The right ovarian ridge, which will henceforth alone engage our attention, has grown very considerably. Its height is now about 0.4 mm. It has in section (vide Pl. 24, fig. 1) a triangular form with constricted base, and is covered by a flat epithelium, except for an area on the outer surface, in length co-extensive with the ovarian ridge, and with a maximum breadth of about 0.25 mm. This area will be spoken of as the ovarian area or region, since the primitive ova are confined to it. The epithelium covering it has a maximum thickness of about 0.05 mm., and thins off rather rapidly on both borders, to become continuous with the general epithelium of the ovarian ridge. Its cells have the same character as before, and are several layers deep. Scattered irregularly amongst them are the primitive ova. The germinal epithelium in the ovarian region is separated by a basement membrane from the adjacent stroma.

In succeeding stages, till the embryo reaches a length of 7 centimetres, no very important changes take place. The ovarian region grows somewhat in breadth, though in this respect different embryos vary considerably. In two embryos of nearly the same age, the breadth of the ovarian epithelium was 0.3 mm. in the one and 0.35 mm. in the other. In the former of these embryos, the thickness of the epithelium was slightly greater than in the latter, viz. 0.09 mm. as compared with 0.08. In both the epithelium was sharply separated from the subjacent stroma. There were relatively more epithelial cells in proportion to primitive ova than at the earlier date, and the individual cells exhibited great variations in shape, some being oval, some angular, others very elongated, and many of them applied to part of an ovum and accommodating themselves to its shape. In some of the more elongated cells very deeply stained nuclei were present, which (in a favourable light and with high powers) exhibited the spindle modification of Strasburger with great clearness, and must therefore be regarded as undergoing division. The ovarian region is at this stage bounded on each side by a groove.

In an embryo of seven centimetres (Pl. 24, fig. 2) the breadth of the ovarian epithelium was 0.5, but its height only 0.06 mm. It was still sharply separated from the subjacent stroma, though a membrane could only be demonstrated in certain parts. The amount of stroma in the ovarian ridge varies greatly in different individuals, and no reliance can be placed on its amount as a test of the age of the embryo. In the base of the ovarian ridge the cells were closely packed, elsewhere they were still embryonic.

My next stage (Pl. 24, fig. 3, and fig. 4), shortly before the time of the hatching of the embryo, exhibits in many respects an advance on the previous one. It is the stage during which a follicular covering derived from the germinal epithelium is first distinctly formed round the ova, in a manner which will be more particularly spoken of in the section devoted to the development of the ovum itself. The breadth of the ovarian region is 0.56 mm., and its greatest height close to the central border, 0.12 mm.--a great advance on the previous stage, mainly, however, due to the larger size of the ova.

The ovarian epithelium is still in part separated from the subjacent stroma by a membrane close to its dorsal and ventral borders, but elsewhere the separation is not so distinct, it being occasionally difficult within a cell or so to be sure of the boundary of the epithelium. The want of a clear line between the stroma and the epithelium is rendered more obvious by the fact that the surface of the latter is somewhat irregular, owing to projections formed by specially large ova, into the bays between which are processes of the stroma. In an ovary about this stage, hardened in osmic acid, the epithelium stains very differently from the subjacent stroma, and the line of separation between the two is quite sharp. A figure of the whole ovarian ridge, shewing the relation between the two parts, is represented on Pl. 24, fig. 5.

The layer of stroma in immediate contact with the epithelium is very different from the remainder, and appears to be destined to accompany the vascular growths into the epithelium, which will appear in the next stage. The protoplasm of the cells composing it forms a loose reticulum with a fair number of oval or rounded nuclei, with their long axis for the most part parallel to the lower surface of the epithelium. It contains, even at this stage, fully developed vascular channels.

The remainder of the stroma of the ovarian ridge has now acquired a definite structure, which remains constant through life, and is eminently characteristic of the genital ridge of both sexes. The bulk of it (Pl. 24, fig. 3, _str_) consists of closely packed polygonal cells, of about 0.014 mm. with large nuclei of about 0.009. These cells appear to be supported by a delicate reticulum. The whole tissue is highly vascular, with the numerous capillaries; the nuclei in the walls of which stand out in some preparations with great clearness.

In the next oldest ovary, of which I have sections, the breadth of the ovarian epithelium is 0.7 mm. and its thickness 0.096. The ovary of this age was preserved in osmic acid, which is the most favourable reagent, so far as I have seen, for observing the relation of the stroma and epithelium. On Pl. 24, fig. 6, is represented a transverse section through the whole breadth of the ovary, slightly magnified to shew the general relations of the parts, and on Pl. 24, fig. 7, a small portion of a section more highly magnified. The inner surface of the ovarian epithelium is more irregular than in the previous stage, and it may be observed that the subjacent stroma is growing in amongst the ova. From the relation of the two tissues it is fairly clear that the growth which is taking place is a definite growth of the stroma into the epithelium, and not a mutual intergrowth of the two tissues. The ingrowths of the stroma are, moreover, directed towards individual ova, around which, outside the follicular epithelium, they form a special vascular investment in the succeeding stages. They are formed of a reticular tissue with comparatively few nuclei.

By the next stage, in my series of ovaries of _Scy. canicula_, important changes have taken place in the constitution of ovarian epithelium. Fig. 8, Pl. 24, represents a portion of the ovarian epithelium, on the same scale as figs. 1, 2, 3, &c., and fig. 9 a section through the whole ovarian ridge slightly magnified. Its breadth is now 1.3 mm., and its thickness 0.3 mm. The ova have grown very greatly, and it appears to me to be mainly owing to their growth that the greater thickness of the epithelium is due, as well as the irregularity of its inner surface (vide fig. 9).

The general relation of the epithelium to the surrounding parts is much the same as in the earlier stage, but two new features have appeared--(1) The outermost cells of the ovarian region have more or less clearly arranged themselves as a kind of epithelial covering for the organ; and (2) the stroma ingrowths of the previous stage have become definitely vascular, and have penetrated through all parts of the epithelium.

The external layer of epithelium is by no means a very marked structure, the character of its cells varies greatly in different regions, and it is very imperfectly separated from the subjacent layer. I shall speak of it for convenience as _pseudo-epithelium_.

The greater part of the germinal epithelium forms anastomosing columns, separated by very thin tracts of stroma. The columns are, in the majority of instances, continuous with the pseudo-epithelium at the surface, and contain ova in all stages of development. Many of the cells composing them naturally form the follicular epithelium for the separate ova; but the majority have no such relation. They have in many instances assumed an appearance somewhat different from that which they presented in the last stage, mainly owing to the individual nuclei being more widely separated. A careful examination with a high power shews that this is owing to an increase in the amount of protoplasm of the individual cells, and it may be noted that a similar increase in the size of the bodies of the cells has taken place in the pseudo-epithelium and in the follicular epithelium of the individual ova.

The stroma ingrowths form the most important feature of the stage. In most instances they are very thin and delicate, and might easily be overlooked, especially as many of the cells in them are hardly to be distinguished, taken separately, from those of the germinal epithelium. These features render the investigation of the exact relation of the stroma and epithelium a matter of some difficulty. I have, however, been greatly assisted by the investigation of the ovary of a young example of _Scyllium stellare_, 16-1/2 centimètres in length, a section of which is represented in Pl. 25, fig. 26. In this ovary, although no other abnormalities were observable, the stroma ingrowths were exceptionally wide; indeed, quite without a parallel in my series of ovaries in this respect. The stroma most clearly divides up the epithelium of the ovary into separate masses, or more probably anastomosing columns, the equivalents of the egg-tubes of Pflüger. These columns are formed of normal cells of the germinal epithelium, which enclose ovarian nests and ova in all stages of development. A comparison of the section I have represented, with those from previous stages, appears to me to demonstrate that the relation of the epithelium and stroma has been caused by an ingrowth or penetration of the stroma into the epithelium, and not by a mutual intergrowth of the two tissues. Although the ovary, of which fig. 26 represents a section was from _Scy. stellare_, and the previous ovaries have been from _Scy. canicula_, yet the thickness of the epithelium may still be appealed to in confirmation of this view. In the previous stage the thickness was about 0.096 mm., in the present one it is about 0.16 mm., a difference of thickness which can be easily accounted for by the growth of the individual ova and the additional tracts of stroma. A pseudo-epithelium is more or less clearly formed, but it is continuous with the columns of epithelium. In the stroma many isolated cells are present, which appear to me, from a careful comparison of a series of sections, to belong to the germinal epithelium.

The thickness of the follicular epithelium on the inner side of the larger ova deserves to be noted. Its meaning is discussed on p. 567.

Quite a different interpretation to that which I have given has been put by Ludwig and Semper upon the parts of the ovary at this stage. My _pseudo-epithelium_ is regarded by them as forming, together with the _follicular epithelium_ of the ova, the sole remnant of the original germinal epithelium; and the masses of cells below the pseudo-epithelium, which I have attempted to shew are derived from the original germinal epithelium, are regarded as parts of the ingrowths of the adjacent stroma.

Ludwig has assumed this interpretation without having had an opportunity of working out the development of the parts, but Semper attempts to bring forward embryological proofs in support of this position.

If the series of ovaries which I have represented be examined, it will not, I think, be denied that the general appearances are very much in favour of my view. The thickened patch of ovarian epithelium can apparently be traced through the whole series of sections, and no indications of its sudden reduction to the thin pseudo-epithelium are apparent. The most careful examination that I have been able to make brings to light nothing tending to shew that the general appearances are delusive. The important difference between us refers to _our views of the nature of the tissue subjacent to the pseudo-epithelium_. If my results be accepted, it is clear that the whole ovarian region is an epithelium interpenetrated by connective tissue ingrowths, so that the region below the pseudo-epithelium is a kind of honeycomb or trabecular net-work of germinal epithelium, developing ova of all stages and sizes, and composed of cells capable of forming follicular epithelium for developing ova. Ludwig figures what he regards as the formation of the follicular epithelium round primitive ova during their passage into the stroma. It is quite clear to me, that his figures of the later stages, 33 and 34, represent fully formed permanent ova surrounded by a follicular epithelium, and that their situation in contact with the pseudo-epithelium is, so to speak, an accident, and it is quite possible that his figures 31 and 32 also represent fully formed ova; but I have little hesitation in asserting that he has not understood the mode of formation of the follicular epithelium, and that, though his statement that it is derived from the germinal epithelium is quite correct, his account of the process is completely misleading. The same criticism does not exactly apply to Semper's statements. Semper has really observed the formation of the follicular epithelium round young ova; but, nevertheless, he appears to me to give an entirely wrong account of the relation of the stroma to the germinal epithelium. The extent of the difference between Semper's and my view may perhaps best be shewn by a quotation from Semper, _loc. cit._, 465:--"In females the nests of primitive ova sink in groups into the stroma. In these groups one cell enlarges till it becomes the ovum, the neighbouring cells increase and arrange themselves around the ova as follicle cells."

Although the histological changes which take place in the succeeding stages are not inconsiderable, they do not involve any fundamental change in the constitution of the ovarian region, and may be described with greater brevity than has been so far possible.

In a half-grown female, with an ovarian region of 3mm. in breadth, and 0.8mm. in thickness, the stroma of the ovarian region has assumed a far more formed aspect than before. It consists (Pl. 24, fig. 10) of a basis in most parts fibrous, but in some nearly homogeneous, with a fair number of scattered cells. Immediately below the pseudo-epithelium, there is an imperfectly developed fibrous layer, forming a kind of tunic, in which are imbedded the relatively reduced epithelial trabeculæ of the previous stages. They appear in sections as columns, either continuous with or independent of the pseudo-epithelium, formed of normal cells of the germinal epithelium, nests of ova, and permanent ova in various stages of development. Below this there comes a layer of larger ova which are very closely packed. A not inconsiderable number of the larger ova have, however, a superficial situation, and lie in immediate contact with the pseudo-epithelium. Some of the younger ova, enclosed amongst epithelial cells continuous with the pseudo-epithelium, are very similar to those figured by Ludwig. It is scarcely necessary to insist that this fact does not afford any argument in favour of his interpretations. The ovarian region is honeycombed by large vascular channels with distinct walls, and other channels which are perhaps lymphatic.

The surface of the ovarian region is somewhat irregular and especially marked by deep oblique transverse furrows. It is covered by a distinct, though still irregular pseudo-epithelium, which is fairly columnar in the furrows but flattened along the ridges. The cells of the pseudo-epithelium have one peculiarity very unlike that of ordinary epithelial cells. Their inner extremities (vide fig. 10) are prolonged into fibrous processes which enter the subjacent tissue, and bending nearly parallel to the surface of the ovary, assist in forming the tunic spoken of above. This peculiarity of the pseudo-epithelial cells seems to indicate that they do not essentially differ from cells which have the character of undoubted connective tissue cells, and renders it possible that the greater part of the tunic, which has apparently the structure of ordinary connective tissue, is in reality derived from the original germinal epithelium, a view which tallies with the fact that in some instances the cells of the tunic appear as if about to assist in forming the follicular epithelium of some of the developing ova. In Raja, the similarity of the pseudo-epithelium to the subjacent tissue is very much more marked than in Scyllium. The pseudo-epithelium appears merely as the superficial layer of the ovarian tunic somewhat modified by its position on the surface. It is formed of columnar cells with vertically arranged fibres which pass into the subjacent layers, and chiefly differ from the ordinary fibres in that they still form parts of the cell-protoplasm enclosing the nucleus. In Pl. 25, fig. 34, an attempt is made to represent the relations of the pseudo-epithelium to the subjacent tissue in Raja. Ludwig's figures of the pseudo-epithelium of the ovary, in the regular form of its constituent cells, and its sharp separation by a basement membrane from the tissue below, are quite unlike anything which I have met with in my sections either of Raja or Scyllium.

Close to the dorsal border of the ovary the epithelial cells of the non-ovarian region have very conspicuous tails, extending into a more or less homogeneous substance below, which constitutes a peculiar form of tunic for this part of the ovarian ridge.

In the full-grown female the stroma of the ovarian region is denser and has a more fibrous aspect than in the younger animal. Below the pseudo-epithelium it is arranged in two or three more or less definite layers, in which the fibres run at right angles. It forms a definite ovarian tunic. The pseudo-epithelium is much more distinct, and the tails of its cells, so conspicuous in previous stages, can no longer be made out.

_Formation of the permanent ova and the follicular epithelium._--In my monograph on the development of Elasmobranch Fishes an account was given of the earliest stages in the development of the primitive ova, and I now take up their development from the point at which it was left off in that work. From their first formation till the stage spoken of in my monograph as P, their size remains fairly constant. The larger examples have a diameter of about 0.035 mm., and the medium-sized examples of about 0.03 mm. The larger nuclei have a diameter of about 0.16 mm., but their variations in size are considerable. If the above figures be compared with those on page 350 of my monograph on Elasmobranch Fishes, it will be seen that the size of the primitive ova during these stages is not greater than it was at the period of their very first appearance.

The ova (Pl. 24, fig. 1) are usually aggregated in masses, which might have resulted from division of a single ovum. The outlines of the individual ova _are always distinct_. Their protoplasm is clear, and their nuclei, which are somewhat passive towards staining reagents, are granular, with one to three nucleoli. I have noticed, up to stage P, the occasional presence of highly refractive spherules in the protoplasm of the primitive ova already described in my monograph (pp. 353, 354, Pl. 12, fig. 15). They seem to occur up to a later period than I at first imagined. Their want of constancy probably indicates that they have no special importance. Professor Semper has described similar appearances in the male primitive ova of a later period.

As to the distribution of the primitive ova in the germinal epithelium, Professor Semper's statement that the larger primitive ova are found in masses in the centre, and that the smaller ova are more peripherally situated is on the whole true, though I do not find this distribution sufficiently constant to lay so much stress on it as he does.

The passive condition of the primitive ova becomes suddenly broken during stage Q, and is succeeded by a period of remarkable changes. It has only been by the expenditure of much care and trouble that I have been able to elucidate to my own satisfaction what takes place, and there are still points which I do not understand.

Very shortly after stage Q, in addition to primitive ova with a perfectly normal nucleus, others may be seen in which the nucleus is apparently replaced by a deeply stained irregular body, smaller than the ordinary nuclei (Pl. 24, fig. 11, _d.n._). This body, by the use of high objectives, is seen to be composed of a number of deeply stained granules, and around it may be noticed a clear space, bounded by a very delicate membrane. The granular body usually lies close to one side of this membrane, and occasionally sends a few fine processes to the opposite side.

The whole body, _i.e._ all within the delicate membrane is, according to my view, a modified nucleus; as appears to me very clearly to be shewn by the fact that it occupies the normal position of a nucleus within a cell body. Semper, on the other hand, regards the contained granular body as the nucleus, which he compares with the spindles of Bütschli, Auerbach, &c.[374]. This interpretation appears to me, however, to be negatived by the position of these bodies. The manner in which Semper may, perhaps, have been led to his views will be obvious when the later changes of the primitive ova are described. The formation of these nuclei would seem to be due to a segregation of the constituents of the original nuclei; the solid parts becoming separated from the more fluid. As a rule, the modified nuclei are slightly larger than the original ones. In stage Q the following two tables shew the dimensions of the parts of three unmodified and of three modified nuclei taken at random.

Footnote 374: _Loc. cit._ p. 361.

_Primitive ova with unmodified nuclei_--

Nuclei.

0.014 mm. 0.012 mm. 0.01 mm.

_Primitive ova with modified nuclei_--

Granular Nuclei. Bodies in nuclei.

0.018 mm. 0.006 mm. 0.018 mm. 0.006 mm. 0.012 mm. 0.009 mm.

For a slightly older stage than Q, the two annexed tables also shew the comparative size of the modified and unmodified nuclei:

_Unmodified nuclei of normal primitive ova--_

0.014 mm. 0.016 mm. 0.014 mm. 0.016 mm. 0.016 mm.

_Nuclei of primitive ova with modified nuclei--_

Granular Nuclei. Bodies in Nuclei.

0.018 mm. 0.008 mm. 0.016 mm. 0.008 mm. 0.016 mm. 0.01 mm. 0.016 mm. 0.018 mm.

These figures bring out with clearness the following points: (1) that the modified nuclei are slightly but decidedly larger on the average than the unmodified nuclei; (2) that the contained granular bodies _are very considerably_ smaller than ordinary nuclei.

Soon after the appearance of the modified nuclei, remarkable changes take place in the cells containing them. Up to the time such nuclei first make their appearance the outlines of the individual ova are very clearly defined, but subsequently, although numerous ova with but slightly modified nuclei are still to be seen, yet on the whole the outlines of all the primitive ova are much less distinct than before; and this is especially the case with the primitive ova containing modified nuclei.

From cases in which three or four ova are found in a mass with modified nuclei, but in which the outline of each ovum is fairly distinct, it is possible to pass by insensible gradations to other cases in which two or three or more modified nuclei are found embedded in a mass of protoplasm in which no division into separate cells can be made out (fig. 14). For these masses I propose to employ the term nests. They correspond in part with the _Ureiernester_ of Professor Semper.

Frequently they are found in hardened specimens to be enclosed in a membrane-like tunic which appears to be of the nature of coagulated fluid. These membranes closely resemble and sometimes are even continuous with trabeculæ which traverse the germinal epithelium. Ovaries differ considerably as to the time and completeness of the disappearance of the outlines marking the separate cells, and although, so far as can be gathered from my specimens, the rule is that the outlines of the primitive ova with modified nuclei soon become indistinct, yet in one of my best preserved ovaries very large nests with modified nuclei are present in which the outline of each ovum is as distinct as during the period before the nuclei undergo these peculiar changes (Pl. 24, fig. 12). In the same ovary other nests are present in which the outlines of the individual ova are no longer visible. The section represented on Pl. 24, fig. 2, is fairly average as to the disappearance of the outlines of the individual ova.

It is clear from the above statements, that in the first instance the nests are produced by the coalescence of several primitive ova into a single mass or syncytium; though of course, the several separate ova of a nest may originally, as Semper believes, have arisen from the division of a single ovum. In any case there can be no doubt that the nests of separate ova increase in size as development proceeds; a phenomenon which is more reasonably explained on the view that the ova divide, than on the view that they continue to be freshly formed. The same holds true for the nests of nuclei and this, as well as other facts, appears to me to render it probable that the nests grow by division of the nuclei without corresponding division of the protoplasmic matrix. I cannot, however, definitely prove this point owing to my having found nests, with distinct outlines to the ova, as large as any without such outlines.

The nests are situated for the most part near the surface of the germinal epithelium. The smaller ones are frequently spherical, but the larger are irregular in form. The former are about 0.05 mm. in diameter; the latter reach 0.1 mm. Scattered generally, and especially in the deeper layers, and at the edges of the germinal epithelium, are still unmodified or only slightly modified primitive ova. These unmodified primitive ova are aggregated in masses, but in these masses the outlines of each ovum, though perhaps less clear than in the earlier period, are still distinct.

When the embryo reaches a length of seven centimètres, and even in still younger embryos, further changes are observable. In the first place many of the modified nuclei acquire fresh characters, and it becomes necessary to divide the modified nuclei into two categories. In both of these the outer boundary of the nucleus is formed by a very delicate membrane, the space within which is perfectly clear except for the granular body. In the variety which now appears in considerable numbers the granular body has an irregular star-like form. The rays of the star are formed of fibres frequently knobbed at their extremities, and the centre of the star usually occupies an eccentric position. Typical examples of this form of modified nucleus, which may be spoken of as the stellate variety, are represented on Pl. 25, fig. 17; between it and the older granular variety there is an infinite series of gradations, many of which are represented on Pl. 24, figs. 12, 14, 15, 16. Certain of the stellate nuclei exhibit two centres instead of one, and in some cases, like that represented on Pl. 25, fig. 19, the stellate body of two nuclei is found united. Both of these forms are possibly modifications of the spindle-like form assumed by nuclei in the act of dividing, and may be used in proving that the nests increase in size by the division of the contained nuclei. In addition to the normal primitive ova, a few of which are still present, there are to be found, chiefly in the deeper layers of the germinal epithelium, larger ova differing considerably from the primitive ova. They form the permanent ova (Pl. 24, fig. 3, _o_). Their average diameter is 0.04 mm., compared with 0.03 mm., the diameter of original primitive ova. The protoplasm of which they are composed is granular, but at first a membrane can hardly be distinguished around them; their nucleus is relatively large, 0.02 - 0.027 mm. in diameter. It presents the characters ascribed by Eimer[375], and many other recent authors[376], to typical nuclei (vide Pl. 24, fig. 3, and Pl. 24, 25, figs. 13, 14, 15, 16, 17, 18). It is bounded by a distinct membrane, within which is a more or less central nucleolus from which a number of radial fibres which stain very deeply pass to the surface; here they form immediately internal to the membrane a network with granules at the nodal points. In some instances the regularity of the arrangement of these fibres is very great, in other instances two central nucleoli are present, in which case the regularity is considerably interfered with. The points in which the youngest permanent ova differ from the primitive may be summed up as follows:--

(1) The permanent ova are larger, the smallest of them being larger than the average primitive ova in the proportion of four to three. (2) They have less protoplasm as compared to the size of the nucleus. (3) Their protoplasm is granular instead of being clear. (4) Their nucleus is clear with exception of a network of fibres instead of being granular as in the primitive ova. It thus appears that the primitive ova and permanent ova are very different in constitution, though genetically related in a way to be directly narrated.

Footnote 375: _Archiv f. micr. Anat._ Vol. XIV.

Footnote 376: Vide especially Klein, _Quart. Journ. of Mic. Sci._ July 1878.

The formation of permanent ova is at its height in embryos of about seven centimètres or slightly larger. The nests at this stage are for the most part of a very considerable size and contain a large number of nuclei, which have probably, as before insisted, originated from a division of the smaller number of nuclei present in the nests at an earlier stage. Figs. 14-18 are representations of nests at this period. The diameter of the nuclei is, on the whole, slightly greater than at an earlier stage. A series of measurements gave the following results:--

0.016 mm. 0.016 mm. 0.018 mm. 0.02 mm. 0.02 mm.

Both varieties of modified nuclei are common enough, though the stellate variety predominates. The nuclei are sometimes in very close contact, and sometimes separated by protoplasm, which in many instances is very slightly granular. In a large number of the nests nothing further is apparent than what has just been described, but in a very considerable number one or more nuclei are present, which exhibit a transitional character between the ordinary stellate nuclei of my second category, and the nuclei of permanent ova as above described; and in these nests the formation of permanent ova is taking place. Permanent ova in the act of development are indicated in my figures by the letters _do_. Many of the intermediate nuclei are more definitely surrounded by granular protoplasm than the other nuclei of the nests, and accordingly have their outlines more sharply defined. Between nuclei of this kind, and others as large as those of the permanent ova, there are numerous transitional forms. The larger ones frequently lie in a mass of granular protoplasm projecting from the nest, and only united with it by a neck (Pl. 24, figs. 14 and 16). For prominences of this kind to become independent ova, it is only necessary for the neck to become broken through. Nests in which such changes are taking place present various characters. In some cases several nuclei belonging to a nest appear to be undergoing conversion into permanent ova at the same time. Such a case is figured on Pl. 25, figs. 17 and 18. In these cases the amount of granular protoplasm in the nest and around each freshly formed ovum is small. In the more usual cases only one or two permanent ova at the utmost are formed at the same time, and in these instances a considerable amount of granular protoplasm is present around the nucleus of the developing permanent ovum. In such instances it frequently happens several of the nuclei not undergoing conversion appear to be in the process of absorption, and give to the part of the nest in which they are contained a very hazy and indistinct aspect (Pl. 24, fig. 15). Their appearance leads me to adopt the view _that while some of the nuclei of each nest are converted into the nuclei of the permanent ova, others break down and are used as the pabulum, at the expense of which the protoplasm of the young ovum grows_.

It should, however, be stated, that after the outlines of the permanent ova have become definitely established, I have only observed in a single instance the inclusion of a nucleus within an ovum (Pl. 25, fig. 24). In many instances normal nuclei of the germinal epithelium may be so observed within the ovum.

The nuclei which are becoming converted into the nuclei of permanent ova gradually increase in size. The following table gives the diameter of four such nuclei:--

0.022 mm. 0.022 mm. 0.024 mm. 0.032 mm.

These figures should be compared with those of the table on page 564.

The ova when first formed are situated either at the surface or in the deeper layers of the germinal epithelium. Though to a great extent surrounded by the ordinary cells of the germinal epithelium, they are not at first enclosed in a definite follicular epithelium. The follicle is, however, very early formed.

My observations lead me then to the conclusion that in a general way the permanent ova are formed by the increase of protoplasm round some of the nuclei of a nest, and the subsequent separation of the nuclei with their protoplasm from the nest as distinct cells--a mode of formation exactly comparable with that which so often takes place in invertebrate egg tubes.

Besides the mode of formation of permanent ova just described, a second one also seems probably to occur. In ovaries just younger than those in which permanent ova are distinctly formed, there are present primitive ova, with modified nuclei of the stellate variety, or nuclei sometimes even approaching in character those of permanent ova, which are quite isolated and not enclosed in a definite nest. The body of these ova is formed of granular protoplasm, but their outlines are very indistinct. Such ova are considerably larger than the normal primitive ova. They may measure 0.04 mm. In a slightly later stage, when fully formed permanent ova are present, isolated ones are not infrequent, and it seems natural to conclude that these isolated ova are the direct descendants of the primitive ova of the earlier stage. It seems a fair deduction that in some cases primitive ova undergo a direct metamorphosis into permanent ova by a modification of their nucleus, and the assumption of a granular character in their protoplasm, without ever forming the constituent part of a nest.

It is not quite clear to me that in all nests the coalescence of the protoplasm of the ova necessarily takes place, since some nests are to be found at all stages in which the ova are distinct. Nevertheless, I am inclined to believe that the fusion of the ova is the normal occurrence.

The mode of formation of the permanent ova may then, according to my observations, take place in two ways:--1. By the formation of granular protoplasm round the nucleus in a nest, and the separation of the nucleus with its protoplasm as a distinct ovum. 2. By the direct metamorphosis of an isolated primitive ovum into a permanent ovum. The difference between these two modes of formation does not, from a morphological point of view, appear to be of great importance.

The above results appear clearly to shew that _the primitive ova in the female are not to be regarded as true ova, but as the parent sexual cells which give rise to the ova_: a conclusion which completely fits in with the fact that cells exactly similar to the primitive ova in the female give rise to the spermatic cells in the male.

Slightly after the period of their first formation the permanent ova become invested by a very distinct and well-marked, somewhat flattened, follicular epithelium (Pl. 24, fig. 3). Where the ova lie in the deeper layers of the germinal epithelium, the follicular epithelium soon becomes far more columnar on the side turned inwards, than on that towards the surface, especially when the inner side is in contact with the stroma (Pl. 24, fig. 7, and Pl. 25, figs. 24 and 26). This is probably a special provision for the growth and nutrition of the ovum.

There cannot be the smallest doubt that the follicular epithelium is derived from the general cells of the germinal epithelium--a point on which my results fully bear out the conclusions of Ludwig and Semper.

The larger ova themselves have a diameter of about 0.06 mm., and their nucleus of about 0.04 mm. The vitellus is granular, and provided with a distinct, though delicate membrane, which has every appearance of being a product of the ovum itself rather than of the follicular epithelium. The membrane would seem indeed to be formed in some instances even before the ovum has a definite investment of follicle cells. The vitellus is frequently vacuolated, but occasionally the vacuoles appear to be caused by a shrinking due to the hardening reagent. The nucleus has the same peculiar reticulate character as at first. Its large size, as compared with the ovum, is very noticeable.

With this stage the embryonic development of the ova comes to a close, though the formation of fresh ova continues till comparatively late in life. I have, however, two series of sections of ovaries preserved in osmic acid, from slightly larger embryos than the one last described, about which it may be well to say a few words before proceeding to the further development of the permanent ova.

The younger of these ovaries was from a Scyllium embryo 10 centimètres long, preserved in osmic acid.

A considerable number of nests were present (Pl. 24, fig. 13), exhibiting, on the whole, similar characters to those just described.

A series of measurements of the nuclei in them were made, leading to the following results:--

0.014 mm. 0.014 mm. 0.016 mm. 0.016 mm. 0.018 mm. 0.018 mm.

Thus, if anything, the nuclei were slightly smaller than in the younger embryo. It is very difficult in the osmic specimens to make out clearly the exact outlines of the various structures, the nuclei in many instances being hardly more deeply stained than in the protoplasm around them. The network in the nuclei is also far less obvious than after treatment with picric acid. The permanent ova were hardly so numerous as in the younger ovary before described. A number of these were measured with the following results:--

Ovum. Nucleus.

0.03 mm. 0.014 mm. 0.034 mm. 0.018 mm. 0.028 mm. 0.016 mm. 0.03 mm. 0.02 mm. 0.04 mm. 0.02 mm. 0.04 mm. 0.02 mm. 0.048 mm. 0.02 mm.

These figures shew that the nuclei of the permanent ova are smaller than in the younger embryo, and it may therefore be safely concluded that, in spite of the greater size of the embryo from which it is taken, the ovary now being described is in a more embryonic condition than the one last dealt with.

Though the permanent ova appeared to be formed from the nests in the manner already described, it was fairly clear from the sections of this ovary that many of the original primitive ova, after a metamorphosis of the nucleus and without coalescing with other primitive ova to form nests, become converted directly into the permanent ova. Many large masses of primitive ova, or at least of ova with the individual outlines of each ovum distinct, were present. The average size of ova composing these was however small, the body measuring about 0.016 mm., and the nucleus 0.012 mm. Isolated ova with metamorphosed nuclei could also be found measuring 0.022, and their nuclei about 0.014 mm.

The second of the two ovaries, hardened in osmic acid, was somewhat more advanced than the ovary in which the formation of permanent ova was at its height. Fewer permanent ova were in the act of being formed, and many of these present had reached a considerable size, measuring as much as 0.07 mm. Nests of the typical forms were present as before, but the nuclei in them were more granular than at the earlier period, and on the average slightly smaller. A series measured had the following diameters:--

0.01 mm. 0.012 mm. 0.014 mm. 0.016 mm.

One of these nests is represented on Pl. 25, fig. 20. Many nests with the outlines of the individual ova distinct were also present.

On the whole it appeared to me, that the second mode of formation of permanent ova, viz. that in which the nest does not come into the cycle of development, preponderated to a greater extent than in the earlier embryonic period.

POST-EMBRYONIC DEVELOPMENT OF THE OVA.--My investigations upon the post-embryonic growth and development of the ova, have for the most part been conducted upon preserved ova, and it has been impossible for me, on this account, to work out, as completely as I should have wished, certain points, more especially those connected with the development of the yolk.

Although my ovaries have been carefully preserved in a large number of reagents, including osmic acid, picric acid, chromic acid, spirit, bichromate of potash, and Müller's fluid, none of these have proved universally successful, and bichromate of potash and Müller's fluid are useless. Great difficulties have been experienced in distinguishing the artificial products of these reagents. My investigations have led me to the result, that in the gradual growth of the ova with the age of the individual the changes are not quite identical with those during the rapid growth which takes place at periods of sexual activity, after the adult condition has been reached--a result to which His has also arrived, with reference to the ova of Osseous Fish. I propose dealing separately with the several constituents of the egg-follicle.

_Egg membranes._--A vitelline membrane has been described by Leydig[377] in Raja, and an albuminous layer of the nature of a chorion[378] by Gegenbaur[379] in Acanthias--the membranes described in these two ways being no doubt equivalent.

Footnote 377: _Rochen u. Haie._

Footnote 378: By _chorion_ I mean, following E. van Beneden's nomenclature, a membrane formed by the follicular epithelium, and, by _vitelline membrane_, one formed by the vitellus or body of the ovum.

Footnote 379: "Bau und Entwicklung d. Wirbelthiereier," &c., _Müll. Archiv_, 1861.

Dr Alex. Schultz[380] has more recently investigated a considerable variety of genera and finds three conditions of the egg membranes. (1) In Torpedo, a homogeneous membrane, which is of the nature of a chorion. (2) In Raja, a homogeneous membrane which is, however, perforated. (3) In Squalidæ, a thick homogeneous membrane, internal to which is a thinner perforated membrane. He apparently regards the perforated inner membrane as a specialised part of the simple membrane found in Torpedo, and states that this membrane is of the nature of a chorion.

Footnote 380: "Zur Entwicklungsgeschichte d. Selachier," _Arch. f. mikr. Anat._ Vol. XI.

My own investigations have led me to the conclusion that though the egg-membranes can probably be reduced to single type for Elasmobranchii, yet that they vary with the stage of development of the ovum. Scyllium (stellare and canicula) and Raja have formed the objects of my investigation. I commence with the two former.

It has already been stated that in Scyllium, even before the follicular epithelium becomes formed, a delicate membrane round the ovum can be demonstrated, which appears to me to be derived from the vitellus or body of the ovum, and is therefore of the nature of a vitelline membrane. It becomes the vitelline membrane of Leydig, the albuminous membrane of Gegenbaur, and homogeneous membrane of Schultz.

In a young fish (not long hatched) with ova of not more than 0.12 mm., this membrane, though considerably thicker than in the embryo, is not thick enough to be accurately measured. In ova of 0.5 mm. from a young female (Pl. 25, fig. 21) the vitelline membrane has a thickness of 0.002 mm. and is quite homogeneous[381]. Internally to it may be observed very faint indications of the differentiation of the outermost layer of the vitellus into the perforated or radially striated membrane of Schultz, which will be spoken of as _zona radiata_.

Footnote 381: The apparent structure in the vitelline membrane in my figure is merely intended to represent the dark colour assumed by it on being stained. The zona radiata has been made rather too thick by the artist.

In an ovum of 1 mm. from the nearly full grown though not sexually mature female, the zona radiata has increased in thickness and definiteness, and may measure as much as 0.004 mm. It is always very sharply separated from the vitelline membrane, but appears to be more or less continuous on its inner border with the body of the ovum, at the expense of which it no doubt grows in thickness.

In ova above 1 mm. in diameter, both vitelline membrane and zona radiata, but especially the latter, increase in thickness. The zona becomes marked off from the yolk, and its radial striæ become easy to see even with comparatively low powers. In many specimens it appears to be formed of a number of small columns, as described by Gegenbaur and others. The stage of about the greatest development of both the vitelline membrane and zona radiata is represented on Pl. 25, fig. 22.

At this time the vitelline membrane appears frequently to exhibit a distinct stratification, dividing it into two or more successive layers. It is not, however, acted on in the same manner by all reagents, and with absolute alcohol appears at times longitudinally striated.

From this stage onwards, both vitelline membrane and zona gradually atrophy, simultaneously with a series of remarkable changes which take place in the follicular epithelium. The zona is the first to disappear, and the vitelline membrane next becomes gradually thinner. Finally, when the egg is nearly ripe, the follicular epithelium is separated from the yolk by an immeasurably thin membrane--the remnant of the vitelline membrane--only visible in the most favourable sections (Pl. 25, fig. 23, _vt._). When the egg becomes detached from the ovary even this membrane is no longer to be seen.

Both the vitelline membrane and the zona radiata are found in Raja, but in a much less developed condition than in Scyllium. The vitelline membrane is for a long time the only membrane present, but is never very thick (Pl. 25, fig. 31). The zona is not formed till a relatively much later period than in Scyllium, and is always delicate and difficult to see (Pl. 25, fig. 32). Both membranes atrophy before the egg is quite ripe; and an apparently fluid layer between the follicular epithelium and the vitellus, which coagulates in hardened specimens, is probably the last remnant of the vitelline membrane. It is, however, much thicker than the corresponding remnant in Scyllium.

Though I find the same membranes in Scyllium as Alexander Schultz did in other Squalidæ, my results do not agree with his as to Raja. Torpedo I have not investigated.

It appears to me probable that the ova in all Elasmobranch Fishes have at some period of their development the two membranes described at length for Scyllium. Of these the inner one, or zona radiata, will probably be admitted on all hands to be a product of the peripheral protoplasm of the egg.

The outer one corresponds with the membrane usually regarded in other Vertebrates as a chorion or product of the follicular epithelium, but, by tracing it back to its first origin, I have been led to reject this view of its nature.

_The follicular epithelium._--The follicular epithelium in the eggs of Raja and Acanthias has been described by Gegenbaur[382]. He finds it flat in young eggs, but in the larger eggs of Acanthias more columnar, and with the cells wedged in so as to form a double layer. These observations are confirmed by Ludwig[383].

Footnote 382: _Loc. cit._

Footnote 383: _Loc. cit._

Alexander Schultz[384] states that in Torpedo, the eggs are at first enclosed in a simple epithelium, but that in follicles of .008 mm. there appear between the original large cells of the follicle (which he describes as granulosa cells and derives from the germinal epithelium) a number of peculiar small cells. He states that these are of the same nature as the general stroma cells of the ovary, and believes that they originate in the stroma. When the eggs have reached 0.1 - 0.15 mm., he finds that the small and large cells have a very regular alternating arrangement.

Footnote 384: _Loc. cit._

Semper records but few observations on the follicular epithelium, but describes in Raja the presence of a certain number of large cells amongst smaller cells. He believes that they may develop into ova, and considers them identical with the larger cells described by Schultz, whose interpretations he does not, however, accept.

My own results accord to a great extent with those of Dr Schultz, as far as the structure of the follicular epithelium is concerned, but I am at one with Semper in rejecting Schultz's interpretations.

In Scyllium, as has already been mentioned, the follicular epithelium is at first flat and formed of a single layer of uniform cells, each with a considerable amount of clear protoplasm and a granular nucleus. It is bounded externally by a delicate membrane--the membrana propria folliculi of Waldeyer--and internally by the vitelline membrane. In the ovaries of very young animals the cells of the follicular epithelium are more columnar on the side towards the stroma than on the opposite side, but this irregularity soon ceases to exist.

In many cases the nuclei of the cells of the follicular epithelium exhibit a spindle modification, which shews that the growth of the follicular epithelium takes place by the division of its cells. No changes of importance are observable in the follicular epithelium till the egg has reached a diameter of more than 1 mm.

It should here be stated that I have some doubts respecting the completeness of the history of the epithelium recorded in the sequel. Difficulties have been met with in completely elucidating the chronological order of the occurrences, and it is possible that some points have escaped my observation.

The first important change is the assumption of a palisade-like character by the follicle cells, each cell becoming very narrow and columnar and the nucleus oval (Pl. 25, fig. 28). In this condition the thickness of the epithelium is about 0.025 mm. The epithelium does not, however, become uniformly thick over the whole ovum, but in the neighbourhood of the germinal vesicle it is very flat and formed of granular cells with indistinct outlines, rather like the hypodermis cells of many Annelida. Coincidently with this change in the follicular epithelium the commencement of the atrophy of the membranes of the ovum, described in the last section, becomes apparent.

The original membrana propria folliculi is still present round the follicular epithelium, but is closely associated with a fibrous layer with elongated nuclei. Outside this there is now a layer of cells, very much like an ordinary epithelial layer, which may possibly be formed of cells of the true germinal epithelium (fig. 28, _fe´_). This layer, which will be spoken of as the secondary follicle layer, might easily be mistaken for the follicular epithelium, and it is possible that it has actually been so mistaken by Eimer, Clark, and Klebs, in Reptilia, and that the true follicular epithelium (in a flattened condition) has been then spoken of as the _Binnenepithel_.

In slightly older eggs the epithelial cells are no longer uniform or arranged as a single layer. The general arrangement of these cells is shewn in Pl. 25, fig. 29. A considerable number of them are more or less flask-shaped, with bulky protoplasm prolonged into a thin stem directed towards the vitelline membrane, with which, in many instances if not all, it comes in contact. These larger cells are arranged in several tiers. Intercalated between them are a number of elongated small cells with scanty protoplasm and a deeply staining nucleus, not very dissimilar to, though somewhat smaller than, the columnar cells of the previous stage. There is present a complete series of cells intermediate between the larger cells and those with a deeply stained nucleus, and were it not for the condition of the epithelium in Raja, to be spoken of directly, I should not sharply divide the cells into two categories. In surface views of the epithelium the division into two kinds of cells would not be suspected. There can, it appears to me, be no question that both varieties of cell are derived from the primitive uniform follicle cells.

The fibrous layer bounding the membrana propria folliculi is thicker than in the last stage, and the epithelial-like layer (_fe´_) which bounds it externally is more conspicuous than before. Immediately adjoining it are vascular and lymph sinuses. The thickness of the follicular epithelium at this stage may reach as much as 0.04 mm., though I have found it sometimes considerably flatter. The cells composing it are, however, so delicate that it is not easy to feel certain that the peculiarities of any individual ovum are not due to handling. The absence of the peculiar columnar epithelium on the part of the surface adjoining the germinal vesicle is as marked a feature as in the earlier stage. When the egg is nearly ripe, and the vitelline membrane has been reduced to a mere remnant, the follicular epithelium is still very columnar (Pl. 25, fig. 23). The thickness is greater than in the last stage, being now about 0.045 mm., but the cells appear only to form a single definite layer. From the character of their nuclei, I feel inclined to regard them as belonging to the category of the smaller cells of the previous stage, and feel confirmed in this view by finding certain bodies in the epithelium, which have the appearance of degenerating cells with granular nuclei, which I take to be the flask-shaped cells which were present in the earlier stage.

I have not investigated the character of the follicular epithelium in the perfectly ripe ovum ready to become detached from the ovary. Nor can I state for the last-described stage anything about the character of the follicular epithelium in the neighbourhood of the germinal vesicle.

As to the relation of the follicular epithelium to the vitelline membrane, and the possible processes of its cells continued into the yolk, I can say very little. I find in specimens teased out after treatment with osmic acid, that the cells of the follicular epithelium are occasionally provided with short processes, which might possibly have perforated the vitelline membrane, but have met with nothing so clear as the teased out specimens figured by Eimer. Nothing resembling the cells within the vitelline membrane, as described by His[385] in Osseous Fish, and Lindgren in Mammalia, has been met with[386].

Footnote 385: _Das Ei bei Knochenfischen._

Footnote 386: _Arch. f. Anat. Phys._ 1877.

My observations in Raja are not so full as those upon Scyllium, but they serve to complete and reconcile the observations of Semper and Schultz, and also to shew that the general mode of growth of the follicular epithelium is fundamentally the same in my representatives of the two divisions of the Elasmobranchii. In very young eggs, in conformity with the results of all previous observers, I find the follicular epithelium approximately uniform. The cells are flat, but extended so as to appear of an unexpected size in views of the surface of the follicle. This condition does not, however, last very long. A certain number of the cells enlarge considerably, others remaining smaller and flat. The differences between the larger and the smaller cells are more conspicuous in sections than in surface views, and though the distribution of the cells is somewhat irregular, it may still be predicted as an almost invariable rule that the smaller cells of the follicle will line that part of the surface of the ovum, near to which the germinal vesicle is situated. On Pl. 25, fig. 30, is shewn in section a fairly average arrangement of the follicle cells. Semper considers the larger cells of such a follicle to be probably primitive ova destined to become permanent ova. This view I cannot accept: firstly, because these cells only agree with primitive ova in being exceptionally large--the character of their nucleus, with its large nucleolus, being not very like that of a primitive ovum. Secondly, because they shade into ordinary cells of the follicle; and thirdly, because no evidence of their becoming ova has come before me, but rather the reverse, in that it seems probable that they have a definite function connected with the nutrition of the egg. To this point I shall return.

In the next stage the small cells have become still smaller. They are columnar, and are wedged in between the larger ones. No great regularity in distribution is as yet attained (Pl. 25, fig. 31). Such a regularity appears in a later stage (Pl. 25, fig. 32), which clearly corresponds with fig. 8 on Pl. 34 of Schultz's paper, and also with the stage of Scyllium in Pl. 25, fig. 29, though the distinction between the two kinds of cells is here far better marked than in Scyllium. The big cells have now become flask-shaped like those in Scyllium, and send a process down to the vitelline membrane. The smaller cells are arranged in two or three tiers, but the larger cells in a single layer. The distribution of the larger and smaller cells is in some instances very regular, as shewn in the surface view on Pl. 25, fig. 33. There can, it appears to me, be no doubt that Schultz's view of the smaller cells being lymph-cells which have migrated into the follicle cannot be maintained.

The thickness of the epithelium at this stage is about 0.04 mm. In the succeeding stages, during which the egg is rapidly growing to the colossal size which it eventually attains, the follicular epithelium does not to any great extent alter in constitution. It grows thicker on the whole, and as the vitelline membrane gradually atrophies, its lower surface becomes irregular, exhibiting somewhat flattened prominences, which project into the yolk. At the greatest height of the prominences the epithelium may reach a thickness of 0.06 mm., or even more. The arrangement of the tissues external to the follicular epithelium is the same in Raja as in Scyllium.

The most interesting point connected with the follicle, both in Scyllium and Raja and presumably in other Elasmobranchii is that its epithelium at the time when the egg is rapidly approaching maturity is composed with more or less of distinctness of two forms of cells. One of these is large flask-shaped and rich in protoplasm, the other is small, consisting of a mere film of protoplasm round a nucleus. Considering that the larger cells appear at the time of rapid growth, it is natural to interpret their presence as connected with the nutrition of the ovum. This view is supported by the observations of Eimer and Braun, on the development of Reptilian ova. In many Reptilian ova it appears from Eimer's[387] observations, that the follicular epithelium becomes several layers thick, and that a differentiation of the cells, similar to that in Elasmobranchii, takes place. The flask-shaped cells eventually undergo peculiar changes, becoming converted into a kind of beaker-cell, with prolongations through the egg membranes, which take the place of canals leading to the interior of the egg. Braun also expresses himself strongly in favour of the flask-shaped cells functioning in the nutrition of the egg[388]. That these cells in the Reptilian ova really correspond with those in Elasmobranchii appears to me clear from Eimer's figures, but I have not myself studied any Reptilian ovum. My reasons for dissenting from both Semper's and Schultz's views on the nature of the two forms of follicular cells have already been stated.

Footnote 387: _Archiv f. mikr. Anat._ Vol. VIII.

Footnote 388: Braun, "Urogenitalsystem d. Amphibien," _Arbeiten a. d. zool.-zoot. Institut Würzburg_, Bd. IV. He says, in reference to the flask-shaped cell, p. 166, "Höchstens würde ich die Funktion der grossen Follikelzellen als _einzellige Drüsen_ mehr betonen."

_The Vitellus and the development of the yolk spherules._--Leydig, Gegenbaur, and Schultz, have recorded important observations on this head. Leydig[389] chiefly describes the peculiar characters of the yolk spherules.

Footnote 389: _Loc. cit._

Gegenbaur[390] finds in the youngest eggs fine granules, which subsequently develop into vesicles, in the interior of which the solid oval spheres, so characteristic of Elasmobranchii, are developed.

Footnote 390: _Loc. cit._

Schultz describes in the youngest ova of Torpedo the minute yolk spherules arranged in a semilunar form around the eccentric germinal vesicle. In older ova they spread through the whole. He also gives a description of their arrangement in the ripe ovum. Dr Schultz further finds in the body of the ovum peculiar protoplastic striæ, arranged as a series of pyramids, with the bases directed outwards. In the periphery of the ovum a protoplastic network is also present, which is continuous with the above-mentioned pyramidal structures.

My observations do not very greatly extend those of Gegenbaur and Schultz with reference to the development of the yolk, and closely agree with what Gegenbaur has given in the paper above quoted more fully for Aves and Reptilia than for Elasmobranchii.

In very young ova the body of the ovum is simply granular, but when it has reached about 0.5 mm. the granules are seen to be arranged in a kind of network, or sponge-work (Pl. 25, fig. 21), already spoken of in my monograph on Elasmobranch Fishes.

This network becomes more distinct in succeeding stages, especially in chromic acid specimens (Pl. 25, fig. 22), probably in part owing to a granular precipitation of the protoplasm. In the late stages, when the yolk spherules are fully developed, it is difficult to observe this network, but, as has been shewn in my monograph above quoted, it is still present after the commencement of embryonic development. An arrangement of the protoplasmic striæ like that described by Schultz has not come under my notice.

The development of the yolk appears to me to present special difficulties, owing to the fact pointed out by His[391] that the conditions of development vary greatly according to whether the ovary is in a state of repose or of active development. I do not feel satisfied with my results on this subject, but believe there is still much to be made out. Observations on the yolk spherules may be made either in living ova, in ova hardened in osmic acid, or in ova hardened in picric or chromic acids. The two latter reagents, as well as alcohol, are however unfavourable for the purpose of this study, since by their action the yolk spherules appear frequently to be broken up and otherwise altered. This has to some extent occurred in Pl. 25, fig. 21, and the peculiar appearance of the yolk of this ovum is in part due to the action of the reagent. On the whole I have found osmic acid the most suitable reagent for the study of the yolk, since without breaking up the developing spherules, it stains them of a deep black colour. The yolk spherules commence to be formed in ova, of not more than 0.06 mm. in the ovaries of moderately old females. In young females they are apparently not formed in such small ova. They arise as extremely minute, highly refracting particles, in a stratum of protoplasm _some little way below the surface, and are always most numerous at the pole opposite the germinal vesicle_. Their general arrangement is very much that figured and described by Allen Thomson in Gasterosteus[392], and by Gegenbaur and Eimer in young Reptilian ova. In section they naturally appear as a ring, their general mode of distribution being fairly typically represented on Pl. 25, fig. 27. The ovum represented in fig. 27 was 0.5 mm. in diameter, and the yolk spherules were already largely developed; in smaller ova they are far less numerous, though arranged in a similar fashion. The developing yolk spherules are not uniformly distributed but are collected in peculiar little masses or aggregations (Pl. 25, fig. 21). These resemble the granular masses, figured by His (_loc. cit._ Pl. 4, fig. 33) in the Salmon, and may be compared with the aggregations figured by Götte in his monograph on _Bombinator igneus_ (Pl. 1, fig. 9). It deserves to be especially noted, that when the yolk spherules are first formed, the _peripheral layer of the ovum_ is entirely free from them, a feature which is however apt to be lost in ova hardened in picric acid (Pl. 25, fig. 21). Two points about the spherules appear clearly to point to their being developed in the protoplasm of the ovum, and not in the follicular epithelium. (1) That they do not make their appearance in the superficial stratum of the ovum. (2) That no yolk spherules are present in the cells of the follicular epithelium, in which they could not fail to be detected, owing to the deep colour they assume on being treated with osmic acid.

Footnote 391: _Das Ei bei Knochenfischen._

Footnote 392: "Ovum" in Todd's _Encyclopædia_, fig. 69.

It need scarcely be said that the yolk spherules at this stage are not cells, and have indeed no resemblance to cells. They would probably be regarded by His as spherules of fatty material, unrelated to the true food yolk.

As the ova become larger the granules of the peripheral layer before mentioned gradually assume the character of the yolk spheres of the adult, and at the same time spread towards the centre of the egg. Not having worked at fresh specimens, I cannot give a full account of the growth of the spherules; but am of opinion that Gegenbaur's account is probably correct, according to which the spheres at first present gradually grow and develop into vesicles, in the interior of which solid bodies (nuclei of His?) appear and form the permanent yolk spheres. When the yolk spheres are still very small they have the typical oblong form[393] of the ripe ovum, and this form is acquired while the centre of the ovum is still free from them.

Footnote 393: The peculiar oval, or at times slightly rectangular and striated yolk spherules of Elasmobranchii are mentioned by Leydig and Gegenbaur (Pl. 11, fig. 20), and myself, _Preliminary Account of Development of Elasmobranch Fishes_, and by Filippi and His in _Osseous Fishes_.

The growth of the yolk appears mainly due to the increase in size and number of the individual yolk spheres. Even when the ovum is quite filled with large yolk spheres, the granular protoplastic network of the earlier stages is still present, and serves to hold together the constituents of the yolk. In the cortical layer of nearly ripe ova, the yolk has a somewhat different character to that which it exhibits in the deeper layers, chiefly owing to the presence of certain delicate granular (in hardened specimens) bodies, whose nature I do not understand, and to special yolk spheres rather larger than the ordinary, provided with numerous smaller spherules in their interior, which are probably destined in the course of time to become free and to form ordinary yolk spheres.

The mode of formation of the yolk spheres above described appears to me to be the normal, and possibly the only one. Certain peculiar structures have, however, come under my notice, which may perhaps be connected with the formation of the yolk. One of these resembles the bodies described by Eimer[394] as "Dotterschorfe." I have only met these bodies in a single instance in ova of 0.6 mm., from the ovary (in active growth) of a specimen of _Scy. canicula_ 23 inches in length. In this instance they consisted of homogeneous clear bodies (not bounded by any membrane) of somewhat irregular shape, though usually more or less oval, and rarely more than 0.02 mm. in their longest diameter. They were very numerous in the peripheral layer of the ovum, but quite absent in the centre, and also not found outside the ovum (as they appear to be in Reptilia). Yolk granules formed in the normal way, and staining deeply by osmic acid, were present, but the "Dotterschorfe" presented a marked contrast to the remainder of the ovum, in being absolutely unstained by osmic acid, and indeed they appeared more like a modified form of vacuole than any definite body. Their general appearance in Scyllium may be gathered from Eimer's figure 8, Pl. 11, though they were much more numerous than represented in that figure, and confined to the periphery of the ovum.

Footnote 394: "Untersuchung über die Eier d. Reptilien," _Archiv f. mikros. Anat._ Vol. VIII.

Dr Eimer describes a much earlier condition of these structures, in which they form a clear shell enclosing a central dark nucleus. This stage I have not met with, nor can I see any grounds for connecting these bodies with the formation of the yolk, and the fact of their not staining with osmic acid is strongly opposed to this view of their function. Dr Eimer does not appear to me to bring forward any satisfactory proof that they are in any way related to the formation of the yolk, but wishes to connect them with the peculiar body, well known as the yolk nucleus, which is found in the Amphibian ovum[395].

Footnote 395: Vide Allen Thomson, article "Ovum," Todd's _Encyclopædia_, p. 95.

Another peculiar body found in the ova may be mentioned here, though it more probably belongs to the germinal vesicle than to the yolk. It has only been met with in the vitellus of some of the medium sized ova of a young female. Examples of this body are represented on Pl. 25, fig. 25A, _x_. As a rule there is only one in each of the ova in which they are present, but there may be as many as four. They consist of small vesicles with a very thick doubly contoured membrane, which are filled with numerous deeply staining spherical granules. At times they contain a vacuole. Some of the larger of them are not very much smaller than the germinal vesicle of their ovum, while the smallest of them present a striking resemblance to the nucleoli (fig. 25B), which makes me think that they may possibly be nucleoli which have made their way out of the germinal vesicle. I have not found them in the late stages or large ova.

The following measurements shew the size of some of these bodies in relation to the germinal vesicle and ovum:--

Diameter of Germinal Diameter of Body in Diameter of Ovum. Vesicle. Vitellus.

0.096 mm. 0.03 mm. 0.009 mm. 0.064 mm. 0.025 mm. 0.012 mm. {0.019 mm. 0.096 mm. 0.03 mm {0.003 mm.

_Germinal vesicle._--Gegenbaur[396] finds the germinal vesicle completely homogeneous and without the trace of a germinal spot. In Raja granules or vesicles may appear as artificial products, and in Acanthias even in the fresh condition isolated vesicles or masses of such may be present. To these structures he attributes no importance.

Footnote 396: _Loc. cit._

Alexander Schultz[397] states that there is nothing remarkable in the germinal vesicle of the Torpedo egg, but that till the egg reaches 0.5 mm., a single germinal spot is always present (measuring about 0.01 mm.), which is absent in larger ova.

Footnote 397: _Loc. cit._

The bodies described by Gegenbaur are now generally recognised as germinal spots, and will be described as such in the sequel. I have very rarely met with the condition with the single nucleolus described by Schultz in Torpedo.

My own observations are confined to Scyllium. In very young females, with ova not larger than 0.09 mm., the germinal vesicle has the same characters as during the embryonic periods. The contents are clear but traversed by a very distinct and deeply staining reticulum of fibres connected with the several nucleoli which are usually present and situated close to the membrane.

In a somewhat older female in the largest ova of about 0.12 mm., the germinal vesicle measures about 0.06 mm., and usually occupies an eccentric position. It is provided with a distinct though delicate membrane. The network, so conspicuous during the embryonic period, is not so clear as it was, and has the appearance of being formed of lines of granules rather than of fibres. The fluid contents of the nucleus remain as a rule, even in the hardened specimens, perfectly clear, though they become in some instances slightly granular. There are usually two, three, or more nucleoli generally situated, as described by Eimer, close to the membrane of the vesicle, the largest of which may measure as much as 0.006 mm. They are highly refracting bodies, containing in most instances a vacuole, and very frequently a smaller spherical body of a similar nature to themselves[398]. Granules are sometimes also present in the germinal vesicle, but are probably only extremely minute nucleoli.

Footnote 398: Compare, with reference to several points, the germinal vesicle at this stage with the germinal vesicle of the frog's ovum figured by O. Hertwig, _Morphologisches Jahrbuch_, Vol. III. pl. 4, fig. 1.

In ova of 0.5 mm. the germinal vesicle has a diameter of 0.12 mm. (Pl. 25, fig. 21). It is usually shrunk in hardened specimens though nearly spherical in the living ovum. Its contents are rendered granular by reagents though quite clear when fresh, and the reticulum of the earlier stages is sometimes with difficulty to be made out, though in other instances fairly clear. In all cases the fibres composing it are very granular. The membrane is thick. Peculiar highly refracting nucleoli, usually enclosing a large vacuole, are present in considerable numbers, and are either arranged in a circle round the periphery, or sometimes aggregated towards one side of the vesicle; and in addition, numerous deeply staining smaller granular aggregations, probably belonging to the same category as the nucleoli (from which in the living ovum they can only be distinguished by their size), are scattered close to the inner side of the membrane over the whole or only a part of the surface of the germinal vesicle. In a fair number of instances bodies like that figured on Pl. 25, fig. 27, are to be found in the germinal vesicle. They appear to be nucleoli in which a number of smaller nucleoli are originating by a process of endogenous growth, analogous perhaps to endogenous cell-formation. The nucleoli thus formed are, no doubt, destined to become free. The above mode of increase for the nucleoli appears to be exceptional. The ordinary mode is, no doubt, that by simple division into two, as was long ago shewn by Auerbach.

Of the later stages of the germinal vesicle and its final fate, I can give no account beyond the very fragmentary statements which have already appeared in my monograph on Elasmobranch Fishes.

_Formation of fresh ova and ovarian nests in the post-embryonic stages._--Ludwig[399] was the first to describe the formation of ova in the post-embryonic periods. His views will be best explained by quoting the following passage:--

"The follicle of Skates and Dog-fish, with the ovum it contains, is to be considered as an aggregation of the cells of the single-layered ovarian epithelium which have grown into the stroma, and of which one cell has become the ovum and the others the follicular epithelium. The follicle, however, draws in with it into the stroma a number of additional epithelial cells in the form of a stalk connecting the follicle with the superficial epithelium. At a later period the lower part of the stalk at its junction with the follicle becomes continuously narrowed, and at the same time a rupture takes place in the cells which form it. In this manner the follicle becomes at last constricted off from the stalk, and so from its place of origin in the superficial epithelium, and subsequently lies freely in the stroma of the ovary."

Footnote 399: _Loc. cit._

He further explains that the separation of the follicles from the epithelium takes place much earlier in Acanthias than in Raja, and that the sinkings of the epithelium into the stroma may have two or three branches each with a follicle.

Semper gives very little information with reference to the post-embryonic formation of ova. He expresses his agreement on the whole with Ludwig, but, amongst points not mentioned by Ludwig, calls attention to peculiar aggregations of primitive ova in the superficial epithelium, which he regards as either rudimentary testicular follicles or as nests similar to those in the embryo.

My observations on this subject do not agree very closely with those either of Ludwig or Semper. The differences between us partly, though not entirely, depend upon the fundamentally different views we hold about the constitution of the ovary and the nature of the epithelium covering it (vide pp. 555 and 556).

In very young ovaries (Pl. 24, fig. 8) nests of ova (in my sense of the term) are very numerous, but though usually superficial in position are also found in the deeper layers of the ovary. They are especially concentrated in their old position, close to the dorsal edge of the organ. In some instances they do not present quite the same appearance as in the embryo, owing to the outlines of the ova composing them being distinct, and to the presence between the ova of numerous interstitial cells derived from the germinal epithelium, and destined to become follicular epithelium. These latter cells at first form a much flatter follicular epithelium than in the embryonic periods, so that the smaller adult ova have a much less columnar investment than ova of the same size in the embryo. A few primitive ova may still be found in a very superficial position, but occasionally also in the deeper layers. I am inclined to agree with Semper that some of these are freshly formed from the cells of the germinal epithelium.

In the young female with ova of about 0.5 mm. nests of ova are still fairly numerous. The nests are characteristic, and present the various remarkable peculiarities already described in the embryo. In many instances they form polynuclear masses, not divided into separate cells, generally, however, the individual ova are distinct. The ova in these nests are on the average rather smaller than during the embryonic periods. The nests are frequently quite superficial and at times continuous with the pseudo-epithelium, and individual ova also occasionally occupy a position in the superficial epithelium. Some of the appearances presented by separate ova are not unlike the figures of Ludwig, but a growth such as he describes has, according to my observations, no existence. The columns which he believes to have grown into the stroma are merely trabeculæ connecting the deeper and more superficial parts of the germinal epithelium; and his whole view about the formation of the follicular epithelium round separate ova certainly does not apply, except in rare cases, to Scyllium. It is, indeed, very easy to see that most freshly formed ova are derived from nests, as in the embryo; and the formation of a follicular epithelium round these ova takes place as they become separated from the nests. A few solitary ova, which have never formed part of a nest, seem to be formed in this stage as in the embryo; but they do not grow into the stroma surrounded by the cells of the pseudo-epithelium, and only as they reach a not inconsiderable size is a definite follicular epithelium formed around them. The follicular epithelium, though not always formed from the pseudo-epithelium, is of course always composed of cells derived from the germinal epithelium.

In all the ova formed at this stage the nucleus would seem to pass through the same metamorphosis as in the embryo.

In the later stages, and even in the full-grown female of Scyllium, fresh ova seemed to be formed and nests also to be present. In Raja I have not found freshly formed ova or nests in the adult, and have had no opportunity of studying the young forms.

_Summary of observations on the development of the ovary in Scyllium and Raja._

(1) The ovary in the embryo is a ridge, triangular in section, attached along the base. It is formed of a core of stroma and a covering of epithelium. A special thickening of the epithelium on the outer side forms the true germinal epithelium, to which the ova are confined (Pl. 24, fig. 1). In the development of the ovary the stroma becomes differentiated into an external vascular layer, especially developed in the neighbourhood of the germinal epithelium, and an internal lymphatic portion, which forms the main mass of the ovarian ridge (Pl. 24, figs. 2, 3, and 6).

(2) At first the thickened germinal epithelium is sharply separated by a membrane from the subjacent stroma (Pl. 24, figs. 1, 2, and 3), but at about the time when the follicular epithelium commences to be formed round the ova, numerous strands of stroma grow into the epithelium, and form a regular network of vascular channels throughout it, and partially isolate individual ova (Pl. 24, figs. 7 and 8). At the same time the surface of the epithelium turned towards the stroma becomes irregular (Pl. 24, fig. 9), owing to the development of individual ova. In still later stages the stroma ingrowths form a more or less definite tunic close to the surface of the ovary. External to this tunic is the superficial layer of the germinal epithelium, which forms what has been spoken of as the pseudo-epithelium. In many instances the protoplasm of its cells is produced into peculiar fibrous tails which pass into the tunic below.

(3) _Primitive ova._--Certain cells in the epithelium lining the dorsal angle of the body-cavity become distinguished as primitive ova by their abundant protoplasm and granular nuclei, at a very early period in development, even before the formation of the genital ridges. Subsequently on the formation of the genital ridges these ova become confined to the thickened germinal epithelium on the outer aspect of the ridges (Pl. 24, fig. 1).

(4) _Conversion of primitive ova into permanent ova._--Primitive ova may in Scyllium become transformed into permanent ova in two ways--the difference between the two ways being, however, of secondary importance.

(_a_) A nest of primitive ova makes its appearance, either by continued division of a single primitive ovum or otherwise. The bodies of all the ova of the nest fuse together, and a polynuclear mass is formed, which increases in size concomitantly with the division of its nuclei. The nuclei, moreover, pass through a series of transformations. They increase in size and form delicate vesicles filled with a clear fluid, but contain close to one side a granular mass which stains very deeply with colouring reagents. The granular mass becomes somewhat stellate, and finally assumes a reticulate form with one more highly refracting nucleoli at the nodal points of the reticulum. When a nucleus has reached this condition the protoplasm around it has become slightly granular, and with the enclosed nucleus is segmented off from the nest as a special cell--a permanent ovum (figs. 13, 14, 15, 16). Not all the nuclei in a nest undergo the whole of the above changes; certain of them, on the contrary, stop short in their development, atrophy, and become employed as a kind of pabulum for the remainder. Thus it happens that out of a large nest perhaps only two or three permanent ova become developed.

(_b_) In the second mode of development of ova the nuclei and protoplasm undergo the same changes as in the first mode; but the ova either remain isolated and never form part of a nest, or form part of a nest in which no fusion of the protoplasm takes place, and all the primitive ova develop into permanent ova. Both the above modes of the formation continue through a great part of life.

(5) _The follicle._--The cells of the germinal epithelium arrange themselves as a layer around each ovum, almost immediately after its separation from a nest, and so constitute a follicle. They are at first flat, but soon become more columnar. In Scyllium they remain for a long time uniform, but in large eggs they become arranged in two or three layers, while at the same time some of them become large and flask-shaped, and others small and oval (fig. 29). The flask-shaped cells have probably an important function in the nutrition of the egg, and are arranged in a fairly regular order amongst the smaller cells. Before the egg is quite ripe both kinds of follicle cells undergo retrogressive changes (Pl. 25, fig. 23).

In Raja a great irregularity in the follicle cells is observable at an early stage, but as the ovum grows larger the cells gradually assume a regular arrangement more or less similar to that in Scyllium (Pl. 25, figs. 30-33).

(6) _The egg membranes._--Two membranes are probably always present in Elasmobranchii during some period of their growth. The first formed and outer of these arises in some instances before the formation of the follicular epithelium, and would seem to be of the nature of a vitelline membrane. The inner one is the zona radiata with a typical radiately striated structure. It is formed from the vitellus at a much later period than the proper vitelline membrane. It is more developed in Scyllium than in Raja, but atrophies early in both genera. By the time the ovum is nearly ripe both membranes are very much reduced, and when the egg (in Scyllium and Pristiurus) is laid, no trace of any membrane is visible.

(7) _The vitellus._--The vitellus is at first faintly granular, but at a later period exhibits a very distinct (protoplasmic) network of fibres, which is still present after the ovum has been laid.

The yolk arises, in the manner described by Gegenbaur, in ova of about 0.06 mm. as a layer of fine granules, which stain deeply with osmic acid. They are at first confined to a stratum of protoplasm slightly below the surface of the ovum, and are most numerous at the pole furthest removed from the germinal vesicle. They are not regularly distributed, but are aggregated in small masses. They gradually grow into vesicles, in the interior of which oval solid bodies are developed, which form the permanent yolk-spheres. These oval bodies in the later stages exhibit a remarkable segmentation into plates, which gives them a peculiar appearance of transverse striation.

Certain bodies of unknown function are occasionally met with in the vitellus, of which the most remarkable are those figured at _x_ on Pl. 25, fig. 25A.

(8) _The germinal vesicle._--A reticulum is very conspicuous in the germinal vesicle in the freshly formed ova, but becomes much less so in older ova, and assumes, moreover, a granular appearance. At first one to three nucleoli are present, but they gradually increase in number as the germinal vesicle grows older, and are frequently situated in close proximity to the membrane.

THE MAMMALIAN OVARY (Pl. 26).

The literature of the mammalian ovary has been so often dealt with that it may be passed over with only a few words. The papers which especially call for notice are those of Pflüger[400], Ed. van Beneden[401], and especially Waldeyer[402], as inaugurating the newer view on the nature of the ovary, and development of the ova; and of Foulis[403] and Kölliker[404], as representing the most recent utterances on the subject. There are, of course, many points in these papers which are touched on in the sequel, but I may more especially here call attention to the fact that I have been able to confirm van Beneden's statement as to the existence of polynuclear protoplasmic masses. I have found them, however, by no means universal or primitive; and I cannot agree in a general way with van Beneden's account of their occurrence. I have found no trace of a germogene (Keimfache) in the sense of Pflüger and Ed. van Beneden. My own results are most in accordance with those of Waldeyer, with whom I agree in the fundamental propositions that both ovum and follicular epithelium are derived from the germinal epithelium, but I cannot accept his views of the relation of the stroma to the germinal epithelium.

Footnote 400: _Die Eierstöcke d. Säugethiere u. d. Menschen_, Leipzig, 1863.

Footnote 401: "Composition et Signification de l'oeuf," _Acad. r. de Belgique_, 1868.

Footnote 402: _Eierstock u. Ei._ Leipzig, 1870.

Footnote 403: _Trans. of Royal Society, Edinburgh_, Vol. XXVII. 1875, and _Quarterly Journal of Microscopical Science_, Vol. XVI.

Footnote 404: _Verhandlung d. Phys. Med. Gesellschaft_, Würzburg, 1875, N. F. Bd. VIII.

In the very interesting paper of Foulis, the conclusion is arrived at, that while the ova are derived from the germinal epithelium, the cells of the follicle originate from the ordinary connective tissue cells of the stroma. Foulis regards the zona pellucida as a product of the ovum and not of the follicle. To both of these views I shall return, and hope to be able to shew that Foulis has not traced back the formation of the follicle through a sufficient number of the earlier stages. It thus comes about that though I fully recognise the accuracy of his figures, I am unable to admit his conclusions. Kölliker's statements are again very different from those of Foulis. He finds certain cords of cells in the hilus of the ovary, which he believes to be derived from the Wolffian body, and has satisfied himself that they are continuous with Pflüger's egg-tubes, and that they supply the follicular epithelium. To the general accuracy of Kölliker's statements with reference to the relations of these cords in the hilus of the ovary I can fully testify, but am of opinion that he is entirely mistaken as to their giving rise to the follicular epithelium, or having anything to do with the ova. I hope to be able to give a fuller account of their origin than he or other observers have done.

My investigations on the mammalian ovary have been made almost entirely on the rabbit--the type of which it is most easy to procure a continuous series of successive stages; but in a general way my conclusions have been controlled and confirmed by observations on the cat, the dog, and the sheep. My observations commence with an embryo of eighteen days. A transverse section, slightly magnified, through the ovary at this stage, is represented on Pl. 26, fig. 35, and a more highly magnified portion of the same in fig. 35A. The ovary is a cylindrical ridge on the inner side of the Wolffian body, composed of a superficial epithelium, the germinal epithelium (_g.e._), and of a tissue internal to this, which forms the main mass of it. In the latter two constituents have to be distinguished--(1) an epithelial-like tissue (_t_), coloured brown, which forms the most important element, and (2) vascular and stroma elements in this.

The germinal epithelium is a layer about 0.03 - 0.04 mm. in thickness. It is (vide fig. 35A, _g.e._) composed of two or three layers of cells, with granular nuclei, of which the outermost layer is more columnar than the remainder, and has elongated rather than rounded nuclei. Its cells, though they vary slightly in size, are all provided with a fair amount of protoplasm, and cannot be divided (as in the case of the germinal epithelium of Birds, Elasmobranchii, &c.), into primitive ova, and normal epithelial cells. Very occasionally, however, a specially large cell, which, perhaps, deserves the appellation primitive ovum, may be seen. From the subjacent tissue the germinal epithelium is in most parts separated by a membrane-like structure (fluid coagulum); but this is sometimes absent, and it is then very difficult to determine with exactness the inner border of the epithelium. The tissue (_t_), which forms the greater mass of the ovary at this stage, is formed of solid columns or trabeculæ of epithelial-like cells, which present a very striking resemblance in size and character to the cells of the germinal epithelium. The protoplasm of these cells stains slightly more deeply with osmic acid than does that of the cells of the germinal epithelium, so that it is rather easier to note a difference between the two tissues in osmic acid than in picric acid specimens. This tissue approaches very closely, and is in many parts in actual contact with the germinal epithelium. Between the columns of it are numerous vascular channels (shewn diagrammatically in my figures) and a few normal stroma cells. This remarkable tissue continues visible through the whole course of the development of the ovary, till comparatively late in life, and during all the earlier stages might easily be supposed to be about to play some part in the development of the ova, or even to be part of the germinal epithelium. It really, however, has nothing to do with the development of the ova, as is easily demonstrated when the true ova begin to be formed. In the later stages, as will be mentioned in the description of those stages, it is separated from the germinal epithelium by a layer of stroma; though at the two sides of the ovary it is, even in later stages, sometimes in contact with the germinal epithelium.

In most parts this tissue is definitely confined within the limits of the ovary, and does not extend into the mesentery by which the ovary is attached. It may, however, be traced _at the anterior end_ of the ovary into connection with the walls of the Malpighian bodies, which lie on the inner side of the Wolffian body (vide fig. 35B), and I have no doubt that it grows out from the walls of these bodies into the ovary. In the male it appears to me to assist in forming, together with cells derived from the germinal epithelium, the seminiferous tubules, the development of which is already fairly advanced by this stage. I shall speak of it in the sequel as tubuliferous tissue. The points of interest in connection with it concern the male sex, which I hope to deal with in a future paper, but I have no hesitation in identifying it with the segmental cords (_segmentalstränge_) discovered by Braun in Reptilia, and described at length in his valuable memoir on their urogenital system[405]. According to Braun the segmental cords in Reptilia are buds from the outer walls of the Malpighian bodies. The bud from each Malpighian body grows into the genital ridge before the period of sexual differentiation, and sends out processes backwards and forwards, which unite with the buds from the other Malpighian bodies. There is thus formed a kind of trabecular work of tissue in the stroma of the ovary, which in the Lacertilia comes into connection with the germinal epithelium in both sexes, but in Ophidia in the male only. In the female, in all cases, it gradually atrophies and finally vanishes, but in the male there pass into it the primitive ova, and it eventually forms, with the enclosed primitive ova, the tubuli seminiferi. From my own observations in Reptilia I can fully confirm Braun's statements as to the entrance of the primitive ova into this tissue in the male, and the conversion of it into the tubuli seminiferi. The chief difference between Reptilia and Mammalia, in reference to this tissue, appears to be that in Mammalia it arises only from a few of the Malpighian bodies at the anterior extremity of the ovary, but in Reptilia from all the Malpighian bodies adjoining the genital ridge. More extended observations on Mammalia will perhaps shew that even this difference does not hold good.

Footnote 405: _Arbeiten a. d. Zool.-zoot. Institut Würzburg_, Bd. IV.

It is hardly to be supposed that this tissue, which is so conspicuous in all young ovaries, has not been noticed before; but the notices of it are not so numerous as I should have anticipated. His[406] states that the parenchyma of the sexual glands undoubtedly arises from the Wolffian canals, and adds that while the cortical layer (Hulle) represents the earlier covering of a part of the Wolffian body, the stroma of the hilus, with its vessels, arises from a Malpighian body. In spite of these statements of His, I still doubt very much whether he has really observed either the tissue I allude to or its mode of development. In any case he gives no recognisable description or figure of it.

Footnote 406: _Archiv f. mikros. Anat._ Vol. I. p. 160.

Waldeyer[407] notices this tissue in the dog, cat, and calf. The following is a free translation of what he says, (p. 141):--"In a full grown but young dog, with numerous ripe follicles, there were present in the vascular zone of the ovary numerous branched elongated small columns (Schläuche) of epithelial cells, between which ran blood-vessels. They were only separated from the egg columns of the cortical layer by a row of large follicles. There can be no doubt that we have here remains of the sexual part of the Wolffian body--the canals of the parovarium--which in the female sex have developed themselves to an extraordinary extent into the stroma of the sexual gland, and perhaps are even to be regarded as _homologues of the seminiferous tubules_ (the italics are my own). I have almost always found the above condition in the dog, only in old animals these seminiferous canals seem gradually to atrophy. Similar columns are present in the cat, only they do not appear to grow so far into the stroma." Identical structures are also described in the calf.

Footnote 407: _Loc. cit._

Romiti gives a very similar description to Waldeyer of these bodies in the dog[408]. Born also describes this tissue in young and embryonic ovaries of the horse as the _Keimlager_[409]. The columns described by Kölliker[410] and believed by him to furnish the follicular epithelium, are undoubtedly my tubuliferous tissue, and, as Kölliker himself points out, are formed of the same tissue as that described by Waldeyer.

Footnote 408: _Archiv f. mikr. Anat._ Vol. X.

Footnote 409: _Archiv f. Anatomie, Physiologie, u. Wiss. Medicin._ 1874.

Footnote 410: _Loc. cit._

Egli gives a very clear and accurate description of this tissue, though he apparently denies its relation with the Wolffian body.

My own interpretation of the tissue accords with that of Waldeyer. In addition to the rabbit, I have observed it in the dog, cat, and sheep. In all these forms I find that close to the attachment of the ovary, and sometimes well within it, a fair number of distinct canals with a large lumen are present, which are probably to be distinguished from the solid epithelial columns. Such large canals are not as a rule present in the rabbit. In the dog solid columns are present in the embryo, but later they appear frequently to acquire a tubular form, and a lumen. Probably there are great variations in the development of the tissue, since in the cat (not as Waldeyer did in the dog) I have found it most developed.

In the very young embryonic ovary of the cat the columns are very small and much branched. In later embryonic stages they are frequently elongated, sometimes convoluted, and are very similar to the embryonic tubuli seminiferi. In the young stages these columns are so similar to the egg tubes (which agree more closely with Pflüger's type in the cat than in other forms I have worked at) that to any one who had not studied the development of the tissue an embryo cat's ovary at certain stages would be a very puzzling object. I have, however, met with nothing in the cat or any other form which supports Kölliker's views.

My next stage is that of a twenty-two days' embryo. Of this stage I have given two figures corresponding to those of the earlier stage (figs. 36 and 36A).

From these figures it is at once obvious that the germinal epithelium has very much increased in bulk. It has a thickness 0.1 - 0.09 mm. as compared to 0.03 mm. in the earlier stage. Its inner outline is somewhat irregular, and it is imperfectly divided into lobes, which form the commencement of structures nearly equivalent to the nests of the Elasmobranch ovary. The lobes _are not_ separated from each other by connective tissue prolongations; the epithelium being at this stage perfectly free from any ingrowths of stroma. The cells constituting the germinal epithelium have much the same character as in the previous stage. They form an outer row of columnar cells internal to which the cells are more rounded. Amongst them a few large cells with granular nuclei, which are clearly primitive ova, may now be seen, but by far the majority of the cells are fairly uniform in size, and measure from 0.01 - 0.02 mm. in diameter, and their nuclei from 0.004 - 0.006 mm. The nuclei of the columnar outer cells measure about 0.008 mm. They are what would ordinarily be called granular, though high powers shew that they have the usual nuclear network. There is no special nucleolus. The rapid growth of the germinal epithelium is due to the division of its cells, and great masses of these may frequently be seen to be undergoing division at the same time. Of the tissue of the ovary internal to the germinal epithelium, it may be noticed that the tubuliferous tissue derived from the Malpighian bodies is no longer in contact with the germinal epithelium, but that a layer of vascular stroma is to a great extent interposed between the two. The vascular stroma of the hilus has, moreover, greatly increased in quantity.

My next stage is that of a twenty-six days' embryo, but the characters of the ovary at this stage so closely correspond with those of the succeeding one at twenty-eight days that, for the sake of brevity, I pass over this stage in silence.

Figs. 37 and 37A are representative sections of the ovary of the twenty-eighth day corresponding with those of the earlier stages.

Great changes have become apparent in the constitution of the germinal epithelium. The vascular stroma of the ovary has grown into the germinal epithelium precisely as in Elasmobranchii. It appears to me clear that the change in the relations between the stroma and epithelium is not due to a mutual growth, but entirely to the stroma, so that, as in the case of Elasmobranchii, the result of the ingrowth is that the germinal epithelium is honeycombed by vascular stroma. The vascular growths generally take the paths of the lines which separated the nests in an earlier condition, and cause these nests to become the egg tubes of Pflüger. It is obvious in figure 37 that the vascular ingrowths are so arranged as imperfectly to divide the germinal epithelium into two layers separated by a space with connective tissue and blood-vessels. The outer part is relatively thin, and formed of a superficial row of columnar cells, and one or two rows of more rounded cells; the inner layer is much thicker, and formed of large masses of rounded cells. The two layers are connected together by numerous trabeculæ, the stroma between which eventually gives rise to the connective tissue capsule, or tunica albuginea, of the adult ovary.

The germinal epithelium is now about 0.19 to 0.22 mm. in thickness. Its cells have undergone considerable changes. A fair number of them (fig. 37A, _p.o._), especially in the outer layer of the epithelium, have become larger than the cells around them, from which they are distinguished, not only by their size, but by their granular nucleus and abundant protoplasm. They are in fact undoubted primitive ova with all the characters which primitive ova present in Elasmobranchii, Aves, &c. In a fairly typical primitive ovum of this stage the body measures 0.02 mm. and the nucleus 0.014 mm. In the inner part of the germinal epithelium there are very few or no cells which can be distinguished by their size as primitive ova, and the cells themselves are of a fairly uniform size, though in this respect there is perhaps a greater variation than might be gathered from fig. 37A. The cells are on the average about 0.016 mm. in diameter, and their nuclei about 0.008 to 0.001 mm., considerably larger, in fact, than in the earlier stage. The nuclei are moreover more granular, and make in this respect an approach to the character of the nuclei of primitive ova.

The germinal epithelium is still rapidly increasing by the division of its cells, and in fig. 37A there are shewn two or three nuclei in the act of dividing. I have represented fairly accurately the appearance they present when examined with a moderately high magnifying power. With reference to the stroma of the ovary, internal to the germinal epithelium, it is only necessary to refer to fig. 37 to observe that the tubuliferous tissue (_t_) forms a relatively smaller part of the stroma than in the previous stage, and is also further removed from the germinal epithelium.

My next stage is that of a young rabbit two days after birth, but to economise space I pass on at once to the following stage five days after birth. This stage is in many respects a critical one for the ovary, and therefore of great interest. Figure 38 represents a transverse section through the ovary (on rather a smaller scale than the previous figures) and shews the general relations of the tissues.

The germinal epithelium is very much thicker than before--about 0.38 mm. as compared with 0.22 mm. It is divided into three obvious layers: (1) an outer epithelial layer which corresponds with the pseudo-epithelial layer of the Elasmobranch ovary, average thickness 0.03 mm. (2) A middle layer of small nests, which corresponds with the middle vascular layer of the previous stage; average thickness 0.1 mm. (3) An inner layer of larger nests; average thickness 0.23 mm.

The general appearance of the germinal epithelium at this stage certainly appears to me to lend support to my view that the whole of it simply constitutes a thickened epithelium interpenetrated with ingrowths of stroma.

The cells of the germinal epithelium, which form the various layers, have undergone important modifications. In the first place a large number of the nuclei--at any rate of those cells which are about to become ova--have undergone a change identical with that which takes place in the conversion of the primitive into the permanent ova in Elasmobranchii. The greater part of the contents of the nucleus becomes clear. The remaining contents arrange themselves as a deeply staining granular mass on one side of the membrane, and later on as a somewhat stellate figure: the two stages forming what were spoken of as the granular and stellate varieties of nucleus. To avoid further circumlocution I shall speak of the nucleus undergoing the granular and the stellate modifications. At a still later period the granular contents form a beautiful network in the nucleus.

The pseudo-epithelium (fig. 38A) is formed of several tiers of cells, the outermost of which are very columnar and have less protoplasm than in an earlier stage. In the lower tiers of cells there are many primitive ova with granular nuclei, and others in which the nuclei have undergone the granular modification. The primitive ova are almost all of the same size as in the earlier stage. The pseudo-epithelium is separated from the middle layer by a more or less complete stratum of connective tissue, which, however, is traversed by trabeculæ connecting the two layers of the epithelium. In the middle layer there are comparatively few modified nuclei, and the cells still retain for the most part their earlier characters. The diameter of the cells is about 0.012 mm., and that of the nucleus about 0.008 mm. In the innermost layer (fig. 38B), which is not sharply separated from the middle layer, the majority of the cells, which in the previous stage were ordinary cells of the epithelium, have commenced to acquire modified nuclei. This change, which first became apparent to a small extent in the young two days after birth, is very conspicuous at this stage. In some of the cells the nucleus is modified in the granular manner, in others in the stellate, and in a certain number the nucleus has assumed a reticular structure characteristic of the young permanent ovum.

In addition, however, to the cells which are becoming converted into ova, a not inconsiderable number may be observed, if carefully looked for, which are for the most part smaller than the others, generally somewhat oval, and in which the nucleus retains its primitive characters. A fair number of such cells are represented in fig. 38B. In the larger ones the nucleus will perhaps eventually become modified; but the smaller cells clearly correspond with the interstitial cells of the Elasmobranch germinal epithelium, and are destined to become converted into the epithelium of the Graafian follicle. In some few instances indeed (at this stage very few), in the deeper part of the germinal epithelium, these cells commence to arrange themselves round the just formed permanent ova as a follicular epithelium. An instance of this kind is shewn in fig. 38B, _o_. The cells with modified nuclei, which are becoming permanent ova, usually present one point of contrast to the homologous cells in Elasmobranchii, in that they are quite distinct from each other, and not fused into a polynuclear mass. They have around them a dark contour line, which I can only interpret as the commencement of the membrane (zona radiata?), which afterwards becomes distinct, and which would thus seem, as Foulis has already insisted, to be of the nature of a vitelline membrane.

In a certain number of instances the protoplasm of the cells which are becoming permanent ova appears, however, actually to fuse, and polynuclear masses identical with those in Elasmobranchii are thus formed (cf. E. van Beneden[411]). These masses become slightly more numerous in the succeeding stages. Indications of a fusion of this kind are shewn in fig. 38B. That the polynuclear masses really arise from a fusion of primitively distinct cells is clear from the description of the previous stages. The ova in the deeper layers, with modified granular nuclei, measure about 0.016 - 0.02 mm., and their nuclei from 0.01 - 0.012 mm.

Footnote 411: _Loc. cit._

With reference to the tissue of the hilus of the ovary, it may be noticed that the tubuliferous tissue (_t_) is relatively reduced in quantity. Its cells retain precisely their previous characters.

The chief difference between the stage of five days and that of two days after birth consists in the fact that during the earlier stage comparatively few modified nuclei were present, but the nuclei then presented the character of the nuclei of primitive ova.

I have ovaries both of the dog and cat of an equivalent stage, and in both of these the cells of the nests or egg tubes may be divided into two categories, destined respectively to become ova and follicle cells. Nothing which has come under my notice tends to shew that the tubuliferous tissue is in any way concerned in supplying the latter form of cell.

In a stage, seven days after birth, the same layers in the germinal epithelium may be noticed as in the last described stage. The outermost layer or pseudo-epithelium contains numerous developing ova, for the most part with modified nuclei. It is separated by a well marked layer of connective tissue from the middle layer of the germinal epithelium. The outer part of the middle layer contains more connective tissue and smaller nests than in the earlier stage, and most of the cells of this layer contain modified nuclei. In a few nests the protoplasm of the developing ova forms a continuous mass, not divided into distinct cells, but in the majority of instances the outline of each ovum can be distinctly traced. In addition to the cells destined to become ova, there are present in these nests other cells, which will clearly form the follicular epithelium. A typical nest from the middle layer is represented on Pl. 26, fig. 39A.

The nests or masses of ova in the innermost layer are for the most part still very large, but, in addition to the nests, a few isolated ova, enclosed in follicles, are to be seen.

A fairly typical nest, selected to shew the formation of the follicle, is represented on Pl. 26, fig. 39B.

The nest contains (1) fully formed permanent ova, completely or wholly enclosed in a follicle. (2) Smaller ova, not enclosed in a follicle. (3) Smallish cells with modified nuclei of doubtful destination. (4) Small cells obviously about to form follicular epithelium.

The inspection of a single such nest is to my mind a satisfactory proof that the follicular epithelium takes its origin from the germinal epithelium and not from the stroma or tubuliferous tissue. The several categories of elements observable in such a nest deserve a careful description.

(1) _The large ova in their follicles._--These ova have precisely the character of the young ova in Elasmobranchii. They are provided with a granular body invested by a delicate, though distinct membrane. Their nucleus is large and clear, but traversed by the network so fully described for Elasmobranchii. The cells of their follicular epithelium have obviously the same character as many other small cells of the nest. Two points about them deserve notice--(_a_) that many of them are fairly columnar. This is characteristic only of the first formed follicles. In the later formed follicles the cells are always flat and spindle-shaped in section. In this difference between the early and late formed follicles Mammals agree with Elasmobranchii. (_b_) The cells of the follicle are much more columnar towards the inner side than towards the outer. This point also is common to Mammals and Elasmobranchii.

Round the completed follicle a very delicate membrana propria folliculi appears to be present[412].

Footnote 412: _Loc. cit._, Waldeyer, p. 23, denies the existence of this membrane for Mammalia. It certainly is not so conspicuous as in some other types, but appears to me nevertheless to be always present.

The larger ova, with follicular epithelium, measure about 0.04 mm., and their nucleus about 0.02 mm., the smaller ones about 0.022 mm., and their nucleus about 0.014 mm.

(2) _Medium sized ova._--They are still without a trace of a follicular epithelium, and present no special peculiarities.

(3) _The smaller cells with modified nuclei._--I have great doubt as to what is the eventual fate of these cells. There appear to be three possibilities.

(_a_) That they become cells of the follicular epithelium; (_b_) that they develop into ova; (_c_) that they are absorbed as a kind of food by the developing ova. I am inclined to think that some of these cells may have each of the above-mentioned destinations.

(4) _The cells which form the follicle._--The only point to be noticed about these is that they are smaller than the indifferent cells of the germinal epithelium, from which they no doubt originate by division. This fact has already been noticed by Waldeyer.

The isolated follicles at this stage are formed by ingrowths of connective tissue cutting off fully formed follicles from a nest. They only occur at the very innermost border of the germinal epithelium. This is in accordance with what has so often been noticed about the mammalian ovary, viz. that the more advanced ova are to be met with in passing from without inwards.

By the stage seven days after birth the ovary has reached a sufficiently advanced stage to answer the more important question I set myself to solve, nevertheless, partly to reconcile the apparent discrepancy between my account and that of Dr Foulis, and partly to bring my description up to a better known condition of the ovary, I shall make a few remarks about some of the succeeding stages.

In a young rabbit about four weeks old the ovary is a very beautiful object for the study of the nuclei, &c.

The pseudo-epithelium is now formed of a single layer of columnar cells, with comparatively scanty protoplasm. In it there are present a not inconsiderable number of developing ova.

A layer of connective tissue--the albuginea--is now present below the pseudo-epithelium, which contains a few small nests with very young permanent ova. The layer of medium sized nests internal to the albuginea forms a very pretty object in well stained sections, hardened in Kleinenberg's picric acid. The ova in it have all assumed the permanent form, and are provided with beautiful reticulate nuclei, with, as a rule, one more especially developed nucleolus, and smaller granular bodies. Their diameter varies from about 0.028 to 0.04 mm. and that of their nucleus from 0.016 to 0.02 mm. The majority of these ova are not provided with a follicular investment, but amongst them are numerous small cells, clearly derived from the germinal epithelium, which are destined to form the follicle (vide fig. 40Aand B). In a few cases the follicles are completed, and are then formed of very flattened spindle-shaped (in section) cells. In the majority of cases all the ova of each nest are quite distinct, and each provided with a delicate vitelline membrane (fig. 40A) In other instances, which, so far as I can judge, are more common than in the previous stages, the protoplasm of two or more ova is fused together.

Examples of this are represented in Pl. 26, fig. 40A. In some of these the nuclei in the undivided protoplasm are all of about the same size and distinctness, and probably the protoplasm eventually becomes divided up into as many ova as nuclei; in other cases, however, one or two nuclei clearly preponderate over the others, and the smaller nuclei are indistinct and hazy in outline. In these latter cases I have satisfied myself as completely as in the case of Elasmobranchii, that only one or two ova (according to the number of distinct nuclei) will develop out of the polynuclear mass, and that the other nuclei atrophy, and the material of which they were composed serves as the nutriment for the ova which complete their development. This does not, of course, imply that the ova so formed have a value other than that of a single cell, any more than the development of a single embryo out of the many in one egg capsule implies that the embryo so developing is a compound organism.

In the innermost layer of the germinal epithelium the outlines of the original large nests are still visible, but many of the follicles have been cut off by ingrowths of stroma. In the still intact nests the formation of the follicles out of the cells of the germinal epithelium may be followed with great advantage. The cells of the follicle, though less columnar than was the case at an earlier period, are more so than in the case of follicles formed in the succeeding stages. The previous inequality in the cells of the follicles is no longer present.

The tubuliferous tissue in the zona vasculosa appears to me to have rather increased in quantity than the reverse; and is formed of numerous solid columns or oval masses of cells, separated by strands of connective tissue, with typical spindle nuclei.

It is partially intelligible to me how Dr Foulis might from an examination of the stages similar to this, conclude that the follicle cells were derived from the stroma; but even at this stage the position of the cells which will form the follicular epithelium, their passage by a series of gradations into obvious cells of the germinal epithelium and the peculiarities of their nuclei, so different from those of the stroma cells, supply a sufficient series of characters to remove all doubt as to the derivation of the follicle cells. Apart from these more obvious points, an examination of the follicle cells from the surface, and not in section, demonstrates that the general resemblance in shape of follicle cells to the stroma cells is quite delusory. They are in fact flat, circular, or oval, plates not really spindle-shaped, but only apparently so in section. While I thus fundamentally differ from Foulis as to the nature of the follicle cells, I am on this point in complete accordance with Waldeyer, and my own results with reference to the follicle cannot be better stated than in his own words (pp. 43, 44).

At six weeks after birth the ovary of the rabbit corresponds very much more with the stages in the development of the ovary, which Foulis has more especially studied, for the formation of the follicular epithelium, than during the earlier stages. His figure (_Quart. Journ. Mic. Sci._, Vol. XVI., Pl. 17, fig. 6) of the ovary of a seven and a half months' human foetus is about the corresponding age. Different animals vary greatly in respect to the relative development of the ovary. For example, the ovary of a lamb at birth about corresponds with that of a rabbit six weeks after birth. The points which may be noticed about the ovary at this age are first that the surface of the ovary begins to be somewhat folded. The appearances of these folds in section have given rise, as has already been pointed out by Foulis, to the erroneous view that the germinal epithelium (pseudo-epithelium) became involuted in the form of tubular open pits. The folds appear to me to have no connection with the formation of ova, but to be of the same nature as the somewhat similar folds in Elasmobranchii. A follicular epithelium is present around the majority of the ova of the middle layer, and around all those of the inner layer of the germinal epithelium. The nests are, moreover, much more cut up by connective tissue ingrowths than in the previous stages.

The follicle cells of the middle layers are very flat, and spindle-shaped in section, and though they stain more deeply than the stroma cells, and have other not easily characterised peculiarities, they nevertheless do undoubtedly closely resemble the stroma cells when viewed (as is ordinarily the case) in optical section.

In the innermost layer many of the follicles with the enclosed ova have advanced considerably in development and are formed of columnar cells. The somewhat heterodox view of these cells propounded by Foulis I cannot quite agree to. He says (_Quart. J. Mic. Sci._, Vol. XVI., p. 210): "The protoplasm which surrounds the vesicular nuclei acts as a sort of cement substance, holding them together in the form of a capsular membrane round the young ovum. This capsular membrane is the first appearance of the membrana granulosa." I must admit that I find nothing similar to this, nor have I met with any special peculiarities (as Foulis would seem to indicate) in the cells of the germinal epithelium or other cells of the ovary.

Figure 41 is a representation of an advanced follicle of a six weeks' rabbit, containing two ova, which is obviously in the act of dividing into two. Follicles of this kind with more than one ovum are not very uncommon. It appears to me probable that follicles, such as that I have figured, were originally formed of a single mass of protoplasm with two nuclei; but that instead of one of the nuclei atrophying, both of them eventually developed and the protoplasm subsequently divided into two masses. In other cases it is quite possible that follicles with two ova should rather be regarded as two follicles not separated by a septum of stroma.

On the later stages of development of the ovary I have no complete series of observations. The yolk spherules I find to be first developed in a peripheral layer of the vitellus. I have not been able definitely to decide the relation of the zona radiata to the first formed vitelline membrane. Externally to the zona radiata there may generally be observed a somewhat granular structure, against which the follicle cells abut, and I cannot agree with Waldeyer (_loc cit._, p. 40) that this structure is continuous with the cells of the discus, or with the zona radiata. Is it the remains of the first formed vitelline membrane? I have obtained some evidence in favour of this view, but have not been successful in making observations to satisfy me on the point, and must leave open the question whether my vitelline membrane becomes the zona radiata or whether the zona is not a later and independent formation, but am inclined myself to adopt the latter view. The first formed membrane, whether or no it becomes the zona radiata, is very similar to the vitelline membrane of Elasmobranchii and arises at a corresponding stage.

_Summary of observations on the mammalian ovary._--The general results of my observations on the mammalian ovary are the following:--

(1) The ovary in an eighteen days' embryo consists of a cylindrical ridge attached along the inner side of the Wolffian body, which is formed of two parts; (_a_) an external epithelium--two or three cells deep (the germinal epithelium); (_b_) a hilus or part forming in the adult the vascular zone, at this stage composed of branched masses of epithelial tissue (tubuliferous tissue) derived from the walls of the anterior Malpighian bodies, and numerous blood-vessels, and some stroma cells.

(2) The germinal epithelium gradually becomes thicker, and after a certain stage (twenty-three days) there grow into it numerous stroma ingrowths, accompanied by blood-vessels. The germinal epithelium thus becomes honeycombed by strands of stroma. Part of the stroma eventually forms a layer close below the surface, which becomes in the adult the tunica albuginea. The part of the germinal epithelium external to this layer becomes reduced to a single row of cells, and forms what has been spoken of in this paper as the pseudo-epithelium of the ovary. The greater part of the germinal epithelium is situated internal to the tunica albuginea, and this part is at first divided up by strands of stroma into smaller divisions externally, and larger ones internally. These masses of germinal epithelium (probably sections of branched trabeculæ) may be spoken of as nests. In the course of the development of the ova they are broken up by stroma ingrowths, and each follicle with its enclosed ovum is eventually isolated by a layer of stroma.

(3) The cells of the germinal epithelium give rise both to the permanent ova and to the cells of the follicular epithelium. For a long time, however, the cells remain indifferent, so that the stages, like those in Elasmobranchii, Osseous Fish, Birds, Reptiles, &c., with numerous primitive ova embedded amongst the small cells of the germinal epithelium, are not found.

(4) The conversion of the cells of the germinal epithelium into permanent ova commences in an embryo of about twenty-two days. All the cells of the germinal epithelium appear to be capable of becoming ova: the following are the stages in the process, which are almost identical with those in Elasmobranchii:--

(_a_) The nucleus of the cells loses its more or less distinct network, and becomes very granular, with a few specially large granules (nucleoli). The protoplasm around it becomes clear and abundant--primitive ovum stage. It may be noted that the largest primitive ova are very often situated in the pseudo-epithelium. (_b_) A segregation takes place in the contents of the nucleus within the membrane, and the granular contents pass to one side, where they form an irregular mass, while the remaining space within the membrane is perfectly clear. The granular mass gradually develops itself into a beautiful reticulum, with two or three highly refracting nucleoli, one of which eventually becomes the largest and forms the germinal spot _par excellence_. At the same time the body of the ovum becomes slightly granular. While the above changes, more especially those in the nucleus, have been taking place, the protoplasm of two or more ova may fuse together, and polynuclear masses be so formed. In some cases the whole of such a polynuclear mass gives rise to only a single ovum, owing to the atrophy of all the nuclei but one, in others it gives rise by subsequent division to two or more ova, each with a single germinal vesicle.

(5) All the cells of a nest do not undergo the above changes, but some of them become smaller (by division) than the indifferent cells of the germinal epithelium, arrange themselves round the ova, and form the follicular epithelium.

(6) The first membrane formed round the ovum arises in some cases even before the appearance of the follicular epithelium, and is of the nature of a vitelline membrane. It seems probable, although not definitely established by observation, that the zona radiata is formed internally to the vitelline membrane, and that the latter remains as a membrane, somewhat irregular on its outer border, against which the ends of the follicle cells abut.

GENERAL OBSERVATIONS ON THE STRUCTURE AND DEVELOPMENT OF THE OVARY.

In selecting Mammalia and Elasmobranchii as my two types for investigation, I had in view the consideration that what held good for such dissimilar forms might probably be accepted as true for all Vertebrata with the exception of Amphioxus.

_The structure of the ovary._--From my study of these two types, I have been led to a view of the structure of the ovary, which differs to a not inconsiderable extent from that usually entertained. For both types the conclusion has been arrived at that the whole egg-containing part of the ovary is really _the thickened germinal epithelium_, and that it differs from the original thickened patch or layer of germinal epithelium, mainly in the fact that it is broken up into a kind of meshwork by growths of vascular stroma. If the above view be accepted for Elasmobranchii and Mammalia, it will hardly be disputed for the ovaries of Reptilia and Aves. In the case also of Osseous Fish and Amphibia, this view of the ovary appears to be very tenable, but the central core of stroma present in the other types is nearly or quite absent, and the ovary is entirely formed of the germinal epithelium with the usual strands of vascular stroma[413]. It is obvious that according to the above view Pflüger's egg-tubes are merely trabeculæ of germinal epithelium, and have no such importance as has been attributed to them. They are present in a more or less modified form in all types of ovaries. Even in the adult Amphibian ovary, columns of cells of the germinal epithelium, some indifferent, others already converted into ova, are present, and, as has been pointed out by Hertwig[414], represent Pflüger's egg-tubes.

Footnote 413: My view of the structure of the ovary would seem to be that held by Götte, _Entwicklungsgeschichte d. Unke_, pp. 14 and 15.

Footnote 414: _Loc. cit._ 36.

_The formation of the permanent ova._--The passage of primitive ova into permanent ova is the part of my investigation to which the greatest attention was paid, and the results arrived at for Mammalia and Elasmobranchii are almost identical. Although there are no investigations as to the changes undergone by the nucleus in other types, still it appears to me safe to conclude that the results arrived at hold good for Vertebrates generally[415]. As has already been pointed out the transformation which the so-called primitive ova undergo is sufficient to shew that _they are not to be regarded as ova but merely as embryonic sexual cells_. A feature in the transformation, which appears to be fairly constant in Scyllium, and not uncommon in the rabbit, is the fusion of the protoplasm of several ova into a syncytium, the subsequent increase in the number of nuclei in the syncytium, the atrophy and absorption of a portion of the nuclei, and the development of the remainder into the germinal vesicles of ova; the vitellus of each ovum being formed by a portion of the protoplasm of the syncytium.

Footnote 415: Since writing the above I have made out that in the Reptilia the formation of the permanent ova takes place in the same fashion as in Elasmobranchii and Mammalia.

As to the occurrence of similar phenomena in the Vertebrata generally, it has already been pointed out that Ed. van Beneden has described the polynuclear masses in Mammalia, though he does not appear to me to have given a complete account of their history. Götte[416] describes a fusion of primitive ova in Amphibia, but he believes that the nuclei fuse as well as the bodies of the ova, so that one ovum (according to his view no longer a cell) is formed by the fusion of several primitive ova with their nuclei. I have observed nothing which tends to support Götte's view about the fusion of the nuclei, and regard it as very improbable. As regards the interpretation to be placed upon the nests formed of fused primitive ova, Ed. van Beneden maintains that they are to be compared with the upper ends of the egg tubes of Insects, Nematodes, Trematodes, &c. There is no doubt a certain analogy between the two, in that in both cases certain nuclei of a polynuclear mass increase in size, and with the protoplasm around them become segmented off from the remainder of the mass as ova, but the analogy cannot be pressed. The primitive ova, or even the general germinal epithelium, rather than these nests, must be regarded as giving origin to the ova, and the nests should be looked on, in my opinion, as connected more with the nutrition than with the origin of the ova. In favour of this view is the fact that as a rule comparatively few ova are developed from the many nuclei of a nest; while against the comparison with the egg tubes of the Invertebrata it is to be borne in mind that many ova appear to develop independently of the nests.

Footnote 416: _Entwicklungsgeschichte d. Unke._

In support of my view about the nests there may be cited many analogous instances from the Invertebrata. In none of them, however, are the phenomena exactly identical with those in Vertebrata. In the ovary of many Hydrozoa (_e.g. Tubularia mesembryanthemum_), out of a large number of ova which develop up to a certain point, a comparatively very small number survive, and these regularly feed upon the other ova. During this process the boundary between a large ovum and the smaller ova is indistinct: in the outermost layer of a large ovum a number of small ova are embedded, the outlines of the majority of which have become obscure, although they can still be distinguished. Just beyond the edge of a large ovum the small ova have begun to undergo retrogressive changes; while at a little distance from the ovum they are quite normal. An analogous phenomenon has been very fully described by Weismann[417] in the case of Leptodera, the ovary of which consists of a germogene, in which the ova develop in groups of four. Each group of four occupies a separate chamber of the ovary, but in summer only one of the four eggs (the third from the germogene) develops into an ovum, the other three are used as pabulum. In the case of the winter eggs the process is carried still further, in that the contents of the alternate chambers, instead of developing into ova, are entirely converted, by a series of remarkable changes, into nutritive reservoirs. Fundamentally similar occurrences to the above are also well known in Insects. Phenomena of this nature are obviously in no way opposed to the view of the ovum being a single cell.

Footnote 417: _Zeit. für wiss. Zool._ Bd. XXVII.

With reference to the origin of the primitive ova, it appears to me that their mode of development in Mammals proves beyond a doubt that they are modified cells of the germinal epithelium. In Elasmobranchii their very early appearance, and the difficulty of finding transitional forms between them and ordinary cells of the germinal epithelium, caused me at one time to seek (unsuccessfully) for a different origin for them. Any such attempts appear to me, however, out of the question in the case of Mammals.

_The egg membranes._--The homologies of the egg membranes in the Vertebrata are still involved in some obscurity. In Elasmobranchii there are undoubtedly two membranes present. (1) An outer and first formed membrane--the albuminous membrane of Gegenbaur--which, in opposition to previous observers, I have been led to regard as a vitelline membrane. (2) An inner radiately striated membrane, formed as a differentiation of the surface of the yolk at a later period. Both these membranes usually atrophy before the ovum leaves the follicle. In Reptilia[418] precisely the same arrangement is found as in Elasmobranchii, except that as a rule the zona radiata is relatively more important. The vitelline membrane external to this (or as it is usually named the chorion) is, as a rule, thin in Reptilia; but in Crocodilia is thick (Gegenbaur), and approaches the condition found in Scyllium and other Squalidæ. It appears, as in Elasmobranchii, to be formed before the zona radiata. A special internal differentiation of the zona radiata is apparently found (Eimer) in many Reptilia. No satisfactory observations appear to be recorded with reference to the behaviour of the two reptilian membranes as the egg approaches maturity. In Birds[419] the same two membranes are again found. The first formed and outer one is, according to Gegenbaur and E. van Beneden, a vitelline membrane; and from the analogy of Elasmobranchii I feel inclined to accept their view. The inner one is the zona radiata, which disappears comparatively early, leaving the ovum enclosed only by the vitelline membrane, when it leaves the follicle. All the large-yolked vertebrate ova appear then to agree very well with Elasmobranchii in presenting during some period of their development the two membranes above mentioned.

Footnote 418: Gegenbaur, _loc. cit._; Waldeyer, _loc. cit._; Eimer, _loc. cit._; and Ludwig, _loc. cit._

Footnote 419: Gegenbaur, Waldeyer, E. van Beneden, Eimer.

Osseous fish have almost always a zona radiata, which it seems best to assume to be equivalent to that in Elasmobranchii. Internal to this is a thin membrane, the equivalent, according to Eimer, of the membrane found by the same author within the zona in Reptilia. A membrane equivalent to the thick vitelline membrane of Elasmobranchii would seem to be absent in most instances, though a delicate membrane, external to the zona, has not infrequently been described; Eimer more especially asserts that such a membrane exists in the perch within the peculiar mucous covering of the egg of that fish.

In Petromyzon, a zona radiata appears to be present[420], which is divided in the adult into two layers, both of them perforated. The inner of the two perhaps corresponds with the membrane internal to the zona radiata in other types. In Amphibia the single late formed and radiately striated (Waldeyer) membrane would appear to be a zona radiata. If the suggestion on page 605 turns out to be correct the ova of Mammalia possess both a vitelline membrane and zona radiata. E. van Beneden[421] has, moreover, shewn that they are also provided at a certain period with a delicate membrane within the zona.

Footnote 420: Carlberla, _Zeit. f. wiss. Zool._ Bd. XXX.

Footnote 421: _Loc. cit._

_The reticulum of the germinal vesicle._--In the course of description of the ovary it has been necessary for me to enter with some detail into the structure of the nucleus, and I have had occasion to figure and describe a reticulum identical with that recently described by so many observers. The very interesting observations of Dr Klein in the last number of this Journal[422] have induced me to say one or two words in defence of some points in my description of the reticulum. Dr Klein says, on page 323, "I have distinctly seen that when nucleoli are present--the instances are fewer than is generally supposed; they are accumulations of the fibrils of the network." I have no doubt that Klein is correct in asserting that nucleoli are fewer than is generally supposed; and that in many of these instances what are called nucleoli are accumulations, "natural or artificial," of the fibrils of the network; but I cannot accept the universality of the latter statement, which appears to me most certainly not to hold good in the case of ova, in which nucleoli frequently exist in the absence of the network.

Footnote 422: [_Quarterly Journal Microscopical Science_, July 1878.]

Again, I find that at the point of intersection of two or more fibrils there is, as a rule, a distinct thickening of the matter of the fibrils, and that many of the dots seen are not merely, as Dr Klein would maintain, optical sections of fibrils.

It appears to me probable that both the network and the nucleoli are composed of the same material--what Hertwig calls nuclear substance--and if Dr Klein merely wishes to assert this identity in the passage above quoted, I am at one with him.

Although a more or less distinct network is present in most nuclei (I have found it in almost all embryonic nuclei) it is not universally so. In the nuclei of primitive ova I have no doubt that it is absent, though present in the unmodified nuclei of the germinal epithelium; and it is present only in a very modified form in the nuclei of primitive ova undergoing a transformation into permanent ova. The absence of the reticulum does not, of course, mean that the substance capable of forming a reticulum is absent, but merely that it does not assume a particular arrangement.

One of the most interesting points in Klein's paper, as well as in those of Heitzmann and Eimer, is the demonstration of a connection between the reticulum of the nucleus and fibres in the body of the cell. Such a connection I have not found in ova, but may point out that it appears to exist between the sub-germinal nuclei in Elasmobranchii and the protoplasmic network in the yolk in which they lie. This point is called attention to in my _Monograph on Elasmobranch Fishes_, page 39[423], where it is stated that "the network in favourable cases may be observed to be in connection with the nuclei just described. Its meshes are finer in the vicinity of the nuclei, and the fibres in some cases appear almost to start from them." The nuclei in the yolk are knobbed bodies divided by a sponge work of septa into a number of areas each with a nucleolar body.

Footnote 423: [This Edition, p. 252.]

EXPLANATION OF PLATES 24, 25, 26.

PLATE 24.

LIST OF REFERENCE LETTERS.

_dn._. Modified nucleus of primitive ovum. _do._. Permanent ovum in the act of being formed. _dv._. Developing blood-vessels. _dyk._ Developing yolk. _ep._ Non-ovarian epithelium of ovarian ridge. _fe._ Follicular epithelium. _gv._ Germinal vesicle. _lstr._ Lymphatic region of stroma. _nn._ Nests of nuclei of ovarian region. _o._ Permanent ovum. _ovr._ Ovarian portion of ovarian ridge. _po._ Primitive ovum. _pse._ Pseudo-epithelium of ovarian ridge. _str._ Stroma ingrowths into ovarian epithelium. _v._ Blood-vessel. _vstr._ Vascular region of stroma adjoining ovarian ridge. _vt._ Vitelline membrane. _x._ Modified nucleus. _yk._ Yolk. _zn._ Zona radiata.

Fig. 1. Transverse section of the ovarian ridge of an embryo of _Scy. canicula_, belonging to stage P, shewing the ovarian region with thickened epithelium and numerous primitive ova. Zeiss C, ocul. 2. _Picric acid._

Fig. 2. Transverse section of the ovarian ridge of an embryo of _Scyllium canicula_, considerably older than stage Q. Zeiss C, ocul. 2. _Picric acid._ Several nests, some with distinct ova, and others with the ova fused together, are present in the section (_nn_), and several examples of modified nuclei in still distinct ova are also represented. One of these is marked _x_. The stroma of the ovarian ridge is exceptionally scanty.

Fig. 3. Transverse section through part of the ovarian ridge, including the ovarian region of an almost ripe embryo of _Scyllium canicula_. Zeiss C, ocul. 2. _Picric acid._ Nuclear nests (_n.n._), developing ova (_d.o._), and ova (_o._), with completely formed follicular epithelium, are now present. The ovarian region is still well separated from the subjacent stroma, and does not appear to contain any cells except those of the original germinal epithelium.

Fig. 4. Section through ovarian ridge of the same embryo as fig. 3, to illustrate the relation of the stroma (_str._) and ovarian region. Zeiss _a a_, ocul. 2. _Picric acid._

Fig. 5. Section through the ovarian ridge of an embryo of _Scyllium canicula_, 10 cm. long, in which the ovary was slightly less advanced than in fig. 3. To illustrate the relation of the ovarian epithelium to the subjacent vascular stroma. Zeiss A, ocul. 2. _Osmic acid._ _y_ points to a small separated portion of the germinal epithelium.

Fig. 6. Section through the ovarian ridge of an embryo of _Scyllium canicula_, slightly older than fig. 5. To illustrate the relation of the ovarian epithelium to the subjacent vascular stroma. Zeiss A, ocul. 2. _Osmic acid._

Fig. 7. More highly magnified portion of the same ovary as fig. 6. To illustrate the same points. Zeiss C, ocul. 2. _Osmic acid._

Fig. 8. Section through the ovarian region (close to one extremity, where it is very small) from a young female of _Scy. canicula_. Zeiss C, ocul. 2. _Picric acid._ It shews the vascular ingrowths amongst the original epithelial cells of the ovarian region.

Fig. 9. Section through the ovarian region of the same embryo as fig. 8, at its point of maximum development. Zeiss A, ocul. 2. _Picric acid._

Fig. 10. Section through superficial part of the ovary of an embryo, shewing the pseudo-epithelium; the cells of which are provided with tails prolonged into the general tissue of the ovary. At _f.e._ is seen a surface view of the follicular epithelium of an ovum. Zeiss C, ocul. 2. _Picric acid._

Fig. 11. Section through part of an ovary of _Scyllium canicula_ of stage Q, with three primitive ova, the most superficial one containing a modified nucleus.

Fig. 12. Section through part of an ovary of an example of _Scyllium canicula_, 8 cm. long. The section passes through a nest of ova with modified nuclei, in which the outlines of the individual ova are quite distinct. Zeiss E, ocul. 2. _Picric acid._

Fig. 13. Section through part of ovary of the same embryo as in fig. 5. The section passes through a nest of nuclei, with at the least two developing ova, and also through one already formed permanent ovum. Zeiss E, ocul. 2. _Osmic acid._

Figs. 14, 15, 16, 17, 18 [Figs. 17 and 18 are on Pl. 25]. Sections through parts of the ovary of the same embryo as fig. 3, with nests of nuclei and a permanent ova in the act of formation. Fig. 14 is drawn with Zeiss D D, ocul. 2. Figs. 15, 16, 17, with Zeiss E, ocul. 2. _Picric acid._

PLATE 25.

LIST OF REFERENCE LETTERS.

_do._ Permanent ovum in the act of being formed. _dyk._ Developing yolk. _fe._ Follicular epithelium. _fe´._ Secondary follicular epithelium. _gv._ Germinal vesicle. _nn._ Nests of nuclei of ovarian region. _o._ Permanent ovum. _pse._ Pseudo-epithelium. _str._ Stroma ingrowths into ovarian epithelium. _vt._ Vitelline membrane. _x._ Modified nucleus. _yk._ Yolk (vitellus). _zn._ Zona radiata.

[Figs. 17 and 18. Vide description of Plate 24.].

Fig. 19. Two nuclei from a nest which appear to be in the act of division. From ovary of the same embryo as fig. 3.

Fig. 20. Section through part of an ovary of the same embryo as fig. 6, containing a nest of nuclei. Zeiss F, ocul. 2. _Osmic acid._

Fig. 21. Ovum from the ovary of a half-grown female, containing isolated deeply stained patches of developing yolk granules. Zeiss B, ocul. 2. _Picric acid._

Fig. 22. Section through a small part of the ovum of an immature female of _Scyllium canicula_, to shew the constitution of the yolk, the follicular epithelium, and the egg membranes. Zeiss E, ocul. 2. _Chromic acid._

Fig. 23. Section through part of the periphery of a nearly ripe ovum of _Scy. canicula_. Zeiss C, ocul. 2. It shews the remnant of the vitelline membrane (_v.t._) separating the columnar but delicate cells of the follicular epithelium (_f.e._) from the yolk (_yk._). In the yolk are seen yolk-spherules in a protoplasmic network. The transverse markings in the yolk-spherules have been made oblique by the artist.

Fig. 24. Fully formed ovum containing a second nucleus (_x_), probably about to be employed as pabulum; from the same ovary as fig. 5. The follicular epithelium is much thicker on the side adjoining the stroma than on the upper side of the ovum. Zeiss F, ocul. 2. _Osmic acid._

Fig. 25. A. Ovum from the same ovary as fig. 21, containing in the yolk three peculiar bodies, similar in appearance to the two small bodies in the germinal vesicle. B. Germinal vesicle of a large ovum from the same ovary, containing a body of a strikingly similar appearance to those in the body of the ovum in A. Zeiss E, ocul. 2. _Picric acid._

Fig. 26. Section of the ovary of a young female of _Scyllium stellare_ 16-1/2 centimetres in length. The ovary is exceptional, on account of the large size of the stroma ingrowths into the epithelium. Zeiss C, ocul. 2. _Osmic acid._

Fig. 27. Ovum of _Scyllium canicula_, 5 mm. in diameter, treated with osmic acid. The figure illustrates the development of the yolk and a peculiar mode of proliferation of the germinal spots. Zeiss A, ocul. 2.

Fig. 28. Small part of the follicular epithelium and egg membranes of a somewhat larger ovum of _Scyllium canicul_a than fig. 22. Zeiss D D, ocul. 2.

Fig. 29. The same parts as in fig. 28, from a still larger ovum. Zeiss D D, ocul. 2.

Fig. 30. Ovum of Raja with follicular epithelium. Zeiss C, ocul. 2.

Fig. 31. Small portion of a larger ovum of Raja than fig. 30. Zeiss D D, ocul. 2.

Fig. 32. Follicular epithelium, &c., from an ovum of Raja still larger than fig. 31. Zeiss D D, ocul. 2.

Fig. 33. Surface view of follicular epithelium from an ovum of Raja of about the same age as fig. 33.

Fig. 34. Vertical section through the superficial part of an ovary of an adult Raja to shew the relation of the pseudo-epithelium to the subjacent stroma. Zeiss D D, ocul. 2.

PLATE 26.

COMPLETE LIST OF REFERENCE LETTERS.

_do._ Developing ovum. _fc._ Cells which will form the follicular epithelium, _fe._ Follicular epithelium. _ge._ Germinal epithelium. _mg._ Malpighian body. _n._ Nest of cells of the germinal epithelium. _nd._ Nuclei in the act of dividing. _o._ Permanent ovum. _ov._ Ovary. _po._ Primitive ovum. _t._ Tubuliferous tissue, derived from Malpighian bodies.

Fig. 35. Transverse section through the ovary of an embryo rabbit of eighteen days, hardened in osmic acid. The colours employed are intended to render clear the distinction between the germinal epithelium (_ge._) and the tubuliferous tissue (_t._), which has grown in from the Wolffian body, and which gives rise in the male to parts of the tubuli seminiferi. Zeiss A, ocul. 2.

Fig. 35A . Transverse section through a small part of the ovary of an embryo from the same female as fig. 35, hardened in picric acid, shewing the relation of the germinal epithelium to the subjacent tissue. Zeiss D D, ocul. 2.

Fig. 35B. Longitudinal section through part of the Wolffian body and the anterior end of the ovary of an eighteen days' embryo, to shew the derivation of tubuliferous tissue (_t._) from the Malpighian bodies, close to the anterior extremity of the ovary. Zeiss A, ocul. 1.

Fig. 36. Transverse section through the ovary of an embryo rabbit of twenty-two days, hardened in osmic acid. It is coloured in the same manner as fig. 35. Zeiss A, ocul. 2.

Fig. 36A. Transverse section through a small part of the ovary of an embryo, from the same female as fig. 36, hardened in picric acid, shewing the relation of the germinal epithelium to the stroma of the ovary. Zeiss D D, ocul. 2.

Figs. 37 and 37A. The same parts of an ovary of a twenty-eight days' embryo as figs. 36 and 36A of a twenty-two days' embryo.

Fig. 38. Ovary of a rabbit five days after birth, coloured in the same manner as figs. 35, 36 and 37, but represented on a somewhat smaller scale. _Picric acid._

Fig. 38A. Vertical section through a small part of the surface of the same ovary as fig. 38. Zeiss D D, ocul. 2.

Fig. 38B. Small portion of the deeper layer of the germinal epithelium of the same ovary as fig. 38. The figure shews the commencing differentiation of the cells of the germinal epithelium into true ova and follicle cells. Zeiss D D, ocul. 2.

Fig. 39A. Section through a small part of the middle region of the germinal epithelium of a rabbit seven days after birth. Zeiss D D, ocul. 2.

Fig. 39B. Section through a small part of the innermost layer of the germinal epithelium of a rabbit seven days after birth, shewing the formation of Graafian follicles. Zeiss D D, ocul. 2.

Figs. 40A and 40B. Small portions of the middle region of the germinal epithelium of a rabbit four weeks after birth. Zeiss D D, ocul. 2.

Fig. 41. Graafian follicle with two ova, about to divide into two follicles, from a rabbit six weeks after birth. Zeiss D D, ocul. 2.

XIII. ON THE EXISTENCE OF A HEAD-KIDNEY IN THE EMBRYO CHICK, AND ON CERTAIN POINTS IN THE DEVELOPMENT OF THE MÜLLERIAN DUCT[424]. BY F. M. BALFOUR AND A. SEDGWICK.

Footnote 424: From the _Quarterly Journal of Microscopical Science_, Vol. XIX. 1879.

(With Plates 27 and 28.)

The following paper is divided into three sections. The first of these records the existence of certain structures in the embryo chick, which eventually become in part the abdominal opening of the Müllerian duct, and which, we believe, correspond with the head-kidney, or "Vorniere" of German authors. The second deals with the growth and development of the Müllerian duct. With reference to this we have come to the conclusion that the Müllerian duct does not develop entirely independently of the Wolffian duct. The third section of our paper is of a more general character, and contains a discussion of the rectifications in the views of the homologies of the parts of the excretory system in Aves, necessitated by the results of our investigations.

We have, as far as possible, avoided entering into the extended literature of the excretory system, since this has been very fully given in three general papers which have recently appeared by Semper[425], Fürbinger[426], and by one of us[427].

Footnote 425: "Das Urogenital-System der Plagiostomen," _Arbeiten a. d. zool.-zoot. Institut. Würzburg_.

Footnote 426: "Zur vergl. Anat. u. Entwick. d. Excretionsorgane d. Vertebraten," _Morphologisches Jahrbuch_, Vol. IV.

Footnote 427: "On the Origin and History of the Urinogenital Organs of Vertebrates," _Journal of Anat. and Phys._, Vol. X. [This Edition No. VII.]

All recent observers, including Braun[428] for Reptilia, and Egli[429] for Mammalia, have stated that the Müllerian duct develops as a groove in the peritoneal epithelium, which is continued backward as a primitively solid rod in the space between the Wolffian duct and peritoneal epithelium.

Footnote 428: _Arbeiten a. d. zool.-zoot. Institut. Würzburg_, Vol. IV.

Footnote 429: _Beitr. zur Anat. u. Entwick. d. Geschlechtsorgane_, Inaug. Diss., Zürich, 1876.

In our preliminary account we stated[430], in accordance with the general view, that the Müllerian duct was formed as a groove, or elongated involution of the peritoneal epithelium adjoining the Wolffian duct. We have now reason to believe that this is not the case. In the earliest condition of the Müllerian duct which we have been able to observe, it consists of three successive open involutions of the peritoneal epithelium, connected together by more or less well-defined ridge-like thickenings of the epithelium. We believe, on grounds hereafter to be stated, that the whole of this formation is equivalent to the head-kidney of the Ichthyopsida. The head-kidney, as we shall continue to call it, takes its origin from the layer of thickened epithelium situated near the dorsal angle of the body-cavity, close to the Wolffian duct, which has been known since the publication of Waldeyer's important researches as the germinal epithelium. The anterior of the three open involutions or grooves is situated some little distance behind the front end of the Wolffian duct. It is simply a shallow groove in the thickest part of the germinal epithelium, and forms a corresponding projection into the adjacent stroma. In front the projection is separated by a considerable interval from the Wolffian duct; but near its hindermost part it almost comes into contact with the Wolffian duct. The groove extends in all for about five of our sections, and then terminates by its walls becoming gradually continued into a slight ridge-like thickening of the germinal epithelium. The groove arises as a simple depression in a linear area of thickened germinal epithelium. The linear area is, however, continued very considerably further forward than the groove, and sometimes exhibits a slight central depression, which might be regarded as a forward continuation of the groove. The passage from the groove to the ridge may best be conceived by supposing the groove to be suddenly filled up, so as to form a solid ridge pointing inwards towards the Wolffian duct.

Footnote 430: _Proceedings of Royal Society_, 1878.

The ridge succeeding the first groove is continued for about six sections, and is considerably more prominent at its posterior extremity than in front. It is replaced by groove number two, which appears as if formed by the reverse process to that by which the ridge arose, viz., by a hollowing out of the ridge on the side towards the body-cavity. The wall of the second groove is, after a few sections, continued into a second ridge or thickening of the germinal epithelium, which, however, is so faintly marked as to be hardly visible in its middle part. In its turn this ridge is replaced by the third and last groove. This vanishes after one or two sections, and behind the point of its disappearance we have failed to find any further traces of the head-kidney. The whole formation extends through about twenty-four of our sections and one and a half segments (muscle-plates).

We have represented (Plate 27, Series A, Nos. 1-10) a fairly complete series of sections through part of the head-kidney of an embryo slightly older than that last described, containing the second and third grooves and accessory parts. The connection between the grooves and the ridges is very well illustrated in Nos. 3, 4, and 5 of this series. In No. 3 we have a prominent ridge, in the interior of which there appears in No. 4 a groove, which becomes gradually wider in Nos. 5 and 6. Both the grooves and ridges are better marked in this than in the younger stage; but the chief difference between the two stages consists in the third groove no longer forming the hindermost limit of the head-kidney. Instead of this, the last groove (No. 7) terminates by the upper part of its walls becoming constricted off as a separate rod, which appears at first to contain a lumen continuous with the open groove. This rod (Nos. 7, 8, 9, 10) situated between the germinal epithelium and Wolffian duct is continued backward for some sections. It finally terminates by a pointed extremity, composed of not more than two cells abreast (Nos. 8-10).

Our third stage, sections of which are represented in series B (Plate 27), is considerably advanced beyond that last described. The most important change which has been effected concerns the ridges connecting the successive grooves. A lumen has appeared in each of these, which seems to open at both ends into the adjacent grooves. At the same time the cells, which previously constituted the ridge, have become (except where they are continuous with the walls of the grooves) partially constricted off from the germinal epithelium. The ridges, in fact, now form ducts situated in the stroma of the ovarian ridge, in the space between the Wolffian duct and the germinal epithelium. The duct continuous with the last groove is somewhat longer than before. In a general way, the head-kidney may now be described as a duct opening into the body-cavity by three groove-like apertures, and continuous behind with the rudiment of the true Müllerian duct. Although the general constitution of the head-kidney at this stage is fairly simple, there are a few features in our sections which we do not fully understand, and a few points about the organ which deserve a rather fuller description than we have given in this general sketch.

The anterior groove (Nos. 1-3, series B, Pl. 27) is at first somewhat separated from the Wolffian duct, but approaches close to it in No. 3. In Nos. 2 and 3 there appears a rod-like body on the outer side of the walls of the groove. In No. 2 this body is disconnected with the walls of the groove, and even appears as if formed by a second invagination of the germinal epithelium. In No. 3 this body becomes partially continuous with the walls of the groove, and finally in No. 4 it becomes completely continuous with the walls of the groove, and its lumen communicates freely with the groove[431].

Footnote 431: A deep focus of the rather thick section represented in No. 3 shewed the body much more nearly in the position it occupies in No. 4.

The last trace of this body is seen on the upper wall of the groove in No. 5. We believe that the body (_r_1) represents the ridge between the first and second grooves of the earlier stage; so that in passing from No. 3 to No. 5 we pass from the first to the second groove. The meaning of the features of the body _r_1 in No. 2 we do not fully understand, but cannot regard them as purely accidental, since we have met with more or less similar features in other series of sections. The second groove becomes gradually narrower, and finally is continued into the second ridge (No. 8). The ridge contains a lumen, and is only connected with the germinal epithelium by a narrow wall of cells. A narrow passage from the body-cavity leads into that wall for a short distance in No. 8, but it is probably merely the hinder end of the groove of No. 7. The third groove appears in No. 11, and opens into the lumen of the second ridge (_r_2) in No. 12. In No. 13 the groove is closed, and there is present in its place a duct (_r_3) connected with the germinal epithelium by a wall of cells. This duct is the further development of the third ridge of the last stage; its lumen opens into the body-cavity through the third and last groove (_gr_3). In the next section this duct (_r_3) is entirely separated from the germinal epithelium, and it may be traced backwards through several sections until it terminates by a solid point, very much as in the last stage.

In the figures of this series (B) there may be noticed on the outer side of the Müllerian duct a fold of the germinal epithelium (_x_) forming a second groove. It is especially conspicuous in the first six sections of the series. This fold sometimes becomes much deeper, and then forms a groove, the upper end of which is close to the grooves of the head-kidney. It is very often much deeper than these are, and without careful study might easily be mistaken for one of these grooves. Fig. C, taken from a series slightly younger than B, shews this groove (_x_) in its most exaggerated form.

The stage we have just described is that of the fullest development of the head-kidney. In it, as in all the previous stages, there appear to be only three main openings into the body-cavity; but we have met in some of our sections with indications of the possible presence of one or two extra rudimentary grooves.

In an embryo not very much older than the one last described the atrophy of the head-kidney is nearly completed, and there is present but a single groove opening into the body-cavity.

In series D (Pl. 28) are represented a number of sections from an embryo at this stage. Nos. 1 and 2 are sections through the hind end of the single groove now present. Its walls are widely separated from the Wolffian duct in front, but approach close to it at the hinder termination of the groove (No. 2). The features of the single groove present at this stage agree closely with those of the anterior groove of the previous stages. The groove is continued into a duct--the Müllerian duct (as it may now be called, but in a previous stage the hollow ridge connecting the first and second grooves of the head-kidney)--which, after becoming nearly separated from the germinal epithelium, is again connected to it by a mass of cells at two points (Nos. 5, 6, and 8). The germinal epithelium is slightly grooved and is much reduced in thickness at these points of contact (_gr_2 and _gr_3), and we believe that they are the remnants of the posterior grooves of the head-kidney present at an earlier stage.

The Müllerian duct has by this stage grown much further backwards, but the peculiarities of this part of it are treated in a subsequent section.

We consider that, taking into account the rudiments we have just described, as well as the fact that the features of the single groove at this stage correspond with those of the anterior groove at an earlier stage, we are fully justified in concluding that _the permanent abdominal opening of the Müllerian duct corresponds with the anterior of our three grooves_.

Although we have, on account of their indefiniteness, avoided giving the ages of the chicks in which the successive changes of the head-kidney may be observed, we may, perhaps, state that all the changes we have described are usually completed between the 90th and 120th hour of incubation.

_The Glomerulus of the Head-Kidney._

In connection with the head-kidney in Amphibians there is present, as is well known, a peculiar vascular body usually described as the glomerulus of the head-kidney. We have found in the chick a body so completely answering to this glomerulus that we have hardly any hesitation in identifying it as such.

In the chick the glomerulus is paired, and consists of a vascular outgrowth or ridge projecting into the body-cavity on each side at the root of the mesentery. It extends from the anterior end of the Wolffian body to the point where the foremost opening of the head-kidney commences. We have found it at a period slightly earlier than that of the first development of the head-kidney. It is represented in figs. E and F, Pl. 28, _gl_, and is seen to form a somewhat irregular projection into the body-cavity, covered by a continuation of the peritoneal epithelium, and attached by a narrow stalk to the insertion of the embryonic mesentery (_me_).

In the interior of this body is seen a stroma with numerous vascular channels and blood corpuscles, and a vascular connection is apparently becoming established, if it is not so already, between the glomerulus and the aorta. We have reason to think that the corpuscles and vascular channels in the glomerulus are developed _in situ_. The stalk connecting the glomerulus with the attachment of the mesentery varies in thickness in different sections, but we believe that the glomerulus is continued unbroken throughout the very considerable region through which it extends. This point is, however, difficult to make sure of owing to the facility with which the glomerulus breaks away.

At the stage we are describing, no true Malpighian bodies are present in the part of the Wolffian body on the same level with the anterior end of the glomerulus, but the Wolffian body merely consists of the Wolffian duct. At the level of the posterior part of the glomerulus this is no longer the case, but here a regular series of primary Malpighian bodies is present (using the term "primary" to denote the Malpighian bodies developed directly out of part of the primary segmental tubes), and the glomerulus of the head-kidney may frequently be seen in the same section as a Malpighian body. In most sections the two bodies appear quite disconnected, but in those sections in which the _glomerulus_ of the Malpighian body comes into view it is seen to be derived from the same formation as the glomerulus of the head-kidney (Pl. 28, fig. F). It would seem, in fact, that the vascular tissue of the glomerulus of the head-kidney grows into the concavity of the Malpighian bodies. Owing to the stage we are now describing, in which we have found the glomerulus most fully developed, being prior to that in which the head-kidney appears, it is not possible to determine with certainty the position of the glomerulus in relation to the head-kidney. After the development of the head-kidney it is found, however, as we have already stated, that the glomerulus terminates at a point just in front of the anterior opening of the head-kidney. It is less developed than before, but is still present up to the period of the atrophy of the head-kidney. It does not apparently alter in constitution, and we have not thought it worth while giving any further representations of it during the later stages of its existence.

_Summary of the development of the head-kidney and glomerulus._--The first rudiment of the head-kidney arises as three successive grooves in the thickened germinal epithelium, connected by ridges, and situated some way behind the front end of the Wolffian duct. In the next stage the three ridges connecting the grooves have become more marked, and in each of them a lumen has appeared, opening at both extremities into the adjoining grooves. Still later the ridges become more or less completely detached from the peritoneal epithelium, and the whole head-kidney then consists of a slightly convoluted duct, with, at the least, three peritoneal openings, which is posteriorly continued into the Müllerian duct. Still later the head-kidney atrophies, its two posterior openings vanishing, and its anterior opening remaining as the permanent opening of the Müllerian duct. The glomerulus arises as a vascular prominence at the root of the mesentery, slightly prior in point of time to the head-kidney, and slightly more forward than it in position. We have not traced its atrophy.

We stated in our preliminary paper that the peculiar structures we had interpreted as the head-kidney had completely escaped the attention of previous observers, though we called attention to a well-known figure of Waldeyer's (copied in the _Elements of Embryology,_ fig. 51). In this figure a connection between the germinal epithelium and the Müllerian duct is drawn, which is probably part of the head-kidney, and may be compared with our figures (Series B, No. 8, and Series D, No. 4). Since we made the above statement, Dr Gasser has called our attention to a passage in his valuable memoir on "The Development of the Allantois[432]," in which certain structures are described which are, perhaps, identical with our head-kidney. The following is a translation of the passage:--

"In the upper region of Müller's duct I have often observed small canals, especially in the later stages of development, which appear as a kind of doubling of the duct, and run for a short distance close to Müller's duct and in the same direction, opening, however, into the body-cavity posterior to the main duct. Further, one may often observe diverticula from the extreme anterior end of the oviduct of the bird, which form blind pouches and give one the impression of being receptacula seminis. Both these appearances can quite well be accounted for on the supposition that an abnormal communication is effected between the germinal epithelium and Müller's duct at unusual places; or else that an attempt at such a communication is made, resulting, however, only in the formation of a diverticulum of the wall of the oviduct."

Footnote 432: _Beiträge zur Entwicklungsgeschichte d. Allantois der Müller'schen Gange u. des Afters._ Frankfurt, 1874.

The statement that these accessory canals are late in developing, prevents us from feeling quite confident that they really correspond with our head-kidney.

Before passing on to the other parts of this paper it is necessary to say a few words in justification of the comparison we have made between the modified abdominal extremity of the Müllerian duct in the chick and the head-kidney of the Ichthyopsida.

For the fullest statement of what is known with reference to the anatomy and development of the head-kidney in the lower types we may refer to Spengel and Fürbringer[433]. We propose ourselves merely giving a sufficient account of the head-kidney in Amphibia (which appears to be the type in which the head-kidney can be most advantageously compared with that in the bird) to bring out the grounds for our determination of the homologies.

Footnote 433: _Loc. cit._

The development of the head-kidney in Amphibia has been fully elucidated by the researches of W. Müller[434], Götte[435], and Fürbringer[436], while to the latter we are indebted for a knowledge of the development of the Müllerian duct in Amphibians. The first part of the urinogenital system to develop is the segmental duct (_Vornieregang_ of Fürbringer), which is formed by a groove-like invagination of the peritoneal epithelium. It becomes constricted into a duct first of all in the middle, but soon in the posterior part also. It then forms a duct, ending in front by a groove in free communication with the body-cavity, and terminating blindly behind. The open groove in front at first deepens, and then becomes partially constricted into a duct, which elongates and becomes convoluted, but remains in communication with the body-cavity by from two to four (according to the species) separate openings. The manner in which the primitive single opening is related to the secondary openings is not fully understood. By these changes there is formed out of the primitive groove an anterior glandular body, communicating with the body-cavity by several apertures, and a posterior duct, which carries off the secretion of the gland, and which, though blind at first, eventually opens into the cloaca. In addition to these parts there is also formed on each side of the mesentery, opposite the peritoneal openings, a very vascular projection into this part of the body-cavity, which is known as the glomerulus of the head-kidney, and which very closely resembles in structure and position the body to which we have assigned the same name in the chick.

Footnote 434: _Jenaische Zeitschrift_, Vol. IX. 1875.

Footnote 435: _Entwicklungsgeschichte d. Unke._

Footnote 436: _Loc. cit._

The primitive segmental duct is at first only the duct for the head-kidney, but on the formation of the posterior parts of the kidney (Wolffian body, &c.) it becomes the duct for these also.

After the Wolffian bodies have attained to a considerable development, the head-kidney undergoes atrophy, and its peritoneal openings become successively closed from before backwards. At this period the formation of the Müllerian duct takes place. It is a solid constriction of the ventral or lateral wall of the segmental duct, which subsequently becomes hollow, and acquires an opening into the body-cavity _quite independent of the openings of the head-kidney_.

The similarity in development and structure between the head-kidney in Amphibia and the body we have identified as such in Aves, is to our minds too striking to be denied. Both consist of two parts--(1) a somewhat convoluted longitudinal canal, with a certain number of peritoneal openings; (2) a vascular prominence at the root of the mesentery, which forms a glomerulus. As to the identity in position of the two organs we hope to deal with that more fully in speaking of the general structure of the excretory system, but may say that one of us[437] has already, on other grounds, attempted to shew that the abdominal opening of the Müllerian duct in the bird is the homologue of the abdominal opening of the segmental duct in Amphibia, Elasmobranchii, &c., and that we believe that this homology will be admitted by most anatomists. If this homology is admitted, the identity in position of this organ in Aves and Amphibia necessarily follows. The most striking difference between Aves and Amphibia in relation to these structures is the fact that in Aves the anterior pore of the head-kidney remains as the permanent opening of the Müllerian duct, while in Amphibia, the pores of the head-kidney atrophy, and an entirely fresh abdominal opening is formed for the Müllerian duct.

Footnote 437: Balfour, "Origin and History of Urinogenital Organs of Vertebrates," _Journal of Anat. and Phys._ Vol. X., and _Monograph on Elasmobranch Fishes._ [This edition Nos. VII. and X.]

II.

_The Growth of the Müllerian Duct._

Although a great variety of views have been expressed by different observers on the growth of the Müllerian duct, it is now fairly generally admitted that it grows in the space between a portion of the thickened germinal epithelium and the Wolffian duct, but quite independently of both of them. Both Braun and Egli, who have specially directed their attention to this point, have for Reptilia and Mammalia fully confirmed the views of previous observers. We were, nevertheless, induced, partly on account of the _à priori_ difficulties of this view, and partly by certain peculiar appearances which we observed, to undertake the re-examination of this point, and have found ourselves unable altogether to accept the general account. We propose first describing, in as matter-of-fact a way as possible, the actual observations we have made, and then stating what conclusions we think may be drawn from these observations.

We have found it necessary to distinguish three stages in the growth of the Müllerian duct. Our first stage embraces the period prior to the disappearance of the head-kidney. At this stage the structure we have already spoken of as the rudiment of the Müllerian duct consists of a solid rod of cells, continuous with the third groove of the head-kidney. It extends through a very few sections, and terminates by a fine point of about two cells, wedged in between the Wolffian duct and germinal epithelium (described above, Nos. 7-10, series A, Plate 27).

In an embryo slightly older than the above, such as that from which series B was taken, but still belonging to our first stage, a definite lumen appears in the anterior part of the Müllerian duct, which vanishes after a few sections. The duct terminates in a point which lies in a concavity of the wall of the Wolffian duct (Plate 27, Nos. 1 and 2, series G). The limits of the Wolffian wall and the pointed termination of the Müllerian duct are in many instances quite distinct; but the outline of the Wolffian duct appears to be carried round the Müllerian duct, and in some instances the terminal point of the Müllerian duct seems almost to form an integral part of the wall of the Wolffian duct.

The second of our stages corresponds with that in which the atrophy of the head-kidney is nearly complete (series D and H, Plate 28).

The Müllerian duct has by this stage made a very marked progress in its growth towards the cloaca, and, in contradistinction to the earlier stage, a lumen is now continued close up to the terminal point of the duct. In the two or three sections before it ends it appears as a distinct oval mass of cells (No. 10, series D, and No. 1, series H), without a lumen, lying between and touching the external wall of the Wolffian duct on the one hand, and the germinal epithelium on the other. It may either lie on the ventral side of the Wolffian duct (series D), or on the outer side (series H), but in either case is opposite the maximum thickening of that part of the germinal epithelium which always accompanies the Müllerian duct in its backward growth.

In the last section in which any trace of the Müllerian duct can be made out (series D, No. 11, and series H, No. 2), it has no longer an oval, well-defined contour, but appears to have completely fused with the wall of the Wolffian duct, which is accordingly very thick, and occupies the space which in the previous section was filled by its own wall and the Müllerian duct. In the following section the thickening in the wall of the Wolffian duct has disappeared (Plate 28, series H, No. 3), and every trace of the Müllerian duct has vanished from view. The Wolffian duct is on one side in contact with the germinal epithelium.

The stage during which the condition above described lasts is not of long duration, but is soon succeeded by our third stage, in which a fresh mode of termination of the Müllerian duct is found. (Plate 28, series I.) This last stage remains up to about the close of the sixth day, beyond which our investigations do not extend.

A typical series of sections through the terminal part of the Müllerian duct at this stage presents the following features:

A few sections before its termination the Müllerian duct appears as a well-defined oval duct lying in contact with the wall of the Wolffian duct on the one hand and the germinal epithelium on the other (series I, No. 1). Gradually, however, as we pass backwards, the Müllerian duct dilates; the external wall of the Wolffian duct adjoining it becomes greatly thickened and pushed in in its middle part, so as almost to touch the opposite wall of the duct, and so form a bay in which the Müllerian duct lies (Plate 28, series I, Nos. 2 and 3). As soon as the Müllerian duct has come to lie in this bay its walls lose their previous distinctness of outline, and the cells composing them assume a curious vacuolated appearance. No well-defined line of separation can any longer be traced between the walls of the Wolffian duct and those of the Müllerian, but between the two is a narrow clear space traversed by an irregular network of fibres, in some of the meshes of which nuclei are present.

The Müllerian duct may be traced in this condition for a considerable number of sections, the peculiar features above described becoming more and more marked as its termination is approached. It continues to dilate and attains a maximum size in the section or so before it disappears. A lumen may be observed in it up to its very end, but is usually irregular in outline and frequently traversed by strands of protoplasm. The Müllerian duct finally terminates quite suddenly (Plate 28, series I, No. 4), and in the section immediately behind its termination the Wolffian duct assumes its normal appearance, and the part of its outer wall on the level of the Müllerian duct comes into contact with the germinal epithelium (Plate 28, series I, No. 5).

We have traced the growing point of the Müllerian duct with the above features till not far from the cloaca, but we have not followed the last phases of its growth and its final opening into the cloaca.

In some of our embryos we have noticed certain rather peculiar structures, an example of which is represented at _y_ in fig. K, taken from an embryo of 123 hours, in which all traces of the head-kidney had disappeared. It consists of a cord of cells, connecting the Wolffian duct and the hind end of the abdominal opening of the Müllerian duct. At the least one similar cord was met with in the same embryo, situated just behind the abdominal opening of the Müllerian duct. We have found similar structures in other embryos of about the same age, though never so well marked as in the embryo from which fig. K is taken. We have quite failed to make out the meaning, if any, of them.

Our interpretation of the appearances we have described in connection with the growth of the Müllerian duct can be stated in a very few words. Our second stage, where the solid point of the Müllerian duct terminates by fusing with the walls of the Wolffian duct, we interpret as meaning that the Müllerian is growing backwards as a solid rod of cells, split off from the outer wall of the Wolffian duct; in the same manner, in fact, as in Amphibia and Elasmobranchii. The condition of the terminal part of the Müllerian duct during our third stage cannot, we think, be interpreted in the same way, but the peculiarities of the cells of both Müllerian and Wolffian ducts, and the indistinctness of the outlines between them, appear to indicate that the Müllerian duct grows by cells passing from the Wolffian duct to it. In fact, although in a certain sense the growth of the two ducts is independent, yet the actual cells which assist in the growth of the Müllerian duct are, we believe, derived from the walls of the Wolffian duct.

III.

_General considerations._

The excretory system of a typical Vertebrate consists of the following parts:--

1. A head-kidney with the characters already described.

2. A duct for the head-kidney--the segmental duct.

3. A posterior kidney--(Wolffian body, permanent kidney, &c. The nature and relation of these parts we leave out of consideration, as they have no bearing upon our present investigations). The primitive duct for the Wolffian body is the segmental duct.

4. The segmental duct may become split into (_a_) a dorsal or inner duct, which serves as ureter (in the widest sense of the word); and (_b_) a ventral or outer duct, which has an opening into the body-cavity, and serves as the generative duct for the female, or for both sexes.

These parts exhibit considerable variations both in their structure and development, into some of which it is necessary for us to enter.

The head-kidney[438] attains to its highest development in the Marsipobranchii (Myxine, Bdellostoma). It consists of a longitudinal canal, from the ventral side of which numerous tubules pass. These tubules, after considerable subdivision, open by a large number of apertures into the pericardial cavity. From the longitudinal canal a few dorsal diverticula, provided with glomeruli, are given off. In the young the longitudinal canal is continued into the segmental duct; but this connection becomes lost in the adult. The head-kidney remains, however, through life. In Teleostei and Ganoidei (?) the head-kidney is generally believed to remain through life, as the dilated cephalic portion of the kidneys when such is present. In Petromyzon and Amphibia the head-kidney atrophies. In Elasmobranchii the head-kidney, so far as is known, is absent.

Footnote 438: I am inclined to give up the view I formerly expressed with reference to the head-kidney and segmental duct, viz. "that they were to be regarded as the most anterior segmental tube, the peritoneal opening of which had become divided, and which had become prolonged backwards so as to serve as the duct for the posterior segmental tubes," and _provisionally_ to accept the Gegenbaur-Fürbringer view which has been fully worked out and ably argued for by Fürbringer (_loc. cit._ p. 96). According to this view the head-kidney and its duct are to be looked on as the primitive and unsegmented part of the excretory system, more or less similar to the excretory system of many Trematodes and unsegmented Vermes. The segmental tubes I regard as a truly segmental part of the excretory system acquired subsequently.--F. M. B.

The development of the segmental duct and head-kidney (when present) is still more important for our purpose than their adult structure.

In Myxine the development of these structures is not known. In Amphibia and Teleostei it takes place upon the same type, viz. by the conversion of a groove-like invagination of the peritoneal epithelium into a canal open in front. The head-kidney is developed from the anterior end of this canal, the opening of which remains in Teleostei single and closes early in embryonic life, but becomes in Amphibia divided into two, three, or four openings. In Elasmobranchii the development is very different.

"The first trace of the urinary system makes its appearance as a knob springing from the intermediate cell-mass opposite the fifth protovertebra. This knob is the rudiment of the abdominal opening of the segmental duct, and from it there grows backwards to the level of the anus a solid column of cells, which constitutes the rudiment of the segmental duct itself. The knob projects towards the epiblast, and the column connected with it lies between the mesoblast and epiblast. The knob and column do not long remain solid, but the former acquires an opening into the body-cavity continuous with a lumen, which makes its appearance in the latter."

The difference in the development of the segmental duct in the two types (Amphibia and Elasmobranchii) is very important. In the one case a continuous groove of the peritoneal epithelium becomes constricted into a canal, in the other a solid knob of cells is continued into a rod, at first solid, which grows backwards without any apparent relation to the peritoneal epithelium[439].

Footnote 439: In a note on p. 50 of his memoir Fürbringer criticises my description of the mode of growth of the segmental duct. The following is a free translation of what he says: "In Balfour's, as in other descriptions, an account is given of a backward growth, which easily leads to the supposition of a structure formed anteriorly forcing its way through the tissues behind. This is, however, not the case, since, to my knowledge, no author has ever detected a sharp boundary between the growing point of the segmental duct (or Müllerian duct) and the surrounding tissues." He goes on to say that "the growth in these cases really takes place by a differentiation of tissue along a line in the region of the peritoneal cavity." Although I fully admit that it would be far easier to homologise the development of the segmental duct in Amphibia and Elasmobranchii according to this view, I must nevertheless vindicate the accuracy of my original account. I have looked over my specimens again, since the appearance of Dr Fürbringer's paper, and can find no evidence of the end of the duct becoming continuous with the adjoining mesoblastic tissues. In the section, before its disappearance, the segmental duct may, so far as I can make out, be seen as a very small but distinct rod, which is much more closely connected with the epiblast than with any other layer. From Gasser's observations on the Wolffian duct in the bird, I am led to conclude that it behaves in the same way as the segmental duct in the Elasmobranchii. I will not deny that it is possible that the growth of the duct takes place by wandering cells, but on this point I have no evidence, and must therefore leave the question an open one.--F. M. B.

The abdominal aperture of the segmental duct in Elasmobranchii, in that it becomes the permanent abdominal opening of the oviduct, corresponds physiologically rather with the abdominal opening of the Müllerian duct than with that of the segmental duct of Amphibia, which, after becoming divided up to form the pores of the head-kidney, undergoes atrophy. Morphologically, however, it appears to correspond with the opening of the segmental duct in Amphibia. We shall allude to this point more than once again, and give our grounds for the above view on p. 640.

The development of the segmental duct in Elasmobranchii as a solid rod is, we hope to shew, of special importance for the elucidation of the excretory system of Aves.

The development of these parts of Petromyzon is not fully known, but from W. Müller's account (_Jenaische Zeitschrift_, 1875) it would seem that an anterior invagination of the peritoneal epithelium is continued backwards as a duct (segmental duct), and that the anterior opening subsequently becomes divided up into the various apertures of the head-kidney. If this account is correct, Petromyzon presents a type intermediate between Amphibia and Elasmobranchii. In certain types, viz. Marsipobranchii and Teleostei, the segmental duct becomes the duct for the posterior kidney (segmental tubes), but otherwise undergoes no further differentiation. In the majority of types, however, the case is different. In Amphibia[440], as has already been mentioned, a solid rod of cells is split off from its ventral wall, which afterwards becomes hollow, acquires an opening into the body-cavity, and forms the Müllerian duct.

Footnote 440: Fürbringer, _loc. cit._

In Elasmobranchii the segmental duct undergoes a more or less similar division. "It becomes longitudinally split into two complete ducts in the female, and one complete duct and parts of a second in the male. The resulting ducts are (1) the Wolffian duct dorsally, which remains continuous with the excretory tubules of the kidney, and ventrally (2) the oviduct or Müllerian duct in the female, and the rudiments of this duct in the male. In the female the formation of these ducts takes place by a nearly solid rod of cells, being gradually split off from the ventral side of all but the foremost part of the original segmental duct, with the short undivided anterior part of which duct it is continuous in front. Into it a very small portion of the lumen of the original segmental duct is perhaps continued. The remainder of the segmental duct (after the loss of its anterior section and the part split off from its ventral side) forms the Wolffian duct. The process of formation of the ducts in the male chiefly differs from that in the female, in the fact of the anterior undivided part of the segmental duct, which forms the front end of the Müllerian duct, being shorter, and in the column of cells with which it is continuous being from the first incomplete."

It will be seen from the above that the Müllerian duct consists of two distinct parts--an anterior part with the abdominal opening, and a posterior part split off from the segmental duct. This double constitution of the Müllerian duct is of great importance for a proper understanding of what takes place in the Bird.

The Müllerian duct appears therefore to develop in nearly the same manner in the Amphibian and Elasmobranch type, as a solid or nearly solid rod split off from the ventral wall of the segmental duct. But there is one important difference concerning the abdominal opening of the duct. In Amphibia this is a new formation, but in Elasmobranchii it is the original opening of the segmental duct. Although we admit that in a large number of points, including the presence of a head-kidney, the urinogenital organs of Amphibia are formed on a lower type than those of the Elasmobranchii, yet it appears to us that this does not hold good for the development of the Müllerian duct.

The above description will, we trust, be sufficient to render clear our views upon the development of the excretory system in Aves.

In the bird the excretory system consists of the following parts (using the ordinary nomenclature) which are developed in the order below.

1. Wolffian duct. 2. Wolffian body. 3. Head-kidney. 4. Müllerian duct. 5. Permanent kidney and ureter.

About 2 and 5 we shall have nothing to say in the sequel.

We have already in the early part of the paper given an account of the head-kidney and Müllerian duct, but it will be necessary for us to say a few words about the development of the Wolffian duct (so called). Without entering into the somewhat extended literature on the subject, we may state that we consider that the recent paper of Dr Gasser[441] supplies us with the best extant account of the development of the Wolffian duct.

Footnote 441: _Arch. für Mic. Anat._ Vol. XIV.

The first trace of it, which he finds, is visible in an embryo with eight protovertebræ as a slight projection from the intermediate cell mass towards the epiblast in the region of the three hindermost protovertebræ. In the next stage, with eleven protovertebræ, the solid rudiment of the duct extends from the fifth to the eleventh protovertebra, from the eighth to the eleventh protovertebra it lies between the epiblast and mesoblast, and is quite distinct from both, and Dr Gasser distinctly states that in its growth backwards from the eighth protovertebra the Wolffian duct never comes into continuity with the adjacent layers.

In the region of the fifth protovertebra, where the duct was originally continuous with the mesoblast, it has now become free, but is still attached in the region of the sixth and to the eighth protovertebra. In an embryo with fourteen protovertebræ the duct extends from the fourth to the fourteenth protovertebra, and is now free between epiblast and mesoblast for its whole extent. It is still for the most part solid though perhaps a small lumen is present in its middle part. In the succeeding stages the lumen of the duct gradually extends backwards and forwards, the duct itself also passes inwards till it acquires its final position close to the peritoneal epithelium; at the same time its hind end elongates till it comes into connection with the cloacal section of the hind-gut. It should be noted that the duct in its backward growth does not appear to come into continuity with the subjacent mesoblast, but behaves in this respect exactly as does the segmental duct in Elasmobranchii (vide note on p. 634).

The question which we propose to ourselves is the following:--What are the homologies of the parts of the Avian urinogenital system above enumerated? The Wolffian duct appears to us morphologically to correspond _in part_ to the segmental duct[442], or what Fürbringer would call the duct of the head-kidney. This may seem a paradox, since in birds it never comes into relation with the head-kidney. Nevertheless we consider that this homology is morphologically established, for the following reasons:--

Footnote 442: The views here expressed about the Wolffian duct are nearly though not exactly those which one of us previously put forward ("Urinogenital Organs of Vertebrates," &c., pp. 45-46) [This edition, pp. 164, 165], and with which Fürbringer appears exactly to agree. Possibly Dr Fürbringer would alter his view on this point were he to accept the facts we believe ourselves to have discovered. Semper's view also differs from ours, in that he believes the Wolffian duct to correspond in its entirety with the segmental duct.

(1) That the Wolffian duct gives rise (vide _supra_, p. 631) to the Müllerian duct as well as to the duct of the Wolffian body. In this respect it behaves precisely as does the segmental duct of Elasmobranchii and Amphibia. That it serves as the duct for the Wolffian body, before the Müllerian duct originates from it, is also in accordance with what takes place in other types.

(2) That it develops in a strikingly similar manner to the segmental duct of Elasmobranchii.

We stated expressly that the Wolffian duct corresponded only in part to the segmental duct. It does not, in fact, in our opinion, correspond to the whole segmental duct, but to the segmental duct minus the anterior abdominal opening in Elasmobranchii, which becomes the head-kidney in other types. In fact, we suppose that the segmental duct and head-kidney, which in the Ichthyopsida develop as a single formation, develop in the Bird as two distinct structures--one of these known as the Wolffian duct, and the other the head-kidney. If our view about the head-kidney is accepted the above position will hardly require to be disputed, but we may point out that the only feature in which the Wolffian duct of the Bird differs in development from the segmental duct of Elasmobranchii is in the absence of the knob, which forms the commencement of the segmental duct, and in which the abdominal opening is formed; so that the comparison of the development of the duct in the two types confirms the view arrived at from other considerations.

The head-kidney and Müllerian duct in the Bird must be considered together. The parts which they eventually give rise to after the atrophy of the head-kidney have almost universally been regarded as equivalent to the Müllerian duct of the Ichthyopsida. By Braun[443], however, who from his researches on the Lizard satisfied himself of the entire independence of the Müllerian and Wolffian ducts in the Amniota, the Müllerian duct of these forms is regarded as a completely new structure with no genetic relations to the Müllerian duct of the Ichthyopsida. Semper[444], on the other hand, though he accepts the homology of the Müllerian duct in the Ichthyopsida and Amniota, is of opinion that the anterior part of the Müllerian duct in the Amniota is really derived from the Wolffian duct, though he apparently admits the independent growth of the posterior part of the Müllerian duct. We have been led by our observations, as well as by our theoretical deductions, to adopt a view exactly the reverse of that of Professor Semper. We believe that the anterior part of the Müllerian duct of Aves, which is at first the head-kidney, and subsequently becomes the abdominal opening of the duct, is developed from the peritoneal epithelium independently of all other parts of the excretory system; but that the posterior part of the duct is more or less completely derived from the walls of the Wolffian duct. This view is clearly in accordance with our account of the facts of development in Aves, and it fits in very well with the development of the Müllerian duct in Elasmobranchii. We have already pointed out that in Elasmobranchii the Müllerian duct is formed of two factors--(1) of the whole anterior extremity of the segmental duct, including its abdominal opening; (2) of a rod split off from the ventral side of the segmental duct. In Birds the anterior part (corresponding to factor No. 1) of the Müllerian duct has a different origin from the remainder; so that if the development of the posterior part of the duct (factor No. 2) were to proceed in the same manner in Birds and Elasmobranchii, it ought to be formed at the expense of the Wolffian (_i.e._ segmental) duct, though in connection anteriorly with the head-kidney. And this is what actually appears to take place.

Footnote 443: "Urogenital-System d. Reptilien," _Arb. aus d. zool.-zoot. Inst. Würzburg_, Vol. IV.

Footnote 444: _Loc. cit._

So far the homologies of the avian excretory system are fairly clear; but there are still some points which have to be dealt with in connection with the permanent opening of the Müllerian duct, and the relatively posterior position of the head-kidney. With reference to the first of these points the facts of the case are the following:--

In Amphibia the permanent opening of the Müllerian duct is formed as an independent opening after the atrophy of the head-kidney.

In Elasmobranchii the original opening of the segmental duct forms the permanent opening of the Müllerian duct and no head-kidney appears to be formed.

In Birds the anterior of the three openings of the head-kidney remains as the permanent opening of the Müllerian duct.

With reference to the difficulties involved in there being apparently three different modes in which the permanent opening of the Müllerian duct is formed, we would suggest the following considerations:

The history of the development of the excretory system teaches us that primitively the segmental duct must have served as efferent duct both for the generative products and kidney secretion (just as the Wolffian duct still does for the testicular products and secretion of the Wolffian body in Elasmobranchii and Amphibia); and further, that at first the generative products entered the segmental duct from the abdominal cavity by one or more of the abdominal openings of the kidney (almost certainly of the head-kidney). That the generative products did not enter the segmental duct at first by an opening specially developed for them appears to us to follow from Dohrn's principle of the transmutation of function (_Functionswechsel_). As a consequence (by a process of natural selection) of the segmental duct having both a generative and a urinary function, a further differentiation took place, by which that duct became split into two--a ventral Müllerian duct and dorsal Wolffian duct.

The Müllerian duct without doubt was continuous with the head-kidney, and so with the abdominal opening or openings of the head-kidney which served as generative pores. At first the segmental duct was probably split longitudinally into two equal portions, but the generative function of the Müllerian duct gradually impressed itself more and more upon the embryonic development, so that, in the course of time, the Müllerian duct developed less and less at the expense of the Wolffian duct. This process appears partly to have taken place in Elasmobranchii, and still more in Amphibia; the Amphibia offering in this respect a less primitive condition than Elasmobranchii; while in Aves it has been carried even further. The abdominal opening no doubt also became specialised. At first it is quite possible that more than one abdominal pore may have served for the generative products; one of which, no doubt, eventually came to function alone. In Amphibia the specialisation of the opening appears to have gone so far that it no longer has any relation to the head-kidney, and even develops after the atrophy of the head-kidney. In Elasmobranchii, on the other hand, the functional opening appears at a period when we should expect the head-kidney to develop. This state is very possibly the result of a differentiation (along a different line to that in Amphibia) by which the head-kidney gradually ceased to become developed, but by which the primitive opening (which in the development of the head-kidney used to be divided into several pores leading into the body-cavity) remained undivided and served as the abdominal aperture of the Müllerian duct. Aves, finally, appear to have become differentiated along a third line; since in their ancestors the anterior pore of the head-kidney appears to have become specialised as the permanent opening of the Müllerian duct.

With reference to the posterior position of the head-kidney in Aves we have only to remark, that a change in position of the head-kidney might easily take place after it acquired an independent development. The fact that it is slightly behind the glomerulus would seem to indicate, on the one hand, that it has already ceased to be of any functional importance; and, on the other, that the shifting has been due to its having a connection with the Müllerian duct.

We have made a few observations on the development of the Müllerian duct in _Lacerta muralis_, which have unfortunately led us to no decided conclusions. In a fairly young stage in the development of the Müllerian duct (the youngest we have met with), no trace of a head-kidney could be observed, but the character of the abdominal opening of the Müllerian duct was very similar to that figured by Braun[445]. As to the backward growth of the Müllerian duct, we can only state that the solid point of the duct in the young stages is in contact with the wall of the Wolffian duct, and the relation between the two is rather like that figured by Fürbringer (Pl. 1, figs. 14-15) in Amphibia.

Footnote 445: _Loc. cit._

DESCRIPTION OF PLATES 27 AND 28.

COMPLETE LIST OF REFERENCE LETTERS.

_ao._ Aorta. _cv._ Cardinal vein. _gl._ Glomerulus. _gr_1. First groove of head-kidney. _gr_2. Second groove of head-kidney. _gr_3. Third groove of head-kidney. _ge._ Germinal epithelium. _mrb._ Malpighian body. _me._ Mesentery. _md._ Müllerian duct. _r_1. First ridge of head-kidney. _r_2. Second ridge of head-kidney. _r_3. Third ridge of head-kidney. _Wd._ Wolffian duct. _x._ Fold in germinal epithelium.

PLATE 27.

SERIES A. Sections through the head-kidney at our second stage. Zeiss 2, ocul. 3 (reduced one-third). The second and third grooves are represented with the ridge connecting them, and the rod of cells running backwards for a short distance.

No. 1. Section through the second groove.

No. 2. Section through the ridge connecting the second and third grooves.

No. 3. Section passing through the same ridge at a point nearer the third groove.

Nos. 4, 5, 6. Sections through the third groove.

No. 7. Section through the point where the third groove passes into the solid rod of cells.

No. 8. Section through the rod when quite separated from the germinal epithelium.

No. 9. Section very near the termination of the rod.

No. 10. Last section in which any trace of the rod is seen.

SERIES B. Sections passing through the head-kidney at our third stage. Zeiss C, ocul. 2. Our figures are representations of the following sections of the series, section 1 being the first which passes through the anterior groove of the head-kidney.

No. 1 SECTION 3. No. 8 SECTION 13. " 2 " 4. " 9 " 15. " 3 " 5. " 10 " 16. " 4 " 6. " 11 " 17. " 5 " 8. " 12 " 18. " 6 " 10. " 13 " 19. " 7 " 11. " 14 " 20.

The Müllerian duct extends through eleven more sections.

The first groove (_gr_1.) extends to No. 3.

The second groove (_gr_2.) extends from No. 4 to No. 7.

The third groove (_gr_3.) extends from No. 11 to No. 13.

The first ridge (_r_1.) extends from No. 2 to No. 5.

The second ridge (_r_2.) extends from No. 8 to No. 11.

The third ridge (_r_3.) extends from No. 13 backwards through twelve sections, when it terminates by a pointed extremity.

FIG. C. Section through the ridge connecting the second and third grooves of the head-kidney of an embryo slightly younger than that from which Series B was taken. Zeiss C, ocul. 3 (reduced one-third).

The fold of the germinal epithelium, which gives rise to a deep groove (_x._) external to the head-kidney is well marked.

SERIES G. Sections through the rod of cells constituting the termination of the Müllerian duct at a stage in which the head-kidney is still present. Zeiss C, ocul. 2.

PLATE 28.

SERIES D. Sections chosen at intervals from a complete series traversing the peritoneal opening of the Müllerian duct, the remnant of the head-kidney, and the termination of the Müllerian duct. Zeiss C, ocul. 3 (reduced one-third).

Nos. 1 and 2. Sections through the persistent anterior opening of the head-kidney (abdominal opening of Müllerian duct). The approach of the Wolffian duct to the groove may be seen by a comparison of these two figures. In the sections in front of these (not figured) the two are much more widely separated than in No. 1.

No. 3. Section through the Müllerian duct, just posterior to the persistent opening.

Nos. 4 and 5. Remains of the ridges, which at an earlier stage connected the first and second grooves, are seen passing from the Müllerian duct to the peritoneal epithelium.

No. 6. Rudiment of the second groove (_gr_2.) of the head-kidney.

Between 6 and 7 is a considerable interval.

No. 7. All traces of this groove (_gr_2.) have vanished, and the Müllerian duct is quite disconnected from the epithelium.

No. 8. Rudiment of the third groove (_gr_3.).

No. 9. Müllerian duct quite free in the space between the peritoneal epithelium and the Wolffian duct, in which condition it extends until near its termination. Between Nos. 9 and 10 is an interval of eight sections.

No. 10. The penultimate section, in which the Müllerian duct is seen. A lumen cannot be clearly made out.

No. 11. The last section in which any trace of the Müllerian duct is visible. No line of demarcation can be seen separating the solid end of the Müllerian duct from the ventral wall of the Wolffian duct.

FIGS. E. and F. Sections through the glomerulus of the head-kidney from an embryo prior to the appearance of the head-kidney. Zeiss B, ocul. 2. A comparison of the two figures shows the variation in the thickness of the stalk of the glomerulus. E. Section anterior to the foremost Malpighian body. F. Section through both the glomerulus of the head-kidney and that of a Malpighian body. The two are seen to be connected.

SERIES H. Consecutive sections through the hind end of the Müllerian duct, from an embryo in which the head-kidney was only represented by a rudiment. (The embryo was, perhaps, very slightly older than that from which Series D was taken.) Zeiss C, ocul. 3 (reduced one-third).

No. 1. Müllerian duct is without a lumen, and quite distinct from the Wolffian wall.

No. 2. The solid end of the Müllerian duct is no longer distinct from the internal wall of the Wolffian duct.

No. 3. All trace of the Müllerian duct has vanished.

SERIES I. Sections through the hinder end of the Müllerian duct from an embryo of about the middle of the sixth day. Zeiss C, ocul. 2 (reduced one-third).

No. 1. The Müllerian duct is distinct and small.

No. 2. Is posterior by twelve sections to No. 1. The Müllerian duct is dilated, and its cells are vacuolated.

No. 3. Penultimate section, in which the Müllerian duct is visible; it is separated by three sections from No. 2.

No. 4. Last section in which any trace of the Müllerian duct is visible; the lumen, which was visible in the previous section, is now absent.

No. 5. No trace of Müllerian duct. Nos. 3, 4, and 5 are consecutive sections.

FIG. K. Section through the hind end of the abdominal opening of the Müllerian duct of a chick of 123 hours. Zeiss C, ocul. 2 (reduced one-third). It illustrates the peculiar cord connecting the Müllerian and Wolffian ducts.

XIV. ON THE EARLY DEVELOPMENT OF THE LACERTILIA, TOGETHER WITH SOME OBSERVATIONS ON THE NATURE AND RELATIONS OF THE PRIMITIVE STREAK[446].

Footnote 446: From the _Quarterly Journal of Microscopical Science_, Vol. XIX. 1879.

(With Plate 29.)

Till quite recently no observations were recorded on the early developmental changes of the reptilian ovum. Not long ago Professors Kupffer and Benecke published a preliminary note on the early development of _Lacerta agilis_ and _Emys Europea_[447]. I have myself also been able to make some observations on the embryo of _Lacerta muralis_. The number of my embryos has been somewhat limited, and most of those which I have had have been preserved in bichromate of potash, which has turned out a far from satisfactory hardening reagent. In spite of these difficulties I have been led on some points to very different results from those of the German investigators, and to results which are more in accordance with what we know of other Sauropsidan types. I commence with a short account of the results of Kupffer and Benecke.

Footnote 447: _Die Erste Entwicklungsvorgänge am Ei der Reptilien_, Königsberg, 1878.

Segmentation takes place exactly as in birds, and the resulting blastoderm, which is thickened at its edge, spreads rapidly over the yolk. Shortly before the yolk is half enclosed a small embryonic shield (area pellucida) makes its appearance in the centre of the blastoderm, which has, in the meantime, become divided into two layers. The upper of these is the epiblast, and the lower the hypoblast. The embryonic shield is mainly distinguished from the remainder of the blastoderm by the more columnar character of its constituent epiblast cells. It is somewhat pyriform in shape, the narrower end corresponding with the future posterior end of the embryo. At the narrow end an invagination takes place, which gives rise to an open sac, the blind end of which is directed forwards. The opening of this sac is regarded by the authors as the blastopore. A linear thickening of epiblast arises in front of the blastopore, along the median line of which the medullary groove soon appears. In the caudal region the medullary folds spread out and enclose between them the blastopore, behind which they soon meet again. On the conversion of the medullary groove into a closed canal the blastopore becomes obliterated. The mesoblast grows out from the lip of the blastopore as four masses. Two of these are lateral: a third is anterior and median, and, although at first independent of the epiblast, soon attaches itself to it, and forms with it a kind of axis-cord. A fourth mass applied itself to the walls of the sac formed by invagination.

With reference to the very first developmental phenomena my observations are confined to two stages during the segmentation[448]. In the earliest of these the segmentation was about half completed, in the later one it was nearly over. My observations on these stages bear out generally the statements of Kupffer and Benecke. In the second of them the blastoderm was already imperfectly divided into two layers--a superficial epiblastic layer formed of a single row of cells, and a layer below this several rows deep. Below this layer fresh segments were obviously being added to the blastoderm from the subjacent yolk.

Footnote 448: For these two specimens, which were hardened in picric acid, I am indebted to Dr Kleinenberg.

Between the second of these blastoderms and my next stage there is a considerable gap. The medullary plate is just established, and is marked by a shallow groove which becomes deeper in front. A section through the embryo is represented in Pl. 29, Series A, fig. 1. In this figure there may be seen the thickened medullary plate with a shallow medullary groove, below which are two independent plates of mesoblast (_me.p._), one on each side of the middle line, very imperfectly divided into somatopleuric and splanchnopleuric layers. Below the mesoblast is a continuous layer of hypoblast (_hy._), which develops a rod-like thickening along the axial line (_ch._). This rod becomes in the next stage the notochord. Although this embryo is not well preserved I feel very confident in asserting the continuity of the notochord with the hypoblast at this stage.

At the hind end of the embryo is placed a thickened ridge of tissue which continues the embryonic axis. In this ridge all the layers coalesce, _and I therefore take it to be equivalent to the primitive streak of the avian blastoderm_. It is somewhat triangular in shape, with the apex directed backward, the broad base placed in front.

At the junction between the primitive streak and the blastoderm is situated a passage, open at both extremities, leading from the upper surface of the blastoderm obliquely forwards to the lower.

The dorsal and anterior wall of this passage is formed of a distinct epithelial layer, continuous at its upper extremity with the epiblast, and at its lower with the notochordal plate, so that it forms a layer of cells connecting together the epiblast and hypoblast. The hinder and lower wall of the passage is formed by the cells of the primitive streak, which only assume a columnar form near the dorsal opening of the passage (vide fig. 4). This passage is clearly the blind sac of Kupffer and Benecke, who, if I am not mistaken, have overlooked its lower opening. As I hope to show in the sequel, it is also the equivalent of the neurenteric passage, which connects the neural and alimentary canals in the Ichthyopsida, and therefore represents the blastopore of Amphioxus, Amphibians, &c.

Series A, figs. 2, 3, 4, 5, illustrate the features of the passage and its relation to the embryo.

Fig. 2 passes through the ventral opening of the passage. The notochordal plate (_ch´._) is vaulted over the opening, and on the left side is continuous with the mesoblast as well as the hypoblast. Figs. 3 and 4 are taken through the middle part of the passage (_ne._), which is bounded above by a continuation of the notochordal plate, and below by the tissue of the primitive streak. The hypoblast (_hy._), in the middle line, is imperfectly fused with the mesoblast of the primitive streak, which is now continuous across the middle line. The medullary groove has disappeared, but the medullary plate (_mp._) is quite distinct.

In fig. 5 is seen the dorsal opening of the passage (_ne._). If a section behind this had been figured, as is done for the next series (B), it would have passed through the primitive streak, and, as in the chick, all the layers would have been fused together. The epiblast in the primitive streak completely coalesces with the mesoblast; but the hypoblast, though attached to the other layers in the middle line, can always be traced as a distinct stratum.

Fig. B is a surface view of my next oldest embryo. The medullary groove has become much deeper, especially in front. Behind it widens out to form a space equivalent to the sinus rhomboidalis of the embryo bird. The amnion forms a small fold covering over the cephalic extremity of the embryo, which is deeply embedded in the yolk. Some somites (protovertebræ) were probably present, but this could not be made out in the opaque embryo.

The woodcut (fig. 1) represents a diagrammatic longitudinal section through this embryo, and the sections belonging to Series B illustrate the features of the hind end of the embryo and of the primitive streak.

As is shown in fig. 1, the notochord (_ch._) has now throughout the region of the embryo become separated from the subjacent hypoblast, and the lateral plates of mesoblast are distinctly divided into somatic and splanchnic layers. The medullary groove is continued as a deepish groove up to the opening of the neurenteric passage, which thus forms a perforation in the floor of the hinder end of the medullary groove (vide Series B, figs. 2, 3, and 4).

The passage itself is somewhat shorter than in the previous stage, and the whole of it is shown in a single section (fig. 4). This section must either have been taken somewhat obliquely, or else the passage have been exceptionally short in this embryo, since in an older embryo it could not all be seen in one section.

The front wall of the passage is continuous with the notochord, which for two sections or so in front remains attached to the hypoblast (figs. 2 and 3). Behind the perforation in the floor of the medullary groove is placed the primitive streak (fig. 5), where all the layers become fused together, as in the earlier stage. Into this part a narrow diverticulum from the end of the medullary groove is continued for a very short distance (vide fig. 5, _mc._).

The general features of the stage will best be understood by an examination of the diagrammatic longitudinal section, represented in woodcut, fig. 1. In front is shown the amnion (_am._), growing over the head of the embryo. The notochord (_ch._) is seen as an independent cord for the greater part of the length of the embryo, but falls into the hypoblast shortly in front of the neurenteric passage. The neurenteric passage is shown at _ne._, and behind it is shown the primitive streak.

In a still older stage, represented in surface view on Pl. 29, fig. C, the medullary folds have nearly met above, but have not yet united. The features of the passage from the neural groove to the hypoblast are precisely the same in the embryo just described, although the lumen of the passage has become somewhat narrower. There is still a short primitive streak behind the embryo.

The neurenteric passage persists but a very short time after the complete closure of the medullary canal. It is in no way connected with the allantois, as conjectured by Kupffer and Benecke, but the allantois is formed, as I have satisfied myself by longitudinal sections of a later stage, in the manner already described by Dobrynin, Gasser, and Kölliker for the bird and mammal.

The general results of Kupffer's and Benecke's observations, with the modifications introduced by my own observations, are as follows:--After the segmentation and the formation of the embryonic shield (area pellucida) the blastoderm becomes distinctly divided into epiblast and hypoblast[449]. At the hind end of the shield a somewhat triangular primitive streak is formed by the fusion of the epiblast and hypoblast with a number of cells between them, which are probably derived from the lower rows of the segmentation cells. At the front end of the streak a passage arises, open at both extremities, leading obliquely forwards through the epiblast to the space below the hypoblast. The walls of the passage are formed of a layer of columnar cells continuous both with epiblast and hypoblast. In front of the primitive streak the body of the embryo becomes first differentiated by the formation of a medullary plate, and at the same time there grows out from the primitive streak a layer of mesoblast, which spreads out in all directions between the epiblast and hypoblast. In the axis of the embryo the mesoblast plate is stated by Kupffer and Benecke to be continuous across the middle line, but this appears very improbable. In a slightly later stage the medullary plate becomes marked by a shallow groove, and the mesoblast of the embryo is then undoubtedly constituted of two lateral plates, one on each side of the median line. In the median line the notochord arises as a ridge-like thickening of the hypoblast, which becomes very soon quite separated from the hypoblast, except at the hind end, where it is continued into the front wall of the neurenteric passage. It is interesting to notice the remarkable relation of the notochord to the walls of the neurenteric passage. More or less similar relations are also well marked in the case of the goose and the fowl (Gasser)[450], and support the conclusion deducible from the lower forms of vertebrata, that the notochord is essentially hypoblastic.

Footnote 449: This appears to me to take place before the formation of the embryonic shield.

Footnote 450: Gasser, _Der Primitivstreifen bei Vogelembryonen_, Marburg, 1878.

The passage at the front end of the primitive streak forms the posterior boundary of the medullary plate, though the medullary groove is not at first continued back to it. The anterior wall of this passage connects together the medullary plate and the notochordal ridge of the hypoblast. In the succeeding stages the medullary groove becomes continued back to the opening of the passage, which then becomes enclosed in the medullary folds, and forms a true neurenteric passage. It becomes narrowed as the medullary folds finally unite to form the medullary canal, and eventually disappears.

I conclude this paper with a concise statement of what appears to me the probable nature of the much-disputed organ, the primitive streak, and of the arguments in support of my view.

In a paper on the primitive streak in the _Quart. Journ. of Mic. Sci._, in 1873 (p. 280) [This edition, p. 45], I made the following statement with reference to this subject:--"It is clear, therefore, that the primitive groove must be the rudiment of some ancestral feature.... It is just possible that it is the last trace of that involution of the epiblast by which the hypoblast is formed in most of the lower animals."

At a later period, in July, 1876, after studying the development of Elasmobranch fishes, I enlarged the hypothesis in a review of the first part of Prof. Kölliker's _Entwicklungsgeschichte_. The following is the passage in which I speak of it[451]:

Footnote 451: _Journal of Anat. and Phys._, Vol. X. pp. 790 and 791. Compare also my _Monograph on Elasmobranch Fishes_, note on p. 68 [This edition, p. 281].

"In treating of the exact relation of the primitive groove to the formation of the embryo, Professor Kölliker gives it as his view that though the head of the embryo is formed independently of the primitive groove, and only secondarily unites with this, yet that the remainder of the body is without doubt derived from the primitive groove. With this conclusion we cannot agree, and the very descriptions of Professor Kölliker appear to us to demonstrate the untenable nature of his results. We believe that the front end of the primitive groove at first occupies the position eventually filled by about the third pair of protovertebræ, but that as the protovertebræ are successively formed, and the body of the embryo grows in length, the primitive groove is carried further and further back, so as always to be situated immediately behind the embryo. As Professor Kölliker himself has shewn it may still be seen in this position even later than the fortieth hour of incubation.

"Throughout the whole period of its existence it retains a character which at once distinguishes it in sections from the medullary groove.

"Beneath it the epiblast and mesoblast are _always fused_, though they are always separate elsewhere; this fact, which was originally shewn by ourselves, has been very clearly brought out by Professor Kölliker's observations.

"The features of the primitive groove which throw special light on its meaning are the following:--

"(1) It does not enter directly into the formation of the embryo.

"(2) The epiblast and mesoblast always become fused beneath it.

"(3) It is situated immediately behind the embryo.

"Professor Kölliker does not enter into any speculations as to the meaning of the primitive groove, but the above-mentioned facts appear to us clearly to prove that the primitive groove is a rudimentary structure, the origin of which can only be completely elucidated by a knowledge of the development of the Avian ancestors.

"In comparing the blastoderm of a bird with that of any anamniotic vertebrate, we are met at the threshold of our investigations by a remarkable difference between the two. Whereas in all the lower vertebrates the embryo is situated at the _edge_ of the blastoderm, it is in birds and mammals situated in the centre. This difference of position at once suggests the view that the primitive groove may be in some way connected with the change of position in the blastoderm which the ancestors of birds must have undergone. If we carry our investigations amongst the lower vertebrates a little further, we find that the Elasmobranch embryo occupies at first the normal position at the edge of the blastoderm, but that in the course of development the blastoderm grows round the yolk far more slowly in the region of the embryo than elsewhere. Owing to this, the embryo becomes left in a bay, the two sides of which eventually meet and coalesce in a linear fashion immediately behind the embryo, thus removing the embryo from the edge of the blastoderm and forming behind it a linear streak not unlike the primitive streak. We would suggest the hypothesis that the primitive groove is a rudiment which gives the last indication of a change made by the Avian ancestors in their position in the blastoderm, like that made by Elasmobranch embryos when removed from the edge of the blastoderm and placed in a central situation similar to that of the embryo bird. On this hypothesis the situation of the primitive groove immediately behind the embryo, as well as the fact of its not becoming converted into any embryonic organ would be explained. The central groove might probably also be viewed as the groove naturally left between the coalescing edges of the blastoderm.

"Would the fusion of epiblast and mesoblast also receive its explanation on this hypothesis? We are of opinion that it would. At the edge of the blastoderm which represents the blastopore mouth of Amphioxus all the layers become fused together in the anamniotic vertebrates. So that if the primitive groove is in reality a rudiment of the coalesced edges of the blastoderm, we might naturally expect the layers to be fused there, and the difficulty presented by the present condition of the primitive groove would rather be that the hypoblast is not fused with the other layers than that the mesoblast is indissolubly united with the epiblast. The fact that the hypoblast is not fused with the other layers does not appear to us to be fatal to our hypothesis, and in Mammalia, where the primitive and medullary grooves present precisely the same relations as in birds, all three layers are, according to Hensen's account, fused together. This, however, is denied by Kölliker, who states that in Mammals, as in Birds, only the epiblast and mesoblast fuse together. Our hypothesis as to the origin of the primitive groove appears to explain in a fairly satisfactory manner all the peculiarities of this very enigmatical organ; it also relieves us from the necessity of accepting Professor Kölliker's explanation of the development of the mesoblast, though it does not, of course, render that explanation in any way untenable."

At a somewhat later period Rauber arrived at a more or less similar conclusion, which, however, he mixes up with a number of opinions from which I am compelled altogether to dissent[452].

Footnote 452: "Primitivrinne u. Urmund," _Morphologisches Jahrbuch_, Band II. p. 551.

The general correctness of my view, as explained in my second quotation, appears to me completely established by Gasser's beautiful researches on the early development of the chick and goose[453], and by my own observations just recorded on the lizard. While at the same time the parallel between the blastopore of Elasmobranchii and of the Sauropsida, is rendered more complete by the discovery of the neurenteric passage in the latter group, which was first of all made by Gasser.

Footnote 453: Gasser, _Der Primitivstreifen bei Vogelembryonen_, Marburg, 1878.

The following paragraphs contain a detailed attempt to establish the above view by a careful comparison of the primitive streak and its adjuncts in the amniotic vertebrates with the blastopore in Elasmobranchii.

In Elasmobranchii the blastopore consists of the following parts:--(1), a section at the end of the medullary plate, which becomes converted into the neurenteric canal[454]; (2), a section forming what may be called the yolk blastopore, which eventually constitutes a linear streak connecting the embryo with the edge of the blastoderm (vide monograph on Elasmobranch fishes, pp. 281 and 296). In order to establish my hypothesis on the nature of the primitive streak, it is necessary to find the representatives of both these parts in the primitive streak of the amniotic vertebrates. The first section ought to appear as a passage from the neural to the enteric side of the blastoderm at the posterior end of the medullary plate. At its front edge the epiblast and hypoblast should be continuous, as they are at the hind end of the embryo in Elasmobranchii, and, finally, the passage should, on the closure of the medullary groove, become converted into the _neurenteric canal_. All these conditions are exactly fulfilled by the opening at the front end of the primitive streak of the lizard (vide woodcut, fig. 1, p. 647). In the chick there is at first no such opening, but, as I hope to shew in a future paper, it is replaced by the epiblast and hypoblast falling into one another at the front end of the primitive streak. At a later period, as has been shewn by Gasser[455], there is a distinct rudiment of the neurenteric canal in the chick, and a complete canal in the goose. Finally, in mammals, as has been shewn by Schäffer[456] for the guinea-pig, there is at the front end of the primitive streak a complete continuity between epiblast and hypoblast. The continuity of the epiblast and hypoblast at the hind end of the embryo in the bird and the mammal is a rudiment of the continuity of these layers at the dorsal lip of the blastopore in Elasmobranchii, Amphibia, &c. The second section of the blastopore in Elasmobranchii or yolk blastopore is, I believe, partly represented by the primitive streak. The yolk blastopore in Elasmobranchii is the part of the blastopore belonging to the yolk sac as opposed to that belonging to the embryo, and it is clear that the primitive streak cannot correspond to the whole of this, since the primitive streak is far removed from the edge of the blastoderm long before the yolk is completely enclosed. Leaving this out of consideration the primitive streak, in order that the above comparison may hold good, should satisfy the following conditions:

Footnote 454: I use this term for the canal connecting the neural and alimentary tract, which was first discovered by Kowalevsky.

Footnote 455: _Loc. cit._

Footnote 456: "A contribution to the history of the development in the Guinea-pig," _Journal of Anat. and Phys._ Vol. XI. pp. 332-336.

1. It should connect the embryo with the edge of the blastoderm.

2. It should be constituted as if formed of the fused edges of the blastoderm.

3. The epiblast of it should eventually not form part of the medullary plate of the embryo, but be folded over on to the ventral side.

The first of these conditions is only partially fulfilled, but, considering the rudimentary condition of the whole structure, no great stress can, it seems to me, be laid on this fact.

The second condition seems to me very completely satisfied. Where the two edges of the blastoderm become united we should expect to find a complete fusion of the layers such as takes place in the primitive streak; and the fact that in the primitive streak the hypoblast does not so distinctly coalesce with the mesoblast as the mesoblast with the epiblast cannot be urged as a serious argument against me.

The growth outwards of the mesoblast from the axis of the primitive streak is probably a remnant of the invagination of the hypoblast and mesoblast from the lip of the blastopore in Amphibia, &c.

The groove in the primitive streak may with great plausibility be regarded as the indication of a depression which would naturally be found along the line where the thickened edges of the blastoderm became united.

With reference to the third condition, I will make the following observations. The neurenteric canal, as it is placed at the extreme end of the embryo, must necessarily, with reference to the embryo, be the hindermost section of the blastopore, and therefore the part of the blastopore apparently behind this can only be so owing to the embryo not being folded off from the yolk sac; and as the yolk sac is in reality a specialised part of the ventral wall of the body, the yolk blastopore must also be situated on the ventral side of the embryo.

Kölliker and other distinguished embryologists have believed that the epiblast of the whole of the primitive streak became part of the neural plate. If this view were correct, which is accepted even by Rauber, the hypothesis I am attempting to establish would fall to the ground. I have, however, no doubt that these embryologists are mistaken. The very careful observations of Gasser shew that the part of the primitive streak adjoining the embryo becomes converted into the tail-swelling, and that the posterior part is folded in on the ventral side of the embryo, and, losing its characteristic structure, forms part of the ventral wall of the body. On this point my own observations confirm those of Gasser. In the lizard the early appearance of the neurenteric canal at the front end of the primitive streak clearly shews that here also the primitive streak can take no share in forming the neural plate.

The above considerations appear to me sufficient to establish my hypothesis with reference to the nature of the primitive streak, which has the merit of explaining, not only the structural peculiarities of the primitive streak, but also the otherwise inexplicable position of the embryo of the amniotic vertebrates in the centre of the blastoderm.

DESCRIPTION OF PLATE 29.

COMPLETE LIST OF REFERENCE LETTERS.

_am._ Amnion. _ch._ Notochord. _ch´._ Notochordal thickening of hypoblast. _ep._ Epiblast. _hy._ Hypoblast. _m.g._ Medullary groove. _me.p._ Mesoblastic plate. _ne._ Neurenteric canal (blastopore). _pr._ Primitive streak.

SERIES A. Sections through an embryo shortly after the formation of the medullary groove. x 120[457].

Footnote 457: The spaces between the layers in these sections are due to the action of the hardening reagent.

Fig. 1. Section through the trunk of the embryo.

Figs. 2-5. Sections through the neurenteric canal.

Fig. B. Surface view of a somewhat older embryo than that from which Series A is taken. x 30.

SERIES B. Sections through the embryo represented in Fig. B. x 120.

Fig. 1. Section through the trunk of the embryo.

Figs. 2, 3. Sections through the hind end of the medullary groove.

Fig. 4. Section through the neurenteric canal.

Fig. 5. Section through the primitive streak.

Fig. C. Surface view of a somewhat older embryo than that represented in Fig. B. x 30.

XV. ON CERTAIN POINTS IN THE ANATOMY OF PERIPATUS CAPENSIS[458].

Footnote 458: From the _Proceedings of the Cambridge Philosophical Society_, Vol. III. 1879.

The discovery by Mr Moseley[459] of a tracheal system in Peripatus must be reckoned as one of the most interesting results obtained by the naturalists of the "Challenger." The discovery clearly proves that the genus Peripatus, which is widely distributed over the globe, is the persisting remnant of what was probably a large group of forms, from which the present tracheate Arthropoda are descended.

Footnote 459: "On the Structure and Development of _Peripatus Capensis_," _Phil. Trans._, Vol. CLXIV. 1874.

The affinities of Peripatus render any further light on its anatomy a matter of some interest; and through the kindness of Mr Moseley I have had an opportunity of making investigations on some well preserved examples of _Peripatus capensis_, a few of the results of which I propose to lay before the Society.

I shall confine my observations to three organs. (1) The segmental organs, (2) the nervous system, (3) the so-called fat bodies of Mr Moseley.

In all the segments of the body, with the exception of the first two or three postoral ones, there are present glandular bodies, apparently equivalent to the segmental organs of Annelids.

These organs have not completely escaped the attention of previous observers. The anterior of them were noticed by Grube[460], but their relations were not made out. By Saenger[461], as I gather from Leuckart's _Bericht_ for the years 1868-9, these structures were also noticed, and they were interpreted as segmental organs. Their external openings were correctly identified. They are not mentioned by Moseley, and no notice of them is to be found in the text-books. The observations of Grube and Saenger seem, in fact, to have been completely forgotten.

Footnote 460: "Bau von _Perip. Edwardsii_," _Archiv f. Anat. u. Phys._ 1853.

Footnote 461: _Moskauer Naturforscher Sammlung_, Abth. Zool. 1869.

The organs are placed at the bases of the feet in two lateral divisions of the body-cavity shut off from the main central median division of the body-cavity by longitudinal septa of transverse muscles.

Each fully developed organ consists of three parts:

(1) A dilated vesicle opening externally at the base of a foot.

(2) A coiled glandular tube connected with this and subdivided again into several minor divisions.

(3) A short terminal portion opening at one extremity into the coiled tube (2) and at the other, as I believe, into the body-cavity. This section becomes very conspicuous in stained preparations by the intensity with which the nuclei of its walls absorb the colouring matter.

The segmental organs of Peripatus, though formed on a type of their own, more nearly resemble those of the Leech than of any other form with which I am acquainted. The annelidan affinities shewn by their presence are of some interest. Around the segmental organs in the feet are peculiar cells richly supplied with tracheæ, which appear to me to be similar to the fat bodies in insects. There are two glandular bodies in the feet in addition to the segmental organs.

The more obvious features of the nervous system have been fully made out by previous observers, who have shewn that it consists of large paired supra-oesophageal ganglia connected with two widely separated ventral cords--stated by them not to be ganglionated. Grube describes the two cords as falling into one another behind the anus--a feature the presence of which is erroneously denied by Saenger. The lateral cords are united by numerous (5 or 6 for each segment) transverse cords.

The nervous system would appear at first sight to be very lowly organised, but the new points I believe myself to have made out, as well as certain previously known features in it appear to me to shew that this is not the case.

The following is a summary of the fresh points I have observed in the nervous system:

(1) Immediately underneath the oesophagus the oesophageal commissures dilate and form a pair of ganglia equivalent to the annelidan and arthropodan sub-oesophageal ganglia. These ganglia are closely approximated and united by 5 or 6 commissures. They give off large nerves to the oral papillæ.

(2) The ventral nerve cords are covered on their ventral side by a thick ganglionic layer[462], and at each pair of feet they dilate into a small but distinct _ganglionic swelling_. From each ganglionic swelling are given off a pair of large nerves[463] to the feet; and the ganglionic swellings of the two cords are connected together by _a pair of commissures containing ganglion cells_[464]. The other commissures connecting the two cords together do not contain ganglion cells.

Footnote 462: This was known to Grube, _loc. cit._

Footnote 463: These nerves were noticed by Milne-Edwards, but Grube failed to observe that they were much larger than the nerves given off between the feet.

Footnote 464: These commissures were perhaps observed by Saenger, _loc. cit._

The chief feature in which Peripatus was supposed to differ from normal Arthropoda and Annelida, viz. the absence of ganglia on the ventral cords, does not really exist. In other particulars, as in the amount of nerve cells in the ventral cords and the completeness of the commissural connections between the two cords, &c., the organisation of the nervous system of Peripatus ranks distinctly high. The nervous system lies within the circular and longitudinal muscles, and is thus not in proximity with the skin. In this respect also Peripatus shews no signs of a primitive condition of the nervous system.

A median nerve is given off from the posterior border of the supra-oesophageal ganglion to the oesophagus, which probably forms a rudimentary sympathetic system. I believe also that I have found traces of a paired sympathetic system.

The organ doubtfully spoken of by Mr Moseley as a fat body, and by Grube as a lateral canal, is in reality a glandular tube, lined by beautiful columnar cells containing secretion globules, which opens by means of a non-glandular duct into the mouth. It lies close above the ventral nerve cords in a lateral compartment of the body-cavity, and extends backwards for a varying distance.

This organ may perhaps be best compared with the simple salivary gland of Julus. It is not to be confused with the slime glands of Mr Moseley, which have their opening in the oral papillæ. If I am correct in regarding it as homologous with the salivary glands so widely distributed amongst the Tracheata, its presence indicates a hitherto unnoticed arthropodan affinity in Peripatus.

XVI. ON THE MORPHOLOGY AND SYSTEMATIC POSITION OF THE SPONGIDA[465].

Footnote 465: From the _Quarterly Journ. of Microscopical Science_, Vol. XIX. 1879.

Professor Schulze's[466] last memoir on the development of Calcareous Sponges, confirms and enlarges Metschnikoff's[467] earlier observations, and gives us at last a fairly complete history of the development of one form of Calcareous Sponge. The facts which have been thus established have suggested to me a view of the morphology and systematic position of the Spongida, somewhat different to that now usually entertained. In bringing forward this view, I would have it understood that it does not claim to be more than a mere suggestion, which if it serves no other function may, perhaps, be of use in stimulating research.

Footnote 466: "Untersuchungen über d. Bau u. d. Entwicklung der Spongien," _Zeit. f. wiss. Zool._ Bd. XXXI. 1878.

Footnote 467: "Zur Entwicklungsgeschichte der Kalkschwämme," _Zeit. f. wiss. Zool._ Bd. XXIV. 1874.

To render clear what I have to say, I commence with a very brief statement of the facts which may be considered as established with reference to the development of _Sycandra raphanus_, the form which was studied by both Metschnikoff and Schulze. The segmentation of the ovum, though in many ways remarkable, is of no importance for my present purpose, and I take up the development at the close of the segmentation, while the embryo is still encapsuled in the parental tissues. It is at this stage lens-shaped, with a central segmentation cavity. An equatorial plane divides it into two parts, which have equal shares in bounding the segmentation cavity. One of these halves is formed of about thirty-two large, round, granular cells, the other of a larger number of ciliated clear columnar cells. While the embryo is still encapsuled a partial invagination of the granular cells takes place, reducing the segmentation cavity to a mere slit; this invagination is, however, quite temporary and unimportant, and on the embryo becoming free, which shortly takes place, no trace of it is visible; but, on the contrary, the segmentation cavity becomes larger, and the granular cells project very much more prominently than in the encapsuled state.

The larva, after it has left the parental tissues, has an oval form and is transversely divided into two areas (fig. 1, A). One of these areas is formed of the elongated, clear, ciliated cells, with a small amount of pigment near the inner ends (_en_), and the other and larger area of the thirty-two granular cells already mentioned (_ec_). Fifteen or sixteen of these are arranged as a special ring on the border of the clear cells. In the centre of the embryo is a segmentation cavity (_cs_) which lies between the granular and the clear cells, but is mainly bounded by the vaulted inner surface of the latter. This stage is known as the amphiblastula stage. After the larva has for some time enjoyed a free existence, a remarkable series of changes takes place, which result in the invagination of the half of it formed of the clear cells, and form a prelude to the permanent attachment of the larva. The entire process of invagination is completed in about half an hour. The whole embryo first becomes flattened, but especially the ciliated half which gradually becomes less prominent (fig. 1, B), and still later the cells composing it undergo a true process of invagination. As a result of this invagination the segmentation cavity is obliterated and the larva assumes a compressed plano-convex form with a central gastrula cavity, and a blastopore in the middle of the flattened surface. The two layers of the gastrula may now be spoken of as ectoderm and entoderm. The blastopore becomes gradually narrowed by the growth over it of the outer row of granular cells. When it has become very small the attachment of the larva takes place by the flat surface where the blastopore is situated. It is effected by protoplasmic processes of the outer ring of ectoderm cells, which, together with the other ectoderm cells, now become amoeboid. At the same time they become clearer and permit a view of the interior of the gastrula. Between the ectoderm cells and the entoderm cells which line the gastrula cavity there arises a hyaline structureless layer, which is more closely attached to the ectoderm than to the entoderm, and is probably derived from the former. A view of the gastrula stage after the larva has become fixed is given in fig. 2.

After invagination the cilia of the entoderm cells can no longer be seen, and are probably absorbed, and their disappearance is nearly coincident with the complete obliteration of the blastopore, an event which takes place shortly after the attachment of the larva. After the formation of the structureless layer between the ectoderm and entoderm, calcareous spicules make their appearance in it as delicate unbranched rods pointed at both extremities. The larva when once fixed rapidly grows in length and assumes a cylindrical form (fig. 3, A). The sides of the cylinder are beset with calcareous spicules which project beyond the surface, and in addition to the unbranched forms, spicules are developed with three and four rays as well as some with a blunt extremity and serrated edge. The extremity of the cylinder opposite the attached surface is flattened, and though surrounded by a ring of four-rayed spicules is itself free from them. At this extremity a small perforation is formed leading into the gastric cavity which rapidly increases in size and forms an exhalent osculum (_os_). A series of inhalent apertures are also formed at the sides of the cylinder. The relative times of appearance of the single osculum and smaller apertures is not constant for the different larvæ. On the central gastrula cavity of the sponge becoming placed in communication with the external water, the entoderm cells lining it become ciliated afresh (fig. 3, B, _en_) and develop the peculiar collar characteristic of the entoderm cells of the Spongida. When this stage of development is reached we have a fully developed sponge of the type made known by Haeckel as Olynthus.

Till the complete development of other forms of Spongida has been worked out it is not possible to feel sure how far the phenomena observable in Sycandra hold good in all cases. Quite recently the Russian embryologist, M. Ganin[468], has given an account, without illustrations, of the development of _Spongilla fluviatilis_, which does not appear reconcileable with that of Sycandra. Considering the difficulties of observation it appears better to assume for this and some other descriptions that the observations are in error rather than that there is a fundamental want of uniformity in development amongst the Spongida.

Footnote 468: "Zur Entwicklung d. Spongilla fluviatilis," _Zoologischer Anzeiger_, Vol. I. No. 9, 1878.

The first point in the development of Sycandra which deserves notice is the character of the free swimming larva. The peculiar larval form, with one half of the body composed of amoeboid granular cells and the other of clear ciliated cells is nearly constant amongst the Calcispongiæ, and widely distributed in a somewhat modified condition amongst the Fibrospongiæ and Myxospongiæ. Does this larva retain the characters of an ancestral type of the Spongida, and if so what does its form mean? It is, of course, possible that it has no ancestral meaning but has been secondarily acquired; I prefer myself to think that this is not the case, more especially as it appears to me that the characters of the larva may be plausibly explained by regarding it as a transitional form between the Protozoa and Metazoa. According to this view the larva is to be considered as a colony of Protozoa, one half of the individuals of which have become differentiated into nutritive forms, and the other half into locomotor and respiratory forms. The granular amoeboid cells represent the nutritive forms, and the ciliated cells represent the locomotor and respiratory forms. That the passage from the Protozoa to the Metazoa may have been effected by such a differentiation is not improbable on _à priori_ grounds, and fits in very well with the condition of the free swimming larva of Spongida, though another and perhaps equally plausible suggestion as to this passage has been put forward by my friend Professor Lankester[469].

Footnote 469: "Notes on Embryology and Classification." _Quarterly Journal of Microscopical Science_, Vol. XVII. 1877. It seems not impossible, if the speculations in this paper have any foundation that while the views here put forward as to the passage from the Protozoon to the Metazoon condition may hold true for the Spongida, some other mode of passage may have taken place in the case of the other Metazoa.

While the above view seems fairly satisfactory for the free swimming stage of the larval Sponge there arises in the subsequent development a difficulty which appears at first sight fatal to it. This difficulty is the invagination of the ciliated cells instead of the granular ones. If the granular cells represent the nutritive individuals of the colony, they and not the ciliated cells ought most certainly to give rise to the lining of the gastrula cavity, according to the generally accepted views of the morphology of the Spongida. The suggestion which I would venture to put forward in explanation of this paradox involves a completely new view of the nature and functions of the germinal layers of adult Sponges.

It is as follows:--When the free swimming ancestor of the Spongida became fixed, the ciliated cells by which its movements used to be effected must have to a great extent become functionless. At the same time the amoeboid nutritive cells would need to expose as large a surface as possible. In these two considerations there may, perhaps, be found a sufficient explanation of the invagination of the ciliated cells, and the growth of the amoeboid cells over them. Though respiration was, no doubt, mainly effected by the ciliated cells, it is improbable that it was completely localised in them, but the continuation of their function was provided for by the formation of an osculum and pores. The ciliated collared cells which line the ciliated chambers, or in some cases the radial tubes, are undoubtedly derived from the invaginated cells, and if there is any truth in the above suggestion, the collared cells in the adult Sponge must be mainly respiratory and not digestive in function, while the normal epithelial cells which cover the surface of the sponge, and in most cases line the greater part of the passages through its substance, must carry on the digestion[470]. If the reverse is the case the whole theory falls to the ground. It has not, so far as I know, been definitely made out where the digestion is carried on. Lieberkühn would appear to hold the view that the amoeboid lining cells of the passages are mainly concerned with digestion, while Carter holds that digestion is carried on by the collared cells of the ciliated chambers.

Footnote 470: That the flat cells which line the greater part of the passages of most Sponges are really derived from ectodermic invaginations appears to me clearly proved by Schulze's and Barrois' observations on the young fixed stages of Halisarea. Ganin appears, however, to maintain a contrary view for Spongilla.

If it is eventually proved by actual experiments on the nutrition of Sponges, that digestion is carried on by the general cells lining the passages, and not by the ciliated cells, it is clear that neither the ectoderm nor entoderm of Sponges will correspond with the similarly named layers in the Coelenterata and the Metazoa. The invaginated entoderm will be the respiratory layer and the ectoderm the digestive and sensory layer; the sensory function being probably mainly localised in the epithelium on the surface, and the digestive one in the epithelium lining the passages. Such a fundamental difference in the germinal layers between the Spongida and the other Metazoa, would necessarily involve the creation of a special division of the Metazoa for the reception of the former group.

XVII. NOTES ON THE DEVELOPMENT OF THE ARANEINA[471].

Footnote 471: From the _Quarterly Journ. of Microscopical Science_, Vol. XX. 1880.

(With Plates 30, 31, 32.)

The following observations do not profess to contain a complete history of the development even of a single species of spider. They are the result of investigations carried on at intervals during rather more than two years, on the ova of _Agelena labyrinthica_; and I should not have published them now, if I had any hope of being able to complete them before the appearance of the work I am in the course of publishing on Comparative Embryology. It appeared to me, however, desirable to publish in full such parts of my observations as are completed before the appearance of my treatise, since the account of the development of the Araneina is mainly founded upon them.

My investigations on the germinal layers and organs have been chiefly conducted by means of sections. To prepare the embryos for sections, I employed the valuable method first made known by Bobretzky. I hardened the embryos in bichromate of potash, after placing them for a short time in nearly boiling water. They were stained as a whole with hæmatoxylin after the removal of the membranes, and embedded for cutting in coagulated albumen.

The number of investigators who have studied the development of spiders is inconsiderable. A list of them is given at the end of the paper.

The earliest writer on the subject is Herold (No. 4); he was followed after a very considerable interval of time by Claparède (No. 3), whose memoir is illustrated by a series of beautiful plates, and contains a very satisfactory account of the external features of development.

Balbiani (No. 1) has gone with some detail into the history of the early stages; and Ludwig (No. 5) has published some very important observations on the development of the blastoderm. Finally, Barrois (No. 2) has quite recently taken up the study of the group, and has added some valuable observations on the development of the germinal layers.

In addition to these papers on the true spiders, important investigations have been published by Metschnikoff on other groups of the Arachnida, notably the scorpion. Metschnikoff's observations on the formation of the germinal layers and organs accord in most points with my own.

The development of the Araneina may be divided into four periods: (1) the segmentation; (2) the period from the close of the segmentation up to the period when the segments commence to be formed; (3) the period from the commencing formation of the segments to the development of the full number of limbs; (4) the subsequent stages up to the attainment of the adult form.

In my earliest stage the segmentation was already completed, and the embryo was formed of a single layer of large flattened cells enveloping a central mass of polygonal yolk-segments.

Each yolk-segment is formed of a number of large clear somewhat oval yolk-spherules. In hardened specimens the yolk-spherules become polygonal, and in ova treated with hot water prior to preservation are not unfrequently broken up. Amongst the yolk-segments are placed a fair number of nucleated bodies of a very characteristic appearance. Each of them is formed of (1) a large, often angular, nucleus, filled with deeply staining bodies (nucleoli?); (2) a layer of protoplasm surrounding the nucleus, prolonged into a protoplasmic reticulum. The exact relation of these nucleated bodies to the yolk-segments is not very easy to make out, but the general tendency of my observations is to shew (1) that each nucleated body belongs to a yolk-sphere, and (2) that it is generally placed not at the centre, but to one side of a yolk-sphere. If the above conclusions are correct each complete yolk-segment is a cell, and each such cell consists of a normal nucleus, protoplasm, and yolk-spherules. There is a special layer of protoplasm surrounding the nucleus, while the remainder of the protoplasm consists of a reticulum holding together the yolk-spherules. Yolk-cells of this character are seen in Pls. 31 and 32, figs. 10-21.

The nuclei of the yolk-cells are probably derived by division from the nuclei of the segmentation rosettes (vide Ludwig, No. 5), and it is probable that they take their origin at the time when the superficial layer of protoplasm separates from the yolk-columns below to form the blastoderm.

The protoplasm of the yolk-cells undergoes rapid division, as is shewn by the fact that there are often two nucleated bodies close together, and sometimes two nuclei in a single mass of protoplasm (fig. 10). It is probable that in some cases the yolk-spheres divide at the same time as the protoplasm belonging to them; the division of the nucleated bodies is, however, in the main destined to give rise to fresh cells which enter the blastoderm.

I have not elucidated to my complete satisfaction the next stage or two in the development of the embryo; and have not succeeded in completely reconciling the results of my own observations with those of Claparède and Balbiani. In order to shew exactly where my difficulties lie it is necessary briefly to state the results arrived at by the above authors.

According to Claparède the first differentiation in Pholcus consists in the accumulation of the cells over a small area to form a protuberance, which he calls the _primitive cumulus_. Owing to its smaller specific gravity the part of the ovum with the cumulus always turns upwards, like the blastodermic pole of a fowl's egg.

After a short time the cumulus elongates itself on one side, and becomes connected by a streak with a white patch, which appears on the surface of the egg, about 90° from the cumulus. This patch gradually enlarges, and soon covers the whole surface of the ovum except the region where the cumulus is placed. It becomes the ventral plate or germinal streak of the embryo, its extremity adjoining the cumulus is the anal extremity, and its opposite extremity the cephalic one. The cumulus itself is placed in a depression on the dorsal surface of the ovum. Claparède compares the cumulus to the dorsal organ of many Crustacea.

Balbiani (No. 1) describes the primitive cumulus in _Tegenaria domestica_, _Epeira diadema_, and _Agelena labyrinthica_, as originating as a protuberance at the centre of the ventral surface, surrounded by a specialised portion of the blastoderm (p. 57), which I will call the ventral plate. In _Tegenaria domestica_ he finds that it encloses the so-called yolk-nucleus, p. 62. By an unequal growth of the ventral plate the primitive cumulus comes to be placed at the cephalic pole of the ventral plate. The cumulus now becomes less prominent, and in a few cases disappears. In the next stage the central part of the ventral plate becomes very prominent and forms the procephalic lobe, close to the anterior border of which is usually placed the primitive cumulus (p. 67). The space between the cumulus and the procephalic lobe grows larger, so that the latter gradually travels towards the dorsal surface and finally vanishes. Behind the procephalic lobe the first traces of the segments make their appearance, as three transverse bands, before a distinct anal lobe becomes apparent.

The points which require to be cleared up are, (1) what is the nature of the primitive cumulus? (2) where is it situated in relation to the embryo? Before attempting to answer these questions I will shortly describe the development, so far as I have made it out, for the stages during which the cumulus is visible.

The first change that I find in the embryo (when examined after it has been hardened)[472] is the appearance of a small, whitish spot, which is at first very indistinct. A section through such an ovum (Pl. 31, fig. 10) shews that the cells of about one half of the ovum have become more columnar than those of the other half, and that there is a point (_pr.c._) near one end of the thickened half where the cells are more columnar, and about two layers or so deep. It appears to me probable that this point is the whitish spot visible in the hardened ovum. In a somewhat later stage (Pl. 30, fig. 1) the whitish spot becomes more conspicuous (_pc._), and appears as a distinct prominence, which is, without doubt, the primitive cumulus, and from it there proceeds on one side a whitish streak. The prominence, as noticed by Claparède and Balbiani, is situated on the flatter side of the ovum. Sections at this stage shew the same features as the previous stage, except that (1) the cells throughout are smaller, (2) those of the thickened hemisphere of the ovum more columnar, and (3) the cumulus is formed of several rows of cells, though not divided into distinct layers. In the next stage the appearances from the surface are rather more obscure, and in some of my best specimens a coagulum, derived from the fluid surrounding the ovum, covers the most important part of the blastoderm. In Pl. 30, fig. 2, I have attempted to represent, as truly as I could, the appearances presented by the ovum. There is a well-marked whitish side of the ovum, near one end of which is a prominence (_pc._), which must, no doubt, be identified with the cumulus of the earlier stages. Towards the opposite end, or perhaps rather nearer the centre of the white side of the ovum, is an imperfectly marked triangular white area. There can be no doubt that the line connecting the cumulus with the triangular area is the future long axis of the embryo, and the white area is, without doubt, the procephalic lobe of Balbiani.

Footnote 472: I was unfortunately too much engaged, at the time when the eggs were collected, to study them in the fresh condition; a fact which has added not a little to my difficulties in elucidating the obscure points in the early stages.

A section of the ovum at this stage is represented in Pl. 31, fig. 11. It is not quite certain in what direction the section is taken, but I think it probable it is somewhat oblique to the long axis. However this may be, the section shews that the whitish hemisphere of the blastoderm is formed of columnar cells, for the most part two or so layers deep, but that there is, not very far from the middle line, a wedge-shaped internal thickening of the blastoderm where the cells are several rows deep. With what part visible in surface view this thickened portion corresponds is not clear. To my mind it most probably corresponds to the larger white patch, in which case I have not got a section through the terminal prominence. In the other sections of the same embryo the wedge-shaped thickening was not so marked, but it, nevertheless, extended through all the sections. It appears to me probable that it constitutes a longitudinal thickened ridge of the blastoderm. In any case, it is clear that the white hemisphere of the blastoderm is a thickened portion of the blastoderm, and that the thickening is in part due to the cells being more columnar, and, in part, to their being more than one row deep, _though they have not become divided into two distinct germinal layers_. It is further clear that the increase in the number of cells in the thickened part of the blastoderm is, _in the main, a result of the multiplication of the original single row of cells_, while a careful examination of my sections proves that it is also partly due to cells, derived from the yolk, having been added to the blastoderm.

In the following stage which I have obtained (which cannot be very much older than the previous stage, because my specimens of it come from the same batch of eggs), a distinct and fairly circumscribed thickening forming the ventral surface of the embryo has become established. Though its component parts are somewhat indistinct, it appears to consist of a procephalic lobe, a less prominent caudal lobe, and an intermediate portion divided into about three segments; but its constituents cannot be clearly identified with the structures visible in the previous stage. I am inclined, however, to identify the anterior thickened area of the previous stage with the procephalic lobe, and a slight protuberance of the caudal portion (visible from the surface) with the primitive cumulus. I have, however, failed to meet with any trace of the cumulus in my sections.

To this stage, which forms the first of the second period of the larval history, I shall return, but it is necessary now to go back to the observations of Claparède and Balbiani.

There can, in the first place, be but little doubt that what I have called the primitive cumulus in my description is the structure so named by Claparède and Balbiani.

It is clear that Balbiani and Claparède have both failed to appreciate the importance of the organ, which my observations shew to be the part of the ventral thickening of the blastoderm where two rows of cells are first established, and therefore the point where the first traces of the future mesoblast becomes visible.

Though Claparède and Balbiani differ somewhat as to the position of the organ, they both make it last longer than I do: I feel certainly inclined to doubt whether Claparède is right in considering a body he figures after six segments are present, to be the same as the dorsal organ of the embryo before the formation of any segments, especially as all the stages between the two appear to have escaped him. In Agelena there is undoubtedly no organ in the position he gives when six segments are found.

Balbiani's observations accord fairly with my own up to the stage represented in fig. 2. Beyond this stage my own observations are not satisfactory, but I must state that I feel doubtful whether Balbiani is correct in his description of the gradual separation of the procephalic lobe and the cumulus, and the passage of the latter to the dorsal surface, and think it possible that he may have made a mistake as to which side of the procephalic lobe, in relation to the parts of the embryo, the cumulus is placed.

Although there appear to be grounds for doubting whether either Balbiani and Claparède are correct in the position they assign to the cumulus, my observations scarcely warrant me in being very definite in my statements on this head, but, as already mentioned, I am inclined to place the organ near the posterior end (and therefore, as will be afterwards shewn, in a somewhat dorsal situation) of the ventral embryonic thickening.

In my earliest stage of the third period there is present, as has already been stated, a procephalic lobe, and an indistinct and not very prominent caudal portion, and about three segments between the two. The definition of the parts of the blastoderm at this stage is still very imperfect, but from subsequent stages it appears to me probable that the first of the three segments is that of the first pair of ambulatory limbs, and that the segments of the cheliceræ and pedipalpi are formed later than those of the first three ambulatory appendages.

Balbiani believes that the segment of the cheliceræ is formed later than that of the six succeeding segments. He further concludes, from the fact that this segment is cut off from the procephalic portion in front, that it is really part of the procephalic lobe. I cannot accept the validity of this argument; though I am glad to find myself in, at any rate, partial harmony with the distinguished French embryologist as to the facts. Balbiani denies for this stage the existence of a caudal lobe. There is certainly, as is very well shewn in my longitudinal sections, a thickening of the blastoderm in the caudal region, though it is not so prominent in surface views as the procephalic lobe.

A transverse section through an embryo at this stage (Pl. 31, fig. 12) shews that there is a ventral plate of somewhat columnar cells more than one row deep, and a dorsal portion of the blastoderm formed of a single row of flattened cells. Every section at this stage shews that the inner layer of cells of the ventral plate is receiving accessions of cells from the yolk, which has not to any appreciable extent altered its constitution. A large cell, passing from the yolk to the blastoderm, is shewn in fig. 12 at _y.c_.

_The cells of the ventral plate are now divided into two distinct layers._ The outer of these is the _epiblast_, the inner the _mesoblast_. The cells of both layers are quite continuous across the median line, and exhibit no trace of a bilateral arrangement.

This stage is an interesting one on account of the striking similarity which (apart from the amnion) exists between a section through the blastoderm of a spider and that of an insect immediately after the formation of the mesoblast. The reader should compare Kowalevsky's (_Mém. Acad. Pétersbourg_, Vol. XVI. 1871) fig. 26, Pl. IX. with my fig. 12. The existence of a continuous ventral plate of mesoblast has been noticed by Barrois (p. 532), who states that the two mesoblastic bands originate from the longitudinal division of a primitive single band.

In a slightly later stage (Pl. 30, fig. 3_a_ and 3_b_) six distinct segments are interpolated between the procephalic and the caudal lobes. The two foremost, _ch_ and _pd_ (especially the first), of these are far less distinct than the remainder, and the first segment is very indistinctly separated from the procephalic lobe. From the indistinctness of the first two somites, I conclude that they are later formations than the four succeeding ones. The caudal and procephalic lobes are very similar in appearance, but the procephalic lobe is slightly the wider of the two. There is a slight protuberance on the caudal lobe, which is possibly the remnant of the cumulus. The superficial appearance of segmentation is produced by a series of transverse valleys, separating raised intermediate portions which form the segments. The ventral thickening of the embryo now occupies rather more than half the circumference of the ovum.

Transverse sections shew that considerable changes have been effected in the constitution of the blastoderm. In the previous stage, the ventral plate was formed of an uniform external layer of epiblast, and a continuous internal layer of mesoblast. The mesoblast has now become divided along the whole length of the embryo, except, perhaps, the procephalic lobes, into two lateral bands which are not continuous across the middle line (Pl. 31, fig. 13, _me_). It has, moreover, become a much more definite layer, closely attached to the epiblast. Between each mesoblastic band and the adjoining yolk there are placed a few scattered cells, which in a somewhat later stage become the splanchnic mesoblast. These cells are derived from the yolk-cells; and almost every section contains examples of such cells in the act of joining the mesoblast.

The epiblast of the ventral plate has not, to any great extent, altered in constitution. It is, perhaps, a shade thinner in the median line than it is laterally. The division of the mesoblast plate into two bands, together, perhaps, with the slight reduction of the epiblast in the median ventral line, gives rise at this stage to an imperfectly marked median groove.

The dorsal epiblast is still formed of a single layer of flat cells. In the neighbourhood of this layer the yolk nuclei are especially concentrated. The yolk itself remains as before.

The segments continue to increase regularly, each fresh segment being added in the usual way between the last formed segment and the unsegmented caudal lobe. At the stage when about nine or ten segments have become established, the first rudiments of appendages become visible. At this period (Pl. 30, fig. 4) there is a distinct median ventral groove, extending through the whole length of the embryo, which becomes, however, considerably shallower behind. The procephalic region is distinctly bilobed. The first segment (that of the cheliceræ) is better marked off from it than in the previous stage, but is without a trace of an appendage, and exhibits therefore, in respect to the development of its appendages, the same retardation that characterised its first appearance. The next five segments, viz. those of the pedipalpi and four ambulatory appendages, present a very well-marked swelling at each extremity. These swellings are the earliest traces of the appendages. Of the three succeeding segments, only the first is well differentiated. The caudal lobe, though less broad than the procephalic lobe, is still a widish structure. The most important internal changes concern the mesoblast, which is now imperfectly though distinctly divided into somites, corresponding with segments visible externally. Each mesoblastic somite is formed of a distinct somatic layer closely attached to the epiblast, and a thinner and less well-marked splanchnic layer. In the appendage-bearing segments the somatic layer is continued up into the appendages.

The epiblast is distinctly thinner in the median line than at the two sides.

The next stage figured (Pl. 30, figs. 5 and 6) is an important one, as it is characterized by the establishment of the full number of appendages. The whole length of the ventral plate has greatly increased, so that it embraces nearly the circumference of the ovum, and there is left uncovered but a very small arc between the two extremities of the plate (Pl. 30, fig. 6; Pl. 31, fig. 15). This arc is the future dorsal portion of the embryo, which lags in its development immensely behind the ventral portion.

There is a very distinctly bilobed procephalic region (_pr.l_) well separated from the segment with the cheliceræ (_ch_). It is marked by a shallow groove opening behind into a circular depression (_st._)--the earliest rudiment of the stomodæum. The six segments behind the procephalic lobes are the six largest, and each of them bears two prominent appendages. They constitute the six appendage-bearing segments of the adult. The four future ambulatory appendages are equal in size: they are slightly larger than the pedipalpi, and these again than the cheliceræ. Behind the six somites with prominent appendages there are four well-marked somites, each with a small protuberance. These four protuberances are provisional appendages. They have been found in many other genera of Araneina (Claparède, Barrois). The segments behind these are rudimentary and difficult to count, but there are, at any rate, five, and at a slightly later stage probably six, including the anal lobe. These fresh segments have been formed by the continued segmentation of the anal lobe, which has greatly altered its shape in the process. The ventral groove of the earlier stage is still continued along the whole length of the ventral plate.

By the close of this stage the full number of post-cephalic segments has become established. They are best seen in the longitudinal section (Pl. 31, fig. 15). There are six anterior appendage-bearing segments, followed by four with rudimentary appendages (not seen in this figure), and six without appendages behind. There are, therefore, sixteen in all. This number accords with the result arrived at by Barrois, but is higher by two than that given by Claparède.

The germinal layers (vide Pl. 31, fig. 14) have by this stage undergone a further development. The mesoblastic somites are more fully developed. The general relations of these somites is shewn in longitudinal section in Pl. 31, fig. 15, and in transverse section in Pl. 31, fig. 14. In the tail, where they are simplest (shewn on the upper side in fig. 14), each mesoblastic somite is formed of a somatic layer of more or less cubical cells attached to the epiblast, and a splanchnic layer of flattened cells. Between the two is placed a completely circumscribed cavity, which constitutes part of the embryonic body-cavity. Between the yolk and the splanchnic layer are placed a few scattered cells, which form the latest derivatives of the yolk-cells, and are to be reckoned as part of the splanchnic mesoblast. The mesoblastic somites do not extend outwards beyond the edge of the ventral plate, and the corresponding mesoblastic somites of the two sides do not nearly meet in the middle line. In the limb-bearing somites the mesoblast has the same general characters as in the posterior somites, but the _somatic_ layer is prolonged as a hollow papilliform process into the limb, so that each limb has an axial cavity continuous with the section of the body-cavity of its somite. The description given by Metschnikoff of the formation of the mesoblastic somites in the scorpion, and their continuation into the limbs, closely corresponds with the history of these parts in spiders. In the region of each procephalic lobe the mesoblast is present as a continuous layer underneath the epiblast, but in the earlier part of the stage, at any rate, is not formed of two distinct layers with a cavity between them.

The epiblast at this stage has also undergone important changes. Along the median ventral groove it has become very thin. On each side of this groove it exhibits in each appendage-bearing somite a well-marked thickening, which gives in surface views the appearance of a slightly raised area (Pl. 30, fig. 5), between each appendage and the median line. These thickenings are the first rudiments of the ventral nerve ganglia. The ventral nerve cord at this stage is formed of two ridge-like thickenings of the epiblast, widely separated in the median line, each of which is constituted of a series of raised divisions--the ganglia--united by shorter, less prominent divisions (fig. 14, _vg_). The nerve cords are formed from before backwards, and are not at this stage found in the hinder segments. _There is a distinct ganglionic thickening for the cheliceræ quite independent of the procephalic lobes._

In the procephalic lobes the epiblast is much thickened, and is formed of several rows of cells. The greater part of it is destined to give rise to the supra-oesophageal ganglia.

During the various changes which have been described the blastoderm cells have been continually dividing, and, together with their nuclei, have become considerably smaller than at first. The yolk cells have in the meantime remained much as before, and are, therefore, considerably larger than the nuclei of the blastoderm cells. They are more numerous than in the earlier stages, but are still surrounded by a protoplasmic body, which is continued into a protoplasmic reticulum. The yolk is still divided up into polygonal segments, but from sections it would appear that the nuclei are more numerous than the segments, though I have failed to arrive at quite definite conclusions on this point.

As development proceeds the appendages grow longer, and gradually bend inwards. They become very soon divided by a series of ring-like constrictions which constitute the first indications of the future joints (Pl. 30, fig. 6). The full number of joints are not at once reached, but in the ambulatory appendages five only appear at first to be formed. There are four joints in the pedipalpi, while the cheliceræ do not exhibit any signs of becoming jointed till somewhat later. The primitive presence of only five joints in the ambulatory appendages is interesting, as this number is permanent in Insects and in Peripatus.

The next stage figured forms the last of the third period (Pl. 30, figs. 7 and 7_a_). The ventral plate is still rolled round the egg (fig. 7), and the end of the tail and the procephalic lobes nearly meet dorsally, so that there is but a very slight development of the dorsal region. There are the same number of segments as before, and the chief differences in appearance between the present and the previous stage depend upon the fact (1) that the median ventral integument between the nerve ganglia has become wider, and at the same time thinner; (2) that the limbs have become much more developed; (3) that the stomodæum is definitely established; (4) that the procephalic lobes have undergone considerable development.

Of these features, the three last require a fuller description. The limbs of the two sides are directed towards each other, and nearly meet in the ventral line. The cheliceræ are two-jointed, and terminate in what appear like rudimentary chelæ, a fact which perhaps indicates that the spiders are descended from ancestors with chelate cheliceræ. The four embryonic post-ambulatory appendages are now at the height of their development.

The stomodæum (Pl. 30, fig. 7, and Pl. 31, fig. 17, _st_) is a deepish pit between the two procephalic lobes, and distinctly in front of the segment of the cheliceræ. It is bordered in front by a large, well-marked, bilobed upper lip, and behind by a smaller lower lip. The large upper lip is a temporary structure, to be compared, perhaps, with the gigantic upper lip of the embryo of Chelifer (cf. Metschnikoff). On each side of and behind the mouth two whitish masses are visible, which are the epiblastic thickenings which constitute the ganglia of the cheliceræ (Pl. 30, fig. 7, _ch.g_).

The procephalic lobes (_pr.l_) now form two distinct masses, and each of them is marked by a semicircular groove, dividing them into a narrower anterior and a broader posterior division.

In the region of the trunk the general arrangement of the germinal layers has not altered to any great extent. The ventral ganglionic thickenings are now developed in all the segments in the abdominal as well as in the thoracic region. The individual thickenings themselves, though much more conspicuous than in the previous stage (Pl. 31, fig. 16, _v.c_), are still integral parts of the epiblast. They are more widely separated than before in the middle line. The mesoblastic somites retain their earlier constitution (Pl. 31, fig. 16). Beneath the procephalic lobes the mesoblast has, in most respects, a constitution similar to that of a mesoblastic somite in the trunk. It is formed of two bodies, one on each side, each composed of a splanchnic and somatic layer (Pl. 31, fig. 17, _sp._ and _so_), enclosing between them a section of the body-cavity. But the cephalic somites, unlike those of the trunk, are united by a median bridge of mesoblast, in which no division into two layers can be detected. This bridge assists in forming a thick investment of mesoblast round the stomodæum (_st_).

The existence of a section of the body-cavity in the præoral region is a fact of some interest, especially when taken in connection with the discovery, by Kleinenberg, of a similar structure in the head of Lumbricus. The procephalic lobe represents the præoral lobe of Chætopod larvæ, but the prolongation of the body-cavity into it does not, in my opinion, necessarily imply that it is equivalent to a post-oral segment.

The epiblast of the procephalic lobes is a thick layer several cells deep, but without any trace of a separation of the ganglionic portion from the epidermis.

The nuclei of the yolk have increased in number, but the yolk, in other respects, retains its earlier characters.

The next period in the development is that in which the body of the embryo gradually acquires the adult form. The most important event which takes place during this period is the development of the dorsal region of the embryo, which, up to its commencement, is practically non-existent. As a consequence of the development of the dorsal region, the embryo, which has hitherto had what may be called a dorsal flexure, gradually unrolls itself, and acquires a ventral flexure. This change in the flexure of the embryo is in appearance a rather complicated phenomenon, and has been somewhat differently described by the two naturalists who have studied it in recent times.

For Claparède the prime cause of the change of flexure is the translation dorsalwards of the limbs. He compares the dorsal region of the embryo to the arc of a circle, the two ends of which are united by a cord formed by the line of insertion of the limbs. He points out that if you bring the middle of the cord, so stretched between the two ends of the arc, nearer to the summit of the arc, you necessarily cause the two ends of the arc to approach each other, or, in other words, if the insertion of the limbs is drawn up dorsally, the head and tail must approach each other ventrally.

Barrois takes quite a different view to that of Claparède, which will perhaps be best understood if I quote a translation of his own words. He says: "At the period of the last stage of the embryonic band (the stage represented in Pl. 31, fig. 7, in the present paper) this latter completely encircles the egg, and its posterior extremity nearly approaches the cephalic region. Finally, the germinal bands, where they unite at the anal lobe (placed above on the dorsal surface), form between them a very acute angle. During the following stages one observes the anal segment separate further and further from the cephalic region, and approach nearer and nearer to the ventral region. This displacement of the anal segment determines, in its turn, a modification in the divergence of the anal bands; the angle which they form at their junction tends to become more obtuse. The same processes continue regularly till the anal segment comes to occupy the opposite extremity to the cephalic region, a period at which the two germinal bands are placed in the same plane and the two sides of the obtuse angle end by meeting in a straight line. If we suppose a continuation of the same phenomenon it is clear that the anal segment will come to occupy a position on the ventral surface, and the germinal bands to approach, but in the inverse way, so as to form an angle opposite to that which they formed at first. This condition ends the process by which the posterior extremity of the embryonic band, at first directed towards the dorsal side, comes to bend in towards the ventral region."

Neither of the above explanations is to my mind perfectly satisfactory. The whole phenomenon appears to me to be very simple, and to be caused by the elongation of the dorsal region, _i.e._ the region on the dorsal surface between the anal and procephalic lobes. Such an elongation necessarily separates the anal and procephalic lobes; but, since the ventral plate does not become shortened in the process, and the embryo cannot straighten itself on account of the egg-shell, it necessarily becomes flexed, and such flexure can only be what I have already called a ventral flexure. If there were but little food yolk this flexure would cause the whole embryo to be bent in, so as to have the ventral surface concave, but instead of this the flexure is confined at first to the two bands which form the ventral plate. These bands are bent in the natural way (Pl. 30, fig. 8_b_), but the yolk forms a projection, a kind of yolk-sack as Barrois calls it, distending the thin integument between the two ventral bands. This yolk-sack is shewn in surface view in Pl. 30, fig. 8, and in section in Pl. 32, fig. 18. At a later period, when the yolk has become largely absorbed in the formation of various organs, the true nature of the ventral flexure becomes apparent, and the abdomen of the young Spider is found to be bent over so as to press against the ventral surface of the thorax (Pl. 30, fig. 9). This flexure is shewn in section in Pl. 32, fig. 21.

At the earliest stage of this period of which I have examples, the dorsal region has somewhat increased, though not very much. The limbs have grown very considerably and _now cross in the middle line_.

The ventral ganglia, though not the supra-oesophageal, have become separated from the epiblast.

The yolk nuclei, each surrounded by protoplasm as before, are much more numerous.

In other respects there are no great changes in the internal features.

In my next stage, represented in Pl. 30, figs. 8_a_, and 8_b_, a very considerable advance has become effected. In the first place the dorsal surface has increased in length to rather more than one half the circumference of the ovum. The dorsal region has, however, not only increased in length, but also in definiteness, and a series of transverse markings (figs. 8_a_ and _b_), which are very conspicuous in the case of the four anterior abdominal segments (the segments with rudimentary appendages), have appeared, indicating the limits of segments dorsally. The terga of the somites may, in fact, be said to have become formed. The posterior terga (fig. 8_a_) are very narrow compared to the anterior.

The caudal protuberance is more prominent than it was, and somewhat bilobed; it is continued on each side into one of the bands, into which the ventral plate is divided. These bands, as is best seen in side view (fig. 8_b_), have a ventral curvature, or, perhaps more correctly, are formed of two parts, which meet at a large angle open towards the ventral surface. The posterior of these parts bears the four still very conspicuous provisional appendages, and the anterior the six pairs of thoracic appendages. The four ambulatory appendages are now seven-jointed, as in the adult, but though longer than in the previous stage they do not any longer _cross or even meet in the middle line_, but are, on the contrary, separated by a very considerable interval. This is due to the great distension by the yolk of the ventral part of the body, in the interval between the two parts of the original ventral plate. The amount of this yolk may be gathered from the section (Pl. 32, fig. 18). The pedipalpi carry a blade on their basal joint. The cheliceræ no longer appear to spring from an independent postoral segment.

There is a conspicuous lower lip, but the upper is less prominent than before. Sections at this stage shew that the internal changes have been nearly as considerable as the external.

The dorsal region is now formed of a (1) flattened layer of epiblast cells, and a (2) fairly thick layer of large and rather characteristic cells which any one who has studied sections of spider's embryos will recognize as derivatives of the yolk. These cells are not, therefore, derived from prolongations of the somatic and splanchnic layers of the already formed somites, but are new formations derived from the yolk. They commenced to be formed at a much earlier period, and some of them are shewn in the longitudinal section (Pl. 31, fig. 15). In the next stage these cells become differentiated into the somatic and splanchnic mesoblast layers of the dorsal region of the embryo.

In the dorsal region of the abdomen the heart has already become established. So far as I have been able to make out it is formed from a solid cord of the cells of the dorsal region. The peripheral layer of this cord gives rise to the walls of the heart, while the central cells become converted into the corpuscles of the blood.

The rudiment of the heart is in contact with the epiblast above, and there is no greater evidence of its being derived from the splanchnic than from the somatic mesoblast; it is, in fact, formed before the dorsal mesoblast has become differentiated into two layers.

In the abdomen three or four transverse septa, derived from the splanchnic mesoblast, grow a short way into the yolk. They become more conspicuous during the succeeding stage, and are spoken of in detail in the description of that stage. In the anterior part of the thorax a longitudinal and vertical septum is formed, which grows downwards from the median dorsal line, and divides the yolk in this region into two parts. In this septum there is formed at a later stage a vertical muscle attached to the suctorial part of the stomodæum.

The mesoblastic somites of the earlier stage are but little modified; and there are still prolongations of the body-cavity into the limbs (Pl. 32, fig. 18).

The lateral parts of the ventral nerve cords are now at their maximum of separation (Pl. 32, fig. 18, _v.g._). Considerable differentiation has already set in in the constitution of the ganglia themselves, which are composed of an outer mass of ganglion cells enclosing a kernel of nerve fibres, which lie on the inner side and connect the successive ganglia. There are still distinct thoracic and abdominal ganglia for each segment, and there is also a pair of separate ganglion for the cheliceræ, which assists, however, in forming the oesophageal commissures.

The thickenings of the præoral lobe which form the supra-oesophageal ganglia are nearly though not quite separated from the epiblast. The semicircular grooves of the earlier stages are now deeper than before, and are well shewn in sections nearly parallel to the outer anterior surface of the ganglion (Pl. 32, fig. 19). The supra-oesophageal ganglia are still entirely formed of undifferentiated cells, and are without commissural tissue like that present in the ventral ganglia.

The stomodæum has considerably increased in length, and the proctodæum has become formed as a short, posteriorly directed involution of the epiblast. I have seen traces of what I believe to be two outgrowths from it, which form the Malpighian bodies.

The next stage constitutes (Pl. 30, fig. 9) the last which requires to be dealt with so far as the external features are concerned. The yolk has now mainly passed into the abdomen, and the constriction separating the thorax and abdomen has begun to appear. The yolk-sack has become absorbed, so that the two halves of the ventral plate in the thorax are no longer widely divaricated. The limbs have to a large extent acquired their permanent structure, and the rings of which they are formed in the earlier stages are now replaced by definite joints. A delicate cuticle has become formed, which is not figured in my sections. The four rudimentary appendages have disappeared, unless, which seems to me in the highest degree improbable, they remain as the spinning mammillæ, two pairs of which are now present. Behind is the anal lobe, which is much smaller and less conspicuous than in the previous stage. The spinnerets and anal lobe are shewn as five papillæ in Pl. 30, fig. 9. Dorsally the heart is now very conspicuous, and in front of the cheliceræ may be seen the supra-oesophageal ganglia.

The indifferent mesoblast has now to a great extent become converted into the permanent tissues. On the dorsal surface there was present in the last stage a great mass of unformed mesoblast cells. This mass of cells has now become divided into a somatic and splanchnic layer (Pl. 32, fig. 22). It has, moreover, in the abdominal region at any rate, become divided up into somites. At the junction between the successive somites the splanchnic mesoblast on each side of the abdomen dips down into the yolk and forms a septum (Pl. 32, fig. 22, _s_). The septa so formed, which were first described by Barrois, are not complete. The septa of the two sides do not, in the first place, quite meet along the median dorsal or ventral lines, and in the second place they only penetrate the yolk for a certain distance. Internally they usually end in a thickened border.

Along the line of insertion of each of these septa there is developed a considerable space between the somatic and splanchnic layers of mesoblast. The parts of the body-cavity so established are transversely directed channels passing from the heart outwards. They probably constitute the venous spaces, and perhaps also contain the transverse aortic branches.

In the intervals between these venous spaces the somatic and splanchnic layers of mesoblast are in contact with each other.

I have not been able to work out satisfactorily the later stages of development of the septa, but I have found that they play an important part in the subsequent development of the abdomen. In the first place they send off lateral offshoots, which unite the various septa together, and divide up the cavity of the abdomen into a number of partially separated compartments. There appears, however, to be left a free axial space for the alimentary tract, the mesoblastic walls of which are, I believe, formed from the septa.

At the present stage the splanchnic mesoblast, apart from the septa, is a delicate membrane of flattened cells (fig. 22, _sp_). The somatic mesoblast is thicker, and is formed of scattered cells (_so_).

The somatic layer is in part converted, in the posterior region of the abdomen, into a delicate layer of longitudinal muscles, the fibres of which are not continuous for the whole length of the body, but are interrupted at the lines of junction of the successive segments. They are not present in the anterior part of the abdomen. The longitudinal direction of these fibres, and their division with myotomes, is interesting, since both these characters, which are preserved in Scorpions, are lost in the abdomen of the adult Spider.

The original mesoblastic somites have undergone quite as important changes as the dorsal mesoblast. In the abdominal region the somatic layer constitutes two powerful bands of longitudinal muscles, inserted anteriorly at the root of the fourth ambulatory appendage, and posteriorly at the spinning mammillæ. Between these two bands are placed the nervous bands. The relation of these parts are shewn in the section in Pl. 32, fig. 20_d_, which cuts the abdomen horizontally and longitudinally. The mesoblastic bands are seen at _m._, and the nervous bands within them at _ab.g_. In the thoracic region the part of the somatic layer in each limb is converted into muscles, which are continued into dorsal and ventral muscles in the thorax (vide fig. 20_c_). There are, in addition to these, intrinsic transverse fibres on the ventral side of the thorax. Besides these muscles there are in the thorax, attached to the suctorial extremity of the stomodæum, three powerful muscles, which I believe to be derived from the somatic mesoblast. One of these passes vertically down from the dorsal surface, in the septum the commencement of which was described in the last stage. The two other muscles are lateral, one on each side (Pl. 31, fig. 20_c_.).

The heart has now, in most respects, reached its full development. It is formed of an outer muscular layer, within which is a doubly-contoured lining, containing nuclei at intervals, which is probably of the nature of an epithelioid lining (Pl. 32, fig. 22, _ht_). In its lumen are numerous blood-corpuscles (not represented in my figure). The heart lies in a space bound below by the splanchnic mesoblast, and to the sides by the somatic mesoblast. This space forms a kind of pericardium (fig. 22, _pc_), but dorsally the heart is in contact with the epiblast. The arterial trunks connected with it are fully established.

The nervous system has undergone very important changes.

In the abdominal region the ganglia of each side have fused together into a continuous cord (fig. 21, _ab.g_). In fig. 20, in which the abdomen is cut horizontally and longitudinally, there are seen the two abdominal cords (_ab.g_) united by two transverse commissures; and I believe that there are at this stage three or four transverse commissures at any rate, which remain as indications of the separate ganglia, from the coalescence of which the abdominal cords are formed. The two abdominal cords are parallel and in close contact.

In the thoracic region changes of not less importance have taken place. The ganglia are still distinct. The two cords formed of these ganglia are no longer widely separated in median line, but meet, in the usual way, in the ventral line. Transverse commissures have become established (fig. 20_c_) between the ganglia of the two sides. There is as little trace at this, as at the previous stages, of an ingrowth of epiblast, to form a median portion of the central nervous system. Such a median structure has been described by Hatschek for Lepidoptera, and he states that it gives rise to the transverse commissures between the ganglia. My observations shew that for the spider, at any rate, nothing of the kind is present.

As shewn in the longitudinal section (Pl. 32, fig. 21), the ganglion of the cheliceræ has now united with the supra-oesophageal ganglion. It forms, as is shewn in fig. 20_b_ (_ch.g._), a part of the oesophageal commissure, and there is no sub-oesophageal commissure uniting the ganglia of the cheliceræ, but the oesophageal ring is completed below by the ganglia of the pedipalpi (fig. 20_c_, _pd.g._).

The supra-oesophageal ganglia have become completely separated from the epiblast.

I have unfortunately not studied their constitution in the adult, so that I cannot satisfactorily identify the parts which can be made out at this stage.

I distinguish, however, the following regions:

(1) A central region containing the commissural part, and continuous below with the ganglia of the cheliceræ.

(2) A dorsal region formed of two hemispherical lobes.

(3) A ventral anterior region.

The central region contains in its interior the commissural portion, forming a punctiform, rounded mass in each ganglion. A transverse commissure connects the two (vide fig. 20_b_).

The dorsal hemispherical lobes are derived from the part which, at the earlier stage, contained the semicircular grooves. When the supra-oesophageal ganglia become separated from the epidermis the cells lining these grooves become constricted off with them, and form part of these ganglia. Two cavities are thus formed in this part of the supra-oesophageal ganglia. These cavities become, for the most part, obliterated, but persist at the outer side of the hemispherical lobes (figs. 20_a_ and 21).

The ventral lobe of the brain is a large mass shewn in longitudinal section in fig. 21. It lies immediately in front of and almost in contact with the ganglia of the cheliceræ.

The two hemispherical lobes agree in position with the fungiform body (_pilzhutförmige Körpern_), which has attracted so much the attention of anatomists, in the supra-oesophageal ganglia of Insects and Crustacea; but till the adult brain of Spiders has been more fully studied it is not possible to state whether the hemispherical lobes become fungiform bodies.

Hatschek[473] has described a special epiblastic invagination in the supra-oesophageal ganglion of Bombyx, which is probably identical with the semicircular groove of Spiders and Scorpions, but in the figure he gives the groove does not resemble that in the Arachnida. A similar groove is found in Peripatus, and there forms, as I have found, a large part of the supra-oesophageal ganglia. It is figured by Moseley, _Phil. Trans._, Vol. CLXIV. pl. lxxv, fig. 9.

Footnote 473: "Beiträge z. Entwick. d. Lepidopteren," _Jenaische Zeit._, Vol. XI. p. 124.

The stomodæum is considerably larger than in the last stage, and is lined by a cuticle; it is a blind tube, the blind end of which is the suctorial pouch of the adult. To this pouch are attached the vertical dorsal, and two lateral muscles spoken of above.

The proctodæum (_pr._) has also grown in length, and the two Malpighian vessels which grow out from its blind extremity (fig. 20_e_, _mp.g._) have become quite distinct. The part now formed is the rectum of the adult. The proctodæum is surrounded by a great mass of splanchnic mesoblast. The mesenteron has as yet hardly commenced to be developed. There is, however, a short tube close to the proctodæum (fig. 20_e_, _mes_), which would seem to be the commencement of it. It ends blindly on the side adjoining the rectum, but is open anteriorly towards the yolk, and there can be very little doubt that it owes its origin to cells derived from the yolk. On its outer surface is a layer of mesoblast.

From the condition of the mesenteron at this stage there can be but little doubt that it will be formed, not on the surface, _but in the interior of the yolk_. I failed to find any trace of an anterior part of the mesenteron adjoining the stomodæum. In the posterior part of the thorax (vide fig. 20_d_), there is undoubtedly no trace of the alimentary tract.

The presence of this rudiment shews that Barrois is mistaken in supposing that the alimentary canal is formed entirely from the stomodæum and proctodæum, which are stated by him to grow towards each other, and to meet at the junction of the thorax and abdomen. My own impression is that the stomodæum and proctodæum have reached their full extension at the present stage, and that both the stomach in the thorax and the intestine in the abdomen are products of the mesenteron.

The yolk retains its earlier constitution, being divided into polygonal segments, formed of large yolk vesicles. The nuclei are more numerous than before. In the thorax the yolk is anteriorly divided into two lobes by the vertical septum, which contains the vertical muscle of the suctorial pouch. In the posterior part of the thorax it is undivided.

I have not yet been able clearly to make out the eventual fate of the yolk. At a subsequent stage, when the cavity of the abdomen is cut up into a series of compartments by the growth of the septa, described above, the yolk fills these compartments, and there is undoubtedly a proliferation of yolk cells round the walls of these compartments. It would not be unreasonable to conclude from this that the compartments were destined to form the hepatic cæca, each cæcum being enclosed in a layer of splanchnic mesoblast, and its hypoblastic wall being derived from the yolk cells. I think that this hypothesis is probably correct, but I have met with some facts which made me think it possible that the thickenings at the ends of the septa, visible in Pl. 32, fig. 22, were the commencing hepatic cæca.

I must, in fact, admit that I have hitherto failed to work out satisfactorily the history of the mesenteron and its appendages. The firm cuticle of young spiders is an obstacle both in the way of making sections and of staining, which I have not yet overcome.

_General Conclusions._

Without attempting to compare at length the development of the spiders with that of other Arthropoda, I propose to point out a few features in the development of spiders, which appear to shew that the Arachnida are undoubtedly more closely related to the other Tracheata than to the Crustacea.

The whole history of the formation of the mesoblast is very similar to that in insects. The mesoblast in both groups is formed by a thickening of the median line of the ventral plate (germinal streak).

In insects there is usually formed a median groove, the walls of which become converted into a plate of mesoblast. In spiders there is no such groove, but a median keel-like thickening of the ventral plate (Pl. 31, fig. 11), is very probably an homologous structure. The unpaired plate of mesoblast formed in both insects and Arachnida is exactly similar, and becomes divided, in both groups, into two bands, one on each side of the middle line. Such differences as there are between Insects and Arachnida sink into insignificance compared with the immense differences in the origin of the mesoblast between either group, and that in the Isopoda, or, still more, the Malacostraca and most Crustacea. In most Crustacea we find that the mesoblast is budded off from the walls of an invagination, which gives rise to the mesenteron.

In both spiders and Myriopoda, and probably insects, the mesoblast is subsequently divided into somites, the lumen of which is continued into the limbs. In Crustacea mesoblastic somites have not usually been found, though they appear occasionally to occur, _e.g._ Mysis, but they are in no case similar to those in the Tracheata.

In the formation of the alimentary tract, again, the differences between the Crustacea and Tracheata are equally marked, and the Arachnida agree with the Tracheata. There is generally in Crustacea an invagination, which gives rise to the mesenteron. In Tracheata this never occurs. The proctodæum is usually formed in Crustacea before or, at any rate, not later than the stomodæum[474]. The reverse is true for the Tracheata. In Crustacea the proctodæum and stomodæum, especially the former, are very long, and usually give rise to the greater part of the alimentary tract, while the mesenteron is usually short.

Footnote 474: If Grobben's account of the development of Moina is correct this statement must be considered not to be universally true.

In the Tracheata the mesenteron is always considerable, and the proctodæum is always short. The derivation of the Malpighian bodies from the proctodæum is common to most Tracheata. Such organs are not found in the Crustacea.

With reference to other points in my investigations, the evidence which I have got that the cheliceræ are true postoral appendages supplied in the embryo from a distinct postoral ganglion, confirms the conclusions of most previous investigators, and shews that these appendages are equivalent to the mandibles, or possibly the first pair of maxillæ of other Tracheata. The invagination, which I have found, of part of a groove of epiblast in the formation of the supra-oesophageal ganglia is of interest, owing to the wide extension of a similar occurrence amongst the Tracheata.

The wide divarication of the ventral nerve cords in the embryo renders it easy to prove that there is no median invagination of epiblast between them, and supports Kleinenberg's observations on Lumbricus as to the absence of this invagination. I have further satisfied myself as to the absence of such an invagination in Peripatus. It is probable that Hatschek and other observers who have followed him are mistaken in affirming the existence of such an invagination in either the Chætopoda or the Arthropoda.

The observations recorded in this paper on the yolk cells and their derivations are, on the whole, in close harmony with the observations of Dohrn, Bobretzky, and Graber, on Insects. They shew, however, that the first formed mesoblastic plate does not give rise to the whole of the mesoblast, but that during the whole of embryonic life the mesoblast continues to receive accessions of cells derived from the cells of the yolk.

_Araneina._

1. Balbiani, "Mémoire sur le Développement des Araneides," _Ann. Sci. Nat._, series v, Vol. XVII. 1873.

2. J. Barrois, "Recherches s. l. Développement des Araignées," _Journal de l'Anat. et de la Physiol._, 1878.

3. E. Claparède, _Recherches s. l'Evolution des Araignées_, Utrecht, 1860.

4. Herold, _De Generatione Araniorum in Ovo_, Marburg, 1824.

5. H. Ludwig, "Ueb. d. Bildung des Blastoderm bei d. Spinnen," _Zeit. f. wiss. Zool._, Vol. XXVI. 1876.

EXPLANATION OF PLATES 30, 31, AND 32.

PLATE 30.

COMPLETE LIST OF REFERENCE LETTERS.

_ch._ Cheliceræ. _ch.g._ Ganglion of cheliceræ. _c.l._ Caudal lobe. _p.c._ Primitive cumulus. _pd._ Pedipalpi. _pr.l._ Præoral lobe. _pp_{1}. _pp_{2}. _etc._ Provisional appendages. _sp._ Spinnerets. _st._ Stomodæum.

I-IV. Ambulatory appendages. 1-16. Postoral segments.

Fig. 1. Ovum, with primitive cumulus and streak proceeding from it.

Fig. 2. Somewhat later stage, in which the primitive cumulus is still visible. Near the opposite end of the blastoderm is a white area, which is probably the rudiment of the procephalic lobe.

Fig. 3_a_ and 3_b_. View of an embryo from the ventral surface and from the side when six segments have become established.

Fig. 4. View of an embryo, ideally unrolled, when the first rudiments of the appendages become visible.

Fig. 5. Embryo ideally unrolled at the stage when all the appendages have become established.

Fig. 6. Somewhat older stage, when the limbs begin to be jointed. Viewed from the side.

Fig. 7. Later stage, viewed from the side.

Fig. 7_a_. Same embryo as fig. 7, ideally unrolled.

Figs. 8_a_ and 8_b_. View from the ventral surface and from the side of an embryo, after the ventral flexure has considerably advanced.

Fig. 9. Somewhat older embryo, viewed from the ventral surface.

PLATES 31 AND 32.

COMPLETE LIST OF REFERENCE LETTERS.

_ao._ Aorta. _ab.g._ Abdominal nerve cord. _ch._ Cheliceræ. _ch.g._ Ganglion of cheliceræ. _ep._ Epiblast. _hs._ Hemispherical lobe of supra-oesophageal ganglion. _ht._ Heart. _l.l._ Lower lip. _m._ Muscles. _me._ Mesoblast. _mes._ Mesenteron. _mp.g._ Malpighian tube. _ms._ Mesoblastic somite. _oe._ OEsophagus. _p.c._ Pericardium. _pd._ Pedipalpi. _pd.g._ Ganglion of pedipalpi. _pr._ Proctodæum (rectum). _pr.c._ Primitive cumulus. _s._ Septum in abdomen. _so._ Somatopleure. _sp._ Splanchnopleure. _st._ Stomodæum. _su._ Suctorial apparatus. _su.g._ Supra-oesophageal ganglion. _th. g._ Thoracic ganglion. _v.g._ Ventral nerve cord. _y.c._ Cells derived from yolk. _yk._ Yolk. _y.n._ Nuclei of yolk cells.

I_g_-IV_g_. Ganglia of ambulatory limbs. 1-16. Postoral segments.

Fig. 10. Section through an ovum, slightly younger than fig. 1. Shewing the primitive cumulus and the columnar character of the cells of one half of the blastoderm.

Fig. 11. Section through an embryo of the same age as fig. 2. Shewing the median thickening of the blastoderm.

Fig. 12. Transverse section through the ventral plate of a somewhat older embryo. Shewing the division of the ventral plate into epiblast and mesoblast.

Fig. 13. Section through the ventral plate of an embryo of the same age as fig. 3, shewing the division of the mesoblast of the ventral plate into two mesoblastic bands.

Fig. 14. Transverse section through an embryo of the same age as fig. 5, passing through an abdominal segment above and a thoracic segment below.

Fig. 15. Longitudinal section slightly to one side of the middle line through an embryo of the same age.

Fig. 16. Transverse section through the ventral plate in the thoracic region of an embryo of the same age as fig. 7.

Fig. 17. Transverse section through the procephalic lobes of an embryo of the same age. _gr._ Section of hemicircular groove in procephalic lobe.

Fig. 18. Transverse section through the thoracic region of an embryo of the same age as fig. 8.

Fig. 19. Section through the procephalic lobes of an embryo of the same age.

Fig. 20_a_, _b_, _c_, _d_, _e_. Five sections through an embryo of the same age as fig. 9. _a_ and _b_ are sections through the procephalic lobes, _c_ through the front part of the thorax. _d_ cuts transversely the posterior parts of the thorax, and longitudinally and horizontally the ventral surface of the abdomen. _e_ cuts the posterior part of the abdomen longitudinally and horizontally, and shews the commencement of the mesenteron.

Fig. 21. Longitudinal and vertical section of an embryo of the same age. The section passes somewhat to one side of the middle line, and shews the structure of the nervous system.

Fig. 22. Transverse section through the dorsal part of the abdomen of an embryo of the same stage as fig. 9.

XVIII. ON THE SPINAL NERVES OF AMPHIOXUS[475].

Footnote 475: From the _Quarterly Journal of Microscopical Science_, Vol. XX. 1880.

In an interesting memoir devoted to the elucidation of a series of points in the anatomy and development of the Vertebrata, Schneider[476] has described what he believes to be motor nerves in Amphioxus, which spring from the anterior side of the spinal cord. According to Schneider these nerves have been overlooked by all previous observers except Stieda.

Footnote 476: _Beiträge z. Anat. u. Entwick. d. Wirbelthiere_, Berlin, 1879.

I[477] myself attempted to shew some time ago that anterior roots were absent in Amphioxus; and in some speculations on the cranial nerves, I employed this peculiarity of the nervous system of Amphioxus to support a view that Vertebrata were primitively provided only with nerves of mixed function springing from the posterior side of the spinal cord. Under these circumstances, Schneider's statement naturally attracted my attention, and I have made some efforts to satisfy myself as to its accuracy. The nerves, as he describes them, are very peculiar. They arise from a number of distinct roots in the hinder third of each segment. They form a flat bundle, of which part passes upwards and part downwards. When they meet the muscles they bend backwards, and fuse with the free borders of the muscle-plates. The fibres, which at first sight appear to form the nerve, are, however, transversely striated, and are regarded by Schneider as muscles; and he holds that each muscle-plate sends a process to the edge of the spinal cord, which there receives its innervation. A considerable body of evidence is requisite to justify a belief in the existence of such very extraordinary and unparalleled motor nerves; and for my part I cannot say that Schneider's observations are convincing to me. I have attempted to repeat his observations, employing the methods he describes.

Footnote 477: "On the Spinal Nerves of Amphioxus," _Journ. of Anat. and Phys._ Vol. X. 1876. [This edition, No. IX. p. 197.]

In the first place, he states that by isolating the spinal cord by boiling in acetic acid, the anterior roots may be brought into view as numerous conical processes of the spinal cord in each segment. I find by treating the spinal cord in this way, that processes more or less similar, but more irregular than those which he figures, are occasionally present; but I cannot persuade myself that they are anything but parts of the sheath of the spinal cord which is not completely dissolved by treatment with acetic acid. By treatment with nitric acid _no such processes are to be seen_, though the whole length and very finest branches of the posterior nerves are preserved.

By treating with nitric acid and clarifying by oil of cloves, and subsequently removing one half of the body so as to expose the spinal cord _in sitû_, the origin and distribution of the posterior nerves is very clearly exhibited. But I have failed to detect any trace of the anterior nerve-roots. Horizontal section, which ought also to bring them clearly into view, failed to shew me anything which I could interpret as such. I agree with Schneider that a process of each muscle-plate is prolonged up to the anterior border of the spinal cord, but I can find no trace of a connection between it and the cord.

Schneider has represented a transverse section in which the anterior nerves are figured. I am very familiar with an appearance in section such as that represented in his figure, but I satisfied myself when I previously studied the nerves in Amphioxus, that the body supposed to be a nerve by Schneider was nothing else than part of the intermuscular septum, and after re-examining my sections I see no reason to alter my view.

A very satisfactory proof that the ventral nerves do not exist would be found, if it could be established that the dorsal nerves contained both motor and sensory fibres. So far I have not succeeded in proving this; I have not, however, had fresh specimens to assist me in the investigation. Langerhans[478], whose careful observations appear to me to have been undervalued by Schneider, figures a branch distributed to the muscles, which passes off from the dorsal roots. Till the inaccuracy of this observation is demonstrated, the balance of evidence appears to me to be opposed to Schneider's view.

Footnote 478: _Archiv f. Mikros. Anatomie_, Vol. XII.

XIX. ADDRESS TO THE DEPARTMENT OF ANATOMY AND PHYSIOLOGY OF THE BRITISH ASSOCIATION, 1880.

In the spring of the present year, Professor Huxley delivered an address at the Royal Institution, to which he gave the felicitous title of '_The coming of age of the origin of species_.' It is, as he pointed out, twenty-one years since Mr Darwin's great work was published, and the present occasion is an appropriate one to review the effect which it has had on the progress of biological knowledge.

There is, I may venture to say, no department of biology the growth of which has not been profoundly influenced by the Darwinian theory. When Messrs Darwin and Wallace first enunciated their views to the scientific world, the facts they brought forward seemed to many naturalists insufficient to substantiate their far-reaching conclusions. Since that time an overwhelming mass of evidence has, however, been rapidly accumulating in their favour. Facts which at first appeared to be opposed to their theories have one by one been shewn to afford striking proofs of their truth. There are at the present time but few naturalists who do not accept in the main the Darwinian theory, and even some of those who reject many of Darwin's explanations still accept the fundamental position that all animals are descended from a common stock.

To attempt in the brief time which I have at my disposal to trace the influence of the Darwinian theory on all the branches of anatomy and physiology would be wholly impossible, and I shall confine myself to an attempt to do so for a small section only. There is perhaps no department of Biology which has been so revolutionised, if I may use the term, by the theory of animal evolution, as that of Development or Embryology. The reason of this is not far to seek. According to the Darwinian theory, the present order of the organic world has been caused by the action of two laws, known as the laws of heredity and of variation. The law of heredity is familiarly exemplified by the well-known fact that offspring resemble their parents. Not only, however, do the offspring belong to the same species as their parents, but they inherit the individual peculiarities of their parents. It is on this that the breeders of cattle depend, and it is a fact of every-day experience amongst ourselves. A further point with reference to heredity to which I must call your attention is the fact that the characters, which display themselves at some special period in the life of the parent, are acquired by the offspring at a corresponding period. Thus, in many birds the males have a special plumage in the adult state. The male offspring is not, however, born with the adult plumage, but only acquires it when it becomes adult.

The law of variation is in a certain sense opposed to the law of heredity. It asserts that the resemblance which offspring bear to their parents is never exact. The contradiction between the two laws is only apparent. All variations and modifications in an organism are directly or indirectly due to its environments; that is to say, they are either produced by some direct influence acting upon the organism itself, or by some more subtle and mysterious action on its parents; and the law of heredity really asserts that the offspring and parent would resemble each other if their environments were the same. Since, however, this is never the case, the offspring always differ to some extent from the parents. Now, according to the law of heredity, every acquired variation tends to be inherited, so that, by a summation of small changes, the animals may come to differ from their parent stock to an indefinite extent.

We are now in a position to follow out the consequences of these two laws in their bearing on development. Their application will best be made apparent by taking a concrete example. Let us suppose a spot on the surface of some very simple organism to become, at a certain period of life, pigmented, and therefore to be especially sensitive to light. In the offspring of this form, the pigment-spot will reappear at a corresponding period; and there will therefore be a period in the life of the offspring during which there is no pigment-spot, and a second period in which there is one. If a naturalist were to study the life-history, or, in other words, the embryology of this form, this fact about the pigment-spot would come to his notice, and he would be justified, from the laws of heredity, in concluding that the species was descended from an ancestor without a pigment-spot, because a pigment-spot was absent in the young. Now, we may suppose the transparent layer of skin above the pigment-spot to become thickened, so as gradually to form a kind of lens, which would throw an image of external objects on the pigment-spot. In this way a rudimentary eye might be evolved out of the pigment-spot. A naturalist studying the embryology of the form with this eye would find that the pigment-spot was formed before the lens, and he would be justified in concluding, by the same process of reasoning as before, that the ancestors of the form he was studying first acquired a pigment-spot and then a lens. We may picture to ourselves a series of steps by which the simple eye, the origin of which I have traced, might become more complicated; and it is easy to see how an embryologist studying the actual development of this complicated eye would be able to unravel the process of its evolution.

The general nature of the methods of reasoning employed by embryologists, who accept the Darwinian theory, is exemplified by the instance just given. If this method is a legitimate one, and there is no reason to doubt it, we ought to find that animals, in the course of their development, pass through a series of stages, in each of which they resemble one of their remote ancestors; but it is to be remembered that, in accordance with the law of variation, there is a continual tendency to change, and that the longer this tendency acts the greater will be the total effect. Owing to this tendency, we should not expect to find a perfect resemblance between an animal, at different stages of its growth, and its ancestors; and the remoter the ancestors, the less close ought the resemblance to be. In spite, however, of this limitation, it may be laid down as one of the consequences of the law of inheritance that every animal ought, in the course of its individual development, to repeat with more or less fidelity the history of its ancestral evolution.

A direct verification of this proposition is scarcely possible. There is ample ground for concluding that the forms from which existing animals are descended have in most instances perished; and although there is no reason why they should not have been preserved in a fossil state, yet, owing to the imperfection of the geological record, palæontology is not so often of service as might have been hoped.

While, for the reasons just stated, it is not generally possible to prove by direct observation that existing forms in their embryonic state repeat the characters of their ancestors, there is another method by which the truth of this proposition can be approximately verified.

A comparison of recent and fossil forms shews that there are actually living at the present day representatives of a considerable proportion of the groups which have in previous times existed on the globe, and there are therefore forms allied to the ancestors of those living at the present day, though not actually the same species. If therefore it can be shewn that the embryos of existing forms pass through stages in which they have the characters of more primitive groups, a sufficient proof of our proposition will have been given.

That such is often the case is a well-known fact, and was even known before the publication of Darwin's works. Von Baer, the greatest embryologist of the century, who died at an advanced age but a few years ago, discussed the proposition at considerable length in a work published between the years 1830 and 1840. He came to the conclusion that the embryos of higher forms never actually resemble lower forms, but only the embryos of lower forms; and he further maintained that such resemblances did not hold at all, or only to a very small extent, beyond the limits of the larger groups. Thus he believed that, though the embryos of Vertebrates might agree amongst themselves, there was no resemblance between them and the embryos of any invertebrate group. We now know that these limitations of Von Baer do not hold good, but it is to be remembered that the meaning _now_ attached by embryologists to such resemblances was quite unknown to him.

These preliminary remarks will, I trust, be sufficient to demonstrate how completely modern embryological reasoning is dependent on the two laws of inheritance and variation, which constitute the keystones of the Darwinian theory.

Before the appearance of the _Origin of Species_ many very valuable embryological investigations were made, but the facts discovered were to their authors merely so many ultimate facts, which admitted of being classified, but could not be explained. No explanation could be offered of why it is that animals, instead of developing in a simple and straightforward way, undergo in the course of their growth a series of complicated changes, during which they often acquire organs which have no function, and which, after remaining visible for a short time, disappear without leaving a trace.

No explanation, for instance, could be offered of why it is that a frog in the course of its growth has a stage in which it breathes like a fish, and then why it is like a newt with a long tail, which gradually becomes absorbed, and finally disappears. To the Darwinian the explanation of such facts is obvious. The stage when the tadpole breathes by gills is a repetition of the stage when the ancestors of the frog had not advanced in the scale of development beyond a fish, while the newt-like stage implies that the ancestors of the frog were at one time organized very much like the newts of to-day. The explanation of such facts has opened out to the embryologist quite a new series of problems. These problems may be divided into two main groups, technically known as those of phylogeny and those of organogeny. The problems of phylogeny deal with the genealogy of the animal kingdom. A complete genealogy would form what is known as a natural classification. To attempt to form such a classification has long been the aim of a large number of naturalists, and it has frequently been attempted without the aid of embryology. The statements made in the earlier part of my address clearly shew how great an assistance embryology is capable of giving in phylogeny; and as a matter of fact embryology has been during the last few years very widely employed in all phylogenetic questions, and the results which have been arrived at have in many cases been very striking. To deal with these results in detail would lead me into too technical a department of my subject; but I may point out that amongst the more striking of the results obtained _entirely_ by embryological methods is the demonstration that the Vertebrata are not, as was nearly universally believed by older naturalists, separated by a wide gulf from the Invertebrata, but that there is a group of animals, known as the Ascidians, formerly united with the Invertebrata, which are now universally placed with the Vertebrata.

The discoveries recently made in organogeny, or the genesis of organs, have been quite as striking, and in many respects even more interesting, than those in phylogeny, and I propose devoting the remainder of my address to a history of results which have been arrived at with reference to the origin of the nervous system.

To render clear the nature of these results I must say a few words as to the structure of the animal body. The body is always built of certain pieces of protoplasm, which are technically known to biologists as cells. The simplest organisms are composed either of a single piece of this kind, or of several similar pieces loosely aggregated together. Each of these pieces or cells is capable of digesting and assimilating food, and of respiring; it can execute movements, and is sensitive to external stimuli, and can reproduce itself. All the functions of higher animals can, in fact, be carried on in this single cell. Such lowly organized forms are known to naturalists as the Protozoa. All other animals are also composed of cells, but these cells are no longer complete organisms in themselves. They exhibit a division of labour: some carrying on the work of digestion; some, which we call nerve-cells, receiving and conducting stimuli; some, which we call muscle-cells, altering their form--in fact, contracting in one direction--under the action of the stimuli brought to them by the nerve-cells. In most cases a number of cells with the same function are united together, and thus constitute a tissue. Thus the cells which carry on the work of digestion form a lining membrane to a tube or sack, and constitute a tissue known as a secretory epithelium. The whole of the animals with bodies composed of definite tissues of this kind are known as the Metazoa.

A considerable number of early developmental processes are common to the whole of the Metazoa.

In the first place every Metazoon commences its existence as a simple cell, in the sense above defined; this cell is known as the ovum. The first developmental process which takes place consists in the division or segmentation of the single cell into a number of smaller cells. The cells then arrange themselves into two groups or layers known to embryologists as the _primary germinal layers_. These two layers are usually placed one within the other round a central cavity. The inner of the two is called the hypoblast, the outer the epiblast. The existence of these two layers in the embryos of vertebrated animals was made out early in the present century by Pander, and his observations were greatly extended by Von Baer and Remak. But it was supposed that these layers were confined to vertebrated animals. In the year 1849, and at greater length in 1859, Huxley demonstrated that the bodies of all the polype tribe or Coelenterata--that is to say of the group to which the common polype, jelly-fish and the sea-anemone belong--were composed of two layers of cells, and stated that in his opinion these two layers were homologous with the epiblast and hypoblast of vertebrate embryos. This very brilliant discovery came before its time. It fell upon barren ground, and for a long time bore no fruit. In the year 1866 a young Russian naturalist named Kowalevsky began to study by special histological methods the development of a number of invertebrated forms of animals, and discovered that at an early stage of development the bodies of all these animals were divided into germinal layers like those in vertebrates. Biologists were not long in recognizing the importance of these discoveries, and they formed the basis of two remarkable essays, one by our own countryman, Professor Lankester, and the other by a distinguished German naturalist, Professor Haeckel, of Jena.

In these essays the attempt was made to shew that the stage in development already spoken of, in which the cells are arranged in the form of two layers enclosing a central cavity has an ancestral meaning, and that it is to be interpreted to signify that all the Metazoa are descended from an ancestor which had a more or less oval form, with a central digestive cavity provided with a single opening, serving both for the introduction of food and for the ejection of indigestible substances. The body of this ancestor was supposed to have been a double-walled sack formed of an inner layer, the hypoblast, lining the digestive cavity, and an outer layer, the epiblast. To this form Haeckel gave the name of gastræa or gastrula.

There is every reason to think that Lankester and Haeckel were quite justified in concluding that a form more or less like that just described was the ancestor of the Metazoa; but the further speculations contained in their essays as to the origin of this form from the Protozoa can only be regarded as suggestive feelers, which, however, have been of great importance in stimulating and directing embryological research. It is, moreover, very doubtful whether there are to be found in the developmental histories of most animals any traces of this gastræa ancestor, other than the fact of their passing through a stage in which the cells are divided into two germinal layers.

The key to the nature of the two germinal layers is to be found in Huxley's comparison between them, and the two layers in the fresh-water polype and the sea-anemone. The epiblast is the primitive skin, and the hypoblast is the primitive epithelial wall of the alimentary tract.

In the whole of the polype group, or Coelenterata, the body remains through life composed of the two layers, which Huxley recognized as homologous with the epiblast and hypoblast of the Vertebrata; but in all the higher Metazoa a third germinal layer, known as the mesoblast, early makes its appearance between the two primary layers. The mesoblast originates as a differentiation of one or of both the primary germinal layers; but although the different views which have been held as to its mode of origin form an important section of the history of recent embryological investigations, I must for the moment confine myself to saying that from this layer there take their origin--the whole of the muscular system, of the vascular system, and of that connective-tissue system which forms the internal skeleton, tendons, and other parts.

We have seen that the epiblast represents the skin or epidermis of the simple sack-like ancestor common to all the Metazoa. In all the higher Metazoa it gives rise, as might be expected, to the epidermis, but it gives rise at the same time to a number of other organs; and, in accordance with the principles laid down in the earlier part of my address, it is to be concluded that _the organs so derived have been formed as differentiations of the primitive epidermis_. One of the most interesting of recent embryological discoveries is the fact that the nervous system is, in all but a very few doubtful cases, derived from the epiblast. This fact was made out for vertebrate animals by the great embryologist Von Baer; and the Russian naturalist Kowalevsky, to whose researches I have already alluded, shewed that this was true for a large number of invertebrate animals. The derivation of the nervous system from the epiblast has since been made out for a sufficient number of forms satisfactorily to establish the generalization that it is all but universally derived from the epiblast.

In any animal in which there is no distinct nervous system, it is obvious that the general surface of the body must be sensitive to the action of its surroundings, or to what are technically called stimuli. We know experimentally that this is so in the case of the Protozoa, and of some very simple Metazoa, such as the freshwater Polype or Hydra, where there is no distinct nervous system. The skin or epidermis of the ancestor of the Metazoa was no doubt similarly sensitive; and the fact of the nervous system being derived from the epiblast implies that the functions of the central nervous system, which were originally taken by the whole skin, became gradually concentrated in a special part of the skin which was step by step removed from the surface, and finally became a well-defined organ in the interior of the body.

What were the steps by which this remarkable process took place? How has it come about that there are nerves passing from the central nervous system to all parts of the skin, and also to the muscles? How have the arrangements for reflex actions arisen by which stimuli received on the surface of the body are carried to the central part of the nervous system, and are thence transmitted to the appropriate muscles, and cause them to contract? All these questions require to be answered before we can be said to possess a satisfactory knowledge of the origin of the nervous system. As yet, however, the knowledge of these points derived from embryology is imperfect, although there is every hope that further investigation will render it less so. Fortunately, however, a study of comparative anatomy, especially that of the Coelenterata, fills up some of the gaps left from our study of embryology.

From embryology we learn that the ganglion-cells of the central part of the nervous system are originally derived from the simple undifferentiated epithelial cells of the surface of the body. We further learn that the nerves are out-growths of the central nervous system. It was supposed till quite recently that the nerves in Vertebrates were derived from parts of the middle germinal layer or mesoblast, and that they only became secondarily connected with the central nervous system. This is now known not to be the case, but the nerves are formed as processes growing out from the central part of the nervous system.

Another important fact shewn by embryology is that the central nervous system, and percipient portion of the organs of special sense, are often formed from the same part of the primitive epidermis. Thus, in ourselves and in other vertebrate animals the sensitive part of the eye, known as the retina, is formed from two lateral lobes of the front part of the primitive brain. The crystalline lens and cornea of the eye are, however, subsequently formed from the skin.

The same is true for the peculiar compound eyes of crabs or Crustacea. The most important part of the central nervous system of these animals is the supra-oesophageal ganglia, often known as the brain, and these are formed in the embryo from two thickened patches of the skin at the front end of the body. These thickened patches become gradually detached from the surface, remaining covered over by a layer of skin. They then constitute the supra-oesophageal ganglia; but they form not only the ganglia, but also the rhabdons or retinal elements of the eye--the parts in fact which correspond to the rods and cones in our own retina. The layer of epidermis or skin which lies immediately above the supra-oesophageal ganglia becomes gradually converted into the refractive media of the crustacean eye. A cuticle which lies on its surface forms the peculiar facets on the surface of the eye, which are known as the corneal lenses, while the cells of the epidermis give rise to lens-like bodies known as the crystalline cones.

It would be easy to quote further instances of the same kind, but I trust that the two which I have given will be sufficient to shew the kind of relation which often exists between the organs of special sense, especially those of vision, and the central nervous system. It might have been anticipated _à priori_ that organs of special sense would only appear in animals provided with a well-developed central nervous system. This, however, is not the case. Special cells, with long delicate hairs, which are undoubtedly highly sensitive structures, are present in animals in which as yet nothing has been found which could be called a central nervous system; and there is every reason to think that the organs of special sense originated _pari passu_ with the central nervous system. It is probable that in the simplest organisms the whole body is sensitive to light, but that with the appearance of pigment-cells in certain parts of the body, the sensitiveness to light became localised to the areas where the pigment-cells were present. Since, however, it was necessary that stimuli received by such organs should be communicated to other parts of the body, some of the epidermic cells in the neighbourhood of the pigment-spots, which were at first only sensitive, in the same manner as other cells of the epidermis, became gradually differentiated into special nerve-cells. As to the details of this differentiation, embryology does not as yet throw any great light; but from the study of comparative anatomy there are grounds for thinking that it was somewhat as follows:--Cells placed on the surface sent protoplasmic processes of a nervous nature inwards, which came into connection with nervous processes from similar cells placed in other parts of the body. The cells with such processes then became removed from the surface, forming a deeper layer of the epidermis below the sensitive cells of the organ of vision. With these cells they remained connected by protoplasmic filaments, and thus they came to form a thickening of the epidermis underneath the organ of vision, the cells of which received their stimuli from those of the organ of vision, and transmitted the stimuli so received to other parts of the body. Such a thickening would obviously be the rudiment of a central nervous system, and it is easy to see by what steps it might become gradually larger and more important, and might gradually travel inwards, remaining connected with the sense organ at the surface by protoplasmic filaments, which would then constitute nerves. The rudimentary eye would at first merely consist partly of cells sensitive to light, and partly of optical structures constituting the lens, which would throw an image of external objects upon it, and so convert the whole structure into a true organ of vision. It has thus come about that, in the development of the individual, the retina or sensitive part of the eye is first formed in connection with the central nervous system, while the lenses of the eye are independently evolved from the epidermis at a later period.

The general features of the origin of the nervous system which have so far been made out by means of the study of embryology are the following:--

(1) That the nervous system of the higher Metazoa has been developed in the course of a long series of generations by a gradual process of differentiation of parts of the epidermis.

(2) That part of the central nervous system of many forms arose as a local collection of nerve-cells in the epidermis, in the neighbourhood of rudimentary organs of vision.

(3) That ganglion cells have been evolved from simple epithelial cells of the epidermis.

(4) That the primitive nerves were outgrowths of the original ganglion cells; and that the nerves of the higher forms are formed as outgrowths of the central nervous system.

The points on which embryology has not yet thrown a satisfactory light are:--

(1) The steps by which the protoplasmic processes, from the primitive epidermic cells, became united together so as to form a network of nerve-fibres, placing the various parts of the body in nervous communication.

(2) The process by which nerves became connected with muscles, so that a stimulus received by a nerve-cell could be communicated to and cause a contraction in a muscle.

Recent investigations on the anatomy of the Coelenterata, especially of jelly-fish and sea-anemones, have thrown some light on these points, although there is left much that is still obscure.

In our own country Mr Romaines has conducted some interesting physiological experiments on these forms; and Professor Schäfer has made some important histological investigations upon them. In Germany a series of interesting researches have also been made on them by Professors Kleinenberg, Claus and Eimer, and more especially by the brothers Hertwig, of Jena. Careful histological investigations, especially those of the last-named authors, have made us acquainted with the forms of some very primitive types of nervous system. In the common sea-anemones there are, for instance, no organs of special sense, and no definite central nervous system. There are, however, scattered throughout the skin, and also throughout the lining of the digestive tract, a number of specially modified epithelial cells, which are no doubt delicate organs of sense. They are provided at their free extremity with a long hair, and are prolonged on their inner side into a fine process which penetrates the deeper part of the epithelial layer of the skin or digestive wall. They eventually join a fine network of protoplasmic fibres which forms a special layer immediately within the epithelium. The fibres of this network are no doubt essentially nervous. In addition to fibres there are, moreover, present in the network cells of the same character as the multipolar ganglion-cells in the nervous system of Vertebrates, and some of these cells are characterized by sending a process into the superjacent epithelium. Such cells are obviously epithelial cells in the act of becoming nerve-cells; and it is probable that the nerve-cells are, in fact, sense-cells which have travelled inwards and lost their epithelial character.

There is every reason to think that the network just described is not only continuous with the sense-cells in the epithelium, but that it is also continuous with epithelial cells which are provided with muscular prolongations. The nervous system thus consists of a network of protoplasmic fibres, continuous on the one hand with sense-cells in the epithelium, and on the other with muscular cells. The nervous network is generally distributed both beneath the epithelium of the skin and that of the digestive tract, but is especially concentrated in the disc-like region between the mouth and tentacles. The above observations have thrown a very clear light on the characters of the nervous system at an early stage of its evolution, but they leave unanswered the questions (1) how the nervous network first arose, and (2) how its fibres became continuous with muscles. It is probable that the nervous network took its origin from processes of the sense-cells. The processes of the different cells probably first met and then fused together, and, becoming more arborescent, finally gave rise to a complicated network.

The connection between this network and the muscular cells also probably took place by a process of contact and fusion.

Epithelial cells with muscular processes were discovered by Kleinenberg before epithelial cells with nervous processes were known, and he suggested that the epithelial part of such cells was a sense-organ, and that the connecting part between this and the contractile processes was a rudimentary nerve. This ingenious theory explained completely the fact of nerves being continuous with muscles; but on the further discoveries being made which I have just described, it became obvious that this theory would have to be abandoned, and that some other explanation would have to be given of the continuity between nerves and muscles. The hypothetical explanation just offered is that of fusion.

It seems very probable that many of the epithelial cells were originally provided with processes the protoplasm of which, like that of the Protozoa, carried on the functions of nerves and muscles at the same time, and that these processes united amongst themselves into a network. By a process of differentiation parts of this network may have become specially contractile, and other parts may have lost their contractility and become solely nervous. In this way the connection between nerves and muscles might be explained, and this hypothesis fits in very well with the condition of the neuro-muscular system as we find it in the Coelenterata.

The nervous system of the higher Metazoa appears then to have originated from a differentiation of some of the superficial epithelial cells of the body, though it is possible that some parts of the system may have been formed by a differentiation of the alimentary epithelium. The cells of the epithelium were most likely at the same time contractile and sensory, and the differentiation of the nervous system may very probably have commenced, in the first instance, from a specialization in the function of part of a network formed of neuro-muscular prolongations of epithelial cells. A simultaneous differentiation of other parts of the network into muscular fibres may have led to the continuity at present obtaining between nerves and muscles.

Local differentiations of the nervous network, which was no doubt distributed over the whole body, took place on the formation of organs of special sense, and such differentiations gave rise to the formation of a central nervous system. The central nervous system was at first continuous with the epidermis, but became separated from it and travelled inwards. Ganglion-cells took their origin from sensory epithelial cells, provided with prolongations, continuous with the nervous network. Such epithelial cells gradually lost their epithelial character, and finally became completely detached from the epidermis.

Nerves, such as we find them in the higher types, originated from special differentiations of the nervous network, radiating from the parts of the central nervous system.

Such, briefly, is the present state of our knowledge as to the genesis of the nervous system. I ought not, however, to leave this subject without saying a few words as to the hypothetical views which the distinguished evolutionist Mr Herbert Spencer has put forward on this subject in his work on Psychology.

For Herbert Spencer nerves have originated, not as processes of epithelial cells, but from the passage of motion along the lines of least resistance. The nerves would seem, according to this view, to have been formed in any tissue from the continuous passage of nervous impulses through it. "A wave of molecular disturbance," he says, "passing along a tract of mingled colloids closely allied in composition, and isomerically transforming the molecules of one of them, will be apt at the same time to form some new molecules of the same type," and thus a nerve becomes established.

A nervous centre is formed, according to Herbert Spencer, at the point in the colloid in which nerves are generated, where a single nervous wave breaks up, and its parts diverge along various lines of least resistance. At such points some of the nerve-colloid will remain in an amorphous state, and as the wave of molecular motion will there be checked, it will tend to cause decompositions amongst the unarranged molecules. The decompositions must, he says, cause "additional molecular motion to be disengaged; so that along the outgoing lines there will be discharged an augmented wave. Thus there will arise at this point something having the character of a ganglion corpuscle."

These hypotheses of Herbert Spencer, which have been widely adopted in this country, are, it appears to me, not borne out by the discoveries to which I have called your attention to-day. The discovery that nerves have been developed from processes of epithelial cells, gives a very different conception of their genesis to that of Herbert Spencer, which makes them originate from the passage of nervous impulses through a tract of mingled colloids; while the demonstration that ganglion-cells arose as epithelial cells of special sense, which have travelled inwards from the surface, admits still less of a reconciliation with Herbert Spencer's view on the same subject.

Although the present state of our knowledge on the genesis of the nervous system is a great advance on that of a few years ago, there is still much remaining to be done to make it complete.

The subject is well worth the attention of the morphologist, the physiologist, or even of the psychologist, and we must not remain satisfied by filling up the gaps in our knowledge by such hypotheses as I have been compelled to frame. New methods of research will probably be required to grapple with the problems that are still unsolved; but when we look back and survey what has been done in the past, there can be no reason for mistrusting our advance in the future.

XX. ON THE DEVELOPMENT OF THE SKELETON OF THE PAIRED FINS OF ELASMOBRANCHII, CONSIDERED IN RELATION TO ITS BEARINGS ON THE NATURE OF THE LIMBS OF THE VERTEBRATA[479].

Footnote 479: From the _Proceedings of the Zoological Society of London_, 1881.

(With Plate 33.)

Some years ago the study of the development of the soft parts of the fins in several Elasmobranch types, more especially in _Torpedo,_ led me to the conclusion that the vertebrate limbs were remnants of two continuous lateral fins[480]. More or less similar views (which I was not at that time acquainted with) had been previously held by Maclise, Humphrey, and other anatomists; these views had not, however, met with much acceptance, and diverge in very important points from those put forward by me. Shortly after the appearance of my paper, J. Thacker published two interesting memoirs comparing the skeletal parts of the paired and unpaired fins[481].

Footnote 480: "Monograph on the Development of Elasmobranch Fishes," pp. 319, 320.

Footnote 481: J. K. Thacker, "Median and Paired Fins; a Contribution to the History of the Vertebrate Limbs," _Trans. of the Connecticut Acad._ Vol. III. 1877. "Ventral Fins of Ganoids," _Trans. of the Connecticut Acad._ Vol. IV. 1877.

In these memoirs Thacker arrives at conclusions as to the nature of the fins in the main similar to mine, but on entirely independent grounds. He attempts to shew that the structure of the skeleton of the paired fins is essentially the same as that of the unpaired fins, and in this comparison lays special stress on the very simple skeleton of the pelvic fin in the cartilaginous Ganoids, more especially in _Acipenser_ and _Polyodon_. He points out that the skeleton of the pelvic fin of _Polyodon_ consists essentially of a series of nearly isolated rays, which have a strikingly similar arrangement to that of the rays of the skeleton in many unpaired fins. He sums up his views in the following way[482]:--

Footnote 482: _Loc. cit._ p. 298.

"As the dorsal and anal fins were specializations of the median folds of _Amphioxus,_ so the paired fins were specializations of the two lateral folds which are supplementary to the median in completing the circuit of the body. These lateral folds, then, are the homologues of Wolffian ridges, in embryos of higher forms. Here, as in the median fins, there were formed chondroid and finally cartilaginous rods. These became at least twice segmented. The orad ones, with more or less concrescence proximally, were prolonged inwards. The cartilages spreading met in the middle line; and a later extension of the cartilages dorsad completed the limb-girdle.

"The limbs of the Protognathostomi consisted of a series of parallel articulated cartilaginous rays. They may have coalesced somewhat proximally and orad. In the ventral pair they had extended themselves mesiad until they had nearly or quite met and formed the hip-girdle; they had not here extended themselves dorsad. In the pectoral limb the same state of things prevailed, but was carried a step further, namely, by the dorsal extension of the cartilage constituting the scapular portion, thus more nearly forming a ring or girdle."

The most important point in Thacker's theories which I cannot accept is the derivation of the folds, of which the paired fins of the Vertebrata are supposed to be specializations, from the lateral folds of _Amphioxus;_ and Thacker himself recognizes that this part of his theory stands on quite a different footing to the remainder.

Not long after the publication of Thacker's paper, an important memoir was published by Mivart in the _Transactions_ of this Society[483]. The object of the researches recorded in this paper was, as Mivart explains, to test how far the hard parts of the limbs and of the azygos fins may have arisen through centripetal chondrifications or calcifications, and so be genetically exoskeletal[484].

Footnote 483: St George Mivart, "On the Fins of Elasmobranchii," _Zoological Trans._ Vol. X.

Footnote 484: Mivart used the term exoskeletal in an unusual and (as it appears to me) inconvenient manner. The term is usually applied to dermal skeletal structures; but the skeleton of the limbs, with which we are here concerned, is undoubtedly not of this nature.

Mivart's investigations and the majority of his views were independent of Thacker's memoir; but he acknowledges that he has derived from Thacker the view that pelvic and pectoral girdles, as well as the skeleton of the limbs, may have arisen independently of the axial skeleton.

The descriptive part of Mivart's paper contains an account of the structure of a great variety of interesting and undescribed types of paired and unpaired fins, mainly of Elasmobranchii. The following is the summary given by Mivart of the conclusions at which he has arrived[485]:--

Footnote 485: _Loc. cit._ p. 480.

"1. Two continuous lateral longitudinal folds were developed, similar to dorsal and ventral median longitudinal folds.

"2. Separate narrow solid supports (radials), in longitudinal series, and with their long axes directed more or less outwards at right angles with the long axis of the body, were developed in varying extents in all these four longitudinal folds.

"3. The longitudinal folds became interrupted variously, but so as to form two prominences on each side, _i.e._ the primitive paired limbs.

"4. Each anterior paired limb increased in size more rapidly than the posterior limb.

"5. The bases of the cartilaginous supports coalesced as was needed, according to the respective practical needs of the different separate portions of the longitudinal folds, _i.e._ the respective needs of the several fins.

"6. Occasionally the dorsal radials coalesced (as in _Notidanus_, &c.) and sought centripetally (_Pristis_, &c.) adherence to the skeletal axis.

"7. The radials of the hinder paired limb did so more constantly, and ultimately prolonged themselves inwards by mesiad growth from their coalesced base, till the piscine pelvic structure arose, as, _e.g._, in _Squatina_.

"8. The pectoral radials with increasing development also coalesced proximally, and thence prolonging themselves inwards to seek a _point d'appui_, shot dorsad and ventrad to obtain a firm support, and at the same time to avoid the visceral cavity. Thus they came to abut dorsally against the axial skeleton, and to meet ventrally together in the middle line below.

"9. The lateral fins, as they were applied to support the body on the ground, became elongated, segmented, and narrowed, so that probably the line of the propterygium, or possibly that of the mesopterygium, became the cheiropterygial axis.

"10. The distal end of the incipient cheiropterygium either preserved and enlarged preexisting cartilages or developed fresh ones to serve fresh needs, and so grew into the developed cheiropterygium; but there is not yet enough evidence to determine what was the precise course of this transformation.

"11. The pelvic limb acquired a solid connection with the axial skeleton (a pelvic girdle) through its need of a _point d'appui_ as a locomotive organ on land.

"12. The pelvic limb became also elongated; and when its function was quite similar to that of the pectoral limb, its structure became also quite similar (_e.g. Ichthyosaurus_, _Plesiosaurus_, _Chelydra_, &c.); but for the ordinary quadrupedal mode of progression it became segmented and inflected in a way generally parallel with, but (from its mode of use) in part inversely to, the inflections of the pectoral limb."

Günther[486] has propounded a theory on the primitive character of the fins, which, on the whole, fits in with the view that the paired fins are structures of the same nature as the unpaired fins. The interest of Günther's views on the nature of the skeleton of the fins more especially depends upon the fact that he attempts to evolve the fin of _Ceratodus_ from the typical Selachian type of pectoral fin. His own statement on this subject is as follows[487]:--

Footnote 486: "Description of _Ceratodus_," _Phil. Trans._ 1871.

Footnote 487: _Loc. cit._ p. 534.

"On further inquiry into the more distant relations of the _Ceratodus_-limb, we may perhaps be justified in recognizing in it a modification of the typical form of the Selachian pectoral fin. Leaving aside the usual treble division of the carpal cartilage (which, indeed, is sometimes simple), we find that this shovel-like carpal forms the base for a great number of phalanges, which are arranged in more or less regular transverse rows (zones) and in longitudinal rows (series). The number of phalanges of the zones and series varies according to the species and the form of the fin; in _Cestracion philippi_ the greater number of phalanges is found in the proximal zones and middle series, all the phalanges decreasing in size from the base of the fin towards the margins. In a Selachian with a long, pointed, scythe-shaped pectoral fin, like that of _Ceratodus_, we may, from analogy, presume that the arrangement of the cartilages might be somewhat like that shewn in the accompanying diagram, which I have divided into nine zones and fifteen series.

"When we now detach the outermost phalanx from each side of the first horizontal zone, and with it the other phalanges of the same series, when we allow the remaining phalanges of this zone to coalesce into one piece (as, in nature, we find coalesced the carpals of _Ceratodus_ and many phalanges in Selachian fins), and when we repeat this same process with the following zones and outer series, we arrive at an arrangement identical with what we actually find in _Ceratodus_."

While the researches of Thacker and Mivart are strongly confirmatory of the view at which I had arrived with reference to the nature of the paired fins, other hypotheses as to the nature of the skeleton of the fins have been enunciated, both before and after the publication of my memoir, which are either directly or indirectly opposed to my view.

Huxley in his memoir on _Ceratodus_, which throws light on so many important morphological problems, has dealt with the nature of paired fins[488].

Footnote 488: T. H. Huxley, "On _Ceratodus Fosteri_, with some Observations on the Classification of Fishes," _Proc. Zool. Soc._ 1876.

He holds, in accordance with a view previously adopted by Gegenbaur, that the limb of _Ceratodus_ "presents us with the nearest known approximation to the fundamental form of vertebrate limb or archipterygium," and is of opinion that in a still more archaic fish than _Ceratodus_ the skeleton of the fin "would be made up of homologous segments, which might be termed pteromeres, each of which would consist of a mesomere with a preaxial and a postaxial paramere." He considers that the pectoral fins of Elasmobranchii, more especially the fin of _Notidanus_, which he holds to be the most primitive form of Elasmobranch fin, "results in the simplest possible manner from the shortening of the axis of such a fin-skeleton as that of _Ceratodus_, and the coalescence of some of its elements." Huxley does not enter into the question of the origin of the skeleton of the pelvic fin of Elasmobranchii.

It will be seen that Huxley's idea of the primitive structure of the archipterygium is not easily reconcilable with the view that the paired fins are parts of a once continuous lateral fin, in that the skeleton of such a lateral fin, if it has existed, must necessarily have consisted of a series of parallel rays.

Gegenbaur[489] has done more than any other living anatomist to elucidate the nature of the fins; and his views on this subject have undergone considerable changes in the course of his investigations. After Günther had worked out the structure of the fin of _Ceratodus_, Gegenbaur suggested that it constituted the most primitive _persisting_ type of fin, and has moreover formed a theory as to the origin of the fins founded on this view, to the effect that the fins, together with their respective girdles, are to be derived from visceral arches with their rays.

Footnote 489: C. Gegenbaur, _Untersuchungen z. vergleich. Anat. d. Wirbelthiere_ (Leipzig 1864-5): erstes Heft, "Carpus u. Tarsus;" zweites Heft, "Brustflosse d. Fische." "Ueb. d. Skelet d. Gliedmaassen d. Wirbelthiere im Allgemeinen u. d. Hintergliedmaassen d. Selachier insbesondere," _Jenaische Zeitschrift_, Vol. V. 1870. "Ueb. d. Archipterygium," _Jenaische Zeitschrift_, Vol. VII. 1873. "Zur Morphologie d. Gliedmaassen d. Wirbelthiere," _Morphologisches Jahrbuch_, Vol. II. 1876.

His views on this subject are clearly explained in the subjoined passages quoted from the English translation of his _Elements of Comparative Anatomy_, pp. 473 and 477.

"The skeleton of the free appendage is attached to the extremity of the girdle. When simplest, this is made up of cartilaginous rods (rays), which differ in their size, segmentation, and relation to one another. One of these rays is larger than the rest, and has a number of other rays attached to its sides. I have given the name of _archipterygium_ to the ground-form of the skeleton which extends from the limb-bearing girdle into the free appendage. The primary ray is the stem of this archipterygium, the characters of which enable us to follow out the lines of development of the skeleton of the appendage. Cartilaginous arches beset with the rays form the branchial skeleton. The form of skeleton of the appendages may be compared with them; and we are led to the conclusion that it is possible that they may have been derived from such forms. In the branchial skeleton of the Selachii the cartilaginous bars are beset with simple rays. In many a median one is developed to a greater size. As the surrounding rays become smaller, and approach the larger one, we get an intermediate step towards that arrangement in which the larger median ray carries a few smaller ones. This differentiation of one ray, which is thereby raised to a higher grade, may be connected with the primitive form of the appendicular skeleton; and as we compare the girdle with a branchial arch, so we may compare the median ray and its secondary investment of rays with the skeleton of the free appendage.

"All the varied forms which the skeleton of the free appendages exhibits may be derived from a ground-form which persists in a few cases only, and which represents the first, and consequently the lowest, stage of the skeleton in the fin--the _archipterygium_. This is made up of a stem which consists of jointed pieces of cartilage, which is articulated to the shoulder-girdle and is beset on either side with rays which are likewise jointed. In addition to the rays of the stem there are others which are directly attached to the limb-girdle.

"_Ceratodus_ has a fin-skeleton of this form; in it there is a stem beset with two rows of rays. But there are no rays in the shoulder-girdle. This biserial investment of rays on the stem of the fin may also undergo various kinds of modifications. Among the Dipnoi, _Protopterus_ retains the medial row of rays only, which have the form of fine rods of cartilage; in the Selachii, on the other hand, the lateral rays are considerably developed. The remains of the medial row are ordinarily quite small, but they are always sufficiently distinct to justify us in supposing that in higher forms the two sets of rays might be better developed. Rays are still attached to the stem and are connected with the shoulder-girdle by means of larger plates. The joints of the rays are sometimes broken up into polygonal plates which may further fuse with one another; concrescence of this kind may also affect the pieces which form the base of the fin. By regarding the free rays, which are attached to these basal pieces, as belonging to these basal portions, we are able to divide the entire skeleton of the fin into three segments--pro-, meso-, and metapterygium.

"The metapterygium represents the stem of the archipterygium and the rays on it. The propterygium and the mesopterygium are evidently derived from the rays which still remain attached to the shoulder-girdle."

Since the publication of the memoirs of Thacker, Mivart, and myself, a pupil of Gegenbaur's, M. v. Davidoff[490], has made a series of very valuable observations, in part directed towards demonstrating the incorrectness of our theoretical views, more especially Thacker's and Mivart's view of the genesis of the skeleton of the limbs. Gegenbaur[491] has also written a short paper in connection with Davidoff's memoir, in support of his own as against our views.

Footnote 490: M. v. Davidoff, "Beiträge z. vergleich. Anat. d. hinteren Gliedmaassen d. Fische, I.," _Morphol. Jahrbuch_, Vol. V. 1879.

Footnote 491: "Zur Gliedmaassenfrage. An die Untersuchungen von Davidoff's angeknüpfte Bemerkungen," _Morphol. Jahrbuch_, Vol. V. 1879.

It would not be possible here to give an adequate account of Davidoff's observations on the skeleton, muscular system, and nerves of the pelvic fins. His main argument against the view that the paired fins are the remains of a continuous lateral fin is based on the fact that a variable but often considerable number of the spinal nerves in front of the pelvic fin are united by a longitudinal commissure with the true plexus of the nerves supplying the fin. From this he concludes that the pelvic fin has shifted its position, and that it may once therefore have been situated close behind the visceral arches. Granting, however, that Davidoff's deduction from the character of the pelvic plexus is correct, there is, so far as I see, no reason in the nature of the lateral-fin theory why the pelvic fins should not have shifted; and, on the other hand, the longitudinal cord connecting some of the ventral roots in front of the pelvic fin may have another explanation. It may, for instance, be a remnant of the time when the pelvic fin had a more elongated form than at present, and accordingly extended further forwards.

In any case our knowledge of the nature and origin of nervous plexuses is far too imperfect to found upon their characters such conclusions as those of Davidoff.

Gegenbaur, in his paper above quoted, further urges against Thacker and Mivart's views the fact that there is no proof that the fin of _Polyodon_ is a primitive type; and also suggests that the epithelial line which I have found connecting the embryonic pelvic and pectoral fins in _Torpedo_ may be a rudiment indicating a migration backwards of the pelvic fin.

With reference to the development of the pectoral fin in the Teleostei there are some observations of 'Swirski[492], which unfortunately do not throw very much light upon the nature of the limb.

Footnote 492: G. 'Swirski, _Untersuch. üb. d. Entwick. d. Schultergürtels u. d. Skelets d. Brustflosse d. Hechts._ Inaug. Diss. Dorpat, 1880.

'Swirski finds that in the Pike the skeleton of the limb is formed of a plate of cartilage continuous with the pectoral girdle, which soon becomes divided into a proximal and a distal portion. The former is subsequently segmented into five basal rays, and the latter into twelve parts, the number of which subsequently becomes reduced.

* * * * *

The observations which I have to lay before the Society were made with the object of determining how far the development of the skeleton of the limbs throws light on the points on which the anatomists whose opinions have just been quoted are at variance.

They were made, in the first instance, to complete a chapter in my work on comparative embryology; and, partly owing to the press of other engagements, but still more to the difficulty of procuring material, my observations are confined to the two British species of the genus _Scyllium_, viz. _Sc. stellare_ and _Sc. canicula_; yet I venture to believe that the results at which I have arrived are not wholly without interest.

Before dealing with the development of the skeleton of the fin, it will be convenient to describe with great brevity the structure of the pectoral and pelvic fins of the adult. The pectoral fins consist of broad plates inserted horizontally on the sides of the body; so that in each there may be distinguished a dorsal and a ventral surface, and an anterior and a posterior border. Their shape may best be gathered from the woodcut (fig. 1); and it is to be especially noted that the narrowest part of the fin is the base, where it is attached to the side of the body. The cartilaginous skeleton only occupies a small zone at the base of the fin, the remainder being formed of a fringe supported by radiately arranged horny fibres[493].

Footnote 493: The horny fibres are mesoblastic products; they are formed, in the first instance, as extremely delicate fibrils on the inner side of the membrane separating the epiblast from the mesoblast.

The true skeleton consists of three basal pieces articulating with the pectoral girdle; on the outer side of which there is a series of more or less segmented cartilaginous fin-rays. Of the basal cartilages one (_pp_) is anterior, a second (_mep_) is placed in the middle, and a third is posterior (_mp_). They have been named by Gegenbaur the _propterygium_, the _mesopterygium_, and the _metapterygium_; and these names are now generally adopted.

The metapterygium is by far the most important of the three, and in _Scyllium canicula_ supports 12 or 13 rays[494]. It forms a large part of the posterior boundary of the fin, and bears rays only on its _anterior_ border.

Footnote 494: In one example where the metapterygium had 13 rays the mesopterygium had only 2 rays.

The mesopterygium supports 2 or 3 rays, in the basal parts of which the segmentation into distinct rays is imperfect; and the propterygium supports only a single ray.

The pelvic fins are horizontally placed, like the pectoral fins, but differ from the latter in nearly meeting each other along the median ventral line of the body. They also differ from the pectoral fins in having a relatively much broader base of attachment to the sides of the body. Their cartilaginous skeleton (woodcut, fig. 2) consists of a basal bar, placed parallel to the base of the fin, and articulated in front with the pelvic girdle.

On its outer border it articulates with a series of cartilaginous fin-rays. I shall call the basal bar the basipterygium. The rays which it bears are most of them less segmented than those of the pectoral fin, being only divided into two; and the posterior ray, which is placed in the free posterior border of the fin, continues the axis of the basipterygium. In the male it is modified in connection with the so-called clasper.

The anterior fin-ray of the pelvic fin, which is broader than the other rays, articulates directly with the pelvic girdle, instead of with the basipterygium. This ray, in the female of _Scyllium canicula_ and in the male of _Scyllium catulus_ (Gegenbaur), is peculiar in the fact that its distal segment is longitudinally divided into two or more pieces, instead of being single as is the case with the remaining rays. It is probably equivalent to two of the posterior rays.

_Development of the paired Fins._--The first rudiments of the limbs appear in _Scyllium_, as in other fishes, as slight longitudinal ridge-like thickenings of the epiblast, which closely resemble the first rudiments of the unpaired fins.

These ridges are two in number on each side--an anterior immediately behind the last visceral fold, and a posterior on the level of the cloaca. In most Fishes they are in no way connected; but in some Elasmobranch embryos, more especially in that of _Torpedo_, they are connected together at their first development by a line of columnar epiblast cells. This connecting line of columnar epiblast, however, is a very transitory structure. The rudimentary fins soon become more prominent, consisting of a projecting ridge both of epiblast and mesoblast, at the outer edge of which is a fold of epiblast only, which soon reaches considerable dimensions. At a later stage the mesoblast penetrates into this fold, and the fin becomes a simple ridge of mesoblast covered by epiblast. The pectoral fins are at first considerably ahead of the pelvic fins in development.

The direction of the original epithelial line which connected the two fins of each side is nearly, though not quite, longitudinal, sloping somewhat obliquely ventralwards. It thus comes about that the attachment of each pair of limbs is somewhat on a slant, and that the pelvic pair nearly meet each other in the median ventral line shortly behind the anus.

The embryonic muscle-plates, as I have elsewhere shewn, grow into the bases of the fins; and the cells derived from these ingrowths, which are placed on the dorsal and ventral surfaces in immediate contact with the epiblast, probably give rise to the dorsal and ventral muscular layers of the limb, which are shewn in section in Plate 33, fig. 1, _m_, and in Plate 33, fig. 7, _m_.

The cartilaginous skeleton of the limbs is developed in the indifferent mesoblast cells between the two layers of muscles. Its early development in both the pectoral and the pelvic fins is very similar. When first visible it differs histologically from the adjacent mesoblast simply in the fact of its cells being more concentrated; while its boundary is not sharply marked.

At this stage it can only be studied by means of sections. It arises simultaneously and continuously with the pectoral and pelvic girdles, and consists, in both fins, of a bar springing at right angles from the posterior side of the pectoral or pelvic girdle, and running parallel to the long axis of the body along the base of the fin. The outer side of this bar is continued into a thin plate, which extends into the fin.

The structure of the skeleton of the fin slightly after its first differentiation will be best understood from Plate 33, fig. 1, and Plate 33, fig. 7. These figures represent transverse sections through the pelvic and pectoral fins of the same embryo on the same scale. The basal bar is seen at _bp_, and the plate at this stage (which is considerably later than the first differentiation) already partially segmented into rays at _br_. Outside the region of the cartilaginous plate is seen the fringe with the horny fibres (_h.f._); and dorsally and ventrally to the cartilaginous skeleton are seen the already well-differentiated muscles (_m_).

The pectoral fin is shewn in horizontal section in Plate 33, fig. 6, at a somewhat earlier stage than that to which the transverse sections belong. The pectoral girdle (_p.g._) is cut transversely, and is seen to be perfectly continuous with the basal bar (_vp_) of the fin. A similar continuity between the basal bar of the pelvic fin and the pelvic girdle is shewn in Plate 33, fig. 2, at a somewhat later stage. The plate continuous with the basal bar of the fin is at first, to a considerable extent in the pectoral, and to some extent in the pelvic fin, a continuous lamina, which subsequently segments into rays. In the parts of the plate which eventually form distinct rays, however, almost from the first the cells are more concentrated than in those parts which will form the tissue between the rays; and I am not inclined to lay any stress whatever upon the fact of the cartilaginous fin-rays being primitively part of a continuous lamina, but regard it as a secondary phenomenon, dependent on the mode of conversion of embryonic mesoblast cells into cartilage. In all cases the separation into distinct rays is to a large extent completed before the tissue of which the plates are formed is sufficiently differentiated to be called cartilage by an histologist.

The general position of the fins in relation to the body, and their relative sizes, may be gathered from Plate 33, figs. 4 and 5, which represent transverse sections of the same embryo as that from which the transverse sections shewing the fin on a larger scale were taken.

During the first stage of its development the skeleton of both fins may thus be described as consisting of _a longitudinal bar running along the base of the fin, and giving off at right angles series of rays which pass into the fin_. The longitudinal bar may be called the basipterygium; and it is continuous in front with the pectoral or pelvic girdle, as the case may be.

The further development of the primitive skeleton is different in the case of the two fins.

_The Pelvic Fin._--The changes in the pelvic fin are comparatively slight. Plate 33, fig. 2, is a representation of the fin and its skeleton in a female of _Scyllium stellare_ shortly after the primitive tissue is converted into cartilage, but while it is still so soft as to require the very greatest care in dissection. The fin itself forms a simple projection of the side of the body. The skeleton consists of a basipterygium (_bp_), continuous in front with the pelvic girdle. To the outer side of the basipterygium a series of cartilaginous fin-rays are attached--the posterior ray forming a direct prolongation of the basipterygium, while the anterior ray is united rather with the pelvic girdle than with the basipterygium. All the cartilaginous fin-rays except the first are completely continuous with the basipterygium, their structure in section being hardly different from that shewn in Plate 33, fig. 1.

The external form of the fin does not change very greatly in the course of the further development; but the hinder part of the attached border is, to some extent, separated off from the wall of the body, and becomes the posterior border of the adult fin. With the exception of a certain amount of segmentation in the rays, the character of the skeleton remains almost as in the embryo. The changes which take place are illustrated by Plate 33, fig. 3, shewing the fin of a young male of _Scyllium stellare_. The basipterygium has become somewhat thicker, but is still continuous in front with the pelvic girdle, and otherwise retains its earlier characters. The cartilaginous fin-rays have now become segmented off from it and from the pelvic girdle, the posterior end of the basipterygial bar being segmented off as the terminal ray.

The anterior ray is directly articulated with the pelvic girdle, and the remaining rays continue articulated with the basipterygium. Some of the latter are partially segmented.

As may be gathered by comparing the figure of the fin at the stage just described with that of the adult fin (woodcut, fig. 2), the remaining changes are very slight. The most important is the segmentation of the basipterygial bar from the pelvic girdle.

The pelvic fin thus retains in all essential points its primitive structure.

_The Pectoral Fin._--The earliest stage of the pectoral fin differs, as I have shewn, from that of the pelvic fin only in minor points (Pl. 33, fig. 6). There is the same longitudinal or basipterygial bar (_bp_), to which the fin-rays are attached, which is continuous in front with the pectoral girdle (_pg_). The changes which take place in the course of the further development, however, are very much more considerable in the case of the pectoral than in that of the pelvic fin.

The most important change in the external form of the fin is caused by a reduction in the length of its attachment to the body. At first (Pl. 33, fig. 6), the base of the fin is as long as the greatest breadth of the fin; but it gradually becomes shortened by being constricted off from the body at its hinder end. In connection with this process the posterior end of the basipterygial bar is gradually rotated outwards, its anterior end remaining attached to the pectoral girdle. In this way this bar comes to form the posterior border of the skeleton of the fin (Pl. 33, figs. 8 and 9), constituting the metapterygium (_mp_). It becomes eventually segmented off from the pectoral girdle, simply articulating with its hinder edge.

The plate of cartilage, which is continued outwards from the basipterygium, or, as we may now call it, the metapterygium, into the fin, is not nearly so completely divided up into fin-rays as the homologous part of the pelvic fin; and this is especially the case with the basal part of the plate. This basal part becomes, in fact, at first only divided into two parts (Pl. 33, fig. 8)--a small anterior part at the front end (_me.p_), and a larger posterior along the base of the metapterygium (_mp_); and these two parts are not completely segmented from each other. The anterior part directly joins the pectoral girdle at its base, resembling in this respect the anterior fin-ray of the pelvic girdle. It constitutes the (at this stage undivided) rudiment of the mesopterygium and propterygium of Gegenbaur. It bears in my specimen of this age four fin-rays at its extremity, the anterior not being well marked. The remaining fin-rays are prolongations outwards of the edge of the plate continuous with the metapterygium. These rays are at the stage figured more or less transversely segmented; but at their outer edge they are united together by a nearly continuous rim of cartilage. The spaces between the fin-rays are relatively considerably larger than in the adult.

The further changes in the cartilages of the pectoral limb are, morphologically speaking, not important, and are easily understood by reference to Pl. 33, fig. 9 (representing the skeleton of the limb of a nearly ripe embryo). The front end of the anterior basal cartilage becomes segmented off as a propterygium (_pp_), bearing a single fin-ray, leaving the remainder of the cartilage as a mesopterygium (_mes_). The remainder of the now considerably segmented fin-rays are borne by the metapterygium.

* * * * *

_General Conclusions._--From the above observations, conclusions of a positive kind may be drawn as to the primitive structure of the skeleton; and the observations have also, it appears to me, important bearings on the theories of my predecessors in this line of investigation.

The most obvious of the positive conclusions is to the effect that the embryonic skeleton of the paired fins consists of a series of parallel rays similar to those of the unpaired fins. These rays support the soft parts of the fins, which have the form of a longitudinal ridge; and they are continuous at their base with a longitudinal bar. This bar, from its position at the base of the fin, can clearly never have been a median axis with the rays on both sides. It becomes the basipterygium in the pelvic fin, which retains its embryonic structure much more completely than the pectoral fin; and the metapterygium in the pectoral fin. The metapterygium of the pectoral fin is thus clearly homologous with the basipterygium of the pelvic fin, as originally supposed by Gegenbaur, and as has since been maintained by Mivart. The propterygium and mesopterygium are obviously relatively _unimportant_ parts of the skeleton as compared with the metapterygium.

My observations on the development of the skeleton of the fins certainly do not of themselves demonstrate that the paired fins are remnants of a once continuous lateral fin; but they support this view in that they shew the primitive skeleton of the fins to have exactly the character which might have been anticipated if the paired fins had originated from a continuous lateral fin. The longitudinal bar of the paired fins is believed by both Thacker and Mivart to be due to the coalescence of the bases of the primitively independent rays of which they believe the fin to have been originally composed. This view is probable enough in itself, and is rendered more so by the fact, pointed out by Mivart, that a longitudinal bar supporting the cartilaginous rays of unpaired fins is occasionally formed; but there is no trace in the embryo Scylliums of the bar in question being formed by the coalescence of rays, though the fact of its being perfectly continuous with the bases of the fin-rays is somewhat in favour of such coalescence.

Thacker and Mivart both hold that the pectoral and pelvic girdles are developed by ventral and dorsal growths of the anterior end of the longitudinal bar supporting the fin-rays.

There is, so far as I see, no theoretical objection to be taken to this view; and the fact of the pectoral and pelvic girdles originating continuously and long remaining united with the longitudinal bars of their respective fins is in favour of it rather than the reverse. The same may be said of the fact that the first part of each girdle to be formed is that in the neighbourhood of the longitudinal bar (basipterygium) of the fin, the dorsal and ventral prolongations being subsequent growths.

On the whole my observations do not throw much light on the theories of Thacker and Mivart as to the genesis of the skeleton of the paired fin; but, so far as they bear on the subject, they are distinctly favourable to those theories.

The main results of my observations appear to me to be decidedly adverse to the views recently put forward on the structure of the fin by Gegenbaur and Huxley, both of whom, as stated above, consider the primitive type of fin to be most nearly retained in _Ceratodus_, and to consist of a central multisegmented axis with numerous lateral rays.

Gegenbaur derives the Elasmobranch pectoral fin from a form which he calls the archipterygium, nearly like that of _Ceratodus_, with a median axis and two rows of rays--but holds that in addition to the rays attached to the median axis, which are alone found in _Ceratodus_, there were other rays directly articulated to the shoulder-girdle. He considers that in the Elasmobranch fin the majority of the lateral rays on the posterior (or median according to his view of the position of the limb) side have become aborted, and that the central axis is represented by the metapterygium; while the pro- and mesopterygium and their rays are, he believes, derived from those rays of the archipterygium which originally articulated directly with the shoulder-girdle.

This view appears to me to be absolutely negatived by the facts of development of the pectoral fin in _Scyllium_--not so much because the pectoral fin in this form is necessarily to be regarded as primitive, but because what Gegenbaur holds to be the primitive axis of the biserial fin is demonstrated to be really the base, and it is only in the adult that it is conceivable that a second set of lateral rays could have existed on the posterior side of the metapterygium. If Gegenbaur's view were correct, we should expect to find in the embryo, if anywhere, traces of the second set of lateral rays; but the fact is that, as may easily be seen by an inspection of figs. 6 and 7, such a second set of lateral rays could not possibly have existed in a type of fin like that found in the embryo. With this view of Gegenbaur's it appears to me that the theory held by this anatomist to the effect that the limbs are modified gill-arches also falls, in that his method of deriving the limbs from gill-arches ceases to be admissible, while it is not easy to see how a limb, formed on the type of the embryonic limb of Elasmobranchii, could be derived from a gill-arch with its branchial rays.

Gegenbaur's older view, that the Elasmobranch fin retains a primitive uniserial type, appears to me to be nearer the truth than his more recent view on this subject; though I hold the fundamental point established by the development of these parts in _Scyllium_ to be that the posterior border of the adult Elasmobranch pectoral fin is the primitive base-line,_i.e._line of attachment of the fin to the side of the body.

Huxley holds that the mesopterygium is the proximal piece of the axial skeleton of the limb of _Ceratodus_, and derives the Elasmobranch fin from that of _Ceratodus_ by the shortening of its axis and the coalescence of some of its elements. The entirely secondary character of the mesopterygium, and its total absence in the young embryo _Scyllium_, appear to me as conclusive against Huxley's view as the character of the embryonic fin is against that of Gegenbaur; and I should be much more inclined to hold that the fin of _Ceratodus_ has been derived from a fin like that of the Elasmobranchii by a series of steps similar to those which Huxley supposes to have led to the establishment of the Elasmobranch fin, but in exactly the reverse order.

There is one statement of Davidoff's which I cannot allow to pass without challenge. In comparing the skeletons of the paired and unpaired fins he is anxious to prove that the former are independent of the axial skeleton in their origin and that the latter have been segmented from the axial skeleton, and thus to shew that an homology between the two is impossible. In support of his view he states[495] that he has satisfied himself, from embryos of _Acanthias_ and _Scyllium_, that the rays of the unpaired fins _are undoubtedly products of the segmentation of the dorsal and ventral spinous processes_.

Footnote 495: _Loc. cit._ p. 514.

This statement is wholly unintelligible to me. From my examination of the development of the first dorsal and the anal fins of _Scyllium_ I find that their rays develop at a considerable distance from, and quite independently of, the neural and hæmal arches, and that they are at an early stage of development distinctly in a more advanced state of histological differentiation than the neural and hæmal arches of the same region. I have also found exactly the same in the embryos of _Lepidosteus_.

I have, in fact, no doubt that the skeleton of both the paired and the unpaired fins of Elasmobranchii and _Lepidosteus_ is in its development independent of the axial skeleton. The phylogenetic mode of origin of the skeleton both of the paired and of the unpaired fins cannot, however, be made out without further investigation.

EXPLANATION OF PLATE 33.[496]

Footnote 496: I employ here the same letters to indicate the stages as in my "Monograph on Elasmobranch Fishes."

Fig. 1. Transverse section through the pelvic fin of an embryo of _Scyllium_ belonging to stage P{1}, magnified 50 diameters. _bp._ basipterygium. _br._ fin ray. _m._ muscle. _hf._ horny fibres supporting the peripheral part of the fin.

Fig. 2. Pelvic fin of a very young female embryo of _Scyllium stellare_, magnified 16 diameters. _bp._ basipterygium. _pu._ pubic process of pelvic girdle (cut across below). _il._ iliac process of pelvic girdle. _fo._ foramen.

Fig. 3. Pelvic fin of a young male embryo of _Scyllium stellare_, magnified 16 diameters. _bp._ basipterygium. _mo._ process of basipterygium continued into clasper. _il._ iliac process of pelvic girdle. _pu._ pubic section of pelvic girdle.

Fig. 4. Transverse section through the ventral part of the trunk of an embryo _Scyllium_ of stage P, in the region of the pectoral fins, to shew how the fins are attached to the body, magnified 18 diameters. _br._ cartilaginous fin-ray. _bp._ basipterygium. _m._ muscle of fin. _mp._ muscle-plate.

Fig. 5. Transverse section through the ventral part of the trunk of an embryo _Scyllium_ of stage P, in the region of the pelvic fin, on the same scale as fig. 4. _bp._ basipterygium. _br._ cartilaginous fin-rays. _m._ muscle of the fins. _mp._ muscle-plate.

Fig. 6. Pectoral fin of an embryo of _Scyllium canicula_, of a stage between O and P, in longitudinal and horizontal section (the skeleton of the fin was still in the condition of embryonic cartilage), magnified 36 diameters. _bp._ basipterygium (eventual metapterygium). _fr._ cartilaginous fin-rays. _pg._ pectoral girdle in transverse section. _fo._ foramen in pectoral girdle. _pe._ epithelium of peritoneal cavity.

Fig. 7. Transverse section through the pectoral fin of a _Scyllium_ embryo of stage P, magnified 50 diameters. _bp._ basipterygium. _br._ cartilaginous fin-ray. _m._ muscle. _hf._ horny fibres.

Fig. 8. Pectoral fin of an embryo of _Scyllium stellare_, magnified 16 diameters. _mp._ metapterygium (basipterygium of earlier stage). _me.p._ rudiment of future pro- and mesopterygium. _sc._ cut surface of a scapular process. _cr._ coracoid process. _fr._ foramen. _hf._ horny fibres.

Fig. 9. Skeleton of the pectoral fin and part of pectoral girdle of a nearly ripe embryo of _Scyllium stellare_, magnified 10 diameters. _mp._ metapterygium. _mes._ mesopterygium. _pp._ propterygium. _cr._ coracoid process.

XXI. ON THE EVOLUTION OF THE PLACENTA, AND ON THE POSSIBILITY OF EMPLOYING THE CHARACTERS OF THE PLACENTA IN THE CLASSIFICATION OF THE MAMMALIA[497].

Footnote 497: From the _Proceedings of the Zoological Society of London_, 1881.

From Owen's observations on the Marsupials it is clear that the yolk-sack in this group plays an important (if not the most important) part, in absorbing the maternal nutriment destined for the foetus. The fact that in Marsupials both the yolk-sack and the allantois are concerned in rendering the chorion vascular, makes it _à priori_ probable that this was also the case in the primitive types of the Placentalia; and this deduction is supported by the fact that in the Rodentia, Insectivora, and Cheiroptera this peculiarity of the foetal membranes is actually found. In the primitive Placentalia it is also probable that from the discoidal allantoic region of the chorion simple foetal villi, like those of the Pig, projected into uterine crypts; but it is not certain how far the umbilical region of the chorion, which was no doubt vascular, may also have been villous. From such a primitive type of foetal membranes divergencies in various directions have given rise to the types of foetal membranes found at the present day.

In a general way it may be laid down that variations in any direction which tended to increase the absorbing capacities of the chorion would be advantageous. There are two obvious ways in which this might be done, viz. (1) by increasing the complexity of the foetal villi and maternal crypts over a limited area, (2) by increasing the area of the part of the chorion covered by the placental villi. Various combinations of the two processes would also, of course, be advantageous.

The most fundamental change which has taken place in all the existing Placentalia is the exclusion of the umbilical vesicle from any important function in the nutrition of the foetus.

The arrangement of the foetal parts in the Rodentia, Insectivora, and Cheiroptera may be directly derived from the primitive form by supposing the villi of the discoidal placental area to have become more complex, so as to form a deciduate discoidal placenta, while the yolk-sack still plays a part, though physiologically an unimportant part, in rendering the chorion vascular.

In the Carnivora, again, we have to start from the discoidal placenta, as evinced by the fact that in the growth of the placenta the allantoic region of the placenta is at first _discoidal_, and only becomes zonary at a later stage. A zonary deciduate placenta indicates an increase both in area and in complexity. The relative diminution of the breadth of the placental zone in late foetal life in the zonary placenta of the Carnivora is probably due to its being on the whole advantageous to secure the nutrition of the foetus by insuring a more intimate relation between the foetal and maternal parts, than by increasing their area of contact. The reason of this is not obvious, but, as shewn below, there are other cases where it is clear that a diminution in the area of the placenta has taken place, accompanied by an increase in the complexity of its villi.

The second type of differentiation from the primitive form of placenta is illustrated by the Lemuridæ, the Suidæ, and _Manis_. In all these cases the area of the placental villi appears to have increased so as to cover nearly the whole subzonal membrane, without the villi increasing to any great extent in complexity. From the diffused placenta covering the whole surface of the chorion, differentiations appear to have taken place in various directions. The placenta of Man and Apes, from its mode of ontogeny, is clearly derived from a diffused placenta (very probably similar to that of Lemurs) by a concentration of the foetal villi, which are originally spread over the whole chorion, to a disk-shaped area, and by an increase in their arborescence. Thus the discoidal placenta of Man has no connexion with, and ought not to be placed in, the same class as those of the Rodentia, Cheiroptera, and Insectivora.

The polycotyledonary forms of placenta are due to similar concentrations of the foetal villi of an originally diffused placenta.

In the Edentata we have a group with very varying types of placenta. Very probably these may all be differentiations within the group itself from a diffused placenta such as that found in _Manis_. The zonary placenta of _Orycteropus_ is capable of being easily derived from that of _Manis_ by the disappearance of the foetal villi at the two poles of the ovum. The small size of the umbilical vesicle in _Orycteropus_ indicates that its discoidal placenta is not, like that of the Carnivora, directly derived from a type with both allantoic and umbilical vascularization of the chorion. The discoidal and dome-shaped placentæ of the Armadillos, _Myrmecophaga_, and the Sloths may easily have been formed from a diffused placenta, just as the discoidal placenta of the Simiidæ and Hominidæ appears to have been formed from a diffused placenta like that of the Lemuridæ.

The presence of zonary placenta in _Hyrax_ and _Elephas_ does not necessarily afford any proof of affinity of these types with the Carnivora. A zonary placenta may be quite as easily derived from a diffused placenta as from a discoidal placenta; and the presence of two villous patches at the poles of the chorion in _Elephas_ very probably indicates that its placenta has been evolved from a diffused placenta.

Although it would not be wise to attempt to found a classification upon the placental characters alone, it may be worth while to make a few suggestions as to the affinities of the orders of Mammalia indicated by the structure of the placenta. We clearly, of course, have to start with forms which could not be grouped with any of the existing orders, but which might be called the Protoplacentalia. They probably had the primitive type of placenta described above: the nearest living representatives of the group are the Rodentia, Insectivora, and Cheiroptera. Before, however, these three groups had become distinctly differentiated, there must have branched off from the primitive stock the ancestors of the Lemuridæ, the Ungulata, and the Edentata.

It is obvious on general anatomical grounds that the Monkeys and Man are to be derived from a primitive Lemurian type; and with this conclusion the form of the placenta completely tallies. The primitive Edentata and Ungulata had no doubt a diffused placenta which was probably not very different from that of the primitive Lemurs; but how far these groups arose quite independently from the primitive stock, or whether they may have had a nearer common ancestor, cannot be decided from the structure of the placenta. The Carnivora were certainly an offshoot from the primitive placental type which was quite independent of the three groups just mentioned; but the character of the placenta of the Carnivora does not indicate at what stage in the evolution of the placental Mammalia a primitive type of Carnivora was first differentiated.

No important light is thrown by the placenta on the affinities of the Proboscidea, the Cetacea, or the Sirenia; but the character of the placenta in the latter group favours the view of their being related to the Ungulata.

XXII. ON THE STRUCTURE AND DEVELOPMENT OF LEPIDOSTEUS[498]. By F. M. BALFOUR and W. N. PARKER.

Footnote 498: From the _Philosophical Transactions of the Royal Society_, 1882.

(With Plates 34-42.)

TABLE OF CONTENTS.

PAGE

INTRODUCTION 739

GENERAL DEVELOPMENT 740

BRAIN-- Adult brain 759 Development of the brain 764 Comparison of the larval and adult brain of _Lepidosteus_, together with some observations on the systematic value of the characters of the Ganoid brain 767

SENSE ORGANS-- Olfactory organ 771 Anatomy of the eye _ib._ Development of the eye 772

SUCTORIAL DISC 774

MUSCULAR SYSTEM 775

SKELETON-- Vertebral column and ribs of the adult 776 Development of the vertebral column and ribs. 778 Comparison of the vertebral column of _Lepidosteus_ with that of other forms 792 The ribs of Fishes 793 The skeleton of the ventral lobe of the tail fin, and its bearing on the nature of the tail fin of the various types of Pisces 801

EXCRETORY AND GENERATIVE ORGANS-- Anatomy of the excretory and generative organs of the female 810 Anatomy of the excretory and generative organs of the male 813 Development of the excretory and generative organs 815 Theoretical considerations 822

THE ALIMENTARY CANAL AND ITS APPENDAGES-- Topographical anatomy of the alimentary canal 828 Development of the alimentary canal and its appendages 831

THE GILL ON THE HYOID ARCH 835

THE SYSTEMATIC POSITION OF LEPIDOSTEUS 836

LIST OF MEMOIRS ON THE ANATOMY AND DEVELOPMENT OF LEPIDOSTEUS 840

LIST OF REFERENCE LETTERS 841

EXPLANATION OF PLATES 842

INTRODUCTION.

The following paper is the outcome of the very valuable gift of a series of embryos and larvæ of _Lepidosteus_ by Professor Alex. Agassiz, to whom we take this opportunity of expressing our most sincere thanks. The skull of these embryos and larvæ has been studied by Professor Parker, and forms the subject of a memoir already presented to the Royal Society.

Considering that _Lepidosteus_ is one of the most interesting of existing Ganoids, and that it is very closely related to species of Ganoids which flourished during the Triassic period, we naturally felt keenly anxious to make the most of the opportunity of working at its development offered to us by Professor Agassiz' gift. Professor Agassiz, moreover, most kindly furnished us with four examples of the adult Fish, which have enabled us to make this paper a study of the adult anatomy as well as of the development.

The first part of our paper is devoted to the segmentation, formation of the germinal layers, and general development of the embryo and larva. The next part consists of a series of sections on the organs, in which both their structure in the adult and their development are dealt with. This part is not, however, in any sense a monograph, and where already known, the anatomy is described with the greatest possible brevity. In this part of the paper considerable space is devoted to a comparison of the organs of _Lepidosteus_ with those of other Fishes, and to a statement of the conclusions which follow from such comparison.

The last part of the paper deals with the systematic position of _Lepidosteus_ and of the Ganoids generally.

GENERAL DEVELOPMENT.

The spawning of _Lepidosteus_ takes place in the neighbourhood of New York about May 20th. Agassiz (No. 1)[499] gives an account of the process from Mr S. W. Garman's notes, which we venture to quote in full.

Footnote 499: The numbers refer to the list of memoirs of the anatomy and development given at the end of this memoir.

"Black Lake is well stocked with Bill-fish. When they appear, they are said to come in countless numbers. This is only for a few days in the spring, in the spawning season, between the 15th of May and the 8th of June. During the balance of the season they are seldom seen. They remain in the deeper parts of the lake, away from the shore, and, probably, are more or less nocturnal in habits. Out of season, an occasional one is caught on a hook baited with a minnow. Commencing with the 20th of April, until the 14th of May we were unable to find the Fish, or to find persons who had seen them during this time. Then a fisherman reported having seen one rise to the surface. Later, others were seen. On the afternoon of the 18th, a few were found on the _points_, depositing the spawn. The temperature at the time was 68° to 69° on the shoals, while out in the lake the mercury stood at 62° to 63°. The _points_ on which the eggs were laid were of naked granite, which had been broken by the frost and heat into angular blocks of 3 to 8 inches in diameter. The blocks were tumbled upon each other like loose heaps of brick-bats, and upon and between them the eggs were dropped. The _points_ are the extremities of small capes that make out into the lake. The eggs were laid in water varying in depth from 2 to 14 inches. At the time of approaching the shoals, the Fish might be seen to rise quite often to the surface to take air. This they did by thrusting the bill out of the water as far as the corners of the mouth, which was then opened widely and closed with a snap. After taking the air, they seemed more able to remain at the surface. Out in the lake they are very timid, but once buried upon the shoals they become quite reckless as to what is going on about them. A few moments after being driven off, one or more of the males would return as if scouting. If frightened, he would retire for some time; then another scout would appear. If all promised well, the females, with the attendant males, would come back. Each female was accompanied by from one to four males. Most often, a male rested against each side, with their bills reaching up toward the back of her head. Closely crowded together, the little party would pass back and forth over the rocky bed they had selected, sometimes passing the same spot half-a-dozen times without dropping an egg, then suddenly would indulge in an orgasm; and, lashing and plashing the water in all directions with their convulsive movements, would scatter at the same instant the eggs and the sperm. This ended, another season of moving slowly back and forth was observed, to be in turn followed by another of excitement. The eggs were excessively sticky. To whatever they happened to touch, they stuck, and so tenaciously that it was next to impossible to release them without tearing away a portion of their envelopes. It is doubtful whether the eggs would hatch if removed. As far as could be seen at the time, upon or under the rocks to which the eggs were fastened there was an utter absence of anything that might serve as food for the young Fishes.

"Other Fishes, Bull-heads, &c., are said to follow the Bill-fish to eat the spawn. It may be so. It was not verified. Certainly the points under observations were unmolested. During the afternoon of the 18th of May a few eggs were scattered on several of the beds. On the 19th there were more. With the spear and the snare, several dozens of both sexes of the Fish were taken. Taking one out did not seem greatly to startle the others. They returned very soon. The males are much smaller than the average size of the females; and, judging from those taken, would seem to have as adults greater uniformity in size. The largest taken was a female, of 4 feet 1-1/2 inch in length. Others of 2 feet 6 inches contained ripe ova. With the 19th of May all disappeared, and for a time--the weather being meanwhile cold and stormy--there were no signs of their continued existence to be met with. Nearly two weeks later, on the 31st of May, as stated by Mr Henry J. Perry, they again came up, not in small detachments on scattered points as before, but in multitudes, on every shoal at all according with their ideas of spawning beds. They remained but two days. During the summer it happens now and then that one is seen to come up for his mouthful of air; beyond this there will be nothing to suggest the ravenous masses hidden by the darkness of the waters."

_Egg membranes._--The ova of _Lepidosteus_ are spherical bodies of about 3 millims. in diameter. They have a double investment consisting of (1) an outer covering formed of elongated, highly refractive bodies, somewhat pyriform at their outer ends (Plate 34, fig. 17, _f.e._), which are probably metamorphosed follicular cells[500], and (2) of an inner membrane, divided into two zones, viz.: an outer and thicker zone, which is radially striated, and constitutes the _zona radiata (z.r.)_, and an inner and narrow homogeneous zone (_z.r´._).

Footnote 500: We have examined the structure of the ovarian ova in order to throw light on the nature of these peculiar pyriform bodies. Unfortunately, the ovaries of our adult examples of _Lepidosteus_ were so badly preserved, that we could not ascertain anything on this subject. The ripe ova in the ovary have an investment of pyriform bodies similar to those of the just laid ova. With reference to the structure of the ovarian ova we may state that the germinal vesicles are provided with numerous nucleoli arranged in close proximity with the membrane of the vesicle.

_Segmentation._--We have observed several stages in the segmentation, which shew that it is complete, but that it approaches the meroblastic type more nearly than in the case of any other known holoblastic ovum.

Our earliest stage shewed a vertical furrow at the upper or animal pole, extending through about one-fifth of the circumference (Plate 34, fig. 1), and in a slightly later stage we found a second similar furrow at right angles to the first (Plate 34, fig. 2). We have not been fortunate enough to observe the next phases of the segmentation, but on the second day after impregnation (Plate 34, fig. 3), the animal pole is completely divided into small segments, which form a disc, homologous to the blastoderm of meroblastic ova; while the vegetative pole, which subsequently forms a large yolk-sack, is divided by a few vertical furrows, four of which nearly meet at the pole opposite the blastoderm (Plate 34, fig. 4). The majority of the vertical furrows extend only a short way from the edge of the small spheres, and are partially intercepted by imperfect equatorial furrows.

_Development of the embryo._--We have not been able to work out the stages immediately following the segmentation, owing to want of material; and in the next stage satisfactorily observed, on the third day after impregnation, the body of the embryo is distinctly differentiated. The lower pole of the ovum is then formed of a mass in which no traces of the previous segments or segmentation furrows could any longer be detected.

Some of the dates of the specimens sent to us appear to have been transposed; so that our statements as to ages must only be taken as _approximately_ correct.

_Third day after impregnation._--In this stage the embryo is about 3.5 millims. in length, and has a somewhat dumb-bell shaped outline (Plate 34, fig. 5). It consists of (1) an outer area (_p.z_) with some resemblance to the area pellucida of the Avian embryo, forming the parietal part of the body; and (2) a central portion consisting of the vertebral and medullary plates and the axial portions of the embryo. In hardened specimens the peripheral part forms a shallow depression surrounding the central part of the embryo.

The central part constitutes a somewhat prominent ridge, the axial part of it being the medullary plate. Along the anterior half of this part a dark line could be observed in all our specimens, which we at first imagined to be caused by a shallow groove. We have, however, failed to find in our sections a groove in this situation except in a single instance (Plate 35, fig. 20, _x_), and are inclined to attribute the appearance above-mentioned to the presence of somewhat irregular ridges of the outer layer of the epiblast, which have probably been artificially produced in the process of hardening.

The anterior end of the central part is slightly dilated to form the brain (_b_); and there is present a pair of lateral swellings near the anterior end of the brain which we believe to be the commencing optic vesicles. We could not trace any other clear indications of the differentiation of the brain into distinct lobes.

At the hinder end of the central part of the embryo a very distinct dilatation may also be observed, which is probably homologous with the tail swelling of Teleostei. Its structure is more particularly dealt with in the description of our sections of this stage.

After the removal of the egg-membranes described above we find that there remains a delicate membrane closely attached, to the epiblast. This membrane can be isolated in distinct portions, and appears to be too definite to be regarded as an artificial product.

We have been able to prepare several more or less complete series of sections of embryos of this stage (Plate 35, figs. 18-22). These sections present as a whole a most striking resemblance to those of Teleostean embryos at a corresponding stage of development.

Three germinal layers are already fully established. The epiblast (_ep._) is formed of the same parts as in Teleostei, viz.:--of an outer epidermic and an inner nervous or mucous stratum. In the parietal region of the embryo these strata are each formed of a single row of cells only. The cells of both strata are somewhat flattened, but those of the epidermic stratum are decidedly the more flattened of the two.

Along the axial line there is placed, as we have stated above, the medullary plate. The epidermic stratum passes over this plate without undergoing any change of character, and the plate is _entirely constituted of the nervous stratum of the epidermis_.

The medullary plate has, roughly speaking, the form of a solid keel, projecting inwards towards the yolk. There is no trace, at this stage at any rate, of a medullary groove; and as, we shall afterwards shew, the central canal of the cerebro-spinal cord is formed in the middle of the solid keel. The shape of this keel varies according to the region of the body. In the head (Plate 35, fig. 18, _m.c._), it is very prominent, and forming, as it does, the major part of the axial tissue of the body, impresses its own shape on the other parts of the head and gives rise to a marked ridge on the surface of the head directed towards the yolk. In the trunk (Plate 35, figs. 19, 20) the keel is much less prominent, but still projects sufficiently to give a convex form to the surface of the body turned towards the yolk.

In the head, and also near the hind end of the trunk, the nervous layer of the epiblast continuous with the keel on each side is considerably thicker than the lateral parts of the layer. The thickening of the nervous layer in the head gives rise to what has been called by Götte[501] "the special sense plate," owing to its being subsequently concerned in the formation of parts of the organs of special sense. We cannot agree with Götte in regarding it as part of the brain.

Footnote 501: "Ueb. d. Entwick. d. Central Nerven Systems d. Teleostier," _Archiv für mikr. Anat._ Vol. XV. 1878.

In the keel itself two parts may be distinguished, viz.: a superficial part, best marked in the region of the brain, formed of more or less irregularly arranged polygonal cells, and a deeper part of horizontally placed flatter cells. The upper part is mainly concerned in the formation of the cranial nerves, and of the dorsal roots of the spinal nerves.

The mesoblast (_ms._) in the trunk consists of a pair of independent plates which are continued forwards into the head, and in the prechordal region of the latter, unite below the medullary keel.

The mesoblastic plates of the trunk are imperfectly divided into vertebral and lateral regions. Neither longitudinal sections nor surface views shew at this stage any trace of a division of the mesoblast into somites. The mesoblast cells are polygonal, and no indication is as yet present of a division into splanchnic and somatic layers.

The notochord (_nc._) is well established, so that its origin could not be made out. It is, however, much more sharply separated from the mesoblastic plates than from the hypoblast, though the ventral and inner corners of the mesoblastic plates which run in underneath it on either side, are often imperfectly separated from it. It is formed of polygonal cells, of which between 40 and 50 may as a rule be seen in a single section. No sheath is present around it. It has the usual extension in front.

The hypoblast (_hy._) has the form of a membrane, composed of a single row of oval cells, bounding the embryo on the side adjoining the yolk.

In the region of the caudal swelling the relations of the germinal layers undergo some changes. This region may, from the analogy of other Vertebrates, be assumed to constitute the lip of the blastopore. We find accordingly that the layers become more or less fused. In the anterior part of the tail swelling, the boundary between the notochord and hypoblast becomes indistinct. A short way behind this point (Plate 35, fig. 21), the notochord unites with the medullary keel, and a neurenteric cord, homologous with the neurenteric canal of other Ichthyopsida, is thus established. In the same region the boundary between the lateral plates of mesoblast and the notochord, and further back (Plate 35, fig. 22), that between the mesoblast and the medullary keel, becomes obliterated.

_Fifth day after impregnation._--Between the stage last described and the next stage of which we have specimens, a considerable progress has been made. The embryo (Plate 34, figs. 6 and 7) has grown markedly in length and embraces more than half the circumference of the ovum. Its general appearance is, however, much the same as in the earlier stage, but in the cephalic region the medullary plate is divided by constrictions into three distinct lobes, constituting the regions of the fore-brain, the mid-brain, and the hind-brain. The fore-brain (Plate 34, fig. 6, _f.b._) is considerably the largest of the three lobes, and a pair of lateral projections forming the optic vesicles are decidedly more conspicuous than in the previous stage. The mid-brain (_m.b._) is the smallest of the three lobes, while the hind-brain (_h.b._) is decidedly longer, and passes insensibly into the spinal cord behind.

The medullary keel, though retaining to a great extent the shape it had in the last stage, is no longer completely solid. Throughout the whole region of the brain and in the anterior part of the trunk (Plate 35, figs. 23, 24, 25) a slit-like lumen has become formed. We are inclined to hold that this is due to the appearance of a space between the cells, and not, as supposed by Oellacher for Teleostei, to an actual absorption of cells, though we must admit that our sections are hardly sufficiently well preserved to be conclusive in settling this point. Various stages in its growth may be observed in different regions of the cerebro-spinal cord. When first formed, it is a very imperfectly defined cavity, and a few cells may be seen passing right across from one side of it to the other. It gradually becomes more definite, and its wall then acquires a regular outline.

The optic vesicles are now to be seen in section (Plate 35, fig. 23, _op._) as flattish outgrowths of the wall of the fore-brain, into which the lumen of the third ventricle is prolonged for a short distance.

The brain has become to some extent separate from the superjacent epiblast, but the exact mode in which this is effected is not clear to us. In some sections it appears that the separation takes place in such a way that the nervous keel is only covered above by the epidermic layer of the epiblast, and that the nervous layer, subsequently interposed between the two, grows in from the two sides. Such a section is represented in Plate 35, fig. 24. Other sections again favour the view that in the isolation of the nervous keel, a superficial layer of it remains attached to the nervous layer of the epidermis at the two sides, and so, from the first, forms a continuous layer between the nervous keel and the epidermic layer of the epiblast (Plate 35, fig. 25). In the absence of a better series of sections we do not feel able to determine this point. The posterior part of the nervous keel retains the characters of the previous stage.

At the sides of the hind-brain very distinct commencements of the auditory vesicles are apparent. They form shallow pits (Plate 35, fig. 24, _au._) of the thickened part of the nervous layer adjoining the brain in this region. Each pit is covered over by the epidermic layer above, which has no share in its formation.

In many parts of the lateral regions of the body the nervous layer of the epidermis is more than one cell deep.

The mesoblastic plates are now divided in the anterior part of the trunk into a somatic and a splanchnic layer (Plate 35, fig. 25, _so._, _sp._), though no distinct cavity is as yet present between these two layers. Their vertebral extremities are somewhat wedge-shaped in section, the base of the wedge being placed at the sides of the medullary keel. The wedge-shaped portions are formed of a superficial layer of palisade-like cells and an inner kernel of polygonal cells. The superficial layer on the dorsal side is continuous with the somatic mesoblast, while the remainder pertains to the splanchnic layer.

The diameter of the notochord has diminished, and the cells have assumed a flattened form, the protoplasm being confined to an axial region. In consequence of this, the peripheral layer appears clear in transverse sections. A delicate cuticular sheath is formed around it. This sheath is probably the commencement of the permanent sheath of later stages, but at this stage it cannot be distinguished in structure from a delicate cuticle which surrounds the greater part of the medullary cord.

The hypoblast has undergone no changes of importance.

The layers at the posterior end of the embryo retain the characters of the last stage.

_Sixth day after impregnation._--At this stage (Plate 34, fig. 8) the embryo is considerably more advanced than at the last stage. The trunk has decidedly increased in length, and the head forms a relatively smaller portion of the whole. The regions of the brain are more distinct. The optic vesicles (_op._) have grown outwards so as to nearly reach the edges of the area which forms the parietal part of the body. The fore-brain projects slightly in front, and the mid-brain is seen as a distinct rounded prominence. Behind the latter is placed the hind-brain, which passes insensibly into the spinal cord. On either side of the mid- and hind-brain a small region is slightly marked off from the rest of the parietal part, and on this are seen two more or less transversely directed streaks, which, by comparison with the Sturgeon[502], we are inclined to regard as the two first visceral clefts (_br.c._). We have, however, failed to make them out in sections, and owing to the insufficiency of our material, we have not even studied them in surface views as completely as we could have wished.

Footnote 502: Salensky, "Recherches s. le Développement du Sterlet." _Archives de Biol._ Vol. II. 1881, pl. XVII. fig. 27.

The body is now laterally compressed, and more decidedly raised from the yolk than in the previous stages. In the lateral regions of the trunk the two segmental or archinephric ducts (_sg._) are visible in surface views: the front end of each is placed at the level of the hinder border of the head, and is marked by a flexure inwards towards the middle line. The remainder of each duct is straight, and extends backwards for about half the length of the embryo. The tail has much the same appearance as in the last stage.

The vertebral regions of the mesoblastic plates are now segmented for the greater part of the length of the trunk, and the somites of which they are composed (Plate 36, fig. 30, _pr._) are very conspicuous in surface views.

Our sections of this stage are not so complete as could be desired: they shew, however, several points of interest.

The central canal of the nervous system is large, with well-defined walls, and in hardened specimens is filled with a coagulum. It extends nearly to the region of the tail.

The optic vesicles, which are so conspicuous in surface views, appear in section (Plate 35, fig. 26, _op._) as knob-like outgrowths of the fore-brain, and very closely resemble the figures given by Oellacher of these vesicles in Teleostei[503].

Footnote 503: "Beiträge zur Entwick. d. Knochenfische," _Zeit. f. wiss. Zool._ Vol. XXIII. 1873, taf. III. fig. IX. 2.

From the analogy of the previous stage, we are inclined to think that they have a lumen continuous with that of the fore-brain. In our only section through them, however, they are solid, but this is probably due to the section merely passing through them to one side.

The auditory pits (Plate 35, fig. 27, _au._) are now well marked, and have the form of somewhat elongated grooves, the walls of which are formed of a single layer of columnar cells belonging to the nervous layer of the epidermis, and extending inwards so as nearly to touch the brain.

In an earlier stage it was pointed out that the dorsal part of the medullary keel was different in its structure from the remainder, and that it was destined to give rise to the nerves. The process of differentiation is now to a great extent completed, and may best be seen in the auditory region (Plate 35, fig. 27, VIII.). In this region there was present during the last stage a great rhomboidal mass of cells at the dorsal region of the brain (Plate 35, fig. 24, VIII.). In the present stage, this, which is the rudiment of the seventh and auditory nerves, is seen growing down on each side from the roof of the hind-brain, between the brain and the auditory involution, and abutting against the wall of the latter.

Rudiments of the spinal nerves are also seen at intervals as projections from the dorsal angles of the spinal cord (Plate 36, fig. 29, _sp.n._). They extend only for a short distance outwards, gradually tapering off to a point, and situated between the epiblast and the dorsal angles of the mesoblastic somites.

The process of formation of the cranial nerves and dorsal roots of the spinal nerves is, it will be seen, essentially the same as that already known in the case of Elasmobranchii, Aves, &c. The nerves arise as outgrowths of a special crest of cells, the _neural crest_ of Marshall, which is placed along the dorsal angle of the cord. The peculiar position of the dorsal roots of the spinal nerves is also very similar to what has been met with in the early stages of these structures by Marshall in Birds[504], and by one of us in Elasmobranchii[505].

Footnote 504: _Journal of Anat. and Physiol._ Vol. XI. p. 491, plates XX. and XXI.

Footnote 505: "Elasmobranch Fishes," p. 156, plates 10 and 13. [This edition, p. 378, pl. 11, 14.]

In the parietal region a cavity has now appeared in part of the trunk between the splanchnic and somatic layers of the mesoblast (Plate 36, fig. 29, _b.c._), the somatic layer (_so._) consisting of a single row of columnar cells on the dorsal side, while the remainder of each somite is formed of the splanchnic layer (_sp._). In many of the sections the somatic layer is separated by a considerable interval from the epiblast.

We have been able to some extent to follow the development of the segmental duct. The imperfect preservation of our specimens has, as in other instances, rendered the study of the point somewhat difficult, but we believe that the figure representing the development of the duct some way behind its front end (Plate 36, fig. 29) is an accurate representation of what may be seen in a good many of our sections.

It appears from these sections that the duct (Plate 36, fig. 29, _sg._) is developed as a hollow ridge-like outgrowth of the somatic layer of mesoblast, directed towards the epiblast, in which it causes a slight bulging. The cavity of the ridge freely communicates with the body-cavity. The anterior part of this ridge appears to be formed first. Very soon, in fact, in an older embryo belonging to this stage, the greater part of the groove becomes segmented off as a duct lying between the epiblast and somatic mesoblast (Plate 36, fig. 28, _sg._), while the front end still remains, as we believe, in communication with the body-cavity by an anterior pore.

This mode of development corresponds in every particular with that observed in Teleostei by Rosenberg and Oellacher.

The structure of the notochord (_nc._) at this stage is very similar to that observed by one of us in Elasmobranchii[506]. The cord is formed of transversely arranged flattened cells, the outer parts of which are vacuolated, while the inner parts are granular, and contain the nuclei. This structure gives rise to the appearance in transverse sections of an axial darker area and a peripheral lighter portion.

Footnote 506: "Elasmobranch Fishes," p. 136, plate 11, fig. 10. [This edition, p. 354, pl. 12.]

The hypoblast retains for the most part its earlier constitution, but underneath the notochord, in the trunk, it is somewhat thickened, and the cells at the two sides spread in to some extent under the thickened portion (Plate 36, fig. 29, _s.nc._). This thickening, as is shewn in transverse sections at the stage when the segmental duct becomes separated from the somatic mesoblast (Plate 36, fig. 28, _s.nc._), is the commencement of the subnotochordal rod.

The tail end of the embryo still retains its earlier characters.

_Seventh day after impregnation._--Our series of specimens of this stage is very imperfect, and we are only able to call attention to the development of a certain number of organs.

Our sections clearly establish the fact that the optic vesicles are now hollow processes of the fore-brain. Their outer ends are dilated, and are in contact with the external skin. The formation of the optic cup has not, however, commenced. The nervous layer of the skin adjoining the outer wall of the optic cup is very slightly thickened, constituting the earliest rudiment of the lens.

In one of our embryos of this day the developing auditory vesicle still has the form of a pit, but in the other it is a closed vesicle, already constricted off from the nervous layer of the epidermis.

With reference to the development of the excretory duct we cannot add much to what we have already stated in describing the last stage.

The duct is considerably dilated anteriorly (Plate 36, fig. 31, _sg._); but our sections throw no light on the nature of the abdominal pore. The posterior part of the duct has still the form of a hollow ridge united with somatic mesoblast (Plate 36, fig. 32, _sg._).

During this stage, the embryo becomes to a small extent folded off from the yolk-sack both in front and behind, and in the course of this process the anterior and posterior extremities of the alimentary tract become definitely established.

We have not got as clear a view of the process of formation of these two sections of the alimentary tract as we could desire, but our observations appear to shew that the process is in many respects similar to that which takes place in the formation of the anterior part of the alimentary tract in Elasmobranchii[507]. One of us has shewn that in Elasmobranchii the ventral wall of the throat is formed _not_ by a process of folding in of the hypoblastic sheet as in Birds, but by a growth of the ventral face of the hypoblastic sheet on each side of and at some little distance from the middle line. Each growth is directed inwards, and the two eventually meet and unite, thus forming a complete ventral wall for the gut. Exactly the same process would seem to take place in _Lepidosteus_, and after the lumen of the gut is in this way established, a process of mesoblast on each side also makes its appearance, forming a mesoblastic investment on the ventral side of the alimentary tract. Some time after the alimentary tract has been thus formed, the epiblast becomes folded in, in exactly the same manner as in the Chick, the embryo becoming thereby partially constricted off from the yolk (Plate 36, figs. 33, 34).

Footnote 507: F. M. Balfour, "Monograph on the Development of Elasmobranch Fishes," p. 87, plate 9, fig. 2. [This edition, p. 303, pl. 10.]

The form of the lumen of the alimentary tract differs somewhat in front and behind. In front, the hypoblastic sheet remains perfectly flat during the formation of the throat, and thus the lumen of the latter has merely the form of a slit. The lumen of the posterior end of the alimentary tract is, however, narrower and deeper (Plate 36, figs. 33, 34, _al._). Both in front and behind, the lateral parts of the hypoblastic sheet become separated from the true alimentary tract as soon as the lumen of the latter is established.

It is quite possible that at the extreme posterior end of the embryo a modification of the above process may take place, for in this region the hypoblast appears to us to have the form of a solid cord.

We could detect no true neurenteric canal, although a more or less complete fusion of the germinal layers at the tail end of the embryo may still be traced.

During this stage the protoplasm of the notochordal cells, which in the last stage formed a kind of axial rod in the centre of the notochord, begins to spread outwards toward the sheath of the notochord.

_Eighth day after impregnation._--The external form of the embryo (Plate 34, fig. 9) shews a great advance upon the stage last figured. Both head and body are much more compressed laterally and raised from the yolk, and the head end is folded off for some distance. The optic vesicles are much less prominent externally. A commencing opercular fold is distinctly seen. Our figure of this stage is not, however, so satisfactory as we could wish.

A thickening of the nervous layer of the external epiblast which will form the lens (Plate 36, fig. 35, _l._) is more marked than in the last stage, and presses against the slightly concave exterior wall of the optic vesicle (_op._). The latter has now a large cavity, and its stalk is considerably narrowed.

The auditory vesicles (Plate 36, fig. 36, _au._) are closed, appearing as hollow sacks one on each side of the brain, and are no longer attached to the epiblast.

The anterior opening of the segmental duct can be plainly seen close behind the head. The lumen of the duct is considerably larger.

The two vertebral portions of the mesoblast are now separated by a considerable space from the epiblast on one side and from the notochord on the other, and the cells composing them have become considerably elongated from side to side (Plate 36, fig. 37, _ms_).

In some sections the aorta can be seen (Plate 36, fig. 37, _ao_) lying close under the subnotochordal rod, between it and the hypoblast, and on either side of it a slightly larger cardinal vein (_cd.v._).

The protoplasm of the notochord has now again retreated towards the centre, shewing a clear space all round. This is most marked in the region of the trunk (Plate 36, fig. 37). The subnotochordal rod (_s.nc._) lies close under it.

A completely closed fore-gut, lined by thickened hypoblast, extends about as far back as the auditory sacks (Plate 36, figs. 35 and 36, _al._). In the trunk the hypoblast, which will form the walls of the alimentary tract, is separated from the notochord by a considerable interval.

_Ninth day after impregnation: External characters._--Very considerable changes have taken place in the external characters of the embryo. It is about 8 millims. in length, and has assumed a completely piscine form. The tail especially has grown in length, and is greatly flattened from side to side: it is wholly detached from the yolk, and bends round towards the head, usually with its left side in contact with the yolk. It is provided with well-developed dorsal and ventral fin-folds, which meet each other round the end of the tail, the tail fin so formed being nearly symmetrical. The head is not nearly so much folded off from the yolk as the tail. At its front end is placed a disc with numerous papillæ, of which we shall say more hereafter. This disc is somewhat bifid, and is marked in the centre by a deep depression.

Dorsal to it, on the top of the head, are two widely separated nasal pits. On the surface of the yolk, in front of the head, is to be seen the heart, just as in Sturgeon embryos. Immediately below the suctorial disc is a slit-like space, forming the mouth. It is bounded below by the two mandibular arches, which meet ventrally in the median line. A shallow but well-marked depression on each side of the head indicates the posterior boundary of the mandibular arch. Behind this is placed the very conspicuous hyoid arch with its rudimentary opercular flap; and in the depression, partly covered over by the latter, may be seen a ridge, the external indication of the first branchial arch.

_Eleventh day after impregnation: External characters._--The embryo (Plate 34, fig. 10) is now about 10 millims. in length, and in several features exhibits an advance upon the embryo of the previous stage.

The tail fin is now obviously not quite symmetrical, and the dorsal fin-fold is continued for nearly the whole length of the trunk. The suctorial disc (Plate 34, fig. 11, _s.d._) is much more prominent, and the papillæ (about 30 in number) covering it are more conspicuous from the surface. It is not obviously composed of two symmetrical halves. The opercular flap is larger, and the branchial arches behind it (two of which may be made out without dissection) are more prominent.

The anterior pair of limbs is now visible in the form of two _longitudinal_ folds projecting in a vertical direction from the surface of the yolk-sack at the sides of the body.

The stages subsequent to hatching have been investigated with reference to the external features and to the habits by Agassiz, and we shall enrich our own account by copious quotations from his memoir.

He states that the first batch were hatched on the eighth[508] day after being laid. "The young Fish possessed a gigantic yolk-bag, and the posterior part of the body presented nothing specially different from the general appearance of a Teleostean embryo, with the exception of the great size of the chorda. The anterior part, however, was most remarkable; and at first, on seeing the head of this young _Lepidosteus_, with its huge mouth-cavity extending nearly to the gill-opening, and surmounted by a hoof-shaped depression edged with a row of protuberances acting as suckers, I could not help comparing this remarkable structure, so utterly unlike anything in Fishes or Ganoids, to the Cyclostomes, with which it has a striking analogy. This organ is also used by _Lepidosteus_ as a sucker, and the moment the young Fish is hatched he attaches himself to the sides of the disc, and there remains hanging immovable; so firmly attached, indeed, that it requires considerable commotion in the water to make him loose his hold. Aërating the water by pouring it from a height did not always produce sufficient disturbance to loosen the young Fishes. The eye, in this stage, is rather less advanced than in corresponding stages in bony Fishes; the brain is also comparatively smaller, the otolith ellipsoidal, placed obliquely in the rear above the gill-opening.... Usually the gill-cover is pressed closely against the sides of the body, but in breathing an opening is seen through which water is constantly passing, a strong current being made by the rapid movement of the pectorals, against the base of which the extremity of the gill-cover is closely pressed. The large yolk-bag is opaque, of a bluish-gray colour. The body of the young _Lepidosteus_ is quite colourless and transparent. The embryonic fin is narrow, the dorsal part commencing above the posterior end of the yolk-bag; the tail is slightly rounded, the anal opening nearer the extremity of the tail than the bag. The intestine is narrow, and the embryonic fin extending from the vent to the yolk-bag is quite narrow. In a somewhat more advanced stage,--hatched a few hours earlier,--the upper edge of the yolk-bag is covered with black pigment cells, and minute black pigment cells appear on the surface of the alimentary canal. There are no traces of embryonic fin-rays either in this stage or the one preceding; the structure of the embryonic fin is as in bony Fishes--previous to the appearance of these embryonic fin-rays--finely granular. Seen in profile, the yolk-bag is ovoid; as seen from above, it is flattened, rectangular in front, with rounded corners, tapering to a rounded point towards the posterior extremity, with re-entering sides."

Footnote 508: This statement of Agassiz does not correspond with the dates on the specimens sent to us--a fact no doubt due to the hatching not taking place at the same time for all the larvæ.

We have figured an embryo of 11 millims. in length, shortly after hatching (Plate 34, fig. 12), the most important characters of which are as follows:--The yolk-sack, which has now become much reduced, forms an appendage attached to the ventral surface of the body, and has a very elongated form as compared with its shape just before hatching. The mouth, as also noticed by Agassiz, has a very open form. It is (Plate 34, fig. 13, _m._) more or less rhomboidal, and is bounded behind by the mandibular arch (_mn._) and laterally by the superior maxillary processes (_s.mx_). In front of the mouth is placed the suctorial disc (_s.d._), the central papillæ of which are arranged in groups. The opercular fold (_h.op._) is very large, covering the arches behind. A well-marked groove is present between the mandibular and opercular arches, but so far as we can make out it is not a remnant of the hyomandibular cleft.

The pectoral fins (Plate 34, fig. 12, _pc.f._) are very prominent longitudinal ridges, which, owing to their being placed on the surface of the yolk-sack, project in a nearly vertical direction: a feature which is also found in many Teleostean embryos with large yolk-sacks.

No traces of the pelvic fins have yet become developed.

The positions of the permanent dorsal, anal, and caudal fins, as pointed out by Agassiz, are now indicated by a deposit of pigment in the embryonic fin.

In an embryo on the sixth day after hatching, of about 15 millims. in length, of which we have also given a figure (Plate 34, fig. 14), the following fresh features deserve special notice.

In the region of the head there is a considerable elongation of the pre-oral part, forming a short snout, at the end of which is placed the suctorial disc. At the sides of the snout are placed the nasal pits, which have become somewhat elongated anteriorly.

The mouth has lost its open rhomboidal shape, and has become greatly narrowed in an antero-posterior direction, so that its opening is reduced to a slit. The mandibles and maxillary processes are nearly parallel, though both of them are very much shorter than in the adult. The operculum is now a very large flap, and has extended so far backwards as to cover the insertion of the pectoral fin. The two opercular folds nearly meet ventrally.

The yolk-sack is still more reduced in size, one important consequence of which is that the pectoral fins (_pc.f._) appear to spring out more or less horizontally from the sides of the body, and at the same time their primitive line of attachment to the body becomes transformed from a longitudinal to a more or less transverse one.

The first traces of the pelvic fins are now visible as slight longitudinal projections near the hinder end of the yolk-sack (_pl.f._).

The pigmentation marking the regions of the permanent fins has become more pronounced, and it is to be specially noted that the ventral part of the caudal fin (the permanent caudal) is considerably more prominent than the dorsal fin opposite to it.

The next changes, as Agassiz points out, "are mainly in the lengthening of the snout; the increase in length both of the lower and upper jaw; the concentration of the sucker of the sucking disc; and the adoption of the general colouring of somewhat older Fish. The lobe of the pectoral has become specially prominent, and the outline of the fins is now indicated by a fine milky granulation. Seen from above, the gill-cover is seen to leave a large circular opening leading to the gill-arches, into which a current of water is constantly passing, by the lateral expansion and contraction of the gill-cover; the outer extremity of the gill-cover covers the base of the pectorals. In a somewhat older stage the snout has become more elongated, the sucker more concentrated, and the disproportionate size of the terminal sucking-disc is reduced; the head, when seen from above, becoming slightly elongated and pointed."

In a larva of about 18 days old and 21 millims. in length, of which we have not given a figure, the snout has grown greatly in length, carrying with it the nasal organs, the openings of which now appear to be divided into two parts. The suctorial disc is still a prominent structure at the end of the snout. The lower jaw has elongated correspondingly with the upper, so that the gape is very considerable, though still very much less than in the adult.

The opercular flaps overlap ventrally, the left being superficial. They still cover the bases of the pectoral fins. The latter are described by Agassiz as being "kept in constant rapid motion, so that the fleshy edge is invisible, and the vibration seems almost involuntary, producing a constant current round the opening leading into the cavity of the gills."

The pelvic fins are somewhat more prominent.

The yolk-sack, as pointed out by Agassiz, has now disappeared as an external appendage.

After the stage last described the young Fish rapidly approaches the adult form. To shew the changes effected we have figured the head of a larva of about a month old and 23 millims. in length (Plate 34, fig. 15). The suctorial disc, though much reduced, is still prominent at the end of the snout. Eventually, as shewn by Agassiz, it forms the fleshy globular termination of the upper jaw.

The most notable feature in which the larva now differs in its external form from the adult is in the presence of an externally heterocercal tail, caused by the persistence of the primitive caudal fin as an elongated filament projecting beyond the permanent caudal (Plate 41, fig. 68).

Delicate dermal fin-rays are now conspicuous in the peripheral parts of all the permanent fins. These rays closely resemble the horny fin-rays in the fins of embryo Elasmobranchii in their development and structure. They appear gradually to enlarge to form the permanent rays, and we have followed out some of the stages of their growth, which is in many respects interesting. Our observations are not, however, complete enough to publish, and we can only say here that their early development and structure proves their homology with the horny fibres or rays in fins of Elasmobranchii. The skin is still, however, entirely naked, and without a trace of its future armour of enamelled scales.

The tail of a much older larva, 11 centims. in length, in which the scales have begun to be formed, is shewn in Plate 34, fig. 16.

We complete this section of our memoir by quoting the following passages from Agassiz as to the habits of the young fish at the stages last described:--

"In the stages intervening between plate iii, fig. 19, and plate iii, fig. 30, the young _Lepidosteus_ frequently swim about, and become readily separated from their point of attachment. In the stage of plate iii, fig. 30, they remain often perfectly quiet close to the surface of the water; but, when disturbed, move very rapidly about through the water.... The young already have also the peculiar habit of the adult of coming to the surface to swallow air. When they go through the process under water of discharging air again they open their jaws wide, and spread their gill-covers, and swallow as if they were choking, making violent efforts, until a minute bubble of air has become liberated, when they remain quiet again. The resemblance to a Sturgeon in the general appearance of this stage of the young _Lepidosteus_ is quite marked."

BRAIN.

I. _Anatomy._

The brain of _Lepidosteus_ has been figured by Busch (whose figure has been copied by Miklucho-Maclay, and apparently by Huxley), by Owen and by Wilder (No. 15). The figure of the latter author, representing a longitudinal section through the brain, is the most satisfactory, the other figures being in many respects inaccurate; but even Wilder's figure and description, though taken from the fresh object, appear to us in some respects inadequate. He offers, moreover, fresh interpretations of certain parts of the brain which we shall discuss in the sequel.

We have examined two brains which, though extremely soft, were, nevertheless, sufficiently well preserved to enable us to study the external form. We have, moreover, made a complete series of transverse sections through one of the brains, and our sections, though utterly valueless from a histological point of view, have thrown some light on the topographical anatomy of the brain.

Plate 38, figs. 47A, B, and C, represent three views of the brain, viz.: from the side, from above, and from below. We will follow in our description the usual division of the brain into fore-brain, mid-brain, and hind-brain.

The fore-brain consists of an anterior portion forming the cerebrum, and a posterior portion constituting the thalamencephalon.

The cerebrum at first sight appears to be composed of (_a_) a pair of posterior and somewhat dorsal lobes, forming what have usually been regarded as the true cerebral hemispheres, but called by Wilder the prothalami, and (_b_) a pair of anterior and ventral lobes, usually regarded as the olfactory lobes, from which the olfactory nerves spring. Mainly from a comparison with our embryonic brains described in the sequel, we are inclined to think that the usual interpretations are not wholly correct, but that the true olfactory lobes are to be sought for in small enlargements (Plate 38. figs. 47A, B, and C, _olf._) at the front end of the brain[509] from which the olfactory nerves spring. The cerebrum proper would then consist of a pair of anterior and ventral lobes (_ce._), and of a pair of posterior lobes (_ce´._), both pairs uniting to form a basal portion behind.

Footnote 509: The homologies of the olfactory lobes throughout the group of Fishes require further investigation.

The two pairs of lobes probably correspond with the two parts of the cerebrum of the Frog, the anterior of which, like that of _Lepidosteus_, was held to be the olfactory lobe, till Götte's researches shewed that this view was not tenable.

The anterior lobes of the cerebrum have a conical form, tapering anteriorly, and are completely separated from each other. The posterior lobes, as is best shewn in side views, have a semicircular form. Viewed from above they appear as rounded prominences, and their dorsal surface is marked by two conspicuous furrows (Plate 38, fig. 47B, _ce´._), which have been noticed by Wilder, and are similar to those present in many Teleostei. Their front ends overhang the base of the anterior cerebral lobes. The basal portion of the cerebrum is an undivided lobe, the anterior wall of which forms the lamina terminalis.

What we have above described as the posterior cerebral lobes have been described by Wilder as constituting the everted dorsal border of the basal portion of the cerebrum.

The portion of the cerebro-spinal canal within the cerebrum presents certain primitive characters, which are in some respects dissimilar to those of higher types, and have led Wilder to hold the posterior cerebral lobes, together with what we have called the basal portion of the cerebrum, to be structures peculiar to Fishes, for which he has proposed the name "prothalami."

In the basal portion of the cerebrum there is an unpaired slit-shaped ventricle, the outer walls of which are very thick. It is provided with a floor formed of nervous matter, in part of which, judging from Wilder's description, a well-marked commissure is placed. We have found in the larva a large commissure in this situation (Plate 37, figs. 44 and 45, _a.c._); and it may be regarded as the homologue of the anterior commissure of higher types. This part of the ventricle is stated by Wilder to be without a roof. This appears to us highly improbable. We could not, however, determine the nature of the roof from our badly preserved specimens, but if present, there is no doubt that it is extremely thin, as indeed it is in the larva (Plate 37, fig. 46B). In a dorsal direction the unpaired ventricle extends so as to separate the two posterior cerebral lobes. Anteriorly the ventricle is prolonged into two horns, which penetrate for a short distance, as _the lateral ventricles_, into the base of the anterior cerebral lobes. The front part of each anterior cerebral lobe, as well as of the whole of the posterior lobes, appears solid in our sections; but Wilder describes the anterior horns of the ventricle as being prolonged for the whole length of the anterior lobes.

In the embryos of all Vertebrates the cerebrum is not at first divided into two lobes, so that the fact of the posterior part of the cerebrum in _Lepidosteus_ and probably other Ganoids remaining permanently in the undivided condition does not appear to us a sufficient ground for giving to the lobes of this part of the cerebrum the special name of prothalami, as proposed by Wilder, or for regarding them as a section of the brain peculiar to Fishes.

The thalamencephalon (_th._) contains the usual parts, but is in some respects peculiar. Its lateral walls, forming the optic thalami, are thick, and are not sharply separated in front from the basal part of the cerebrum; between them is placed the third ventricle. The thalami are of considerable extent, though partially covered by the optic lobes and the posterior lobes of the cerebrum. They are not, however, relatively so large as in other Ganoid forms, more especially the Chondrostei and _Polypterus_.

On the roof of the thalamencephalon is placed a large thin-walled vesicle (Plate 38, figs. 47A and B, _v.th._), which undoubtedly forms the most characteristic structure connected with this part of the brain. Owing to the wretched state of preservation of the specimens, we have found it impossible to determine the exact relations of this body to the remainder of the thalamencephalon; but it appears to be attached to the roof of the thalamencephalon by a narrow stalk only. It extends forwards so as to overlap part of the cerebrum in front, and is closely invested by a highly vascular layer of the pia mater.

No mention is made by Wilder of this body; nor is it represented in his figures or in those of the other anatomists who have given drawings of the brain of _Lepidosteus_. It might at first be interpreted as a highly-developed pineal gland, but a comparison with the brain of the larva (vide p. 764) shews that this is not the case, but that the body in question is represented in the larva by a special outgrowth of the roof of the thalamencephalon. The vesicle of the roof of the thalamencephalon is therefore to be regarded as a peculiar development of the tela choroidea of the third ventricle.

How far this vesicle has a homologue in the brains of other Ganoids is not certain, since negative evidence on this subject is all but valueless. It is possible that a vesicular sack covering over the third ventricle of the Sturgeon described by Stannius[510], and stated by him to be wholly formed of the membranes of the brain, is really the homologue of our vesicle.

Footnote 510: "Ueb. d. Gehirn des Störs," Müller's _Archiv_, 1843, and _Lehrbuch d. vergl. Anat. d. Wirbelthiere_. Cattie, _Archives de Biologie_, Vol. III. 1882, has recently described in _Acipenser sturio_ a vesicle on the roof of the thalamencephalon, whose cavity is continuous with the third ventricle. This vesicle is clearly homologous with that in _Lepidosteus_. (June 28, 1882.)

Wiedersheim[511] has recently described in _Protopterus_ a body which is undoubtedly homologous with our vesicle, which he describes in the following way:--

"Dorsalwärts ist das Zwischenhirn durch ein tiefes, von Hirnschlitz eingenommenes Thal von Vorderhirn abgesetzt; dasselbe ist jedoch durch eine häutige, mit der Pia mater zusammenhängende Kuppel oder Kapsel überbrückt."

Footnote 511: R. Wiedersheim, _Morphol. Studien_, 1880, p. 71.

This "Kuppel" has precisely the same relations and a very similar appearance to our vesicle. The true pineal gland is placed behind it. It appears to us possible that the body found by Huxley[512] in _Ceratodus_, which he holds to be the pineal gland, is in reality this vesicle. It is moreover possible that what has usually been regarded as the pineal gland in _Petromyzon_ may in reality be the homologue of the vesicle we have found in _Lepidosteus_.

Footnote 512: "On _Ceratodus Forsteri_," &c., _Proc. Zool. Soc._ 1876.

We have no observations on the pineal gland of the adult, but must refer the reader for the structure and relations of this body to the embryological section.

The infundibulum (Plate 38, fig. 47A, _in._) is very elongated. Immediately in front of it is placed the optic chiasma (Plate 38, figs. 47A and C, _op.ch._) from which the optic fibres can be traced passing along the sides of the optic thalami and to the optic lobes, very much as in Müller's figure of the brain of _Polypterus_.

On the sides of the infundibulum are placed two prominent bodies, the lobi inferiores (_l.in._), each of which contains a cavity continuous with the prolongation of the third ventricle into the infundibulum. The apex of the infundibulum is enlarged, and to it is attached a pituitary body (_pt._).

The mid-brain is of considerable size, and consists of a basal portion connecting the optic thalami with the medulla, and a pair of large optic lobes (_op.l._). The iter a tertio ad quartum ventriculum, which forms the ventricle of this part of the brain, is prolonged into each optic lobe, and the floor of each prolongation is taken up by a dome-shaped projection, the homologue of the torus semicircularis of Teleostei.

The hind-brain consists of the usual parts, the medulla oblongata and the cerebellum. The medulla presents no peculiar features. The sides of the fourth ventricle are thickened and everted, and marked with peculiar folds (Plate 38, figs. 47A and B, _m.o._).

The cerebellum is much larger than in the majority of Ganoids, and resembles in all essential features the cerebellum of Teleostei. In side views it has a somewhat S-shaped form, from the presence of a peculiar lateral sulcus (Plate 38, fig. 47A, _cb._). As shewn by Wilder, its wall actually has in longitudinal section this form of curvature, owing to its anterior part projecting forwards into the cavity of the iter[513]. This forward projection is not, however, so conspicuous as in most Teleostei. The cerebellum contains a large unpaired prolongation of the fourth ventricle.

Footnote 513: In Wilder's figure the walls of the cerebellum are represented as much too thin.

II. _Development._

The early development of the brain has already been described; and, although we do not propose to give any detailed account of the later stages of its growth, we have thought it worth while calling attention to certain developmental features which may probably be regarded as to some extent characteristic of the Ganoids. With this view we have figured (Plate 37, figs. 44, 45) longitudinal sections of the brain at two stages, viz.: of larvæ of 15 and 26 millims., and transverse sections (Plate 37, figs. 46A-G) of the brain of a larva at about the latter stage (25 millims.).

The original embryonic fore-brain is divided in both embryos into a cerebrum (_ce._) in front and a thalamencephalon (_th_) behind. In the younger embryo the cerebrum is a single lobe, as it is in the brains of all Vertebrate embryos; but in the older larva it is anteriorly (Plate 37, fig. 46A) completely divided into two hemispheres. The roof of the undivided posterior part of the cerebrum is extremely thin (Plate 37, fig. 46B). Near the posterior border of the base of the cerebrum there is a great development of nervous fibres, which may probably be regarded as in part equivalent to the anterior commissure (Plate 37, figs. 44, 45 _a.c._).

Even in the oldest of the two brains the olfactory lobes are very slightly developed, constituting, however, small lateral and ventral prominences of the front end of the hemispheres. From each of them there springs a long olfactory nerve, extending for the whole length of the rostrum to the olfactory sack.

The thalamencephalon presents a very curious structure, and is relatively a more important part of the brain than in the embryo of any other form which we know of. Its roof, instead of being, as usual, compressed antero-posteriorly[514], so as to be almost concealed between the cerebral hemispheres and the optic lobes (mid-brain), projects on the surface for a length quite equal to that of the cerebral hemispheres (Plate 37, figs. 44 and 45, _th._). In the median line the roof of the thalamencephalon is thin and folded; at its posterior border is placed the opening of the small pineal gland. This body is a papilliform process of the nervous matter of the roof of this part of the brain, and instead of being directed forwards, as in most Vertebrate types, tends somewhat backwards, and rests on the mid-brain behind (Plate 37, figs. 44, 45, and 46C and D, _pn._). The roof of the thalamencephalon immediately in front of the pineal gland forms a sort of vesicle, the sides of which extend laterally as a pair of lobes, shewn in transverse sections in Plate 37, figs. 46C and D, as _th.l._ This vesicle becomes, we cannot doubt, the vesicle on the roof of the thalamencephalon which we have described in the adult brain. Immediately in front of the pineal gland the roof of the thalamencephalon contains a transverse commissure (Plate 37, fig. 46C, _z._), which is the homologue of a similarly situated commissure present in the Elasmobranch brain[515], while behind the pineal gland is placed the posterior commissure. The sides of the thalamencephalon are greatly thickened, forming the optic thalami (Plate 37, figs. 46C and D, _op.th._), which are continuous in front with the thickened outer walls of the hemispheres. Below, the thalamencephalon is produced into a very elongated infundibulum (Plate 37, figs. 44, 45, 46E, _in._), the apex of which is trilobed as in Elasmobranchii and Teleostei. The sides of the infundibulum exhibit two lobes, the lobi inferiores (Plate 37, fig. 46E, _l.in._), which are continued posteriorly into the crura cerebri.

Footnote 514: Vide F. M. Balfour, _Comparative Embryology_, Vol. II. figs. 248 and 250.

Footnote 515: Vide F. M. Balfour, _Comparative Embryology_, Vol. II. pp. 355-6 [the original edition], where it is suggested that this commissure is the homologue of the grey commissure of higher types.

The pituitary body[516] (Plate 37, figs. 44, 45, 46E, _pt._) is small, not divided into lobes, and provided with a very minute lumen.

Footnote 516: We have not been able to work out the early development of the pituitary body as satisfactorily as we could have wished. In Plate 37, fig. 40, there is shewn an invagination of the oral epithelium to form it; in Plate 37, figs. 41 and 42, it is represented in transverse section in two consecutive sections. Anteriorly it is still connected with the oral epithelium (fig. 41), while posteriorly it is free. It is possible that an earlier stage of it is shewn in Plate 36, fig. 35. Were it not for the evidence in other types of its being derived from the epiblast we should be inclined to regard it as hypoblastic in origin.

In front of the infundibulum is the optic chiasma (Plate 37, fig. 46D, _op.ch._), which is developed very early. It is, as stated by Müller, a true chiasma.

The mid-brain (Plate 37, figs. 44 and 45, _m.b._) is large, and consists in both stages of (1) a thickened floor forming the crura cerebri, the central canal of which constitutes the iter a tertio ad quartum ventriculum; and (2) the optic lobes (Plate 37, figs. 46E, F, G, _op.l._) above, each of which is provided with a cavity continuous with the median iter. The optic lobes are separated dorsally and in front by a well-marked median longitudinal groove. Posteriorly they largely overlap the cerebellum. In the anterior part of the optic lobes, at the point where the iter joins the third ventricle, there may be seen slight projections of the floor into the lumen of the optic lobes (Plate 37, fig. 46E). These masses probably become in the adult the more conspicuous prominences of the floor of the ventricles of the optic lobes, which we regard as homologous with the tori semicirculares of the brain of the Teleostei.

The hind-brain is formed of the usual divisions, viz.: cerebellum and medulla oblongata (Plate 37, figs. 44 and 45, _cb._, _md._). The former constitutes a bilobed projection of the roof of the hind-brain. Only a small portion of it is during these stages left uncovered by the optic lobes, but the major part extends forwards for a considerable distance under the optic lobes, as shewn in the transverse sections (Plate 37, figs. 46F and G, _cb._); and its two lobes, each with a prolongation of its cavity, are continued forwards beyond the opening of the iter into the fourth ventricle.

It is probable that the anterior horns of the cerebellum are equivalent to the prolongations of the cerebellum into the central cavity of the optic lobes of Teleostei, which are continuous with the so-called fornix of Göttsche.

III. _Comparison of the larval and adult brain of Lepidosteus, together with some observations on the systematic value of the characters of the Ganoid brain._

The brain of the older of the two larvæ, which we have described, sufficiently resembles in most of its features that of the adult to render material assistance in the interpretation of certain of the parts of the latter. It will be remembered that in the adult brain the parts usually held to be olfactory lobes were described as the anterior cerebral lobes. The grounds for this will be apparent by a comparison of the cerebrum of the larva and adult. In the larva the cerebrum is formed of (1) an unpaired basal portion with a thin roof, and (2) of a pair of anterior lobes, with small olfactory bulbs at their free extremities.

The basal portion in the larva clearly corresponds in the adult with the basal portion, together with the two posterior cerebral lobes, which are merely special outgrowths of the dorsal edge of the primitive basal portion. The pair of anterior lobes have exactly the same relations in the larva as in the adult, except that in the former the ventricles are prolonged for their whole length instead of being confined to their proximal portions. If, therefore, our identifications of the larval parts of the brain are correct, there can hardly be a question as to our identifications of the parts in the adult. As concerns these identifications, the comparison of the brain of our two larvæ appears conclusive in favour of regarding the anterior lobes as parts of the cerebrum, as distinguished from the olfactory lobes, in that they are clearly derived from the undivided anterior portion of the cerebrum of the younger larva.

The comparison of the larval brain with that of the adult again appears to us to leave no doubt that the vesicle attached to the roof of the thalamencephalon in the adult is the same structure as the bilobed outgrowth of this roof in the larva; and since there is in addition a well-developed pineal gland in the larva with the usual relations, there can be no ground for identifying the vesicle in the adult with the pineal gland.

Müller, in his often quoted memoir (No. 13), states that the brains of Ganoids are peculiar and distinct from those both of Teleostei and Elasmobranchii; but in addition to pointing out that the optic nerves form a chiasma he does not particularly mention the features, to which he alludes in general terms. More recently Wilder (No. 15) has returned to this subject; and though, as we have already had occasion to point out, we cannot accept all his identifications of the parts of the Ganoid brain, yet he has called attention to certain characteristic features of the cerebrum which have an undoubted systematic value.

The distinctive characters of the Ganoid brain are, in our opinion, (1) the great elongation of the region of the thalamencephalon; and (2) the unpaired condition of the posterior part of the cerebrum, and the presence of so thin a roof to the ventricle of this part as to cause it to appear open above.

The immense length of the region of the thalamencephalon is a feature in the Ganoid brain which must at once strike any one who examines figures of the brains of Chondrostei, _Polypterus_, or _Amia_. It is less striking in the adult _Lepidosteus_, though here also we have shewn that the thalamencephalon is really very greatly developed; but in the larva of _Lepidosteus_ this feature is still better marked, so that the brain of the larva may be described as being more characteristically Ganoid than that of the adult.

The presence of a largely developed thalamencephalon at once distinguishes a Ganoid brain from that of a Teleostean Fish, in which the optic thalami are very much reduced; but _Lepidosteus_ shews its Teleostean affinities by a commencing reduction of this part of the brain.

The large size of the thalamencephalon is also characteristic of the Ganoid brain in comparison with the brain of the Dipnoi; but is not however so very much more marked in the Ganoids than it is in some Elasmobranchii.

On the whole, we may consider the retention of a large thalamencephalon as a primitive character.

The second feature which we have given as characteristic of the Ganoid brain is essentially that which has been insisted upon by Wilder, though somewhat differently expressed by him.

The simplest condition of the cerebrum is that found in the larva of _Lepidosteus_, where there is an anterior pair of lobes, and an undivided posterior portion with a simple prolongation of the third ventricle, and a very thin roof. The dorsal edges of the posterior portion, adjoining the thin roof, usually become somewhat everted (cf. Wilder), and in _Lepidosteus_ these edges have in the adult a very great development, and form (vide Plate 38, fig. 47A-C, _ce´._) two prominent lobes, which we have spoken of as the posterior cerebral lobes.

These characters of the cerebrum are perhaps even more distinctive than those of the thalamencephalon.

In Teleostei the cerebrum appears to be completely divided into two hemispheres, which are, however, all but solid, the lateral ventricles being only prolonged into their bases. In Dipnoi again there is either (_Protopterus_, Wiedersheim[517]) a completely separated pair of oval hemispheres, not unlike those of the lower Amphibia, or the oval hemispheres are not completely separated from each other (_Ceratodus_, Huxley[518], _Lepidosiren_, Hyrtl[519]); in either case the hemispheres are traversed for the whole length by lateral ventricles which are either completely or nearly completely separated from each other.

Footnote 517: _Morphol. Studien_, III. Jena, 1880.

Footnote 518: "On _Ceratodus Forsteri_," _Proc. Zool. Soc._ 1876.

Footnote 519: _Lepidosiren paradoxa._ Prag. 1845.

In Elasmobranchii the cerebrum is an unpaired though bilobed body, but traversed by two completely separated lateral ventricles, and without a trace of the peculiar membranous roof found in Ganoids.

Not less interesting than the distinguishing characters of the Ganoid brain are those cerebral characters which indicate affinities between _Lepidosteus_ and other groups. The most striking of these are, as might have been anticipated, in the direction of the Teleostei.

Although the foremost division of the brain is very dissimilar in the two groups, yet the hind-brain in many Ganoids and the mid-brain also in _Lepidosteus_ approaches closely to the Teleostean type. The most essential feature of the cerebellum in Teleostei is its prolongation forwards into the ventricles of the optic vesicles as the valvula cerebelli. We have already seen that there is a homologous part of the cerebellum in _Lepidosteus_; Stannius also describes this part in the Sturgeon, but no such part is represented in Müller's figure of the brain of _Polypterus_, or described by him in the text.

The cerebellum is in most Ganoids relatively smaller, and this is even the case with _Amia_; but the cerebellum of _Lepidosteus_ is hardly less bulky than that of most Teleostei.

The presence of tori semicirculares on the floor of the mid-brain of _Lepidosteus_ again undoubtedly indicates its affinities with the Teleostei, and such processes are stated by Stannius to be absent in the Sturgeon, and have not, so far as we are aware, been described in other Ganoids. Lastly we may point to the presence of well-developed lobi inferiores in the brain of _Lepidosteus_ as an undoubted Teleostean character.

On the whole, the brain of _Lepidosteus_, though preserving its true Ganoid characters, approaches more closely to the brain of the Teleostei than that of any other Ganoid, including even _Amia_.

It is not easy to point elsewhere to such marked resemblances of the Ganoid brain, as to the brain of the Teleostei.

The division of the cerebrum into anterior and posterior lobes, which is found in _Lepidosteus_, probably reappears again, as already indicated, in the higher Amphibia. The presence of the peculiar vesicle attached to the roof of the thalamencephalon has its parallel in the brain of _Protopterus_, and as pointing in the same direction a general similarity in the appearance of the brain of _Polypterus_ to that of the Dipnoi may be mentioned.

There appears to us to be in no points a close resemblance between the brain of Ganoids and that of Elasmobranchii.

SENSE ORGANS.

_Olfactory organ._

_Development._--The nasal sacks first arise during the late embryonic period in the form of a pair of thickened patches of the nervous layer of the epiblast on the dorsal surface of the front end of the head (Plate 37, fig. 39, _ol._). The patches very soon become partially invaginated; and a small cavity is developed between them and the epidermic layer of the epiblast (Plate 37, figs. 42 and 43, _ol._). Subsequently, the roof of this space, formed by the epidermic layer of the epiblast, is either broken through or absorbed; and thus open pits, _lined entirely by the nervous layer of the epidermis_, are formed.

We are not acquainted with any description of an exactly similar mode of origin of the olfactory pits, though the process is almost identical with that of the other sense organs.

We have not worked out in detail the mode of formation of the double openings of the olfactory pits, but there can be but little doubt that it is caused by the division of the single opening into two.

The olfactory nerve is formed very early (Plate 37, fig. 39, I), and, as Marshall has found in Aves and Elasmobranchii, it arises at a stage prior to the first differentiation of an olfactory bulb as a special lobe of the brain.

_The Eye._

_Anatomy._--We have not made a careful histological examination of the eye of _Lepidosteus_, which in our specimens was not sufficiently well preserved for such a purpose; but we have found a vascular membrane enveloping the vitreous humour on its retinal aspect, which, so far as we know, is unlike anything which has so far been met with in the eye of any other adult Vertebrate.

The membrane itself is placed immediately outside the hyaloid membrane, _i.e._ on the side of the hyaloid membrane bounding the vitreous humour. It is easily removed from the retina, to which it is only adherent at the entrance of the optic nerve. In both the eyes we examined it also adhered, at one point, to the capsule of the lens, but we could not make out whether this adhesion was natural, or artificially produced by the coagulation of a thin layer of albuminous matter. In one instance, at any rate, the adhesion appeared firmer than could easily be produced artificially.

The arrangement of the vessels in the membrane is shewn diagrammatically in Plate 38, fig. 49, while the characteristic form of the capillary plexus is represented in Plate 38, fig. 50.

The arterial supply appears to be derived from a vessel perforating the retina close to the optic nerve, and obviously homologous with the artery of the processus falciformis and pecten of Teleostei and Birds, and with the arteria centralis retinæ of Mammals. From this vessel branches diverge and pursue a course towards the periphery. They give off numerous branches, the blood from which enters a capillary plexus (Plate 38, figs. 49 and 50) and is collected again by veins, which pass outwards and finally bend over and fall into (Plate 38, fig. 49) a circular vein (_cr.v._) placed at the outer edge of the retina along the insertion of the iris (_ir_). The terminal branches of some of the main arteries appear also to fall directly into this vein.

The membrane supporting the vessels just described is composed of a transparent matrix, in which numerous cells are embedded (Plate 38, fig. 50).

_Development._--In the account of the first stages of development of _Lepidosteus_, the mode of formation of the optic cup, the lens, &c., have been described (vide Plates 35 and 36, figs. 23, 26, 35). With reference to the later stages in the development of the eye, the only subject with which we propose to deal is the growth of the mesoblastic processes which enter the cavity of the vitreous humour through the choroid slit.

_Lepidosteus_ is very remarkable for the great number of mesoblast cells which thus enter the cavity of the vitreous humour, and for the fact that these cells are _at first unaccompanied by any vascular structures_ (Plate 37, fig. 43, _v.h_). The mesoblast cells are scattered through the vitreous humour, and there can be no doubt that during early larval life, at a period however when the larva is certainly able to see, every histologist would consider the vitreous humour to be a tissue formed of scattered cells, with a large amount of intercellular substance; and the fact that it is so appears to us to demonstrate that Kessler's view of the vitreous humour being a mere transudation is not tenable.

In the larva five or six days after hatching, and about 15 millims. in length, the choroid slit is open for its whole length. The edges of the slit near the lens are folded, so as to form a ridge projecting into the cavity of the vitreous humour, while nearer the insertion of the optic nerve they cease to exhibit any such structure. The mesoblast, though it projects between the lips of the ridge near the lens, only extends through the choroid slit into the cavity of the vitreous humour in the neighbourhood of the optic nerve. Here it forms a lamina with a thickened edge, from which scattered cells in the cavity of the vitreous humour seem to radiate.

At a slightly later stage than that just described, blood-vessels become developed within the cavity of the vitreous humour, and form the vascular membrane already described in the adult, placed close to the layer of nerve-fibres of the retina, but separated from this layer by the hyaloid membrane (Plate 38, fig. 48, _v.sh._). The artery bringing the blood to the above vascular membrane is bound up in the same sheath as the optic nerve, and passes through the choroid slit very close to the optic nerve. Its entrance into the cavity of the vitreous humour is shewn in Plate 38, fig. 48 (_vs._); its relation to the optic nerve in Plate 37, fig. 46, C and D (_vs._).

The above sheath has, so far as we know, its nearest analogue in the eye of _Alytes_, where, however, it is only found in the larva.

The reader who will take the trouble to refer to the account of the imperfectly-developed processus falciformis of the Elasmobranch eye in the treatise _On Comparative Embryology_, by one of us[520], will not fail to recognize that the folds of the retina at the sides of the choroid slit, and the mesoblastic process passing through this slit, are strikingly similar in _Lepidosteus_ and Elasmobranchii; and that, if we are justified in holding them to be an imperfectly-developed processus falciformis in the one case, we are equally so in the other.

Footnote 520: Vol. II. p. 414 [the original edition].

Johannes Müller mentions the absence of a processus falciformis as one of the features distinguishing Ganoids and Teleostei. So far as the systematic separation of the two groups is concerned, he is probably perfectly justified in this course; but it is interesting to notice that both in Ganoids and Elasmobranchii we have traces of a structure which undergoes a very special development in the Teleostei, and that the processus falciformis of Teleostei is therefore to be regarded, not as an organ peculiar to them, but as the peculiar modification within the group of a primitive Vertebrate organ.

SUCTORIAL DISC.

One of the most remarkable organs of the larval _Lepidosteus_ is the suctorial disc, placed at the front end of the head, to which we have made numerous allusions in the first section of this memoir.

The external features of the disc have been fully dealt with by Agassiz, and he also explained its function by observations on the habits of the larva. We have already quoted (p. 755) a passage from Agassiz' memoir shewing how the young Fishes use the disc to attach themselves firmly to any convenient object. The discs appear in fact to be highly efficient organs of attachment, in that the young Fish can remain suspended by them to the sides of the jar, even after the water has been lowered below the level at which they are attached.

The disc is formed two or three days before hatching, and from Agassiz' statements, it appears to come into use immediately the young Fish is liberated from the egg membranes.

We have examined the histological structure of the disc at various ages of its growth, and may refer the reader to Plate 34, figs. 11 and 13, and Plate 37, figs. 40 and 44. The result of our examination has been to shew that the disc is provided with a series of papillæ often exhibiting a bilateral arrangement. The papillæ are mainly constituted of highly modified cells of the mucous layer of the epidermis. These cells have the form of elongated columns, the nucleus being placed at the base, and the main mass of the cells being filled with a protoplasmic reticulum. They may probably be regarded as modified mucous cells. In the mesoblast adjoining the suctorial disc there are numerous sinus-like vascular channels.

It does not appear probable that the disc has a true sucking action. It is unprovided with muscular elements, and there appears to be no mechanism by which it could act as a sucking organ. We must suppose, therefore, that its adhesive power depends upon the capacity of the cells composing its papillæ to pour out a sticky secretion.

MUSCULAR SYSTEM.

There is a peculiarity in the muscular system of _Lepidosteus_, which so far as we know has not been previously noticed. It is that the lateral muscles of each side are not divided, either in the region of the trunk or of the tail, into a dorso-lateral and ventro-lateral division.

This peculiarity is equally characteristic of the older larvæ as of the adult, and is shewn in Plate 41, figs. 67, 72, and 73, and Plate 42, figs. 74-76. In the Cyclostomata the lateral muscles are not divided into dorsal and ventral sections; but except in this group such a division has been hitherto considered as invariable amongst Fishes.

This character must, without doubt, be held to be the indication of a very primitive arrangement of the muscular system. In the embryos of all Fishes with the usual type of the lateral muscles, the undivided condition of the muscles precedes the divided condition; and in primitive forms such as the Cyclostomata and Amphioxus the embryonic condition is retained, as it is in _Lepidosteus_.

SKELETON.