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ERABADMRE be hte A beth ME BOM MSY Pies Dy dh BLA Pe Bae he 6 Sidod ABE at <8, e a Sa aaies Ld wa ee earre ae ELE BA Hat be ENB aOR EA BAER SEM OE ML BM ABRAM DRAMA EE ECO er AE MOE EMM PR BMD her th ikon Mh te hobo h Ksbod) GAMA AE Bote 0 Btn OR BG RRS BNI BE BD Ce eh OE Mat gE Ome Bok Mh id Pot A dbok bn Ke MAY OL ELD BD Fal ON PPE EIR RAAB LE EBONM DOLD i NE end oad tel Pe Te EBT ae Oe hie BAPE MOTE OER Oe FPP AD RAPA LRA PAOLA RAL 2 ADA BYURS YALE ARLE EK BM DAKO DDD E tb DSLR A BL Bd A Aa bP BEAR DM hd sah A bh A OD bg AMF ee ee oe ee DL DAO EDT OOF id Ce ee ed oi ae weap eee ere eee es MOP PE De AAS ee eS Se es ee ee YAS 5 PAL IRE MENA OL EPPS EA DBA ALMA HAS MAPS ERM OARS MK BUELL oe RA Ma EEL atte rene ee ee eee ee ee ee “n ee ee ee ie ee ee) AMMO OMS AE RAM BS Dh PM PMO ASA SNO SII PAR AHS th , i ye , aw aw? ut ) BA ‘ j f ne | ! ’ ; a, ti ey ita j i i \ r a il : 4 : " vy ( 7 j An aT ih Ae + Oe fat 7) ¥ Les, Ai y eae ; J ‘ f ' Potty” i" i. ie ae ad 7 Wai i at ig 7 by ACM, I ’ , ite iP fy fy i. f 4 ay GaP RC i i vi Lyn it iL Li : j aft vA hay j fi 5 ae ane ey ty | ses Aud ate vy he eT OTN A aA Lt i j a A ak iy “i iy bi me =, g bSEOS oo 1Oe0 O WWW 10 HM/181N + a Bee Me Zz AMPHIOXUS AND THE ANCES PRY OF THE VERTEBRATES Columbia Gnibersity Biological Series. a EDITED BY HENRY FAIRFIELD OSBORN. 1. FROM THE GREEKS TO DARWIN. By Henry Fairfield Osborn, Sc.D. Princeton. 2. AMPHIOXUS AND THE ANCESTRY OF THE VERTEBRATES. By Arthur Willey, B.Sc. Lond. Univ. 3. FISHES, LIVING AND FOSSIL. An Introductory Study. By Bashford Dean, Ph.D. Columbia. 4. THE CELL IN DEVELOPMENT AND INHERITANCE. By Edmund B. Wilson, Ph.D. J.H.U. ee : i ‘ ad a * ay ; Z) » iT | v . $ 7 \ ‘ A eee are > v; 1 a, f : i a 7 all ae : i. e. Wie: ha y ¥ * " uy ~ J ‘ » ~ } =. : 7) if o R be \ » 7 ’ ’ i , ‘ % x; ’ t ee | gl » . ~~ ~ rd a ri 222191] UOLT ‘(VNISSHJ) ONVY LY ONVINVG FHL I SOXOTHINY COLUMBIA UNIVERSITY BIOLOGICAL SERIES. J. AMPHIOXUS AND THE ANCESTRY OF THE VERTEBRATES BY ARTHUR, WILLEY, BSc Tutor In BioLtocy, CotumpiA CoLLEGE; BALFouR STUDENT OF THE UNIVERSITY OF CAMBRIDGE WITH A PREFACE BY HENRY FAIRFIELD OSBORN New Work MACMILLAN AND GO. AND LONDON 1894 All rights reserved CopyrIGHT, 1894, By MACMILLAN AND Co, 16% Norioood Jress. J. S. Cushing & Co.— Berwick & Smith. Boston, Mass., U.S.A. Dedicated IN GRATITUDE AND ESTEEM TO PROPESSOR, ES RAY -LANKESTER, EER. s- BY HIS FORMER PUPIL THE AUTHOR = ty ial Se ce ae Ve he Tee | Deeg ; i . SLs idee | * tg oat) ‘ ; ° , * t . i , ’ . - . . * ' ‘oy Yi. 5 ; ¢ : * ft ay PRP eae B: Tuis volume originated in a course of University lec- tures prepared at my suggestion by the author. It seemed important that he should bring within the reach of students and of specialists among other groups, his own extensive observations upon Amphioxus and other remote ancestors of the Vertebrates, as well as the general litera- ture upon this group. While our detailed knowledge of the structure and habits of these animals has been rapidly increasing in recent years, it is still in the main very widely scattered in monographs and special papers. Probably no single group illustrates more beautifully the principles of transformism ; for the Protochordates in their embryonic development exhibit remarkable reminis- cences of past adaptations, and, in their adult develop- ment, the most varied present adaptations to pelagic, deep-sea, littoral, free-swimming, and sessile life. As Lankester has shown, the Ascidians alone give us a whole chapter in Darwinism. But degeneration and change of function constitute only one side of their history. In vii Vill PREFACE. progressive development some of these types have come to so closely resemble, superficially, certain of the larger groups of Invertebrates, such as the Molluscs and Worms, that it is only at a comparatively recent date they have found their way out of these groups into the Protochor- data. Many of these misleading resemblances are now interpreted as parallels of structure springing from parallels in life habit, seen not only in the general body form, but in special organs, such as the breathing apparatus of the Ascidians and Molluscs. By the side of parallelisms are real invertebrate and vertebrate affinities; so that the problem of resolving these various cases of original and acquired likeness in their bearing upon descent has become one of the most fascinating which modern Zodlogy affords. For example, among the real invertebrate ties of the Protochordates are the ciliated embryos of Balanoglossus and Amphioxus, the Tornaria larva and ciliated ectoderm of Balanoglossus. The nervous system of Balanoglossus presents both ver- tebrate and invertebrate characters; the respiratory sys- tem is identical with that of Amphioxus, while in the embryonic development there are many resemblances zz¢er se. In short, in Balanoglossus and the Ascidians the invertebrate type of structure, whether original or ac- quired, predominates. But in Amphioxus the balance is far on the other or vertebrate side of the scale, and this, with its resemblances to lower forms, gives us the con- PREFACE. ix necting link between Protochordate and Chordate organ- isation. Before entering into any of these discussions, the author has given a thorough systematic and structural treatment, especially of Amphioxus. This exquisite form, Amphioxus, is of almost world-wide distribution and has enjoyed the attention of every great zoologist for over half a century, yet the most recent studies upon it have been among the most productive of discovery. Its interest and value as an object of biologi- cal education has steadily increased with the knowledge that in contrast with all the related forms, it stands as a persistent specialised but not degenerate type, perhaps not far from the true ancestral line of the Vertebrates. Hoi ©: try q Qe OD F4 & J = CON TENTS: : FNS OIONOKG IG COINS = ao 6 aa eet cen kor 1 on Boa co ok llon oor ad al I, JNSZMIROIMAY ONY ANGE HOD AUISTS Sy Mio 8 tn 6 sd some tld we < 7 JE UISUO UCN Cred G94 -Mbkinc. Goro to. e Geo, kor. 0) © “a ou owe 5 JBL A GES) AUSIO) IDMSTAMESUANIONT§ 5 5 65 0 6 of 56 HY om 8 5 6 © 9 IDRGNDINPNE, AON UGC Me ye cee gine wpce Ge (Bea Dk LOLF OhOR Qu val dow iq a (Crbaiutn cyxel Sensors. go % 6 6 6 o Bb 6 ao 6 iy INE RIN AT ANNTAT © Miven ie tuts teu W Pom ay eerell ts mesa ey scat eee wn tne nec2 AttrialCavity: ape Wes “tye WM Mf jj a ‘ f & V i; ip V % Gail) x Figs. 36 and 37. — Schematic transverse sections through post-pharyngeal region, illustrating mode of origin of atrial chamber. (After LANKESTER and WILLEY.) ao. Aorta. 6.c.Ccelom. +m and lm. Right and left metapleural folds. s.a.7. Sub- atrial ridges, which fuse together to form the floor of a/, the atrium. if. Aliment- ary canal. s.z.v. Sub-intestinal vein. SS meeting and coalescence of these sudatrial ridges that the atrial cavity becomes enclosed as a small median tube lined by ectoderm. As soon as it has become closed off from the exterior, the atrial tube commences to grow in size, and it gradually INTERNAL ANATOMY. ha expands laterally and also in an upward direction, propor- tionately reducing the extent of the ccelom as it does so (Fig. 37; cf. also Figs 26): At its posterior extremity the atrial tube does not become closed in, but remains perma- nently open as the atriopore. It is a curious fact that the fusion of the subatrial ridges to enclose the atrial tube takes place gradually from behind forwards, so that for a long time the latter has the form of a canal open to the exterior at both ends. The chief feat- ures in the formation of the atrium are shown diagrammat- ically in Fig. 38, A, B, and C. In Fig. 38 A the atrial tube has not begun to be closed in, but the two metapleural folds are seen running side by side for some distance. Anteriorly the development of the right metapleur is in advance of that of the left, and it is seen to bend round to the right side of the body in correspondence with the asymmetry of the gill- slits (vide infra). Having ar- rived at the front end of the pharynx, the right metapleur bends sharply inwards to the gradually dies out in front. Fig. 38.— Three plastic diagrams of larvze of Amphioxus from the ven- tral aspect, illustrating the mode of enclosure of the atrial tube from be- hind forwards. The atrium is still entirely unclosed in 4; partially closed in &; and almost completely closed in C. (After LANKESTER and WILLEY.) p.s. Primary gill-slits. 7. Right metapleur. 7.2. Przeoral pit. 0. Mouth. at.p. Atriopore. mid-ventral line and then In Fig. 38 B the subatrial ridges have met and fused for a short distance behind the 78 ANATOMY OF AMPHIOXUS. pharynx, so as to enclose a tube which corresponds to that portion of the future atrial cavity which lies between the atriopore and the hinder end of the pharynx. Finally, in Fig. 38 C, the closure of the atrial tube has advanced forwards over the gill-slits almost to the anterior extremity of the pharynx, still leaving, however, one or two gill-slits open directly to the exterior in front. Meanwhile, the floor of the atrium has increased in width, and the meta- pleural folds are separated by a wider interval than before (Fig. 38 C). Eventually the atrium closes up completely in front, so that the gill-slits no longer open directly to ENG vexXt Emon, Remembering that the atrium of Amphioxus arises as an unpaired median tube (see below, IV.), while the pro- nephric duct is always paired, the following are some of the reasons for supposing a partial homology between the two structures :— (a) They are both derived, either wholly (atrium), or in a large measure (pronephric duct), from the ectoderm.® (8) They both receive and carry away the excretory prod- ucts from the pronephric tubules; and (y), they are both, to a greater or less extent, lined by an epithelium, which is itself glandular and excretory.’ Comparison between the Excretory System of Amphioxus and that of the Annelids. Having considered the relation existing between the pronephric system of Amphioxus and the corresponding system in the embryonic and larval stages of the higher Vertebrates, we will now pass on to a brief comparison with the excretory system of the Invertebrates. The excretory system of a typical Annelid presents INTERNAL ANATOMY. 79 certain resemblances to that of Amphioxus, in that it occurs in the form of distinct segmental tubules, or nephridia, each possessing a funnel-shaped opening into the body-cavity, and an opening to the exterior at the sur- face of the body. It was, in fact, the recognition, some twenty years ago, by SEMPER and BALFour, of the resemblance between the arrangement of the nephri- dia of the Annelids and the primary segmental ori- gin of the kidney of the Craniota that was chiefly instrumental in placing the Annelid-theory of Verte- brate descent on a tempo- rarily firm basis. A dissection of the an- terior portion of the body of an earthworm, exposing Fig. 39.—Anterior portion of earth- the nephridial tubules, is shown in Fig. 39. A pair of such convoluted tubules occurs in each segment, or worm dissected open from above to show the nephridia and nervous system. (From W. T. SEDGWICK and E. B. WILSON’s General Biology.) pr. Prostomium (preoral lobe). c.g. Cerebral ganglion, which has receded from the prostomium from the ectoderm of which it arose. com. Circumcoesophageal commissure surrounding the buccal tube (latter not represented). v.x.c. Ventral nerve-cord. 2. Segmental nerves. zh. Nephridia. sf. Dissepiments. ring, of the body, com- mencing from the third. Physiologically, of course, they are directly com- parable to the renal tubules of the Chordata, and in their general features, allowing for the absence of a common duct, the similarity in the two cases is striking enough. But when this undoubted similarity is used as an argument for deriving the Vertebrate excretory system directly from that of the Annelids, we tread on very uncertain ground. SO ANATOMY OF AMPHIOXUS. If we were to consider the excretory system apart from the rest of the organisation, this would be the only course to follow. But when the whole organisation is taken into account, the only justifiable conclusion seems to be, not that the Vertebrate renal system is to be derived from that of the Annelids, but that, as Riickert suggests, both may possibly have been evolved from a common starting-point. It is eminently probable that, in respect to this and the other systems of organs, as well as the segmentation of the body, the Annelids and Vertebrates present an in- stance of parallel evolution. This will become more evi- dent as we proceed. Those who uphold the so-called Annelid-theory have no cause to complain of the absence of a common duct to the nephridia, since this has been found in some cases to occur. In 1884 EpuARD MEYER discovered that in certain marine Annelids (Lanzce conchilega and Loimta medusa) belonging to the family of the Terebellida, the nephridia of each side were joined together by longitudinal ducts, which he compared, though with great reserve, to the mesonephric ducts of the Vertebrata.* In these worms the nephridia do not occur in all the segments of the body, but are confined to the anterior so-called thoracic region, their number being very limited. In the thorax, the dissepi- ments which typically divide the segments from one another are absent, so that the body-cavity would here form a continuous uninterrupted space, were it not that it is divided into two chambers, an anterior and a posterior, of which the latter is the larger, by a muscular diaphragm. In the anterior thoracic chamber (Fig. 40) there are three pairs of nephridia which are united together on each side by a short duct opening to the exterior by a single aperture. * This discovery was also made later but independently by J. T. CuNNING- HAM for Lance conchilega. INTERNAL ANATOMY. SI In the posterior chamber there are four pairs of much larger nephridia, which are similarly joined together by a prominent longitudinal duct from which short processes corresponding in number The duct itself ends blindly at external apertures. both ends, but is prolonged posteriorly far beyond the region of the nephridia (Fig. 40). (thes spresence ors sthis longitudinal duct in these worms is a very remark- able circumstance, but it is undoubtedly an expression of the same phenomenon as the anastomoses between successive nephridia which have been described by Ersic for the Capitellidz, as well as the complicated series of anastomoses which convert the entire nephri- dial system into a marvel- lous network of tubules dis- covered by A. G. BouRNE in the marine leech, Povz- tobdella, and by BEDDARD in the curious earthworm, Pericheta. to the nephridia lead to the Fig. 40. — Schematic lateral view of anterior end of Lanice conchilega to show the nephridia. (After EDUARD MEYFR from Hatschek’s Lehrbuch's der Zoologie.\ The ventral side of the body is to the left of the figure. d. Longitudinal ducts of the nephridia. ¢.0. Position of external openings. /£ Nephridial funnel (=ccelomic opening of nephridium). a. Position of mouth; bounded by two prominent lateral lobes, and fringed by a great number of “feelers,” which are cut short in the figure. ?. Branchial tentacles (three on each side of the body). The present state of our knowledge does not admit of an attempt to specify the particular type of nephridial system from which that of the Annelids, on the one hand, §2 ANATOMY OF AMPHIOXUS. and that of the Vertebrates, on the other, took their origin. In view of the apparent absence of nephridial tubules in Balanoglossus and the fact that in the Ascidians the renal organs are special structures peculiar to this group, it is extremely difficult to associate the Vertebrate type of excretory system with that of any Invertebrate. Since the Annelid-theory precludes the possibility of Amphioxus being regarded as an ancestral form, and yet if, nevertheless, it is, as we believe, primitive and not essentially degenerate, the discovery of the excretory tubules in Amphioxus happily releases us not only from necessity, but also from the possibility of referring the Vertebrate excretory system back to that of the Annelids. ° Nervous System. The central nervous system of Amphioxus consists of a closed thick-walled tube lying along the dorsal side of the body above the notochord. Viewed externally, it is a perfectly plain, more or less cylinder-shaped structure, without any constrictions or enlargements whatever. Its largest diameter in the adult occurs about the middle of its course, and not at its anterior end. Posteriorly it is nearly coextensive with the notochord, and, like it, tapers down almost to a point.* Anteriorly it terminates abruptly some distance behind the front end of the notochord. (Cf. Figs. 3 and 11.) If the dorsal nerve-cord be removed from the body and * The extreme posterior end of the nerve-cord is usually swollen out into a small ampulla-like dilatation. (PouCcHET, ROHON, RETzIUs.) RETzIUS has observed that occasionally the nerve-cord is prolonged beyond the dilata- tion and actually bends round the posterior end of the notochord. INTERNAL ANATOMY. 83 examined from above, its general appearance will be as shown in Fig. 41. In front there is a pair of nerves which proceed symmetrically from the sides of the nerve- tubes. “Harther back there: is another pair of nerves which arise more dorsally than the anterior pair, but are likewise placed symmetrically one oppo- Behind this second pair of nerves the spinal site the other. nerve-roots are no longer dis- posed symmetrically, but alter- nate with one another, in cor- similar respondence with a alternation of the myotomes, the alternation becoming more and more pronounced as we proceed backwards. Again, be- hind the second pair of nerves there are two kinds of spinal nerve-roots, dorsal and ventral. The former leave the nerve-cord from its dorsal surface, and the latter from the margins of its ventral side. In the dorsal roots the nerve-fibrils are collected together to form a single com- pact nerve round which the sheath of the nerve-cord is con- tinued, while in the ventral roots 4 i Fig. 41.— Anterior portion of spinal cord of Amphioxus; seen from above. (After SCHNEIDER.) Between the first pair of cranial nerves is seen the eye-spot; one of the branches of the second pair of cranial nerves sometimes arises directly from the spinal cord as shown on the right; farther back are seen the pigment spots of the nerve-cord. 4 VIO Taeeie Fr e> oO e upon four larger blastomeres (ma- meres again into two. In this cromeres). Radial type of cleavage. way the stage with four equal (A** ® B. Wirson.) blastomeres in one plane is produced. Next follows an equatorial cleavage, by which eight blastomeres are pro- duced, the four upper cells at the animal pole being some- 108 DEVELOPMENT OF AMPHIOXUS. what smaller than the four lower cells at the vegetative pole, since the latter contain a greater quantity of the yolk-spherules (Fig. 55). The next cleavage giving rise to an embryo of sixteen cells is meridional. Then the eight upper and the eight lower cells become respectively divided by equatorial cleavages, and so the thirty-two cell stage is reached (Fig. 56). The embryo is now known as a dblastula, and consists of a mulberry-like mass of cells sur- rounding a central cavity called Fig. 56.— Thirty-two cell stage, : : consisting of four rows of eight cells, the segmentation-cavity OY blas- each ranged around a central seg- mentation cavity (blastoccel). The polar body is still visible at the ani- From this point of the de- mal pole. (After HATSCHEK.) tocal. velopment the blastomeres go on dividing with more or less regularity, until the wall of the blastula consists of a great number of cells arranged in a single layer about the central cavity. The segmentation of the egg of Amphioxus, however, by no means follows the uniform and stereotyped plan that has been hitherto supposed. It has recently been discovered by Professor E. B. Witson that Amphioxus presents an example of a polymorphic cleavage. Instead of following one type, it follows three types of cleavage ; namely, a vadial type (Figs. 55 and 56), a dzlateral type (Fig. 57), and a spiral type (Fig. 58). These three types of cleavage are reducible to a common basis, and are con- nected together by all possible intermediate gradations. Wilson points out that in the bilateral type of cleavage Amphioxus shows a close correspondence with the Ascid- ian embryo. EMBRYONIC DEVELOPMENT. 109 The segmentation or cleavage of the ovum results in the formation of a spherical blastula, closed at all points, Fig. 5'7.-— Three stages in the segmentation of the ovum, according to the bilateral type. From the lower pole. (After E. B. WILSON.) A. Eight-cell stage. A, B, CD. The four macromeres, above which are seen portions of the four micromeres. /-/. Plane of first cleavage, with respect to which the cells divide in such a way as to become arranged in a bilaterally symmetrical manner. //-II, Plane of second cleavage. 2B. Transition to the sixteen-cell stage. C. Sixteen-cell stage. The line in each cell indicates the direction in which the next division of the cell would take place. and consisting of some 256 cells surrounding a spacious cavity, the blastoccel. The stages of development lead- ing up to the blastula are known as the segmentation stages. At their completion, although, of course, cell-division continues to take place actively, yet other pro- cesses supervene which render the _ Fig. 58.— Eight-cell stage pgs Sears from the upper pole, illlustrat- mere division of the individual cells jing the spiral type of cleavage. of minor morphological importance. (fer E- B- W11s0N.) Gastrulation. The next phase of the development is known as the gastrulation of the embryo. The cells forming the lower or vegetative side of the blastula remain, throughout the segmentation stages, somewhat larger than the rest of the IIO DEVELOPMENT OF AMPHIOXUS. blastula-wall. This side now becomes flattened, as shown in Fig. 59 A. Next, the flattened side of the blastula becomes gradually tucked up or invaginated into the blastoceel (Fig. 59 4) until, finally, the segmentation cavity is completely obliter- ated, and the invaginated (After HATSCHEK.) layer of cells becomes tightly fitted up against the outer layer (Fig. 50°C). The embryo, now known as the gastrula, is a double- layered sac, the cavity of optical transverse section. C, The invagination is completed and the blastocoel is obliterated; optical longitudinal section. which was produced by in- ZN vagination, and is known as the premetive gastral cavity, a KD . a Ss Ma Rd : fa or archenteron. This cavity is widely open to the ex- A Fig. 59. — Three stages in the gastrulation of Amphioxus, seen in optical section. A. Blastula with flattened vegetative surface; optical transverse section. et s od) OY a. Os terior by the orifice of invagi- nation, or d/astopore, which in German is designated by the expressive term Uvmund. The two layers of cells which constitute the wall of the gastrula are the pvr7zmztive germ-layers ; the outer layer is the primitive ectoderm, B. Lower pole becomes invaginated into the blastoccel ; and the inner layer, sur- rounding the gastral cavity, is the primitive endoderm ; the two layers are continuous with one another round the margin of the blastopore. The blastopore is at first a very wide oval opening, but it soon becomes narrowed down to a small aperture EMBRYONIC DEVELOPMENT. Li by the continued deepening of the archenteric cavity (Hig: (60): It is now a well-established fact that all multicellular animals (Metazoa) pass through a gastrula-stage in the course of their development, although the form of the gastrula is often extremely modified and difficult to recog- nise. The significance of this fact, as was long since pointed Cor out by Huxley, Haeckel, Lan- kester, and others, is very great when it is remembered oon that the embryonic character- 853 istics of the gastrula are essentially identical with the permanent features of the : ° Fig. 60.— Optical longitudinal sec- organisation of the Ccelen- tion of later gastrula. Cilia (flagella) terata (Hydra, etc. ). have been protuded from the ectoderm f cells, and the embryo at this stage Returning to the gastrula begins to rotate within the follicle. 5 ¢ (After HATSCHEK.) of Amphioxus, in the course of the further differentiation which goes hand in hand with the progressive growth and development, we shall find that the primitive ectoderm gives rise to (1) the central nervous system and (2) the definitive ectoderm ,; the primitive endoderm gives rise to (1) the mesoderm, which is usually regarded as a third or intermediate germ-layer ; (2) the xotochord; and (3) the definitive endoderm, which forms the lining mucous epithelium of the alimentary canal; finally, the primitive gastral cavity or archenteron will become subdivided into (1) the dody-cavity or calom, and (2) the definitive gut or alimentary canal. The embryo shown in optical section in Fig. 60 repre- sents the stage reached at midnight of the first night of development. It will be noticed that one side is convex, 177 DEVELOPMENT OF AMPHIOXUS. while the opposite side is flattened; this is an indication that dorso-ventral differentiation has taken place, since the flattened side marks the dorsum or back of the embryo, while the convex side is ventral. It may be seen further that the blastopore is inclined towards the dorsal side of the embryo. The dorsal inclination of the blastopore is eminently characteristic of the vertebrate gastrula from the Ascidians up to the highest craniate forms. In the Inverte- brates (Annelids, Molluscs, etc.) GED the blastopore acquires a ventral petals inclination.* Be a At the stage represented in Fig. Sean 60 the embryo commences to ro- Boise He tate within the vitelline membrane, oe Sari ed a) each ectodermic cell being now ES provided with a vibratile cilium. Fig. 61. — Elongated gas- 6 trula. Optical longitudinal sec- The embryo next begins to elon- tion. The cilia are omitted gate, and the blastopore becomes) 20m ho) ecod- antes : ; HATSCHEK.) still narrower (Fig. 61). A comparison of the accompanying figures will show that the narrowing of the blastopore is effected by the downward and backward growth of its dorsal border, while the ventral lip remains stationary. The dorsal ecto- derm, which is converted into the medullary plate, now shows indications of a shallow longitudinal groove. This is the beginning of the medullary groove which leads on to the formation of the central nervous system. * For a discussion of the phylogenetic relation of the blastopore or proto- stoma (Hatschek) to the mouth and anus, the following works should be consulted: ADAM SBDGWICK, Ox the Origin of Metameric Segmentation, etc., Quarterly Jour. Micro. Sc., XXIV., 1884, and by the same author, (Votes on Llasmobranch Development, 1b. Vol. XXXIII., 1891-92. Finally, BERTHOLD HATSCHEK, Lehrbuch der Zoologie, Jena, 1888-91. EMBRYONIC DEVELOPMENT. 113 Growth of Free-swimming Embryo. Between 4 and 5 A.M. in the first morning of develop- ment, z.e. at about the eighth hour, the embryo has reached the stage represented in Fig. 62, and it now bursts through the vitelline membrane and becomes free, swimming by means of its cilia at the surface of the sea, or aquarium. The fact that Amphioxus has a free-swimming, ciliated embryo is important as providing a general connecting link between the Vertebrates and the Invertebrates, since eS BES foto Saris! oa Qrle--s=35 Ine i | to re tS] au aut Fig. 62.— Embryo of Amphioxus at the stage at which it ruptures the follicle and becomes free-swimming. A. Seen from above as a semi-opaque object. (After KOWALEVSKY.) B&B. Seen in sagittal (optical) section. (After HATSCHEK.) arc. Archenteron. m.~. Medullary plate. wy.c. Myoccelomic pouches of archenteron. 7.7.c. Posterior neurenteric canal. the possession of a ciliated ectoderm is very common among Invertebrate embryos, but entirely unknown among the craniate Vertebrates. The medullary plate is now being closed off from the outer surface. This is effected by the co-operation of two factors. The ectoderm which bounds the medullary plate laterally, grows over it, and simultaneously the ectoderm of the posterior or ventral lip of the blastopore grows for- ward over the medullary plate so as to shut in the blasto- pore from the exterior (Fig. 62 A and 4). The archenteric I14 DEVELOPMENT OF AMPHIOXUS. cavity therefore no longer opens by the blastopore to the exterior, but it communicates with the medullary tube. The blastopore has, in fact, become converted into the neurenteric canal, joining the canal of the central nervous system with the cavity of the alimentary system. This remarkable condition of things was first discovered by KOowWALEVSKY, who also found it in the Ascidians and in a number of the higher Vertebrates. It has since been found to occur in all classes of Vertebrates, including man. Hitherto the body-wall of the embryo has consisted of only two primary germ-layers, ectoderm and endoderm. At the stage now under consideration, a third interme- diate layer, the mesoderm, has begun to put in its appear- ance. The mesoderm arises in the first instance as a series of paired lateral pouches of the archenteron. In Fig. 62 the first two or three archenteric pouches are distinctly visible. Before proceeding, however, to a more detailed account of the origin of the nervous system and of the mesoderm, we will trace briefly the changes in external appearance which the embryos undergo up to the time of the formation of the mouth. As the embryos are very transparent, the external appearance necessarily involves a good deal of the inter- nal structure. The period of embryonic development may be defined as commencing with the first cleavage of the ovum, and end- ing with the perforation of the mouth, thus comprising approximately the first thirty-six hours. During this period the embryo does not take up independent nourish- ment, but lives on the original food-yolk which was con- tained in the egg. During the first few hours of its pelagic or free-swim- EMBRYONIC DEVELOPMENT. I1l5 ming existence, the embryo keeps rigidly to the surface of the water. After its escape from the vitelline membrane, it grows rapidly in length. Fresh archenteric pouches are added to those already formed, one after the other, in metameric order. The medullary plate (z.e. the fore-cast of the nerve- tube) becomes completely closed in beneath the superficial ectoderm except at its anterior extremity, where it remains open to the exterior in the mid-dorsal line by an aperture known as the zxeuropore (Fig. 63 A, B, C). Finally, the notochord becomes differentiated from the primitive endo- derm. According to Hatschek the number of mesodermic somites which arise as diverticula from the archenteron is fourteen pairs. Those which are subsequently added to these arise at the hinder end of the body by prolifera- tion from the cells which lie behind, and at the sides of the neurenteric canal, or in that region, so that they never appear as actual outgrowths from the archenteron.” In Fig. 63 C the embryo has undergone some radical changes in form. Its body, previously cylindrical, has become laterally compressed, the ectoderm cells of the hinder end of the body have begun to elongate so as to form the rudiment of a provisional caudal fin, and the front end of the body has grown out into the shape of a snout. In connexion with the latter there are two remarkable structures which arise as a pair of outgrowths from the anterior region of the archenteron, and were first described by Hatschek as a pair of anterior intestinal diverticula. These we shall return to later. Near the front end of the alimentary canal a curious sac-like structure has appeared (Fig. 63 C). It arose as a transverse groove in the floor of the gut in the region 116 DEVELOPMENT OF AMPHIOXUS. of the first myotome, extending from the right side under- neath to the left side of the body. (Cf. Figs. 63 A and 71.) The groove deepened, and its margins coalesced, and so it rea mute, my / Fe Nae] of x v7 Tres ee at a SIE elpla o orator ole or arate ee seas = Le bat acs =, ; PS LI mn os S WY] Beso ; eles 8 a & Aer Be we a a wer me oS ees Ses ate SIC Se eS Se aa ee rem ae war niet ae eae ae es ae ky ors ~——- tnt Fig. 63.— Growth of the ciliated embryo of Amphioxus. ghtly altered.) K, sli (After HATSCHE with nine pairs of myoccelomic pouches ; from left side. &. Same stage from dorsal side. A. Stage, with fifteen pairs of myotomes; from Cc. Stage the right side. notochord. Vacuoles have appeared in cells of LS. c.s.g. Club-shaped gland. Notochord. Rudiment of first gill-slit. ch. S066 Za.d. Left head-cavity (left anterior intestinal diverticulum). int, Intestine. my.c. Myoccelomic (archenteric) pouches. np. Neu- gment granules Pi p.m.c. Posterior neuren- PS: n.t. Medullary tube. in floor of medullary tube. ropore. teric canal. r.a.d. Right head-cavity (right anterior intestinal diverticulum). became constricted from the gut, and now forms a hollow sac closed at present at both ends. shaped gland. It is known as the clué- Immediately behind it, in Fig. 63 C, is seen EMBRYONIC DEVELOPMENT. 17 a shallow depression in the floor of the gut. This is the indication of the first gill-slit which becomes perforated at this point later. From this stage it is an easy tran- sition to the stage which marks the close of the embryonic and the com- mencement of the J/arval period of development. In the embryo shown in Fig. 64, the mouth appears as an oval aperture placed asymmetrically on the left side. At its first origin it is relatively much smaller than shown in the figure. A disc-shaped thickening of the ectoderm appears on the left side, in the region of the first myotome. The subjacent endoderm fuses with the thickening, and then the centre of the disc becomes perforated, and so the mouth is formed. The club-shaped gland has acquired an opening to the exterior immediately below the mouth, on the left side; while the body of the gland lies on the right side. Behind the club-shaped gland on the Fig. 64.—Stage in which the external apertures of the body, przeoral pit, mouth, first gill-slit, and anus have become perforated. Age about 36 hours, From the left side. (After HATSCHEK.) ai, Alimentary canal. az. Anus. 4.c. Body-cavity. ch. Notochord. ed. Endostyle. ./. Club-shaped gland, which has acquired an opening to the exterior on the left side below the mouth. g.s’. First primary gill-slit. Fig. 64. m. Mouth. #.c. Nerve- tube; the neurenteric canal has closed up, but the nerve-tube still curves round the hinder end of the notochord. mp. Neuropore. 7.0.c. Przeoral coelom (right head-cavity). #.%. Przeoral pit (left head-cavity). 7 Provisional caudal fin. 118 DEVELOPMENT OF AMPHIOXUS. right side is the first gill-slit, opening directly to the exterior. At the time of its actual perforation it lies near the mid-ventral line of the body, but as it increases in size it becomes shifted up to the right side. The neurenteric canal is closed up, and the nerve-tube ends blindly behind, being curved round the hinder end of the notochord. Immediately in front of and below the point where the neurenteric canal formerly existed, the anus has now made its appearance, approximately, if not exactly, in the mid-ventral line * (Fig. 64). We will now return to consider more closely the exact development of the mesodermic somites, the notochord, and the nerve-cord. Development of Central Nervous System. As in the craniate Vertebrates, so in Amphioxus the medullary plate arises as a median unpaired longitudinal specialised portion of the dorsal ectoderm. The way in which it becomes separated from the superficial ectoderm has already been indicated above, but it can best be studied in transverse sections. In the sections shown in Figs. 65 and 66, the separation of the medullary plate from the ectoderm, and its subse- quent conversion into a closed tube, is so clearly illus- trated, that further description is unnecessary. A unique feature in connexion with the formation of the central nervous system of Amphioxus is, that the medullary plate sinks below and becomes covered over by the superficial ectoderm before it takes on the form of a closed tube, so that for some time it exists as a half-canal open dorsally * According to Hatschek, the anus breaks through slightly to the left of the middle line. 119 EMBRYONIC DEVELOPMENT. ‘sainSy ayy woy readde pynom sev 901vos aie ‘pauayey Ajawiayxa awosaq sey YoryM “wNIEyIC ‘aqny puv ajeid Areynpay, “2% qys-is Areurid ysany "se ‘purls podeys-qni9 7.0" ay} nq ‘(1Xe} Jy} UL O} Pedajet sy} oie fay) H 0} wo (“MAHOSLVH J0yV) SUOHYIII PUL UOTVUIIOJ JO BpOU tj} Suneaysnyt ‘sede juesayip yo sokaquio Y melee NAAN Wy 4 x a < ‘ysnoua oo1vos ale Tajonu eT, * a o1mrojaoo oy} JO Tajonu ay} sainsy OM} ysv[ 94} Ul—"A'N *(sayTWos OTWIaposaut) sayonod orwo *payiuo AjjeyUaploow a1OM S19}9] ay A[BAYNIISUOD patajj}s] UPEq ‘O19 ‘SaPWIOS SIULIapOsatu ‘p1loyoo}ou ‘gqn} Alv|[npaur ay} Jo Snoiyy suoyoas asisasuvsy,— “SO “BIA 120 DEVELOPMENT OF AMPHIOXUS. against the ectoderm. Later the dorsal margins of this half-canal meet and fuse in the middle line, and so produce the medullary tube * (Fig. 66). Origin of Mesoderm and Celom. In consequence of the flattening and incurving of the medullary plate, pressure is brought to bear on the dorsal wall of the archenteron, and the dorso-lateral bor- ders of the latter acquire the form of two longitudinal grooves (Figs. 65 A and 4). It is from these grooves that the archenteric pouches are split off. The grooves deepen, and in doing so become divided up into a series of pouches. Eventually the pouches become shut off from the archenteron gradually from before backwards, and then appear as closed cavities on either side of the notochord, which has, in the meantime, been developing (Fig. 65 /). In the higher Vertebrates the mesoderm arises as two solid, lateral, longitudinal bands, which are split off from the primitive endoderm. These mesodermic bands are at first unsegmented, and might be taken to correspond with the longitudinal grooves of the archenteron of Amphioxus, as described above. Later, only the dorsal portion of the mesodermic bands undergoes segmentation, while the ventral portion, which becomes hollowed out to form the general body-cavity, is never segmented in the crani- ate Vertebrates. (Cf. Fig. 33.) In Amphioxus the whole of the mesoderm is contained in the archenteric pouches, and is, therefore, at first entirely segmented. As soon as the pouches have lost their primitive con- * In the Ascidian embryo the formation of the medullary tube takes place after the manner typical of craniate Vertebrates (see below, IV.). EMBRYONIC DEVELOPMENT. I2I nexion with the archenteron, they commence to extend dorsally and ventrally between the ectoderm and the in- ternal organs (Fig. 66). inner or visceral wall of the pouch adjacent to the noto- Meanwhile the cells forming the chord elongate transversely and longitudinally, and begin to form the plate-like muscle-fibres of the myotome. The cells producing these fibres are arranged in such a way that each of them gives rise to a muscle-fibre extending from the anterior to the pos- terior limit of a myotome.* The muscles are at first closely approximated to the notochord and project freely into the cavity of the pouch. The latter gradually grows downwards, until it meets its fellow of the other side; the two fuse together, and so the cavity 1s made con- tinuous from side to side, Fig. 66.— Transverse section through the middle of the body of an embryo, with ten pairs of somites, to show the closure of medullary tube and the dorsal and ventral extension of the mesodermic somites. (After HATSCHEK.) al, Alimentary canal. ch. Notochord, in the cells of which vacuoles have com- menced to form. d.#. Commencing for- mation of longitudinal muscle-plates from the cells forming the inner wall of the somite. my.c. Myoccelomic cavity, below the intestine. When this occurs, the primarily single cavity of each archenteric pouch becomes divided into two portions ; namely, a dorsal portion, the somfe proper or myocel, and a ventral portion, the cw/om, by a transverse partition, which arises through a fusion between the parietal and * Already in the embryo shown in Fig. 63 C, and even at a somewhat ear- lier stage, the muscles are so far developed that the body can be bent and jerked. By the time the mouth has broken through, muscular locomotion effectually replaces the primitive cz/ary locomotion, although the cilia persist to a late stage. U2Z DEVELOPMENT OF AMPHIOXUS. visceral walls of the cavity, at about the level of the base of the notochord (Fig. 67). The dissepiments between the myotomes are formed from the contiguous walls of the successive pouches, but ventrally, in the region of the ccelom, they break down, so that the latter then becédmes a continuous unseg- mented cavity. On account of the fact that the archen- teric pouches give rise both to the cavity of the somites (myocel) and to the general body-cavity (ccelom proper or splanchnoce!), they are often spoken of as the myo- celomic pouches. The cav- ity of the original archen- Fig. 67. Scheme of a transverse section through the body of a larva with five gill-slits, to show the division between myoccel and_ splanchnoccel. (After HATSCHEK.) z.c. Spinal cord (medullary tube). ch. Notochord. dm. Muscles. my. Myo- cel. sc. Rudiment of sclerotome. al, Alimentary canal. s.z.v. Sub-intestinal teric pouches is known as the primitive the epithelial walls of which caelom, constitute the szesoderm. As differentiation and or- ganogeny proceed, the meso- vein. sp. Splanchnoccel. . : derm gives rise to (1) the musculature, (2) the connective tissue, (3) the blood-vessels, (4) the reproductive organs, (5) the calomic epithelium or lining of body-cavity, also called the perztoneum, and (6) the excretory tubules. The development of the last- named structures has, however, not yet been worked out in Amphioxus. The parietal layer of the mesoderm applies itself closely against the ectoderm, and gives rise to the cutis of the body-wall. The connective tissue-sheath of the notochord and EMBRYONIC DEVELOPMENT. 123 nerve-cord, together with the internal sheath or fascia of the muscles of the myotome, arises from the walls of a pouch-like diverticulum of myoccel which grows up be- tween the muscles and the notochord and nerve-cord. (Cf. Figs. 67 and 68.) The myoccel also grows downwards between the somatic layer of the peritoneum and the ecto- derm (Fig. 68). According to Hatschek the dorsal and ventral fin-spaces are also derived from the myoccel.? The diverticulum of the myoccel which has just been described is known as the sclerotome, since it gives rise to the fibrous sheath of the notochord and_ nerve-cord, which (z.e. the sheath) in most of the higher forms becomes replaced by carti- lage, and finally by bone. In the craniate Vertebrates nha) GbesGdhene @? | Gate the sclerotome arises as a Section through region between atriopore : : ; and anus, of a young Amphioxus shortly solid proliferation of cells after the metamorphosis. (After HatT- ; ; ; SCHEK.) from the visceral wall at the d.f.c. Dorsal fin-space. my. Myoccel. base of the somite. This 5¢.Sclerotome. ao. Aorta. ad, Intestine. i , : s z.m. Intercoelic membrane. 5.2.v. Sub-in- solid proliferation is un- testinal vein. sf. Splanchnoccel. v. fc. ; : Ventral fin- b douptedly-a medication of “°C re a hollow diverticulum, involving, as it does, only the visceral wall of the somite, precisely as we find it in Amphioxus.* (Cf. Fig. 33.) On their outer surface the muscles of the myotomes are not provided with a sheath of connective tissue (fascia), standing, in this respect, in contrast to the condition which obtains in the Craniota. 124 DEVELOPMENT OF AMPHIOXUS. Origin of the Notochora. The notochord is formed from the endodermic cells which lie between the mesodermic pouches and constitute the dorsal wall of the archenteron. The dorsal wall of the archenteron at an early stage becomes converted into a shallow longitudinal groove whose concavity is turned towards the archenteric cavity (Fig. 65 D). This groove gradually deepens (Fig. 65 £), and eventually its walls become closely appressed to one another so as to obliter- ate the lumen (Fig. 65 /). Finally the adjoining cells of the archenteric wall grow across the gap occasioned by the formation of the notochord, and joining together, shut off the latter from any participation in the enteric wall (Fig. 66). In this way is the notochord separated from the endoderm gradually from before backwards. Poste- riorly it remains for a considerable time fused with the endoderm at the point where the anterior wall of the neu- renteric canal becomes continuous with the dorsal wall of the archenteron. We have indicated above that the differentiation of the notochord takes place from before backwards. At its anterior extremity a very noteworthy exception to this rule is presented. In the region of the first myotome the notochord retains an open communication with the archenteron after its lumen has already been obliterated behind this point. Moreover, in the embryo, with eight pairs of myoccelomic pouches (Fig. 68 dzs), the front end of the notochord lies some distance behind the front end of the body, while the anterior portion of the archenteron extends beyond the notochord. Eventually the notochord is continued to the front end of the body by becoming constricted off from the dorsal wall of the anterior sec- EMBRYONIC DEVELOPMENT. I25 tion of the archenteron in the usual way. This retarded growth of the notochord anteriorly indicates that its exten- sion to the tip of the snout is a secondary phenomenon. Ancestrally we are bound to assume it did not extend so far forwards. The forward . oO ‘e] extension of the notochord SEE J is, as noted above, obviously HY a> ho . : pense aay A 772 useful to Amphioxus in ren- ASSN dering its pointed snout Cassese mee _-Ch sufficiently resistant to en- Eat ee | A able it to burrow in the ee 45 7 Oisesese, Seno eo ‘a sand. When it wants to ace ees yay 5 5 . Spray Bere reas (a os LL f bury itself in the sand, it Poets an ; ; Pees Kooea tort eee) == 45 has not to take pains to dig Pes sta e8 a Be, Psy en ee YO | a hole, but darts in in the po geeagore8 . 49 fraction of a second. gy The histological differen- = ihe Cro, tiation of the notochord Caan commences soon after the sides of the chordal groove Fig. 68 47s.— Embryo of Amphioxus, ‘= with eight pairs of somites to show the have come together so as to primary relations of the anterior end of F the notochord. From above. After obliterate the lumen. The yarscuex.) ( cells composing the noto- p.c. Preechordal portion of archen- 5 teron, which becomes converted into the chord are, at the first ap- head-cavities. .f. Neuropore. ch, Noto- chord; over which lies the neural tube. my. Myoccelomic pouches. ze. Neuren- themeroove; placcas ena to) mc. canal. 2 ae : : N.B.—In this and other figures of end, but soon begin to inter- Amphioxus embryos here reproduced 1 ‘th th after Hatschek, the so-called mesoder- ace with one another across nic pole cells have been omitted in the middle line (Fig 65 F) accordance with the observations of = WILSON and LwoFF. and finally each cell comes to occupy the whole width of the notochord (Fig. 66). Meanwhile vacuoles begin to appear in the cells (Fig. 66). The vacuolisation of its component cells is an extremely proximation of the walls of 126 DEVELOPMENT OF AMPHIOXUS. characteristic feature of the notochordal tissue throughout the group of the Vertebrates. It is carried on to such an extent in Amphioxus as to obscure the original cellular We: WK i it i N MY structure of the notochord. 5 The cells anastomose with one another in the longitu- dinal direction, and so pro- duce a reticulum the meshes Fig. 5 sagittal section of of which represent the vacu- notochord of a young Amphioxus of oles whose first origin is 8 mm., to show the vacuolar character : of the notochordal tissue and the dis- shown in Fig. 66. Most of placement of the nuclei to the dorsal and - ventral borders. (After LWOFF.) the nuclei become eventually displaced from the centre of the notochord, and are, in the adult, almost exclusively confined to its dorsal and ventral aspects (Fig. 69). The Preoral “ Head-cavities” of Amphioxus. Before leaving the embryonic period of the development it is necessary to consider the origin and fate of what may be called the “ead-cavities of Amphioxus as made known to us by the work of Hatschek. They arise symmetrically as a pair of diverticula from the anterior portion of the archenteron, which lies at first partly in front of the notochord (Fig. 68 47s) and completely in front of the myoccelomic pouches (Fig. 70). They begin to appear at the stage in which some eight pairs of pouches are already present. Their origin there- fore, in point of time and the subsequent modifications which they undergo, show that they do not belong to the metameric series of the mesodermic pouches, but are structures saz generis. EMBRYONIC DEVELOPMENT. 127. The archenteron extends at first to the front end of the body. Its anterior portion, after the formation of several mesoblastic somites, becomes marked off from the hinder region by a slight constriction, which gradually becomes deeper and deeper (Fig. 70), until finally the whole of this portion of the archenteron is divided into two separate sacs, which eventually lose all connexion with the ar- chenteron (Fig. 71). The ali- mentary canal now no longer ERTOToIo > > Se ai reaches to the anterior ex- ny tremity of the body. Very SEAT Loans Sista SAS irae st alcloial sola ios soon after their separation from the archenteron these 5 wea EOE. a \ on ae sacs enter upon a series of changes by which their origi- nally symmetrical disposi- tion is entirely destroyed. Already in Fig. 71 it can be noticed that the right sac is becoming larger than Fig. 70.— Embryo, with nine pairs of the left, and the epithelium primitive somites, seen in optical section Ave fi , i ; from the ventral surface, to show the lining its walls is losing its origin of the head-cavites. (After Har- ioinval bical iy SCHEK.) origimal cubical C aracter, y.a.d. Right head-cavity. /a.d. Left the inner ends of the cells head-cavity. zy.c. Myoccelomic pouches : . (primtive somites). arc. Archenteron. are rounding off, and in fact it is being converted from a cubical toa flattened pavement epithelium (Figs. 63 C and 64). The left sac, on the con- trary, retains its original form and dimensions for a long time. During the asymmetrical changes affecting the two sacs, which take place coincidently with the formation of the snout, the left one comes to lie transversely below the notochord, while the right sac becomes greatly enlarged 128 DEVELOPMENT OF AMPHIOXUS. and constitutes the cavity of the snout lying below the notochord (Fig. 63 C). Shortly after the breaking through of the mouth the left sac acquires an opening to the exterior on the left side of the body (Fig. 64). The right sac becomes the preoral body-cavity or coelom of the “head,” while the left sac is known as the preoral pit. It is necessary to emphasise the fact that these two structures which are so different in their fully formed con- dition are at first perfectly 4 eA Lad i ; similar and symmetrical and form a pair of “head-cavi- Ultimately, as we have seen, only one of them ERCEOGEEG. BEEQOECED ties.” as statoto aa a Fig. 71. — Anterior portion of em- bryo, with thirteen primitive somites, actually persists as a head- cavity; namely, the right one. The entire conversion of from the ventral side in optical section. (After HATSCHEK.) r.a.d. and /.a.d. Right and left head- cavities. c.s.g¢. Rudiment of club-shaped gland. the left sac into the przoral pit is probably to be regarded as a secondary or cenoge- netic phenomenon, but the acquirement of an opening to. the exterior is probably not secondary, since a similar opening (the proboscis-pore) occurs in Balanoglossus. In addition to the above-described peculiarities which sufficiently distinguish the head-cavities from the myocce- lomic pouches, must be mentioned the fact that at no point of their epithelial walls are muscles developed. It is probable that the praeoral head-cavities of Amphi- oxus are homologous with the premandibular cavities of the higher Vertebrates, from the walls of which the greater number of the eye-muscles are developed.* This view is * This is also the opinion of Kupffer. Singularly enough van Wijhe has advanced the view that only the right head-cavity of Amphioxus is to be EMBRYONIC DEVELOPMENT. 129 strongly confirmed by the mode of development of the premandibular cavities in the Cyclostomes. In these fishes, VON Kupr- FER has shown that they actually appear in the form of a pair of diverticula from the anterior extremity of the archenteron (Fig. 72). If a comparison be made between Figs. 70 and 72, it will be at once manifest how close the correspondence is } us Fig. 72. — Horizontal projection of between the mode of de- pharynx and przoral endodermic exten- - sion of a young Ammocetes planeri of velopment of the head-cavi- 3% mm., reconstructed from a series of ties in Amphioxus and in transverse sections. (After KUPFFER.) p.e. Przeoral endodermic extension Ammoceetes. In the Se- (preorale Endodermtasche). gm. and m. leva st th iailacit -_ Praemandibular and mandibular portions achnlans € similarity 1S of head-cavities. ph. Cavity of pharynx. hardly less strikine.® 7, 2, 3. First three pairs of gill-pouches. 2 N.B.— Kupffer considers it probable that the mandibular as well as the pre- . mandibular cavities arise from the single Endostyle and Pigment pair of endodermic diverticula. In the Granules. course of the following pages I have referred chiefly to the praemandibular In Fig. 64 there is to be cavities alone so as to avoid complica- tions. noticed a vertically placed structure lying in front of and contiguous with the club- shaped gland. It is a tract of very high cylindrical cells forming part of the right wall of the alimentary canal in homologised with the preemandibular cavity (see below, V.). Kupffer regards the premandibular and mandibular head-cavities as rudimentary or meta- morphosed gill-pouches. This deduction is entirely foreign to the standpoint which I have adopted. The conclusion may seem plausible from the con- ditions observed in Acipenser alone; but when these are regarded from a comparative point of view, the deduction is, to my mind, unjustified. It should be added that Kupffer has shown that the head-cavities (premandibular and mandibular) of Acipenser also arise as endodermic pouches. 130 DEVELOPMENT OF AMPHIOXUS. this region. (Cf. Figs. 65 Gand 75.) I have shown that this epithelial tract is the rudiment of the exdostyle (vide anpra). It is a curious fact that the first trace of pigment to appear in the nerve-tube is not the eye-spot, but that at a constant point in the region of the fifth somite a black pigment-spot is deposited in a cell in the ventral wall of the medullary tube. This is followed by another smaller pigment granule slightly posterior to the first (Fig. 63 Cy The eye-spot appears at the end of the embryonic period. LARVAL DEVELOPMENT. Formation of Primary Gill-slits, etc. With the establishment of the definite relations oi the head-cavities, the mouth, club-shaped gland, first gill-slit, and anus, the embryo enters upon the larval phase of the development. It is no longer, or only very rarely, to be taken from the surface of the sea, but descends to a depth of several fathoms. When kept in aquaria, the larve can often be observed to be suspended vertically, and apparently quite motionless in the water. This suspension is, no doubt, effected by the movement of the long cilia, or flagella, with which the ectoderm is provided, each cell possessing one flagellum.® The principal changes which take place during the early stages of this phase of the development are the addition of new myotomes, the formation of new gill-slits, in meta- meric order, in an unpaired series on the right side of the larva, to the number of from twelve to fifteen, or even sixteen (the more usual number being fourteen), and the origin of the atrial cavity. LARVAL DEVELOPMENT. 131 Each gill-slit breaks through in, or slightly to the right of, the mid-ventral line, and then grows well up on the right side of the body. A larva with three gill-slits and the indication of a fourth is represented in Fig. 73. The originally circular mouth has grown to a much larger size, and extends on the left side anterior to the endostylar Fig. 73. — Larva of Amphioxus, with three gill-slits and the rudiment of a fourth; from the left side. (After LANKESTER and WILLEY.) p.p. Preeoral pit. ed. Endostyle lying on right side, seen through the wide lateral mouth. g/. Position of external aperture of club-shaped gland. 4.s’. First primary gill-slit. az. Anus. N.B.— Actual length of larva, nearly 1% mm. tract (which is on the right wall of the pharynx) and posterior to the first gill-slit. The oral opening later attains to relatively gigantic dimensions, and forms one of the most striking features of the larva. The anus is now displaced from its original ventral position to the left side in consequence of the increased development of the provisional caudal fin. The latter consists of elongated ectodermal cells, in which a certain amount of brown pigment is deposited. Later, when the dermal expansion, which has been described above as the definitive caudal fin, begins to grow out, it pushes the cells composing the provisional fin before it, so that they form a fringe round its border. Eventually the provisional fin disappears entirely. The gill-slits now go on adding to their number, one after the other, until the larva reaches the stage shown in Fig. 74. In this larva there are fourteen primary unpaired gill-slits, lying, for the most part, on the right side of the 132 DEVELOPMENT OF AMPHIOXUS. pharynx, although the more posterior slits bend under the pharynx, while the most posterior have a median ventral position. In front the gill-slits still open directly to the exterior, but the right metapleural fold is seen to be hanging over the tops of them; while the hinder slits now open into the partially formed atrium, which has already closed in . psi pss Se Fig. '74.— Anterior portion of larva, with fourteen primary gill-slits and rudi- ments of the secondary gill-slits; viewed as a transparent object from the right side. (After WILLEY.) 5.0. Sense-organ of przoral pit (groove of Hatschek). e. Endostyle. 7. In- ternal opening of club-shaped gland. s,s. Rudiments of secondary gill-slits. 7.518, p.si4, Thirteenth and fourteenth primary gill-slits. The lower margin of the mouth is seen through the anterior gill-slits. Total length of larva, nearly 342 mm. posteriorly, as described above. The larva is remarkably transparent, so that its internal organisation can be seen as clearly as possible through the outer body-wall. The long axis of the primary gill-slits is approximately at right angles to the long axis of the body. They are not more numerous than the myotomes in the correspond- ing region of the body, so that the drvanchiomery at this stage coincides with the muscular metamery. In Fig. 73 the first gill-slit was somewhat larger than the second and third. At about that stage, however, its further growth became arrested, and now it is seen to be considerably smaller than those which immediately follow it. In addition to its external opening on the left side, be- LARVAL DEVELOPMENT. 133 lat <3, SSS Sy a> maa OE) poe i L] Li Go w o rem \y Fig. 75. — Transverse sections through the region of the mouth of larve of Amphioxus, to show the endostyle and the external and internal openings of club- shaped gland. (After LANKESTER and WILLEY.) A. Section passing through the anterior corner of the mouth of a larva, with eleven gill-slits. &B. Section passing through the middle of the mouth of a larva, with twelve gill-slits. al, Pharyngeal cavity. 4.c. Coelom (splanchnoccel). 67.e. Branchial epithelium. e.a. Branchial artery. ezd. Endostyle. ex.o. External opening of club-shaped gland. fc. Dorsal fin-space. .g/. Lower portion of club-shaped gland. g.s’. First gill-slit. zz. Interccelic membrane. 7.0, Internal opening of club-shaped gland. Za. Left aorta; there is no corresponding right aorta in the larva. ». Mouth. 7m. Rudiment of right metapleur; a mere ectodermic thickening in 4; a solid thickening of the cutis in 2, in which two of the original enlarged ectoderm cells have become imbedded. These curious cells occur over a long stretch of the metapleural folds during this phase of the development, disappearing eventually. In B, the left metapleur is indicated by an ectodermic thickening immediately below the gill-slit. 2. So-called nephridium of Hatschek. 134 DEVELOPMENT OF AMPHIOXUS. low the mouth (see Fig. 64), the club-shaped gland has now acquired an opening at its upper extremity, on the right side, into the pharynx.’ The gland lies, as usual, behind, and closely approximated to, the endostylar tract, which is bent forwards on itself at its upper end (Figs. 75 A and B&). Pigment-spots are present in great numbers at the base of the neural canal. The pigment is deposited in special vn Pro er TI A i HT NMA » > hs U ATr7Tz A.r77z Fig. '76.— Transverse sections through the region of the przeoral pit. (After LANKESTER and WILLEY.) A, Through a larva, with twelve gill-slits and no atrium. 4. Through a larva, in which the atrium was closed in over all the gill-slits except the first two. (Cf. Fig. 38 C.) arm. Anterior median portion of right metapleur. .0.c. Praeoral body-cavity (right head-cavity) ; this cavity becomes much reduced after the metamorphosis, and is largely filled up by gelatinous tissue. #.%. Praeoral pit. s.0. Sense-organ of preeoral pit (groove of Hatschek). 20.4. Rudiment of left half of oral hood. my'. Sclerotome (diverticulum of myoccel my). Other letters as above. Section 4 is taken through a plane slightly posterior to section A. LARVAL DEVELOPMENT. 135 cells, the pzgment-cells, which arise as modified epithelial cells of the central canal. These cells send out several branching processes, which lose themselves in the fibrous tract of the spinal cord. Already in the youngest larva — namely, that shown in Fig. 64 — the preoral pit had become subdivided into two portions, which, however, retained a free communication with one another. In the course of the changes which the left head-cavity had to undergo in its conversion into the przoral pit it had come to lie transversely below the notochord. Sub- sequently it extended itself, in the form of an offshoot, dorsally to the right of the base of the notochord. This offshoot from the przeoral pit appears to serve as a special sense-organ lying ultimately, as mentioned above, in the roof of the oral hood, whose function is possibly to test the water as it enters the mouth (Figs. 76 A and 3B, amde big 745 (ete. ). Formation of Secondary Gull-slits. Above the primary gill-slits in Fig. 74, and like them, on the right side of the body, is to be observed a longitudinal ridge provided with a series of nodal enlargements which alternate with the primary gill-openings, the first of them lying above and between the third and fourth primary slits. Each of these enlargements represents a thickening in the wall of the pharynx, which has undergone fusion with the bedy-wall beneath the right metapleural fold, in the angle formed by the latter with the body-wall. These metameric fusions of the pharyngeal wall with the body-wall are the forecast of a second row of gill-slits, whose relation to the primary row will become clear as we pro- 136 DEVELOPMENT OF AMPHIOXUS. ceed. With their appearance, the larva enters upon that phase of its development which has been called the later larval period. It is the period of the metamorphosis of the larva, during which the pronounced asymmetrical arrangement of the parts is exchanged for the partial, but not absolute, symmetry which we have noted in the adult. The metamorphosis, therefore, consists largely in the sym- metrisation of the larva. The simultaneous appearance of the six nodal thicken- ings in the exact position, shown in Fig. 74, is very constant. Shortly afterwards a minute perforation appears in the centre of each thickening almost simultaneously, except in the case of the first, which usually becomes perforated rather later than the others. The originally small circular openings of the secondary gill-clefts gradually increase in size and become oval in shape, their long axes being parallel to the long axis of the body, instead of at right angles to it as in the case of the primary slits. Next, the upper borders of the secondary slits begin to flatten, and later to show signs of curving downwards. The changes in shape, which affect the secondary slits at the stages now under consideration, may be expressed by saying that they are at first shaped like a biconvex lens, then like a plano-convex lens with the flat surface directed upwards and the convex surface downwards, and finally like a concavo-convex lens with the concavity directed upwards (Fig. 77). During these changes, which do not take place in all the secondary slits at the same time, the last one especially retaining for a long time its primitive shape, the walls of the successive slits become sharply rounded off and distinct from one another, and anew perforation makes its appear- ance in front, above, and between the second and third LARVAL DEVELOPMENT. 137 primary slits. This new slit constitutes the definitive first slit of the secondary series (Fig. 77). The larva shown in Fig. 77 presents a very different aspect from that shown in Fig. 74; the transition from one stage to the other is, of course, gradual, and all intermediate steps can be observed. In the stage which we are now considering (Fig. 77), the atrial cavity has become com- pletely closed up in front, so that now none of the gill-slits open directly to the exterior. None of the primary slits now he entirely on the right side, but they have become bent under the pharynx, and EPH ee - ee _ e Ss . : - YAS SS > / <0 psr SST 0G eae Vv | : gl 4 psi .© Fig. 77.— Anterior portion of larva, in which the secondary slits have become perforated, and the primary slits are passing round to the left side. From the right side. (After WILLEY.) 5.0. Sense-organ of przoral pit. v. Right half of velum. e. Endostyle, grow- ing beyond the club-shaped gland g/. g.s5’. First primary slit, much reduced in size. 5.5’. First secondary slit. 7.512, Twelfth primary slit, behind which is to be seen a vestige of the thirteenth slit. t i a thus extend round to the left side. This bodily migration of the primary slits from one side to the other occurs in correlation with the increase in size of the secondary slits, which, as they continue to grow, push, as it were, the primary slits before them, and so cause the latter to bend under the pharynx in the way described. The peculiar growth by which the primary gill-slits are gradually carried from the right to the left side, may be described as a trans- verse or rotatory growth affecting the pharynx 7 ¢ofo in 138 DEVELOPMENT OF AMPHIOXUS. the region of the secondary slits. Such of the primary slits as occur behind this region are not affected by the rotatory method of growth, and retain their original position in the mid-ventral line of the pharynx. It is to be noted also that there are only twelve primary gill-slits at this stage. Assuming that in the particular larva here figured there were originally fourteen primary slits, the fourteenth has closed up and vanished without leaving a trace, while a vestige of the thirteenth can still be recognised. The actual process involved in the closure and disappearance of a certain number of the primary slits can, as we shall see, be readily observed in the living larva. Club-shaped Gland and Endostyle. The internal aperture of the club-shaped gland into the pharynx is exceptionally plain at this stage, and its refring- ent walls and relatively large size give it a curiously slit- like appearance. We shall find that the gland subsequently atrophies, but the most persistent part of it — that is to say, the last part of it to disappear —is precisely the internal opening with its refringent border. The endostyle, whose primary position, as we have seen, was immediately in front of the club-shaped gland, now presents a remarkable condition. It has begun to grow backwards and downwards, being probably pulled down, so to speak, by the general rotatory growth of which we have spoken above; and so the club-shaped gland no longer lies behind it, but upon it. The gland itself being disconnected with the wall of the pharynx, except at its upper end where it opens into the latter, is not affected by the complicated changes to which the pharyngeal wall, including gill-slits, mouth, and endostyle, is subjected, so LARVAL DEVELOPMENT. 139 that it forms a convenient punctum fixum with relation to which the growth of neighbouring structures, particularly that of the endostyle, can be determined. The upper and lower limbs of the endostyle are inclined to one another at an acute angle, and may be said to form two unequal sides of a triangle, the apex of which is directed backwards between the rows of secondary and the primary gill-clefts (Fig. 77). Between the two rows of slits on the right side of the body there is a blood-vessel, representing the anterior continuation of the sub-intestimal vessel, which ends blindly in front above the first primary slit. This is the future ventral branchial artery, with which we are already ac- quainted. When its final situation in the mid-ventral line below the endostyle is remembered, its position in the larva high up on the right side, as in Fig. 74, will appear very striking. Continued Migration of Primary Gill-slits. The secondary slits now go on growing in size, and the primary slits gradually tend to disappear entirely from the right side until, as in Fig. 78, only the original upper por- e tn 705° Fig. 78. — Anterior portion of larva from right side, to show the backward growth of the endostyle between the primary and secondary gill-slits. (After WILLEY.) s.o. Sense-organ of preoral pit. #.s'. First primary slit. 7. Internal opening of club-shaped gland. e. Endostyle. 7.4, Peripharyngeal ciliated band. 140 DEVELOPMENT OF AMPHIOXUS. tions of them are visible from this side. In some of the secondary slits the dorsal margin, which had previously begun to curve downwards, has now reached the ventral margin and fused with it (Fig. 78, third secondary slit). In this way is the tongue-bar formed, and the primitively simple gill-opening is divided into two distinct halves. The formation of the tongue-bars occurs in the secondary slits considerably in advance of the primary, both actually and relatively, since the latter have existed all through the earlier period of the larval development without a trace of tongue-bars. Peripharyngeal Bands. The endostyle has now grown a long distance behind the club-shaped gland, and extends backwards between the two rows of slits as far as the middle of the second secondary slit. From the anterior part of the upper half of the endostyle, which is now nearly equal in length to the lower half, arises an epithelial tract in the wall of the pharynx, which appears in the form of a band of ciliated cells, and proceeds backwards below the notochord to the end of the pharynx. A corresponding ciliated band occurs in the left wall of the pharynx, proceeding from a similar point in the lower limb of the endostyle. In their course below the notochord the two bands take part in forming the hyperpharyngeal (dorsal) groove of the pharynx. Atrophy of First Primary Gill-sit and Club-shaped Gland, etc. We have already seen indications of a reduction in the size of the first primary slit. This reduction has advanced considerably in the stage we are now describing (Fig. 78), where the slit in question is only recognisable in side view as a small groove, LARVAL DEVELOPMENT. I4I The next stage to be considered is characterised above all by the simultaneous atrophy, closure, and disappearance of the club-shapea gland, and the first primary gill-slit (Fig. 70). At this stage the mmerease in size of the secondary slits has progressed to such an extent that the primary slits have been displaced entirely from their original position, and are no longer to be seen from the Fig. 79.— Anterior portion of larva from right side after the disappearance of the club-shaped gland. (After WILLEY.) s.o. Sense-organ. e. Endostyle. 7.6. Peripharyngeal band. s,s’, First secondary slit. right side, except in the case of the hindermost slits of the series, which remain, as mentioned above, in a median ventral position until their disappearance. A larva seen from below, so as to show the relative positions of the gill-slits and endostyle, etc., at this stage, is represented in Fig. 80. It is obvious, from what has been said above, that in the passage of the primary slits from their original position on the right side of the body to their final position on the left side, their dorsal and ventral margins are reversed. What was at first the dorsal edge of a primary slit becomes its ventral edge, and vice versa. In other words, what is actually the dorsal border of the primary slits in Fig. 74 is morphologically the ventral border ; and conversely, what is actually the latter is morphologically the former ; and it is 142 DEVELOPMENT OF AMPHIOXUS. from the latter, towards the completion of the rotatory growth, which carries the slits from one side to the other, that the tongue-bars arise (Fig. 80). The vertical and longitudinal axes of most of the slits, both primary and secondary, are now almost equal, but the original difference in this respect, which we noted above, is still to be observed in the case of the foremost and hindmost slits of the two series. (Cf. Fig. 80, s.st and g.s*, and s.s§ and fs.) The first primary slit has 5 7 2 : : i ois pis p.s® Saree La es eee ad Revs: z eis : BBL eens 2h SE SSS as Fig. 80.— Anterior portion of larva of same age as in Fig. 79, seen from the ventral surface. The pharynx is flattened out. (After WILLEY.) ch. Notochord. mm, Entrance to mouth. v. Velum. 7.51. Vestige of first primary slit. 4.52. Secondary primary slit. .s!9, Tenth primary slit. 7.512. Ves- tige of twelfth primary slit. s.sl. First secondary slit. e. Endostyle. s.s. Eighth secondary slit. a. Atrium, pressed aside. now completely closed up, and its former existence is barely indicated by a loose granular appearance at the place it formerly occupied. The alternation of the gill-slits of the two series comes out very clearly in Fig. 80. In most of the secondary slits the formation of the tongue-bars is completed; but not so in any of the primary slits, where it is only be- ginning. There are now eight secondary slits, an additional one having been added behind, alternating with the ninth and tenth primary slits. Usually the formation of secondary slits stops at this point, no more being formed until the LARVAL DEVELOPMENT. 143 number of primary slits is reduced to the same number ; namely, eight. Since it is usual for the primary slits to break through in the first instance to the number of fourteen, no less than six of them must close up and disappear before the stage with only eight gill-slits on each side of the body is arrived at. The six slits which are to close include the first and the five posterior primary slits. In the larva shown in Fig. 80, the tenth and eleventh primary slits would have to close at a later stage ; the twelfth is on the point of closure, and its walls present the characteristic coarsely granular appearance spoken of above, while the thirteenth and fourteenth slits have entirely vanished. In addition to the fact of the closure of these primary slits, it is important also to emphasise the fact that they disappear without leaving a trace behind. In the higher Vertebrates there are a number of structures not only di- rectly connected at some stage of development with the pharyngeal wall, but also at some distance removed from it, which various morphologists have interpreted as the remnants of ancestral gill-clefts, without sufficiently con- sidering the question whether gill-clefts were in the habit of leaving their mark behind them. In Amphioxus, at all events, they do not. The Adjustment of the Mouth, etc. While the gill-slits have been adjusting themselves to their definitive positions, the mouth has also been sub- jected to a peculiar kind of growth, which results in its bending round the front end of the pharyngeal wall, and ultimately assuming an anterior and median position, as we find it in the adult. 144 DEVELOPMENT OF AMPHIOXUS. In Fig. 81, a larva corresponding in age approximately to that of Fig. 74 is represented as seen from the left side. As noted above, the posterior primary slits bend nor- mally under the pharynx at this stage, and some of them extend as much on one side of the body as on the other, being continued across the ventral side of the pharynx. The great feature of this larva is the relatively prodigious mouth, through which the upper portions of the first four primary slits can be seen. From this side we look into the depths of the przoral pit, having only seen it by transparency in the preceding olf i wt he { ul ? t 7b rhe Fig. 81.— Anterior portion of larva, with thirteen gill-slits, from the left side. (After WILLEY.) olf. Olfactory pit, communicating with neuropore. +x. ‘“‘ Nephridium” of Hat- schek. 2.¢. Spinal cord. ch. Notochord. 7.f. Przeoral pit. ex. External open- ing of club-shaped gland. cz. Rudiment of buccal cirri. 7.6. Peripharyngeal band. m. Mouth. 7.518. Thirteenth primary slit. i je CL figures. It is continued backwards into a ciliated groove, which abuts on the dorsal margin of the mouth. Prob- ably most of the food which enters the mouth passes along this groove. Below the pointed anterior extremity of the mouth is to be seen the external aperture of the club-shaped gland, and a short distance behind this is a round, refringent body, which has become differentiated from the gelatinous LARVAL DEVELOPMENT. 145 connective tissue lying below the epidermis, and repre- sents the rudiment of the first element of the cartilagi- nous skeleton of the buccal cirri. Running parallel with the lower margin of the mouth, and curving gently upwards to the dorsal wall of the pharynx, is a ciliated band proceeding from the lower limb of the endostyle, and corresponding to the one on the other side, which we found in connexion with the upper portion of the endostyle. Its course on the left side is somewhat different anteriorly from that of the right side, owing to the position and size of the mouth. (Cf. Figs. 78 and 81.) The so-called olfactory pit, which arose at a much earlier stage as an ectodermic depression above and in connexion with the neuropore, no longer lies in the mid-dorsal line as in Fig. 64, but it has been displaced to the left side by the upgrowth of the dorsal fin (Fig. 81). Here, as in the case of the anus, the development of a median fin has no other effect on the aperture in question than to cause it to forsake its primitively median and symmetrical position and to assume an asymmetrical position on the left side of the body. This is important to bear in mind, as the asym- metrical position of the mouth will be explained below on an analogous basis. For the present it is sufficient to call attention to the fact that, with the exception of the gill-slits, whose primary unpaired character is due to the retarded or /atent develop- ment of their antimeres, the unpaired openings in the body-wall—namely, neuropore, preoral pit, external aper- ture of club-shaped gland, mouth, and anus —all lie on the left side of the body. At a slightly later stage than the preceding, the front end of the mouth is found to be no longer pointed, but to have become rounded off, and, moreover, to lie at a deeper 146 DEVELOPMENT OF AMPHIOXUS. level than previously (Fig. 82). The posterior groove of the przeoral pit which we described in the last stage, seems to be preparing the way for the mouth to dip inwards towards the right wall of the pharynx, which, in fact, it has actually begun to do. At a still later stage, corresponding to that shown in Fig. 77, the shape of the mouth has become entirely altered (Fig. 83). It has now the form of a triangle with the apex directed backwards and the base standing vertically in front. But the apex and the base are not in the same tangential plane, Ki . x ‘ » £ , { M 5 s ‘ Fig Cx CA%cz in fre Fig. 82.— Anterior portion of larva somewhat older than preceding, to show commencing adjustment of the mouth. (After WILLEY.) e. Endostyle seen through the mouth. Other letters as above. the former being on the left side of the body, and the latter much deeper inwards; in fact, just below the skin on the right side of the body. (Cf. Fig. 77.) We see, therefore, that the longitudinal diameter of the larval mouth is gradually shortening. It is eventually reduced to zero when the right and left sides of the mouth or velum come to lie opposite to one another, the velum ultimately attaining a circular form and a median sub- vertical position underneath the oral hood. When the larva has reached the age to which Fig. 11 refers, the right LARVAL DEVELOPMENT. 147 half of the velum is nearly but not even yet quite opposite to the left half (Fig. 93). In the preceding stage (Fig. 82) there were several additional buccal cartilages added to the first one which we described. In the present stage these have begun to grow outwards so as to produce small notches in the integument, which is now commencing at this point to form the right half of the oral hood. The left half of the latter arises as a downgrowth of the integument from the upper margin of the przeoral pit and its posterior continua- tion, the above-mentioned ciliated groove. (Cf. Figs. 81, 82, and 83.) The hinder portion of this fold is at first on Fig. 83. — Anterior portion of still older larva, from the left side, to show change in shape and position of the mouth. (After WILLEY.) Letters as above. The left half of the oral hood is now growing down over the mouth and preeoral pit. a level with the dorsal margin of the mouth, and in fact merges into the latter, but subsequently grows over it, extending to its posterior extremity, where it meets the right half of the oral hood. It is obvious from the above description and figures that a large part of the right wall of the oral hood is derived from the original wall of the snout below the preoral pit, and so an explanation is afforded of the fact noted in the first chapter that the right half of the oral hood is continu- ous round the anterior extremity of the notochord with the cephalic expansion of the dorsal fin.9 148 DEVELOPMENT OF AMPHIOXUS. The przoral pit itself is absorbed, as it were, into the oral hood, so that it eventually loses its independent exist- ence as a pit, although the sense-organ of the przoral pit persists in the adult as a deep groove in the dorsal wall of the oral hood to the right of the base of the notochord. The remaining ciliated epithelium of the original przoral pit increases in extent, and grows out into the finger- shaped tracts which we have already described as being characteristic of the inner surface of the oral hood, consti- tuting the so-called “ Raderogan.” (Cf. Fig. 3.) Equalisation of the Gill-stits. In the stage next succeeding that of which a ventral view is given in Fig. 80, the first eight primary slits—that is to say, from the original second to the ninth inclusive — Fig. 84.— Larva toward the close of the metamorphosis, from the left side. (After WILLEY.) o. Olfactory pit. v.Velum. 7.4. Peripharyngeal band. e. Endostyle. 9.52. Second primary slit, the first having closed up. m. Left metapleur. s.a. Floor of atrium. p.s!2, 7.518, Vestiges of the twelfth and thirteenth primary slits. have become definitely established on the /ef¢ side, their longitudinal and vertical axes are equalised, and in most of them the tongue-bars are completely formed (Fig. 84). No tongue-bar is formed in the first slit on either side, and this slit apparently remains as a rule simple throughout life. LARVAL DEVELOPMENT. 149 In Fig. 84 the last indications of the twelfth and thir- teenth primary slits are to be observed as slight depres- sions in the floor of the pharynx in the mid-ventral line. The tenth and eleventh slits would close up later. It should be pointed out that the closure of the poste- rior primary slits does not proceed in perfect correspond- ence with the age of the larva, but takes place sometimes at an earlier and sometimes at a later stage than here depicted. The gill-slits of both sides now begin to elongate in the vertical direction (Fig. 93), and eventually a very well- marked stage is reached, which is characterised by the presence of eight pairs of gill-clefts. This latter stage would appear to have a considerable duration, and, as it stands on the borderland between the larva and the adult, and forms the commencement of what may be called the adolescent period of the development, it may well be regarded as a critical stage. By this time the young Amphioxus has given up its free pelagic life in the open sea, and has commenced to burrow in the sand, which it continues to do for the rest of its life.* Further Growth of Endostyle, etc. At the point at which we left the endostyle, its two halves were in the relation to one another of upper and lower. During the steps in the metamorphosis which we have recorded above, the upper half of the endostyle is brought down to the same level as the lower half on the right side of it, and so the definite form of the endostyle is established by the conjunction of its right and left halves. It then proceeds to grow backwards along the * The duration of the larval development of Amphioxus may be estimated at about three months. 150 DEVELOPMENT OF AMPHIOXUS. base of the pharynx between the two rows of gilt-slits, but does not reach the posterior end of the pharynx until a much later period.” : The features in the development of the endostyle which ought to be especially emphasised are, firstly, its direc- tion of growth from before backwards, and secondly, its primary anterior position in the wall of the pharynx in front of all the gill-slits. In connexion with the modification in the shape and position of the mouth, as described above, it is important to insist on the fact that the mouth of the larva is directly converted into the velum of the adult, while the oral hood which grows over the mouth is a new formation. During the period of the metamorphosis the larva does not increase in length. It is rather a readjustment of parts which is then taking place than an increase in bulk which is the symbol of active growth. From the time of the first indication of the secondary slits (Fig. 74) till after the completion of the passage of the primary slits from the right to the left side of the body, the average length of the larva may be taken as approximately 3.5 mm. The adolescent period is essentially the period of active growth in bulk and maturity. The increase in length during this period does not, however, depend on the addition of new myotomes to those already formed, but merely on the progressive growth in size of the latter. The full complement of myotomes was developed during the early larval period, and is present in the larva repre- sented in Pig. 74: LARVAL DEVELOPMENT. I5I Development of Reproductive Organs. One of the most interesting events which we have now to chronicle is the development of the reproductive organs. This commences when the young Amphioxus has reached the length of about 5 mm. Our knowledge of the details of the processes involved in the formation of the genital organs is again due to the work of BovERI, who has made the discovery that the Fig. 85. — Transverse section through the pharyngeal region of a young individual of 5 mm., to show place of origin of sexual elements. (After BOVERI.) f Fascia. e.c. Portion of ccelom, which will form the endostylar ccelom, ug. Primitive sexual cells in the lower angle of the myoccel. Other letters as above. primitive sexual cells arise in the cavity of the myotome by differentiation of certain of the epithelial cells lining the myoccel. It had previously been assumed that they were derivatives 152 DEVELOPMENT OF AMPHIOXUS. of the peritoneal epithelium lining the general body-cavity. The fact that they arise in the way shown by Boveri is one of great morphological importance. In a transverse section of a young individual 5 mm. in length, the primitive sexual cells are to be recognised as a closely packed group of cells, with large nuclei in the lower angle of the myotome ; that is, in the angle formed by the membrane which divides the myoccel from the splanchnoceel, which we may call the zxztercelic membrane, with the cutis (Fig. 85). Since the myotomes of one side alternate with those of the other, so do the centres of Fig. 86.— Longitudinal views of the developing gonads, obtained by dissecting out the ventral borders of the myotomes. (After BOVERI.) u.g. Primitive sexual cells arising from the myoccelic epithelium; the nuclei scattered about the surface of the preparations also belong to the myoccelic epithelium. formation of the primitive sexual cells, and in a given section, as in Fig. 85, only one such centre is to be observed on the right or left side of the section, as the case may be. Its actual position in the longitudinal aspect of the myo- tome is shown in Fig. 86 A, B, and C. The formative centres of the primitive sexual cells lie at first in the angle mentioned above, but applied to the posterior faces of the dissepiments between the myotomes (Fig. 86 A). At a somewhat later stage, having slightly increased in bulk, they begin to push the dissepiments before them LARVAL DEVELOPMENT. 153 so as to make a projection into the myoceel in front (Fig. 86 42, C). This projection of the primitive gonad into the myoccel next in front of that to which it originally belonged, is gradually carried to such an extent that the gonad becomes entirely shut off from its original myoccel and hangs freely into the next one, being connected by a short stalk with the azterior face — Fig. 87.— Similar prepara- f : tion as the preceding, showing of the dissepiment and surrounded 4 jater stage in the development of the primitive gonad. (After by a membrane which is obviously Bove derived from, and for some time continuous with, the original dissepiment (Fig. 87). In correlation with the increase in size of the primitive gonad, an evagination of the basal wall of the myoccel in which it now lies, takes place, and by the time the young Amphi- Fig. 88. — Preparation showing the rhomboidal pouches of the myoccel which project into the atrial cavity. (After BOVERI.) This condition is found in individuals of 13-14 mm. oxus has attained a length of 13 or 14 mm. there is, in connexion with each primitive gonad, a wide rhomboidal expansion of the lower portion of each corresponding myoccel projecting into the atrial cavity (Fig. 88). The cavity of these sacs, to the wall of which the gonads are at this stage still united by a stalk, constitutes the so- called perigonadial celom,4 or cavity of the gonadic pouches, which, at the time of sexual maturity, is entirely filled up by the sexual elements. 154 DEVELOPMENT OF AMPHIOXUS. The gonadic pouches next become gradually constricted off from the myoccelic spaces, and eventually lose all com- munication with them. In the midst of the at first solid mass of primitive sexual cells a cavity subsequently appears, and the gonad be- comes a hollow sac (Fig. 89). In the course of its fur- ther growth the gonadic sac (not to be confused with the gonadic pouch in which it lies) grows out into a num- Fig. 89.— Portion of transverse sec- tion through an individual of 13 mm., ber of lappets, and so_be- to explain the conditions observed in comes a racemose reproduc- preceding preparation. (After BOVERI.) é.v. Blood-vessel. go. Gonadic sac. tive eland (Langerhans). pouch). tm Tramvene muscles, ‘The He primitive sexual cells index line to which there is no letter remain for a considerable indicates the fold by which the gonadic pouch becomes constricted off from the length of time in an abso- ee lutely indifferent condition, and it is impossible to distinguish the male from the female. According to LANGERHANS, sexual differentiation does not begin to take place until the individuals have reached a length of 17 mm., and sometimes it does not occur until a much later period. It is inaugurated by the commence- ment of the processes of spermatogenesis and ovogenesis. There are no accessory sexual characters in Amphioxus, and the sex can only be determined by an examination of the reproductive glands. The segmental arrangement of the formative centres of the reproductive organs at the base of the myotomes is again met with in the embryonic development of the Selachians, as shown by RuckeErt (Fig. 90). Here, also, LARVAL DEVELOPMENT. 155 the primitive sexual cells make their first appearance in the segmented area of the trunk at the base of the somites. Later on, by differential growth, they come to lie on the dorsal wall of the unsegmented peritoneal cavity, and their primitive segmental origin is entirely obscured ; while in Amphioxus the primitive segmentation of the gonads is maintained throughout lite. Fig. 90. — Horizontal section through This forms another most the ventral portion of six consecutive h , mesodermic somites of an embryo of interesting example of the pristiurus, to show the segmental origin the adult of the sexual elements. (After RUCKERT). ) c. Cavities of somites. g.c. Sexual Amphioxus, in the details cells. : : : This observation of Riickert’s has of its organisation, €SSe€Nn- recently been doubted, with how much : a ] th _ justice it is difficult to say, by MINOT tially resembles mite (Gegen das Gonotom. Anat. Anz. IX. © bryos of the higher types. 1894. pp. 210-213). way in which GENERAL CONSIDERATIONS. We will now pass on to give a general interpretation of some of the principal phenomena which are presented to us in the development of Amphioxus. Larval Asymmetry. By far the most prominent feature of the fully formed larva is its astounding asymmetry, and it is extremely important, from a morphological point of view, to form a just conception of it. I 56 DEVELOPMENT OF AMPHIOXUS. The phenomenon of asymmetry manifests itself in the larva of Amphioxus under several very different aspects, and is occasioned by various causes. For convenience we may classify the forms of asymmetry which we have to consider under three main divisions, according to the type of organs involved. 1. Median Asymmetry. — This relates to such structures as lie normally in the middle line, whether dorsal or ven- tral, but which have been mechanically or correlatively dis- placed from their primitive position by the differential growth of neighbouring parts. Such are the olfactory pit and neuropore, the anus, the mouth, and the endostyle. All these are essentially and primordially median and unpaired structures. We have already dealt with the neuropore and anus, while the mouth and endostyle will be con- sidered below. 2. Bilateral Asymmetry. — This refers to the alternation of paired structures, such as myotomes, spinal nerves, gill- slits, and gonads, which we have already noted in the adult organisation. Primarily the organ of one side lies opposite to its antimere of the other side. By a secondary displace- ment it comes to alternate with it.* 3. Unilateral Asymmetry. — Next to the asymmetrical mouth, this is perhaps the most striking form of asym- metry which the larva of Amphioxus exhibits. It relates to those structures which belong to the category of paired organs, but which, in the course of the larval development, appear unpaired on one side of the body. Such are the * When the myoccelomic pouches first appear in the embryo they are placed symmetrically. At an early stage, however (see Fig. 63 B), the alter- nation sets in. This involves such later-appearing structures as the spinal nerves and gonads, so that they alternate from the time of their first origin. The alternation of the gill-slits would seem to be independent of that of the myotomes. LARVAL DEVELOPMENT. 157 gill-slits and the praeoral pit. As described in the fore- going pages the asymmetry of the przoral pit is a second- ary occurrence, since it arises at first as one of a pair of symmetrically disposed head-cavities, or anterior intestinal diverticula, while the unilateral asymmetry of the gill-slits is ontogenetically primary. The unilateral gonads of the species of Amphioxus from the Bahamas and Torres Straits also belong to this category. Although, on account of their essentially azygous nature, the mouth and endostyle have been separated from the gill-slits in the above classification, it is obvious that their asymmetrical position in the larva must be ascribed to one and the same cause. In the succeeding pages we shall endeavour to demonstrate what this cause was. Explanation of Asymmetry of Mouth and Gill-shits. It is quite evident that the primary gill-slits which appear on the right side of the larva belong primitively, or ancestrally, to the left side, to which, in fact, they are eventually transferred. Meanwhile, the left side of the larval pharyngeal region is largely occupied by the huge oral aperture. We may figure to ourselves the primitively left-side gill- slits being carried over to the right side by a semi-rotation from left to right of the pharyngeal wall. The primitive right side of the pharynx would thus be crowded out, so to speak, and the right-side gill-slits would be temporarily obliterated owing to lack of room, while the original mid- ventral line would be carried high up on the right side, where, in point of fact, it is plainly indicated by the bran- chial artery, which lies actually above the primary gill-slits in the larva (Fig. 74, etc.). I 58 DEVELOPMENT OF AMPHIOXUS. Thus the actual topographical conditions in the larva do not by any means coincide with the morphological rela- tions of parts, since the morphological mid-ventral line of the pharynx lies high up on the right side of the body. It should be carefully noted that the form of asymmetry which we are now considering only affects the anterior portion of the larval body. The same semi-rotation of the pharyngeal region which converted the primitive left side of the larva into the actual right side caused the primitively median mouth to take up its position on the actual left side. But since, as we have noted, the rotation occurred from left to right, the mouth must have been originally situated in the median dorsal line. In postulating a virtual semi-rotation of the ancestral pharynx, we do not, of course, mean to suggest the prob- ability of an actual movement in bulk about the longi- tudinal axis, but merely that the formative centres of the various structures belonging to this region of the body (gill-slits, mouth, endostyle, etc.) have, by the correlated interaction of their component cell-groups, been diverted from their ancestral relations through the intercalation, in the course of the progressive evolution of the organism, of a new and disturbing element. We are now in a position to say what this disturbing element is. It is the secondary forward extension of the notochord beyond the limits of the dorsal nerve-tube to the tip of the snout. As already stated, there is direct evidence to show that this is a secondary and not an an- cestral feature, inasmuch as in the young embryo (Fig. 68 dzs) the notochord is removed from the anterior extrem- ity of the body by a very appreciable interval, which is oc- cupied by that portion of the archenteron which gives rise LARVAL DEVELOPMENT. 159 to the head-cavities. Moreover, as was pointed out above, the dorsal groove of the archenteron, which gives rise to the notochord, remains open into the archenteric cavity in the region of the first myotome, and even somewhat behind the level of the neuropore, for some time after its walls have approximated to form the solid notochord behind this region. The forward extension of the notochord in Amphioxus is, therefore, de facto, to a large extent an ontogenetic phenomenon, although, from the very beginning, it shows what may be described as a precocious tendency to extend beyond the nerve-tube. We shall also find that there is every reason to suppose that it is a cenogenetic, and not a palingenetic, feature. Since we know for an actual fact that the primary gill- slits of the larva belong ancestrally to the left side, it fol- lows as an absolute topographical necessity that the mouth has been brought to one side from an originally median dorsal position, by the same semi-rotation of the pharynx (in the sense explained above) which has demonstrably carried the primitive left-side gill-slits under the pharynx up to the right side of the larva. But this is not the only criterion by which we can judge of the ancestral position of the mouth. In the larvz of the Ascidians, the nearest existing rel- atives of Amphioxus, there is a praoral lobe and a neuro- pore, which opens at first to the exterior in the mid-dorsal line, just as in Amphioxus. But in contrast to the latter form the notochord does not extend forwards into the re- gion of the przoral lobe, but it stops short behind the cerebral vesicle. Immediately in front of the neuropore, in the Ascidian larva, the wall of the pharynx comes into contact with the 160 DEVELOPMENT OF AMPHIOXUS. ectoderm and fuses with it, and then at the point of fusion a perforation takes place, and the mouth is established in the mid-dorsal line. During the formation of the mouth the neuropore temporarily closes up, but subsequently it reopens — zzZo the mouth. In Amphioxus we can only assume that in correlation with the forward extension of the notochord, the mouth was compelled to forsake its primitive relations to the neuropore and to move to one side so as to make way for the notochord. The growth of the latter to the front end of the body obviously prevents the wall of the pharynx from coming into contact with the ectoderm in the mid- dorsal line, while it leaves the neuropore unaffected, since the nerve-tube is essentially dorsal to the notochord, and the pharynx, on the other hand, essentially ventral to it. This explains the fact that the hypophysis (olfactory pit) of Amphioxus opens dorsally directly to the exterior instead of into the mouth as it does in the Ascidian. The secondary gill-slits — that is, those belonging to the primitive right side of the body — present an interesting instance of retarded or latent development. This is due to the fact that their own side of the body is at first usurped by their primitive antimeres, the so-called primary slits, as a result of which they have themselves been temporarily crowded out as mentioned above. In con- sequence of their retardation, when they do appear to inaugurate the process of symmetrisation, they do not conform to the method in which metameric structures are normally produced, but most of them —namely, from the second to the seventh inclusive—arise simultaneously while the first and the eighth arise somewhat later. LARVAL DEVELOPMENT. I6I Larval Asymmetry not Adaptive and not Advantageous ; Forward Extension of Notochord Adaptive and Advan- lageous. The conclusion to be drawn from the above considera- tions is that the remarkable asymmetry of the larva of Amphioxus, in respect of the pharynx and the parts con- nected with it, is of no specific advantage whatever to the larva, but is merely a stage, which has been preserved in the ontogeny, of a topographical readjustment of parts necessitated by the removal of the mouth from its primi- tive mid-dorsal position in consequence of the secondary forward extension of the notochord, which has thus caused a virtual semi-rotation of the pharyngeal region of the body. On the other hand, the forward extension of the notochord is a distinct advantage in later life, since, by giving resistancy to the snout, it enables the animal to burrow its way into the sand with such astonishing facility, while the fact that it grows to the front end of the body at a very early stage in the embryonic development, long before it comes to be put to this definite use, must be regarded as an instance of precoczous development of which there are numerous and otherwise inexplicable examples in the field of comparative embryology. The larval asymmetry of Amphioxus is therefore a purely secondary or cenogenetic feature, and has no directly ances- tral or palingenetic significance, although, as we have shown above, it serves indirectly as a clue to what the ancestral condition was. At the same time it is a primary feature in the actual ontogeny; that is to say, the asymmetrical structures (mouth and gill-slits) arise zz sztw, and are not removed in the individual development from a primary 162 DEVELOPMENT OF AMPHIOXUS. symmetrical to a secondary asymmetrical position, as is the case, for instance, with the neuropore. It may appear paradoxical, but is nevertheless correct, to say that in the ontogeny the mouth and gill-slits appear primarily in a secondary position. It is quite evident that the asymmetry of the larva of Amphioxus is of a totally different character to the well- known asymmetry of the flat-fishes or Pleuronectide (turbot, sole, plaice, halibut, flounder, etc.). The latter are hatched as perfectly symmetrical larve with eyes quite opposite to one another. Then, in adaptation toa life at the bottom of the sea, after a short pelagic existence they turn over on one side, in some species the right side, and in others the left, and the eye of that side moves over the snout, sometimes even through the snout, to the other side, and so the eyes come to lie on the same side. In this case, therefore, the asymmetry, which is secondary in every sense of the word, is the result of a special adaptation to a particular habit of life, and is accordingly of the greatest advantage to the fishes which possess it. On the other hand, its extraordinary asymmetry is of no conceivable advantage to the larva of Amphioxus, and does not represent an adaptation to any peculiar mode of existence whatever. It is rather the mechanical, incidental, accessory, and subsidiary accompaniment of another organic change which is both advantageous and adaptive, namely, the forward extension of the notochord; and while the excessive asym- metry is indifferent to the pelagic larva, it would be posi- tively detrimental to the adult. Thus in all respects the larval asymmetry of Amphioxus is the precise converse of the adult asymmetry of the Pleuronectidee.® AMPHIOXUS AND AMMOCGTES. I 63 AMPHIOXUS AND AMMOCCETES. We will now pass on to consider what new light the larval development of Amphioxus throws on its relation- ship to the craniate Vertebrates. As a type of the latter with which to make the com- parison, we will select Aszmocetes, the larva of the lamprey, Petromyzon, which is the nearest relative of Amphioxus among the Craniota. Nervus Branchialis Vagt. Although Ammoccetes possesses an organisation which, especially in virtue of its nervous system and _ sense- organs, entitles it to an undoubted place among the craniate Vertebrates, yet, on the whole, its structural ele- ments remain in such a relatively simple condition of elaboration that it readily adapts itself to a comparison with Amphioxus. At the same time the system of ganglia and peripheral cranial nerves indicated in Fig. 91 will show what a great gap there is between the two forms. Nevertheless, a nerve corresponding to that which lies over the gill-slits in Fig. g1, the nxervus branchialts vagi, has recently been discovered in Amphioxus by van WHE, so that there need be no difficulty in comparing the pharyngeal tract of Ammoccetes with that of Amphioxus. It may be added here that the nerve-supply of the pharynx of Amphioxus was described as a branchial plexus by Rouon and Fusari, but the origin of the nerves which gave rise to the plexus was not satisfactorily determined, beyond the fact that they arose from the rami viscerales of the dorsal spinal nerves. Van WIjJHE also was not 164 DEVELOPMENT OF AMPHIOXUS. able to determine the precise origin of the longitudinal nerve discovered by him. This nerve, which lies on either side at the place where the ligamentum denticulatum passes into the gelatinous lamella derived from the inter- coelic membrane, gives off the branches which form the “branchial plexus.” Van Wijhe states that the origin of the “ramus branchialis vagi” of Amphioxus is to be sought in the eighth to the tenth dorsal spinal nerves. Fig. 91.— Anterior portion of young Ammoccetes of 4 mm., to show extension of brain, origin of endostyle (thyroid), relations of branchial nerves, etc. (After KUPFFER.) I, ll, Ill, 1V. The so-called ‘‘Hauptganglia.” / and //. Trigeminus. III, Acustico-facialis. /V. Glossopharyngeus. V. Vagus. au. Auditory capsule. ch. Notochord. ¢. Endostyle (hypobranchial groove, thyroid). Ay. Hypophysis, in front of which is the nasal groove. ./. Nervus lateralis. 2.67. Nervus branchialis. 0.f. Eye. . Pineal body (epiphysis). ~.m. Przeoral endodermic pouch (median portion of przemandibular cavity. st. Stomodceum. /, V///, First and eighth gill-pouches; the small circles behind the gill-pouches indicate the positions of the external openings of the gill-pouches, which will become perforated later. The small black spots in front of the (later appearing) external openings represent the so-called ganglia pretrematica. He found that the nerve curved ventralwards in front and passed downwards:through the interccelic membrane until it reached the level of the ventral transverse muscles in front of the visceral branch of the eleventh spinal nerve. He was unable to follow it further in the complex nerve- plexus which lies on the surface of the muscles. It is probable, however, that the branchial nerve arises from AMPHIOXUS AND AMMOCGTES. 165 the visceral branch of the eighth, ninth, or tenth spinal nerve.* Stomodeum, Hypophysis, and Gill-slits. It is a common fact that the time and order of forma- tion of corresponding parts differ greatly in the develop- ment of different species. Thus in Ammoceetes, at the stage shown in Fig. g1, the definitive mouth, correspond- ing to the velum in Amphioxus, has not yet formed, but the equivalent of the oral hood is already present in the form of a deep in-pushing of the ectoderm which, at its blind end, is closely applied to the anterior endodermic wall. The mouth will break through later in the middle of the area of contact between ectoderm and endoderm. This ectodermic invagination, whose cavity is probably the homologue of the vestibule formed by the oral hood which leads into the mouth in Amphioxus, is known as the stomodeum. Immediately in front of the stomodceum is another ectodermic involution which is in contact with the front of the brain, and is known as the hypfophysis or pituitary body." It will appear later that this is the probable equivalent of the so-called olfactory pit of Amphioxus. In the wall of the pharynx of Ammoccetes there are, at this stage, the indications of eight pairs of gill-slits. They have not yet, however, broken through to the exterior, but consist of a succession of hollow outgrowths of the phar- ynx stretching towards the ectoderm with which they will eventually fuse (Fig. 92 A, B, C). In the case, however, of the first pair of gill-pouches, * It is not impossible that many of the rami viscerales may send up branches to the branchial plexus, as was indeed described by Rohon. In this case, Van Wijhe’s nerve would be of the nature of a codlector. 166 DEVELOPMENT OF AMPHIOXUS. it does not come to a fusion with the ectoderm; but in- stead they begin to undergo a retrogressive development and eventually flatten completely out (Fig. 92 £). They are thus shown to be rudimentary structures, morphologi- cally representing the first pair of gill-clefts, but never achieving their full development. Fig. 92. — Horizontal sections through the pharyngeal region of Ammoccetes, to show the relation of the first pair of gill-pouches to the peripharyngeal grooves. (After DOHRN.) A, Two days after hatching; first pair of gili-pouches well developed. &B. Six days after hatching; first pair of gill-pouches flattened out. Cc. Nine days after hatching; appearance of peripharyngeal grooves. I-VI, Gill-pouches. 6.w. Body-wall. oes. (Esophagus. gf. Pharynx. ph.g. Peripharyngeal groove. sf, Stomodceum. ve/. Velum. As to their position, they occupy the extreme anterior angles of the pharynx formed by its lateral walls with the anterior transverse wall against which the stomodceum is applied. Whatever may be the reason for it, the atrophy of the first pair of gill-pouches in Ammoceetes is of pre- cisely the same nature as the atrophy of the first gill-slit in Amphioxus, with the distinction that the latter actually opens to the exterior for a time. AMPHIOXUS AND AMMOCGTES. 167 Endostyle or Hypobranchial Groove. At a stage in the development of Ammoccetes which precedes the flattening out of the anterior gill-pouches, a median depression occurs in the extreme anterior region of the ventral wall of the pharynx between the first pair of gillpouches. In its production the wall of the pharynx at this region projects itself ventrally and slightly forward. This groove, which is known as the hypobranchial groove, develops in the direction from before backwards, and eventually extends backwards as a longi- tudinal groove as far as the fifth pair of gill-pouches (Fig. 91). WILHELM MULLER showed that it was the homologue of the exdostyle of Ascidians and Amphioxus, and he has been amply confirmed by Dourn. It agrees with the lat- ter structure in its origin at the anterior extremity of the pharynx and subsequent growth backwards and in its histological structure, the most marked feature of the lat- ter being the four longitudinal rows of gland-cells which were noted above in the endostyle of Amphioxus. (Cf. Fig. 13.) Like the latter, also, it is a slime-secreting gland. In Ammoccetes the hypobranchial groove becomes largely shut off from the cavity of the pharynx by the gradual ingrowth of a diaphragm-like lamella which pro- ceeds from the angle made by the groove in front with the anterior wall of the pharynx (Fig. 91). Subsequently a similar diaphragm grows in from the posterior margin of the groove, and finally the latter only communicates with the pharynx by a small aperture in the mid-ventral line between the fourth pair of gill-pouches. 168 DEVELOPMENT OF AMPHIOXUS. Peripharyngeal Cilitated Bands of Ammocetes. Corresponding with the right and left peripharyngeal ciliated bands which we described as proceeding from the anterior borders of the endostyle in Amphioxus there is a pair of ciliated grooves in the pharyngeal wall of Ammo- coetes which proceed from the anterior lip of the hypo- branchial groove after the latter has become to a large extent shut off from the pharynx by the above-mentioned diaphragm. These grooves curve forwards and upwards SS Che Fig. 93. — Young Amphioxus, after the metamorphosis, having eight gill-slits on each side. From the right side. (After WILLEY.) ?.6. Peripharyngeal band. v. Velum; shown separately below the main figure, with rudiments of four velar tentacles. e¢. Endostyle, extending backwards to the level of the fourth gill-slit. 7.7. Right metapleur. in front of the gill-clefts (after the obliteration of the first pair of gill-pouches), and then proceed backwards on either side of the dorsal middle line of the pharynx as far as the commencement of the cesophagus. Here they appear to curve downwards again, and uniting together, extend for- wards as a median ventral groove to the posterior lip of the hypobranchial aperture. AMPHIOXUS AND AMMOCGTES. 169 The last-mentioned median ciliated groove would appear to be unrepresented in Amphioxus, but the downward curvature of the ciliated bands of the latter behind the gill- slits can be observed (Fig. 93). In Ammoceetes the ciliated peripharyngeal grooves, where they curve upwards in front along the anterior wall of the pharynx, apparently occupy the same position which was previously occupied by the first pair of gill-pouches. Since the latter have already entirely disappeared, there is nothing in the way of their occupying this position (Fig. g2 C). In Amphioxus, where the corresponding gill-slit remains open for a long time, the peripharyngeal band exists without connexion of any sort with the portion of the wall occupied by the slit, and when the latter closes up, it leaves no trace behind. Thyroid Gland. When the metamorphosis of Ammoccetes into Petromy- zon takes place (which happens after a larval existence of some two years’ duration), the hypobranchial groove loses all connexion with the pharynx and becomes broken up by the ingrowth of connective tissue into a number of separate capsules which collectively constitute the thyroid gland of Petromyzon. The thyroid gland is one of those enigmatical ductless glands which form such a curious and constant feature of the Vertebrate organisation. There is considerable doubt as to the specific physio- logical function which it has to perform, but at the same time it is a necessary factor in the Vertebrate economy, and is of great importance from a pathological point of view. In the higher forms it is attached to the lower side of the larynx, and appears to have received its name on 170 DEVELOPMENT OF AMPHIOXUS. account of its close proximity to the thyroid cartilage of the latter, the older anatomists assuming a functional relation between the two structures. We know perhaps more about the morphological than about the physiological significance of the thyroid gland, since it is the vestige of the very actively functional endo- style or hypobranchial groove of the Ascidians, Amphioxus, and Ammoceetes. Morphology of Club-shaped Gland of Amphioxus. In describing above the formation of the second row of gill-slits in Amphioxus, we found that the first secondary slit paired with the second primary slit. It now remains to consider what has become of the antimere of the first primary slit. The probability is that, unlike the antimeres of the suc- ceeding primary slits, that of the first has not suffered a retardation of development, but is present from the very beginning of the larval development, although in a some- what modified form. I refer to the club-shaped gland. The club-shaped gland fulfils the requirements of a gill- slit in so far as it opens at one end into the pharynx, and at the other to the exterior. Since, as we have shown, the morphological mid-ventral line lies high up on the right side, immediately above the primary gill-slits, it is evident that its anterior continuation would pass through the en- dostyle precisely at the point where the latter is redoubled upon itself. But the internal opening of the club-shaped gland lies above the upper limb of the endostyle, and therefore it is placed not only on the actual right side of the larva, but in opposition to the first primary slit, on the morphological right side as well. AMPHIOXUS AND AMMOCGTES. Ld It must be supposed that the original gill-slit, from which the club-shaped gland is derived, acquired, for some reason or other, a tubular form. A familiar precedent for gill-slits being drawn out into elongated tubes, the effect of which is to separate the in- ternal from the external opening by a long interval, is presented by the hag-fish, JZyazze. Myxine also shows us that, in correlation with the canalisation of the gill-slits, their external apertures may enter into new relations dif- fering considerably from the primitive condition. As is well known, the elongated tubular gill-clefts of Myxine do not open separately to the exterior, but fuse together at their distal extremities, so as to give rise to a longitudinal duct on each side, which opens to the exterior some dis- tance behind the gill-region. It is only on some such supposition as this—namely, that the external aperture of the gill-slit represented by the club-shaped gland of Amphioxus has assumed new topo- graphical relations in correlation with the canalisation of the original slit — that its position on the opposite (left) side of the body to the internal opening of the gland is ren- dered intelligible. The position of the internal opening furnishes the criterion by which to judge of the primitive relations of the original gill-slit. With the above point of view, therefore, we may signal- ise the following facts to prove that the club-shaped gland is the antimere of the first primary gill-slit. 1. They arise simultaneously in the embryo as grooves in the ventral wall of the pharynx. 2. They come to lie on opposite sides of the morphological median line —the first gill-slit entirely so, and the club-shaped gland in respect of its internal opening into the pharynx. 172 DEVELOPMENT OF AMPHIOXUS. 3. They atrophy and disappear simultaneously during the metamorphosis of the larva. 4. No secondary gill-slit ever arises to pair with the first primary slit. As the stage represented in Fig. 64 marks such a vital turning-point in the development of the individual, being the stage at which the embryo becomes a larva and the struggle for existence in obtaining independent nourish- ment genuinely sets in, it is important to be able to define it accurately. In view of the above considerations, we arrive at the conclusion that the larva is at this stage possessed morphologically of a pair of gill-slits. It should be pointed out that this opening stage of the larval development appears to be of the nature of a vest- ang phase, during which the larva accumulates energy for future growth. Preoral “ Nephridium”’ of Hatschek. In the larvee of Amphioxus there is a structure lying at the base of the notochord on the left side, immediately above the preoral pit, which we have not yet consid- ered. (Cf. Figs. 81 and 82, x.) According to Hatschek, who first described it, it arises in the larva as a mesodermal, ciliated funnel and canal in front of the mouth, in the region of the first metamere. It lies in a narrow division or prolongation of the body-cavity, beneath the left aorta. (CE Fig. 768.) At its hinder end it opens into the pharynx. Hatschek interprets this structure as a nephridium. Its true physiological, and especially its morphological, sig- nificance is, however, very perplexing and requires further study. AMPHIOXUS AND AMMOCCTES. 173 Ancestral Number of Gill-slits. The unlimited number of gill-slits in the adult Amphi- oxus has led to a good deal of controversy as to the ap- proximate number present in the ancestral Vertebrate, some authorities being of the opinion that Amphioxus presents the primitive condition in this respect, and others that the multiplication of gill-slits in this form was a secondary phenomenon. Sometimes as many as fourteen pairs of gill-clefts are found in a remarkable cyclostome fish from the Pacific, allied to Myxine, and called Sdellostoma.* With this ex- ception, no true fishes, recent or fossil, are known which possess more gill-slits than the existing sharks belonging to the family of the JVotzdanide. Of these the genus Heptanchus possesses eight gill-clefts (ze. seven plus the spiracle) on each side, and Heranchus seven. In Ammo- coetes, as we have seen, there are at one time indications of eight pairs of gill-slits. The first pair of these, how- ever, never breaks through to the exterior, and eventually disappears, but Dohrn has shown that the primary rela- tion in which the seventh pair of cranial nerves stands to it, indicates that it is the homologue of the spzvacle of the higher forms. Moreover, in the larval development of Amphioxus several facts combine to produce the impression that the indefinite number of gill-slits in the adult is a secondary acquirement. First of all, there is the series of primary gill-slits which, while varying within narrow limits, usually numbers fourteen. Their unpaired unilateral character is merely incidental, as explained above, and it may be stated * For a recent account of Bdellostoma, consult HowARD AYERS, No. 69, bibliography. 174 DEVELOPMENT OF AMPHIOXUS. that they are potentially paired, the first of them in all probability being actually paired (with the club-shaped gland). In the second place, after the closure of a number of the primary slits, the so-called crztzcal stage occurs with eight pairs of gill-slits. This is another resting phase in the development, and marks the turning-point from the Jarval to the adolescent period. Subsequently the addi- tion of new gill-slits behind those already present com- mences and goes on indefinitely throughout life. Counting in the first pair of slits (z.e. first primary slit plus club-shaped gland) which is destined to atrophy, we must regard it as probable that the proximate common ancestor of Amphioxus and the higher Vertebrates was characterised by the presence of from xzne to fourteen pairs of gill-clefts, although it is also probable that there was a variable tendency to add to this number by fresh perforations. NOTES. I. (p. 105.) It is unaccountable how there can have been conflicting statements as to the ejection of the genital products (male and female) through the atriopore. It was first observed by DE QUATREFAGES in 1845, and his observations have since been fully confirmed by PauL Bert, A. WILLEY, and E. B. WiLson. On the other hand, both Kowatevsky and Hartscuexk affirm that they are discharged through the mouth. It is to be regretted that two such eminent observers should have committed this error, since it is difficult to eradicate it from the text-books. 2. (p. 115.) The primitive endoderm cells in the neighbour- hood of the neurenteric canal apparently retain an undifferentiated character, until the completion of the myotome-formation. In the young embryo they are to be observed in transverse section in pro- cess of division, numbers of karyokinetic figures being present. But the cells divide without regard to the median plane of sym- NOTES. 175, metry, and the recent researches of E. B. Witson and Lworr lead to the conclusion that the so-called mesoblastic pole-cells, which were described by HarTscHEk, have no real independent existence. 3. (p. 123.) Whether the dorsal and ventral fin-spaces are actually derived from the original myoccel, as described by Hat- schek, or do not rather arise by a splitting of an originally solid thickening of the gelatinous connective tissue which surrounds them, must remain doubtful. The cavity of the metapleural folds certainly arises as a schizocel, t.e. by a hollowing out of a solid thickening. Even in case the fin-spaces also arise as schizoccels, Hatschek’s interpretation of their morphological significance might still hold good. 4. (p. 123.) A transitory pouch-like diverticulum of the myo- coel has been observed in connexion with the formation of the sclerotome in the Selachian embryo by Rast and H. E. ZIecver. 5. (p. 129.) Since the work of BaLrour on the development of Elasmobranch fishes (Selachians), it has been known that the paired praemandibular head-cavities communicate with one another across the median line in the embryo. ‘The important results obtained by the researches of KupFFER (Petromyzon, Acipenser), KAsTSCHENKO (Selachian), and Juia Piatr (Selachian), not only established the fact that the praeemandibular cavities arose essen- tially as anterior archenteric pouches (cf. Fig. 72), but also that the median cavity which effected their communication across the middle line, from side to side, arose by constriction from the front end of the archenteron (using the latter term with some latitude), and that, therefore, the wnzon of the right and left premandibular cavities in the embryo of the craniate Vertebrates is primary, and not secondary, as was previously supposed. For an excellent historical and critical summary of our knowl- edge of the origin of the head-cavities in the craniate Vertebrates, the reader may consult Froriep. (See bibliography.) 6. (p. 130.) The ciliation of the ectoderm in the larva of Amphioxus continuing, as it does, long after the muscles have been fully differentiated, and when the cilia are therefore no longer required for purposes of locomotion, should be especially noted as evidence of a very archaic organisation. We shall find in the last chapter that the possession of a ciliated 176 DEVELOPMENT OF AMPHIOXUS. ectoderm is a prime characteristic of Bal/anoglossus and many of the lower worms (e.g. /Vemertines). In none of the craniate Vertebrates is the ectoderm at any time ciliated. 7. (p. 134.) The exact stage at which the club-shaped gland reopens into the pharynx must remain an open question. It is, very probably, subject to a good deal of variation in this respect, occurring now earlier, now later. Experiments to determine the physiological role of this gland are much needed. 8. (p. 143.) In accordance with Dohrn’s conception of the principle of the change of function (Das Princip des Functions- qwechsels), the number and nature of the organs of the Vertebrate body, which have been interpreted as modified gill-clefts, are truly astonishing. First and foremost, Dohrn supposed that the Verte- brate mouth arose by the fusion of two gill-slits across the middle line, the old Annelid-mouth, which perforated the central nervous system, having been lost. A great many forcible arguments have been brought forward in support of this hypothesis. Dohrn him- self would probably admit that it is only tenable on his further hypothesis that Amphioxus is a form which has undergone a retro- gressive evolution from the craniate Vertebrates. This was a better hypothesis than that of Semper, who, perceiving that Amphioxus would not fall in with the Annelid-theory, declared, “er sel kein Wirbelthier ; also, auch kein Fisch.” Besides the mouth, many other structures have similarly been referred back to modified gill-slits, among which may be mentioned the nose, hypophysis, thyroid gland, lens of the eye, and the anus. None of these comparisons is supported by the facts of develop- ment and anatomy of either Amphioxus or the Tunicates, while most of them would appear to be definitely disproved by these facts. 9. (p. 147.) Since the right metapleural fold bends round to the median ventral line of the snout, as shown in Fig. 38, and since, further, at a later period, the right half of the oral hood is similarly continued round the front end of the body into the dorsal fin, it is clear that the right half of the oral hood must arise essentially in continuity with the right metapleur. On the contrary, the left half of the oral hood arises entirely independently of the left metapleur. It is possible that this discontinuity of NOTES. 177 development of the left half of the oral hood and the left meta- pleur has been secondarily brought about. 10. (p. 150.) The study of transverse sections has led me to the conclusion that the backward extension of the endostyle is effected by interstitial growth, and not by the conversion of the cells which form the primary floor of the pharynx into endostylar elements. These cells are probably disintegrated and absorbed by the endostyle as it grows backward. It. (p. 153.) For a comparison between the perigonadial cavities of Amphioxus and the mesonephric tubules of the craniates the reader should consult Boveri’s original memoirs. (See bibliography.) 12. (p.159.) The following definition of the so-called bio- genetic law of recapitulation (Haeckel’s biogentisches Grund- gesetz) will explain the meaning of Haeckel’s terms “ cenogenesis ” and “ palingenesis.” According to this law: The development of the individual (onfogeny) is a compressed summary of the gradual modifications which have resulted in the evolution of the species, or type (phylogeny = Stammesgeschichte) ; this recapitulation (summary, or Auszug) of the phylogenetic stages in the ontogeny is the more perfect according as the ancestral development (Palingenesis, Auszugsentwicklung) has been the less disturbed or falsified through secondary or “recent” adaptation (ceno- genesis, Stdrungsentwickelung) of the embryo or larva to a new environment. 13. (p. 162.) The explanation of the asymmetry of the larva of Amphioxus given in the text was first suggested by me in 1891. It may be well to state that it has not as yet received very general recognition in the more recent literature on the subject. It was, however, fortunate enough to receive the endorsement of the late Professor MitNes MarsHALt in his text-book of Vertebrate Em- bryology. When the pelagic larvae of Amphioxus are confined in glass jars, after a certain lapse of time they sink to the bottom, like all other pelagic organisms. When they arrive at the bottom, they fall over on to one side, owing to a physical impossibility to rest in any other position, just as was described above for the adult. It ought not to require to be emphasised that their inci- dentally lying on one side is not due to a pressing desire or I 78 DEVELOPMENT OF AMPHIOXUS. instinct to assume that position, but rather because they cannot help it. It is apparently in consequence of a misunderstanding of this observation that KorscHetr and Hetper ascribe the larval asymmetry of Amphioxus to the same causes which brought about the asymmetry of the Pleuronectidz. Another, and, as it appears, a still more impossible view, has recently been expressed by vAN WijHE. According to van Wijhe, the left-sided mouth occupies its normal and primitive position in the larva of Amphioxus, and in that position it represents a gill-slit, whose antimere is the club- shaped gland. Van Wijhe arrived at this view as a result of his very important discoveries as to the musculature and innervation of the adult mouth. These discoveries may be summarised as follows : — 1. The outer muscle of the oral hood represents the anterior continuation of the /e/t half only of the transverse and subatrial muscles. 2. The inner nerve-plexus of the oral hood is formed on both sides, exclusively from nerves which arise from the left side of the central nervous system. 3. The velum is innervated entirely from nerves of the left side. From these observations van Wijhe concludes that the mouth of Amphioxus, even in the adult, is essentially an organ of the left side, and is neither homologous with the Ascidian nor with the craniate mouth. It would seem, however, that the more obvious and justifiable interpretation of these facts is that the asymmetrical musculature and innervation described by van Wijhe are merely the partial persistence in the adult of the more complete asymmetry of the larva. Van Wijhe’s observations, therefore, do not affect the question of the cause of the asymmetry in any degree. 14. (p. 165.) As first shown by Dohrn, the hypophysis of Ammoceetes first arises from the roof of the stomodceum, from which it is subsequently removed to the dorsal surface of the head by the enormous development of the upper lip. 15. (p. 169.) The ciliated tracts in the pharynx of Ammo- coetes were first described and figured by ANTON SCHNEIDER in NOTES. 179 1879. In 1886 DouRN thought he had proved that the anterior portion of them, which bends upwards on either side of the pharynx and forms the sevipharyngeal grooves, represented the last traces of the aborted first pair of gill-clefts. Although they appear at the place which was formerly occupied by these rudi- mentary gill-pouches, yet, according to Dohrn’s own account, they do not appear until after the gill-pouches have completely flattened out. Under these circumstances, but above all, in view of the relations of the homologous peripharyngeal bands in Amphioxus which exist both before and after the disappearance of the first pair of gill-clefts (¢.e. first primary gill-cleft and club-shaped gland), it must be assumed that Dohrn’s interpretation, though most natural, was nevertheless somewhat at fault. LV. THE ASCIDIANS: Tue Ascidians, Tunicates, or sea-squirts, as they are indifferently called, constitute one of the most clearly defined and yet most heterogeneous groups of animals which it is possible to imagine. There is a great variety of families, genera, and species occurring all the world over, and in all depths of the ocean from the tide-marks to the profoundest depths. Most of them are sedentary animals, remaining fixed all their lifetime on one spot, whether attached to rocks, stones, shells, or sea-weeds, from which they are incapable of moving. There are, however, several very extraordi- nary genera of Ascidians which swim or float about per- petually in the open ocean, and have become adapted in the extremest manner to a purely pelagic environment. These pelagic Ascidians have become so modified in adap- tation to their oceanic existence, and their development diverges, as a rule, so much from the normal, that they will hardly enter at all into the present discussion, with the exception of one family, the Appendiculariea. Just as there are two kinds of sessile Ascidians, szwple and compound or colonial, so there are two analogous kinds of pelagic Ascidians. In some of the latter, however, where there is an alternation of generations, one genera- tion, namely, the asexual generation, is a solitary form, while the sexual generation is a colonial form, as, for example, the solztary Salpa and the chain-Salpa. 180 ANATOMY AND DEVELOPMENT. I8I For convenience, the Ascidians, as a whole, may be arranged as follows :— SESSILE ASCIDIANS. SIMPLE. COLONIAL. e.g. Ascidia. e.g. Clavelina. Phallusia. LBotryllus. Ciona. Aniaroucium. Molgula. Distaplia. Cynthia. Didemnum. PELAGIC ASCIDIANS. SIMPLE. COLONIAL e.g. Appendicularia. (or capable of producing a colony by budding). e.g. Pyrosonma. Salpa. Dotiolum. The compound sessile Ascidians consist of colonies of individuals or asczdiozooids produced by budding from a parent individual. Such colonies are often brilliantly coloured and of massive proportions, as Amaroucium and Fragarium. Others form thin encrusting expansions on the surfaces of marine plants and shells, as Botryllus and L epto- clinum. In others, again, the individuals are entirely separate, except at the base, where they are connected together by a common creeping stolon from which new buds are periodically produced, as Clavelina and Perophora. STRUCTURE OF A SIMPLE ASCIDIAN. Test, Mantle, Atrium, Branchial Sac. The simple or solitary Ascidians which do not produce buds, present hardly less striking differences among the different families than do the compound, but their general shape is much more uniform. 182 THE ASCIDIANS. An average simple Ascidian, as Phallusta or Cynthia, has been aptly compared to a leather bottle provided with two spouts. The spouts occur in the form of two funnel- like prominences projecting from the surface of the body and bearing at their free extremities the zzcurrent or buc- cal and excurrent or cloacal apertures respectively, the latter usually occurring at a lower level than the former. The most prominent and, apart from the two apertures, the only external feature of a simple Ascidian, is the char- acteristic ¢azc or zest which surrounds the whole body. As a rule, all Ascidians of whatever kind possess this external tunic, and it is one of their chief diagnostic characters. According to the species this test may be of a cartilagi- nous, coriaceous, fibrous, or membranous consistency, usually opaque, but sometimes hyaline and transparent, as in Corella, Salpa, etc. Its outer surface may be smooth, wrinkled, or rough, capillated, papillated, or mammillated. In 1845 Kart ScHMIDT made the discovery that the test of the Ascidians was largely composed of the substance which forms the cell-walls in plant tissues ; namely, ce//u- Jose. When treated with the proper chemical reagents, it gives the cellulose-reaction. This is interesting as show- ing the fundamental identity of protoplasm whether it occurs in animal- or in plant-cells, since in both cases it - is capable of depositing cellulose. Judging by external appearances an ordinary Ascidian resembles nothing so little as Amphioxus, and yet it is probably more closely related to the latter than is the lamprey larva, Ammoccetes, whose external resemblance to Amphioxus is incomparably greater. It is only in its internal organisation that we meet with structures which remind us strongly of corresponding parts in Amphioxus. ANATOMY AND DEVELOPMENT. 183 A schematic representation of a dissection of a typical Ascidian after Professor W. A. HERDMAN, whose reports on the Ascidians collected during the voyage of H. M. S. Challenger have done so much to advance our knowledge of the group, is given in Fig. 94. The greater part of the thick cartilaginoid test (also called tunic, outer mantle, or cellulose mantle), 4 is supposed to be removed from the right side, and its cut edge can be traced all the way round. Below the test comes the inner or muscular mantle, 72, which is the true body-wall, to which the external tunic is secondarily superadded.t The muscular mantle is limited externally (below the test) by the epidermis, and beneath the latter are the interlacing muscle-fibres which compose the bulk of the mantle. Beneath the mantle is an extensive cavity surrounding to a large extent the viscera. This is the perzbranchial or atrial cavity which communicates with the exterior by the atrial or cloacal aperture, az.s. The mouth, os, leads into the pharynx or branchial sac, ph, which is of surprising dimensions, and stretches nearly to the posterior end of the body. The walls of the bran- chial sac are perforated by innumerable gill-openings, the so-called stzgmata, arranged in successive transverse rows, through which the water which enters at the mouth passes out of the sac into the atrial cavity. Dorsal Lamina, Endostyle, and Peripharyngeal Band. On cutting through its right wall we open into the cavity of the branchial sac along the dorsal side of which a fold is seen projecting freely into the cavity, the so-called dorsal lamina corresponding to the dorsal groove in the pharynx of Amphioxus, while along its ventral side is a 184 THE ASCIDIANS. \\ unl nas uN ai ; mee F \ gaqnsnnel Too coo sr Fig. 94. — Diagram of a dissection of Asczdia, from the right side. (After HERDMAN.) The peribranchial cavity is indicated by the black shading. an. Anus. at.s. Atrial siphon. c.g. Cerebral ganglion, beneath which is the subneural gland and its duct. a@./. Dorsal lamina. ewzd. Endostyle. 2. Gonad. g.d. Genital duct. zz¢. Intestine. 2. Muscular mantle. oes. Aperture, leading from branchial sac into cesophagus. o7.s. Buccal siphon. ff. Branchial sac. st. Stomach. ¢# Test or cellulose mantle. ‘7. Buccal or coronary tentacles. ty. Typhlosole; internal fold of intestinal wall, to increase the digestive surface. ANATOMY AND DEVELOPMENT. 185 well-defined groove with white glistening walls, which is the exdostyle. The groove of the endostyle is deeper here than in Amphioxus, but its epithelial walls have the same histological differentiation, with the two rows of gland- cells on each side of the middle line, the latter being occupied by a median group of cells carrying very long cilia. The food which enters the mouth together with the water does not pass out of the pharynx into the atrial chamber, but is caught up by the slime secreted by the endostyle and is then carried forwards along the endostyle, and, having arrived at the anterior extremity of the latter at the base of the buccal tube, is carried round along a circular ciliated groove which surrounds the base of the mouth at the entrance to the branchial sac, until it reaches the dorsal side of the animal, when it is led backwards by the ciliary action of the cells of the dorsal lamina in the form of a cord of slime in which the food-particles (micro- scopic organisms, vegetable débris) are imbedded. The ciliated groove round the base of the buccal tube connecting the anterior extremity of the endostyle with the dorsal lamina is known as the peripharyngeal band or pericoronal groove. Ne have already made the acquaint- ance of the homologue of this structure both in Amphi- oxus and in Ammoccetes. It forms a complete circle round the base of the buccal tube and is indicated in Fig. 94 by the black line which limits the pharyngeal wall anteriorly. It is still better shown in Fig. 96, which represents a young individual of Clavelina. The cord of slime containing the food passes backwards along the dorsal lamina to the opening of the cesophagus, which lies near the posterior end of the branchial sac, in the dorsal middle line, through which it passes into the stomach. The dorsal lamina is continued to one side of 186 THE ASCIDIANS. the oesophageal aperture, as a low ridge, which joins the posterior extremity of the endostyle.* Visceral Anatomy. Except in its most anterior region, the dorsal border of the pharynx lies freely in the atrial chamber. On the contrary, along its ventral border, throughout the whole section (After HERDMAN.) ‘The muscular mantle is indicated by the black shading. a, Peribranchial cavity traversed by numerous vascular trabeculze, through which the blood flows into the branchial bars. 47.5. Branchial sac. é.v. “ Blood-vessels.” d./. Dorsallamina. e. Endo- style. ec. Ectoderm. gy. Gonad. gd. Double genital duct. 7. Intestine, with tvphlosole. 7. Rec- tum, 7.0. Renal vesicles. 7# Test. Fig. 95. — Diagrammatic transverse through the middle of the body of Asczdza. length of the endo- style, it is attached to the muscular mantle. In other words, the right and left halves of the atrial cavity are continuous round the dorsal side of the pharynx, but)! Gare separated from one another ventrally by the concrescence of the endostyle with the mantle. (Cf. Fig. 95.) In Amphioxus, as we have seen, the opposite condition ob- tains. There, the dor- sal wall of the pharynx ° is closely applied to the notochord, while the endostylar tract * Compare the above with the description of the course of the ciliated tracts in the pharynx of Ammoccetes, given on p. 168. ANATOMY AND DEVELOPMENT. 187 is free, so that the right and left halves of the atrial cavity are continuous ventrally, instead of dorsally. In order to see the stomach and intestine, it is necessary to cut through the left wall of the pharynx, since the vis- cera lie, at least in the genus Ascidia (or Phallusia), on the left side of the pharynx. It should be pointed out that the topographical arrangements vary considerably among the different genera of Tunicates. In Clavelina, for example, the viscera lie behind the pharynx, as shown in Fig. 96. On the left side of the pharynx (Fig. 94) the short cesophagus leads into the dilated stomach, which again narrows down to the looped intestine, and finally the lat- ter bends sharply forwards into the rectum, which opens by the anus into the atrial cavity, the excrement being carried to the exterior by the constant stream of water which flows out through the atrial or cloacal aperture. Instead of being straight, as in Amphioxus, the aliment- ary canal is here doubled round upon itself. This U-shaped character of the alimentary canal of Ascidians is shown with great clearness in the case of Clavelina (Fig. 96), where there are no secondary convolutions in the course of the intestine. The Ascidians are one and all hermaphrodite, and the reproductive glands frequently lie between the loops of the intestine, while two ducts, ovzduct and vas deferens, which often present the appearance of a single duct with a double lumen, proceed forwards by the side of the rec- tum, to open into the cloacal region of the atrial cavity near the anus (Fig. 94, ¢ and gd). The ovary and testis, though quite separate in the adult, originate, according to the account given by the Belgian zoologists, EDOUARD VAN BENEDEN and CHARLES JULIN, 188 THE ASCIDIANS. from a common centre of formation, which subsequently undergoes a division into two portions, one of which be- comes the ovary, and the other the testis. Similarly the oviduct and vas deferens are derived by division of a primarily single structure, which arises in continuity with, and in fact as an outgrowth from, the primitive sexual gland. In spite of their hermaphroditism, it would appear that not all the Ascidians are self-fertilising, although many, if not most of them, are. In some cases it is supposed that in different individuals the male and female organs attain maturity at different times, so that in a given individual, when the ovary is ripe the testis is unripe, so that it must be fertilised from another individual, in which the testis is - ripe, but the ovary unripe, and so on. Nervous System and Hypoplysts. (Neurohypophysial Systent.) The central nervous system of an Ascidian usually bears a ridiculously small proportion to the bulk of the organ- ism. Its main constituent is a ganglion which lies im- bedded in the thickness of the mantle, between the oral and the atrial siphons, the two latter structures being innervated by nerves proceeding from the ganglion. As belonging to the central nervous system must also be mentioned a solid nerve-cord which runs along the dorsal border of the branchial sac from the cerebral ganglion to the visceral region (Fig. 96). This was discovered by van Beneden and Julin, and is derived from a persistent portion of the central nervous system of the larva. Beneath the cerebral ganglion is a lobulated glandular organ known as the sabneural gland. It is provided with ANATOMY AND DEVELOPMENT. 189 a duct which runs forward and opens at the end of a ciliated funnel-shaped dilatation into the branchial sac at the base of the buccal tube (Figs. 94, 96, and g7) in front of the peripharyngeal band. The branchial opening of the duct of the subneural gland appears primarily as a simple circular orifice, but it does not usually retain this character in the adult. Generally it assumes a crescentic form by the in- curving of its anterior or posterior lip, and then in many cases the horns of the crescent so formed become coiled over and over con- centrically, and usually in the plane, so that the lips of approximately same the aperture assume a very complicated appearance and constitute the so-called dor- sal tubercle (Fig. 97). It has taken a long time and the work of a great many zoologists to achieve our (which present knowledge is by no means Fig. 96. — Young Clavelina, shortly after the metamorphosis, from the right side. (After VAN BENEDEN and JULIN.) at. Atrial opening. afc. Atrial cav- ity. 3.5. Blood-sinus. ed. Endostyle. ep. Epicardium; outgrowth from bran- chial sac behind endostyle, which grows down into the creeping stolon, forming a septum in the latter, and being the chief element in the production of buds. f Lobes of the fixing organ, which give rise to the creeping stolon. .. Ganglion. gs. Stigmata. 4. Heart. Ay. Hypophysis (dorsal tubercle). 7zz/. Intestine. 7. Mouth. oes. Esophagus. 4.0. Periphar- yngeal band. fc. Pericardium. 7 Re- mains of tail, withdrawn into the body. v.m, Visceral nerve. complete) of the subneural gland of Ascidians and its duct. Fig. 97. — Hypophysis of Phad/usia mentula, prepared out and seen from the inside. (After JULIN.) g- Subneural gland, above which may be seen the outline of the ganglion and its nerves. ad. Duct of the subneural gland. z. Dorsal tubercle, the opening of the hypophysis into the branchial sac. The actual opening is indicated in black. pc. Peripharyngeal groove. ep. Epi- branchial groove. d./. Dorsal lamina, slightly displaced, to show the duct of the subneural gland above it. N.B.—In this species, the atrial and buccal siphons are widely separated, and the duct of the subneural gland is very long. THE ASCIDIANS. The dorsal tubercle was discovered by the celebrated SAVIGNY in 1816, and was for a long time supposed to be an independent sense- organ of an olfactory nature. The subneural gland was detected not as a gland, but aS an enigmatical structure lying below the brain by the English naturalist Han- COCK in 1868. character was demonstrated by NassonorF and Ussow in 1874-75, the last-named author showing its connex- ion by means of the duct with the dorsal tubercle. In 1881 JULIN produced an admirable memoir on the Its glandular subneural gland and its duct, and strongly urged its ho- mology with the pituitary body or hypophysis cerebri of the higher Vertebrates. The same suggestion was made in a more tentative form in the same year by BaLrour. We shall have to consider this question Suffice it to say at present that Julin’s sugges- later. tion has been accepted to ANATOMY AND DEVELOPMENT. IOI the extent that the subneural organ of the Ascidians is frequently spoken of as the Aypophyszs. Circulatory System. With regard to the circulatory system the Ascidians differ markedly from Amphioxus in the possession of a well-defined eart which lies in a distinct pericardium. The heart lies ventrally and usually in the neighbourhood of the stomach. (Cf. Fig. 96.) Its wall is muscular, but consists only of a single layer of cells whose deeper portions (2.c. towards the cavity of the heart) are drawn out into striated muscular fibres, while the outer portions of the cells containing the nuclei project into the cavity of the pericardium. There is therefore no true endothelial lining to the heart, and the cells which build up its wall offer a most interest- ing example of epithelio-muscular tissue, as was first pointed out by Edouard van Beneden. This type of muscular tis- sue, in which the muscle-fibres occur as basal prolonga- tions of cells which still retain their epithelial character, is found, as is well known, in the case of the body-muscles of the Nematode or thread-worms, and is above all character- istic of the Coelenterata (Hydroids and Medusz). There are no true blood-vessels in Ascidians, but the passages along which the blood percolates are merely lacunze in the connective tissue and musculature of the body and between the viscera. They are not lined by an endothelium, and are more correctly described as. b/ood- sinuses. They are often irregular in their outline, as shown in the transverse section represented in Fig. 95, but often again they simulate the appearance of true blood-vessels, as in the case of those branches which pass from the mantle into the substance of the test, as well as the tubes 192 THE ASCIDIANS. which traverse the wall of the branchial sac in every direction. In the second chapter it was pointed out that the Vertebrate heart arose as a specialisation of a portion of the primitive sub-intestinal blood-vessel whose calibre was originally uniform throughout, and that in Amphioxus the cardiac region of the vascular system retains its primitive tubular character. Very different is the actual origin of the Ascidian heart ; although it is simply a dilated tubular structure, yet it arises entirely independently of and prior to the rest of the vascular system at a time, in fact, before the formation of the muscular mantle and before the atrial cavity has so far extended itself as to almost entirely replace the original body-cavity. The blood-sinuses of the Ascidians are rem- nants of the latter. With the formation and growth of the atrial cavity, the perforation of the stigmata, and the development of the muscular mantle, the original body-cavity becomes reduced to a system of narrow canal-like spaces which constitute the above-mentioned blood-sinuses. The general distribu- tion of the blood-sinuses can be made out from Fig. 95. There are two main longitudinal sinuses, one below the endostyle and another above the dorsal lamina, while others are scattered irregularly in the muscular mantle ; others again lie in amongst the viscera forming the inter- spaces between the various parts; and finally the bran- chial bars between the stigmata are all hollow, and their cavities are placed in communication with the system of sinuses at intervals as shown in Fig. 95. The periodic contraction of the heart of Ascidians takes place on a highly characteristic and unique plan. Each systole occurs as a peristaltic wave of contraction passing ANATOMY AND DEVELOPMENT. 193 from one end of the heart to the other; but the chief peculiarity in connexion with it is, that after a certain number of contractions in one direction the heart makes a brief pause and then commences to contract again in the opposite direction, and so it goes on contracting now in one direction and now in the other. This phenomenon of the periodic reversal of the direction of contraction of the Tunicate heart is known as the vecurrent action of the heart, and was discovered in 1824 by vAN HASSELT. The discovery was first made in the case of Salpa, but it has since been found to hold good for all Tunicates. When the heart contracts from its posterior to its an- terior extremity, that is to say, in the postero-anterior direction, —the blood is thereby propelled forwards into the blood-sinus which lies below the endostyle, and from this it passes into sinuses which run transversely into the bran- chial bars. In the basket-work formed by the intercross- ing of the branchial bars, the blood has a complicated and irregular course, and is finally collected into the dorsal sinus which lies above the dorsal lamina. Here it flows backwards, and after passing in amongst the viscera arrives back to the heart. (Other branches of the sinuses pass into the test, where they end in curious knob-like dilata- tions.) On the contrary, when the heart contracts in the reversed or antero-posterior direction, the blood which has already been oxygenated in its passage through the branchial bars is sent to the viscera direct, and from there it collects into the dorsal sinus, from which it is distributed over the branchial sac, and so into the sub-endostylar or ventral sinus, in which, it flows backwards to the heart. On account of the above peculiarities relating to its independent origin, the histological structure of its wall, 194. THE ASCIDIANS. and its recurrent action, the Tunicate heart would appear to be a unique organ peculiar to the group of the Ascid- ians and analogous but not homologous, or only incom- pletely so, with the heart of the Vertebrates. Again, the vascular system of an Ascidian is only func- tionally comparable to that of Amphioxus, since true vessels provided with an endothelial lining are entirely absent, their place being taken by sinuses which arose by reduction from the original body-cavity. Renal Organs. The renal organs of the Ascidians have no apparent morphological relation to those of Amphioxus, and therefore need not detain us. They consist of a group of bladder-like vesicles with cellular walls lying around the intestine. The products of excretion (uric acid, etc.) are deposited inside the vesicles in the form of solid concretions. There is no excretory duct. In Molgula, there is a single large cylin- drical renal sac closed at both ends and lying on the right side of the body, behind the heart, known as the organ of Bojanus. Comparison between an Ascidian and Amphioxus. Having sketched in rough outline the organisation of an adult Ascidian, we are now in a position to consider in what respects it resembles and in what it differs from that of Amphioxus. We shall see that some of the most funda- mental differences will be made good by the structure of the larva,— such as the absence of a dorsal nerve-tube and of a notochord. Let us first consider the resemblances between an adult Ascidian and Amphioxus. ANATOMY AND DEVELOPMENT. 195 In both cases the pharynx is perforated by a great num- ber of g2/l-apertures (gill-slits, stigmata), converting it into a branchial sac and opening into an atrial or peribranchial cavity instead of directly to the exterior. At the base of the pharynx there is a longitudinal gland consisting of a groove open throughout its whole length towards the cavity of the pharynx, and known as the exdostyle, whose histo- logical character is closely similar in the two cases. From the anterior extremity of the endostyle a ciliated band of columnar cells passes round the wall of the pharynx on each side, in front of the gill-openings, and abuts on the dor- sal border of the pharynx, along which it is continued back- wards in connexion with the dorsal lamina in the one case and the kyperpharyngeal groove in the other. This band forms a circlet round the pharynx behind the velum, and is the peripharyngeal band.* We shall find also that the Ascidian hypophysis is essentially homologous with the olfactory pit of Amphioxus. In the Ascidians there are sphincter muscles round the buccal and atrial siphons, and inside the former, in front of the peripharyngeal band (pericoronal groove), there is a circlet of tentacles corresponding perhaps to the velar tentacles of Amphioxus. (Cf. Fig. 94, ¢z.) The differences between the structure of an adult Ascid- ian and of Amphioxus may appear to outweigh the resem- blances, but it must be remembered that they are all correlated with and accessory to the one great difference in the mode of existence of the respective types. An Ascidian is sessile ; Amphioxus is free. The former, as it were, builds its house upon a rock and is immovable ; the latter lives in the shifting sands, and is capable of extremely active locomotion. * As mentioned above, this band is usually grooved in the Ascidians. 196 THE ASCIDIANS. In correlation with this sessile habit of existence we find that the Ascidians, in contrast to Amphioxus, are hermaph- rodite, —an almost universal condition among sessile organ- isms of every description. They are unsegmented, the muscles not being divided up into myotomes ; and none of their organs (gonads, renal organs, etc.) are metamerically repeated, unless we regard the successive transverse rows of stigmata in the wall of the branchial sac as evidence of metamerism. It is, however, of a totally different nature from the metamerism of the gill-slits of Amphioxus, and we shall see that only in the earlier stages of their devel- opment can the stigmata of the Ascidians be compared with the former. Another of the most characteristic accompaniments of a sessile mode of life is the U-shaped alimentary canal. Instead of being a straight tube with a posteriorly directed anus as in Amphioxus, the alimentary canal of the Ascid- ians is doubled up upon itself, the rectum is directed for- wards, and the anus opens into the atrial cavity. The absence of a dorsal nerve-tube and notochord in the adult Ascidian has been indicated above. t In spite of these great differences, the presence of the endostyle and the perforated wall of the pharynx in the adult, and above all the features in the embryonic and larval development, entitle the Ascidians to be defined as more or less Amphioxus-like creatures which have become adapted to a sessile habit of existence. DEVELOPMENT OF ASCIDIANS. The first accurate and detailed account of the embryonic development of Ascidians was the classical memoir pub- lished in 1867 by Kowa.evsxky in the Mémoires de l Académie impériale des Sciences de St. Pétersbourg. ANATOMY AND DEVELOPMENT. 197 The Ascidian larva was known long before this time, and the external features of its metamorphosis were de- scribed in 1828 jointly by AupouIN and MILNE-Epwarps, to whom the discovery of the free-swimming larva is due. Furthermore, the internal structure of the tailed larva, and even the histological structure of the axial rod of the tail, was described with some accuracy by KRouN in 1852, but in ignorance of the details of the embryonic devel- opment, he was unable to give the right morphological interpretation to the various parts, and did not identify the axial rod with the notochord of the higher forms. Segmentation and Gastrulation. The segmentation of the egg, the formation of a hollow one-cell-layered blastula, and the flattening and_ subse- quent invagination of one side of the blastula to form the two-cell-layered gastrula, take place on a plan so essentially similar to what has been described above for Amphioxus that it is not necessary to dwell at length upon them here. Suffice it to point out that the segmen- tation of the Ascidian egg takes place typically, according to VAN BENEDEN and JULIN, on a strictly bilateral plan. That is to say, when the ovum has divided into two blastomeres, right and left, each blastomere represents and will give rise to the corresponding half of the larval body, and the descendants of the first two blastomeres can be distinguished for a remarkably long time on each side of the middle line of the embryo, —a fact which is highly characteristic of Ascidian development. After the gastrula has begun to elongate, and the blas- topore has been narrowed down by the approximation of its lips to a small aperture situated at the posterior dorsal extremity of the embryo, the formation of the medullary plate occurs. 198 THE .ASCIDIANS, Formation of Medullary Tube and Notochord. Here, as in Amphioxus, the dorsal wall of the embryo flattens, while the ventral remains convex, and the ecto- dermic cells on the dorsal side become marked off from the rest by their larger size and columnar shape. The medullary plate extends nearly to the front end of the embryo, while posteriorly its cells form a ring round the blastopore. In the formation of the medullary tube, however, there is an important difference, and the Ascidian embryo con- forms in this point more to the mode of development Fig. 98.— Transverse sections through embryo of Clavelina Rissoana, to show mode of formation of medullary tube and mesoderm. (After DAVIDOFF.) A. Through anterior region of embryo, with medullary groove still open. 8. Through posterior region, with closed medullary tube. ch. Rudiment of notochord. ec. Ectoderm. ez, Endoderm, mes. Mesoderm. m.g. Medullary groove. m.¢, Medullary tube. which is typical of the higher Vertebrates than does Amphioxus. In the latter the medullary plate sinks bodily below the level of the surrounding ectoderm, which then grows over it. Subsequently while underneath the ectoderm the medullary plate assumes the form of a half-canal open towards the ectoderm, and eventually its margins come together and so form a complete tube. In the Ascidian embryo the overgrowth of the surround- ing ectoderm and the folding up of the margins of the ANATOMY AND DEVELOPMENT. 199 medullary plate occur simultaneously, so that when the latter has the form of a half-canal it is not closed over by a layer of ectoderm, but is open to the exterior (Fig. 98). At a somewhat later stage the two medullary folds meet together and fuse in the middle line (Fig. 98 4), and this, combined with a slight forward growth of the posterior lip of the blastopore, leads to the inclusion of the latter in the medullary tube, so that we arrive at the condition already de- scribed for Amphioxus, in which the nerve-tube opens in front to the exterior by the zewropore and behind into the ar- nteron t - chentero by he blasto Fig. 99. — A. Embryo of Phallusia mam- pore, which has now miata seen in optical section from above, to : show notochord. become converted into B, Section through: tail of older embryo the xeurenteric canal. of Phallusia mammillata. (After KOWALEV- : SKY.) Meanwhile the cells ch. Notochord. ec. Ectoderm. ez, Endo- derm. mes. Mesoderm. z¢. Medullary tube. forming the dorsal wall of the archenteron in its posterior two-thirds begin to gather themselves together to form the notochord (Figs. 98 and gg). The cells forming the notochord are at first arranged end to end (Fig. 99), and subsequently interlace in the manner described above for Amphioxus. Origin of Mesoderm. At about the same time in which the formation of the medullary tube and notochord is going on, the mesoderm begins to put in its appearance, and this is the first event in the development in which there is an important dif- 200 THE ASCIDIANS. ference between the Ascidian and Amphioxus. The mesoderm in the Ascidian embryo does not arise as a series of archenteric pouches, but is produced on each side by a solid proliferation of cells from the primitive endoderm which lines the archenteric cavity. This solid proliferation begins in the middle region of the embryo near the an- terior limit of the notochord, and extends backwards (Figs. 98 and 100). It takes place from the dorso-lateral cells of the endoderm, in a posi- tion corresponding to that at which the mesoblastic pouches of Amphioxus grow out from the archenteron. The mesoderm of the As- cidian embryo therefore Fig. 100.— Embryo of Clavelina Ris- agrees with that of the em- soana seen from above, to show the re- : ¥ lation of parts. (Simplified after van bryo of Amphioxus in being Se ane EN) derived from the primitive np. Neuropore. ez, Endoderm. ent.c. Enteric cavity. .f. Medullary tube. endoderm, but differs in be- mes. Mesodermic band. ch. Notochord. . : ec. Ectoderm. ing solid and unsegmented.* * For a recent and elaborate discussion of the origin of the mesoderm in the Ascidians see VON DAVIDOFF’s Untersuchungen zur Entwicklungsgeschichte der Distaplia magnilarva, etc., I. Allgemeine Entw. der Keimblitter. Mitth. Zool. Stat. Neapel, IX. 1891. pp. 533-651. As shown by van Beneden and Julin in Clavelina, the primary mesoderm of the Ascidian embryo can be detected at a much earlier stage of development than in Amphioxus. I have studied the origin of the mesoderm in Cynthia papillosa and found that the primary mesoderm cells are to be distinguished, by their poverty in food-yolk, from the remaining endoderm, at the commencement of gastrula- tion (at the so-called p/akula-stage). They occur in the form of a crescent round the posterior margin of the blastopore, and are carried in by the invagi- nation, and then increase in number by mitotic division. In Cynthia, these ANATOMY AND DEVELOPMENT. 201 We thus have two solid longitudinal mesodermic bands inserted between the ectoderm and endoderm. Anteriorly the mesodermic bands consist of several layers of cells super- imposed one above the other (Fig. 98), but farther back they consist of only one layer of cells. Both portions of the mesoderm—namely, the anterior two- or three-layered and the posterior one-layered portions —arise in continuity with one another, but they have different fates, the former eventually breaking up into loose cells which float about in the body-cavity and constitute the so-called mesenchyme, the latter, on the other hand, becoming converted into the musculature of the tail; whence the former is spoken of as the gastra/ and the latter as the caudal mesoderm. Outgrowth of Tart. In Amphioxus, at the stage corresponding to that of which we have been speaking — namely, when the embryo has an oval or sub-elliptical shape—it bursts through the vitelline membrane inside which it has already been rotat- ing for some time by means of the cilia of the ectoderm, and escapes into the open sea. This is not the case, however, with the Ascidian embryo. The latter is never ciliated externally, and it remains enclosed within the fol- licular membrane throughout the whole of the emédryonic period of development. After the stage in question, the growth in the length of the embryo is accompanied by a ventral curvature, owing to the confined space in which it is contained. Moreover, the increase in length is not due to a simple elongation of the entire body of the embryo, as is the case with Amphi- primary mesoderm cells appear to give rise almost exclusively to the caudal mesoderm, while the gastral mesoderm appears to be added in front by prolifera- tion from the primitive endoderm as described above. 202 THE ASCIDIANS. oxus, but it is merely due to the outgrowth of the tail from the body of the embryo (Fig. 101). The structures involved in the outgrowing tail are the dorsal nerve-tube, the notochord, the caudal mesoderm, which lies on each side of the notochord, and will give rise to the muscles of the tail, and finally a solid cord of endo- derm consisting of two rows of cells placed side by side below the notochord (Fig. 99 £). As soon as the tail begins to grow out, the neu- 53 ‘ss Sree cos SOW ea g SOR x a FS Sek @ renteric canal becomes ob- ld ee : oe literated, and shortly after- wards the anterior neuropore Fig. 101. — aes of Phallusia closes up temporarily. Ata mammillata in side view, to show com- ] 5 mencing outgrowth of tail. (After ater period, as we shall see, KOWALEVSKY.) S : reo ch. Notochord. ec. Ectoderm. ev. En- at ie Pe no NORTE to doderm. mes. Mesoderm; the cells in- the exterior, but into the dicated by dark outlines, beneath which 5 1 may be seen the notochord and caudal uccal tube. endoderm. 2.9. Neuropore. 2.¢. Medul- As the tail grows in lary tube. length, it becomes coiled round about the body of the embryo, attaining two or three times the length of the latter. The cord of endoderm cells in the tail of the Ascidian larva has been supposed to represent a rudimentary intes- tine homologous with the straight intestine of Amphioxus, the larval tail being on this view equivalent to the post-branchial portion of the trunk in Amphioxus. This view, however, is probably not correct, although there is something to be said in favour of it. The probability is that the tail of the Ascidian larva or tadpole, as it is often called, is an organ which has been specially elaborated in the course of its evolution for the particular benefit of the Ascidians, since (exclusive of the pelagic forms) it is their ANATOMY AND DEVELOPMENT. 203 sole organ of locomotion, and hence of transportation from place to place; this only being possible during the larval period. As arule, the larval phase of an Ascidian’s existence is a remarkably brief one, and there is on this account all the more need for an effective propelling organ, which will enable the larva to arrive at a suitable resting-place. In Amphioxus, as described above, locomotion is ef- fected by serpentine movements of the whole trunk in virtue of its muscle-segments, and there is therefore no need for a tail in addition; but there is, nevertheless, a short post-anal extension of the body, which alone can be regarded as the homologue of the tail of the Ascidian larva. In the latter (¢.g. Ciona, Phallusia, etc.) the muscles are entirely confined to the tail, none being formed in the body proper, until after the resorption of its caudal appendage. On the view which I am endeavouring to make clear, it follows that the tail of the Tunicate tadpole is of the same nature as that of the Amphibian tadpole, and, in fact, of the craniate Vertebrates generally, and, as has just been said, is only represented by the short post-anal section of the trunk in Amphioxus. The solid cord of endoderm in the tail is not, therefore, a rudiment of a primitive intestine, but it is analogous to, even if not, as first suggested by BaLrour, homologous with, the so-called post-anal gut which occurs in the em- bryos of the higher Vertebrates, and bears a similar rela- tion to the formation of the tail that the endoderm-cord in the Ascidian embryo does. Thus in the typical Ascidian embryo the elongation of the trunk (body proper) does not take place to any consid- erable extent during the embryonic or even larval period, but only after the metamorphosis. 204. THE ASCIDIANS. With the formation of the tail the enteric cavity be- comes confined as a closed sac to the anterior portion of the embryo. It is bounded dorsally by the nerve-tube, which is somewhat dilated in this region, and in front, at the sides and below, it is in close contiguity with the ectoderm. Formation of the Adhesive Papille. At a much later stage than that represented in Fig. Ior, the ectoderm bounding the convex anterior extremity of the body becomes raised up into three prominences, whose relations to one another are those of the corners of a tri- angle. They are due to the ectodermic cells at the respec- tive points assuming a high columnar shape. They become eventually raised very much above the adjoining surface of the ectoderm, and become the adhesive papille or fixing glands of the larva. The cells composing them acquire the power of secreting a viscid substance, by which the larva can fix itself to any favourable surface (Fig. 102). Cerebral Vesicle and its Sense-organs. We have spoken above of the dilated anterior portion of the nerve-tube. This is the part of the central nervous system which undergoes the most striking subsequent changes. By a gradual widening of its cavity, accom- panied by a local thinning out of its wall, this portion of the neural tube lying in front of the notochord becomes transformed into a spacious sub-spherical vesicle, known as the cerebral vesicle (Fig. 102). While the anterior portion of the neural tube is enlarg- ing to form the cerebral vesicle, granules of black pigment are deposited by certain cells in the dorsal wall of the vesicle. The granules are at first scattered about in the ANATOMY AND DEVELOPMENT. interior of the cells. 205 The most anterior of the cells con- taining the pigment is at first distinguished from the others solely on account of the fact that the pig- ment-granules which it contains are somewhat larger than those in the succeeding cells, (Cf. Fig. 103.) Later on, however, the first pigmented cell is seen to separate itself eret.c Fig. 102. — Embryo of Ascidia mentula shortly before hatching; from the right side. (After WILLEY.) ch. Notochord, undergoing vacuolisation. e. Eye. ent.c. Enteric cavity. Adhesive papilla. #7. Anterior portion of nerve-tube (spinal cord). 0. Otocyst, lying on the floor of the cerebral vesicle and projecting up freely into its cavity. 7.a. Right atrial involu- tion. s#, Stomodcoeum, from the others, and it becomes gradually trans- ferred by a differential growth of the wall of the vesicle down the right wall to its final position in the ventral wall of the vesicle (Figs. 102, 103). This cell is the ofocyst, and the pigment-granules become consolidated together to form the ofo/ith. The latter is apparently Fig. 103.— Optical sections through cerebral vesicle of embryos of Ascidia mentula, to show mode of origin of eye and otocyst. (After WILLEY.) e. Eye. o. Otocyst. extruded from the cell (otocyst) in which it was originally formed, and the latter assumes a cup-shape, in the hollow of which the otolith lies. The two structures together form the so-called auditory organ, whose function may be not so much of an auditory nature as that of an equilibrat- ing apparatus. 206 THE ASCIDIANS. The other pigment-cells of the dorsal wall of the cerebral vesicle collect themselves together and form a slight pro- tuberance in the right dorso-lateral corner of the vesicle, while the pigment-granules, which were at first scattered about in the interior of the cells, become concentrated at their converging extremities towards the cavity of the vesicle. And in this way is formed the single eye of the Ascidian tadpole; the original pigment-producing cells constitute the vetzza, which retains its primitive position as part of the epithelial wall of the brain.* Subsequently two or three cells from the adjoining wall of the vesicle take up a position, one above the other, in front of the mass of pigment and, having previously, by an alteration in the character of their protoplasmic con- tents, acquired a high refractive index, constitute the J/ens of the eye, which projects obliquely downwards into the cavity of the vesicles (Gh) Big, 105-45) The cerebral vesicle of the Ascidian tadpole is the un- doubted homologue of the corresponding, but less pro- nounced, structure in Amphioxus. It differs from the latter in lying wholly in front of the anterior extremity of the notochord, in possessing a more highly organised eye, provided with a cellular lens, and in the presence of an otocyst, which, as we have seen, is evolved from the same group of cells which gave rise to the eye. The eye of the Tunicate tadpole agrees fundamentally with the type of eye peculiar to the Vertebrates, in that the retina is derived from the wall of the brain. On this * The fact that the lens of the Tunicate eye as well as the retina and the otocyst arise by differentiation of one and the same epithelial layer of the primitive cerebral vesicle, has recently been described by SALENSKy for the larva of Distaplia, magnilarva. (W.SALENSKY. JMorphologische Studien an Tunicaten: TI. Ueber das Nervensystem der Larven u. Embryonen vor Distaplia magnilarva. Morph. Jahrb. XX. 1893. pp. 48-74.) ANATOMY AND DEVELOPMENT. 207 account it is called a myelonic eye. In the typical Inverte- brate eye, on the contrary, the retinal cells are differen- tiated from the external ectoderm. Comparison of Tuntcate Eye with the Pineal Eye. The Tunicate eye, however, differs essentially from the paired eyes of the craniate Vertebrates in that the lens, as well as the retina, is derived from the wall of the brain. The lens of the lateral eye of the Vertebrates is derived by an invagination of the external ectoderm, which meets and fits in with the retinal cup at the end of the optic vesicle. It is, therefore, an extremely interesting fact which was pointed out by BALDWIN SPENCER, that the Tunicate eye agrees, in respect of the origin of its lens, with the parietal or pineal eye of the Lacertilia, in which the lens is likewise derived from cells which form part of the wall of the: cerebral outgrowth which gives rise to the pineal body. The pineal body is another of those remarkable rudi- mentary structures whose constant presence in all groups of Vertebrates forms such an eminently characteristic feature of their organisation. It develops as a_ hollow median outgrowth from the dorsal wall of the brain (thalamencephalon), the distal extremity of which dilates into a vesicle and becomes separated from the proximal portion.* For a long time the pineal body was a persistent enigma * According to the most recent work on the subject the distal vesicle be- comes entirely constricted off from the primary epiphysial (pineal) outgrowth of the brain, and the parietal nerve does not represent the primitive connex- ion of the pineal eye with the roof of the brain, but it arises quite inde- pendently of the proximal portion of the epiphysis. See A. KLINCKOwsTROM, Bettrage zur Kenntniss des Parietulauges. Zoologische Jahrbiicher (Anat. Abth.), VII. 1893. pp. 249-280. 208 THE ASCIDIANS. and the subject of much speculation, one of the most cele- brated hypotheses with regard to its significance being that of DrscarTEs, who regarded it as the seat of the soul. More recently it has been shown to represent a rudi- mentary, unpaired eye. Although in most cases, curiously enough, it exhibits in existing forms no trace of an eye- structure, it has been shown by DE GRAAF and SPENCER that, as a matter of fact, in many lizards the distal vesicle does actually become converted into an eye which, though of a rudimentary character, is possessed of a retina, pig- ment, and lens. In these forms the pineal body pierces the roof of the cranium, occasioning the parietal foramen, which is so characteristic of the Lacertilian skull, and the pineal eye lies outside the cranium immediately below the skin, through which it can be distinguished in external view by the presence of a modified scale placed above it. In the animals below the lizards in the scale of organi- sation (Amphibians and Fishes), as well as in those above them, the distal vesicle of the pineal body apparently does not become so far differentiated as to be recognised as an actual eye, except in the case of the Cyclostome fishes, where, as shown by BEarpD, it presents the three essential elements of an eye; namely, retina, pigment, and lens, lying, however, inside the cartilaginous cranium. The facts in our possession would seem to indicate that the remote ancestors of the Vertebrates possessed a median, unpaired, myelonic eye, which was subsequently replaced in function by the evolution of the paired eyes. It would, however, be premature either to assert this or to express it as a definite opinion, especially since, in refer- ring to the evolution of the paired eyes of Vertebrates, we are bordering on ground upon which I have no imme- ANATOMY AND DEVELOPMENT. 209 diate intention of treading. The pineal eye may not have been primitively so much an organ of vision as a light- perceiving organ, as is no doubt the case with the eye of the Tunicate tadpole. We may at least conclude that there can be no doubt that the Tunicate eye is the functional homologue of the pineal eye of the higher Vertebrates, as Spencer sug- gested. Stomodeal and Atrial [Involutions. By the time that the cerebral vesicle of the Ascidian embryo with its contained sense-organs (eye and otocyst) is approaching the completion of its full development, no less than three ectodermic invaginations occur in the body of the embryo. One of these is situated immediately in front of and in contact with the anterior wall of the cere- bral vesicle, the blind end of the involution pressing against the subjacent endoderm. This is the stomodeum., and its formation is preliminary to the perforation of the mouth which takes place later, and places the stomodceum in open communication with the portion of the enteric cavity which will become the branchial sac (Fig. 102). It should be emphatically noted that the stomodceal invagi- nation occurs in the dorsal middle line immediately adja- cent to the anterior extremity of the central nervous system. The other two ectodermic invaginations occur symmetri- cally, one to the right and the other to the left of the dorsal middle line, behind the region of the cerebral vesicle, and constitute the pair of atrial involutions, which, by their subsequent growth and modification, give rise to the atrial or peribranchial cavity. We see, therefore, that the epi- thelium which forms the lining membrane of this cavity is, as in Amphioxus, derived from the external ectoderm. 210 THE ASCIDIANS. For some considerable time after the metamorphosis the young Ascidian possesses two separate atrial cavities, right and left, each opening to the exterior by its own atrial aperture. Eventually the two cavities extend round the branchial sac dorsally, so that their walls come into contact in the dorsal middle line, and finally the dividing line breaks down, and they become continuous one with another dorsally, remaining separated ventrally, as described above. At the same time that the two atrial cavities grow towards one another, their external apertures become in- volved in the same process of growth, and, moving together, finally fuse in the dorsal middle line, and so form the single atrial or cloacal aperture of the adult.* Beyond agreeing in its ectodermal origin, there might appear to be not much in common between the mode of development of the atrial cavity in the Ascidians and in Amphioxus. No morphologist would recognise a fundamental differ- ence in the fact that the right and left halves of the atrial cavity in Amphioxus arise by a single median involution of the ectoderm, instead of from a pair of involutions, and that they are from the first continuous with one another instead of becoming so secondarily (Fig. 104). In like manner, the fact that the two halves of the atrial cavity are continuous with one another ventrally in Amphi- oxus and dorsally in the Ascidians, is easily brought into correlation with the other differences in the organisation of the two types, which have been described above, and is no bar to our regarding the atrial cavity of the one as being homologous with that of the other. * The time at which the atrial cavities fuse together varies very much in different genera. In Molgula manhattensis, for instance, whose stigmata develop on a similar plan to those of Ciona (see below), there is a single atrial aperture at the moment of the metamorphosis. ANATOMY AND DEVELOPMENT. 201 One feature in connexion with the formation of the atrial cavity, in which the Ascidians stand in marked contrast to Amphioxus, does, however, require a special explanation. Whereas in Amphioxus the atrial involution has the form of a longitudinal groove, in the Ascidians it occurs on each side, as a local inpushing of the ectoderm with a minute circular orifice of invagination.” The fact has already been stated above that the elonga- tion of the body proper of an Ascidian embryo or larva does not, in the main, take place until after the metamorphosis. Fig. 104. — Diagrammatic transverse sections, to illustrate the mode of forma- tion of the atrium in (A) an Ascidian and (2) Amphioxus. (After WILLEY.) The atrial involutions occur at a time when the tail is rapidly increasing its length; the body proper, on the con- trary, remaining stationary so far as increase in size is concerned, and retaining at this stage approximately the dimensions which it possessed when the tail first began to grow out. Moreover, they occur defore the appearance of any gill-clefts in the wall of the branchial sac, so that in the Ascidians the gill-slits never open directly to the exterior. In Amphioxus, on the other hand, there is no such delay in the elongation of the body of the embryo, but it goes on continuously till the full complement of myotomes has been 212 THE ASCIDIANS. formed. The post-anal portion of the body, which we sup- pose to be the homologue of the tail of the Ascidian tad- pole, does not appear until a somewhat late period in the development. There is very little of it present in the larva with three gill-slits (Fig. 73). The reason of this, as explained above, is that the post- anal section of the trunk is of only minor functional sig- nificance in Amphioxus, but is all-important to the Ascidian larva, and consequently, as is the case with many other structures of great functional importance in the various groups of the animal kingdom, it exhibits a precocious development. Not only, therefore, has the elongation of the body of Amphioxus already taken place before the occurrence of the atrial involution, but the primary gill-slits have also broken through the wall of the pharynx, and open freely to the exterior before the atrium begins to be closed in. In Amphioxus, then, the atrial involution has been drawn out into the form of a longitudinal groove because it occurs subsequently to the elongation of the body and the perforation of the gill-slits. In the Ascidian embryo the (paired) atrial involution has the form of a simple pit with a circular margin, be- cause it arises before the elongation of the body proper of the embryo and before the perforation of the gill-clefts, so that no influence has been at work to draw it out into the form of a groove. We see, therefore, that a great many of the differences between the Ascidian tadpole and the larva of Amphi- oxus can be explained sufficiently to allow of their being brought into genetic relation with one another, by consid- ering the relative time at which corresponding develop- mental processes take place in the two cases. ANATOMY AND DEVELOPMENT. 213 The following table will help to make this matter clearer. ORDER OF ASCIDIAN. AMPHIOXUS. OccuURRENCE. It Gastrulation. Gastrulation. 2s Oval embryo with medullary | Oval embryo with medullary tube, neurenteric canal, tube, neurenteric canal, notochord, and mesoblast. | notochord, and mesoblast. (Last two commencing. ) (Last two commencing.) Be Outgrowth of tail. Commencing elongation of body of embryo, and escape from vitelline membrane. 4. Continued growth of tail. Continued elongation of em- bryo. ie Formation of stomodceum and | Formation of mouth, and com- atrial involutions. mencing perforation of gill- clefts. 6. Escape from vitelline mem- | Continued formation of gill- brane. clefts and outgrowth of tail (z.e. post-anal section of trunk). le Commencing perforation of | Formation of longitudinal atrial gill-clefts. involution. 8. Metamorphosis and commenc- | Metamorphosis. ing elongation of body proper. Of course the above table has no concern with the actual time (hours and days) from the commencement of the development at which such and such an event occurs. The type of Ascidian referred to in the above description is a simple Ascidian like Czova or Phallusia. The above table also shows how the development of the Ascidian and of Amphioxus moves along parallel lines up to a certain point, and then at the time of the outgrowth of the tail in the embryo of the former and the hatching of the embryo of the latter, divergences set in. 214 THE ASCIDIANS. It has long been recognised that the development of an Ascidian is much abbreviated in comparison with that of Amphioxus, since in the former it neither comes to the formation of a ciliated embryo nor to the production of archenteric pouches for the mesoderm. One of the chief evidences, however, of abbreviation in the Ascidian devel- opment is the precocious formation of the larval tail. Formation of Alimentary Canal and Hatching of Larva. When the enteric cavity of the Ascidian embryo begins to grow in length so as to give rise to the stomach and intestine, which it does shortly after the appearance of the atrial involutions, there is only one resource open to it on account of the limited space in which it lies, and that is to double round upon itself. This it accordingly does. As the growth progresses, the posterior dorsal angle of the enteric cavity bends sharply downwards on the right side, and then upwards and slightly forwards on the left side, ending at first blindly in the vicinity of the left atrial sac. In this way the four divisions of the alimentary canal become established ; namely, pharynx or branchial sac, cesophagus, stomach, and intestine. (Cf. Fig. 105.) By the time these changes have taken place, the embry- onic development is at an end, and the larva is ready to hatch. By spasmodic jerkings of its tail, the larva finally succeeds in bursting the egg-follicle or vitelline membrane in which it has been hitherto enclosed, and so escapes into the open sea. Clavelina and Ciona. While the development of most forms of Tunicata is re- ducible to a common type, yet the details vary within very wide limits in different genera. The tendency here, as ANATOMY AND DEVELOPMENT. 215 elsewhere, is to abbreviate the development by omitting certain ontogenetic processes, and so arriving at the de- sired end, as it were, by a short cut. One of the most impressive instances of such an abbre- viated development, and one which can be demonstrated with the utmost certainty, is afforded by the genus Clave- fina, in contrasting it with the closely allied genus Czona. Clavelina (see Fig. 96) is an Ascidian, provided at its base with creeping processes or stolons containing a lumen continuous with the body-cavity, by which it adheres to rocks and weeds. Buds are formed from the stolon, which grow up into new individuals precisely like the parent form which developed from the egg, and soa colony is produced. Ciona also has similar basal processes of the test, con- taining prolongations of the original body-cavity, but no buds are produced. In Clavelina, the embryonic development, up to the time of the hatching of the larva, takes place inside the peri- branchial chamber of the parent, which becomes converted into a kind of brood-pouch. In Ciona, the eggs are extruded into the water, where they are fertilised by the simultaneous extrusion of sper- matozoa from the same individual. Finally, in Clavelina the egg is much larger and contains more food-yolk than that of Ciona. We see, therefore, that in these two genera the egg is at the outset subjected to different sets of conditions, both internally and externally. METAMORPHOSIS OF CIONA INTESTINALIS. Three stages in the metamorphosis of the larva of Czona entestinalis are shown in Fig. 105. First, there is the free- swimming larva, which, after a pelagic existence of one or 216 THE ASCIDIANS. perhaps two days’ duration, is on the point of fixing itself to a foreign object by means of the sticky secretion of its three adhering papille. This larva possesses features which we have not yet considered. Let us give our attention in the first place to the tail. Vacuoltsation of the Notochord. The vacuolisation of the notochordal tissue, which was described above for Amphioxus, has already proceeded to such an extent that there is no longer any trace of cellu- lar structure in the centre of the notochord. It is entirely filled with a perfectly colourless substance, probably of gelatinous consistency, while the nuclei have been dis- placed entirely from the centre and can be seen to lie closely pressed against the dorsal and ventral sides of the sheathing membrane of the notochord (Fig. 105 A). There is one respect in which the above vacuolisation of the cells of the notochord differs considerably from the corresponding process in Amphioxus and the higher Vertebrates. Whereas in the latter forms the vacuoles appear inside the individual cells, —in other words, are zztrace//ular, — in the Ascidian tadpole they occur between the cells, and are therefore zztercellular. This was first made out by Kowalevsky, and can readily be observed. (Cf. Fig. 102.) The intercellular spaces separate the cells which were previously fitted accurately together, end to end, and, gradually increasing in size, they eventually flow together and so constitute a continuous space, while the cells with their nuclei become thrust aside. Assuming that the vacuoles contain a more or less fluid substance secreted by the protoplasm of the cells, the ANATOMY AND DEVELOPMENT. 217 above difference in the vacuolisation of the notochordal tissue in Amphioxus and the Ascidian larva would resolve itself into saying that the secretion was retained inside the cells in the one case, and deposited outside them in the other. Mesenchyme and Body-cavity. The endoderm cells of the tail, which formed at first a solid cord below the notochord, have now become con- verted into loose corpuscles, which have mostly floated out of the tail into the hinder portion of the body-cavity, and have become indistinguishable from the mesoderm- cells. The latter are beginning to lose their compact dis- position in the form of the two mesodermic bands, espe- cially in the hinder region, and to be scattered about in the body-cavity. The body-cavity of the young Ascidian is not unre- servedly homologous with that of Amphioxus, on account of this remarkable behaviour of the mesoderm. The cavity does not arise in the midst of the mesoderm by a splitting apart of its component cells, but it is simply produced by a separation of the endoderm from the ecto- derm, the two layers being at first in contact at the sides and below; in fact, everywhere, except where the dorsal nerve-tube intervenes. In the cavity thus produced between ectoderm and endoderm the mesodermic bands at first lie freely, and then their component cells break away from their compact association and float about the cavity in the form of scattered corpuscles, known collectively as mesenchyme. This mesenchyme later gives origin to the muscula- ture of the body proper of the Ascidian, and also to the definitive blood-corpuscles, genital organs, and renal 218 THE ASC/DIANS. vesicles.* All these structures are differentiated from the loose mesenchyme cells, all of which at first course round about the body of the young Ascidian like blood, being kept in motion by the beating of the heart. In the stage shown in Fig. 105 A the mesodermic bands are still fairly compact in front, having extended them- selves anteriorly at the sides of the enteron by interstitial growth. Preoral Body-cavity and Preoral Lobe. When the larva first hatches, the endoderm and ecto- derm are in contact with one another at the anterior extremity of the body, just as they are in the earlier stages. (Cf. Fig. 102.) Soon, however, the ectoderm, with the adhering papille, springs away from the endo- derm at this point, leaving a space into which the two lateral mesodermic bands force their way. In this way a special anterior portion of the body-cavity, preoral and preenteric, is produced, and is at first com- pletely filled by a compact mass of rounded cells derived from the mesodermic bands. — The end of the body of the larva at which the adhering papilla are placed of course corresponds to the tip of the snout in Amphioxus. Just as Amphioxus burrows into the sand with its snout, so the Ascidian larva fixes itself to the surface of a rock or weed by its snout. The anterior or przoral portion of the body-cavity, of which we have just traced the origin, is, and subsequently becomes in a still more pronounced way, the cavity of the snout, or preoral lobe. * The pericardium arises ventrally from the endodermic wall of the bran- chial sac, and the heart is formed by an infolding of the dorsal wall of the pericardium. ANATOMY AND DEVELOPMENT. 219 50 99° 0 7 > 10% % 32% ' - ——— ot. : ‘a She EN % 1 } ay ed Fig. 105.— Metamorphosis of Czona intestinalis; above is represented the anterior portion of the free-swimming larva from the left side; on the left, the larva, shortly after fixation, from the right side; and on the right, the stage at which the change of axis commences, from the left side. (After WILLEY.) a. Atrial aperture. 4. Branchial sac. ch. Notochord. e. Endostyle. # Organ of fixation. g. Ganglion. %. Neuropore (having reopened into branchial sac). z, Intestine. 72 Pyloric gland. mm. Mouth. 2. Nerve-tube. oe, (Esophagus. ob. Eye. of. Otocyst. g. Pericardium. s.Stomach, s¢. Stigmata. 4 Tail. 220 LHE ASCIDIANS. Body-cavity of an Ascidian and Celom of Amphioxus. We must now endeavour to show how the body-cavity of the Ascidian can be brought into genetic relationship with the cclom of Amphioxus. The question of the absence of metamerism in connexion with the origin of mesoblast in the Ascidians need not detain us, since it is so obviously correlated with their mode of life. It may safely be asserted that the Ascidian mesoderm, as a whole, is homologous with that of Amphioxus as a whole, but in the details of its origin and fate it is widely different. If we figure to ourselves the coelomic epithelium of Amphioxus losing its character as a membrane and break- ing up into its constituent cells, which would then lie loosely in the body-cavity, we should have essentially the same condition of things as in the Ascidians. There are numer- ous precedents in the animal kingdom for such a disinte- gration of an epithelial membrane. A most perfect instance of it has been described by Dr. R. von ERLANGER* in connexion with the origin of the mesoderm in the fresh-water snail, Paludina vivipara. Here the mesoderm appears at first in the form of a median bilobed archenteric pouch of relatively large dimensions. Soon, however, the cells forming the wall of the pouch begin to assume irregular shapes, and so disturb the contour of the epithelium, and eventually they break apart entirely and fill every nook and corner of the available space with a loose mesenchyme. Similar out-wanderings of cells from an epithelial wall, though not often of such a complete character as the instance above cited, are by no means infrequent. * Zur Entwicklung der Paludina vivipara, Parts I. and II. Morpholo- gisches Jahrbuch, XVII. 1891. ANATOMY AND DEVELOPMENT. Zo A striking example is afforded by the body-cavity of the worm-like Balanoglossus, of which we shall speak later. Here, according to Bateson, the cells lining the cavity are continually budding off daughter-cells, which fall into the cavity, and eventually almost entirely fill it up with mesenchymatous tissue. In this case, therefore, mesen- chyme and an epithelial wall coexist. Similarly, the epzthelial sclerotome of Amphioxus is rep- resented by a mesenchymatous sclerotome in the higher Vertebrates. It is not necessary to multiply instances, but many others could be adduced. If, now, this disintegration of partetal and visceral layers of the mesoderm, which we have imagined above to take place in the ontogeny of an animal like Amphioxus, be supposed to be thrown back in the development, or, in other words, abbreviated to such an extent that the pre- liminary formation of a continuous ccelomic epithelium no longer takes place, we should have precisely those condi- tions which we actually find in existing Ascidians. As in the cases above quoted for purposes of illustra- tion, so in the Ascidians the mesenchymatous condition undoubtedly originated ancestrally from what we may call an epithelial condition. In the Ascidians we may conclude, therefore, that while ontogenetically the mesenchymatous condition is to all intents and purposes primary, from a phylogenetic point of view it is pre-eminently secondary or cenogenetic. Having made the reservations implied in the above statements, we may confidently assert that as a whole the body-cavity of the Ascidians is homologous with the coelom of Amphioxus, and we may define the former as a ccelom in which the cells, instead of associating together 222 THE ASCIDIANS. to form a lining membrane round the cavity, remain independent of one another and scattered about inside the cavity. Fixation of the Ascidian Larva. When the larva first fixes itself to some available surface, the tail remains for a time stretched straight out and almost motionless, giving perhaps an occasional twitch. Soon the tail is observed to become shorter and to finally disappear, having been drawn within the body proper of the young Ascidian. The entire tail, with the whole of the notochord, musculature, and caudal portion of nerve- tube, becomes thus retracted and invaginated into the posterior region of the body-cavity, wnere it forms a coiled amorphous mass, which goes through a gradual series of histolytic changes, and is finally absorbed by being dissolved in the fluid of the body-cavity (Fig. 105 2). By the time the tail has been completely drawn up into the body, the organ of fixation or snout, as we have called it above, becomes drawn out into a long probosciform structure in a line with the long axis of the body. Its cavity is no longer completely filled with mesoderm-cells as it was at first (Fig. 105 A), but it has become so volu- minous that its contained cells are loosely scattered about (Fig. 105 B). In the concluding chapter we shall endeav- our to show, what has been already implied, namely, that the organ of fixation is seen to the best possible advantage from a morphological point of view in the species now under consideration, viz. Czona intestinalis, and that it is homologous with the przoral lobe (snout) of Amphioxus, including under that term both the przoral body-cavity and the preoral pit, and further that it is homologous with the proboscis of Balanoglossus. ANATOMY AND DEVELOPMENT. 223 At the stage shown in Fig. 105 A, the lumen of the alimentary canal is extremely reduced, and in many places, as in the region of the endostyle, ¢, its opposite walls are in actual apposition, so that the lumen at these points is almost obliterated. This temporary reduction of the lumen of the alimentary canal is due to the narrow space into which it has to be compressed, combined above all with the relatively enor- mous size of the cerebral vesicle, which exercises a great pressure on the subjacent dorsal wall of the branchial sac. It may be added that the larva of Ciona does not take in food independently until after fixation. Reopening of Neuropore; Degeneration of Cerebral Vesicle ; Formation of Definitive Ganglion. One of the most obvious features of the metamorphosis is the rapid expansion undergone by the enteric and body cavities and the no less rapid degeneration of the cerebral vesicle. This expansion, by relieving the crowded char- acter of the various parts, facilitates greatly the study of the changes which take place in the internal organisation. The neuropore, which we have described above as having closed up at an early period, now reopens again and places the neural tube —that is to say, as much of it as remains after the atrophy of the tail—in open communication with the base of the buccal tube (Fig. 105 4, 7). The spacious cavity of the cerebral vesicle has vanished, and its walls have undergone disintegration, and, except for a portion of the dorsal wall which becomes converted into another channel, are now represented by a mass of histolytic residua filling the original cavity of the vesicle and lying below the anterior portion of the nerve-tube. pogpHyal THE ASCIDIANS. This remnant of the cerebral vesicle of the larva with its sense-organs becomes eventually absorbed, and the eye and otolith may often be found floating about the body-cavity with the ordinary mesenchyme-cells, and occasionally they can be seen actually passing through the heart. The anterior portion of the nerve-tube itself, which now opens into the base of the buccal tube or stomodceum,* is derived from a portion of the dorsal wall of the original cerebral vesicle which was constricted off from the latter in the form of a narrow tube slightly to the left of the mid- dorsal line (Fig. 105 B, 7). Subsequently the cells forming the dorsal wall of this portion of the nerve-tube proliferate and form a solid thickening which becomes the definitive ganglion of the adult (Figs. 105 C, 106, and 107, g). The lumen of the nerve-tube behind the region of the definitive ganglion finally becomes obliterated by the mutual approximation of its constituent cells, and that portion of the primitive nerve-tube which in the larva lay between the cerebral vesicle and the root of the tail is thus represented in the adult by a solid “cordon ganglionnatre viscéral”’ (van Beneden and Julin) which starts from the posterior end of the adult cerebral ganglion, and, proceed- ing along the dorsal side of the pharynx above the dorsal lamina, becomes lost among the viscera. (Cf. Figs. 96, 105, and 107.) Below and in front of the definitive ganglion, which finally becomes quite separate from the dorsal wall of the neural tube, the lumen of the latter persists and becomes * According to renewed observations on Ciona, I find that the neuropore reopens into the buccal tube precisely in the line of junction of the stomo- dceum with the wall of the branchial sac, so that its upper margin is continu- ous with the (ectodermic) stomodceal epithelium, and its lower margin with the (endodermic) branchial epithelium. (See below, V.) ANATOMY AND DEVELOPMENT. 225 by subsequent extension the lumen of the subneural gland and its duct. Thus the anterior portion of the primitive neural tube, having become constricted off from the cerebral vesicle of the larva, and having given rise by proliferation from its dorsal wall to the definitive ganglion, becomes bodily converted into that structure which we shall call, in agree- ment with JuLin, the “ypophysts. The opening of the latter into the base of the buccal tube becomes the dorsal tubercle of the adult. Finally, at a much later stage, the glandular portion of the hypophy- sis arises by proliferation of spongy tissue from the ven- tral wall of that portion of the xeuro-hypophysial tube which lies immediately be- low the ganglion. A section through the cerebral vesicle of a larva of Distaplia, a colony-build- ing Ascidian, showing the hypophysis in process of being constricted off from the vesicle, is given in Fig. 100. dition of things generally is In this genus the con- very different from what obtains in Ciona, but it is introduced to show the Fig. 106. — Frontal section through cerebral vesicle of a larva of Distaplia magnilarva, to show the origin of the ganglion and hypophysis. (After HJORT; combination of two figures.) In the larva of Distaplia, the hypophy- sis opens into the branchial sac _ be- hind the stomodoeum. c.v. Cerebral vesicle. ec. Ectoderm. en. Endoderm. gg. Ganglion. Ay. Hy- pophysis (neuro-hypophysial tube). essential similarity in the mode of origin of the hypophy- sis in this form, as observed by Dr. JoHAN Hyjorv. In Distaplia, as is also the case to a less extent in Clavelina, the ganglion begins to develop from the wall 226 THE ASCIDIANS. of the neuro-hypophysial tube while the latter is still in connexion with, and therefore before the atrophy of, the cerebral vesicle, thus indicating a hastening in the devel- opment as compared with Czona. The convexity caused in the dorsal wall of the branchial sac by the pressure of the cerebral vesicle persists as the anterior portion of the dorsal lamina, and in many or most simple Ascidians becomes grooved, forming the efzbran- chial groove of JULIN (Fig. 97). At present it is merely a ridge, the epzbranchial ridge. In Fig. 105 C the proximal (oral) end of the endostyle, eé, is seen to be connected with the epibranchial ridge by the peripharyngeal band, which we have already described in the adult. It apparently arises 27 sztw by simple spe- cialisation of the cells forming the epithelial wall of the pharynx at this point. Primary Topographical Relations and Change of Axis. It must be especially noted that the long axis of the young Ciona for some time after fixation is identical with that of the tailed larva, and therefore the primary topo- graphical relations of the various parts are maintained at the stage shown in Fig. 105 4, and we can accordingly make use of this stage in which different structures are much clearer than in the free-swimming larva for the purpose of describing the primary topography, which is of the utmost importance when it is desired to institute a comparison with Amphioxus. Since, as we have seen, the details of the embryogenetic processes differ in many respects widely from what occurs in Amphioxus, we are inevitably compelled to rely to a very large extent on topographical relations in order to estimate the homology of this or that structure in the ANATOMY AND DEVELOPMENT. 227 Ascidians and in Amphioxus. Fortunately there is one structure as to whose complete homology, in the Urochorda (Tunicata), on the one hand, and the Cephalochorda, on the other, no one entertains a doubt, and that is the exdostyle. We thus have in the endostyle a firm basis upon which to ground our deductions. In the larva and in the young Ascidian before the primary long axis has been disturbed in the way which we shall shortly describe, the endostyle is the most anterior endodermic structure in the body, and lies dorso-ventrally at right angles to the long axis of the body (Fig. 105 A and B, e). As described above in the larvz of Amphioxus, particu- larly in the younger larvae (see Figs. 64 and 73), the endo- style, though lying asymmetrically on the right side, being involved in the general asymmetry of the larva, is quite anterior in position, in front of all the gill-slits and partly in front of, though also partly opposite, the mouth (on account of its asymmetry), and almost at right angles (see especially Fig. 64) to the long axis of the body. As there is only a short stretch of simple endoderm in front of the endostyle in the larva of Amphioxus, we may describe it as the most anterior differentiated endodermic structure in the larva, thus corresponding with remarkable precision to the condition described above in the larval and newly fixed Ascidian. In the middle of the wall of the branchial sac in Fig. 105 4 are seen, somewhat in front of and below the atrial aperture, a, of this side, two lens-shaped structures whose slightly concave sides face each other. These are the borders of the two first-formed primary branchial stigmata or gill-clefts. Their actual openings into the atrial chamber are at present so small that they can hardly be seen in 228 THE ASCIDIANS. surface-view, but they are situated at the inner or con- cave sides of the two thickenings. Fig. 107. — Young Ciona intestinalis after the completion of the change of axis; from the left side. (After WILLEY.) 7, 7V. Primary stigmata. a. Anus, situated immediately below the left atrial aperture. end. Endostyle. ££ Organ of fixation. .. Ganglion. hy. Hypophysis. 7zw¢. Intestine. Zaz, Left atrial aperture. /.. Longitudinal muscle. 2. Mouth. oes. CXsophagus. 7.6. Peripharyngeal band. py. Pyloric gland. st. Stomach. ¢ Coronary tentacles. v.7. Visceral nerve (cordon ganglion- naire viscéral). On either side of the latter can be seen the ordinary cavity of the pharynx proceeding to- wards the cesophagus. At a later stage the openings of the two first-formed stigmata become distinctly visi- ble(Fig. 105 C). Mean- while a change of axis is taking place in the body of the Ascidian. During the extraor- young dinary change of axis which we are about to describe the probos- lobe (snout, organ of fixa- ciform przoral tion) remains station- ary, and the rest of the body actually rotates through an angle of 90 degrees, using the or- gan of fixation as a pivot about which it turns. Invbigs Tos G the which takes place very gradu- rotation ally is only half performed; while in Fig. 107 it is complete. The method of growth by which this rotation takes place ANATOMY AND DEVELOPMENT. 229 is of a very singular character, and it is difficult to define it in precise terms. In this way then the endostyle (and branchial sac generally) comes to be placed at right angles to its primary position. Since in Amphioxus the endostyle altered its primary axis by a process of independent growth while the long axis of the pharynx was constant throughout the develop- ment, we find that here again, as in so many previous instances, the details by which similar end-results are arrived at are widely dissimilar. This complete change of axis by which the przoral lobe (organ of fixation) becomes placed at the posterior extrem- ity of the body can only be regarded as a cenogenetic feature.* It is therefore chiefly to the primary relations which the various structures bear to one another, before the change of axis, that we must turn for purposes of comparison. If we do this, we find that the following sequence of organs obtains as well in the larva of Amphioxus as in the newly fixed larva of Ciona; namely: 1, praeoral lobe; 2, endo- style ; 3, mouth; 4, gill-clefts. Formation of Additional Branchial Stigmata. After the change of axis of the body, the long axes of the stigmata lie transversely. In their further growth they go on elongating in the same (transverse) direction, and after they have attained a certain size their ventral ends — that is to say, the ends nearest the endostyle— bend round towards each other, and from each of the two first- * Tt goes without saying that the primary long axis of the Ascidian larva is homologous with the long axis of Amphioxus. 230 THE ASCIDIANS. formed stigmata a minute portion becomes gradually con- stricted or nipped off. Thus between and cut off from the two original stigmata, there come to lie two intermediate stigmata of much smaller size. (Cf. Fig. 107.) In this way, then, in Ciona, we arrive at the stage with four branchial stigmata on each side of the pharynx. For convenience we shall refer to these by the Roman nu- merals, I., 11, Til“andihV. Wts a remarkable fact that II. and III. do not arise by new perforations, but are cut off from I. and IV. respectively. On account of the close relations which the two first- formed stigmata, I. and IV., bear to one another during the production of the intermediate stigmata, their ventral extremities coming into contact and apparently some- times fusing together so that II. and III. might almost be described as a joint production of I. and IV. rather than as entirely independent offshoots, one is forced to the conclusion that the two first-formed stigmata themselves, though they actually appear simultaneously as separate perforations, in reality represent the two halves of a single primitive gill-slit divided into two by a tongue- bar. If, moreover, we examine the exact origin of these two stigmata (I. and IV.) by means of transverse and horizontal sections, we may become convinced that such is indeed the case; namely, that they represent the two halves of a primitive gill-slit which, on account of the precocious formation of the tongue-bar between them, become perforated separately. For the formation of any two or more consecutive gill- slits, we usually expect to find separate endodermic pockets or pouches of greater or less depth growing out towards the ectoderm. (Cf. Figs. 72 and 92.) We ought to find something analogous to this in Ciona ANATOMY AND DEVELOPMENT. 231 if the two first-formed stigmata had the value of indepen- dent gill-slits. Instead, however, of anything approaching to two endo- dermic outgrowths, we find at the base of the atrial invo- lution a single endodermic ingrowth making its appearance (Fig. 108). The angles made by this ingrowth with the neighbour- ing wall of the branchial sac remain in contact with the floor of the atrium, then fuse with it, and finally become a A os at MALLILILSELLIELULLIMLILIUIEUA Ey | pp t Uy 19) UN LLL LL YW WLLL 2b at Sa C D i, LLL LLL ILL. PE TILL ped STE Ys 75 Fig. 108. — Diagrams illustrating the mode of origin of the two first-formed branchial stigmata in Ciona. (After WILLEY.) at, Atrial involution. ec. Ectoderm. em. Endoderm. g.s. Stigmata. 726. ‘Tongue-bar. perforated (Fig. 108). This is the way in which the stig- mata, I. and IV., arise, and it is difficult, if not impossible, to interpret the above-mentioned endodermic ingrowth otherwise than as a precocious tongue-bar. Even in Amphioxus it was seen how the tongue-bars of the secondary slits arose relatively much earlier than those of the primary slits. If they arose still a trifle earlier, we should have the two halves of each slit becom- ing separately perforated, just as it happens in Ciona. In a species of Balanoglossus an analogous precocious 232 THE ASCIDIANS. formation of tongue-bars, before the perforation of the slits, has been described by Professor T. H. Morean. From what has been said above, we conclude that the first four pairs of primary branchial stigmata of Ciona (and this probably applies equally to many species of Phallusia) represent and are derivatives of one pair of primitive, ancestral gill-slits. After a comparatively long interval, during which the intermediate stigmata, II. and III., increase in length transversely, two more stigmata, V. and VL., arise at inter- vals, one after the other, by sepa- rate perforations behind those already formed (Fig. 109). On account of the independent origin of V. and VI., it might be supposed that they would have the morphological value of dis- tinct gill-slits, and that we had before us three pairs of ancestral Fig. 109.— Primary branchial gill-slits represented by six pairs sigmata sof the might isidevet a) of: | primany, joranchialstiematas young Ciona. (After WILLEY.) For this interpretation to hold good, we should expect to find that in other forms in which six primary branchial stigmata were produced, their origin was either the same or reducible to the same type as that of the branchial stigmata of Ciona. This, however, is not the case, since I have found that in Molgula manhattensis,* a simple Ascidian which occurs in great numbers at New Bedford, Mass., the six primary stigmata, corresponding precisely to those in * My observations on the development of Molgula manhattensis were made at the Marine Biological Laboratory, at Woods Holl, Mass., in the summer of 1893. ANATOMY AND DEVELOPMENT. 233 Ciona, have a somewhat different mode of origin. The two first-formed stigmata (=I. and IV. in Ciona) appear simultaneously as in Ciona. Then after growing to a cer- tain size, they curve round at their ventral ends, not in opposite directions so as to meet each other as they do in Ciona, but in the same direction (Fig. 110). The recurved ends then become constricted off from the parent stig- mata. Later on, a fifth gill-opening arises behind the first four stigmata by independent perforation, and after Fig. 110. — Diagram illustrating the mode of origin of the six primary bran- chial stigmata of Molgula manhattensis. The numbers are placed at the ventral ends of the slits. The figure is a combination of several hitherto unpublished drawings of different stages in the development. /, ///, and V arose by separate perforation. attaining a certain size, it, in its turn, curves round at its ventral end, and eventually the sixth stigmatic opening is constricted off from the fifth. Since the first six primary stigmata have such different origins in two different species, it is obvious that in attempting to make a comparison with Amphioxus we can only use the two first-formed stigmata, because they agree in the above-mentioned species, and in many others in 234 THE ASCIDIANS. arising simultaneously, and in representing, in all proba- bility, the two halves of a primitive gill-slit, cut in two by a tongue-bar. The stigmata which are added to these must, therefore, be regarded as secondary modifications, hardly comparable to the successive formation of new gill-slits in Amphioxus. In the Ascidians, therefore, we can only detect the representatives of one pair of primitive gill-slits, and there is every reason for supposing them to be homologous with the first pair of gill-slits in Amphioxus as defined above. The six primary stigmata of each side give rise, by re- peated subdivision, to the innumerable stigmata of the adult, both in Ciona and Molgula. The following de- scription, however, applies more particularly to Ciona. In the first place, the primary stigmata grow to a sur- prising transverse length, and then commence to divide into two equal portions by small tongue-like projections, which grow across the aperture indifferently from the anterior or posterior walls of the respective stigmata, and, fusing with the opposite wall, divide the transversely elongated slit into two completely separated halves. Then each of the latter divides again in the same manner, and so the process of subdivision of existing stigmata goes on. In this way six transverse rows of stigmata arise. These may be distinguished as secondary stigmata, since they arise by division from the primary. Gradually, by a peculiar process of growth, the long axes of the secondary stigmata change their direction, and instead of lying transversely they become directed antero- posteriorly. This is their definitive position, and the stigmata now go on rapidly dividing again, and the num- ber of transverse rows of stigmata is in this way doubled, trebled, quadrupled, etc., and we thus arrive at the adult ANATOMY AND DEVELOPMENT. 235 condition. Out of the multitude of stigmata which are present in the adult Ciona only four arise by independent perforation; namely, the primary stigmata I. and IV. (which we regard as the two halves of a primitively single slit) and V. and VI. First Appearance of Musculature. By the time the change of axis of the entire body of the young Ciona has been effected the musculature characteristic of the adult begins to put in an appear- ance. In Fig. 107 circular sphincter muscles are present round the buccal and atrial apertures. The latter are still paired, but are carried by differential growth dorsalwards at a later stage, and finally coalesce together in the dorsal middle line to produce the single atrial aperture of the adult. One strand of the longitudinal muscles of the later muscular mantle is likewise to be seen in Fig. 107. It tends to branch dichotomously. Posteriorly it is inserted on the inner surface of the organ of fixation near the point where it joins on to the body. Later new muscle-bands arise similar to the first, and become distributed over the body-wall in a spreading fan-like fashion, but posteriorly they are all inserted in the same region of the organ of fixation. Alimentary Canal and Pyloric Gland. The course of the alimentary canal can be gathered so plainly from the accompanying figures (Figs. 105 and 107) that it hardly needs a verbal description. From the posterior dorsal corner of the branchial sac the cesophagus leads into the wide stomach, and from the latter, again, the intestine, which often possesses a strangulated appear- 236 THE ASCIDIANS. ance, doubles up obliquely forwards to the left atrial chamber, into which it opens by the anus (Fig. 107). In the angle made by the outgoing intestine with the stomach, a blind diverticulum arises. It is at first a sim- ple coecum, but soon begins to branch (Fig. 105 C), and finally forms an arborescent growth embracing the in- testine (Fig. 107). This is the so-called pyloric gland, and it is probably homologous with the hepatic caecum of Amphioxus. Appendicularia. It is generally agreed among those who have a voice in the matter, that most of the pelagic Ascidians (Salpa, Doliolum, Pyrosoma) are highly modified forms, spe- cially adapted to a pelagic life, one of the results of which is that their repro- duction is marked by a complicated alternation of generations. It would, therefore, not assist us in Our comparison with Amphioxus to describe these types. There is, however, one family of pelagic Ascidians, the Appendicularie, with re- spect to which there are two widely different opinions. Fig. 111.— Appendicularia (Fritil- The Appendicularize are Zaria) furcata, from the ventral surface. 5 F : A (After LANKESTER.) pelagic, free-swimming As- a. Anus. g?/. Unicellular glands. gs. jj; whose adult condi- Gill-slits. %. Dorsal hood-like fold of cidians, See integument. 2. Mouth. 4. Tail. tion is so far similar to the ANATOMY AND DEVELOPMENT. 237 larval condition of the fixed Ascidians, that they retain the tail as their organ of locomotion throughout life (Fig. 111). The tail is inserted in the middle of the ventral surface of the body proper, and is obviously a mere appendage of the latter. The mouth is terminal or sub-terminal. There is a sin- gle pair of branchial stigmata, which open into a pair of tubular atrial cavities, whose separate external apertures are seen in front, on the ventral surface behind the mouth. The alimentary canal is U-shaped, and the anus opens on the ventral surface to the right of the middle line, some- times behind and some- times (according to the species) in front of the stigmata (Figs. III, Piz); ~dWhevendostyle is always quite anterior in position, and some- times, as in Fig. 112, removed by a consider- able interval from the Fig. 112. — Diagram of the organisation of stigmata. a species of Appendicularia, from the right side. I 7 _ (After HERDMAN.) n the posterior cx a. Anus; the index line was accidentally tremity of the body drawn about % of an inch in front of the anus. 1 é.s. Branchial sac. ch. Notochord. e. Endostyle. are p aced the gonads, g. Ganglion, from which the nerve-cord proceeds male and female. in backwards to the tail, passing to the right of the ? alimentary canal. g.s. Gill-slit. 4. Heart. zmz. close proximity to one Intestine. 7. Mouth. 2c. Nerve-cord, with h i‘ .. ganglionic enlargements in the tail. of. Otocyst; another, the testis 1M feneath which the hypophysis opens into the front and the ovary branchial sac. ov. Ovary. 7.6. Peripharyngeal band. s¢.Stomach. Ze. Testis. behind. The heart, as described by LANKESTER, is a unique example of a func- tional organ reduced to the lowest possible level of histo- logical structure. It consists simply of two cells placed 238 THE ASCIDIANS. opposite one another and connected together by contractile protoplasmic threads, which keep up a pulsating motion. The tail is, as might be expected, more elaborately or- ganised than that of the Ascidian larva. The dorsal nerve- cord is solid, and proceeds backwards from the ganglion, passing to the 7zgfz¢ of the alimentary canal until it reaches the tail, along which it is continued, lying to the /eft of the notochord; it possesses ganglionic enlargements at intervals in the tail, from which nerves pass out. The caudal musculature also shows somewhat doubtful traces of being segmented in correspondence with the ganglionic swellings of the nerve-cord. In connexion with the cerebral ganglion there is a sense-organ in the form of an otocyst, with an enclosed otolith, and below this a ciliated pit opens into the ante- rior region of the branchial sac, corresponding to the hypophysis, or sub-neural organ, of the fixed Ascidians. According to one view, Appendicularia is the living rep- resentative of the free-swimming ancestor of the Ascidians. According to the other view, it is less primitive than the fixed Ascidians, and was derived from the latter by the gradual increase, from generation to generation, of the du- ration of the pelagic existence of the larva, until they ceased to metamorphose, and so retained the larval struct- ure throughout life, becoming at the same time sexually mature.® These two views are, of course, antagonistic, and the former of them is held by a number of well-known author- ities. As we are ignorant of the development of Appen- dicularia, it is impossible to decide definitely between them. With the facts which are at our disposal, however, the second view — namely, that the Appendiculariz represent Ascidian larvze which have become secondarily adapted to ANATOMY AND DEVELOPMENT. 239 a pelagic life, and have acquired the faculty of attaining sexual maturity — would be more in harmony with what we know of the relation of Amphioxus to the Ascidians. And it would seem that this affinity can be better demon- strated through the comparison of Amphioxus, both adult and larva, with a fixed Ascidian like Ciona than with Appendicularia.® On the latter view, therefore, the so-called metamerism of the tail of Appendicularia, on which so much stress has been laid, would be simply a secondary elaboration of the tail for the purpose of serving as a permanent locomotor organ. The dorsal nerve-cord of Appendicularia was regarded by Fo. as a simple peripheral nerve. We have described above how a portion of the primitive nerve-tube in Ciona and other Ascidians becomes reduced to a solid nerve. It would be of the greatest interest to discover the mode of origin of this nerve-cord in Appendicularia. Abbreviated Ontogeny of Clavelina. In order to demonstrate clearly the relatively primitive character of the development of Ciona it is sufficient to enumerate a few facts drawn from the development of Clavelina as described by Dr. OswaLp SEELIGER. As mentioned above, Clavelina is a near relative of Ciona, and in the adult condition resembles it very closely in many respects. The development of Clavelina was formerly regarded as being of a primitive character, but is in reality, more especially in the later stages, abbreviated and hastened to a remarkable extent. Like Ciona it possesses in the adult numerous trans- verse rows of stigmata. Each opening, however, arises by 240 THE ASCIDIANS. an independent perforation, so that all those preliminary ontogenetic processes which precede the establishment of the transverse rows of stigmata in Ciona are dropped out of the development of Clavelina.* In Clavelina, again, the change of axis of the body proper occurs in the unhatched larva; so does the fusion of the two atrial apertures to form the dorsal cloacal siphon. The longitudinal muscles of the body proper commence to appear in the free-swimming larva, while the caudal muscles are enjoying their highest functional activity. The vacuolisation of the notochord does not proceed so far as in Ciona, since the cells are never actu- ally removed from the centre of the notochord, but remain as thin discs stretching across the latter, so that the vacuolar spaces do not become continuous. The behaviour of the organ of fixation in the larva of Clavelina is such that it could hardly be recognised as a przoral lobe except in the light of Ciona. NOTES. 1. (p. 183.) The test or cellulose mantle of the Ascidians con- tains great numbers of cells of various kinds. These were formerly supposed to be derived from the subjacent ectoderm of the body- wall. Kowa.evsky has recently shown, however, that the cells of the outer (cellulose) mantle of the Ascidians are derived from wandering mesenchyme-cells which wander from the body-cavity through the ectoderm (either de/ween the ectodermic cells or actually passing ¢hvough the individual cells) into the mantle. * A mode of formation of the branchial stigmata, intermediate between that of Clavelina and Ciona or Molgula, has been described by GARSTANG for Botryllus. In this genus, the primary branchial stigmata all arise by in- dependent perforations, and then later become divided up into the transverse rows of stigmata. (W. GARSTANG. Ox the development of the stigmata in Ascidians. Proc. Roy. Soc., Vol. LI. 1892.) NOTES. 241 2. (p. 211.) In Clavefina the atrial involutions do not merely arise as minute circular invaginations of the ectoderm, but at first they appear as short, though quite distinct, longitudinal grooves. Compare also the remarkable longitudinal atrial tubes of Pyvosoma. 3. (p. 238.) There is another possible way of interpreting the structure and systematic position of Appendicularia which may perhaps be nearer the truth than either of the views mentioned in the text. It is not absolutely necessary to suppose that the ancestors of Appendicularia were fixed Ascidians; but both Appendicularia and the fixed Ascidians may have descended from a common free-swimming stock, and have undergone certain modifications in common, such as loss of true vascular system and celom. Then, while the Ascidians proper became adapted toa sessile existence, Appendicularia may be supposed to have gone to the opposite extreme, and have become adapted to an absolutely pelagic existence. In becoming adapted to such a purely pelagic or oceanic environment as that of Appendicularia, it is eminently conceivable that an animal would have to undergo as radical a modification of structure as it would in becoming adapted to a sessile existence. (Compare Sa/a, Doliolum, etc.) V. fAE PROTOCHORDATAIIN THEIR RELADION TO THE -PROBLEM OF VERTEBRATE, DES SCE Nae “ Den Schlissel richtigen Verstandnisses gibt nicht das Hineinpressen neuer Thatsachen in eine alte Schablone, sondern das Aufsuchen des genetischen Zusammenhangs der Erscheitnungen.” — WEISMANN. BALANOGLOSSUS. External Features. Or the free-living protochordates, the lowest type of organisation is undoubtedly presented by the Exteropneusta (Hemichorda), the group to which Balanoglossus belongs. Balanoglossus is a remarkable worm-like creature which lives buried in the sand or mud of the sea-shore. By means of numerous unicellular integumentary glands which are distributed over the surface of the body, it secretes a mucous substance to which particles of sand adhere, and so makes for itself tubes of sand in which it lives at about the level of the low tide-mark. It possesses such a characteristic external form and odour (like iodoform) as to render it peculiarly easy of recognition. In front there is a long and extremely sensitive proboscis which is capable of great contraction and extension, and is, in the living animal, of a brilliant yellow or orange colour. Behind the proboscis follows a well-marked col/lar-region, 242 BALANOGLOSSUS. 243 consisting externally of a collar-like expansion of the integument, with free anterior and posterior margins over- lapping the base of the proboscis in front and the anterior portion of the gz//-slzts behind. In the ventral middle line, at the base of the proboscis and concealed by the collar, is situated the mouth (Fig. 113). Following behind the collar is the region of the trunk or body proper, which, in the adult of some species, reaches a relatively enormous length, even extending to Fig. 113. — Larva of Balanoglossus Kowalevskii, with five pairs of gill-slits ; from the right side. (After BATESON.) a. Anus. a.~. Temporary pedicle of attachment. c¢. Collar. cz. Notochord. g.s. Gill-slits. mm. Mouth. gr. Proboscis. two or three feet. The ectodermal covering of the body consists in general of ciliated cells, among which are scat- tered unicellular mucous glands ; the cilia, however, appear to be more prominent on the proboscis than elsewhere. In the region of the trunk, which immediately follows upon the collar region, there are a great number of paired 244 THE PROTOCHORDATA. openings on the dorsal side of the body, placing the anterior portion of the digestive tract in communication with the outer world. Theseare the gz//-s/zts, and they are arranged strictly in consecutive or metameric pairs to the number of upwards of fifty in the adult. In their structure, and more especially in the possession of tongue-bars, they bear a remarkable resemblance to the gill-slits of Amphioxus. This is particularly striking in young individuals. As the adult form is approached in the development, the bulk of the gill-slits sinks below the surface, only opening at the latter by small slit-like pores, and thus their true character is obscured in a superficial view. Projecting into the interior of the proboscis is a rod-like structure which arises as an outgrowth from the alimentary canal dorsal to the mouth. The lumen of this endodermic diverticulum becomes narrowed down and, in fact, partially obliterated, while the cells constituting its walls give rise to a spongy vacuolar tissue which strongly resembles the notochordal tissue of Amphioxus and the higher Verte- brates. On account of its dorsal position above the mouth, its endodermic origin, and the vacuolisation of its cells, this structure was identified by BaTEson in 1885 as the zofo- chord. Nervous System and Gonads. The nervous system of Balanoglossus presents many features of the utmost interest and suggestiveness. It consists essentially of an ectodermal network of nerve-fibres forming the inner layer of the skin (ectoderm) all over the body. In this primitive nervous sheath, which envelops the whole body, there are certain definite local thickenings. Two of these thickenings occur respectively along the whole length of the dorsal and ventral middle lines in the trunk-region, thus producing the dorsal and ventral median BALANOGLOSSUS. 245 longitudinal nerve-cords. In the region of the collar the dorsal nerve-cord becomes entirely separated from the ectoderm, and this portion of it contains, at least in young individuals, a central canal which, from its origin and relations, was shown by BATEsoN, and more recently by MorGan, to be homologous with the central canal of the vertebrate spinal cord. Anteriorly the dorsal nerve-cord becomes continuous with a specially dense tract of the general nerve-plexus at the inner posterior surface of the ov bc* Com Fig. 114.— Diagram of the organisation of Balanoglossus, from the left side. (From a drawing kindly lent by Professor T. H. MORGAN.) al. Alimentary canal. 6cl1. Coelom of proboscis (anterior or preeoral body- cavity). 6c?. Coelom of collar. 4c3. Coelom of trunk. 4.v. Blood-vessel, proceed- ing from the so-called heart (which lies at base of proboscis above the noto- chord) to the ventral blood-vessel. ch.,Notochord. com. Commissure, between dorsal and ventral nerve-cords. dz. Dorsal nerve-cord, separated from the integu- ment in the collar-region. d.d.v. Dorsal blood-vessel. g/. Proboscis-gland ; modified ccelomic epithelium surrounding heart and front end of notochord. m. Mouth. #.v. Pulsating vesicle, lying inside the “heart.” v.4.v, Ventral blood- vessel. v.2. Ventral nerve-cord. proboscis (Fig. 114). This proboscidian plexus thins out somewhat towards the anterior extremity, but nevertheless forms a complete nerve-sheath for the proboscis and indi- cates the sensitive character of the latter (Fig. 115). The ventral nerve-cord does not extend into the region of the collar, but from the point where the collar joins on to the trunk the ventral cord is connected with the dorsal nerve-cord by a commissure-like thickening of the integu- mentary plexus, which passes in the skin on each side round the hinder end of the collar-region (Fig. 114). 2406 Fig. 115.— Diagrammatic transverse sec- tion through hinder region of proboscis of Balanoglossus. (From a drawing kindly lent by Professor T. H. MORGAN.) D. Dorsal. V. Ventral. cl, Proboscis- cavity, almost filled up by mesenchymatous and muscular tissue,* proliferated from the THE PROTOCHORDATA. The testes or ovaries, accord- genital organs, ing to the sex of the individual, occur as a paired metameric series of pouch-like bodies or gonadic sacs which ex- tend backwards far be- yond the region of the gill-slits. sacs are suspended in the The gonadic body-cavity by solid cords attached to the dorsal which _ be- come perforated in the integument, spawning season to ad- original coelomic epithelial layer (indicated by the black line below the ectoderm). p.v. Pulsating vesicle. #4. Heart. ch. Noto- chord. .s. Integumentary nerve-plexus. mit of the expulsion of the reproductive elements. Metamerism. Although there is no muscular metamerism in Balano- glossus, yet we have seen that other organs (gill-slits and gonads) are arranged metamerically. And in point of fact, among those Invertebrates which are not included under the phylum of the Articulata, if there is one pecu- liarity of organisation more sporadic in its occurrence than another, it is metamerism. It may affect the most differ- ent organs of the body either collectively or individually, and nothing is more patent than the fact that the meta- meric repetition of parts has arisen independently over and over again in different groups of animals.1 * This tissue is not represented in Figs. 114 and 116, although it is present throughout the body-cavity. BALANOGLOSSUS. 247 Far from assuming as a self-evident fact that the extreme metamerism of the Annelids and Arthropods is genetically identical with that of the Vertebrates, we have every reason to suppose that it has been elaborated entirely independently in the two cases, and that the apparent simi- larity is due, as already intimated, to a parallel evolution. Body-cavities ; Proboscts-pore ; Collar-pores. Corresponding to the three regions into which the body of Balanoglossus is divided, — namely, probos- cis, collar, and trunk, —the body-cavity is divided up into three systems of cavities. These are (a) the anterior body-cavity or cavity of the iam ae) I < proboscis, (8) a pair of collar- “t 1 1 cavities, and (y) a pair of body-cavities which form the unsegmented ccelom of the trunk (Figs. 114, 115). These cavities arise essen- WNW comm | ANdHANEAANADLDADINAT THI mM || tially as pouches from the @Ooe® @® Le RAaannn archenteron (Fig. 117), al- though their actual develop- , : : Fig. 116. — Diagram of the organisa- ment differs considerably IN tion of Balanoglossus, from the dorsal different species (MorGAN). side. (From a drawing kindly lent by : ° ’ Professor T. H. MORGAN.) lie proboscis-cavity 1S c.p. Collar-pores. go. Gonads. g.s. . : : Gill-slits; the dark lines converging be- placed in communication hind indicate the superficial portions of with the exterior by an open- the gill-slits; below the surface are seen . : the free ends of the tongue-bars. ff. ing th rough the posterior Pproboscis-pore. Other letters as above. 248 THE PROTOCHORDATA. wall of the proboscis known as the proboscis-pore. In B. Kowalevskii this pore lies asymmetrically to the left of the dorsal middle line (Fig. 115), while in B. Kupfferi a corresponding opening occurs Fig. 117. — Diagrammatic horizontal section through an embryo of Balanoglos- sus (type of the direct development), to show the origin of the body-cavities as archenteric pouches. (After BATESON.) ap. Tuft of cilia at the apical pole (indication of an apical plate). écl. Probos- cis-cavity. dc. Collar-cavities. cé3, Trunk- cavities. cd. Circular band of cilia. to the right of the middle line, so that in this species there are two proboscis- pores constituting a sym- metrical pair. The left proboscis-pore of Balanoglossus is obvi- ously to be compared with the przoral pit of Amphi- OXUS. The collar-cavities also open to the exterior by pores, one on each side underneath the dorsal pos- terior. “free, fold yor the collar, and on a level with the opening of the first gill-slit. These are the funnel-shaped col/ar-pores. SPENGEL states that water is taken in through the collar-pores into the cavity of the collar in order to swell the latter up, so that it may serve as an accessory organ of locomotion in so far as an alternate inflation and collapse of the collar would assist the animal in its slow burrowings in the sand. BALANOGLOSSUS. 249 Alimentary Canal. The mouth cannot be closed, as there is no sphincter muscle, and accordingly, as the animal progresses through the sand, it swallows a large quantity of the latter in which food-particles (unicellular organisms, etc.) may also be involved. As the sand passes through the intestine, it becomes enveloped in the mucous secretion of the intes- tinal epithelium, and is ejected through the anus in a cord of slime. The alimentary canal is a straight tube between mouth and anus. In its hinder portion it is usually sacculated, z.e. provided with paired lateral saccular dilatations comparable to the so-called intestinal ceca of the Ne- mertine worms. (See below.) In the region of the pharynx the lumen of the alimentary canal is incompletely divided by lateral constrictions into y Fig. 118. — Transverse section through two portions, an upper Or the gill-region of Balanoglossus. (After he peal fi : SPENGEL.) vanchiat portion Carrying al, Digestive portion of gut. 47. the gill-slits, and a lower or Branchial portion of gut. bc8, Third . ; ; e body-cavity (trunk ccelom) ; this is also digestive portion (Fig. I 18). nearly obliterated in the adult by the pro- liferation of mesenchyme or “paren- The latter was compared by chyme” from its walls. d@.c. Dorsal GEGENBAUR* to the endo- nerve-cord. d.d.v. Dorsal blood-vessel. ahs go. Gonad. g.s. Gill-slit. £4. Tongue- style of the Ascidians, but bar. v.4.v. Ventral blood-vessel. v.1.c. ae | nerve-cord. it is probable that this com- Vem"! nervecord parison, although a very natural and useful one at the time at which it was made, will not hold good, since there is * CARL GEGENBAUR, Elements of Comparative Anatomy. Translated by F. Jeffrey Bell. London, 1878. 250 THE PROTOCHORDATA. nothing in the structure or development of this part of the alimentary tract in Balanoglossus which will bear compari- son with the endostyle.* As indicated in the larvae of Amphioxus and the Ascidians, it would seem that the endostyle first became evolved or differentiated at the anterior end of the pharynx, zz front of the gill-slits, in correlation with the dorsal position of the mouth. Development; the Tornaria Larva. The development of Lalanoglossus Kowalevskit as made known to us by the admirable work of BATESON is what is knownas a strictly direct development; that is to say, the embryonic, larval, and adult stages follow one another by gradual transitions concomitantly with the simple progres- sive growth of the individual and without any striking metamorphosis. In other species of Balanoglossus the larval form is remarkably different from the adult, and becomes transformed into the latter by a very distinct metamorphosis. The extraordinary larval form here re- ferred to was discovered in 1848 by JOHANNES MULLER, who named it 7ornzaria, and regarded it, as did his succes- sors KrouHn, ALEXANDER AGAssiZz, and Fritz MULLER, as the larva of an Echinoderm (Starfish). It was not until 1869 that its true character as the larva * A ciliated tract in the floor of the cesophagus of a Tornaria from the Pacific has recently been compared to the endostyle by W. E. Ritrer. (Ox a New Balanoglossus Larva from the Coast of California and its Possession of an Endostyle. Zool. Anz. XVII. 1894. pp. 24-30.) The comparison is at present somewhat doubtful. More recently GARSTANG has suggested that the endostyle is derived from the adoral ciliated band of the Echinoderm larva. (See Fig. 119.) The suggestion is an interesting one, but Garstang’s idea of the relations of the preeoral lobe is very different to the one here set forth. (WALTER GARSTANG, Preliminary Note on a New Theory of the Phylogeny of the Chordata. Zool. Anz. XVII. pp. 122-125.) BALANOGLOSSUS. 251 of a species of Balanoglossus was demonstrated by Extras METSCHNIKOFF. Shortly afterwards, Metschnikoff’s dis- covery was confirmed and amplified by ALEXANDER AGASSIZ. The superficial likeness between Tornaria and such Echi- noderm larve as Bipinnaria or Auricularia is astonishing, and a renewed study of the detailed organisation of Tornaria, recently made by MorGan, appears to have established the fact, originally insisted upon by Metschni- koff, that this resemblance can only be accounted for on the ground of genetic affinity. In Figs. 119 and 120 two types of larve, Tornaria and Auricularia, are shown side by side; and although unfortunately they are not figured from exactly the same aspect, yet it is obvious at a glance that, in spite of certain differences which will be enumerated below, they both belong to the same category of larval forms. A highly characteristic feature of these larve is the remarkable ectodermal ciliated band which constitutes a perfectly symmetrical but somewhat complicated undulat- ing seam round the body. The larve are strictly pelagic, and swim about in the open sea by means of their cilia ; but the latter, instead of being distributed evenly over the whole surface of the body, are concentrated in the region of the ciliated bands which are composed of thickened ectoderm. In Tornaria there are two ciliated bands, viz.: 1) the above-mentioned undulating seam which is usually known as the cercumoral or longitudinal ciliated band, and 2)a postoral circular ciliated band. Only the former is present in Auricularia, and the absence of the circular band in this form constitutes one of the chief differences between the two larvae. 252 THE PROTOCHORDATA. From a morphological point of view a more striking resemblance between the two larve than that furnished by the longitudinal ciliated bands exists in connexion with the anterior body-cavity or exteroce/l. In the Echinoderm eeircibietlctne eI frene mee . ‘ Figs. 119 and 120.— Auricularia, larva of Synapta (after SEMON); and Tornaria, larva of Balanoglossus. (After MORGAN.) a. Anus. aj. Apical plate. cl, Anterior body-cavity, communicating with exterior by the water-pore. 4c?, 6c8, Second and third body-cavities of Tornaria. ¢.6, Circular ciliated band of Tornaria. c.c. Contractile cord between apical plate and anterior body-cavity of Tornaria. gf. Gill-pouches. 4.c. Hydroccel of Auricularia (anterior body-cavity). /.c.6. Longitudinal (circumoral) ciliated band. Ze. Left enteroccel (body-cavity). . Mouth. z. Lateral (paired) nerve-band of Auricularia. 7e. Right enteroccel. sf. Calcareous spicules. s¢, Stomach. wp. Water-pore. N.B.—In Auricularia, the margin of the mouth is surrounded by a ciliated band discovered by SEMON, and known as the adoral ciliated band. The poste- rior, V-shaped portion of this band lies inside on the ventral floor of the larval cesophagus. larva this cavity arises as a median pouch of the archen- teron, and there is every reason to suppose that it has a similar origin in Tornaria, although this point has not yet BALANOGLOSSUS. 253 been determined. The primary anterior enteroccel in the Echinoderm larva is not quite the same as the correspond- ing cavity in Tornaria, since it contains also the elements of the general body-cavity. Apart from slight differences, the collar-cavities and general body-cavities arise essen- tially in the same way in Tornaria as they do in the case of the direct developing larva of Balanoglossus (see above).* In the Echinoderm larva, however, the paired body- cavities do not arise as independent archenteric pouches, but they become constricted off from the anterior entero- ceel. Making allowance for these deviations in the origin of the body-cavities, — deviations which are by no means fundamental, since in both cases the body-cavities are ultimately reducible to archenteric pouches, —it is an extremely striking fact that both in Tornaria and Auricu- laria the anterior enteroccel acquires an opening to the exterior on the dorsal surface to the left of the middle line. This opening is called the waterpore, since it forms the outlet (possibly both outlet and inlet) of the water-vascular system of the Echinoderm. In Tornaria it persists after the metamorphosis as the proboscis-pore, which has been described above. The Larva of Asterias vulgaris; Water-pores and Preoral Lobe. In view of what was said above as to the occurrence of paired proboscis-pores in B. Kupfferi, it is interesting to note that sometimes there are two water-pores, a right and a left, in Echinoderm larvae. This has been observed by * As to the origin of the body-cavities in different species of Balanoglos- sus, MORGAN summarises his observations as follows: “They may arise as enteric diverticula, as endodermal proliferations, or even arise from mesenchy- matous beginnings.” (See Morcan. No. 125 bibliog.) 254 THE PROTOCHORDATA. Brooks and G. W. FIELp in the larvz of a common star- fish, Asterzas vulgaris. In this case the primary enteroccel becomes constricted off from the archenteron in the form of two equal pouches. The right and left enteroccelic sacs then take up a symmetrical position on each side of the larval cesophagus, and each sac next opens to the exterior by a water-pore. The pore in connexion with the right sac (Fig. 121) is, however, of a transitory, rudimentary character, and soon closes up, while the left pore per- sists as the definitive water- pore. As in Tornaria, so here, the cavity of the larval body generally, and of the preeoral region (pre@oral lobe) in particular, is the primary body-cavity or blastoccel, and contains scattered mes- enchyme-cells. At a later Fig. 121. — Young larva of Asterias stage In the larva of As- vulgaris, from the dorsal side. (After terjag the right and left G. W. FIELD.) : ; ; pl. Preoral lobe. 4.c.5. Circumoral enteroccelic sacs, having in- (longitudinal) ciliated band. oes, G¢soph- : agus. ve. and Ze. Right and left en- creased greatly mM length, teroccelic sacs, each opening bya“water- meet one another in the pore” to the exterior. sf. Stomach. zz. Aperture, leading from stomach into in- region of the preeoral lobe en and fuse together, thus put- ting their two cavities into communication across the median line. The median portion of the enteroccel thus produced extends up into the przoral lobe, and so the primary blastoccelic cavity of the latter is replaced by a secondary ingrowth of the enteroccel (Fig. 122). Similarly with the metamorphosis of Tornaria, the anterior enteroccel, which is at first of very inconsid- BALANOGLOSSUS. 255 erable extent (Fig. 120), increases greatly in size, and assumes its definite position and proportions as the cavity of the preeoral lobe (z.e. proboscis), thus replacing the original blastoccelic space, while the water-pore remains as the proboscis-pore. As described in the previ- ous chapter, the cavity of the preoral lobe (fixing stolon) of the Ascidian tad- pole is of the nature of a blastocoel or primary body- cavity, containing loose mes- enchyme-cells, and it is 2 Fig. 122.— Older larva (Bipinnaria) therefore of great IMPOT- of Asterias vulgaris, from the ventral tance to note that whether side. (After G. W. FIELD.) a By a fusion of the two przeoral loops the cavity of the preoral of the ciliated band across the apex of the o 5 przeoral lobe, followed by a separation in lobe is a 6lastocwl or an the transverse direction, the originally enterocel, the morphological single circumoral band (cf. Figs. 119 and 5 121) has become divided into two bands, value of the structure itself a preoral ciliated band #.c.6. and a post- remains the same oral longitudinal ciliated band /.c.6. The ; posterior transverse portion of the pra- oral ciliated band has undergone a fusion Apical Plate of Tornaria. With the front end of the originally dis- tinct adoral band (cf. Fig. 119). 7.2. Prae- At the anterior end of oral lobe, into which the enterocoel has extended. mm. Mouth. me. and Ze. Right the body, or, in other words, and left enteroccelic cavities. s¢, Stomach, at the apex of the praoral “ ees lobe, in Tornaria, there is an ectodermic thickening in which nerve-cells and nerve-fibres and a pair of simple eyes have become differentiated. This is the so-called apical plate, and it constitutes the central nervous system of the larva. It can be recognised for some time after the metamorphosis at the tip of the proboscis, but eventually disappears completely. A similar apical plate occurs in 256 THE PROTOCHORDATA. a great number of Invertebrate larvae, and is especially characteristic of the free-swimming larve (Trochophores, or Trochospheres) of Annelids and Molluscs. We shall return to this later. In Tornaria a single contractile cord passes from the apical plate to the anterior enteroccel. There is no apical plate in Auricularia, nor in most of the other Echinoderm larve ; but there is reason to sup- pose that it has been secondarily lost, since a transitory ectodermal thickening at the apical pole can frequently be observed in the course of their development ; and, moreover, in what is probably the most primitive Echino- derm larva known (viz. the larva of the Crinoid, Axzedon), there is a well-developed apical plate. Metamorphosis of Tornaria. The metamorphosis of Tornaria, as originally described by Alexander Agassiz, takes place with relative sudden- ness. According to the more recent account of the meta- morphosis given by MorGan, a marked diminution in size occurs; the internal organs are drawn together in such a way that the larval cesophagus, with the gill-pouches (see Fig. 120), is drawn backwards into the body, and the anterior enteroccel, as already described, is carried for- wards into the przeoral lobe. The longitudinal (circum- oral) ciliated band, which was the first to develop, is also the first to disappear, while the posterior circular band persists to a somewhat later stage. The Nemertines. It is thus evident that Balanoglossus, especially through its Tornaria larva, shows undoubted marks of affinity to NEMERTINES. 257 the Echinoderms. It will next be shown that there are certain features in the adult anatomy which apparently indicate a distinct genetic relationship to another group of the Invertebrates ; namely, the Vemertine worms. The Nemertines are elongated, flattened, or cylindrical worms, with a smooth cz/zated skin and no external seg- mentation, occurring, as a rule, in a closely similar habitat to that of Balanoglossus, buried in the sand or mud of the sea-shore. Like Balanoglossus, they also possess unicellular integu- mentary glands, by means of which they secrete a mucous substance, to which frequently sand-grains adhere, thus producing a tube of sand round the body. Some of them reach an enormous length, and one at least must be measured in yards (Lzneus longissimus exceeding three yards in length). The chief anatomical features which offer material for direct comparison between the Nemertines and Balano- glossus relate to the ectoderm, proboscis, nervous system, mesenchymatous tissue, the reproductive organs, and the alimentary canal. As for the ectoderm, considered apart from the nervous system, it need only be repeated that in both cases it is composed of ciliated cells and scattered mucous glands. The proboscis of the Nemertines is one of the most characteristic organs of this group of animals. It is not permanently protruded, and does not serve as an organ of locomotion, as in Balanoglossus, but is usually carried about entirely withdrawn within the body of the animal, from which it can be shot out with great force and rapidity when the occasion demands it. During the process of extrusion it is turned completely inside out, and conversely, during the process of zxtroverszon, the retraction takes 258 THE PROTOCHORDATA. place from the tip backwards by the in-rolling of its walls. According to the graphic description of HuBRECHT, it is retracted ‘fin the same way as the tip of a glove finger would be if it were pulled backwards by a thread situated in the axis and attached to the tip.” When at rest within the body the proboscis lies freely within a hollow cylinder, the wall of which is thick and muscular, and constitutes the prodoscis-sheath (Fig. 123). Fig. 123. — Diagrammatic transverse section through the middle of the body of a Nemertine. (After LANG, Text-b00k of Comp. Anat.) én, Basement-membrane. c.m. Circular muscles. d.z. Dorsal or “ medullary” nerve. d.v. Dorsal blood-vessel. g. Gonads. wz. Intestine. /.m. Longitudinal muscles. 7. Lateral nerves. ¢.v. Lateral blood-vessel. ~. Proboscis. #.s. Pro- boscis-sheath. Sometimes beneath the ectodermal epithelium of the Nemertine proboscis there is a continuous sheath of nerve- fibres, comparable to the nervous plexus in the proboscis of Balanoglossus. Partly, therefore, on account of its structure, and partly on account of its topographical relations when extruded, we are led to suppose that a certain homology exists NEMERTINES. 259 between the retractile proboscis of the Nemertines and the non-retractile proboscis of Balanoglossus (BATESON). In the most primitive Nemertines the nervous system consists essentially of a somewhat complicated pair of cerebral ganglia and a diffuse nerve-plexus, with nerve- cords lying at the base of the ectoderm.* As the cerebral ganglia probably belong to the same category as the cere- bral ganglia of all other typical Invertebrates, and are not represented in Balanoglossus, we can afford to neglect them at present. Confining our attention to the ecto- dermal nerve-plexus, we find occurring in it, along definite lines, local thickenings, after the same principle, but not all on the same lines, as was described above for Balano- glossus. Directly comparable with the dorsal longitudinal nerve-cord of Balanoglossus, there is a similar thickening or concentration of the integumentary nerve-plexus in some of the Nemertines, in the dorsal middle line (Car- inina, Cephalothrix). Hubrecht, who discovered this, calls it the medullary nerve. There is, however, no correspond- ing ventral nerve-cord in the Nemertines, but, instead of this, there is a pair of lateral thickenings, constituting the well-known /ateral nerves of the Nemertines (Fig. 124). It is usually supposed that the lateral nerves of the Nemertines are homologous with the two halves of the ven- tral nerve-cord in the Annelids. In the Annelids the primitive lateral nerves (which are so typical of the Platy- helminths, or flat-worms) have approached one another in the mid-ventral line, and have often undergone intimate fusion together. In some cases, however, they are separated from one another by a wide interval (Sabellaria, etc.). * HUBRECHT compared the lobes of the cerebral ganglia of a Nemertine to the cranial ganglia of the Vertebrates, the lateral nerves to the Rami laterales vagi, and the proboscis-sheath to the notochord. 260 THE PROTOCHORDATA. In the Annelids, in contrast to the Nemertines, the gan- glion-cells are not distributed uniformly along the whole length of the nerve-cord, but are collected together to form definite ganglionic swellings. It is, therefore, very significant that in the Nemertines we have a median dorsal ‘medullary”’ nerve, in addition to the elements which constitute the ventral nerve-cord of the Annelids. In many Nemertines the dorsal and lateral nerve-cords do not continue to lie in the ectoderm throughout life, but Fig. 124.— Diagrammatic view of anterior portion of a Nemertine, from the left side. (After HUBRECHT, from LANG.) a./. Anterior lobe of brain. £./. Posterior lobe of brain. . Opening of pro- boscis. . Mouth. d.#. Dorsal nerve. /. Lateral nerve. 7.7. Ring-nerves. sink deeper into the body, and so come to be separated from the ectoderm, first by the basement membrane, and then by one or more muscular layers of the body-wall. In the Hoplonemertea (those in which the proboscis is armed with stylets) the medullary nerve is absent. In all cases, however, the longitudinal nerve-cords remain connected with one another by a more or less plexiform arrangement of nerve-fibres ; although sometimes a more definite con- nexion, by means of metameric ring-nerves, has been observed by Huprecut (Fig. 124). NEMERTINES. 261 There is no true ccelom in the Nemertines, and the space between the alimentary canal and body-wall is oc- cupied by a gelatinous mesenchyme, containing muscular and connective tissue elements. In Balanoglossus the cav- ity of the coelom becomes largely obliterated in the adult, by the proliferation of cells from the epithelium of its walls, thus filling up the cavities with a more or less solid parenchymatous tissue. Like Balanoglossus, the Nemertines have a straight all- mentary canal, provided with paired lateral outgrowths or tntestinal ceca, and a terminal anus. The gonadic sacs of the Nemertines offer a striking re- semblance to those of Balanoglossus. They occur as a metameric series of paired sacs, which alternate with the above-mentioned intestinal cceca, and communicate with the exterior by short tubes, which are at first solid, as in Balanoglossus, subsequently becoming hollowed out and opening above the lateral cords (Fig. 124). Finally it should be pointed out that, while excretory organs, in the form of a well-developed single pair of elongated nephridia, provided with numerous internal ‘“end-sacs,” are present in the Nemertines, nothing of the kind has yet been detected in Balanoglossus. CEPHALODISCUS AND RHABDOPLEURA. It is interesting to note that there are some remarkable animals which stand in a similar relation to Balanoglossus that the Ascidians do to Amphioxus. While Balano- glossus is free-living, does not produce buds, and has a straight alimentary canal, these creatures, of which only two genera are at present known, Cephalodiscus and Rhab- dopleura, \ead a sessile existence, produce buds, and have 262 THE PROTOCHORDATA. a U-shaped alimentary canal. Both are deep-sea forms, Cephalodiscus having been dredged during the Challenger Expedition, from the Straits of Magellan, at a depth of 245 fathoms ; while Rhabdopleura was first dredged indepen- dently, off the Shetland Islands, at go fathoms, by the Rev. Wins EN ‘i ih \ —\ ‘ Fig. 125. — Cephalodiscus dodecalophus, from the ventral side. (After M’'INTOSH.) Actual length of polypide from extremity of branchial plumes to the tip of the pedicle is about 2 mm. 4.s. Buccal shield; the shading on its surface indicates pigment-markings. At the tip of the pedicle, buds are produced. Canon NormMav, and off the Lofoten Islands, at 200 fath- oms, by Professor G. O. Sars (1866-68). Rhabdopleura is the name given by ALLMAN (1869), who published a short account of it; and it has since been described by Sars, LANKESTER, and G. H. FOWLER. CEPHALODISCUS. 263 The account which we possess of Cephalodiscus forms one of the Challenger Reports, and was written by Pro- fessor W. C. M’Inrosa, who made out the main features of its anatomy. It was further treated, from a morpholog- ical standpoint, by SipNey F. HARMER, who pointed out its remarkably close affinity to Balanoglossus. The most important morphological features in the anat- omy of Cephalodiscus are shown in Figs. 125-127. The individuals live in colonies, in a “house” or ce@n@ecium, which consists of a ramifying and anastomosing system of tubes, the walls of which are composed of a semi-trans- parent, gelatinous material, whose outer surface is covered with spinous projections. The walls of the coencecium are furthermore perforated by numerous apertures, which allow of the ingress and egress of water. The adult members of a colony have no organic con- nexion between themselves, but each one is independent and free to wander about the tunnels of the coencecium. Although Cephalodiscus has not been studied in the living condition, there is every reason to suppose that it moves about in its tube by means of the large duccal shield (Fig. 125) overhanging the mouth, by which it can attach itself to the inner surface of the tube, and then help itself along by the curious pedicle which occurs ventrally near the hinder end. It thus seems probable that this pedicle can be used as a sucker, but its chief function lies in the production of buds which grow out from it, and eventually become detached. Bateson has described a somewhat similar sucker at the hinder end of the body in young individuals of Balanoglossus (Fig. 113). Behind and above the buccal shield there is a row of twelve tentacles or branchial plumes, each possessing a central stem or shaft which carries numerous lateral 2064 THE PROTOCHORDATA. pinne. An important function of these plumes is to produce currents of water by the action of their cilia, which vibrate in such a direction that the water with food-particles is led into the mouth. The superfluous water is led out from the proximal portion of the aliment- ary canal by a single pair of gz//-sits which are not visible in surface view, since they are overhung by a fold of the integument known as the post-oral lamella or operculum, corresponding to the posterior free fold of the collar in Balanoglossus (Pigs126): In its internal organisa- tion, 1f due allowance be made for its U-shaped ali- mentary canal, Cephalodis- cus greatly resembles Bala- noglossus (Figs. 126, 127). The buccal shield of the Fig. 126. — Longitudinal frontal (right former is obviously the and left) section through an adult Cephalo- equivalent of the probos- discus. (After HARMER.) bc2, Second portion of body-cavity cis of the latter, and the (collar-coelom). 4c8. Third portion of : igh S ai body-cavity (trunk ccelom). 67. Pharynx. cavity whic it contains cp. Collar-pores. .g.s. Gill-slits. z¢, In- corresponds to the probos- testine. 7.5. Nervous system. of. Oper- : culum, oes. (Esophagus. sf. Stomach. C1S-Cavity. Moreover, the zt. Base of tentacle. proboscis-cavity in Cephalo- discus (z.e. the cavity of the buccal shield) communicates with the exterior by ¢wo proboscis-pores placed right and left of the dorsal middle line. Following behind the buccal shield is the collar-region, from which the branchial plumes arise dorsally, while CEPHALODISCUS. 265 laterally and ventrally it is produced into a free fold to form the above-mentioned operculum. The collar-region contains a section of the coelom which is precisely homolo- AB Fig. 127. — Longitudinal sagittal section through an adult Cephalodiscus. (After HARMER.) The section is supposed to be taken sufficiently to one side of the middle line to allow of the representation of one of the ovaries and one of the proboscis-pores. a. Anus. 6.c. Trunk-ccelom. c¢.c. Collar-ccelom. chk. Notochord. zm#f¢, Intes- tine. mm. Mouth. .s. Nervous system. of. Postoral lamella (operculum). ov. Ovary; the oviduct is deeply pigmented. f.c. Preeoral ccelom (cavity of buccal shield). £#%. Pharynx. 7.f. Proboscis-pore. fed. Base of pedicle. st, Stomach. gous with the collar-cavities of Balanoglossus. As in the latter form, it communicates with the exterior by a pair of collar-pores which open at the level of the gill-slits. 266 THE PROTOCHORDATA. The collar-ccelom is continued posteriorly into the opercu- lum, and anteriorly into the twelve tentacular appendages. Finally, behind the collar comes the region of the body containing the viscera, which are surrounded by the third section of the ccelom. Only the female reproductive organs have been observed up to the present time in Cephalodiscus. They occur as a pair of gonadic sacs, opening to the exterior on each side of the dorsal middle line between the anus and the central nervous system. The latter is very simple, being represented merely by a dorsal thickening of the ectoderm, with nerve-fibres in the region of the collar and posterior portion of proboscis. Finally, a short notochordal diverticulum projects into the base of the buccal shield as in Balanoglossus. Rhabdopleura differs considerably from Cephalodiscus in many respects, but, nevertheless, has some fundamen- tal characteristics in common with it. In Rhabdopleura the individuals of a colony are not independent, but are connected with each other by a common cord or cawlus, which represents the remains of the contractile stalks of the polyps. As the growth of the colony proceeds, the distal portions of the stalks (z.e. the portions farthest away from the animals) become shrunken and hard. The buds arise from the soft portions of the caulus, and never be- come detached as they do in the case of Cephalodiscus. There is only a single pair of tentacular plumes in Rhab- dopleura. Fow Ler has recently shown that in Rhabdopleura the cceelom, whose existence was first established by Lan- KESTER, exhibits the same subdivisions as have been mentioned above for Cephalodiscus; namely, (1) the cavity of the large buccal shield, (2) the collar-cavity opening PRAZORAL LOBE. 267 to the exterior by a pair of dorsally placed collar-pores, and (3) the body-cavity proper surrounding the alimentary canal. According to Fowler, who has recently described them in Rhabdopleura, the nervous system and notochord have essentially similar relations to those which obtain in Cephalodiscus, but there are no proboscis-pores and no gill-slits. THE PRAORAL LOBE OF ECHINODERM LARV#. In the previous pages a good deal of stress has been laid on the existence of a przoral lobe in the various types considered. We have recognised it in the snout of Am- phioxus (przeoral coelom + przeoral pit), in the proboscis of Balanoglossus, the fixing organ of the Ascidian tadpole, and in the buccal shield of Cephalodiscus and Rhabdo- pleura. From a morphological standpoint the przoral lobe is probably one of the most important, as it is certainly one of the oldest, structures of the body of bilateral animals, and it becomes, therefore, a matter of the first moment to be able to trace the modifications which it has undergone along the different lines of evolution which have culmi- nated in the existing types of animal life. The subject is a very large one, and can only be treated here in its broadest outlines. It is now very generally admitted by zodlogists that the Echinoderms (star-fishes, sea-urchins, etc.) owe the vadzal symmetry, which is one of the most obvious characteristics of their organisation, to their having been derived from bilaterally symmetrical ancestors, which became adapted to a fixed or sessile existence. If this view is correct, and there is good reason for supposing that it is, it follows that the majority of living Echingderms have secondarily 268 THE PROTOCHORDATA. lost their sessile mode of existence, and have again become free-living, retaining, however, their radial symmetry. At the present time the fixed habit of life is only retained by the members of one of the subdivisions of the Echino- derm class ; namely, the Crznozdea. Most genera of Crinoids (Akzzocrinus, Pentacrinus, etc.) remain fixed by a long, jointed stalk throughout life; but the well-known “feather-star,” a ' * R 1 pos Siok Vor * apt 4 ; Hl 4 Jn ie ye %, + 7 TA a at ’ 7 | i ; i ~.e% 5 . [ E > f. ' : ky . j ‘ zg -_ t he < M, o4 = * “93 ‘ Ms f a? : ’ “A wu ( Y \ , “4 " ' +AAY Rta +) 7 y ’ Le . vi : vi." 7 j >. & & A ipod, ee | 5 os | fits of} uy ui P} : He / Rate tn Ek % he Fett yea Pe ee athS PT t's “he oe > aed, : i : . a 7 ‘ s/t} : WIAALG f ; Ry ail iy years eh , Fil ie ) : * ye a Pa ‘ . * f o- % “ne = ’ i \ p NG tr: a>). ; oy Dt vA 7 > ioe f ‘ 5? ahs ype L Plt ol ane he ic ra : aie i" a HK ! pe ‘ 7 by, i Fe ; , . » “ rou 7 . mi vie LE: by ve) ve Grukis % { wees A - “ae , ied cs 7 4, wise ye ; r Ve 7 r F : i i vrs ra a ay “i BAYES dts ‘hes «th ‘+ 8 ‘te ¥ ‘ or an Apu er ¥ Fade oes Bh he tae at eae JED A, (oer es We ; : ‘ m4 =P cies i r a > ae » . hae 9 Pe f we i nm re re. % : Vie iy hi Mil t Anh ‘ : : ey 1. ASP PT AA Ba ON A 5 tha dig if ie Mi AD) . ’ ’ M F “ Dy. tg Pa. oat c : ie. H m2 r 7: a j a al ; Sie eed ee (eee NC IO coy Wy A a ‘ i j 7 a ee he 4 TAs Ruud St NAL ih ne we ig Nai whet rot fii | Aaa ‘4 ; e c sal an: 7 Vale aA. ee pea vel, thas Sa aps ‘i at we : Lae sve " aa uy ; ¢ Lobe) gat . PS, ‘4 < 4 f ‘ . A Ave 7 it pie o ay - o ‘ee i ba rae : A lirev, ah i Ha mn a a8 ar bu t : ; fe ¥ _ tN REFERENCES: INTRODUCTION. Carus, J. Victor. Geschichte der Zoologie. Miinchen, 1872. DoOHRN, ANTON. Der Ursprung der Wirbelthiere und das Prin- cip des Functionswechsels. Leipzig, 1875. HAECKEL, ERNST. Anxnthropogenie oder Entwickelungsgeschichte des Menschen. Leipzig, 1874; 4th Edit., 1891. LANKESTER, E. Ray. Article “Vertebrata.” Encycl. Brit., gth Edit. Republished in “ Zoodlogical Articles,” London, 1891. PERRIER, EDMOND. La Philosophie Zoologigue avant Darwin, 2d Edit. Paris, 1886. SEMPER, CARL. De Verwandtschafisbeziehungen der gegheder- ten There. Parts I. toIII. Wiirzburg, 1875-76. I. AND II. ANATOMY OF AMPHIOXUS.* ANDREWS, E. A. The Bahama Amphioxus (preliminary ac- count). Johns Hopkins University Circulars, Vol. XII. p. 104. June, 1893. ANDREWS, E. A. An Undescribed Acraniate: Asymmetron lucayanum. Studies from the Biol. Lab. Johns Hopkins Uni- versity, Vol. V. No. 4. 1893. pp. 213-247. Plates XIII.- XIV. Contains bibliography of systematic and faunistic works on Amphioxus. ANTIPA, GR. Ueber die Beziehungen der Thymus zu den soge- nannten Kiemenspaltenorganen bet Selachiern. Anat. Anz. VII. 1892. pp. 690-692. One figure in text. * This bibliography does not by any means include all that has been written on the anatomy of Amphioxus. Some of the older and shorter works, as well as some of those relating to special points of histological detail, have been omitted, as they are fully dealt with in many of the memoirs here cited. 295 296 | fe) 1 12 8. 14 15 16 17 18 19 20 REFERENCES. BALFourR, F. M. A Preliminary Account of the Development of the Elasmobranch Fishes. Quarterly Jour. Micro. Sc. XIV. N.S. 1874. pp. 323-364. Plates 13-15. Paper in which Balfour first published his discovery of the seg- mental origin of excretory tubules. This was made out also in the same year by Semper and. Schultz. (Vide infra, Schultz.) BALFour, F. M. On the Origin and History of the Urino- genital Organs of Vertebrates. Jour. of Anat. and Physiol. X. 1875. pp. 17-48. Eight figures in text. Amplification of his pre- vious work, with bibliography up to date. BALFour, F. M. The Development of Elasmobranch Fishes. Development of the Trunk. Jour. of Anat. and Physiol. XI. 1876. pp. 128-172. Plates 5 and 6. First account of origin of paired limbs from continuous epiblastic thickenings. BALFour, F. M. A Monograph on the Development of Elasmo- branch Fishes. London, 1878. BEDDARD, FRANK EvERS. Ox the Occurrence of Numerous Nephridia in the Same Segment in Certain Earthworms, and on the Relationship between the Excretory System in the Annelida and in the Platyhelminths. Quarterly Jour. Micro. Sc. XXVIII. N.S. 1888. pp. 397-411. Plates 30-31. Contains discovery of neph- ridial network in Pericheta. BENHAM, W. BLAXLAND. The Structure of the Pharyngeal Bars of Amphioxus. Quarterly Jour. Micro. Sc. XXXV.N.S. 1893. pp. 97-118. Plates 6-7. BOURNE, ALFRED GIBBS. Contributions to the Anatomy of the Hirudinea. Quarterly Jour. Micro. Sc. XXIV. N.S. 1884. pp- 419-506. Plates 24-34. Contains discovery of nephridial network in Pontobdella. BOvERI, THEODOR. Ueber die Niere des Amphioxus. Miin- chener Medicin. Wochenschrift. No. 26. 1890. Sep. Abd. pp. I-13. Two figures in text. (Preliminary note.) BOVERI, THEODOR. Die Mierencandalchen des Amphioxus. Ein Beitrag zur Phylogenie des Urogenitalsystems der Wirbelthiere. Zoolog. Jahrbiicher. Abth. fiir Morphol. V. 1892. pp. 429-510. Taf. 31-34 and five figures in text. Costa, O. GABRIELE. Cenni zoologict ossia descrizione som- maria delle specie nuove adi aninrali discoperti in diverse contrade del regno nell’ anno 1834. Napoli, 1834. See also Fauna del regno di Napoli. 1839-50. CuENoT, L. Etudes sur le sang et les glandes lymphatiques dans la série animale. Archives de zool. expérimentale, XIX. 1891. Amphioxus. pp. 55-56. iS) ta 23 24 25 26 27 28 ‘29 REFERENCES. 297 Notes absence of blood-corpuscles in Amphioxus. Those described by previous authors must therefore require another ex- planation. DOHRN, ANTON. Studien zur Urgeschichte des Worbelthier- korpers. IV. Section 5. Entstehung und Bedeutung der Thymus der Selachier. Mitth. Zool. Stat. Neapel. V. 1884. pp. 141-151. Taf. 8. Figs. 1 and 2. E1sic, HuGco. Die Segmentalorgane der Capitelliden. Mitth. Zool. Stat. Neapel. I. 1879. pp. 93-118. Taf. IV. Discovery of numerous nephridia in single segments and an- astomoses between successive nephridia. EMERY, CARLO. Le specie del genere Fierasfer nel Golfo di Napoli. 2d Monograph in the “ Fauna und Flora des Golfes von Neapel.” Leipzig, 1880. EMERY, CARLO. Zur Morphologie der Kopfniere der Teleostier. Biologisches Centralblatt, I. 1881. pp. 527-529. See also Zoologischer Anzeiger, VIII. 1885. pp. 742-744. FuSARI, ROMEO. Settrag zum Studium des peripherischen Nervensystems von Amphioxus lanceolatus. Internationale Mo- natsschrift fiir Anatomie und Physiologie, VI. 1889. pp. 120-140. Taf. VII.-VIII. GoopsIR, JOHN. Ox the Anatomy of Amphioxus lanceolatus. Transactions of the Royal Society of Edinburgh, Vol. XV. Part I. 1841. pp. 241-263. GRENACHER, H. Settradge zur nahern Kenntniss der Muscu- latur der Cyclostomen und Leptocardier. (Leptocardia proposed by Haeckel as a classificatory name on account of the simple tubular “heart” of Amphioxus.) Zeitschr. fiir Wiss. Zoologie, XVII. 1867. pp. 577-597. Taf. XXXVI. First isolation of muscle-plates of Amphioxus. GUNTHER, ALBERT. Synopsis of Genus Branchiostoma. In Report on Zool. Collections of H. M.S. Alert. 1881-82. pp. 31- 33. London, 1884. ; HATSCHEK, BERTHOLD. Die Metamerie des Amphioxus und des Ammocetes. Verh. Anat. Gesellschaft, 6th Versammlung. Wien, 1892. pp. 137-161. Eleven figures in text. 29 61s. HATSCHEK, BERTHOLD. Zur Metamerie der Wirbelthiere. - 30 Anat. Anz. VII. Dec. 1892. pp. 89-91. Hux ey, T. H. Preliminary Note upon the Brain and Skull of Amphioxus lanceolatus. Proceedings of the Royal Society, XXIII. 1874. pp. 127-132. Points out that in Myxine and Ammoceetes a velum is present separating the buccal (stomodceal) from the branchial cavity. 298 31 32 33 34 35 36 37 38 REFERENCES. The resemblance of the buccal cavity and tentacles (cirri) of Ammoceetes to the corresponding parts in Amphioxus is so close that there can hardly be any doubt the two are homologous. The anterior end of the nerve-tube of Amphioxus corresponds to the lamina terminalis of the craniate Vertebrates. Huxtey, T. H. Ox the Classification of the Animal Kingdom. Journal of the Linnzan Society (London), XII. 1876. pp. 199- 226. (Read 3d Dec., 1874.) Section on “efical,” p. 216 et seg. Atrial cavity of Amphi- oxus and Ascidians is an epiccel like the opercular cavity of the Amphibian tadpole. KOLLIKER, ALBERT. Ueber das Geruchsorgan von Amphioxus. Miiller’s Archiv fiir Anat. Physiol., etc. 1843. pp. 32-35. Taf. Ey igs Discovery of olfactory pit and first description of the spermatozoa of Amphioxus. KOPPEN, Max. Settrage zur vergleichenden Anatomie des Centralnervensystems der Worbelthiere. Zur Anatomie des Lidechsengehirns. Morphologische Arbeiten (Schwalbe), I. 1892. pp- 496-515. Taf. 22-24. Contains discovery of giant-fibres in caudal portion of spinal cord of Lacerta viridis. KouHL, K. Lenzge Bemerkungen iiber Sinnesorgane des Amphi- oxus lanceolatus. Zool. Anz. 1890. pp. 182-185. States that sometimes there is a shallow olfactory groove on the right side as well as that in the left. Such grooves are often due to artificial crumpling, and the observation requires confirmation. KRUKENBERG, C. FR. W. Zur Kenntnts des chemischen Baues von Amphioxus lanceolatus und der Cephalopoden. Zool. Anz. 1881. pp. 64-66. See also HOPPE-SEYLER’S reply. pp. 185-187. Compare also CUENOT (supra). KUPFFER, CARL VON. Studien zur vergleichende Entwick- lunesgeschichte des Kopfes der Kranioten,l. Die Entwicklung des Kopfes von Acipenser sturio an Medianschnitten untersucht. 95 pp. 8°. g Tafeln. Miinchen und Leipzig, 1893. Contains also a chapter on brain of Amphioxus, with figures. LANGERHANS, PAUL. Zur Anatomie des Amphioxus lanceolatus. Archiv fiir mikroskopische Anatomie, XII. 1876. pp. 290-348. Taf. XII.-XV. Standard work on the histology of Amphioxus. LANKESTER, E. Ray. On Some New Points in the Structure of Amphioxus and their Bearing on the Morphology of Vertebrata. Quarterly Jour. Micro. Sc. XV. N.S. 1875. pp. 257-267. 39 40 4I REFERENCES. 299 LANKESTER, E. Ray. Contributions to the Knowledge of Amphi- oxus lanceolatus, Yarrell. \b., Vol. XXIX. 1889. pp. 365-408. Five plates. Lworr, Basitius. Uber den Zusammenhang von Markrohr una Chorda beim Amphioxus und dahnliche Verhdaltnisse bet Anneliden. Zeitschrift fur wiss. Zoologie. Bd. 65. 1893. pp. 299-308. Taf. XVII. Describes those supporting fibres of the spinal cord of Amphi- oxus which descend in successive paired groups to the notochordal sheath and penetrate the latter in order to insert themselves on the inner surface of the sheath. The openings in the notochordal sheath of Amphioxus, through which the ventral supporting fibres pass, were first observed by WILHELM MULLER in 1871. (W. MULLER, Ueber den Bau der Chorda dorsalis. Jenaische Zeit- schrift, VI. 1871. pp. 327-354.) See also PLATT (infra) and LworF (88). Latter contains complete bibliography of literature relating to structure of notochord. Mayer, Paut. Uber die Entwicklung des Herzens und der grossen Gefassstamme bet den Selachiern. Mitth. Zool. Stat. Neapel. VII. 1887. pp. 338-370. Taf. 11-12. MEYER, EDUARD. Studien iiber den Korperbau der Anneliden. Mitth. Zool. Stat. Neapel. VII. 1887. pp. 592-741. Taf. 22-27. 42 bts. MOREAU, CAMILLE. echerches sur la Structure de la Corde 43 pods 46 dorsale de VAmphioxus. Bull. Acad. Belg. Tome 39. No. 3. 1875. 22pp. One plate. MULLER, WILHELM. Ueber die Stammesentwicklung des Sehorgans der Wirbelthiere. 76 pp. Five plates. 4°. Leipzig, 1874. MULLER, WILHELM. Ueber das Urogenitalsystem des Amphi- oxus und der Cyclostomen. Jenaische Zeitschr. fiir Naturwissen- schaft, Bd. II. (neve Folge). 1875. Sep. Abdruck. pp. 1-38. Two plates. This is the important work in which the pronephros and mesonephros were for the first time clearly distinguished from one another. The author was, however, in error regarding Johannes Miiller’s renal papillae of Amphioxus. MULier, JOHANNES. Uber den Bau und die Lebenserscheinun- gen des Branchiostoma lubricum Costa, Amphioxus lanceolatus, Yarrell. Berlin, 1844. 4°. 40 pp. Five plates. Read at the konigl. Akademie, 1841. NANSEN, FriptTjJoF. The Structure and Combination of the His- tological Elements of the Central Nervous System. Bergens Museums Aarsberetning for 1886. Bergen, 1887. 48 49 REFERENCES. OWSJANNIKOW, PHILIP. Ueber das Centralnervensystem des Amphioxus lanceolatus. Bulletin de Acad. imp. des Sciences de St. Pétersbourg, Tome XII. 1868. pp. 287-302, with one plate. Also in Mélanges Biologiques, T. VI. pp. 427-450. Introduced a method of maceration by which he was able to shake out the central nervous system and thus isolate it from the body. In this way he was able to correct the erroneous descrip- tions of de Quatrefages and others (who stated that there were ganglionic enlargements in the spinal cord), and to discover the alternate arrangement of the spinal nerves. PLATT, JULIA B. /72bres connecting the Central Nervous System and Chorda in Amphioxus. Anat. Anz. VII. 1892. pp. 282- 284. Three figures in text. PoLuarRD, E. C. A Mew Sforozoin in Amphioxus. Quarterly Jour. Micro. Sc... XXXIV. N.S. 1893. pp. 311-316. Plate XXIX. Unicellular parasites in intestinal epithelium. 49 bts. POUCHET, GEORGES. On the Laminar Tissue of Amphioxus. 5oO Le! 52 Quarterly Jour. Micro. Sc. XX. N.S. pp. 421-430. Plate XXIX. DE QUATREFAGES, ARMAND. J/émotre sur le systéme nerveux et sur Vhistologie du Branchiostome ou Amphioxus. Annales des ~ sciences nat. Zoologie. 3d series. IV. 1845. pp. 197-248. Plates 10-13. First observation of passage of ova through atriopore; and discovery of the peripheral ganglion-cells in connexion with the cranial nerves. RATHKE, HEINRICH. Semerkungen iiber den Bau des Ampht- oxus lanceolatus, eines Fisches aus der Ordnung der Cyclostomen. Konigsberg, 1841. 4°. pp. 1-38. One plate. REtzius, GusTAvV. Zur Kenntniss des centralen Nervensystems von Amphioxus lanceolatus. Biologische Untersuchungen. Neue Folge II. pp. 29-46. Taf. XI.-XIV. Stockholm, 1890. 52 d2s. RetTzius, Gustav. Das hintere Ende des Ruckenmarks und 53 54 sein Verhalten zur Chorda dorsalis bet Amphioxus lanceolatus. Verh. Biol. Vereins. (Biologiska Foreningens Forhandlingar.) Stockholm. Bd. IV. pp. 10-15. 9g figs. 1891. RouvDE, EMIL. Héstologische Untersuchungen tiber das Nerven- system von Amphioxus lanceolatus. In Anton Schneider’s Zoo- logische Beitrage. Bd.II., Heft 2. Breslau, 1888. pp. 169-211. Plates XV.—XVI. Standard work on the central nervous system of Amphioxus. Rowon, JOSEF Victor. Untersuchungen iiber Amphioxus lanceolatus. Ein Beitrag zur vergleichenden Anatomie der Wur- 55 56 57 58 59 - 60 61 REFERENCES. 301 belthiere. In Denkschriften der Math.-Naturwiss. Classe der kais. Akad. der Wissenschaften. Bd. XLV. Wien, 1882. 64 pp. 4°. Six plates. Relates chiefly to nervous system. Describes also the smooth muscle-fibres in wall of pharynx, etc. Finds that the majority of sensory nerve-fibres to the skin end freely between the cells of the ectoderm in bush-like ramifications. For the rest, see NANSEN ROHDE, RETzIUS, and FUSARI. RotpH, W. Untersuchungen iiber den Bau des Amphioxus lanceolatus. Morphologisches Jahrbuch, II. 1876. pp. 87-164. Taf. V.-VII.; also figures in text. RUCKERT, JOHANNES. L£xtwickelung der Excretionsorgane. Ergebnisse der Anatomie und Entwicklungsgeschichte (Merkel und Bonnet), I. 1891. pp. 606-695. Includes an extensive bibli- ography. SCHNEIDER, ANTON. Settrage zur vergleichenden Anatomie und Entwicklungsgeschichte der Wéorbelthiere. L. Amphioxus lanceolatus. pp. 3-31. Taf. XIV.-XVI. 4°. Berlin, 1879. SCHULTZ, ALEXANDER. Zur Entwickelungsgeschichte des Sela- chieretes. Archiv. fiir Mikr. Anat. XI. 1875. pp. 569-580. Mat. 34. Preliminary notes of both Sewer and Schultz, regarding the segmental origin of the excretory tubules, were published in the Centralblatt fiir Medicinische Wissenschaft, 1874. SEMON, RICHARD. Studien tiber den Bauplan des Urogenital- systems der Wirbelthiere; dargelegt an der Entwickelung dieses Organsystems bet Ichthyophis glutinosus. Jenaische Zeitschrift, XXVI. 1891. pp. 89-203. Taf. I.-XIV. SPENGEL, J.W. Beitrag zur Kenntniss der Kiemen des Amphi- oxus. Zool. Jahrbiicher. Abth. fiir Morphol. IV. 1890. pp. 257- 296. Taf. 17-18. SPENGEL, J. W. Benham’s Kritik meiner Angaben iiber die temen des Amphioxus. Anat. Anz. VIII. 1893. pp. 762-765. STIEDA, LupwiG. Studien iiber den Amphioxus lanceolatus. Mém. de l’Acad. Impériale des Sciences de St. Pétersbourg, 7th series, Vol. XIX. No.7. 7opp. Four plates. 1873. Contains some good observations on the central nervous system. First to show that the split-like structure above central canal did not correspond to the posterior fissure of the vertebrate spinal cord, but was a portion of the original central canal itself, the lumen of which had been partially obliterated by approximation of its walls. First identification of ventral (motor) roots of spinal nerves in Amphioxus. 302 63 64 65 66 67 68 69 REFERENCES. THACHER, JAMES K. Median and Paired Fins ; a Contribution to the History of Vertebrate Limbs. Transactions Connecticut Academy, III. No. 7. 1877. pp. 281-310. Plates 49-60. WEIsS, F. Ernest. L2xcretory Tubules in Amphioxus lanceolatus. Quarterly Jour. of Micro. Sc. XXXI. N.S. 1890. pp. 489-497. Plates 34-35. VAN WIJHE, J].W. Ueber Amphioxus. Anat. Anz. VIII. 1893. pp. 152-172. VAN WIJHE, J. W. Die Kopfregion der Crantoten beim Amphi- oxus, nebst Bemerkungen iiber die Wirbeltheorie des Schadels. Anat. Anz. IV. 1889. pp. 558-566. VAN WIJHE, J. W. Ueber die Mesodermsegmente des Rumpfes und die Entwicklung des Excretionssystems bet Selachiern. Archiv. f. Mikr. Anat. XXXIII. 1889. pp. 461-516. Taf. 30-32. WILLEY, ARTHUR. Report on a Collection of Amphioxus, made by Professor A. C. Haddon, in Torres Straits, 1888-89. Quarterly Jour. Micro. Sc. XXXV.N.S. January, 1894. pp. 361-371. One figure in text. Branchiostoma cultellum. Peters. TT: DEVELOPMENT OF AMPHIOXUS. AYERS, HowarpD. Sdellostoma Dombeyt, Lac. A Study from the Hopkins Marine Laboratory. Biological Lectures, Marine Biological Laboratory, Woods Holl. 1893. No. VII. Boston, 1894. 69 dts. BERT, PAUL. Ox the Anatomy and Physiology of Amphioxus. Annals and Mag. of Nat. Hist., 3d Series. Vol. XX. 1867. pp- 302-304. (Translated from Comptes Rendus. Aug. 26th, 1867. pp. 364-367.) Breeding season of Amphioxus at Arcachon is from March to May. Was the first to observe the ejection of the sperm through the atriopore. Calls attention to remarkable lack of regenerative power in Amphioxus. Individuals cut in two will live for several days, but will not regenerate. “If the extremity of the body of an Amphioxus be cut off, the wound does not cicatrize; on the contrary, the tissues become gradually disintegrated. I have seen animals, with only the tail mutilated, become gradually eaten away up to the middle of the branchial region, and live thus without any intestines, without abdominal walls, and without branchiz for several days.” These observations of Paul Bert are 7O 71 72 73 74 75 76 77 78 79 REFERENCES. 303 capable of easy confirmation, and should be borne in mind in view of the extraordinary regenerative power which Wilson dis- covered in the segmentation stages of the embryo. Boveri, THEODOR. Uder die Bildungsstitte der Geschlechts- driisen und die Entstehung der Genitalkammern beim Amphi- oxus. Anat. Anz. VII. 1892. pp. 170-81. Twelve figures. DoHRN, ANTON. Studien zur Urgeschichte des Wirbelthier- korpers. Ill. Die Entstehung und Bedeutung der Hypophysis bei Petromyzon Planert. Mitth. Zool. Stat. Neapel. IV. 1882. Dourn, ANTON. Studien, VIII. De Thyreoidea bei Petromy- zon, Amphioxus und Tunicaten. Ib. VI. 1885. Dohrn lays unnecessary stress upon the fact that often in transverse section, especially in the anterior region of the pharynx, the endostyle of Amphioxus projects up into the cavity of the pharynx in the form of a convex lens-shaped ridge. This is merely due to the muscular contraction of the pharynx, which almost invariably takes place when Amphioxus is placed in a killing reagent. It is, therefore, not an anatomical feature of any significance. DouRN, ANTON. Studien, XII. Zhyreotdea und Hypobran- chialrinne, Spritzlochsack und Pseudobranchialrinne bet Fischen, Ammocetes und Tuntkaten. \b. VII. 1887. Dourn, ANTON. Studien, XIII. Uber Nerven und Gefiisse bet Ammocetes und Petromyzon Planerz. \b. VIII. 1888. FRORIEP, AuGUST. ELxtwickelungsgeschichte des Kopfes. Ergebnisse der Anat. und Entwickelungsgesch (Merkel und Bonnet), I. 1891. pp. 561-605. Eleven figures. Includes an extensive bibliography. HATSCHEK, BERTHOLD. Studien iiber Entwicklung des Amphi- oxus. Arbeiten a. d. Zool. Institute. Wein, 1881. 88 pp. Nine plates. HATSCHEK, BERTHOLD. J/@tttheilungen iiber Amphioxus. Zoologischer Anzeiger, VII. 1884. pp. 517-520. Olfactory pit, sense-organ of preoral pit, anterior preoral “ nephridium.” HATSCHEK, BERTHOLD. Uber den Schichtenbau von Amphi- oxus. Anat. Anz. III. 1888. pp. 662-667. Five figures. Origin of sclerotome, etc. KASTSCHENKO, N. Zur Entwicklungsgeschichte des Selachier- embryos. Anat. Anz. III. 1888. pp. 445-467. One of the first to bring forward definite embryological facts to prove that the anterior (pra-auditory) head-cavities of VAN WIJHE (Ueber die Mesodermsegmente, etc., des Selachierkopfes. Amster- 304 80 81 83 84 85 86 REFERENCES. dam, 1882) are not homodynamous with the true somites. He was followed in this respect by RABL (Theorie des Mesoderms. Morphologisches Jahrbuch, XV. 1889). KORSCHELT, E., und HEIDER, K. Lehrbuch der vergleichen- den Entwicklungsgeschichte der wirbellosen Thiere. 3a Heft. Jena, 1893. KOWALEVSKY, ALEXANDER. Lutwicklungsgeschichte des Am- phioxus lanceolatus. Mém. de l’Acad. Imp. des Sciences de St. Pétersbourg..’ VIT?* Series) F2> XI> No: 4. 16672)7 dehree plates. IKOWALEVSKY, ALEXANDER. lWWeztere Studien iiber die Ent- wicklungsgeschichte des Amphioxus lanceolatus, nebst einem Beitrage zur Homologie des Nervensystems der Wiirmer und Wirbelthiere. Arch. f. Mikr. Anat. XIII. 1877. pp. 181-204. Two plates. Among the definite discoveries communicated by Kowalevsky in these two memoirs may be mentioned the following: General features of segmentation and gastrulation, origin of mesoderm from archenteric pouches, unique method of formation of nerve-tube (see text), origin of notochord, neurenteric canal, asymmetrical origin of gill-slits and mouth, and zz fart the metamorphosis. KUPFFER, CARL VON. Die Entwicklung von Petromyzon Planerz. Arch. f. Mikr. Anat. XXXV. 1890. pp. 469-558. Six plates. Origin of head-cavities, hypophysis, etc. KUPFFER, CARL VON. Die Entwicklung der Kopfnerven der Vertebraten. Verhandl. Anat. Gesellschaft in Miinchen. 1891. pp. 22-55. Eleven figures. (Erganzungsheft zum Anat. Anz. VIEW SIcor.) Ammoceetes (see Fig. 92 in text). KUPFFER, CARL VON. Studien zur vergleichende Entwick- lungsgeschichte des Kopfes der Kranioten I. Die Entwicklung des Kopfes von Acipenser sturio an Medianschnitten untersucht. pp. 95. Nine plates. Seven figures in text. Miinchen and Leipzig, 1893. Important contribution to the delimitation of the wall of the brain. On page 84 is a reconstruction of head-cavities of Am- moccetes (see Fig. 72). Figs. 21 and 22 in the plates repre- sent cerebral vesicle of Amphioxus. (Cf. Fig. 51.) LANKESTER, E. Ray, and WILLEY, A. The Development of the Atrial Chamber of Amphioxus. Quarterly Jour. Micro. Sc. XXXI. 1890. pp. 445-466. Four plates. 87 88 89 go / 9I g2 93 REFERENCES. 305 LEUCKART, RUDOLPH, und PAGENSTECHER, ALEX. Untfer- suchungen tiber niedere Seethiere. Amphioxus Jlanceolatus. Miiller’s Archiv f. Anat. u. Physiol. 1858. pp. 558-569. Taf. XVIII. Description of larve of Amphioxus taken off Heligoland. Drew attention to larval asymmetry, and to the existence of the brain-ventricle (cerebral vesicle). In absence of knowledge of early development their interpretation of many of the structures (especially preoral pit, mouth, and _ gill-slits) was incorrect. Latter applies also to Schultze’s observations. Lworr, Basitius. Uber Bau und Entwicklung der Chorda von Amphioxus. Mittheilungen a. d. Zool. Station. Neapel. IX. 1891. pp. 483-502. One plate. Consult this memoir for previous literature on histology of notochord. Lworr, BasILius. Ueber einige wichtige Punkte in der Ent- wicklung des Amphioxus. Biologisches Centralblatt, XII. 1892. pp- 729-744. Eight figures. Notes absence of mesodermal “ pole-cells.” From frequency of mitoses in dorsal ectoderm of gastrula, concludes that the material destined to form dorsal wall of archenteron, from which notochord and myoccelomic pouches arise, grows in from the ectoderm round dorsal lip of blastopore. Hence notochord and mesoderm are essentially derived from ectoderm! MARSHALL, A. MILNES. Vertebrate Embryology. London, 1893. MULLER, JOHANNES. Uber die Fugendzustande einiger See- thiere. _Monatsbericht der konigl. preuss. Akad. der Wissen- schaften zu Berlin. 1851. pp. 468-474. First accurate description of larva of Amphioxus, p. 474. In 1847 Johannes Miiller obtained a young Amphioxus of 2} mm. at Helsingfors. He says that the appearance of the gill-slits was peculiar, in that there were two rows of slits in the pharyngeal wall, placed one above the other. In the upper row were five round slits, while the lower slits were vertically elongated and were fourteen in number. Headds that it was doubtful whether it represented the young “ Branchiostoma lubricum ” or belonged to a new species. MULLER, WILHELM. Ueber die Hypobranchialrinne der Tunt- katen und deren Vorhandensein bet Amphioxus und den Cyklo- stomen. Jenaische Zeitschrift f. Naturwiss. VII. 1873. pp. 327-332. PLATT, JuLIA B. Further Contribution to the Morphology of the Vertebrate Head. Anat. Anz. VI. 1891. pp. 251-265. 96 97 98 99 I0o Io! 103 REFERENCES. RaB1, CarL. Uber die Differenzierung des Mesoderms. Anat. Anz. III. 1888. pp. 667-673. Eight figures. Discovery of the sclerotome-diverticulum in embryo of Pristiurus. RIcE, HENRY J. Observations upon the Habits, Structure, and Development of Amphioxus lanceolatus. American Nat. XIV. 1880. pp. 171-210. Plates 14 and 15. Author was the first to find Amphioxus in Chesapeake Bay. With regard to development, he gives some fairly good figures of larvee, and observed some of the more obvious features of the metamorphosis, as already described by Kowalevsky. RUCKERT, JOHANNES. Ueber der Entstehung der Excretions- organe bet Selachiern. Arch. fiir Anat. u. Physiol. (Anatomische Abtheilung). 1888. pp. 205-278. Three plates. Contains also the discovery of segmental origin of gonads. SCHNEIDER, ANTON. JSezttrage zur vergleichenden Anatomie und Entwicklungsgeschichte der Wirbelthiere, IT. Anatomie und Entwickl. von Petromyzon und Ammocetes. 4°. Ten plates. Berlin, 1879. Figure of the ciliated grooves in pharynx of Ammoccetes, at page 84. SCHULTZE, MAx. Seobachtung junger Exemplare von Amphi- oxus. Zeit. f. Wiss. Zool. III]. 1851-2. pp. 416-419. Two larve from Heligoland. Good description of structure of notochord. VAN WIJHE, J. W. Ueber Amphioxus. Anat. Anz. VIII. 1893. pp- 152-172. WILLEY, A. Ox the Developmeut of the Atrial Chamber of Amphioxus. (Preliminary communication.) Proceedings of the Royal Society, XLVIII. 1890. pp. 80-89. Wittey, A. Zhe Later Larval Development of Amphioxus. Quarterly Jour. Micro. Sc. XXXII. 1891. pp. 183-234. Three plates. WILSON, EDMUND B. On Multiple and Partial Devolopment in Amphioxus. Anat. Anz. VII. 1892. pp. 732-740. Eleven figures. In this and the following more detailed paper, the author describes and interprets a remarkable series of experiments on the artificial production of twins and dwarfs. Besides this. there are many important observations on the normal cleavage of the egg. WILson, EDMUND B. Amphioxus and the Mosaic Theory of Development. Journal of Morphology, VIII. 1893. pp. 579- 638. Ten plates. 104 REFERENCES. 307 ZIEGLER, H. Ernst. Der Ursprung der mesenchymatischen Gewebe bei den Selachiern. Archiv f. Mikr. Anat. XXXII. 1888. pp. 378-400. One plate. Independent discovery of sclerotome-diverticulum. (See Rabl.) IV. ASCIDIANS. For bibliography relating to the Ascidians, see Professor W. A. HERD- 105 ~ 106 107 108 109 IIo MAN’S Reports on the Tunicata collected during the ‘‘ Challenger” expedition — Parts I.-III. 1882-88; and also KORSCHELT und HEIDER, “ Lerhbuch der vergleichenden Entwicklungsgeschichte der wirbellosen Thiere.” Heft III. Jena, 1893. Vi PROTOGHORDATES, -ETC. AYERS, HOWARD. Concerning Vertebrate Cephalogenesis. Jour. Morph. IV. 1890-91. pp. 221-245. BATESON, WILLIAM. JJemoirs on the Development of Balano- glossus. Quarterly Jour. Micro. Sc. Vols. XXIV.-XXVI. 1884-86. Brooks, W. K. The Systematic Affinity of Salpa in its Relation to the Conditions of Primitive Pelagic Life ; the Phylogeny of the Tunicata ; and the Ancestry of the Chordata. Part II. of Monograph of the Genus Salpa. Johns Hopkins University. Baltimore, 1893. BURCKHARDT, RUDOLF. De Homologieen des Zwischenhirn- daches und thre Bedeutung fiir die Morphologie des Hirns bet niederen Vertebraten. Anat. Anz. IX. 1894. pp. 152-155 and 320-324. Relates to neuropore of craniate Vertebrates. Author calls the lobus olfactorius impar of Kupffer, the recessus neuroporicus. CLAPP, CORNELIA M. Some Points in the Development of the Toad-fish (Batrachus Tau). Jour. Morph. V. 1891. pp. 494- gol. Observations on the double origin of mouth, made in 1889, not published in this paper. DaviporF, M. von. Ueber den “Canalis neurentericus anterior bet den Ascidien.” Anat. Anz. VIII. 1893. pp. 301-303. 308 REFERENCES. 1s DOHRN, ANTON. Studien zur Urgeschichte des Worbelthier- korpers, I. Der Mund der Knochenfische. Mitth. Zool. Stat. Neapel. III. 1881-2. pp. 253-263. ‘ 112 FIELD, GEORGE W. The Larva of Asterias vulgaris. Quarterly Jour. Micro. Sc. XXXIV. 1892. pp. 105-128. 113 FowLerR, G. HERBERT. The Morphology of Rhabdopleura Normant Allman. Festschrift fiir Rudolf Leuckart. pp. 293-297. Leipzig, 1892. II4 HARMER, S. F. See M’INTOSH. O05 HERDMAN, W. A. Article ‘‘ Tunicata.” Ency. Brit. gth ed., republished in “ Zoological Articles” by Lankester, etc. 116 HUuBRECHT, A. A. W. Article “ Nemertines.” Ency. Brit. oth ed., republished in “ Zodlogical Articles” by Lankester, etc. 116 67s. HUBRECHT, A. A. W. On the Ancestral Form of the Chordata. Quarterly Jour. Micro. Sc. XXIII. 1883. pp. 349-368. For later works on this subject see Notes to Chap. V. 117 KUPFFER, C. VON. Lntwickelungsgeschichte des Kopfes. In Merkeland Bonnet’s Ergebnisse der Anatomie und Entwickelungs- geschichte, II. 1893. pp. 501-564. 118 LANG, ARNOLD. Zum Verstandnis der Organisation von Cephalodiscus dodecalophus M°’Znt. Jenaische Zeitschrift f. Naturwiss. XXV._ 1891. 119 LANG, ARNOLD. Ueber den Einfluss der festsitzenden Lebens- weise auf die Thiere. Jena, 1888. 120 LANKESTER, E. Ray. Degeneration: a Chapter in Darwinism. Nature Series. London, 1880. Republished in “ The Advance- ment of Science; Occasional Essays and Addresses.” London, 1890. I2I LANKESTER, E. Ray.