On the Ciliary Mechanisms and Interrelationships of Lamellibranchs

On the Ciliary Mechanisms and Interrelationships of Lamellibranchs. PAET IV: Cuticular Fusion, with special reference to the Fourth Aperture in certain Lamelli- branchs. By Daphne Atkins, B.Sc. Marine Biological Laboratory, Plymouth.

With 11 Text-figures.

CONTENTS. PAGE INTRODUCTION 423 HISTOLOGICAL STRUCTURE OF THE JUNCTION .... 424 CUTICULAK FUSION IN RELATION TO THE FOURTH APERTURE IN CERTAIN LAMELLIBRANCHS ...... 436 Origin of the Fourth Aperture ...... 442 SUMMARY ...... 444

INTBODUCTION. A PECULIAR form of fusion, involving the cuticle only, has been found in a number of Lamellibranchs, but has been chiefly studied in the Solenidae. The positions so far discovered in which this type of fusion occurs are between the dorsal edges of the ascending lamellae of the outer demibranchs and the mantle or the visceral mass in Solen marginatus Montagu (=vagina), Ensis siliqua (L.), Ensis arcuatus (Jeffreys), Cultellus pellucidus (Pennant), Solecurtus scopula (Turton) (=candidus), Lutraria lutraria (L.) (=elliptica), and Tellina tenuis da Costa; between the dorsal edges of the ascending lamellae of the two inner demibranchs in Barnea parva (Pennant); and between the mantle lobes in the region between the pedal and fourth apertures in Ensis siliqua, Ensis arcuatus, and Cultellus pellucidus. 424 D. ATKINS

HlSTOLOGICAL STRUCTURE OF THE JUNCTION. Material was fixed in Bouin-Duboscq's fluid with the following formula: saturated picric acid in 90 per cent, alcohol, 2 parts; saturated corrosive sublimate (water), 3 parts; 40 per cent, formalin, 1 part; glacial acetic acid, 2 parts. The stains used were Heidenhain's iron haematoxylin counterstained with acid

m.f

TEXT-FIG. 1 A. Transverse section showing the method of junction of the mantle lobes in the mid-ventral region between the pedal and fourth apertures. A. Ensis siliqua (adult). X980. fuchsin; Mallory's triple stain; and eosin and light (Gatenby, 1928, p. 432): the latter was preferred as it clearly differentiated the cuticle. The epithelium of the parts concerned in this cuticular type of junction has generally, in fixed material, a distinctive appearance, the cells being hourglass-shaped owing to the development of spaces between them (Text-figs. 1, 3, 4). It is uncertain, however, to what extent this may be due to shrinkage on fixation: THE CILIARY MECHANISMS OF LAMELLIBRANCHS 425 intercellular spaces were rare in the specimen of Cultellus pellucidus sectioned (Text-fig. 2A). Extremely fine muscle- fibrils penetrate the cells, giving to them an appearance rather similar to that of ciliated cells in which the ciliary rootlets are clearly visible (cf. Text-figs. 1 and 11): ciliated epithelia sometimes also show intercellular spaces.

e. c. cu.

TEXT-FIG. 1 B. B. Ensis arcuatus (young specimen, 4 cm. long), cu., cuticle: showing a dark median line in (A); ex., hourglass-shaped epithelial cell; m.f., muscle-fibres; sp., space between the cells. Eosin and light green. x980.

The cells appear to increase in height with age, for in an Ensis siliqua, 1-8 cm. long, the cells of the epithelium concerned in the junction of the mantle lobes between the pedal and fourth aperture were 5 to 6^ high; in an Ensis arcuatus, 4 cm. long, they were 7 to 10/u. high (see Text-fig. 1 B) ; and in an adult Ensis siliqua, about 16 cm. long, they were 18 to 30/x high (see Text-fig. 1 A). The combined layers of cuticle f:A J.EXT-ITO. Z. Cultellus pellucidus. Transverse sections of the junction of the mantle lobes in the mid-ventral region between the pedal and fourth apertures. A. About 216 p anterior to the fourth aperture. B. Showing breaking down of the cells; about 60 p. in front of the fourth aperture, cu., cuticle; e.c, epithelial cell; m.f., muscle- fibres ; o.s.m., outer surface of mantle; sp., space between cells. Eosin and light green, x 980. THE CILIARY MECHANISMS OF LAMELLIBRANCHS 427 also increase in thickness with age. This latter specimen had been in the laboratory four days, and the mantle had begun to rupture, so it is possible that the stretched appearance of the cells may be due to some extent to the fact that they had suffered tension owing to the gaping of the valves, and did not recover in the piece removed for fixing. However, in the junction between the dorsal edge of the ascending lamella of the outer demibraneh and the mantle (Text-figs. 3, 4) in Ensis arcuatus (4 cm. long) the epithelial cells were rather higher than those of the junction between the mantle lobes in the same specimen, though in the former position they would suffer relatively less stress. It is noteworthy that the mantle is pro- duced into a ridge of varying height along the line of attachment of the demibranchs (Text-fig. 3): a similar ridge is found in Solen marginatus. In an adult Solen marginatus the cells of the epithelium concerned in the union between the outer demibranchs and the mantle were high and the fibrils running through them especially clear: this was also found in the same position in Solecurtus s c o p u 1 a. The epithelial cells of the junction between the two inner demibranchs in Barnea parva were tall and slender and the cuticle clearly visible. In Tellina tenuis though the junction between the upturned outer demibranchs and the visceral mass is of the cuticular type, at least in the posterior region, yet the cells are small, have not the distinctive hourglass- shape, and the cuticle between the two parts is extremely thin and difficult to distinguish. The chief interest of this type of junction lies in the cuticle of the two opposed surfaces which appear to have undergone fusion, the sinuosity of the cuticle of one side being faithfully followed by that of the opposite side (Text-figs. 1, 2, 4). A small cavity between the two layers in the material of Ensis siliqua sectioned was probably pathological, for it contained some amorphous material. In some instances a darker median line was visible in the cuticle between the two opposed epithelial layers, when stained with eosin and light green. This was observed in the junction of the mantle lobes between the pedal and fourth apertures in an adult Ensis siliqua (Text-fig. 1 A) 428 D. ATKINS

TEXT-FIG. 3. Ensis arcuatus (4cm.long). Section showing the attachment of the dorsal edge of the ascending lamella of the outer demibranch to the ridge of the mantle, asc.l., ascending lamella; m., mantle; m./., muscle-fibres; m.r., mantle ridge. Eosin and light green. X 450. and in the union of the outer demibranchs and the mantle in an Ensis arcuatus 4 cm. long (Text-fig. 4): curiously enough it was not visible in the junction between the mantle lobes in this small specimen, the appearance being as indicated THE CILIABY MECHANISMS OF LAMELLIBRANCHS 429 in Text-fig. 1 B, the median region of the cuticle staining less heavily than that on each side. A tendency for the cuticle to be faintly striated, with striae at right angles to the fused surface, and to separate in blocks corresponding to the cells is also evident. From the examination of sections, and from observations on living animals which will be given later, it is evident that the

— m.f

TEXT-FIG. 4. Ensis areuatus. Enlarged sketch of the junction of the dorsal edge of the ascending lamella of the outer demibranch and the mantle shown in Text-fig. 3. asc.L, ascending lamella; cu., cuticle, showing a dark median line; e.c., epithelial cell; m./., muscle-fibres; m.r., mantle ridge; sp., space between cells. Eosin and light green. X980. cuticular type of junction does not separate normally; when separation does occur it is due to accident or to tissue deterioration on the approach of death of the animal. Rupture seems to be usually due to breaking down of the cells (Text-fig. 2 B) and not to separation of the two cuticular layers. Rupture of the junction between the outer demibranehs and the mantle, or the visceral mass, and between the two inner demibranehs has not been observed, only of that between the mantle lobes in Ensis and Cultellus on unhealthy gaping of the valves, when the mantle in the ventral region would be liable to suffer considerable strain. In Ensis siliqua the ascending lamella 430 D. ATKINS of the outer demibranch could not be separated from the mantle by pulling the demibranch. The difference in the form of the junction between the mantle lobes, and in the arrangement of the pallial muscles, in C u 11 e 1 -

B TEXT-FIG. 0. Cultellus pellucid u a. Transverse sections of the ventral region of the mantle to show the general appearance: A. About 200/x anterior to the fourth aperture. B. About 150 /J. posterior to the aperture (ciliated grooves are not present in sections close to the aperture), m.l., longitudinal muscles; m.r., radial muscles; m.t., transverse muscles; n.p., pallial nerve; p., periostracum. Eosin and light green, x 70. lus pellucidus in the mid-ventral line in front of, and behind, the fourth aperture is evident from Text-fig. 5; A being a section about 200/LI anterior and B one about 150^. posterior to it. Anteriorly the junction is cuticular, the epithelium of the two opposed surfaces being clearly visible, and the muscles of the one lobe not penetrating into the other; while posteriorly THE CILIAEY MECHANISMS OF LAMELLIBRANCHS 481 fusion is complete, no trace of fused epithelial surfaces being discernible, and muscles crossing the ventral region from one side to the other. A similar state of affairs is found in E n s i s si 1 iqua and Ensis arcuatus. Rupture of the mantle occurs only anterior to the fourth aperture where fusion is of the cuticular type, and not posterior to it where tissue fusion is complete. The weakness of the cuticular type of fusion would seem to lie not in the actual fusion of the cuticle, but in the arrangement of the muscles (contrast Text-fig. 5 A and B). In Solen marginatus, in which a fourth aperture is absent, the mantle lobes being united along the entire ventral surface, the fusion (Text-fig. 6) is throughout of the type found posterior to the fourth aperture in Ensis and Cultellus, that is, it is true tissue fusion. In this bivalve rupturing of the mantle in the mid-ventral line apparently does not occur when the animal becomes feeble and the valves gape, for specimens died with it still intact, but separating from the inner surface of the valves. Two Lamellibranchs with the fourth aperture near the posterior end of the animal, just below the inhalent siphon, have been examined, namely Lutraria lutraria and Thracia villosiuscula (Macgillivray). Both these species die with gaping valves, without rupture of the mantle in the mid-ventral line. In a Lutraria lutraria, 9 cm. long, the position of fusion of the mantle lobes was clearly visible in transverse sections of the ventral region taken about 10 mm. in front of the fourth aperture (Text-fig. 7). Contraction and contortion of the mantle in this region occurred to such an extent on fixation that the epithelium along the line of junction was thrown into complicated folds, and the cuticle could not be followed as a continuous line: Text-fig. 8 gives some idea of the appearance. In a small specimen less than 2 cm. long, the epithelial cells and their nuclei were somewhat easier to distinguish and the cuticle was clearly visible for short distances. The fusion of the mantle lobes between the fourth and pedal apertures in Lutraria lutraria seems to have proceeded much further, and is evidently much stronger, than in Ensis and Cultellus. It appears that fine muscle-fibres from one 432 D. ATKINS mantle lobe cross the junction into the opposite lobe, but it is impossible to be certain of this owing to the folding previously mentioned. In living Lutraria the position of the line of junction is visible as a double white line, stretching from the fourth to the pedal aperture.

TEXT-FIO. 6. Solen marginatus. Photograph of transverse section of the ventral region of the mantle about 10 mm. posterior to the pedal aperture. Heidenhain's iron haematoxylin and acid fuohsin. X ca. 37J. In Thraeia villosiuscula true tissue fusion of the mantle lobes is complete, transverse sections of the ventral region of the mantle just anterior to the fourth aperture of a specimen 23 mm. long showing no trace of fused epithelial surfaces (Text-fig. 9). It is curious that in Cultellus pellucidus and Solen TEXT-FIG. 7. Lutraria lutraria (9 cm. long). Photograph of transverse section of the ventral region of the mantle about 10 mm. anterior to the fourth aperture to show the line of fusion of the mantle lobes. Eosin and light green. X ca. 127J. 434 D. ATKINS marginatus in which the union of the dorsal edges of the ascending lamellae of the outer demibranchs and the mantle is by cuticular fusion, yet where organic fusion occurs between

cutcle; eph., epitheu; m.L, longitudinal verse muscles. Eosin and light green, x 450. the two inner demibranchs, that is from just posterior to the foot to the end of the gills, the fusion is complete with no indication of where it has occurred. All trace is absent even in the region where the organic junction is giving place to the THE CILIARY MECHANISMS OF LAMELLIBRANOHS 435 ciliary union which obtains between the two inner deniibranchs for a short distance behind the foot (for about 10 mm. in an adult Solen marginatus) and between them and the foot. A section through the transition region in Solen masrginatus is shown in Text-fig. 10. The bridge joining the two inner demi-

TEXT-EIG. 9. Thracia villosiuscula (23 mm. long). Transverse section of the ventral region of the mantle just anterior to the fourth aperture. n.p., pallial nerve; p., periostraoum. Heidenhain's iron haematoxylin. X 93J. branchs is narrow in Cultellus pellucidus, and it is apparently because of this that Eschrich (1931, pp. 573-4) found it easy to pull them apart. The union between the inner demibranchs and the foot, and inter se, is normally entirely ciliary in Ensis siliqua and Ensis arcuatus, but a small specimen of the latter species showed on sectioning slight local organic fusion, which was true tissue fusion, and not mere fusion of the cuticle (Text-fig. 11). From the state of affairs found between the inner demibranchs of Solen marginatus, Cultellus pellucidus, and the small specimen of Ensis arcuatus there is no indication NO. 315 F f 436 D. ATKINS that cuticular fusion is preceded in development by ciliary junction. It is probable that when the cuticular layers of the surfaces in contact are first secreted and probably in a semi- liquid or gelatinous state, strong permanent fusion or adhesion of the two layers takes place; a study of development of these bivalves would probably settle the question.

Solen marginatus (adult). Section through the junction of the dorsal edges of the ascending lamellae of the inner demibranchs in the region where ciliary junction is passing into organic junction. I.-B.C, infra- and S.-B.C, supra-branchial chamber. Eosin and light green, x 70.

CUTICULAR FUSION IN RELATION TO THE FOURTH APERTURE IN CERTAIN LAMELLIBRANCHS. Perhaps the most interesting occurrence of cuticular fusion is in those Solenidae which possess a fourth aperture, where the mantle lobes fuse between this and the pedal aperture. Certain references to the form of the union of the mantle lobes in this region in Cultellus pellucidus occur in the literature. Eschrich (1981, pp. 539-40) stated: 'Wenn wir von dem bereits erwahnten Spritzloch ausgehen, so ist der Mantel von hier aus nach vorn unverwachsen. Die Bander legen sich aber, einen hermetischen Versehluss bildend, dicht THE CILIARY MECHANISMS OF LAMBLLIBEANCHS 437

TEXT-FIO. 11. Ensis arcuatus (4 cm. long). Section through the ciliary junction of the dorsal edges of the ascending lamellae of the inner demibranchs. Slight organic union is present ventrally, but this is local and unusual. I.-B.C., infra- and S.-B.C, supra-branchial chamber. Eosin and light green. x980. 438 D. ATKINS aneinander. Vielleicht ist auch hier eine Verklebung der Kutikularschichten eingetreten.' And according to Graham (1934, p. 176) in this same species: 'Between the anterior end of this aperture (i.e. the fourth) and the anterior end of the bivalve itself is a region where the amount of fusion of the edges of the mantle folds is variable: occasionally specimens are en- countered in which this region gapes widely, but normally the edges are closely adpressed; it will be noted that this is mere contiguity: there is no real tissue connexion. The fourth aperture is therefore continuous with the rest of the pedal

According to Stenta (1903, p. 237) in Ensis siliqua between the fourth and pedal aperture 'sind die Mantelrander jederseits frei und stehen in loser Beruhrung miteinander'. From Haas's (1934, pp. 605-8) discussion of the form of the union described by Eschrich (1931) in Cultellus pellucid u s in relation to rupture along the mid-ventral line in this region in the Solenidae, and to the different conceptions of the structure of the inhalent siphon in L e d a (== N u c u 1 a n a)1 (Deshayes, 1857; Pelseneer, 1891, p. 168; 1911, p. 5; Stempell, 1898, p. 350), it is evident that he.considered that the adhesion of the cuticular layers is of such a nature that the junction may be easily dissolved without injury to the tissues, in fact he remarked that in the Solenidae ' das Reissen der scheinbaren Bauchnaht ist eben nichts weiter als die Loslosung beider verklebter Mantelrandlappen voneinander'. It would seem from sections of Cultellus pellucid us and Ensis siliqua that this rarely, if ever, occurs, but that the tearing is through the epithelial cells (see Text-fig. 2 B), sometimes close to the cuticle which is then left adhering entirely to the epithelium of the opposite mantle lobe. The fusion of the cuticular layers is of such a nature that it apparently can with- stand a greater stress than the cell-walls. In fact the separation of the mantle lobes in those Solenidae with a fourth mantle opening is not a reversible process; if it should by some accident

1 It has been shown in Part I (Atkins, 1936) that in Nuoulana (=Leda) minuta the junction of the two halves of the inhalent siphon in the mid-ventral line is a ciliary one, and not of the cuticular type. THE CILIARY MECHANISMS OF LAMELLIBRANCHS 439 occur in a healthy animal under normal conditions, it must be considered as an injury and as such needing repair. A number of observations have been made on the condition of the mantle between the fourth and pedal apertures in E n s i s siliqua and Ensis arcuatus. In these species the fourth aperture which is always present and bordered with tentacles— those of opposite sides interdigitatingwhen the aperture is closed —is roughly 8 to 10 mm. long, in specimens about 18 cm. long. On April 11, 1933, forty-four specimens of the two species of Ensis (siliqua and arcuatus), which had been gathered that afternoon from the Salcombe Estuary, South Devon, were received in the evening at the laboratory. Of this number nine showed an additional opening between the pedal and fourth apertures: it was found that in most of these the foot had been entirely or partly torn out in removing the animals from the sand. In five of these the fourth aperture was of normal extent (i.e. throughout its length it was bordered with tentacles), but the pedal aperture was enlarged along the ventral surface for distances between 2-5 and 8 cm. This was probably due to the end of the foot thrust in the sand not being in line with the length of the shell as the animal was pulled from the ground and thus tearing through the mantle lobes. In two, a split in the mantle occurred between the fourth and pedal apertures, but confluent with neither. In two, the fourth and pedal apertures were confluent, the valves gaping badly in one of these. The thirty-five Ensis with the fourth aperture normal were placed in bowls under circulation; after four days fourteen only retained it in this condition. Of twenty-five of the original number only six retained it in this condition after six days. On June 8, 1933, twelve Ensis (siliqua and arcuatus) were collected from Drake's Island in Plymouth Sound: they were mostly small, the largest being about 12 cm. long. The mantle between the fourth and pedal apertures was entire in all specimens. They were placed in bowls under circulation and in eight days all had suffered rupture of the mantle in this region. On October 20 of the same year a large number of Ensis (siliqua and arcuatus) were received from Salcombe Estuary. Seventy with the fourth aperture normal in size were 440 D. ATKINS placed in bowls under circulation. After seven days about twenty-eight retained it in this condition; these were mostly small specimens, but three (13, 14, and 15 cm. long) were especially heavy looking and brown in colour. The E n s i s obtained in October were more active than those of April and June, and perhaps owing to the cooler weather withstood laboratory conditions rather better than the two previous batches. The highest air temperature recorded in the laboratory during the week beginning April 11 was 66-8°F.; during that beginning June 8 was 73-6° F.; and during that beginning October 20 was 62-8° P. From the foregoing observations it would seem to be evident that in normal healthy Ensis the fourth aperture is small, and entirely bordered by tentacles; if it is enlarged beyond the region of these, then this is abnormal. The mid-ventral region of the mantle between the fourth and pedal apertures is a weak place in the Solenidae, as becomes evident when the animals are removed from their natural habitat. Ensis, unless provided with sand in which to burrow, does not live well in captivity, in fact it becomes unhealthy, and may even die, in a very few days; death appears to be hastened by rupturing of the mantle. Solen marginatus, which lacks a fourth aperture and the weak line in the mid- ventral region, when deprived of sand in which to burrow lives considerably longer than does Ensis. After a week or more the valves may gape slightly, but the mantle does not rupture and the animal lives on, though obviously unhealthy. The ease with which the mantle splits in the mid-ventral line between the pedal and fourth apertures on unhealthy gaping of the valves is no doubt responsible for statements as to the variation in the presence and extent of the fourth aperture in the Solenidae (Bloomer, 1902, p. 133; 1903, p. 44; Graham, 1931, p. 726). It seems possible that bivalves living more or less permanently buried in sand, mud, &c, rely to a certain extent on the pressure of the surrounding soil to keep the valves from gaping, and in consequence the adductor muscles, and those across the ventral region of the mantle where the margins are united, are not THE CILIARY MECHANISMS OF LAMELLIBRANCHS 441 sufficiently strong to prevent gaping of the valves on enfeeble- ment of the animals: the position also of the adductor muscles close to the hinge is one in which they are less efficient than they would be nearer the margins of the valves; burrowing bivalves, living in a sheltered position, have no need of tightly closing the shell (Douville, 1907, p. 97). It is difficult otherwise to account for their withstanding laboratory conditions far better if provided with sand, &c, in which to burrow. Under such conditions a small Ensis arcuatus, 4-5 cm. long, lived forty weeks; three small Solecurtusscopula (= c a n d i - dus), 3-3 cm., 3-5 cm., and 2-8 cm. long, eight, fourteen, and fifteen weeks respectively; a small Solecurtus chamasolen (= antiquatus) 2-9 cm. long, twenty-two weeks; and two small Lutraria lutraria, 5 cm. and 3-5 cm. long, thirty- seven and forty-five weeks. Solecurtus, like Ensis, when unprovided with sand supports laboratory conditions badly; Lutraria though somewhat more hardy does not live long. A Mya truncata, on the other hand, has lived in the laboratory for sixty weeks, although unburied, and was apparently healthy when killed. Cultellus pellucidus is especially delicate; frequently a good proportion of those dredged by S.S. Salpa, on arrival at the laboratory, have the mantle between the fourth and pedal apertures ruptured to a varying extent. The ease with which the mantle splits in this position in 0 u 1 - tellus pellucidus has been previously noted by Bloomer (1902, p. 133; 1903, p. 44). The specimen from which Text-fig. 2 was drawn was fixed directly on arrival, and though the mantle appeared intact, deterioration had already set in in some places, as evidenced by the stretching and breaking down of the cells (Text-fig. 2 B). These burrowing bivalves under laboratory conditions, and unprovided with sand in which to hide, soon become ' hockley' (see Orton, 1934, p. 101, for definition of term); the adductor muscles lose tone, and—owing to the elasticity of the ligament— the valves tend to gape, and in Ensis and Cultellus pellucidus the mantle splits along the line of weakness.1 1 In moribund specimens of E n s i s the foot is occasionally found inserted into the infra-branchial chamber, the anterior end pointing backwards. 442 D. ATKINS The shell differences which Graham (1981, p. 726) found to be correlated with variation in extent of the fourth aperture in Ensis siliqua might perhaps be better stated in terms of strength of the shell ligament. In 'shells of lighter build' the ligament is possibly less strong than in those of 'more robust construction' and therefore in the former rupturing does not occur so readily—and the fourth aperture will tend to be distinct. Morse (1913, p. 270) has described in Solenomya velum, a burrowing Protobranch, rupturing of the mantle in the mid- ventral line between the pedal and the single posterior exhalent aperture (there is no fourth aperture in this genus), as the animal becomes enfeebled. He stated that ' There is a median suture in this ventral membrane and in one rupture the suture became dislocated, showing there was a strain upon it. In another case a small rupture appeared on each side of the median suture. That this membrane limits the expansion of the valves is shown by cutting the membrane, when the valves immediately open wider in much the same way as when the adductors are severed in other lamellibranchs.' Prom Drew's figure (1900, fig. 10, p. 264) of a transverse section of the ventral margins of the mantle and cuticle of Solenomya velum it appears that the fusion of the two lobes is complete and not of the cuticular type: sections of Solenomya togata also failed to reveal any trace of a line of fusion, but fixation was not good. The rupturing which Morse described is evidently due to the thinness of the mantle in the ventral region, and the strength and arrangement of the pallial muscles. Origin of the Fourth Aperture. As to the origin of the fourth aperture in the Solenidae, Bloomer (1903, pp. 44-5) held that ' From a morphological point of view, it is reasonable to infer that a portion of the pedal aperture first became specialized by developing a tentacular fringe; then this fringe extended posteriorly, and the pallial walls coalesced, separating the fourth from the pedal aperture, and finally the fourth aperture gradually proceeded farther posteriorly, until it attained a position favourable for the function it originated for', which in Ensis siliqua and Ensis ensis he considered to be that of 'an accessory food-providing organ, and THE CILIARY MECHANISMS OP LAMELLIBRANCHS 443 an exhalent orifice for ejecting water or foreign matter, though the latter function is probably a secondary one'. This view of its origin he based chiefly on the fact that in Ceratosolen (=Pharus) legumen (L.)—which has no fourth aperture— 'the dorsal and ventral surfaces of the pedal aperture carry a tentacular fringe, and extend farther posteriorly' than in Sdlen vagina (= mar gin at us), which also has no fourth aperture; and that in Cultellus pellucidus the fourth aperture is situated more anteriorly (at about the posterior end of the first quarter of the animal) than in Ensis ensis and Ensis siliqua, where it is about the centre of the ventral surface. He held that Solen marginatus is a more primitive form than Ceratosolen legumen; Cerato- solen legumen than Cultellus pellucidus; and Cultellus pellucidus than Ensis siliqua or Ensis ensis. While this view of its origin may be right, it must be observed that the presence of tentacles on the mantle margin is perhaps the rule in Lamellibranchs—in the closely allied family Solecurtidae they are found in Solecurtus scopula on the external surface of the mantle edge throughout its extent (see Graham, 1934, p. 180)—and that their absence round the pedal aperture in the Solenidae might on the contrary be considered as due to specialization of the mantle margins in this region to form the muscular pedal flaps or valves. It seems simpler to imagine that an originally large pedal opening was gradually reduced by fusion of the mantle lobes—as the foot increased in length and came to be protruded from the anterior end of the shell—leaving a small aperture posteriorly, functioning normally as an additional inhalent aperture, and, on sudden closure of the valves, for the ejection of unwanted material. The fourth aperture in Lutraria and Thracia appears to have originated at an earlier period than in the Solenidae (see also Bloomer, 1903, p. 45), judging from the condition of the mantle in the mid-ventral line between this aperture and the pedal (see p. 431); and in Thracia before Lutraria. In Lutraria it has been found that under experimental conditions at least the aperture seems to be generally closed, but on occasions the entrance of particles has been observed, as 444 D. ATKINS indeed must happen through any open aperture leading into the inhalent chamber of any bivalve, if the lateral cilia of the gills are active. Its chief use, however, appears to be, as Bloomer (1903) observed, for the sudden ejection of unwanted material. In Lutraria and the Anatinacea the fourth aperture is situated just below the inhalent siphon (Bloomer, 1903; Pel- seneer, 1890; 1911, pp. 74-5), and according to Kellogg's figures of Mytilimeria nuttallii Conrad and Lyonsia saxicola Baird (1915, figs. 22, 26, 27, 28) the bay in the mantle, in which material brought by the mantle currents collects, is just posterior to it. It is perhaps easier for material to be expelled by way of the fourth aperture than by the inhalent siphon. In the Protobranch Nuculana minuta (see Atkins, 1936, Part I) it was observed that the mantle collections were expelled just ventral to the inhalent siphon, rather than by way of the siphon itself. In Lutraria and Thracia the fourth aperture is closer to the inhalent siphon than apparently in Mytilimeria nuttallii and Lyonsia saxicola.

SUMMARY. A form of fusion involving only the cuticle has been found in certain Lamellibranchs between the outer demibranchs and the mantle or the visceral mass; between the two inner demibranchs; and, in forms possessing a fourth aperture, between the mantle lobes in the mid-ventral line between this and the pedal aperture. The histological structure of the junction is described, and is especially considered in relation to the condition of the fourth aperture in the Solenidae.

KEFERENCES. Atkins, D. (1936).—"Ciliary Mechanisms and Interrelationships of Lamellibranohs, Part I", 'Quart. Journ. Micr. Sci.', vol. lxxix. Bloomer, H. H. (1901).—"Anatomy of the British Species of the Genus Solen, I", 'Journ. Malaeol.', vol. viii. (1902).—"Anatomy of the British Species of the Genus Solen, IV", ibid., vol. ix. (1903).—"Origin and Function of the Fourth Aperture in some Pelecypoda", ibid., vol. x. THE CILIAEY MECHANISMS OF LAMELLIBRANCHS 445

Bloomer, H. H. (1912).—"Anatomy of Species of Cultellus and Azor", 'Proc. Malac. Soc. Lond.', vol. x. Deshayes, G. P. (1857).—"Traite elementaire de Conchyliologie', torn, ii, p. 264 (not seen personally). Douvill6, H. (1907).—"Lamellibranchea cavicoles ou Desmodontes", 'Bull. Soc. Geol. France', 4e ser., torn. vii. Drew, G. A. (1900).—"Locomotion in Solenomya and its Relatives", 'Anat. Anz.', Bd. xvii. (1907).—"Habits and Movements of the Razor-shell Clam, Ensis directus", 'Biol. Bull. Woods Hole', vol. xii, 1906-7. Eschrich, H. (1931).—"Cultellus pellucidus. Eine monographische Dar- stellung", 'Zool. Jahrb.', Bd. liii. Gatenby, J. B. (1928).—'Bolles Lee's Microtomist's Vade-Mecum', 9th ed., London. Ghosh, E. (1920).—"Taxonomic Studies on the soft parts of the Solenidae", 'Rec. Indian Mus.', vol. xix. Graham, A. (1931).—"Morphology, Feeding Mechanisms, and Digestion of Ensis siliqua", 'Trans. Roy. Soc. Edin.', vol. lvi. (1934).—"Structure and Relationships of Lamellibranchs possessing a Cruciform Muscle", 'Proc. Roy. Soc. Edin.', vol. liv, 1933-4. Haas, P. (1934).—"Bivalvia" in H. G. Bronn's 'Klassen und Ordnungen des Tierreichs'. Kellogg, J. L. (1915).—"Ciliary Mechanisms of Lamellibranchs, with descriptions of anatomy", 'Journ. Morph.', vol. xxvi. Morse, E. S. (1913).—"Observations on Living Solenomya (velum and borealis)", 'Biol. Bull. Woods Hole', vol. xxv. Orton, J. H. (1934).—"Bionomical Studies on Cardium edule, with special reference to Mortality in 1933", 'James Johnstone Memorial Volume', Univ. Press, Liverpool. Pelseneer, P. (1890).—"Sur le quatrieme orifice palleal des Pelecypodes", 'C. R. Acad. Sci.', torn. ex. (1891).—"Contribution a 1'etude des Lamellibranches", 'Arch. Biol.', torn. xi. (1911).—'Lamellibranches de l'Expedition du Siboga', Partie Ana- tomique, Monographic liii a, Siboga-Expeditie. Stempell, W. (1898).—"Beitr. z. Kenntniss der Nuculiden", 'Zool. Jahrb.', Suppl. iv. Stenta, M. (1903).—"Stromungen im Mantelraume der Lamellibranchia- ten", 'Arb. Zoolog. Inst. Univ. Wien', torn. xiv.