The morphology and evolution of the ventral gill arch skeleton in batoid fishes (Chondrichthyes: Batoidea) TSUTOMU MIYAKE AND JOHN D. MCEACHRAN Department of Biology, Dalhousie University, Halifax, Nova Scotia, B3H 431, Canada and Department of Wildlqe and Fisheries Sciences, Texas AdYM University, College Station, TX 77843, U.S.A. Received July 1988, reuised manuscript accepted October I990 The ventral gill arch skeleton was examined in some representatives of batoid fishes. The homology of the components was elucidated by comparing similarities and differences among the components of the ventral gill arches in chondrichthyans, and attempts were made to justify the homology by giving causal mechanisms of chondrogenrsis associated with the vcntral gill arch skeleton. The ceratohyal is present in some batoid fishes, and its functional replacement, the pseudohyal, seems incomplete in most groups of batoid fishes, except in stingrays. The medial fusion of the pseudohyal with successive ceratobranchials occurs to varying degrees among stingray groups. The ankylosis between the last two ceratobranchials occurs uniquely in stingrays, and it serves as part of the insertion of the last pair of coracobranchialis muscles. ‘The basihyal is possibly independently lost in electric rays, the stingray genus Urotrygan (except U. dauzesz) and pelagic myliohatoid stingrays. ‘I‘he first hypobranchial is oriented anteriorly or anteromedially, and it varies in shape and size among batoid fishes. It is represented by rami projecting posterolaterally from the basihyal in sawfishes, guitarfishes and skates. It consists of a small piece ofcartilage which extends anteromedially from the medial end of the first ccratobranchial in electric rays. It is a large cartilaginous plate in most of stingrays. It is absent in pelagic myliobatoid stingrays. The remaining hypobranchial cartilages also vary in shape and size among batoid fishes. Torpedo and possibly the Jurassic Be1emnobali.r and Spathobatis possess the generalized or typical chondrichthyan ventral gill arch structure in which the hypobranchials form a X-shaped pattern. In the electric ray Hypnos and narkinidid and narcinidid electric rays, the hypobranchial components are oriented longitudinally along the mid-portion of the ventral gill arches. They form a single cartilaginous plate in the narkinidid electric rays, Narcine and Diplobatis. In guitarfishes and skates, the second hypobranchial is unspecialized, and in skates, it does not have a direct contact with the second ceratobranchial. In both groups, the third and fourth hypobranchials are composed of a small cartilage which forms a passage for the afferent branches of the ventral aorta and serve as part of the insertion of the coracobranchialis muscle. In sawfishes and stingrays, the hypobranchials appear to be included in the medial platc. In sawfishes, the second and third components separately chondrify in adults, but the fourth component appears to be fused with the middle medial plate. In stingrays, a large medial plate appears to include thr second through to the last hypohranchial and most of the basibranchial copulae. The medial plate probahly develops independently in sawfishes and stingrays. Because thc last basibranchial copula appears to be a composite of one to two hypobranchials and at least two basibranchial copulae, the medial plate may be formed by several developmental processes of chondrogenesis. More detailed comparative anatomical and developmental studies are needed to unveil morphogenesis and patternings of the ventral gill arch skeleton in batoid fishes. KEY WORDS:-- Batoids - ehondrichthyans ~ ontogeny - homology 75 0024-4082/91/050075 + 26 $03.00/0 0 1991 The Linnean Society of London 76 T. MIYAKE AND J. D. MCEACHRAN CONTENTS Introduction ...... 76 Material and methods ...... 76 Development of ventral gill arch skeleton ...... 77 Ventral gill arch skeleton in holocephalans and sharks ...... 78 Ventral gill arch skeleton in batoid fishes ...... 81 Discussion ...... 93 Acknowledgements ...... 97 References ...... 98 Abbreviations used in figures ...... 99 Appendix ...... 99 INTRODUCTION Batoid fishes comprise 400 to 500 living species in five subgroups (pristoids, rhinobatoids, rajoids, torpedinoids and myliobatoids) within recent elasmobranchs (Compagno, 1973, 1977; McEachran, 1982). The earliest known batoids are rhinobatoid-like fishes from the Lower Jurassic (Saint-Seine, 1949; Cappetta, 1987; Carroll, 1988). Anatomical studies of the group have been extensive (Gegenbaur, 1865, 1872; Parker, 1879; Garman, 1913; Allis, 1923), and they have provided an important basis for studying phylogenetic interrelationships of batoid fishes (White, 1937). However, recent phylogenetic studies have not produced a consensus of interrelationships of the taxa (Maisey, 1984a, 1986) and there is disagreement as to whether batoid fishes are monophyletic (Compagno, 1973, 1977) or diphyletic (Jarvik, 1977). Lack of consensus among these studies (Compagno, 1973, 1977; Heemstra & Smith, 1980; Maisey, 1984b) appears to be due in part to their lack of comprehensive anatomical comparisons of the major taxa. The gill arch skeleton has been well studied in fishes and has been used to infer phylogenetic interrelationships of osteichthyan fishes (Nelson, 1969; Lauder & Liem, 1983). In chondrichthyans, the ventral gill arch skeleton has been described by Gegenbaur ( 1872), Garman ( 19 13), Holmgren ( 1940, 1941), Hamdy (1956, 1957, 1973) and Hamdy & Khalil (1973). These studies revealed that chondrichthyans are unique in possessing a C-shaped pattern of hypobranchial cartilages and that the pattern of batoid fishes is considerably modified from the generalized condition in chondrichthyans (Nelson, 1969; Miyake, 1988). Thus, specialization of the ventral gill arch skeleton in batoid fishes makes it difficult to homologize the components throughout chondrichthyans (Miyake, 1988). Preliminary results of studies of the ventral gill arch skeleton in batoid fishes are presented here. Attempts were made to trace homologous components of the ventral gill arch skeleton throughout the major taxa of batoid fishes, and causal explanations of developmental processes are given to support our discussions on homology of the ventral gill arch skeleton in batoid fishes. MATERIAL AND METHODS Structures of the ventral gill arch skeleton in chondrichthyans were examined from gross dissections, from cleared and stained specimens and from X-radiographs of specimens. Embryos and juvenile specimens were cleared and double-stained with Alcian Blue (Kodak 14091) and Alizarin (Alizarin sodium EVOLU'I'ION OF BATOIDS 77 sulphonate, Fisher Scientific Company) for cartilage and calcified cartilage, respectively (Dingerkus & Uhler, 1977). All specimens illustrated in the figures are listed in the Appendix. Homologies of the components of the ventral gill arch skeleton in batoid fishes were elucidated by comparing the components of the ventral gill arch skeleton among chondrichthyans. Causal explanations of developmental processes are offered to support our perceptions of homology of the ventral gill arch skeleton in chondrichthyans based on our conceptual framework of homology (Miyake el al., in press). DEVELOPMENT OF VENTRAL GILL ARCH SKELETON The pattern of the ventral gill arch skeleton in recent elasmobranchs can be elucidated from the blastematic condensation and subsequent chondrification of the skeleton in the spiny dogfish Squalus acanthias embryos (Holmgren, 1940; El-Toubi, 1952; Jollie, 1971) (Fig. 1) and in some embryos of batoid fishes (Holmgren, 1940). The first horizontal blastema forms medially between the blastemas of the presumptive ceratohyal or pseudohyal. Because this portion of the skeleton articulates with the ceratohyal in adult holocephalans and sharks (Fig. 2) and with either the ceratohyal or pseudohyal in adult batoid fishes, it is considered to be the basihyal component. However, it either comprises the basihyal and the first hypobranchial (Miyake, 1988) or merely represents the first basibranchial copula (Nelson, 1969). According to Jollie (1971) the lateral longitudinal portion of the blastema is a component of the presumptive first hypobranchial. This blastema is continuous with the diagonally oriented first ceratobranchial blastema. In most sharks, the first pair of hypobranchials is either the distolateral ramus of the basihyal, as seen in Squalus acanthias, or is not chondrified. In some sharks the first hypobranchial is represented by a small piece of cartilage between the basihyal and the first ceratobranchial. In batoid fishes, the first pair of hypobranchials is either the distolateral ramus of the basihyal or a separate cartilage. In the 50 mm embryo of Raja (Raja)clavata, the basihyal and the first pair of hypobranchials are present as separate components A B C D Figure 1. Early development of the ventral gill arch skeleton in Squalus aranlhias Linnacus. A, 35-37 mm TL; B, 48 mm TL; C, 67 mm TL; D, 102 mm TL. A from Jollie (1971: fig. 8); B from Holmgren (1940: fig. 68); C and D from El-Toubi (1952: figs 3, 5). The star indicates where the blastema of the first hypobranchial develops. Arrow heads indicate a topographic relationship of hypobranchials with their corresponding ceratobranchials. 7a T. MIYAKE AND J. D. MCEACHRAN (Holmgren, 1940: fig. 138). At later stages, however, the two components fuse to form a Y-shaped cartilage. Posterior to the first ceratobranchial blastema in Squalus acanthius embryos, there are three or four pairs of blastemas which maintain one-to-one relationships with their corresponding ceratobranchial components. They are here identified as hypobranchial blastemas. The topographic relationship of the second to at least the fourth ceratobranchial blastemas with the corresponding hypobranchial ones is maintained during development (Fig. 1B-D) and leads to the Z-shaped pattern of the hypobranchials in adults. The initial chondrification of the hypobranchials appears to occur at the rami of the blastema and proceeds laterally toward the ceratobranchial blastemas and medially toward the midline (Fig. 1A). These lateral and medial directions of chondrification create a pair of hypobranchials which is incompletely or completely segmented as the lateral and medial portions of the cartilages in adults of some batoid fishes, as discussed below. There is generally a space between the basihyal and the second hypobranchial in both blastematic and adult stages in recent elasmobranchs (Nelson, 1969) (Fig. 1A) and the Palaeozoic Xenacanthus (Maisey, 1986). This space is thought to represent the developmental field of the second basibranchial which appears to be almost completely suppressed. Occasionally a small piece of cartilage representing a remnant of the second basibranchial is seen in Heterodontus japonicus (Gegenbaur, 1872: table 19, fig. 3). The third to the fifth and/or sixth basibranchials lie between successive paired arch components (Nelson, 1969). The blastemas of these basibranchials are continuous with the hypobranchial blastemas. As the chondrification proceeds, the hypobranchial components tend to be separated from the median components which have their own centre of chondrification (Holmgren, 1940; El-Toubi, 1952). The median components thereafter become the basibranchial copulae. The number of the copulae varies among chondrichthyans. VENTRAL GILL ARCH SKELETON IN HOLOCEPHALANS AND SHARKS In holocephalans, the ceratohyal articulates tightly with the small basihyal (Fig. 2B). There are five separate pairs of ceratobrnnchials (Fig. 2A, B). Each of the first three pairs of ceratobranchials articulates with its corresponding hypobranchial. The last two pairs articulate with a single cartilage which is oriented anterolaterally (Fig. 2A, B). The different orientations of the hypobranchials may either represent the pattern of the chondrification which leads to the separation of the lateral and medial portions of the hypobranchials or reflect ontogenetic fusion of the last two hypobranchials, as seen in electric rays. There are at least four basibranchial copulae (Fig. 2A, B). In sharks, a large ceratohyal articulates with a relatively large basihyal (Fig. 2C-I). In Sguatina, there are two cartilages of unknown origin associated with the ceratohyal and basihyal. One of these cartilages lies along the anteromedial margin of the ceratohyal and articulates with the basihyal (Fig. 3). A similar cartilage is found in Datatias licha (Gegenbaur, 1872: table 19, fig. 2) (Fig. 2G). Topographic relationship of this cartilage with the ceratohyal suggests that it may represent a developmentally equivalent form to the hypohyal. The other cartilage lies dorsal to the ceratohyal, and spans the medial end of the first A B C D C c bl c bl cb E F G H I bh C. bs Figure 2. Ventral view of' the ventral gill arch skeleton in holoccphalans and sharks. A, Hydro1ngu.r rolliei (Lay & Bennett) (from Rosrn F/ nl., 1981: fig. 47); B. C,'dl~dp~ht~~sp. (from Edgeworth, 1935: fig. 80); C, ClzlnnyduselnchuJ nnguineuJ Garman (from Allis, 1923: fig. 36); D, Heptrnnchiasperlo [Bonnaterre) (Trom Gegenbaur, 1872: tahlc 18. 6s. 1 ); K, Hderodontus jnponicus (Maclay & Maclray) [from Grgenbaur. 1872: table 19, fig. 3); F, LImop/erus spinnx (Linnaeus) (from Grgenbaur, 1872: tahlr 18. fig. 6): G. I)nlo/in.\ lirhn (Bonnaterrr) (from Cegenbaur, 1872: table 19, fig. 2); H, I~zstzus brn~iliensis(Quay & Gaimard) (lrom Shirai, 1985: lig. 4); I. Prkliophoru\ jnponmJ Gunthrr [lkom Gdrrnin. 1913: pl. 64, fig. 1). 'l'he arrow indicates a blind spare between the first horizontal har and the second gill arrh components. WU 80 7’. MIYAKE ANDJ. D. MCEACHRAN A Figure 3. Ventral view of the ventral gill arch skeleton in Squatzna dumeril LeSueur (TCWC 4214.2). ceratobranchial and the lateral end of the basihyal. The position of this cartilage suggests that it may be a component of the first hypobranchial. In Pristiophorus, a pair of cartilages is located lateral to the basihyal (Fig. 21). It represents the first hypobranchial because a similar cartilage, identified as the first hypobranchial, is found in the 50 mm embryo of the skate Raja (Raja) clavata (Holmgren, 1940: fig. 138). Most sharks have five pairs of ceratobranchials (Fig. 2E-I). However, Hexanchus and Chlamydoselachus have six pairs (Fig. 2C); Heptranchias and Notorhynchus have seven pairs (Fig. 2D). When the hypobranchials are present, they articulate with their corresponding ceratobranchials. In Isistius, at least third, fourth and fifth ceratobrachials articulate with a single hypobranchial (Shirai, 1985) (Fig. 2H). A small pair of cartilages, which may represent part of the first hypobranchials, is only occasionally present, i.e. in some species of Heptranchias (Gegenbaur, 1872) (Fig. ZD), Scapanorhynchus (Garman, 1913: pl. 51, fig. 3), Heterodontus (Gegenbaur, 1872) (Fig. ZE), Scyliorhinus (Gegenbaur, 1872: table 18, fig. 4; Holmgren, 1940: fig. 117; El-Toubi, 1952: fig. 7), Ginglymostoma, Parascyllium, Brachaelurus, Orectolobus and Chiloscylliurn (Dingerkus, 1986: fig. 4; Dingerkus & DeFino, 1983: figs 51-55), Sphyrna (Hamdy & Khalil, 1973: fig. 3) and Dalatias (Gegenbaur, 1872) (Fig. 2G). The first hypobranchial may also be represented by the rami projecting distolaterally from the basihyal, as seen in Squalas acanthias. In most sharks, the second to the last hypobranchials articulate with their corresponding ceratobranchials. However, in Isistius, the second to the fifth hypobranchial components appear to be fused into a single anterolaterally oriented cartilage (Shirai, 1985) (Fig. 2H). In Squatina, the fifth hypobranchial EVOLUTION OF BA?’OIDS 81 may consist of the rami projecting posterolaterally from the basibranchial copula (Fig. 3). The number of basibranchial copulae varies in sharks, and depends on the pattern of chondrification in the hypobranchial and basibranchial blastemas. As stated above, the development of the second basibranchial is probably suppressed but it is present in Heterodontus (Fig. 2E). In Squatina, the third to sixth basibranchials are represented by a single plate. In some sharks, e.g. some Chiloschyllium and Hemischyllium (Dingerkus & DeFino, 1983: figs 5 1-56), Scyliorhinus (Gegenbaur, 1872: table 18, fig. 4; Holmgren, 1940: fig. 117; El-Toubi, 1952: fig. 7), Galeorhinus (Gegenbaur, 1872: table 19, fig. 4), Sphyrna (Hamdy & Khalil, 1973: fig. 3), Isistius (Shirai, 1985) (Fig. 2H) and Pristiophorus (Fig. 21), there is also only a single basibranchial copula representing the fifth and/or sixth basibranchial components. VENTRAL GILL ARCH SKELETON IN BATOID FISHES Batoid fishes are distinguished from holocephalans and sharks by the articulation of the last pair of ceratobranchials with the anteromedial aspect of the scapulocoracoid (Heemstra & Smith, 1980). The presence of a pseudohyal has been claimed to be a derived character of batoid fishes (Holmgren, 1941; Compagno, 1973; 1977) but this cartilage also occurs in sharks (de Beer, 1932). It seems likely that the ventral gill arch skeleton of batoid fishes consisted primitively of a basihyal, a pair of certohyals and/or possibly a pair of pseudohyals, five pairs of ceratobranchials, four or five pairs of hypobranchials and one or two basibranchial copulae (Saint-Seine, 1949; Nelson, 1969). However, uncertainties remain in identifying specific components as illustrated in the ventral gill arch skeleton of the Jurassic guitarfish-like Belemnobatis and Spathobatis. Because the two genera are proposed to be allied to guitarfishes (Cappetta, 1987) or to sawfishes (Maisey, 1984a), we contend that the overall morphological pattern of the ventral gill arches in both fossil genera resembles that of recent batoid fishes, and thus that clarification is needed regarding the identity of the components. We base the identity of each component of the ventral gill arches in both fossil genera on three assumptions: (1) the first ceratobranchial always has a physical contact with a centrally located horizontal bar because of early blastematic relationships between the two compenents; (2) there is one-to-one relationship of the second to the last ceratobranchial with each corresponding hypobranchial; (3) the horizontal bar, therefore, is a composite of basihyal and the first hypobranchial cartilages. In both fossil genera, the centrally located large horizontal bar is not the basihyal cartilage (Fig. 4A, D: bh) as Saint-Seine (1949) proposed, but a composite of basihyal and the first hypobranchial cartilages (Fig. 4B, C, E, F: bh+ hpl). Four pairs of hypobranchial cartilages would thus be considered the second to fifth paired hypobranchials (Fig. 4B, C, E, F: hpZ-hp5), not the first to fourth (Fig. 4A, D: hp 1-hp4). However, as shown below, because batoid fishes possess a pair of pseudohyal cartilages with or without a small pair of putative ceratohyal cartilages, there are at least two possible alternative identities of the pairs of laterally located cartilages in both Belemnobatis and Spalhobatis. In the first alternative, the second to fifth paired cartilages (Fig. 4B, E: cb2-cb5) would have a physical contact 82 T. MIYAKE AND J. D. MCEACHRAN D E F Y co Figure 4. Three possible interpretations of the identity of the ventral gill arch skeletal components in the Jurassic Belemnobatzs (A-C) and Spathobatis (D-F).A, D,A proposal of Saint-Seine (1949). The first pair of medially located cartilages was considered the first hypobranchial, and four pairs of hypobranchials are assumed to be in close contact with the second through the fifth pairs of laterally located cartilages. B, E, The first alternative proposal in this study, assuming that the second to fifth laterally located pairs of cartilages (cb2-cb5) have a physical contact with each corresponding hypobranchial. C, F, The second alternative proposal in this study, assuming that the last four pairs of laterally located cartilages (cb2-cb5) have a physical contact with the second to the fifth hypobranchials See text for details with each corresponding hypobranchial. In this case, the last small cartilage projecting from the coracoid bar (Fig. 4B, E: cb6?) may be considered the sixth ceratobranchial in both genera. In Belemnobatis, the first pair of laterally located cartilages would be the first ceratobranchial (Fig. 4B: cbl),and the small paired cartilages between the horizontal bar and the first pair of ceratobranchials may be the pseudohyal (Fig. 4B: pseuhy?). The first pair of laterally located cartilages in Spathabatis, on the other hand, are either the first ceratobranchial or pseudohyal (Fig. 4E: cbl or pseuhy?). The second alternative is to assume that the last four pairs of laterally located cartilages have close physical contacts with the second to the fifth hypobranchials in both fossil genera (Fig. C, F: cb2-cb5). In Belemnobatis, a small paired cartilage lateral to the horizontal bar (a composite of the basihyal and the first hypobranchial) would be the ceratohyal cartilage (Fig. 4C: cert), whereas the cartilage is missing in Spathobatis. The first two pairs of laterally located cartilages would thus be identified as the pseudohyal and the first ceratobranchials in both genera, respectively (Fig. 4C, F: pseuhy, cbl). Saint-Seine’s proposal and our first alternative both introduce unusual patterns of the skeletal components of the ventral gill arches in batoid fishes: the medial position of the first hypobranchial and uncertain identity of the first pair of laterally located cartilages, either ceratohyal or pseudohyal (Saint-Seine’s proposal) (Fig. 4A, D) and six pairs of ceratobranchials (our proposal) (Fig. 4B, E). Our second alternative, however, is identical to the pattern of the ventral gill arch skeleton in recent batoid fishes as well as other elasmobranch fishes EVOLUI'ION OF BATOIDS 83 (Fig. 46, F), and it seems to be the most parsimonious interpretation. It would thus be assumed that in both fossil genera, a physical contact takes place between the second to fifth hypobranchials and the corresponding ceratobranchials, as illustrated in Fig. 4C and F, respectively. Sawfishes possess a pseudohyal but lack a ceratohyal. The pseudohyal articulates with the ventral aspect of the first ceratobranchial, and its anteromedial margin is in contact with the ramus of the basihyal (the first hypobranchial) (Fig. 5A). There are five separate pairs of ceratobranchials. The first ceratobranchial articulates with a concavity in the ramus of the basihyal. The second and third ceratobranchials articulate separately with two small cartilages, the second and third hypobranchial components, lateral to the anterior medial plate (Fig. 5A). The fourth ceratobranchial articulates with the distolateral aspect of the middle medial plate whereas the fifth ceratobranchial tightly ankyloses with a concavity in the last basibranchial copula (Fig. 5A). The basihyal consists of a horizontal cartilaginous bar. On the assumption that the second to fifth ceratobranchials articulate with the corresponding hypobranchial, the rami of the basihyal are considered the first hypobranchials. The three tightly articulated medial plates located posterior to the basihyal are the basibranchial components. The anterior plate is not strongly calcified, and its anterior margin is fibrous. Because it is located between the second arch components, the plate is considered at least the third basibranchial copula. The middle plate is rectangular and possesses a bridge through which the fourth, fifth and sixth afferent branches of the ventral aorta pass (Fig. 5B). It may consist of at least the fourth basibranchial copula and the fourth hypobranchial component. The posterior plate, the last basibranchial copula, is spade-shaped and may represent the fifth and/or sixth copulae of basibranchials and the fifth hypobranchial component. The guitarfish, Rhinobatos halaui (Hamdy, 1956, 1957), R. granulatus (Hamdy & Khalil, 1973) and R. productus (Miyake, 1988) (Fig. 5C) and Rhynchobalus 4iddensis (Fig. 5D) possess both a ceratohyal and pseudohyal. On the other hand, Platyrhina siensis (Garman, 1 9 13) and Zupteryx brevirostris (Miyake, 1988) possess only the pseudohyal. When present, the ceratohyal is much smaller than the pseudohyal (Hamdy, 1957), and excludes the more posteriorly located pseudohyal from contact with the ramus of the basihyal (the first hypobranchial) (Fig. 5D). When the ceratohyal is absent, the pseudohyal is in close contact with the ramus of the basihyal (Fig. 5C). There are five separate pairs of ceratobranchials in guitarfishes. The first ceratobranchial articulates with the distal ramus of the basihyal (the first hypobranchial). The second through the fourth ceratobranchials articulate with three corresponding hypobranchial components (Fig. 5C, D). The fifth ceratobranchial always articulates with the lateral side of the basibranchial copula. The basihyal of guitarfishes is inverted U-shaped with the distolateral rarni (Fig. 5C, D). The rarni may represent the first hypobranchials. There are three pairs of hypobranchials anterior to the basibranchial copula. In most species of Rhinobatos examined, the second hypobranchial is arched and extends from the anterior margin of the copula of basibranchials to the medial aspect of the second ceratobranchial (Fig. 5C). In Rhynchobatus, the second hypobranchial is incompletely or completely divided into the lateral and medial portions (El-Toubi & Hamdy, 1959; Miyake, 1988) (Fig. 5D). The lateral portion is 84 T. MIYAKE AND J. D. MCEACHRAN A ?hpl+bh B A epr-hvm C D l+bh E Figure 5. Ventral view of the ventral gill arch skeleton in three batoid taxa. A, B, Prihperlinatus Latham (MCZ 36960); C, Rhinobatos produdus Girard (TCWC 6183.1); D, R/ynchohatus djiddensis (Forskal) (MCZ 806); E, Raja (Leucuraja) garmani Whitely (TCWC 1888.1). B shows thc pattern or the afferent branches of the ventral aorta in Pristis pcclinatus. The arrow indicates the gap betwecn the second hypobranchial and the second ceratobranchial. EVOLUTION OF BATOIDS 85 expanded anterolaterally and overlaps the basihyal medially. The third and fourth hypobranchial components (Fig. 5C, D) are small cartilages lying lateral to the medial portion of the second hypobranchial. In Rhinobatos percellens the three hypobranchials on each side are fused together (Garman, 19 13). Both the ceratohyal and pseudohyal are found in some skates, i.e. Pseudoraja jischeri and Sympterygia brevircaudata (McEachran & Miyake, 1990) and Raja (Leudoraja) circularis (Khalil & Hassan, 1973), but only the pseudohyal is present in others, i.e. Raja (Leudoraja) garmani (Fig. 5E), R. (L.) erinacea (Garman, 1913) and Sympteygia acuta (Garman, 1913). The large pseudohyal lies lateral to the lateral ramus of the basihyal (the first hypobranchial) (Fig. 5E). There are five separate pairs of ceratobranchials in skates. The first ceratobranchial articulates with the ramus of the basihyal (the first hypobranchial). The second ceratobranchial possesses a large anteromedial ramus which overlies a portion of the first ceratobranchial. The third and fourth hypobranchials articulate with the small third and fourth hypobranchial cartilages. The fifth ceratobranchial articulates with the anterolateral margin of the basibranchial copula. The basihyal constitutes a horizontal or an arched bar in skates which either has or lacks anterolateral projections (McEachran & Miyake, 1990). The rami projecting posterolaterally or posteriorly from the bar possibly represent the first hypobranchial. The second hypobranchial consists of a bifurcated pair of cartilages which do not have a direct contact with the second ceratobranchial. The gap between the second hypobranchial and the second ceratobranchial suggests that the lateral portion of the second hypobranchial is missing. During the early development of Raja (Leucoraja) circularis (Khalil & Hassan, 1973) and R. (L.)erinacea (McEachran & Miyake, unpublished data), a small cartilage lies medial to the second ceratobranchial and is connected with the tip of the second hypobranchial by means of connective tissue. This small cartilage seems to represent a remnant of the lateral portion of the second hypobranchial. The second hypobranchial is always fused with the basibranchial copula (McEachran, unpublished data). The third and fourth hypobranchials are small plate-like cartilages and separately chondrify from the basibranchial copula, as in guitarfishes. However, the two cartilages occasionally fuse to form a large plate-like cartilage in Raja (Raja) clavata and Symptevgia brevicaudafa (McEachran, unpublished data; McEachran & Miyake, 1990). The basibranchial copula is triangular- shaped and possesses a distal projection. There are several types of patterns of the gill arch skeleton in electric rays, although all electric rays examined lack a basihyal. Torpedo and Hypnos lack a ceratohyal but possess a pseudohyal which is in close contact with the distoventral aspect of the first hypobranchial (Fig. 6A, B). There are five separate pairs of ceratobranchials. In Torpedo, the first ceratobranchial is fused with the first hypobranchial (Fig. 6A). The second, third and fourth ceratobranchials articulate with their corresponding hypobranchials (Fig. 6A). The fifth ceratobranchial tightly articulates with anterolateral margin of the basibranchial copula. In Hypnos, the first ceratobranchial is a separate cartilage that medially articulates with the second of three longitudinally oriented cartilages (Fig. 6B). The second ceratobranchial articulates with the second of three longitudinally oriented cartilages. The third ceratobranchial is in close contact with two or three small cartilages lying between it and the third of three longitudinally A C D cor-h Pa P W+ E F G H P 5 Figure 6. Ventral view of the ventral gill arch skeleton in electric rays. A, Torpedo ml~/brnzncnAyrrs (MCZ 43); B, Hvpiios monopkg@un (Shaw & Noddcr) (MCZ 38602): C, .Vnrke jnponicn (Schlegcl) (MCZ 1339); D, Typhlonarke qsoni (Hamilton) (FKSKU 46477); E, Nnrcine brnsiliuniiJ (Oikrs) (TCWC uncat., 235 mm 'I'L); F, Disco/Lvgp hchridii (Heckel) (FRSKU 105043); G, Benlhobnlis mnrcidn Bran & Weed ('I'CWC 1903.1); H, Diplohnlih piclus Palrnrr ('I'CWC 1900.1). EVOLUTION OF BATOIDS 87 oriented cartilages (Fig. 6B). The fourth ceratobranchial is attached to the anterolateral margin of the basibranchial copula by means of connective tissue. The fifth ceratobranchial ankyloses with a concavity along the lateral margin of the basibranchial copula. The first hypobranchial in Torpedo and Hypnos is arched medially from the anterior margin of the first ceratobranchial. In Torpedo the second through the fourth hypobranchials form U-shaped arches which articulate with the corresponding ceratobranchials. The second and third hypobranchials are segmented, and the gaps between two successive segments are filled with uncalcified cartilage. In Hypnos the hypobranchials are represented by a series of three cartilages located on either side of the midline anterior to the basibranchial copula (Fig. 6B). Two larger cartilages are tentatively regarded as the second and third hypobranchial components, but the identity of the small cartilage is not clear. The pattern of orientation of the three hypobranchials is totally different from that of Torpedo and resembles those of narkinidid and narcinidid electric rays, as described below. The basibranchial copula in Torpedo and Hypnos is large and occupies about one-half of the space in the ventral gill arches. Narkinidid electric rays, flarke and Typhlonarke, possess both a ceratohyal and pseudohyal (Fig. 6C, D). The ceratohyal is larger than the pseudohyal and lies lateral to the first hypobranchial and anterior to the pseudohyal. The ceratohyal is connected to the hyomandibular by ligamentous tissue (Compagno, 1977). There are five separate pairs of ceratobranchials in Narke and Typhlonarke. The first, second and third ceratobranchials articulate or are in close contact with a large longitudinally oriented cartilage. The fourth and fifth ceratobranchials both articulate with the posterolateral aspect of the basibranchial copula (Fig. 6C, D). In Narke and Typhlonarke, the hypobranchial cartilages are represented by a series of two cartilages located on either side of the midline anterior to the basibranchial copula. The smaller cartilage lying posterior to the ceratohyal is regarded as the first hypobranchial. The larger cartilage is provisionally considered to represent the second and third hypobranchial components. The basibranchial copula is a T-shaped cartilage (Fig. 6C, D). Both the ceratohyal and pseudohyal are found in the narcinidid electric rays Benthobatis and Diplobatis (Fig. 6G, H), but the ceratohyal is lacking in Narcine and Discopyge (Fig. 6E, F). In the former genera the ceratohyal is in close contact with the lateral aspect of the first hypobranchial, but only in Discopyge does the pseudohyal articulate with the first hypobranchial (Fig. 6F). In the three other genera the pseudohyal is not in contact with other cartilages. Narcinidid electric rays possess five separate pairs of ceratobranchials. The first ceratobranchial articulates with the small first hypobranchial. In Nurcine and D$lobatis, the second, third and/or fourth ceratobranchials articulate with a longitudinally oriented cartilaginous plate (Fig. 6E, H) whereas in Discopyge and Benthobatis, the third and fourth ceratobranchials each articulate with a separate cartilage (Fig. 6F, G). The fifth ceratobranchial always articulates with the posterolateral aspect of the basibranchial copula. The longitudinal components located on either side of the midline of the ventral gill arches in Hypnos and narkinidid and narcinidid electric rays are unique within batoid fishes and therefore difficult to identify unequivocally. A piece of evidence for considering them hypobranchial components in the present study comes from the early 88 T. MIYAKE AND J. D. MCEACHRAN A cert 5 Figure 7. Early development of hybranchial cartilages in Narcine bmsiliensi~(Olkrsj ('I'CWC uncat. j. A, 34 mm TL; B, 36 rnm TL. Arrows connect the hypobranchials with their corresponding ceratobranchials. development of Narcine brasiliensis (Fig. 7). At the 34 mm embryonic stage of Narcine bransiliensis, there are four pairs of separately chondrified components on either side of the midline of the ventral gill arches. The distolateral end of each component appears to have a relationship with one of the ceratobranchials (Fig. 7A). This relationship suggests that these four pairs of cartilages represent the hypobranchials. At the 36 mm embryonic stage, the posterior three components fuse to form a longitudinally oriented cartilaginous plate (Fig. 7B), as seen in adult Narcine brasiliensis (Fig. 6E). Occasionally, only two of these three components are fused in adults. Therefore, the large cartilaginous plate in Narcine and Diplobatis represents the second, third and/or fourth hypobranchial components. The pair of plate-like cartilages located medial to the second and third hypobranchials in Benthobatis are of unknown origin (Fig. 6G). Stingrays lack a ceratohyal but possess a pseudohyal which articulates with the distal end of the first hypobranchial. There are five pairs of ceratobranchial in most stingrays, but six pairs in Hexatrygon (Fig. 8F). The ceratobranchials articulate with small rami along the lateral margin of the medial plate (Fig. 8). In most stingrays, the pseudohyal is fused with the first ceratobranchial (Fig. 8). However, Davatis kuhlii (El-Toubi & Hamdy, 1959), Taeniuraforskaii (Hamdy & Khalil, 1973) and Hexatrygon beckulii (Heemstra & Smith, 1980) lack this fusion. The pseudohyal and the first two ceratobranchials are fused in Dasyatis americana (Fig. 8H) and Paratrygon aieraba (Rosa, 1985). The pseudohyal and the first three ceratobranchials are fused in some of potamotrygonids (Rosa, 1985) and Aetobatus narinari (Garman, 19 13; Hamdy & Khalil, 1973). The pseudohyal and the first four ceratobranchials are fused in Potamotrygon constetlata (Fig. 8G), Gymnura micrura (Fig. 8K), G. altauela (Hamdy, 1973) and Aetomyleus ariluus (Garman, 1913). The pseudohyal and all the ceratobranchials are fused in Mobula hypostoma (Fig. 8M). Rosa et al. (1987) regarded the medial fusion of the pseudohyal and two successive ceratobranchials as one of the synapomorphies of Potamotrygon and Plesiotrygon. However, this fusion occurs more universally within stingray taxa, EVOLU'IION OF BATOIDS 89 D E Y F H I L M Figure 8. Ventral view of the ventral gill arch skeleton in stingrays. A, Lhtrypon munda Gill iUSNM 220612); B, Urotygon chilensis (Gunther) (LACM 7013); C, Urolophus halleri Cooper (SIO uncat.); U, Urolophus aurantiacus Miiller & Henle (TCWC uncac.); E, Urotygon dauiesi Wallace (BPBM 24578); F, Hexatygon beckulii Heemstra & Smith (from Hwmstra & Smith, 1980: fig. 10); G, Potnmotvgon constellata (Vaillant) (from Garman, 1913: pl. 70, fig. 2); H, Dasyatis nmerirana Hildebrand & Schroeder (TCWC 5820.1); I, Himanturn irnpricato (Block & Schneider) (MCZ 59269);J, Taeniuro lymma (Forskal) (TCWC 5276.1); K, (iymnura rnzcrura (Bloch & Schneider) (TCWC uncat.); L, Myt'iobatis peruuianus Garman (from Garman, 1913: pl. 73, fig. 2); M. Mobuta hypostorno (Bancroft) [from Garman, 1913: pl. 75, fig. 2).The arrow and circle indicate the site of ankylosis of the last two ceratobranchials. 90 T. MIYAKE AND J. D. MCEACHRAN A B C D E G H I J K EVO1,UTION OF BA'I'OIDS 91 Figure 9. Ventral view of the skeleton of ventral gill arches in Urolophidae. A, Crrolrygon dauiesi Wallace (BPBM 24578); B, Urotygon microphthalmurn Delsman (USNM 222692); C, Urolrygon uenezuefae Schultz (TBT 76-34); D, Urotygon rnunda Gill (USNM 220612); E, Urohygon nann Miyake & McEachran (SIO 65-167); E', Urolyson reticulala Miyake & McEachran (USNM 22264); C, Urotvgon sirnulalrix Miyake & McEachran (GCRL 13064); H, Urolygon chilensis Giinther (LACM 7013); I, Urotvgon roger.ri (Jordan & Starks) (LACM 50-57); J, Urolrygon aspidura (Jordan & Gilbert) (CAS 51834); K, Urolophusjamaicensis (Cuvier) (TCWC 0815.1); L, Urolophus halferi Cooper (SIO uncat.); M, Urolophus concentricus (Osburn & Nichols) (LACM 31771); N, Urolophus paucimaculalus Dixon (FSFL ED189); 0, Urolophus crucialus Laceprde (TCWC uncat.); P, Urolophus buccufentus Macleay (FSFL EC361); Q, Urofophus auranliacus Muller & Henle (TCWC uncat.). The arrow indicates the ankylosis of the fourth and fifth ceratobranchials. and varies among the genera of stingrays (Miyake, 1988). The last two ceratobranchials of stingrays are strongly ankylosed and form part of the insertion of the coracobranchialis muscle (Miyake, 1988) (Fig. 8). However, Heemstra & Smith (1980) did not state whether or not this ankylosis is present in the six-gilled stingray Hexatvgon. The basihyal is variably developed in stingrays. It is absent in the genus Urotrygon (Miyake, 1988) (Fig. 8A, B) except in U. dauiesi (Fig. 8E). In the pelagic myliobatoid stingrays, both the basihyal and the first hypobranchial are absent (Fig. 8L, M). In the remaining groups of stingrays both the basihyal and the first hypobranchial exist as separate components, or comprise a single component, i.e. some individuals of Urolophus jamaicensis (Miyake, 1988). In the former case, the basihyal is occasionally segmented into small cartilages (Fig. 8C, G-J). In stingrays, the mid-ventral portion of the ventral gill arches consists of a 92 T. MIYAKE AND J. D. MCEACHRAN single medial plate (Figs 8, 9). The plate is composed of the anterior and posterior portions. The anterior portion of the medial plate possesses a small to large medial projection on the anterior margin in most stingrays (Figs 8, 9) except in Urotrygon daviesi (Figs 8E, 9A) and pelagic myliobatoid stingrays (Fig. 8L, M). The anterior portion of the medial plate possesses a bridge accommodating the passage of the ventral aorta and its afferent branches in some stingrays (Figs 8-10), i.e. some species of Urotrygon and Urolophus, some species of freshwater potamotrygonids (Rosa, 1985), Hexatrygon (Heemstra & Smith, 1980) (Figs 8F, 9) and Gymnura (Figs 8K, 9). Neither dasyatidid nor pelagic myliobatoid stingrays possess a bridge (Miyake, 1988) (Fig. 8). In Hexatrygon (Figs 8F, 9A), the bridge resembles that of Urotrygon daviesi (Figs 8E, 9A), Urolophus bucculenlus (Fig. 9P) and Gymnura (Fig. 8K) in having no cartilaginous roof medially. Figure 9 shows a variety of forms of the bridge on the medial plate within the stingray family Urolophidae, and Fig. 10 shows some examples of the passage of the ventral aorta and its afferent branches through the bridge. Within the genus Urotrygon, there are four types of bridge: (1) bridge covering the anterior to posterior end in the anterior portion of the medial plate without a medial roof, as in Urotrygon daviesi (Fig. 9A) and Urolophus bucculentus (Fig. 1OP); (2) bridge covering the anterior half of the medial plate where at least the fourth branch of the ventral aorta passes through (Fig. 10B), as in some species of Urotygon (Fig. 9D, E, I, J); (3) small bridge consisting of a cartilaginous passage for the anterior portion of the ventral aorta leading to the second and third branches (Fig. lOC), as in several species of Urotrygon (Fig. 9B, C, G, H) and amphi- American Urolophus (Fig. 9K-M); (4)bridge covering the entire ventral surface on the anterior portion of the medial plate where the fourth to sixth branches of the ventral aorta pass through (Fig. lOD), as in Australian Urolophus paucimaculatus (Fig. 9N) and Urolophus cruciatus (Fig. 90) and Indo-Pacific Urolophus aurantiacus (Fig. 9Q). The posterior portion of the medial plate, which probably represents the basibranchial copula, is ventrally concave and tapers posteriorly. It forms the dorsal wall of the pericardial cavity. Because of the formation of the medial plate A B C D Figure 10. Topographic relationships of afferent branches of ventral aorta with four types of medial plate of ventral gill arches in Urolophidae. A, Urotiygon daviesi Wallace (BPBM 24578); B, Urofiygorz munda Gill (USNM 220612); C, Urolophus jamaicensis (Cuvier) (TCWC 0815.1); D, Urolophus paucimaculatus Dixon (FSFL ED 189). EVOLUTION OF BATOIDS 93 in stingrays, Nishida (2nd International Conference on Indo-Pacific Fishes, Tokyo, Japan, 1985) proposed that there are no hypobranchials in stingrays. However, in UrotTgon daviesi, there are three pairs of the rami projecting posterolaterally from the medial plate which articulate with the second, third and fourth ceratobranchials, respectively (Figs 8E, 9A). These rami suggest that components of the hypobranchials are either ontogenetically fused with the medial plate or chondrify as part of the medial plate and project laterally toward ceratobranchials. In other stingrays, the plate possesses small projections along the side of the medial plate which are intimately associated with the corresponding ceratobranchials and are thought to represent hypobranchials (Miyake, 1988). DISCUSSION The ventral gill arch skeleton of chondrichthyans is unique in having a large space between the basihyal and the second hypobranchial and developing C-shaped hypobranchial cartilages. However, because of a variety of forms of the hypobranchials in batoid fishes, as shown above, it is not clear whether the pattern of blastemas, as seen in Squalus acanthias, actually represents those in batoid fishes. Nelson ( 1969) discussed segmentations of the basibranchials in both chondrichthyans and osteichthyans, arguing that the segmentation of the basibranchials is ontogenetically secondary and that it may have arisen independently in each group. His discussion implies that the ancestral pattern of the skeleton of at least the basibranchials in both the groups may have been continuous without segmentations. However, it still remains uncertain as to what developmental processes gave rise to the segmented basibranchial patterns: changes in pattern of blastemas, changes in pattern of chondrification, possible remodelling, or all of these. As seen in the development of hypobranchials in Narcine brasiliensis (Fig. 7), each hypobranchial chondrifies posterolaterally in relation to its corresponding ceratobranchial, but at later stages the medial part of the second and third hypobranchial grows more posteriorly, rather than medially, and fuses to form a composite of the second to fourth hypobranchials. Although this growth pattern is in part attributed to the change in pattern of chondrification, whether the underlying blastemas also differ from those of Squalus acanthias remains to be elucidated. Although our study shows that the segmental nature of basibranchials exists in some batoid fishes (Nelson, 1969), it is not clear how the segmental development of basibranchials takes place, as discussed above. In addition, as Nelson (1969) pointed out, there is a large space between the hyoid arch and first gill arch in the ventral aspect of chondrichthyans. Following Nelson’s proposal for homologous components of basibranchials in chondrichthyans (Nelson, 1969), the second basibranchial would have been developed in this blind space. As shown above, some sharks possess a small cartilage which develops at an analogous position to that of the second basibranchial, whereas in batoid fishes there is no such cartilage except in sawfishes and some stingrays where the development of the medial plate may cover the space. It is speculated that the second basibranchial component may have existed in earlier radiations of chondrichthyan fishes, although, as Nelson (1969) stated, there is no definite evidence for why the blind space exists. The condition is more exaggerated in 94 T MIYAKE AND J. D. MCEACHRAN stingrays in which there is a greater distance between the basihyal and the pseudohyal than in other batoid fishes, and this condition may relate causally to the anterior growth of the first hypobranchial. Therefore, considering the occurrence of the medial plate in sawfishes and stingrays, the development of basibranchials and hypobranchials provides an interesting study which would reveal underlying epigenetic mechanisms for intricate pattern formation of these cartilages. The components of the ventral gill arch skeleton in batoid fishes are identical to those of other chondrichthyans, but they are distinct in both patterns and forms. The basihyal seems to be lost independently in electric rays, Urotrygon stingrays (except U. daviesi) and myliobatoids (Fig. 10). The loss of the basihyal in electric rays and Urotrygon does not result in a concomitant loss of the first hypobranchial. The first hypobranchial exists as a small cartilage in torpedinid and narcinidid electric rays (Fig. 6). In benthic stingrays, the first hypobranchial is a long cartilage which articulates anteriorly with the basihyal if the latter cartilage is present. In myliobatoids, the basihyal and the first hypobranchial are absent. In some individuals of Urolophus jamaicensis, the basihyal and the first hypobranchial form a large continuous cartilaginous plate (Miyake, 1988). We therefore propose that the horizontal bar in sawfishes, guitarfishes, skates and some stingrays represents a composite of the basihyal and the first hypobranchial, with the distolateral rarni of the horizontal bar which represent part or all of the first hypobranchial, as Nelson (1969) suggested. If this is the case, the same condition may hold true in the Jurassic guitarfish Belemnobatis and Spathobatis in which the first horizontal bar may represent a composite of the basihyal and the first hypobranchial. In most guitarfishes and probably the Jurassic Belemnobatis and Spathobatis, the second hypobranchial retains a generalized pattern. However, in Rhynchobatus, the cartilage is subdivided into a lateral and medial portion. In skates, apparently the lateral portion does not chondrify, and this condition is found invariably in skates (McEachran, unpublished data). The third and fourth hypobranchials of guitarfishes and skates are unique in being block-like cartilages which are either separate components or a single component. The concavity and small cartilaginous ridges formed on the ventral surface of these cartilages serve as the passage of the fourth, fifth and sixth afferent branches of the ventral aorta and part of the insertion of the coracobranchialis muscle (Miyake, 1988). It is not known whether these cartilages are the product of the lateral, the medial or both portions of the hypobranchials. The structure of the hypobranchials in electric rays is more variable than in the other four groups of batoid fishes. The Z-shaped pattern of the hypobranchials is retained only in the genus Torpedo. In the genus Hypnos, which has been placed in the same suborder as in Torpedo, the arrangement of the hypobranchials is longitudinal to the axis of the body, as seen in narkinidid and narcinidid electric rays. In Narcine the longitudinal arrangement is arrived at secondarily by fusion of the individual hypobranchial components. However, this observation does not explain underlying developmental processes for this fusion: the size and pattern of blastemas, pattern of chondification or remodelling of initial cartilages. The longitudinal arrangement of the second to the last hypobranchial can be considered a synapomorphy of HTpnos, narkinidid and narcinidid electric rays or a homoplasious (convergent) event between EVOLUI’ION OF BATOIDS 95 Hypnos and narkinidid and narcinidid electric rays. However, it is likely that the underlying processes of this arrangement are different among the three subgroups of electric rays. In sawfishes and stingrays, the development of the second through the last hypobranchial components is ontogenetically associated with the formation of the medial plate. In sawfishes, the second and third hypobranchial components chondrify separately, but the fourth may be included in the middle medial plate. In stingrays, the medial plate possesses a series of small projections along the lateral sides where ceratobranchials articulate with the plate. These projections are more obvious in the medial plate of Urotrygon duuiesi (Figs 8E, 9A). Based on these observations, we propose that the rami projecting posterolaterally from the medial plate in Urotrygon daviesi represent at least the lateral portion of hypobranchials, and that the medial plate is a composite of hypobranchials and basibranchials. In most chondrichthyans, the last hypobranchial (the fifth hypohranchial when five sets of gill arches are present) does not develop as a separate component. In Sguatina and probably the Jurassic Belemnobatzs and Spathobutis, the basibranchial copula possesses a pair of the rami projecting posterolaterally which articulate with the fifth ceratobranchials, suggesting that the rami may represent the lateral portion of the fifth hypobranchials. It is likely that the last basihranchial copula in most chondrichthyans is a composite of several differen c cartilages. It may represent the fifth and sixth basibranchials and one or two hypobranchials. The bridges on the medial plate for passage of the afferent branches of the ventral aorta are peculiar to sawfishes and some stingrays. The phylogenetic significance of this character state is difficult to evaluate because it is highly unlikely that sawfishes and stingrays constitute a clade (Compagno, 1977; Maisey, 198413) and because the presence and state of development of the bridges vary among species and within species of Urotrypon (Miyake, unpublished data). However, since the bridges serve as sites of the insertion for the coracohyo- mandibularis and coracobranchialis muscles (Miyake, 1988), their formation may result from interactions between the muscles and cartilages. When the bridges are not present, the muscles insert on cartilaginous projections on the medial plate (Miyake, 1988). Understanding the interactions between these tissues may elucidate the ontogeny of the medial plate. The functional replacement of the ceratohyal with the pseudohyal has been proposed as one of the synapomorphies of batoid fishes (Maisey, 1984a; Carroll, 1988). However, both the ceratohyal and pseudohyal are found in several groups of batoid fishes, and the pseudohyal is much smaller than the ceratohyal in narkinidid and narcinidid electric rays. Also de Beer (1932) stated that the pseudohyal cartilage is found in sharks. Therefore, the replacement of the ceratohyal with pseudohyal cannot be considered a synapomorphy of batoid fishes. Maisey (1984b) agreed with Compagno (1977) in considering the electric rays the most primitive batoid fishes because in the genus Nurke, the ceratohyal articulates with the hyomandibular. However, the ceratohyal, when present, is connected by ligamentous tissue to the latter cartilage (Miyake, 1988), as Compagno (1977) originally stated. This connection may alternatively be considered a primitive character state for batoid fishes or a derived character 96 T. MIYAKE AND J. D. MCEACHRAN state for electric rays. If it is a primitive state for batoid fishes because the condition is found in other chondrichthyan groups, it is of no use in assessing the phylogenetic position of electric rays within electric ray groups. If it is a derived state for the genus Narke, it is of use in assessing the phylogenetic position of the genus within electric ray groups. The medial fusion of the pseudohyal and the ceratobranchials is unique to stingrays. The degree of the fusion appears to be taxon-specific, although the extent of the fusion is occasionally variable in the species of Urotrygon and Urolophus (Miyake, 1988). The ankylosis of the last two ceratobranchials which serve as part of the insertion of the coracobranchialis muscle appears to be unique to stingrays though the condition is not known for Hexatrygon. Regardless of the phylogenetic significance of fusion of the pseudohyal with ceratobranchials and ankylosis of the last two ceratobranchials, these structures may be of great structural and functional significance. Both structural modifications may be related to the rigid formation of the pectoral fin in stingrays., i.e. the double articulation of the propterygium with the scapulocoracoid (Miyake, 1988), a rigid articulation of the propterygium with the neurocranium by the antorbital cartilage (Garman, 1913; Compagno, 1973, 1977) and possibly the development of the medial plate in the ventral gill arches (Miyake, 1988). Identification of the components, the pattern and evolution of the skeleton of the ventral gill arches in batoid fishes resides in developmental studies, and requires more detailed comparisons of the skeleton throughout chondrichthyans. The examination of the early stages of blastematic condensation would clarify early pattern formation of the components in the ventral gill arch skeleton. The mechanisms by which blastematic condensation leads to the formation of hypobranchials and basibranchials remain unknown. However, based on what has been known about the skeletogenesis of higher vertebrates, it can be speculated that, for instance, a variety of patterns of hypobranchial and basibranchial copulae in batoid fishes as well as in other chondrichthyans is attributed to several underlying developmental mechanisms. The initial size of blastematic condensations for these cartilages (Hall, 1978; Ede, 1983; Thorogood, 1983) and their subsequent modifications by segmentation and bifurcation of the condensations (Shubin & Alberch, 1986; Oster & Murary, 1989) would establish their initial shapes and sizes. Patterns of the chondrification, after the initial condensations of blastemas, would play a role in subsequent patternings of hypobranchial and basibranchial components, especially as seen in the formation of fused hypobranchial and basibranchial components and incomplete or complete separation of the lateral and medial portions of hypobranchials. Processes responsible for patterns include directions of chondrification, necrotic activity or remodelling of the cartilages by absorption (Hinchliffe, 1982; Thorogood, 1983; Hinchliffe & Griffiths, 1984). Thus, developmental studies covering these areas of skeletal research hopefully will in turn aid in identifying several of the components of the ventral gill arch skeleton, i.e. nature and relationships of the basihyal and the first hypobranchial, separate or fused states or both in hypobranchials and different configurations of hypobranchials and basibranchial copulae. Because the patterning of the ventral gill arch skeleton seems to be also closely associated with the pattern of muscles and vascular system (Miyake, 1988), it is imperative to examine epigenetic interactions of the formation of these different structures (Hinchliffe & Johnson, EVOLUTION OF BATOIDS 97 1980; Kean & Houghton, 1987). Alberch & Kollar (1988) concluded, as a workshop report of the Meeting of Craniofacial Development held at the University of Bath in 1987, that “a synthesis between an early patterning process coupled with a more labile set of morphogenetic and inductive interactions provides a suggestive framework to study both the development and evolution of the vertebrate head”. ACKNOM’LEDGEMENTS This study was part of a dissertation preparation in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Texas A&M University by the senior author. We are greatly indebted to the following individuals for the loan of specimens for this study: J. E. Randall and A. Suzumoto (BPBM); T. Iwamoto (California Academy of Science, San Francisco) and A. E. Anderson (currently J.L.B. Smith Institute of Ichthyology, Rhodes University, Grahamstown); I. Nakamura (FRSKU); R. K. Johnson (currently Grice Biological Laboratory, Charleston), D. J. Stewart (currently Center for Limnology, University of Wisconsin, Madison) and T. Grande (Field Museum of Natural History, Chicago); H. Hatanaka and S. Kawahara (Far Seas Fishery Research Laboratory, Distant-water Trawl Resources Section, Japan) ; C. E. Dawson and S. G. Poss (Gulf Coast Research Laboratory Museum, Ocean Springs); R. I. Lavenberg, C. C. Swift and J. A. Seigel (LACM); K. F. Liem, M. L. J. Stiassny (currently American Museum of Natural History, New York) and K. E. Hartel (MCZ); R. Rosenblatt and H. J. Walker, Jr. (SIO); T. B. Thorson (University of Nebraska, Lincoln); R. S. Rosa (Universidade Federal da Paraiba, Brasil); V. G. Springer, S. L. Jewett and L. P. Norrod (USNM). We are also grateful to K. F. Liem, M. L. J. Stiassny and K. E. Hartel for their hospitality and assistance during the visit of the senior author to Museum of Comparative Zoology, Harvard University. We acknowledge Gulf Specimen Marine Laboratories, Inc., Florida, for their donation of embryos and adult specimens of Narcine brasiliensis. T. B. Thorson and H.-J. Chung are also acknowledged for the donation of specimens of Poturnotrygon rnagdulena and Urolophus aurantiacus. Thanks go to M. Rezter, assistant curator of Texas Cooperative Wildlife Collection, Texas A&M University, College Station (currently Department of Ecology, Ethology & Evolution, University of Illinois, Urbana, Illinois), for his curatorial assistance. We acknowledge Brian K. Hall for numerous discussions on development and evolution with the senior author and for his financial support to the senior author. Finally, we thank J. G. Maisey (American Museum of Natural History, New York) and another anonymous reviewer for their comments and suggestions on the first draft of our manuscript. This study was supported in part by the National Science Foundation (INT84-05423) and (BSRB87-00292) to JDM and by the Mini-grants of College of Agriculture and Department of Wildlife and Fisheries Sciences, Texas A&M University and the Ernst Mayr Grant of Museum of Comparative Zoology, Harvard University to TM. The manuscript was completed with support from the Izzac Walton Killam Postdoctoral Fellowship to TM and from Natural Sciences and Engineering Research Council of Canada Operating Grant to B. K. Hall. 98 T. MIYAKE AND J. D. MCEACHRAN REFERENCES ALBERCH, P. & KOLLAR, E. 1988. Strategies of head development: workshop report. Devclopmmt (Supplement), 103: 25-30. ALLIS, E. P., 1923. The cranial anatomy of Chlamydoseluchus anguineus. Acta zoologica, Stockholm, 4: 123-221. CAPPETTA, H., 1987. Chondrichthyes 11: Mesozoic and Cenozoid elasmobranchs. In H.-P. Schultz (Ed.), Handbook of Paleoichthyology, Val. 3B. Stuttgart: Gustav Fischer Verlag. CARROLL, R. L., 1988. Vertebrate Paleontology and Evolution. New York: W. H. Freeman and Company. COMPAGNO, L. J. P., 1973. Interrelatiomhips of living elasmohranchs. In P. H. Greenwood, R. S. Miles & C. Patterson (Eds), Interrelatiomhips of Fishes: 15-61. London: Academic Press. COMPAGNO, L. J. P., 1977. Phyletic relationships of living sharks and rays. American ~oologist,17: 303-322. DE BEER, G. R., 1932. On the skeleton of the hyoid arch in rays and skates. Quarterh Journal of Microscopical Science, 75: 307-320. DINGERKUS, G., 1986. Interrelationships of orectolobilform sharks (Chondrichthyes: Selachii). In T. Uyeno, R. Arai, T. Taniuchi & K. Matsuura (Eds), Indo-Pacijc Fish Biology: Proceedings of the Second International Conference on Indo-Pa+ Fishes: 227-245. Tokyo: Ichthyological Society of Japan. DINCERKUS, G. & DEFINO, T. C., 1983. A revision of the orectolohiform shark family Hemiscyllidae (Chondrichthyes, Selachii). Bulletin of the American Museum of Natural History, 176: 1-94. DINGERKUS, G. & ULHER, D. L., 1977. Enzyme clearing of alcian blue stained whole small vertebrates for demonstration of cartilage. Stain Technology, 52: 229-232. EDE, D. A,, 1983. Cellular condensations and chondrogenesis. In B. K. Hall (Ed.),Cartilage, 2; 143-185. New York: Academic Press. EDGEWORTH, F. H., 1935. The Cranial Muscle of Vertebrates. Cambridge: Cambridge University Press. EL-TOUBI, M. R., 1952. The development of the chondrocranium of the sting dogfish Acanthias uulgaris (Squalus acanthias). Part 11: Branchial arches and extravisceral cartilages. Journal of Morphology, 90: 33-64. EL-TOUBI, M. R. & HAMDY, A. R., 1959. Studies on the head skeleton of Rhinobatus halavi, Rhynchobatus djiddensis and Trygon kuhlii. Publications of Marine Biological Station, Al-Ghardaqa (Red Sea), Cairo University, 10: 3-40. GARMAN, S., 1913. The Plagiostoma. Memoirs of the Museum of Comparative Zoology at Harvard College, 36. Cambridge: Harvard College. GEG ENBAUR, C., 1865. Untersuchungen zur Vergleichenden Anatomie der Wirbelthiere. zweites Heft. I. Schultergurtel der Wirbelthiere 2. Brustjosse der Fische. Leipzig: Wilhelm Englemann. GEGENBAUR, C., 1872. Untersuchungen zur Vergleichenden Anatomie der Wirbelthiere. Drittes Heft. Das Kopfskelet der Seladier, ein Beitrag zur Erkenntniss der Genese des Kopfskeletes der Wirbelthiere. Leipzig: Wilhelm Engelmann. HALL, B. K., 1978. Developmental and Cellular Skeletal Biology. New York: Academic Press. HAMDY, A. R., 1956. The development of the extravisceral cartilages of Rhinobatus halavi. Proceedings of Egyptian Academy of Sciences, 11: 79-83. HAMDY, A. R., 1957. The development of the hyoid arch in Rhinobatidae. Proceedings of Egyptian Academy of Sciences, 12: 58-7 1. HAMDY, A. R., 1973. Studies on the batoid features in the skull of Pteroplatea altavela-the viscerocranium. Proceedings of Egyptian Academy of Sciences, 24: 15-18. HAMDY, A. R. & KHALIL, M. S., 1973. Studies on the arrangement of hypobranchials in Selachii. Proceedings of Egyptian Academy of Sciences, 24: 5-9. HEEMSTRA, P. C. & SMITH, M. M., 1980. Hexatrygonidae, a new family of stingray (Myliobatiforrnes: Batoidea) from South Africa. Ichthyological Bulletin of the 3. L. B. Smith Institute of Ichthyology, Rhodes Universily, 43: 1-17. HINCHLIFFE, J. R., 1982. Cell death in vertebrate limb morphogenesis. In R. J. Harrison & V. Navaratnam (Eds), Progress in Anatomy, 2: 1-1 7. Cambridge: Cambridge University Press. HINCHLIFFE, J. R. & GRIFFITHS, P. J., 1984. Experimental analysis of the control of cell death in chick limb development. In I. Davies & D. C. Sigee (Eds), Cell Aging and Cell Death, 2: 223-242. Cambridge: Cambridge University Press. HINCHLIFFE, J. R. &JOHNSON, D. R., 1980. The Development of the Vertebrate Limb. Oxford: Clarendon Press. HOLMGREN, N., 1940. Studies on the head in fishes. Embryological, morphological and phylogenetic researches. Part I. Development of the skull in sharks and rays. Acta zoologica, 22: 51-267. HOLMGREN, N., 1941. Studies on the head in fishes. Embryological, morphological and phylogenetic researches. Part 11. Comparative anatomy of the adult selachian skull, with remarks on the dorsal fins in sharks. Acta zoologica, 22; 1-100. JARVIK, E., 1977. The systematic position of acanthodian fishes. In S. M. Andrews, R. S. Miles & A. D. Walker (Eds), Problem in Vertebrate Evolution: 199-225. London: Academic Press. JOLLIE, M., 1971. Some developmental aspects of the head skeleton of the 37 mm Squalus acanthias foetus. Journal of Morphology, 133: 17-40, KEAN, M. R. & HOUGHTON, P., 1987. The role of function in the development of human craniofacial form-a perspective. Anatomical Record, 218: 107-1 10. KHALIL, M. S. & HASSAN, S. H., 1973. Studies on the viscerocranium ofRaja circularis. Bulletin of Faculty of Science, Cairo University, 48: 167-185. EVOLUTION OF BATOIDS 99 LAUDER, G. V. & LIEM, K. F., 1983. The evolution and interrelationships of the actinopterygian fishes. Bulletin of Museum of Comparative ,Zoology, Haruard Uniuersily, 150: 95-1 97. LEVITON, A. E., GIBBS, R. H., JR., HEAL, E. & DAWSON, C. E., 1985. Standards in herpetology and ichthyology: Part 1. Standard symbolic codes for institutional resource collections in herpetology and ichthyology. Copeia, 2985(3): 802-832. MAISEY, J. G., 1984a. Chondrichthyan phylogeny: A look at the evidence. Journal of Vertebrate Paleontology, 4(3): 359-371. MAISEY, J. G., 1984b. Higher elasmobranch phylogeny and biostratigraphy. ABBREVIATIONS USED IN FIGURES aff afferent branches of ventral aorta bdg bridge on medial plate hh basihyal cartilage hs b hasibranchial cartilagr cert ceratohyal cartilage cob coracoid bar of pectoral girdle cor-hym coracohyomandibular muscle exhra extrabisceral gill rays hp hypobranchial cartilage HI heart hyh hypohyal cartilage mp medial plate cartilage pseuhy pseudohyal cartilage APPENDIX Material examined Acronyms of museums or institutions housing specimens examined and used for figures in this study follow Leviton et a/. (1985): BPBM: Bernice P. Bishop Museum, Honolulu, Hawaii; FRSKU: Fisheries Research 100 T. MIYAKE AND J. D. MCEACHRAN Station, Faculty of Agriculture, Kyoto University, Maizuru, Japan; FSFL: Far Seas Fisheries Research Laboratory, Distant-water Trawl Resources Section, Japanese Fisheries Agency, Shimizu, Japan; LACM: Los Angeles County Museum of Natural History, Los Angeles, California; MCZ: Museum of Comparative Zoology, Harvard University, Cambridge, Massachusetts; SIO: Scripps Institution of Oceanography, LaJolla, California; TCWC: Texas Cooperative Wildlife Collection, Texas A&M University, College Station, Texas; USNM: National Museum of Natural History, Smithsonian Institution, Washington, D.C. Squatinomorphii Squatiniformes Squatinidae Squatina dumeril LeSueur (TCWC 4214.2, 275 mm TL) Batoidei Pristiformes Pristidae Pristispectinatus Latham (MCZ 36960, 1028 mm TL) Rajiformes Rhinobatoidei Rhinobatidae Rhinobatosproductus Girard (TCWC 6183. I, 554 mm TL); Rhynchobalus djiddensis (Forskal) (MCZ 806, 490 mm TL) Rajoidei Rajiidae Raja (Leucoraja) garmani Whitely (TCWC 1888.1, 283 mm TL) Torpediniformes Torpedinoidea Torpedinidae Torpedo ~alifonticaAyres (MCZ 43, 334 mm TL) H ypnidae Hypnos monopterygium (Shaw & Nodder) (MCZ 38602, 282 mm TL) Narcinoidea Narkinididae Narke japonica (Schlegel) (MCZ 1339, 270 mm TL); Typhlonarke aysoni (Hamilton) (FRSKU 46477, 317 mm TL) Narcinididae Narcine bradiensis (Olfers) (TCWC uncat., 253 mm, 34 mm, 36 mm TL); Discopyge tschudii (Heckel) (FRSKU 105043, 394 mm TL); Benthobatis marcida Bean & Weed (TCWC 1903.1, 137 rnrn TL); Diplobatispictus Palmer (TCWC 1900.1, 119 mm TL) Myliobatiformes Dasyatoidea Urolophidae Urotygon microphthalmum Delsman (USNM 222692, 244 mm TL); Urotygon venezuelae Schultz (TBT 76-34, 235 mm TL); Urotrygon munda Gill (USNM 220612, 201 mm TL); Urotygon nana Miyake & McEachran (SIO 65-167 152 mm TL); Urotrygon reticulata Miyake & McEachran (USNM 222644, 188 mm TL); Urotygon simulatrix Miyake & McEachran (GCRL 13064, 267 mrn TL); Urotrygon chilensis Giinther (LACM 7013, 352 mm TL); Urotrygon rogersi (Jordan & Starks) (LACM 50-57, 355 mm TL); Urotrygon aspidura (Jordan & Gilbert) (CAS 51834, 284 rnrn TL); Urotrygon daviesi Wallace (BPBM 24578, 481 mm TL); Urolophus jamaicensis (Cuvier) (TCWC 0815.1, 285 mm TL); Urolophus halleri Cooper (SIO uncat., 381 mm TL); Urolophus concentricus (Osburn & Nichols) (LACM 31771, 278 mm TL); Urolophus paucimaculatus Dixon (FSFL ED189, 407 rnm TL); Urolophus cruciatus Lacepede (TCWC uncat., 231 mm TL); Urolophus bucculentus Macleay (FSFL EC361, 538 mm TL); Urolophus aurantiacus Muller & Henle (TCWC uncat., 251 mm TL) Dasyatididae Dasyatis americana Hildebrand & Schroeder (TCWC 5820.1, 614 mm TL); Himantura zmpricata (Bloch & Schneider) (MCZ 59269, 401 mm TL); laeniura lymma (Forskal) (TCWC 5276.1, 442 mm TL) Gymnuridae Gyrnnura micrura (Bloch & Schneider) (TCWC uncat., 203 mm TL)