590.5 FI N.S. 99 Jun 28, 2002 Biology OpiTJv the l hannel Cattish ophysi: Siluriformes) ieldiana 2 (Permanence of Paper). FIELDIANA Zoology NEW SERIES, NO. 99 Morphology and Development of the Postcranial Skeleton in the Channel Catfish Ictalurus punctatus (Ostariophysi: Siluriformes) Terry Grande Associate Professor Department of Biology Loyola University 6525 North Sheridan Road Chicago, Illinois 60626 U.S.A.* Research Associate Department of Geology Field Museum of Natural History 1400 South Lake Shore Drive Chicago, Illinois 60605-2496 U.S.A. * Address for correspondence. Judith D. Shardo Department of Biological Sciences University of South Alabama 124 Life Sciences Building Mobile, Alabama 36688 U.S.A. Accepted January 16, 2002 Published June 28, 2002 Publication 1518 S/0L0GY LIBRARY W Bimu HAH NOV 1 9 2002 PUBLISHED BY FIELD MUSEUM OF NATURAL HISTORY © 2002 Field Museum of Natural History ISSN 0015-0754 PRINTED IN THE UNITED STATES OF AMERICA Table of Contents Abstract 1 Introduction 2 Methods 2 Skeletal Preparation 2 Histological and SEM Preparation 3 Developmental Staging Method 3 Materials 3 Developmental Series Examined 3 Adult Materials Examined 4 Abbreviations 4 Results 4 Developmental Staging 5 Discussion 14 Overview of Median Fin Fold Formation and Development 14 Overview of Paired Fin Development 15 Development of the Caudal Fin Skeleton 18 Development of the Weberian Apparatus/ Dorsal Fin Unit 21 Variation in Development 27 Acknowledgments 28 Literature Cited 28 2. Early formation of vertebral proto- centra 9 3. Development of the caudal fin skeleton 9 4. Development of the anal fin skeleton .... 13 5. Development of the median fins and axial skeleton 16 6. Development of the vertebral column .... 17 7. Formation of caudal fin epurals and neural spines 19 8. Intraspecific variation in caudal fin structure 20 9. Development of the Weberian appara- tus/dorsal fin unit 22 10. Development of the dorsal fin spines .... 23 1 1 . Adult anterior vertebral region, lateral view 25 12. Adult anterior vertebral region, dorsal and ventral views 26 List of Tables List of Illustrations 1 . Scanning electron micrograph of the development of Ictalurus punctatus 1 . Summary of specimen sampling corre- lated with the first occurrence of defin- ing criteria 5 2. List of defining and concurrent charac- ters for stages 1-18 6 111 Morphology and Development of the Postcranial Skeleton in the Channel Catfish Ictalurus punctatus (Ostariophysi: Siluriformes) Terry Grande1 Judith D. Shardo2 Abstract The morphology and development of the postcranial skeleton of three independent series of the channel catfish Ictalurus punctatus were studied using a combination of techniques (histology, SEM, skeletal clearing and staining). More than 2,000 specimens ranging in developmental stage from a fertilized egg to individuals about 400 mm TL were examined. Our results show individual variation in the onset of the hatching and foraging periods, and in the appearance or number of several skeletal structures (e.g., number of hypurals). Our results also show that regardless of this variation, the sequence of development of the postcranial structures is consistent within the series studied, and that the development of these structures is correlated more with the size of the fish than with age. Because of the consistent pattern of postcranial skeletal development observed, we were able to construct an ontogenetic staging scheme consisting of 18 developmental stages, each characterized by one defining criterion. Additional and more variable characters that occur concurrently with each of the 18 defining criteria are identified as concurrent features. This staging method facilitates future comparisons with the developmental patterns of other fish taxa, and is independent of age. As part of this study, careful developmental descriptions of the Weberian apparatus, vertebral column, and paired and median fins were made. A primary goal of the study was to better understand the developmental relationship between the Weberian apparatus and the dorsal fin skeleton. Together they form an extremely unusual anatomical complex whose development and function are tightly linked. During development, the fourth neural spine of the Weberian apparatus forms a tight articulation with the first two proximal radials of the dorsal fin. In catfishes that exhibit a similar modification of the dorsal fin, sound production has been im- plicated. Through detailed anatomical descriptions, this study examined contested homologies of the vertebral column and caudal fin. Such homologies include the caudal fin epurals, which in Ictalurus punctatus form as independent elements that later fuse to the neural spines of posterior vertebrae. New terminology is suggested for several skeletal structures to reflect their devel- opmental origin. 1 Associate Professor, Department of Biology, Loyola University, 6525 North Sheridan Road, Chicago, Illinois 60626; Research Associate, Department of Geology, Field Museum of Natural History, Roosevelt Road at Lake Shore Drive, Chicago, Illinois 60605-2496. 2 Department of Biological Sciences, University of South Alabama, 124 Life Sciences Building, Mobile, Alabama 36688. FIELDIANA: ZOOLOGY, N.S., NO. 99, JUNE 28, 2002, PP. 1-30 1 Introduction Until recently, researchers examining patterns of vertebrate development, and more specifically actin- opterygian development, focused on a handful of "model species" (e.g., the zebrafish Danio rerio) to make generalizations about the formation of mor- phological structures such as the neural tube and vertebral column (Kuwada et al., 1990; Kimmel et al., 1995). However, studies such as those of Schul- tze and Arratia (1988, 1989), Bemis and Grande (1992), Arratia and Schultze (1992), Shardo (1995), and Grande and Bemis (1998) have illustrated sig- nificant variation in skeletal development within Ac- tinopterygii. Obviously, no one fish species can be used as a model for all of Actinopterygii. This study describes the development of the postcranial skeleton (post early cleavage stages) of Ictalurus punctatus, commonly known as the channel catfish, one of the most common ictal- urids in North America. Phylogenetically, /. punctatus is considered by many researchers (e.g., Lundberg & Baskin, 1969; Arratia, 1992; de Pinna, 1996; Coburn & Grubach, 1998) to be a relatively basal siluriform because it exhibits a primitive caudal fin and a relatively primitive Weberian apparatus in comparison to other cat- fishes. On the other hand, as a siluriform, /. punctatus exhibits many modifications and bone fusions not present in outgroup ostariophysans such as cypriniforms, characiforms, and gon- orhynchiforms (Rosen & Greenwood, 1970; Fink & Fink, 1981, 1996; Grande & Poyato- Ariza, 1999). Although specimens of adult catfishes are common in North American museum collec- tions, developmental material and consequen- tially developmental studies are rare (e.g., Armstrong, 1962; Arratia, 1990, 1992; Kobay- akawa, 1992; Adriaens, 1998; Coburn & Gru- bach, 1998). Studies of the development of /. punctatus are rarer still, and have focused only on specific parts of the skeleton. For example, Kindred (1919) and Eaton (1937) examined skull development, while Al-Rawi (1966) ex- amined the early development of the Weberian apparatus. The current study examined the de- velopment of the entire postcranial skeleton (vertebral column, Weberian apparatus, dorsal and caudal fins, and pectoral and pelvic fins) using three independent developmental series, each from a single spawn, for a total of more than 2,000 specimens. We found that, contrary to Lundberg and Baskin (1969), the caudal fin skeleton of /. punctatus exhibits considerable morphological variation within and between the developmental series examined. Despite this variation, consistent features of axial skeleton formation can also be identified, and the de- tailed developmental timing of particular skel- etal elements appears to correlate more closely with the size of the fish than with its age. Hypotheses about the evolution, development, function, and homologies of different elements of the Weberian apparatus have been proposed by many authors (e.g., Krumholz, 1943; Martin, 1963; Alexander, 1964; Rosen & Greenwood, 1970; Gayet, 1986; Chardon & Vandewalle, 1997). Some of these hypotheses are in conflict with each other. For example, Coburn and Futey (1996) argue that the claustrum in otophysans is derived from the first supraneural, while Fink and Fink (1981) argue that the claustrum forms from a disassociated part of the first neural arch. As part of our study we investigated various hypoth- eses of Weberian apparatus formation in light of the new morphological data we obtained. Our ob- servations show that the development of the We- berian apparatus in /. punctatus is functionally linked to the development of the dorsal fin skel- eton. We term this functional unit the Weberian apparatus/dorsal fin unit, and explore some impli- cations of this unit as a stabilizer for the dorsal fin spine. We also comment on its possible role in sound transmission. Methods Skeletal Preparation Specimens were preserved in either 10% buff- ered formalin or 4% buffered paraformaldehyde. Skeletal material was prepared using a modified version of Dingerkus and Uhler's (1977) tech- nique for staining and counterstaining bone and cartilage. In this method, bone is stained with aliz- arin red and cartilage is stained with alcian blue. Trypsin was used to render the soft tissue trans- parent. An ethyl alcohol series was substituted for the KOH step in the standard Dingerkus and Uhler method because the larval specimens are fragile. Once cleared and stained, the specimens were stored in glycerin. Specimens were dissect- ed, examined, and drawn under a Wild MZ8 dis- secting microscope. Total lengths (TL) were taken from all specimens. Standard lengths (SL) were FIELDIANA: ZOOLOGY recorded for specimens with defined caudal fin supports. Histological and SEM Preparation Formalin-fixed specimens ranging in size from 5 mm to 20 mm TL were prepared for histological examination using one of two methods. To help establish the onset of vertebral column ossifica- tion, specimens were decalcified in De-cal (Na- tional Laboratories), dehydrated in an ethyl alco- hol series, cleared in xylene, embedded in paraf- fin, and cut in 10-u,m-thick transverse or sagittal sections. To examine the morphology of the We- berian apparatus and vertebral column, specimens were prepared using the low-viscosity nitrocellu- lose (LVN) embedding technique of Thomas (1983). Specimens were decalcified in formic acid, dehydrated in an alcohol series, and embed- ded in a graded concentration series of LVN. Transverse sections 40 u,m thick were cut using an American Optical 860 sliding microtome. Both thin and thick histological sections were stained with a modified version of Humason's (1972) he- matoxylin and picro-ponceau procedure. Embryonic (i.e., prehatch) and yolk sac larval specimens were studied with scanning electron microscopy (SEM). For embryonic specimens, the chorion was removed as the first step. Dechorion- ated embryos and yolk sac larvae were washed in sodium cacodylate buffer and then postfixed in an aqueous solution of 2% osmium tetroxide. Each specimen was then dehydrated through an alcohol series, treated with Peldri II, dried in air, coated with 400 A of gold/palladium (Young et al., 1995), and examined in a Cambridge 240 SEM. Developmental Staging Method Early ontogeny consists of growth and a se- quence of developmental changes over time. These rates of growth and developmental change are not necessarily constant or correlated with each other. Thus, age or length does not consis- tently correspond to a level of development, par- ticularly among different species (Fowler, 1970; Reimchen & Nelson, 1987). For purposes of com- parison, in this study the developmental sequence of the axial skeleton is divided into morphological stages. Each stage is characterized by one mor- phological feature that acts as the defining crite- rion (Shardo, 1995). Additional and more variable characters that occur concurrently with the defin- ing criterion of a particular stage but are not nec- essarily linked to the defining character are de- fined as concurrent features (Shardo, 1995). This method of developmental staging allows for com- parisons with other species, regardless of the length or age of the individuals being compared. Materials Developmental Series Examined Three independent developmental series of channel catfish were examined. Two separate se- ries were spawned from eggs obtained from Osage Catfisheries (Osage Beach, Mo.). The third series was spawned from eggs obtained from the Catfish Genetics Research Unit (U.S. Department of Agriculture, Agriculture Research Services, Stoneville, Miss.). Each developmental series was raised at a different location but under approxi- mately the same conditions (i.e., simulated stream facilities at about 26°C). Series A was raised by Grande in a greenhouse facility with natural light- ing at Loyola University, Chicago. Series B was raised by Bemis laboratory personnel working at the Osage Catfish facility and obtained from W. E. Bemis (University of Massachusetts, Amherst). Series C was raised by Shardo at the Stoneville Catfish Facility. A total of 2,293 specimens (985 specimens in series A, 258 specimens in series B, 1,050 specimens in series C) were collected from these series, ranging from newly fertilized eggs to foraging subadults with ossified axial skeletons. Samples of at least seven specimens were col- lected for 18 days (series B) and 30 days (series A and C). At least four samples were preserved each day within the collecting period. The number of samples and the frequency of collecting de- creased in all three series toward the last days of collecting, after the fish had achieved a total length of 35-40 mm or were foraging freely. All fish from series A are deposited in the fish collection at Loyola University, Chicago (LU D081090). Illustrated specimens were assigned Loyola University (LU) catalogue numbers. Se- ries B is deposited in the University of Massa- chusetts Ichthyological Collection, Amherst (UAM Fl 1257), and series C is housed at the Uni- versity of South Alabama, Mobile (USA 040645). GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS Adult Materials Examined Materials examined to assess the morphological condition of the postcranial skeleton in larger specimens and related taxa are as follows: Clarias liberiensis: 1 specimen (SL = 60 mm): FMNH 50272 (cleared and stained). Corxdoras aeneus: 3 specimens (SL = 42-69 mm): FMNH 54832 (cleared and stained). Doras carinatus: 1 specimen (SL = 1 10 mm): FMNH 53192 (cleared and stained). Helogenes marmoratus: 1 specimen (SL = 60 mm): FMNH 701 15 (cleared and stained). Ictalurus punctatus: 9 specimens (SL = 160-400 mm): FMNH 16711. 86267. 6430. 63699 (skele- tons): LU F.082256. F082257. F082258. F.082259. F082260 (cleared and stained, alcohol). Ictalurus balsanus: 2 specimens: FMNH 51269 (dis- articulated skeletons). Ictalurus furcatus: 1 specimen: FMNH 73900 (very large disarticulated skeleton). Noturus gyrinus: 1 specimen: (SL = 56 mm): FMNH 42269 (cleared and stained). Rhamdia nicaraguensis: 1 specimen: (SL = 150 mm): FMNH 5904 (cleared and stained). Rhamdia nasuta: 1 specimen: (SL = 75 mm): FMNH 35319 (cleared and stained). Trachelyopterichthys taeniatus: 1 specimen: (SL — 65 mm): FMNH 69929 (cleared and stained). Trichomycterus banneaui: 2 specimens: (SL = 65-71 mm): FMNH 70014 (cleared and stained). Trichomycterus laticeps: 2 specimens: (SL = 38-45 mm): FMNH 79128 (cleared and stained). Abbreviations The following abbreviations are used for insti- tutions and anatomical terminology: FMNH Field Museum of Natural History. Chi- cago LU Loyola University. Chicago UAM University of Massachusetts. Amherst act actinotrichia anp anterior nuchal plate ar anterior radial (= supraneural of Grande & Lundberg. 1988: pterygiop- hore of Fink & Fink. 1996) boc basioccipital br bony ridge chc chordacentrum (sensu Schultze & Ar- ratia. 1986) cl claustrum dare dorsal arcocentrum {sensu Schultze & Arratia. 1986: = basidorsal of Patter- son, 1968; Schultze & Arratia. 1986; Grande & Bemis. 1998) df dorsal fin dfl dorsal fin spine 1 df2 dorsal fin spine 2 dl dorsal lamina drl-7 dorsal radials 1-7 ep epural ha hemal arch hyl-6 hypural 1-6 in intercalarium lpdt lepidotrichia na neural arch nc notochord ns neural spine ns4 neural spine 4 nt neural tube OS os suspensorium Pf pectoral fin bud phy parhypural pnp posterior nuchal plate pr proximal radial pu preural centrum p4a anterior part of the transverse process p4p posterior part of the transverse process rn radial nodule SC scaphium sn supraneural soc supraoccipital tp4 transverse process 4 tr tripus ul-2 ural centrum 1-2 un uroneural vl-10 vertebra 1-10 varc ventral arcocentrum (sensu Schultze & Arratia. 1986: = basiventral of Patter- son, 1968: Schultze & Arratia. 1986: Grande & Bemis. 1998) Results Although some individual variation between and within particular series of /. punctatus is ev- ident (such as in the caudal fin skeleton), the basic pattern and sequence of bone ossification is con- sistent among the three series. As modified from Bemis and Grande (1992) and Shardo (1995). we divide the development of /. punctatus into three major periods: the embryonic period (defined as the period from fertilization to hatching), the yolk sac larval period, and the foraging period. The onset of the particular developmental pe- FIELDIANA: ZOOLOGY Table 1. Summary of samples exhibiting first occurrence of defining criteria. Series A Series B Series C Stage (LU D.081090) (UAM F11257) (USA 040645) 1 (Day 2, sample 7) (Day 2, sample 8) (Days ?-5, samples ?-40) 2 7.6 mm (day 3, sample 10) 7.6 mm (day 4, sample 14) 8.2-9.5 mm (days 6-7, samples 41-50, hatching) 3 8.1-10 mm (day 4, sample 15) 8.6 mm (day 4, sample 17) 8.4-10.2 mm (day 7, sample 51) 4 9.4-10.7 mm (day 6, sample 20) 9.0 mm (day 5, sample 18, 9.9-12.2 mm (days 8-10, hatching) samples 52-55) 5 9.8-11 mm (day 6, sample 24, 9.7-10.0 mm (day 5, sample 20) 1 1.9-12.6 mm (day 1 1, sample hatching) 56) 6 10.2-11.5 mm (day 7, sample 10.5-1 1.0 mm (day 6, samples 11.5-13.6 (days 11-12, samples 28) 22-23) 56-57) 7 9.7-12.1 mm (day 8, samples 10.9-12.2 mm (day 7, samples 11.5-13.6 mm (day 12, sample 31-35) 24-26) 57) 8 12.0-13.6 mm (day 9, sample 12.5-13.5 mm (days 8-9, 11.5-13.6 mm (day 12, sample 39) samples 28-29) 57) 9 12.6-14.4 mm (day 10, sample 14.2-14.3 mm (day 10, sample 13.9-14.4 mm (day 13, sample 44) 32) 58) 10 14.4 mm (day 10, sample 46) 14.0-14.4 mm (day 11, sample 13.4-16.1 mm (days 14-15, 33) samples 59-60) 11 15.0 mm (day 11, sample 51) 14.8-15.0 mm (day 12, sample 16.2-17.6 mm (day 17, sample 34) 62) 12 15.0-15.5 mm (day 13, sample 15.6-16.0 mm (day 13, sample 16.9 mm (day 17, sample 62) 53, foraging) 35, foraging) 13 15.5-16.1 mm (day 14, sample 16.5-17.5 mm (day 15, sample 27.1-31.2 mm (day 36, sample 60) 36) 81, foraging) 14 16.9-18.1 mm (day 18, sample 17.5-18.3 mm (day 17, sample 27.1-31.2 mm (day 36, sample 78) 37) 81) 15 23.0-26.0 mm (day 27, sample 21.7-24.0 mm (day 23, sample No samples 89) 39) 16 40.0 mm (day ?, sample 93) 30.5-32.9 mm (day ?, sample 40) No samples No samples 17 45.0-50.0 mm (day ?, sample 95) 50.0-60 mm (day ?, sample 95) No samples 18 No samples No samples riods varied among the three series. This is un- surprising, because variations in the timing of the three developmental periods (e.g., number of days to hatching) could reflect the slightly different wa- ter temperatures or light regimens in the environ- ments in which the fish were raised (Fowler, 1970). The sampling time periods for each catfish series are listed below and in Table 1. More im- portant, our results show that the development of particular structures in /. punctatus is closely cor- related with the length of the fish. This correlation is consistent with the findings of Faustino and Power (1998) for Spams aurata. The development of the axial skeleton is further divided into 18 stages (Table 2). The defining cri- teria proposed here are characters that show little or no variation in developmental sequence and are common to teleosts (e.g., the presence of hypural 1) or more specifically to ostariophysans. Defining criteria were easily observed in all three devel- opmental series. The concurrent features listed in Table 2 include both postcranial and cranial char- acters that first appear in that stage in at least one of the three series. Any variation in the timing of the concurrent features among the series is men- tioned in the description of each stage. Age and length of the specimens in a single stage varied somewhat among the three series and are treated as concurrent features. Developmental Staging Embryonic Period. The embryonic period for /. punctatus extends from fertilization to hatching. In the three series examined, this developmental period lasted about five to seven days. We cannot more precisely determine the length of the em- bryonic period for series A and B because the GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS Table 2. Defining and concurrent criteria for stages 1-18. Stage Defining criteria Concurrent characters 10 11 12 13 14 Presence of 4 visceral arches; at least 50 somites formed Presence of hypural 1 of caudal fin Presence of at least 4 cartilaginous hypurals Protocentra mineralize in anterior region of vertebral column Presence of ural centrum 1 (uT) Formation of ural centrum 2 (u2) Neural arches meet and begin to ossify at dorsal midline posterior to dorsal fin Mineralization of all vertebral centra Fusion of vertebral centra 2 and 3 First 2 dorsal fin radials enlarge and articulate with spine of neural arch 4 of Weberian complex Fusion of vertebrae 2, 3. and 4 Transverse process of vertebra 4 of Weberian apparatus articulates with pectoral girdle All Weberian ossicles are ossified All hypurals are at least two-thirds ossified Notochord present but unconstricted; pectoral fin buds present; brain with three distinct divisions; presence of 1 continuous fin fold Parhypural and hypural 2 often but not always formed; actinotrichia formed in caudal fin; basidorsal forming; mandible, hyomandibula, otic capsule, and cranial floor forming in cartilage; utricular and saccular otoliths formed; opercular facet ossified; notochord constricted anteriorly Basidorsals and basiventrals formed; orbital and epiphyseal bar forming; pterygoids and visceral arches 1-5 formed in cartilage: supracleithra and cleithra formed; pelvic fin buds present; dentaries begin to ossify in series A and B Formation of autogenous epural in caudal fin skeleton; formation of anal radials begins; presence of at least 7 cartilaginous proximal radials in dorsal fin; dentaries ossify in series C Uroneural 1 (un 1 ) formed in series C; parasphenoid begins to ossify; teeth on dentary. maxilla forming; presence of lateral line on dor- sal skull roof marking the beginning of frontal bone formation in series A and B; basioccipital and exoccipitals ossifying; early ossification of hyomandibular in series C First appearance of Weberian apparatus ossicles (i.e., tripus, scaphium, intercalarium) and transverse process of vertebra 4; presence of all proximal dorsal fin radials; initial ossification of parhypural and hypurals in series C; formation of sixth hypural in large specimens Ossification of Weberian ossicles begins; formation of teeth on epibranchial 4; presence of a supraneural anterior to the dorsal fin; formation of teeth on epibranchial 4; formation of first dorsal fin spine; formation of distal radials in anal fin in series C Uroneural 1 elongates to the distal margin of u2; second dorsal fin spine forms; lateral line forms around orbit and preopercular region; ossification of parhypural in series A and B Ossification of all branchiostegal rays; formation of distal radials of anal fin in series A and B; premaxilla with 1 row of teeth First appearance of a cartilaginous claustrum; formation of "epurals" associated with posterior 7 preural centra; tooth formation on both ventral and dorsal gill arches continues; presence of 5 distal radials of dorsal fin; formation of basal segment of pectoral fin spine Elongation of "epurals"; formation of the basal segment of pectoral fin spine in series A and B; ossification of pectoral girdle and fins in series C Hyomandibula begins to ossify in series A and B; neural and hemal arches are ossified Each half of the neural arch of vertebra 5 meets along dorsal midline All "epurals" are at least on-half ossified; most skull bones (e.g., supraoccipital, pterygoids) have formed and are well-ossified; caudal fin forks and looks like adult structure FIELDIANA: ZOOLOGY Table 2. Continued. Stage Defining criteria Concurrent characters 1 5 Proximal radials of anal fin ossify 1 6 Completion of Weberian/dorsal fin complex 17 Hypurals 3 and 4 form a unit with u2 18 Fusion of pul, ul, unl, and hypurals 1 and 2, forming compound centrum of caudal fin Pectoral girdle ossified in series A and B; all but last dorsal fin radial are ossified Fusion of "epurals" with corresponding neural spines in series A and B Completion of skull lateral line system; continued ossification of skull and growth Fusion of hemal spines to corresponding centra, anterior to pu2; adult coloration observed Note: Defining criteria and concurrent characters are explained in the text. precise time of fertilization was not recorded, al- though it is known that fertilization occurred about one day prior to the first collection. A more precise fertilization time is known for series C because the eggs were fertilized by sperm in the laboratory. Series A: July 9-July 13/14 (stages 1-5, day 6) Series B: May 29-June 3 (stages 1-4, day 5) Series C: June 25-June 30 (stages 1-2, day 6-7) Yolk Sac Larval Period. This period extends from hatching to the complete depletion of the yolk sac. During this time all postcranial axial skeletal elements are formed, although not com- pletely ossified (e.g., caudal fin hypurals). The starting date for the yolk sac larval period was determined when the majority of the fishes in each series had hatched. Series A: July 1 5— July 19 (stages 6-11, ending on day 1 1 ) Series B: June 4-June 11 (stages 5-11, ending on day 12) Series C: June 30-July 15 (stages 3-12, ending on day 17) Foraging Period. The start of the foraging pe- riod is characterized by the disappearance of the yolk sac. The fishes are now foraging on their own. It is a time of continued skeletal ossification and growth. Foraging was determined either by observing food in the gut tract of a fish or by direct observation of a fish feeding. Series A: July 20-Sept. 15 (collections termi- nated) (stages 12-18) Series B: June 12-June 22 (collections termi- nated) (stages 12-16) Series C: July 16-July 30 (collections terminat- ed) (stages 13-14) Stage 1 This study is concerned with the development and ossification of the postcranial skeleton, not with early cleavage stages. We thus begin by de- scribing the level of development of specimens just prior to the formation of elements in the post- cranial skeleton. These specimens already exhibit head lift (Fig. 1A). At this stage the optic vesicles are formed, the division of the brain into three primary brain regions (prosencephalon, mesen- cephalon, and rhombencephalon) is obvious, the branchiomeres have undercut the head laterally, four visceral arches have formed, at least 50 so- mites are present, Meckel's cartilage and the max- illary barbels are beginning to form, and the pec- toral fin buds first appear. Also in stage 1, the fishes exhibit an unconstricted notochord and a continuous fin fold. Neural and hemal arches, hy- purals, and median fin pterygiophores are not yet present. Stage 2 This stage is characterized by the presence of a cartilaginous hypural 1 in the caudal fin skel- eton. In most specimens examined the parhy- pural and hypural 2 are also present in cartilage. The appearance of hypural 1 is quickly followed by the formation of additional hypurals. We found only a few specimens of stage 2 in which hypural 1 was the only hypural in the caudal region. Actinotrichia are present in the ventral part of the caudal fin only. The notochord shows a series of constrictions resulting in a series of protocentra (i.e., centra precursors; Arratia, 1991; Grande & Bemis, 1998). In the skull, Meckel's cartilage plus the cartilaginous hy- GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS Fig. 1. SEM images of early developmental stages of Ictalurus pimctatus. A. Stage 1. showing the early development of the pectoral fin buds, head lift, and the early formation of the maxillary barbel. Scale bar = 1 mm. B. Stage 3. Scale bar = 1 mm. C and D. Stage 4. showing the development of the pectoral fin buds, the dentary barbels, and the early formation of the dorsal fin. Scale bar = 2 mm in C. 1 mm in D. For abbreviations, see p. 4. omandibula and the otic capsule are forming. The utricular and saccular otoliths have formed, and skull dermal bone formation begins w ith the opercular facet of the opercular bone (i.e.. the articular surface of the opercle for the articu- lation with the hyomandibula). Stage 3 At least four cartilaginous hypurals are formed at this stage (Fig. 3A). In addition, cartilaginous dorsal arcocentra are formed along the length of the constricted notochord and cartilaginous ven- tral arcocentra are found only posterior to the yolk sac. As discussed in Arratia and Schultze (1992). arcocentra will bear neural and hemal arches, re- spectively, and correspond to the basidorsals and basiventrals. respectively, of Schultze and Arratia (1986) and Grande and Bemis (1998). They thus form the chondral components of the vertebrae. Other internal characters observable at this stage include the presence of a cartilaginous hy- omandibula. pterygoids, and ceratobranchials 1- 5. as well as a cartilaginous orbital and epiphyseal bar and lateral expansion of Meckel's cartilage. Ossification of the dentary was observed in stage 3 of series A and B but was delayed to stage 4 in series C. Ossification of the maxillae at the base of the maxillary barbel, supracleithra and cleithra. and enlargement of the opercle are also evident. Both the utricular and saccular otoliths are clearly visible in specimens at this stage. Externally, the body and tail are still coiled around the yolk (Fig. IB). An additional set of barbels has formed median to the maxillary bar- bels, which have elongated. The pelvic fin buds appear as small protrusions immediately poste- rior to the yolk sac. The pectoral fins have en- larged slightly with the formation of the basal scapulocoracoid cartilage. Distal to the scapu- locoracoid cartilage is a sheet of condensing cartilage that will form the radials of the pec- toral fin. The head is primarily free from the yolk, attached only in the region of the opercle, thus freeing the lower jaw and allowing the mouth to open. FIELDIANA: ZOOLOGY Fig. 2. A. Sagittal histological sections showing the first sign of protocentra mineralization. Arrows point to individual protocentra in a specimen 9 mm TL (stage 3- 4). Anterior is to the left. B, Photograph of cleared and stained specimen (13 mm TL, LU F.082289, stage 8) showing mineralization of the chordacentra from within the notochordal sheath. Arrows point to the outer margin of the notochordal sheath. Anterior is to the left. Fig. 3. Development of the caudal fin skeleton from an unconstricted notochord stage to the formation of the compound terminal centrum. A, Embryo, 9.2 mm TL (stage 3) (LU F.082275). B, Embryo, 10.4 mm TL (stage 5), showing the formation of ural centrum 1 (ul). Note that ul begins to form before the posterior preural centra (LU F.082276). C, Yolk sac larva, 11.5 mm TL (stage 6), showing the formation of ural centrum 2 (LU E082277). D, Yolk sac larva, 13.1 mm TL (stage 8), showing the formation of the compound centrum, the autogenous epural, the median "epurals," and ossifica- tion of the parhypural. Only the first five median "epur- als" are illustrated (LU F082278). E, Foraging juvenile, 20 mm Tl (stage 14-15), showing further development of caudal skeleton (LU F.082279). F, Foraging juvenile, 40 mm TL (late stage 15), showing the association of hypurals 3 and 4 with u2 (LU F082280). G, Subadult, 168 mm TL (no stage), showing a fully formed caudal fin skeleton (LU F082281). Anterior is to the left. Os- sification of hypurals and mineralization of centra are shown with stippling. Unstippled areas are cartilaginous. For abbreviations, see p. 4. ha Phy D hy3-6 hy3-6 -— _ __hy3+hy4+u2 pul+ul+unl+ phy+hyl-2 •4+u2 pul+u2+unl+ phy+hy 1 -2 GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS Stage 4 Stage 4 marks the beginning of vertebral centra formation. Fishes at this stage in series A and B are 5-6 days old and have not yet hatched, but they are 8-10 days old and post-hatching in series C. After the notochord has constricted and the ar- cocentra have formed, chordacentra ("ring cen- tra," Lundberg & Baskin, 1969; Collette et al., 1983; Schultze & Arratia, 1989; Arratia, 1991) begin to develop from within the notochordal sheath of each protocentrum (Laerm, 1982; ter- minology of Grande & Bemis, 1998). In /. punc- tatus, mineralization of dorsal and ventral chor- dacentra occurs simultaneously. Each dorsal and ventral pair of chordacentra quickly enlarges, fus- es, and forms a ring around each protocentrum (Figs. 2A and B). Although at this stage in de- velopment only a thin mineralized surface layer is present on each protocentrum (most protocentra have mineralized in specimens of 10-11 mm TL), the chordacentra will quickly thicken, and auto- centra, which arise as direct ossifications, will sur- round each chordacentrum. The autocentra (ob- served in individuals about 13 mm TL; see Stage 6) will in turn enlarge and complete the formation of the vertebral centra. Chordacentrum formation in /. punctatus is unlike that reported by Arratia and Schultze (1992) for Oncorhynchus and sev- eral other salmonids, which primarily exhibit only ventral chordacentra. In salmonids, chordacentra enlarge dorsally and are eventually supplanted by autocentra. Not surprisingly, variation in chorda- centrum formation is present among teleosts, but the dorsal-ventral pattern observed in /. punctatus seems fixed among ostariophysans. Chordacen- trum formation in /. punctatus is accompanied by a straightening of the trunk and tail (Figs. 1C and D). Concurrent features present at this stage include an autogenous cartilaginous epural in the caudal fin skeleton, lepidotrichia in the entire caudal lobe, a minimum of seven cartilaginous proximal radials in the dorsal fin, rudimentary ventral ar- cocentra above the yolk sac, and a third pair of barbels under the chin. In the pectoral fins the scapulocoracoid cartilage is enlarging and three proximal radials are present. The medialmost pec- toral radial appears continuous with the scapulo- coracoid cartilage. With the exception of the an- terior radial in the dorsal fin, radials in all fins form and ossify from rostrad to caudad. During this time the cartilaginous proximal anal fin radials form. In the specimens examined the proximal anal fin radials begin to form in an an- terior to posterior direction before the anal fin lobe forms. In siluriforms the middle radials of both the anal and dorsal fins are absent as separate elements. It has been suggested that the middle radials are lost (Fink & Fink, 1981) or fuse with the proximal radials (Grande, 1987). In the spec- imens observed, the proximal radials formed as single and solid forms. Slightly later in develop- ment, the distal radials form between the right and left halves of the lepidotrichial bases. Stage 5 Stage 5 is defined by the presence of ural cen- trum 1 (ul). As observed in Amia calva (Grande & Bemis, 1998) and Oncorhynchus mykiss (Ar- ratia & Schultze, 1992), ural centrum 1 in /. punc- tatus begins to mineralize before mineralization of the abdominal protocentra is complete (Figs. 3B and C). Ural centrum 1, however, mineralizes dif- ferently from the other centra. Instead of the si- multaneous dorsal-ventral pattern of the abdomi- nal centra, ural centrum 1 mineralizes from the ventral side of the protocentrum. Growth of ven- tral chordacentra forming the ural centra appears to be common among primitive teleosts (Schultze & Arratia, 1988, 1989). Ural centrum 1 through- out its development is associated with hypurals 1 and 2 only. An unmineralized protocentrum lies dorsal to the parhypural at this stage and is pre- sumed to be the precursor to preural centrum 1 (pul). Concurrent characters include the complete mineralization around the anteriormost protocen- tra in series A and B, but around over two-thirds of the total protocentra (beginning anteriorly) in series C. Also in series C, uroneural 1 (unl) was observed extending from the completed postero- dorsal corner of ural centrum 1 . This single uro- neural eventually extends along the dorsolateral margin of the notochord. It thickens and straight- ens into a rodlike structure and. together with the epural, stiffens the epaxial portion of the caudal fin (Lauder, 1989). The early formation of uro- neurals (i.e., membrane versus cartilage bone) varies among teleosts (Greenwood, 1966; Patter- son, 1968; Schultze & Arratia, 1989; Arratia & Schultze, 1992). Our observations show that uro- neural 1 in /. punctatus forms as membrane bone without a cartilaginous precursor (Fig. 3D). This pattern of formation of uroneural 1 in /. punctatus 10 FIELDIANA: ZOOLOGY may be a synapomorphy of ictalurids and possibly siluriforms. In the pectoral fins, the scapulocoracoid carti- lage has elongated both dorsally and anteroven- trally along the medial surface of the cleithra. For- mation of pectoral fin rays has begun. In the more slowly developing pelvic fins, six fin rays extend out from a small cartilaginous bar in the pelvic fin buds. The parasphenoid begins to ossify, and by now at least one-half of the dentary is ossified and supports a single row of teeth. Four bran- chiostegal rays have ossified on each side of the skull, and the opercle continues to grow and os- sify. In series A and B, lateral line canals were observed on the dorsal skull roof, marking the start of frontal bone formation. Fully formed fron- tal bones appeared in stage 6 in series C. The basioccipital and exoccipitals ossify at the anterior end of the notochord. Stage 6 This stage is characterized by the mineraliza- tion of ural centrum 2 (u2). Ural centrum 2 forms from a ventral chordacentrum, as does ural cen- trum 1, and is closely associated with cartilagi- nous hypurals 3 and 4. In a few specimens of series C, cartilaginous hypurals 4 and 5 articulate with ural centrum 2. In most specimens examined ural centrum 2 never fuses with ural centrum 1. Concurrently in series C (11.5-13.6 mm TL), ossification of the central portions of the parhy- pural and hypurals 1-5 begins. Ossification of the parhypural in series A and B does not occur until later in development (about stage 8), at around 13.5 mm TL. Hypurals in series A and B do not begin ossifying until after the parhypural begins to ossify. Larger specimens in all series often con- tain a sixth cartilaginous hypural. From this stage onward, a gradually increasing percentage of specimens examined contain a sixth hypural, which is very slow to ossify. In many specimens examined, specifically in series A and B, hypural 6 never develops. At this stage (10.7-13.6 mm TL) in all series, all dorsal fin radials are present, as well as at least six dorsal fin rays. In general, the skull bones exhibit slightly more ossification than in stage 5, but more important, this stage marks the first appearance of Weberian apparatus ossicles (i.e., tripus, scaphium, and intercalarium) and the transverse processes of vertebra 4. These elements, although preformed in cartilage as basi- dorsals and basiventrals, respectively, ossify very rapidly, suggesting the importance of a function- ing Weberian apparatus in larval catfishes. Stage 7 Neural arches posterior to the dorsal fin meet along the dorsal midline and begin to ossify prox- imally. The formation of a neural arch bridge is typical for ostariophysans (Fink & Fink, 1981). Neural arches 5-11, which lie ventral to the dorsal fin radials, do not meet along the dorsal midline to form neural spines as do the more posterior arches. Instead they are separated and prevented from meeting by the elongation and intervention of the dorsal fin radials as the Weberian apparatus/ dorsal fin complex forms. The Weberian ossicles begin to ossify at this stage in fishes about 1 1 mm TL, starting with the tripus. Not surprisingly, the Weberian ossicles os- sify before ossification of the more posterior neu- ral or hemal arches is complete. During this stage hemal arches also meet in the midline, forming hemal spines posterior to the anus. Additional concurrent characters include the presence of teeth on epibranchial 4 of the gill arches, and for- mation of the first dorsal fin spine. The first spine will become the locking mechanism for the sec- ond fin spine in the fully formed dorsal fin. In addition, a supraneural forms anterior to neural arch 3 of the Weberian apparatus. This supra- neural will eventually fuse with arch 3 and be- come a component of the Weberian apparatus. Stage 8 Stage 8 is marked by the completion of all chordacentra (i.e., mineralization of all protocen- tra). During this stage the compound centrum of the caudal fin, consisting of preural centrum 1 and ural centrum 1 , forms before mineralization of the abdominal and preural protocentra is complete (Figs. 3B and C). Eventually, as the abdominal centra form, in an anterior to posterior direction, they catch up with the formation of the ural centra (Fig. 3D). Histological cross sections show that most chordacentra have thickened, and that the anterior neural arches and parapophyses are for the most part fused with their corresponding cen- tra. This is achieved by perichondral ossification of the dorsal and ventral arcocentra and their fu- sion to the autocentra by a thin superficial ossifi- cation (Schultze & Arratia, 1992). Arcocentra and GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS 11 autocentra in these vertebrae are now indistin- guishable, and the vertebrae exhibit the adult con- dition (i.e., well-ossified structures developmen- tally formed both chondral and perichordal bone; Grizzle & Rogers, 1985). Concurrently in series A and B, uroneural 1 forms at the distal margin of ural centrum 1 . In- terestingly, the formation of uroneural 1 in series A and B occurs later than in series C but at ap- proximately the same TL, 11.5-13.6 mm. A lengthening of uroneural 1 to the distal margin of ural centrum 2 was observed in all series. At this stage the second dorsal fin spine forms. In ictalurids the base of the second fin spine forms an ectoconvex surface that grips its cartilaginous proximal radial. When the radial ossifies, it fuses with the lepidotrichia and becomes the base of the definitive spine (Reed, 1924). Although hypural and parhypural formation oc- curs early in development, ossification of the cau- dal fin supports occurs later. At this stage the par- hypural begins to ossify at its midpoint in series A and B. This ossification began during stage 6 in series C. The parhypural, however, does not fuse to the compound centrum, and the distal end of this element does not ossify until much later. Stage 9 between the bones. The functional significance of this is discussed in a subsequent section. This stage also marks the appearance of the car- tilaginous claustrum. The claustrum is the last of the Weberian ossicles to form and the last to os- sify. Concurrent characteristics observed at this stage include an increase in number of teeth on the ventral and dorsal gill arches and the forma- tion of five distal radials in the dorsal fin. This stage also marks the formation of a series of car- tilaginous median elements associated with the posterior six free preural vertebrae (Fig. 3D). The formation of these elements has not been previ- ously reported for /. punctatus. In the course of their development, each of the median elements will fuse undetectably to its corresponding neural spine. This condition and the possible homology of these elements with epurals are addressed fur- ther in the Discussion. In the pectoral fins, the ventral ends of the sca- pulocoracoid cartilages continue to elongate and meet midventrally. Of the eight to nine fin rays present, the one on the leading edge of the fin has become thickened, and in series C the basal por- tion ossifies and forms small projecting hooks. In the pelvic fins, the small cartilaginous bars have expanded into triangular plates with seven to eight projecting rays. Fishes at this stage exhibit a complete fusion of vertebral centra 2 and 3. The Weberian appa- ratus in ictalurids consists of ossified elements supported by a compound centrum that is a fusion of vertebrae 2, 3, and 4. These vertebrae fuse with each other sequentially. The fusion of these centra begins ventrally and ends with their final fusion at the dorsal margins. Concurrently occurring features include ossifi- cation of all branchiostegal rays, the formation of teeth on the premaxilla, and the formation of car- tilaginous distal radials in the anal fin in series A and B (occurring earlier, in stage 7, in series C). Stage 10 Stage 10 is characterized by the articulation of the dorsal fin with the Weberian apparatus via neural spine 4. In ictalurids the proximal dorsal fin radials enlarge and expand ventrally, articulat- ing with the spine of neural arch 4 of the Weber- ian apparatus. In adult specimens examined, this articulation is very strong, with virtually no play Stage 11 Stage 1 1 is defined by the complete fusion of vertebra 4 with the combined vertebra of 2 + 3 to form the foundation of the Weberian apparatus. Although vertebra 5 is associated with the We- berian apparatus and partially articulates with it ventrally, as seen in many specimens examined (Al-Rawi, 1966), it is not considered part of the Weberian apparatus in ictalurids (Fink & Fink, 1981, 1996; Coburn & Grubach, 1998). Additional characteristics observed at this stage include an elongation of the autogenous epural, the median elements associated with posterior neural spines, plus the formation of the basal seg- ment of the pectoral fin spines in series A and B. In series C the basal ossification of the pectoral fin spine that began in stage 10 has extended dis- tally to two-thirds of the spine. In series C, all of the elements of the pectoral girdle and fins are ossified except for a small central portion of the scapulocoracoid associated with the radials and the distal portion of the spine. 12 FIELDIANA: ZOOLOGY Fig. 4. Development of the anal fin skeleton. A, Foraging juvenile, 13 mm TL (stage 9). B, Adult. 30 mm TL (no stage). Anterior is to the left. For abbreviations, see p. 4. Stage 12 Stage 12 is defined by the articulation of the anterior part of the transverse process of vertebra 4 (tp4) with the suspensorium of the pectoral gir- dle. Also in this stage the uroneural extends to the distal margin of the hypurals. Its length is com- plete, and in subsequent stages it will enlarge only in girth. All neural and hemal arches are ossified, although parapophyses posterior to hemal arch 9 are cartilaginous. In the skull, dermal bones en- large in size and the lateral and medial walls of the hyomandibula begin to ossify. In series C, the first sign of ossification in the hyomandibula oc- curs much earlier, in stage 5. Teeth are present on ceratobranchial 5 of the ventral gill arches. Stage 13 Stage 13 is defined by the complete ossification of all Weberian ossicles. The last ossicle to ossify is the claustrum. The two halves of the neural arch of vertebra 5 meet along the dorsal midline. The joining of these arches does not form a neural spine (there are no neural spines on arches 5-1 1). In fact, the only reason they are able to join is that neural arch 5 is always positioned between the elongated dorsal fin radials 2 and 3. In other words, the modified dorsal fin radials never inter- fere with the formation of this arch as they do with the more posterior arches. The anterolateral processes of the pelvic basipterygia are formed. Stage 14 All hypurals are at least two-thirds ossified. In most cases, only the distal rim of each hypural remains cartilaginous. This rim remains cartilag- inous for a long time, and is still present in fish of 40 mm TL. In series C, hypurals 1-5 are os- sified, except for the distal rims, but hypural 6 has only now reached two-thirds ossification. All pos- terior median elements associated with neural arches, which include the autogenous epural, are at least one-half ossified. Likewise, skull bones such as the opercular series, pterygoids, dentary, hyomandibula, supraoccipital, pterotics, and ba- sioccipital, are ossified. The frontal bones contin- ue to enlarge, and this enlargement is correlated with the development of the lateral line system. The caudal fin is now deeply forked, as is char- acteristic of the adult condition in this species. Stage 15 At this stage the radials of the anal fin are well ossified (Figs. 4A and B). As stated previously, the middle radials are most likely lost in ictalur- ids. The proximal radial equivalent of these radi- GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS 13 als ossifies first, followed by the middle radial equivalent. The distal radials of the anal fin ossify last (Fig. 4B). By this stage the pectoral girdle has ossified in series A and B (this event occurred in stage 1 1 in series C). and all but the last dorsal fin pterygiophore is bone. Hemal arches anterior to preural centrum 2 are fused with their corre- sponding centra, as is characteristic of most os- tariophysans (Fink & Fink. 1981: Fig. 3G). Stage 16 Stage 16 is defined by the completion of the Weberian apparatus dorsal fin complex. In addi- tion, the once cartilaginous median elements at- tached posteriorly to the preural neural spines have fused completely with the corresponding spines in fish of 40 mm TL in series A and B. It is possible that this fusion occurred earlier in se- ries C. In fish 40 mm TL and larger it is impos- sible to distinguish these elements from their neu- ral spines. By this stage only one autogenous epural remains positioned dorsal to the compound caudal centrum. The fusion of posterior elements such as these with their neural spines strengthens the caudal fin and adds support for musculature (Lauder. 1989). Stage 17 Hypurals 3 and 4 become articulated with ural centrum 2 and form a unit. According to Lund- berg and Baskin (1969). ural centrum 2 never fus- es with the compound centrum, but a fusion of ural centrum 2 and hypurals 3 and 4 does occur. We agree with Lundberg and Baskin (1969) that a structural unit does form between ural centrum 2 and hypurals 3 and 4. but we do not agree that a fusion of the three elements is always the case. We have found that in some cases, hypurals 3 and 4 fuse with each other, but in other specimens of the same size they only articulate. In the majority of the developmental material examined (speci- mens 50 mm TL and smaller), fusion of the three elements does not occur. It appears that only in very large specimens does such a fusion occur. It is possible that the fusion of hypurals 3 and 4 with each other and with ural centrum 2 is a very late developmental occurrence. We also have ob- served, although rarely (LU F082282). fusion of the ural centrum 2 unit with the compound cen- trum in series A. In fish about 45 mm TL. the skulls are com- pletely ossified, although the right and left frontal bones still do not suture completely in the dorsal midline. The basipterygia of the pelvic fins are beginning to ossify. The last structures to ossify are the distal radials of the dorsal and anal fins. At this stage the development of the lateral line is complete. Stage 18 In state 18. the compound centrum in the cau- dal fin forms, consisting of preural centrum 1, ural centrum 1. uroneural 1. and parhypural and hy- purals 1 and 2. This is the last stage in the de- velopment of the postcranial skeleton. This stage occurs quite late in development and is seen only in fish longer than 50 mm TL. Also in this stage all hemal spines anterior to preural centrum 2 fuse with their corresponding centra. Fishes at this stage exhibit adult pigmentation. Discussion In this section the development of the postcra- nial skeleton of /. punctatus is examined in dis- crete developmental units — the median fin folds, the pectoral and pelvic fins, the Weberian appa- ratus/dorsal fin complex, and the caudal fin skel- eton. The results of our study are discussed in conjunction with the findings of other researchers in an attempt to better understand the develop- ment of the postcranial skeleton in ictalurids. De- bates and controversies involving the homologies of certain skeletal structures (e.g.. the anterior nu- chal plate) are addressed. Finally, we discuss the variation within and between each of the devel- opmental series examined. An understanding of variation is key to developmental studies such as this one, and to anatomical studies in general. Overview of Median Fin Fold Formation and Development The sequence of median fin formation and the order of development of their constituent radials, rays, and spines varies among actinopterygians (Dunn. 1983). The median fins of /. punctatus form from a thin continuous ridge or fold that extends from behind the head around the posterior 14 FIELDIANA: ZOOLOGY tip of the notochord and ventrally to the anal opening. No secondary fin folds are present. By the end of the yolk sac period all median fins have formed, and the fins resemble their adult shapes. Interestingly, the sequence of median fin forma- tion does not parallel the sequence of skeletal sup- port formation of these same fins. In other words, the first fin lobe to form is the dorsal, followed by the caudal and then the anal. The first fin sup- ports to form, however, are in the caudal fin, fol- lowed closely by those in the dorsal fin and then in the anal fin. The sequence of fin formation in /. punctatus differs from the sequences described for paddlefishes (a lower actinopterygian) by Be- mis and Grande ( 1 999) in that an anterior to pos- terior direction of fin formation was not observed. On the other hand, the fin formation sequence in /. punctatus directly parallels that described by Fuiman (1983) for other ostariophysans (e.g., Hy- pentelium etowanum), and for esocoids and scom- broids as described by Martin (1983) and Collette et al. (1983), respectively. It is possible that the sequence of fin formation seen in /. punctatus is typical for ostariophysans and possibly teleosts in general, but not for lower actinopterygians. Figure 5 illustrates the formation of the median fins and skeletal supports in four developmentally different specimens of /. punctatus from series A and B. Although specimens of series C followed the same sequence of defining criteria as series A and B, they were generally larger when they achieved each stage (see Table 1). In embryonic fishes about 7 mm TL (not illustrated), the first fin supports to form (i.e., the parhypural and hy- purals 1-2) are in the caudal region below the upturned notochord. The notochord extends al- most to the posterodorsal margin of the continu- ous median fin fold. No other fin supports are pre- sent at this time. In embryonic fishes about 10 mm TL (Fig. 5A), the dorsal fin fold begins to appear, and seven dorsal radials are discernible. A distinct caudal fin lobe is not evident at this time, although one autogenous epural is formed above the notochord, and five hypurals with at least 13 fin rays are present below it. On the ventral side, 18 anal fin radials have formed in an anterior to posterior direction in fishes of this size, even though no anal fin has formed. The median fin fold now appears to be discontinuous between the dorsal fin and the remainder of the fin fold. In fishes about 1 1 .5 TL (Fig. 6A) the dorsal fin has enlarged and fin rays are present in a one-to-one pattern with their corresponding radials. Ural cen- trum 1 is formed in the caudal fin. By the time the now yolk-sac larval fish have reached about 12-14 mm TL (Figs. 5B and 6B), the dorsal ra- dials have elongated toward the developing We- berian apparatus, all caudal fin skeletal supports are formed in cartilage, the caudal fin rays are segmented, and the fin begins to take on its char- acteristic forked appearance. In the anal fin, the proximal radials have elongated and the distal ra- dials begin to form. Also in this stage the adipose fin forms. Remnants of the median fin fold are present between the adipose and caudal fins. From this point on (foraging period), as illustrated in Figures 5C and D and 6C and D, median fin de- velopment is devoted to the ossification of the skeletal supports, the formation of the dorsal fin spines, and the forking of the caudal fin. In fishes 25 mm TL (Fig. 5D), no sign of the median fin fold remains. The formation of the median fins in /. puncta- tus, and possibly other ostariophysans, seems to be functionally correlated. The caudal fin sup- ports, first to form, are necessary for locomotion and are possibly involved in propulsive escape maneuvers from predators. The dorsal fin and its spines, second in the series to develop, aid in the defense of the fish. Both fins enhance the surviv- ability of the free-swimming yet immature fish and thus are strategically important to form first, before the anal and pelvic fins. Overview of Paired Fin Development The formation of the median fins is clearly in- dependent of the formation of the pectoral and pelvic fins. The pectoral fin buds appear first in fishes about 6.8 mm TL (Fig. 1A). They are not attached to the body but appear to emerge from the yolk sac covering. As the fish's body increases in height the yolk sac gets smaller and the pec- toral fin buds move closer to the sides of the body, eventually to become part of the flank when the yolk sac is depleted and the ventral body wall forms. Based on the development of series C, in- ternal skeletal support begins with the formation of the scapulocoracoid cartilage posterior to the cleithrum in specimens of 8.5-10 mm TL. Pos- terior to the scapulocoracoid is a layer of cartilage that will form three radials when the fishes reach 10-12 mm TL. The anteriormost radial forms as a part of the scapulocoracoid cartilage; the other two are separate. As the larvae exceed 1 2 mm TL, the six or seven fin rays present begin to ossify, and the scapulocoracoid cartilage elongates dor- GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS 15 r-i—~- Fig. 5. Series of cleared and double-stained specimens showing the development of the median fins and postcranial axial skeleton. Arrows point to the formation of the median fins. A. Embryonic larvae, 10 mm TL (stage 3-4). B. Yolk sac larvae. 15.2 mm TL (stage 11-12). C. Foraging juvenile. 15.2 mm TL (stage 12). D. Foraging juvenile, 25 mm TL (stage 14-15). sally and anteroventrally along the medial side of the cleithrum. In specimens of 13-16 mm TL. the coracoid portions have elongated sufficiently to meet midventrally. The anteriormost fin ray has a thick ossified basal portion and will become the spine. This spine becomes two-thirds ossified and has projecting hooks as larvae increase to 17.5 mm TL. At this point ossification has begun in all 16 FIELDIANA: ZOOLOGY B Fig. 6. Series of alizarin-stained specimens showing the development of the vertebral column, median fin supports, and dermal skull bones. Arrows mark the dorsal fin fold. A, Yolk sac larvae, 11.5 mm TL (stage 4). B, Yolk sac larvae, 13.7 mm TL (stage 8). C, Foraging juvenile, 19 mm TL (stage 14). D, Foraging juvenile, 23.5 mm TL (stage 15). of the pectoral girdle and fins except for a small portion of the scapulocoracoid cartilage associat- ed with the radials and tips of the radials. Pelvic fins form later in development, when the larvae are about 8.5-10 mm TL (series C). Pelvic fin buds form along two membranous creases formed by the attachment of the posterodorsal portion of the yolk sac to the body. The buds lie lateral to the fin fold and just anterior to the anal opening. By the time the fishes have reached 12 mm TL, the basipterygia have condensed as small cartilage rods, which further expand into trian- gular shapes as the fishes grow to 16-17.5 mm TL. Seven to eight fin rays form, and both fin rays and basipterygia are ossified in specimens 32 mm TL and larger. GRANDE & SHARDO: POSTCRANIAL SKELETON IN 1CTALURUS PUNCTATUS Development of the Caudal Fin Skeleton Our observations of the development of the caudal fin skeleton of /. pimctatus show some fun- damental consistencies in the three series exam- ined, as well as considerable individual variation between and within the series. Our observations differ from those of Lundberg and Baskin (1969), who found little individual variation among the specimens of /. pimctatus they examined. In this study we highlight the different types of caudal skeleton variation observed but will focus on the general developmental patterns found in all three series. The sizes apply to series A and B. As discussed in Francois (1966), Monod (1968), Laerm (1976), Schultze and Arratia (1989). Arratia and Schultze (1992), and Grande and Bemis (1998), the first vertebral elements to form in actinopterygians are the cartilaginous dor- sal and ventral arcocentra (sensu Arratia & Schul- tze, 1992). From within a constricted notochord, chordacentra form, followed by the perichondral ossification of the arcocentra and their fusion with the autocentra, which in turn surround the chor- dacentra. In /. pimctatus, the arcocentra (i.e., neu- ral and hemal arch precursors) have formed in conjunction with the parhypural and hypurals in fishes about 7 mm TL (Fig. 3A, stage 2). The protocentra do not begin to mineralize until the fish reaches about 9-10 mm TL. As described earlier in the discussion of stages 4. 5, and 6. min- eralization of the protocentra begins in the ante- rior region of the vertebral column, just behind the skull, and continues posteriorly. Before the mineralization process reaches the posterior cen- tra, however, the first ural centrum begins to form along its ventral margin (Fig. 3B), associated only with hypurals 1 and 2. As ural centrum 1 contin- ues to develop, it enlarges and, in specimens of about 1 1 mm TL. is attached to preural centrum 1 (Fig. 3C). Interestingly, in no specimen exam- ined was an autogenous preural centrum 1 ob- served. As illustrated in Figures 3E and F, some of the cleared and stained specimens show verti- cal demarcations within the compound centrum, presumably between preural centrum 1 and ural centrum 1. We infer from these specimens, and agree with Lundberg and Baskin (1969). that the compound centrum is formed from two protocen- tra (i.e., preural centrum 1 and ural centrum 1 pul + u 1 ) that have developed together, with ural cen- trum 1 forming first. Ural centrum 2 remains as- sociated with hypurals 3 and 4, and only in a few specimens examined became fused into the com- pound centrum. In specimens of 13 mm TL, a thin, single uro- neural (unl) has formed and is fused to the pos- terodorsal margin of the compound centrum (Fig. 3D). Uroneural 1 has a wavelike appearance at this point, but it will become characteristically rigid as it enlarges. Although uroneurals in prim- itive teleosts are preformed in cartilage (Schultze & Arratia, 1989; Arratia & Schultze, 1992), we found that uroneural 1 in /. punctatus develops from membrane bone. Additionally, Arratia (per- sonal communication, 2000) found that in speci- mens of /. pimctatus that she has examined, os- sification of uroneural 1 begins caudally and con- tinues anteriorly, surrounding the developing ural neural arch, and later fuses with the compound centrum. We were not able to corroborate her ob- servations. In our developmental material it ap- pears that uroneural 1 begins to form rostrally. In specimens of increasing size and age, uroneural 1 increases in length and eventually reaches the posterior margin of hypural 6 (e.g., specimens of 20 mm TL). It is likely that variation in uroneural formation occurs, but, in the specimens we ex- amined, uroneural 1 does not appear to begin its development caudally and ossify rostrally. Other caudal fin characteristics include the presence of an enlargement of ural centrum 2, and the first sign of bone in the parhypural. The siluriform caudal skeleton as described by Lundberg and Baskin (1969) has a single epural positioned above the neural arch of pu 1 + u 1. We found that this is true for adults, but the adult condition has a complex ontogenetic origin in /. punctatus. We observed the formation of six car- tilaginous median elements positioned and usually attached to the posterior edge of the posteriormost neural spines (Figs. 3 and 7). Although in most specimens examined these elements are loosely attached posteriorly to the neural spines, in LU F082278 and LU F082279 two and three autog- enous elements were observed (Figs. 3D and E). We recognize that the condition in these speci- mens may be an anomaly. On the other hand, it may indicate separate ossification centers for these elements which differ from the ossification centers of the neural arches or spines. During de- velopment, all of these elements elongate, ossify, and completely fuse with their corresponding neu- ral spines (Figs. 3E-G and Fig. 7). With the ex- ception of the last epural in the adult channel cat- fish, there is no indication that cartilaginous ele- ments were ever associated with posterior neural 18 FIELDIANA: ZOOLOGY Fig. 7. Photograph of cleared and stained specimens showing the formation of the autogenous epural and the epurals associated with neural spines. A, Yolk sac lar- vae, 11.0 mm TL (stage 4-5). B, Yolk sac larvae, 15 spines. The homology of these elements is debat- able, but, based on the development of these el- ements (i.e., as median elements that elongate and ossify in the same pattern as the autogenous epur- al) and their placement in the body plane (i.e., in sequence with the autogenous epural), we enter- tain the possibility that these elements are serial homologues of the autogenous epural (Monod, 1968). We acknowledge that epurals associated with preural centra 3-6 have not been reported in ostariophysans, but we argue that until a more thorough survey of skeletal development in prim- itive teleosts is conducted, this interpretation can- not be ruled out. At the very least, the "epural" condition in /. punctatus may provide insight into the much debated origins and homologies of epur- als. As defined by Goodrich (1930), epurals are modified radials. Schultze and Arratia (1989) and Arratia and Schultze (1992) argue that epurals are detached neural spines, while Patterson (1968) and Grande and Bemis (1991) argue that epurals are serial homologues of supraneurals. In /. punc- tatus, these structures are clearly associated with neural spines. The more anterior ones abut and eventually become incorporated into their spine, while the autogenous epural is positioned directly above a stunted neural spine of the compound centrum in the caudal skeleton. With this said, we also observed that these median structures form as cartilaginous condensations after the formation and almost complete ossification of their corre- sponding neural spines. In other words, the for- mation of these elements seems to be secondary to the formation of neural spines, and to occur from separate ossification centers. As discussed by Arratia and Schultze (1992), epurals form in different ways, and those that arise from indepen- dent cartilages can be interpreted as neural spines that have lost their arches. If their interpretation is correct, then our observations would support the connection between epurals and neural spines. As the ossification of the caudal fin skeleton continues (Fig. 3E), the "epurals" elongate and ossify as discussed above, the hypurals ossify ex- cept at their distal margins, and hypurals 1 and 2 begin to show signs of fusion with the compound mm TL (stage 11). C, Foraging juvenile, 26.3 mm TL (stage 16). Note that in B, a second autogenous epural is present (marked by an arrow), corresponding to the neural spine of preural 2 (pu2) (LU F.082289). For ab- breviations, see p. 4. GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS 19 centrum. Uroneural 1 has elongated, extending past the distal edges of the hypurals. Internally (as seen only in cleared and stained material), speci- mens of about 20 mm TL still show an articula- tion line between pul and ul (Figs. 3E and F). In specimens of 20-50 mm TL, hypurals 3 and 4 are separate, and although they are in contact with ural centrum 2, they are not fused with it. Figure 3F shows that ural centrum 2 forms a cap around the base of the articulating hypurals 3 and 4 in fishes of about 40 mm TL. As reported by Lund- berg and Baskin (1969) and Fink and Fink (1981), ural centrum 2 is fused with the bases of hypurals 3 and 4 in most siluroids (in Diplomystidae, hy- purals 3 and 4 remain separate elements: Arratia, 1987). This unit in turn articulates with but never fuses with the compound centrum. Our observa- tions corroborate these findings in most specimens examined (Fig. 3G). Our results indicate, how- ever, that the caudal fin skeleton continues ossi- fying, and that additional fusions of skeletal ele- ments occur long into the adult stage. Although the fusion of hypurals 3 and 4 with each other and with ural centrum 2 seems to be the common condition, we have also found variation in com- parably sized adult specimens in which hypurals 3 and 4 never fuse with ural centrum 2. Ural cen- trum 2 remains as a cap around the bases of hy- purals 3 and 4, as illustrated in Figure 3F. In other specimens examined, hypurals 3 and 4 do fuse with ural centrum 2, and ural centrum 2 fuses into the compound centrum (e.g., LU F082282). An additional caudal fin anomaly observed and worth reporting is the amount of intraspecific var- iation in hypural number (Fig. 8). Intraspecific and interspecific variation in hypural loss and fu- sion has been reported by Lundberg and Baskin (1969) and Arratia (1983), and although interspe- cific variation is common in catfishes, intraspecif- ic variation in hypural number is rare. Lundberg and Baskin (1969) found considerable intraspecif- ic variation in a population of Noturus gyrinus, which they argued is the result of hypural fusions, but they found none in the 13 species of Ictalurus they also examined. Arratia (1983) found consid- erable interspecific variation in hypural number within Trichomycteridae (subfamily Trichomyc- terinae), but found intraspecific variation only in the Andean trichomycterids (e.g., Trichomyeterus laucaensis). Ictalurus punctatus specimens of about 10 mm TL and smaller had only five or fewer hypurals. If a sixth hypural forms, it forms in fishes of more than 1 1 mm TL. Although ictalurids are diag- Fig. 8. Cleared and stained specimens showing varia- tion in the number of caudal fin hypurals. A, Adult, 30 mm TL, with five hypurals (LU F.082284). B. Adult, 27 mm TL. with six hypurals (LU F.082283). nosed in part by the presence of six hypurals (Lundberg & Baskin, 1969; Nelson, 1994, p. 00), over one-third of the specimens we examined in our developmental series had only five. We dis- count the possibility of early hypural fusions as an explanation for this phenomenon and agree with Lundberg and Baskin (1969) that hypural fu- sion does not necessarily take place with increas- ing size. We also think that it is hypural 6 that has been lost, because when a hypural is absent, it is always in the position of the sixth, which is the last of the series to develop. In no specimen examined with five hypurals were rudiments of the sixth hypural observed, and in no specimen with six hypurals were hypurals 5 and 6 ever fused or partially fused. The occurrence of five hypurals instead of six is widespread among all three developmental series examined and, like the 20 FIELDIANA: ZOOLOGY different observed configurations of hypurals 3 and 4 with ural centrum 2, seems to be random. We therefore suggest caution when using hypural number as a caudal fin character to diagnose /. punctatus. It is possible that the number of hy- purals may not be a reliable character to diagnose species within Ictaluridae. Development of the Weberian Apparatus/ Dorsal Fin Unit The Weberian apparatus, diagnostic of the Oto- physi (Rosen & Greenwood, 1970), consists of a series of modified anterior centra, neural arches, and pleural ribs that connects the gasbladder to the back of the skull. When the gasbladder pul- sates in a sound field, high-frequency vibrations are transmitted from it via the Weberian ossicles to the back of the skull and then to the inner ear (Alexander, 1964, 1965). This system, along with the lateral line system, enables the fish to receive a wide range of sound frequencies. Although the Weberian apparatus is assumed to function in the same basic way among otophysan subgroups, morphological variation among these subgroups is apparent. This variation, or possible specializa- tion, is exemplified by the Siluroidei and includes, at the primitive level, a fusion of vertebral centra 2-4, loss of the articular process of the interca- larium, and modifications of the tripus, os suspen- sorium, and the transverse process of the fourth centrum (Fink & Fink, 1981, 1996). The Weberian apparatus of /. punctatus and many other catfishes is functionally complex be- cause of its close association with the dorsal fin skeleton (Chardon, 1968). In /. punctatus, the an- terior two proximal radials of the dorsal fin are expanded, elongated, and tightly articulated with the neural spine of the fourth vertebra of the We- berian apparatus. In adult /. punctatus, the We- berian apparatus and the dorsal fin skeleton es- sentially form an interconnected unit whose de- velopment can be considered together; this in turn poses some interesting functional questions. This study begins with the first appearance of identifiable Weberian ossicles in catfishes of about 10 mm TL (stage 6, series A and B). At this point in development, all of the basidorsals (i.e., dorsal arcocentra) have formed along the vertebral col- umn. Basiventrals (i.e., ventral arcocentra) poste- rior to the second centra have also formed. Basi- ventrals 1 and 2 are absent in siluroids (Fink & Fink, 1981). Hypotheses concerning the deriva- tion and homologies of Weberian ossicles are many. Watson (1939), Bamford (1948), and Ro- sen and Greenwood (1970) argue that the sca- phium is derived from basidorsal 1 that consists of two processes, one dorsally oriented and an- other anteriorly oriented. Matveive (1929), Rad- ermaker et al. (1989), and Vandewalle et al. (1990), however, argue that the scaphium is formed from basidorsal 1 plus an ossification of mesenchyme. Lai Hora (1922) states that cartilag- es from the skull contribute to the formation of the scaphium. Basidorsal 2, according to most au- thors (e.g., Watson, 1939; Fink & Fink, 1981; Vandewalle et al., 1990; Chardon & Vandewalle, 1997), will become the intercalarium, which at this point in the development of /. punctatus re- mains relatively indistinct, as are basidorsals 3 and 4. The claustrum, which some researchers ar- gue is derived from neural arch 1 (e.g., Fink and Fink, 1981), or is derived from mesenchyme (e.g., Watson, 1939), or is homologous with the first supraneural (e.g., Gayet, 1982; Coburn & Futey, 1996), is not present in /. punctatus at this time. Basiventral 3 and possibly the third pleural rib form the tripus (Fink & Fink, 1981, 1996). In /. punctatus, the tripus consists of a dorsal part that will extend anteriorly and, when fully formed, will flatten to articulate with the intercalarium. The more ventral part of the tripus, the transfor- mator process, is long and threadlike in 10-mm fish. It will form a crescent-like structure on the ventral side of the complex centrum of the We- berian apparatus and become embedded in the tu- nica externa of the gasbladder. Basiventral 4 de- velops at a rapid rate. It is already thicker and more elongate relative to the other basiventrals. It will form the transverse process of the fourth cen- trum, and its anterior projection will articulate with the suspensorium of the shoulder girdle (Lundberg, 1975). The os suspensorium, although not present until later in development, forms from the ventral side of the transverse process and is embedded in the gasbladder. The first signs of ossification in the Weberian ossicles occur in fish of about 12 or 13 mm TL (Fig. 9A). The first ossicle to begin ossifying is the tripus (i.e., dorsal component), followed by the leading tip of the transverse process of basi- ventral 4. The transformator process of the tripus and the proximal end of the transverse process ossify later. Basidorsal 4 has grown considerably larger than the other basidorsals, and its dorsal tip begins to bend posteriorly, approaching the first proximal radial of the dorsal fin. Basidorsals 3 and GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS dr3-7 df2 df l ^ >/ A Fig. 9. Development of the Weberian apparatus/dorsal fin unit. A. Yolk sac larvae, 12.3 mm TL (stage 7) (LU 4 will form neural arches that join to an enlarged supraneural that projects forward and articulates with the ventral part of the supraoccipital (Fink & Fink, 1996). This large supraneural forms above vertebrae 2 and 3 and is present in fish of about 1 1 mm TL. Although in /. punctatus it initially forms as one crescent-shaped structure (Fig. 9A), some researchers argue for multiple ossification centers for this element (e.g., Coburn & Futey. 1996). Gayet and Chardon "(1987) and Chardon and Vandewalle (1997) suggest that this supra- neural represents a combination of supraneurals 2 and 3. Arratia (1987) argues that in Diplomystes chilensis this supraneural is actually formed from supraneurals 3 and 4, and that in specimen MNHN B. 584. these supraneurals were observed as separate elements (Arratia, 1987; Fig. 9). Our study was not able to shed light on this interpre- tation. Like Rosen and Greenwood's (1970) study of B rye on meeki, we observed only one center of ossification in all relevant specimens of /. punc- tatus examined. At this stage in development (about 1 1 mm TL). all of the vertebrae are autog- enous. In the dorsal fin, seven cartilaginous radials are formed. Although the first two radials will be- come incorporated into the Weberian apparatus/ dorsal fin complex, the dorsal fin is completely separate from the basidorsals at this point. The first and most proximal segment of lepidotrichia has formed in the dorsal fin. Of these dorsal fin rays, the anterior two will develop into fin spines. Catfish spines form first as soft rays in discrete lepidotrichial segments. This is followed by the addition of concentric layers of dermal bone over the surface. The addition of these bony deposits causes the margins of these segments to gradually approach each other, and finally to fuse (Reed. 1924; Fig. 10). In fishes of about 15-18 mm TL. the tripus is ossified and its anterior process has elongated to F.082285). B. Yolk sac larvae. 13.6 mm TL (stage 9- early stage 10). showing the connection between the first two dorsal fin radials and neural arch 4 (LU F.082286). Also note that "anterior radial" (ar) is new terminology to reflect the serial homology of this element with the other dorsal fin radials. C. Foraging juvenile. 16.3 mm TL (stage 13). showing further development and the for- mation of the dorsal fin spines (LU F.082287). D. For- aging juvenile. 38.5 mm TL (no stage), showing a con- nection with the back of the skull via the anterior radial and the Weberian apparatus (LU F.082288). Anterior is to the left. For abbreviations, see p. 4. 22 FIELDIANA: ZOOLOGY Fig. 10. Development of the first and second dorsal fin spines. A, Yolk sac larvae, 1 1 .5 mm TL. B, Yolk sac larvae, 13.0 mm TL. C, Foraging juvenile, 17 mm TL. D, Foraging juvenile, 25.6 mm TL. For abbreviations, see p. 4. the anterior margin of vertebra 1. The transverse process of vertebra 4 is also ossified and continues to expand (Fig. 9B). The os suspensorium has be- gun to form from the ventral side of the transverse process, near its connection of vertebra 4 with vertebra 5, starting with the dorsal lamina (Al- Rawi's terminology, 1966). The dorsal lamina ex- tends anteromesially between the transformator process of the tripus and the postcardinal vein, and terminates in the radial nodule. The radial nodule in the adult catfish is surrounded by the crescent-shaped transformator process of the tri- pus. Both the dorsal lamina and the radial nodule are homologous with the fourth pleural rib and parapophysis of vertebra 4 (Fink & Fink, 1981). Additionally, the intercalarium is completely os- sified, and the scaphium begins to exhibit its char- acteristic shape as its horizontal process extends toward the exoccipitals. The large supraneural (often called the neural complex; Rosen & Green- wood, 1970) has enlarged and, although still car- tilaginous, articulates with the expanded neural arches of vertebrae 3 and 4. Also by this time (stage 11) vertebrae 2-4 are fused, thus complet- ing the formation of the complex centrum. The fusion of vertebrae 2 and 3 occurs in fish aver- aging 13 mm TL (stage 9). Vertebra 1 remains independent and never fuses with the Weberian complex centrum. In fishes of about 15 mm TL, the cartilaginous claustrum is seen for the first time above the sca- phium (Fig. 9B). The homology and derivation of the claustrum have been debated recently. Fink and Fink (1981) hypothesized that the claustrum forms from a dissociated dorsomedial portion of the first neural arch. Coburn and Futey (1996) ar- gued that the claustrum is derived from supra- neural 1. Their hypothesis is based on their ex- amination of several cyprinids (e.g., Luxilus) and catostomids (e.g., Ictiobus), in which supraneural 2 appears to form early in development from paired structures, fusing later in development. They argue that because supraneural 2 forms as a paired element, so might supraneural 1, and if that is true, then it is conceivable that the claustrum forms from supraneural 1 . They also found no ev- idence for a dorsal extension of neural arch 1. Although our observations, based on histology GRANDE & SHARDO: POSTCRANIAL SKELETON IN 1CTALURUS PUNCTATUS 23 and cleared and stained specimens of /. punctatus, cannot support Coburn and Futey's (1996) obser- vations of paired supraneurals, we also cannot support the hypothesis that the claustrum forms from neural arch 1, as does the scaphium. The claustrum in /. punctatus is the last Weberian os- sicle to form. It appears in cartilage after the sca- phium has ossified, and we can see no dorsal ex- tension connecting the scaphium with the claus- trum, or any claustrum precursor, at any time dur- ing development. It is possible that if supraneurals begin development as paired structures, as Coburn and Futey (1996) argue, then they very quickly fuse, resulting in median elements. If this hap- pens, then we may have missed our small window of opportunity to observe this supraneural form- ing from essentially two halves in the specimens observed. Additional ontogenetic material of oth- er siluriforms and closer sampling times may be necessary to resolve this. Continuing with Figure 9B, the early stages in the development of the Weberian apparatus/dorsal fin unit were observed. The first two proximal ra- dials of the dorsal fin lengthen ventrally; the first makes contact with neural arch 4 and is positioned between the two halves of this arch. The first dor- sal fin spine is complete while the second spine continues to ossify along with its corresponding dorsal fin radial. A median cartilaginous element, called a nuchal plate by some authors (e.g., Teu- gels, 1996), a supraneural by others (e.g., Lund- berg, 1982; Lundberg & McDade, 1986; Grande & Lundberg, 1988; Grande & de Pinna, 1998), and a radial or pterygiophore by others (e.g., Brown & Ferraris, 1988; Fink & Fink, 1996), forms anterior to the first dorsal fin radial. This structure, when ossified, articulates with the su- praoccipital bone, forming a connection between the dorsal fin and the skull. The homology of this structure is at times perplexing. Lundberg (1982) and Grande and Lundberg (1988) consider this structure to be a serial homologue of the dorsal fin radials, not a nuchal plate, which is involved in the locking mechanism of the second dorsal fin spine; nor do they consider it to be a supraneural. We agree with their assessment. In /. punctatus this element forms almost directly dorsal to the supraneurals associated with the Weberian appa- ratus and in the same plane as the dorsal fin ra- dials. It therefore does not seem possible for this element also to be a supraneural. Additionally, we agree with Mabee (1988) and Grande and Bemis (1991), who reviewed the homology of radials and supraneurals in centrarchid and polyodontid fishes and concluded that radials are not homol- ogous with supraneurals. Mabee (1988) also showed that the ossification of radials does not always occur in an anterior to posterior direction (e.g., anal fin radials in centrarchids). In /. punc- tatus this anterior element ossifies last. Thus, be- cause of the placement of this element directly dorsal to and in addition to the supraneurals found in /. punctatus, and because this element is in di- rect sequence with the dorsal fin radials, we refer to it as an anterior radial, following Fink and Fink (1996) and reflecting its derivation. As development of Weberian apparatus/dorsal fin complex continues (Fig. 9C), the supraoccip- ital crest, which will articulate with the anterior radial, begins to form posteriorly. The claustrum is at least 50% ossified; the horizontal process of the scaphium is rounded anteriorly and will artic- ulate with the back of the skull via the exoccipi- tals. The first dorsal fin radial is partially ossified and sits between the two halves of neural arch 4. This arch has elongated into a spine that articu- lates with the first dorsal fin spine and the anterior nuchal plate. The second dorsal fin radial is par- tially ossified, and its spine consists of two seg- ments bounded by lepidotrichia. All other dorsal fin radials have elongated but remain cartilagi- nous. As the developing catfish enters the foraging period, the remaining developmental stages are devoted to further ossifying already formed struc- tures (i.e., making the final attachments among the Weberian ossicles, the dorsal fin skeleton, and the skull) and growth. As seen in fishes of 30 mm TL and larger, the transverse process of vertebra 4 develops two expansions. The anterior part of the transverse process is massive and articulates with the ventral process of the mesial limb of the su- pracleithrum (Lundberg, 1975). The posterior part of the transverse process (p4p) is thinner, shorter, and positioned horizontally. The posteromedial margin of p4p forms a crescent-like indentation and articulates with the anterior margin of the transverse process of the fifth vertebra (Figs. 1 1 A Fig. 1 1 . Photographs of skeletons of /. punctatus showing in A the connection between the anterior part of the Weberian apparatus and the back of the skull (FMNH 73900), and in B the fully developed and articulated Weberian 24 FIELDIANA: ZOOLOGY apparatus/dorsal fin unit (FMNH 16711). In B, note the tight connection between neural arch 4 and the first two dorsal fin radials. Anterior is to the left. For abbreviations, see p. 4. GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS 25 and B). The neural arches of the compound cen- trum enlarge and fuse with the anterior supra- neural. As shown in Figure 9D, complex ossifications are associated with the dorsal fin. The anterior and posterior nuchal plates have ossified in fishes of more than 35 mm TL. The anterior radial has os- sified fully and has made contact with both the anterior nuchal plate and the supraoccipital. Os- sification of the first proximal and distal radials is complete, and the first proximal radial forms a tight connection with the posterior margin of the neural arch complex of the Weberian apparatus. The first dorsal fin spine helps form a locking mechanism for the second fin spine. The second proximal radial is ossified, and its distal tip artic- ulates with the Weberian apparatus. Eventually the entire second radial will abut the first, which in turn is connected with the Weberian apparatus. In fishes of about 40 mm TL, proximal radials 3— 6 are ossified (proximal radial 7 and distal radials 3-7 are still cartilaginous). They have elongated and extend between the two halves of neural arch- es 5-9, preventing them from meeting at the dor- sal midline. Instead of the typical midline closure of the arches, as seen with the more posterior ones, an ossified horizontal rod connects the two halves of each arch at least one-third the length of the neural arch. The distal tip of each radial is positioned between the two halves of each arch but above the rod connecting the two halves. In a fully ossified Weberian apparatus/dorsal fin complex (Figs. 12A and B) a solid bony connec- tion is made between the Weberian apparatus and the dorsal fin skeleton via the spine of neural arch 4 and the anterior dorsal fin radials. It is interest- ing that the anterior radial articulates with the su- praoccipital, forming a connection with the skull (not shown in Figs. 12A and B). When fully os- sified, the Weberian apparatus/dorsal fin complex forms at least three connections with the back of the skull. The first one, as just stated, is the artic- ulation of the anterior radial and the supraoccipital crest; the second is by means of an articulation between anterior neural arch 3 and the ventral side of the supraoccipital (Fig. 9D); and the third is via the Weberian ossicles (i.e., scaphium, claus- trum, and anterior process of the tripus) with the exoccipitals. The functional significance of a connection be- tween the Weberian apparatus and the dorsal fin is not certain. Reed (1924) argued that since cat- fish spines remain in the structural and functional state of a soft ray far into the foraging stage, the Fig. 12. A. Anterior vertebral column of the specimen figured in Figure 9D, 38.5 mm TL, in dorsal view. B. Illustration of specimen figured in Figure 9D in ventral view. Anterior is directed upward. For abbreviations, see p. 4. 26 FIELDIANA: ZOOLOGY dorsal fin can scarcely be considered a weapon or an element of active defense. Alexander (1965) argued that the principal function of catfish spines is protection, and that the locking mechanism of the dorsal fin spine must be firmly mounted and reinforced by a strong skeleton in order for it to be effective. Essentially, the added skeletal sup- port of the ossified anterior radial and the attach- ment of the dorsal fin to the skull and Weberian apparatus are aids to stabilize the locking of the second dorsal fin spine. Alexander (1965) and Ad- riaens (1998) suggested, however, that the locking mechanism of the spine is a pre-adaptation for sound production. Although stridulation by means of pectoral fins has been demonstrated in /. punc- tatus (Fine et al., 1996), the dorsal fin of/, punc- tatus does not possess the necessary opposable serrated pterygiophores commonly associated with sound production. We thus do not consider sound production a possible function for this unit. We do, however, entertain the possibility of sound perception or transmission as a viable, although admittedly inefficient, function for the Weberian apparatus/dorsal fin attachment. The Weberian ap- paratus transmits high-frequency vibrations from the gasbladder to the inner ear. The possibility of transmitting low-frequency vibrations from the water to the inner ear via this unit is theoretically plausible and intriguing. Physiological experi- ments are necessary to explore this hypothesis. Variation in Development The availability of such a large developmental sample size has afforded us the rare opportunity to examine individual variation within and be- tween series. The overall pattern of development shown by the 18 stages is very similar for all three series. The rate of development is faster in the earlier stages and then slows in the later stages, about stage 12 for series C and stage 13 for series A and B. The pattern, of course, depends on the stage criteria, which were determined by our se- lection, but nonetheless accurately reflects the rapid changes occurring in early development of the axial skeleton relative to later development. The second point is that although the three se- ries follow the same development in sequence as represented by the 18 stages, there is a distinct difference in the age profile of series C compared with series A and B. At every stage up to 16, specimens of series C are older than specimens of series A and B. In contrast, the ages of specimens of A and B are similar at each stage. This age differential appears to result from the develop- ment of the axial skeleton beginning (stage 2) at an earlier age in series A and B than in series C (see Table 1). Although the magnitude of the dif- ference in ages at each consecutive stage fluctu- ates somewhat, the differential is maintained until stage 16 (from stage 16 to stage 18, age data are not available for all three series). The most likely explanation for this age difference would be in- cubation and rearing conditions, particularly tem- perature. However, all series were reared under approximately the same conditions, with temper- ature about 26°C. Series A was reared in a green- house in Chicago, series B at a private hatchery in Missouri, and series C in a Department of Ag- riculture research unit in Mississippi. The poten- tial for uncontrolled environmental effects is high but cannot account for both the similarity in age profiles between series A and B and the dissimi- larity in profiles between series C and series A/B. The origins of the three series offer another pos- sibility. Both series A and B were spawned at a hatchery in Missouri, although several years apart. The hatchery series and the research unit series are possibly separate populations with small inherent differences in the timing of developmen- tal events. Size as measured by total length also varies among the three series. Again, the differences in size are greater between series C and series A/B than between series A and series B, but the mag- nitude of the differences is not large, and fre- quently the size ranges for the three series overlap (see Table 1). In general, series C tends to be slightly larger at each stage than series A or B, possibly the result of being slightly older. We defined concurrent features as additional variable characters that occur during a stage but that are not restricted to appearing only in that stage in all specimens. Clearly, age and growth in length vary in timing in relation to specific mor- phological developmental events (stage criteria) and can be classified as concurrent features. In this study, however, size is a slightly better pre- dictor of morphological development than age. We found a number of other concurrent features that also exhibit individual variation within and between series. Among sensory structures, the pairs of barbels appear in earlier stages in series C than in series A and B, but the appearance of the lateral line marking the beginning of the fron- tal bone occurs one stage later (stage 6) in series GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS 27 C than in series A and B. Among other skull el- ements, the dentary bone forms one stage later (stage 4) and the hyomandibula begins to ossify almost seven stages earlier (stage 5) in series C than in series A and B. In the paired fins, the pectoral girdle and fin are completely ossified at an earlier stage in series C (stage 1 1 ) than in se- ries A and B (stage 15). Of the medial fins, the distal radials of the anal fin form at an earlier stage in series C (stage 7) than in series A and B (stage 9). There are several variations in the cau- dal fin. The parhypural and hypurals 1-5 begin to ossify (stage 6 versus stage 11) and uroneural 1 forms (stage 5 versus stage 8) at earlier stages in series C than in series A and B. In respect to five structures — the hyomandibu- la, pectoral girdle/fin, anal fin radials, hypural/par- hypural. and uroneural 1 — the larvae in series C were smaller than their counterparts in series A and B. In respect to only two structures, the hy- omandibula and pectoral girdle/fin, were the lar- vae of series C younger than their counterparts in series A and B when the concurrent features oc- curred. There is no obvious pattern to the varia- tion; it does not seem to track age, size, or mor- phological development as defined by stage cri- teria. Only in the caudal fin does the variation occur in a consistent manner, which might provide a slightly earlier advantage in escape behavior for yolk sac larvae in series A and B. But much of this variation is probably the result of different structures, or of systems having different devel- opmental rates that are responsive to a variety of factors. Acknowledgments Special thanks to William E. Bemis for provid- ing developmental series B used in this study. Thanks to Joseph Schluep for his photographic expertise, Warren Jones for his help with the his- tology, and Chris Serpico and Andrew Woodard for helping to collect and raise many of the fishes in this study. Mary Anne Rogers and Mark West- neat of the Field Museum made available that in- stitution's catfish material on loan. Many thanks to Gloria Arratia, Lance Grande. Mario de Pinna, and an unnamed reviewer for their comments and suggestions during the preparation of this manu- script. Support for this project came from the Na- tional Science Foundation (grant No. DEB 9707530) to Judith D. Shardo, the Totogany Creek Fund for Research of Vertebrate Morphology to Terry Grande, and the National Science Founda- tion (grant No. DEB 9903533) to Terry Grande. Literature Cited Adriaens. D. 1998. On how a larva becomes an adult catfish: A functional morphological approach to the cranial ontogeny of the African catfish. Clarius gar- iepinus (Siluriformes. Clariidae). Ph.D. diss.. Univer- sity of Gent, Belgium. Alexander. R. M. 1964. The structure of the Weberian apparatus in the Siluri. Proceedings of the Zoological Society of London. 142: 419-440. . 1965. Structure and function in the catfish. Journal of Zoology. 148: 88-152. Al-Rawi. A. H. A. 1966. The development of the We- berian apparatus and swim bladder in the channel cat- fish, Ictalurus punctatus (Rafinesque). Ph.D. diss.. University of Oklahoma. Norman. Armstrong. P. B. 1962. Stages in the Development of Ictalurus nebulosus. Syracuse University Press. Syra- cuse. New York. Arratia. G. 1983. The caudal skeleton of ostariophysan fishes (Teleostei): Intraspecific variation in Tricho- mycteridae (Siluriformes). Journal of Morphology. 177: 213-229. . 1987. Description of the primitive family Dip- lomystidae (Siluriformes. Teleostei. Pisces): Morphol- ogy, taxonomy and phylogenetic implications. Bonner Zoologische Monographien. 24: 1-123. 1990. Development and diversity of the sus- pensorium of trichomycterids and comparison with loricarioids (Teleostei: Siluriformes). Journal of Mor- phology. 205: 193-218. . 1991. The caudal skeleton of Jurassic teleosts: A phylogenetic analysis, pp. 249-340. In Chang. M.- M.. Y. H. Liu. and G. R. Zang. eds.. Early Vertebrates and Related Problems in Evolutionary Biology. Sci- ence Press. Beijing. 1992. Development and variation of the sus- pensorium of primitive catfishes (Teleostei: Ostario- physi) and their phylogenetic relationships. Bonner Zoologische Monographien. 32: 1-149. Arratia. G.. and H.-P. Schultze. 1992. Reevaluation of the caudal skeleton of certain actinopterygian fish- es: III. Salmonidae. Homologization of caudal skeletal structures. Journal of Morphology. 214: 187-249. Bamford, T. W. 1 948. Cranial development of Galeich- th\s felis. Proceedings of the Zoological Society of London. 118: 364-391. Bemis. W. E.. and L. Grande. 1992. Early development of the actinopterygian head. I. External development and staging of the paddlefish Pohodon spathula. Jour- nal of Morphology. 213: 47-83.' . 1999. Development of the median fins on the North American paddlefish {Pohodon spathula), and a reevaluation of the lateral fin-fold hypothesis, pp. 41-68. /// Arratia. G.. and H.-P. Schultze. eds.. Sys- tematics and Fossil Record. Verlag Dr. Pfeil. Munich. 28 FIELDIANA: ZOOLOGY Brown, B. A., and C. J. Ferraris. 1988. Comparative osteology of the Asian catfish family Chacidae, with the description of a new species from Burma. Amer- ican Museum Novitates, 2907: 1-16. Chardon, M. 1968. Anatomis comparee de l'appareil de Weber et des structures connexes chez les Silurifor- mes. Annales du Musee Royale de l'Afrique Centrale, serie 8, 169: 1-277. Chardon, M., and P. Vandewalle. 1997. Evolutionary trends and possible origin of the Weberian apparatus. Netherlands Journal of Zoology, 47: 383-403. Coburn, M. M., and L. M. Futey. 1996. The ontogeny of supraneurals and neural arches in the cypriniform Weberian apparatus (Teleostei: Ostariophysi). Zoolog- ical Journal of the Linnean Society, 116: 333-346. Coburn, M. M., and P. G. Grubach. 1998. Ontogeny of the Weberian apparatus in the armored catfish Co- rydoras paleatus (Siluriformes: Callichthyidae). Cop- eia, 1998: 301-311. COLLETTE, B. B., T. POTTHOFF, W. J. RICHARDS, S. UeY- angi, J. L. Russo, AND Y. Nishikawa. 1983. Pp. 591— 620. In Moser, H. G., W J. Richards, D. M. Cohen, M. P. Fahay, A. W Kendall, and S. L. Richardson, eds., Ontogeny and Systematics of Fishes. American Society of Ichthyologists Special Publication 1. de Pinna, M. C. C. 1996. A phylogenetic analysis of the Asian catfish families Sisoridae, Akysidae, and Am- blycipitidae, with a hypothesis on the relationships of Neotropical Aspredinidae (Teleostei, Ostariophysi). Fieldiana, Zoology, n.s., 84: 1-82. Dingerkus, G., and L. D. Uhler. 1977. Enzyme clear- ing of alcian blue stained whole vertebrates for dem- onstration of cartilage. Journal of Stain Technology, 52: 299-232. Dunn, J. R. 1983. Developmental osteology, pp. 48-50. In Moser, H. G., W J. Richards, D. M. Cohen, M. P. Fahay, A. W Kendall, and S. L. Richardson, eds., On- togeny and Systematics of Fishes. American Society of Ichthyologists Special Publication 1 . Eaton, T. H. 1937. Form and function in the head of the channel catfish, Ictalurus lacustris punctatus. Jour- nal of Morphology, 83: 181-194. Faustino, M., and D. M. Power. 1998. Development of osteological structures in the sea bream: Vertebral col- umn and caudal fin complex. Journal of Fish Biology, 52: 11-22. Fine, M. L., D. McElroy, J. Rafi, C. B. King, K. E. Loesser, and S. Newton. 1996. Lateralization of pec- toral stridulation sound production in the channel cat- fish. Physiological Behavior, 60: 753-757. Fink, S. V., and W L. Fink. 1981. Interrelationships of ostariophysan fishes (Teleostei). Zoological Journal of the Linnean Society, 72: 297-353. . 1996. Interrelationships of ostariophysan fishes (Teleostei), pp. 209-249. In Stiassny, M., L. R. Par- enti, and G. D. Johnson, eds., Interrelationships of Fishes. Academic Press, San Diego. Fowler, J. A. 1970. Control of vertebral number in tel- eosts an embryological problem. Quarterly Review of Biology, 45: 148-167. Francois, Y. 1966. Structure et developpement de la vertebre de Salmo et des Teleosteens. Archives de Zoologie Experimental et Generate, 107: 283-325. Fuiman, L.A. 1983. Ostariophysi: Development and re- lationships, pp. 126-137. In Moser, H. G., W. J. Rich- ards, D. M. Cohen, M. P. Fahay, A. W Kendall, and S. L. Richardson, eds., Ontogeny and Systematics of Fishes. American Society of Ichthyologists Special Publication 1. Gayet, M. 1982. Consideration sur la phylogenie et la paleobiogeographie des Ostariophysaires. Geobios, 6: 39-52. 1986. Probleme de l'origine des osselets de We- ber. Oceanis, 12: 357-366. Gayet, M., and M. Chardon. 1987. Possible otophysic connections in some fossil and living ostariophysan fishes. Proceedings of the European Ichthyological Congress of Stockholm, 1985: 31-42. Goodrich, E. S. 1930. Studies on the Structure and De- velopment of Vertebrates. Macmillan Co., London. Grande, L. 1987. Redescription of fHypsidoris farso- nensis (Teleostei: Siluriformes) with a reassessment of its phylogenetic relationships. Journal of Vertebrate Paleontology, 7: 24-54. Grande, L., and W E. Bemis. 1991. Osteology and phy- logenetic relationships of fossil and Recent paddle- fishes (Polyodontidae) with comments on the interre- lationships of Acipenseriformes. Journal of Vertebrate Paleontology, 11 (suppl. to vol. 1): 1-121. . 1998. A comprehensive phylogenetic study of amiid fishes (Amiidae) based on comparative skeletal anatomy: An empirical search for interconnected pat- terns of natural history. Society of Vertebrate Pale- ontology Memoir, 4, 690 pp.; supplement to Journal of Vertebrate Paleontology, 18. Grande, L., and M. C. C. de Pinna. 1998. Description of a second species of the catfish ~\Hypsidoris and a reevaluation of the genus and family f Hypsidoridae. Journal of Vertebrate Paleontology, 18: 451-474. Grande, L., and J. G. Lundberg. 1988. Revision and redescription of the genus Astephus (Siluriformes: Ic- taluridae) with a discussion of its phylogenetic rela- tionships. Journal of Vertebrate Paleontology, 8: 139— 171. Grande, T, and F. J. Poyato-Ariza. 1999. Phylogenetic relationships of fossil and Recent gonorynchiform fishes (Teleostei: Ostariophysi). Zoological Journal of the Linnean Society, 125: 197-238. Greenwood, H. P. 1966. The caudal fin skeleton in os- teoglossoid fishes. Annals and Magazine of Natural History, London, series 13, 9: 581-597. Grizzle, J. M., and W A. Rogers. 1985. Anatomy and Histology of the Channel Catfish, 3rd ed. Craftmaster Printers, Inc., Opelika, Alabama. Humason, G. L. 1972. Animal Tissue Techniques. W. H. Freeman, San Francisco. Kimmel, C. B., W W Ballard, S. R. Kimmel, B. Ull- man, and T F. Schilling. 1995. Stages of embryonic development of the zebrafish. Developmental Dynam- ics, 203: 253-310. Kindred, J. 1919. The skull of Amieurus. Illinois Bio- logical Monographs, 5: 1-120. Kobayakawa, M. 1992. Comparative morphology and development of bony elements in the head region in three species of Japanese catfishes (Silurus: Siluridae: GRANDE & SHARDO: POSTCRANIAL SKELETON IN ICTALURUS PUNCTATUS Siluriformes). Japanese Journal of Ichthyology, 39: 25-36. Krumholz, L. A. 1943. A comparative study of the We- berian ossicles in North American ostariophysine fish- es. Copeia, 1943: 33-40. Kuwada, J. Y., R. R. Bernhardt, and A. B. Chitins. 1990. Pathfinding by identified growth cones in the spinal cord of zebrafish embryos. Journal of Neuro- science Research, 10: 1299-1308. Laerm, J. 1976. The development, function, and design of amphicoelus vertebrae in teleost fishes. Zoological Journal of the Linnean Society, 58: 237-254. . 1982. The origin and homology of the neopter- ygian vertebral centrum. Journal of Paleontology, 56: 191-202. Lal Hora, S. 1922. The homology of the Weberian os- sicles. Journal of the Asiatic Society of Bengal, 19: 1-4. Lauder, G. 1989. Caudal fin locomotion in ray-finned fishes: Historical analyses. American Zoologist, 29: 85-102. Lundberg, J. G. 1975. Homologies of the upper shoul- der girdle and temporal region bones in catfishes (Or- der Siluriformes), with comments on the skull of Hel- ogeneidae. Copeia, 1975: 66-74. . 1982. The comparative anatomy of the toothless blindcat, Trogloglanis pattersoni Eigenmann, with a phylogenetic analysis of the ictalurid catfishes. Mis- cellaneous Publications of the Museum of Zoology, University of Michigan, 163: 1-85. Lundberg, J. G., and J. N. Baskin. 1969. The caudal skeleton of the catfishes, order Siluriformes. American Museum Novitates, 2398: 1-49. Lundberg, J. G., and L. A. McCade. 1986. On the South American catfish Brachyrhamdia imitator My- ers (Siluriformes, Pimelodidae), with phylogenetic ev- idence for a large intrafamilial lineage. Notulae Na- turae, 463:1-24. Mabee, P. M. 1988. Supraneural and predorsal bones in fishes: Development and homologies. Copeia, 1988: 827-838. Martin, F. D. 1983. Esocoidei: Development and rela- tionships, pp. 140-142. In Moser, H. G., W. J. Rich- ards, D. M. Cohen, M. P. Fahay, A. W. Kendall, and S. L. Richardson, eds., Ontogeny and Systematics of Fishes. American Society of Ichthyologists Special Publication 1. Martin, R. L. 1963. A possible evolutionary pathway for the development of the Weberian ossicles. The Bi- ologist, 45: 41-54. Matveive, B. 1929. Die Entwicklung der vorderen Wir- bel und des weberschen Apparates bei Cyprinidae. Zoologischer Jahrbucher, 51: 463-534. Monod, T. 1968. Le complexe urophore des poissons teleosteens. Memoires de l'lnstitut Francais d'Afrique Noire, 81: 1-705. Nelson, J. S. 1994. Fishes of the World, 3rd ed. John Wiley & Sons, New York. Patterson, C. 1968. The caudal skeleton in Lower Li- assic pholidophorid fishes. Bulletin of the British Mu- seum (Natural History), Geology, 16: 210-239. Radermaker, F, C. Surlemont, P. Sanna, M. Chardon, and P. Vandewalle. 1989. Ontogeny of the Weberian apparatus of Clarias gariepinus (Pisces Siluriformes). Canadian Journal of Zoology, 67: 2090-2097. Reed, H. D. 1924. The morphology and growth of the spines of siluroid fishes. Journal of Morphology, 38: 431-451. Reimchen, T E., and J. S. Nelson. 1987. Habitat and morphological correlates to vertebral number as shown in a teleost Gasterosteus aculeatus. Copeia, 1987: 868-874. Rosen, D. E., and P. H. Greenwood. 1970. Origin of the Weberian apparatus and the relationships of the ostariophysan and gonorynchiform fishes. American Museum Novitates, 2428: 1-25. Schultze, H.-P, and G. Arratia. 1986. Reevaluation of the caudal skeleton of actinopterygian fishes: I. Lepisosteus and Amia. Journal of Morphology, 190: 215-241. . 1988. Reevaluation of the caudal fin skeleton of some actinopterygian fishes. II. Hiodon, Elops, and Albula. Journal of Morphology, 195: 257-303. 1989. The composition of the caudal skeleton of teleosts (Acinopterygii: Osteichthyes). Zoological Journal of the Linnean Society, 97: 189-231. Shardo, J. D. 1995. Comparative embryology of tele- ostean fishes. I. Development and staging of the American shad, Alosa sapidissima (Wilson, 1811). Journal of Morphology, 225: 125-167. Teugels, G. G. 1996. Taxonomy, phylogeny and bio- geography of catfishes (Ostariophysi, Siluridei): an overview. Aquatic Living Resources, 9: 9-34. Thomas, K. 1983. A nitrocellulose embedding technique for vertebrate morphologists. Herpetology Review, 14: 80-81. Vandewalle, P., F Radermaker, C. Surlemont, and M. Chardon. 1990. Apparition of the Weberian char- acters in Barbus barbus (Linne, 1758) (Pisces Cypri- nidae) Zoologica Anzeiger, 225: 262-376. Watson, J. M. 1939. The development of the Weberian ossicles and anterior vertebrae in the goldfish. Pro- ceedings of the Royal Society of London, series B, 127: 452-472. Young, B., F Sheft, and B. Yost. 1995. Sound pro- duction in Pituophis melanoleucus (Serpentes: Colu- bridae) with the first description of a vocal cord in snakes. Journal of Experimental Zoology, 273: 472- 481. 30 FIELDIANA: ZOOLOGY '..!'•? LT i' Ji in NeotropjpS) Man mail in il son and Robe t M. Timm, Fieldiana , .'- itic Re\ ew of Philippine Macaques (Prima • ' :reopithe By Jack Foocien. Fiet ; n.s.. no 64 1991 \ Kej to the Bats of the Philippine Islands i ■-. Nina R Zooh. no. 69. 1992. 44 pages. 60 ill is., ^ tanies. PiihJifatfsori \ IHihfcaiH Systematic Reviev o Southeast \sian Longtail M caques Macaco, fas ■ uhu H . Jack Foodc i Fieldian Zoology, n.s., no. 81. 1995 106 \ iges. 31, illus., : table: Hie Birds of Sibuyan Island Romblon Province. Philippines with Particulai Referent Util ;: Distribution and Biogeographic A ! i ; o By Stephen M Goodman Davi< Willai Gonzales. Fieldiana: Z/!- . n.s., no. 8 2 1995. ~-": pages, 12 illus . :--<:- Publication 147 I . Frogs of Vietnam: A Report on New Co1 led ions, 13} Robert F. Ingei el al. Fieldi m ■' 'log . n.s 92. 1999. 46 page . 18 illus . I 1 tables. Pnhlk-alioi! '< iS9. , 0 0 Systematic Review o1 the Rhesus Macaque. Ma aca mulatto fZti net- ian. ; SO) By la m . Fieldiana: Zoology, n.s . no. 96. 2000. 180 pages. 22 illus . ! I ables Publication 1509. $65 ■\ Floral and Fauna! Inventory of the Part National de Marojejy. Vladag; .car VVi \ ! ! : :nce \ Elevational Variation. I dited b) Stephen M. Goodman. Fieldiana: Zopiog} pages, with illus and tables. }<.- irdei / ieldiana. please address correspo d Peter I ortsas : ortsas Book- , Ltd. 5 \:- ■■ '• forth I i\-i ,'V Chicago, 11. 60630 ! i . \iiana(a;aol. UMVERHTY OF ILLiNOlS-UKBANA