A three‐dimensional placoderm (stem‐group gnathostome) pharyngeal skeleton and its implications for primitive gnathostome pharyngeal architecture

The pharyngeal skeleton is a key vertebrate anatomical system in debates on the origin of jaws and gnathostome (jawed vertebrate) feeding. Furthermore, it offers considerable potential as a source of phylogenetic data. Well‐preserved examples of pharyngeal skeletons from stem‐group gnathostomes remain poorly known. Here, we describe an articulated, nearly complete pharyngeal skeleton in an Early Devonian placoderm fish, Paraplesiobatis heinrichsi Broili, from Hunsrück Slate of Germany. Using synchrotron light tomography, we resolve and reconstruct the three‐dimensional gill arch architecture of Paraplesiobatis and compare it with other gnathostomes. The preserved pharyngeal skeleton comprises elements of the hyoid arch (probable ceratohyal) and a series of branchial arches. Limited resolution in the tomography scan causes some uncertainty in interpreting the exact number of arches preserved. However, at least four branchial arches are present. The final and penultimate arches are connected as in osteichthyans. A single median basihyal is present as in chondrichthyans. No dorsal (epibranchial or pharyngobranchial) elements are observed. The structure of the pharyngeal skeleton of Paraplesiobatis agrees well with Pseudopetalichthys from the same deposit, allowing an alternative interpretation of the latter taxon. The phylogenetic significance of Paraplesiobatis is considered. A median basihyal is likely an ancestral gnathostome character, probably with some connection to both the hyoid and the first branchial arch pair. Unpaired basibranchial bones may be independently derived in chondrichthyans and osteichthyans.     

Development of the skull and pectoral girdle in Siberian sturgeon,Acipenser baerii,and Russian sturgeon,Acipenser gueldenstaedtii (Acipenseriformes: Acipenseridae)

The head is considered the major novelty of the vertebrates and directly linked to their evolutionary success. Its form and development as well as its function, for example in feeding, is of major interest for evolutionary biologists. In this study, we describe the skeletal development of the cranium and pectoral girdle in Siberian (Acipenser baerii ) and Russian sturgeon (A. gueldenstaedtii ), two species that are commonly farmed in aquaculture and increasingly important in developmental studies. This study comprises the development of the neuro‐, viscero‐ and dermatocranium and the dermal and chondral components of the pectoral girdle, from first condensation of chondrocytes in prehatchlings to the early juvenile stage and reveals a clear pattern in formation. The otic capsules, the parachordal cartilages, and the trabeculae cranii are the first centers of chondrification, at 8.4mm TL. These are followed by the mandibular, then the hyoid, and later the branchial arches. Teeth form early on the dentary, dermopalatine, and palatopterygoid, and then appear later in the buccal cavity as dorsal and ventral toothplates. With ongoing chondrification in the neurocranium a capsule around the brain and a strong rostrum are formed. Dermal ossifications start to form before closure of the dorsal neurocranial fenestrae. Perichondral ossification of cartilage bones occurs much later in ontogeny. Our results contribute data bearing on the homology of elements such as the lateral rostral canal bone that we regard homologous to the antorbital of other actinopterygians based on its sequence of formation, position and form. We further raise doubts on the homology of the posterior ceratobranchial among Actinopteri based on the formation of the hyoid arch elements. We also investigate the basibranchials and the closely associated unidentified gill‐arch elements and show that they are not homologous. J. Morphol. 278:418–442, 2017. © 2017 Wiley Periodicals, Inc.     

Functional ecology of feeding in elasmobranchs

Feeding behavior in the species of captive chondrichthyans is studied to clarify the functional mechanisms responsible for feeding ecology. Kinematics and pressure in the buccal, hyoid and pharyngeal regions were quantified in Squalus acanthias, Chiloscyllium plagiosum and Leucoraja erinacea using sonomicrometry and pressure transducers. Means and coefficients of variation were analyzed by species and by behavior to test for stereotypy and flexibility in the feeding mechanism. Several instances of mechanical stereotypy as well as flexibility were found in the feeding kinematics and pressure of the three chondrichthyan species. In general, Squalus acanthias shows more stereotyped feeding behavior than C. plagiosum and L. erinacea. Different aspects of feeding behavior stand out among the three species. Chiloscyllium plagiosum generates lowest pressures, S. acanthias achieves the greatest area changes, and L. erinacea has longer durations for manipulating prey. Capture events are functionally and behaviorally stereotyped while processing events are functionally and behaviorally flexible with the ability to use suction or compression to process the same food item. Squalus acanthias is a functional specialist and C. plagiosum is functionally a generalist, with both species exhibiting behavioral flexibility. Leucoraja erinacea is a functional and behavioral generalist. Using functional morphology to explain mechanical stereotypy and flexibility in the feeding behavior of three suction feeding chondrichthyan species has allowed a better understanding of specialist and generalist trophic behaviors.     

The suction mechanism of the pipid frog,Pipa pipa (Linnaeus, 1758)

Most suction‐feeding, aquatic vertebrates create suction by rapidly enlarging the oral cavity and pharynx. Forceful enlargement of the pharynx is powered by longitudinal muscles that retract skeletal elements of the hyoid, more caudal branchial arches, and, in many fish, the pectoral girdle. This arrangement was thought to characterize all suction‐feeding vertebrates. However, it does not exist in the permanently aquatic, tongueless Pipa pipa , an Amazonian frog that can catch fish. Correlating high‐speed (250 and 500 fps) video records with anatomical analysis and functional tests shows that fundamental features of tetrapod body design are altered to allow P. pipa to suction‐feed. In P. pipa , the hyoid apparatus is not connected to the skull and is enclosed by the pectoral girdle. The major retractor of the hyoid apparatus arises not from the pectoral girdle but from the femur, which lies largely within the soft tissue boundaries of the trunk. Retraction of the hyoid is coupled with expansion of the anterior trunk, which occurs when the hypertrophied ventral pectoral elements are depressed and the urostyle and sacral vertebra are protracted and slide forward on the pelvic girdle, thereby elongating the entire trunk. We suggest that a single, robust pair of muscles adduct the cleithra to depress the ventral pectoral elements with force, while modified tail muscles slide the axial skeleton cranially on the pelvic girdle. Combined hyoid retraction, axial protraction, and pectoral depression expand the buccopharyngeal cavity to a volume potentially equal to that of the entire resting body of the frog. Pipa may be the only tetrapod vertebrate clade that enlarges its entire trunk during suction‐feeding.     

The fate of cranial neural crest cells in the Australian lungfish, Neoceratodus forsteri

The cranial neural crest has been shown to give rise to a diversity of cells and tissues, including cartilage, bone and connective tissue, in a variety of tetrapods and in the zebrafish. It has been claimed, however, that in the Australian lungfish these tissues are not derived from the cranial neural crest, and even that no migrating cranial neural crest cells exist in this species. We have earlier documented that cranial neural crest cells do migrate, although they emerge late, in the Australian lungfish. Here, we have used the lipophilic fluorescent dye, DiI, to label premigratory cranial neural crest cells and follow their fate until stage 43, when several cranial skeletal elements have started to differentiate. The timing and extent of their migration was investigated, and formation of mandibular, hyoid and branchial streams documented. Cranial neural crest was shown to contribute cells to several parts of the head skeleton, including the trabecula cranii and derivatives of the mandibular arch (e.g., Meckel's cartilage, quadrate), the hyoid arch (e.g., the ceratohyal) and the branchial arches (ceratobranchials I-IV), as well as to the connective tissue surrounding the myofibers in cranial muscles. We conclude that cranial neural crest migration and fate in the Australian lungfish follow the stereotyped pattern documented in other vertebrates.     

Temporal requirement of Hoxa2 in cranial neural crest skeletal morphogenesis

Little is known about the spatiotemporal requirement of Hox gene patterning activity in vertebrates. In Hoxa2 mouse mutants, the hyoid skeleton is replaced by a duplicated set of mandibular and middle ear structures. Here, we show that Hoxa2 is selectively required in cranial neural crest cells (NCCs). Moreover, we used a Cre-ERT2 recombinase system to induce a temporally controlled Hoxa2 deletion in the mouse. Hoxa2 inactivation after cranial NCC migration into branchial arches resulted in homeotic transformation of hyoid into mandibular arch skeletal derivatives, reproducing the conventional Hoxa2 knockout phenotype, and induced rapid changes in Alx4, Bapx1, Six2 and Msx1 expression patterns. Thus, hyoid NCCs retain a remarkable degree of plasticity even after their migration in the arch, and require Hoxa2 as an integral component of their morphogenetic program. Moreover, subpopulations of postmigratory NCCs required Hoxa2 at discrete time points to pattern distinct derivatives. This study provides the first temporal inactivation of a vertebrate Hox gene and illustrates Hox requirement during late morphogenetic processes.     

Development of the muscles associated with the mandibular and hyoid arches in the Siberian sturgeon,Acipenser baerii (Acipenseriformes: Acipenseridae)

The skeleton of the jaws and neurocranium of sturgeons (Acipenseridae) are connected only through the hyoid arch. This arrangement allows considerable protrusion and retraction of the jaws and is highly specialized among ray‐finned fishes (Actinopterygii). To better understand the unique morphology and the evolution of the jaw apparatus in Acipenseridae, we investigated the development of the muscles of the mandibular and hyoid arches of the Siberian sturgeon, Acipenser baerii. We used a combination of antibody staining and formalin‐induced fluorescence of tissues imaged with confocal microscopy and subsequent three‐dimensional reconstruction. These data were analyzed to address the identity of previously controversial and newly discovered muscle portions. Our results indicate that the anlagen of the muscles in A. baerii develop similarly to those of other actinopterygians, although they differ by not differentiating into distinct muscles. This is exemplified by the subpartitioning of the m. adductor mandibulae as well as the massive m. protractor hyomandibulae, for which we found a previously undescribed portion in each. The importance of paedomorphosis for the evolution of Acipenseriformes has been discussed before and our results indicate that the muscles of the mandibular and the hyoid may be another example for heterochronic evolution.     

A novel pharyngeal expansion mechanism in the yellow-spotted fanray, Platyrhina tangi (Elasmobranchii: Batoidea), with special reference to the function of the fifth ceratobranchial cartilage in batoids

Expansion of the ‘pharynx’ during breathing or capturing prey in fishes generally involves posteroventral retraction of the hyoid arch. However, the hyoid arch structure of batoid fishes (skates, rays, guitarfishes, and sawfishes) is unique, and how they expand the pharyngeal cavity is poorly understood. To investigate the mechanism of pharyngeal expansion during breathing in the yellow-spotted fanray, Platyrhina tangi, we conducted anatomical and kinematic investigations of the pharyngeal region. Our study revealed that the yellow-spotted fanray and sharks have different skeletal linkage systems for pharyngeal expansion. During pharyngeal expansion in the yellow-spotted fanray, the hyoid bar and branchial apparatus rotate ventrally around the hinge joint between the fifth ceratobranchial cartilage and the pectoral girdle. This pharyngeal expansion mechanism appears to be widespread among batoid fishes and is unique among cartilaginous fishes (sharks, batoids, and holocephalans). Batoid fishes possibly developed this pharyngeal expansion mechanism during early batoid evolution.     

Development of the mandibular, hyoid arch and gill arch skeleton in the Chinese barb Puntius semifasciolatus: comparisons of ossification sequences among Cypriniformes

The morphogenesis and sequence of ossification and chondrification of skeletal elements of the jaws, and hyoid arch and gill arches of Puntius semifasciolatus are described. These data provide a baseline for further studies and enable comparisons with other described cypriniforms. Some general patterns of ossification in the hyoid arch and branchial arches in cypriniforms were notable. First, the overall development is from anterior to posterior, with the exception of the fifth ceratobranchial bone, which ossifies first. Second, where ossification of iterated elements is sequential, it tends to proceed from posterior to anterior, even when more posterior chondrifications are the smallest in the series. Ossification of the ceratobranchial, epibranchial and pharyngobranchial bones tends to proceed from ventral to dorsal. The comparisons revealed small sets of skeletal elements whose ossification sequence appears to be relatively conserved across cyprinid cypriniforms. Several potentially key timing changes in the ossification sequence of the jaws, hyoid arch and gill arches were identified, such as the accelerated timing of ossification of the fifth ceratobranchial bone, which may be unique to cypriniforms.     

Regeneration of amphioxus oral cirri and its skeletal rods: implications for the origin of the vertebrate skeleton

The oral cirri of amphioxus function as the first filter during feeding by eliminating unwanted large or noxious particulates. In this study, we were able to regenerate cirri following artificial amputation. This is the first firm observation of regeneration in amphioxus. Using this regeneration system, we studied skeletogenesis of the cellular skeleton of amphioxus oral cirri. During regeneration, the skeletal cells showed expression of fibrillar collagen and SoxE genes. These observations suggest that an evolutionarily conserved genetic regulatory system is involved in amphioxus cirrus and vertebrate cartilage skeletogenesis. In addition, Runx and SPARC/osteonectin expression were observed in regenerating cirral skeletal cells, indicating that cirral skeletogenesis is similar to vertebrate osteogenesis. We propose that the common ancestors of chordates possessed a genetic regulatory system that was the prototype of chondrogenesis and osteogenesis in vertebrates. Genome duplications caused divergence of this genetic regulatory system resulting in the emergence of cartilage and mineralized bone. The development of the vertebrate skeleton is an example of the functional segregation and subsequent recruitment of unique genetic materials that may account for the evolutionary diversification of novel cell types.     

Comparative genomics of chondrichthyan Hoxa clusters

Background  

The chondrichthyan or cartilaginous fish (chimeras, sharks, skates and rays) occupy an important phylogenetic position as the sister group to all other jawed vertebrates and as an early lineage to diverge from the vertebrate lineage following two whole genome duplication events in vertebrate evolution. There have been few comparative genomic analyses incorporating data from chondrichthyan fish and none comparing genomic information from within the group. We have sequenced the complete Hoxa cluster of the Little Skate (Leucoraja erinacea) and compared to the published Hoxa cluster of the Horn Shark (Heterodontus francisci) and to available data from the Elephant Shark (Callorhinchus milii) genome project.     

An investigation on the effect of photoperiod and temperature on vertebral band deposition in little skate Leucoraja erinacea

An investigation was undertaken to determine whether photoperiod or temperature have an effect on the timing of vertebral opaque–transluscent band‐pair deposition in captive young‐of‐the‐year (YOY) little skate Leucoraja erinacea. The experimental design consisted of a randomized complete block split plot design with two factors: temperature and light. Temperature was nested within light and therefore four variables were tested: 1) constant light, 2) constant temperature, 3) seasonal light and 4) seasonal temperature. For 18 months, L. erinacea experienced accelerated seasonal conditions of temperature and light to mimic 3 years of growth. This study provides primary and supporting evidence that seasonal photoperiod and temperature, respectively, have no effect on timing of vertebral band‐pair deposition in captive L. erinacea. Vertebral analysis of surviving L. erinacea (n = 6, time = 18 months) showed that all produced 1–1·5 band pairs, while centrum edge analysis (n = 56) showed timing of winter and summer band deposition were similar regardless of treatment. The winter band (translucent) appeared in February 2007 and January 2008 while the summer band (opaque) showed up in July for both 2007 and 2008 and mimicked patterns observed in the wild. While temperature and photoperiod appear to have no effect on timing of band‐pair deposition in YOY L. erinacea, other mechanisms which may influence band deposition should be investigated including the effect of food ration and the presence of a circa‐annual rhythm and hormone secretion.     

The morphology and evolution of the ventral gill arch skeleton in batoid fishes (Chondrichthyes: Batoidea)

The ventral gill arch skeleton was examined in some representatives of batoid fishes. The homology of the components was elucidated by comparing similarities and differences among the components of the ventral gill arches in chondrichthyans, and attempts were made to justify the homology by giving causal mechanisms of chondrogenesis associated with the ventral gill arch skeleton. The ceratohyal is present in some batoid fishes, and its functional replacement, the pseudohyal, seems incomplete in most groups of batoid fishes, except in stingrays. The medial fusion of the pseudohyal with successive ceratobranchials occurs to varying degrees among stingray groups. The ankylosis between the last two ceratobranchials occurs uniquely in stingrays, and it serves as part of the insertion of the last pair of coracobranchialis muscles. The basihyal is possibly independently lost in electric rays, the stingray genus Urotrygon (except U. daviesi) and pelagic myiiobatoid stingrays. The first hypobranchial is oriented anteriorly or anteromedially, and it varies in shape and size among batoid fishes. It is represented by rami projecting posterolaterally from the basihyal in sawfishes, guitarfishes and skates. It consists of a small piece of cartilage which extends anteromedially from the medial end of the first ccratobranchial in electric rays. It is a large cartilaginous plate in most of stingrays. It is absent in pelagic myliobatoid stingrays. The remaining hypobranchial cartilages also vary in shape and size among batoid fishes. Torpedo and possibly the Jurassic Belemnobalis and Spathobatis possess the generalized or typical chondrichthyan ventral gill arch structure in which the hypobranchials form a Σ-shaped pattern. In the electric ray Hypnos and narkinidid and narcinidid electric rays, the hypobranchial components are oriented longitudinally along the mid-portion of the ventral gill arches. They form a single cartilaginous plate in the narkinidid electric rays, Narcine and Diplobatis. In guitarfishes and skates, the second hypobranchial is unspecialized, and in skates, it does not have a direct contact with the second ceratobranchial. In both groups, the third and fourth hypobranchials are composed of a small cartilage which forms a passage for the afferent branches of the ventral aorta and serve as part of the insertion of the coracobranchialis muscle. In sawfishes and stingrays, the hypobranchials appear to be included in the medial plate. In sawfishes, the second and third components separately chondrify in adults, but the fourth component appears to be fused with the middle medial plate. In stingrays, a large medial plate appears to include the second through to the last hypobranchial and most of the basibranchial copulae. The medial plate probably develops independently in sawfishes and stingrays. Because the last basibranchial copula appears to be a composite of one to two hypobranchials and at least two basibranchial copulae, the medial plate may be formed by several developmental processes of chondrogenesis. More detailed comparative anatomical and developmental studies are needed to unveil morphogenesis and patternings of the ventral gill arch skeleton in batoid fishes.