Cranial neural crest cells contribute to connective tissue in cranial muscles in the anuran amphibian, Bombina orientalis.

The contribution of cranial neural crest cells to the development and patterning of cranial muscles in amphibians was investigated in the phylogenetically basal and morphologically generalized frog, Bombina orientalis. Experimental methods included fluorescent marking of premigratory cranial neural crest and extirpation of individual migratory streams. Neural crest cells contributed to the connective tissue component, but not the myofibers, of many larval muscles within the first two branchial arches (mandibular and hyoid), and complex changes in muscle patterning followed neural crest extirpation. Connective tissue components of individual muscles of either arch originate from the particular crest migratory stream that is associated with that arch, and this relationship is maintained regardless of the segmental identity-or embryonic derivation-of associated skeletal components. These developmental relations define a pattern of segmentation in the head of larval anurans that is similar to that previously described in the domestic chicken, the only vertebrate that has been thoroughly investigated in this respect. The fundamental role of the neural crest in patterning skeleton and musculature may represent a primitive feature of cranial development in vertebrates. Moreover, the corresponding developmental processes and cell fates appear to be conserved even when major evolutionary innovations-such as the novel cartilages and muscles of anuran larvae-result in major differences in cranial form.     

Cranial neural-crest migration and chondrogenic fate in the oriental fire-bellied toad Bombina orientalis: Defining the ancestral pattern of head development in anuran amphibians

We assess cranial neural-crest cell migration and contributions to the larval chondrocranium in the phylogenetically basal and morphologically generalized anuran Bombina orientalis (Bombinatoridae). Methods used include microdissection, scanning electron microscopy, and vital dye labeling, in conjunction with confocal and fluorescence microscopy. Cranial neural-crest cells begin migrating before neural-fold closure and soon form three primary streams. These streams contribute to all cranial cartilages except two medial components of the hyobranchial skeleton (basihyal and basibranchial cartilages), the posterior portion of the trabecular plate, and the otic capsule, the embryonic origin of which is unknown. Chondrogenic fate is regionalized within the cranial neural folds, with the anterior regions contributing to anterior cartilages and the posterior regions to posterior cartilages. A neural-crest contribution also was consistently observed in several cranial nerves and the connective tissue component of many cranial muscles. Notwithstanding minor differences among species in the initial configuration of migratory streams, cranial neural-crest migration and chondrogenic potential in metamorphosing anurans seem to be highly stereotyped and evolutionarily conservative. This includes a primary role for the neural crest in the evolutionary origin of the paired suprarostral and infrarostral cartilages, two prominent caenogenetic features of the rostral skull unique to anuran larvae. Our results provide a model of the ancestral pattern of embryonic head development in anuran amphibians. This model can serve as a basis for examining the ontogenetic mechanisms that underlie the diversity of cranial morphology and development displayed by living frogs, as well as the evolutionary consequences of this diversity. © 1996 Wiley-Liss, Inc.     

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.     

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.     

Cephalic muscle development in the Australian lungfish,Neoceratodus forsteri

Lungfishes are the extant sister group of tetrapods. As such, they are important for the study of evolutionary processes involved in the water to land transition of vertebrates. The evolution of a true neck, that is, the complete separation of the pectoral girdle from the cranium, is one of the most intriguing morphological transitions known among vertebrates. Other salient changes involve new adaptations for terrestrial feeding, which involves both the cranium and its associated musculature. Historically, the cranium has been extensively investigated, but the development of the cranial muscles much less so. Here, we present a detailed study of cephalic muscle development in the Australian lungfish, Neoceratodus forsteri, which is considered to be the sister taxon to all other extant lungfishes. Neoceratodus shows several developmental patterns previously described in other taxa; the tendency of muscles to develop from anterior to posterior, from their region of origin toward insertion, and from lateral to ventral/medial (outside‐in), at least in the branchial arches. The m.protractor pectoralis appears to develop as an extension of the most posterior m.levatores arcuum branchialium, supporting the hypothesis that the m.cucullaris and its derivatives (protractor pectoralis, levatores arcuum branchialium) are branchial muscles. We present a new hypothesis regarding the homology of the ventral branchial arch muscles (subarcualis recti and obliqui, transversi ventrales) in lungfishes and amphibians. Moreover, the morphology and development of the cephalic muscles confirms that extant lungfishes are neotenic and have been strongly influenced via paedomorphosis during their evolutionary history.     

Expression ofHoxDGenes in Developing and Regenerating Axolotl Limbs

Hoxgenes play a critical role in the development of the vertebrate axis and limbs, and previous studies have implicated them in the specification of positional identity, the control of growth, and the timing of differentiation. Axolotl limbs offer an opportunity to distinguish these alternatives because the sequence of skeletal differentiation is reversed along the anterior–posterior axis relative to that of other tetrapods. We report that during early limb development, expression patterns ofHoxDgenes in axolotls resemble those in amniotes and anuran amphibians. At later stages, the anterior boundary ofHoxd-11expression is conserved with respect to morphological landmarks, but there is no anterior–distal expansion of the posterior domain ofHoxd-11expression similar to that observed in mice and chicks. Since axolotls do not form an expanded paddle-like handplate prior to digit differentiation, we suggest that anterior expansion of expression in higher vertebrates is linked to the formation of the handplate, but is clearly not necessary for digit differentiation. We also show that the 5′HoxDgenes are reexpressed during limb regeneration. The change in the expression pattern ofHoxd-11during the course of regeneration is consistent with the hypothesis that the distal tip of the regenerate is specified first, followed by intercalation of intermediate levels of the pattern. BothHoxd-8andHoxd-10are expressed in non-regenerating wounds, butHoxd-11is specific for regeneration. It is also expressed in the posterior half of nerve-induced supernumerary outgrowths.     

The developmental patterns of the cartilaginous elements in vertebrate ontogeny]

Comparative embryological data presented in the paper support an idea that chondrification of the mesenchyme does not begin until the latter becomes condensated. Size and density of the skeletogenous rudiments are not the same in different vertebrates. As a rule, in animals with much of the mesenchyme (chondrichthyans, amniotes), the prochondral condensations contain more cells and have both greater mass and density. The distribution pattern of the mesenchyme is also significant for the future development of the cartilaginous elements which at the earlier stages grow largely by the recruitment of surrounding mesenchymal cells. Such kind of the growth mode is probably most similar to the cartilage fusion mode: both processed take place in the absence of the perichondrium. The non-skeletal dense structures influence on the development of the cartilaginous skeleton primarily by determining distribution pattern of the mesenchyme, particularly the condensation of skeletogenous cells. THe growing cartilages themselves can influence mechanically on the surrounding organs.     

Histology of the heterostracan dermal skeleton: Insight into the origin of the vertebrate mineralised skeleton

Living vertebrates are divided into those that possess a fully formed and fully mineralised skeleton (gnathostomes) versus those that possess only unmineralised cartilaginous rudiments (cyclostomes). As such, extinct phylogenetic intermediates of these living lineages afford unique insights into the evolutionary assembly of the vertebrate mineralised skeleton and its canonical tissue types. Extinct jawless and jawed fishes assigned to the gnathostome stem evidence the piecemeal assembly of skeletal systems, revealing that the dermal skeleton is the earliest manifestation of a homologous mineralised skeleton. Yet the nature of the primitive dermal skeleton, itself, is poorly understood. This is principally because previous histological studies of early vertebrates lacked a phylogenetic framework required to derive evolutionary hypotheses. Nowhere is this more apparent than within Heterostraci, a diverse clade of primitive jawless vertebrates. To this end, we surveyed the dermal skeletal histology of heterostracans, inferred the plesiomorphic heterostracan skeleton and, through histological comparison to other skeletonising vertebrate clades, deduced the ancestral nature of the vertebrate dermal skeleton. Heterostracans primitively possess a four‐layered skeleton, comprising a superficial layer of odontodes composed of dentine and enameloid; a compact layer of acellular parallel‐fibred bone containing a network of vascular canals that supply the pulp canals (L1); a trabecular layer consisting of intersecting radial walls composed of acellular parallel‐fibred bone, showing osteon‐like development (L2); and a basal layer of isopedin (L3). A three layered skeleton, equivalent to the superficial layer L2 and L3 and composed of enameloid, dentine and acellular bone, is possessed by the ancestor of heterostracans + jawed vertebrates. We conclude that an osteogenic component is plesiomorphic with respect to the vertebrate dermal skeleton. Consequently, we interpret the dermal skeleton of denticles in chondrichthyans and jawless thelodonts as independently and secondarily simplified. J. Morphol. 276:657–680, 2015. © 2015 The Authors Journal of Morphology Published by Wiley Periodicals, Inc.     

Evolutionary mechanisms of rib loss in anurans: a comparative developmental approach

ABSTRACT The presence of free ribs is presumed to be a primitive morphological character observed only in a few families of Recent anurans, whereas the absence of ribs has been considered to be a derived condition that is widespread within this order. A comparative study of rib development based on representatives of several anuran lineages (Alytes, Bombina, Bufo, Discoglossus, Hyla, Pelobates, Pelodytes, Rana, and Xenopus) reveals a previously undetected diversity of developmental features in the formation and interaction between neural arches and ribs. The absence of free ribs at premetamorphic or later stages is verified in some groups, but we present for the first time evidence of the existence of larval rib rudiments in others, both in the anterior (Rana, Hyla) and posterior (Bufo, Discoglossus, Pelobates) presacral regions. Heterochrony seems to have played a major role in the processes underlying rib reduction. The intracolumnar differences between anterior (V(2)-V(4)) and posterior (V(5)-V(8)) regions are based on perturbations in the timing of early differentiation. Furthermore, a clear shift in the relative timing of ossification among evolutionary lineages was detected. In this respect Xenopus has a highly derived condition. The use of the morphological character of "rib loss" in phylogenetic analyses must be reconsidered due to the different convergent developmental paths described here. The phylogenetic analysis of a "sequence units" matrix of rib development is compared with current anuran phylogenies. Some evolutionary information appears to be clearly present in the ontogenetic data of this "missing morphology," but its value for evolutionary inferences is rather limited.     

Skull development during anuran metamorphosis: I. Early development of the first three bones to form--the exoccipital, the parasphenoid, and the frontoparietal

In anuran amphibians, cranial bones typically first form at metamorphosis when they rapidly invest or replace the cartilaginous larval skull. We describe early development of the first three bones to form in the Oriental fire-bellied toad, Bombina orientalis--the parasphenoid, the frontoparietal, and the exoccipital--based on examination of serial sections. Each of these bones is fully differentiated by Gosner stage 31 (hindlimb in paddle stage) during premetamorphosis. This is at least six Gosner developmental stages before they are first visible in whole-mount preparations at the beginning of prometamorphosis. Thus, developmental events that precede and mediate the initial differentiation of these cranial osteogenic sites occur very early in metamorphosis--a period generally considered to lack significant morphological change. Subsequent development of these centers at later stages primarily reflects cell proliferation and calcified matrix deposition, possibly in response to increased circulating levels of thyroid hormone which are characteristic of later metamorphic stages. Interspecific differences in the timing of cranial ossification may reflect one or both of these phases of bone development. These results may qualify the use of whole-mount preparations for inferring the sequence and absolute timing of cranial ossification in amphibians.     

The mechanism of cartilage subdivision in the reorganization of the zebrafish pectoral fin endoskeleton

A cartilaginous pectoral fin endoskeleton in zebrafish (Danio rerio) develops early, after which the cartilage of the larval fin endoskeleton undergoes a complete transformation into the adult morphology. This transformation includes multiple subdivisions of a single cartilaginous disk. The type of cartilage subdivision is unique to teleost fish. In this study, we present the timing and the developmental features of these subdivisions and we discuss variation in this process, caused by differences in growth rate. We establish that the cartilage subdivisions are developmentally linked to the formation of lepidotrichia in the fin fold. At the cellular level, we show that neither apoptosis nor resorption by chondroclasts and/or macrophages contributes to the cartilage subdivision. Ultrastructural observations show dedifferentiation of chondrocytes in subdivision zones. Different from forelimb development in other vertebrates, dedifferentiation is an important mechanism in the development of the adult pectoral fin skeleton. We here provide further support for the idea that the phenotype of skeletal tissues is not terminal and that plasticity of differentiated connective tissues can play an important role in various developmental and homeostatic processes.     

Limb development in a "nonmodel" vertebrate, the direct-developing frog Eleutherodactylus coqui.

Mechanisms that mediate limb development are regarded as highly conserved among vertebrates, especially tetrapods. Yet, this assumption is based on the study of relatively few species, and virtually none of those that display any of a large number of specialized life-history or reproductive modes, which might be expected to affect developmental pattern or process. Direct development is an alternative life history found in many anuran amphibians. Many adult features that form after hatching in metamorphic frogs, such as limbs, appear during embryogenesis in direct-developing species. Limb development in the direct-developing frog Eleutherodactylus coqui presents a mosaic of apparently conserved and novel features. The former include the basic sequence and pattern of limb chondrogenesis, which are typical of anurans generally and appear largely unaffected by the gross shift in developmental timing; expression of Distal-less protein (Dlx) in the distal ectoderm; expression of the gene Sonic hedgehog (Shh) in the zone of polarizing activity (ZPA); and the ability of the ZPA to induce supernumerary digits when transplanted to the anterior region of an early host limb bud. Novel features include the absence of a morphologically distinct apical ectodermal ridge, the ability of the limb to continue distal outgrowth and differentiation following removal of the distal ectoderm, and earlier cessation of the inductive ability of the ZPA. Attempts to represent tetrapod limb development as a developmental "module" must allow for this kind of evolutionary variation among species.     

Development of the ethmoidal structures of the endocranium in Discoglossus pictus (Anura: Discoglossidae)

We use histological techniques and computer‐aided three‐dimensional reconstructions made from serial histological sections to describe the ontogeny of the ethmoidal endocranium of discoglossid frog Discoglossus pictus. We identify a pattern of development for the suprarostral cartilage that differs from previous findings and probably represents the ancestral anuran pattern. The nasal cartilages, including the inferior prenasal cartilage, are de novo adult structures. The only larva‐derived structures of the adult nasal capsules are the posterior aspects of the solum nasi and septum nasi. We also identify patterns of development for the ethmoid plate and postnasal wall that occur during early in ontogenesis. These patterns are associated with development events during metamorphic climax. The pattern of timing of chondrification of the anterior nasal cartilages more closely coincides with that of the neobatrachian species than that recorded for the pelobatid frog Spea. In addition, this study supports a sister taxon relationship between Discoglossus and Alytes. J. Morphol. 271:1078‐1093, 2010. © 2010 Wiley‐Liss, Inc.     

Retrofitting larval neuromuscular circuits in the metamorphosing frog

Maturation of vertebrate neuromuscular systems typically occurs in a continuous, orderly progression. After an initial period of developmental adjustment by means of cell death and axonal pruning, relatively stable relationships, with only subtle modifications, are maintained between motoneurons and their appropriate targets throughout life. However, among a restricted group of vertebrates (amphibians and especially the anuran amphibians) the sequential maturation of neuromuscular systems is altered by an abrupt reordering of the basic body plan that encompasses cellular changes in all tissues from skeleton to nervous system. Many anuran amphibians possess neuromuscular circuits that are remarkable by virtue of their complete reorganization during the brief span of metamorphosis. During this period motor systems initially designed for the behavioral patterns of aquatic tadpoles are adjusted to meet the drastically different motor activities of postmetamorphic terrestrial life. This adjustment involves the deletion of neural elements mediating larval specific activities, the accelerated maturation of neural circuits eliciting adult-specific activities and the retrofitting of larval neuromuscular components to serve postmetamorphic behaviors. This review focuses on the cellular events associated with the neuromuscular adaptation in the jaw complex during metamorphosis of the leopard frog, Rana pipiens. As part of the metamorphic reorganization of the jaw apparatus there is a complete turnover of the myofiber complement of the adductor mandibulae musculature. Trigeminal motoneurons initially deployed to the larval myofibers are redirected to new muscle fibers. Simultaneously the cellular geometry and synaptic input to these motoneurons is revamped. These changes suggest that trigeminal neuromuscular circuitry established during embryogenesis is updated during metamorphosis and reused to provide the basis for adult jaw motor activity that is far different than its larval counterpart.     

Cranial neural crest cell migration in the Australian lungfish, Neoceratodus forsteri

SUMMARY A crucial role for the cranial neural crest in head development has been established for both actinopterygian fishes and tetrapods. It has been claimed, however, that the neural crest is unimportant for head development in the Australian lungfish ( Neoceratodus forsteri   ), a member of the group (Dipnoi) which is commonly considered to be the living sister group of the tetrapods. In the present study, we used scanning electron microscopy to study cranial neural crest development in the Australian lungfish. Our results, contrary to those of Kemp, show that cranial neural crest cells do emerge and migrate in the Australian lungfish in the same way as in other vertebrates, forming mandibular, hyoid, and branchial streams. The major difference is in the timing of the onset of cranial neural crest migration. It is delayed in the Australian lungfish in comparison with their living sister group the Lissamphibia. Furthermore, the delay in timing between the emergence of the hyoid and branchial crest streams is very long, indicating a steeper anterior-posterior gradient than in amphibians. We are now extending our work on lungfish head development to include experimental studies (ablation of selected streams of neural crest cells) and fate mapping (using fluoresent tracer dyes such as DiI) to document the normal fate as well as the role in head patterning of the cranial neural crest in the Australian lungfish.     

Ossification patterns in the tetrapod limb – conservation and divergence from morphogenetic events

Two different patterns of the condensation and chondrification of the limbs of tetrapods are known from extensive studies on their early skeletal development. These are on the one hand postaxial dominance in the sequential formation of skeletal elements in amniotes and anurans, and on the other, preaxial dominance in urodeles. The present study investigates the relative sequence of ossification in the fore‐ and hindlimbs of selected tetrapod taxa based on a literature survey in comparison to the patterns of early skeletal development, i.e. mesenchymal condensation and chondrification, representing essential steps in the late stages of tetrapod limb development. This reveals the degree of conservation and divergence of the ossification sequence from early morphogenetic events in the tetrapod limb skeleton. A step‐by‐step recapitulation of condensation and chondrification during the ossification of limbs can clearly be refuted. However, some of the deeper aspects of early skeletal patterning in the limbs, i.e. the general direction of development and sequence of digit formation are conserved, particularly in anamniotes. Amniotes show a weaker coupling of the ossification sequence in the limb skeleton with earlier condensation and chondrification events. The stronger correlation between the sequence of condensation/chondrification and ossification in the limbs of anamniotes may represent a plesiomorphic trait of tetrapods. The pattern of limb ossification across tetrapods also shows that some trends in the sequence of ossification of their limb skeleton are shared by major clades possibly representing phylogenetic signals. This review furthermore concerns the ossification sequence of the limbs of the Palaeozoic temnospondyl amphibian Apateon sp. For the first time this is described in detail and its patterns are compared with those observed in extant taxa. Apateon sp. shares preaxial dominance in limb development with extant salamanders and the specific order of ossification events in the fore‐ and hindlimb of this fossil dissorophoid is almost identical to that of some modern urodeles.