Miniaturization and its effects on cranial morphology in plethodontid salamanders, genus Thorius (Amphibia: Plethodontidae). I. Osteological variation

Cranial skeletal morphology, ontogeny and variation are examined in five species of Thorius , a genus of diminutive plethodontid salamanders that are among the smallest, extant, tailed tetrapods. The skull of adull Thorius is characterized by: (1) limited development or absence of several ossified elements and dentition; (2) increased inter-and intraspecific variability; (3) novel morphological configurations of the braincase and jaw suspensorium. Posthatching cranial mineralization in all species of Thorius is truncated precociously with respect to that typical of larger and more generalized plethodontid genera, such as Pseudoeurycea. These features implicate paedomorphosis as a predominant mechanism responsible for the evolution of decreased size in Thorius from larger plethodontid ancestors. Interspecific differences in cranial morphology are evident; species may be characterized by greater or lesser degrees of truncated development. However, there is no consistent relationship between degree of paedomorphosis and mean adult body size in interspecific comparisons. Adult morphology of several individual elements represent potentially useful taxonomic characters for distinguishing species.
Reduction, increased variability, and morphological novelty are common to many lineages of dwarfed taxa. They represent a null hypothesis for examination of the developmental mechanisms and morphological consequences of miniaturization in other groups.     

The skull and jaw musculature as guides to the ancestry of salamanders

The fossil record provides no evidence supporting a unique common ancestry for frogs, salamanders and apodans. The ancestors of the modern orders may have diverged from one another as recently as 250 million years ago, or as long ago as 400 million years according to current theories of various authors. In order to evaluate the evolutionary patterns of the modern orders it is necessary to determine whether their last common ancestor was a rhipidistian fish, a very primitive amphibian, a labyrimhodom or a ‘lissamphibian’. The broad cranial similarities of frogs and salamanders, especially the dominance of the braincase as a supporting element, can be associated with the small size of the skull in their immediate ancestors. Hynobiids show the most primitive cranial pattern known among the living salamander families and “provide a model for determining the nature of the ancestors of the entire order. Features expected in ancestral salamanders include: (1) Emargination of the cheek; (2) Movable suspensorium formed by the quadrate, squamosal and pterygoid; (3) Occipital condyle posterior to jaw articulation; (4) Distinct prootic and opisthotic; (5) Absence ol otic notch; (6) Stapes forming a structural link between braincase and cheek. In the otic region, cheek and jaw suspension, the primitive salamander pattern (resembles most closely the microsaurs among known Paleozoic amphibians, and shows no significant features in common with either ancestral frogs or the majority of labyrinth odonts. The basic pattern of the adductor jaw musculature is consistent within both frogs and salamanders, but major differences are evident between the two groups. The dominance of the adductor mandibulae externus in salamanders can be associated with the open cheek in all members of that order, and the small size of this muscle in frogs can be associated with the large otic notch. The spread of different muscles over the otic capsule, the longus head ol the adductor mandibulae posterior in frogs and the superficial head of the adductor mandibulae internus in salamanders, indicates that fenestration of the skull posterodorsal to the orbit occurred separately in the ancestors of the two groups. Reconstruction of the probable pattern of the jaw musculature in Paleozoic amphibians indicates that frogs and salamanders might have evolved from a condition hypothesized for primitive labyrinthodonts, but the presence of a large otic notch in dissorophids suggests specialization toward the anuran, not the urodele condition. The presence of either an einarginated cheek or an embayment of the lateral surface of the dentary and the absence of an otic notch in microsaurs indicate a salamander-like distribution of die adductor jaw muscles. The ancestors of frogs and salamanders probably diverged from one another in the early Carboniferous, Frogs later evolved from small labyrinthodonts and salamanders from microsaurs. Features considered typical of lissamphibians evolved separately in the two groups in the late Permian andTriassic.     

Distinct spatiotemporal roles of hedgehog signalling during chick and mouse cranial base and axial skeleton development

The cranial base exerts a supportive role for the brain and includes the occipital, sphenoid and ethmoid bones that arise from cartilaginous precursors in the early embryo. As the occipital bone and the posterior part of the sphenoid are mesoderm derivatives that arise in close proximity to the notochord and floor plate, it has been assumed that their development, like the axial skeleton, is dependent on Sonic hedgehog (Shh) and modulation of bone morphogenetic protein (Bmp) signalling. Here we examined the development of the cranial base in chick and mouse embryos to compare the molecular signals that are required for chondrogenic induction in the trunk and head. We found that Shh signalling is required but the molecular network controlling cranial base development is distinct from that in the trunk. In the absence of Shh, the presumptive cranial base did not undergo chondrogenic commitment as determined by the loss of Sox9 expression and there was a decrease in cell survival. In contrast, induction of the otic capsule occurred normally demonstrating that induction of the cranial base is uncoupled from formation of the sensory capsules. Lastly, we found that the early cranial mesoderm is refractory to Shh signalling, likely accounting for why development of the cranial base occurs after the axial skeleton. Our data reveal that cranial and axial skeletal induction is controlled by conserved, yet spatiotemporally distinct mechanisms that co-ordinate development of the cranial base with that of the cranial musculature and the pharyngeal arches.     

Head size,weaponry, and cervical adaptation: Testing craniocervical evolutionary hypotheses in Ceratopsia

The anterior cervical vertebrae form the skeletal connection between the cranial and postcranial skeletons in higher tetrapods. As a result, the morphology of the atlas‐axis complex is likely to be shaped by selection pressures acting on either the head or neck. The neoceratopsian (Reptilia:Dinosauria) syncervical represents one of the most highly modified atlas‐axis regions in vertebrates, being formed by the complete coalescence of the three most anterior cervical vertebrae. In ceratopsids, the syncervical has been hypothesized to be an adaptation to support a massive skull, or to act as a buttress during intraspecific head‐to‐head combat. Here, we test these functional/adaptive hypotheses within a phylogenetic framework and critically examine the previously proposed methods for quantifying relative head size in the fossil record for the first time. Results indicate that neither the evolution of cranial weaponry nor large head size correlates with the origin of cervical fusion in ceratopsians, and we, therefore, reject both adaptive hypotheses for the origin of the syncervical. Anterior cervical fusion has evolved independently in a number of amniote clades, and further research on extant groups with this peculiar anatomy is needed to understand the evolutionary basis for cervical fusion in Neoceratopsia.     

Miniaturization in plethodontid salamanders (Gaudata: Plethodontidae) and its consequences for the brain and visual system

In seven species of plethodontid salamanders (Desmognathus ochrophaeus, Eurycea bislineata, Plethodon cinereus, Batrachoseps attenuatus, Hydromantes italicus, Thorius narisovalis and Bolitoglossa subpalmata), absolute and relative volumes of the eye, the brain, major regions of the brain, and regions containing the major visual and visuomotor centres (i.e. thalamus, praetectum, tectum and tegmentum mesencephali), and the density and number of neurons in these regions were determined. The seven species range from moderately large to extremely small in body size and from the smallest to the largest genome sizes found in terrestrial salamanders. The following processes were observed in miniaturized salamanders with intermediate to large genome and cell sizes (Batrachoseps, Thorius) as compared to small and medium-sized salamanders with small genome and cell sizes: (1) increase in the relative size of the brain, from 3.9 to 12.4% of head volume; (2) reduction in relative size of the ventricles from 10.9 to 5.8% of brain volume; (3) increase in relative volume of those brain regions containing the major visual and visuomotor centres from 29.2 to 37% of brain volume; (4) increase in volume of grey matter relative to white matter, from 33.2 to 44.4% of midbrain volume; (5) increase in volume of tectal relative to tegmental grey matter, from 54.8 to 76.8% of total midbrain volume; (6) increase in neuron packing density in the regions containing the visual centres, from 16 to 31.5%. Because of these compensatory processes, Thorius, the smallest species with a head 1/27 and a brain 1/9 the size of that of the largest one, Hydromantes, has 1/3 as many central visual neurons (58 000 vs. 187 000). Some of these processes found in miniaturized salamanders, such as increase in tectal cell density, also occur in large salamanders with very large genome and cell sizes, viz. in Bolitoglossa (25%) and Hydromantes (29%). Thus, increase in genome size and cell size seem to pose functional problems similar to miniaturization; both cases involve an increase in cell size relative to overall organismal structure.     

Development and evolution of the tetrapod skull–neck boundary

The origin and evolution of the vertebrate skull have been topics of intense study for more than two centuries. Whereas early theories of skull origin, such as the influential vertebral theory, have been largely refuted with respect to the anterior (pre‐otic) region of the skull, the posterior (post‐otic) region is known to be derived from the anteriormost paraxial segments, i.e. the somites. Here we review the morphology and development of the occiput in both living and extinct tetrapods, taking into account revised knowledge of skull development by augmenting historical accounts with recent data. When occipital composition is evaluated relative to its position along the neural axis, and specifically to the hypoglossal nerve complex, much of the apparent interspecific variation in the location of the skull–neck boundary stabilizes in a phylogenetically informative way. Based on this criterion, three distinct conditions are identified in (i) frogs, (ii) salamanders and caecilians, and (iii) amniotes. The position of the posteriormost occipital segment relative to the hypoglossal nerve is key to understanding the evolution of the posterior limit of the skull. By using cranial foramina as osteological proxies of the hypoglossal nerve, a survey of fossil taxa reveals the amniote condition to be present at the base of Tetrapoda. This result challenges traditional theories of cranial evolution, which posit translocation of the occiput to a more posterior location in amniotes relative to lissamphibians (frogs, salamanders, caecilians), and instead supports the largely overlooked hypothesis that the reduced occiput in lissamphibians is secondarily derived. Recent advances in our understanding of the genetic basis of axial patterning and its regulation in amniotes support the hypothesis that the lissamphibian occipital form may have arisen as the product of a homeotic shift in segment fate from an amniote‐like condition.     

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.     

Metamorphosis of cranial design in tiger salamanders (Ambystoma tigrinum): A morphometric analysis of ontogenetic change

While ontogenetic analyses of skull development have contributed to our understanding of phylogenetic patterns in vertebrates, there are few studies of taxa that undergo a relatively discrete and rapid change in morphology during development (metamorphosis). Morphological changes occurring in the head at metamorphosis in tiger salamanders (Ambystoma tigrinum) were quantified by a morphometric analysis of cranial osteology and myology to document patterns of change during metamorphosis. We employed a cross-sectional analysis using a sample of larvae just prior to metamorphosis and a sample of transformed individuals just after metamorphosis, as well as larvae undergoing metamorphosis. There were no differences in external size of the head among the larval and transformed samples. The hyobranchial apparatus showed many dramatic changes at metamorphosis, including shortening of ceratobranchial 1 and the basibranchial. The subarcualis rectus muscle increased greatly in length at metamorphosis, as did hypobranchial length and internasal distance. A truss analysis of dorsal skull shape showed that at metamorphosis the snout becomes wider, the maxillary and squamosal triangles rotate posteromedially, and the neurocranium shortens (while maintaining its width), resulting in an overall decrease in skull length at metamorphosis. These morphometric differences are interpreted in light of recent data on the functional morphology of feeding in salamanders. Morphological reorganization of the hyobranchial apparatus and shape changes in the skull are related to the acquisition of a novel terrestrial feeding mode (tongue projection) at metamorphosis. Metamorphic changes (both internal and external) that can be used to judge metamorphic condition are discussed.     

Constructional morphology within the head of hammerhead sharks (sphyrnidae)

The study of functional trade‐offs is important if a structure, such as the cranium, serves multiple biological roles, and is, therefore, shaped by multiple selective pressures. The sphyrnid cephalofoil presents an excellent model for investigating potential trade‐offs among sensory, neural, and feeding structures. In this study, hammerhead shark species were chosen to represent differences in head form through phylogeny. A combination of surface‐based geometric morphometrics, computed tomography (CT) volumetric analysis, and phylogenetic analyses were utilized to investigate potential trade‐offs within the head. Hammerhead sharks display a diversity of cranial morphologies where the position of the eyes and nares vary among species, with only minor changes in shape, position, and volume of the feeding apparatus through phylogeny. The basal winghead shark, Eusphyra blochii, has small anteriorly positioned eyes. Through phylogeny, the relative size and position of the eyes change, such that derived species have larger, more medially positioned eyes. The lateral position of the external nares is highly variable, showing no phylogenetic trend. Mouth size and position are conserved, remaining relatively unchanged. Volumetric CT analyses reveal no trade‐offs between the feeding apparatus and the remaining cranial structures. The few trade‐offs were isolated to the nasal capsule volume's inverse correlation with braincase, chondrocranial, and total cephalofoil volume. Eye volume also decreased as cephalofoil width increased. These data indicate that despite considerable changes in head shape, much of the head is morphologically conserved through sphyrnid phylogeny, particularly the jaw cartilages and their associated feeding muscles, with shape change and morphological trade‐offs being primarily confined to the lateral wings of the cephalofoil and their associated sensory structures. J. Morphol. 276:526–539, 2015. © 2015 Wiley Periodicals, Inc.     

The influence of feeding on the evolution of sensory signals: a comparative test of an evolutionary trade‐off between masticatory and sensory functions of skulls in southern African Horseshoe bats (Rhinolophidae)

The skulls of animals have to perform many functions. Optimization for one function may mean another function is less optimized, resulting in evolutionary trade‐offs. Here, we investigate whether a trade‐off exists between the masticatory and sensory functions of animal skulls using echolocating bats as model species. Several species of rhinolophid bats deviate from the allometric relationship between body size and echolocation frequency. Such deviation may be the result of selection for increased bite force, resulting in a decrease in snout length which could in turn lead to higher echolocation frequencies. If so, there should be a positive relationship between bite force and echolocation frequency. We investigated this relationship in several species of southern African rhinolophids using phylogenetically informed analyses of the allometry of their bite force and echolocation frequency and of the three‐dimensional shape of their skulls. As predicted, echolocation frequency was positively correlated with bite force, suggesting that its evolution is influenced by a trade‐off between the masticatory and sensory functions of the skull. In support of this, variation in skull shape was explained by both echolocation frequency (80%) and bite force (20%). Furthermore, it appears that selection has acted on the nasal capsules, which have a frequency‐specific impedance matching function during vocalization. There was a negative correlation between echolocation frequency and capsule volume across species. Optimization of the masticatory function of the skull may have been achieved through changes in the shape of the mandible and associated musculature, elements not considered in this study.     

An ontogenetic perspective on the study of sexual dimorphism, phylogenetic variability, and allometry of the skull of European ground squirrel, Spermophilus citellus (Linnaeus, 1766)

Most studies of morphological variability in or among species are performed on adult specimens. However, it has been proven that knowledge of the patterns of size and shape changes and their covariation during ontogeny is of great value for the understanding of the processes that produce morphological variation. In this study, we investigated the patterns of sexual dimorphism, phylogenetic variability, and ontogenetic allometry in the Spermophilus citellus with geometric morphometrics applied to cross-sectional ontogenetic data of 189 skulls from three populations (originating from Burgenland, Banat, and Dojran) belonging to two phylogenetic lineages (the Northern and Southern). Our results indicate that sexual dimorphism in the ventral cranium of S. citellus is expressed only in skull size and becomes apparent just before or after the first hibernation because of accelerated growth in juvenile males. Sexes had the same pattern of ontogenetic allometry. Populations from Banat and Dojran, belonging to different phylogroups, were the most different in size but had the most similar adult skull shape. Phylogenetic relations among populations, therefore, did not reflect skull morphology, which is probably under a significant influence of ecological factors. Populations had parallel allometric trajectories, indicating that alterations in development probably occur prenatally. The species’ allometric relations during cranial growth showed characteristic nonlinear trajectories in the two northern populations, with accelerated shape changes in juveniles and continued but almost isometric growth in adults. The adult cranial shape was reached before sexual maturity of both sexes and adult size after sexual maturity. The majority of shape changes during growth are probably correlated with the shift from a liquid to a solid diet and to a lesser degree due to allometric scaling, which explained only 20 % of total shape variation. As expected, viscerocranial components grew with positive and neurocranial with negative allometry.     

Geographic Variation in the Skull Morphometry of Four Populations of Batrachuperus karlschmidti(Urodela: Hynobiidae)

Geographic variation of morphology is an important topic of evolutionary biology, and research on geographic variation can provide insights on the formation, evolution, and adaptation of species and subspecies. The vertebrate skull is a developmentally and functionally complex morphological structure with multiple functions, that is susceptible to vary according to selection pressure. In this study, geographic variations in skull morphology of Batrachuperus karlschmidti from four different geographic populations(Shade, Gexi,Shangluokema, and Xinduqiao) were examined via geometric morphometrics. No significant differences were found among these populations with regard to skull size; however, significant variation was found in skull shape. The most notable shape changes are the relative sizes and positions of the frontal, maxilla,pterygoid, and vomer. Skull shape changes were not related to allometry. However, due to limitation of sample populations and size, the results of this study need to be further verified by more sample populations and individuals in the future. The results of this study contribute to our knowledge about these aspects of morphological variability in this species as well as in hynobiid salamanders.     

Early development of chondrocranium in the tailed frog Ascaphus truei (Amphibia: Anura): Implications for anuran palatoquadrate homologies

Chondrocranial development in Ascaphus truei was studied by serial sectioning and graphical reconstruction. Nine stages (21–29; 9–18 mm TL) were examined. Mesodermal cells were distinguished from ectomesenchymal (neural crest derived) cells by retained yolk granules. Ectomesenchymal parts of the chondrocranium include the suprarostrals, pila preoptica, anterior trabecula, and palatoquadrate. Mesodermal parts of the chondrocranium include the orbital cartilage, posterior trabecula, parachordal, basiotic lamina, and otic capsule. Development of the palatoquadrate is as follows. The pterygoid process first connects with the trabecula far rostrally; their fusion progresses caudally. The ascending process connects with a mesodermal bar that extends from the orbital cartilage to the otic capsule, and forms the ventral border of the dorsal trigeminal outlet. This bar is the “ascending process” of Ascaphus adults; it is a neurocranial, not palatoquadrate structure. The basal process chondrifies in an ectomesenchymal strand running from the quadrate keel to the postpalatine commissure. Later, the postpalatine commissure and basal process extend anteromedially to contact the floor of the anterior cupula of the otic capsule, creating separate foramina for the palatine and hyomandibular branches of the facial nerve. Based on these data, and on comparison with other frogs and salamanders, the anuran anterior quadratocranial commissure is homologized with the pterygoid process of salamanders, the anuran basal process (=“pseudobasal” or “hyobasal” process) with the basal process of salamanders, and the anuran otic ledge with the basitrabecular process of salamanders. The extensive similarities in palatoquadrate structure and development between frogs and salamanders, and lacking in caecilians, are not phylogenetically informative. Available information on fossil outgroups suggests that some of these similarities are primitive for Lissamphibia, whereas for others the polarity is uncertain. J. Morphol. 231:63-100, 1997. © 1997 Wiley-Liss, Inc.     

Tongue evolution in lungless salamanders, family plethodontidae. III. Patterns of peripheral innervation

Innervation of the tongue and associated musculature in plethodontid salamanders was studied using Palmgren stained sectioned materials, fresh dissection, and whole mounts of experimental specimens treated with horseradish peroxidase (HRP). Species studied were chosen to represent modes of tongue projection recognized by Lombard and Wake ('77). Special attention was given to species of the genera Plethodon, Batrachoseps, Pseudoeurycea, and Hydromantes, but representatives of other genera were investigated. As expected we found that cranial nerves IX and X and spinal nerve 1 supplied the muscles involved in tongue movement. The peripheral courses of the nerves were traced, and both functionally related and phylogenetically determined routes were found. As relative projection length increases, the nerves supplying the tongue tip also increase in length. When the tongue is at rest the long nerves are stored in coils. The coil of ramus lingualis lies between the ceratobranchials, but that of ramus hypoglossus is more variable, although constant within a species. Ramus hypoglossus bifurcates into separate branches to tongue and anterior musculature of the floor of the mouth. In generalized, presumably primitive, modes the bifurcation and coiling are far anterior. In most of the tongue projection modes bifurcation is relatively posterior, but in one, bifurcation is anterior, but coiling is relatively posterior in position. The most unusual condition is in Hydromantes, in which bifurcation is relatively posterior and a coiled ramus hypoglossus joins a coiled ramus lingualis to form a unique, coiled common ramus to the tongue tip. Hydromantes has the greatest projection distance of any salamander.     

Chondrocranial development in larval Rana sylvatica (Anura: Ranidae): morphometric analysis of cranial allometry and ontogenetic shape change

This study provides baseline quantitative data on the morphological development of the chondrocranium in a larval anuran. Both linear and geometric morphometric methods are used to quantitatively analyze size-related shape change in a complete developmental series of larvae of the wood frog, Rana sylvatica. The null hypothesis of isometry was rejected in all geometric morphometric and most linear morphometric analyses. Reduced major axis regressions of 11 linear chondrocranial measurements on size indicate a mixture of allometric and isometric scaling. Measurements in the otic and oral regions tend to scale with negative allometry and those associated with the palatoquadrate and muscular process scale with isometry or positive allometry. Geometric morphometric analyses, based on a set of 11 chondrocranial landmarks, include linear regression of relative warp scores and multivariate regression of partial warp scores and uniform components on log centroid size. Body size explains about one-quarter to one-third of the total shape variation found in the sample. Areas of regional shape transformation (e.g., palatoquadrate, otic region, trabecular horns) are identified by thin-plate spline deformation grids and are concordant with linear morphometric results. Thus, the anuran chondrocranium is not a static structure during premetamorphic stages and allometric patterns generally follow scaling predictions for tetrapod cranial development. Potential implications regarding larval functional morphology, cranial development, and chondrocranial evolution in anurans are discussed.     

Component analysis of craniologic characters of silver foxes (Vulpes fulvus Desm.) and their changes arising under domestication]

The data on cranial measurements performed in silver foxes indicate that there are differences in sizes measured between the farm foxes--bred population and the population selected for domestication. A method of principal components was used to analyse the cranial measurements and their changes under domestication. The first component covers about 50% of cranial diversity, which is interpreted as variation in the total skull size. This component clearly separates the two sexes, but not different populations. The second component presumably reflects the growth rate allometry between the skull length and width. The third and fourth components are measurements of skull width; the fifth one reflects the sizes of brain skull. None of these components clearly separate the foxes from farm--bred and domesticated populations. However, some differences in distribution are observed.