Comparative development of limb musculature in phylogenetically and ecologically divergent lizards.

BACKGROUND: Squamate reptiles (lizards, snakes, and amphisbaenians) exhibit incredible diversity in their locomotion, behavior, morphology, and ecological breadth. Although they often are used as models of locomotor diversity, surprisingly little attention has been given to muscle development in squamate reptiles. In fact, the most detailed examination was conducted almost 80 years ago and solely focused on the proximal limb regions. Herein, we present forelimb and hindlimb muscle morphogenesis data for three lizard species with different locomotion and feeding strategies: the desert grassland whiptail lizard, the central bearded dragon, and the veiled chameleon. This study fills critical gaps in our understanding of muscle morphogenesis in squamate reptiles and presents a comparative and temporospatial analysis of muscle development. RESULTS: Our results reveal a conserved pattern of early muscle development among lizards with different adult morphologies and ecologies. The variations that exist are concentrated in distal regions, particularly the specialized autopodia of chameleons, where differentiation of muscles associated with the digits is delayed. CONCLUSIONS: The chameleon autopod provides an example of major evolutionary modifications to the skeleton with only minor disruption of the conserved order and pattern of limb muscle development. This robustness of muscle patterning facilitates the evolution of extreme yet functional phenotypes.

A new heart for a new head in vertebrate cardiopharyngeal evolution.

It has been more than 30 years since the publication of the new head hypothesis, which proposed that the vertebrate head is an evolutionary novelty resulting from the emergence of neural crest and cranial placodes. Neural crest generates the skull and associated connective tissues, whereas placodes produce sensory organs. However, neither crest nor placodes produce head muscles, which are a crucial component of the complex vertebrate head. We discuss emerging evidence for a surprising link between the evolution of head muscles and chambered hearts - both systems arise from a common pool of mesoderm progenitor cells within the cardiopharyngeal field of vertebrate embryos. We consider the origin of this field in non-vertebrate chordates and its evolution in vertebrates.

Vertebral architecture in the earliest stem tetrapods.

The construction of the vertebral column has been used as a key anatomical character in defining and diagnosing early tetrapod groups. Rhachitomous vertebrae--in which there is a dorsally placed neural arch and spine, an anteroventrally placed intercentrum and paired, posterodorsally placed pleurocentra--have long been considered the ancestral morphology for tetrapods. Nonetheless, very little is known about vertebral anatomy in the earliest stem tetrapods, because most specimens remain trapped in surrounding matrix, obscuring important anatomical features. Here we describe the three-dimensional vertebral architecture of the Late Devonian stem tetrapod Ichthyostega using propagation phase-contrast X-ray synchrotron microtomography. Our scans reveal a diverse array of new morphological, and associated developmental and functional, characteristics, including a possible posterior-to-anterior vertebral ossification sequence and the first evolutionary appearance of ossified sternal elements. One of the most intriguing features relates to the positional relationships between the vertebral elements, with the pleurocentra being unexpectedly sutured or fused to the intercentra that directly succeed them, indicating a 'reverse' rhachitomous design. Comparison of Ichthyostega with two other stem tetrapods, Acanthostega and Pederpes, shows that reverse rhachitomous vertebrae may be the ancestral condition for limbed vertebrates. This study fundamentally revises our current understanding of vertebral column evolution in the earliest tetrapods and raises questions about the presumed vertebral architecture of tetrapodomorph fish and later, more crownward, tetrapods.

Musculoskeletal anatomy of the pelvic fin of Polypterus: implications for phylogenetic distribution and homology of pre- and postaxial pelvic appendicular muscles.

As a member of the most basal clade of extant ray-finned fishes (actinopterygians) and of one of the most basal clades of osteichthyans (bony fishes + tetrapods), Polypterus can provide insights into the ancestral anatomy of both ray-finned and lobe-finned fishes, including those that gave rise to tetrapods. The pectoral fin of Polypterus has been well described but, surprisingly, neither the bones nor the muscles of the pelvic fin are well known. We stained and dissected the pelvic fin of Polypterus senegalus and Polypterus delhezi to offer a detailed description of its musculoskeletal anatomy. In addition to the previously described adductor and abductor muscles, we found preaxial and postaxial muscles similar to those in the pectoral fin of members of this genus. The presence of pre- and postaxial muscles in both the pectoral and pelvic fins of Polypterus, combined with recent descriptions of similar muscles in the lobe-finned fishes Latimeria and Neoceratodus, suggests that they were present in the most recent common ancestor of bony fishes. These results have crucial implications for the evolution of appendicular muscles in both fish and tetrapods.

An experimental and morphometric test of the relationship between vertebral morphology and joint stiffness in Nile crocodiles (Crocodylus niloticus).

Despite their semi-aquatic mode of life, modern crocodylians use a wide range of terrestrial locomotor behaviours, including asymmetrical gaits otherwise only found in mammals. The key to these diverse abilities may lie in the axial skeleton. Correlations between vertebral morphology and both intervertebral joint stiffness and locomotor behaviour have been found in other animals, but the vertebral mechanics of crocodylians have not yet been experimentally and quantitatively tested. We measured the passive mechanics and morphology of the thoracolumbar vertebral column in Crocodylus niloticus in order to validate a method to infer intervertebral joint stiffness based on morphology. Passive stiffness of eight thoracic and lumbar joints was tested in dorsal extension, ventral flexion and mediolateral flexion using cadaveric specimens. Fifteen measurements that we deemed to be potential correlates of stiffness were taken from each vertebra and statistically tested for correlation with joint stiffness. We found that the vertebral column of C. niloticus is stiffer in dorsoventral flexion than in lateral flexion and, in contrast to that of many mammals, shows an increase in joint stiffness in the lumbar region. Our findings suggest that the role of the axial column in crocodylian locomotion may be functionally different from that in mammals, even during analogous gaits. A moderate proportion of variation in joint stiffness (R(2)=0.279-0.520) was predicted by centrum width and height, neural spine angle and lamina width. These results support the possible utility of some vertebral morphometrics in predicting mechanical properties of the vertebral column in crocodiles, which also should be useful for forming functional hypotheses of axial motion during locomotion in extinct archosaurs.

Locomotor characteristics of the ground-walking chameleon Brookesia superciliaris.

Understanding the locomotor characteristics of early diverging ground-walking chameleons (members of the genera Brookesia, Rhampholeon, Palleon, and Rieppeleon) can help to explain how their unique morphology is adapted to fit their environment and mode of life. However, nearly all quantitative studies of chameleon locomotion thus far have focused on the larger "true arboreal" chameleons. We investigated kinematics and spatiotemporal gait characteristics of the Brown Leaf Chameleon (Brookesia superciliaris) on different substrates and compared them with true arboreal chameleons, nonchameleon lizards, and other small arboreal animals. Brookesia exhibits a combination of locomotor traits, some of which are traditionally arboreal, others more terrestrial, and a few that are very unusual. Like other chameleons, Brookesia moved more slowly on narrow dowels than on broad planks (simulating arboreal and terrestrial substrates, respectively), and its speed was primarily regulated by stride frequency rather than stride length. While Brookesia exhibits the traditionally arboreal trait of a high degree of humeral protraction at the beginning of stance, unlike most arboreal tetrapods, it uses smaller shoulder and hip excursions on narrower substrates, possibly reflecting its more terrestrial habits. When moving at very slow speeds, Brookesia often adopts an unusual footfall pattern, lateral-sequence lateral-couplets. Because Brookesia is a member of one of the earliest-diverging groups of chameleons, its locomotion may provide a good model for an intermediate stage in the evolution of arboreal chameleons. Thus, the transition to a fully arboreal way of life in "true arboreal" chameleons may have involved changes in spatiotemporal and kinematic characteristics as well as morphology.

Evolution of Hindlimb Muscle Anatomy Across the Tetrapod Water-to-Land Transition, Including Comparisons With Forelimb Anatomy.

Tetrapod limbs are a key innovation implicated in the evolutionary success of the clade. Although musculoskeletal evolution of the pectoral appendage across the fins-to-limbs transition is fairly well documented, that of the pelvic appendage is much less so. The skeletal elements of the pelvic appendage in some tetrapodomorph fish and the earliest tetrapods are relatively smaller and/or qualitatively less similar to those of crown tetrapods than those of the pectoral appendage. However, comparative and developmental works have suggested that the musculature of the tetrapod forelimb and hindlimb was initially very similar, constituting a "similarity bottleneck" at the fins-to-limbs transition. Here, we used extant phylogenetic bracketing and phylogenetic character optimization to reconstruct pelvic appendicular muscle anatomy in several key taxa spanning the fins-to-limbs and water-to-land transitions. Our results support the hypothesis that transformation of the pelvic appendages from fin-like to limb-like lagged behind that of the pectoral appendages. Compared to similar reconstructions of the pectoral appendages, the pelvic appendages of the earliest tetrapods had fewer muscles, particularly in the distal limb (shank). In addition, our results suggest that the first tetrapods had a greater number of muscle-muscle topological correspondences between the pectoral and pelvic appendages than tetrapodomorph fish had. However, ancestral crown-group tetrapods appear to have had an even greater number of similar muscles (both in terms of number and as a percentage of the total number of muscles), indicating that the main topological similarity bottleneck between the paired appendages may have occurred at the origin of the tetrapod crown group. Anat Rec, 2018. © 2018 Wiley Periodicals, Inc. Anat Rec, 303:218-234, 2020. © 2018 American Association for Anatomy.

Evolutionary parallelisms of pectoral and pelvic network-anatomy from fins to limbs.

Lobe-fins transformed into limbs during the Devonian period, facilitating the water-to-land transition in tetrapods. We traced the evolution of well-articulated skeletons across the fins-to-limbs transition, using a network-based approach to quantify and compare topological features of fins and limbs. We show that the topological arrangement of bones in pectoral and pelvic appendages evolved in parallel during the fins-to-limbs transition, occupying overlapping regions of the morphospace, following a directional trend, and decreasing their disparity over time. We identify the presence of digits as the morphological novelty triggering topological changes that discriminated limbs from fins. The origin of digits caused an evolutionary shift toward appendages that were less densely and heterogeneously connected, but more assortative and modular. Disparity likewise decreased for both appendages, more markedly until a time concomitant with the earliest-known tetrapod tracks. Last, we rejected the presence of a pectoral-pelvic similarity bottleneck at the origin of tetrapods.

Reconstructing pectoral appendicular muscle anatomy in fossil fish and tetrapods over the fins-to-limbs transition.

The question of how tetrapod limbs evolved from fins is one of the great puzzles of evolutionary biology. While palaeontologists, developmental biologists, and geneticists have made great strides in explaining the origin and early evolution of limb skeletal structures, that of the muscles remains largely unknown. The main reason is the lack of consensus about appendicular muscle homology between the closest living relatives of early tetrapods: lobe-finned fish and crown tetrapods. In the light of a recent study of these homologies, we re-examined osteological correlates of muscle attachment in the pectoral girdle, humerus, radius, and ulna of early tetrapods and their close relatives. Twenty-nine extinct and six extant sarcopterygians were included in a meta-analysis using information from the literature and from original specimens, when possible. We analysed these osteological correlates using parsimony-based character optimization in order to reconstruct muscle anatomy in ancestral lobe-finned fish, tetrapodomorph fish, stem tetrapods, and crown tetrapods. Our synthesis revealed that many tetrapod shoulder muscles probably were already present in tetrapodomorph fish, while most of the more-distal appendicular muscles either arose later from largely undifferentiated dorsal and ventral muscle masses or did not leave clear correlates of attachment in these taxa. Based on this review and meta-analysis, we postulate a stepwise sequence of specific appendicular muscle acquisitions, splits, and fusions that led from the ancestral sarcopterygian pectoral fin to the ancestral tetrapod forelimb. This sequence largely agrees with previous hypotheses based on palaeontological and comparative work, but it is much more comprehensive in terms of both muscles and taxa. Combined with existing information about the skeletal system, our new synthesis helps to illuminate the genetic, developmental, morphological, functional, and ecological changes that were key components of the fins-to-limbs transition.

First anatomical network analysis of fore- and hindlimb musculoskeletal modularity in bonobos, common chimpanzees, and humans.

Studies of morphological integration and modularity, and of anatomical complexity in human evolution typically focus on skeletal tissues. Here we provide the first network analysis of the musculoskeletal anatomy of both the fore- and hindlimbs of the two species of chimpanzee and humans. Contra long-accepted ideas, network analysis reveals that the hindlimb displays a pattern opposite to that of the forelimb: Pan big toe is typically seen as more independently mobile, but humans are actually the ones that have a separate module exclusively related to its movements. Different fore- vs hindlimb patterns are also seen for anatomical network complexity (i.e., complexity in the arrangement of bones and muscles). For instance, the human hindlimb is as complex as that of chimpanzees but the human forelimb is less complex than in Pan. Importantly, in contrast to the analysis of morphological integration using morphometric approaches, network analyses do not support the prediction that forelimb and hindlimb are more dissimilar in species with functionally divergent limbs such as bipedal humans.

Anatomical network analysis of the musculoskeletal system reveals integration loss and parcellation boost during the fins-to-limbs transition.

Tetrapods evolved from within the lobe-finned fishes around 370 Ma. The evolution of limbs from lobe-fins entailed a major reorganization of the skeletal and muscular anatomy of appendages in early tetrapods. Concurrently, a degree of similarity between pectoral and pelvic appendages also evolved. Here, we compared the anatomy of appendages in extant lobe-finned fishes (Latimeria and Neoceratodus) and anatomically plesiomorphic amphibians (Ambystoma, Salamandra) and amniotes (Sphenodon) to trace and reconstruct the musculoskeletal changes that took place during the fins-to-limbs transition. We quantified the anatomy of appendages using network analysis. First, we built network models-in which nodes represent bones and muscles, and links represent their anatomical connections-and then we measured network parameters related to their anatomical integration, heterogeneity, and modularity. Our results reveal an evolutionary transition toward less integrated, more modular appendages. We interpret this transition as a diversification of muscle functions in tetrapods compared to lobe-finned fishes. Limbs and lobe-fins show also a greater similarity between their pectoral and pelvic appendages than ray-fins do. These findings on extant species provide a basis for future quantitative and comprehensive reconstructions of the anatomy of limbs in early tetrapod fossils, and a way to better understand the fins-to-limbs transition.

Bonobo anatomy reveals stasis and mosaicism in chimpanzee evolution, and supports bonobos as the most appropriate extant model for the common ancestor of chimpanzees and humans.

Common chimps and bonobos are our closest living relatives but almost nothing is known about bonobo internal anatomy. We present the first phylogenetic analysis to include musculoskeletal data obtained from a recent dissection of bonobos. Notably, chimpanzees, and in particular bonobos, provide a remarkable case of evolutionary stasis for since the chimpanzee-human split c.8 Ma among >120 head-neck (HN) and forelimb (FL) muscles there were only four minor changes in the chimpanzee clade, and all were reversions to the ancestral condition. Moreover, since the common chimpanzee-bonobo split c.2 Ma there have been no changes in bonobos, so with respect to HN-FL musculature bonobos are the better model for the last common ancestor (LCA) of chimpanzees/bonobos and humans. Moreover, in the hindlimb there are only two muscle absence/presence differences between common chimpanzees and bonobos. Puzzlingly, there is an evolutionary mosaicism between each of these species and humans. We discuss these data in the context of available genomic information and debates on whether the common chimpanzee-bonobo divergence is linked to heterochrony.

Comparative musculoskeletal anatomy of chameleon limbs, with implications for the evolution of arboreal locomotion in lizards and for teratology.

Chameleon species have recently been adopted as models for evo-devo and macroevolutionary processes. However, most anatomical and developmental studies of chameleons focus on the skeleton, and information about their soft tissues is scarce. Here, we provide a detailed morphological description based on contrast enhanced micro-CT scans and dissections of the adult phenotype of all the forelimb and hindlimb muscles of the Veiled Chameleon (Chamaeleo calyptratus) and compare these muscles with those of other chameleons and lizards. We found the appendicular muscle anatomy of chameleons to be surprisingly conservative considering the remarkable structural and functional modifications of the limb skeleton, particularly the distal limb regions. For instance, the zygodactyl autopodia of chameleons are unique among tetrapods, and the carpals and tarsals are highly modified in shape and number. However, most of the muscles usually present in the manus and pes of other lizards are present in the same configuration in chameleons. The most obvious muscular features related to the peculiar opposable autopodia of chameleons are: (1) presence of broad, V-shaped plantar and palmar aponeuroses, and absence of intermetacarpales and intermetatarsales, between the digits separated by the cleft in each autopod; (2) oblique orientation of the superficial short flexors originating from these aponeuroses, which may allow these muscles to act as powerful adductors of the "super-digits"; and (3) well-developed abductor digiti minimi muscles and abductor pollicis/hallucis brevis muscles, which may act as powerful abductors of the "super-digits."

Characteristic tetrapod musculoskeletal limb phenotype emerged more than 400 MYA in basal lobe-finned fishes.

Previous accounts of the origin of tetrapod limbs have postulated a relatively sudden change, after the split between extant lobe-finned fish and tetrapods, from a very simple fin phenotype with only two muscles to the highly complex tetrapod condition. The evolutionary changes that led to the muscular anatomy of tetrapod limbs have therefore remained relatively unexplored. We performed dissections, histological sections, and MRI scans of the closest living relatives of tetrapods: coelacanths and lungfish. Combined with previous comparative, developmental and paleontological information, our findings suggest that the characteristic tetrapod musculoskeletal limb phenotype was already present in the Silurian last common ancestor of extant sarcopterygians, with the exception of the autopod (hand/foot) structures, which have no clear correspondence with fish structures. Remarkably, the two major steps in this long process - leading to the ancestral fin anatomy of extant sarcopterygians and limb anatomy of extant tetrapods, respectively - occurred at the same nodes as the two major similarity bottlenecks that led to the striking derived myological similarity between the pectoral and pelvic appendages within each taxon. Our identification of probable homologies between appendicular muscles of sarcopterygian fish and tetrapods will allow more detailed reconstructions of muscle anatomy in early tetrapods and their relatives.

Comparative Myology and Evolution of Marsupials and Other Vertebrates, With Notes on Complexity, Bauplan, and "Scala Naturae".

Opossums are frequent subjects of developmental studies because marsupials share developmental features not seen in placentals and because Didelphimorpha is the sister-group of other extant Marsupialia. But is the adult marsupial muscular system markedly different from that of placentals or is it, like the skeletal system, very similar? We provide, for the first time, a brief description of all head and limb muscles of Didelphis virginiana based on our dissections and using a unifying nomenclature by integrating the data gathered in our long-term project on the development, homologies, and evolution of the muscles of all major vertebrate taxa. Our data indicate that there were many more muscle synapomorphic changes from the last common ancestor (LCA) of amniotes to the mammalian LCA (63) and from this LCA to the LCA of extant therians (48) than from this latter LCA to the LCA of extant placentals (10 or 11). Importantly, Didelphis is anatomically more plesiomorphic (only 14 changes from LCA of extant therians) than are rats (37 changes) and humans (63 changes), but its musculature is more complex (193 muscles) than that of humans (only 180 muscles). Of the 194 muscles of Didelphis, 172 (89%) are present in rats, meaning that their adult muscle anatomy is indeed very similar. This similarity supports the existence of a common, easy recognizable therian Bauplan, but one that is caused by developmental constraints and by evolutionary change driven by the needs of the embryos/neonates, rather than by a "goal" toward a specific adult plan/"archetype," as the name Bauplan suggests. Anat Rec, 299:1224-1255, 2016. © 2016 Wiley Periodicals, Inc.

Morphological and functional changes in the vertebral column with increasing aquatic adaptation in crocodylomorphs.

The lineage leading to modern Crocodylia has undergone dramatic evolutionary changes in morphology, ecology and locomotion over the past 200+ Myr. These functional innovations may be explained in part by morphological changes in the axial skeleton, which is an integral part of the vertebrate locomotor system. Our objective was to estimate changes in osteological range of motion (RoM) and intervertebral joint stiffness of thoracic and lumbar vertebrae with increasing aquatic adaptation in crocodylomorphs. Using three-dimensional virtual models and morphometrics, we compared the modern crocodile Crocodylus to five extinct crocodylomorphs: Terrestrisuchus, Protosuchus, Pelagosaurus, Steneosaurus and Metriorhynchus, which span the spectrum from terrestrial to fully aquatic. In Crocodylus, we also experimentally measured changes in trunk flexibility with sequential removal of osteoderms and soft tissues. Our results for the more aquatic species matched our predictions fairly well, but those for the more terrestrial early crocodylomorphs did not. A likely explanation for this lack of correspondence is the influence of other axial structures, particularly the rigid series of dorsal osteoderms in early crocodylomorphs. The most important structures for determining RoM and stiffness of the trunk in Crocodylus were different in dorsoventral versus mediolateral bending, suggesting that changes in osteoderm and rib morphology over crocodylomorph evolution would have affected movements in some directions more than others.

The anatomy and ontogeny of the head, neck, pectoral, and upper limb muscles of Lemur catta and Propithecus coquereli (primates): discussion on the parallelism between ontogeny and phylogeny and implications for evolutionary and developmental biology.

Most anatomical studies of primates focus on skeletal tissues, but muscular anatomy can provide valuable information about phylogeny, functional specializations, and evolution. Herein, we present the first detailed description of the head, neck, pectoral, and upper limb muscles of the fetal lemuriforms Lemur catta (Lemuridae) and Propithecus coquereli (Indriidae). These two species belong to the suborder Strepsirrhini, which is often presumed to possess some plesiomorphic anatomical features within primates. We compare the muscular anatomy of the fetuses with that of infants and adults and discuss the evolutionary and developmental implications. The fetal anatomy reflects a phylogenetically more plesiomorphic condition in nine of the muscles we studied and a more derived condition in only two, supporting a parallel between ontogeny and phylogeny. The derived exceptions concern muscles with additional insertions in the fetus which are lost in adults of the same species, that is, flexor carpi radialis inserts on metacarpal III and levator claviculae inserts on the clavicle. Interestingly, these two muscles are involved in movements of the pectoral girdle and upper limb, which are mainly important for activities in later stages of life, such as locomotion and prey capture, rather than activities in fetal life. Accordingly, our findings suggest that some exceptions to the "ontogeny parallels phylogeny" rule are probably driven more by ontogenetic constraints than by adaptive plasticity.