Identification of multiple transcription factor genes potentially involved in the development of electrosensory versus mechanosensory lateral line organs.

In electroreceptive jawed vertebrates, embryonic lateral line placodes give rise to electrosensory ampullary organs as well as mechanosensory neuromasts. Previous reports of shared gene expression suggest that conserved mechanisms underlie electroreceptor and mechanosensory hair cell development and that electroreceptors evolved as a transcriptionally related "sister cell type" to hair cells. We previously identified only one transcription factor gene, Neurod4, as ampullary organ-restricted in the developing lateral line system of a chondrostean ray-finned fish, the Mississippi paddlefish (Polyodon spathula). The other 16 transcription factor genes we previously validated in paddlefish were expressed in both ampullary organs and neuromasts. Here, we used our published lateral line organ-enriched gene-set (arising from differential bulk RNA-seq in late-larval paddlefish), together with a candidate gene approach, to identify 25 transcription factor genes expressed in the developing lateral line system of a more experimentally tractable chondrostean, the sterlet (Acipenser ruthenus, a small sturgeon), and/or that of paddlefish. Thirteen are expressed in both ampullary organs and neuromasts, consistent with conservation of molecular mechanisms. Seven are electrosensory-restricted on the head (Irx5, Irx3, Insm1, Sp5, Satb2, Mafa and Rorc), and five are the first-reported mechanosensory-restricted transcription factor genes (Foxg1, Sox8, Isl1, Hmx2 and Rorb). However, as previously reported, Sox8 is expressed in ampullary organs as well as neuromasts in a catshark (Scyliorhinus canicula), suggesting the existence of lineage-specific differences between cartilaginous and ray-finned fishes. Overall, our results support the hypothesis that ampullary organs and neuromasts develop via largely conserved transcriptional mechanisms, and identify multiple transcription factors potentially involved in the formation of electrosensory versus mechanosensory lateral line organs.

Snakebite epidemiology, outcomes and multi-cluster risk modelling in Eswatini.

BACKGROUND: Halving snakebite morbidity and mortality by 2030 requires countries to develop both prevention and treatment strategies. The paucity of data on the global incidence and severity of snakebite envenoming causes challenges in prioritizing and mobilising resources for snakebite prevention and treatment. In line with the World Health Organisation's 2019 Snakebite Strategy, this study sought to investigate Eswatini's snakebite epidemiology and outcomes, and identify the socio-geographical factors associated with snakebite risk. METHODOLOGY: Programmatic data from the Ministry of Health, Government of Eswatini 2019-2021, was used to assess the epidemiology and outcomes of snakebite in Eswatini. We developed a snake species richness map from the occurrence data of all venomous snakes of medical importance in Eswatini that was subjected to niche modelling. We formulated four risk indices using snake species richness, various geospatial datasets and reported snakebites. A multivariate cluster modelling approach using these indices was developed to estimate risk of snakebite and the outcomes of snakebite in Eswatini. PRINCIPAL FINDINGS: An average of 466 snakebites was recorded annually in Eswatini. Bites were recorded across the entire country and peaked in the evening during summer months. Two cluster risk maps indicated areas of the country with a high probability of snakebite and a high probability of poor snakebite outcomes. The areas with the highest rate of snakebite risk were primarily in the rural and agricultural regions of the country. SIGNIFICANCE: These models can be used to inform better snakebite prevention and treatment measures to enable Eswatini to meet the global goal of reducing snakebite morbidity and mortality by 50% by 2030. The supply chain challenges of antivenom affecting southern Africa and the high rates of snakebite identified in our study highlight the need for improved snakebite prevention and treatment tools that can be employed by health care workers stationed at rural, community clinics.

Delays, fears and training needs: Perspectives of health workers on clinical management of snakebite revealed by a qualitative study in Kitui County, Kenya.

Key aims of the WHO Strategy to halve snakebite morbidity and mortality include health system strengthening and training of health workers. This requires knowledge of local health system needs and capacity, health worker training needs, and factors influencing health worker decision-making in snakebite management. This study explored health worker experiences and perceptions of snakebite management, both individually and in the context of their local health system. We used a qualitative study design with semi-structured interviews (n = 14) and focus group discussions (n = 4). We employed a combination of sampling strategies aiming to achieve maximum variation among key informants within resource limitations. We recruited health workers (n = 33) of varying roles from purposively selected tier 2, 3 and 4 health facilities (n = 12) and the community (tier 1) in four sub-counties in Kitui County, Kenya. We conducted inductive thematic analysis of all transcripts. The results identified that health workers recognised snake envenoming as a time-critical emergency in which delay in care seeking, sometimes exacerbated by health system referral delays, was a major barrier to effective management of patients. Clinicians strongly voiced a need for training in snakebite management, diagnosis and antivenom administration. Unexpressed needs for training were demonstrated in traditional remedy ineffectiveness, syndromic management, and critical appraisal of treatment effectiveness. Under-resourcing in antivenom, other medication, equipment, infrastructure and staffing also challenged management. Fear of snakebite and fear of antivenom, both linked to past experiences, influenced clinical decision-making. Our findings clearly indicate a need in Kitui County for training programmes that equip health workers for clinical decision-making in snakebite management. We further identify community intervention needs to facilitate prompt presentation to healthcare, including practical affordable transport solutions, and systematic health system resourcing needs. In addition, we recommend supportive supervision and further research in response to the emotional stress resulting from managing difficult cases in under-resourced settings.

Insights into olfactory ensheathing cell development from a laser-microdissection and transcriptome-profiling approach.

Olfactory ensheathing cells (OECs) are neural crest-derived glia that ensheath bundles of olfactory axons from their peripheral origins in the olfactory epithelium to their central targets in the olfactory bulb. We took an unbiased laser microdissection and differential RNA-seq approach, validated by in situ hybridization, to identify candidate molecular mechanisms underlying mouse OEC development and differences with the neural crest-derived Schwann cells developing on other peripheral nerves. We identified 25 novel markers for developing OECs in the olfactory mucosa and/or the olfactory nerve layer surrounding the olfactory bulb, of which 15 were OEC-specific (that is, not expressed by Schwann cells). One pan-OEC-specific gene, Ptprz1, encodes a receptor-like tyrosine phosphatase that blocks oligodendrocyte differentiation. Mutant analysis suggests Ptprz1 may also act as a brake on OEC differentiation, and that its loss disrupts olfactory axon targeting. Overall, our results provide new insights into OEC development and the diversification of neural crest-derived glia.

Olfactory ensheathing cells abutting the embryonic olfactory bulb express Frzb, whose deletion disrupts olfactory axon targeting.

We and others previously showed that in mouse embryos lacking the transcription factor Sox10, olfactory ensheathing cell (OEC) differentiation is disrupted, resulting in defective olfactory axon targeting and fewer gonadotropin-releasing hormone (GnRH) neurons entering the embryonic forebrain. The underlying mechanisms are unclear. Here, we report that OECs in the olfactory nerve layer express Frzb-encoding a secreted Wnt inhibitor with roles in axon targeting and basement membrane breakdown-from embryonic day (E)12.5, when GnRH neurons first enter the forebrain, until E16.5, the latest stage examined. The highest levels of Frzb expression are seen in OECs in the inner olfactory nerve layer, abutting the embryonic olfactory bulb. We find that Sox10 is required for Frzb expression in OECs, suggesting that loss of Frzb could explain the olfactory axon targeting and/or GnRH neuron migration defects seen in Sox10-null mice. At E16.5, Frzb-null embryos show significant reductions in both the volume of the olfactory nerve layer expressing the maturation marker Omp and the number of Omp-positive olfactory receptor neurons in the olfactory epithelium. As Omp upregulation correlates with synapse formation, this suggests that Frzb deletion indeed disrupts olfactory axon targeting. In contrast, GnRH neuron entry into the forebrain is not significantly affected. Hence, loss of Frzb may contribute to the olfactory axon targeting phenotype, but not the GnRH neuron phenotype, of Sox10-null mice. Overall, our results suggest that Frzb secreted from OECs in the olfactory nerve layer is important for olfactory axon targeting.

Neural crest Notch/Rbpj signaling regulates olfactory gliogenesis and neuronal migration.

The neural crest-derived ensheathing glial cells of the olfactory nerve (OECs) are unique in spanning both the peripheral and central nervous systems: they ensheathe bundles of axons projecting from olfactory receptor neurons in the nasal epithelium to their targets in the olfactory bulb. OECs are clinically relevant as a promising autologous cell transplantation therapy for promoting central nervous system repair. They are also important for fertility, being required for the migration of embryonic gonadotropin-releasing hormone (GnRH) neurons from the olfactory placode along terminal nerve axons to the medial forebrain, which they enter caudal to the olfactory bulbs. Like Schwann cell precursors, OEC precursors associated with the developing olfactory nerve express the glial marker myelin protein zero and the key peripheral glial transcription factor Sox10. The transition from Schwann cell precursors to immature Schwann cells is accelerated by canonical Notch signaling via the Rbpj transcription factor. Here, we aimed to test the role of Notch/Rbpj signaling in developing OECs by blocking the pathway in both chicken and mouse. Our results suggest that Notch/Rbpj signaling prevents the cranial neural crest cells that colonize the olfactory nerve from differentiating as neurons, and at later stages contributes to the guidance of GnRH neurons.

Insights into Electroreceptor Development and Evolution from Molecular Comparisons with Hair Cells.

The vertebrate lateral line system comprises a mechanosensory division, with neuromasts containing hair cells that detect local water movement ("distant touch"); and an electrosensory division, with electrosensory organs that detect the weak, low-frequency electric fields surrounding other animals in water (primarily used for hunting). The entire lateral line system was lost in the amniote lineage with the transition to fully terrestrial life; the electrosensory division was lost independently in several lineages, including the ancestors of frogs and of teleost fishes. (Electroreception with different characteristics subsequently evolved independently within two teleost lineages.) Recent gene expression studies in a non-teleost actinopterygian fish suggest that electroreceptor ribbon synapses employ the same transmission mechanisms as hair cell ribbon synapses, and show that developing electrosensory organs express transcription factors essential for hair cell development, including Atoh1 and Pou4f3. Previous hypotheses for electroreceptor evolution suggest either that electroreceptors and hair cells evolved independently in the vertebrate ancestor from a common ciliated secondary cell, or that electroreceptors evolved from hair cells. The close developmental and putative physiological similarities implied by the gene expression data support the latter hypothesis, i.e., that electroreceptors evolved in the vertebrate ancestor as a "sister cell-type" to lateral line hair cells.

Striking parallels between carotid body glomus cell and adrenal chromaffin cell development.

Carotid body glomus cells mediate essential reflex responses to arterial blood hypoxia. They are dopaminergic and secrete growth factors that support dopaminergic neurons, making the carotid body a potential source of patient-specific cells for Parkinson's disease therapy. Like adrenal chromaffin cells, which are also hypoxia-sensitive, glomus cells are neural crest-derived and require the transcription factors Ascl1 and Phox2b; otherwise, their development is little understood at the molecular level. Here, analysis in chicken and mouse reveals further striking molecular parallels, though also some differences, between glomus and adrenal chromaffin cell development. Moreover, histology has long suggested that glomus cell precursors are 'émigrés' from neighbouring ganglia/nerves, while multipotent nerve-associated glial cells are now known to make a significant contribution to the adrenal chromaffin cell population in the mouse. We present conditional genetic lineage-tracing data from mice supporting the hypothesis that progenitors expressing the glial marker proteolipid protein 1, presumably located in adjacent ganglia/nerves, also contribute to glomus cells. Finally, we resolve a paradox for the 'émigré' hypothesis in the chicken - where the nearest ganglion to the carotid body is the nodose, in which the satellite glia are neural crest-derived, but the neurons are almost entirely placode-derived - by fate-mapping putative nodose neuronal 'émigrés' to the neural crest.

Notch and Fgf signaling during electrosensory versus mechanosensory lateral line organ development in a non-teleost ray-finned fish.

The lateral line system is a useful model for studying the embryonic and evolutionary diversification of different organs and cell types. In jawed vertebrates, this ancestrally comprises lines of mechanosensory neuromasts over the head and trunk, flanked on the head by fields of electrosensory ampullary organs, all innervated by lateral line neurons in cranial lateral line ganglia. Both types of sense organs, and their afferent neurons, develop from cranial lateral line placodes. Current research primarily focuses on the posterior lateral line primordium in zebrafish, which migrates as a cell collective along the trunk; epithelial rosettes form in the trailing zone and are deposited as a line of neuromasts, within which hair cells and supporting cells differentiate. However, in at least some other teleosts (e.g. catfishes) and all non-teleosts, lines of cranial neuromasts are formed by placodes that elongate to form a sensory ridge, which subsequently fragments, with neuromasts differentiating in a line along the crest of the ridge. Furthermore, in many non-teleost species, electrosensory ampullary organs develop from the flanks of the sensory ridge. It is unknown to what extent the molecular mechanisms underlying neuromast formation from the zebrafish migrating posterior lateral line primordium are conserved with the as-yet unexplored molecular mechanisms underlying neuromast and ampullary organ formation from elongating lateral line placodes. Here, we report experiments in an electroreceptive non-teleost ray-finned fish, the Mississippi paddlefish Polyodon spathula, that suggest a conserved role for Notch signaling in regulating lateral line organ receptor cell number, but potentially divergent roles for the fibroblast growth factor signaling pathway, both between neuromasts and ampullary organs, and between paddlefish and zebrafish.

Evolution of the hypoxia-sensitive cells involved in amniote respiratory reflexes.

The evolutionary origins of the hypoxia-sensitive cells that trigger amniote respiratory reflexes - carotid body glomus cells, and 'pulmonary neuroendocrine cells' (PNECs) - are obscure. Homology has been proposed between glomus cells, which are neural crest-derived, and the hypoxia-sensitive 'neuroepithelial cells' (NECs) of fish gills, whose embryonic origin is unknown. NECs have also been likened to PNECs, which differentiate in situ within lung airway epithelia. Using genetic lineage-tracing and neural crest-deficient mutants in zebrafish, and physical fate-mapping in frog and lamprey, we find that NECs are not neural crest-derived, but endoderm-derived, like PNECs, whose endodermal origin we confirm. We discover neural crest-derived catecholaminergic cells associated with zebrafish pharyngeal arch blood vessels, and propose a new model for amniote hypoxia-sensitive cell evolution: endoderm-derived NECs were retained as PNECs, while the carotid body evolved via the aggregation of neural crest-derived catecholaminergic (chromaffin) cells already associated with blood vessels in anamniote pharyngeal arches.

Insights into electrosensory organ development, physiology and evolution from a lateral line-enriched transcriptome.

The anamniote lateral line system, comprising mechanosensory neuromasts and electrosensory ampullary organs, is a useful model for investigating the developmental and evolutionary diversification of different organs and cell types. Zebrafish neuromast development is increasingly well understood, but neither zebrafish nor Xenopus is electroreceptive and our molecular understanding of ampullary organ development is rudimentary. We have used RNA-seq to generate a lateral line-enriched gene-set from late-larval paddlefish (Polyodon spathula). Validation of a subset reveals expression in developing ampullary organs of transcription factor genes critical for hair cell development, and genes essential for glutamate release at hair cell ribbon synapses, suggesting close developmental, physiological and evolutionary links between non-teleost electroreceptors and hair cells. We identify an ampullary organ-specific proneural transcription factor, and candidates for the voltage-sensing L-type Cav channel and rectifying Kv channel predicted from skate (cartilaginous fish) ampullary organ electrophysiology. Overall, our results illuminate ampullary organ development, physiology and evolution.

Constitutively active Notch1 converts cranial neural crest-derived frontonasal mesenchyme to perivascular cells in vivo.

Perivascular/mural cells originate from either the mesoderm or the cranial neural crest. Regardless of their origin, Notch signalling is necessary for their formation. Furthermore, in both chicken and mouse, constitutive Notch1 activation (via expression of the Notch1 intracellular domain) is sufficient in vivo to convert trunk mesoderm-derived somite cells to perivascular cells, at the expense of skeletal muscle. In experiments originally designed to investigate the effect of premature Notch1 activation on the development of neural crest-derived olfactory ensheathing glial cells (OECs), we used in ovo electroporation to insert a tetracycline-inducible NotchΔE construct (encoding a constitutively active mutant of mouse Notch1) into the genome of chicken cranial neural crest cell precursors, and activated NotchΔE expression by doxycycline injection at embryonic day 4. NotchΔE-targeted cells formed perivascular cells within the frontonasal mesenchyme, and expressed a perivascular marker on the olfactory nerve. Hence, constitutively activating Notch1 is sufficient in vivo to drive not only somite cells, but also neural crest-derived frontonasal mesenchyme and perhaps developing OECs, to a perivascular cell fate. These results also highlight the plasticity of neural crest-derived mesenchyme and glia.

The origin and evolution of cell types.

Cell types are the basic building blocks of multicellular organisms and are extensively diversified in animals. Despite recent advances in characterizing cell types, classification schemes remain ambiguous. We propose an evolutionary definition of a cell type that allows cell types to be delineated and compared within and between species. Key to cell type identity are evolutionary changes in the 'core regulatory complex' (CoRC) of transcription factors, that make emergent sister cell types distinct, enable their independent evolution and regulate cell type-specific traits termed apomeres. We discuss the distinction between developmental and evolutionary lineages, and present a roadmap for future research.

Evidence for a Notch1-mediated transition during olfactory ensheathing cell development.

Olfactory ensheathing cells (OECs) are a unique glial population found in both the peripheral and central nervous system: they ensheath bundles of unmyelinated olfactory axons from their peripheral origin in the olfactory epithelium to their central synaptic targets in the glomerular layer of the olfactory bulb. Like all other peripheral glia (Schwann cells, satellite glia, enteric glia), OECs are derived from the embryonic neural crest. However, in contrast to Schwann cells, whose development has been extensively characterised, relatively little is known about their normal development in vivo. In the Schwann cell lineage, the transition from multipotent Schwann cell precursor to immature Schwann cell is promoted by canonical Notch signalling. Here, in situ hybridisation and immunohistochemistry data from chicken, mouse and human embryos are presented that suggest a canonical Notch-mediated transition also occurs during OEC development.

The evolution of the vertebrate cerebellum: absence of a proliferative external granule layer in a non-teleost ray-finned fish.

The cerebellum represents one of the most morphologically variable structures in the vertebrate brain. To shed light on its evolutionary history, we have examined the molecular anatomy and proliferation of the developing cerebellum of the North American paddlefish, Polyodon spathula. Absence of an external proliferative cerebellar layer and the restriction of Atonal1 expression to the rhombic lip and valvular primordium demonstrate that transit amplification in a cerebellar external germinal layer, a prominent feature of amniote cerebellum development, is absent in paddlefish. Furthermore, expression of Sonic hedgehog, which drives secondary proliferation in the mouse cerebellum, is absent from the paddlefish cerebellum. These data are consistent with what has been observed in zebrafish and suggest that the transit amplification seen in the amniote cerebellum was either lost very early in the ray-finned fish lineage or evolved in the lobe-finned fish lineage. We also suggest that the Atoh1-positive proliferative valvular primordium may represent a synapomorphy (shared derived character) of ray-finned fishes. The topology of valvular primordium development in paddlefish differs significantly from that of zebrafish and correlates with the adult cerebellar form. The distribution of proliferative granule cell precursors in different vertebrate taxa is thus the likely determining factor in cerebellar morphological diversity.

The development of lateral line placodes: taking a broader view.

The lateral line system of anamniote vertebrates enables the detection of local water movement and weak bioelectric fields. Ancestrally, it comprises neuromasts - small sense organs containing mechanosensory hair cells - distributed in characteristic lines over the head and trunk, flanked on the head by fields of electroreceptive ampullary organs, innervated by afferent neurons projecting respectively to the medial and dorsal octavolateral nuclei in the hindbrain. Given the independent loss of the electrosensory system in multiple lineages, the development and evolution of the mechanosensory and electrosensory components of the lateral line must be dissociable. Nevertheless, the entire system arises from a series of cranial lateral line placodes, which exhibit two modes of sensory organ formation: elongation to form sensory ridges that fragment (with neuromasts differentiating in the center of the ridge, and ampullary organs on the flanks), or migration as collectives of cells, depositing sense organs in their wake. Intensive study of the migrating posterior lateral line placode in zebrafish has yielded a wealth of information concerning the molecular control of migration and neuromast formation in this migrating placode, in this cypriniform teleost species. However, our mechanistic understanding of neuromast and ampullary organ formation by elongating lateral line placodes, and even of other zebrafish lateral line placodes, is sparse or non-existent. Here, we attempt to highlight the diversity of lateral line development and the limits of the current research focus on the zebrafish posterior lateral line placode. We hope this will stimulate a broader approach to this fascinating sensory system.

A fate-map for cranial sensory ganglia in the sea lamprey.

Cranial neurogenic placodes and the neural crest make essential contributions to key adult characteristics of all vertebrates, including the paired peripheral sense organs and craniofacial skeleton. Neurogenic placode development has been extensively characterized in representative jawed vertebrates (gnathostomes) but not in jawless fishes (agnathans). Here, we use in vivo lineage tracing with DiI, together with neuronal differentiation markers, to establish the first detailed fate-map for placode-derived sensory neurons in a jawless fish, the sea lamprey Petromyzon marinus, and to confirm that neural crest cells in the lamprey contribute to the cranial sensory ganglia. We also show that a pan-Pax3/7 antibody labels ophthalmic trigeminal (opV, profundal) placode-derived but not maxillomandibular trigeminal (mmV) placode-derived neurons, mirroring the expression of gnathostome Pax3 and suggesting that Pax3 (and its single Pax3/7 lamprey ortholog) is a pan-vertebrate marker for opV placode-derived neurons. Unexpectedly, however, our data reveal that mmV neuron precursors are located in two separate domains at neurula stages, with opV neuron precursors sandwiched between them. The different branches of the mmV nerve are not comparable between lampreys and gnatho-stomes, and spatial segregation of mmV neuron precursor territories may be a derived feature of lampreys. Nevertheless, maxillary and mandibular neurons are spatially segregated within gnathostome mmV ganglia, suggesting that a more detailed investigation of gnathostome mmV placode development would be worthwhile. Overall, however, our results highlight the conservation of cranial peripheral sensory nervous system development across vertebrates, yielding insight into ancestral vertebrate traits.

Olfactory ensheathing glia are required for embryonic olfactory axon targeting and the migration of gonadotropin-releasing hormone neurons.

Kallmann's syndrome is caused by the failure of olfactory axons and gonadotropin-releasing hormone (GnRH) neurons to enter the embryonic forebrain, resulting in anosmia and sterility. Sox10 mutations have been associated with Kallmann's syndrome phenotypes, but their effect on olfactory system development is unknown. We recently showed that Sox10 is expressed by neural crest-derived olfactory ensheathing cells (OECs). Here, we demonstrate that in homozygous Sox10(lacZ/lacZ) mouse embryos, OEC differentiation is disrupted; olfactory axons accumulate in the ventromedial olfactory nerve layer and fewer olfactory receptor neurons express the maturation marker OMP (most likely owing to the failure of axonal targeting). Furthermore, GnRH neurons clump together in the periphery and a smaller proportion enters the forebrain. Our data suggest that human Sox10 mutations cause Kallmann's syndrome by disrupting the differentiation of OECs, which promote embryonic olfactory axon targeting and hence olfactory receptor neuron maturation, and GnRH neuron migration to the forebrain.

The evolution and development of vertebrate lateral line electroreceptors.

Electroreception is an ancient vertebrate sense with a fascinating evolutionary history involving multiple losses as well as independent evolution at least twice within teleosts. We review the phylogenetic distribution of electroreception and the morphology and innervation of electroreceptors in different vertebrate groups. We summarise recent work from our laboratory that has confirmed the homology of ampullary electroreceptors in non-teleost jawed vertebrates by showing, in conjunction with previously published work, that these are derived embryonically from lateral line placodes. Finally, we review hypotheses to explain the distribution of electroreception within teleosts, including the hypothesis that teleost ampullary and tuberous electroreceptors evolved via the modification of mechanosensory hair cells in lateral line neuromasts. We conclude that further experimental work on teleost electroreceptor development is needed to test such hypotheses.

Developmental evidence for serial homology of the vertebrate jaw and gill arch skeleton.

Gegenbaur's classical hypothesis of jaw-gill arch serial homology is widely cited, but remains unsupported by either palaeontological evidence (for example, a series of fossils reflecting the stepwise transformation of a gill arch into a jaw) or developmental genetic data (for example, shared molecular mechanisms underlying segment identity in the mandibular, hyoid and gill arch endoskeletons). Here we show that nested expression of Dlx genes--the 'Dlx code' that specifies upper and lower jaw identity in mammals and teleosts--is a primitive feature of the mandibular, hyoid and gill arches of jawed vertebrates. Using fate-mapping techniques, we demonstrate that the principal dorsal and ventral endoskeletal segments of the jaw, hyoid and gill arches of the skate Leucoraja erinacea derive from molecularly equivalent mesenchymal domains of combinatorial Dlx gene expression. Our data suggest that vertebrate jaw, hyoid and gill arch cartilages are serially homologous, and were primitively patterned dorsoventrally by a common Dlx blueprint.