Development and regeneration of the crushing dentition of skates (Rajidae)

Sharks and rays (elasmobranchs) have the remarkable capacity to continuously regenerate their teeth. The polyphyodont system is considered the ancestral condition of the gnathostome dentition. Despite this shared regenerative ability, sharks and rays exhibit dramatic interspecific variation in their tooth morphology. Ray (batoidea) teeth typically constitute crushing pads of flattened teeth, whereas shark teeth are pointed, multi-cuspid units. Although recent research has addressed the molecular development of the shark dentition, little is known about that of the ray. Furthermore, how dental diversity within the elasmobranch lineage is achieved remains unknown. Here, we examine dental development and regeneration in two Batoid species: the thornback skate (Raja clavata) and the little skate (Leucoraja erinacea). Using in situ hybridization and immunohistochemistry, we examine the expression of a core gnathostome dental gene set during early development of the skate dentition and compare it to development in the shark. Elasmobranch tooth development is highly conserved, with sox2 likely playing an important role in the initiation and regeneration of teeth. Alterations to conserved genes expressed in an enamel knot-like signalling centre may explain the morphological diversity of elasmobranch teeth, thereby enabling sharks and rays to occupy diverse dietary and ecological niches

Development and evolution of dentition pattern and tooth order in the Skates and Rays (Batoidea; Chondrichthyes)

Shark and ray (elasmobranch) dentitions are well known for their multiple generations of teeth, with isolated teeth being common in the fossil record. However, how the diverse dentitions characteristic of elasmobranchs form is still poorly understood. Data on the development and maintenance of the dental patterning in this major vertebrate group will allow comparisons to other morphologically diverse taxa, including the bony fishes, in order to identify shared pattern characters for the vertebrate dentition as a whole. Data is especially lacking from the Batoidea (skates and rays), hence our objective is to compile data on embryonic and adult batoid tooth development contributing to ordering of the dentition, from cleared and stained specimens and micro-CT scans, with 3D rendered models. We selected species (adult and embryonic) spanning phylogenetically significant batoid clades, such that our observations may raise questions about relationships within the batoids, particularly with respect to current molecular-based analyses. We include developmental data from embryos of recent model organisms Leucoraja erinacea and Raja clavata to evaluate the earliest establishment of the dentition. Characters of the batoid dentition investigated include alternate addition of teeth as offset successional tooth rows (versus single separate files), presence of a symphyseal initiator region (symphyseal tooth present, or absent, but with two parasymphyseal teeth) and a restriction to tooth addition along each jaw reducing the number of tooth families, relative to addition of successor teeth within each family. Our ultimate aim is to understand the shared characters of the batoids, and whether or not these dental characters are shared more broadly within elasmobranchs, by comparing these to dentitions in shark outgroups. These developmental morphological analyses will provide a solid basis to better understand dental evolution in these important vertebrate groups as well as the general plesiomorphic vertebrate dental condition

An ancient dental gene network regulates development and continuous regeneration of teeth in sharks

The appearance of toothed vertebrates has proven a major determinant of the overall success of this lineage. This is most apparent in sharks and rays (elasmobranchs), which further retain the capacity for life-long tooth regeneration. Given their comparatively basal phylogenetic position, elasmobranchs therefore offer the opportunity for crucial insights into putative ancestral characters of tooth development, yet despite their evolutionary significance this remains poorly understood. Using the established chondrichthyan model, the catshark (Scyliorhinus sp.), we identified the expression of genes representative of conserved signaling pathways during stages of early dental competence, tooth initiation and regeneration. The expression patterns of β-catenin, shh, bmp4, pax9, pitx1/2, and the stem cell marker Sox2, characterise an ancestrally conserved gene set deployed during initiation of the elasmobranch dentition, suggesting that all vertebrate dentitions are defined by the expression of this core set of genes. These findings provide novel evidence to support the conservation in deep evolutionary time of a core set of dental patterning genes, therefore further defining the evolutionary trajectory of tooth development. We show how these genes facilitate the emergence of the shark dentition and offer insights into their deployment during development of the dental lamina, a sheet of dental epithelial cells that are responsible for continuous tooth regeneration. This study further promotes a specific experimental agenda to further characterise the roles of these core developmental genes during vertebrate tooth development, and importantly dental regeneration

25th annual computational neuroscience meeting: CNS-2016

The same neuron may play different functional roles in the neural circuits to which it belongs. For example, neurons in the Tritonia pedal ganglia may participate in variable phases of the swim motor rhythms [1]. While such neuronal functional variability is likely to play a major role the delivery of the functionality of neural systems, it is difficult to study it in most nervous systems. We work on the pyloric rhythm network of the crustacean stomatogastric ganglion (STG) [2]. Typically network models of the STG treat neurons of the same functional type as a single model neuron (e.g. PD neurons), assuming the same conductance parameters for these neurons and implying their synchronous firing [3, 4]. However, simultaneous recording of PD neurons shows differences between the timings of spikes of these neurons. This may indicate functional variability of these neurons. Here we modelled separately the two PD neurons of the STG in a multi-neuron model of the pyloric network. Our neuron models comply with known correlations between conductance parameters of ionic currents. Our results reproduce the experimental finding of increasing spike time distance between spikes originating from the two model PD neurons during their synchronised burst phase. The PD neuron with the larger calcium conductance generates its spikes before the other PD neuron. Larger potassium conductance values in the follower neuron imply longer delays between spikes, see Fig. 17.Neuromodulators change the conductance parameters of neurons and maintain the ratios of these parameters [5]. Our results show that such changes may shift the individual contribution of two PD neurons to the PD-phase of the pyloric rhythm altering their functionality within this rhythm. Our work paves the way towards an accessible experimental and computational framework for the analysis of the mechanisms and impact of functional variability of neurons within the neural circuits to which they belong

Sox2+ progenitors in sharks link taste development with the evolution of regenerative teeth from denticles

Teeth and denticles belong to a specialized class of mineralizing epithelial appendages called odontodes. Although homology of oral teeth in jawed vertebrates is well supported, the evolutionary origin of teeth and their relationship with other odontode types is less clear. We compared the cellular and molecular mechanisms directing development of teeth and skin denticles in sharks, where both odontode types are retained. We show that teeth and denticles are deeply homologous developmental modules with equivalent underlying odontode gene regulatory networks (GRNs). Notably, the expression of the epithelial progenitor and stem cell marker sex-determining region Y-related box 2 (sox2) was tooth-specific and this correlates with notable differences in odontode regenerative ability. Whereas shark teeth retain the ancestral gnathostome character of continuous successional regeneration, new denticles arise only asynchronously with growth or after wounding. Sox2+ putative stem cells associated with the shark dental lamina (DL) emerge from a field of epithelial progenitors shared with anteriormost taste buds, before establishing within slow-cycling cell niches at the (i) superficial taste/tooth junction (T/TJ), and (ii) deep successional lamina (SL) where tooth regeneration initiates. Furthermore, during regeneration, cells from the superficial T/TJ migrate into the SL and contribute to new teeth, demonstrating persistent contribution of taste-associated progenitors to tooth regeneration in vivo. This data suggests a trajectory for tooth evolution involving cooption of the odontode GRN from nonregenerating denticles by sox2+ progenitors native to the oral taste epithelium, facilitating the evolution of a novel regenerative module of odontodes in the mouth of early jawed vertebrates: the teeth

Tooth addition in early ontogeny of <i>Raja clavata</i>.

<p>A-D. Volume rendered and segmented developing teeth in <i>Raja clavata</i> embryos (either VG Studio Max, Drishti, or Avizo). A-B. Embryo 85mm TL, volume rendered upper jaw showing developing teeth as tooth germs under the skin labial to the numerous papillary projections and the symphysis of the palatoquadrate cartilages (S). B. Upper jaw with false colour to highlight the sub-epithelial tooth germs. C-F. Embryo 104mm TL (C, D, stained with iodine and potassium, I2M. Density volume rendered, E, F, unstained). C. Labial view of segmented developing tooth germs in the upper and lower jaws (Avizo); symphyseal tooth (st, red) present in initial row of both jaws, (high density of the connective tissue enhanced at symphyseal junction between cartilages). D. Higher magnification of upper jaw, of lingually rotated jaw to show symphyseal junction (high density tissue) and 3^rd row teeth with one at the symphysis (S). E-F. Volume rendered images of the developing teeth (Drishti). Note that the lateral edges of the images are the limits of the render and not the full extent of the teeth. In both jaws there is a symphyseal tooth (st) in the first row and third row of the lower jaw, but the tooth positions may be shifted with growth and not regular. In the upper jaw there is a symphyseal tooth in the first tooth row. Teeth in second row are developing at later times (arrow heads, start of mineralization) relative to the symphyseal tooth Note in E, cusps have a different orientation in 1^st and 2^nd rows as tooth germs change their developmental positions. F. Lingual view, tooth roots have not started to form and the pulp cavity is open. Scale bars = 500 μm.</p

Phylogeny of the Batoidea and selected outgroups.

<p>Four selected phylogenies showing the varying topologies and the differing positions of clades such as the Torpediniformes. A. After [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.ref010" target="_blank">10</a>] with batoids as derived sharks. B. After [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.ref007" target="_blank">7</a>] with Torpediniformes in a basal position. C. After [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.ref006" target="_blank">6</a>] with the Torpediniformes as a sister group to the Myliobatiformes, but phylogenetic analysis did not include the Rajidae. D. After [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.ref005" target="_blank">5</a>] with polyphyly of the ‘rhinobatids’ and Torpediniformes as a sister group to the Platyrhinidae. Profile images not to scale, slightly modified from FAO publications under a Creative Commons Attribution-Noncommercial 3.0 Unported License. Species studied are: Rajidae; <i>Amblyraja doellojurdoi</i>, <i>Amblyraja frerichsi</i>, <i>Atlantoraja castelnaui</i>, <i>Bathyraja griseocauda</i>, <i>Bathyraja scaphiops</i>, <i>Dipturus batis</i>, <i>Dipturus binoculata</i>, <i>Dipturus chilensis</i>, <i>Leucoraja circularis</i>, <i>Leucoraja erinacea</i>, <i>Leucoraja naevus</i>, <i>Psammobatis normani</i>, <i>Psammobatis rudis</i>, <i>Raja brachyura</i>, <i>Raja clavata</i>, <i>Raja microocellata</i>, <i>Raja undulata</i>, <i>Rioraja agassizi</i>, <i>Sympterygia acuta</i>. Platyrhinidae; <i>Platyrhinidis triseriata</i>. Torpediformes: <i>Discopyge tschudii</i>, <i>Narcine</i> sp., <i>Torpedo puelcha</i>. Zapteryx: <i>Zapteryx brevirostris</i>. Pristidae: <i>Anoxypristis cuspidata</i>, <i>Pristis perotetti</i>. Glaucostegus: <i>Glaucostegus typus</i>. Rhynchobatus: <i>Rhynchobatus djiddensis</i> s.s., <i>Rhynchobatus</i> ex. gr. <i>djiddensis</i>. Rhina: <i>Rhina ancylostoma</i>. Rhinobatos: <i>Rhinobatos horkelii</i>, <i>Trygonorrhina fasciata</i>. Dasyatidae: <i>Dasyatis brevis</i>, <i>Dasyatis</i>? <i>macrophthalma</i>, <i>Himantura uarnak</i>, <i>Himantura</i> sp. 1., <i>Himantura</i> sp. 2., <i>Neotrygon kuhlii</i>, <i>Pastinachus sephen</i>, <i>“Taeniura” lymma</i>, <i>Taeniura meyeni</i>. Myliobatidae and other derived Myliobatiformes: <i>Aetobatus</i> ex. gr. <i>narinari</i>, <i>Aetomylaeus maculatus</i>, <i>Myliobatis aquila</i>, <i>Myliobatis australis</i>, <i>Myliobatis californica</i>, <i>Myliobatis goodei</i> s.s., Myliobatis spp., <i>Rhinoptera javanica</i>, <i>Mobula</i> sp.</p

Phylogeny of the Batoidea and selected outgroups with positions of dental development type.

<p>Tooth development type as seen in different batoid clades (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.g001" target="_blank">Fig. 1E</a>).</p

Development of batoid tooth crown size co-ordinated with roots.

<p>A. Composite volume render of the basal surface of the lower dentition of a female <i>Raja clavata</i> showing the bilobed tooth roots typical of batoids. B. Composite volume render of an embryo of <i>Discopyge tschudii</i> where the initial upper teeth show the presence of bilobed roots. C. Render of a basal section through the lower teeth of an embryo of <i>Myliobatis</i> sp. showing the presence of one or two, poorly developed grooves within the teeth (white arrows). D. Composite volume render of the basal surface of the lower dentition of a neonate specimen of <i>Myliobatis</i> sp. showing up to seven grooves in the roots of the symphyseal teeth and fewer in the other teeth. Many of the grooves are to a greater or lesser extent roofed over. E. Composite volume render of the basal surface of the lower dentition of an adult specimen of <i>Myliobatis</i> sp. showing multiple, well developed, grooves in the roots and in the top row their relationship to the crown. All scale bars are 1mm except where indicated.</p

Morphological variation in batoid dentitions.

<p>A, B. Labial and lingual views, volume rendered scan of the jaws of an adult female <i>Raja clavata</i> (BMNH 2015.1.25.1). C. Symphyseal region of the lower dentition of the ‘rhinobatid’ <i>Glaucostegus typus</i> (BMNH 2015.1.25.3), showing alternate row pattern and massive numbers of small teeth. D. Lower dentition of the myliobatid <i>Aetobatus</i> ex. gr. <i>narinari</i> (BMNH 2015.1.25.4), in which only enlarged symphyseal teeth are present. E. Whole, articulated jaw of <i>Rhina ancylostoma</i> (BMNH 2015.1.25.5) showing convoluted pattern of the teeth and a region of malformed teeth (black arrow, see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.g003" target="_blank">Fig. 3D</a> in [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122553#pone.0122553.ref001" target="_blank">1</a>]). F. Upper jaw of a female <i>Raja clavata</i>, showing homodont dentition of low crowned teeth. G. Upper jaw of a male <i>Raja clavata</i> (BMNH 2015.1.25.2), showing a heterodont dentition with tall cusped teeth. H. Upper dentition of a young female of the dasyatid <i>Neotrygon kuhlii</i> (BMNH 2015.1.25.6) with enlarged ‘caniniform’ teeth, in S+9 position on the jaw. In this and other figures, symphyseal teeth are labelled ‘st’, the jaw symphysis as ‘S’. All scale bars are 1cm unless marked otherwise.</p