Clare V. H. Baker

The origins of the neural crest. Part II: an evolutionary perspective

The neural crest and cranial ectodermal placodes are traditionally thought to be unique to vertebrates; however, they must have had evolutionary precursors. Here, we review recent evidence suggesting that such ancestral cell types can be identified in modern non-vertebrate chordates, such as amphioxus (a cephalochordate) and ascidians (urochordates). Hence, migratory neuroectodermal cells may well have been present in the common ancestor of the chordates, such that the possibility of their existence in non-chordate deuterostomes (hemichordates and echinoderms) must also be considered. Finally, we discuss the various non-neuronal cell types produced by the neural crest in order to demonstrate that it is plausible that these different cell types evolved from an ancestral population that was neuronal in nature.

Neural crest origin of olfactory ensheathing glia

Olfactory ensheathing cells (OECs) are a unique class of glial cells with exceptional translational potential because of their ability to support axon regeneration in the central nervous system. Although OECs are similar in many ways to immature and nonmyelinating Schwann cells, and can myelinate large-diameter axons indistinguishably from myelination by Schwann cells, current dogma holds that OECs arise from the olfactory epithelium. Here, using fate-mapping techniques in chicken embryos and genetic lineage tracing in mice, we show that OECs in fact originate from the neural crest and hence share a common developmental heritage with Schwann cells. This explains the similarities between OECs and Schwann cells and overturns the existing dogma on the developmental origin of OECs. Because neural crest stem cells persist in adult tissue, including skin and hair follicles, our results also raise the possibility that patient-derived neural crest stem cells could in the future provide an abundant and accessible source of autologous OECs for cell transplantation therapy for the injured central nervous system.

The origins of the neural crest. Part I: embryonic induction.

Neural crest cells form at the border between the neural plate and the epidermis. The tissue interactions that underlie neural crest cell induction have been investigated primarily by heterotopic grafting experiments in vivo and by conjugating different tissues in vitro. Three models have been proposed to explain the induction of neural crest cells at the neural plate border, i.e. (1) the influence of signals from the mesoderm, (2) changes in ectodermal competence and (3) local interactions between neural and non-neural ectoderm. The weight of the evidence supports the last model, although there are data that suggest a role for signals from the mesoderm. FGFs seem to be necessary but not sufficient for neural crest cell induction. BMP-4 is sufficient to induce neural crest cells from chick neural explants in vitro and intermediate levels of BMP-4-signalling induce neural crest cell markers in Xenopus animal cap assays. These data suggest a gradient model in which neural crest cells are induced by a particular range of BMP-4 activity, although a single-signal model may be too simplistic. Neural crest cell induction may be an ongoing process, in which an initial induction at the neural plate border is followed by further induction within the dorsal neural tube.

Wnt5a Is Required for Cardiac Outflow Tract Septation in Mice

Lack of septation of the cardiac outflow tract (OFT) results in persistent truncus arteriosus (PTA), a form of congenital heart disease. The outflow myocardium expands through addition of cells originating from the pharyngeal mesoderm referred to as secondary/anterior heart field, whereas cardiac neural crest (CNC) cell–derived mesenchyme condenses to form an aortopulmonary septum. We show for the first time that a mutation in Wnt5a in mice leads to PTA. We provide evidence that Wnt5a is expressed in the pharyngeal mesoderm adjacent to CNC cells in both mouse and chicken embryos and in the myocardial cell layer of the conotruncus at the time when CNC cells begin to form the aortopulmonary septum in mice. Although expression domains of secondary heart field markers are not altered in Wnt5a mutant embryos, the expression of CNC cell marker PlexinA2 is significantly reduced. Stimulation of CNC cells with Wnt5a protein elicits Ca2+ transients, suggesting that CNC cells are capable of responding to Wnt5a. We propose a novel model in which Wnt5a produced in the OFT by cells originating from the pharyngeal mesoderm signals to adjacent CNC cells during formation of the aortopulmonary septum through a noncanonical pathway via localized intracellular increases in Ca2+.

Fine-grained fate maps for the ophthalmic and maxillomandibular trigeminal placodes in the chick embryo.

Vertebrate cranial ectodermal placodes are transient, paired thickenings of embryonic head ectoderm that are crucial for the formation of the peripheral sensory nervous system: they give rise to the paired peripheral sense organs (olfactory organs, inner ears and anamniote lateral line system), as well as the eye lenses, and most cranial sensory neurons. Here, we present the first detailed spatiotemporal fate-maps in any vertebrate for the ophthalmic trigeminal (opV) and maxillomandibular trigeminal (mmV) placodes, which give rise to cutaneous sensory neurons in the ophthalmic and maxillomandibular lobes of the trigeminal ganglion. We used focal DiI and DiO labelling to produce eight detailed fate-maps of chick embryonic head ectoderm over approximately 24 h of development, from 0-16 somites. OpV and mmV placode precursors arise from a partially overlapping territory; indeed, some individual dyespots labelled both opV and mmV placode-derived cells. OpV and mmV placode precursors are initially scattered within a relatively large region of ectoderm adjacent to the neural folds, intermingled both with each other and with future epidermal cells, and with geniculate and otic placode precursors. Although the degree of segregation increases with time, there is no clear border between the opV and mmV placodes even at the 16-somite stage, long after neurogenesis has begun in the opV placode, and when neurogenesis is just beginning in the mmV placode. Finally, we find that occasional cells in the border region between the opV placode and mmV placode express both Pax3 (an opV placode specific marker) and Neurogenin1 (an mmV placode specific marker), suggesting that a few cells are responding to both opV and mmV placode-inducing signals. Overall, our results fill a large gap in our knowledge of the early stages of development of both the opV and mmV placodes, providing an essential framework for subsequent studies of the molecular control of their development.

Specification of GnRH-1 neurons by antagonistic FGF and retinoic acid signaling

A small population of neuroendocrine cells in the rostral hypothalamus and basal forebrain is the key regulator of vertebrate reproduction. They secrete gonadotropin-releasing hormone (GnRH-1), communicate with many areas of the brain and integrate multiple inputs to control gonad maturation, puberty and sexual behavior. In humans, disruption of the GnRH-1 system leads to hypogonadotropic gonadism and Kallmann syndrome. Unlike other neurons in the central nervous system, GnRH-1 neurons arise in the periphery, however their embryonic origin is controversial, and the molecular mechanisms that control their initial specification are not clear. Here, we provide evidence that in chick GnRH-1 neurons originate in the olfactory placode, where they are specified shortly after olfactory sensory neurons. FGF signaling is required and sufficient to induce GnRH-1 neurons, while retinoic acid represses their formation. Both pathways regulate and antagonize each other and our results suggest that the timing of signaling is critical for normal GnRH-1 neuron formation. While Kallmanns syndrome has generally been attributed to a failure of GnRH-1 neuron migration due to impaired FGF signaling, our findings suggest that in at least some Kallmann patients these neurons may never be specified. In addition, this study highlights the intimate embryonic relationship between GnRH-1 neurons and their targets and modulators in the adult.

The amniote paratympanic organ develops from a previously undiscovered sensory placode

The paratympanic organ (PTO), a mechanosensory hair cell-containing pouch in the amniote middle ear, was first described 100 years ago yet its origins remain unresolved. Homology with the anamniote spiracular organ is supported by association with homologous skeletal elements and similar central targets of afferent neurons, suggesting it might be a remnant of the water-dependent lateral line system, otherwise lost during the amniote transition to terrestrial life. However, this is incompatible with studies suggesting it arises from the first epibranchial (geniculate) placode. Here we show that a previously undiscovered Sox2-positive placode, immediately dorsal to the geniculate placode, forms the PTO and its afferent neurons, which are molecularly and morphologically distinct from geniculate neurons. These data remove the only obstacle to accepting the homology of the PTO and spiracular organ. We hypothesise that the PTO/spiracular organ represents an ancient head ectoderm module, developmentally and evolutionarily independent of both lateral line and epibranchial placodes.

Activation of Pax3 target genes is necessary but not sufficient for neurogenesis in the ophthalmic trigeminal placode.

Vertebrate cranial neurogenic placodes are relatively simple model systems for investigating the control of sensory neurogenesis. The ophthalmic trigeminal (opV) placode, for which the earliest specific marker is the paired domain homeodomain transcription factor Pax3, forms cutaneous sensory neurons in the ophthalmic lobe of the trigeminal ganglion. We previously showed that Pax3 expression in avian opV placode cells correlates with specification and commitment to a Pax3+, cutaneous sensory neuron fate. Pax3 can act as a transcriptional activator or repressor, depending on the cellular context. We show using mouse Splotch(2H) mutants that Pax3 is necessary for the normal neuronal differentiation of opV placode cells. Using an electroporation construct encoding a Pax3-Engrailed fusion protein, which represses Pax3 target genes, we show that activation of Pax3 target genes is required cell-autonomously within chick opV placode cells for expression of the opV placode markers FGFR4 and Ngn2, maintenance of the preplacodal marker Eya2, expression of Pax3 itself (suggesting that Pax3 autoregulates), neuronal differentiation and delamination. Mis-expression of Pax3 in head ectoderm is sufficient to induce FGFR4 and Ngn2 expression, but neurons do not differentiate, suggesting that additional signals are necessary to enable Pax3+ cells to differentiate as neurons. Mis-expression of Pax3 in the Pax2+ otic and epibranchial placodes also downregulates Pax2 and disrupts otic vesicle closure, suggesting that Pax3 is sufficient to alter the identity of these cells. Overall, our results suggest that activation of Pax3 target genes is necessary but not sufficient for neurogenesis in the opV placode.

The evolution and elaboration of vertebrate neural crest cells

Vertebrate neural crest cells are embryonic neuroepithelial cells that undergo an epithelial-mesenchymal transition, migrate throughout the embryo and form a wide variety of derivatives, including peripheral neurons and glia, pigment cells, and craniofacial cartilage, bone and teeth. Neural crest cell evolution and elaboration is intimately bound up with vertebrate evolution: the most primitive living vertebrates, lampreys and hagfishes, have most but not all neural crest derivatives. A torrent of recent molecular information has changed our understanding of vertebrate phylogenetic relationships, expanded our understanding of the gene circuitry underlying neural crest development, and given interesting information on the deployment of homologues of these genes in invertebrate relatives such as ascidians and amphioxus. New molecular insights into the evolutionary origin of cartilage, as well as into the nature and evolution of the cells and genes involved in tooth and bone formation, enable tentative hypotheses to be framed for the evolution of skeletal neural crest derivatives.

Electrosensory ampullary organs are derived from lateral line placodes in cartilaginous fishes.

Ampullary organ electroreceptors excited by weak cathodal electric fields are used for hunting by both cartilaginous and non-teleost bony fishes. Despite similarities of neurophysiology and innervation, their embryonic origins remain controversial: bony fish ampullary organs are derived from lateral line placodes, whereas a neural crest origin has been proposed for cartilaginous fish electroreceptors. This calls into question the homology of electroreceptors and ampullary organs in the two lineages of jawed vertebrates. Here, we test the hypothesis that lateral line placodes form electroreceptors in cartilaginous fishes by undertaking the first long-term in vivo fate-mapping study in any cartilaginous fish. Using DiI tracing for up to 70 days in the little skate, Leucoraja erinacea, we show that lateral line placodes form both ampullary electroreceptors and mechanosensory neuromasts. These data confirm the homology of electroreceptors and ampullary organs in cartilaginous and non-teleost bony fishes, and indicate that jawed vertebrates primitively possessed a lateral line placode-derived system of electrosensory ampullary organs and mechanosensory neuromasts.