Fate map and morphogenesis of presumptive neural crest and dorsal neural tube

In contrast to the classical assumption that neural crest cells are induced in chick as the neural folds elevate, recent data suggest that they are already specified during gastrulation. This prompted us to map the origin of the neural crest and dorsal neural tube in the early avian embryo. Using a combination of focal dye injections and time-lapse imaging, we find that neural crest and dorsal neural tube precursors are present in a broad, crescent-shaped region of the gastrula. Surprisingly, static fate maps together with dynamic confocal imaging reveal that the neural plate border is considerably broader and extends more caudally than expected. Interestingly, we find that the position of the presumptive neural crest broadly correlates with the BMP4 expression domain from gastrula to neurula stages. Some degree of rostrocaudal patterning, albeit incomplete, is already evident in the gastrula. Time-lapse imaging studies show that the neural crest and dorsal neural tube precursors undergo choreographed movements that follow a spatiotemporal progression and include convergence and extension, reorientation, cell intermixing, and motility deep within the embryo. Through these rearrangement and reorganization movements, the neural crest and dorsal neural tube precursors become regionally segregated, coming to occupy predictable rostrocaudal positions along the embryonic axis. This regionalization occurs progressively and appears to be complete in the neurula by stage 7 at levels rostral to Hensen's node.     

Isolation and culture of neural crest cells from embryonic murine neural tube

The embryonic neural crest (NC) is a multipotent progenitor population that originates at the dorsal aspect of the neural tube, undergoes an epithelial to mesenchymal transition (EMT) and migrates throughout the embryo, giving rise to diverse cell types. NC also has the unique ability to influence the differentiation and maturation of target organs. When explanted in vitro, NC progenitors undergo self-renewal, migrate and differentiate into a variety of tissue types including neurons, glia, smooth muscle cells, cartilage and bone. NC multipotency was first described from explants of the avian neural tube. In vitro isolation of NC cells facilitates the study of NC dynamics including proliferation, migration, and multipotency. Further work in the avian and rat systems demonstrated that explanted NC cells retain their NC potential when transplanted back into the embryo. Because these inherent cellular properties are preserved in explanted NC progenitors, the neural tube explant assay provides an attractive option for studying the NC in vitro. To attain a better understanding of the mammalian NC, many methods have been employed to isolate NC populations. NC-derived progenitors can be cultured from post-migratory locations in both the embryo and adult to study the dynamics of post-migratory NC progenitors, however isolation of NC progenitors as they emigrate from the neural tube provides optimal preservation of NC cell potential and migratory properties. Some protocols employ fluorescence activated cell sorting (FACS) to isolate a NC population enriched for particular progenitors. However, when starting with early stage embryos, cell numbers adequate for analyses are difficult to obtain with FACS, complicating the isolation of early NC populations from individual embryos. Here, we describe an approach that does not rely on FACS and results in an approximately 96% pure NC population based on a Wnt1-Cre activated lineage reporter. The method presented here is adapted from protocols optimized for the culture of rat NC. The advantages of this protocol compared to previous methods are that 1) the cells are not grown on a feeder layer, 2) FACS is not required to obtain a relatively pure NC population, 3) premigratory NC cells are isolated and 4) results are easily quantified. Furthermore, this protocol can be used for isolation of NC from any mutant mouse model, facilitating the study of NC characteristics with different genetic manipulations. The limitation of this approach is that the NC is removed from the context of the embryo, which is known to influence the survival, migration and differentiation of the NC.     

Interpreting neural computations by examining intrinsic and embedding dimensionality of neural activity

The ongoing exponential rise in recording capacity calls for new approaches for analysing and interpreting neural data. Effective dimensionality has emerged as an important property of neural activity across populations of neurons, yet different studies rely on different definitions and interpretations of this quantity. Here, we focus on intrinsic and embedding dimensionality, and discuss how they might reveal computational principles from data. Reviewing recent works, we propose that the intrinsic dimensionality reflects information about the latent variables encoded in collective activity while embedding dimensionality reveals the manner in which this information is processed. We conclude by highlighting the role of network models as an ideal substrate for testing more specifically various hypotheses on the computational principles reflected through intrinsic and embedding dimensionality.     

Global rhythmic activities in hippocampal neural fields and neural coding

Global oscillations of the neural field represent some of the most interesting expressions of the hippocampal activity, being related also to learning and memory. To study oscillatory activities of the CA3 field in theta range, a model of this sub-field of Hippocampus has been formulated. The model describes the firing activity of CA3 neuronal populations within the frame of a kinetic theory of neural systems and it has been used for computer simulations. The results show that the propagation of activities induced in the neural field by hippocampal afferents occurs only in narrow time windows confined by inhibitory barrages, whose time-course follows the theta rhythm. Moreover, during each period of a theta wave, the entire CA3 field bears a firing activity with peculiar space-time patterns, a sort of specific imprint, which can induce effects with similar patterns on brain regions driven by the hippocampal formation. The simulation has also demonstrated the ability of medial septum to influence the global activity of the CA3 pyramidal population through the control of the population of inhibitory interneurons. At last, the possible involvement of global population oscillations in neural coding has been discussed.     

Microfabricated nerve–electrode interfaces in neural prosthetics and neural engineering

Neural interfaces and implants are finding more clinical applications and there are rapid technological advances for more efficient and safe design, fabrication and materials to establish high-fidelity neural interfaces. In this review paper, we highlight new developments of the microfabricated electrodes and substrates with regard to the design, materials, fabrication and their clinical applications. There is a noticeable trend towards integration of microfluidic modules on a single neural platform. In addition to the microelectrodes for neural recording and stimulation, microfluidic channels are integrated into a nerve–electrode interface to explore the rich neurochemistry present at the neural interface and exploit it for enhanced electrochemical stimulation and recording of the central and peripheral nervous system.     

Coupling of a neural filter and a neural controller for improvement of fermentation performance

The noise associated with fermentation processes is normally minimised by a filtering technique. However, sometimes the noise may be beneficial if it is properly regulated. For recombinant -galactosidase production in a fed-batch fermentation subject to Gaussian disturbances, it is shown that a neural network trained to act as a noise filter can allow disturbances of only a particular variance which maximizes -galactosidase synthesis. By coupling such a filter with a neural controller, the productivity may be enhanced beyond what is possible with static filtering and either proportional-integral-derivative or neural control.     

Differential distribution of competence for panplacodal and neural crest induction to non-neural and neural ectoderm

It is still controversial whether cranial placodes and neural crest cells arise from a common precursor at the neural plate border or whether placodes arise from non-neural ectoderm and neural crest from neural ectoderm. Using tissue grafting in embryos of Xenopus laevis, we show here that the competence for induction of neural plate, neural plate border and neural crest markers is confined to neural ectoderm, whereas competence for induction of panplacodal markers is confined to non-neural ectoderm. This differential distribution of competence is established during gastrulation paralleling the dorsal restriction of neural competence. We further show that Dlx3 and GATA2 are required cell-autonomously for panplacodal and epidermal marker expression in the non-neural ectoderm, while ectopic expression of Dlx3 or GATA2 in the neural plate suppresses neural plate, border and crest markers. Overexpression of Dlx3 (but not GATA2) in the neural plate is sufficient to induce different non-neural markers in a signaling-dependent manner, with epidermal markers being induced in the presence, and panplacodal markers in the absence, of BMP signaling. Taken together, these findings demonstrate a non-neural versus neural origin of placodes and neural crest, respectively, strongly implicate Dlx3 in the regulation of non-neural competence, and show that GATA2 contributes to non-neural competence but is not sufficient to promote it ectopically.     

Apical accumulation of Rho in the neural plate is important for neural plate cell shape change and neural tube formation

Although Rho-GTPases are well-known regulators of cytoskeletal reorganization, their in vivo distribution and physiological functions have remained elusive. In this study, we found marked apical accumulation of Rho in developing chick embryos undergoing folding of the neural plate during neural tube formation, with similar accumulation of activated myosin II. The timing of accumulation and biochemical activation of both Rho and myosin II was coincident with the dynamics of neural tube formation. Inhibition of Rho disrupted its apical accumulation and led to defects in neural tube formation, with abnormal morphology of the neural plate. Continuous activation of Rho also altered neural tube formation. These results indicate that correct spatiotemporal regulation of Rho is essential for neural tube morphogenesis. Furthermore, we found that a key morphogenetic signaling pathway, the Wnt/PCP pathway, was implicated in the apical accumulation of Rho and regulation of cell shape in the neural plate, suggesting that this signal may be the spatiotemporal regulator of Rho in neural tube formation.     

Lunatic fringe causes expansion and increased neurogenesis of trunk neural tube and neural crest populations

Both neurons and glia of the PNS are derived from the neural crest. In this study, we have examined the potential function of lunatic fringe in neural tube and trunk neural crest development by gain-of-function analysis during early stages of nervous system formation. Normally lunatic fringe is expressed in three broad bands within the neural tube, and is most prominent in the dorsal neural tube containing neural crest precursors. Using retrovirally-mediated gene transfer, we find that excess lunatic fringe in the neural tube increases the numbers of neural crest cells in the migratory stream via an apparent increase in cell proliferation. In addition, lunatic fringe augments the numbers of neurons and upregulates Delta-1 expression. The results indicate that, by modulating Notch/Delta signaling, lunatic fringe not only increases cell division of neural crest precursors, but also increases the numbers of neurons in the trunk neural crest.     

Reverberations and control of neural networks

Summary The simulation of neural networks, such as the brain cortex, which have a diffuse and rather uniform structure quite unlike the simple block-structure of extant computers, leads naturally to the study of functions and principles which only in part fall within the scope of Automata Theory. Systems of decision equations must be studied with a view especially to obtaining practical means for the prevision and computation of diffuse reverberations of wanted general characteristics, with the exclusion of all others. This amounts to deriving constraints on the allowed variability of the couplings among elements during learning processes, failing which the behavior of the simulator would become uncontrollable for practical purposes. A simple mathematical treatment is presented, which essentially linearizes these problems by an appropriate use of matrix algebra and permits a straightforward study of the wanted conditions, as well as of the controlling elements which may have to be added to the network.This work has been performed in part at the Laboratoire de Physique Théorique et Hautes Energies, Faculté des Sciences de Paris.This work has been performed with the joint sponsorship of the U.S.A.F. and their European Office of Aerospace Research under contracts no. AF EOAR 66-25 and AF 33(615)-2786.We wish to express our sincere thanks to Dr. F. Lauria for many illuminated discussions; and to Prof. M. Lévy for his kind hospitality at the Laboratoire de physique Théorique, in Paris, where part of this research was made.     

Behavioral and neural analysis of extinction

The neural mechanisms by which fear is inhibited are poorly understood at the present time. Behaviorally, a conditioned fear response may be reduced in intensity through a number of means. Among the simplest of these is extinction, a form of learning characterized by a decrease in the amplitude and frequency of a conditioned response when the conditioned stimulus that elicits it is repeatedly nonreinforced. Because clinical interventions for patients suffering from fear dysregulation seek to inhibit abnormal, presumably learned fear responses, an understanding of fear extinction is likely to inform and increase the efficacy of these forms of treatment. This review considers the behavioral, cellular, and molecular literatures on extinction and presents the most recent advances in our understanding while identifying issues that require considerable further research.     

The neural crest and neural crest cells: discovery and significance for theories of embryonic organization

The neural crest has long fascinated developmental biologists, and, increasingly over the past decades, evolutionary and evolutionary developmental biologists. The neural crest is the name given to the fold of ectoderm at the junction between neural and epidermal ectoderm in neurula-stage vertebrate embryos. In this sense, the neural crest is a morphological term akin to head fold or limb bud. This region of the dorsal neural tube consists of neural crest cells, a special population(s) of cell, that give rise to an astonishing number of cell types and to an equally astonishing number of tissues and organs. Neural crest cell contributions may be direct — providing cells — or indirect — providing a necessary, often inductive, environment in which other cells develop. The enormous range of cell types produced provides an important source of evidence of the neural crest as a germ layer, bringing the number of germ layers to four — ectoderm, endoderm, mesoderm, and neural crest. In this paper I provide a brief overview of the major phases of investigation into the neural crest and the major players involved, discuss how the origin of the neural crest relates to the origin of the nervous system in vertebrate embryos, discuss the impact on the germ-layer theory of the discovery of the neural crest and of secondary neurulation, and present evidence of the neural crest as the fourth germ layer. A companion paper (Hall, Evol. Biol. 2008) deals with the evolutionary origins of the neural crest and neural crest cells.     

Role of DNMT3B in the regulation of early neural and neural crest specifiers

The de novo DNA methyltransferase DNMT3B functions in establishing DNA methylation patterns during development. DNMT3B missense mutations cause immunodeficiency, centromere instability and facial anomalies (ICF) syndrome. The restriction of Dnmt3b expression to neural progenitor cells, as well as the mild cognitive defects observed in ICF patients, suggests that DNMT3B may play an important role in early neurogenesis. We performed RNAi knockdown of DNMT3B in human embryonic stem cells (hESCs) in order to investigate the mechanistic contribution of DNMT3B to DNA methylation and early neuronal differentiation. While DNMT3B was not required for early neuroepithelium specification, DNMT3B deficient neuroepithelium exhibited accelerated maturation with earlier expression, relative to normal hESCs, of mature neuronal markers (such as NEUROD1) and of early neuronal regional specifiers (such as those for the neural crest). Genome-wide analyses of DNA methylation by MethylC-seq identified novel regions of hypomethylation in the DNMT3B knockdowns along the X chromosome as well as pericentromeric regions, rather than changes to promoters of specific dysregulated genes. We observed a loss of H3K27me3 and the polycomb complex protein EZH2 at the promoters of early neural and neural crest specifier genes during differentiation of DNMT3B knockdown but not normal hESCs. Our results indicate that DNMT3B mediates large-scale methylation patterns in hESCs and that DNMT3B deficiency in the cells alters the timing of their neuronal differentiation and maturation.