miR-126、miR-31在运动神经元与神经干细胞分化所得胆碱能神经元间表达情况的比较

目的了解运动神经元和神经干细胞诱导分化所得胆碱能神经元间miR-126和miR-31的差异表达情况,并以此来探讨两种细胞之间的差异。方法应用ABI公司的TaqMan MicroRNA Assays real-time PCR技术,观察miR-126和miR-31在运动神经元与神经干细胞分化所得胆碱能神经元中的表达情况。结果 miR-126在神经干细胞分化所得胆碱能神经元中的表达是在运动神经元中的0.002倍(P<0.05)。miR-31在神经干细胞分化所得胆碱能神经元中的表达是在运动神经元中的56.444倍(P<0.05)。结论 miR-126和miR-31在运动神经元与神经干细胞分化所得胆碱能神经元中的表达存在差异,对二者预测靶基因参与的生物学过程分析,暗示两种细胞可能在信号传导和发育上存在有差别。     

Differentiation state determines neural effects on microvascular endothelial cells

Growing evidence indicates that nerves and capillaries interact paracrinely in uninjured skin and cutaneous wounds. Although mature neurons are the predominant neural cell in the skin, neural progenitor cells have also been detected in uninjured adult skin. The aim of this study was to characterize differential paracrine effects of neural progenitor cells and mature sensory neurons on dermal microvascular endothelial cells. Our results suggest that neural progenitor cells and mature sensory neurons have unique secretory profiles and distinct effects on dermal microvascular endothelial cell proliferation, migration, and nitric oxide production. Neural progenitor cells and dorsal root ganglion neurons secrete different proteins related to angiogenesis. Specific to neural progenitor cells were dipeptidyl peptidase-4, IGFBP-2, pentraxin-3, serpin f1, TIMP-1, TIMP-4 and VEGF. In contrast, endostatin, FGF-1, MCP-1 and thrombospondin-2 were specific to dorsal root ganglion neurons. Microvascular endothelial cell proliferation was inhibited by dorsal root ganglion neurons but unaffected by neural progenitor cells. In contrast, microvascular endothelial cell migration in a scratch wound assay was inhibited by neural progenitor cells and unaffected by dorsal root ganglion neurons. In addition, nitric oxide production by microvascular endothelial cells was increased by dorsal root ganglion neurons but unaffected by neural progenitor cells.     

Delta/Notch signaling promotes formation of zebrafish neural crest by repressing Neurogenin 1 function

In zebrafish, cells at the lateral edge of the neural plate become Rohon-Beard primary sensory neurons or neural crest. Delta/Notch signaling is required for neural crest formation. ngn1 is expressed in primary neurons; inhibiting Ngn1 activity prevents Rohon-Beard cell formation but not formation of other primary neurons. Reducing Ngn1 activity in embryos lacking Delta/Notch signaling restores neural crest formation, indicating Delta/Notch signaling inhibits neurogenesis without actively promoting neural crest. Ngn1 activity is also required for later development of dorsal root ganglion sensory neurons; however, Rohon-Beard neurons and dorsal root ganglion neurons are not necessarily derived from the same precursor cell. We propose that temporally distinct episodes of Ngn1 activity in the same precursor population specify these two different types of sensory neurons.     

Delta signaling mediates segregation of neural crest and spinal sensory neurons from zebrafish lateral neural plate

We examined the role of Delta signaling in specification of two derivatives in zebrafish neural plate: Rohon-Beard spinal sensory neurons and neural crest. deltaA-expressing Rohon-Beard neurons are intermingled with premigratory neural crest cells in the trunk lateral neural plate. Embryos homozygous for a point mutation in deltaA, or with experimentally reduced delta signalling, have supernumerary Rohon-Beard neurons, reduced trunk-level expression of neural crest markers and lack trunk neural crest derivatives. Fin mesenchyme, a putative trunk neural crest derivative, is present in deltaA mutants, suggesting it segregates from other neural crest derivatives as early as the neural plate stage. Cranial neural crest derivatives are also present in deltaA mutants, revealing a genetic difference in regulation of trunk and cranial neural crest development.     

Zebrafish narrowminded suggests a genetic link between formation of neural crest and primary sensory neurons.

In the developing vertebrate nervous system, both neural crest and sensory neurons form at the boundary between non-neural ectoderm and the neural plate. From an in situ hybridization based expression analysis screen, we have identified a novel zebrafish mutation, narrowminded (nrd), which reduces the number of early neural crest cells and eliminates Rohon-Beard (RB) sensory neurons. Mosaic analysis has shown that the mutation acts cell autonomously suggesting that nrd is involved in either the reception or interpretation of signals at the lateral neural plate boundary. Characterization of the mutant phenotype indicates that nrd is required for a primary wave of neural crest cell formation during which progenitors generate both RB sensory neurons and neural crest cells. Moreover, the early deficit in neural crest cells in nrd homozygotes is compensated later in development. Thus, we propose that a later wave can compensate for the loss of early neural crest cells but, interestingly, not the RB sensory neurons. We discuss the implications of these findings for the possibility that RB sensory neurons and neural crest cells share a common evolutionary origin.     

Pax3-expressing trigeminal placode cells can localize to trunk neural crest sites but are committed to a cutaneous sensory neuron fate

The cutaneous sensory neurons of the ophthalmic lobe of the trigeminal ganglion are derived from two embryonic cell populations, the neural crest and the paired ophthalmic trigeminal (opV) placodes. Pax3 is the earliest known marker of opV placode ectoderm in the chick. Pax3 is also expressed transiently by neural crest cells as they emigrate from the neural tube, and it is reexpressed in neural crest cells as they condense to form dorsal root ganglia and certain cranial ganglia, including the trigeminal ganglion. Here, we examined whether Pax3+ opV placode-derived cells behave like Pax3+ neural crest cells when they are grafted into the trunk. Pax3+ quail opV ectoderm cells associate with host neural crest migratory streams and form Pax3+ neurons that populate the dorsal root and sympathetic ganglia and several ectopic sites, including the ventral root. Pax3 expression is subsequently downregulated, and at E8, all opV ectoderm-derived neurons in all locations are large in diameter, and virtually all express TrkB. At least some of these neurons project to the lateral region of the dorsal horn, and peripheral quail neurites are seen in the dermis, suggesting that they are cutaneous sensory neurons. Hence, although they are able to incorporate into neural crest-derived ganglia in the trunk, Pax3+ opV ectoderm cells are committed to forming cutaneous sensory neurons, their normal fate in the trigeminal ganglion. In contrast, Pax3 is not expressed in neural crest-derived neurons in the dorsal root and trigeminal ganglia at any stage, suggesting either that Pax3 is expressed in glial cells or that it is completely downregulated before neuronal differentiation. Since Pax3 is maintained in opV placode-derived neurons for some considerable time after neuronal differentiation, these data suggest that Pax3 may play different roles in opV placode cells and neural crest cells.     

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.     

Effect of lectins and sugars on primary sperm attachment in the horseshoe crab, Limulus polyphemus L

The in vitro differentiation of quail neural crest cells into serotoninergic neurons is reported. Serotoninergic neurons were identified by two independent methods, formaldehyde-induced histofluorescence and indirect staining with antiserotonin antibodies. Serotonin-positive cells first appeared on the third day in culture, simultaneously, or slightly prior to the first pigmented cells and adrenergic neurons. Comparable numbers of serotoninergic cells were found in crest cell cultures derived from vagal, thoracic/upper lumbar, and lumbosacral levels of the neuraxis. The neural crest origin of the serotonin neurons was further corroborated by the demonstration that cultures of somites, notochords, and neural tubes (three tissues adjacent to the neural crest and thus the most likely contaminants of crest cell cultures) did not contain serotonin-producing cells, and that mast cells were absent in crest cell cultures. The identification of serotoninergic neurons in quail neural crest cell cultures makes an important addition to the number of neural crest derivatives that are capable of differentiating in culture. Furthermore, it suggests that the in vitro culture system will prove a valid approach to the elucidation of the cellular and molecular mechanisms that govern neural crest cell differentiation.     

Differential development of cholinergic neurons from cranial and trunk neural crest cells in vitro

Several studies have suggested that the development of cholinergic properties in cranial parasympathetic neurons is determined by these cells' axial level of origin in the neural crest. All cranial parasympathetic neurons normally derive from cranial neural crest. Trunk neural crest cells give rise to sympathetic neurons, most of which are noradrenergic. To determine if there is an intrinsic difference in the ability of cranial and trunk neural crest cells to form cholinergic neurons, we have compared the development of choline acetyltransferase (ChAT)-immunoreactive cells in explants of quail cranial and trunk neural crest in vitro. Both cranial and trunk neural crest explants gave rise to ChAT-immunoreactive cells in vitro. In both types of cultures, some of the ChAT-positive cells also expressed immunoreactivity for the catecholamine synthetic enzyme tyrosine hydroxylase. However, several differences were seen between cranial and trunk cultures. First, ChAT-immunoreactive cells appeared two days earlier in cranial than in trunk cultures. Second, cranial cultures contained a higher proportion of ChAT-immunoreactive cells. Finally, a subpopulation of the ChAT-immunoreactive cells in cranial cultures exhibited neuronal traits, including neurofilament immunoreactivity. In contrast, neurofilament-immunoreactive cells were not seen in trunk cultures. These results suggest that premigratory cranial and trunk neural crest cells differ in their ability to form cholinergic neurons.     

The B30 ganglioside is a cell surface marker for neural crest-derived neurons in the developing mouse

We have previously reported the isolation of a monoclonal antibody, mAb B30, that recognizes two minor gangliosides specifically expressed in a small subset of neurons in the developing mouse central nervous system (Stainier and Gilbert, 1989). B30 labels mesencephalic trigeminal neurons shortly after differentiation until about 2 weeks after birth. Postnatally, it also labels two specific monolayers of cerebellar neurons. In this study, we have characterized the B30 immunoreactivity in the developing peripheral nervous system of the mouse. We report that B30 is a marker for neural crest-derived neurons and have used it to follow the neuronal differentiation of neural crest cells in a serum-free chemically defined culture system. Within hours after plating, neural crest cells migrate away from the neural tube explant on a fibronectin or laminin substrate and by 24 hr, up to 15% of them have differentiated into morphologically identifable neurons. In vitro as in vivo, undifferentiated mouse neural crest cells express the GD3 ganglioside which is recognized by mAb B33, and neural crest-derived neurons can be labeled by mAbs B33, B30, and also E1.9, a specific neuronal cytoskeletal marker. We also show the unique biochemical specificity of mAb B30 and provide experimental evidence for the role of the B30 ganglioside in the cellular adhesion process.     

神经干细胞向神经元分化调控的研究进展

神经干细胞是一类具有分裂潜能和自更新能力的母细胞,它可以通过对称分裂和不对称分裂方式产生神经组织的各类细胞,包括神经元、星形胶质细胞和少突胶质细胞。中枢神经系统受到损伤后,神经元和胶质细胞的损伤导致了临床症状,内源性神经干细胞的修复作用不大,原因是干细胞的数量有限,微环境的不允许。移植的神经干细胞进入体内后,由于受到多种因素的影响,常保持未分化状态或大部分分化为胶质细胞。神经干细胞向神经元分化的调控机制及其影响因素直接决定神经干细胞源性神经元的比例和神经元之间功能性突触的数量。现就其研究进展做一综述。     

Bmp activity establishes a gradient of positional information throughout the entire neural plate.

Bone morphogenetic proteins (Bmps) are key regulators of dorsoventral (DV) patterning. Within the ectoderm, Bmp activity has been shown to inhibit neural development, promote epidermal differentiation and influence the specification of dorsal neurons and neural crest. In this study, we examine the patterning of neural tissue in mutant zebrafish embryos with compromised Bmp signalling activity. We find that although Bmp activity does not influence anteroposterior (AP) patterning, it does affect DV patterning at all AP levels of the neural plate. Thus, we show that Bmp activity is required for specification of cell fates around the margin of the entire neural plate, including forebrain regions that do not form neural crest. Surprisingly, we find that Bmp activity is also required for patterning neurons at all DV levels of the CNS. In swirl/bmp2b(-) (swr(-)) embryos, laterally positioned sensory neurons are absent whereas more medial interneuron populations are hugely expanded. However, in somitabun(-) (sbn(-)) embryos, which probably retain higher residual Bmp activity, it is the sensory neurons and not the interneurons that are expanded. Conversely, in severely Bmp depleted embryos, both interneurons and sensory neurons are absent and it is the most medial neurons that are expanded. These results are consistent with there being a gradient of Bmp-dependent positional information extending throughout the entire neural and non-neural ectoderm.     

Genetic schemes and schemata in neurophysiology

Information in nervous systems is often carried by neural ensembles--groups of neurons in transient functional linkage--and written in a code that involves the spatial locations of active neurons or synapses and the times at which activity occurs. Even in favorable neuroanatomical circumstances, studying neural ensemble function presents a serious experimental challenge. One recent strategy to overcome this challenge relies on protein-based sensors that provide direct optical images of neural activity, and on protein-based effectors that interfere with it. Because these molecules are encodable in DNA, they can be introduced into intact animals by genetic manipulation, and their expression pattern can be tailored to include--exclusively and at the same time comprehensively--the neurons of interest. Circumscribed populations of neurons can thus be studied in virtual isolation at defined stages of intact neural pathways.     

Energy coding in neural network with inhibitory neurons

This paper aimed at assessing and comparing the effects of the inhibitory neurons in the neural network on the neural energy distribution, and the network activities in the absence of the inhibitory neurons to understand the nature of neural energy distribution and neural energy coding. Stimulus, synchronous oscillation has significant difference between neural networks with and without inhibitory neurons, and this difference can be quantitatively evaluated by the characteristic energy distribution. In addition, the synchronous oscillation difference of the neural activity can be quantitatively described by change of the energy distribution if the network parameters are gradually adjusted. Compared with traditional method of correlation coefficient analysis, the quantitative indicators based on nervous energy distribution characteristics are more effective in reflecting the dynamic features of the neural network activities. Meanwhile, this neural coding method from a global perspective of neural activity effectively avoids the current defects of neural encoding and decoding theory and enormous difficulties encountered. Our studies have shown that neural energy coding is a new coding theory with high efficiency and great potential.     

Retinal neural progenitors express topographic map markers

Transplantation of neural stem cells for replacing neurons after neurodegeneration requires that the transplanted stem cells accurately reestablish the lost neural circuits in order to restore function. Retinal ganglion cell axons project to visual centers of the brain forming circuits in precise topographic order. In chick, dorsal retinal neurons project to ventral optic tectum, ventral neurons to dorsal tectum, anterior neurons to posterior tectum and posterior neurons to anterior tectum; forming a continuous point-to-point map of retinal cell position in the tectal projection. We found that when stem cells derived from ventral retina were implanted in dorsal host retina, the stem cells that became ganglion cells projected to dorsal tectum, appropriate for their site of origin in retina but not appropriate for their site of implant in retina. This led us to ask if retinal progenitors exhibit topographic markers of cell position in retina. Indeed, retinal neural progenitors express topographic markers: dorsal stem cells expressed more Ephrin B2 than ventral stem cells and, conversely, ventral stem cells expressed more Pax-2 and Ventroptin than dorsal stem cells. The fact that neural progenitors express topographic markers has pertinent implications in using neural stem cells in cell replacement therapy for replacing projecting neurons that express topographic order, e.g., analogous neurons of the visual, auditory, somatosensory and motor systems.     

Neural integration of reward, arousal, and feeding: Recruitment of VTA, lateral hypothalamus, and ventral striatal neurons

The ability to control neuronal activity using light pulses and optogenetic tools has revealed new properties of neural circuits and established causal relationships between activation of a single genetically defined population of neurons and complex behaviors. Here, we briefly review the causal effect of activity of six genetically defined neural circuits on behavior, including the dopaminergic neurons DA in the ventral tegmental area (VTA); the two main populations of medium-sized spiny neurons (D1- and D2-positive) in the striatum; the giant Cholinergic interneurons in the ventral striatum; and the hypocretin- and MCH- expressing neurons in the lateral hypothalamus. We argue that selective spatiotemporal recruitment and coordinated spiking activity among these cell type-specific neural circuits may underlie the neural integration of reward, learning, arousal and feeding.     

Immunohistochemical Analysis of the Development of the Floor Plate- and Notochord-Deprived Neural Tube

To analyze the characteristics of neurons and the ectopic fibers that occur in the floor plate-deprived neural tube, the neural tube was separated ipsilaterally from the floor plate and notochord in chick embryos at H-H stage 12. After fixation, operated embryos were labeled with several monoclonal antibodies for detecting cell types and defining the regional characteristics of the neural tube. On the operated side, the basement membrane of the neural tube showed characteristics similar to that of the alar plates. Many neurons had axons that extended outside of the neural tube but which lacked the antigen normally associated with motoneurons. Fibers from the dorsal root ganglia also displayed an atypical distribution within the neural tube. These observations suggest that the neurons in the alar plate can develop independently from the influence(s) of the floor plate and/or notochord and send their axons outside of the neural tube despite the fact that neurons developed in the alar plate do not send axons into the periphery during normal development. It is likely that inhibitory mechanisms, which normally function to restrict axonal growth to within the neural tube, either do not develop or are prevented from functioning in the basal plate lacking environment.     

Differentiation of the zebrafish enteric nervous system and intestinal smooth muscle

Development of the enteric nervous system is critical for normal functioning of the digestive system. In vertebrates, enteric precursors originate from the neural crest and migrate into the digestive system. Enteric neurons enable the digestive system to sense and respond to local conditions without the need for central nervous system input. Here we describe major steps in differentiation of the zebrafish enteric nervous system. During migration and neural differentiation of enteric precursors, we identify regions of the enteric nervous system in different phases of differentiation. Early in migration, a small group of anterior enteric neurons are first to form. This is followed by an anterior to posterior wave of enteric neural differentiation later in the migratory phase. Enteric precursors continue proliferating and differentiating into the third day of embryogenesis. nNOS neurons form early while serotonin neurons form late toward the end of enteric neural differentiation. Numbers of enteric neurons increase gradually except during periods of circular and longitudinal intestinal smooth muscle differentiation.