Intravital imaging of neuronal development

描述(申请人提供)：神经元发育涉及细胞命运分配、细胞增殖和细胞迁移的精心设计的组合。这些过程的协调导致了中枢神经系统中大脑的刻板印象区域和周围神经系统中的一组模式神经节。面对这种交叉过程研究的复杂性和分子生物学的巨大进步，外周神经系统的研究得到了发育中生物体内高分辨率成像研究的帮助。例如，激光扫描共聚焦显微镜对神经脊内标记的细胞克隆进行了直接评估，并直接跟踪了图案化的迁移；当与分子方法相结合时，这导致了对周围神经系统发育的分子和细胞机制的新见解。拟议中的实验的目标是使用斑马鱼的小脑在脊椎动物的中枢神经系统中进行这样的协调分析。小脑一直是中枢神经系统中研究最深入的区域之一。在胚胎发育过程中，神经前体沿着特定的迁移路径进行长距离迁移，形成了在物种之间保存良好的明确的小脑层状结构。小脑形成的位置允许在活的斑马鱼胚胎内以亚细胞分辨率成像前体细胞及其完整的迁移路径。拟议的研究将建立在我们早期对形成斑马鱼小脑内细胞的成像实验的基础上，以评估神经元引导的分子基础。已定义的信号通路的扰动将被用来探索它们在前体细胞迁移的每个阶段中的作用(S)。由于小脑的结构在物种之间保存得很好，这些对低等脊椎动物的研究将为高等脊椎动物和人类神经元的发育提供重要的见解。

DESCRIPTION (provided by applicant): Neuronal development involves a carefully choreographed combination of cell fate assignment, cell proliferation and cell migration. The coordination of these processes results in the stereotyped regions of the brain in the central nervous system and the patterned set of ganglia in the peripheral nervous system. Faced with the complexity of studying such interleaved processes and the dramatic advances in molecular biology, research in the peripheral nervous system has been assisted by high resolution imaging studies within the developing organism. For example, laser scanning confocal microscopy of labeled clones of cells within the neural crest has allowed the cell lineages to be directly assessed, and the patterned migration to be directly followed; when combined with molecular approaches, this has resulted in new insights to the molecular and cellular mechanisms of peripheral nervous system development. The goal of the proposed experiments is to perform such a coordinated analysis in the central nervous system of a vertebrate, using the zebrafish cerebellum. The cerebellum has been one of the most intensely studied regions of the central nervous system. The defined laminar structure of the cerebellum, which is well-conserved between species, emerges during embryogenesis from the long distance migration of neuronal precursors along specific migratory pathways. The cerebellum form in a position that permits the precursor cells and their complete migration pathway to be imaged with subcelllular resolution within the living zebrafish embryo. The proposed studies will build upon our earlier imaging experiments of cells within the forming zebrafish cerebellum to assess the molecular basis of neuronal guidance. Defined perturbations of defined signaling pathways will be used to explore their role(s) during each phase of the precursor cell migration. Because the structure of the cerebellum is well-conserved between species, these studies on lower vertebrates will offer important insights into the higher vertebrate and human neuronal development.