Resolving Spatiotemporally-Specific Multicellular Dynamics In Vivo During Olfactory Neurogenesis

项目摘要 虽然已知许多神经缺陷和障碍是由特定的发育障碍引起的 细胞类型和/或链接到研究得很好的信号通路，即多个细胞的系统级协调 在神经发生过程中，种群及其信号输出的时空特异性仍然很差 明白了。这项提议的长期目标是利用可及和快速的 发展斑马鱼嗅觉上皮以定量表征细胞-细胞信号通路和 驱动脊椎动物复杂神经元群体组装的多细胞行为。建议 实验将检验Notch和Wnt信号通路在空间和时间上提供 引导干细胞迁移到嗅觉上皮并调节感觉神经发生的敏感线索。 一小部分细胞将在活体内以亚细胞空间分辨率和 具有亚分钟的时间分辨率，从而确定干细胞迁移的系统级协调， 正常信号和中断信号过程中的规范和区分。首先，新的工具和技术将 用于在嗅觉发育的特定时间和位置干扰体内的Wnt信号，并 用算法模拟并解释祖细胞的迁移行为。接下来，将操纵Notch信令 以量化其在嗅觉系统全神经发生中的作用，并将确定靶基因 是神经元指定和/或分化所必需的。最后，转录因子insm1a将是 类似的研究，以确定其在Notch信号介导的嗅神经发生中的作用。这些方法 在这个提案中，将直接分析体内数据，以了解和预测多细胞行为，而不会减少 生物学的复杂性，并有助于揭示表型，这可能会促进我们对系统范围的广泛理解 神经元在体内的分化和组装。

Project Summary While many neurological defects and disorders are known to result from developmental perturbations in specific cell types and/or are linked to well-studied signaling pathways, the system-level coordination of multiple cell populations and the spatiotemporal specificity of their signaling outputs during neurogenesis remains poorly understood. The long-term objective of this proposal is to take advantage of the accessible and rapidly developing zebrafish olfactory epithelium to quantitatively characterize cell-cell signaling pathways and multicellular behaviors that drive the assembly of complex neuronal populations in vertebrates. Proposed experiments will test the hypothesis that the Notch and Wnt signaling pathways provide spatially- and temporally- sensitive cues to guide stem cell migration into the olfactory epithelium and regulate sensory neurogenesis. Small subsets of cells will be manipulated and analyzed completely in vivo, at subcellular spatial resolution and with sub-minute temporal resolution, so as to determine the system-level coordination of stem cell migration, specification, and differentiation during both normal and disrupted signaling. First, new tools and techniques will be used to perturb Wnt signaling in vivo at specific times and locations during olfactory development and to algorithmically model and explain progenitor cells’ migratory behavior. Next, Notch signaling will be manipulated in vivo to quantitate its effects on olfactory system-wide neurogenesis, and target genes will be identified that are required for neuronal specification and/or differentiation. Finally, the transcription factor insm1a will be similarly investigated to determine its role in Notch signaling-mediated olfactory neurogenesis. The approaches in this proposal will directly analyze in vivo data to understand and predict multicellular behavior without reducing biological complexity and help uncover phenotypes that may advance our broad understanding of system-wide neuronal differentiation and assembly in vivo.