Mechanisms of Target-Specific Axon Regeneration

项目摘要 神经损伤是一种常见的疾病，会导致感觉和/或运动障碍。恢复涉及到 神经纤维内受损的轴突必须重新延伸到适当的靶点的再生过程 组织，在一个被称为靶标特异性再生的过程中。这一过程在人类身上经常失败，留下患者 有慢性健康问题。改善临床结果需要更好地理解如何针对特定的靶点 再生是受调控的。我们知道神经支持支架的组件可以引导轴突重新伸展 沿着简单的小路。然而，当轴突到达神经分支点时，它们需要更具体的指导。 区分多条路径并选择正确路径的机制。我们对此了解甚少 指导这些决定的是什么环境线索，以及轴突再生如何恰当地解释这些决定。 这项提议的目的是确定调节轴突靶向的细胞和分子机制。 促进特定目标再生的决定。 我已经建立了斑马鱼迷走神经的模型，以阐明靶向特异性轴突的机制。 再生。再生的迷走神经轴突在五个神经分支之间选择，以强有力地重新支配正确的 靶组织，尽管他们是如何做到这一点的尚不清楚。我假设有两种不相互排斥的机制 调节靶点特异性再生：1)化学感觉，再生轴突可以在空间上解释 环境中指导其生长的图案化化学引导线索；2)丛生，其中 再生轴突可以识别未受损的轴突，这些轴突正在支配其预期的目标，并将它们用作 用于定向生长的衬底。这项提案的三个目标将全面确定轴突如何生长 在靶标特异性再生过程中，它们与环境在细胞、生物学和分子水平上相互作用。在……里面 目标1，我将结合一种新的单细胞嵌合体再生实验与活体成像和遗传学和 建立对活体轴突环境如何的概念性理解的药物操作 互动指导目标决策。在目标2中，我将结合一种新的方法来标记和分离活神经元 基于他们的神经支配靶点，用体内和体外技术来精确测量每个 这五个神经支配靶群与环境中的其他轴突和化学信号相互作用。在AIM 3、我将神经支配靶点特异性神经元分离与RNAseq和突变分析相结合，以实现无偏 识别调节五个神经支配靶点群中每一个靶点选择的分子。本研究 将极大地增强我们对轴突如何重新支配其靶组织的基本理解 并为开发改进的神经损伤治疗方法提供了重要的知识基础。

Project Summary Nerve damage is a common affliction that causes sensory and/or motor deficits. Recovery involves a regenerative process in which damaged axons within a nerve fiber must re-extend to the appropriate target tissues, in a process known as target-specific regeneration. This process often fails in humans, leaving patients with chronic health problems. Improving clinical outcomes requires a better understanding of how target-specific regeneration is regulated. We know that components of the nerve support scaffold can guide axon re-extension along simple paths. However, when axons reach nerve branch points, they require more specific guidance mechanisms to differentiate between multiple paths and select the correct one. We have little understanding of what environmental cues guide these decisions, and how they are appropriately interpreted by regrowing axons. The objective of this proposal is to identify cellular and molecular mechanisms that regulate axon targeting decisions to promote target-specific regeneration. I have established the zebrafish vagus nerve as a model to elucidate mechanisms of target-specific axon regeneration. Regenerating vagus axons select between five nerve branches to robustly re-innervate the correct target tissue, although how they do so is not known. I hypothesize that two non-mutually-exclusive mechanisms regulate target-specific regeneration: 1) chemosensation, in which a regenerating axon can interpret spatially patterned chemical guidance cues in the environment that direct its growth; 2) fasciculation, in which a regenerating axon can recognize undamaged axons that are innervating its intended target and use them as a substrate for directed growth. The three aims of this proposal will comprehensively identify how growing axons interact with their environment at the cell biological and molecular levels during target-specific regeneration. In Aim 1, I will combine a novel single-cell chimera regeneration assay with live imaging and genetic and pharmacological manipulations to establish a conceptual understanding of how in vivo axon-environment interactions guide targeting decisions. In Aim 2, I will combine a novel method to label and isolate live neurons based on their innervation target with in vivo and in vitro techniques to precisely measure how axons of each of the five innervation target groups interact with other axons, and with chemical signals, in the environment. In Aim 3, I will combine innervation target-specific neuron isolation with RNAseq and mutant analysis for unbiased identification of molecules that regulate target selection in each of the five innervation target groups. This study will greatly enhance our fundamental understanding of how axons reinnervate their target tissues during regeneration, and provide an important knowledge base to develop improved treatments for nerve damage.