REGULATION OF NEURONAL MOTILITY

神经元运动的调节

• 批准号: 2714486
• 负责人: PAUL FORSCHER
• 金额: $ 25.26万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-08-01 至 2002-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2714486
• 关键词: Aplysia; biological signal transduction; cell adhesion; cell motility; developmental neurobiology; digital imaging; fluorescence; growth cones; intermolecular interaction; lasers; membrane proteins; membrane structure; micromanipulator; neural cell adhesion molecules; neuronal guidance; photoactivation; tissue /cell culture

DESCRIPTION (Adapted from Applicant's Abstract): The molecular events that underlie the process of growth cone guidance and axonal pathfinding are one of the major challenges for Neurobiology in the future. Recently, research in developmental neurobiology has moved decisively into the molecular arena with the discovery and characterization of molecules that act as extracellular pathfinding cues and specific receptors on growth cones and axons for recognizing these cues. Some of these receptors interact with extracellular matrix components that may serve as spatial cues, still others recognize cell adhesion molecules (CAMs) on other cell surfaces. Although there has been intense interest in molecules involved in neuronal guidance, little is actually known about how adhesion protein or receptor occupation lead to the rapid alterations of structure and motility that underlie pathfinding decisions nor how the forces involved are transduced. The proposed research attempts to fill this gap in our knowledge: (1) by characterizing the cytoskeletal protein dynamics underlying growth cone motility (2) by characterizing signal transduction mechanisms and cell surface recognition processes involved in converting this motility into guidance. The results of this work have direct implications for clinical interpretation of developmental brain disorders involving aberrant neuronal pathway formation and will extend our understanding of nerve regeneration. To accomplish these aims the applicant has developed a model system for studying growth cone interactions with pseudotarget substrates that mimic stereotypic morphological alterations observed when growth cones interact with native targets. Pseudotargets are constructed by derivatizing latex or silica microbeads with test ligands such as cell adhesion proteins or antibodies. When placed on the growth cone surface, they react with targeted membrane proteins and elicit an array of ligand dependent responses that are hypothesized to be related to mechanisms of cell adhesion and target recognition. Single beam gradient optical traps (laser tweezers) are used to facilitate bead placement on the growth cone surface. High resolution video enhanced DIC imaging and digital image processing techniques allow the tracking of bead movements and the analysis of structural changes with high spatial fidelity and temporal resolution. To address molecular substrates underlying these structural changes, fluorescent cytoskeletal and regulatory protein analogs are made and injected into cells. Their movements are being assessed using several fluorescence imaging techniques as well as photoactivation of fluorescence.

描述（改编自申请人的摘要）：分子事件 生长锥引导和轴突寻路过程的基础是其中之一 神经生物学未来面临的主要挑战。 最近，研究 发育神经生物学已决定性地进入分子领域 随着分子的发现和表征 细胞外寻路线索和生长锥上的特定受体 轴突识别这些线索。 其中一些受体与 可作为空间线索的细胞外基质成分，还有其他 识别其他细胞表面上的细胞粘附分子 (CAM)。 虽然 人们对参与神经元引导的分子产生了浓厚的兴趣， 关于粘附蛋白或受体如何占据实际上知之甚少 导致基础结构和运动性的快速改变 寻路决策或所涉及的力量如何转换。 这 拟议的研究试图填补我们知识中的这一空白：（1） 表征生长锥下方的细胞骨架蛋白动力学 通过表征信号转导机制和细胞的运动性 (2) 表面识别过程涉及将这种运动转化为 指导。 这项工作的结果对临床有直接影响 涉及异常神经元的脑发育障碍的解释 通路的形成并将扩展我们对神经再生的理解。 为了实现这些目标，申请人开发了一个模型系统 研究生长锥与模仿的假靶基底的相互作用 生长锥相互作用时观察到的刻板形态变化 与本机目标。 伪目标是通过衍生乳胶或 带有测试配体（例如细胞粘附蛋白）的二氧化硅微珠或 抗体。 当放置在生长锥表面时，它们与 靶向膜蛋白并引发一系列配体依赖性反应 推测与细胞粘附机制有关 目标识别。 单光束梯度光阱（激光镊子）是 用于促进珠子放置在生长锥表面上。 高的 分辨率视频增强 DIC 成像和数字图像处理 技术允许跟踪珠子运动并分析 具有高空间保真度和时间分辨率的结构变化。 到 解决这些结构变化背后的分子底物， 荧光细胞骨架和调节蛋白类似物的制备和 注入细胞。 他们的行动正在通过多种方式进行评估 荧光成像技术以及荧光光激活。