REGULATION AND MECHANOCHEMISTRY OF NEURONAL MOTILITY

在神经元发育的寻路阶段，生长锥 作为一种特殊的传感器，能够引导延伸的轴突 远距离的目标地点。生长锥显示出高水平的 肌动蛋白为基础的运动，在一个尚未明确的方式，支持这一点， 指导过程。生长锥表面的特异性受体也会出现 参与到生长锥的引导中。其中一些受体 细胞外基质的成分可能作为空间线索。 尽管人们对表征分子有着浓厚的兴趣， 参与神经元引导，对实际信号知之甚少 涉及的转导过程;例如，受体占据如何导致 细胞骨架结构和运动性的动态改变， 是寻路决策的基础。这项研究试图填补这一空白。 我们的知识差距：（1）通过表征信号转导机制 参与生长锥运动和结构的调节，以及（2）通过 研究维持这种现象的基本机械化学过程， 能动性 这项工作的结果应直接影响到 异常脑发育障碍的临床解释 神经元通路的形成，也可能超出我们的理解， 神经再生的过程。分子底物诊断探针 参与发育和再生神经元疾病的人， 从拟议的研究结果。 一种采用高分辨率数字视频成像的模型系统， 活神经元中的微观膜表面探针将用于 研究生长锥中信号转导机制 指导被认为参与免疫反应的受体的特异性探针 将制定指导和目标识别程序。这些 膜探针将被用来研究生长锥的变化 细胞骨架结构和运动性，似乎发生在响应 适当的细胞外引导信号 信号的最终目的 本项目的转导部分是详细描述受体 调节膜-细胞骨架相互作用的介导过程， 生长锥中基于肌动球蛋白的运动性，并将这些分子事件 宏观的生长锥制导问题。 生长锥如何产生动力尚不清楚;因此，第二个 该提案的阶段涉及机械化学的表征 过程和细胞骨架动力学基础的生长锥运动。一 更清楚地了解生长锥机械化学将补充 信号转导的问题上面概述。身份，空间 神经元肌球蛋白样分子的定位和生化特性 参与肌动蛋白为基础的运动在生长锥将进行调查。 神经肌动蛋白-肌球蛋白的相互作用将通过尝试 在体外去膜细胞中重建神经元肌动蛋白-肌球蛋白运动 细胞模型系统完整细胞中肌动蛋白动力学的进一步表征 生长锥也将使用荧光类似物 细胞化学

During the pathfinding phase of neuronal development, the growth cone functions as a specialized sensor, capable of guiding extending axons toward distant target sites. Growth cones display a high level of actin-based motility which, in an as yet unspecified manner, supports this guidance process. Specific receptors on the growth cone surface also appear to be involved in growth cone guidance. Some of these receptors interact with components of the extracellular matrix that may serve as spatial cues. Although there has been intense interest in characterizing molecules involved in neuronal guidance, very little is known about the actual signal transduction processes involved; for example, how receptor occupation leads to dynamic alterations of cytoskeletal structure and motility likely to underlie pathfinding decisions. The proposed research attempts to fill this gap in our knowledge: (1) by characterizing signal transduction mechanisms involved in regulation of growth cone motility and structure and (2) by investigating the basic mechanochemical processes that sustain this motility. The results of this work should have direct implications for clinical interpretation of developmental brain disorders involving aberrant neuronal pathway formation and may also exceed our understanding of the process of nerve regeneration. Diagnostic probes for molecular substrates involved in developmental and regenerative neuronal disorders could also result from the proposed research. A model system employing high resolution digital-video imaging of microscopic membrane surface probes in living neurons will be used to investigate signal transduction mechanisms involved in growth cone guidance. Specific probes for receptors thought to be involved in the processes of guidance and target recognition will be developed. These membrane probes will then be used to investigate alterations of growth cone cytoskeletal structure and motility that appear to occur in response to appropriate extracellular guidance cues. The ultimate goal of the signal transduction section of this project is to characterize in detail receptor mediated processes that regulate membrane-cytoskeletal interactions and actomyosin-based motility in growth cones and relate these molecular events to the macroscopic problem of growth cone guidance. How growth cones generate locomotive force is not known; thus, the second phase of this proposal involves characterization of the mechanochemical processes and cytoskeletal dynamics underlying growth cone motility. A clearer understanding of growth cone mechanochemistry will compliment the signal transduction questions outlined above. The identity, spatial localization and biochemical properties of neuronal myosin-like molecules involved in actin-based movements in growth cones will be investigated. Neuronal actin-myosin interactions will be approached by attempting to reconstitute neuronal actin-myosin motility in vitro in a demembranated cell model system. Further characterization of actin dynamics in intact growth cones will also be undertaken using fluorescent analog cytochemistry.