Single Molecule Imaging of Guided Axonal Development using Plasmon Nanorulers

描述（由申请人提供）：亲神经生长因子（如神经生长因子（NGF）和嗜中性蛋白）通过配体诱导的神经特异性膜受体酪氨酸激酶（nsmRTK）二聚化刺激神经细胞（NC）的前神经信号，并调节神经细胞中的几个关键过程，包括轴突特化和伸长。 然而，目前还不清楚这些生物分子调节剂如何协调高度复杂的形态发生神经回路网络的形成。 为了理解具有高时空复杂性的轴突发育的分子机制，需要在具有多变量分析能力的单分子水平上实时监测神经细胞。 在这里，我们提出了一种创新的方法，在单分子水平上使用原位表面等离子体暗视野显微镜研究轴突发育的分子机制。 与传统的显微镜，只提供了一个静态的评估细胞状态，建议的表面等离子体显微镜允许我们可视化信号分子的单分子事件连续轴突生成期间以及形态发生的发展。 通过这种新技术，具体来说，我将挑战以下主题：1。 TrKA的原位监测 单个神经细胞的二聚化及其与不同形态发生（轴突特化和伸长）的相关性。 2. 聚焦单个细胞的神经突激活及其与周围细胞的通讯。 3. 测试人工磁镊系统用于引导轴突发育的可行性。

DESCRIPTION (provided by applicant): Neurotropic growth factors such as nerve growth factor (NGF) and neutrophins stimulate proneuritic signals of neural cells (NCs) via ligand-induced dimerizations of neural specific membrane receptor- tyrosine kinases (nsmRTKs) and modulate several key processes in neural cells including axon specification and elongation. However, it is still unclear how these biomolecular regulators coordinate highly complex morphogenesis for the neural circuit network formation. To understand molecular mechanisms of axonal development with high spatio-temporal complexity, it is required to monitor neural cells in real-time at the single molecule level with multivariate analytical capability. Here, we propos an innovative approach to investigate molecular mechanisms of axonal development at the single molecule level using in situ surface plasmon dark field microscopy. Unlike conventional microscopy that only provides a static assessment of cell status, the proposed surface plasmon microscopy allows us to visualize single molecule events of the signaling molecules continuously over the axon generation period as well as morphogenetic development. With this new technique, specifically, I will challenge the following topics: 1. In situ monitoring of TrKA dimerization of individual nerual cells and its correlation with distinct morphogenesis (axon specification and elongation). 2. Focused neurite activation of a single cell and its communication with surrounding cells. 3. Test the feasibility of an artificial magnetic tweezer system for the guided axonal development.