Dynamic Microdomains in Brain Extracellular Space

大脑细胞外空间的动态微域

• 批准号: 7047739
• 负责人: SABINA HRABETOVA
• 金额: $ 10.26万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2006-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7047739
• 关键词: biological transport; brain morphology; brain stem; cerebellum; charge coupled device camera; computer simulation; diffusion; electron microscopy; extracellular; fluorescent dye /probe; glia; hypothalamus; ionophores; laboratory rat; macromolecule; neocortex; neurophysiology; neuroregulation; nontherapeutic iontophoresis; quaternary ammonium compound; tissue /cell culture

DESCRIPTION (provided by applicant): In brain, intercellular communication, nutrient and metabolite trafficking, and the delivery of drugs takes place in the extracellular space (ECS). Diffusion, the major transport mechanism mediating these processes, is governed by two structural parameters of the ECS, tortuosity and volume fraction. The tortuosity (?) represents the hindrance imposed on the diffusing molecules by the tissue in comparison with an obstacle-free medium, whereas the volume fraction (a) is the proportion of tissue volume occupied by the ECS. A fundamental question remains unanswered: what hinders molecules traveling through the brain? In healthy brain, ? extracted from the diffusion of small ECS markers is about 1.6 but increases to 1.9 in pathologies where cells swell. It had been thought that ? can be explained by circumnavigation of markers around cells. However, ? derived theoretically or obtained from simulations in media composed of convex cells exhibits an upper limit of about 1.23. Obviously, some other significant factor determines ? in the brain. The central hypothesis of this proposal is that the ECS contains pocket-like microdomains that can significantly slow down the diffusion process, and that these microdomains are formed and regulated by glia. The iontophoresis-based tetramethylammonium (TMA) method and integrative optical imaging (IOI) will quantify diffusion of TMA+ and fluorescent macromolecules, respectively. Rat neocortical slices will be the main preparation but some experiments will examine the cerebellum, hypothalamus or brainstem because of the unique morphological features of these regions. There are three specific aims. Aim 1. Establish that pocket-like microdomains hinder diffusion in brain ECS. The ECS structure will be altered by background macromolecules that fill the pockets and ? will be measured. Trapping of macromolecules will be characterized by the IOI and electron microscopy will localize the entrapment sites. Aim 2. Show that glia form and regulate microdomains. Tortuosity will be measured in the cerebellum where glial wrappings are abundant, in the hypothalamus where withdrawal of glial processes from the supraoptic nucleus will be induced pharmacologically, and in the neocortex where glial toxins will swell glia. Aim 3. Measure diffusion within a microdomain. Diffusion will be measured within a microdomain model, the giant calyx of Held synapse in the brainstem. Computer simulations of diffusion will complement experimental work on Aims 1 and 3. This proposal is focused on basic research with major implications for transport of substances in the nervous system. It identifies a novel role for glia in regulation of diffusion in the ECS.

描述（由申请人提供）：在脑中，细胞间通讯、营养和代谢物运输以及药物递送发生在细胞外间隙（ECS）。扩散，介导这些过程的主要运输机制，是由两个结构参数的ECS，曲折度和体积分数。弯曲度（？）表示与无障碍介质相比由组织施加在扩散分子上的障碍，而体积分数（a）是ECS占据的组织体积的比例。一个基本的问题仍然没有答案：是什么阻碍了分子在大脑中的传播？在健康的大脑中，？从小ECS标记物的扩散中提取的平均值约为1.6，但在细胞膨胀的病理中增加到1.9。有人这么认为吗？可以通过细胞周围标记的环绕来解释。不过，？从理论上导出的或从由凸单元组成的介质中的模拟获得的值显示出约1.23的上限。显然，还有其他重要因素决定？在大脑中。该建议的中心假设是，ECS包含口袋样的微区，可以显着减缓扩散过程，这些微区的形成和调节神经胶质细胞。 基于离子电渗的四甲基铵（TMA）方法和集成光学成像（IOI）将分别量化TMA+和荧光大分子的扩散。大鼠新皮层切片将是主要的制备材料，但一些实验将检查小脑、下丘脑或脑干，因为这些区域具有独特的形态学特征。有三个具体目标。目标1.建立口袋状微区阻碍脑ECS中的扩散。ECS结构将被背景大分子填充的口袋和改变？将被衡量。大分子的捕获将通过IOI表征，并且电子显微镜将定位捕获位点。目标二。表明胶质细胞形成和调节微区。将在小脑中测量迂曲度，其中胶质包裹物丰富，在下丘脑中将诱导胶质过程从视上核中撤出，以及在新皮质中，其中胶质毒素将使胶质细胞膨胀。目标3.测量微域内的扩散。将在微区模型（脑干中Held突触的巨大萼）内测量扩散。扩散的计算机模拟将补充目标1和3的实验工作。该提案侧重于对神经系统中物质运输具有重大影响的基础研究。它确定了一个新的作用，神经胶质细胞在调节扩散的ECS。