Cerebellar malfunction and damage during ischemia

描述(由申请者提供)：这项提案的主要目标是研究在脑缺血期间可能涉及小脑损伤和功能障碍的机制。脑缺血发生在心脏骤停、中风和围产期窒息期间，是导致死亡和长期残疾的主要原因。小脑是卒中的常见靶点，而小脑浦肯野细胞是最容易受到缺血性损伤的脑细胞之一。然而，对于浦肯野细胞如何对缺血做出反应并受到损伤，人们知之甚少。这种信息的缺乏是有问题的，因为在其他大脑区域运作的许多缺血机制涉及浦肯野细胞中不表达或具有不寻常配置的分子过程。在这项提议中，将使用脑片模型来模拟体外脑缺血，并将使用膜片钳记录、共聚焦荧光成像和药物操作来研究小脑缺血损伤的机制。模拟缺血引起浦肯野细胞严重去极化，这种去极化是由非NMDA型离子亲和性谷氨酸受体(可能还有其他谷氨酸受体/转运体)的激活所介导的，但GABAA受体的激活延缓了这种去极化的发生。这些电生理反应与广泛的组织肿胀和随后发生的坏死性和凋亡性细胞死亡非常相似，与体内模型观察到的情况非常相似。模拟缺血也会严重干扰通过小脑的电信号传导，这种干扰在缺血发作终止后会持续很长一段时间。确定导致细胞损伤和信号处理中断的机制应该会为治疗的发展提供有用的信息。该建议的具体目的是：1)确定导致浦肯野细胞周围谷氨酸积聚的机制，阐明哪些受体介导浦肯野细胞的反应并确定它们对细胞损伤的贡献；2)确定GABAA受体激活延迟浦肯野细胞去极化开始的机制，并确定这种延迟是有益的还是有害的；以及3)确定小脑电路中的哪里以及通过什么机制破坏电信号转导。

DESCRIPTION (provided by applicant): ' The primary goal of this proposal is to investigate mechanisms that may be involved in cerebellar damage and malfunction during brain ischemia. Brain ischemia, which occurs during cardiac arrest, stroke and perinatal asphyxia, is a leading cause of death and long term disability. The cerebellum is a frequent target of stroke, and cerebellar Purkinje cells are one of the most susceptible brain cells to ischemic damage. However, relatively little is known about how Purkinje cells respond to and are damaged by ischemia. This lack of information is problematic because many of the ischemic mechanisms operating in other brain regions involve molecular processes that are either not expressed or have an unusual configuration in Purkinje cells. For this proposal, a brain slice model will be used to simulate brain ischemia in vitro and patch-clamp recording, confocal fluorescence imaging and pharmacological manipulations will be used to investigate mechanisms of cerebellar ischemic damage. Simulated ischemia induces a severe depolarization of Purkinje cells which is mediated by activation of non-NMDA ionotropic glutamate receptors (and possibly other glutamate receptors/transporters) but its onset is delayed by activation of GABAA receptors. These electrophysiological responses are associated with extensive tissue swelling and the subsequent development of necrotic and apoptotic cell death very similar to that observed with in vivo models. Simulated ischemia also severely disrupts electrical signal transduction through the cerebellum and the disruption persists for long periods after the ischemic episode is terminated. Determining the mechanisms that underlie both cellular damage and disrupted signal processing should provide useful information for the development of therapies. The specific aims of this proposal are: 1) To determine the mechanisms that lead to glutamate accumulation around Purkinje cells, elucidate which receptors mediate the Purkinje cell response and determine their contribution to cell damage; 2) To determine the mechanisms by which GABAA receptor activation delays the onset of Purkinje cell depolarization and to determine if the delay is beneficial or damaging; and 3) To determine where in the cerebellar circuitry and by what mechanism electrical signal transduction is disrupted.