GLUTAMATE RELEASE AND UPTAKE AT CEREBELLAR SYNAPSES

描述：(申请人摘要) 细胞外的谷氨酸水平被非常精确地控制 在时间和空间上，实现高效和选择性的突触兴奋 并通过兴奋性毒性防止神经元死亡。一方面，谷氨酸 突触间隙中的浓度必须迅速上升到毫微克 确保突触后离子亲电受体激活的浓度。在……上面 另一方面，平均而言，亚微摩尔水平的谷氨酸 维持在细胞外空间以防止细胞死亡。这些要求 满足于谷氨酸的爆炸性胞吐释放和高容量 和由家族提供的高亲和力谷氨酸摄取系统 依赖于钠的谷氨酸转运体。当渲染这些传输器时 无效的，无论是在药物上，转基因小鼠，还是自发的 人类突变(如肌萎缩侧索硬化症)，谷氨酸紧张性水平升高和减慢 突触释放部位周围的清除可导致癫痫发作，增强 对缺血侮辱的易感性，以及神经元和器官的死亡。 这项提议的目的是确定有多少谷氨酸从 释放后的突触裂隙，离谷氨酸释放部位有多远 达到足以激活受体的浓度，摄取的速度有多快 系统隔离谷氨酸，以及这些过程是如何受到 谷氨酸释放量的生理变化包括 囊泡充盈过程中的多囊释放和变异性。我们将使用完整的 大鼠急性小脑切片细胞膜片钳和外向式膜片钳方法 解决离子型和代谢型谷氨酸受体的后果 突触裂隙内外的激活，包括 邻近的神经胶质细胞和神经元。

DESCRIPTION: (Applicant's Abstract) Extracellular levels of glutamate are controlled with great precision both temporally and spatially, achieving efficient and selective synaptic excitation and preventing neuronal death by excitotoxicity. On one hand, glutamate concentrations in the synaptic cleft must rise rapidly to millimolar concentrations to ensure activation of postsynaptic ionotropic receptors. On the other hand, sub-micromolar levels of glutamate, on average, must be maintained in the extracellular space to prevent cell death. These requirements are met by the explosive exocytotic release of glutamate and the high capacity and high affinity glutamate uptake system provided by the family of Na-dependent glutamate transporters. When these transporters are rendered ineffective, either pharmacologically, in transgenic mice, or by spontaneous mutation in humans (e.g., ALS), elevated tonic levels of glutamate and slowed clearance around synaptic release sites can result in seizures, enhanced susceptibility to ischemic insults, and neuronal and organismal death. The aim of this proposal is to determine how much glutamate escapes from the synaptic cleft following release, how far from the release site glutamate attains concentrations sufficient to activate receptors, how rapidly the uptake system sequesters glutamate, and how these processes are affected by physiological alterations in the amount of glutamate released including multivesicular release and variability in vesicular filling. We will use whole cell and outside-out patch clamp methods in acute rat cerebellar slices to address the consequences of ionotropic and metabotropic glutamate receptor activation inside and outside of the synaptic cleft, including receptors on neighboring glia and neurons.