MECHANISMS OF EXCITATORY AMINO ACID ACTIONS AND TOXICITY

兴奋性氨基酸可能是最重要的一类 大脑中的兴奋性递质，但过量会导致死亡 神经细胞的。至少有三种兴奋性氨基酸 受体，由“特定”激动剂N-甲基-D-天冬氨酸定义 (NMDA)，Quisquate和Kainate，每种都能引起兴奋性毒性， 虽然海人酸是毒性最大的。尽管最近取得了进展，但 兴奋性毒性的机制尚不清楚，而且可能不同 三种类型的受体。目前的建议是继续提供支持 用于兴奋性毒性研究。我们将测试四种可能的机制 毒性，包括a)细胞内钙积累超过 细胞缓冲它的能力，通过钙进入 激动剂激活的通道或电压依赖的钙通道； B)过度后继发的渗透和浓度梯度干扰 钠和氯化物的进入；c)自由基的形成，可能 其次是钙的积累，导致蛋白水解酶的激活； 以及d)缺乏受体脱敏，尤其是红藻氨酸 受体，导致损伤继发于上述一个或多个因素。 我们建议对急性分离的患者进行全细胞贴片记录 梨状和海马神经元，并研究电生理学 梨状皮质脑片细胞内毒性指标 以及沐浴应用激动剂的群体反应记录。 我们将研究三种特定的激动剂以及BMAA和BOAA，这两种独特的 与人类疾病有关的氨基酸。在补丁研究中，我们将 比较胰蛋白酶和机械分离的神经元在 脱敏和电流-电压关系，然后机械地使用 分离的神经元研究二价阳离子在携带或 阻断激动剂激活电流，分析其特性 如果发生脱敏，并确定两种受体的特征 独特的氨基酸。在切片研究中，我们将评估 游离钙、钠和氯的浓度及抑制物 自由基产生或清道夫对每一种兴奋性的丧失 给激动剂。拟议的研究有可能既有可能 有助于了解兴奋性行为的基本知识 氨基酸，并使人们更好地了解 兴奋性毒性的机制。

The excitatory amino acids are probably the most important class of excitatory transmitters in the brain, but in excess they cause the death of neurons. There are at least three types of excitatory amino acid receptors, defined by the "specific" agonists N-methyl-Daspartate (NMDA), quisqualate and kainate, and each can cause excitotoxicity, although kainate is the most toxic. In spite of recent advance the mechanisms of excitotoxicity are unclear and may not be the same at the three types of receptors. The present proposal is for continued support for studies on excitotoxicity. We will test four possible mechanisms of toxicity, including a) accumulation of intracellular calcium beyond the ability of the cell to buffer it, with the calcium entering through either agonist-activated channels or voltage-dependent calcium channels; b) osmotic and concentration gradient disruption secondary to excessive entry of sodium and chloride; c) free radical formation, possibly secondary to calcium accumulation leading to activation of proteases; and d) lack of receptor desensitization, especially of the kainate receptor, leads to damage secondary to one or more of the above factors. We propose to perform whole cell patch recordings from acute dissociated piriform and hippocampal neurons, and study electrophysiologic indicators of toxicity in piriform cortex slices, using intracellular and population response recordings with bath application of agonists. We will study the three specific agonists and BMAA and BOAA, two unique amino acids associated with human disease. In the patch studies we will compare trypsin and mechanically dissociated neurons for differences in desensitization and current-voltage relations, then use mechanically dissociated neurons to study the role of divalent cations in carrying or blocking agonist-activated currents, analyze the properties of desensitization if it occurs and characterize receptors for the two unique amino acids. In the slice studies we will evaluate the effect of calcium, sodium and chloride concentrations and inhibitors of free radical production or scavengers on loss of excitability for each of the give agonists. The proposed studies have the potential to both contribute to the fundamental knowledge of the actions of the excitatory amino acids, and to lead to increased understanding of the variety of mechanisms responsible for excitotoxicity.