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 的神经元。 至少有三种兴奋性氨基酸 受体，由“特异性”激动剂N-甲基-D门冬氨酸定义 （NMDA）、使君子酸和红藻氨酸，并且每一种都可以引起兴奋性毒性， 尽管红藻氨酸盐是毒性最强的。 尽管最近取得了进展， 兴奋性毒性的机制尚不清楚，并且可能在 三种受体 目前的建议是继续支持 用于兴奋性毒性的研究 我们将测试四种可能的机制， 毒性，包括a）细胞内钙的积累超过 细胞的缓冲能力，钙离子通过 激动剂激活的通道或电压依赖性钙通道; B）继发于过量的渗透和浓度梯度破坏 钠和氯化物的进入; c）自由基的形成，可能 继发于钙积累，导致蛋白酶活化; 和d）缺乏受体脱敏，特别是红藻氨酸盐 受体，导致上述一种或多种因素引起的损伤。 我们建议进行全细胞斑片记录从急性解离 梨状核和海马神经元，并研究电生理 梨状皮质切片中的毒性指标，使用细胞内 和群体反应记录。 我们将研究三种特异性激动剂和BMAA和BOAA，两种独特的 与人类疾病有关的氨基酸。 在贴片研究中，我们将 比较胰蛋白酶和机械分离的神经元的差异， 脱敏和电流-电压关系，然后使用机械 分离的神经元，以研究二价阳离子在携带或 阻断激动剂激活电流，分析 脱敏，如果它发生和表征受体的两个 独特的氨基酸 在切片研究中，我们将评估 钙、钠和氯化物浓度和游离钙离子抑制剂 自由基的产生或清除剂的兴奋性损失的每一个 给予激动剂。 拟议的研究有可能同时 有助于对兴奋性神经元活动的基本认识。 氨基酸，并导致增加对各种 负责兴奋性毒性的机制。

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.