Mechanism and Treatment of nerve agent-induced seizures

描述（由申请人提供）：该提案旨在开发用于治疗有机磷酸盐（OP）神经毒剂诱导癫痫发作的新型、基于机制的疗法。OP神经毒剂，如沙林、梭曼和VX，通过抑制中枢和外周胆碱酯酶而起作用，并增强胆碱能传递。在实验动物中，所有已知的神经毒剂都会在接触后几分钟内产生惊厥性癫痫发作和癫痫持续状态。在两伊战争期间暴露于神经毒剂中毒的人中也观察到癫痫发作，在东京地铁袭击中使用了沙林和VX。OP诱导癫痫发作的机制仍不确定。我们实验室的初步研究已经使用OP，对氧磷，作为替代剂来研究OP诱导的癫痫发作。我们证明，低剂量的对氧磷注入海马引起长时间的癫痫发作。此外，我们证明，浓度的对氧磷，导致癫痫发作的海马脑片可以增强兴奋性神经传递刺激谷氨酸释放突触前末梢。这些发现导致了指导这一提议的中心假设的形成：OP神经毒剂的惊厥浓度增强了谷氨酸能神经传递。该提案旨在确认关于OP剂对兴奋性传递的影响的初步研究结果，并测试减少突触前末梢谷氨酸释放的候选化合物作为对抗胆碱能药物诱导的癫痫发作的潜在疗法。实验有三个目标，每组都有具体的治疗干预里程碑。我们提出了量化的结果标准，代表“去不去”的决策点。目的1中概述的实验试图使用膜片钳电生理学和FM染料技术证实和扩展OP胆碱酯酶抑制剂增强海马突触前末梢谷氨酸释放的观察结果。目的2中提出的实验试图使用膜片钳技术和FM染料技术测试两类化合物，生长抑素及其类似物，以及腺苷及其类似物，在对照静息条件期间和OP刺激后减少谷氨酸释放的能力。两类化合物，甘丙肽和神经肽Y，被保留，并将进行测试的情况下，任何一个两个拟议的代理人不能减少谷氨酸释放30%。目的3：测试两类化合物在控制锂/毛果芸香碱和OP对氧磷诱导的癫痫持续状态中的抗惊厥作用。保留两类化合物，神经肽Y和甘丙肽，并将在两种拟议药物中的任何一种未能控制50%实验动物的地西泮难治性癫痫发作的情况下进行测试。通过研究OP诱导癫痫发作的机制，我们提出了确定新的治疗OP诱导癫痫发作的治疗靶点。其次，我们建议开发基于新的治疗靶点的疗法。此外，我们开发了一种OP诱导癫痫发作的模型，可用于民用

DESCRIPTION (provided by applicant): This proposal proposes to develop novel, mechanism-based therapies for the treatment of organophosphate (OP) nerve agent-induced seizures. OP nerve agents, such as sarin, soman and VX act by central and peripheral cholinesterase inhibition and enhance cholinergic transmission. In experimental animals, all known nerve agents, produce convulsive seizures and status epilepticus within minutes of exposure. Seizures were also observed in humans exposed to nerve agent poisoning during the Iran-Iraq war, and in the Tokyo subway attacks where sarin and VX were used. The mechanism of OP-induced seizures remains uncertain. Preliminary studies in our laboratory have used the OP, paraoxon, as a surrogate agent to study OP-induced seizures. We demonstrate that low doses of paraoxon infused into the hippocampus cause prolonged seizures. Furthermore, we demonstrate that the concentrations of paraoxon that cause seizures in hippocampal slices can enhance excitatory neurotransmission by stimulating glutamate release from presynaptic terminals. These findings lead to the formulation of the central hypothesis guiding this proposal: convulsant concentrations of OP nerve agents enhance glutamatergic neurotransmission. This proposal seeks to confirm preliminary findings regarding the effect of OP agent on excitatory transmission and to test candidate compounds that diminish glutamate release from presynaptic terminals as potential therapy against seizures induced by cholinergic agents. Experiments are proposed within three aims, each group has specific milestones toward therapeutic interventions. We present quantitative outcomes criteria that represent "go-no go" decision points. Experiments outlined in Aim 1 seek to confirm and extend the observation that OP cholinesterase inhibitors enhance glutamate release from presynaptic terminals in the hippocampus, using patch clamp electrophysiology and FM dye technique. Experiments proposed in Aim 2 seek to test two classes of compounds, somatostatin and its analogs, and adenosine and its analogs, for their ability to diminish glutamate release during control resting conditions and following OP stimulation, using patch clamp technique and FM dye technique. Two classes of compounds, galanin and neuropeptide Y, are held in reserve and will be tested in case either one of the two proposed agents fail to reduce glutamate release by 30%. Aim 3: To test anticonvulsant action of two classes of compounds in controlling lithium/pilocarpine and OP paraoxon-induced status epilepticus. Two classes of compounds, neuropeptide Y and galanin are held in reserve and will be tested in case either one of the two proposed agents fail to control diazepam refractory seizures in 50% of experimental animals. By studying the mechanisms of OP-induced seizures, we are proposing to identify novel therapeutic targets for the treatment of OP-induced seizures. Second, we propose to develop therapies based on novel therapeutic targets. Furthermore, we develop a model of OP-induced seizures that can be used in civilian