TBI epileptogenesis: pathologic hippocampal L-glut synaptic plasticity

描述(由申请人提供)： 项目摘要创伤性脑损伤(TBI)患者发展为创伤性癫痫(PTE)的分子机制尚不清楚。已有研究表明，脑外伤后PTE的发生是恢复早期点燃癫痫和受损神经组织残留刺激诱发癫痫的共同结果。谷氨酸是中枢神经系统中的主要兴奋性神经递质，必须受到严格的调控，以便在不产生过度兴奋的情况下，实现适当的神经元信号传递。脑创伤后细胞外谷氨酸释放增加，兴奋性氨基酸转运体重新摄取谷氨酸的能力下降，导致谷氨酸蓄积，继发损伤级联反应，产生异常的神经元信号。我们认为，脑损伤后海马区谷氨酸水平升高的部分原因是神经分泌机制的改变，以及圈套调节和/或再隔离过程。如果不加以控制，这些变化成为脑外伤后癫痫发生的驱动因素，导致PTE。海马区固有谷氨酸能突触可塑性的病理适应性可促进脑损伤后癫痫发生的假说将通过三个特定目标进行验证：(1)利用新型微阵列电极系统，对接受过侧向液压冲击伤(LFPI)的动物(和适当对照)进行遥测监测，直到有脑电(EEG)证据表明癫痫样活动为止：(1)通过使用新型微阵列电极系统，测量中度LFPI后癫痫和非癫痫动物海马结构中的紧张性谷氨酸水平和诱发的谷氨酸释放。在这些动物的海马区组织中，我们将使用Western blotting技术来量化(2)神经递质释放过程的组件和(3)某些谷氨酸转运体。研究异常谷氨酸神经元胞吐/重排序与PTE之间的这种可能的关系，将有助于深入了解谷氨酸神经传递的潜在调控机制，以及癫痫发生的分子事件，并为治疗干预提供潜在的靶点。该项目的长期目标是恢复与脑损伤相关的癫痫发生相关的正常异常机制。动物研究表明，FDA批准并积极使用的几种新的抗癫痫药物，如左乙拉西坦和乳糖胺，也可能通过影响谷氨酸神经传递而起到抗癫痫药物的作用。如果本文建议的研究表明谷氨酸释放的慢性失调与癫痫的发生有关，这些药物将是进一步的动物机制评估和PTE患者临床研究的合适化合物。如果这些研究证实谷氨酸转运体异常与PTE的发生有关，用最近开发的GLT-1转运体的腺相关病毒预转染以增强谷氨酸再摄取，可能会提供另一种新的方法来靶向涉及谷氨酸调节的特定机制。 公共卫生相关性： 除了影响平民人口外，创伤性脑损伤(TBI)已被贴上当前伊拉克和阿富汗战斗的标志性损伤的标签。脑外伤是美国发病率和死亡率的主要原因，也是创伤后癫痫(PTE)的危险因素。研究脑损伤引起的谷氨酸神经递质释放和再摄取的失调，可能揭示重要的机制，确定潜在的治疗靶点，并准确地确定治疗窗口，以消除或降低发生PTE的可能性。识别新疗法的分子靶点将消除脑外伤诱发的癫痫发作，这将减轻平民、士兵和退伍军人回归平民生活时的人类痛苦，并减少癫痫给美国医疗保健系统、军方和退伍军人管理局带来的长期经济成本。

DESCRIPTION (provided by applicant): Project Summary The molecular mechanism that underlies evolution to post-traumatic epilepsy (PTE) in individuals with traumatic brain injury (TBI) is unknown. It has been suggested that the development of PTE following TBI is the combined result of kindling epileptogenesis during the early stages of recovery and triggering by residual irritation from damaged neural tissue. Glutamate, the major excitatory neurotransmitter (NT) in the central nervous system, must be tightly regulated to allow for proper neuronal signaling without creating excessive excitation. TBI produces an increase in glutamate release into the extracellular space and a concurrent decrease in the ability of excitatory amino acid transporters to reuptake glutamate that may lead to its accumulation, propagating secondary damage cascades and producing aberrant neuronal signaling. We suggest that the enhanced hippocampal glutamate levels seen post-TBI are in part due to changes of the neurosecretory machinery and SNARE regulators and/or re-sequestration process. Left unchecked, these changes become drivers of post-TBI epileptogenesis leading to PTE. The hypothesis that pathologic adaptation of inherent hippocampal glutamatergic synaptic plasticity promotes post-TBI epileptogenesis will be tested through three specific aims in animals (and appropriate controls) that have been administered a lateral fluid percussion injury (LFPI) and then monitored by telemetry until electroencephalographic (EEG) evidence of epileptiform activity: (1) Measure of tonic glutamate levels and evoked glutamateamate release in the hippocampal formation of epileptic and non-epileptic animals after moderate LFPI through use of a novel microarray electrode system. In tissue from hippocampal formation of these same animals we will use Western blotting techniques to quantify (2) components of the neurotransmitter release process and (3) certain glutamate transporters. Investigation of this putative relationship between aberrant glutamate neuronal exocytosis/resequestration and PTE will provide insight into potential mechanisms by which glutamate neurotransmission is regulated and into the molecular events of epileptogenesis as well as provide potential foci for therapeutic intervention. The long-term goal of this project is to restore to normal aberrant mechanisms associated with TBI-associated epileptogenesis. There are indications from animal studies that several newer FDA-approved and actively used antiepileptic drugs, e.g. levetiracetam and lacosamide, may also act as antiepileptogenic agents through effect on glutamate neurotransmission. Such agents would be appropriate compounds for further mechanistic evaluation in animals and clinical studies in PTE patients if the studies proposed here suggest that chronic dysregulation of glutamatergic release is associated with epileptogenesis. Pretransfection with a recently developed adeno-associated virus for the GLT-1 transporter to enhance glutamate reuptake, currently under investigation by one of us, may provide yet another, novel approach to target a specific mechanism involved in glutamate regulation should these studies confirm a glutamate transporter abnormality is associated with development of PTE. PUBLIC HEALTH RELEVANCE: In addition to affecting civilian populations, traumatic brain injury (TBI) has been labeled the signature injury of the current combat in Iraq and Afghanistan. TBI is a leading cause of morbidity and mortality in the United States and is a risk factor for post-traumatic epilepsy (PTE). Examining TBI-induced dysregulation of glutamate neurotransmitter release and reuptake, the specific aims of this proposal, may reveal important mechanisms, identify potential therapeutic targets, and pinpoint the therapeutic window to eliminate or reduce the probability of developing PTE. Recognition of molecular targets for new therapies that would extinguish TBI- induced epileptogenesis would alleviate the human suffering of civilians, soldiers and veterans as they return to civilian life, as well as diminish the concomitant long-term economic costs of epilepsy to the US health care system, the military and the Veterans Administration.