Role of microglia in a novel model of temporal lobe epilepsy

项目总结：中枢神经系统（CNS）的病毒感染与 癫痫发作、癫痫持续状态（SE）和慢性癫痫发展的风险。我们的合作小组有 开发了第一个病毒诱导癫痫的动物模型。接受脑内注射的小鼠（C57 BL/6） 注射泰勒氏鼠脑脊髓炎病毒（TMEV）显示急性自发性癫痫发作， 感染后30天，在最初的感染中存活下来，并继续发展为自发性复发性癫痫发作。 此外，我们的数据来自使用具有各种细胞因子或细胞因子的C57 BL/6小鼠的最后奖励期。 受体敲除表明，TNFα系统中的操作可以显著改变病理性 TMEV注射后观察到后遗症。因此，我们建议测试我们的总体假设， TMEV感染后，小胶质细胞TNFα的产生和释放呈钙依赖性增加， 激活神经元TNFαR1通路，促进癫痫发作。拟议的实验将 导致更好地了解病毒和免疫对急性癫痫发作的作用，改变 神经元和小胶质细胞功能以及癫痫发生。我们将使用多学科方法来测试我们的 假设，包括体内和体外最新技术水平双光子显微镜、钙成像、新型 转基因小鼠模型，慢性视频脑电图监测和脑切片电生理学，以：1）确定 身体变化的时间过程和程度、运动性和自发性钙瞬变的发展 在TMEV感染小鼠的小胶质细胞中，在急性感染期使用GCAMP 5G选择性表达， 2）确定急性脑缺血期间活化的小胶质细胞中钙瞬变的潜在机制。 感染期和钙瞬变在细胞因子产生中的作用;和3）确定是否通过 神经元TNFαR1通路是TMEV后海马兴奋性和癫痫发作活动的基础 感染我们预计这些实验将为TNFα的作用提供重要的新见解， 受体，TNFαR1在细胞死亡，突触传递和癫痫发生中的作用，并为 开发新的治疗干预措施，以预防感染诱发的癫痫。

PROJECT SUMMARY: Viral infections of the central nervous system (CNS) are associated with an increased risk for seizures, status epilepticus (SE), and the development of chronic epilepsy. Our collaborative group has developed the first animal model of viral-induced epilepsy. Mice (C57BL/6) who receive intra-cerebral injections of Theiler's Murine Encephalomyelitis Virus (TMEV) display acute spontaneous seizures several days after infection, survive the initial infection and go on to develop spontaneous recurrent seizures. Furthermore, our data from the last award period using C57BL/6 mice with various cytokines or cytokine receptors knocked-out have shown that manipulations in the TNFα system can significantly alter the pathologic sequelae observed following TMEV injection. Therefore, we propose to test our overall hypothesis that following TMEV infection, calcium dependent increases in production and release of TNFα from microglia activates the neuronal TNFαR1 pathway, contributing to seizure generation. The proposed experiments will lead to a greater understanding of the role of viral and immune contributions to acute seizures, altered neuronal and microglial function, and epileptogenesis. We will use a multidisciplinary approach to test our hypothesis, including, state of the art in vivo and in vitro 2 photon microscopy, calcium imaging, novel transgenic mouse models, chronic video-EEG monitoring and brain slice electrophysiology to: 1) Determine the time course and extent of physical changes, motility, and the development of spontaneous calcium transients in microglia in TMEV infected mice during the acute infection period using GCAMP5G selectively expressed in microglia; 2) Determine the mechanisms underlying calcium transients in activated microglia during the acute infection period and the role of calcium transients in cytokine production; and 3) Determine if signaling through the neuronal TNFαR1 pathway underlies hippocampal excitability and seizure activity following TMEV infection. We anticipate that these experiments will provide important new insight into the role of TNFα and it's receptor, TNFαR1 in cell death, synaptic transmission and epileptogenesis and set the stage for the development of novel therapeutic interventions for the prevention of infection induced epilepsy.