GABA Receptors and Hilar Neuron Survival in Epilepsy

描述（由申请人提供）：齿状回门部神经元的丢失是颞叶癫痫患者和相关获得性癫痫模型中最一致的形态学发现之一。两组主要的肝门神经元经常受损，gaba能生长抑素中间神经元和谷氨酸能苔藓细胞。尽管人们多年来一直对门门神经元感兴趣，但重要的问题仍然存在。为什么神经元对兴奋性毒性损伤如此脆弱？如何保护他们？这些神经元亚群的丢失和保存对功能的影响是什么？现在，一系列新技术、试剂和小鼠将以令人兴奋的新方式探索这些问题。拟议的研究将使用在这些神经元中表达cre重组酶的小鼠，通过cre激活的病毒基因表达来选择性地操纵它们。结合光学和电镜免疫组织化学方法将用于评估变化，功能相关性将通过电生理方法确定。广泛的目标是确定在这些神经元中增加强直性gaba能抑制是否可能具有神经保护作用，并在一组单独的研究中，绘制正常和易发作动物体内苔藓细胞或生长抑素神经元在操纵后激活的功能电路。特异性目的1将验证两组门门神经元缺乏GABAA受体（GABAAR）的大量亚基表达和低水平的强直抑制的假设。特异性目的2将验证外源性GABAA亚基的表达，该亚基通常参与小脑的强直抑制，将导致功能性GABAA受体的形成，并增加两组门级神经元的强直抑制。特异性目的3将测试假设，即增加门静脉神经元的强直抑制将保护它们免受癫痫持续状态后的损伤。特异性目的4将通过光遗传学方法选择性地操纵神经元，然后通过Fos标记识别激活的神经元，研究活体门门神经元的功能电路。将比较正常和易发作动物的激活模式，以验证在体内刺激剩余的苔藓细胞或沉默剩余的生长抑素中间神经元将导致癫痫小鼠颗粒细胞激活增加的假设。通过允许对完整大脑中的hila神经元进行选择性操作，这些研究将为其在正常和改变的神经元回路中的功能提供独特的视角。

DESCRIPTION (provided by applicant): Loss of neurons in the hilus of the dentate gyrus is one of the most consistent morphological findings in humans with temporal lobe epilepsy and related models of acquired epilepsy. Two major groups of hilar neurons are frequently damaged, GABAergic somatostatin interneurons and glutamatergic mossy cells. Despite years of interest in hilar neurons, important questions persist. Why are the neurons so vulnerable to excitotoxic damage? How could they be protected? What are the functional effects of both loss and preservation of subgroups of these neurons? A set of new technologies, reagents and mice will now allow probing these questions in exciting new ways. The proposed studies will use mice with Cre-recombinase expression in these neurons to selectively manipulate them through Cre-activated viral gene expression. A combination of light and electron microscopic immunohistochemical methods will be used to evaluate the changes, and functional correlates will be determined with electrophysiological methods. The broad goals are to determine if increasing tonic GABAergic inhibition in these neurons could be neuroprotective and, in a separate set of studies, to map the functional circuits that are activated following manipulation o either mossy cells or somatostatin neurons in normal and seizure-prone animals in vivo. Specific Aim 1 will test the hypothesis that both groups of hilar neurons lack substantial expression of the ¿ subunit of the GABAA receptor (GABAAR) and have low levels of tonic inhibition. Specific Aim 2 will test the hypothesis that expressing an exogenous GABAAR subunit, which is normally involved in tonic inhibition in the cerebellum, will lead to the formatin of functional GABAA receptors and increase tonic inhibition in both groups of hilar neurons. Specific Aim 3 will test the hypothesis that increasing tonic inhibition in hilar neurons will protct them from damage following status epilepticus. Specific Aim 4 will examine the functional circuitry of hilar neurons in vivo by using optogenetic methods to manipulate the neurons selectively and then identifying the activated neurons by Fos labeling. The patterns of activation in normal and seizure-prone animals will be compared to test the hypotheses that stimulating remaining mossy cells or silencing remaining somatostatin interneurons in vivo will lead to increased granule cell activation in the epileptic mice. By allowing selective manipulation of hila neurons in the intact brain, these studies will provide unique views of their function within norma and altered neuronal circuits. PUBLIC HEALTH RELEVANCE: This proposed research is relevant to public health because it will provide new information about neurons that are frequently damaged in epilepsy, ischemia and traumatic brain injury. Loss of specific groups of neurons in the hippocampus is one of the most consistent findings in acquired epilepsy in humans and related animal models. These studies will evaluate the effects of increasing inhibition in these vulnerable neurons with the goa of finding new ways to protect the neurons from seizure-induced damage and will also identify the functional effects of manipulating these neurons in the intact brain.