Project 3 - Development and investigation of murine models of channelopathy-associated epilepsy

项目摘要--项目3 癫痫是一种常见的神经系统疾病，影响着300多万美国人，并有大量的遗传因素 对其病因学的贡献。电压门控离子通道基因突变(通道病)，特别是 电压门控钠(NAV)和钾(KV)通道基因已成为早发的主要原因 癫痫性脑病。这些严重的癫痫综合征通常很难用现有的治疗方法来治疗。 并与不利的神经发育后遗症有关，使它们成为改善的高度优先事项 治疗方法包括精准医学。少数癫痫的功能特征- 异源表达系统中相关的电压门控离子通道突变表明 一系列的功能障碍，但目前很难推断这些结果对体内的影响。的一个主要目标 我们的中心是确定体外细胞模型预测神经元功能障碍和药理学的效果如何 在一个完整的大脑中做出反应。为了实现这一目标，Project 3将专注于一系列具有代表性的鼠标 带有NAV和KV通道变异的模型，导致典型的功能障碍模式。我们假设 神经元内不同类型细胞中特定通道对兴奋性的相对贡献的差异 网络决定了兴奋-抑制平衡的净效应，并影响药理反应。 小鼠模型提供了评估通道变异在整个动物细胞中的作用的机会 和网络层面，以及调查药理反应。在目标1中，我们将开发鼠标 研究NAV和KV通道变异与早发性癫痫脑病相关的模型。 小鼠品系将在体内进行神经学表型和药理反应的评估。在目标2中，我们 将确定NAV和KV通道变异对通道特性和固有细胞兴奋性的影响 从小鼠模型中分离出急性分离的神经元，然后确定 使这些神经元的通道活动和/或细胞兴奋性正常化的药物。这些结果 将与来自异源表达系统(项目1)和患者特定的类似记录进行比较 IPSC来源的神经元(项目2)在体外和体内模型之间建立重要的相关性。在……里面 目的3，我们将确定NAV和KV通道变异对神经元内在特性的影响和 继而影响脑片中的网络活动，进而决定其药理作用的有效性 使异常的细胞和网络兴奋性正常化的药物。项目3的结果将提供机械性 洞察完整大脑中经络功能障碍的影响，并确定治疗策略 使兴奋-抑制平衡正常化，预防/减少癫痫发作。本项目与项目之间的协同效应 1和2包括相同通道病相关癫痫变种的跨平台比较， 将有助于将结果转化为有价值的信息，以便在 常见的神经性疾病。

PROJECT SUMMARY – PROJECT 3 Epilepsy is a common neurological disorder that affects over 3 million Americans and has a substantial genetic contribution to its etiology. Mutation of voltage-gated ion channel genes (‘Channelopathies’), particularly voltage-gated sodium (NaV) and potassium (KV) channel genes, have emerged as a major cause of early onset epileptic encephalopathies. These severe epilepsy syndromes are often difficult to treat with existing therapies and are associated with adverse neurodevelopmental sequelae, making them a high priority for better treatment approaches like precision medicine. Functional characterization of a small number of epilepsy- associated voltage-gated ion channel mutations in heterologous expression systems have demonstrated a range of dysfunction, but it is presently difficult to extrapolate these results to in vivo effects. A major goal of our Center is to determine how well in vitro cellular models predict neuronal dysfunction and pharmacological responses in an intact brain. To accomplish this goal, Project 3 will focus on a series of representative mouse models with NaV and KV channel variants that cause prototypical patterns of dysfunction. We hypothesize that differences in the relative contribution of specific channels to excitability in various cell types within neuronal networks determine the net effect on excitation-inhibition balance and influence pharmacological response. Mouse models provide the opportunity to evaluate the effect of channel variants at the whole animal, cellular and network levels, as well as to investigate pharmacological responses. In Aim 1, we will develop mouse models to investigate NaV and KV channel variants associated with early onset epileptic encephalopathy. Mouse lines will be evaluated for neurological phenotypes and pharmacological response in vivo. In Aim 2, we will determine the impact of NaV and KV channel variants on channel properties and intrinsic cell excitability in acutely dissociated neurons isolated from mouse models, and then determine the effectiveness of pharmacological agents at normalizing channel activity and/or cell excitability in these neurons. These results will be compared with similar recordings from heterologous expression systems (Project 1) and patient-specific iPSC-derived neurons (Project 2) to establish important correlations between in vitro and in vivo models. In Aim 3, we will determine the impact of NaV and KV channel variants on intrinsic properties of neurons and consequent effects on network activity in brain slices, and then determine the effectiveness of pharmacological agents at normalizing aberrant cellular and network excitability. Results from Project 3 will provide mechanistic insight into the effects of channel dysfunction in intact brains, and determine therapeutic strategies that normalize excitation-inhibition balance and prevent/reduce seizures. Synergy between this project and Projects 1 and 2 include cross-platform comparisons of the same channelopathy-associated epilepsy variants, which will facilitate translation of results into valuable information for implementation of precision medicine in this common neurological disorder.