Characterization of the involvement of the cerebellum in animal models of C9orf72 ALS/FTD

项目摘要： 肌萎缩侧索硬化症（Amyotrophic lateral sclerosis，ALS）是一种以神经系统退行性疾病， 大脑和脊髓中的运动神经元。额颞叶痴呆（FTD）是一种早发性痴呆， 额叶和颞叶的退化导致认知缺陷以及行为和语言障碍 异常FTD与ALS具有相同的病理生理学特征。一个六核苷酸重复扩增在一个 C9 orf 72的内含子是这两种疾病最常见的遗传原因。大部分 C9 orf 72 ALS/FTD和其他类型ALS的病理生理学研究迄今为止一直集中在上 和下运动神经元。最近，几项功能研究发现，大脑中的灰质体积较低， 症状前C9 orf 72重复扩增携带者的小脑，以及C9 orf 72的解剖学证据 疾病小鼠模型已经证明了小脑变性。此外，病理报告使用 尸检或患者细胞已显示出内含物、RNA灶、二肽重复蛋白， 小脑，尤其是浦肯野细胞中C9 orf 72 mRNA和蛋白水平，提示小脑 C9 orf 72 ALS/FTD发病机制中的功能障碍。然而，在动物疾病方面的研究很少， 专注于小脑的模型。我们对C9 orf 72敲除小鼠的强有力的初步数据显示， 缺陷和改变浦肯野细胞活性。小脑在C9 orf 72 ALS/FTD中的功能意义 发病机制尚不清楚，更不用说详细的机制了。这些未知因素阻碍了人们了解 C9 orf 72 ALS/FTD的病理生理学，并为患者开发有效的治疗方法。的总目标 我们的研究是使用转基因小鼠来研究C9 orf 72 ALS/FTD的病理生理学。的具体目标 本申请旨在表征C9 orf 72 ALS/FTD小鼠模型，以确定小脑在 ALS和FTD的发病机制。我们假设在症状前阶段，C9 orf 72突变体 小鼠在协调和平衡方面有运动缺陷，小脑的解剖和功能缺陷， 特别是在浦肯野细胞中，由于C9 orf 72蛋白功能的丧失和离子通道的变化。 我们计划用以下具体目的来检验我们的假设：（1）我们将检查C9 orf 72突变小鼠， 行为测试电池，（2）我们将表征神经元形态，被动膜特性， 浦肯野细胞的内在兴奋性，和（3）我们将进行急性分离的浦肯野细胞记录， 蛋白质印迹分析，以确定参与浦肯野细胞放电改变的离子通道。成功 目的的完成将扩大目前关于小脑参与的知识， C9 orf 72介导的ALS/FTD。这将大大增加我们对糖尿病的病理生理学的理解。 C9 orf 72 ALS/FTD，并有助于其他遗传性ALS的病理生理学研究。而且 研究结果将支持小脑作为治疗C9 orf 72 ALS/FTD的潜在治疗靶点。 .

Project Summary: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by the progressive loss of motor neurons in the brain and spinal cord. Frontotemporal dementia (FTD) is early-onset dementia caused by degeneration of the frontal and temporal lobes with cognitive deficits and behavioral and language abnormalities. FTD shares pathophysiological features with ALS. A hexanucleotide repeat expansion in an intron of C9orf72 is the most frequently reported genetic cause of both diseases. Most of the pathophysiological studies of C9orf72 ALS/FTD and other types of ALS so far have been focused on upper and lower motor neurons. Recently, several functional studies have detected lower gray matter volume in the cerebellum in pre-symptomatic C9orf72 repeat expansion carriers, and anatomical evidence in C9orf72 disease mouse models have demonstrated cerebellar degeneration. Furthermore, pathology reports using postmortem or patient cells have demonstrated inclusions, RNA foci, dipeptide repeat proteins, and decreased C9orf72 mRNA and protein levels in the cerebellum, especially Purkinje cells, implicating cerebellar dysfunction in the pathogenesis of C9orf72 ALS/FTD. However, there are very few studies in animal disease models focusing on the cerebellum. Our strong preliminary data on C9orf72 knockout mice has revealed motor deficits and altered Purkinje cell activity. The functional significance of the cerebellum in the C9orf72 ALS/FTD pathogenesis is not known, let alone the detailed mechanisms. These unknowns hamper efforts to understand the pathophysiology of C9orf72 ALS/FTD and to develop effective therapies for patients. The overall goal of our research is to use transgenic mice to study the pathophysiology of C9orf72 ALS/FTD. The specific goal of this application is to characterize mouse models of C9orf72 ALS/FTD to determine the role of cerebellum in the pathogenesis of ALS and FTD. We hypothesize that at the presymptomatic stage, the C9orf72 mutant mice have motor deficits in coordination and balance, anatomical and functional deficits in the cerebellum, especially in the Purkinje cells due to the loss of function of C9orf72 protein and changes in the ion channels. We plan to test our hypothesis with the following Specific Aims: (1) we will examine C9orf72 mutant mice in the behavioral test battery, (2) we will characterize neuronal morphology, passive membrane properties, and intrinsic excitabilities of Purkinje cells, and (3) we will perform acutely dissociated Purkinje cell recording and Western blot analysis to determine ion channels involved in the altered firing of Purkinje cells. The successful completion of the Aims will expand the current knowledge concerning the involvement of the cerebellum in C9orf72-mediated ALS/FTD. This should significantly increase our understanding of the pathophysiology of C9orf72 ALS/FTD and contribute to the pathophysiological studies of other genetic ALS. Furthermore, the outcome of the research will support cerebellum as a potential therapeutic target to treat C9orf72 ALS/FTD. .