Glial huntingtin and neuronal excitotoxicity

描述（由申请人提供）：亨廷顿病（HD）是由亨廷顿蛋白（htt）N-末端区域的多聚谷氨酰胺（polyQ）结构域扩增引起的。虽然突变htt在大多数类型的细胞中普遍表达，但它导致选择性神经变性，其特征在于HD患者脑纹状体中严重的神经元损失。谷氨酸兴奋性毒性一直是一个长期存在的理论来解释选择性变性的中型棘神经元（MSN）在纹状体。MSN接收丰富的谷氨酸能输入，并且对谷氨酸刺激过敏。由于细胞外谷氨酸大部分被星形胶质细胞（支持神经元存活并保护免受神经元兴奋性毒性的主要类型的神经胶质细胞）去除，因此星形胶质细胞去除细胞外谷氨酸的能力的任何降低都可能增加MSN对谷氨酸毒性的脆弱性。然而，胶质细胞在HD神经病理学中的作用仍有待充分表征。我们最近的研究表明突变htt在星形胶质细胞中表达，在细胞核中形成聚集体。我们还发现星形胶质细胞中的突变htt降低谷氨酸转运体GLT-1的表达和谷氨酸的摄取。此外，神经元-胶质细胞共培养显示，正常的胶质细胞保护htt介导的神经毒性，而胶质细胞表达突变htt促进神经元的脆弱性谷氨酸兴奋性毒性。我们假设突变htt在神经胶质细胞中的表达影响神经胶质细胞的功能，从而导致神经递质摄取的改变并导致神经病理学的发生。为了验证这一假设，我们将集中在星形胶质细胞中的突变htt的影响，并检查是否突变htt在体外和体内阻碍星形胶质细胞的重要功能。在目的1中，我们将使用培养的原代胶质细胞来研究突变htt如何影响胶质细胞中谷氨酸摄取和神经元功能。目的2：利用转基因小鼠研究星形胶质细胞中突变htt对神经元兴奋性毒性的影响。公共卫生相关性：亨廷顿病是由亨廷顿蛋白中的多聚谷氨酰胺扩增引起的，亨廷顿蛋白是一种在神经元和非神经元细胞中普遍表达的蛋白质。尽管突变亨廷顿蛋白的广泛表达，但该疾病的特征是脑神经元的选择性变性。脑中神经元的功能和存活在很大程度上受神经胶质细胞的调节，神经胶质细胞是脑中的大多数细胞。然而，突变亨廷顿蛋白在神经胶质细胞中的作用及其对神经病理学的贡献尚不清楚。本申请将使用小鼠遗传和分子神经生物学方法来研究神经胶质细胞中的亨廷顿蛋白是否以及如何影响神经元的活力和功能。这些研究将有助于开发治疗亨廷顿病的替代策略。

DESCRIPTION (provided by applicant): Huntington's disease (HD) is caused by the expansion of a polyglutamine (polyQ) domain in the N-terminal region of the protein huntingtin (htt). While mutant htt is expressed ubiquitously in most types of cells, it leads to selective neurodegeneration that is characterized by severe neuronal loss in the brain striatum of HD patients. Glutamate excitotoxicity has been a long- standing theory to account for the selective degeneration of medium spiny neurons (MSNs) in the striatum. MSNs receive abundant glutamatergic input and are hypersensitive to glutamate stimulation. Since extracellular glutamate is largely removed by astrocytes, a major type of glial cell that supports neuronal survival and protects against neuronal excitotoxicity, any reduction in the ability of astrocytes to remove extracellular glutamate may increase the vulnerability of MSNs to glutamate toxicity. Yet the role of glia in HD neuropathology remains to be fully characterized. Our recent studies show that mutant htt is expressed in astrocytes, forming aggregates in their nuclei. We also found that mutant htt in astrocytes reduces both the expression of the glutamate transporter GLT-1 and the uptake of glutamate. Furthermore, neuron-glia cocultures revealed that normal glia protect against htt-mediated neurotoxicity, whereas glia expressing mutant htt promote neuronal vulnerability to glutamate excitotoxicity. We hypothesize that the expression of mutant htt in glia affects glial function, which leads to altered neurotransmitter uptake and contributes to neuropathology. To test this hypothesis, we will focus on the effects of mutant htt in astrocytes and examine whether mutant htt impedes the important functions of astrocytes in vitro and in vivo. In Aim 1, we will use cultured primary glial cells to examine how mutant htt affects glutamate uptake and neuronal function in glial cells. In Aim 2, we will use transgenic mice expressing mutant htt in astrocytes to study how mutant htt in astrocytes contributes to neuronal excitotoxicity. PUBLIC HEALTH RELEVANCE: Huntington disease is caused by a polyglutamine expansion in huntingtin, a protein that is ubiquitously expressed in neuronal and non-neuronal cells. Despite the widespread expression of mutant huntingtin, the disease is characterized by selective degeneration in neurons in the brain. The function and survival of neurons in the brain are largely regulated by glial cells, which are the majority of cells in the brain. However, the role of mutant huntingtin in glial cells and its contribution to neuropathology are unclear. This application will use mouse genetic and molecular neurobiology approaches to investigate whether and how huntingtin in glial cells affects neuronal viability and function. These studies will help develop alternative strategies for treating Huntington disease.