Dysregulated intracellular calcium in microglia during Alzheimer's disease

项目摘要 小胶质细胞，大脑的免疫细胞，执行基本的稳态功能，包括碎片清除， 通过释放生长因子和其他分子来吞噬和维持神经元。但在 在阿尔茨海默病（AD）中，小胶质细胞聚集在淀粉样蛋白斑块附近，在那里它们获得激活的 表型，失去其稳态表型，并采取激活表型，可以诱导 神经毒性小胶质细胞的细胞内钙调节它们从这种稳态静息 状态到激活的免疫效应物状态。小胶质细胞内钙协调功能，如 吞噬作用、增殖、迁移、神经营养因子的产生、细胞因子的产生和释放，以及 神经毒性活性氧的形成。细胞内钙调节异常出现在活化的，非- AD小鼠模型中淀粉样蛋白β（Aβ）斑块附近的稳态小胶质细胞。我们的总体假设是 AD中细胞内钙离子失调改变了小胶质细胞表型。我们的初步数据表明， L型电压依赖性钙通道（L-VDCCs）可以使小胶质细胞向更稳态的方向移动， 表型。所提出的实验验证了L-VDCCs可以驱动小胶质细胞钙的假设 AD病理过程中的失调。我们将通过使用L-VDCCs的药理学拮抗作用来测试这一点 结合在AD病理存在下小胶质细胞特异性L-VDCC的基因消融，以确定 L-VDCCs对小胶质细胞表型和功能的作用。对于本建议的培训方面，我们将使用1）在 体内多光子成像以评估小胶质细胞功能和钙动力学，以及2）单核RNA 测序以确定体内L-VDCC拮抗作用的细胞特异性作用。如果我们的假设是正确的，我- VDCC功能将是联系小胶质细胞钙调节异常和小胶质细胞功能表型的机制 在AD期间。这一发现与许多年龄相关的神经退行性疾病有关， 观察到小胶质细胞表型的变化。

Project Summary Microglia, the immune cells of the brain, perform essential homeostatic functions including debris clearance by phagocytosis and maintenance of neurons by release of growth factors and other molecules. However, during Alzheimer’s disease (AD), microglia accumulate near amyloid plaques where they acquire an activated phenotype, lose their homeostatic phenotype, and take on an activated phenotype that can induce neurotoxicity. Intracellular calcium in microglia regulates their transformation from this homeostatic resting state to an activated immune-effector state. Microglia intracellular calcium orchestrates functions such as phagocytosis, proliferation, migration, generation of neurotrophic factors, cytokine production and release, and formation of neurotoxic reactive oxygen species. Intracellular calcium dysregulation appears in activated, non- homeostatic microglia near amyloid β (Aβ) plaques in mouse models of AD. Our overall hypothesis that dysregulated intracellular calcium shifts microglia phenotype in AD. Our preliminary data suggest that blocking L-type voltage dependent calcium channels (L-VDCCs) can shift microglia towards a more homeostatic phenotype. The proposed experiments test the hypothesis that L-VDCCs can drive microglia calcium dysregulation during AD pathology. We will test this by using pharmacological antagonism of L-VDCCs combined with genetic ablation of microglia-specific L-VDCCs in the presence of AD pathology to define the role of L-VDCCs on microglia phenotype and function. For the training aspect of this proposal, we will use 1) in vivo multiphoton imaging to assess microglia function and calcium dynamics and 2) single-nuclei RNA sequencing to define the cell-specific effects of L-VDCC antagonism in vivo. If our hypothesis is correct, L- VDCC function will be a mechanistic link microglia calcium dysregulation and microglia functional phenotype during AD. This finding would be relevant to numerous age-associated neurodegenerative disorders where changes in microglia phenotype are observed.