Mechanisms of Dantrolene Neuroprotection in Alzheimer's Disease

摘要 针对淀粉样蛋白治疗阿尔茨海默病（AD）的临床试验已经 基本上失败了。Ca 2+调节异常可能是一种替代机制。在家族性AD（FAD）中， 钙稳态被NMDA（NMDAR）和兰尼碱（RYR）的过度激活破坏 受体，导致细胞溶质钙增加和随后的认知功能障碍， 神经病理学APOE 4是散发性AD（SAD）的主要危险因素，也可导致Ca 2 + 与NMDAR/RYR过度激活相关的失调。丹曲林，一种RYR拮抗剂， 临床上可获得的药物，已经显示减轻淀粉样蛋白病理，神经变性， FAD动物模型中突触和记忆丧失。我们的初步研究表明， 鼻内丹曲林给药消除了5XFAD小鼠的记忆丧失。此外，委员会认为， 丹曲林促进来自人的诱导多能干细胞（iPSC）的神经元分化 通过抑制RYR/NMDAR过度表达， activation.我们的长期目标是研究丹曲林治疗AD的疗效。整体 本研究的目的是研究丹曲林对大鼠脑缺血再灌注损伤的作用及其机制。 人和啮齿动物SAD和FAD细胞中的成体神经发生，以及鼻内给药的影响 丹曲林给药对AD转基因小鼠成年神经发生和认知功能的影响。 我们的中心假设是，鼻内丹曲林抑制过度激活 RYR和NMDAR在AD中促进成人神经发生，改善认知功能 和减少AD神经病理学。我们将通过以下具体目标来检验这一假设。 具体目标1。确定丹曲林对RYR和NMDAR表达的影响 以及AD患者的细胞溶质、线粒体和ER Ca 2+浓度和AD生物标志物 使用来自患有SAD或SAD的成纤维细胞患者的诱导多能干细胞（iPSC） （APOE 4风险因子）或FAD（PSEN 1突变），以及分离的神经祖细胞（NPC 来自E4 FAD（APOE 4 +5XFAD）和5XFAD转基因Specific Aim 2。为了检验 丹曲林对RYR和NMDAR活性、神经发生、增殖和细胞凋亡的影响 从AD细胞衍生的神经元和神经胶质的功能。使用与SA 1中相同的细胞。 具体目标3。为了确定丹曲林对成人神经发生、认知功能和神经发育的影响， 功能和神经病理学。我们认为丹曲洛林 促进AD小鼠成年期神经发生和恢复认知，减轻AD病理 通过减轻RYR和/或NMDAR的过度激活，特别是使用 鼻内途径。

Abstract Clinical trials for the treatment of Alzheimer's disease (AD) targeting amyloid have largely failed. Ca2+ dysregulation may be an alternative mechanism. In familial AD (FAD), calcium homeostasis is disrupted by over activation of NMDA (NMDAR) and ryanodine (RYR) receptors leading to increased cytosolic calcium and subsequent cognitive dysfunction and neuropathology. APOE4, a major risk factor for sporadic AD (SAD), also causes Ca2+ dysregulation related to NMDAR/RYR over activation. Dantrolene, a RYR antagonist and clinically available drug, has been shown to mitigate amyloid pathology, neurodegeneration, synaptic and memory loss in a FAD animal model. Our preliminary studies suggested that intranasal dantrolene administration abolished memory loss in 5XFAD mice. Furthermore, dantrolene promoted neuronal differentiation in induced pluripotent stem cells (iPSC) from patients with either SAD with APOE4 or familial AD (FAD) by inhibition of RYR/NMDAR over- activation. Our long term goal is to examine the efficacy of dantrolene to treat AD. The overall objective of this study is to investigate the effects and underlying mechanisms of dantrolene on adult neurogenesis in human and rodent SAD and FAD cells, as well as the effects of intranasal dantrolene administration on adult neurogenesis and cognitive function in AD transgenic mice. Our central hypothesis is that intranasal dantrolene inhibits the excessive activation of RYRs and NMDARs in AD and promotes adult neurogenesis, improved cognitive function and reduced AD neuropathology. We will test the hypothesis by the following specific aims. Specific Aim 1. To determine the effects of dantrolene on RYR and NMDAR expression and on cytosolic, mitochondrial, and ER Ca2+ concentrations and AD biomarkers in AD stem cells using induced pluripotent stem cells (iPSC) from fibroblasts patients with either SAD (APOE4 risk factor) or FAD (PSEN1 mutations), as well as neuroprogenitor cells (NPC) isolated from E4FAD (APOE4+5XFAD) and 5XFAD transgenic Specific Aim 2. To examine the effects of dantrolene on RYR and NMDAR activity, neurogenesis, proliferation, and the cellular function of derived neurons and glia from AD cells. using the same cells as in SA1. Specific Aim 3. To determine the effects of dantrolene on adult neurogenesis, cognitive function, and neuropathology in animal models of AD. We expect that dantrolene will promote adult neurogenesis and restore cognition and reduce AD pathology in AD mouse models by alleviating the excessive activation of RYRs and/or NMDARs, especially with the intranasal approach.