Investigating the Neuronal Signals Initiating Synapse Loss in Aging and Alzheimer's Disease

研究衰老和阿尔茨海默病中引发突触丢失的神经信号

• 批准号: 10671547
• 负责人: Elizabeth Ka-yoon Woo
• 金额: $ 3.25万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10671547
• 关键词: 3-Dimensional; Acute; Age; Aging; Agonist; Alzheimer' s Disease; Biochemistry; Brain; CASP3 gene; Calcium; Calcium Channel; Calcium Signaling; Cell Death; Chronic; Clinical; Cognition; Cognitive deficits; Complement; Complement 1q; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytosol; Data; Dementia; Dendrites; Disease Progression; Dose; Early Intervention; Elderly; Ensure; Event; Excision; Exposure to; Generations; Human; ITPR1 gene; Image; Image Analysis; Immunoelectron Microscopy; Immunofluorescence Immunologic; Immunohistochemistry; Impaired cognition; In Situ; In Vitro; Individual; Inositol; Late Onset Alzheimer Disease; Lead; Life; Macaca; Macaca mulatta; Mediating; Methods; Microglia; Mitochondria; Modeling; Molecular; Morphology; Mus; Neurofibrillary Tangles; Neurons; Pathogenesis; Pathway interactions; Patients; Phenotype; Phosphatidylserines; Phosphorylation; Population; Process; Quality of life; Research; Resolution; Risk Factors; Ryanodine Receptor Calcium Release Channel; Signal Transduction; Smooth Endoplasmic Reticulum; Synapses; Testing; Therapeutic Intervention; Time; Tissues; age related; aged; association cortex; cost; experimental study; familial Alzheimer disease; in vitro Model; insight; interest; mitochondrial dysfunction; mouse model; neurodevelopment; neurotransmission; phosphoric diester hydrolase; preempt; quantitative imaging; receptor; reconstruction; response; superresolution microscopy; therapeutic target; tripolyphosphate

Abstract: Late-onset Alzheimer’s disease (LOAD) targets the association cortices to cause profound dementia, and available treatments do not alter disease progression. The greatest risk factor for LOAD is advanced age, yet it is unknown why the aging association cortices are vulnerable to degeneration. Cognitive impairment tightly correlates with synapse loss in the dorsolateral prefrontal association cortex (dlPFC), which subserves higher-order cognition. Mouse models have shown that microglia can remove synapses by interacting with molecular “tags” on neurons, including complement (C1q) and phosphatidylserine (PS). However, it is unclear what upstream changes within neurons trigger the generation of these molecules, and whether these mechanisms are activated in the aging dlPFC. The proposed research will utilize an aged rhesus macaque model with mechanistic in vitro experiments, to identify the intraneuronal mechanisms that contribute to synapse loss in the aging association cortex. Aging rhesus macaques have an expanded dlPFC and naturally develop cognitive deficits, plaques and tangles, complement C1q expression, and region-specific synapse loss. Aged macaques also develop an abnormal mitochondrial phenotype termed “Mitochondria-on-a- string” (MOAS) that is seen in human LOAD. I hypothesize that MOAS may arise from chronic calcium (Ca2+) overload of mitochondria, generating molecules participating in synapse removal, including activated C1q, PS, and Caspase 3. Synapses in dlPFC are especially vulnerable to Ca2+ dysregulation as they express cAMP- protein kinase A (PKA) signaling to magnify internal Ca2+ release through ryanodine receptor 2 (RyR2) and inositol triphosphate receptor type 1 (IP3R1). This process is regulated by the phosphodiesterase PDE4D in young brain, which is lost with advancing age. I hypothesize that sustained elevations in cytosolic calcium leads to Ca2+ overload of mitochondria and the induction of MOAS with advancing age, leading to the expression of molecules (C1q, PS, and cleaved caspase 3) that mediate synapse removal by nearby microglia. Aims 1 and 2 will utilize high resolution immuno-Electron Microscopy (EM) and 3D EM reconstruction to elucidate the interactions between MOAS, and (Aim 1) molecules known to mediate synapse removal, and (Aim 2) markers of Ca2+ dysregulation in the dlPFC of young vs. aged macaques. Preliminary data indicate that MOAS preferentially associate with C1q and are more frequent under conditions when PDE4D expression is absent. Aim 3 will use primary murine cortical neuron cultures, immunofluorescence, super-resolution microscopy, and biochemistry to model Ca2+ dysregulation in vitro and test whether chronically elevated Ca2+ levels can induce the MOAS phenotype and generate the molecules mediating synapse removal. Relevant in vitro findings will be cross-validated in the macaque tissue. Identification of the intraneuronal events that lead to synapse loss in the vulnerable aging cortex will provide key insights into how advancing age contributes to LOAD pathogenesis, and help identify potential targets for early therapeutic interventions.

摘要：晚发性阿尔茨海默氏病（LOAD）以皮质联合作用为目标，导致严重的后果。 痴呆症，并且可用的治疗不会改变疾病进展。 LOAD 的最大危险因素是。 年事已高，但为何衰老相关皮层容易发生认知退化尚不清楚。 损伤与背外侧前额叶联合皮层 (dlPFC) 的突触损失密切相关， 促进高阶认知 小鼠模型表明，小胶质细胞可以通过消除突触。 与神经元上的分子“标签”相互作用，包括补体 (C1q) 和磷脂酰丝氨酸 (PS)。 然而，目前尚不清楚神经元内的哪些上游变化触发了这些分子的产生，并且 这些机制是否在老化的 dlPFC 中被激活。拟议的研究将利用老化的 dlPFC。 恒河猴模型与机制体外实验，以确定神经元内机制 导致衰老联合皮层突触损失的因素 衰老的恒河猴的 dlPFC 会扩大。 并自然形成认知缺陷、斑块和缠结、补充 C1q 表达和区域特异性 老年猕猴也会出现一种异常的线粒体表型，称为“线粒体”。 人类负载中出现的“绳子”（MOAS）我认为MOAS可能是由慢性钙（Ca2+）引起的。 线粒体超载，产生参与突触去除的分子，包括激活的 C1q、PS、 和 Caspase 3。 dlPFC 中的突触特别容易受到 Ca2+ 失调的影响，因为它们表达 cAMP- 蛋白激酶 A (PKA) 信号通过兰尼碱受体 2 (RyR2) 放大内部 Ca2+ 释放， 肌醇三磷酸受体 1 型 (IP3R1) 该过程受磷酸二酯酶 PDE4D 调节。 年轻的大脑随着年龄的增长而丧失，我一直在努力维持细胞质钙的持续升高。 随着年龄的增长，导致线粒体 Ca2+ 超载并诱导 MOAS，从而导致 介导附近小胶质细胞去除突触的分子（C1q、PS 和 cleaved caspase 3）的表达。 目标 1 和 2 将利用高分辨率免疫电子显微镜 (EM) 和 3D EM 重建来 阐明 MOAS 和（目标 1）已知介导突触去除的分子之间的相互作用，以及 （目标 2）年轻猕猴 dlPFC 中 Ca2+ 失调的标志物初步数据表明： MOAS 优先与 C1q 结合，并且在 PDE4D 表达低的条件下更频繁。 目标 3 将使用原代小鼠皮质神经元培养物、免疫荧光、超分辨率。 显微镜和生物化学在体外模拟 Ca2+ 失调并测试 Ca2+ 是否长期升高 水平可以诱导 MOAS 表型并产生介导突触去除的分子。 体外研究结果将在猕猴组织中进行交叉验证，以鉴定导致的神经元内事件。 脆弱的老化皮质中的突触损失将提供关键的见解，以了解年龄的增长如何促进 LOAD 发病机制，并帮助确定早期治疗干预的潜在目标。