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.

翻译后摘要：迟发性阿尔茨海默病（LOAD）的目标是协会皮层，造成深刻的 痴呆症，可用的治疗不会改变疾病的进展。LOAD的最大风险因素是 尽管老年人的大脑皮层已经进入老年，但尚不清楚为什么衰老的联合皮层容易退化。认知 损伤与背外侧前额叶联合皮层（dlPFC）的突触丢失密切相关， 有助于高阶认知。小鼠模型显示，小胶质细胞可以通过以下方式移除突触： 与神经元上的分子“标签”相互作用，包括补体（C1 q）和磷脂酰丝氨酸（PS）。 然而，目前还不清楚神经元内的上游变化触发了这些分子的产生， 这些机制是否在老化的dlPFC中被激活。这项研究将利用一个古老的 恒河猴模型与机械体外实验，以确定神经元内机制， 会导致老化的联合皮层中的突触丢失。衰老的恒河猴有一个扩大的dlPFC 并自然发展认知缺陷，斑块和缠结，补体C1 q表达，以及区域特异性 突触丧失老年猕猴也会出现一种异常的线粒体表型，称为“线粒体上的”。 在人类的LOAD中看到的字符串”（MOAS）。我推测MOAS可能是由慢性钙（Ca 2+）引起的。 线粒体超载，产生参与突触清除的分子，包括激活的C1 q，PS， Caspase 3 dlPFC中的突触特别容易受到Ca 2+失调的影响，因为它们表达cAMP- 蛋白激酶A（PKA）信号传导通过兰尼碱受体2（RyR 2）放大内部Ca 2+释放， 肌醇三磷酸受体1型（IP 3R 1）。这一过程由磷酸二酯酶PDE 4D调节， 年轻的大脑，随着年龄的增长而丢失。我推测细胞内钙离子的持续升高 导致线粒体Ca 2+超载和MOAS的诱导，导致 通过附近的小胶质细胞介导突触移除的分子（C1 q、PS和裂解的半胱天冬酶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， 阐明MOAS和（目标1）已知介导突触清除的分子之间的相互作用， (Aim 2）年轻与老年猕猴dlPFC中Ca 2+失调的标志物。初步数据表明 MOAS优先与C1 q相关，并且在PDE 4D表达被抑制的条件下更常见。 无托叶目的3将使用原代小鼠皮层神经元培养，免疫荧光，超分辨率 显微镜和生物化学来模拟体外Ca 2+失调，并测试慢性升高的Ca 2+是否 水平可以诱导MOAS表型并产生介导突触去除的分子。适用于 将在猕猴组织中交叉验证体外发现。识别导致神经元内事件 在脆弱的老化皮层突触损失将提供关键的见解如何推进年龄有助于 LOAD发病机制，并帮助确定早期治疗干预的潜在目标。