Substrates of selective neuronal vulnerability in aging: neocortical pyramidal neurons and their surrounding neuropil environment in visual versus frontal cortex of young and aged rhesus monkeys

灵长类动物的正常衰老通常会导致认知功能受损，特别是在工作记忆方面， 在中年开始下降。我们的团队和其他人已经确定，与年龄相关的认知障碍 不是由于神经元的明显死亡，而是与一系列亚致死性变化有关， 神经元，特别是在层（L3），如脊柱，突触和髓鞘损失和随之而来的改变， 突触和内在电生理特性。重要的是，这些结构和功能变化 在前额叶皮层（PFC）的神经元和白色物质通路中， 在工作记忆中起关键作用的大脑区域。相比之下，初级视皮层（V1）的特性 锥体神经元在正常衰老期间大部分被保留。我们目前缺乏一个机械的理解， 为什么这两个大脑区域的锥体神经元在正常衰老过程中的脆弱性不同， PFC单细胞和通路水平的相关变化影响网络功能，从而影响工作记忆 性能该项目的总体假设是，神经元的选择性脆弱性和相关的 老化过程中LPFC中的网络与V1相比，是由于固有特性和 在这两个领域的神经元，神经元在LPFC的增加更大的敏感性， 氧化应激和炎症。我们提出了一种新的实验方法-多重 免疫组织化学结合高分辨率结构分析， 单个神经元-比较单个LPFC和V1锥体神经元的特性， 年轻和年老的恒河猴的周围神经元。这些猴子也会被评估 认知状态，通路完整性和CSF促炎细胞因子水平作为其他现有NIH- 资助的项目。本课题有两个目的：1）评价植物生理形态学特性， 特征L3锥体神经元LPFC和V1的年轻和老年猴。我们将评估树突状细胞 拓扑结构和树突棘亚型的数量和密度，并将这些数据与现有数据相关联， 30种不同的生理特性。2)为了表征规范属性， 衰老对目标1中研究的相同L3锥体神经元的影响。我们将 对同一组织样本进行约20种蛋白质靶标的原位免疫荧光多重检测， 确定充满生物细胞素的第3层锥体神经元的分子表型。该项目的一个主要成果是 将能够定量地指定两个L3锥体神经元之间不同的那些参数， 高度不同的大脑区域以及哪些参数组合最能预测衰老中的认知障碍。 这项研究将成为未来一系列更大规模研究的基础，以调查关系和相互依赖 在正常衰老过程中， 与恒河猴的认知能力有关。

Normal aging in primates often leads to impaired cognitive function, particularly in working memory, which begins to decline in middle-age. Our group and others have established that age-related cognitive impairment is not due to overt death of neurons but rather is associated with a constellation of sublethal changes to neurons particularly in layer (L3), such as spine, synapse and myelin loss and consequent alterations to synaptic and intrinsic electrophysiological properties. Importantly these structural and functional changes have been abundantly observed with aging in neurons and white matter pathways in the prefrontal cortex (PFC), a brain area that is a key player in working memory. By contrast, the properties of primary visual cortex (V1) pyramidal neurons are largely spared during normal aging. We currently lack a mechanistic understanding of why pyramidal neurons in these two brain areas are differentially vulnerable in normal aging or how age- related changes at the single-cell and pathway level in PFC impact network function and thus working memory performance. The overall hypothesis of this project is that selective vulnerability of neurons and associated networks in LPFC compared to V1 during aging is due to key differences in both the intrinsic properties and the neuropil context of neurons in the two areas, and a greater susceptibility of neurons in LPFC to increases in oxidative stress and inflammation. We propose a novel experimental approach -multiplexed immunohistochemistry combined with high resolution structural analyses of physiologically characterized individual neurons- to compare the properties of individual LPFC and V1 pyramidal neurons in the context of their surrounding neuropil in young and aged rhesus monkeys. These monkeys will also have been assessed for cognitive status, pathway integrity, and CSF pro-inflammatory cytokine levels as part of other existing NIH- funded projects. This project has two aims: 1) To assess the morphological properties of physiologically characterized L3 pyramidal neurons in LPFC and V1 of young and aged monkeys. We will assess dendritic topology and the number and density of dendritic spine subtypes and correlate these data with existing data on 30 different physiological properties of these same cells. 2) To characterize the normative properties and effects of aging on the same L3 pyramidal neurons studied in Aim 1 in the context of the neuropil. We will perform in situ immunofluorescence multiplexing of ~20 protein targets on the same tissue sample to determine the molecular phenotype of biocytin-filled layer 3 pyramidal neurons. A major outcome of this project will be the ability to quantitatively specify those parameters that differ between L3 pyramidal neurons in two highly distinct brain areas and which combination of parameters best predict cognitive impairment in aging. This study will form the basis of future series of larger studies to investigate relationships and co-dependence of age-related cellular changes in a variety of cell types, laminae and cortical areas during normal aging that can be correlated with cognitive performance in rhesus monkeys.