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，特别是工作记忆，这 在中年时开始下降。我们的团队和其他人已经确定了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L3层的神经元，如脊髓、突触和髓鞘的丢失以及由此引起的 突触和内在电生理特性。重要的是，这些结构和功能的变化 随着前额叶皮质(PFC)神经元和白质通路的老化而被大量观察到 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 在正常老化过程中，锥体神经元在很大程度上是幸免的。我们目前缺乏机械性的理解 为什么这两个脑区的锥体神经元在正常衰老或多大年龄时具有不同的脆弱性- PFC中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。这个项目的总体假设是神经元和与之相关的选择性脆弱性 在老化过程中，LPFC中的网络与V1中的网络相比，是由于在固有属性和 这两个区域神经元的神经束背景，以及LPFC内神经元对 氧化应激和炎症。我们提出了一种新的实验方法--多路传输 免疫组织化学结合高分辨结构分析的生理特征 单个神经元-比较单个LPFC和V1锥体神经元在 幼年恒河猴和老年恒河猴周围的神经纤维。这些猴子也将被评估 认知状态、通路完整性和脑脊液促炎细胞因子水平作为其他现有NIH- 受资助的项目。本项目有两个目的：1)评估植物的形态特征 对幼猴和老年猴LPFC和V1的L3锥体神经元进行了观察。我们将评估树枝状结构 以及树突棘亚型的数量和密度，并将这些数据与现有的数据进行关联 这些相同细胞的30种不同的生理特性。2)确定规范性属性的特征， 老化对目标1中的相同L3锥体神经元的影响是在神经束的背景下研究的。我们会 在同一组织样本上进行~20个蛋白质靶点的原位免疫荧光多路复用 测定充满生物细胞素的第3层锥体神经元的分子表型。这个项目的一个主要成果 将能够定量地指定两个L3锥体神经元之间的不同参数 大脑区域高度不同，哪种参数组合最能预测衰老过程中的认知损害。 这项研究将形成未来一系列更大的研究的基础，以调查关系和相互依赖 在正常衰老过程中各种细胞类型、板层和皮质区中与年龄相关的细胞变化 可以与恒河猴的认知表现相关。

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.