ANIMAL MODEL AND FUNCTIONAL CHANGES IN AGING

项目2的研究涉及总体方案的三个方面 目标. 作为项目目标#1的组成部分，行为研究 将进行评估老年大鼠的性能测试， 注意，这是敏感的胆碱能免疫毒性病变 年轻大鼠基底前脑的神经元。 这种行为 表征将为神经生物学研究提供背景， 其他项目（项目1，5和6），以及在这些任务中测试的大鼠将 在本项目目标2下，也可用于研究 即刻早期基因对胆碱能激活的影响。 为此目的， 将使用c-Fos mRNA的杂交组织化学（ISHH）进行作图 激活后，低剂量的胆碱能激动剂（毛果芸香碱）作为一个 功能和行为障碍。 该分析还将讨论 第三个计划目标是研究信号转导 机制等 为了实现这一目标，我们还将 检查蛋白激酶C（目标#1）。 我们将评估几种基因的mRNA， 用ISHH法测定海马PKC亚型。 在第二系列研究中， 我们将研究PKC在胞浆和膜组分中的定位 用Western blotting。 这些实验将在两个 行为情境：在一段确定的空间学习之后， 在海马中诱导行为相关的可塑性后， 阵 其余实验（目标3和4）旨在 解决关于糖皮质激素在 海马老化（计划目标#4）。 这些研究将首先 应用溶液杂交技术研究肾上腺皮质激素受体mRNA 以确定老年海马体的变化是否与 行为障碍和海马其他相关改变 mRNA，即神经胶质细胞酸性磷酸酶mRNA表达增加 蛋白（GFAP）和β淀粉样前体蛋白（BAPP）。 除了 表征这些mRNA在完整的年轻和老年大鼠中的状态， 将进行实验以确定糖皮质激素暴露是否 在mRNA的出现中起着必要和/或充分的作用 海马老化和海马依赖性行为特征 损伤 最后，设计实验来检查可能的基础 海马糖皮质激素受体（GR）的年龄依赖性差异 功能 我们的研究将集中在AP-1及其组分Jun和Fos， 它可以确定GR调控的基因是否被激活， 压抑 我们将检验一个或多个Jun mRNA是 与老年海马中的Fos mRNA相比，Fos mRNA升高，这种情况可能导致 导致GR功能从抑制“转换”为激活。 我们将 然后确定衰老和/或认知障碍是否与 与Jun-related蛋白的结合增加（Fos减少）， GFAP-AP 1和/或增殖素复合元件的表达。

The research in Project 2 addresses three of the Overall Program Objectives. As a component of Program Objective #1, behavioral studies will be conducted to assess the performance of aged rats on tests of attention, which are sensitive to immunotoxic lesions of cholinergic neurons in the basal forebrain in young rats. This behavioral characterization will provide a background for neurobiological studies in other projects (Projects 1, 5, and 6), and rats tested on these tasks will also be used under Aim #2 in this project to study the response of an immediate early gene to cholinergic activation. For this purpose in situ hybridization histochemistry (ISHH) for c-Fos mRNA will be used to map activation after a low dose of cholinergic agonist (pilocarpine) as a function of and behavioral impairment. This analysis will also address the 3rd Program Objective aimed at the study of signal transduction mechanisms. To achieve that program objective, we will in addition examine protein kinase C (Aim #1). We will assess mRNAs for several isoforms of PKC in hippocampus using ISHH. In a second series of studies, we will examine localization of PKC in cytosol and membrane fractions using Western blotting. These latter experiments will be conducted in two behavioral contexts: after a defined episode of spatial learning and following induction of behaviorally relevant plasticity in the hippocampal formation. The remaining experiments (in Aims 3 & 4) are designed to address specific hypotheses about the role of glucocorticoids in hippocampal aging (Program Objective #4). These studies will initially focus on adrenocorticoid receptor mRNA using solution hybridization assays to determine whether alterations in the aged hippocampus are related to behavioral impairment and to other associated alterations in hippocampal mRNAs, i.e. increased expression of mRNAs for glial fibrillary acidic protein (GFAP) and beta amyloid precursor protein (BAPP). In addition to characterizing the status of these mRNAs in intact young and aged rats, experiments will be conducted to determine whether glucocorticoid exposure plays a necessary and/or sufficient role in the emergence of mRNA signatures of hippocampal aging and hippocampal-depend behavioral impairment. Finally, experiments are designed to examine a possible basis for age-dependent differences in hippocampal glucocorticoid receptor (GR) function. Our studies will focus on AP-1 and its components Jun and Fos, which can determine whether genes regulated by GR are activated or repressed. We will examine the hypothesis that one or more Jun mRNAs are elevated compared to Fos mRNA in aged hippocampus, a condition that could lead to 'switching' GR function from repression to activation. We will then determine whether aging and/or cognitive impairment is associated with increased binding of Jun-related proteins (decreased Fos) to the GFAP-AP1 and/or proliferin composite element using gel shift assays.