Synaptic substrates of age-dependent memory deficits

 描述（由申请人提供）：虽然衰老本身是阿尔茨海默病（AD）的最大风险因素，但许多有记忆问题的老年人最终不会患AD。在这样的人，病理学，值得AD诊断是不存在的，尽管重叠的认知障碍。这项研究计划的主要目标是在大鼠中模拟这种“正常”的与年龄相关的认知异质性，然后使用高分辨率技术来揭示与年龄相关的记忆形式下降的突触和树突基质，这些记忆形式需要海马体。作为海马的主要输出，CA 1区一直是我们和其他人研究的主要目标。事实上，过去的研究表明，虽然主要的年龄相关的突触和神经元损失是在CA 1缺席，更微妙的年龄相关的变化合谋修改神经元的活动模式和行为可塑性。重要的是，这种改变只存在于老年大鼠的一个子集，与其他相同的实足年龄显示年轻成人样的收购莫里斯水迷宫（MWM）的学习和跟踪眨眼条件反射（TEBC）。因此，老年大鼠的认知异质性为揭示与年龄相关的记忆衰退的基质以及记忆保存的基质提供了重要的机会。本次更新申请中提出的实验将继续揭示与年龄相关的认知异质性的突触和树枝状基底，但这是通过建立在我们过去的研究结果的基础上，具有概念和技术的复杂性来实现的。具体而言，在目标1中，我们将行为特征的年轻成年和老年F344 xBN F1杂种大鼠MWM学习和TEBC，然后探针突触Ca 2+信号与2-光子激光扫描（2 PLSM）Ca 2+成像和2-光子谷氨酸（2 Pglutamate）uncaging到个别基底，斜，簇树突的离散微域的棘。在这种高分辨率功能探测之后，目标2和3将使用免疫金场发射扫描电子显微镜（iFESEM）和免疫荧光阵列断层扫描（iAT）来形态学重建成像的树突，然后蛋白质组学重建树突的信号网络。目标2将集中在一个参与突触和树突电压信号和整合的离子通道的相互作用组。目标3将探测离子通道的成像树突参与一个相互作用体，支配突触和树突钙信号。利用这些目标提供的信息，目标4将通过病毒转染来拯救与年龄相关的记忆障碍和信号异常，以上调或下调与认知障碍或保存相关的离子通道的表达。这种“拯救”将通过行为测试、2 PLSM Ca 2+成像、2 P谷氨酸解笼以及使用iFESEM和iAT的重建蛋白质组显微镜来探测。总之，拟议目标中的实验将以分子精度确定老年大鼠树突状信号传导和记忆功能的关键调节因子，揭示对“成功”和“不成功”认知衰老的深入了解，并提供有关可行药物靶点的关键信息，这些靶点可以减缓或预防与衰老相关的认知衰退。

 DESCRIPTION (provided by applicant): Though aging itself is the biggest risk factor for Alzheimer's disease (AD), many aged persons with memory problems do not ultimately suffer from AD. In such persons, pathology that would merit AD diagnosis is absent, despite the overlapping cognitive symptomatology. The principal goal of this research program is to model such "normal" age-related cognitive heterogeneity in rats and then use high-resolution techniques to reveal the synaptic and dendritic substrates of age-associated decline in forms of memory that require the hippocampus. As the primary output of the hippocampus, region CA1 has been a primary target of such investigation by us and others. Indeed, past studies have revealed that while major age-related synapse and neuron loss are absent in CA1, more subtle aging-linked changes collude to modify neuronal activity patterns and behavioral plasticity. Importantly, such alterations are present in only a subset of aged rats, with others the same chronological age showing young adult-like acquisition on Morris Water Maze (MWM) learning and trace eyeblink conditioning (TEBC). Such cognitive heterogeneity among aged rats, then, provides an important opportunity to reveal both the substrates of age-related memory decline as well as the substrates of memory preservation. The experiments proposed in this renewal application will continue to reveal the synaptic and dendritic substrates of age-related cognitive heterogeneity, but do so by building on our past findings with conceptual and technical complexity. Specifically, in Aim 1, we will behaviorally characterize young adult and aged F344xBN F1 hybrid rats with MWM learning and TEBC and then probe synaptic Ca2+ signaling with 2-photon laser scanning (2PLSM) Ca2+ imaging and 2-photon glutamate (2Pglutamate) uncaging onto spines in discrete microdomains of individual basal, oblique, and tuft dendrites. Following this high-resolution functional probing, Aims 2 and 3 will use immunogold field emission scanning electron microscopy for backscattered electrons (iFESEM) and immunofluorescence array tomography (iAT) to morphologically reconstruct the imaged dendrites and then proteomically reconstruct the dendrites' signaling networks. Aim 2 will focus on an interactome of ion channels involved in synaptic and dendritic voltage signaling and integration. Aim 3 will probe the imaged dendrites for ion channels involved in an interactome that governs synaptic and dendritic Ca2+ signaling. Using the information provided by these Aims, Aim 4 will rescue age-related memory impairments and signaling abnormalities with viral transfection to up or down-regulate expression of ion channels linked to cognitive impairment or preservation. Such "rescue" will be probed with behavioral testing, 2PLSM Ca2+ imaging, 2Pglutamate uncaging, and reconstructive proteomic microscopy with iFESEM and iAT. Together, the experiments in the proposed Aims will identify the key regulators of dendritic signaling and memory function in aged rats with molecular precision, reveal deep insight into both "successful" and "unsuccessful" cognitive aging, and provide critical information about viable drug targets that could slow or prevent aging-linked cognitive decline.