Oxidative damage to receptor: G-protein coupling in the aged hippocampus

描述（由申请人提供）：衰老与许多认知领域的下降有关，特别是记忆力。由于到2040年，65岁以上的人将占美国人口的20%以上，因此迫切需要阐明衰老损害记忆的机制，并确定改进的治疗干预途径。重要的是，对人类的神经心理学评估表明，并非所有老年人都会被临床诊断为记忆障碍，这表明时间和认知老化不是同义的过程。与阿尔茨海默病不同，与年龄相关的记忆变化与广泛的神经元损失无关;相反，海马内突触数量选择性减少，突触后受体的反应性降低。这些行为和神经元的后遗症明显的老年人可以成功地模拟老年大鼠的空间位置的记忆测试。本研究将利用啮齿类动物的老化模型，结合时序老化和认知老化的各个方面，来研究突触前和突触后基质的相关变化。对海马神经元、胆碱能和γ-氨基丁酸能回路和受体的平行分析将确定变化是否是特定神经递质系统的特异性变化，还是老年大脑中更普遍的变化。通过免疫组织化学染色技术观察海马神经支配的变化，并使用无偏体视学方法进行定量。M1毒蕈碱和mGluR 5代谢型谷氨酸受体介导的GTP结合（受体偶联的功能性测量）将使用其同源G蛋白1亚基Gq/11特异性闪烁邻近计数技术选择性测量。这些变化将通过GABAB受体介导的结合的测量进行对比，GABAB受体介导的结合不随年龄而恶化，而是通过Gi/Go级联信号。在没有明显的受体和G-蛋白的损失，它已被假设，与氧化应激相关的分子过程中的老年海马可能会干扰正常的信号转导。为了验证这一假设，将通过免疫沉淀和质谱分析受体和G蛋白，以确定这些特定的蛋白质是否受到氧化损伤。总的来说，该项目将评估与正常记忆功能密切相关的三种神经递质系统中与年龄相关的变化，并确定这些回路和突触在衰老中发生变化的机制。从这些实验中获得的数据不仅将大大增强对大脑衰老和认知的理解，而且还将阐明下一代药物治疗对潜在治疗调节感兴趣的特定神经基质的变化。此外，为解决这些重点研究工作而提出的实验方法的多样性将为申请人提供出色的博士前培训经验，以支持他所陈述的职业目标和目的。 公共卫生相关性：到2040年，65岁以上的人将占美国人口的20%以上，因此了解老年人记忆力下降的机制是迫切的科学和临床问题。本研究项目将测量神经信号的突触前和突触后方面，以确定与记忆缺陷相关的老年海马体中代谢型受体和G蛋白的具体变化。该项目的发现将增强我们对老年大脑的理解，并为治疗治疗潜力提供新的见解。

DESCRIPTION (provided by applicant): Aging is associated with declines in a number of cognitive domains, particularly memory. As people over the age of 65 will comprise more than 20% of the US population by the year 2040, there is a pressing need to elucidate the mechanisms by which aging impairs memory and to identify improved routes of therapeutic intervention. Importantly, neuropsychological assessments of humans demonstrate that not all aged individuals will be clinically diagnosed with memory impairment, demonstrating that chronological and cognitive aging are not synonymous processes. Unlike Alzheimer's disease, age-related changes in memory are not associated with wide-spread neuronal loss; rather there is a selective decrease in the numbers of synapses within the hippocampus and decreased responsivity of post-synaptic receptors. These behavioral and neuronal sequelae apparent in aged humans can be successfully modeled in aged rats tested for memory of spatial locations. This project will make use of a rodent model of aging integrating aspects of chronological and cognitive aging to investigate associated changes in presynaptic and postsynaptic substrates. Parallel analysis of glutamatergic, cholinergic and GABAergic hippocampal circuits and receptors will determine if changes are specific to a particular neurotransmitter system or more general changes in the aged brain. Changes to innervation of the hippocampus will be visualized by immunohistochemical staining techniques and quantified using unbiased stereological approaches. M1 muscarinic- and mGluR5 metabotropic glutmate receptor-mediated GTP-binding, a functional measure of receptor coupling, will be selectively measured using a scintillation proximity counting technique specific to their cognate G-protein 1-subunit, Gq/11. These changes will be contrasted by measures of GABAB receptor-mediated binding which do not deteriorate with age, but signal via a Gi/Go cascade. In the absence of overt loss of receptor and G-proteins, it has been hypothesized that molecular processes associated with oxidative stress in the aged hippocampus may interfere with normal signal transduction. To test this hypothesis, receptors and G-proteins will be analyzed by immunoprecipitation and mass spectrometry to determine if these specific proteins are oxidatively damaged. Collectively, this project will evaluate age-related changes in three neurotransmitter systems closely tied to normal memory function and determine the mechanisms by which these circuits and synapses are changed in aging. The data derived from these experiments will not only greatly enhance understanding of brain aging and cognition, but elucidate changes to specific neural substrates that are of interest for potential therapeutic modulation by the next generation of pharmaceutical treatments. Furthermore, the diversity of experimental approaches proposed to address these focused research efforts will provide for an excellent pre-doctoral training experience to the applicant in support of his stated career goals and objectives. PUBLIC HEALTH RELEVANCE: As people over the age of 65 will comprise more than 20% of the U.S. population by 2040, appreciating the mechanisms by which memory declines in the elderly is matter of pressing scientific and clinical concern. This research project will measure presynaptic and postsynaptic aspects of neural signaling to determine specific changes to metabotropic receptors and G-proteins in the aged hippocampus associated with memory deficits. The findings of this project will enhance our understanding of the aged brain and provide new insights for therapeutic treatment potential.