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衰退有关，特别是记忆力。到2040年，65岁以上的人口将占美国人口的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，并不是所有的老年人都会在临床上被诊断为记忆障碍，这表明时间衰老和认知衰老不是同义的过程。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丢失无关；相反，海马体内突触数量的选择性减少和突触后受体的响应性降低。这些在老年人身上明显的行为和神经元后遗症可以在老年大鼠的空间位置记忆测试中成功地建模。这个项目将利用一个结合了时序和认知衰老方面的啮齿动物衰老模型来研究突触前和突触后底物的相关变化。对谷氨酸、胆碱和GABA能的海马回路和受体的平行分析将确定变化是特定于特定的神经递质系统还是老年人大脑中更普遍的变化。海马区神经支配的变化将通过免疫组织化学染色技术可视化，并使用无偏见的体视学方法进行量化。M1和mGluR5代谢性谷氨酸受体介导的GTP结合是受体偶联的一种功能指标，将使用针对其同源G蛋白1亚单位GQ/11的闪烁邻近计数技术选择性地测量。这些变化将与GABAB受体介导的结合指标进行对比，GABAB受体介导的结合不随年龄恶化，但通过GI/GO级联信号传递。在没有明显的受体和G蛋白丢失的情况下，已经假设与衰老的海马区的氧化应激相关的分子过程可能干扰了正常的信号转导。为了验证这一假设，受体和G蛋白将通过免疫沉淀和质谱仪进行分析，以确定这些特定蛋白是否受到氧化损伤。总而言之，该项目将评估与正常记忆功能密切相关的三个神经递质系统中与年龄相关的变化，并确定这些电路和突触在衰老过程中发生变化的机制。来自这些实验的数据不仅将极大地增强对大脑老化和认知的理解，还将阐明特定神经底物的变化，这些变化对下一代药物治疗的潜在治疗调节有意义。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极好的博士前培训经验，以支持其所述的职业目标和目的。 与公共健康相关：到2040年，65岁以上的人口将占美国人口的20%以上，了解老年人记忆力下降的机制是迫切需要关注的科学和临床问题。这项研究项目将测量神经信号的突触前和突触后方面，以确定与记忆缺陷有关的老化海马区代谢性受体和G蛋白的特定变化。这一项目的发现将增进我们对老龄大脑的了解，并为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.