GLIAL ACTIVATION IN AGED MEMORY IMPAIRED RATS

本申请的重点是PAR-98-021中列出的主题#23)神经系统中的非神经细胞。PI是一名新的研究员，他正试图在老龄化研究领域建立自己的职业生涯。本研究旨在研究衰老过程中可能导致氧化应激的神经胶质细胞激活，并探讨线粒体DNA(MtDNA)损伤与老年啮齿动物认知功能障碍等生物标志物的关系。这一建议与我之前的研究有关，即老年Long-Evans大鼠b-淀粉样蛋白前体(APP)、锰超氧化物歧化酶(Mn-SOD)和胶质纤维酸性蛋白(GFAP)的基因表达水平。所有上述指标均与该模型动物的记忆障碍程度相关。这些结果表明，在衰老过程中存在氧化应激和神经胶质细胞的激活。我们的初步实验还显示，线粒体DNA在老年记忆受损的大鼠中受损。线粒体DNA是一种裸露的环状双链DNA，由于在复制过程中缺乏校对，以及DNA修复系统缺乏组织性，它极易受到氧化应激的影响。考虑到衰老过程中氧化应激的产生，线粒体DNA损伤的积累可能在衰老和疾病的神经退变过程中发挥重要的病理生理作用。已知激活的神经胶质细胞会产生氧化应激，同时它们通过产生神经因子来支持神经元的活动。事实上，我们观察到，在一些老年记忆受损的大鼠中，一种不可磨灭的一氧化氮基因的表达水平要高得多。因此，我们假设，在衰老过程中，胶质细胞的激活通过氧化损伤而导致记忆损伤。为了研究这一假设，我们提出了以下研究。我们将检测年轻和老年大鼠胶质细胞活化前后GFAP、Mn-SOD和APP基因的表达。我们将向脑室注射脂多糖(LPS)来激活胶质细胞。注射前后用Morris水迷宫进行空间记忆测试。我们还将使用定量聚合酶链式反应检测这些动物的线粒体DNA损伤。这种定量PCR检测DNA损伤的方法是基于这样的证据，即几个DNA损伤能够阻止Taq DNA聚合酶的进展，从而导致PCR扩增减少。我们将对这些生物和行为数据进行线性回归分析和方差分析。我们预计，衰老和神经胶质细胞的激活会增加候选基因的表达和线粒体DNA的损伤，生物标志物的水平可能与记忆障碍有关。这一建议使我们能够准备初步的数据，以扩大研究，以调节衰老过程中的胶质细胞活动。随后，如果我们在该动物模型中建立记忆障碍与神经胶质细胞激活之间的关系，将是发展基于神经胶质细胞活性调节的NOEL疗法的一个很好的工具。

This application is focused on the topic #23) non-neuronal cells in the nervous system listed in PAR-98-021. The PI is a new investigator who is trying to establish his career in the aging study field. This proposal is to study the glial activation which may cause oxidative stress during the aging process and to seek the relationship of mitochondrial DNA (mtDNA) damage to cognitive impairment and other biological markers in aged rodent models. This proposal is related to my previous study , the levels of gene expression for b-amyloid protein precursor (APP), manganese super oxide dismutase (Mn-SOD) and glial fibrillary acidic protein (GFAP) in aged Long-Evans rats. All of the above markers were correlated with the degree of memory impairment in this model animal. These results indicate the presence of oxidative stress and glial activation during the aging process. Our preliminary experiment also revealed mtDNA damaged in aged memory impaired rats. MtDNA is a naked circular double-stranded DNA and due to the lack of proofreading during the replication and a less organized DNA repair system, it is extremely vulnerable to oxidative stress. Considering the production of oxidative stress during aging, accumulation of mtDNA damage may play an important pathophysiological role in the neurodegenerative process of aging and diseases. The activated glial cells are known to produce oxidative stress, while they support neuronal activity by the production of the nerve factors. In fact, we observed a much higher level of an indelible type nitric oxide gene expression in some of the aged memory impaired rats. Thus, we hypothesized that glial activation causes memory impairment by oxidative damaged during the aging process. To investigate this hypothesis, we propose the following studies. We will measure the GFAP, Mn-SOD and APP genes expression in young and aged rats with/without glial activation. We will inject lipopoly saccharide (LPS) into the cerebro-ventricle to activate the glia. The spatial memory will be tested by the Morris water maze before and after the injection. We will also measure mtDNA damage in these animals using quantitative PCR. This quantitative PCR assay to detect DNA damage is based on the evidence that several DNA lesions are able to block the progression of Taq DNA polymerase which results in a decreased PCR amplification. We will analyze these biological and behavioral data by linear regression analysis and ANOVA. We would expect that the candidate genes expression and the mtDNA damage are increased by the aging and glial activation and that the level of biological markers may be correlated with the memory impairment. This proposal allows us to prepare the preliminary data to expand the study to regulate the glial activity during the aging process. Subsequently, if we establish a relationship between memory impairment and glial activation in this animal model, it will be a good tool to develop noel therapeutics based on regulation of glial activity.