ACh neuroprotection against excitotoxicity in RGCs

描述（由申请方提供）：兴奋性毒性是中枢神经系统中过量兴奋性神经递质释放通过细胞凋亡机制破坏神经元的过程。在哺乳动物视网膜中，过量的谷氨酸释放已被证明参与视网膜神经节细胞（RGC）死亡，并与几种视网膜疾病状态相关，包括青光眼、糖尿病性视网膜病变和视网膜缺血。最近，该实验室的研究表明，乙酰胆碱（ACh）通过激活烟碱乙酰胆碱受体（nAChRs）在离体成年猪视网膜神经节细胞中保护谷氨酸诱导的兴奋性毒性。然而，参与神经保护的nAChR的类型以及将nAChR的激活与神经保护联系起来的机制尚不清楚。基于从该实验室获得的初步结果，将在本提案中测试的假设是，猪RGC上的nAChR的激活启动信号级联，以减少由谷氨酸诱导的兴奋性毒性引起的细胞死亡。本研究的目的有三：1）鉴定和鉴定乙酰胆碱（ACh）对谷氨酸兴奋性毒性的神经保护作用相关的nAChR亚单位; 2）为了检验通过nAChR的钙渗透是发生神经保护所必需的假设，和3）为了检验第二信使途径将nAChR活化与谷氨酸的神经保护联系起来的假设，诱发兴奋性毒性。使用免疫细胞化学，药理学，电生理和钙成像技术相结合，从这项研究中获得的结果应确定类型的乙酰胆碱受体亚基负责乙酰胆碱在猪视网膜神经节细胞的神经保护，并将表明第二信使级联参与乙酰胆碱的神经保护作用。理解视网膜中AChR神经保护的机制可以最终导致减少由于兴奋性毒性引起的细胞死亡和治疗视网膜中的兴奋性毒性相关疾病，例如青光眼，以及CNS中的兴奋性毒性相关疾病，例如阿尔茨海默病。在哺乳动物视网膜中，过量的谷氨酸释放已被证明参与视网膜神经节细胞死亡，并与某些视网膜疾病状态相关，包括青光眼、糖尿病性视网膜病变和视网膜缺血。本研究旨在确定和分析视网膜神经保护机制，防止视网膜神经节细胞因兴奋性毒性而死亡。了解神经保护机制对兴奋性毒性是必不可少的任何未来的治疗干预，导致治疗兴奋性毒性在视网膜和其他地区的中枢神经系统。神经保护治疗最终可以改善世界上数百万人的生活质量。

DESCRIPTION (provided by applicant): Excitotoxicity is the process by which excess excitatory neurotransmitter release in the central nervous system destroys neurons through an apoptotic mechanism. In the mammalian retina, excess glutamate release has been shown to be involved in retinal ganglion cell (RGC) death and is associated with several retinal disease states, including glaucoma, diabetic retinopathy and retinal ischemia. Recently, studies from this lab have demonstrated that acetylcholine (ACh) protects against glutamate-induced excitotoxicity in isolated adult pig retinal ganglion cells through activation of nicotinic acetylcholine receptors (nAChRs). However, the types of nAChRs involved in neuroprotection and the mechanism linking activation of nAChRs to neuroprotection is unknown. Base on preliminary results obtained from this lab, the hypothesis that will be tested in this proposal is that activation of nAChRs on pig RGCs initiates a signaling cascade to reduce cell death caused by glutamate-induced excitotoxicity. Three specific aims have been designed to address this hypothesis: 1) To identify and characterize the nAChR subunits associated with ACh's neuroprotective effect against glutamate-induced excitotoxicity in isolated adult pig RGCs; 2) To test the hypothesis that calcium permeation through nAChRs is required for neuroprotection to occur and 3) To test the hypothesis that second messenger pathways link nAChR activation to neuroprotection of glutamate-induced excitotoxicity. Using a combination of immunocytochemical, pharmacological, electrophysiological and calcium imaging techniques, the results obtained from this study should identify the types of nAChR subunits responsible for ACh neuroprotection in pig retinal ganglion cells and will indicate which second messenger cascades are involved in ACh's neuroprotective effect. Understanding the mechanism of AChR neuroprotection in the retina can ultimately lead to reduced cell death due to excitotoxicity and treatment for excitotoxicity- linked diseases in the retina, such as glaucoma, as well as excitotoxic-linked diseases in the CNS, such as Alzheimer's disease. In the mammalian retina, excess glutamate release has been shown to be involved in retinal ganglion cell death and is associated with certain retinal disease states including glaucoma, diabetic retinopathy and retinal ischemia. This study proposes to identify and analyze a mechanism of neuroprotection in the retina that prevents retinal ganglion cells from dying due to excitotoxicity. Understanding the mechanism of neuroprotection against excitotoxicity is essential for any future therapeutic intervention leading to treatment of excitotoxicity in the retina and other regions of the CNS. Neuroprotection treatment can ultimately improve the quality of life for millions of people in the world.