Reduced vesicular catecholamine storage mediates oxidant-induced neurotoxicity

描述（由申请人提供）：帕金森病（PD）是一种使人衰弱的神经退行性疾病，其特征是壳核和黑质产生DA的神经元中多巴胺（DA）的丧失，导致运动障碍，如震颤、僵硬、姿势不稳定和运动迟缓。目前认为帕金森病是由环境和遗传因素共同引起的。目前的治疗方法包括左旋多巴，MAOB抑制剂和DA激动剂，这些都增加了大脑中DA信号的数量。正常情况下，DA在神经元内受到高度调控；多巴胺转运体（DAT）介导突触对多巴胺的选择性摄取，调节神经元信号的大小和持续时间。然后，囊泡单胺转运蛋白（VMAT2）将胞质DA泵入囊泡，供胞外释放。VMAT2在介导神经元对细胞损伤的易感性方面很重要，特别是通过预防氧化应激。VMAT2通常会隔离儿茶酚胺，保护神经元免受损伤；然而，当VMAT2表达减少时，DA和NE在细胞质中被氧化，引起内源性氧化损伤。在DA处理功能障碍期间，细胞抗氧化防御被激活；但当这些防御失效时，就会发生神经变性。为了研究VMAT2在PD中的作用，许多小组试图生成VMAT2 KO小鼠，但完全消融VMAT2是致命的。之前，我们的实验室已经证明，VMAT2 （VMAT2 LO）基因减少95%的小鼠表现出纹状体DA的进行性丧失、左旋多巴反应性行为缺陷、突触核蛋白聚集和黑质多巴胺细胞丧失。我们假设，随着小鼠年龄的增长，VMAT2缺陷小鼠的进行性损伤是由于氧化应激增加。本研究的目的是阐明VMAT2在PD发展中的作用，更具体地说，VMAT2在儿茶酚胺介导的毒性存在下调节神经元氧化还原状态的能力。我将通过以下方法来验证这一假设：1)确定VMAT2 LO小鼠中是否存在年龄依赖性氧化还原状态的改变，2)研究左旋多巴是否会加剧这种氧化还原状态的改变，以及3)确定NE在VMAT2 LO小鼠中氧化诱导的神经变性中的作用。鉴于氧化应激相关研究的重要性日益增加，特别是与神经退行性变有关，本研究有可能阐明人类PD发展的可能内源性机制。此外，通过本研究发现的任何氧化机制都可以作为PD疾病进展中遗传和环境因素之间的联系。

DESCRIPTION (provided by applicant): Parkinson's disease (PD) is a debilitating neurodegenerative disorder, which is characterized by the loss of dopamine (DA) in the putamen and DA producing neurons of the substantia nigra, resulting in motor disturbances such as tremor, rigidity, postural instability and bradykinesia. PD is currently thought to be caused by a combination of environmental and genetic factors. Current therapies include L-DOPA administration, MAOB inhibitors, and DA agonists, which all increase the amount of DA signaling in the brain. Normally, DA is highly regulated within the neuron; the dopamine transporter (DAT) mediates selective uptake of dopamine from the synapse, adjusting the magnitude and duration of neuronal signaling. The vesicular monoamine transporter (VMAT2) then pumps cytosolic DA into vesicles for exocytotic release. VMAT2 is important in mediating neuronal susceptibility to cellular damage, particularly via the prevention of oxidative stress. VMAT2 normally sequesters catecholamines, protecting against neuronal damage; however, when VMAT2 expression is reduced, DA and NE can become oxidized in the cytosol, inducing endogenous oxidative damage. During periods of DA handling dysfunction, cellular antioxidant defenses are activated; but when these defenses fail, neurodegeneration can occur. In order to study the role of VMAT2 in PD, many groups have tried to generate VMAT2 KO mice, but completely ablating VMAT2 is lethal. Previously, our laboratory has shown that mice with a 95% genetic reduction in VMAT2 (VMAT2 LO) display progressive loss of striatal DA, L-DOPA responsive behavioral deficits, synuclein aggregation, and nigral dopamine cell loss. We hypothesize that the progressive damage in the VMAT2 deficient mice is due to increased oxidative stress as the mice age. The goal of this research is to elucidate the role of VMAT2 in the development of PD, more specifically, the ability of VMAT2 to modulate neuronal redox state in the presence of catecholamine mediated toxicity. I will test this hypothesis by 1) determining if there is an agedependent altered redox state in VMAT2 LO mice, 2) investigating if L-DOPA administration exacerbates this altered redox status, and 3) determining the role of NE in oxidant-induced neurodegeneration in the VMAT2 LO mice. Given the increasing importance of oxidative stress related research, especially as it pertains to neurodegeneration, this research has the potential to illuminate possible endogenous mechanism for the development of PD in humans. Furthermore, any oxidative mechanisms that are uncovered through this research can serve as the link between genetic and environmental factors in PD disease progression.