Pittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh, Pittsburgh, PA, USA; Department of Neurology, University of Pittsburgh, Pittsburgh, PA, USA.
Center for Biologic Imaging, University of Pittsburgh, Pittsburgh, PA, USA; Department of Cell Biology, University of Pittsburgh, Pittsburgh, PA, USA.
Redox Biol. 2020 Oct;37:101695. doi: 10.1016/j.redox.2020.101695. Epub 2020 Aug 22.
Convergent evidence implicates impaired mitochondrial function and α-Synuclein accumulation as critical upstream events in the pathogenesis of Parkinson's disease, but comparatively little is known about how these factors interact to provoke neurodegeneration. We previously showed that α-Synuclein knockdown protected rat substantia nigra dopaminergic neurons from systemic exposure to the mitochondrial complex I inhibitor rotenone. Here we show that motor abnormalities prior to neuronal loss in this model are associated with extensive α-Synuclein-dependent cellular thiol oxidation. In order to elucidate the underlying events in vivo we constructed novel transgenic zebrafish that co-express, in dopaminergic neurons: (i) human α-Synuclein at levels insufficient to provoke neurodegeneration or neurobehavioral abnormalities; and (ii) genetically-encoded ratiometric fluorescent biosensors to detect cytoplasmic peroxide flux and glutathione oxidation. Live intravital imaging of the intact zebrafish CNS at cellular resolution showed unequivocally that α-Synuclein amplified dynamic cytoplasmic peroxide flux in dopaminergic neurons following exposure to the mitochondrial complex I inhibitors MPP or rotenone. This effect was robust and clearly evident following either acute or prolonged exposure to each inhibitor. In addition, disturbance of the resting glutathione redox potential following exogenous hydrogen peroxide challenge was augmented by α-Synuclein. Together these data show that α-Synuclein is a critical determinant of the redox consequences of mitochondrial dysfunction in dopaminergic neurons. The findings are important because the mechanisms underlying α-Synuclein-dependent reactive oxygen species fluxes and antioxidant suppression might provide a pharmacological target in Parkinson's disease to prevent progression from mitochondrial dysfunction and oxidative stress to cell death.
汇集的证据表明，线粒体功能障碍和α-突触核蛋白积累是帕金森病发病机制中的关键上游事件，但关于这些因素如何相互作用引发神经退行性变，人们知之甚少。我们之前表明，α-突触核蛋白敲低可保护大鼠黑质多巴胺能神经元免受系统性暴露于线粒体复合物 I 抑制剂鱼藤酮的影响。在这里，我们表明，在该模型中神经元丢失之前出现的运动异常与广泛的α-突触核蛋白依赖性细胞硫醇氧化有关。为了阐明体内的潜在事件，我们构建了新型转基因斑马鱼，在多巴胺能神经元中共同表达：（i）水平不足以引发神经退行性变或神经行为异常的人α-突触核蛋白；（ii）用于检测细胞质过氧化物通量和谷胱甘肽氧化的遗传编码比率荧光生物传感器。在完整的斑马鱼中枢神经系统中以细胞分辨率进行的活体体内成像毫不含糊地表明，α-突触核蛋白在暴露于线粒体复合物 I 抑制剂 MPP 或鱼藤酮后放大了多巴胺能神经元中的动态细胞质过氧化物通量。这种作用是强大的，并且在每种抑制剂的急性或长期暴露后都非常明显。此外，α-突触核蛋白增强了外源性过氧化氢挑战后静止谷胱甘肽氧化还原电位的干扰。这些数据共同表明，α-突触核蛋白是多巴胺能神经元中线粒体功能障碍的氧化还原后果的关键决定因素。这些发现很重要，因为α-突触核蛋白依赖性活性氧通量和抗氧化剂抑制的机制可能为帕金森病提供药理学靶点，以防止从线粒体功能障碍和氧化应激进展到细胞死亡。
