VGLUT2调节谷氨酸-谷氨酰胺循环在锰诱导的帕金森病模型的作用研究

Parkinson’s disease (PD) is a chronic, progressive, disabling neurodegenerative disorder that begins in mid to late life and is characterized by motor impairment, olfactory disturbances, sleep disorders, autonomic dysfunction and depression. The cause of PD is unknown, currently considered to be associated with aging, environment and genetic factors. Epidemiologic studies have found increased risk of PD associated with exposure to environmental toxicants such as pesticides, solvents, metals, and other pollutants. Excess accumulation of manganese (Mn) in the brain results in a neurological syndrome like PD. Recently, scientists have debated the possibility that Mn may have an etiological role in PD or that it may accelerate the expression of PD. In this study, we use the chronic Mn exposure mice to explore the pathogenesis of Mn neurotoxicity and found that VGLUT2 showed a dramatic increase in the Mn exposure mice. Recombinant retroviral vectors and drugs are constructed to regulate VGLUT2 levels both in vivo and in vitro, to investigate the mechanism of VGLUT2 regulated glutamate-glutamine cycle in the neurotoxicity of Mn. These findings will provide theoretical basis for the pathogenesis of Mn neurotoxicity and PD, give new insight into the treatment of these diseases.

帕金森病（Parkinson’s disease，PD）是一种好发于中老年的常见神经系统退行性疾病，临床表现包括运动障碍、嗅觉障碍、睡眠障碍、自主神经功能障碍及情绪障碍等。PD发病机制迄今尚不明确，可能与老龄化、环境和遗传因素密切相关。流行病学发现，暴露于环境毒物（如农药，有机溶剂，金属和其他污染物）能使PD患病风险增加。锰在脑内的蓄积能导致类似帕金森病的症状。最近有研究指出脑内锰的蓄积可能是PD的直接病因或加速PD进展的重要危险因素。本研究以慢性锰中毒PD小鼠为模型，拟探讨锰的神经毒性损伤的机制。前期工作中我们发现了囊泡谷氨酸转运体2（VGLUT2）在锰中毒小鼠中显著高表达。本研究拟从细胞和动物两个层面，通过构建重组慢病毒干预VGLUT2基因的表达或相关药物处理，探讨VGLUT2调控谷氨酸-谷氨酰胺循环在锰的神经毒性损伤的机制，为探索锰中毒及PD的发病机制提供了理论依据和新的治疗靶点。

成年腹侧被盖区多巴胺（Dopamine, DA）神经元部分子集表达囊泡谷氨酸转运体2（Vesicular Glutamate Transporters 2, VGLUT2），并在纹状体释放谷氨酸作为第二种神经递质。最近的研究表明，由MPTP、 6-羟基多巴胺、鱼藤酮等神经毒素产生的细胞应激可上调存活DA神经元中的VGLUT2，表明其可能在DA神经元存活中发挥作用，推测可能与VGLUT2和谷氨酸共释放对线粒体功能的保护机制有关。在前期工作中我们发现在转录水平VGLUT2在锰诱导的帕金森病（Parkinson’s disease，PD）模型小鼠中显著高表达。本研究中，一方面我们进一步探索锰诱导帕金森模型小鼠的致病机制，以及猛神经毒性损伤后存活的DA神经元中VGLUT2在各脑区的表达及其潜在的作用机制，结果表明锰诱导PD小鼠模型组中脑黑质部位酪氨酸羟化酶(Tyrosine Hydroxylase, TH)阳性神经元显著减少，锰诱导PD小鼠中脑黑质内VGLUT2表达较对照组增高，而在纹状体内表达量无明显差异，且PD模型小鼠中脑组织谷氨酸（Glutamate, Glu）及γ-氨基丁酸（γ-Aminobutyric Acid, GABA）水平较对照组增高；另一方面我们在亚慢性锰中毒PD小鼠建模前予以选择性VGLUTs抑制剂CSB6B，进一步探索DA神经元VGLUT2表达和谷氨酸共释放对选择性DA神经元脆弱性影响，我们发现从DA神经元中选择性抑制VGLUT2表达并没有显著改变中脑细胞外DA和Glu的基础水平，但减弱了猛神经毒性诱导的细胞外DA和Glu水平的增加。这些发现表明VGLUT2在猛神经毒性诱导的 DA神经元的存活发挥了重要作用，DA神经元的谷氨酸共释放可能参与了VGLUT2的保护机制。进一步阐明这些机制可能有助于我们寻找增强DA神经元恢复力的方法，从而减缓甚至预防DA神经退行性变。