Raj Aishwarya, Kaushal Alka, Datta Indrani
Department of Biophysics, National Institute of Mental Health and Neurosciences, Institute of National Importance, Bengaluru, India.
J Neurosci Res. 2022 Feb;100(2):681-706. doi: 10.1002/jnr.24994. Epub 2021 Dec 13.
Serving as a source of glutathione and up-taking and metabolizing glutamate are the primary supportive role of astrocytes for the adjacent neurons. Despite the clear physical association between astrocytes and α-synuclein, the effect of extracellular α-synuclein on these astrocytic functions has not yet been elucidated. Hence, we aim to assess the effect of various forms of α-synuclein on antioxidant mechanism and glutamate metabolism. Wild-type and A53T/A30P double-mutant α-synuclein, both in monomeric and aggregated forms, were added extracellularly to media of midbrain rat astrocyte culture, with their survival, oxidative, and nitrative stress, glutathione and glutamate content, expression of enzymes associated with oxidative stress and glutamate metabolism, glutamate and glutathione transporters being assessed along with the association/engulfment of these peptides by astrocytes. A30P/A53T peptide associated more with astrocytes, and low-extracellular K concentration showed prominent reduction in the engulfment of the monomeric forms, suggesting that the association of the aggregated forms was greater with the membrane. The peptide-associated astrocytes showed lower survival and increased oxidative stress generation, owing to the decrease in nuclear localization of Nrf2 and increase in iNOS, and further aggravated by the decrease in glutathione content and related enzymes like glutathione synthetase, glutathione peroxidase, and glutathione reductase. Glutamate uptake increased in aggregate-treated cells due to the increase in GLAST1 expression, de novo synthesis of glutamate by pyruvate carboxylase, and/or glutamine synthase, bolstered by the differential glutamate dehydrogenase enzyme activity. We thus show for the first time that extracellular α-synuclein exposure leads to astrocytic dysfunction with respect to the antioxidant mechanism and glutamate metabolic profile. The impact was higher in the case of the aggregated and mutated peptide, with the highest dysfunction for the mutant aggregated α-synuclein treatment.
作为谷胱甘肽的来源以及摄取和代谢谷氨酸是星形胶质细胞对相邻神经元的主要支持作用。尽管星形胶质细胞与α-突触核蛋白之间存在明显的物理关联，但细胞外α-突触核蛋白对这些星形胶质细胞功能的影响尚未阐明。因此，我们旨在评估各种形式的α-突触核蛋白对抗氧化机制和谷氨酸代谢的影响。将野生型和A53T/A30P双突变α-突触核蛋白的单体和聚集形式细胞外添加到中脑大鼠星形胶质细胞培养介质中，同时评估它们的存活、氧化和硝化应激、谷胱甘肽和谷氨酸含量、与氧化应激和谷氨酸代谢相关的酶的表达、谷氨酸和谷胱甘肽转运体以及星形胶质细胞对这些肽的结合/吞噬情况。A30P/A53T肽与星形胶质细胞的结合更多，低细胞外钾浓度显示单体形式的吞噬显著减少，表明聚集形式与膜的结合更强。与肽结合的星形胶质细胞显示存活率降低和氧化应激产生增加，这是由于Nrf2核定位减少和诱导型一氧化氮合酶增加所致，谷胱甘肽含量和谷胱甘肽合成酶、谷胱甘肽过氧化物酶和谷胱甘肽还原酶等相关酶的减少进一步加剧了这种情况。由于GLAST1表达增加、丙酮酸羧化酶和/或谷氨酰胺合成酶从头合成谷氨酸，并受到谷氨酸脱氢酶活性差异的支持，聚集处理细胞中的谷氨酸摄取增加。我们首次表明，细胞外暴露于α-突触核蛋白会导致星形胶质细胞在抗氧化机制和谷氨酸代谢谱方面出现功能障碍。在聚集和突变肽的情况下影响更高，突变聚集α-突触核蛋白处理导致的功能障碍最为严重。
