Huang Hui-Ting, Liao Chih-Kai, Chiu Wen-Tai, Tzeng Shun-Fen
Institute of Life Sciences, College of Bioscience and Biotechnology, National Cheng Kung University, Tainan, Taiwan.
Department of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan.
Int J Biochem Cell Biol. 2017 May;86:42-53. doi: 10.1016/j.biocel.2017.03.008. Epub 2017 Mar 18.
Astrocytes, a stellate-shape glial population in the central nervous system (CNS), maintain glutamate homeostasis in adult CNS by undergoing glutamate uptake at the synapse through their glutamate transporter-1 (GLT-1). Peroxisome proliferator-activated receptor-α (PPARα) can be activated by endogenous saturated fatty acids to regulate astrocytic lipid metabolism and functions. However, it is unclear if PPARα can exert the regulatory action on GLT-1 expression in astrocytes. This study showed that treatment with palmitic acid (PA) and the other two PPARα agonists (GW 7647 and WY 14,643) caused no change in the morphology of astrocytes, whereas membranous GLT-1 protein levels in astrocytes were significantly decreased by PA and PPARα agonists. Through lentivirus-mediated overexpression of GLT-1 tagged with red fluorescent protein (GLT-1-RFP), we also observed that GLT-1-RFP puncta in the processes of astrocytes were inhibited by the PPARα agonists. This reduction was prevented by the addition of the PPARα antagonist, GW6471. GLT-1-RFP was co-localized to the early endosome marker-EEA1 in astrocytes treated with the PPARα agonists. Moreover, PPARα-induced inhibition in membranous GLT-1 expression was abolished by the addition of dynamin inhibitor (dynasore). Furthermore, the co-treatment of astrocytes with PPARα agonists and dynasore, or with PPARα agonists and protein kinase C (PKC) inhibitor bis-indolylmaleimide 1 (BIS1), prevented the endocytosis of GLT-1-RFP. Based on the results, we conclude that the PPARα agonists increased GLT-1 endocytosis in astrocytes possibly through the PKC signaling pathway. In addition, our findings provide important information of PPARα involvement in the downregulation of astrocytic glutamate uptake via the promoted GLT-1 endocytosis.
星形胶质细胞是中枢神经系统（CNS）中的一种星形胶质群体，通过其谷氨酸转运体-1（GLT-1）在突触处摄取谷氨酸来维持成年CNS中的谷氨酸稳态。过氧化物酶体增殖物激活受体-α（PPARα）可被内源性饱和脂肪酸激活，以调节星形胶质细胞的脂质代谢和功能。然而，尚不清楚PPARα是否能对星形胶质细胞中GLT-1的表达发挥调节作用。本研究表明，用棕榈酸（PA）和其他两种PPARα激动剂（GW 7647和WY 14,643）处理不会改变星形胶质细胞的形态，而PA和PPARα激动剂可使星形胶质细胞中的膜性GLT-1蛋白水平显著降低。通过慢病毒介导的红色荧光蛋白标记的GLT-1（GLT-1-RFP）过表达，我们还观察到PPARα激动剂可抑制星形胶质细胞突起中的GLT-1-RFP斑点。添加PPARα拮抗剂GW6471可阻止这种减少。在用PPARα激动剂处理的星形胶质细胞中，GLT-1-RFP与早期内体标记物EEA1共定位。此外，添加发动蛋白抑制剂（dynasore）可消除PPARα诱导的膜性GLT-1表达抑制。此外，将星形胶质细胞与PPARα激动剂和dynasore共同处理，或与PPARα激动剂和蛋白激酶C（PKC）抑制剂双吲哚马来酰亚胺1（BIS1）共同处理，可阻止GLT-1-RFP的内吞作用。基于这些结果，我们得出结论，PPARα激动剂可能通过PKC信号通路增加星形胶质细胞中GLT-1的内吞作用。此外，我们的研究结果提供了重要信息，即PPARα通过促进GLT-1内吞作用参与星形胶质细胞谷氨酸摄取的下调。
