Western Michigan University, Department of Biological Sciences, Kalamazoo, MI 49008, USA.
Neuropharmacology. 2013 Oct;73:111-21. doi: 10.1016/j.neuropharm.2013.05.020. Epub 2013 May 29.
The objective of this study was to determine the neuroprotective role of tropisetron on retinal ganglion cells (RGCs) as well as to explore the possible mechanisms associated with alpha7 nAChR-induced neuroprotection. Adult pig RGCs were isolated from all other retinal tissue using a two-step panning technique. Once isolated, RGCs were cultured for 3 days under control untreated conditions, in the presence of 500 μM glutamate to induce excitotoxicity, and when tropisetron was applied before glutamate to induce neuroprotection. 500 μM glutamate decreased RGC survival by an average of 62% compared to control conditions. However, RGCs pretreated with 100 nM tropisetron before glutamate increased cell survival to an average of 105% compared to controls. Inhibition studies using the alpha7 nAChR antagonist, MLA (10 nM), support the hypothesis that tropisetron is an effective neuroprotective agent against glutamate-induced excitotoxicity; mediated by α7 nAChR activation. ELISA studies were performed to determine if signaling cascades normally associated with excitotoxicity and neuroprotection were up- or down-regulated after tropisetron treatment. Tropisetron had no discernible effects on pAkt levels but significantly decreased p38 MAPK levels associated with excitotoxicity from an average of 15 ng/ml to 6 ng/ml. Another mechanism shown to be associated with neuroprotection involves internalization of NMDA receptors. Double-labeled immunocytochemistry and electrophysiology studies provided further evidence that tropisetron caused internalization of NMDA receptor subunits. The findings of this study suggest that tropisetron could be an effective therapeutic agent for the treatment of degenerative disorders of the central nervous system that involves excitotoxicity.
本研究旨在确定托烷司琼对视网膜神经节细胞（RGC）的神经保护作用，并探讨与α7 nAChR 诱导的神经保护相关的可能机制。使用两步淘选技术从所有其他视网膜组织中分离出成年猪 RGC。一旦分离出来，RGC 在未处理的对照条件下培养 3 天，在存在 500μM 谷氨酸的情况下诱导兴奋性毒性，并且在谷氨酸之前应用托烷司琼以诱导神经保护。与对照条件相比，500μM 谷氨酸使 RGC 存活率平均降低了 62%。然而，在用谷氨酸预处理之前用 100nM 托烷司琼预处理的 RGC 将细胞存活率提高到平均 105%，与对照相比。使用α7 nAChR 拮抗剂 MLA（10nM）进行的抑制研究支持这样的假设，即托烷司琼是一种有效的神经保护剂，可抵抗谷氨酸诱导的兴奋性毒性；通过α7 nAChR 激活介导。进行 ELISA 研究以确定在托烷司琼处理后与兴奋性毒性和神经保护相关的信号级联是否上调或下调。托烷司琼对 pAkt 水平没有明显影响，但显著降低了与兴奋性毒性相关的 p38 MAPK 水平，从平均 15ng/ml 降低到 6ng/ml。与神经保护相关的另一种机制涉及 NMDA 受体的内化。双标记免疫细胞化学和电生理学研究进一步提供了证据表明托烷司琼导致 NMDA 受体亚基内化。这项研究的结果表明，托烷司琼可能是治疗涉及兴奋性毒性的中枢神经系统退行性疾病的有效治疗剂。
