Department of Pharmacy, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Acta Pharmacol Sin. 2013 Jul;34(7):886-92. doi: 10.1038/aps.2013.42. Epub 2013 May 20.
To investigate the actions of the muscarinic agonist carbachol on glutamate-induced neurotoxicity in PC12 cells, and the underlying mechanisms.
PC12 cells were treated with different concentrations of glutamate for 24 or 48 h. The cell viability was measured using MTT assay, and the expression and activation of GSK-3β were detected with Western blot. β-Catenin translocation was detected using immunofluorescence. Luciferase reporter assay and real-time PCR were used to analyze the transcriptional activity of β-catenin.
Glutamate (1, 3, and 10 mmol/L) induced PC12 cell death in a dose-dependent manner. Moreover, treatment of the cells with glutamate (1 mmol/L) caused significant overactivation of GSK-3β and prevented β-catenin translocation to the nucleus. Pretreatment with carbachol (0.01 μmol/L) blocked glutamate-induced cell death and GSK-3β overactivation, and markedly enhanced β-catenin transcriptional activity.
Activation of muscarinic receptors exerts neuroprotection in PC12 cells by attenuating glutamate-induced GSK-3β overactivation, suggesting potential benefits of muscarinic agonists for Alzheimer's disease.
研究毒蕈碱激动剂卡巴胆碱对 PC12 细胞中谷氨酸诱导的神经毒性的作用及其机制。
用不同浓度的谷氨酸处理 PC12 细胞 24 或 48 小时。用 MTT 法检测细胞活力，用 Western blot 检测 GSK-3β的表达和激活。用免疫荧光法检测 β-连环蛋白易位。用荧光素酶报告基因检测和实时 PCR 分析 β-连环蛋白的转录活性。
谷氨酸（1、3 和 10mmol/L）呈剂量依赖性诱导 PC12 细胞死亡。此外，用谷氨酸（1mmol/L）处理细胞会导致 GSK-3β过度激活，并阻止 β-连环蛋白向核内易位。用卡巴胆碱（0.01μmol/L）预处理可阻断谷氨酸诱导的细胞死亡和 GSK-3β过度激活，并显著增强 β-连环蛋白的转录活性。
毒蕈碱受体的激活通过减轻谷氨酸诱导的 GSK-3β过度激活对 PC12 细胞发挥神经保护作用，提示毒蕈碱激动剂可能对阿尔茨海默病有益。
