【摘要】 利用不同电刺激条件模拟神经电活动 ,研究C2C12细胞 (小鼠骨骼肌成肌细胞系 )内Ca2 + 激活的不同形式及其对nAChR基因表达活性的影响 .电刺激分化 1d的C2C12细胞 ,用共聚焦显微镜记录不同电刺激参数引起的细胞内Ca2 + 信号变化 ,共观察到 3种不同形式的Ca2 + 信号 ,称之为整体Ca2 + 振荡、局部Ca2 + 振荡和局部Ca2 + 振荡诱发的整体Ca2 + 振荡 .在此基础上 ,又构建nAChRγ亚基启动子萤光素酶报告基因质粒并转染C2C12细胞 ,进一步检测不同形式Ca2 + 引起细胞萤光素酶活性的变化 ,测定细胞内PKC活性的改变 .不同Ca2 + 振荡引起细胞内PKC活性升高幅度不同 ;电刺激C2C12细胞可导致nAChRγ基因表达活性降低 ,3种Ca2 + 振荡对nAChRγ基因启动子活性的影响无显著差异 .结果表明 ,电刺激可引起肌细胞产生不同形式Ca2 + 信号 ,但不同形式Ca2 + 信号对nAChRγ启动子活性的抑制无明显差异 ,提示Ca2 + 依赖的PKC途径可能不是nAChR基因表达下调的唯一途径更多还原

【Abstract】 By using different patterns of electrical stimulation to mimic electricity of nerve, the types of calcium oscillation and their effects on nAChR gene expression activity were observed in differentiated C2C12 cells (mouse skeletal myoblastic cell line) C2C12 cells were loaded with the Ca 2+ sensitive indicator Fluo 3 and imaged with a confocal microscope to record calcium oscillation Electrical stimuli were applied to induce different patterns of calcium oscillation of differentiated C2C12 cells transfected with pGL3 γ,which was the expression vector containing the luciferase reporter gene driven by the control region of the nAChRγ subunit gene The promoter activity of the nAChRγ subunit was examined The results showed that (1) three types of calcium oscillation in differentiated C2C12 cells were induced by different electrical stimulation: global calcium oscillation, transmission of local calcium oscillation and local calcium oscillation induced global calcium oscillation; (2) calcium oscillation enhanced protein kinase C (PKC) activities;(3) the promoter activity of the nAChRγ subunit was suppressed after electrical stimulation, but there was no difference among three patterns of electrical stimulation These results suggest that electrical stimulation can suppress nAChRγ subunit promoter activity, but there is no close relation between nAChRγ gene activities and the patterns of stimulation It is likely that calcium activated protein kinase C (PKC) dependent pathway is not the only way in the regulation of nAChRγ gene expression更多还原