新生大鼠心肌细胞培养及电生理特性观察

Method for culturing neonatal rat cardiac myocytes and observation of its electrophysiological properties

目的应用原代细胞培养技术,以获取大量具有正常电生理特性的心肌细胞。方法用胶原酶溶液处理1 d龄SD大鼠的心脏,差速贴壁法纯化获得的单个心肌细胞,CO2培养箱中培养8 d后,以膜片钳技术记录心肌细胞的动作电位和各跨膜离子流。结果细胞培养中,改进了酶溶液的成分、浓度和经典的差速贴壁法作用时间,获得了大量的高纯度单个心肌细胞。在此细胞上,采用电流钳技术,可引导出心室肌动作电位。运用心肌细胞离子通道的电压依赖性和特异阻断剂,在不同的钳制电位和指令电位下,可记录到INa,ICa,Ito,IK等多种离子流。结论改进的细胞培养技术可获得大量的具有正常电生理特性的心肌细胞。

Objective To harvest a great number of cultured cardiac myocytes with normal electrophysiological properties. Methods The hearts of neonatal rats （ SD rats, 1 d） were processed by the collagenase-contained solution. Twice-centrifugations were used to separate the myocytes from the cell suspension. After incubation for 8 days, the action potentials and transmembrance ionic current were recorded by the patch clamp technique. Results After modifying the collagenase-contained solution and time for centrifugation, large quantity of single isolated myocytes were obtained. The patch clamp experiment demonstrated that these cells possessed various ion channels, such as Na^＋ , K^＋ , Ca^2＋ and transient outward current （I10） , the activities of which led to the formation of normal action potentials. Conclusion The modified cell-culture mothod can provide large number of cardiac myocytes that bear the normal electrophysiological characteristics.