大鼠肺动脉平滑肌细胞的分离及电压门控性钾电流的记录方法

Isolation and Recording of Voltage Gated Potassium Channel of Pulmonary Arterial Smooth Muscle Cells in Rat

目的介绍一种大鼠肺动脉平滑肌细胞(PASMCs)的急性分离方法并观察电压门控性钾电流电生理特性。方法应用胶原酶和木瓜蛋白酶联合消化法获得大鼠PASMCs,利用全细胞膜片钳技术记录PASMCs膜上的电压门控性钾通道(Kv)电流。结果在相差显微镜下观察大鼠PASMCs呈舒展梭形,边界清晰,有完整的细胞膜,胞浆均匀,数量多,活性好。结论用酶急性分离的大鼠PASMCs,容易进行全细胞膜片钳记录,方法简单、稳定、可靠。

Objective: To introduce a simple method of acutely preparing of isolated pulmonary arterial smooth muscle cells and to study the electrophysiological characteristics of voltage-gated potassium channels.Method: Single isolated pulmonary arterial smooth muscle cells were obtained by collagenase and papain digestion method and the whole cell patch clamp recording technique was used to record the voltage-gated potassium channels.Results: The morphological of isolated cells appeared to be typical fusiform shape by phase-contrast microscope. The cell membrane was integrity and nucleus was slick. The isolating method could produce adequate cells in normal form. The cells with smooth borders were chosen for whole-cell patch clamp recording.Conclusion: The established method of isolation was simple, stable and reliable.