大鼠肺动脉平滑肌细胞的分离及钾电流的记录

Isolation of pulmonary arterial smooth muscle cells and record of potassium channel currents in rats

目的分离大鼠肺动脉平滑肌细胞(PASMCs)并建立记录电压门控性钾离子通道(KV)和钙离子激活性钾通道(KCa)电流的方法。方法急性酶解法分离出单个PASMC,采用全细胞膜片钳技术记录KV和KCa电流。结果成功获得单个PASMC。在+60mV时,4-氨基吡啶5mmol/L可明显抑制KV电流,电流密度从(134.69±4.73)pA/pF减少到(35.02±5.64)pA/pF(P<0.05)。在+60mV时,四乙铵1mmol/L可明显抑制KCa电流,电流密度从(15.10±1.62)pA/pF减少到(3.82±0.72)pA/pF(P<0.05)。结论急性酶解法成功获得活性良好的PASMCs,并建立了KV、KCa电流的记录方法,为研究肺动脉高压时钾通道的电生理学改变奠定基础。

Objective To isolate pulmonary arterial smooth muscle cells （PASMCs） and establish a method for recording currents of voltage-gated potassium channel （Kv） and calcium- activated potassium channel（KCa） in rats. Methods Single PASMC was isolated by enzyme digestion and the currents of Kv and Kca were recorded by whole-cell patch-clamp technique. Results Single PASMC was successfully obtained. 4-Aminopyridine 5 mmol/L significantly inhibited Kv currents, density of which was reduced from （134. 69±4.73）pA/pF to （35.02±5.64）pA/pF at 60mV （P〈0. 05）. Tetraethylammonium i mmol/L inhibited K~ currents, density of which was reduced from （15.10±1.62） pA/pF to （3.82±0.72） pA/pF at 60 mV（P〈0.05）. Conclusion High-quality PASMCs are obtained by enzyme digestion, and the method for recording currents of Kv and Kcais established, which offers an effective cell model to explore the pathogenesis of pulmonary artery diseases.