关附庚素对HERG钾通道电流及通道动力学的影响

目的 用膜片钳全细胞记录法评价关附庚素（GFG）对HERG基因表达的快速激活延迟整流钾通道电流（IKr）及通道动力学的影响。方法 使用脂质体介导的瞬时转染法把野生型HERG基因转染入人胚肾细胞（HEK293）,采用标准的全细胞膜片钳技术记录IKr通道电流,观察不同浓度（3、12.5、50、200 和1000 Umol/L）的GFG对IKr和通道动力学的作用。 结果GFG对IKr的尾电流（Itail）具有浓度依赖性抑制作用,半数最大抑制浓度为17.9滋mol/L,Hill常数为0.75。 50 Umol/L GFG使得刺激电流（Istep）和Itail的最大峰值电位前移,呈电压依赖性,但不改变激活电位。 50 Umol/L GFG对IKr具有时间依赖性阻断效应,时间延长抑制效果增强。 50 Umol/L GFG使激活曲线左移,半数激活电压从（-2.15±1.94） mV改变为（-9. 22±2.43） mV（n = 5,P〈0. 05）,但对曲线的斜率因子k无显著影响;50 Umol/L GFG使得稳态失活曲线左移,半数失活电压从（-56.9±1.2） mV改变为（-64.2±1.5） mV（n=6,P〈0. 05）,但不影响曲线斜率因子k。50 Umol/L GFG加快通道的失活,在-60 mV以上能加快通道的失活后恢复时间,并可显著减慢通道的去激活时间。 结论 GFG对HERG基因表达的IKr通道电流具有浓度、电压和时间依赖性阻断作用,主要是通过结合通道的激活态和失活态发挥抑制效应。

The effect of Guanfu Base G on potassium current encoded by the HERG gene.

Objective To evaluate the electrophysiogical effects and of Guanfu Base G （GFG） on the rapid form of delayed rectifier potassium current （IKr） encoded by the HERG gene using patch clamp whole cell recording techniques. Methods Wild-type HERG eDNA plasmids were transfected into human embryonic kidney （HEK293） cells by lipofectamine method. The whole cell patch method was used to record the effects of GFG on the HERG potassium channel and kinesics of channel gating. Results GFG produced a concentration-dependent and voltage-dependent blockage of the Itail with IC50（ 50% inhibitory concentration） of 17.9 μmoL/L, and the Hill coefficient of 0.75. GFG degraded peak current potential of HERG and Itail without altered activation threshold potential. GFG blocked the HERG channel in a time-dependent manner. GFG shifted the activation curve in a negative direction and slowed channel deactivation. Moreover, GFG shifted the inactivation curve in a negative direction, accelerated channel recovery from inactivation and accelerated channel inactivation. Conclusion GFG blocked HERG channel currents and the blockade is dependence on open and inactivated channel states.