硫化氢对大鼠结肠收缩活动的影响及其离子通道机制

Effects of hydrogen sulfide on colonic contraction of rats and ion channel mechanisms

目的 探讨硫化氢（H2S）对大鼠结肠收缩活动的影响及其离子通道机制.方法 取30只雄性Wistar大鼠远端结肠制备纵行肌条（LMS）及环形肌条（CMS）；张力换能器及多通道生理信号采集处理系统记录H2S供体硫氢化钠（NaHS）对LMS和CMS自发性收缩活动的影响及河豚毒素（TTX）孵育后NaHS对LMS和CMS收缩活动的影响；采用酶消化法分离近端结肠平滑肌细胞,运用全细胞膜片钳技术记录NaHS对平滑肌细胞L型钙通道电流（ICa.L）和大电导钙激活钾电流（IBKCa）的影响.结果 NaHS[（1～ 12）× 10-5mol/L]浓度依赖性促进结肠LMS和CMS的收缩活动,其中（2 ～ 12）×10-5mol/L NaHS使LMS和CMS收缩显著增加（均P＜0.05）,且该促进作用不被TTX阻断；NaHS（6 ×10-5和12×10-5 mol/L）浓度依赖性促进ICa.L,使0 mV处ICa,L由（-3.16 ±0.47）pA/pF分别增加至（-3.33 ±0.54）、（-3.65±0.66）pA/pF（均P＜0.05）；NaHS（12×10-5 mol/L）作用前后,最大峰电流均在0 mV处,不改变L型钙通道的电压依赖特性,亦不影响L型钙通道的激活态；NaHS（12×10-5 mol/L）对L型钙通道的促进作用具有时间依赖性、可逆性；NaHS（6×10-5和12×10-5 mol/L）浓度依赖性抑制IBKCa,使60 mV处IBKCa由（16.68±1.23） pA/pF分别降低至（15.26±2.67）、（13.80±3.04）pA/pF（均P＜0.05）.结论 低浓度H2S可能通过开放平滑肌细胞L型钙通道,抑制大电导钙激活钾通道,进而促进大鼠结肠平滑肌收缩.

Objective To explore the effects of hydrogen sulfide （H2S） on colonic contraction of rats at low concentration and ion channel mechanisms.Methods Organ bath recordings were used to examine the contraction of colonic smooth muscle strips.The whole-cell patch-clamp technique was used to record the currents of L-type calcium and large conductance Ca2＋-activated K＋ （BKCa） channels in smooth muscle cells isolated from 30 male Wistar rats.Results The H2S donor NaHS （（1-12） × 10-5mol/L） increased the spontaneous contractions of longitudinal and circular muscle strips in a dose-dependent manner （all P 〈0.05） and the effect could not be blocked by tetrodotoxin.NaHS （6 × 10-5 and 12 × 10-5 mol/L） reversibly increased the L-type calcium current （ICa,L） in a dose-dependent manner.And the peak of ICa,L at 0 mV increased from （-3.16 ± 0.47） to （-3.33 ± 0.54） and （-3.65 ± 0.66） pA/pF respectively （n =6,both P 〈 0.05）.Current-voltage （Ⅰ-Ⅴ） curve had no shift after NaHS treatment and H2S donor caused no change in the curves of steady-state activation.Likewise BKCa channel was significantly inhibited by NaHS （6 × 10-5 and 12 × 10-5 mol/L） in a dose-dependent manner.And IBKCa at 60 mV decreased from （16.68 ± 1.23） to （15.26 ± 2.67） and （13.80 ± 3.04） pA/pF respectively （both P 〈 0.05）.Conclusions H2S at low concentrations increases the spontaneous contraction of rat colonic smooth muscle in a dose-dependent manner.Such an effect may be due to a direct activation of L-type calcium channel and an inhibition of BKCa channel in smooth muscle cells.