hERG1鉀離子通道門閥受季銨鹽衍生物調控之機制

Abstract

心臟跳動的節率被許多離子通道所調節，其中 human ether-à-go-go-related gene-1 (hERG1) 鉀離子通道扮演著重要的角色。hERG1通道回返電流的發生可促使心肌動作電位進行到再極化的階段，使心肌電位回到靜止膜電位，進而行使心臟正常的生理功能。不過，當hERG1通道發生點突變時，便會造成Long QT syndrome。Long QT syndrome 是很嚴重的疾病，一旦發生時很容易導致人體產生心律不整、甚至是死亡的危險。過去的研究指出，有些藥物可以抑制hERG1通道之電流，並因而有機會引發Long QT syndrome。最近的研究也報導，在hERG1通道外部添加常見的鉀離子通道抑制劑季銨鹽衍生物 (quaternary ammonium derivatives, QAs)，會依據QAs不同的疏水性，而對hERG1通道電流的抑制效果及其inactivation有不同程度的影響。不過，過去還沒有研究藉由內部添加QAs，來探討hERG1通道內部孔洞的性質。因此，我們此篇論文的研究目的便是藉由分別添加QAs在hERG1通道的外部和內部，來探討hERG1通道外部孔洞和內部孔洞性質上的差異，並嘗試著解釋QAs如何抑制hERG1通道和影響其gating。我們發現內部添加QAs對於hERG1通道回返電流的抑制效果，比從外部添加快很多，而且抑制作用與復極化電位及其前置之去極化電位皆有相關。我們也發現，不同大小的QAs對於hERG1通道的作用有顯著的不同，各QA中以tetrahexylammonium (THxA) 最強，較大或較小之QA作用皆減弱。若改由細胞外投給QA，亦可見對hERG1 channel有抑制作用，其中亦以THxA最強，不過其親和力低於細胞內投與者的十倍以上。我們同時也發現，細胞內之多胺離子 (例如spermine) 對於hERG1 通道電流亦有顯著之抑制作用。這些發現顯示hERG1通道之內口區與外口區，在其開關以致產生回返電流之過程中，應該都有重要的型態變化。

The human ether-à-go-go-related gene-1 (hERG1) potassium channel plays an important role in the regulation of cardiac rhythm. The resurgent currents through hERG1 channel contributes significantly to the repolarization phase of the cardiac action potential. Mutations or block of hERG1 channel may cause Long QT syndrome, a severe form of cardiac arrhythmia with a high incidence of sudden death. It has been shown that externally applied quaternary ammonium derivatives (QAs), a group of well-known potassium channel blockers, inhibit hERG1 channel current and affect inactivation of hERG1 channel according to their hydrophobicity. However, the effect of internally applied QAs has not been explored despite that some key gating conformational changes may involve the inner part of the pore. We first applied QAs on outside of hERG1 channel to confirm the previous findings on external QA block of hERG1 channel. We then perfused QAs to the internal side of the membrane with the inside-out patch configuration. We found that the blocking effect of internally applied QAs on the hERG1 channel happens much faster than external QAs. Moreover, the blocking effect of internal QAs is closely dependent on the voltage of the repolarization as well as the preceding depolarization phase. There are different inhibitory effects of QAs of different size. Tetrahexylammonium (THxA) has the strongest effect and the other QAs of the smaller or larger size all show weaken effect on hERG1 currents. For the externally applied QAs, THxA also has the sharpest effect. However, the blocking potency is in generally 10-fold smaller than internal QA. In the meanwhile, we demonstrate that internal polyamines (e.g. spermine) also have an inhibitory effect on the resurgent hERG1 currents. We conclude that there must be significant gating conformational changes associated with the genesis of the resurgent hERG1 currents in both the internal and external pore mouths of the hERG1 channel.