BLOCK OF THE INACTIVATING POTASSIUM CHANNEL BY CLOFILIUM AND HYDROXYLAMINE DEPENDS ON THE SEQUENCE OF THE PORE REGION

Cardiac antiarrhythmic compounds are a diverse group divided into clas ses that differ in their mechanisms of action. Recent attention has fo cused on class III compounds, which prolong the action potential by bl ocking K+ channels. The purpose of this study was to characterize the mechanisms of actions of a class iii compound, clofilium, and a simple analog, hydroxylamine, on an inactivating K+ channel. The defined sys tem used a cloned inactivating K+ channel (Shaker-B) expressed in Xeno pus oocytes. This channel is similar in physiological properties and c ore sequence to the inactivating K+ channel cloned from mammalian hear t. Results presented here demonstrate that clofilium (100 mu M) and hy droxylamine (10 mM) can cause use-dependent block, depending on the se quence of the pore region. A mutation of the pore known to influence s electivity and tetraethylammonium binding (threonine-441 to serine) co nfers use-dependent sensitivity to hydroxylamine and clofilium. Hybrid channels were formed from the coinjection of wild-type and mutant cha nnel mRNAs; the analysis of block with the hybrid channels suggests th at binding of hydroxy[amine involves all subunits of the tetrameric ch annel, whereas clofilium affects channels containing as few as one mut ant subunit. The simplest interpretation is that all four subunits con tribute to an internal binding site for blockers such as clofilium and hydroxylamine and threonine-441 influences this binding site. The eff ectiveness of clofilium, unlike hydroxylamine, on the hybrid channels may reflect its structural complexity, which could allow interaction w ith a broader receptor site. Future studies will test this idea using other class III-related compounds.