Mechanism of block and identification of the verapamil binding domain to HERG potassium channels

Calcium channel antagonists have diverse effects on cardiac electrophysiolo gy. We studied the effects of verapamil, diltiazem, and nifedipine on HERG K+ channels that encode I-Kr in native heart cells. In our experiments, ver apamil caused high-affinity block of HERO current (IC50 = 143.0 nmol/L), a value close to those reported for verapamil block of L-type Ca2+ channels, whereas diltiazem weakly blocked HERG current (IC50=17.3 mu mol/L), and nif edipine did not block HERG current. Verapamil block of HERG channels was us e and frequency dependent, and verapamil unbound from HERG channels at volt ages near the normal cardiac cell resting potential or with drug washout. B lock of HERG current by verapamil was reduced by lowering pH(o), which decr eases the proportion of drug in the membrane-permeable neutral form. N-meth yl-verapamil, a membrane-impermeable permanently charged verapamil analogue , blocked HERG channels only when applied intracellularly. Verapamil antago nized dofetilide block of HERG channels, which suggests that they may share a common binding site. The C-type inactivation-deficient mutations, Ser620 Thr and Ser631Ala, reduced verapamil block, which is consistent with a role for C-type inactivation in high-affinity drug block, although the Ser620Th r mutation decreased verapamil block 20-fold more than the Ser631Ala mutati on. Our findings suggest that verapamil enters the cell membrane in the neu tral form to act at a site within the pore accessible from the intracellula r side of the cell membrane, possibly involving the serine at position 620. Thus, verapamil shares high-affinity HERG channel blocking properties with other class III antiarrhythmic drugs, and this may contribute to its antia rrhythmic mechanism.