An envelope of tails test was used to show that the delayed rectifier K § current (Iv.) of guinea pig ventricular myocytes results from the activation of two outward K § currents. One current was specifically blocked by the benzenesulfonamide antiarrhythmic agent, E-4031 (IC50 = 397 nM). The drug-sensitive current, "I~" exhibits prominent rectification and activates very rapidly relative to the slowly activating drug-insensitive current, "Ivs." Iv, was characterized by a delayed onset of activation that occurs over a voltage range typical of the classically described cardiac I K. Fully activated Ivs, measured as tail current after 7.5-s test pulses, was 11.4 times larger than the fully activated I~. Ig r was also blocked by d-sotalol (100 #M), a less potent benzenesulfonamide Class III antiarrhythmic agent. The activation curve of lv~ had a steep slope (+7.5 mV) and a negative half-point (-21.5 mV) relative to the activation curve of Ivs (slope = + 12.7 mV, half-point = + 15.7 mV). The reversal potential (Erev) of I~ (-93 mV) was similar to EK (--94 mV for [K+]o ----4 mM), whereas Er~ of Ivs was -77 mV. The time constants for activation and deactivation of Iv~ made up a bell-shaped function of membrane potential, peaking between -30 and -40 mV (170 ms). The slope conductance of the linear portion of the fully activated Ig,-V relation was 22.5 S/F. Inward rectification of this relation occurred at potentials > -50 mV, resulting in a voltage-dependent decrease in peak Iv~ at test potentials > 0 InV. Peak Iw at 0 mV averaged 0.8 pA/pF (n -----21). Although the magnitude ofl~ was small relative to fully activated lw, the two currents were of similar magnitude when measured during a relatively short pulse protocol (225 ms) at membrane potentials (-20 to + 20 mV) typical of the plateau phase of cardiac action potentials.
Class III antiarrhythmic agents act by selective prolongation of cardiac action potential duration (APD). Methanesulfonanilide class III agents (e.g., E-4031 and dofetilide) are extremely potent and lengthen action potentials in a "reverse" rate-dependent manner; i.e., effects are greater at low compared with high rates of stimulation. By using the whole-cell current-clamp technique in isolated guinea pig ventricular myocytes, APD was shortened by rapid pacing (244 +/- 16 msec at 30 pulses per minute, 166 +/- 8 msec at 240 pulses per minute; n = 8). Dofetilide (1 microM) prolonged APD more when cells were stimulated at the rate of 30 pulses per minute (44 +/- 10-msec increase) than at 240 pulses per minute (21 +/- 5-msec increase). We investigated the mechanism of APD prolongation using voltage-clamp techniques. Dofetilide selectively inhibited IKr (IC50, 31.5 nM), defined as the rapidly activating inward rectifying component of net delayed rectifier K+ current (IK), without effects on the larger but more slowly activating component of IK (IKs) or on the inward rectifier K+ current (IK1). To examine the rate-dependent effects of dofetilide on APD, trains of conditioning pulses to 0 mV (200-msec duration) were applied at either 30 or 240 pulses per minute to mimic the action potential experiments. Test pulses or ramps were given after the conditioning train to quantitate changes in IK1, IKr, or IKs. The magnitude of neither IK1 nor IKr was dependent on the rate of the preceding train of depolarizations. Sensitivity to block of IKr by dofetilide was rate independent.(ABSTRACT TRUNCATED AT 250 WORDS)
The mechanism by which isoproterenol (ISO) prevents the prolongation of action potential duration (APD) and refractory period (RP) by the class III antiarrhythmic agent E-4031 was studied. E-4031 (1 microM) increased RP by 50% with no effect on contractile force in papillary muscles isolated from guinea pig heart. ISO (1 microM) increased force of contraction more than fivefold and decreased RP by 25%. The prolongation of RP by E-4031 was prevented by pretreatment of muscles with ISO. The prolongation of APD in isolated guinea pig ventricular myocytes by 5 microM E-4031 also was antagonized by prior exposure of the cells to 1 microM ISO. Instantaneous currents and delayed rectifier K+ currents, IK, were measured in isolated myocytes using the suction microelectrode voltage-clamp technique. Currents were measured in response to 225-msec depolarizing pulses from a holding potential of -40 mV. Previous studies have demonstrated that IK in these cells results from activation of two distinct outward K+ currents, IKs and IKr (specifically blocked by E-4031). ISO doubled the magnitude of IKs without significant effect on IKr. The instantaneous current, putatively identified as a Cl- current, also was doubled by ISO but was unaffected by E-4031. The augmented conductance of IKs and instantaneous current by ISO results in a decrease in RP. The small effect of E-4031 on APD and RP in the presence of ISO results from the smaller contribution of IKr relative to the augmented repolarizing currents.
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