Mechanism of inhibition of delayed rectifier K+ current by 4-aminopyridine in rabbit coronary myocytes.

1. The mechanisms involved in the 4-aminopyridine (4-AP)-induced block of delayed rectifier K+ current (IK(V)) in vascular smooth muscle cells were studied in cells enzymatically isolated from the rabbit coronary artery. 2. 4-AP inhibited slowly inactivating IK(V) in a dose-dependent manner (concentration producing half-maximal inhibition, K1/2, = 1.37 mM), and shifted the steady-state activation and inactivation curves of IK(V) by +9 and +16 mV, respectively. 3. The time constant of activation was significantly increased by 4-AP at +20 mV; deactivation kinetics were unaffected upon repolarization to -40 mV. The fast (tau f approximately 1 s) and slow (tau s approximately 5 s) time constants of inactivation (0 and +20 mV), and the recovery kinetics (tau r approximately 6 s) at -60 mV were not significantly affected by 0.5 mM 4-AP. However, tau f disappeared in the presence of 2 mM 4-AP while tau s remained unaffected. 4. Use-dependent unblock of IK(V) was revealed at potentials > or = -10 mV from analyses of the voltage dependence of 4-AP-sensitive currents and the frequency-dependent changes ('reverse use dependence') of IK(V) during the application of repetitive steps (-60 to +20 mV for 250 ms at a rate of 0.25 Hz) in control conditions, in the presence of 0.5 mM 4-AP, and after washout of the drug. These results suggested that 4-AP preferentially binds to the channel in the closed state, and unbinding is promoted by transitions to the open state. 5. The channel was modelled as a simple three-state mathematical loop model incorporating single closed, open and inactivated states. The block by 4-AP was modelled as a state-dependent interaction with 4-AP primarily binding to the closed state. Computer simulations support the hypothesis that 4-AP-induced block of the delayed rectifier K+ (KV) channel in the closed state is relieved during membrane depolarization. 6. Closed state binding of 4-AP to the KV channel depolarizes vascular smooth muscle cells by shifting the activation curve of these channels to more positive potentials.