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 (I-K(V)) in vascular smooth muscle cells were studied in cells enzymatically isolated from the rabbit coronary artery. 2. 4-AP inhibited slowly inactivating I-K(V) in a dose-dependent manner (concentration producing half-maximal inhibition, K-1/2, = 1.37 mM), and shifted the steady-state activation and inactivation curves of I-K(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) approximate to 1 s) and slow (tau(s) approximate to 5 s) time constants of inactivation (0 and + 20 mV), and the recovery kinetics (tau(r) approximate to 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 I-K(V) was revealed at potentials greater than or equal to - 10 mV from analyses of the voltage dependence of 4-AP-sensitive currents and the frequency-dependent changes ('reverse use dependence') of I-K(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+ (K,) channel in the closed state is relieved during membrane depolarization. 6. Closed state binding of 4-AP to the K-V channel depolarizes vascular smooth muscle cells by shifting the activation curve of these channels to more positive potentials.