CONTROL OF RESTING MEMBRANE-POTENTIAL BY DELAYED RECTIFIER POTASSIUM CURRENTS IN FERRET AIRWAY SMOOTH-MUSCLE CELLS

1. In order to determine the physiological role of specific potassium currents in airway smooth muscle, potassium currents were measured in freshly dissociated ferret trachealis cells using the nystatin-permeabilized, whole-cell method, at 35-degrees-C. 2. The magnitude of the outward currents was markedly increased as bath temperature was increased from 22 to 35-degrees-C. This increase was primarily due to the increase in maximum potassium conductance (g(K,max)), although there was also a small leftward shift in the relationship between g(K) and voltage at higher temperatures. The maximum conductance and the kinetics of current activation and inactivation were also temperature dependent. At 35-degrees-C, gating of the current was steeply voltage dependent between -40 and 0 mV. Current activation was well fitted by fourth-order kinetics; the mean time constants of activation (30 mV clamp step) were 1.09+/-0.17 and 1.96+/-0.27 ms at 35 and 22-degrees-C, respectively. 3. Outward currents using the nystatin method were qualitatively similar to delayed rectifier currents recorded in dialysed cells with high calcium buffering capacity solutions. 4-Aminopyridine (4-AP; 2 mM), a specific blocker of delayed rectifier potassium channels in this tissue, inhibited over 80% of the outward current evoked by voltage-clamp steps to between -10 and +20 mV (n = 6). Less than 5% of the outward current was blocked over the same voltage range by charybdotoxin (100 nM; n = 15), a specific antagonist of large-conductance, calcium-activated potassium channels in this tissue. 4. The degree to which delayed rectifier and calcium-activated potassium conductances control resting membrane potential was examined in current-clamp experiments. The resting membrane potential of current clamped cells was -33.6+/-1.0 mV (n = 62). Application of 4-AP (2 mM) resulted in a 14.4+/-1.0 mV depolarization (n = 8) and an increase in input resistance. Charybdotoxin (100 nM) had no effect on resting membrane potential (n = 6). 5. Force measurements were made in isolated strips of trachealis muscle to determine the effect of pharmacological blockade of individual potassium conductances on resting tone. In the presence of tetrodotoxin (1 muM) and atropine (1 muM), 4-AP increased baseline tension in a dose-dependent manner, with an EC50 of 1.8 mM (n = 13); application of 5 mM 4-AP increased tone to 86.8+/-8.1% of that produced by 1 muM methacholine, and this tone was almost completely inhibited by nifedipine (1 muM). Conversely, antagonism of calcium-activated potassium (K(Ca)+) channels with charybdotoxin (100 nM) resulted in only a slight increase in resting tone (4.9+/-3.0% of methacholine response). 6. We conclude that delayed rectifier potassium channels play a critical role in maintaining resting membrane potential in ferret trachealis smooth muscle. Calcium-activated potassium channels appear to comprise a smaller component of the resting potassium conductance, and accordingly are less important in defining the passive behaviour of this muscle.