Tracheal smooth muscle cells were enzymatically isolated from guinea-pig trachea. These cells contracted in response to acetylcholine (0.01-10 microM) in a concentration-dependent fashion. Under current-clamp conditions with 140 mM K+ in the pipette solution, the membrane potential oscillated spontaneously at around -30 mV. Under voltage-clamp conditions, there appeared spontaneous but steady oscillations of outward current (IO). On depolarization from a holding potential at -40 mV, three components of outward current were elicited: transient outward current (IT), steady-state outward current (IS) and IO. These three components of outward current reversed around the K+ equilibrium potential and were abolished by Cs+ in the pipette, indicating that K+ was the major charge carrier of these outward currents. All these three components were completely suppressed by extracellular tetraethylammonium (10 mM). Both IT and IO were depressed by quinidine (1 mM), 4-aminopyridine (10 mM) and nifedipine (100 nM), but IS was not affected. IT and IO were suppressed by a Ca2(+)-free perfusate with less than 1 nM Ca2+ in the pipette, while with 10 nM Ca2+ in the pipette, only IO was suppressed. In both conditions, IS was not affected by the Ca2(+)-free perfusate. Therefore, it is suggested that IO, IT and IS are separate types of K+ current. With Cs+ in the pipette, K+ currents were almost completely suppressed and a transient inward current was observed during depolarizing pulses. The inward current was not affected by tetrodotoxin and increased when the concentration of extracellular Ca2+ was raised, indicating that the current is a Ca2+ channel current. Even with a holding potential of -80 mV, the low-threshold inward current could not be observed. The high-threshold Ca2+ current was abolished by nifedipine (100 nM) and was enhanced by Bay K 8644 (100 nM). The order of permeation of divalent cations through the Ca2+ channel was Ba2+ greater than Sr2+ approximately Ca2+. Cd2+ blocked the Ca2+ current more effectively than Ni2+. These results may indicate that the Ca2+ current of tracheal smooth muscle cells is mainly composed of the current through an L-type Ca2+ channel.