The effect of pancuronium on alamethicin-induced currents was studied in negatively charged lipid bilayer membranes. Pancuronium induces inactivation of the alamethicin-induced current. Inactivation is only observed if this compound is added to the compartment containing alamethicin. Moreover, the process of inactivation is reduced or abolished if pancuronium is added to the alamethicin-free side of the membrane. The time needed to recover from inactivation is greatly reduced if the aqueous solution in the alamethicin-free compartment is stirred. These data suggest that pancuronium permeates through the membrane when the alamethicin-induced conductance is "turned on," binds to the other membrane surface, and changes the surface potential.