Voltage-dependent Ba2+ block of K+ channels in apical membrane of frog skin

Abstract

The characteristics of the blockage of apical K+ channels in the frog skin (Rana temporaria) by Ba2+ were investigated with current-voltage measurements and current-fluctuation analysis. Inward K+ currents were recorded with a transepithelial K+ concentration gradient oriented from mucosa to serosa. When Ba2+ (5-10,000 microM) was added to the mucosal solution, the inward K+ current was progressively depressed. This blockage was strongly voltage dependent. The apparent macroscopic Michaelis-Menten constant decreased exponentially with increasing transepithelial voltage (Vt) (mucosa positive). According to Eyring rate theory, the binding site for the Ba2+ ion in the K+ channel was found to be located at a relative electrical distance of 0.72 of the apical membrane potential field, measured from the cytosolic side. From current-voltage (I-V) measurements and Michaelis-Menten kinetics, the Ba2+ dissociation constant was calculated: KBa = 57 microM (Vt = 0 mV). The voltage dependence of the on- and off-rate of the Ba2+-receptor interaction was obtained from the analysis of current fluctuations induced by Ba2+. An exponential relationship between the rates and Vt was obtained, as was found for KBa from the I-V measurements. KBa (at Vt = 0 mV) calculated from the noise experiments was 66 microM.