Adult ventricular myocytes lose T-tubules over few days in culture, which causes the loss of about 60% of the cell membrane capacitance (Cm) (Mitcheson et al., 1996). In this study, we have measured, in whole-cell voltage-clamped rabbit right ventricular myocytes at 0, 1, 2 and 3-5 days of culture (nine to 20 myocytes at each age) in a defined Dulbecco's modified Eagle's medium, the value of Cm and the magnitudes of the background inward rectifier current (IK(1)) and of the 2,4-dinitrophenol-induced ATP-sensitive potassium current (IK(ATP)). Cm, IK(1) and IK(ATP) all had decreased significantly by 51, 83 and 88%, respectively after 4 days of culture. Analysis using a single exponential decay function of time gave time constants of, 2.6+/-0.2, 2.2+/-0.5 and 2.4+/-0.4 days, respectively. Linear regressions of IK(1) and IK(ATP) versus Cm had regression coefficients of 0.93 and 0. 98, respectively. These observations are consistent with a strong link of the decay of IK(1) and IK(ATP) currents to that of Cm. Furthermore, the time course of changes in IK(1) when an external blocker (100 microm BaCl(2)) was applied and washed by local perfusion (95% change in 50 ms) agrees with a model including a diffusion time constant of 300 ms. This value is consistent with the known kinetics of diffusion of divalent cations in the T-tubules. Taken together, these results could be explained by the localization of a major part of the IK(1) and IK(ATP) currents of ventricular cardiomyocytes in the T-tubules. As a consequence, transient accumulation of K(+) ions in cardiac T-tubules may take place and modulate excitation-contraction coupling.