Negative shift of cardiac Na+ channel kinetics in cell-attached patch recordings

Na+ channel kinetics were studied by recording single-channel currents in the cell-attached patch configuration of the patch-clamp technique in single ventricular cells isolated from guinea pig hearts. The inactivation time course of ensemble currents was accelerated, and the peak amplitude increased temporarily and then decreased within a few minutes after the gigaohm seal formation. After reaching a new steady state, the inactivation-voltage relation was found to have shifted to more negative potentials. The potential of half-maximal inactivation was more negative by 20-31 mV from the resting potential or between -96 and -112 mV. The voltage dependency of the channel activation also shifted. Although the cell membrane was depolarized using the whole cell patch-clamp electrode and single-channel currents were recorded with an independent cell-attached electrode, the shift of the inactivation curve was also evident. Complete removal of Ca2+ using 5 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid in the pipette solution failed to prevent the shift. Increasing Ca2+ to 10 mM, however, reduced magnitude of the shift significantly. Involvement of an increased membrane fluidity and surface potential of the glass pipette to the shift is discussed.