Abstract
Membrane currents through the Ca2+ channel were studied in a hybridoma cell line (MAb‐7B) constructed by fusion of S194 myeloma cells and splenic B lymphocytes from the mouse. The whole‐cell variation of the patch‐electrode voltage‐clamp technique was used. When [Ca2+]o = 2.5 mM, [Na+]o = 150 mM and [Na+]i = 155 mM, the current reversed from inward to outward at 20.9 +/‐ 2.4 mV (mean +/‐ S.D., n = 62). Both inward and outward currents showed voltage‐dependent inactivation with the same membrane potential dependence of steady‐state inactivation. The decay time constant of the current decreased from about 27 ms at ‐44 mV to a saturation value of 16 ms at about ‐20 mV, and remained at this value even when the current became outward. From the above results both the inward and outward currents were considered to flow through Ca2+ channels. The inward current showed no change when the external Na+ was replaced with Cs+ or tetraethylammonium and increased when [Ca2+]o was increased. Also, the reversal potential became more positive with increasing [Ca2+]o with a slope of 29 mV/decade change of [Ca2+]o. Effects of different divalent cations examined at 10 mM concentration showed the reversal potential to become more positive in the order of Mn2+, Sr2+ approximately equal to Ba2+ and Ca2+ whereas the relative maximum amplitudes of peak inward current were 1.0 for Ca2+, 1.24 for Sr2+, 0.99 for Ba2+ and 0.07 for Mn2+. When [Ca2+]o or [Mg2+]o was reduced by chelators, monovalent cations became capable of carrying inward current through the Ca2+ channel. These monovalent currents share common kinetic properties with the Ca2+ current, as judged from the steady‐state inactivation and the decay time constant of the current. The monovalent cation current was blocked by divalent cations in a voltage‐dependent manner. The half‐blocking concentrations of Ca2+ and Mg2+ at ‐45 mV were 2.0 X 10(‐6) M and 3.0 X 10(‐5) M respectively. The same voltage‐dependent binding mechanism can explain the outward current carried by monovalent cations at large positive potentials at normal Ca2+ concentrations. The suppression of the monovalent currents by Ca2+ and Mg2+ showed different voltage dependences. The suppression by Ca2+ increased and then decreased as the membrane potential was made negative, whereas the suppression by Mg2+ increased monotonically. This difference can be explained by considering the fact the Ca2+ is permeant and Mg2+ is impermeant through the Ca2+ channel.