Determination of Pure Voltage-Dependent Ca2+ Current In Paramecium Caudatum and Its Inhibition By Divalent Cations

Voltage-dependent Ca2+ currents in Paramecium caudatum were studied under voltage clamp conditions. To separate Caz+ inward currents from concomitant K+ outward currents, the voltage-dependent Ca2+ conductance was temporarily inactivated by a preceding depolarization. The remaining currents were then subtracted from the overall currents measured in the absence of a prepulse. In this way pure Ca2+ currents could be obtained up to a depolarization of 100 mV, which is about 50 mV below the theoretical Ca2+ equilibrium potential (ECa). Ca2+ currents were maximal at a depolarization of 35 mV and declined with further approach to ECa, but they did not reverse sign in the voltage range tested. In the presence of Mg2+, Co2+, Mr2+ or Ni2+, the Ca2+ inward currents decreased to a different extent. From experiments where these cations were added at different concentrations and from measurements at different Ca2+ concentrations in the absence of other divalent cations the following ratio of apparent dissociation constants could be derived: kNi: kCo: kCa: kMn: kMg = 1:3:4·3–4·7: 5:6·5. With a confidence of 95% the absolute value of kCa lies between 40 and 130μmoll−1. These results indicate that Ca2+ and other divalent cations compete for binding sites at the Ca-channel and thus determine excitability. Indirect effects due to changes of the surface potential are discussed.