Sarcolemmal Ca influx through L-type Ca channels in ventricular myocytes of a teleost fish

Abstract

The whole cell patch clamp method was used to measure Ca current through L-type Ca channels in enzymatically isolated ventricular myocytes of crucian carp (Carassius carassius L.) heart. Fish were acclimated to 22 degrees C for more than 4 wk, and properties of Ca current were measured at room temperature (21 +/- 1 degrees C). Depolarizing voltage steps from -50 mV evoked rapidly activating Ca currents, which exhibited a bell-shaped voltage dependence with peak amplitude at 0 mV. The currents were suppressed by nifedipine (5 microM), verapamil (2.5 microM), and Cd2+ (175 microM). The current amplitude was increased by 67.5 +/- 17.2% (n = 5) in the presence of 1 microM isoproterenol. Steady-state inactivation and activation curves showed half-maximal inactivation at -31.3 +/- 0.95 mV, with a slope factor of 5.88 +/- 0.51, and half-maximal activation at -10.6 +/- 1.65 mV, with a slope factor of 7.84 +/- 0.54 (n = 9). The overlap of inactivation and activation curves suggests the presence of a small window current, which is maximally 4% of the peak current at -27 mV. The density of L-type Ca current was 6.95 +/- 0.79 pA/pF at 0 mV (n = 35). A total increment in cellular Ca contributed by L-type Ca current during a 500-ms voltage clamp pulse was calculated from the integral of Ca current and cell volume. The charge transfer through L-type Ca current was 0.325 +/- 0.023 pC/pF, and the mean cell volume was 1,377 +/- 44 microns3. The increment in total cellular Ca by Ca influx through L-type Ca channels was calculated to be 39.3 +/- 2.8 microM. These findings imply that Ca influx through L-type Ca channels can contribute significantly to the activation of contraction in the ventricular myocytes of fish heart.