Dibucaine displaceable sarcolemmal Ca2+ fraction in guinea-pig cardiac myocytes

In previous work we found that Ca2+ bound to the internal leaflet of sarcolemma of the neonatal cultured rat cardiomyocytes may be displaced by a local anesthetic, dibucaine (DBC). This resulted in inhibition of Na/Ca exchange. Now we found that the DBC displaceable Ca2+ fraction may be demonstrated also in single, enzymatically isolated cardiomyocytes of adult guinea pigs. Only cell length was used as an index of the free sarcoplasmic Ca2+ concentration since we found that DBC is fluorescent when irradiated with UV light, the wave length overlapping that of Indo 1 fluorescence. DBC (0.5 mM) induced a contracture whose amplitude ranged from 30%-100% of that of the electrically stimulated twitch. Blocking of Na/Ca exchange with 0.5 mM Ni2+, or 0Na+, 0Ca2+ solution did not affect the amplitude nor the time course of the contracture. Caffeine (15 mM) superfused during exposure of the cell to DBC initiated contraction which relaxed despite continued caffeine superfusion. DBC also initiated contracture in cells pretreated with thapsigargin, a blocker of sarcoplasmic reticulum Ca2+ uptake. DBC did not initiate contracture in skinned myocytes and decreased their sensitivity to Ca2+. These results suggest that DBC displaces Ca2+ from sarcolemma and that the displacement results in inhibition of Na/Ca exchange. Inhibition may however, be overcome by high subsarcolemmal Ca2+ concentration induced by its SR release with caffeine. DBC rapidly blocked sodium and calcium currents in voltage clamped cells. Calcium current recovered within about a minute upon washout of DBC, whereas recovery of sodium current started after about 8 min and was completed within about 12 min. Rapid depolarisation from a holding potential of -80 mV to 0 or +20 mV potentiated the response of the cell to DBC. This suggests that the Ca2+ fraction which is displaced by DBC is distinct from that released by depolarisation and therefore does not contribute to activation of a normal, electrically stimulated contraction.