REMOVAL OF CA CURRENT INACTIVATION IN DIALYZED GUINEA-PIG ATRIAL CARDIOBALLS BY CA CHELATORS

Ca currents [ICa] flowing during voltage clamp depolarizations were studied in cultured guinea-pig atrial cardioballs by means of single low resistance patch clamp pipettes. The pipettes were filled with solutions containing Cs+ as major cation in order to block K+ currents and high concentrations of various Ca chelating agents (EGTA [ethylene glycol bis(.beta.-amino ethylether)N,N,N''N'',-tetraacetic acid], nitrilotriacetic acid [NTA] citrate, dipicolinic acid [DPA]) to prevent rises of the intracellular Ca-activity by Ca-entry. ICa of myocytes loaded with 20 mM of either EGTA or NTA display a biphasic time course of inactivation at membrane potentials between -25 and +45 mV. The fast phase is reduced with increasingly positive membrane potentials. In cells loaded with either citrate or DPA inactivation is negligible or absent for small depolarizations. In the range of membrane potentials where maximum current flows (0 to +10 mV) a monophasic slow time course of inactivation is observed. At more positive membrane potentials inactivation is slowed. The amount of inactivation under this condition is related to the current density of the cell. Conditions, which for a given membrane potential reduce the amplitude of ICa such as extracellular application of blocking ions (Co2+, Cd2+), a conditioning depolarization, or rundown of Ca-channels lead to a slowing or a complete removal of inactivation in cells dialyzed with citrate or DPA, respectively. Cells loaded with these Ca chelators did not show any symptom of voltage dependent inactivation of ICa. Under the conditions described action potentials were recorded in the current clamp mode. Upon dialysis with EGTA the typical triangular shaped atrial action potential develops a plateau of 500-800 ms in duration. With citrate-containing pipette solutions the action potential duration usually is several seconds. Evidently, inactivation of cardiac ICa can be considerably slowed or even removed. Inactivation of this current depends on Ca entry rather than membrane potential. The fast phase of inactivation observed with EGTA (NTA) possibly reflects the slow kinetics of the binding reaction of this type of Ca chelators.