RATE OF NA+ CA2+ EXCHANGE ACROSS THE PLASMA-MEMBRANE OF SYNAPTOSOMES MEASURED USING THE FLUORESCENCE OF CHLOROTETRACYCLINE - IMPLICATIONS TO CALCIUM HOMEOSTASIS IN SYNAPTIC TERMINALS/

It is shown that the fluorescence of chlorotetracycline (CTC) can be u sed to continuously monitor Ca2+ fluxes mediated by the Na+/Ca2+-excha nger of the plasma membrane of synaptosomes. The kinetics of Ca2+ upta ke can be followed from the kinetics of the increase of CTC fluorescen ce with external Ca2+ concentrations in the micromolar range. Since th e fluorescence of CTC is not sensitive to Ca2+ concentration below 20 mu M this avoids any significant contribution of Ca2+ flux through Ca2 + channels to CTC fluorescence. By replacing KCl by choline chloride i n the buffer to avoid plasma membrane depolarization it is shown that the amplitude of the CTC fluorescence change is dependent upon the Na-gradient preimposed across the plasma membrane, and the rate constant of the kinetic process is dependent upon the Ca2+ concentration, The rate constant of the Ca2+ influx measured with depolarized and non-dep olarized synaptic plasma membrane vesicles at 37 degrees C and pH 7.4 were 0.55 +/- 0.10 and 0.25 +/- 0.02 min(-1), respectively. The overal l rate of Na+/Ca2+ exchange calculated under conditions close to physi ological Na+ and Ca2+ gradients and membrane resting potential ranged from 15 to 25% of the activity of the plasma membrane Ca2+ pump under these experimental conditions. The results also point out that membran e depolarization increases approx. 2-fold the rate of Na+/Ca2+ exchang e in synaptic plasma membrane vesicles.