Abstract
The role of intracellular Ca 2+ as essential activator of the Na +Ca 2+ exchange carrier was explored in membrane vesicles containing 67% right-side-out and 10% inside-out vesicles, isolated from squid optic nerves. Vesicles containing 100 μM free calcium exhibited a 2-fold increase in the initial rate of Na i +-dependent Ca 2+ uptake as compared with vesicles where intravesicular calcium was chelated by 2 mM EGTA or 10 mM HEDTA. The activatory effect exerted by intravesicular Ca 2+ on the reverse mode of Na +Ca 2+ exchange (i.e. Na i +Ca 0 2+ exchange) is saturated at about 100 μM Ca i 2+ and displays an apparent K 1 2 of 12 μM. Intravesicular Ca 2+ produced activation of Na i +Ca 0 2+ exchange activity rather than an increase in Ca 2+ uptake due to Ca 2+Ca 2+ exchange. The presence of Ca i 2+ was essential for the Na i +-dependent Na + influx, a partial reaction of the Na +Ca 2+ exchanger. In fact, the Na + influx levels in vesicles loaded with 2 mM EGTA were close to those expected from diffusional leak while in vesicles containing Ca i 2+ an additional Na +Na + exchange was measured. The results suggest that in nerve membrane vesicles Ca 2+ at the inner aspect of the membrane acts as an activator of the Na +Ca 2+ exchange system.
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