The dynamics of mitochondrial Ca fluxes

We have investigated the kinetics of mitochondrial Ca²⁺ influx and efflux and their dependence on cytosolic Ca²⁺] and Na⁺] using low-Ca²⁺-affinity aequorin. The rate of Ca²⁺ release from mitochondria increased linearly with mitochondrial Ca²⁺] (Ca²⁺]_M). Na⁺-dependent Ca²⁺ release was predominant al low Ca²⁺]_M but saturated at Ca²⁺]_M around 400 μM, while Na⁺-independent Ca²⁺ release was very slow at Ca²⁺]_M below 200 μM, and then increased at higher Ca²⁺]_M, perhaps through the opening of a new pathway. Half-maximal activation of Na⁺-dependent Ca²⁺ release occurred at 5-10 mM Na⁺], within the physiological range of cytosolic Na⁺]. Ca²⁺ entry rates were comparable in size to Ca²⁺ exit rates at cytosolic Ca²⁺] (Ca²⁺]_c) below 7 μM, but the rate of uptake was dramatically accelerated at higher Ca²⁺]_c. As a consequence, the presence of Na⁺] considerably reduced the rate of Ca²⁺]_M increase at Ca²⁺]_c below 7 μM, but its effect was hardly appreciable at 10 μM Ca²⁺]_c. Exit rates were more dependent on the temperature than uptake rates, thus making the Ca²⁺]_M transients to be much more prolonged at lower temperature. Our kinetic data suggest that mitochondria have little high affinity Ca²⁺ buffering, and comparison of our results with data on total mitochondrial Ca²⁺ fluxes indicate that the mitochondrial Ca²⁺ bound/Ca²⁺ free ratio is around 10- to 100-fold for most of the observed Ca²⁺]_M range and suggest that massive phosphate precipitation can only occur when Ca²⁺]_M reaches the millimolar range.