Cytosolic free Ca²⁺ concentration and intracellular calcium distribution of Ca²⁺ -tolerant isolated heart cells

The cytosolic free Ca²⁺ concentration of calcium-tolerant rat myocytes has been measured by the null point titration technique using arsenazo III as a Ca²⁺ indicator and digitonin to permeabilize the plasma membrane. The mean value obtained for 8 separate preparations was 270 ± 35 nM. The distribution of releasable calcium between the mitochondrial and sarcoplasmic reticular compartments was measured by the successive additions of uncoupler and A23187 to cells pretreated with ruthenium red. The relative distribution of calcium in each pool was independent of the cell calcium content up to the maximum value of releasable calcium investigated (4.5 nmol/mg of cell dry weight) and was distributed in the approximate ratio of 2:1 in favor of the sarcoplasmic reticulum. The cells contained 1 nmol of calcium/mg of cell dry weight in a form nonreleasable by A23187, which was independent of the total cell calcium content as measured by atomic absorption spectroscopy. It is calculated that the calcium content of mitochondria in heart under physiological conditions is about 5 nmol/mg of mitochondrial protein. At this level, the mitochondria are likely to provide effective buffering of the cytosolic free Ca²⁺ concentration of quiescent heart cells. The corresponding intramitochondrial free Ca²⁺ is in a range above values needed to regulate the activity of Ca²⁺ -dependent enzymes of the citric acid cycle in heart. The physiological calcium content of the sarcoplasmic reticulum in heart cells is estimated to be about 2.5 nmol/mg of cell dry weight, which is at least 5-fold greater than the amount of calcium release calculated to cause maximum tension development of cardiac muscle.