Intracellular calcium content of human erythrocytes: relation to sodium transport systems

To study the possible role of intracellular Ca (Cai) in controlling the activities of the Na+-K+ pump, the Na+-K+ cotransport and the Na+/Li+ exchange system of human erythrocytes, a method was developed to measure the amount of Ca embodied within the red cell. For complete removal of Ca associated with the outer aspect of the membrane, it proved to be essential to wash the cells in buffers containing less than 20 nM Ca. Ca was extracted by HClO4 in Teflon vessels boiled in acid to avoid Ca contaminations and quantitated by flameless atomic absorption. Cai of fresh human erythrocytes of apparently healthy donors ranged between 0.9 and 2.8 mumol/liter cells. The mean value found in females was significantly higher than in males. The interindividual different Ca contents remained constant over periods of more than one year. Sixty to 90% of Cai could be removed by incubation of the cells with A23187 and EGTA. The activities of the Na+-K+ pump, of Na+-K+ cotransport and Na+/Li+ exchange and the mean cellular hemoglobin content fell with rising Cai; the red cell Na+ and K+ contents rose with Cai. Ca depletion by A23187 plus EGTA as well as chelation of intracellular Ca2+ by quin-2 did not significantly enhance the transport rates. It is concluded that the large scatter of the values of Cai of normal human erythrocytes reported in the literature mainly results from a widely differing removal of Ca associated with the outer aspect of the membrane.