Control (AA) and
sickle cell anemia (SS) erythrocytes were loaded with Ca-
chelator (
Quin2 or
Benz2) to increase the cellular exchangeable Ca2+ pool and to measure the Ca2+ exchange fluxes and the cytosolic ionized Ca2+ ([Ca]i) (Lew et al., 1982, Nature, 298, 478). The
chelator incorporation induced a decrease in the
ATP content which was smaller in SS than in AA cells and partially reversible upon reincubation in a
chelator-free medium. The amount of trapped
chelator was determined by two methods: 45Ca binding to the
chelator in Ca-
ionophore treated cells in Ca-
EGTA buffers and [3H]
Quin2 incorporation. A slight over-estimation of the
chelator content was found with the second method but incorporation was the same in both types of cells. The kinetics of 45Ca equilibration and 45Ca release were used to measure Ca2+ fluxes and [Ca]i in oxygenated
chelator-loaded cells. SS cells, as compared to AA cells, exhibited a moderate increase in Ca2+ fluxes (30-75%) but [Ca]i remained in the same range (about 20 nM). Thus the excess of Ca2+ found in SS cells is not available for the Ca2+ pump or the K+ channel a conclusion in agreement with that of Bookchin et al. (1984, Cell
Calcium, 5, 277). Analysis of the 45Ca kinetics showed that in AA cells, exchangeable Ca2+ behaved as one compartment. In SS cells, the existence of a second slowly-exchangeable Ca2+ compartment was demonstrated. This latter (3-5 mumol/l cells) was independent of the concentration of the
chelator and thus could represent exchangeable Ca2+ enclosed within the intracellular inside-out vesicles recently observed in SS cells (Williamson et al., 1984, J. Cell. Biol., 99, 430a). Alternatively, these two kinetic pools could reflect heterogeneity of the SS cell population.