Characterization of voltage-dependent calcium influx in human erythrocytes by fura-2

Thus far, the methods used to determine erythrocyte Ca²⁺ influx have not allowed the assessment of the kinetics of ion uptake. To overcome this drawback, we studied a new method, using the fluorescent Ca²⁺-chelator fura-2, which directly quantifies intracellular Ca²⁺ changes in human erythrocytes. This method has the advantage over previous techniques that it monitors continuously cellular Ca²⁺ levels. The Ca²⁺ influx is modulated by cellular membrane potential in the presence of a transmembrane Ca²⁺ concentration gradient and exhibits a first slow increase of the intracellular Ca²⁺ concentration, followed, after the reachment of a threshold value of 125 ± 13 nM Ca²⁺, by a faster increase until a plateau is reached. The influx rate is inhibited by dihydropyridines in the micromolar range. These findings support the hypothesis that erythrocyte Ca²⁺ influx is mediated by a carrier similar to the slow Ca²⁺ channels and is dependent on membrane depolarization.