Calmidazolium leads to an increase in the cytosolic Ca²⁺ concentration in Dictyostelium discoideum by induction of Ca²⁺ release from intracellular stores and influx of extracellular Ca²⁺

The Ca²⁺ stores of Dictyostelium discoideum amoebae take part in control of homoeostasis of the cytosolic free Ca²⁺ concentration ([Ca²⁺]_i) and the cyclic-AMP-induced [Ca²⁺]_i-signalling cascade. In order to characterize regulatory mechanisms of these stores, we incubated cells with the calmodulin antagonist calmidazolium. Measurement of permeabilized and intact cells in suspension with a Ca²⁺-sensitive electrode revealed that calmidazolium induced Ca²⁺ release from intracellular stores, influx of Ca²⁺ across the plasma membrane and subsequent efflux. In single fura-2-loaded cells calmidazolium evoked rapid and global transient elevations of [Ca²⁺]_i. Other calmodulin antagonists (trifluoperazine, chlorpromazine, fendiline and W7) also induced transient elevations of [Ca²⁺]_i, which were, however, slower and observed in fewer cells. The calmidazolium-induced influx of extracellular Ca²⁺ was inhibited by preincubation with 2,5-di-(t-butyl)-1,4-hydroquinone (BHQ) and 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl), both known to interact with pumps of the inositol 1,4,5-trisphosphate (IP₃)-sensitive store, and by the V-type H⁺-ATPase inhibitor bafilomycin A₁, which affects the acidosomal Ca²⁺ store. Incubation with pump inhibitors did not itself induce changes in [Ca²⁺]_i. We conclude that the effects of calmidazolium are, at least in part, mediated by its calmodulin-antagonizing properties, that it acts by inducing Ca²⁺ release from filled storage compartments, and that its target of action is both the IP₃-sensitive store and the acidosome; emptying of these stores leads to influx of extracellular Ca²⁺.