Multiple effects of econazole on calcium signaling: Depletion of thapsigargin-sensitive calcium store, activation of extracellular calcium influx, and inhibition of capacitative calcium entry

The effect of econazole on intracellular calcium levels ([Ca²⁺](i)) in Madin Darby canine kidney cells was investigated using fura-2 fluorimetry. Econazole increased [Ca²⁺](i) dose-dependently at 5-50 μM. The Ca²⁺ signal consisted of an initial rise, a gradual decay and a sustained plateau. Extracellular Ca²⁺ removal partially reduced the econazole response. Mn²⁺ quench of fura-2 fluorescence confirmed econazole-induced Ca²⁺ influx. The econazole-sensitive intracellular Ca²⁺ store overlaps with that sensitive to thapsigargin, an inhibitor of the endoplasmic reticulum Ca²⁺ pump, because 25 μM econazole depleted the thapsigargin-sensitive store, and conversely, thapsigargin abolished the econazole response. Econazole (25-50 μM) partially inhibited capacitative Ca²⁺ entry induced by cyclopiazonic acid, another endoplasmic reticulum Ca²⁺ pump inhibitor, measured by depleting internal Ca²⁺ store in Ca²⁺-free medium followed by adding 10 mM CaCl₂. Econazole induced capacitative Ca²⁺ entry itself. Pretreatment with La³⁺ (100 μM) partially inhibited 25 μM econazole-induced Mn²⁺ quench of fura-2 fluorescence, and La³⁺ immediately reduced 20 μM econazole-induced Ca²⁺ signal when added at the peak of the signal, suggesting that econazole induced Ca²⁺ influx via two separate pathways: one is sensitive to La³⁺, the other is not. La³⁺ enlarged 25 μM econazole-induced [Ca²⁺](i) transient during the decay phase. The econazole response was not altered when the cytosolic level of inositol 1,4,5-trisphosphate was inhibited by the phospholipase C inhibitor U73122. Copyright (C) 1999 Elsevier Science B.V.