Imidazole‐induced elevations of intracellular calcium in HL‐60 cells: effect of inhibition of phospholipase C by the steroidal maleimide U73122

Abstract A variety of agents, including imidazoles, such as miconazole, elicit elevations of calcium in human leukemic HL‐60 cells through a stimulation of influx of calcium and/or a release of intracellular calcium. Release of intracellular calcium by such agents may involve stimulation of IP 3 formation. The steroidal maleimide U73122, a potent phospholipase C (PLC) inhibitor, at 3–10 µM blocked elevations of calcium elicited in HL‐60 cells by ATP. At a 10&!hyphen; µM concentration, U73122 either blocked or altered elevations of calcium elicited by imidazoles, such as miconazole, SKF 96365, and clotrimazole that are often used to block capacitative calcium entry, and by calmidazolium, a quaternary imidazole that can trigger massive influx of calcium in HL‐60 cells. Elevations of calcium elicited by thapsigargin were not affected by 3 µM U73122, but were reduced by higher concentrations. Remarkably, U73122 at 10 µM did not block, but instead slowed and potentiated the elevation of calcium elicited by higher concentrations of imidazoles, an effect that may involve inhibition of Ca 2+ ‐ATPases of the endoplasmic reticulum. The steroidal succinimide U73343 had minimal effects on calcium elevations in HL‐60 cells. The results indicate that U73122 requires cautious use in assessing the role of PLC in controlling calcium levels in HL‐60 cells, in particular when used with imidazoles, such as SKF 96365, which are used to block capacitative calcium entry. The effects of U73122 on cell calcium are reviewed. In toto, the results suggest that many lipophilic agents may affect intracellular calcium by activating PLC through unknown mechanisms. Drug Dev. Res. 67:519–534, 2006. Published 2006 Wiley‐Liss, Inc.