Staurosporine effects calcium homeostasis in cultured bovine adrenal chromaffin cells

Abstract

These studies show that the potent, non-specific, protein kinase inhibitor, staurosporine, disrupts Ca 2+ homeostasis in cultured bovine adrenal chromaffin cells. Staurosporine treatment reduces basal and A23187-stimulated catecholamine releases from chromaffin cells, but does not inhibit activated Ca 2+ influx. Furthermore, pretreatment with staurosporine also reduces Ca 2+-stimulated catecholamine release from digitonin-permeabilized cells ( t 1 2 , 40.6 min; IC 50, 66.0 nm). However, staurosporine does not inhibit the rise in intracellular Ca 2+ ([Ca 2+] i) in response to nicotine stimulation as measured by fura-2 photometry. These studies demonstrate that staurosporine interferes with the secretory process at some step at or after the rise in [Ca 2+] i in adrenal chromaffin cells. Examination of the effects of staurosporine on 45Ca 2+ movement shows that staurosporine produces a slowly developing basal 45Ca 2+ accumulation; after 30 min no significant change is observed, but by 120 min, 45Ca 2+ accumulation is increased by 29.5%. Thapsigargin and 2,5-di( tert-butyl)-1,4-benzohydroquinone (tBHQ), inhibitors of Ca 2+ ATPases, were used to determine whether staurosporine induced 45Ca 2+ accumulation results from sequestration of 45Ca 2+ within intracellular stores. While thapsigargin has no significant effect, concomitant treatment with tBHQ prevents the increase in 45Ca 2+ uptake associated with staurosporine treatment. Therefore, the tBHQ-sensitive Ca 2+ store, but not the thapsigargin/inositol 1,4,5-triphosphate-sensitive Ca 2+ store, appears to be staurosporine-sensitive. Overall, these studies indicate that staurosporine reduces catecholamine release by interfering with Ca 2+ homeostasis. Furthermore, this work suggests that a staurosporine-sensitive phosphoprotein(s) is involved with the regulation of Ca 2+ homeostasis in bovine adrenal chromaffin cells.