Mechanisms involved in ATP-evoked Ca 2+ oscillations in isolated human granulosa-luteal cells

Abstract

Using single-cell microfluorimetry, we have shown that ATP evoked repetitive Ca 2+ oscillations in intact Fura-2 loaded human granulosa-luteal cells (hG/LCs) in the absence of extracellular Ca 2+. Sustained increases in [Ca 2+] i required extracellular Ca 2+ and ATP depleted stores were refreshed by brief (2 min) incubation with external Ca 2+ Basal [Ca 2+] i was unaffected by caffeine (1 mM), but 20 mM caffeine inhibited ATP-evoked Ca 2+ release in the absence of external Ca 2+. Thimerosal (10 μM) evoked repetitive Ca 2+ spikes, under Ca 2+-free conditions, which fused to form an elevated plateau when external Ca 2+ was replaced. Thimerosal-induced changes in [Ca 2+] i were reversibly inhibited by the thiol reducing agent dithiothreitol (1 mM). The periodicity and amplitude of the [Ca 2+] i oscillations produced by thimerosal and ATP differ. ATP- or thimerosal-evoked changes in [Ca 2+] i were unaffected by dantrolene sodium (10 μM). The Ca 2+-ATPase inhibitor thapsigargin (1 μM) increased [Ca 2+] i and attenuated subsequent ATP-evoked changes in [Ca 2+] i. We conclude that ATP stimulates an oscillatory release of Ca 2+ from InsP 3-sensitive stores in hG/LCs.