Histamine-evoked Ca²⁺ oscillations in HeLa cells are sensitive to methylxanthines but insensitive to ryanodine

Abstract

The relative contribution of inositol-trisphosphate(InsP ₃)-sensitive and InsP ₃-insensitive Ca²⁺ stores to the agonist-evoked oscillatory release of Ca²⁺ in HeLa cells was investigated using fura-2 cytosolic Ca²⁺ measurements and whole-cell recordings of Ca²⁺-activated K⁺ currents [K(Ca²⁺)]. The experimental approach chosen consisted in studying the effects on Ca²⁺ oscillations of a variety of pharmacological agents such as ryanodine, ruthenium red, caffeine and theophylline, which are known to affect the Ca²⁺ channels responsible for Ca²⁺-induced Ca²⁺ release (CICR) in excitable cells. The results obtained essentially indicate (a) that neither ryanodine nor ruthenium red affects the generation of periodic K(Ca²⁺) current pulses in whole-cell experiments, and (b) that histamine-induced Ca²⁺ oscillations are inhibited by caffeine and theophylline in a dose-dependent manner. However, these methylxanthines were unable, at concentrations ranging from 0.1 mM to 10 mM, either to mobilize Ca²⁺ from internal stores or to block the initial Ca²⁺ rise evoked by histamine. In addition, both methylxanthines showed at high concentrations (10–20 mM) a moderate inhibitory action on the production of InsP ₃ induced by histamine. This effect was not essential to the action of caffeine on the oscillatory release of Ca²⁺, since an inhibition by caffeine of InsP ₃-induced Ca²⁺ oscillations was still observed in whole-cell experiments where the InsP ₃ concentration was kept constant. The results also show (c) that the application of either caffeine or theophylline during histamine stimulation leads systematically to an increased Ca²⁺ sequestration in InsP ₃-sensitive Ca²⁺ pools, the effect observed with theophylline being stronger than that resulting from the application of caffeine, and finally (d) that the action of caffeine and theophylline is not related to an increase in cAMP concentration since neither forskolin (10–50 μM) nor 8-Br-cAMP (1 mM) caused an inhibition of the InsP ₃-induced Ca²⁺ oscillations. It is concluded on the basis of these results that the agonist-evoked Ca²⁺ oscillations in HeLa cells do not involve directly or indirectly a ryanodine-sensitive Ca²⁺-release channel with CICR properties, but rather arise from a control by Ca²⁺ of the InsP ₃ Ca²⁺-release process.