Characterization of a plasma membrane calcium oscillator in rat pituitary somatotrophs

In excitable cells, oscillations in intracellular free calcium concentrations ([Ca2+](i)) can arise from action-potential-driven Ca2+ influx, and such signals can have either a localized or global form, depending on the coupling of voltage-gated Ca2+ influx to intracellular Ca2+ release pathway. Here we show that rat pituitary somatotrophs generate spontaneous [Ca2+](i) oscillations, which rise from fluctuations in the influx of external Ca2+ and propagate within the cytoplasm and nucleus. The addition of caffeine and ryanodine, modulators of ryanodine-receptor channels, and the depletion of intracellular Ca2+ stores by thapsigargin and ionomycin did not affect the global nature of spontaneous [Ca2+](i) signals. Bay K 8644, an L-type Ca2+ channel agonist, initiated [Ca2+](i) signaling in quiescent cells, increased the amplitude of [Ca2+](i) spikes in spontaneously active cells, and stimulated growth hormone secretion in perifused pituitary cells. Nifedipine, a blocker of L-type Ca2+ channels, decreased the amplitude of spikes and basal growth hormone secretion, whereas Ni2+, a blocker of T-type Ca2+ channels, abolished spontaneous [Ca2+](i) oscillations, Spiking was also abolished by the removal of extracellular Na+ and by the addition of 10 mM Ca2+, Mg2+, or Sr2+, the blockers of cyclic nucleotide-gated channels. Reverse transcriptase-polymerase chain reaction and Southern blot analyses indicated the expression of mRNAs for these channels in mixed pituitary cells and purified somatotrophs, Growth hormone-releasing hormone, an agonist that stimulated cAMP and cGMP productions in a dose-dependent manner, initiated spiking in quiescent cells and increased the frequency of spiking in spontaneously active cells, These results indicate that in somatotrophs a cyclic nucleotide-controlled plasma membrane Ca2+ oscillator is capable of generating global Ca2+ signals spontaneously and in response to agonist stimulation. The Ca2+-signaling activity of this oscillator is dependent on voltage-gated Ca2+ influx but not on Ca2+ release from intracellular stores.