DYNAMIC VIDEO IMAGING OF CYSTOLIC CA-2+ IN THE ALPHA-T3-1, GONADOTROPE-DERIVED CELL-LINE

Studies with pituitary cell cultures have revealed Ca2+ as a mediator of gonadotropin-releasing hormone (GnRH) action. Here we have used dynamic video imaging to characterize Ca2+ metabolism in the gonadotrope-derived, αT3-1 cell line focusing on stimuli that influence Ca2+ metabolism of gonadotropes in primary culture. At least 90% of the cells imaged responded to 100 nMGnRH with an increase in cytosolic Ca2+ (typically 10- to 20-fold). GnRH (10-1000 nM) caused a biphasic "spikeplateau" increase, whereas the spike phase was not seen with 0.01-1 nM GnRH. The plateau response was blocked by Ca2+-free medium and inhibited by 10 μM nifedipine (a dihydropyridine Ca2+ channel antagonist) but not by blockade of voltage-sensitive Na+ channels with 10 μM tetrodotoxin. In Ca2+-free medium GnRH caused only a transient Ca2+ increase. Endothelin-1 (400 nM) also increased Ca2+ although only 30-50% of the cells responded and the average response was less than that with 100 nM GnRH. Depolarization with 30 mM KCl increased Ca2+ in >90% of the cells with an average effect comparable to the 100 nM GnRH effect. Protein kinase C activation approximately doubled the Ca2+ concentration and induced Ca2+ oscillations in some cells. The close parallels between responses of these cells and those observed with primary cultures suggests that αT3-1 cells will provide a valuable model for further studies of the involvement of Ca2+ in GnRH action. © 1992.