Changes in cytosolic ionized calcium induced by activators of secretion in GH3 cells

Cytosolic ionized calcium (Cai) was measured in GH3 cells with aequorin incorporated by hypoosmotic shock treatment. Cai rose from 0.4 to 1.3 microM after the administration of 10(-7) M thyrotropin-releasing hormone (TRH) and from 0.3 to 1.1 microM after depolarization caused by high-K concentration (60 mM). Repeated TRH stimulations decreased the magnitude of the Cai response. This magnitude was directly related to the interval between stimuli. Incubations in low-calcium media (0.13 mM Ca) for 0.5 h did not alter the Cai response to TRH, but incubations in Ca-free media reduced the response. These results suggest that TRH mobilized calcium from an endogenous pool of Ca that can be depleted and replenished. The rise in Cai induced by K depolarization was slower than that evoked by TRH (time constant, t = 9 s for TRH and t = 111 s for K depolarization). Even when Cai was elevated by K depolarization, it could still be further increased by TRH, suggesting that these two stimuli mobilize calcium from two different pools and by different mechanisms.