Evidence that guanosine 5′-[γ-thio]triphosphate stimulates plasma membrane Ca2+ inflow when introduced into hepatocytes

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Abstract
1. Slowly hydrolysable analogues of GTP were introduced into hepatocytes by incubating the cells in the absence of Mg2+ and in the presence of ATP4-. Experiments using guanosine 5′-[gamma-[35S]thio]triphosphate (GTP[35S])indicated that about 50% of the GTP[S] loaded into the cells was subsequently hydrolysed. 2. In cells loaded with GTP[S] and incubated in the absence of added extracellular Ca2+ (Ca2+o), the rate of activation of glycogen phosphorylase observed after addition of 1.3 mM-Ca2+o was 250% greater than the rate observed in unloaded cells. Smaller effects (130%) were observed in cells loaded with either guanyl-5′-yl imidodiphosphate or guanosine 5-[beta-thio]diphosphate (GDP[S]). Cells loaded with adenosine 5′-[gamma-thio]triphosphate showed no increase in glycogen phosphorylase activity on addition of Ca2+o. 3. The effect of a submaximal concentration of GTP[S] on the Ca2+-induced activation of glycogen phosphorylase was additive with that of a half-maximally effective concentration of vasopressin. GTP[S] did not increase the effect of a maximally effective concentration of the hormone. 4. Cells loaded with GTP[S] exhibited an increased initial rate of 45Ca2+ exchange measured at 1.3 mM-Ca2+o. 5. GTP[S] did not affect the amount of 45Ca2+ exchanged by cells incubated at 0.1 mM-Ca2+o or the ability of vasopressin to release 45Ca2+ from these cells. 6. It is concluded that the introduction of slowly hydrolysable analogues of GTP to the liver cell cytoplasmic space stimulates the inflow of Ca2+ across the plasma membrane through a channel similar to that activated by vasopressin.

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