Intracellular mechanisms involved in the stimulation of growth hormone release from rat anterior pituitary cells by Russell's viper venom

Abstract

ABSTRACT We have previously shown that a heat-stable component of Russell's viper venom (RVV) releases GH in a dose-dependent manner from cultured rat anterior pituitary cells. We have now investigated the intracellular mechanisms involved in RVV-stimulated GH release by concomitant administration of RVV with known intracellular mediators in rat pituitary cells. 3-Isobutyl-1-methylxanthine (IBMX; 0·5 mmol/l), added to cultured rat anterior pituitary cells simultaneously with RVV, at concentrations up to a maximally effective dose of 10 μg/ml, increased GH release (3·7-fold, 4·0-fold and 2·0-fold; P < 0·001) compared with the effect of venom alone. These effects were additive, indicating that RVV and IBMX stimulate through different intracellular messengers. RVV failed to increase the formation of basal or IBMX-stimulated intracellular cyclic AMP (cAMP), confirming that RVV affects GH release through a cAMP-independent pathway. 12-0-Tetradecanoylphorbol-13-acetate (TPA; 0·1 μmol/l), added simultaneously with various doses of RVV (0·1–10 μg/ml), did not increase GH release beyond the maximal effect of RVV. This result indicates that RVV might be stimulating GH release through a similar mechanism to that of TPA (by activating protein kinase C). When pituitary cells were perifused with Ca2+-free medium or verapamil (50 μmol/l), RVV-stimulated GH release was inhibited by 65 and 42% respectively. This reflects the recognized requirement of Ca2+ for secretory processes. However, RVV (10 μg/ml) had no significant effect on intracellular free Ca2+ concentrations as measured using the fluorescent Ca2+ probe quin-2. These findings indicate that the mechanism of action of RVV on GH release is independent of a change in both cAMP levels and intracellular free Ca2+ concentrations, and is dependent upon protein kinase C. Journal of Endocrinology (1990) 127, 111–117