Veratridine and ouabain stimulate calcium-dependent prolactin release

We have investigated the effects of veratridine, a Na+ channel activator, and ouabain, an inhibitor of Na+-K+-ATPase, on short term (1-h) PRL release from primary cultures of rat anterior pituitary cells and from the rat anterior pituitary cell line GH4C1 in culture. Both compounds should increase intracellular Na+. Veratridine (20-500 microM) and ouabain (0.1-3 mM) stimulated PRL release from normal cells. The stimulation was inhibited by the omission of Ca++ from the release buffer or by preincubation with the calcium channel blocker D600 (20-500 microM), suggesting a role for Ca++ in the action of these compounds. Ouabain (1 mM), but not veratridine (200 microM), stimulated PRL release from GH4C1 cells, an effect that was also inhibited by calcium channel blockers. In the presence of the dopaminergic agonist bromocriptine (30 nM), the amount of stimulated release by veratridine (200 microM) and ouabain (1 mM) was reduced by 50%. The veratridine effect was only partially inhibited by preincubation of the cells with the Na+ channel blocker tetrodotoxin (1 and 10 microM), but the effect was inhibited completely when Na+ in the buffer was replaced by choline, suggesting that the action of veratridine requires extracellular Na+. The results of this study indicate that 1) ouabain- and veratridine-stimulated PRL release are largely dependent on Ca++; 2) veratridine appears to act through a tetrodotoxin-insensitive mechanism; and 3) stimulation of PRL release by these compounds is similar to that by 50 mM KCl and cAMP in its sensitivity to bromocriptine.