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Na⁺ Dependence of Gonadotropin-Releasing Hormone Action: Characterization of the Na⁺/H⁺ Antiport in Pituitary Gonadotropes

Institute for Hormone and Fertility Research 2000 Hamburg 54, Germany; and Nacogdoches, Texas 75963-1548 (E.J.C.)

Abstract

GnRH stimulates LH release from gonadotropes in a Ca²⁺-dependent manner. Because of the apparent relationship between cellular Ca²⁺ metabolism and Na⁺-driven antiports, we investigated their influence on GnRH action. We also assessed the influence of bicarbonate, because its transport may alter effects of Na⁺/H⁺ exchange on intracellular pH. In pituitary cell cultures without bicarbonate, GnRH-stimulated LH release was reduced by Na⁺ omission, by amiloride, and by amiloride analogs that selectively block Na⁺/H⁺ exchange. The Na⁺ dependence of amiloride action (EC6o, 14 and 100 nM in medium with 20 and 135 mM NaCl, respectively, and no effect in Na⁺-free medium) and the order of potency of these analogs, indicated specific inhibition of Na⁺/H⁺ exchange. 5-(N,N-Di- methyl)amiloride (DMA; a potent Na⁺/H⁺ exchange inhibitor) reduced GnRH-stimulated LH release, but not GnRH receptor binding or Ca²⁺ ionophore (A23187)-stimulated LH release, suggesting inhibition at a locus beyond receptor occupancy but before exocytosis. Amiloride analogs that selectively inhibit Na⁺/Ca²⁺ exchange also modestly reduced GnRH-stimulated LH release. Bicarbonate (10 mM) reduced the inhibitory effects of DMA and Na⁺ omission (but not the effects of the Na⁺/Ca²⁺ exchange inhibitors or of a Ca²⁺ channel antagonist), and the effect of bicarbonate was inhibited by a blocker of bicarbonatedependent antiports. These observations reveal the Na⁺ dependence of GnRH action and that gonadotropes possess a Na⁺/H⁺ exchanger. The Na⁺ dependence of GnRH-stimulated LH release appears to reflect at least in part dependence upon this antiport. Prevention of the Na⁺/H⁺ exchange inhibitor effects by bicarbonate supports the specificity of their action, but suggests regulation of this antiport as an unlikely means of controlling LH release in vivo. (Endocrinology 128: 771–778, 1991)

Received June 4, 1990.