3′,5′-cyclic adenosine monophosphate augments intracellular Ca2+ concentration and gonadotropin-releasing hormone (GnRH) release in immortalized GnRH neurons in an Na+-dependent manner

In GT1-7 cells, cAMP increases the intracellular Ca2+ concentration ([Ca2+](i)) through activation of the voltage-gated Ca2+ channels, thereby facilitating GnRH release. To activate these channels, the membrane potential must be depolarized. In the present study we hypothesize that cAMP depolarizes the cells by increasing the membrane Na+ permeability, as in the case of somatotrophs and pancreatic beta-cells. To examine this, we analyzed [Ca2+](i) and [Na+](i) in GT1-7 cells by an intracellular ion-imaging technique along with cAMP assay by RIA. Forskolin, a direct activator of adenylyl cyclase, increased [Ca2+](i) and [Na+](i) via cAMP formation. The forskolin-induced increase in [Ca2+](i) depended on the presence of Ca2+ and Na+ in the extracellular solution. This response was blocked by the voltage-gated Ca2+ channel blocker, nifedipine; the nonselective cation channel blocker, gadolinium (Gd3+); and the cyclic nucleotide-gated channel blocker, l-cis-diltiazem. In contrast, the forskolin-induced increase in [Na+](i) depended only on extracellular Na+, not on Ca2+. Gd3+ and l-cis-diltiazem also blocked the increase in [Na+](i). Furthermore, the forskolin-induced increase in GnRH release was blunted in both low Ca2+ and low Na+ media. The results indicate that cAMP increases the membrane Na+ permeability, probably through nonselective cation, channels on GT1-7 cells, thereby promoting GnRH release.