SIGNALING PATHWAYS INVOLVED IN GNRH SECRETION IN GT(1) CELLS

The GT(1) GnRH neuronal cell lines exhibit highly differentiated properties of GnRH neurons. We have used GT(1-1) cells to study the role of the cyclic AMP/protein kinase A, cyclic GMP/protein kinase G and Ca2+/protein kinase C signaling pathways in the regulation of GnRH secretion. Superfusion of GT(1-1) cells with the cyclic AMP analog 8-Br-cyclic AMP (0.5 and 2.5 mM) or the adenylate cyclase activator forskolin (1 and 10 mu M) for 100 min increased the amplitude of GnRH secretion 2- to 35-fold. The cyclic GMP analog 8-Br-cyclic GMP (2.5 mM) also stimulated the amplitude of GnRH release from superfused GT(1-1) cells, although to a much lesser extent (1.5- to 3-fold). The amplitude of GnRH pulses was also stimulated (5- to 50-fold) by the protein kinase C activator TPA (1 mu M). Increasing intracellular Ca2+ with an ionophore (ionomycin, 1 mu M) or by the Ca2+ channel activator Bay K 8644 (10 mu M) also stimulated GnRH release, while secretion was markedly decreased and spontaneous pulsatility abolished by the L-type Ca2+ channel blocker methoxyverapamil(10 mu M). These results demonstrate that in GT(1) cells the protein kinase A, protein kinase G and protein kinase C pathways are functionally coupled to regulation of GnRH secretion. Furthermore, pulsatile GnRH secretion is coupled to the entry of extracellular Ca2+ via L-type Ca2+ channels.