PARTIAL CHARACTERIZATION OF K+-INDUCED INCREASE IN [CA2+](CYT) AND GNRH RELEASE IN GT1-7 NEURONS

Secretion of pituitary gonadotropins is regulated centrally by the hypothalamic decapeptide gonadotropin releasing hormone (GnRH). Using the immortalized hypothalamic GT1-7 neuron, we characterized pharmacologically the dynamics of cytosolic Ca2+ and GnRH release in response to K+-induced depolarization of GT1-7 neurons. Our results showed that K+ concentrations from 7.5 to 60 mM increased [Ca2+](cyt) in a concentration-dependent manner. Resting [Ca2+](cyt) in GT1-7 cells was determined to be 69.7 +/- 4.0 nM (mean +/- S.E.M.; n = 69). K+-induced increases in [Ca2+](cyt) ranged from 58.2 nM at 7.5 mM [K+] to 347 nM at 60 mM [K+]. K+-induced GnRH release ranged from about 10 pg/ml at 7.5 mM [K+] to about 60 pg/ml at 45 mM [K+]. K+-induced increases in [Ca2+](cyt) and GnRH release were enhanced by 1 mu M BayK 8644, an L-type Ca2+ channel agonist. The BayK enhancement was completely inhibited by 1 mu M nimodipine, an L-type Ca2+ channel antagonist. Nimodipine (1 mu M) alone partially inhibited K+-induced increases in [Ca2+](cyt) and GnRH release. Conotoxin (1 mu M) alone had no effect on K+-induced GnRH release or [Ca2+](cyt), but the combination of conotoxin (1 mu M) and nimodipine (1 mu M) inhibited K+-induced increase in [Ca2+](cyt) significantly more (p < 0.02) than nimodipine alone, suggesting that N-type Ca2+ channels exist in GT1-7 neurons and may be part of the response to Kf. The response of [Ca2+](cyt) to K+ was linear with increasing [K+] whereas the response of GnRH release to increasing [K+] appeared to be saturable. K+-induced increase in [Ca2+](cyt) and GnRH release required extracellular [Ca2+]. These experiments suggest that voltage dependent N- and L-type Ca2+ channels are present in immortalized GT1-7 neurons and that GnRH release is, at least in part, dependent on these channels for release of GnRH.