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To examine the role of calcium signaling in the joint modulation of gonadotropin-releasing hormone (GnRH) secretion by gamma-aminobutyric acid (GABA) and glutamate, cytoplasmic calcium ([Ca2+]i) responses to the two transmitters were analyzed in monolayer networks of the GT1-7 line of immortalized GnRH neurons. [Ca2+]i was increased by GABA and the GABAA receptor agonist, muscimol, and these responses were inhibited by the GABAA receptor antagonist, bicuculline. In contrast, the GABAB receptor agonist, baclofen, and the GABAB receptor antagonist, phaclofen, had no effect on basal and GABA- and glutamate-induced Ca2+ levels in GT1-7 neurons. The GABA- and muscimol-induced responses consisted of a spike increase in [Ca2+]i followed by a decrease to a plateau; both the increase and the subsequent decrease in [Ca2+]i depended on agonist concentration. Glutamate, N-methyl-D-aspartate (NMDA), and kainate also increased [Ca2+]i, but were less effective than GABA. GABA-, glutamate-, NMDA-, and kainate-induced [Ca2+]i responses were almost abolished in Ca(2+)-free medium and were markedly attenuated by nifedipine. The [Ca2+]i response to GABA was unaffected by prior application of glutamate, and vice versa. This additive effect of glutamate on the GABA-induced [Ca2+]i response was mimicked by prior or simultaneous application of low (10 mM) KCl. The [Ca2+]i response to simultaneous application of GABA and glutamate was also equal to the sum of the individual responses, whereas the GnRH secretory response was larger. However, the secretory responses to GABA and glutamate applied individually or together, were markedly attenuated by nifedipine.(ABSTRACT TRUNCATED AT 250 WORDS)

Glutamate and its receptors are present in the hypothalamus and have been proposed to participate in neuroendocrine regulation, including the control of GnRH secretion. To address the mechanism of glutamate action, we measured [Ca2+]i, inositol phosphate, and secretory responses to glutamate receptor subtype agonists and antagonists in the immortalized GT1-7 cell line of GnRH-secreting hypothalamic neurons. Glutamate, N-methyl-D-aspartate (NMDA), kainate, and trans-(+/-)-1-amino-(1S,3R)-cyclopentanedicarboxylic acid increased GnRH secretion. In monolayer cultures of GT1-7 cells, L- but not D-glutamate induced a moderate, concentration-dependent rise in [Ca2+]i. The action of glutamate on [Ca2+]i was mimicked by NMDA, alpha-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropanoic acid (AMPA), and kainate. Responses to NMDA were potentiated by the coagonist, glycine, and were inhibited by an antagonist of the glycine site on the NMDA receptor, 5,7-dichlorokynurenic acid (DCKA). NMDA-induced [Ca2+]i responses were also inhibited by Mg2+ and by the NMDA receptor antagonist, (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,1 0-imine hydrogen maleate (MK-801), but not by the AMPA/kainate antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). In contrast, responses to AMPA and kainate were inhibited by CNQX but not by Mg2+, DCKA, or MK-801. Responses to glutamate were more inhibited by MK-801 plus CNQX than by either antagonist alone. All [Ca2+]i responses were nearly abolished in Ca(2+)-free solution. None of the agonists stimulated inositol phosphate formation.(ABSTRACT TRUNCATED AT 250 WORDS)