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Intracellular antagonistic interaction between GnRH and gonadotrophin surge-inhibiting/attenuating factor bioactivity downstream of second messengers involved in the self-priming process

The antagonistic control of LH concentrations by GnRH and the putative ovarian factor gonadotrophin surge-inhibiting or -attenuating factor (GnSIF/AF) was studied by perifusion of female rat pituitary glands in vitro. LH release and GnRH self-priming were measured in response to five (0.32–10 nmol l^–1) or three (10 nmol l^–1) 10 min pulses of GnRH. In the latter case, pulses were preceded by five 10 min pulses of either 1 mmol 8-bromo-cAMP l^–1 plus 10 mmol theophylline l^–1 or 1 µmol phorbol 12-myristate 13-acetate l^–1. The stimulations were carried out in the presence or absence of steroid-free bovine follicular fluid, which possesses GnSIF/AF bioactivity, to study the effect of GnSIF/AF on the self-priming process during successive stimulations by GnRH. First, the effect of follicular fluid was studied on GnRH-induced LH release from pituitary glands collected during the ovarian cycle. Only when a clear self-priming effect was evident, as on day 2 of dioestrus and the day of pro-oestrus, follicular fluid antagonized the self-priming effect of GnRH. Second, when glands on day 2 of dioestrus were used, self-priming was displayed by various combinations of GnRH and follicular fluid. The pulses with the lowest concentrations of GnRH together with a high concentration of follicular fluid, however, led to stable low amplitude LH pulses. Finally, priming of the pituitary LH response to GnRH with 8-bromo-cAMP plus theophylline or phorbol 12-myristate 13-acetate was inhibited by follicular fluid. These results confirm the control of LH release by GnRH and GnSIF/AF bioactivity. The effect of GnSIF/AF in follicular fluid is most pronounced on the days before the LH surge, showing evidence of its important role in maintaining low LH concentrations during this period. GnSIF/AF neutralizes downstream actions of second messengers involved in GnRH self-priming.