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Unlabelled: Mediobasal hypothalamic slices of adult ovariectomized (OVX) rats treated or not with 17 beta-estradiol (E2) were superfused in buffered (pH 7.2) Locke medium containing bacitracin. A 6-min pulse of K+ (56 m M) was less effective in releasing luteinizing hormone releasing hormone (LH-RH) from mediobasal hypothalamic slices sampled from OVX rats than from OVX animals treated subcutaneously with either E2 or stilbestrol implants for 5 days; in contrast, the basal release of the neuropeptide was identical in both cases. Direct addition to the superfusion medium of 17 beta-estradiol (10(-10) to 10(-7) M) or stilbestrol (10(-8) M) potentiated the K+-induced LH-RH release from slices of OVX animals. The K+-induced LH-RH release observed after in vivo E2 implantation was not further amplified by in vitro addition of the hormone. Tamoxifen and hydroxytamoxifen, estrogen antagonists, were ineffective by themselves, but reversed the E2 facilitation of K+-evoked LH-RH release. In contrast, 17 alpha-estradiol, progesterone, or cholesterol (10(-8) or 10(-9) M) hat no effect on either basal or stimulated release of the neurohormone. Somatostatin release measured under identical conditions was not affected by castration or by in vitro addition of the steroid., In Conclusion: (1) estradiol appears selectively and specifically involved in the process coupling, nerve endings depolarization, and LH-RH release, and (2) the effect is receptor-mediated and does not appear to require nuclear translocation of the steroid or transcription processes, since it can be readily elicited upon addition of the hormone to nerve endings disconnected from their cell bodies.

Mediobasal hypothalamic (MBH) slices of adult ovariectomized (OVX) rats with or without 17 beta-estradiol (E2) pretreatment, were superfused in buffered (pH 7.2) oxygenated Locke medium containing bacitracin. Pulsatile or continuous administration of progesterone (10(-7) or 10(-8) M) produced a marked increase in luteinizing hormone-releasing hormone (LHRH) release provided the animals had received E2 prior to sacrifice. Omission of Ca++ in the medium, or addition of a Ca++ channel blocker (D-600, 10(-4) M), of a calmodulin inhibitor (trifluoperazine, 30 microM) or of a calmodulin-dependent tubulin kinase inhibitor (phenytoin, 50 microM), antagonized the stimulatory effect of progesterone. When sodium channels were blocked by tetrodotoxin (5 X 10(-7) M), the stimulatory effect of the steroid was completely abolished. The amplitude of the K+-induced LHRH release was slightly increased in the presence of progesterone (10(-7) M) but only from MBH slices of OVX-E2-treated rats. These results indicate that the secretory response of LHRH to progesterone requires priming with estradiol, is Ca++-dependent and involves mediation of calmodulin and a calmodulin-dependent kinase system.

The phasic luteinizing hormone (LH) release observed in ovariectomized (OVX), estrogen-implanted rats was further amplified and advanced when progesterone (P) was given 4 h prior to the gonadotropin surge. In contrast, an inhibitory effect of P on the daily LH surge was observed when P was administered 16-36 h prior to LH peak. In order to determine whether this biphasic action of P is primarily exerted on the release of luteinizing hormone releasing hormone (LHRH), on the pituitary response to LHRH, or on both, mediobasal hypothalamic slices or pituitary fragments of adult OVX rats or of OVX rats pretreated with estrogen alone or in combination with P were tested in a perifusion system. Mediobasal hypothalamic slices were perifused in buffered (pH 7.2) oxygenated Locke's medium containing bacitracin (2 X 10(-5) M). In the absence of estrogen pretreatment, high (56 mM) concentrations of K+ were barely effective in releasing LHRH. Subcutaneous implantation of 17 beta-estradiol for 5 days markedly increased the amplitude of the LHRH secretory response to K+ depolarization. Additional administration of P (25 mg/rat s.c.) 4 h before sacrifice further amplified the K+-induced LHRH release. In contrast, the K+-evoked LHRH secretion was significantly inhibited when P was given 16 or 36 h before. Estradiol thus appears to facilitate the LHRH secretory response to depolarizing stimuli, whereas P either enhances or blocks the induced LHRH release depending upon its time of administration. At the pituitary level, the sensitivity of LHRH-induced LH release was also increased after estrogen pretreatment.(ABSTRACT TRUNCATED AT 250 WORDS)