EFFECT OF LUTEINIZING HORMONE RELEASING HORMONE
ON ACCUMULATION OF PITUITARY CYCLIC AMP AND GMP IN VITRO G. L. RIGLER, G. T. PEAKE AND A. RATNER Departments of Physiology and Medicine, University of New Mexico School of Medicine, Albuquerque, New Mexico 87131, U.S.A.
(Received 19 August 1977)
Cyclic AMP has been increasingly implicated as an important intracellular mediator regulating the secretion of peptide hormones (Vale, Grant Guillemin, 1973), but there is controversy concerning its role in the regulation of the luteinizing hormone releasing hormone (LH-RH)-stimulated secretion of luteinizing hormone (LH; Labrie, Pelletier, Borgeat, Drouin, Ferland & Belanger, 1976; Ratner, Wilson, Srivastava & Peake, 1976; Sundberg, Fawcett & McCann, 1976). Although less clearly established, the guanyl cyclase\p=n-\cyclicGMP system may also play an important role in regulating cellular events (Goldberg, O'Dea & Haddox, 1973). Studies with a highly purified growth hormone (GH) releasing factor suggested the involvement of cyclic GMP in the mediation of GH release from the somatotrope (Wilson, Steiner, Dhariwal & Peake, 1974). This communication describes studies which suggest a role for the guanylate cyclase\p=n-\cyclicGMP system in the regulation of the LH-RH-stimulated secretion of LH. Male Sprague-Dawley rats were decapitated and the anterior pituitary glands removed and incubated as described previously (Wilson et al. 1974). Each incubation flask contained four hemipituitary glands. After pre-incubation for 60 min, the medium from both control and experimental flasks was replaced with 1 ml fresh medium and 20 ng LH-RH was added to the experimental flasks. After incubation, tissue extracts were assayed for cyclic AMP and GMP by radioimmunoassay as described previously (Steiner, Kipnis, Utiger & Parker, 1969) and medium was stored at —20 °C until assayed for LH (reagents for radioimmuno¬ assay of LH kindly provided by NIAMDD). The present test system was validated by determining the dose-related release of LH in response to 0-8, 4, 20 or 100 ng LH-RH/ml after incubation for 120 min. Since 20 ng LH-RH/ml was shown to be a reproducible stimulus (214% of control) for secretion of LH in vitro, this dose was chosen for studies of the effect of LH-RH on the levels of cyclic nucleotides in tissue. The secretion of LH was significantly increased after incubation with LH-RH for 60 min and a significant increase in the pituitary content of cyclic AMP (control, 1-34 + 0-2 (s.e.m.) pmol AMP/mg; LH-RH-treated, 2-60 ±0-2 pmol AMP/mg; &
ON ACCUMULATION OF PITUITARY CYCLIC AMP AND GMP IN VITRO G. L. RIGLER, G. T. PEAKE AND A. RATNER Departments of Physiology and Medicine, University of New Mexico School of Medicine, Albuquerque, New Mexico 87131, U.S.A.
(Received 19 August 1977)
Cyclic AMP has been increasingly implicated as an important intracellular mediator regulating the secretion of peptide hormones (Vale, Grant Guillemin, 1973), but there is controversy concerning its role in the regulation of the luteinizing hormone releasing hormone (LH-RH)-stimulated secretion of luteinizing hormone (LH; Labrie, Pelletier, Borgeat, Drouin, Ferland & Belanger, 1976; Ratner, Wilson, Srivastava & Peake, 1976; Sundberg, Fawcett & McCann, 1976). Although less clearly established, the guanyl cyclase\p=n-\cyclicGMP system may also play an important role in regulating cellular events (Goldberg, O'Dea & Haddox, 1973). Studies with a highly purified growth hormone (GH) releasing factor suggested the involvement of cyclic GMP in the mediation of GH release from the somatotrope (Wilson, Steiner, Dhariwal & Peake, 1974). This communication describes studies which suggest a role for the guanylate cyclase\p=n-\cyclicGMP system in the regulation of the LH-RH-stimulated secretion of LH. Male Sprague-Dawley rats were decapitated and the anterior pituitary glands removed and incubated as described previously (Wilson et al. 1974). Each incubation flask contained four hemipituitary glands. After pre-incubation for 60 min, the medium from both control and experimental flasks was replaced with 1 ml fresh medium and 20 ng LH-RH was added to the experimental flasks. After incubation, tissue extracts were assayed for cyclic AMP and GMP by radioimmunoassay as described previously (Steiner, Kipnis, Utiger & Parker, 1969) and medium was stored at —20 °C until assayed for LH (reagents for radioimmuno¬ assay of LH kindly provided by NIAMDD). The present test system was validated by determining the dose-related release of LH in response to 0-8, 4, 20 or 100 ng LH-RH/ml after incubation for 120 min. Since 20 ng LH-RH/ml was shown to be a reproducible stimulus (214% of control) for secretion of LH in vitro, this dose was chosen for studies of the effect of LH-RH on the levels of cyclic nucleotides in tissue. The secretion of LH was significantly increased after incubation with LH-RH for 60 min and a significant increase in the pituitary content of cyclic AMP (control, 1-34 + 0-2 (s.e.m.) pmol AMP/mg; LH-RH-treated, 2-60 ±0-2 pmol AMP/mg; &
