Inhibitory effects of antagonistic analogs of GHRH on GH3 pituitary cells overexpressing the human GHRH receptor

• Published in: Journal of endocrinology Vol. 175; no. 2; pp. 425 - 434
• Main Authors: Kovacs, M, Schally, AV, Lee, EJ, Busto, R, Armatis, P, Groot, K, Varga, JL
• Format: Journal Article
• Language: English
• Published: Colchester BioScientifica 01.11.2002; Portland Press
• Subjects: Animals; Biological and medical sciences; Cyclic AMP - analysis; Cyclic AMP - biosynthesis; Fundamental and applied biological sciences. Psychology; Gene Expression - genetics; Growth Hormone - analysis; Growth Hormone - metabolism; Growth Hormone-Releasing Hormone - analogs & derivatives; Growth Hormone-Releasing Hormone - antagonists & inhibitors; Growth Hormone-Releasing Hormone - genetics; Hormones and neuropeptides. Regulation; Humans; Hypothalamus. Hypophysis. Epiphysis. Urophysis; Pituitary Gland - cytology; Pituitary Gland - physiology; Prolactin - analysis; Rats; Receptors, Somatotropin - genetics; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; Tumor Cells, Cultured; Vertebrates: endocrinology; Hypothalamic hormone; Pituitary gland; Antagonist; Hormone releasing factor; Somatotropin releasing factor; Hormonal receptor
• ISSN: 0022-0795; 1479-6805
• DOI: 10.1677/joe.0.1750425

GH3 rat pituitary tumor cells produce GH and prolactin (PRL), but lack the GHRH receptor (GHRH-R). We expressed human GHRH-R (hGHRH-R) in GH3 cells using recombinant adenoviral vectors and studied the effects of GHRH antagonists. The mRNA expression of the GHRH-R gene in the cells was demonstrated by RT-PCR. An exposure of the GH3 cells infected with hGHRH-R to 10(-10), 10(-9) and 10(-8) m hGHRH for 1 or 2 h in culture caused a dose-dependent elevation of the intracellular cAMP concentration and the cAMP efflux. Exposure to hGHRH also elicited dose-dependent increases in GH and PRL secretion from these cells. Neither the uninfected nor the antisense hGHRH-R-infected control cells exhibited cAMP, GH and PRL responses to GHRH stimulation. GHRH antagonists JV-1-38 and jv-1-36 applied at 3x10(-8) m for 3 h, together with 10(-9) m GHRH, significantly inhibited the GHRH-stimulated cAMP efflux from the hGHRH-R-infected cells by 36 and 80% respectively. The more potent antagonist JV-1-36 also decreased the intracellular cAMP levels in these cells by 55%. Exposure to JV-1-36 for 1 h nullified the stimulatory effect of GHRH on GH secretion and significantly inhibited it by 64 and 77% after 2 and 3 h respectively. In a superfusion system, GHRH at 10(-10), 10(-9) and 10(-8) m concentrations induced prompt and dose-related high cAMP responses and smaller increases in the spontaneous GH secretion of the hGHRH-R-infected cells. Antagonists JV-1-36 and JV-1-38 applied at 3x10(-8) m for 15 min, together with 10(-9) m GHRH, inhibited the GHRH-stimulated cAMP response by 59 and 35% respectively. This work demonstrates that GHRH antagonists can effectively inhibit the actions of GHRH on the hGHRH-R. Our results support the view that this class of compounds would be active clinically.