Cryptic gonadotropin-releasing hormone receptors of rat pituitary cells in culture are unmasked by epidermal growth factor.
Protein kinase activators as well as several neuropeptides are able to increase the GnRH-binding capacity of cultured adenohypophyseal cells. To determine whether such up-regulation of GnRH-binding sites can be achieved by a substance(s) endogenous to the pituitary, binding experiments were performed after exposure of cells to increasing amounts of medium conditioned by incubation with primary cultures of adenohypophyseal cells for 4 days. Addition of the conditioned medium elicited a 50% increase in GnRH binding. Characterization of the agent(s) responsible for the effect was attempted by submitting the conditioned medium to molecular sieve filtration, adding or immunoprecipitating endogenous substances, and comparing the susceptibilities of the responses to various inhibitors of transduction processes. Fractionation of the medium indicated that active molecules were of a proteic nature, with M(r) ranging from 5,000-10,000. Among major endogenous moieties corresponding to these criteria [epidermal] growth factor (EGF), transforming growth factor-alpha, and insulin-like growth factors I and II), only the first two exhibited properties similar to those of the conditioned medium. EGF stimulated binding with an EC50 of 3.6 +/- 0.8 pM. Immunoprecipitation of EGF, but not transforming growth factor-alpha, inactivated the conditioned medium. The effects of both conditioned medium and EGF were inhibited by herbimycin, a tyrosine kinase inhibitor; U73122, a phospholipase C inhibitor; and prior desensitization of protein kinase C. In contrast, both were insensitive to pertussis toxin pretreatment. In parallel, EGF did not increase LH secretion by itself, but potentiated its response to GnRH in a concentration range of 1 pM to 1 nM, resulting in a shift of the curve toward lower values of GnRH. It is concluded that EGF is able to control the accessibility of binding sites to GnRH and to potentiate the responsiveness of gonadotropes to the decapeptide.
Leblanc
P
P
Unité de Dynamique des Systèmes Neuroendocriniens, INSERM U-159, Centre Paul Broca, Paris, France.
L'Héritier
A
A
Kordon
C
C
D000818:Animals; D016227:Benzoquinones; D002478:Cells, Cultured; D002850:Chromatography, Gel; D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism; D007163:Immunosorbent Techniques; D047029:Lactams, Macrocyclic; D007986:Luteinizing Hormone / Q000378:metabolism; D008297:Male; D010903:Pituitary Gland, Anterior / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011809:Quinones / Q000494:pharmacology; D051381:Rats; D011966:Receptors, LHRH / Q000378:metabolism*; D017828:Rifabutin / Q000031:analogs & derivatives; D016211:Transforming Growth Factor alpha / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors
D016227:Benzoquinones; D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D047029:Lactams, Macrocyclic; D011809:Quinones; D011966:Receptors, LHRH; D016211:Transforming Growth Factor alpha; D017828:Rifabutin; D007987:Gonadotropin-Releasing Hormone; D004815:Epidermal Growth Factor; C020754:herbimycin; D007986:Luteinizing Hormone; D011505:Protein-Tyrosine Kinases; D010738:Type C Phospholipases