Stimulation of the release of [32P]guanosine 5'-diphosphate from G proteins in the rat anterior pituitary lobe by gonadotrophin-releasing and thyrotrophin-releasing hormones.
Plasma membranes (1-2 mg protein) purified from the anterior pituitary lobes of adult male rats were incubated with 0.6 mumol [alpha-32P]guanosine 5'-triphosphate (GTP) l-1 in an ATP-regenerating buffer at 37 degrees C for 60 min; during this incubation the [32P]GTP was hydrolysed and the nucleotide that was predominantly bound to the membranes was [32P]GDP. The release of [32P]GDP from the membranes was monitored at 37 degrees C; the amount released was proportional to the protein concentration and increased as a function of time. 5'-Guanylylimidodiphosphate (Gpp(NH)p) increased [32P]GDP release by up to 30% at 0.1 mumol l-1. Although 10 nmol Gpp(NH)p l-1 had no effect on GDP release, it appeared to stabilize the hormonal effect by blocking further GDP-GTP exchange. Gonadotrophin-releasing hormone (GnRH) agonist and thyrotrophin-releasing hormone (TRH), at 0.1 mumol l-1 caused a maximum increase in the release of [32P]GDP of 31-38%. The GnRH agonist (0.1 mumol l-1) stimulated GDP release by 21%, 24%, 17% and 14% at 30 s, 1, 2 and 5 min, respectively. TRH (0.1 mumol l-1) stimulated GDP release by 38%, 30%, 17% and 16% at 30 s, 1, 2 and 5 min, respectively. A GnRH antagonist also stimulated [32P]GDP release, albeit less effectively than GnRH agonist; the antagonist did not inhibit agonist stimulation of GDP release. These results indicate that ligand binding to the GnRH and TRH receptors results in interaction of the receptor with a guanine-nucleotide-dependent transducer protein (G protein) and activation of GTP-GDP exchange.(ABSTRACT TRUNCATED AT 250 WORDS)
Ravindra
R
R
Department of Medicine, East Carolina University, Greenville 27858-4354.
Aronstam
R S
RS
D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology*; D006153:Guanosine Diphosphate / Q000096:biosynthesis*; D008297:Male; D010903:Pituitary Gland, Anterior / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D013268:Stimulation, Chemical; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology*
D006153:Guanosine Diphosphate; D007987:Gonadotropin-Releasing Hormone; D013973:Thyrotropin-Releasing Hormone; D019204:GTP-Binding Proteins