STABLE TRANSFECTION OF GH(3) CELLS WITH RAT GONADOTROPIN-RELEASING-HORMONE RECEPTOR COMPLEMENTARY DEOXYRIBONUCLEIC-ACID RESULTS IN EXPRESSION OF A RECEPTOR-COUPLED TO CYCLIC ADENOSINE 3',5'-MONOPHOSPHATE-DEPENDENT PROLACTIN-RELEASE VIA A G-PROTEIN

GH(3) cells, which normally release PRL in response to stimulation by TRH, have been stably transfected with rat GnRH receptor complementary DNA (GGH(3)-1 ' cells). Unlike the parent line, GGH(3)-1 ' cells expr ess GnRH receptor, which can be measured in a radioligand assay using a metabolically stable GnRH analog. The number of receptors (11,000 +/ - 2,800 receptors/cell; n = 3) and Kd (4.1 +/- 1.0 X 10(-8) M; n = 3), determined using a radioiodinated GnRH agonist, as well as binding in hibition values for GnRH agonists and antagonists and for unrelated su bstances suggest that this receptor is similar to those expressed in c ell cultures derived from rat pituitaries, although the binding affini ty is about 1 log lower in the former. Unlike GnRH-stimulated release of gonadotropins from primary pituitary cultures, which does not requi re protein synthesis and is not coupled to cAMP production, GnRH-stimu lated PRL release from the transfected cell line is absolutely depende nt on protein synthesis, and cAMP fulfills the requirements of a secon d messenger. The receptor appears to be coupled to adenylate cyclase-m ediated PRL release through a cholera toxin-sensitive G-protein. These studies provide functional evidence to support the view that the clon ed receptor is the physiological receptor for the releasing hormone, a nd that this receptor can differentially couple to G-proteins dependin g on their availability and accessibility in the target cell.