COUPLING OF CORTICOTROPIN-RELEASING HORMONE RECEPTORS TO ADENYLYL-CYCLASE IN HUMAN Y-79 RETINOBLASTOMA CELLS

In human Y-79 retinoblastoma cells, corticotropin-releasing hormone (C RH) stimulates adenylyl cyclase activity and increases cyclic AMP accu mulation. Different CRH analogues mimic the CRH stimulation of adenyly l cyclase and show similar sensitivity to the CRH receptor antagonist alpha-helical CRH(9-41). Vasoactive intestinal peptide (VIP) also incr eases the enzyme activity but less potently than CRH, and its effect i s counteracted by the VIP receptor antagonist [D-p-Cl-Phe(6),Leu(17)] VIP. The VIP antagonist does not affect the response to CRH. The CRH-s timulated adenylyl cyclase activity is amplified by Mg2+, is inhibited by submicromolar concentrations of Ca2+, and requires GTP. Moreover, the CRH stimulation is reduced by pretreatment of cells with cholera t oxin and by incubation of membranes with the RM/1 antibody, which reco gnizes the C-terminus of the alpha subunit of G(s). In immunoblots, th e RM/1 antibody identifies a doublet of 45 and 52 kDa. Two proteins of similar molecular weights are ADP-ribosylated by cholera toxin. These data demonstrate that in human Y-79 retinoblastoma cells, specific CR H receptors stimulate cyclic AMP formation by interacting with G, and by affecting a Ca2+-inhibitable form of adenylyl cyclase.