TREATMENT OF INTACT STRIATAL NEURONS WITH CHOLERA-TOXIN OR 8-BROMOADENOSINE 3',5'-(CYCLIC)PHOSPHATE DECREASES THE ABILITY OF PERTUSSIS TOXIN TO ADP-RIBOSYLATE THE ALPHA-SUBUNITS OF INHIBITORY AND OTHER GUANINE-NUCLEOTIDE-BINDING REGULATORY PROTEINS, GI AND GO - EVIDENCE FOR 2 DISTINCT MECHANISMS

Using primary cultures of striatal neurones from the mouse embryo, we showed that treatment of intact cells with cholera toxin (5-mu-g/ml, 22 h) decreases the subsequent ADP-ribosylation of the alpha subunit of the guanine-nucleotide-binding regulatory protein G(o) (G(o)alpha) and the alpha subunit of the inhibitory guanine-nucleotide-binding regulatory protein (G(i)alpha) of adenylate cyclase, which is catalyzed in vitro on neuronal membranes by pertussis toxin. The inhibitory effect of cholera toxin could not only be attributed to an increased production of cAMP in neurones. Treatment of cells with 0.1-mu-M 8-bromoadenosine 3',5'-(cyclic)phosphate (Br8cAMP) for 16 h, or with 0.1 mM Br8cAMP for 5 min, mimicked the effect of cholera toxin on the ADP-ribosylation of G(o)alpha and G(i)alpha in vitro. However, the two agents seem to act through distinct mechanisms. The protein kinase inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine prevented the action of Br8cAMP but not that of cholera toxin. In addition, measurements of the pI of the G(o)alpha, deduced from immunoblots of two-dimensional gels performed using a specific antibody directed against G(o)alpha, suggest that treatment of neurones with cholera toxin induces ADP-ribosylation of G(o)alpha in intact cells, while Br8cAMP does not.