Effects of pertussis toxin on cAMP and cGMP responses to carbamylcholine in N1E-115 neuroblastoma cells.
As noted previously, in N1E-115 neuroblastoma cells, carbamylcholine, a muscarinic cholinergic agonist, increased cGMP over 15-fold and decreased basal and prostaglandin E1 (PGE1)-stimulated cAMP content. In contrast to the stimulatory effects of PGE1 on cAMP, which were immediate, the carbamylcholine-induced decrease in basal and PGE1-stimulated cAMP exhibited a delay. The delay in carbamylcholine inhibition was independent of the extent of adenylate cyclase activation. Although basal cAMP content was suppressed within 30 sec after addition of carbamylcholine, inhibition was not maximal for at least 2 min following agonist addition; the delay was similar in cells exposed to PGE1 for 10 min prior to carbamylcholine but could be eliminated by incubation of the cells with muscarinic cholinergic agonist for 5 min prior to addition of prostaglandin. N1E-115 neuroblastoma cells possess a 41,000-Da membrane protein believed to be a component of the inhibitory GTP-binding protein of adenylate cyclase that is ADP ribosylated by pertussis toxin. Incubation of the cells with pertussis toxin prior to the addition of carbamylcholine reduced the maximal extent of inhibition of cAMP content and prevented the [32P]ADP-ribosylation of a 41,000-Da protein by toxin and [32P]NAD in membrane preparations from these cells. Incubation of cells with pertussis toxin, however, did not significantly alter the dose-response curve for carbamylcholine effects on cGMP. Even high concentrations of carbamylcholine, effective in stimulating cGMP, had minimal effects on cAMP content in toxin-treated cells; thus, ADP-ribosylation of Gi converts the adenylate cyclase but not the guanylate cyclase system to an agonist-insensitive state.
Bruni
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Burns
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Hewlett
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Moss
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D037361:Adenylate Cyclase Toxin; D000527:Alprostadil; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D002217:Carbachol / Q000494:pharmacology*; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000502:physiology*; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism; D037342:Pertussis Toxin; D011458:Prostaglandins E / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000502:physiology*; D010566:Virulence Factors, Bordetella
D037361:Adenylate Cyclase Toxin; D001427:Bacterial Toxins; D011458:Prostaglandins E; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D002217:Carbachol; D000242:Cyclic AMP; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D000527:Alprostadil; D006152:Cyclic GMP