Guanine nucleotide-sensitive, high affinity binding of carbachol to muscarinic cholinergic receptors of 1321N1 astrocytoma cells is insensitive to pertussis toxin.
Activation of muscarinic cholinergic receptors of 1321N1 human astrocytoma cells attenuates cyclic AMP accumulation. This effect results from an activation of phosphodiesterase with no direct inhibition of adenylate cyclase activity. In spite of this lack of coupling of muscarinic receptors to adenylate cyclase, guanine nucleotides reduce the apparent binding affinity of the agonist carbachol in a washed membrane preparation of 1321N1 cells. The order of potency for this effect is guanosine 5'-O-(3-thiotriphosphate) greater than 5'-guanylyl-imidodiphosphate = GTP = GDP; ATP has no effect. The occurrence of a Mr = 41,000 protein labeled in the presence of [32P]NAD and pertussis toxin as well as the occurrence of guanine nucleotide-mediated inhibition of forskolin-stimulated adenylate cyclase activity indicate that the functional inhibitory guanine nucleotide regulatory component of adenylate cyclase (Ni) is present in 1321N1 cells. Pertussis toxin pretreatment of NG108-15 neuroblastoma X glioma cells, which express muscarinic receptors that link through Ni to inhibit adenylate cyclase, blocked the GTP-sensitive, high affinity binding of carbachol. In contrast, pretreatment of 1321N1 cells with a concentration of pertussis toxin that blocked [32P]ADP ribosylation of the Mr = 41,000 substrate and GTP-mediated inhibition of forskolin-stimulated adenylate cyclase activity had no effect on GTP-sensitive high affinity binding of carbachol. These results suggest that muscarinic cholinergic receptors of 1321N1 cells couple to a guanine nucleotide regulatory protein that is distinct from Ni.
Evans
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Martin
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Hughes
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Harden
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D037361:Adenylate Cyclase Toxin; D000262:Adenylyl Cyclases / Q000378:metabolism; D001254:Astrocytoma / Q000378:metabolism*; D001427:Bacterial Toxins / Q000494:pharmacology*; D002217:Carbachol / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D006150:Guanine Nucleotides / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D037342:Pertussis Toxin; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000378:metabolism*; D013329:Structure-Activity Relationship; D010566:Virulence Factors, Bordetella
D037361:Adenylate Cyclase Toxin; D001427:Bacterial Toxins; D006150:Guanine Nucleotides; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D002217:Carbachol; D000242:Cyclic AMP; D037342:Pertussis Toxin; D000262:Adenylyl Cyclases