title: Reaction of quinacrine mustard with the acetylcholine receptor from Torpedo californica.
abstract: Amines with local anesthetic activity are typically also noncompetitive inhibitors of the agonist-induced increase in cation permeability mediated by the nicotinic acetylcholine receptor. Quinacrine is such an agent, and we have synthesized tritiated quinacrine mustard, a derivative capable of reacting with nucleophiles. Quinacrine mustard was reacted with receptor-rich membrane from torpedo electric tissue, excess reagent was removed by partition into liposomes, and the modified receptor was extracted and reconstituted with exogenous phospholipid. After reaction of the native membrane with 10 microM quinacrine mustard for 5 min, binding of cobratoxin to the acetylcholine binding sites is inhibited 15%; in contrast, receptor-mediated 86Rb uptake in the reconstituted vesicles is inhibited 70%. When the reaction with quinacrine mustard is carried out in the presence of 10 microM carbamylcholine or 10 microM d-tubocurarine, there is no block of the acetylcholine binding sites; nevertheless, the inhibition of Rb uptake is greater than that resulting from reaction in the absence of acetylcholine binding site ligands. Conversely, when the reaction is carried out in the presence of either 100 microM quinacrine or 100 microM proadifen (also a potent noncompetitive inhibitor), either with or without carbamylcholine or d-tubocurarine, the inhibition of 86Rb uptake is about 70% smaller. Under the same conditions that we used in the functional studies, quinacrine mustard reacts with the four types of chains that constitute the receptor complex, alpha 2 beta gamma delta. The presence of the acetylcholine binding site ligands, however, results in increased reaction with the alpha and beta chains, while the presence of the noncompetitive inhibitors, with or without the acetylcholine binding site ligands, results in decreased reaction with the alpha and beta chains. We conclude that the alpha and beta chains contribute to one or more functionally significant binding sites for noncompetitively inhibiting amines.
mesh_terms: D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects; D003039:Cobra Neurotoxin Proteins / Q000378:metabolism; D004557:Electric Organ / Q000032:analysis*; D007473:Ion Channels / Q000187:drug effects; D011335:Proadifen / Q000494:pharmacology; D011796:Quinacrine / Q000031:analogs & derivatives* / Q000494:pharmacology; D011797:Quinacrine Mustard / Q000494:pharmacology*; D011868:Radioisotopes; D011950:Receptors, Cholinergic / Q000187:drug effects* / Q000378:metabolism; D012413:Rubidium / Q000378:metabolism; D014101:Torpedo; D014316:Tritium; D014403:Tubocurarine / Q000494:pharmacology
chemical_list: D003039:Cobra Neurotoxin Proteins; D007473:Ion Channels; D011868:Radioisotopes; D011950:Receptors, Cholinergic; D014316:Tritium; D011797:Quinacrine Mustard; D002217:Carbachol; D011335:Proadifen; D011796:Quinacrine; D012413:Rubidium; D014403:Tubocurarine