title: Biochemical interactions of carbamates and ecothiophate with the activated conformation of nicotinic acetylcholine receptor.
abstract: Purified Torpedo nobiliana electric organ acetylcholine receptor (AChR) was reconstituted into membranes containing natural phospholipids supplemented with cholesterol (25% w/w). The reconstituted system facilitates the study of the effects of drugs on the regulation of the AChR channel complex under both resting and carbachol (carb)-stimulated conditions. Neostigmine (Neo) was the only carbamate to induce activation of [3-H]-phencyclidine ([3-H]-PCP) binding to the channel sites, acting as a weak agonist. The activation of [3-H]-PCP binding is dependent upon the nature of the reconstituted systems, with carb/Neo activation ratios of 8, 3, and 1 for the intact purified AChR vesicles fraction (PVF), the PVF reconstituted in phospholipid/cholesterol (CRPVF), and the PVF reconstituted in phospholipid (RPVF), respectively. The carbamates Neo, physostigmine (Physo), and pyridostigmine (Pyrido) inhibited carb-activated [3-H]-PCP binding with Ki values of 10, 20, and 1,600 microM, respectively. The inhibition was mixed competitive-noncompetitive in nature. The characteristic response of CRPVF to carb-stimulated [22-Na] influx was inhibited by the three carbamates, with IC-50 values of 6, 50, and 1,000 microM for Neo, Physo, and Pyrido, respectively. The quaternary ammonium organophosphate ecothiophate (Eco) inhibited carb-stimulated [22-Na] influx with potency similar to that of Neo. Preincubation of AChR preparation with the carbamates and ecothiophate caused a reduction in the binding of [125-I]-alpha-bungarotoxin ([125-I]-alpha-BGT) with the following decreasing order of potency: Neo less than Physo less than Eco less than Pyrido. Calcium has a direct modulatory role on the time-course inhibition of [125-I]-alpha-BGT binding by these drugs. While we observed a high potency of Neo and Physo in inhibiting [125-I]-alpha-BGT binding, it was undetectable for the carbamate insecticide 2-methyl-2-(methylthio)propionaldehyde-O-(methylcarbamoyl)oxime (aldicarb). These data suggest that the potent anticholinesterase carbamate agents interact differently with the AChR and its ionic channel. Their interactions with the nicotinic AChR channel system can be described as (a) weakly agonist, (b) directly acting on the open conformation of the channel, and (c) blocking the AChR-binding sites.
mesh_terms: D001711:Biotransformation; D002038:Bungarotoxins / Q000378:metabolism; D002219:Carbamates / Q000494:pharmacology*; D004456:Echothiophate Iodide / Q000494:pharmacology; D009388:Neostigmine / Q000494:pharmacology; D010622:Phencyclidine / Q000378:metabolism; D010830:Physostigmine / Q000494:pharmacology; D011729:Pyridostigmine Bromide / Q000494:pharmacology; D011978:Receptors, Nicotinic / Q000187:drug effects / Q000378:metabolism*
chemical_list: D002038:Bungarotoxins; D002219:Carbamates; D011978:Receptors, Nicotinic; D009388:Neostigmine; D010830:Physostigmine; D004456:Echothiophate Iodide; D010622:Phencyclidine; D011729:Pyridostigmine Bromide