The muscarinic antagonists aprophen and benactyzine are noncompetitive inhibitors of the nicotinic acetylcholine receptor.

Certain muscarinic antagonists (e.g., atropine, aprophen, and benactyzine) are used as antidotes for the treatment of organophosphate poisoning. We have studied the interaction of aprophen and benactyzine, both aromatic esters of diethylaminoethanol, with nicotinic acetylcholine receptor (AChR) in BC3H-1 intact muscle cells and with receptor-enriched membranes of Torpedo californica. Aprophen and benactyzine diminish the maximal carbamylcholine-elicited sodium influx into muscle cells without shifting Kact (carbamylcholine concentration eliciting 50% of the maximal 22Na+ influx). The concentration dependence for the inhibition of the initial rate of 22Na+ influx by aprophen and benactyzine occurs at lower concentrations (Kant = 3 and 50 microM, respectively) than those needed to inhibit the initial rate of [125I]-alpha-bungarotoxin binding to the agonist/antagonist sites of the AChR (Kp = 83 and 800 microM, respectively). The effective concentration for atropine inhibition of AChR response (Kant = 150 microM in BC3H-1 cells) is significantly higher than those obtained for aprophen and benactyzine. Both aprophen and benactyzine interact with the AChR in its desensitized state in BC3H-1 cells without further enhancing agonist affinity. Furthermore, these ligands do not alter the value of Kdes (equilibrium concentration of agonist which diminishes 50% of the maximal receptor response) in BC3H-1 muscle cells. The affinity of aprophen and benactyzine for the allosterically coupled noncompetitive inhibitor site of the AChR in Torpedo was determined using [3H]phencyclidine as a probe. Both compounds were found to preferentially associate with the high affinity (desensitized) state rather than the resting state of Torpedo AChR. There is a 14- to 23-fold increase in the affinity of aprophen and benactyzine for the AChR (KD = 0.7 and 28.0 microM in the desensitized state compared to 16.4 and 384 microM in the resting state, respectively). These data indicate that aprophen and benactyzine binding are allosterically regulated by the agonist sites of Torpedo AChR. Thus, aprophen and benactyzine are effective noncompetitive inhibitors of the AChR at concentrations of 1-50 microM, in either Torpedo or mammalian AChR. These concentrations correspond very well with the blood level of these drugs found in vivo to produce a therapeutic response against organophosphate poisoning.