Modulation of central muscarinic receptor binding in vitro by ultralow levels of the organophosphate paraoxon

Homogenates of calf caudate nuclei were found to contain at least three distinct subclasses of cholinergic, muscarinic receptors. These subtypes, labeled with 3H]quinuclidinyl benzilate (QNB), can be separated by rapid filtration with the use of the selective ligands, pirenzepine, AF-DX116, and 4-DAMP which have high affinity for the M1, M2, and M3 subtypes, respectively. Paraoxon was found to modulate 3H]QNB binding in a noncompetitive manner at concentrations below those needed to affect acetylcholinesterase activity. Pretreatment of the membrane protein with high concentrations of both the M2 selective antagonist, AF-DX116, and the M3 selective antagonist, 4-DAMP, protected against paraoxon inhibition of 3H]QNB binding, while the M1 selective antagonist pirenzepine did not. Paraoxon sensitive sites, M2 and M3, are found predominantly on presynaptic neurons in the central nervous system. It is postulated that blockade of these sites may interfere with negative feedback inhibition of acetylcholine release and facilitate the development of behavioral and motor deficits that may be associated with chronic exposure to low levels of organophosphates.