SUBTYPE-SELECTIVE POSITIVE COOPERATIVE INTERACTIONS BETWEEN BRUCINE ANALOGS AND ACETYLCHOLINE AT MUSCARINIC RECEPTORS - RADIOLIGAND BINDING-STUDIES

We studied the interactions of strychnine, brucine, and three of the N -substituted analogues of brucine with [H-3]N-methylscopolamine (NMS) and unlabeled acetylcholine at m1-m5 muscarinic receptors using equili brium and nonequilibrium radioligand binding studies. The results were consistent with a ternary allosteric model in which both the primary and allosteric ligands bind simultaneously to the receptor and modify the affinities of each other. The compounds had K-d values in the subm illimolar range, inhibited [H-3]NMS dissociation, and showed various p atterns of positive, neutral, and negative cooperativity with [H-3]NMS and acetylcholine, but there was no predictive relationship between t he effects. Acetylcholine affinity was increased similar to 2-fold by brucine at m1 receptors, similar to 3-fold by N-chloromethyl brucine a t m3 receptors, and similar to 1.5-fold by brucine-N-oxide at m4 recep tors. The existence of neutral cooperativity, in which the compound bo und to the receptor but did not modify the affinity of acetylcholine, provides the opportunity for a novel form of drug selectivity that we refer to as absolute subtype selectivity: an agent showing positive or negative cooperativity with the endogenous ligand at one receptor sub type and neutral cooperativity at the other subtypes would exert funct ional effects at only the one subtype, regardless of the concentration of agent or its affinities for the subtypes. Our results demonstrate the potential for developing allosteric enhancers of acetylcholine aff inity at individual subtypes of muscarinic receptor and suggest that m inor modification of a compound showing positive, neutral, or low nega tive cooperativity with acetylcholine may yield compounds with various patterns of cooperativity across the receptor subtypes.