Different agonist-receptor active conformations for rat brain M1 and M2 muscarinic receptors that are separately coupled to two biochemical effector systems.

In dissociated cellular preparations of adult rat cortex, M1 and M2 muscarinic receptors were shown to mediate phosphoinositide metabolism and cAMP inhibition, respectively. Additionally, in dissociated striatum, an M2 receptor was shown to inhibit the level of cAMP. The components of "receptor reserve" in these three receptor-effector systems were evaluated by the method of partial receptor inactivation and the dissociation constants for the full agonist carbachol were determined. In dissociated cellular preparations of cortex, and in the presence of 10 mM lithium ion, carbachol (EC50 = 116 microM) activated an M1 receptor subtype (atropine Ki = 0.9 nM; pirenzepine Ki = 9.3 nM) to elicit up to a 7-fold release over the basal level of [3H]inositol 1-phosphate. Carbachol was 100-fold more potent (EC50 approximately 1 microM) in the inhibition of forskolin-elevated [3H]cAMP levels in both the cortex (maximally 28%) and striatum (maximally 49%). Pirenzepine blocked the [3H]cAMP inhibition responses to carbachol in cortex and striatum with Ki values of 334 nM and 313 nM, respectively, which indicated that cortical and striatal M2 receptors mediate [3H]cAMP inhibition. The equilibrium dissociation constants for the full agonist carbachol in mediating these two biochemical responses were determined after partial receptor inactivation with propylbenzilylcholine mustard. The results indicate that cortical M1 receptor-mediated phosphoinositide metabolism is elicited by carbachol through a low affinity agonist-receptor complex (carbachol Kd = 90 microM). However, the cortical and striatal M2 receptor-mediated inhibition of [3H]cAMP is mediated by a high affinity agonist-receptor complex (carbachol Kd = 8.5 microM and 2.9 microM, respectively). Thus, the agonist is bound in a low affinity active conformation of the M1 receptor but the agonist is bound in a high affinity active conformation of the M2 receptor. In contrast to the cortical M1-phosphoinositide system, the central M2 receptors exhibited a significant receptor reserve in their mediation of [3H]cAMP inhibition, as elicited by the full agonist carbachol. Whereas the ratio of Kd/EC50 for carbachol was 0.9 at the cortical M1 receptor, this ratio was 9.4 and 3.2 at the cortical M2 and striatal M2 receptors, respectively.