Identification of rat brain muscarinic M₄ receptors coupled to cyclic AMP using the selective antagonist muscarinic toxin 3

In membranes of olfactory tubercle and striatum, the selective muscarinic M₄ receptor antagonist muscarinic toxin 3 completely antagonized the acetylcholine-induced inhibition of forskolin- and dopamine D₁ receptor-stimulated cyclic AMP formation with K(i) values of 7 and 4 nM, respectively. In olfactory bulb, where acetylcholine stimulated basal adenylyl cyclase activity and inhibited forskolin-stimulated enzyme activity, muscarinic toxin 3 caused a partial antagonism of both acetylcholine effects with high potencies (K(i) values=4-6 nM). In frontal cortex, muscarinic toxin 3 counteracted the acetylcholine-induced potentiation of corticotropin-releasing hormone-stimulated cyclic AMP with a K(i) of 58 nM, which is close to the toxin affinity for the muscarinic M₁ receptor. In the same brain region, the acetylcholine inhibition of forskolin-stimulated enzyme activity was not affected by muscarinic toxin 3. In microdissected regions of the hippocampus, a significant portion (33-48%) of the acetylcholine inhibition of forskolin-stimulated adenylyl cyclase activity was blocked by muscarinic toxin 3 with K(i) values (6-8 nM) consistent with the involvement of muscarinic M₄ receptors. These data show that muscarinic toxin 3 discriminates between adenylyl cyclase-coupled muscarinic receptors and demonstrate the utility of the toxin in identifying the relative contribution by the muscarinic M₄ receptor subtype. Copyright (C) 1998 Elsevier Science B.V.