The role of muscarinic receptor subtypes in acetylcholine release from urinary bladder obtained from muscarinic receptor knockout mouse

We investigated the subtype of prejunctional muscarinic receptors associated with inhibition of acetylcholine (ACh) released from the mouse bladder. We measured endogenous ACh release in the bladder obtained from the wild-type mice and muscarinic 1–5 (M1–M5) receptor knockout (KO) mice. Electrical field stimulation increased ACh release in all bladder preparations obtained from wild-type and M1–M5 receptor KO mice. The amount of ACh released from M1–M3 and M5 receptor KO mice was equal to that in the wild-type mice. In contrast, the amount of electrical field stimulation–induced ACh release in M4 receptor KO mice was significantly larger than that in the wild-type mice, but the extent of increase was small. Atropine increased electrical field stimulation–induced ACh release to levels found in wild-type mice in all M1–M5 receptor KO mice. In M2/M4 receptor double KO mice, the amount of electrical field stimulation–induced ACh release was equivalent to that in the M4 receptor KO mice. The cholinergic component of electrical field stimulation–induced contraction (in the presence of α,β-methylene ATP) in the detrusor of M4 receptor KO mice was no different from that in the detrusor of wild-type mice. M4 receptor immunoreactivity was located between smooth muscle cells, colocalized with choline acetyltransferase immunoreactivity. These results indicate that the prejunctional inhibitory muscarinic receptors are of the M4 and non-M2 receptor subtypes. The nature of the non-M2 receptors remains unknown.