Immunological localization of m1-m5 muscarinic acetylcholine receptors in peripheral tissues and brain.

Knowledge of the distributions and functions of native m1-m5 muscarinic acetylcholine receptors in tissues is limited. To characterize the family of m1-m5 proteins directly, a panel of subtype-selective antibodies was generated against divergent i3 loop-fusion proteins. Each antibody was shown to bind a single cloned receptor specifically. In peripheral tissues and brain, four receptor proteins (m1-m4) were found to account for the vast majority of the muscarinic binding sites using immunoprecipitation studies with the subtype-specific antibodies. The subtypes were differentially distributed, although most tissues were comprised of a complex mixture of receptors. Moreover, within tissues there were major differences in the precise localization of the subtypes, as determined by immunocytochemistry. The immunological methods described offer a novel approach with exquisite sensitivity and specificity for delineating the distribution of m1-m5 receptors in animal and human tissues.