The presence of an M₄ subtype muscarinic receptor in the bovine adrenal medulla revealed by mRNA and receptor binding analyses

The muscarinic receptor subtype present in the bovine adrenal medulla was characterized. Hybridization of RNA to highly specific m₁-m₅ muscarinic receptor cDNA probes detected the presence of only m₄ subtype mRNA in this tissue. Muscarinic receptor binding studies using the non-selective ligand [³H]N-methyl-scopolamine showed a single class of binding sites with a maximum density of 19.8 fmol/mg protein and a dissociation constant (K_D) of 220 pM in the adrenal medulla, while the M₁ selective ligand [³H]telenzepine did not bind detectably. Competition of specific antagonists with [³H]N-methyl-scopolamine for binding to the membranes produced a rank order of potencies with a profile that fitted either the cloned m₃ or m₄ receptor. In further comparative studies, the adrenal gland of the rat showed the presence of m₄ subtype mRNA in addition to the m₃ subtype.