MUSCARINIC RECEPTOR SUBTYPES IN HUMAN NASAL-MUCOSA - CHARACTERIZATION, AUTORADIOGRAPHIC LOCALIZATION, AND FUNCTION-INVITRO

Muscarinic receptors play important roles in the regulation of glandular secretion and vasomotor tone in human nasal mucosa. M1, M2, and M3 muscarinic receptor subtypes were pharmacologically characterized in human inferior turbinates by receptor-binding assays using [H-3](-)quinuclidinyl benzilate (QNB, identifies total muscarinic receptors) and [H-3]-pirenzepine (PZ). Receptors were localized by autoradiography, and their function examined in vitro by assaying mucus secretion from cultured nasal mucosal explants. In competition assays, PZ was employed as a selective muscarinic antagonist for M1 receptors, gallamine and AF-DX 116 for M2 receptors, and 4-DAMP for M3 receptors. These ligands are selective at low nanomolar concentrations, but can interact with other muscarinic receptors at higher concentrations. It is not known if they can interact with putative M4 and M5 muscarinic receptor subtypes. Using [H-3](-)QNB, total muscarinic receptor binding was 688.4 +/- 49.6 fmol/mg protein (B(max)), with a K(d) of 1.47 +/- 0.13 nM. [H-3]-PZ bound to 45 % of the total QNB binding sites. In competition experiments, 4-DAMP displaced [H-3](-)QNB with the lowest IC50, followed by PZ and AF-DX 116. Autoradiograms demonstrated that [H-3](-)QNB binding was completely displaced by 4-DAMP, partially displaced by PZ, but not displaced by gallamine or AF-DX 116, and suggested that M1 and M3 subtypes coexist in submucosal glands. The localization of M1 receptors on submucosal glands was confirmed by direct labeling with [H-3]-PZ. [H-3]-PZ also labeled vessels, but with a low silver grain density. Autoradiographic [H-3]-QNB binding was displaced by 4-DAMP and atropine, but not by PZ, gallamine, or AF-DX 116. In studies of mucus secretion in vitro, 4-DAMP significantly inhibited methacholine-induced secretion. Although less effective, PZ also had significant inhibitory effects. Neither gallamine nor AF-DX 116 had any inhibitory effect. M1 receptors (PZ binding sites) may regulate glandular secretion while M3 receptors (4-DAMP binding sites) may regulate glandular secretion and vasomotor tone in human nasal mucosa.