Binding of [3H]quinuclidinyl benzilate to intestinal mucus. An artifact in identification of epithelial cell muscarinic receptors.
The widely used muscarinic receptor ligand [3H]quinuclidinyl benzilate ([3H]QNB) was found to bind in a site-specific but artifactual manner to rat intestinal mucus, obscuring specific binding to muscarinic receptors on intestinal epithelial cells. Atropine inhibited [3H]QNB binding to mucus with an apparent IC50 of 2.1 x 10(-7) M, compared to an IC50 of 1.4 x 10(-8) M obtained with a homogenate of intestinal epithelial cells. Unlabeled QNB also inhibited binding of [3H]QNB to mucus but the apparent IC50 (4 x 10(-7) M) was about 300-fold greater than the IC50 determined with a control tissue, heart muscle (IC50, 1.2 x 10(-9) M). [3H]QNB binding was saturable over the concentration range of 1-7 nM in the heart, with an apparent KD of 0.76 nM. As expected from the high IC50 for QNB in the mucus binding experiments, binding to mucus was not saturable over the 1-15 nM concentration range. Based on pH profiles and temperature dependency of binding, it seems unlikely that mucin, the primary component of mucus, was responsible for [3H]QNB binding to the mucus. The findings have implications for studies which involve binding of [3H]QNB in particular and other ligands in general to mucus-secreting epithelial tissues.
D000818:Animals; D001665:Binding Sites; D004848:Epithelium / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000378:metabolism*; D008297:Male; D009077:Mucins / Q000378:metabolism; D009093:Mucus / Q000378:metabolism*; D011485:Protein Binding; D011812:Quinuclidines / Q000378:metabolism*; D011813:Quinuclidinyl Benzilate / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D011950:Receptors, Cholinergic / Q000302:isolation & purification*; D011976:Receptors, Muscarinic / Q000302:isolation & purification*; D013696:Temperature
D009077:Mucins; D011812:Quinuclidines; D011950:Receptors, Cholinergic; D011976:Receptors, Muscarinic; D011813:Quinuclidinyl Benzilate