title: Muscarinic binding sites on bovine pulmonary arterial endothelial cells in culture.
abstract: We have investigated the presence and nature of muscarinic binding sites on membranes from cultured bovine pulmonary arterial endothelial cells (BPAE). BPAE were harvested and subcultured nonenzymatically; experiments were performed 3-5 days postconfluence and between 10 and 25 passage numbers. Utilizing radioligand binding techniques with the muscarinic receptor antagonists [3H]3-quinuclidinyl benzilate ([3H]QNB) and [3H]N-methylscopolamine ([3H]MS) as probes, we identified a small population of atropine-sensitive muscarinic sites (1,800-2,000 sites/cell or 7-8 fmol/mg protein). Muscarinic binding sites on BPAE membranes resembled classical muscarinic receptors in that (a) the binding of 2 nM [3H]QNB was inhibited by muscarinic agonists and antagonists, (b) [3H]QNB binding was 30 times more sensitive to R(-)- than to S(+)-QNB, (c) binding of the muscarinic receptor agonist carbamylcholine involved high and low affinity components, (d) the stable GTP analog, Gpp(NH)p (100 microM) shifted agonist binding curves to the right by a factor of three, and (e) the high affinity binding of the agonist [3H]oxotremorine-M to muscarinic receptors was depressed by Gpp(NH)p. On the other hand, gallamine, which allosterically regulates muscarinic receptor binding in other tissues, did not affect the rates of dissociation of [3H]QNB, [3H]MS or [3H]oxotremorine-M from BPAE binding sites. We concluded that BPAE in culture exhibit muscarinic binding sites which possess many but not all of the properties associated with classical muscarinic receptors.
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta 30912-2300.
authors.forename: R S
authors.initials: RS
authors.lastname: Aronstam
authors.forename: U S
authors.initials: US
authors.lastname: Ryan
authors.forename: J D
authors.initials: JD
authors.lastname: Catravas
mesh_terms: D000495:Allosteric Site; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D018727:Muscarinic Antagonists; D019832:N-Methylscopolamine; D010095:Oxotremorine / Q000378:metabolism; D010276:Parasympatholytics / Q000378:metabolism*; D010277:Parasympathomimetics / Q000378:metabolism* / Q000494:pharmacology; D011651:Pulmonary Artery / Q000166:cytology / Q000378:metabolism*; D011813:Quinuclidinyl Benzilate / Q000378:metabolism; D011869:Radioligand Assay; D011976:Receptors, Muscarinic / Q000378:metabolism*; D012602:Scopolamine Derivatives / Q000378:metabolism*
chemical_list: D018727:Muscarinic Antagonists; D010276:Parasympatholytics; D010277:Parasympathomimetics; D011976:Receptors, Muscarinic; D012602:Scopolamine Derivatives; D010095:Oxotremorine; D011813:Quinuclidinyl Benzilate; D019832:N-Methylscopolamine