Muscarinic binding sites on bovine pulmonary arterial endothelial cells in culture

Pharmacology. 1992;44(6):324-33. doi: 10.1159/000138937.

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Allosteric Site
  • Animals
  • Binding Sites
  • Cattle
  • Cells, Cultured
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / metabolism*
  • Muscarinic Antagonists
  • N-Methylscopolamine
  • Oxotremorine / metabolism
  • Parasympatholytics / metabolism*
  • Parasympathomimetics / metabolism*
  • Parasympathomimetics / pharmacology
  • Pulmonary Artery / cytology
  • Pulmonary Artery / metabolism*
  • Quinuclidinyl Benzilate / metabolism
  • Radioligand Assay
  • Receptors, Muscarinic / metabolism*
  • Scopolamine Derivatives / metabolism*

Substances

  • Muscarinic Antagonists
  • Parasympatholytics
  • Parasympathomimetics
  • Receptors, Muscarinic
  • Scopolamine Derivatives
  • Oxotremorine
  • Quinuclidinyl Benzilate
  • N-Methylscopolamine