Archives of Pharmacal Research

, Volume 17, Issue 6, pp 443–451 | Cite as

Selectivity of oxomemazine for the M1 muscarinic receptors

  • Shin Woong Lee
  • Chang Woo Woo
  • Jeung Gu Kim
Research Articles


The binding characteristics of pirenzepine and oxomemazine to muscarinic receptor were studied to evaluate the selectivity of oxomemazine for the muscarinic receptor subtypes in rat cerebral microsomes. Equilibrium dissociation constant (KD) of (−)-[3H]quinuclidinyl benzilate([3H]QNB) determined from saturation isotherms was 64 pM. Analysis of the pirenzepine inhibition curve of [3H]QNB binding to cerebral microsome indicated the presence of two receptor subtypes with high (Ki=16 nM, M1 receptor) and low (Ki=400 nM, M3 receptor) affinity for pirenzepine. Oxomemazine also identified two receptor subtypes with about 20-fold difference in the affinity for high (Ki=84nM, OH receptor) and low (Ki=1.65 μM, OL receptor) affinity sites. The percentage populations of M1 and M3 receptors to the total receptors were 61∶39, and those of OH and OL receptors 39∶61, respectively. Both pirenzepine and oxomemazine increased the KD value for [3H]QNB without affecting the binding site concentrations and Hill coefficient for the [3H]QNB binding, Oxomemazine had a 10-fold higher affinity at M1 receptors than at M3 receptors, and pirenzepine a 8-fold higher affinity at OH receptors than at OL receptors. Analysis of the shallow competition binding curves of oxomemazine for M1 receptors and pirenzepine for OL receptors yielded that 69% of M1 receptors were of OH receptors and the remaining 31% of OL receptors, and that 29% of OL receptors were of M1 receptors and 71% of M3 receptors. However, M3 for oxomemazine and OH for pirenzepine were composed of a uniform population. These results suggest that oxomemazine could be classified as a selective drug for M1 receptors and also demonstrate that rat cerebral microsomes contain three different subtypes of M1, M3 and the other site which is different from M1, M2 and M3 receptors.

Key words

Muscarinic receptor subtype Oxomemazine Pirenzepine Receptor binding Rat cerebrum 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References Cited

  1. Ambre, J. J., Bennett, D. R., Cranston, J. W., Dickinson, B. D., Evans, R. M., Glade, M. J., Kosman, M. E., Lampe, K. F., McCann, M. A., Pang, D. C., Proudfit, C. M., Rapoza, N. P., Ratko, T. A., Seidenfeld, J., Smith, S. J. and Weaver, R., Drug evaluations annual 1991.American Medical Association, pp. 1601–1604 (1991).Google Scholar
  2. Barlow, R. B., Berry, K. J., Glenton, P. A. M., Nikolaou, N. M. and Soh, K. S., A comparison of affinity constants for muscarinic-sensitive acetylcholine receptors in guinea-pig atrial pacemaker cells at 29°C. and ileum at 37°C.Br. J. Pharmacol., 58, 613–620 (1976).PubMedGoogle Scholar
  3. Barlow, R. B., Burston, K. N. and Vis, A., Three types of muscarinic receptors?Br. J. Pharmacol., 68, 141–142 (1980).Google Scholar
  4. Berrie, C. P., Birdsall, N. J. M., Burgen, A. S. V. and Hulme, E. C., The binding of muscarinic receptors in the lacrimal gland: Comparision with the cerebral cortex and myocardium.Br. J. Pharmacol., 78, 66P (1983).Google Scholar
  5. Birdsall, N. J. M. and Hulme, E. C., Biochemical studies on muscarinic acetylcholine receptors.J. Neurochem., 27, 7–16 (1976).PubMedCrossRefGoogle Scholar
  6. Bolden, C., Cusack, B. and Richelson, E., Clozapine is a potent and selective muscarinic antagonist at the five cloned human muscarinic cholinergic receptors expressed in CHO-K1 cells.Eur. J. Pharmacol., 192, 205–206 (1991).PubMedCrossRefGoogle Scholar
  7. Bolden, C., Cusack, B. and Richelson, E., Antagonism by antimuscarinic and neuroleptic compounds at the five cloned human muscarinic cholinergic receptors expressed in Chinese hamster ovary cells.J. Pharmacol. Exp. Ther., 260, 576–580 (1992).PubMedGoogle Scholar
  8. Bonner, T. I., The molecular basis of muscarinic receptor density.Trends Neurol. Sci., 12, 148–151 (1989).CrossRefGoogle Scholar
  9. Bonner, T. I., Buckley, N. J., Young, A. C. and Brann, M. R., Identification of a family of muscarinic acetylcholine receptor genes.Science, 237, 527–532 (1987).PubMedCrossRefGoogle Scholar
  10. Bonner, T. I., Young, A. C., Brann, M. R. and Buckley, N. J., Cloning and expression of the human and rat M5 muscarinic acetylcholine receptor genes.Neuron, 1, 403–410 (1988).PubMedCrossRefGoogle Scholar
  11. Brown, D. A., Forward, A. and Marsh, S., Antagonist discrimination between ganglionic and ileal muscarinic receptors.Br. J. Pharmacol., 71, 362–364 (1980).PubMedGoogle Scholar
  12. Buckley, N. J., Bonner, T. I. and Brann, M. R., Localization of a family of muscarinic receptor mRNAs in rat brain.J. Neurosci., 8, 4646–4652 (1988).PubMedGoogle Scholar
  13. Buckley, N. J., Bonner, T. I., Buckley, C. M. and Brann, M. R., Antagonist binding properties of five cloned muscarinic receptors expressed in CHO-K1 cells.Mol. Pharmacol., 35, 469–476 (1989).PubMedGoogle Scholar
  14. Cheng, Y. C. and Prusoff, W. H., Relationship between the inhibition constant (Ki) and the concentration of inhibitor which causes 50 percent inhibition (IC50) of an enzymatic reaction.Biochem. Pharmacol., 22, 3099–3108 (1973).PubMedCrossRefGoogle Scholar
  15. Dauphin, F. and Hamel, E., Identification of multiple muscarinic binding site subtypes in cat and human cerebral vasculature.J. Pharmacol. Exp. Ther., 260, 660–667 (1992).PubMedGoogle Scholar
  16. Doods, H. N., Mathy, M. J., Davidesko, D., Van, Charldorp, K. J., De, Jonge, A. and Van, Zwieten, P. A., Selectivity of muscarinic antagonists in radioligand and in vivo experiments for the putative M1, M2 and M3 receptors.J. Pharmacol. Exp. Ther., 242, 257–262 (1987).PubMedGoogle Scholar
  17. Delmendo, R. E., Michel, A. D. and Whiting, R. L., Affinity of muscarinic receptor antagonists for three putative muscarinic receptor binding sites.Br. J. Pharmacol., 96, 457–464 (1989).PubMedGoogle Scholar
  18. Dörje, F., Wess, J., Lambrecht, G., Tacke, R., Mutschler, E. and Brann, M. R., Antagonist binding profiles of five cloned human muscarinic receptor subtypes.J. Pharmacol. Exp. Ther., 256, 727–733 (1991).PubMedGoogle Scholar
  19. El-Fakahany, E. E., Cioffi, C. L., Abdellatif, M. M. and Miller, M. M., Competitive interaction of pirenzepine with rat brain muscarinic acetylcholine receptors.Eur. J. Pharmacol., 131, 237–247 (1986).PubMedCrossRefGoogle Scholar
  20. Giachetti, A., Micheletti, R. and Montagna, E., Cardioselective profile of AF-DX 116, a muscarinic M2 receptor antagonist.Life Sci., 38, 1663–1672 (1986).PubMedCrossRefGoogle Scholar
  21. Giraldo, E., Hammer, R. and Ladinsky, H., Distribution of muscarinic receptor subtypes in rat brain as determined in binding studies with AF-DX 116 and pirenzepine.Life Sci., 40, 833–840 (1987).PubMedCrossRefGoogle Scholar
  22. Goyal, R. K. and Rattan, S., Neurohumoral, hormonal, and drug receptors for the lower esophageal sphincter.Gastroenterology, 74, 598–618 (1978).PubMedGoogle Scholar
  23. Hammer, R., Muscarinic receptors in the stomach.Scand. J. Gastroenterol., 15 (Suppl. 66), 5–11 (1980).Google Scholar
  24. Hammer, R., Berrie, C. P., Birdsall, N. J. M., Burgen, A. S. V. and Hulme, E. C., Pirenzepine distinguishes between different subclasses of muscarinic receptors.Nature (Lond.), 283, 90–92 (1980).CrossRefGoogle Scholar
  25. Hammer, R. and Giachetti, A., Muscarinic receptor subtypes: M1 and M2 biochemical and functional characterization.Life. Sci., 31, 2991–2998 (1982).PubMedCrossRefGoogle Scholar
  26. Hammer, R. and Giachetti, A., Selective muscarinic receptor antagonists.Trends Pharmacol. Sci., 5, 18–20 (1984).CrossRefGoogle Scholar
  27. Hammer, R., Giraldo, E., Schiavi, G. B., Monferini, E. and Ladinsky, H., Binding profile of a novel cardioselective muscarinic receptor antagonist, AF-DX 116, to membrane of peripheral tissues and brain in the rat.Life Sci., 38, 1653–1662 (1986).PubMedCrossRefGoogle Scholar
  28. Hirschowitz, B. I. R., Hammer, R., Giachetti, A., Kierns, J. J. and Levine, R. R., Subtypes of muscarinic receptors.Trends Pharmacol. Sci., 5 (suppl.), pp. VII (1984).Google Scholar
  29. Kloog, Y., Egozi, Y. and Sokolovskey, M., Characterization of muscarinic acetylcholine receptors from mouse brain: evidence for regional heterogeneity and isomerization.Mol. Pharmacol., 15, 545–548 (1979).PubMedGoogle Scholar
  30. Lazareno, S. and Roberts, F. F., Functional and binding studies with muscarinic M2 subtype selective antagonists.Br. J. Pharmacol., 98, 309–317 (1989).PubMedGoogle Scholar
  31. Lee, S. W., Park, Y. J., Park, I. S. and Lee, J. S., Interaction of antihistaminics with muscarinic receptor (II): Action on the cerebral muscarinic M1 receptor.Yakhak Hoeji, 34(4), 224–237 (1990).Google Scholar
  32. Lee, S. W. and Kim, J. G., Binding profiles of oxomemazine to the muscarinic receptor subtypes.Korean J. Pharmacol., 30, 49–57 (1994).Google Scholar
  33. Lowry, O. H., Rosebrough, N. J., Farr, A. L. and Randall, R. J., Protein measurement with the Folin phenol reagent.J. Biol. Chem., 193, 265–275 (1951).PubMedGoogle Scholar
  34. Luthin, G. R. and Wolfe, B. B., Comparison of [3H] pirenzepine and [3H]quinuclidinylbenzilate binding to muscarinic cholinergic receptors in rat brain.J. Pharmacol. Exp. Ther., 228, 648–655 (1984).PubMedGoogle Scholar
  35. Micheletti, R., Montagna, E. and Giachetti, A., AF-DX 116, a cardioselective muscarinic antagonist.J. Pharmacol. Exp. Ther., 241, 628–634 (1987).PubMedGoogle Scholar
  36. Michel, A. D. and Whiting, R. L., Methoctramine reveals heterogeneity of M2 muscarinic receptors in longitudinal ileal smooth muscle membranes.Eur. J. Pharmacol., 145, 305–311 (1988).PubMedCrossRefGoogle Scholar
  37. Munson, P. J. and Rodbard, D., LIGAND: A versitile computerized approach for characterization of ligand-binding systems.Anal. Biochem., 107, 220–239 (1980).PubMedCrossRefGoogle Scholar
  38. Peralta, E. G., Ashkenazi, A., Winslow, J. W., Smith, D. H., Ramachandran, J. and Caron, D., Distinct primary structures, ligand binding properties and tissue specific expression of four human muscarinic acetylcholine receptors.EMBO J., 6, 3923–3929 (1987).PubMedGoogle Scholar
  39. Scatchard, G., The attractions of proteins for small molecules and ions.Ann. N. Y. Acad. Sci., 51, 660–672 (1949).CrossRefGoogle Scholar
  40. Watson, M., Yamamura, H. I. and Roeske, W. R., A unique regulatory profile and regional distribution of [3H]pirenzepine binding in the rat provide evidence for distinct M1 and M2 muscarinic receptor subtypes.Life Sci., 32, 3001–3011 (1983).PubMedCrossRefGoogle Scholar
  41. Wess, J., Lambrecht, G., Mutschler, E., Brann, M. R. and Dörje, F., Selectivity profile of the novel muscarinic antagonist UH-AH 37 determined by the use of cloned receptors and isolated tissue preparations.Br. J. Pharmacol., 102, 246–250 (1991).PubMedGoogle Scholar

Copyright information

© The Pharmaceutical Society of Korea 1994

Authors and Affiliations

  • Shin Woong Lee
    • 1
  • Chang Woo Woo
    • 1
  • Jeung Gu Kim
    • 1
  1. 1.College of PharmacyYeungnam UniversityGyongsanKorea

Personalised recommendations