Skip to main content
PMC home page
British Journal of Pharmacology logoLink to British Journal of Pharmacology
. 1992 Aug;106(4):1019–1027. doi: 10.1111/j.1476-5381.1992.tb14450.x

Characterization and autoradiographical localization of non-adrenoceptor idazoxan binding sites in the rat brain.

N J Mallard 1, A L Hudson 1, D J Nutt 1
PMCID: PMC1907664  PMID: 1356565

Abstract

1. In rat whole brain homogenates, saturation analysis revealed that both [3H]-idazoxan and [3H]-RX821002, a selective alpha 2-adrenoceptor ligand, bound with high affinity to an apparent single population of sites. However, the Bmax for [3H]-idazoxan was significantly (P less than 0.01) greater than that for [3H]-RX821002. 2. In competition studies, (-)-adrenaline displaced 3 nM [3H]-idazoxan binding with an affinity consistent with [3H]-idazoxan labelling alpha 2-adrenoceptors. However, this displacement was incomplete since 23.68 +/- 1.11% of specific [3H]-idazoxan binding remained in the presence of an excess concentration (100 microM) of (-)-adrenaline. In contrast, unlabelled idazoxan promoted a complete displacement of [3H]-idazoxan binding with a Hill slope close to unity and an affinity comparable with its KD determined in saturation studies. 3. Displacement of [3H]-idazoxan binding by the alpha 2-adrenoceptor antagonists yohimbine, RX821002 (2-(2-methoxy-1,4-benzodioxan-2-yl)-2-imidazoline) and RX811059 (2-(2-ethoxy-1,4-benzodioxan-2-yl)-2-imidazoline) was more complex, with Hill slopes considerably less than unity, and best described by a two-site model of interaction comprising a high and low affinity component. The proportion of sites with high affinity for each antagonist was similar (60-80%). 4. The rank order of antagonist potency for the high affinity component in each displacement curve (RX821002 greater than RX811059 greater than yohimbine) is similar to that determined against the binding of [3H]-RX821002 to rat brain, suggesting that these components reflect the inhibition of [3H]-idazoxan binding to alpha 2-adrenoceptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Full text

PDF
1019

Images in this article

1023Figure 4
on p.1023
1024Figure 5
on p.1024
1025Figure 6
on p.1025

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Benavides J., Quarteronet D., Imbault F., Malgouris C., Uzan A., Renault C., Dubroeucq M. C., Gueremy C., Le Fur G. Labelling of "peripheral-type" benzodiazepine binding sites in the rat brain by using [3H]PK 11195, an isoquinoline carboxamide derivative: kinetic studies and autoradiographic localization. J Neurochem. 1983 Dec;41(6):1744–1750. doi: 10.1111/j.1471-4159.1983.tb00888.x. [DOI] [PubMed] [Google Scholar]
  2. Blundell J. Pharmacological approaches to appetite suppression. Trends Pharmacol Sci. 1991 Apr;12(4):147–157. doi: 10.1016/0165-6147(91)90532-w. [DOI] [PubMed] [Google Scholar]
  3. Boyajian C. L., Loughlin S. E., Leslie F. M. Anatomical evidence for alpha-2 adrenoceptor heterogeneity: differential autoradiographic distributions of [3H]rauwolscine and [3H]idazoxan in rat brain. J Pharmacol Exp Ther. 1987 Jun;241(3):1079–1091. [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Bricca G., Dontenwill M., Molines A., Feldman J., Belcourt A., Bousquet P. The imidazoline preferring receptor: binding studies in bovine, rat and human brainstem. Eur J Pharmacol. 1989 Mar 14;162(1):1–9. doi: 10.1016/0014-2999(89)90597-9. [DOI] [PubMed] [Google Scholar]
  6. Broadhurst A. M., Wyllie M. G. A reassessment of the binding of [3H]rauwolscine to membranes from the rat cortex. Neuropharmacology. 1986 Mar;25(3):287–295. doi: 10.1016/0028-3908(86)90254-6. [DOI] [PubMed] [Google Scholar]
  7. Brown C. M., MacKinnon A. C., McGrath J. C., Spedding M., Kilpatrick A. T. Alpha 2-adrenoceptor subtypes and imidazoline-like binding sites in the rat brain. Br J Pharmacol. 1990 Apr;99(4):803–809. doi: 10.1111/j.1476-5381.1990.tb13010.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Brüning G., Kaulen P., Baumgarten H. G. Quantitative autoradiographic localization of alpha 2-antagonist binding sites in rat brain using [3H]idazoxan. Neurosci Lett. 1987 Dec 29;83(3):333–337. doi: 10.1016/0304-3940(87)90110-8. [DOI] [PubMed] [Google Scholar]
  9. Chapleo C. B., Myers P. L., Butler R. C., Doxey J. C., Roach A. G., Smith C. F. alpha-adrenoreceptor reagents. 1. Synthesis of some 1,4-benzodioxans as selective presynaptic alpha 2-adrenoreceptor antagonists and potential antidepressants. J Med Chem. 1983 Jun;26(6):823–831. doi: 10.1021/jm00360a008. [DOI] [PubMed] [Google Scholar]
  10. Cheng Y., Prusoff W. H. Relationship between the inhibition constant (K1) and the concentration of inhibitor which causes 50 per cent inhibition (I50) of an enzymatic reaction. Biochem Pharmacol. 1973 Dec 1;22(23):3099–3108. doi: 10.1016/0006-2952(73)90196-2. [DOI] [PubMed] [Google Scholar]
  11. Convents A., De Keyser J., De Backer J. P., Vauquelin G. [3H]rauwolscine labels alpha 2-adrenoceptors and 5-HT1A receptors in human cerebral cortex. Eur J Pharmacol. 1989 Jan 17;159(3):307–310. doi: 10.1016/0014-2999(89)90163-5. [DOI] [PubMed] [Google Scholar]
  12. Coupry I., Atlas D., Podevin R. A., Uzielli I., Parini A. Imidazoline-guanidinium receptive site in renal proximal tubule: asymmetric distribution, regulation by cations and interaction with an endogenous clonidine displacing substance. J Pharmacol Exp Ther. 1990 Jan;252(1):293–299. [PubMed] [Google Scholar]
  13. De Vos H., Convents A., De Keyser J., De Backer J. P., Van Megen I. J., Ebinger G., Vauquelin G. Autoradiographic distribution of alpha 2 adrenoceptors, NAIBS, and 5-HT1A receptors in human brain using [3H]idazoxan and [3H]rauwolscine. Brain Res. 1991 Dec 6;566(1-2):13–20. doi: 10.1016/0006-8993(91)91675-q. [DOI] [PubMed] [Google Scholar]
  14. Doxey J. C., Lane A. C., Roach A. G., Virdee N. K. Comparison of the alpha-adrenoceptor antagonist profiles of idazoxan (RX 781094), yohimbine, rauwolscine and corynanthine. Naunyn Schmiedebergs Arch Pharmacol. 1984 Feb;325(2):136–144. doi: 10.1007/BF00506193. [DOI] [PubMed] [Google Scholar]
  15. Doxey J. C., Roach A. G., Smith C. F. Studies on RX 781094: a selective, potent and specific antagonist of alpha 2-adrenoceptors. Br J Pharmacol. 1983 Mar;78(3):489–505. doi: 10.1111/j.1476-5381.1983.tb08809.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ernsberger P., Giuliano R., Willette R. N., Granata A. R., Reis D. J. Hypotensive action of clonidine analogues correlates with binding affinity at imidazole and not alpha-2-adrenergic receptors in the rostral ventrolateral medulla. J Hypertens Suppl. 1988 Dec;6(4):S554–S557. doi: 10.1097/00004872-198812040-00174. [DOI] [PubMed] [Google Scholar]
  17. Ernsberger P., Giuliano R., Willette R. N., Reis D. J. Role of imidazole receptors in the vasodepressor response to clonidine analogs in the rostral ventrolateral medulla. J Pharmacol Exp Ther. 1990 Apr;253(1):408–418. [PubMed] [Google Scholar]
  18. Ernsberger P., Meeley M. P., Mann J. J., Reis D. J. Clonidine binds to imidazole binding sites as well as alpha 2-adrenoceptors in the ventrolateral medulla. Eur J Pharmacol. 1987 Jan 28;134(1):1–13. doi: 10.1016/0014-2999(87)90125-7. [DOI] [PubMed] [Google Scholar]
  19. Ernsberger P., Meeley M. P., Reis D. J. An endogenous substance with clonidine-like properties: selective binding to imidazole sites in the ventrolateral medulla. Brain Res. 1988 Feb 16;441(1-2):309–318. doi: 10.1016/0006-8993(88)91409-6. [DOI] [PubMed] [Google Scholar]
  20. Feldman J., Tibiriça E., Bricca G., Dontenwill M., Belcourt A., Bousquet P. Evidence for the involvement of imidazoline receptors in the central hypotensive effect of rilmenidine in the rabbit. Br J Pharmacol. 1990 Jul;100(3):600–604. doi: 10.1111/j.1476-5381.1990.tb15853.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Galitzky J., Larrouy D., Berlan M., Lafontan M. New tools for human fat cell alpha-2A adrenoceptor characterization. Identification on membranes and on intact cells using the new antagonist [3H]RX821002. J Pharmacol Exp Ther. 1990 Jan;252(1):312–319. [PubMed] [Google Scholar]
  22. Ghika J., Tennis M., Hoffman E., Schoenfeld D., Growdon J. Idazoxan treatment in progressive supranuclear palsy. Neurology. 1991 Jul;41(7):986–991. doi: 10.1212/wnl.41.7.986. [DOI] [PubMed] [Google Scholar]
  23. Gomez R. E., Ernsberger P., Feinland G., Reis D. J. Rilmenidine lowers arterial pressure via imidazole receptors in brainstem C1 area. Eur J Pharmacol. 1991 Mar 26;195(2):181–191. doi: 10.1016/0014-2999(91)90534-w. [DOI] [PubMed] [Google Scholar]
  24. Hamilton C. A., Yakubu M. A., Jardine E., Reid J. L. Imidazole binding sites in rabbit kidney and forebrain membranes. J Auton Pharmacol. 1991 Aug;11(4):277–283. doi: 10.1111/j.1474-8673.1991.tb00325.x. [DOI] [PubMed] [Google Scholar]
  25. Jackson H. C., Dickinson S. L., Nutt D. J. Exploring the pharmacology of the pro-convulsant effects of alpha 2-adrenoceptor antagonists in mice. Psychopharmacology (Berl) 1991;105(4):558–562. doi: 10.1007/BF02244380. [DOI] [PubMed] [Google Scholar]
  26. Jackson H. C., Griffin I. J., Nutt D. J. The effects of idazoxan and other alpha 2-adrenoceptor antagonists on food and water intake in the rat. Br J Pharmacol. 1991 Sep;104(1):258–262. doi: 10.1111/j.1476-5381.1991.tb12416.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lachaud-Pettiti V., Podevin R. A., Chrétien Y., Parini A. Imidazoline-guanidinium and alpha 2-adrenergic binding sites in basolateral membranes from human kidney. Eur J Pharmacol. 1991 Jan 25;206(1):23–31. doi: 10.1016/0922-4106(91)90142-5. [DOI] [PubMed] [Google Scholar]
  28. Langin D., Lafontan M., Stillings M. R., Paris H. [3H]RX821002: a new tool for the identification of alpha 2A-adrenoceptors. Eur J Pharmacol. 1989 Aug 11;167(1):95–104. doi: 10.1016/0014-2999(89)90751-6. [DOI] [PubMed] [Google Scholar]
  29. Langin D., Lafontan M. [3H]idazoxan binding at non-alpha 2-adrenoceptors in rabbit adipocyte membranes. Eur J Pharmacol. 1989 Jan 10;159(2):199–203. doi: 10.1016/0014-2999(89)90707-3. [DOI] [PubMed] [Google Scholar]
  30. Langin D., Paris H., Dauzats M., Lafontan M. Discrimination between alpha 2-adrenoceptors and [3H]idazoxan-labelled non-adrenergic sites in rabbit white fat cells. Eur J Pharmacol. 1990 Apr 25;188(4-5):261–272. doi: 10.1016/0922-4106(90)90010-u. [DOI] [PubMed] [Google Scholar]
  31. Lehmann J., Koenig-Bérard E., Vitou P. The imidazoline-preferring receptor. Life Sci. 1989;45(18):1609–1615. doi: 10.1016/0024-3205(89)90270-1. [DOI] [PubMed] [Google Scholar]
  32. MacKinnon A. C., Brown C. M., Spedding M., Kilpatrick A. T. [3H]-idazoxan binds with high affinity to two sites on hamster adipocytes: an alpha 2-adrenoceptor and a non-adrenoceptor site. Br J Pharmacol. 1989 Dec;98(4):1143–1150. doi: 10.1111/j.1476-5381.1989.tb12658.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Michel M. C., Brodde O. E., Schnepel B., Behrendt J., Tschada R., Motulsky H. J., Insel P. A. [3H]idazoxan and some other alpha 2-adrenergic drugs also bind with high affinity to a nonadrenergic site. Mol Pharmacol. 1989 Mar;35(3):324–330. [PubMed] [Google Scholar]
  34. Michel M. C., Insel P. A. Are there multiple imidazoline binding sites? Trends Pharmacol Sci. 1989 Sep;10(9):342–344. doi: 10.1016/0165-6147(89)90002-3. [DOI] [PubMed] [Google Scholar]
  35. Munson P. J., Rodbard D. Ligand: a versatile computerized approach for characterization of ligand-binding systems. Anal Biochem. 1980 Sep 1;107(1):220–239. doi: 10.1016/0003-2697(80)90515-1. [DOI] [PubMed] [Google Scholar]
  36. Osman O. T., Rudorfer M. V., Potter W. Z. Idazoxan: a selective alpha 2-antagonist and effective sustained antidepressant in two bipolar depressed patients. Arch Gen Psychiatry. 1989 Oct;46(10):958–959. doi: 10.1001/archpsyc.1989.01810100100021. [DOI] [PubMed] [Google Scholar]
  37. Petrash A. C., Bylund D. B. Alpha-2 adrenergic receptor subtypes indicated by [3H]yohimbine binding in human brain. Life Sci. 1986 Jun 9;38(23):2129–2137. doi: 10.1016/0024-3205(86)90212-2. [DOI] [PubMed] [Google Scholar]
  38. Stillings M. R., Chapleo C. B., Butler R. C., Davis J. A., England C. D., Myers M., Myers P. L., Tweddle N., Welbourn A. P., Doxey J. C. Alpha-adrenoreceptor reagents. 3. Synthesis of some 2-substituted 1,4-benzodioxans as selective presynaptic alpha 2-adrenoreceptor antagonists. J Med Chem. 1985 Aug;28(8):1054–1062. doi: 10.1021/jm00146a013. [DOI] [PubMed] [Google Scholar]
  39. Tesson F., Prip-Buus C., Lemoine A., Pegorier J. P., Parini A. Subcellular distribution of imidazoline-guanidinium-receptive sites in human and rabbit liver. Major localization to the mitochondrial outer membrane. J Biol Chem. 1991 Jan 5;266(1):155–160. [PubMed] [Google Scholar]
  40. Vigne P., Lazdunski M., Frelin C. Guanabenz, guanochlor, guanoxan and idazoxan bind with high affinity to non-adrenergic sites in pig kidney membranes. Eur J Pharmacol. 1989 Jan 31;160(2):295–298. doi: 10.1016/0014-2999(89)90503-7. [DOI] [PubMed] [Google Scholar]
  41. Walter D. S., Flockhart I. R., Haynes M. J., Howlett D. R., Lane A. C., Burton R., Johnson J., Dettmar P. W. Effects of idazoxan on catecholamine systems in rat brain. Biochem Pharmacol. 1984 Aug 15;33(16):2553–2557. doi: 10.1016/0006-2952(84)90623-3. [DOI] [PubMed] [Google Scholar]
  42. Welbourn A. P., Chapleo C. B., Lane A. C., Myers P. L., Roach A. G., Smith C. F., Stillings M. R., Tulloch I. F. Alpha-adrenoreceptor reagents. 4. Resolution of some potent selective prejunctional alpha 2-adrenoreceptor antagonists. J Med Chem. 1986 Oct;29(10):2000–2003. doi: 10.1021/jm00160a033. [DOI] [PubMed] [Google Scholar]
  43. Wikberg J. E. High affinity binding of idazoxan to a non-catecholaminergic binding site in the central nervous system: description of a putative idazoxan-receptor. Pharmacol Toxicol. 1989 Jan;64(1):152–155. doi: 10.1111/j.1600-0773.1989.tb00620.x. [DOI] [PubMed] [Google Scholar]
  44. Wikberg J. E., Uhlén S., Chhajlani V. Medetomidine stereoisomers delineate two closely related subtypes of idazoxan (imidazoline) I-receptors in the guinea pig. Eur J Pharmacol. 1991 Feb 14;193(3):335–340. doi: 10.1016/0014-2999(91)90148-j. [DOI] [PubMed] [Google Scholar]
  45. Wikberg J. E., Uhlén S. Further characterization of the guinea pig cerebral cortex idazoxan receptor: solubilization, distinction from the imidazole site, and demonstration of cirazoline as an idazoxan receptor-selective drug. J Neurochem. 1990 Jul;55(1):192–203. doi: 10.1111/j.1471-4159.1990.tb08838.x. [DOI] [PubMed] [Google Scholar]
  46. Yablonsky F., Dausse J. P. Non-adrenergic binding sites for the "alpha 2-antagonist" [3H]idazoxan in the rabbit urethral smooth muscle. Pharmacological and biochemical characterization. Biochem Pharmacol. 1991 Mar 1;41(5):701–707. doi: 10.1016/0006-2952(91)90069-h. [DOI] [PubMed] [Google Scholar]

Articles from British Journal of Pharmacology are provided here courtesy of The British Pharmacological Society

RESOURCES