Naunyn-Schmiedeberg's Archives of Pharmacology

, Volume 391, Issue 4, pp 361–369 | Cite as

Carrier-mediated uptake of clonidine in cultured human lung cells

  • Marc Le Vée
  • Elodie Jouan
  • Olivier Fardel
Original Article


The lung is a preferential organ site for accumulation of lipophilic basic amine drugs, so-called pneumophilic drugs and belonging to various pharmacological classes, which can result in lung toxicity. In order to investigate the mechanism involved in such pulmonary accumulation of drugs, uptake of clonidine, used here as a prototypical basic amine drug, was characterized in cultured human lung cells. Clonidine accumulation in lung alveolar A549 cells was found to be temperature- and pH-dependent; it was saturable, with a Michaelis–Menten affinity constant (Km) value of 569.4 μM. Various pneumophilic drugs, including amitriptyline, verapamil, propranolol, chlorpromazine, imipramine, and quinidine, markedly cis-inhibited clonidine uptake in A549 cells, in a dose-dependent manner for at least some of them. They additionally trans-stimulated clonidine efflux from A549 cells, thus suggesting that they are substrates for the putative clonidine transporter. In addition to alveolar A549 cells, bronchial epithelial BEAS-2B cells as well as lung endothelial HULEC-5a cells were found to exhibit clonidine accumulation abrogated by amitriptyline, verapamil, and chlorpromazine. Taken together, these data likely provided evidence for carrier-mediated uptake of clonidine in human lung cells. This carrier, which remains to be molecularly identified, interacts with various pneumophilic drugs, suggesting that it may contribute to lung accumulation of these drugs in a notable way.


Clonidine Uptake Carrier Transport Lung Pneumophilic drugs 


Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

210_2018_1467_MOESM1_ESM.pdf (303 kb)
ESM 1 (PDF 302 kb)


  1. Ala-Kokko TI, Pienimaki P, Lampela E, Hollmen AI, Pelkonen O, Vahakangas K (1997) Transfer of clonidine and dexmedetomidine across the isolated perfused human placenta. Acta Anaesthesiol Scand 41(2):313–319. CrossRefPubMedGoogle Scholar
  2. Anderson MW, Orton TC, Pickett RD, Eling TE (1974) Accumulation of amines in the isolated perfused rabbit lung. J Pharmacol Exp Ther 189(2):456–466PubMedGoogle Scholar
  3. Andre P, Debray M, Scherrmann JM, Cisternino S (2009) Clonidine transport at the mouse blood-brain barrier by a new H+ antiporter that interacts with addictive drugs. J Cereb Blood Flow Metab 29(7):1293–1304. CrossRefPubMedGoogle Scholar
  4. Atlas D (1991) Clonidine-displacing substance (CDS) and its putative imidazoline receptor. New leads for further divergence of alpha 2-adrenergic receptor activity. Biochem Pharmacol 41(11):1541–1549. CrossRefPubMedGoogle Scholar
  5. Bend JR, Serabjit-Singh CJ, Philpot RM (1985) The pulmonary uptake, accumulation, and metabolism of xenobiotics. Annu Rev Pharmacol Toxicol 25(1):97–125. CrossRefPubMedGoogle Scholar
  6. Berezhnov AV, Soutar MP, Fedotova EI, Frolova MS, Plun-Favreau H, Zinchenko VP, Abramov AY (2016) Intracellular pH modulates autophagy and Mitophagy. J Biol Chem 291(16):8701–8708. CrossRefPubMedPubMedCentralGoogle Scholar
  7. Boutroy MJ, Gisonna CR, Legagneur M (1988) Clonidine: placental transfer and neonatal adaption. Early Hum Dev 17(2-3):275–286. CrossRefPubMedGoogle Scholar
  8. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72(1-2):248–254. CrossRefPubMedGoogle Scholar
  9. Brown EA (1974) THe localization, metabolism and effects of drugs and toxicants in lung. Drug Metab Rev 3(1):33–87CrossRefPubMedGoogle Scholar
  10. Camus P, Coudert B, D'Athis P, Dumas M, Escousse A, Jeannin L (1990) Pharmacokinetics of amiodarone in the isolated rat lung. J Pharmacol Exp Ther 254(1):336–343PubMedGoogle Scholar
  11. Chapy H, Andre P, Decleves X, Scherrmann JM, Cisternino S (2015a) A polyspecific drug/proton antiporter mediates diphenhydramine and clonidine transport at the mouse blood-retinal barrier. Br J Pharmacol 172(19):4714–4725. CrossRefPubMedPubMedCentralGoogle Scholar
  12. Chapy H, Goracci L, Vayer P, Parmentier Y, Carrupt PA, Decleves X, Scherrmann JM, Cisternino S, Cruciani G (2015b) Pharmacophore-based discovery of inhibitors of a novel drug/proton antiporter in human brain endothelial hCMEC/D3 cell line. Br J Pharmacol 172:4888–4904CrossRefPubMedPubMedCentralGoogle Scholar
  13. Cody V, DeTitta GT (1979) The molecular conformation of clonidine hydrochloride, an a-adrenergic agonist. Journal of Crystal and Molecular Structure 9(1):33–43. CrossRefGoogle Scholar
  14. Conway EL, Jarrott B (1980) Clonidine distribution in the rat: temporal relationship between tissue levels and blood pressure response. Br J Pharmacol 71(2):473–478. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Dahlin KL, Lastbom L, Blomgren B, Ryrfeldt A (1997) Acute lung failure induced by tricyclic antidepressants. Toxicol Appl Pharmacol 146(2):309–316. CrossRefPubMedGoogle Scholar
  16. Daniel WA, Wojcikowski J (1997) Contribution of lysosomal trapping to the total tissue uptake of psychotropic drugs. Pharmacol Toxicol 80(2):62–68. CrossRefPubMedGoogle Scholar
  17. Ernsberger P, Graves ME, Graff LM, Zakieh N, Nguyen P, Collins LA, Westbrooks KL, Johnson GG (1995) I1-imidazoline receptors. Definition, characterization, distribution, and transmembrane signaling. Ann N Y Acad Sci 763:22–42CrossRefPubMedGoogle Scholar
  18. Fischer W, Metzner L, Hoffmann K, Neubert RH, Brandsch M (2006) Substrate specificity and mechanism of the intestinal clonidine uptake by Caco-2 cells. Pharm Res 23(1):131–137. CrossRefPubMedGoogle Scholar
  19. Fischer W, Neubert RH, Brandsch M (2007) Clonidine accumulation in human neuronal cells. Eur J Pharm Sci 32(4-5):291–295. CrossRefPubMedGoogle Scholar
  20. Foth H (1995) Role of the lung in accumulation and metabolism of xenobiotic compounds--implications for chemically induced toxicity. Crit Rev Toxicol 25(2):165–205. CrossRefPubMedGoogle Scholar
  21. Grafe F, Wohlrab W, Neubert R, Brandsch M (2004) Carrier-mediated transport of clonidine in human keratinocytes. Eur J Pharm Sci 21(2-3):309–312. CrossRefPubMedGoogle Scholar
  22. Heytler PG (1979) Uncouplers of oxidative phosphorylation. Methods Enzymol 55:462–442. CrossRefPubMedGoogle Scholar
  23. Insel PA, Sanda M (1979) Temperature-dependent changes in binding to beta-adrenergic receptors of intact S49 lymphoma cells. Implications for the state of the receptor that activates adenylate cyclase under physiological conditions. J Biol Chem 254(14):6554–6559PubMedGoogle Scholar
  24. Isaac L (1980) Clonidine in the central nervous system: site and mechanism of hypotensive action. J Cardiovasc Pharmacol 2(Suppl 1):S5–19. CrossRefPubMedGoogle Scholar
  25. Ito Y, Sato S, Son M, Kume H, Takagi K, Yamaki K (2002) Bioelectric toxicity caused by chlorpromazine in human lung epithelial cells. Toxicol Appl Pharmacol 183(3):198–206. CrossRefPubMedGoogle Scholar
  26. Iven H (1977) The pharmacokinetics and organ distribution of ajmaline and quindine in the mouse. Naunyn Schmiedeberg's Arch Pharmacol 298(1):43–50. CrossRefGoogle Scholar
  27. Jarrott B, Louis WJ, Summers RJ (1979) The characteristics of [3H]-clonidine binding to an alpha-adrenoceptor in membranes from guinea-pig kidney. Br J Pharmacol 65(4):663–670. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Jigorel E, Le Vee M, Boursier-Neyret C, Bertrand M, Fardel O (2005) Functional expression of sinusoidal drug transporters in primary human and rat hepatocytes. Drug Metab Dispos 33:1418–1422CrossRefPubMedGoogle Scholar
  29. Jonker JW, Schinkel AH (2004) Pharmacological and physiological functions of the polyspecific organic cation transporters: OCT1, 2, and 3 (SLC22A1-3). J Pharmacol Exp Ther 308(1):2–9. CrossRefPubMedGoogle Scholar
  30. Kubo Y, Seko N, Usui T, Akanuma S, Hosoya K (2016) Lysosomal trapping is present in retinal capillary endothelial cells: insight into its influence on cationic drug transport at the inner blood-retinal barrier. Biol Pharm Bull 39:1319–1324CrossRefPubMedGoogle Scholar
  31. Kubo Y, Tsuchiyama A, Shimizu Y, Akanuma S, Hosoya K (2014) Involvement of carrier-mediated transport in the retinal uptake of clonidine at the inner blood-retinal barrier. Mol Pharm 11:3747–3753CrossRefPubMedGoogle Scholar
  32. Le Vee M, Noel G, Jouan E, Stieger B, Fardel O (2013) Polarized expression of drug transporters in differentiated human hepatoma HepaRG cells. Toxicol in Vitro 27(6):1979–1986. CrossRefPubMedGoogle Scholar
  33. Li JX (2017) Imidazoline I2 receptors: An update. Pharmacol Ther 178:48–56. CrossRefPubMedGoogle Scholar
  34. MacGregor TR, Relihan GL, Keirns JJ (1985) Pharmacokinetics of oral sustained release clonidine in humans. Arzneimittelforschung 35(1A):440–446PubMedGoogle Scholar
  35. Mayati A, Moreau A, Denizot C, Stieger B, Parmentier Y, Fardel O (2017) beta2-adrenergic receptor-mediated in vitro regulation of human hepatic drug transporter expression by epinephrine. Eur J Pharm Sci 106:302–312. CrossRefPubMedGoogle Scholar
  36. Misinzo G, Delputte PL, Nauwynck HJ (2008) Inhibition of endosome-lysosome system acidification enhances porcine circovirus 2 infection of porcine epithelial cells. J Virol 82(3):1128–1135. CrossRefPubMedGoogle Scholar
  37. Mizugaki M, Hishinuma T, Kimura K, Nakamura H, Aso H, Ishii F, Nishikawa M, Itoh K, Tomioka Y, Ishiwata S, Moritani H, Uyama T, Ido T (1994) The distribution of [11C]cocaine in normal and cocaine-sensitization mice. Nucl Med Biol 21(6):793–799. CrossRefPubMedGoogle Scholar
  38. Moreau A, Le Vee M, Jouan E, Parmentier Y, Fardel O (2011) Drug transporter expression in human macrophages. Fundam Clin Pharmacol 25:743–752CrossRefPubMedGoogle Scholar
  39. Muller J, Neubert R, Brandsch M (2004) Transport of clonidine at cultured epithelial cells (JEG-3) of the human placenta. Pharm Res 21:692–694CrossRefPubMedGoogle Scholar
  40. Nickel S, Clerkin CG, Selo MA, Ehrhardt C (2016) Transport mechanisms at the pulmonary mucosa: implications for drug delivery. Expert Opin Drug Deliv 13(5):667–690. CrossRefPubMedGoogle Scholar
  41. Orton TC, Anderson MW, Pickett RD, Eling TE, Fouts JR (1973) Xenobiotic accumulation and metabolism by isolated perfused rabbit lungs. J Pharmacol Exp Ther 186(3):482–497PubMedGoogle Scholar
  42. Paalzow LK, Edlund PO (1979) Pharmacokinetics of clonidine in the rat and cat. J Pharmacokinet Biopharm 7(5):481–494. CrossRefPubMedGoogle Scholar
  43. Regunathan S, Reis DJ (1996) Imidazoline receptors and their endogenous ligands. Annu Rev Pharmacol Toxicol 36(1):511–544. CrossRefPubMedGoogle Scholar
  44. Roerig DL, Kotrly KJ, Dawson CA, Ahlf SB, Gualtieri JF, Kampine JP (1989) First-pass uptake of verapamil, diazepam, and thiopental in the human lung. Anesth Analg 69:461-466CrossRefGoogle Scholar
  45. Ryan CL, Pappas BA (1990) Prenatal exposure to antiadrenergic antihypertensive drugs: effects on neurobehavioral development and the behavioral consequences of enriched rearing. Neurotoxicol Teratol 12(4):359–366. CrossRefPubMedGoogle Scholar
  46. Sakamoto A, Matsumaru T, Yamamura N, Suzuki S, Uchida Y, Tachikawa M, Terasaki T (2015) Drug transporter protein quantification of immortalized human lung cell lines derived from tracheobronchial epithelial cells (Calu-3 and BEAS2-B), bronchiolar-alveolar cells (NCI-H292 and NCI-H441), and alveolar type II-like cells (A549) by liquid chromatography-tandem mass spectrometry. J Pharm Sci 104:3029–3038CrossRefPubMedGoogle Scholar
  47. Salomon JJ, Ehrhardt C, Hosoya K (2014) The verapamil transporter expressed in human alveolar epithelial cells (A549) does not interact with beta2-receptor agonists. Drug Metab Pharmacokinet 29(1):101–104. CrossRefPubMedGoogle Scholar
  48. Salomon JJ, Gausterer JC, Yahara T, Hosoya K, Huwer H, Hittinger M, Schneider-Daum N, Lehr CM, Ehrhardt C (2015) Organic cation transporter function in different in vitro models of human lung epithelium. Eur J Pharm Sci 80:82–88. CrossRefPubMedGoogle Scholar
  49. Sugano K, Kansy M, Artursson P, Avdeef A, Bendels S, Di L, Ecker GF, Faller B, Fischer H, Gerebtzoff G, Lennernaes H, Senner F (2010) Coexistence of passive and carrier-mediated processes in drug transport. Nat Rev Drug Discov 9(8):597–614. CrossRefPubMedGoogle Scholar
  50. Svens K, Ryrfeldt A (2001) A study of mechanisms underlying amitriptyline-induced acute lung function impairment. Toxicol Appl Pharmacol 177(3):179–187. CrossRefPubMedGoogle Scholar
  51. Takano M, Kamei H, Nagahiro M, Kawami M, Yumoto R (2017) Nicotine transport in lung and non-lung epithelial cells. Life Sci 188:76–82CrossRefPubMedGoogle Scholar
  52. Tamai I (2013) Pharmacological and pathophysiological roles of carnitine/organic cation transporters (OCTNs: SLC22A4, SLC22A5 and Slc22a21). Biopharm Drug Dispos 34(1):29–44. CrossRefPubMedGoogle Scholar
  53. Uhr M, Grauer MT, Yassouridis A, Ebinger M (2007) Blood-brain barrier penetration and pharmacokinetics of amitriptyline and its metabolites in p-glycoprotein (abcb1ab) knock-out mice and controls. J Psychiatr Res 41(1-2):179–188. CrossRefPubMedGoogle Scholar
  54. Zhang J, Abdel-Rahman AA (2006) Nischarin as a functional imidazoline (I1) receptor. FEBS Lett 580(13):3070–3074. CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Institut de Recherches en Santé, Environnement et Travail (IRSET), UMR INSERM U1085, Université de Rennes 1RennesFrance
  2. 2.Pôle Biologie, Centre Hospitalier UniversitaireRennesFrance

Personalised recommendations