, Volume 2, Issue 1, pp 63–72 | Cite as

Brain-to-blood transporters for endogenous substrates and xenobiotics at the blood-brain barrier: An overview of biology and methodology



In the past decade, research into P-glycoprotein involving the blood-brain barrier (BBB) has seen a shift in the concept of the BBB as a structural barrier to that of a functional barrier for xenobiotics and changed simultaneously the strategy for the discovery and development of drugs acting in the CNS. As far as making advances in neurotherapeutics are concerned, the brain-to-blood transport function at the BBB will be one of the most important issues. Knowing the limitations of thein vivo andin vitro methods for BBB efflux research, it is essential to adopt a multidisciplinary approach in investigating the true physiological role of the BBB. Among several methods, the Brain Efflux Index method and the use of conditionally immortalized brain capillary endothelial cell lines, established from transgenic rats harboring temperature-sensitive simian virus 40 large T-antigen gene, are likely to be very useful tools for the BBB efflux transport research. According to our recent findings using these methods, several transporters in the brain capillary endothelial cells appear to play an important role in reducing the brain level of hydrophilic endogenous substrates produced either in the brain or peripheral organs, e.g., neurotransmitters, neuromodulators, metabolites of neurotransmitters, and uremic toxins. It has been reported also that large hydrophilic molecules, such as IgG, apo-transferrin, and amyloid-β peptide, are susceptible to brain-to-blood efflux transport. In the light of the latest findings, we have formed the hypothesis that the BBB acts as a CNS detoxifying system for both endogenous substrates and xenobiotics in the brain. A fuller understanding of the physiological role of BBB efflux transporters will provide rational insights to assist in the development of safer neurotherapeutics.

Key Words

Blood-brain barrier efflux transport brain efflux index method conditionally immortalized cell line MDR1 ABCG2/BCRP amyloid peptide 


  1. 1.
    Reese TS, Karnovsky MJ. Fine structural localization of blood-brain banier to exogenous peroxidase.J Cell Biol 34: 207–217, 1967.PubMedCrossRefGoogle Scholar
  2. 1a.
    Reese TS, Feder N, Brightman MW. Electron microscopic study of the blood-brain and blood-cerebrospinal fluid barriers with microperoxidase.J Neuropathol Exp Neurol 30: 137–138, 1971.PubMedGoogle Scholar
  3. 1b.
    Cordon-Cardo C, O’Brien JP, Casals D, Rittman-Grauer L, Biedler JL, Melamed MR, et al. Multidrug-resistance gene (P-glycoprotein) is expressed by endothelial cells at blood-brain barrier sites.Proc Natl Acad Sci USA 86: 695–698, 1989.PubMedCrossRefGoogle Scholar
  4. 2.
    Tsuji A, Terasaki T, Takabatake Y, Tenda Y, Tamai I, Yamashima T, et al. P-glycoprotein as the drug efflux pump in primary cultured bovine brain capillary endothelial cells.Life Sci 51: 1427–1437, 1992.PubMedCrossRefGoogle Scholar
  5. 3.
    Schinkel AH, Smit JJ, van Tellingen O, Beijnen JH, Wagenaar E, van Deemter L, et al. Disruption of the mouse mdrla P-glycoprotein gene leads to a deficiency in the blood-brain barrier and to increased sensitivity to drugs.Cell 77: 491–502, 1994.PubMedCrossRefGoogle Scholar
  6. 4.
    Terasaki T, Hosoya K. The blood-brain barrier efflux transporters as a detoxifying system for the brain.Adv Drug Deliv Rev 36: 195–209, 1999.PubMedCrossRefGoogle Scholar
  7. 5.
    Kakee A, Terasaki T, Sugiyama Y. Brain efflux index as a novel method of analyzing efflux transport at the blood-brain barrier.J Pharmacol Exp Ther 277: 1550–1559, 1996.PubMedGoogle Scholar
  8. 5a.
    Zhang Y, Schuetz JD, Elmquist WF, Miller DW. Plasma membrane localization of multidrug resistance-associated protein (MRP) homologues in brain capillary endothelial cells.J Pharmacol Exp Ther 311: 449–455, 2004.PubMedCrossRefGoogle Scholar
  9. 6.
    Mori S, Takanaga H, Ohtsuki S, Deguchi T, Kang YS, Hosoya K, et al. Rat organic anion transporter 3 (rOAT3) is responsible for brain-to-blood efflux of homovanillic acid at the abluminal membrane of brain capillary endothelial cells.J Cereb Blood Flow Metab 23: 432–440, 2003.PubMedCrossRefGoogle Scholar
  10. 7.
    Terasaki T, Ohtsuki S, Hori S, Takanaga H, Nakashima E, Hosoya K. New approaches to in vitro models of blood-brain barrier drug transport.Drug Discov Today 8: 944–954, 2003.PubMedCrossRefGoogle Scholar
  11. 8.
    Gaillard PJ, Voorwinden LH, Nielsen JL, Ivanov A, Atsumi R, Engman H, et al. Establishment and functional characterization of an in vitro model of the blood-brain barrier, comprising a co-culture of brain capillary endothelial cells and astrocytes.Eur J Pharm Sci 12: 215–222, 2001.PubMedCrossRefGoogle Scholar
  12. 9.
    Gaillard PJ, van der Sandt IC, Voorwinden LH, Vu D, Nielsen JL, de Boer AG, et al. Astrocytes increase the functional expression of P-glycoprotein in an in vitro model of the blood-brain barrier.Pharm Res 17: 1198–1205, 2000.PubMedCrossRefGoogle Scholar
  13. 10.
    Hosoya K, Tetsuka K, Nagase K, Tomi M, Saeki S, Ohtsuki S, et al. Conditionally immortalized brain capillary endothelial cell lines established from a transgenic mouse harboring temperature—sensitive simian virus 40 large T-antigen gene.AAPS PharmSci 2: E27, 2000.PubMedCrossRefGoogle Scholar
  14. 11.
    Hosoya K, Takashima T, Tetsuka K, Nagura T, Ohtsuki S, Takanaga H, et al. mRNA expression and transport characterization of conditionally immortalized rat brain capillary endothelial cell lines; a new in vitro BBB model for drug targeting.J Drug Target 8: 357–370, 2000.PubMedCrossRefGoogle Scholar
  15. 12.
    Hori S, Ohtsuki S, Hosoya K, Nakashima E, Terasaki T. A pericyte-derived angiopoietin-1 multimeric complex induces occludin gene expression in brain capillary endothelial cells through Tie-2 activation in vitro.J Neurochem 89: 503–513, 2004.PubMedCrossRefGoogle Scholar
  16. 13.
    Hori S, Ohtsuki S, Tachikawa M, Kimura N, Kondo T, Watanabe M, et al. Functional expression of rat ABCG2 on the luminal side of brain capillaries and its enhancement by astrocyte-derived soluble factor(s).J Neurochem 90: 526–536, 2004.PubMedCrossRefGoogle Scholar
  17. 14.
    Choi TB, Pardridge WM. Phenylalanine transport at the human blood-brain barrier. Studies with isolated human brain capillaries.J Biol Chem 261: 6536–6541, 1986.PubMedGoogle Scholar
  18. 15.
    O’Kane RL, Martinez-Lopez I, DeJoseph MR, Vina JR, Hawkins RA. Na+-dependent glutamate transporters (EAAT1, EAAT2, and EAAT3) of the blood-brain barrier. A mechanism for glutamate removal.J Biol Chem 274: 31891–31895, 1999.PubMedCrossRefGoogle Scholar
  19. 16.
    Sanchez del Pino MM, Peterson DR, Hawkins RA. Neutral amino acid transport characterization of isolated luminal and abluminal membranes of the blood-brain barrier.J Biol Chem 270: 14913–14918, 1995.PubMedCrossRefGoogle Scholar
  20. 17.
    Yamazaki M, Neway WE, Ohe T, Chen I, Rowe JF, Hochman JH, et al. In vitro substrate identification studies for p-glycoprotein-mediated transport species difference and predictability of in vivo results.J Pharmacol Exp Ther 296: 723–735, 2001.PubMedGoogle Scholar
  21. 18.
    Kakee A, Takanaga H, Terasaki T, Naito M, Tsuruo T, Sugiyama Y. Efflux of a suppressive neurotransmitter, GABA, across the blood-brain barrier.J Neurochem 79: 110–118, 2001.PubMedCrossRefGoogle Scholar
  22. 19.
    Tetsuka K, Takanaga H, Ohtsuki S, Hosoya K, Terasaki T. The L-isomer-selective transport of aspartic acid is mediated by ASCT2 at the blood-brain barrier.J Neurochem 87: 891–901, 2003.PubMedCrossRefGoogle Scholar
  23. 20.
    Takanaga H, Ohtsuki S, Hosoya K, Terasaki T. GAT2/BGT-1 as a system responsible for the transport of γ-aminobutyric acid at the mouse blood-brain barrier.J Cereb Blood Flow Metab 21: 1232–1239, 2001.PubMedCrossRefGoogle Scholar
  24. 21.
    Hardebo JE, Owman C. Characterization of the in vitro uptake of monoamines into brain microvessels.Acta Physiol Scand 108: 223–229, 1980.PubMedCrossRefGoogle Scholar
  25. 22.
    Wakayama K, Ohtsuki S, Takanaga H, Hosoya K, Terasaki T. Localization of norepinephrine and serotonin transporter in mouse brain capillary endothelial cells.Neurosci Res 44: 173–180, 2002.PubMedCrossRefGoogle Scholar
  26. 23.
    Hosoya K, Sugawara M, Asaba H, Terasaki T. Blood-brain barrier produces significant efflux of L-aspartic acid but not D-aspartic acid in vivo evidence using the brain efflux index method.J Neurochem 73: 1206–1211, 1999.PubMedCrossRefGoogle Scholar
  27. 24.
    Betz AL, Goldstein GW. Polarity of the blood-brain barrier neutral amino acid transport into isolated brain capillaries.Science 202: 225–227, 1978.PubMedCrossRefGoogle Scholar
  28. 25.
    Takanaga H, Tokuda N, Ohtsuki S, Hosoya K, Terasaki T. ATA2 is predominantly expressed as system A at the blood-brain barrier and acts as brain-to-blood efflux transport for L-proline.Mol Pharmacol 61: 1289–1296, 2002.PubMedCrossRefGoogle Scholar
  29. 26.
    Emanuelsson BM, Paalzow L, Sunzel M. Probenecid-induced accumulation of 5-hydroxyindoleacetic acid and homovanillic acid in rat brain.J Pharm Pharmacol 39: 705–710, 1987.PubMedCrossRefGoogle Scholar
  30. 27.
    Kim CS, Roe CR, Mann JD, Breese GR. Octanoic acid produces accumulation of monoamine acidic metabolites in the brain interaction with organic anion transport at the choroid plexus.J Neurochem 58: 1499–1503, 1992.PubMedCrossRefGoogle Scholar
  31. 28.
    Morrison PF, Morishige GM, Beagles KE, Heyes MP. Quinolinic acid is extruded from the brain by a probenecid-sensitive carrier system a quantitative analysis.J Neurochem 72: 2135–2144, 1999.PubMedCrossRefGoogle Scholar
  32. 29.
    Kusuhara H, Sekine T, Utsunomiya-Tate N, Tsuda M, Kojima R, Cha SH, et al. Molecular cloning and characterization of a new multispecific organic anion transporter from rat brain.J Biol Chem 274: 13675–13680, 1999.PubMedCrossRefGoogle Scholar
  33. 30.
    Mori S, Ohtsuki S, Takanaga H, Kikkawa T, Kang YS, Terasaki T. Organic anion transporter 3 is involved in the brain-to-blood efflux transport of thiopurine nucleobase analogs.J Neurochem 90: 931–941, 2004.PubMedCrossRefGoogle Scholar
  34. 31.
    Asaba H, Hosoya K, Takanaga H, Ohtsuki S, Tamura E, Takizawa T, et al. Blood-brain barrier is involved in the efflux transport of a neuroactive steroid, dehydroepiandrosterone sulfate, via organic anion transporting polypeptide 2.J Neurochem 75: 1907–1916, 2000.PubMedCrossRefGoogle Scholar
  35. 32.
    Gao B, Stieger B, Noe B, Fritschy JM, Meier PJ. Localization of the organic anion transporting polypeptide 2 (Oatp2) in capillary endothelium and choroid plexus epithelium of rat brain.J Histochem Cytochem 47: 1255–1264, 1999.PubMedCrossRefGoogle Scholar
  36. 33.
    Hosoya K, Asaba H, Terasaki T. Brain-to-blood efflux transport of estrone-3-sulfate at the blood-brain barrier in rats.Life Sci 67: 2699–2711, 2000.PubMedCrossRefGoogle Scholar
  37. 34.
    Deguchi T, Kusuhara H, Takadate A, Endou H, Otagiri M, Sugiyama Y. Characterization of uremic toxin transport by organic anion transporters in the kidney.Kidney Int 65: 162–174, 2004.PubMedCrossRefGoogle Scholar
  38. 35.
    Ohtsuki S, Asaba H, Takanaga H, Deguchi T, Hosoya K, Otagiri M, et al. Role of blood-brain barrier organic anion transporter 3 (OAT3) in the efflux of indoxyl sulfate, a uremic toxin its involvement in neurotransmitter metabolite clearance from the brain.J Neurochem 83: 57–66, 2002.PubMedCrossRefGoogle Scholar
  39. 36.
    Muting D. Studies on the pathogenesis of uremia. Comparative determinations of glucuronic acid, indican, free and bound phenols in the serum, cerebrospinal fluid, and urine of renal diseases with and without uremia.Clin Chim Acta 12: 551–554, 1965.PubMedCrossRefGoogle Scholar
  40. 37.
    Deguchi Y, Yokoyama Y, Sakamoto T, Hayashi H, Naito T, Yamada S, et al. Brain distribution of 6-mercaptopurine is regulated by the efflux transport system in the blood-brain barrier.Life Sci 66: 649–662, 2000.PubMedCrossRefGoogle Scholar
  41. 38.
    Terasaki T, Pardridge WM. Restricted transport of 3′-azido-3′-deoxythymidine and dideoxynucleosides through the blood-brain barrier.J Infect Dis 158: 630–632, 1988.PubMedCrossRefGoogle Scholar
  42. 39.
    Takasawa K, Terasaki T, Suzuki H, Sugiyama Y. In vivo evidence for carrier-mediated efflux transport of 3′-azido-3′-deoxythymidine and 2′,3′-dideoxyinosine across the blood-brain barrier via a probenecid-sensitive transport system.J Pharmacol Exp Ther 281: 369–375, 1997.PubMedGoogle Scholar
  43. 40.
    Zhang Y, Han H, Elmquist WF, Miller DW. Expression of various multidrug resistance-associated protein (MRP) homologues in brain microvessel endothelial cells.Brain Res 876: 148–153, 2000.PubMedCrossRefGoogle Scholar
  44. 41.
    Zhang Y, Schuetz JD, Elmquist WF, Miller DW. Plasma membrane localization of multidrug resistance-associated protein (MRP) homologues in brain capillary endothelial cells.J Pharmacol Exp Ther 311: 449–455, 2004.PubMedCrossRefGoogle Scholar
  45. 42.
    Sugiyama D, Kusuhara H, Lee YJ, Sugiyama Y. Involvement of multidrug resistance associated protein 1 (Mrp1) in the efflux transport of 17β estradiol-D-17β-glucuronide (E217βG) across the blood-brain barrier.Pharm Res 20: 1394–1400, 2003.PubMedCrossRefGoogle Scholar
  46. 43.
    Cooray HC, Blackmore CG, Maskell L, Barrand MA. Localisation of breast cancer resistance protein in microvessel endothelium of human brain.Neuroreport 13: 2059–2063, 2002.PubMedCrossRefGoogle Scholar
  47. 44.
    Kage K, Tsukahara S, Sugiyama T, Asada S, Ishikawa E, Tsuruo T, et al. Dominant-negative inhibition of breast cancer resistance protein as drug efflux pump through the inhibition of S-S dependent homodimerization.Int J Cancer 97: 626–630, 2002.PubMedCrossRefGoogle Scholar
  48. 45.
    Cisternino S, Mercier C, Bourasset F, Roux F, Scherrmann JM. Expression, up-regulation, and transport activity of the multidrug-resistance protein Abcg2 at the mouse blood-brain barrier.Cancer Res 64: 3296–3301, 2004.PubMedCrossRefGoogle Scholar
  49. 46.
    Zhang Y, Pardridge WM. Mediated efflux of IgG molecules from brain to blood across the blood-brain barrier.J Neuroimmunol 114: 168–172, 2001.PubMedCrossRefGoogle Scholar
  50. 47.
    Zhang Y, Pardridge WM. Rapid transferrin efflux from brain to blood across the blood-brain barrier.J Neurochem 76: 1597–1600, 2001.PubMedCrossRefGoogle Scholar
  51. 48.
    Shibata M, Yamada S, Kumar SR, Calero M, Bading J, Frangione B, et al. Clearance of Alzheimer’s amyloid-ss(1–40) peptide from brain by LDL receptor-related protein-1 at the blood-brain barrier.J Clin Invest 106: 1489–1499, 2000.PubMedCrossRefGoogle Scholar
  52. 49.
    Lam FC, Liu R, Lu P, Shapiro AB, Renoir JM, Sharom FJ, et al. β-Amyloid efflux mediated by p-glycoprotein.J Neurochem 76: 1121–1128, 2001.PubMedCrossRefGoogle Scholar
  53. 50.
    Vogelgesang S, Cascorbi I, Schroeder E, Pahnke J, Kroemer HK, Siegmund W, et al. Deposition of Alzheimer’s β-amyloid is inversely correlated with P-glycoprotein expression in the brains of elderly non-demented humans.Pharmacogenetics 12: 535–541, 2002.PubMedCrossRefGoogle Scholar
  54. 51.
    Bard F, Cannon C, Barbour R, Burke RL, Games D, Grajeda H, et al. Peripherally administered antibodies against amyloid β-peptide enter the central nervous system and reduce pathology in a mouse model of Alzheimer disease.Nat Med 6: 916–919, 2000.PubMedCrossRefGoogle Scholar
  55. 52.
    DeMattos RB, Bales KR, Cummins DJ, Dodart JC, Paul SM, Holtzman DM. Peripheral anti-Aβ antibody alters CNS and plasma Aβ clearance and decreases brain Aβ burden in a mouse model of Alzheimer’s disease.Proc Natl Acad Sci USA 98: 8850–8855, 2001.PubMedCrossRefGoogle Scholar
  56. 53.
    Terasaki T, Hosoya K. The brain efflux index method (BEI). In: Alfred Benzon Symposium 45, Brain Barrier Systems (Palson OB, Knudsen GM, Moos T, eds). Cophenhagen, Denmark: Munksgaad, 1998.Google Scholar

Copyright information

© The American Society for Experimental NeuroTherapeutics, Inc 2005

Authors and Affiliations

  1. 1.New Industry Creation Hatchery Center and Graduate School of Pharmaceutical SciencesTohoku UniversitySendaiJapan
  2. 2.CREST & SORSTJapan Science and Technology AgencyJapan

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