Abstract
The blood brain barrier regulates the transport of chemicals from entering and leaving the brain. Brain capillaries establish the barrier and restrict transport into the brain by providing a physical and chemical barrier. The physical barrier is due to tight membrane junctions separating the capillary endothelial cells resulting in limited paracellular transport. The chemical barrier is due to the expression of multidrug transporters that mediate the efflux of a broad range of hydrophobic chemicals. Because of the unusual nutrient demands of the brain, this limited permeability is compensated by the expression of a large number of transporters that are responsive to the metabolic demands of the brain. Consequently, the blood brain barrier indirectly regulates brain function by directly controlling the uptake of nutrients. Two widely used methods for studying the blood brain are a cell culture model using rat, pig, or cow brain endothelial cells and isolated microvessels. The cell culture model is more popular likely because it is easier to use and less costly compared to isolated microvessels. In some laboratories, brain endothelial cells are cocultured with astrocyte- or astroglial-conditioned media. The endothelial cells express many of the transporters displayed in vivo but not all. Although cell culture models vary, none express the tight barrier observed in vivo. Because microvessels are isolated directly from the brain, they express all of the transporters displayed in vivo. Their disadvantage is that the preparation is laborious, requires animals, and has a shorter lifespan in vitro. We present an approach in which transport is first verified in isolated microvessels, and then the mechanism is studied in cell culture.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Rubin, L. L. & Staddon, J. M. (1999). The cell biology of the blood-brain barrier. Annu Rev Neurosci 22, 11–28.
Zeller, K., Rahner-Welsch, S. & Kuschinsky, W. (1997). Distribution of Glut1 glucose transporters in different brain structures compared to glucose utilization and capillary density of adult rat brains. J Cereb Blood Flow Metab 17, 204–9.
Gordon, G. R., Mulligan, S. J. & MacVicar, B. A. (2007). Astrocyte control of the cerebrovasculature. Glia 55, 1214–21.
Begley, D. J. (2004). ABC transporters and the blood-brain barrier. Curr Pharm Des 10, 1295–312.
Kalaria, R. N. & Harik, S. I. (1987). Blood-brain barrier monoamine oxidase: enzyme characterization in cerebral microvessels and other tissues from six mammalian species, including human. J Neurochem 49, 856–64.
Renkawek, K., Murray, M. R., Spatz, M. & Klatzo, I. (1976). Distinctive histochemical characteristics of brain capillaries in organotypic culture. Exp Neurol 50, 194–206.
Wolff, J. E., Belloni-Olivi, L., Bressler, J. P. & Goldstein, G. W. (1992). Gamma-glutamyl transpeptidase activity in brain microvessels exhibits regional heterogeneity. J Neurochem 58, 909–15.
Nicolazzo, J. A., Charman, S. A. & Charman, W. N. (2006). Methods to assess drug permeability across the blood-brain barrier. J Pharm Pharmacol 58, 281–93.
Roux, F. & Couraud, P. O. (2005). Rat brain endothelial cell lines for the study of Âblood-brain barrier permeability and transport functions. Cell Mol Neurobiol 25, 41–58.
Bobilya, D. J. A model for transport studies of the blood-brain barrier. Methods Mol Biol 637, 149–63.
Gumbleton, M. & Audus, K. L. (2001). Progress and limitations in the use of in vitro cell cultures to serve as a permeability screen for the blood-brain barrier. J Pharm Sci 90, 1681–98.
Perriere, N., Yousif, S., Cazaubon, S., Chaverot, N., Bourasset, F., Cisternino, S., Decleves, X., Hori, S., Terasaki, T., Deli, M., Scherrmann, J. M., Temsamani, J., Roux, F. & Couraud, P. O. (2007). A functional in vitro model of rat blood-brain barrier for molecular analysis of efflux transporters. Brain Res 1150, 1–13.
Boado, R. J. & Pardridge, W. M. (1994). Measurement of blood-brain barrier GLUT1 glucose transporter and actin mRNA by a quantitative polymerase chain reaction assay. J Neurochem 62, 2085–90.
Kido, Y., Tamai, I., Nakanishi, T., Kagami, T., Hirosawa, I., Sai, Y. & Tsuji, A. (2002). Evaluation of blood-brain barrier transporters by co-culture of brain capillary endothelial cells with astrocytes. Drug Metab Pharmacokinet 17, 34–41.
Dolman, D., Drndarski, S., Abbott, N. J. & Rattray, M. (2005). Induction of aquaporin 1 but not aquaporin 4 messenger RNA in rat primary brain microvessel endothelial cells in culture. J Neurochem 93, 825–33.
Butt, A. M., Jones, H. C. & Abbott, N. J. (1990). Electrical resistance across the Âblood-brain barrier in anaesthetized rats: a developmental study. J Physiol 429, 47–62.
Smith, Q. R. & Rapoport, S. I. (1986). Cerebrovascular permeability coefficients to sodium, potassium, and chloride. J Neurochem 46, 1732–42.
Crone, C. (1984). Lack of selectivity to small ions in paracellular pathways in cerebral and muscle capillaries of the frog. J Physiol 353, 317–37.
Yang, J., Mutkus, L. A., Sumner, D., Stevens, J. T., Eldridge, J. C., Strandhoy, J. W. & Aschner, M. (2001). Transendothelial permeability of chlorpyrifos in RBE4 monolayers is modulated by astrocyte-conditioned medium. Brain Res Mol Brain Res 97, 43–50.
Stewart, P. A. & Wiley, M. J. (1981). Developing nervous tissue induces formation of blood-brain barrier characteristics in invading endothelial cells: a study using quail – chick transplantation chimeras. Dev Biol 84, 183–92.
Gaillard, P. J., Voorwinden, L. H., Nielsen, J. L., Ivanov, A., Atsumi, R., Engman, H., Ringbom, C., de Boer, A. G. & Breimer, D. D. (2001). 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, 215–22.
Wolburg, H., Neuhaus, J., Kniesel, U., Krauss, B., Schmid, E. M., Ocalan, M., Farrell, C. & Risau, W. (1994). Modulation of tight junction structure in blood-brain barrier endothelial cells. Effects of tissue culture, second messengers and cocultured astrocytes. J Cell Sci 107 ( Pt 5), 1347–57.
Roux, F., Durieu-Trautmann, O., Chaverot, N., Claire, M., Mailly, P., Bourre, J. M., Strosberg, A. D. & Couraud, P. O. (1994). Regulation of gamma-glutamyl transpeptidase and alkaline phosphatase activities in immortalized rat brain microvessel endothelial cells. J Cell Physiol 159, 101–13.
Rubin, L. L., Hall, D. E., Porter, S., Barbu, K., Cannon, C., Horner, H. C., Janatpour, M., Liaw, C. W., Manning, K., Morales, J. & et al. (1991). A cell culture model of the blood-brain barrier. J Cell Biol 115, 1725–35.
Torok, M., Huwyler, J., Gutmann, H., Fricker, G. & Drewe, J. (2003). Modulation of transendothelial permeability and expression of ATP-binding cassette transporters in cultured brain capillary endothelial cells by astrocytic factors and cell-culture conditions. Exp Brain Res 153, 356–65.
Hoheisel, D., Nitz, T., Franke, H., Wegener, J., Hakvoort, A., Tilling, T. & Galla, H. J. (1998). Hydrocortisone reinforces the blood-brain barrier properties in a serum free cell culture system. Biochem Biophys Res Commun 244, 312–6.
Brown, R. C., Morris, A. P. & O’Neil, R. G. (2007). Tight junction protein expression and barrier properties of immortalized mouse brain microvessel endothelial cells. Brain Res 1130, 17–30.
Goldstein, G. W., Wolinsky, J. S., Csejtey, J. & Diamond, I. (1975). Isolation of metaÂbolically active capillaries from rat brain. J Neurochem 25, 715–7.
Dallaire, L., Tremblay, L. & Beliveau, R. (1991). Purification and characterization of metabolically active capillaries of the blood-brain barrier. Biochem J 276 (Pt 3), 745–52.
Betz, A. L. & Goldstein, G. W. (1986). Specialized properties and solute transport in brain capillaries. Annu Rev Physiol 48, 241–50.
Miller, D. S., Nobmann, S. N., Gutmann, H., Toeroek, M., Drewe, J. & Fricker, G. (2000). Xenobiotic transport across isolated brain microvessels studied by confocal microscopy. Mol Pharmacol 58, 1357–67.
Li, J. Y., Boado, R. J. & Pardridge, W. M. (2001). Blood-brain barrier genomics. J Cereb Blood Flow Metab 21, 61–8.
Ohtsuki, S., Yamaguchi, H., Katsukura, Y., Asashima, T. & Terasaki, T. (2008). mRNA expression levels of tight junction protein genes in mouse brain capillary endothelial cells highly purified by magnetic cell sorting. J Neurochem 104, 147–54.
Yousif, S., Marie-Claire, C., Roux, F., Scherrmann, J. M. & Decleves, X. (2007). Expression of drug transporters at the Âblood-brain barrier using an optimized isolated rat brain microvessel strategy. Brain Res 1134, 1–11.
Moro, V., Kacem, K., Springhetti, V., Seylaz, J. & Lasbennes, F. (1995). Microvessels isolated from brain: localization of muscarinic sites by radioligand binding and immunofluorescent techniques. J Cereb Blood Flow Metab 15, 1082–92.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2011 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Vernon, H., Clark, K., Bressler, J.P. (2011). In Vitro Models to Study the Blood Brain Barrier. In: Costa, L., Giordano, G., Guizzetti, M. (eds) In Vitro Neurotoxicology. Methods in Molecular Biology, vol 758. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-170-3_10
Download citation
DOI: https://doi.org/10.1007/978-1-61779-170-3_10
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-61779-169-7
Online ISBN: 978-1-61779-170-3
eBook Packages: Springer Protocols