Abstract
Studies initiated to investigate the distribution of cytochrome P450 2B (CYP2B) isoenzymes in rat brain cells revealed significant activity of CYP2B-dependent 7-pentoxyresorufin-O-dealkylase (PROD) in microsomes prepared from both, cultured rat brain neuronal and glial cells. Neuronal cells exhibited 2-fold higher activity of PROD than the glial cells. RT-PCR and immunocytochemical studies demonstrated significant constitutive mRNA and protein expression of CYP2B in cultured neuronal and glial cells. Induction studies with phenobarbital (PB), a known CYP2B inducer, revealed significant concentration dependent increase in the activity of PROD in cultured brain cells with glial cells exhibiting greater magnitude of induction than the neuronal cells. This difference in the increase in enzyme activity was also observed with RT-PCR and immunocytochemical studies indicating differences in the induction of CYP2B1 and 2B2 mRNA as well as protein expression in the cultured brain cells. Furthermore, a greater magnitude of induction was observed in CYP2B2 than CYP2B1 in the brain cells. Our data indicating differences in the expression and sensitivity of the CYP2B isoenzymes in cultured rat brain cells will help in identifying and distinguishing xenobiotic metabolizing capability of these cells and understanding the vulnerability of the specific cell types toward neurotoxins.
Similar content being viewed by others
References
Guengerich FP, Shimada T, Umbenhauer DR et al (1986) Structure and function of cytochrome P-450. Adv Exp Med Biol 197:83–94
Dannan GA, Guengerich FP, Kaminsky LS et al (1983) Regulation of cytochrome P-450. Immunochemical quantitation of eight isozymes in liver microsomes of rats treated with polybrominated biphenyl congeners. J Biol Chem 258:1282–1288
Christou M, Wilson NM, Jefcoate CR (1987) Expression and function of three cytochrome P-450 isozymes in rat extrahepatic tissues. Arch Biochem Biophys 258:519–534
Waxman DJ, Azaroff L (1992) Phenobarbital induction of cytochrome P-450 gene expression. Biochem J 281:577–592
Kapitulnik J, Gelboin HV, Guengerich FP et al (1987) Immunohistochemical localization of cytochrome P-450 in rat brain. Neurosci 20:829–833
Kohler C, Eriksson LG, Hansson T et al (1988) Immunohistochemical localization of cytochrome P-450 in the rat brain. Neurosci Lett 84:109–114
Naslund BM, Glaumann H, Warner M et al (1988) Cytochrome P-450 b and c in the rat brain and pituitary gland. Mol Pharmacol 33:31–37
Hodgson AV, White TB, White JW et al (1993) Expression analysis of the mixed function oxidase system in rat brain by the polymerase chain reaction. Mol Cell Biochem 120:171–179
Strobel HW, Thompson CM, Antonovic L (2001) Cytochromes P450 in brain: function and significance. Curr Drug Metab 2:199–214
Parmar D, Dayal M, Seth PK (2003) Expression of cytochrome P450s (P450s) in brain: physiological, pharmacological and toxicological consequences. Proc Ind Nat Acad Sci 6:905–928
Parmar D, Dhawan A, Seth PK (1998) Evidence for O-dealkylation of 7-pentoxyresorufin by cytochrome P450 2B1/2B2 isoenzymes in brain. Mol Cell Biochem 189:201–205
Ravindranath V, Anandatheerthavarada HK, Shankar SK (1989) Xenobiotic metabolism in human brain—presence of cytochrome P-450 and associated mono-oxygenases. Brain Res 496:331–335
Anandatheerthavarada HK, Shankar SK, Ravindranath V (1990) Rat brain cytochromes P-450: catalytic, immunochemical properties and inducibility of multiple forms: preparation of brain microsomes with cytochrome P450 activity using calcium aggregation method NADPH cytochrome P-450 reductase in rat, mouse and human brain. Brain Res 536:339–343
Farin FM, Omiecinski CJ (1993) Regiospecific expression of cytochrome P-450s and microsomal epoxide hydrolase in human brain tissue. J Toxicol Environ Health 40:317–335
Schilter B, Omiecinski CJ (1993) Regional distribution and expression modulation of cytochrome P-450 and epoxide hydrolase mRNAs in the rat brain. Mol Pharmacol 44:990–996
Hedlund E, Gustafsson JA, Warner M (1998) Cytochrome P450 in the brain: 2B or not 2B. Trends Pharmacol Sci 19:82–85
Ghersi-Egea JF, Leninger-Muller B, Suleman G et al (1994) Localization of drug-metabolizing enzyme activities to blood-brain interfaces and circumventricular organs. J Neurochem 62:1089–1096
Miksys S, Hoffmann E, Tyndale RF (2000) Regional and cellular induction of nicotine-metabolizing CYP2B1 in rat brain by chronic nicotine treatment. Biochem Pharmacol 59:1501–1511
Upadhya SC, Chinta SJ, Pai HV et al (2002) Toxicological consequences of differential regulation of cytochrome P450 isoforms in rat brain regions by phenobarbital. Arch Biochem Biophys 399:56–65
Dhawan A, Parmar D, Das M et al (1990) Cytochrome P-450 dependent monooxygenases in neuronal and glial cells: inducibility and specificity. Biochem Biophys Res Commun 170:441–447
Kapoor N, Pant AB, Dhawan A et al (2006) Differences in sensitivity of cultured rat brain neuronal and glial cytochrome P450 2E1 to ethanol. Life Sci 79:1514–1522
Warner M, Kohler C, Hansen T et al (1988) Regional distribution of cytochrome P450 in the rat brain: spectral quatification and contribution of P450b,e and P450c,d. J Neurochem 50:1057–1065
Johri A, Dhawan A, Singh RL et al (2006) Effect of prenatal exposure of deltamethrin on the ontogeny of xenobiotic metabolizing Cytochrome P450s in the brain and liver of offsprings. Tox Appl Pharmacol 214:279–289
Omiecinski CJ, Hassett C, Costa P (1990) Developmental expression and in situ localization of the phenobarbital inducible rat hepatic mRNAs for the cytochromes CYP2B1, 2B2, 2C6 and 3A1. Mol Pharmacol 38:462–470
Soh Y, Rhee HM, Dong HS et al (1996) Immunological detection of CYP2E1 in fresh rat lymphocytes and its pretranslational induction by fasting. Biochem Biophys Res Commun 227:541–546
Kempermann G, Knoth R, Gebicke-Haerter PJ et al (1994) Cytochrome P450 in rat astrocytes in vivo and in vitro: intracellular localization and induction by phenytoin. J Neurosci Res 39:576–588
Ravindranath V (1998) Metabolism of xenobiotic in the central nervous system: implications and challenges. Biochem Pharmacol 56:547–551
Baes M, Gulick T, Choi HS et al (1994) A new orphan member of the nuclear hormone receptor superfamily that interacts with a subset of retinoic acid response elements. Mol Cell Biol 14:1544–1552
Choi HS, Chung M, Tzameli I et al (1997) Differential transactivation by two isoforms of the orphan nuclear hormone receptor CAR. J Biol Chem 272:23565–23571
Souidi M, Gueguen Y, Linard C et al (2005) In vivo effects of chronic contamination with depleted uranium on CYP3A and associated nuclear receptors PXR and CAR in the rat. Toxicol 214:113–122
Rosenbrock H, Hagemeyer CE, Ditter M et al (2001) Expression and localization of the CYP2B subfamily predominantly in neurones of rat brain. J Neurochem 76:332–340
Volk B, Meyer RP, von Lintig F et al (1995) Localization and characterization of cytochrome P450 in the brain. In vivo and in vitro investigations on phenytoin- and phenobarbital-inducible isoforms. Toxicol Lett 82–83:655–662
Lucas D, Berthou F, Girre C et al (1993) High-performance liquid chromatographic determination of chlorzoxazone and 6-hydroxychlorzoxazone in serum: a tool for indirect evaluation of cytochrome P4502E1 activity in humans. J Chromatogr 622:79–86
Roberge C, Beaudet MJ, Anderson A (2004) GABA(A)/central benzodiazepine receptor and peripheral benzodiazepine receptor ligands as inducers of phenobarbital-inducible CYP2B and CYP3A. Biochem Pharmacol 68:1383–1389
Schilter B, Andersen MR, Acharya C et al (2000) Activation of cytochrome P450 gene expression in the rat brain by phenobarbital-like inducers. J Pharmacol Exp Ther 294:916–922
Ibach B, Appel K, Gebicke-Haerter P et al (1998) Effect of phenytoin on cytochrome P450 2B mRNA expression in primary rat astrocyte cultures. J Neurosci Res 54:402–411
Makar TK, Nedergaard M, Preuss A et al (1994) Vitamin E, ascorbate, glutathione, glutathione disulfide, and enzymes of glutathione metabolism in cultures of chick astrocytes and neurons: evidence that astrocytes play an important role in antioxidative processes in the brain. J Neurochem 62:45–53
Meyer RP, Knoth R, Schiltz E et al (2001) Possible function of astrocyte cytochrome P450 in control of xenobiotic phenytoin in the brain: in vitro studies on murine astrocyte primary cultures. Exp Neurol 167:376–384
Hagemeyer CE, Rosenbrock H, Ditter M et al (2003) Predominantly neuronal expression of cytochrome P450 isoforms CYP3A11 and CYP3A13 in mouse brain. Neuroscience 117:521–529
Acknowledgment
Authors are grateful to Director, ITRC for his keen interest in the work. The financial assistance of Council of Science & Technology, U.P and CSIR (CMM-0018) for carrying out the above studies is gratefully acknowledged. The help of Dr. Pankaj Seth, National Brain Research Centre (NBRC), Manesar, Gurgaon, India, in the immunocytochemical analysis is also gratefully acknowledged. The technical assistance of Mr. B.S. Pandey and Mr. Rajesh Misra is gratefully acknowledged. ITRC Manuscript Communication Number: 2458.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Kapoor, N., Pant, A.B., Dhawan, A. et al. Differences in the expression and inducibility of cytochrome P450 2B isoenzymes in cultured rat brain neuronal and glial cells. Mol Cell Biochem 305, 199–207 (2007). https://doi.org/10.1007/s11010-007-9544-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11010-007-9544-0