Cytochromes P450 involved in the bioactivation of cocaine in the rat.

Persistent Link:
http://hdl.handle.net/10150/187215
Title:
Cytochromes P450 involved in the bioactivation of cocaine in the rat.
Author:
Poet, Torka Sue.
Issue Date:
1995
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Cytochrome P450 enzyme inducters and inhibitors were used to elucidate the isoforms involved in the bioactivation of cocaine in the rat. Phenobarbital and dexamethasone induction were used to induce P450s 2B and 3A, among others, respectively. Isoform-specific inhibitions of P450s 2B and 3A were then used to determine the relative contributions of these enzymes toward cocaine bioactivation. The site-specific hydroxylation of androstenedione was compared to cocaine toxicity in rat liver slices and cocaine and norcocaine toxicity in isolated hepatocytes. Cocaine toxicity determined in rat liver slices, and androstenedione 16$\beta$-hydroxylations in microsomes prepared from coincubated slices, revealed a correlation between P450 2B marker activities and cocaine-mediated toxicity following inhibition of P450 2B activities by chloramphenicol. While cocaine N-demethylation rates were increased by both phenobarbital- and dexamethasone-induction, toxicity in isolated hepatocytes from phenobarbital-induced rats were much greater than in hepatocytes from dexamethasone-induced rats. Further, P450 2B inactivation by chloramphenicol protected against cocaine- and norcocaine-mediated toxicity in hepatocytes from phenobarbital-induced rats, whereas P450 3A inhibition by troleandomycin had no protective effects. While P450s 3A have been observed by other researchers to be important to the oxidative metabolism of cocaine in mice and humans, the importance of P450s 3A in rats is shown here to be limited to the first oxidation step of cocaine which produces norcocaine. Strong differences in the abilities P450s 2B to N-demethylate cocaine in other species were observed. Phenobarbital induction of rats and dogs resulted in increased microsomal cocaine N-demethylation rates commensurate with results from reconstituted purified 2B enzymes from these species. Cocaine N-demethylation rates were low in purified rabbit 2B4 and expressed human 2B6 enzyme was unable to oxidize cocaine. In the guinea pig, which constitutively express P450 2B but is only moderately induced by phenobarbital treatment, the rate of cocaine N-demethylation was high in microsomes from non-induced animals and was not remarkably increased by phenobarbital induction. In agreement with this, neither cocaine nor norcocaine exerted any toxicity in lymphoblastoid cells expressing human P450 2B6.
Type:
text; Dissertation-Reproduction (electronic)
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Pharmacology and Toxicology; Graduate College
Degree Grantor:
University of Arizona
Committee Chair:
Halpert, James R.

Full metadata record

DC FieldValue Language
dc.language.isoenen_US
dc.titleCytochromes P450 involved in the bioactivation of cocaine in the rat.en_US
dc.creatorPoet, Torka Sue.en_US
dc.contributor.authorPoet, Torka Sue.en_US
dc.date.issued1995en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractCytochrome P450 enzyme inducters and inhibitors were used to elucidate the isoforms involved in the bioactivation of cocaine in the rat. Phenobarbital and dexamethasone induction were used to induce P450s 2B and 3A, among others, respectively. Isoform-specific inhibitions of P450s 2B and 3A were then used to determine the relative contributions of these enzymes toward cocaine bioactivation. The site-specific hydroxylation of androstenedione was compared to cocaine toxicity in rat liver slices and cocaine and norcocaine toxicity in isolated hepatocytes. Cocaine toxicity determined in rat liver slices, and androstenedione 16$\beta$-hydroxylations in microsomes prepared from coincubated slices, revealed a correlation between P450 2B marker activities and cocaine-mediated toxicity following inhibition of P450 2B activities by chloramphenicol. While cocaine N-demethylation rates were increased by both phenobarbital- and dexamethasone-induction, toxicity in isolated hepatocytes from phenobarbital-induced rats were much greater than in hepatocytes from dexamethasone-induced rats. Further, P450 2B inactivation by chloramphenicol protected against cocaine- and norcocaine-mediated toxicity in hepatocytes from phenobarbital-induced rats, whereas P450 3A inhibition by troleandomycin had no protective effects. While P450s 3A have been observed by other researchers to be important to the oxidative metabolism of cocaine in mice and humans, the importance of P450s 3A in rats is shown here to be limited to the first oxidation step of cocaine which produces norcocaine. Strong differences in the abilities P450s 2B to N-demethylate cocaine in other species were observed. Phenobarbital induction of rats and dogs resulted in increased microsomal cocaine N-demethylation rates commensurate with results from reconstituted purified 2B enzymes from these species. Cocaine N-demethylation rates were low in purified rabbit 2B4 and expressed human 2B6 enzyme was unable to oxidize cocaine. In the guinea pig, which constitutively express P450 2B but is only moderately induced by phenobarbital treatment, the rate of cocaine N-demethylation was high in microsomes from non-induced animals and was not remarkably increased by phenobarbital induction. In agreement with this, neither cocaine nor norcocaine exerted any toxicity in lymphoblastoid cells expressing human P450 2B6.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplinePharmacology and Toxicologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.chairHalpert, James R.en_US
dc.contributor.committeememberBrendel, Klausen_US
dc.contributor.committeememberGandolfi, A. Jayen_US
dc.contributor.committeememberLaird, Hugh E.en_US
dc.contributor.committeememberRegan, John W.en_US
dc.identifier.proquest9603361en_US
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