Biochemical Pharmacology, Vol. 31, No. 15, pp. 2523-2529. 1982 Printed in Great Britain. 0~10~2952/82/152523~7 $03.0010 0 1982 Pergamon Press Ltd. zyxwvut INDUCTION OF LIVER MICROSOMAL CYTOCHROME P-450 AND ASSOCIATED MONOOXYGENASES BY OCTACHLOROSTYRENE IN INBRED STRAINS OF MICE LACK OF CORRELATION WITH THE MURINE zyxwvutsrqponmlkjihgfedcbaZY Ah LOCUS J@RN A. HOLME and ERIK DYBING Department of Toxicology, National Institute of Public Health, Postuttak. Oslo 1, Norway (Receiued 19 November 1981; accepred 3 March 1982) Abstract-Single intraperitoneal injections of octachlorostyrene (OCS) and hexachlorobenzene in genetically polycyclic aromatic hydrocarbon ‘responsive’ C57iBLi6 (B6) mice led to a time- and dose- dependent increase in the levels of liver microsomal cytochromes P-450 and bs as well as in the activities of NADPH cytochrome P-450 (cytochrome c) reductase, ethylmorphine (EM) N-demethylase, 4- nitroanisole (PNA) 0-demethylase and acetanilide 4-hydroxylase (AcA hydroxylase). No, or only a very moderate, increase in the activity of aryl hydrocarbon hydroxylase was seen after OCS and HCB, respectively. Pretreatments with phenobarbital (PB) or 3-methylcholanthrene (MC) both increased AcA hydroxylase activity to a similar degree, whereas pretreatment with polychlorinated biphenyls (Aroclor 1254) had an effect equal to the sum of PB and MC. Judged from sodium dodecylsulfate polyacrylamide gel electrophoresis studies, OCS and HCB predominantly increased a microsomal polypeptide of apparent mol. wt 52,000, similar to PB. A reduced response was seen after OCS or HCB treatment of aromatic hydrocarbon ‘non-responsive’ DBAR (D2) mice compared to B6 mice, both with respect to AcA hydroxylase as well as EM demethylase and PNA demethylase activities. OCS treatment of B6D2Fl mice resulted in a doubling of AcA hydroxylase activity, but in mice of the (B6D2)D2 backcross no distinct subgroupings of individual AcA hydroxylase activities were apparent. These results demonstrate that OCS is an inducer of the PB-type in mice and that induction of AcA hydroxylase by OCS is not regulated by the Ah locus. Octachlorostyrene (OCS; Fig. 1) is a byproduct in the manufacture of many chlorinated hydrocarbons. Together with hexachlorobenzene (HCB)*, OCS has been found to be a major organochlorine contami- nant in fish caught in the Grenlandfjord region in Norway [l, 2] and in the Great Lakes in the U.S.A. [31. Like HCB, a structurally related chlorinated hydrocarbon with a demonstrated carcinogenic effect in laboratory animals [4,5], OCS has been shown to cause liver hypertrophy, megalocytosis, porphy- ria, increased heme biosynthesis as well as induction of liver microsomal cytochrome P-450 and associated monooxygenases in rats [6,7]. Several reports have stated that HCB is a mixed type of inducer, giving a pattern of induced micro- somal enzyme activities similar to the sum of the effects of PB- and MC-type of inducers [8-10]. In a recent work [7] we found that intraperitoneal or dietary administration of OCS or HCS to rats of both sexes seemed to increase primarily the PB-types of the cytochromes. AcA has been used by Guenthner and Nebert [ll] * Abbreviations used: PCB, polychlorinated biphenyls (Aroclor 1254); HCB, hexachlorobenzene; OCS, octach- lorostyrene; PB, phenobarbital; MC, 3-methylcholan- threne; cytochrome P-450, a collective term for all forms of the cytochromes P-450; EM, ethylmorphine; PNA, 4- nitroanisole; AcA, acetanilide; AHH, aryl hydrocarbon (benzo[a]pyrene) hydroxylase; B6, the inbred C57BLi6J mouse strain; D2, the inbred DBARJ mouse strain. as a selective substrate for cytochrome P-448 metab- olism. However, PB-treatment has also been found to increase AcA hydroxylase activity [7]. Therefore, it was not possible to decide whether the increase observed in the AcA hydroxylase activity after OCS- and HCB-treatment was due to an increase in a MC-type or PB-type of cytochrome. Studies with crosses and backcrosses of mice showing genetic differences in aromatic hydrocarbon responsiveness (for review see Ref. 12) might answer this question. It was therefore of interest to characterize the effects of OCS on the cytochrome P-450 system in such mice. MATERIALSAND METHODS Chemicals. OCS was a generous gift from Norsk Hydro (Porsgrunn, Norway). The compound was at least 98% pure as judged by gas chromatography on a Varian 1400 instrument equipped with the electrochemical detector. [‘4C]Acetanilide (uni- formly ring-labelled, sp. act. 10.5 mCi/mmole) was purchased from California Bionuclear Corporation LI 1 LI CI Fig. 1. Octachlorostyrene. 2523