Page 186 - The Toxicology of Fishes

166 The Toxicology of Fishes

some tropical fishes from the Bermuda Archipelago; however, great differences were found in aminopy-
rene-N-demethylase and pentoxyresorufin-O-depentylase activities among species, with the Bermuda
chub (Kyphosous sectatrix) and sergeant major (Abudefduf saxatilis) displaying the highest activities. In
addition, polyclonal antibody (pAb) against rat CYP2B1 cross-reacted most strongly with hepatic
microsomal proteins in tomtate (Haemulon aurolineatum), pinfish (Lagodan rhomboides), Bermuda
chub, and sergeant major (Stegeman et al., 1997). The reason for the observed differences in CYP2B-
like expression among species is not known. Earlier, it was suggested that natural dietary compounds
may be causing these differences, as higher CYP2B-like protein levels were observed in butterfly fish
(Chaetodon capistratus) that consumed gorgonians (containing high levels of allelochemicals) compared
to butterfly fish that avoided gorgonians (Vrolijk et al., 1994). CYP2N mRNA expression in Chaetodon
xanthurus (CYP2N7) was significantly higher than in the facultative coralline-feeding butterfly fish C.
kleini, C. auriga (CYP2N6), or C. vagabundus, as well as an obligate coralline feeding species (C.
punctofasciatus) from the Great Barrier Reef in Australia (DeBusk, 2001). When each species, including
C. xanthurus, was gavaged with gorgonian extracts from Sinnularia maxima for 3 days, with the exception
of C. punctofasciatus, CYP2N mRNA expression was diminished (DeBusk, 2001). In the Bermuda
species investigated, herbivorous fish had higher CYP levels (including CYP2B-like proteins) compared
to carnivorous fish (Stegeman et al., 1997). It remains to be shown if natural dietary chemicals may act
as inducers of CYP2B-like forms in fish or if other mechanisms are involved.
Phenobarbital (PB) and 1,4-bis(2-[3,5-dichloropyridyloxy])benzene (TCPOBOP) are powerful PB-
type inducers of CYP2B genes in mammals (Poland et al., 1981). In mammals, induction of CYP2B by
PB-type inducers proceeds through activation of the constitutive androstane receptor (CAR) followed
by nuclear translocation, dimerization with the retinoid X receptor (RXR), and binding to phenobarbital
response elements (PBREMs) in the promoter region of the CYP2B genes (Honkakoski et al., 1997,
1998a,b).
In fish, however, an apparent lack of response to PB-type inducers has been observed (Buhler and
Williams, 1989; Eisele et al., 1984; Goksøyr et al., 1987; Haasch et al., 1994; Iwata et al., 2002; Stegeman,
1981), although a CAR immunoreactive protein was detected in scup liver cytosol and nucleus using
antibodies against human CAR. No induction of CYP protein levels, including scup P450B, or catalytic
activities were seen in scup injected with TCPOBOB. In fact, TCPOBOB treatment had no effect on
translocation of the cytosolic CAR-immunoreactive protein in scup liver (Iwata et al., 2002). This study
points to functional differences, possibly in receptor activation or translocation, between fish and mam-
mals. Recently, a single piscine CAR/PXR gene was identified (fr078207) when searching the pufferfish
genome; however, this receptor was more related to PXR family members and hence a probable functional
analog of PXR (Maglich et al., 2003). Thus, CAR may have diverged from the pregnane X receptor
(PXR) at a later point in vertebrate evolution, or CAR may have been lost in some or all teleost lines
(Maglich et al., 2003). The apparent lack of a piscine CAR receptor may be one explanation for the
observed lack of PB-type as well as diminished CYP3A (see below) induction in fish.

CYP2E-Like Forms—The possible existence of a CYP2E form in fish (Poeciliopsis monacha-lucida)
was proposed based on hybridization with a rat CYP2E1 49-base oligonucleotide, antibodies to rat
CYP2E1, as well as responsiveness to ethanol treatment. This CYP form was suggested to be involved
in the CYP-mediated dealkylation of the fish carcinogen diethylnitrosamine (Kaplan et al., 1991).
Furthermore, hepatic microsomal metabolism of the mammalian CYP2E substrate chlorzoxazone in
winter flounder (Pleuronectes americanus) and in viviparous Poeciliopsis monacha and Poeciliopsis
viriosa is indicative of the presence of CYP2E-like enzymes (Kaplan et al., 2001; Wall and Crivello,
1998). A piscine CYP2E gene ortholog, however, has so far not been reported.

The CYP2K Subfamily—A CYP protein, denoted LMC2, was isolated from rainbow trout liver (Miranda
et al., 1989). It was subsequently cloned and assigned as CYP2K1 (Buhler et al., 1994). CYP2K1 is
one of the dominant CYP forms expressed in liver and trunk kidney, and it displays sexually dimorphic
expression, with higher levels in sexually mature males compared to females (Buhler et al., 1994). In
addition to liver and trunk kidney, CYP2K1 also is expressed, though at lower levels, in blood cells,
upper intestine, head kidney, stomach, heart, gonads, and male muscle (Cok et al., 1998). Heterologous