title: Quantitative PCR analysis of CYP1A induction in Atlantic salmon (Salmo salar).
abstract: Environmental pollutants are hypothesized to be one of the causes of recent declines in wild populations of Atlantic salmon (Salmo salar); across Eastern Canada and the United States. Some of these pollutants, such as polychlorinated biphenyls and dioxins, are known to induce expression of the CYP1A subfamily of genes. We applied a highly sensitive technique, quantitative reverse transcription-polymerase chain reaction (RT-PCR), for measuring the levels of CYP1A induction in Atlantic salmon. This assay was used to detect patterns of CYP1A mRNA levels, a direct measure of CYP1A expression, in Atlantic salmon exposed to pollutants under both laboratory and field conditions. Two groups of salmon were acclimated to 11 and 17 degrees C, respectively. Each subject then received an intraperitoneal injection (50 mg kg(-1)) of either beta-naphthoflavone (BNF) in corn oil (10 mg BNF ml(-1) corn oil) or corn oil alone. After 48 h, salmon gill, kidney, liver, and brain were collected for RNA isolation and analysis. All tissues showed induction of CYP1A by BNF. The highest base level of CYP1A expression (2.56 x 10(10) molecules/microg RNA) was found in gill tissue. Kidney had the highest mean induction at five orders of magnitude while gill tissue showed the lowest mean induction at two orders of magnitude. The quantitative RT-PCR was also applied to salmon sampled from two streams in Massachusetts, USA. Salmon liver and gill tissue sampled from Millers River (South Royalston, Worcester County), known to contain polychlorinated biphenyls (PCBs), showed on average a two orders of magnitude induction over those collected from a stream with no known contamination (Fourmile Brook, Northfield, Franklin County). Overall, the data show CYP1A exists and is inducible in Atlantic salmon gill, brain, kidney, and liver tissue. In addition, the results obtained demonstrate that quantitative PCR analysis of CYP1A expression is useful in studying ecotoxicity in populations of Atlantic salmon in the wild.
authors.affiliation: Department of Fisheries and Wildlife, 13 Natural Resources Building, Michigan State University, East Lansing, MI 48824, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: B  Rees
authors.forename: Stephen D
authors.initials: SD
authors.lastname: McCormick
authors.forename: John P
authors.initials: JP
authors.lastname: Vanden Heuvel
authors.forename: Weiming
authors.initials: W
authors.lastname: Li
mesh_terms: D000818:Animals; D017756:Aquaculture; D001483:Base Sequence; D019363:Cytochrome P-450 CYP1A1 / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics; D004147:Dioxins / Q000009:adverse effects; D004781:Environmental Exposure; D004785:Environmental Pollutants / Q000009:adverse effects; D004790:Enzyme Induction / Q000187:drug effects; D005880:Gills / Q000201:enzymology; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D009206:Myocardium / Q000201:enzymology; D011078:Polychlorinated Biphenyls / Q000009:adverse effects; D012333:RNA, Messenger / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D020317:Salmo salar / Q000378:metabolism*; D014018:Tissue Distribution; D014874:Water Pollutants, Chemical / Q000009:adverse effects*
chemical_list: D004147:Dioxins; D004785:Environmental Pollutants; D012333:RNA, Messenger; D014874:Water Pollutants, Chemical; D011078:Polychlorinated Biphenyls; D019363:Cytochrome P-450 CYP1A1