title: Role of zebrafish cytochrome P450 CYP1C genes in the reduced mesencephalic vein blood flow caused by activation of AHR2.
abstract: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) causes various signs of toxicity in early life stages of vertebrates through activation of the aryl hydrocarbon receptor (AHR). We previously reported a sensitive and useful endpoint of TCDD developmental toxicity in zebrafish, namely a decrease in blood flow in the dorsal midbrain, but downstream genes involved in the effect are not known. The present study addressed the role of zebrafish cytochrome P450 1C (CYP1C) genes in association with a decrease in mesencephalic vein (MsV) blood flow. The CYP1C subfamily was recently discovered in fish and includes the paralogues CYP1C1 and CYP1C2, both of which are induced via AHR2 in zebrafish embryos. We used morpholino antisense oligonucleotides (MO or morpholino) to block initiation of translation of the target genes. TCDD-induced mRNA expression of CYP1Cs and a decrease in MsV blood flow were both blocked by gene knockdown of AHR2. Gene knockdown of CYP1C1 by two different morpholinos and CYP1C2 by two different morpholinos, but not by their 5 nucleotide-mismatch controls, was effective in blocking reduced MsV blood flow caused by TCDD. The same CYP1C-MOs prevented reduction of blood flow in the MsV caused by β-naphthoflavone (BNF), representing another class of AHR agonists. Whole-mount in situ hybridization revealed that mRNA expression of CYP1C1 and CYP1C2 was induced by TCDD most strongly in branchiogenic primordia and pectoral fin buds. In situ hybridization using head transverse sections showed that TCDD increased the expression of both CYP1Cs in endothelial cells of blood vessels, including the MsV. These results indicate a potential role of CYP1C1 and CYP1C2 in the local circulation failure induced by AHR2 activation in the dorsal midbrain of the zebrafish embryo.
authors.affiliation: Department of Toxicology, School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu 069-8501, Japan. akubota@whoi.edu
authors.forename: Akira
authors.initials: A
authors.lastname: Kubota
authors.forename: John J
authors.initials: JJ
authors.lastname: Stegeman
authors.forename: Bruce R
authors.initials: BR
authors.lastname: Woodin
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Iwanaga
authors.forename: Ryo
authors.initials: R
authors.lastname: Harano
authors.forename: Richard E
authors.initials: RE
authors.lastname: Peterson
authors.forename: Takeo
authors.initials: T
authors.lastname: Hiraga
authors.forename: Hiroki
authors.initials: H
authors.lastname: Teraoka
mesh_terms: D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental; D008636:Mesencephalon / Q000098:blood supply*; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity; D014176:Protein Biosynthesis / Q000187:drug effects; D018336:Receptors, Aryl Hydrocarbon / Q000502:physiology*; D012039:Regional Blood Flow; D015027:Zebrafish / Q000196:embryology / Q000235:genetics*; D029961:Zebrafish Proteins / Q000502:physiology*; D019324:beta-Naphthoflavone / Q000633:toxicity
chemical_list: C474372:AHR2 protein, zebrafish; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D029961:Zebrafish Proteins; D019324:beta-Naphthoflavone; D003577:Cytochrome P-450 Enzyme System