title: Transient suppression of AHR activity in early red seabream embryos does not prevent the disruption of peripheral nerve projection by 2,3,7,8-tetrachlorodibenzo-p-dioxin.
abstract: The toxicity of dioxins such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is mainly mediated by an aryl hydrocarbon receptor (AHR), which regulates the transcription of multiple target genes including cytochrome P450 1A (CYP1A). Our pervious study identified the presence of TCDD-induced defects of peripheral nerve projection in red seabream (Pagrus major) embryos. However, it remains unclear whether the TCDD-induced peripheral neurotoxicity is mediated by the AHR. To assess the contribution of the red seabream AHR (rsAHR) signaling pathway to the neuronal toxicity, red seabream embryos at 10h post-fertilization (hpf) were treated for 80 min with TCDD (0, 0.3, 5.3, and 37 nM in seawater) alone or in combination with CH223191 (500 nM in seawater), which is an AHR antagonist. A preliminary in vitro reporter gene assay confirmed that TCDD-induced transcriptional activity via rsAHR1 and rsAHR2 was suppressed by CH223191 treatment in a dose-dependent manner. CYP1A mRNA expression in embryos was determined by 2-step real time quantitative-polymerase chain reaction at 24 and 120 hpf and in situ hybridization at 48, 72, 96 and 120 hpf. The morphology of the peripheral nerve system (PNS) was also microscopically observed by fluorescent staining using an anti-acetylated tubulin antibody at 120 hpf. CYP1A mRNA expression was dose-dependently induced by TCDD at all of the examined developing stages. The suppression of TCDD-induced CYP1A expression by CH223191 treatment was observed in embryos at 24 and 48 hpf, while the effect of the rsAHR antagonist disappeared at 96 and 120 hpf. This phenomenon indicated the transient suppression of TCDD-induced rsAHR activation by CH223191 treatment. The immunostaining of peripheral nerves at 120 hpf demonstrated that the projections of the craniofacial nerve were altered in TCDD-treated embryos, and the frequency of TCDD-induced abnormal projections was not prevented by co-treatment with CH223191. These results indicate that the transient suppression of TCDD-induced rsAHR activation during the early developing stages of the red seabream does not influence the abnormal projection of peripheral nerves. In conclusion, transient rsAHR activation in the early stages of development is not involved in the neurotoxicity.
authors.affiliation: Center for Marine Environmental Studies (CMES), Ehime University, Bunkyo-cho 2-5, Matsuyama 790-8577, Japan.
authors.forename: Midori
authors.initials: M
authors.lastname: Iida
authors.affiliation: Department of Life and Nanopharmaceutical Science, Kyung Hee University, Seoul 130-701, Republic of Korea; Department of Biology, Kyung Hee University, Seoul 130-701, Republic of Korea.
authors.forename: Su-Min
authors.initials: SM
authors.lastname: Bak
authors.affiliation: Graduate School of Science and Engineering, Ehime University, Matsuyama 790-8577, Japan.
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Murakami
authors.affiliation: Department of Life and Nanopharmaceutical Science, Kyung Hee University, Seoul 130-701, Republic of Korea; Department of Biology, Kyung Hee University, Seoul 130-701, Republic of Korea.
authors.forename: Eun-Young
authors.initials: EY
authors.lastname: Kim
authors.affiliation: Center for Marine Environmental Studies (CMES), Ehime University, Bunkyo-cho 2-5, Matsuyama 790-8577, Japan. Electronic address: iwatah@agr.ehime-u.ac.jp.
authors.forename: Hisato
authors.initials: H
authors.lastname: Iwata
mesh_terms: D000818:Animals; D001391:Azo Compounds / Q000494:pharmacology; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects; D017933:Peripheral Nervous System / Q000187:drug effects*; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity*; D011485:Protein Binding / Q000187:drug effects; D011720:Pyrazoles / Q000494:pharmacology; D018336:Receptors, Aryl Hydrocarbon / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D021541:Sea Bream / Q000196:embryology* / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014874:Water Pollutants, Chemical / Q000633:toxicity*
chemical_list: C511621:2-methyl-2H-pyrazole-3-carboxylic acid (2-methyl-4-o-tolylazophenyl)amide; D001391:Azo Compounds; D000072317:Polychlorinated Dibenzodioxins; D011720:Pyrazoles; D018336:Receptors, Aryl Hydrocarbon; D014874:Water Pollutants, Chemical; D019363:Cytochrome P-450 CYP1A1
keywords: Aryl hydrocarbon receptor; CH223191; Neurotoxicity; Red seabream; TCDD