Project description:Polychlorinated biphenyls (PCBs) are persistent and ubiquitously distributed environmental pollutants. Based on their chemical structure, PCBs are classified into non-ortho substituted and ortho-substituted congeners. Non-ortho-substituted PCBs are structurally similar to dioxin or TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) and their mode of action and toxic effects are well established. In contrast, much less is known about the effects of ortho-substituted PCBs. Studies conducted so far have focused on tissue-specific effects but there is limited knowledge about the effects on the whole organism, particularly the sensitive developmental stages in vertebrates. Hence, in this study we investigated the effects of exposure to an environmentally relevant ortho-substituted PCB (2,2’,4,4’,5,5’-hexachlorobiphenyl; PCB153) on zebrafish embryos. We exposed zebrafish embryos to either DMSO (0.1%; solvent control) or three different concentrations of PCB153 (0.1, 1 and 10 μM) from 4 hours post-fertilization (hpf) to 120 hpf. At the end of the exposure, larvae were sampled for determining transcriptional changes (RNA sequencing) and the remaining embryos were maintained in contaminant-free environment. At 7 and 14 days post-fertilization (dpf), zebrafish larvae were assessed for locomotory behavior. We did not observe any overt phenotypes during the exposure period, but observed a spinal phenotype in the 10μM PCB153 treated group starting at 7 dpf. This phenotype was observed in a dose-dependent manner and majority of the embryos with this phenotype died by 14 dpf. RNA sequencing of 5 dpf larvae exposed to PCB153 also revealed dose-dependent changes in gene expression patterns. A total of 633, 2227, and 3378 differentially expressed genes were observed in 0.1, 1 and 10 μM PCB153 treated embryos, respectively. Among these, 301 genes were common to all treatment groups, and KEGG pathway analysis revealed enrichment of genes related to circadian rhythm, FOXO signaling and insulin resistance pathways. We are currently investigating the functions of genes that are uniquely altered by different PCB153 concentrations. Overall, these results suggest that developmental exposure to PCB153, a PCB congener highly prevalent in the environment, targets multiple physiological processes including photoperiod regulation and metabolism. [Funded by NIH P01ES021923 and NSF OCE-1314642].

Project description:The phenylpyrazole fipronil is a widely used insecticide designed to inhibit γ -amino-butyric acid (GABA) receptors, the major inhibitory neurotransmitter in the central nervous system. Fipronil has been detected in some water systems in the ng/L range, and is reported to be neurotoxic. To address the risks associated with fipronil exposure, we measured morphological, physiological, and molecular responses in zebrafish (Danio rerio) embryos following a 48 hour exposure (20 ng/L – 2 mg/L). Survival was not different than controls following treatments below 200 µg fipronil/L but was ~20% higher with concentrations above 200 µg fipronil/L. Once the embryos hatched, they underwent a 7 day depuration phase. At 9 days post-fertilization (9 dpf), body length and notochord length were not different than controls for any dose. To assess sub-lethal effects, transcriptome profiling was conducted in 9 dpf larvae following 48 hour exposure + 7 dpf depuration to environmentally relevant concentrations of fipronil (200 ng fipronil/L), as well as two higher concentrations of the pesticide (200 µg fipronil/L and 2 mg fipronil/L). Transcriptome profiling revealed that all three concentrations affected pathways related to chromosome condensation and the metabolism of estrogens and androgens as well as genes related to methylation. In addition, 200 ng fipronil/L down-regulated genes related to the circadian clock, histone and DNA methylation, and histone acetylation, while the highest dose increased networks related to immune function (e.g. lectin-induced complement pathway and the alternative complement pathway). The two highest concentrations of fipronil increased the expression of transcriptional networks associated with mitochondrial respiratory chain dysfunction and mitochondrial protein transport. As such, we exposed 24 hpf embryos to fipronil for 24 hours and measured oxygen consumption rate to assess mitochondrial function. There were no differences in basal and maximal respiration in the embryos nor ATP production, and fipronil did not affect mitochondrial bioenergetics. This study suggests that fipronil at environmentally relevant concentrations does not adversely affect the survival or morphology of fish embryos, however sub-lethal endpoints should be examined to more fully characterize the long term effects of fipronil exposure in larval fish.

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Danio rerio

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