AHR2 required for normal behavioral responses and proper development of the skeletal and reproductive systems in zebrafish

Garcia, Gloria R.; Bugel, Sean M.; Truong, Lisa; Spagnoli, Sean; Tanguay, Robert L.

doi:10.1371/journal.pone.0193484

Cited by 47 publications

(72 citation statements)

References 60 publications

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“…However, analysis of zebrafish behavior is far more variable between laboratories. Nonetheless, consistent with previous reports that TCDD is teratogenic (Carney et al, 2006) and causes abnormal behavior in larval zebrafish (Garcia et al, 2018), we detected TCDD as a hit for both endpoints. Similarly, in accordance with Noyes et al (2015), we identified BDE-99, BDE-47, BPDP, TPhP, TCEP, and TCPP as causing hypoactivity in the photomotor behavior assay at 5 dpf.…”

Section: Comparison Of Hits To Existing Zebrafish Datasupporting

confidence: 92%

Teratological and Behavioral Screening of the National Toxicology Program 91-Compound Library in Zebrafish (Danio rerio)

Dach

Yaghoobi

Schmuck

et al. 2018

Toxicological Sciences

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To screen the tens of thousands of chemicals for which no toxicity data currently exists, it is necessary to move from in vivo rodent models to alternative models, such as zebrafish. Here, we used dechorionated Tropical 5D wild-type zebrafish embryos to screen a 91-compound library provided by the National Toxicology Program (NTP) for developmental toxicity. This library contained 86 unique chemicals that included negative controls, flame retardants, polycyclic aromatic hydrocarbons (PAHs), drugs, industrial chemicals, and pesticides. Fish were exposed to 5 concentrations of each chemical or an equal amount of vehicle (0.5% DMSO) in embryo medium from 6 h post-fertilization (hpf) to 5 days post-fertilization (dpf). Fish were examined daily for mortality and teratogenic effects and photomotor behavior was assessed at 4 and 5 dpf. Of the 5 negative control compounds in the library, none caused mortality/teratogenesis, but two altered behavior. Chemicals provided in duplicate produced similar outcomes. Overall, 13 compounds caused mortality/teratology but not behavioral abnormalities, 24 only affected behavior, and 18 altered both endpoints, with behavior affected at concentrations that did not cause mortality/teratology (55/86 hits). Of the compounds that affected behavior, 52% caused behavioral abnormalities at either 4 or 5 dpf. Compounds within the same functional group caused different behavioral abnormalities, while similar behavioral patterns were caused by compounds from different groups. Our data suggest that behavior is a sensitive endpoint for developmental toxicity screening that integrates multiple modes of toxic action and is influenced by the age of the larval fish at the time of testing.

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Section: Comparison Of Hits To Existing Zebrafish Datasupporting

confidence: 92%

Teratological and Behavioral Screening of the National Toxicology Program 91-Compound Library in Zebrafish (Danio rerio)

Dach

Yaghoobi

Schmuck

et al. 2018

Toxicological Sciences

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“…Maternal zygotic embryos could be derived from homozygous ahr1 single and double mutant parents with natural spawning. In contrast, natural spawning was not observed in ahr2 mutant embryos, consistent with previous studies reporting reduced fertility in ahr2 mutants (Garcia et al, 2018). ahr2 mutant embryos were therefore generated from incrosses of heterozygous ahr2 mutant parents, which demonstrated normal fecundity.…”

Section: Generation Of Aryl Hydrocarbon Receptor Mutant Zebrafishsupporting

confidence: 89%

“…ahr2 mutant embryos were therefore generated from incrosses of heterozygous ahr2 mutant parents, which demonstrated normal fecundity. Previously published zebrafish ahr mutants are highlighted in Figure 1 for comparison (Garcia et al, 2018;Goodale et al, 2012;Sugden et al, 2017).…”

Section: Generation Of Aryl Hydrocarbon Receptor Mutant Zebrafishmentioning

confidence: 99%

“…Zebrafish ahr2 is thought to be the primary functional homolog of the human AHR based on studies using recombinant Ahr2 protein in vitro and ahr2 mutant zebrafish. Ahr2 binds ligands like TCDD, forms a heterodimer with a zebrafish ARNT paralog (arnt2) on XRE DNA and induces expression of AHR target genes like CYP1A1 (Andreasen, Hahn, et al, 2002;Andreasen, Spitsbergen, et al, 2002;Garcia et al, 2018;Goodale et al, 2012;Prasch et al, 2003;Tanguay et al, 1999Tanguay et al, , 2000.…”

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confidence: 99%

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ahr2, But Not ahr1a or ahr1b, Is Required for Craniofacial and Fin Development and TCDD-dependent Cardiotoxicity in Zebrafish

Souder

Gorelick

2019

Toxicological Sciences

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The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that binds environmental toxicants and regulates gene expression. AHR also regulates developmental processes, like craniofacial development and hematopoiesis, in the absence of environmental exposures. Zebrafish have 3 paralogs of AHR: ahr1a, ahr1b, and ahr2. Adult zebrafish with mutations in ahr2 exhibited craniofacial and fin defects. However, the degree to which ahr1a and ahr1b influence ahr2 signaling and contribute to fin and craniofacial development are not known. We compared morphology of adult ahr2 mutants and ahr1a;ahr1b single and double mutant zebrafish. We found that ahr1a;ahr1b single and double mutants were morphologically normal whereas ahr2 mutant zebrafish demonstrated fin and craniofacial malformations. At 5 days post fertilization, both ahr1a;ahr1b and ahr2 mutant larvae were normal, suggesting that adult phenotypes are due to defects in maturation or maintenance. Next, we analyzed the function of zebrafish AHRs activated by environmental ligands. The prototypical AHR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), induces toxicity in humans and rodents via AHR and causes cardiotoxicity in zebrafish embryos. It has been shown that embryos with mutations in ahr2 are resistant to TCDD toxicity, yet it is unclear whether ahr1 receptors are required. Furthermore, though AHR was shown to interact with estrogen receptor alpha following TCDD treatment, it is not known whether this interaction is constitutive or context-dependent. To determine whether estrogen receptors are constitutive cofactors for AHR signaling, we used genetic and pharmacologic techniques to analyze TCDD-dependent toxicity in estrogen receptor and ahr mutant embryos. We found that embryos with mutations in ahr1a;ahr1b or estrogen receptor genes are susceptible to TCDD toxicity whereas ahr2 mutant embryos are TCDD-resistant. Moreover, pharmacologic blockade of nuclear estrogen receptors failed to prevent TCDD toxicity. These findings suggest that ahr1 genes do not have overlapping functions with ahr2 in fin and craniofacial development or TCDD-dependent toxicity, and that estrogen receptors are not constitutive partners of ahr2.

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“…To determine significant differences between three treatment group data were analyzed using a one-way ANOVA with Tukey post hoc test (P-value < 0.05, statistically significant) with GraphPad Prism 4 software (La Jolla, CA). Significant differences were calculated with two-way ANOVA and Tukey post hoc test or a repeated measures two-way ANOVA and Tukey post-hoc test (P-value < 0.05, statistically significant) when both treatment and another condition, like water condition, zone of tank, or a behavioral stimulus, was present 38 . For the shuttle box assay a linear regression was fitted to the data for each treatment to generate initial time and rate of learning graphs, while a separate analysis of variance (AOV) followed by a Tukey's statistical difference was used to calculate statistical significance amongst the groups 34 .…”

Section: Discussionmentioning

confidence: 99%

Nitrate and nitrite exposure increases anxiety-like behavior and alters brain metabolomic profile in zebrafish

García-Jaramillo

Beaver

Truong

et al. 2020

Preprint

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IntroductionDietary nitrate lowers blood pressure and improves athletic performance in humans, yet data supporting observations that it may increase cerebral blood flow and improve cognitive performance are mixed. Here we tested the hypothesis that nitrate and nitrite treatment would improve indicators of learning and cognitive performance in a zebrafish (Danio rerio) model. We also explored the extent to which nitrate and nitrite treatment affected the brain metabolome in order to understand how nitrate and nitrite supplementation may affect indices of cognitive function.MethodsFish were exposed to sodium nitrate (606.9 mg/L), sodium nitrite (19.5 mg/L), or control water for 2-4 weeks and free swim, startle response, innate predator avoidance, social cohesion, and shuttle box assays were performed.ResultsNitrate and nitrite treatment did not change fish weight, length, predator avoidance, or distance and velocity traveled in an unstressed environment. Nitrate- and nitrite-treated fish initially experienced more negative reinforcement and increased time to decision in the shuttle box assay, which is consistent with a decrease in associative learning or executive function however, over multiple trials, all treatment groups demonstrated behaviors associated with learning. Nitrate and nitrite treatment significantly increased anxiety-like behavior but did not alter epinephrine, norepinephrine or dopamine levels. Targeted LC-MS/MS analysis revealed no significant increase in brain nitrate or nitrite concentrations with treatment. An untargeted metabolomics analysis found 47 metabolites whose abundance was significantly altered in the brain with nitrate and nitrite treatment including an 18-19% reduction in the neurotransmitter γ-aminobutyric acid (GABA), and 17-22% reduction in its precursor, glutamine, which may contribute to the increased anxiety-like behavior.ConclusionNitrate and nitrite treatment did not adversely affect multiple parameters of zebrafish health but was associated with mild anxiety-like behavior, changes in the brain metabolome, and caused a short-term decrease in executive function or associative learning.

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AHR2 required for normal behavioral responses and proper development of the skeletal and reproductive systems in zebrafish

Cited by 47 publications

References 60 publications

Teratological and Behavioral Screening of the National Toxicology Program 91-Compound Library in Zebrafish (Danio rerio)

Teratological and Behavioral Screening of the National Toxicology Program 91-Compound Library in Zebrafish (Danio rerio)

ahr2, But Not ahr1a or ahr1b, Is Required for Craniofacial and Fin Development and TCDD-dependent Cardiotoxicity in Zebrafish

Nitrate and nitrite exposure increases anxiety-like behavior and alters brain metabolomic profile in zebrafish

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