Thyroid hormones (THs) regulate neurodevelopment, thus TH disruption is widely posited as a mechanism of developmental neurotoxicity for diverse environmental chemicals. Zebrafish have been proposed as an alternative model for studying the role of TH in developmental neurotoxicity. To realize this goal, it is critical to characterize the normal ontogenetic expression profile of TH signaling molecules in the developing zebrafish and determine the sensitivity of these molecules to perturbations in TH levels. To address these gaps in the existing database, we characterized the transcriptional profiles of TH transporters, deiodinases (DIOs), receptors (TRs), nuclear coactivators (NCOAs), nuclear corepressors (NCORs), and retinoid X receptors (RXRs) in parallel with measurements of endogenous TH concentrations and tshβ mRNA expression throughout the first five days of zebrafish development. Transcripts encoding these TH signaling components were identified and observed to be upregulated around 48-72 h post fertilization (hpf) concurrent with the onset of larval production of T4. Exposure to exogenous T4 and T3 upregulated mct8, dio3-b, trα-a, trβ, and mbp-a levels, and downregulated expression of oatp1c1. Morpholino knockdown of TH transporter mct8 and treatment with 6-propyl-2-thiouracil (PTU) was used to reduce cellular uptake and production of TH, an effect that was associated with downregulation of dio3-b at 120 hpf. Collectively, these data confirm that larval zebrafish express orthologs of TH signaling molecules important in mammalian development and suggest that there may be species differences with respect to impacts of TH disruption on gene transcription.

BackgroundHyperthyroidism is the most common endocrine disorder observed in domestic felines; however, its etiology is largely unknown. Two classes of persistent organic pollutants, polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs) are known to interfere with thyroid hormone (TH) signaling and regulation; thus, it is postulated that they contribute to the etiopathogenesis of feline hyperthyroidism and pose a risk to humans and other species. In this case-control study, the concentrations of 13 PBDE and 11 PCB congeners were measured by gas chromatography mass spectrometry in serum or plasma samples from 20 hyperthyroid and 31 control cats in order to investigate the association between concentration of PBDE and PCB congeners and feline hyperthyroidism. Logistic regression analysis was used to determine whether elevated concentrations of individual congeners were associated with a higher risk of feline hyperthyroidism.ResultsHyperthyroid cats had higher concentrations of four PBDE congeners (BDE17, BDE100, BDE47, and BDE49) and five PCB congeners (PCB131, PCB153, PCB174, PCB180, and PCB196), compared to control cats. In addition, the sum of both PBDE and PCB congener concentrations were elevated in the hyperthyroid group compared to control cats; however, only the increased PCB concentrations were statistically significant. The sum total PBDE concentrations in our feline samples were approximately 50 times greater than concentrations previously reported in human populations from a geographically similar area, whereas sum total PCB concentrations were comparable to those previously reported in humans.ConclusionsThese observational findings support the hypothesis that PBDEs and PCBs may contribute to the etiopathogenesis of hyperthyroidism in felines. As domestic house cats are often exposed to higher concentrations of PBDEs than humans, they may serve as sentinels for the risk of TH disruption that these pollutants pose to humans and other species.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-017-1031-6) contains supplementary material, which is available to authorized users.

High throughput in vitro, in silico, and computational approaches have identified numerous environmental chemicals that interfere with thyroid hormone (TH) activity, and it is posited that human exposures to such chemicals are a contributing factor to neurodevelopmental disorders. However, whether hits in screens of TH activity are predictive of developmental neurotoxicity (DNT) has yet to be systematically addressed. The zebrafish has been proposed as a second tier model for assessing the in vivo DNT potential of TH active chemicals. As an initial evaluation of the feasibility of this proposal, we determined whether an endpoint often used to assess DNT in larval zebrafish, specifically photomotor behavior, is altered by experimentally induced hyper-and hypothyroidism. Developmental hyperthyroidism was simulated by static waterborne exposure of zebrafish to varying concentrations (3-300 nM) of thyroxine (T4) or triiodothyronine (T3) beginning at 6 h post-fertilization (hpf) and continuing through 5 days post-fertilization (dpf). Teratogenic effects and lethality were observed at 4 and 5 dpf in fish exposed to T4 or T3 at concentrations > 30 nM. However, as early as 3 dpf, T4 ( > 3 nM) and T3 ( > 10 nM) significantly increased swimming activity triggered by sudden changes from light to dark, particularly during the second dark period (Dark 2). Conversely, developmental hypothyroidism, which was induced by treatment with 6-propyl-2-thiouracil (PTU), morpholino knockdown of the TH transporter mct8, or ablation of thyroid follicles in adult females prior to spawning, generally decreased swimming activity during dark periods, although effects did vary across test days. All effects of developmental hypothyroidism on photomotor behavior occurred independent of teratogenic effects and were most robust during Dark 2. Treatment with the T4 analog, Tetrac, restored photomotor response in mct8 morphants to control levels. Collectively, these findings suggest that while the sensitivity of photomotor behavior in larval zebrafish to detect TH disruption is influenced by test parameters, this test can distinguish between TH promoting and TH blocking activity and may be useful for assessing the DNT potential of TH-active chemicals.

Environmental toxicants that interfere with thyroid hormone (TH) signaling can impact growth and development in animals and humans. Zebrafish represent a model to study chemically induced TH disruption, prompting the need for sensitive detection of THs. Simultaneous quantification of 3,3',5-triiodo-l-thyronine (T3), thyroxine (T4), 3,3',5'-triiodo-l-thyronine (rT3), 3,5-diiodo-l-thyronine (3,5-T2) and 3,3'-diiodo-l-thyronine (3,3'-T2) in zebrafish larvae was achieved by ultra-performance liquid chromatography-tandem mass spectrometry in positive ion mode. Solid-phase extraction with SampliQ cartridges and derivatization with 3 m hydrochloric acid in n-butanol reduced matrix effects. Derivatized compounds were separated on an Acquity UPLC BEH C column with mobile phases consisting of 0.1% acetic acid in deionized water and 0.1% acetic acid in methanol. The limits of detection ranged from 0.5 to 0.6 pg injected on column. The method was validated by evaluating recovery (77.1-117.2%), accuracy (87.3-123.9%) and precision (0.5-12.4%) using diluted homogenized zebrafish embryos spiked with all target compounds. This method was then applied to zebrafish larvae collected after 114 h of exposure to polychlorinated biphenyls (PCBs), including PCB 28, PCB 66 and PCB 95, or the technical mixture Aroclor 1254. Exposure to PCB 28 and PCB 95 increased the T4:T3 ratio and decreased the T3:rT3 ratio, demonstrating that this method can effectively detect PCB-induced alterations in THs.

Thyroid hormone (TH) is essential for brain development. While disruption of TH signaling by environmental chemicals has been discussed as a mechanism of developmental neurotoxicity (DNT) for more than a decade, there remains a paucity of information linking specific TH disrupting chemicals to adverse neurodevelopmental outcomes. This data gap reflects, in part, the fact that the molecular machinery of TH signaling is complex and varies according to cell type and developmental time. Thus, establishing a baseline of the ontogenetic profile of expression of TH signaling molecules in relevant cell types is critical for developing in vitro and alternative systems-based models for screening TH disrupting chemicals for DNT. Here, we characterize the transcriptomic profile of molecules critical to TH signaling across three species–human, rat, and zebrafish–in vitro and in vivo across different stages of neurodevelopment. Our data indicate that while cultured human and rat neural progenitor cells, primary cultures of rat cortical cells, and larval zebrafish all express a fairly comprehensive transcriptome of TH signaling molecules, the spatiotemporal expression profiles as well as the responses to TH vary across species and developmental stages. The data presented here provides a roadmap for identifying appropriate in vitro and in simpler systems-based models for mechanistic studies and screening of chemicals that alter neurodevelopment via interference with TH action.