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Impaired swim bladder inflation in early-life stage fathead minnows exposed to a deiodinase inhibitor, iopanoic acid (presentation)
Cavallin, J., G. Ankley, B. Blackwell, C. Blanksma, K. Fay, K. Jensen, M. Kahl, D. Knapen, Pat Kosian, S. Poole, E. Randolph, A. Schroeder, L. Vergauwen, AND Dan Villeneuve. Impaired swim bladder inflation in early-life stage fathead minnows exposed to a deiodinase inhibitor, iopanoic acid (presentation). SETAC Midwest Chapter, Minneapolis, MN, March 20 - 22, 2017.
The fish early life stage (FELS) toxicity test (OECD 210; OCSPP 850.1400) is the most widely used chronic ecotoxicity test. There is strong interest in development of alternatives to reduce costs, increase efficiency, and minimize animal use. Development of adverse outcome pathways (AOPs) that link pathway perturbations that could be measured either using in vitro systems or in short-term high throughput fish embryo tests to potential adverse effects on fish growth or survival have been viewed as a means to support reduction, refinement, and/or replacement of FELS testing. The present study contributes to the overall goal, while at the same time addressing the topic of thyroid axis disruption, a critical concern relative to implementation of the legislatively-mandated Endocrine Disruptor Screening Program (EDSP). A screening assay for deoiodinase inhibition, a molecular initiating event associated with thyroid axis disruption, is under development by the CSS High Throughput Toxicology project. This poster describes two experiments in which embryo-larval fathead minnows were exposed to iopanoic acid, a model deiodinase inhibitor. Effects on swim bladder inflation and morphology, as critical developmental events that impact young of year survival in fishes, were examined. Additionally, key events establishing a linkage between deiodinase inhibition as a mode of action, altered ratios of T3/T4 in developing fish, and the adverse outcome were demonstrated. These results provide supporting evidence for a series of AOP descriptions that have been deposited to the internationally harmonized AOP wiki and directly support the aims of the CSS Adverse Outcome Pathway Discovery and Development Task. This research enhances the utility of molecular screening assays for deiodinase inhibition as a tool to inform risk assessment and regulatory decision making concerning potential impacts on fish. It complements additional work in the same project focused on effects of deiodinase inhibition on amphibian metamorphosis and developmental outcomes in mammals.
The present study investigated whether inhibition of deiodinase, the enzyme which converts thyroxine (T4) to the more biologically-active form, 3,5,3'-triiodothyronine (T3), would impact inflation of the posterior and/or anterior chamber of the swim bladder, processes previously demonstrated to be thyroid-hormone regulated. Two experiments were conducted using a model deiodinase inhibitor, iopanoic acid (IOP). In the first study, fathead minnow (Pimephales promelas) embryos were exposed to 0.6, 1.9, or 6.0 mg IOP/L or control water in a flow-through system until reaching 6 days post-fertilization (dpf) at which time posterior swim bladder inflation was assessed. To examine effects on anterior swim bladder inflation, a second study was conducted with 6 dpf larvae exposed to the same IOP concentrations until reaching 21 dpf. Fish from both studies were sampled for T4/T3 measurements, gene transcription analyses, and thyroid histopathology. In the embryo study, incidence and length of inflated posterior swim bladders were significantly reduced in the 6.0 mg/L treatment at 6 dpf. Incidence of inflation and length of anterior swim bladder in larval fish were significantly reduced in all IOP treatments at 14 dpf, but inflation recovered by 18 dpf. Throughout the larval study, whole body T4 concentrations were significantly increased and T3 concentrations were significantly decreased in all IOP treatments. Consistent with hypothesized compensatory responses, significant up-regulation of deiodinase-2 mRNA was observed in the larval study, and down-regulation of thyroperoxidase mRNA was observed in all IOP treatments in both studies. Taken together, these results support the hypothesized adverse outcome pathways linking inhibition of deiodinase activity to impaired swim bladder inflation.