Impaired swim bladder inflation in early life stage fathead minnows exposed to a deiodinase inhibitor, iopanoic acid

Abstract

Inflation of the posterior and/or anterior swim bladder is a process previously demonstrated to be regulated by thyroid hormones. We investigated whether inhibition of deiodinases, which convert thyroxine (T4) to the more biologically active form, 3,5,3'-triiodothyronine (T3), would impact swim bladder inflation. Two experiments were conducted using a model deiodinase inhibitor, iopanoic acid (IOP). First, fathead minnow embryos were exposed to 0.6, 1.9, or 6.0 mg/L or control water until 6 d postfertilization (dpf), at which time posterior swim bladder inflation was assessed. To examine anterior swim bladder inflation, a second study was conducted with 6-dpf larvae exposed to the same IOP concentrations until 21 dpf. Fish from both studies were sampled for T4/T3 measurements and gene transcription analyses. 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 were significantly reduced in all IOP treatments at 14 dpf, but inflation recovered by 18 dpf. Throughout the larval study, whole-body T4 concentrations increased and T3 concentrations decreased in all IOP treatments. Consistent with hypothesized compensatory responses, deiodinase-2 messenger ribonucleic acid (mRNA) was up-regulated in the larval study, and thyroperoxidase mRNA was down-regulated in all IOP treatments in both studies. These results support the hypothesized adverse outcome pathways linking inhibition of deiodinase activity to impaired swim bladder inflation. Environ Toxicol Chem 2017;36:2942-2952. Published 2017 Wiley Periodicals Inc. on behalf of SETAC. This article is a US government work and, as such, is in the public domain in the United States of America.

Keywords: Adverse outcome pathway; Aquatic toxicology; Developmental toxicity; Endocrine-disrupting compounds; Thyroid disruption.

Figure 1

Effects of iopanoic acid (IOP) exposure on fathead minnow embryo thyroperoxidase (tpo) mRNA transcript abundance at 6 days post-fertilization. Bars represent mean ± SE of n=4 pooled whole body samples per treatment. Different letters indicate significant differences (p < 0.05) among treatments.

Figure 2

Effects of iopanoic acid exposure (IOP; 0 mg/L, 0.6 mg/L, 1.9 mg/L, and 6.0 mg/L) on fathead minnow larval expression of mRNA transcripts for (A) thyroperoxidase (tpo), (B) deiodinase 1 (dio1), (C) deiodinase 2 (dio2), (D) deiodinase 3 (dio3) at 10, 14, 18, and 21 days post-fertilization (dpf). Data points represent the mean ± SE of n=4 pooled whole body larval samples per treatment. * indicates significant differences (p < 0.05) between control and IOP treatments.

Figure 3

Effects of iopanoic acid exposure (IOP; 0 mg/L, 0.6 mg/L, 1.9 mg/L, and 6.0 mg/L) on whole-body thyroid hormone concentrations in pooled fathead minnows at 4 and 6 days post-fertilization (dpf). (A) 3,5,3′-triiodothyronine (T3) concentrations; (B) thyroxine (T4) concentrations. Data points represent the mean ± SE of n=4 pooled whole body samples per treatment. Different letters indicate significant differences (p < 0.05) among treatments.

Figure 4

Effects of iopanoic acid exposure (IOP; 0 mg/L, 0.6 mg/L, 1.9 mg/L, and 6.0 mg/L) on whole-body thyroid hormone concentrations in pooled fathead minnow larvae at 6 (T0), 10, 14, 18, and 21 days post-fertilization (dpf). (A) 3,5,3′-triiodothyronine (T3) concentrations; (B) thyroxine (T4) concentrations. Data points represent the mean ± SE of n=4 pooled whole body samples per treatment. Different letters indicate significant differences (p < 0.05) among treatments.

Figure 5

Effects of iopanoic acid exposure (IOP; 0 mg/L, 0.6 mg/L, 1.9 mg/L, and 6.0 mg/L) on the posterior swim bladder in fathead minnows at 6 days post-fertilization. (A) mean percent inflation of posterior swim bladders (n=4 reps of 25 fish); (B) mean length of inflated posterior swim bladders (n=86–99). Bars represent mean ± SE. Different letters indicate significant differences (p < 0.05) among treatments.

Figure 6

Effects of iopanoic acid exposure (IOP; 0 mg/L, 0.6 mg/L, 1.9 mg/L, and 6.0 mg/L) on the mean percent inflation of anterior swim bladders in fathead minnow larvae at (A) 14 days post-fertilization (dpf) (n=4 reps of 16 fish), and (B) 18 dpf (n=4 reps of 12 fish). Bars represent mean ± SE. Different letters indicate significant differences (p < 0.05) among treatments.

Figure 7

Effects of iopanoic acid exposure (IOP; 0 mg/L, 0.6 mg/L, 1.9 mg/L, and 6.0 mg/L) on the mean length of inflated anterior (black bars) and posterior (white bars) swim bladders in fathead minnow larvae at (A) 14 days post-fertilization (dpf) (n=27–64), (B) 18 dpf (n=36–48), and (C) 21 dpf (n=8). Bars represent mean ± SE. Significant (p < 0.05) differences between treatments are indicated with lower case letters for anterior swim bladders and upper case letters for posterior swim bladders. Swim bladder lengths for 14 and 18 dpf samples were obtained via live measurements with a camera, and 21 dpf swim bladder lengths were obtained through histological measurements due to lack of transparency of the fish at 21 dpf.