Elsevier

Aquatic Toxicology

Volume 94, Issue 2, 31 August 2009, Pages 87-93
Aquatic Toxicology

Effects of exposure to acetochlor on the expression of thyroid hormone related genes in larval and adult rare minnow (Gobiocypris rarus)

https://doi.org/10.1016/j.aquatox.2009.06.002Get rights and content

Abstract

Acetochlor is a commonly used herbicide in agricultural environments, which can accelerate T3-induced metamorphosis in amphibians. However, effects of acetochlor on the thyroid system in fish are still unclear at environmentally relevant concentrations. In this study, expression profiles of thyroid hormone receptor alpha (trα), deiodinase (d1 and d2), malic enzyme (me), and sodium iodide symporter (nis) genes were determined in larval and adult rare minnow (Gobiocypris rarus) after exposure to different levels of acetochlor (20, 200, and 2000 ng/l) for 21 days, respectively. Furthermore, plasma thyroid hormones (THs) levels in adults were evaluated simultaneously. The results showed that d1, d2, me, and nis mRNA levels were significantly down-regulated in the larvae. Similar expression changes of these genes in female brains were found. However, in adult livers, these gene expressions had no significantly effects, except the trα mRNA level significantly up-regulated. These results indicated that exposure to acetochlor could result in tissue-specific alternative expression of TH-related genes in adults. Moreover, the expression of d2 and me showed a positive correlation with plasma T4 levels in female brains. Therefore, larval development and adult brain of rare minnow could be affected by acetochlor at environmentally relevant concentrations.

Introduction

In recent years, a lot of endocrine disrupting chemicals (EDCs) have been released into the aquatic environment (Petrovic et al., 2004). Previous studies confirmed that some EDCs could cause disturbance of thyroid hormone homeostasis, hypothyroidism, and thyroid gland abnormalities in fish (Arukwe and Jenssen, 2005). As a significant portion of EDCs, several pesticides are potential endocrine disrupters, which alter the normal functioning of the endocrine system (Charlier and Plomteux, 2002). The use of pesticides on agricultural land may result in contamination of adjacent surface waters and thus pose a potential risk to a range of aquatic organisms such as fish (Guo et al., 2008). However, studies on disruption of the thyroid system in fish are limited (van der Ven et al., 2006).

Acetochlor (2-chloro-N-(ethoxymethyl)-N-(2-ethyl-6-methylphenyl)-acetamide) is a common herbicide all over the world (Lengyel and Foldenyi, 2003), which has been widely used in China (Ye, 2003). Previous studies have demonstrated the presence of acetochlor in surface water (Hladik et al., 2008), soil (Chao et al., 2007), and sediment (Xue et al., 2005). Measurable concentrations of acetochlor ranged from 0.05 to 2.5 μg/l in surface water (Boyd, 2000, Hladik et al., 2008, Kolpin et al., 1996). Acetochlor has been classified by the US EPA as a B-2 carcinogen (U.S. EPA, 1994). Furthermore, acetochlor can accelerate T3-dependent metamorphosis in amphibians (Cheek et al., 1999, Crump et al., 2002, Helbing et al., 2006). However, the effects of acetochlor alone on the thyroid system of fish are unclear.

The thyroid hormones (THs) triiodothyronine (T3) and thyroxine (T4) have a wide range of biological effects in physiological processes of vertebrates (Power et al., 2001). In fish, THs have been implicated as being important regulators in differentiation, growth, metabolism, and salinity adaptation (Crane et al., 2004, Orozco et al., 2002). The effects of exposure to EDCs may be multiple targets with the complex regulatory network of thyroid hormone metabolism and action: TH receptors (TR), mediating gene regulation in response to T3 (Marchand et al., 2001); deiodinase enzyme (D1 and D2), outer ring deiodination of T4 to the biologically active T3 (Orozco and Valverde, 2005); malic enzyme (ME), having a central role providing NADPH for fatty acid biosynthesis lipid metabolism and being a well-characterised endpoint of THs action (Rosebrough et al., 2006); and the sodium iodide symporter (NIS), an integral plasma membrane glycoprotein of thyroid hormone synthesis (Dohan et al., 2003). Previous studies indicated that the gene expressions of TH receptors and deiodinases were sensitive molecular biomarkers for thyroid disruption in fish exposed to environmental contaminants (Picard-Aitken et al., 2007, Scholz and Mayer, 2008). However, little is known about the me and nis genes as potential biomarkers in fish.

Rare minnow (Gobiocypris rarus) is considered to be an appropriate species for the assessment of endocrine disrupting chemicals due to its small size, ease of culture, short life cycle and prolific egg production with high fertilization and hatching rates (Zha et al., 2007). Rare minnow is an ideal model fish in the laboratory. We have been focused on changes of the hypothalamic–pituitary–gonad (HPG) axis in Chinese rare minnow by environment pollutants (Zha et al., 2008, Zhang et al., 2008). However, the potential disruption of hypothalamic–pituitary–thyroid (HPT) axis is largely unknown.

With the advances in molecular approaches, the application of toxicogenomics could be a powerful tool for evaluating the effects and discovering molecular mechanisms underlying toxic response (Nie et al., 2006). Therefore, in this study, we suppose that the expressions of trα, d1, d2, me, and nis genes can be potential biomarkers to study on disruption of the thyroid system. Our studies aimed at investigating the effects of environmentally relevant concentrations of acetochlor on plasma THs levels and the expression of THs related genes of larvae and adult rare minnow, and discussing the possible molecular mechanisms underlying toxic response.

Section snippets

Chemicals

Acetochlor was purchased from Sigma–Aldrich Chemical Co. (USA). Stock solution of acetochlor was prepared by dilution in dimethylsulfoxide (DMSO) (Absolute grade), which was purchased from Tedia Company Inc. (Fairfield, OH, USA). To obtain the final concentration for exposure, the appropriate amount of the stock solution was added to the aquarium water via a combination of polytetrafluorethylene (PFTE) and isoversinic tubes (Abimed, Langenfeld, Germany). A vehicle treatment containing a

Mortality and growth

No mortality was observed in treatment and control groups during the exposure period. At the end of acetochlor exposure, there was no significant difference in growth of larvae (Table 2).

Quantitation of trα, d1, d2, me, and nis mRNA at the larval stage

The mRNA expressions of trα, d1, d2, me, and nis in whole larvae were determined by real-time PCR after the 21-day exposure to acetochlor (Fig. 1). The trα mRNA levels were significantly increased at 20 ng/l (p < 0.05), but significantly decreased at 200 ng/l. The d1, d2, and nis mRNA levels were significantly

Discussion

Synthetic organic pesticides have been detected frequently in ground water, surface water, aquatic biota and sediment, and atmosphere in decades (Barbash et al., 2001). As a common pre-emergent herbicide all over the world, measurable concentrations of acetochlor can accelerate T3-induced metamorphosis in amphibians. In this study, effects of environmentally relevant concentrations of acetochlor on the expression of trα, d1, d2, me, and nis genes were investigated in larvae and adults rare

Conclusion

The gene expressions of THs related genes in larvae were sensitive to acetochlor. These results indicated that normal development of larvae might be affected at environmentally relevant concentrations of acetochlor. Exposure to acetochlor can decrease circulating TH levels of adults, and result in tissue-specific alternations of THs related genes expression in adults. The present data suggested that exposure to acetochlor may affect normal brain development, especially in females. However,

Acknowledgements

This work was supported by Chinese Academy of Sciences (KZCX2-YW-Q02-05), National Basic Research Program of China (2009CB421605), and the National Natural Science Foundation of China (20737003, 20877089).

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