Aquatic Toxicology 140–141 (2013) 279–287 Contents lists available at SciVerse ScienceDirect Aquatic Toxicology jou rn al hom ep age: www.elsevier.com/locate/aquatox Waterborne exposure of zebrafish embryos to micromole concentrations of ioxynil and diethylstilbestrol disrupts thyrocyte development M.A. Campinho ∗ , D.M. Power Comparative and Molecular Endocrinology Group, Centro de Ciências do Mar (CCMAR), Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal a r t i c l e i n f o Article history: Received 16 January 2013 Received in revised form 11 June 2013 Accepted 14 June 2013 Keywords: Development Diethylstilbestrol Endocrine disruption Ioxynil Thyrocytes Zebrafish a b s t r a c t The herbicide ioxynil (IOX) and synthetic estrogen diethylstilbestrol (DES) are common aquatic con- taminants with an endocrine disrupting action. In juvenile teleost fish IOX and DES disrupt the hypothalamic–pituitary–thyroid (HPT) axis. To assess how IOX and DES influence the developing HPT axis prior to establishment of central regulation of thyroid hormones, zebrafish embryos were exposed to low concentrations of the chemicals in water. IOX and DES (1 and 0.1 M) exposure failed to modify hypothalamic development but had a negative effect on thyrocyte development. Specifically, IOX and DES caused a significant (p < 0.05) reduction in the size of the thyroid anlagen by decreasing the mRNA expression field of both nk2.1a and thyroglobulin (Tg) genes. Inhibition of thyroid gland development by IOX and DES (0.1 M) was strongly associated with altered heart morphology. To test if the effect of IOX and DES on the thyroid was a consequence of altered cardiac development a morpholino (MO) against zebrafish cardiac troponin I (zcTnI) was microinjected. The zcTnI morphants had modified heart function, a small thyroid anlagen and a reduction in the mRNA expression of nk2.1a and Tg genes similar to that of zebrafish exposed to IOX (1 and 0.1 M) and DES (0.1 M). Collectively the data indicate that IOX and DES alter thyroid development in zebrafish and chemicals that alter heart development and function can have an indirect endocrine disrupting action on the thyroid in teleosts. © 2013 Elsevier B.V. All rights reserved. 1. Introduction The hypothalamic–pituitary–thyroid (HPT) axis is a central regulator of vertebrate physiology. Its leading systemic signal is thyroid hormones (THs) produced by the thyroid gland. THs include thyroxine (T4), the main product of thyrocytes that is mainly deiodinated in the periphery to produce the active hormone tri- iodothyronine (T3) (Bianco and Larsen, 2005). Thyroid Stimulating Hormone (TSH) liberated from the pituitary controls most, if not all, steps of TH production by thyrocytes and is regulated through neg- ative feedback by serum T4. Hypothalamic neurons sense serum levels of T4 and regulate TSH secretion by pituitary thyrotrophs. In birds and mammals, thyrotrophin releasing hormone (TRH) is the main hormone controlling pituitary TSH production but in amphibians and teleosts (Denver et al., 2002) corticotrophin releas- ing hormone (CRH) assumes this role (de Jesus et al., 1991, 1990; Denver, 1997). Given the cellular and molecular actions of THs ∗ Corresponding author. Tel.: +351 800900x7375; fax: +351 289800051. E-mail addresses: macampinho@ualg.pt (M.A. Campinho), dpower@ualg.pt (D.M. Power). in vertebrates the HPT-axis has an important regulatory role in a diversity of processes encompassing basal cellular respiration to sexual differentiation in vertebrates. Disruption of the HPT axis may lead to a systemic response in which several physiological imbalances coexist, leading to decreased viability of individuals (Boas et al., 2012; Carr and Pati ˜ no, 2011). It is well established that the HPT axes in aquatic verte- brates, namely amphibians and teleosts, are subject to alterations after waterborne exposure to anthropogenic endocrine-disrupting chemical (EDCs) (Brar et al., 2009; Leatherland, 1998; Leatherland and Sonstegard, 1977; Mukhi and Pati ˜ no, 2007). Contamination of aquatic systems by the widely used herbicide ioxynil (IOX) and synthetic estrogen diethylstilbestrol (DES) is common as they are present in agricultural run-offs and municipal waste water, and are also released into the environment by the chemical and medical industry. These chemicals are characterized by low water solubil- ity and a tendency to bioaccumulate (European-Comission, 2004; Linders et al., 1994; Yang et al., 2008). In fish IOX exhibits an acute LC50 at 3.5–8.5 mg/L and a long term “no observed effect concen- tration” (NOEC) of 3.2 mg/L (European-Comission, 2004). In adult Carassius auratus water concentrations of 10 g/L DES modify liver and testis histology (Yang et al., 2008). Concentrations of 0.9 g/l 0166-445X/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.aquatox.2013.06.014