Endocrine Abstracts

Atrazine (ATR) is a widely used herbicide, with known effects as an endocrine disrupting chemical. Several studies have implicated ATR in causing disorders of sex development in reptiles, and chronic exposure can cause an increase incidence of intersex in fish, suggesting that the hypothalamo-pituitary–gonadal axis is a major site of ATR action. In this study, we investigated whether acute exposure (hours to days) to ATR could cause abnormalities in the development of Zebrafish larvae. Wild-type AB zebrafish larvae were exposed to ATR (1–10 μM) within 2 h post-fertilisation (hpf), and examined at 24, 48 and 72 hpf. Morphometric analyses revealed that ATR caused a concentration-dependent increase in pericardial oedema and hatching rates (***P<0.001) compared with DMSO-treated controls, but significantly reduced eye diameter (**P<0.01). In addition, ATR exposure caused a modest, but highly significant reduction in body length (***P<0.001). Subsequent in situ hybridisation analyses of these larvae at each of the indicated time-points revealed altered expression of pomc, prl and l h in ATR-treated Zebrafish. To establish whether the growth alterations observed in Zebrafish were also observed in pituitary cell lines, crystal violet assays were performed on αT3-1, LβT2 (gonadotroph) and GH3 (somatotroph) cells. ATR exposure for up to 72 h failed to significantly alter cell proliferation in any of the cell lines examined. RT-PCR analyses of pituitary gene expression in αT3-1 and LβT2 cells revealed that ATR enhanced expression of Egr1. However, the expression of Nr5a1 was inhibited in αT3-1 cells, but enhanced in LβT2. Finally, as ATR is thought to interact with phosphodiesterase enzymes, we examined the effect of ATR on cAMP accumulation in αT3-1 and LβT2 cells. Surprisingly, ATR failed to significantly alter cAMP in either cell line, whereas the diterpine, Forskolin, dramatically enhanced cAMP in both cell types (***P<0.001). These data reveal that ATR may have rapidly acting effects during development in Zebrafish, and that disruption to pituitary gene expression in particular could contribute to endocrine disorders in later life.

Declaration of funding: This work was supported by a Wellcome Trust CVRT award (to J S).