Abstract title:
The cyp11c1-GFP transgenic zebrafish model to study the expression and the perturbation of a steroidogenic gene by stress and endocrine disrupting chemicals.


Abstract text:
The 11β-hydroxylase enzyme, coded by the cyp11c1 gene, is involved in the biosynthesis of cortisol in the zebrafish interrenal cells. Cortisol plays a critical role in important physiological processes such as stress response. The present work reviews the data recently obtained on the cyp11c1-GFP transgenic model developed to study the expression and the perturbation of the cyp11c1 gene. This new transgenic line expresses the Green Fluorescent Protein (GFP) under the control of the zebrafish promoter cyp11c1. Using in vivo fluorescence imaging techniques on whole organisms, it was demonstrated that the fluorescent reporter can be detected as soon as 4 days post-fertilization (dpf) and until 7 dpf, although further in development pigmentation is a limit to measure the expression of GFP. The signal is located in the anatomical region of the interrenal cells in the head kidney. Tissue sections and immunostaining with specific antibodies shown that the GFP expression mimics the endogenous protein expression. The expression pattern corresponds to previous literature concerning steroidogenesis enzymes and Star expression in wild-type zebrafish (To et al., 2007). In order to assess the response of this model to stress, 6 dpf cyp11c1-GFP larvae were exposed to osmotic shock. As a pharmacological approach, agonist and antagonist ligands of the glucocorticoids receptor (GR) has been tested on the line once the hypothalamo-pituitary-interrenal axis is functional. The GR is indeed involved in cortisol biosynthesis in the interrenal cells and is expressed at early developmental stages. Exposure to osmotic stress revealed an induction of cyp11c1 expression, implying that this gene is involved in the stress response and that this model is receptive to stress. Exposure to the GR antagonist Mifepristone shown a downregulation of cyp11c1 expression, suggesting that the GR might be involved in the regulation of the endogenous cyp11c1 expression in a constitutive manner. The GR agonist Dexamethasone downregulated the gene expression and the coexposure of both compounds rescued the basal expression of the gene, implying that Dexamethasone might induce a negative feedback on the gene expression, possibly via the HPI axis. Exposure to Forskolin, a compound known to induce steroidogenesis, seemed to upregulate the gene, and the glucocorticoids antagonist Pregnenolone seemed to induce a downregulation of cyp11c1 expression. This newly developed transgenic line provides an interesting model to acquire new data on the expression of the key steroidogenic gene cyp11c1 in zebrafish and its perturbation by endocrine active compounds or stress factors, and it could be used to further study the effects of compounds on the HPI axis, which is critical for cortisol homeostasis.

Track and session:
B - Aquatic and terrestrial ecotoxicology
Fish model species in environmental toxicology