Abstract : In fish, 11B-hydroxylase (11BH) is the enzyme responsible for the biosynthesis of cortisol, the main corticosteroid in fish, which is implicated in the regulation of multiple physiological processes including stress response, energy metabolism, hydromineral balance, reproduction and the immune system. The gene coding for 11BH, namely cyp11c1, is expressed early in interrenal cells during the development participating to the emergence of the corticosteroid axis. To further investigate the expression of cyp11c1 and its potential deregulation by stressors and pharmaceutical ligands, we established a transgenic zebrafish line that expresses Green Fluorescent Protein (GFP) under the control of the zebrafish cyp11c1 promoter. Using in vivo fluorescence imaging techniques on whole larvae, we showed an expression of the reporter gene from 4 to 7 days postfertilization (dpf) in an anatomical region where interrenal cells are known to be present. Tissue sections and immunostaining with specific 11BH antibodies revealed that GFP is co-localized with the endogenous cyp11c1 protein in interrenal cells. This well agrees with the expression and localization of other steroidogenic interrenal marker genes such as star and cyp11a1 in wild-type zebrafish at this stage of development (To et al. 2007). To further characterize the cyp11c1-GFP zebrafish model, GFP was quantified in 6-dpf old zebrafish after exposure to glucocorticoids receptor (GR) agonists (Dexamethasone, DEX) and antagonists (Mifepristone, MIF). Exposure to DEX (0.1uM, 1uM and 10uM from 4 to 6 dpf) increased GFP intensity in a concentration dependent manner, suggesting that GR is involved in the transcriptional regulation of cyp11c1 during development. This effect was abolished when zebrafish larvae were co-exposed with MIF. Interestingly, exposure to DEX induced whole-body cortisol in a concentration-dependent manner which is consistent with the effect seen on 11BH. Altogether, these data suggest that the newly developed transgenic cyp11c1-GFP zebrafish is an interesting model to study the expression of 11B-hydroxylase gene in interrenal cells and its perturbation by endocrine active compounds acting on the HPI axis, which is critical for cortisol homeostasis.
