Mineralocorticoid Receptor Mediates Cortisol Regulation of Ionocyte Development in Tilapia (<i>Oreochromis mossambicus</i>)

Cortisol is the predominant corticosteroid in ray-finned fish since it does not possess the aldosterone synthase necessary to produce specific mineralocorticoids. Cortisol is traditionally believed to function as a fish mineralocorticoid. However, the effects of cortisol are mediated through corticosteroid receptors in other vertebrates, and there is an ongoing debate about whether cortisol acts through the glucocorticoid receptor (GR) or the mineralocorticoid receptor (MR) in teleosts. To investigate this issue, we conducted a study using euryhaline Mozambique tilapia (<i>Oreochromis mossambicus</i>) as the experimental species. The experiment was designed to investigate the effect of cortisol on ionocyte development at both the cellular and gene expression levels in tilapia. We administered exogenous cortisol and receptor antagonists, used immunohistochemistry to quantify ionocyte numbers, and performed real-time PCR to assess the expression of the differentiation factor tumor protein 63 (P63) mRNA, an epidermal stem cell marker. We observed that cortisol increased the number of Na⁺-K⁺-ATPase (NKA)-immunoactive ionocytes (increased by 1.6-fold) and promoted the gene expression of <i>P63</i> mRNA (increased by 1.4-fold). Furthermore, we found that the addition of the mineralocorticoid receptor antagonist Spironolactone inhibited the increase in the number of ionocytes (decreased to the level of the control group) and suppressed the gene expression of <i>P63</i> (similarly decreased to the level of the control group). We also provided evidence for <i>gr</i>, <i>mr</i>, and <i>p63</i> localization in epidermal cells. At the transcript level, <i>mr</i> mRNA is ubiquitously expressed in gill sections and present in epidermal stem cells (cells labeled with <i>p63</i>), supporting the antagonism and functional assay results in larvae. Our results confirmed that cortisol stimulates ionocyte differentiation in tilapia through the MR, rather than the GR. Therefore, we provide a new direction for investigating the dual action of osmotic regulation and skin/gill epithelial development in tilapia, which could help resolve previously inconsistent and conflicting findings.