The existence of an interaction between the adrenal/interrenal axis and the thyroidal axis has since long been established in vertebrates, including fish. However, in contrast to mammals, birds and amphibians, no effort was made in fish to expand these studies beyond the level of measuring plasma thyroid hormones. We therefore set out to examine the acute effects of a single dose of dexamethasone (DEX) on plasma thyroxine (T(4)) and 3,5,3'-triiodothyronine (T(3)) levels, as well as on the activity and mRNA expression of the different iodothyronine deiodinases in liver, gills, kidney and brain in Nile tilapia. To take into account the effect of handling stress, this treatment was compared both to a non-treated and to a saline injected group. In general, the observed changes were acute (3 and 6h) while values had returned to control levels by 24h post-injection. Only DEX administration caused an acute drop in circulating T(3) levels compared to non-treated animals, while none of the treatments affected plasma T(4) levels. This indicates that the DEX induced decrease in plasma T(3) levels was not due to a lowered thyroidal hormone production and secretion. DEX injection provoked a decrease in peripheral T(3) production capacity via a decrease in hepatic outer ring deiodination activity (both D1 and D2), whereas T(3) clearance increased by induction of the inner ring deiodinating D3 pathway in liver and in gills. Deiodination activities in kidney and brain were not affected. Effects of saline injection were only observed in liver, where D1 activity decreased and D3 activity increased as in the DEX group, but to a lesser extent. Real-time PCR showed that the changes in hepatic D3 were clearly regulated at the pretranslational level, while this was not confirmed for the other changes. Our results show that both handling stress and DEX injection acutely disturb peripheral deiodination activity in Nile tilapia. However, the effects of the long acting glucocorticoid analogue are more pronounced and result in a decrease in circulating T(3) availability.