The effect of elevated oocyte triiodothyronine content on development of rainbow trout embryos and expression of mRNA encoding for thyroid hormone receptors

Abstract

The ability of developing rainbow trout Oncorhynchus mykiss embryos to compensate for elevated oocyte triiodothyronine (T3) content and whether elevation of oocyte T3 content within a physiologically meaningful range affects growth rates of the embryo or the expression of genes encoding for thyroid hormone receptors α(TRα) and β(TRβ) were examined. Oocytes were immersed in ovarian fluid alone (control) or T3‐enriched ovarian fluid prior to fertilization and water hardening, to induce a dose‐dependant increase in oocyte T3 content of c. 3 (control), c. 30 (LT3) or c. 110 ng egg−1(HT3). To examine the interaction of embryo somatic growth with altered thyroid state more effectively, the embryos were reared at two ambient temperatures (8·5 and 5·5°C ) to induce different growth rates. A significant decline in whole embryo T3 content was measured in the T3‐treatment groups reared at both water temperatures by 3 weeks post‐fertilization (dpf), and may have reflected the action of outer ring monodeiodinase, which was present in microsomes prepared from embryos 23 dpf. Whole embryo T3 levels in the HT3 group, however, remained higher than controls until phase 2 of development [the onset of endogenous thyroid hormone (TH) release]. This suggested that the embryos exerted some control over their response to exogenous TH, but that there was a limit to the level of control exerted by the embryonic tissues. Reverse transcriptase‐polymerase chain reaction (RT‐PCR) revealed the presence of mRNA encoding for the two TR isoforms as early as 26 dpf, and quantitative real‐time RT‐PCR (qPCR) was used to examine the effect of elevated oocyte T3 content on the expression of these TR genes in embryos raised at 8·5 and 5·5° C, and sampled at similar developmental stages prior to the onset of embryonic TH synthesis, to ensure that the oocyte T3 was the only source of TH exposure to the embryo. There was a suppression of the TRα gene expression in the control 5·5° C group relative to the control 8·5° C group. In addition, both TRα and TRβ mRNA accumulation was lower, relative to the controls, in the LT3 treatment group reared at 8·5° C suggesting a suppressive effect of the lower level of T3 treatment on the TR gene expression. Conversely, there were no differences from controls in the HT3 treatment group, possibly indicating that this level of exposure overrides the down‐regulating capacity of the embryo. Similar patterns were seen for TRα and TRβ mRNA accumulation in embryos reared at 5·5° C, but because of the temperature suppressed level of TRα mRNA in the controls, significant affects of the LT3 treatment were only found for TRβ. There were no measurable effects of T3 treatment on oocyte fertility or embryo somatic growth for either temperature treatment group, nor was somatic growth hormone content (measured only in the 8·5° C treatment group) apparently related to in ovo T3 levels. The results suggest that altered in ovo T3 levels, within the ranges used here, do not induce marked affects on embryo development, probably because of the ability of the embryo to maintain the integrity of its TH milieu.