The thyroid phenotype of tshr morphants is partially rescued by tshr mRNA overexpression. For phenotypic rescue, tshr morphants were coinjected with pTol2hsp70:tshr plasmid and transposase mRNA and heat shocked from 2 d postfertilization onwards. A–L, Results from in situ hybridization analyses of thyroid marker expression in heat-shocked embryos at 100 hpf. Ventral (A–H) and lateral views (I–L) of the thyroid region are shown with all embryos being oriented with anterior to the left. Arrows point to thyroid tissue. Compared with noninjected tshr morphants, pTol2hsp70:tshr-injected tshr morphants showed a partial rescue of slc5a5 (A–D) and nkx2.1a (I–L) mRNA expression. Expression of tg mRNA in pTol2hsp70:tshr-injected morphants after heat shock was indistinguishable from control embryos (E–H). Boxplots in M and N summarize results from quantification of the number of follicles immunoreactive for TG and T4 in 120 hpf Tg(fli1a:EGFP) embryos, respectively. Immunostaining was evaluated in control embryos (co), tshr morphants (tshr-MO), and tshr morphants rescued by coinjection of pTol2hsp70:tshr (MO-R). Black dots mark outliers, and the number of larvae analyzed is indicated in parentheses. Asterisks denote statistically significant differences (Dunn's test: *, P < 0.05; **, P < 0.01; ***, P < 0.001). n.s., Not significant.