Maturation of thyroid-axis gene regulation under altered TH conditions. (a) Changes in relative tshb, thrb, and dio3b mRNA levels during the first month of development as measured by RT-qPCR. (b) Changes in relative tshb, thrb, and dio3b mRNA levels after treatment with different doses of TH (T4 challenge). At 7 dpf (left), tshb mRNA levels responded appropriately to TH (decreased) with the addition of 2.5 nM T4, whereas thrb and dio3b mRNA levels responded appropriately (increased) with the addition of 100 nM T4 (black arrows). At 14 dpf (right), tshb mRNA levels were reduced with the addition of only 1 nM T4, suggesting increased sensitivity to TH (arrow). Similarly, thrb mRNA levels increased at lower T4 doses (arrow) at 14 dpf compared with 7 dpf. At 14 dpf, dio3b mRNA levels increased much more dramatically in response to doses of T4 higher than 12.5 nM than at 7 dpf (compare orange, green, and black arrows). (c) Effects of early PTU treatments on sensitivity to TH levels (T4 challenge). Larvae were grown in 0.65 mM PTU for 7 days prior to being challenged with 0.25 to 25 nM T4. Values are shown as the mean ± SEM. Student t test: *P < 0.05; **P < 0.01; ***P < 0.001.