We have previously demostrated that there are developmental changes in proximal tubule Na⁺/H⁺ exchanger (NHE) activity. There is a maturational increase in postnatal brush border membrane (BBM) NHE3 protein abundance and decrease in NHE8 protein abundance. This study determined if thyroid hormone plays a role in the rat renal maturational isoform switch from NHE8 to NHE3 and if thyroid hormone regulates NHE8. Administration of thyroid hormone to neonatal rats, prior to the normal postnatal increase in serum thyroid hormone levels, resulted in a premature increase in NHE3/-actin BBM protein abundance and mRNA abundance. Thyroid hormone caused a premature decrease in BBM NHE8/-actin protein abundance, while there was no change in mRNA expression. Rats made hypothyroid from birth were studied at 28 days. In these hypothyroid adult rats, the maturational increase in BBM NHE3 protein abundance and NHE3 mRNA expression was prevented. In contrast the developmental decrease in BBM NHE8 protein abundance was prevented in hypothyroid adults, but mRNA expression was unchanged in hypothyroid rats. To determine if the effect of thyroid hormone was due to a direct epithelial effect, we studied NRK (normal rat kidney) cells in culture. Thyroid hormone caused a decrease in surface expression of NHE8, determined by biotinylation. NHE8 activity, measured as the sodium dependent rate of intracellular pH recovery from an acid load, was less with thyroid treatment than control. In conclusion, thyroid hormone plays a potential role in the developmental isoform change from NHE8 to NHE3 and decreases NHE8 activity.