Renal proximal tubule cells overexpress aromatic L-amino acid decarboxylase (MD) to produce dopamine, which inhibits salt absorption in the hypertonic environment. We examined the effect of TonEBP on AAD expression in human proximal tubule epithelial cells, HK-2 cell line. Confocal microscopy showed that after 2 h of exposure to the hypertonic medium, TonEBP accumulation in nuclei increased as compared to the isotonic control. The activated TonEBP enhanced the mRNA expression of the representative downstream genes (i.e., SMIT and TauT). Meanwhile, AAD protein abundance also increased with TonEBP activation. EMSA and luciferase reporter assay showed that TonEBP was involved in transcriptional regulation of AAD upon hypertonic stress. Inactivation of TonEBP by the p38 inhibitor SB203580, or TonEBP shRNA significantly reduced AAD expression, which was rescued by re-expressing Myc-tagged TonEBP. Up-regulation of AAD increased dopamine synthesis, and dopamine inhibited NKA activity in hypertonic condition. These results suggested that TonEBP played an important role in the epithelial cells of renal proximal tubule upon hypertonic stress by enhancing AAD expression, which could promote dopamine secretion to negative regulate NKA activity. The elucidation of a new mechanism described in this study combined with previous findings provides more insights into this issue. (C) 2011 Elsevier Inc. All rights reserved.