Tbr-1, a neuron-specific T-box transcription factor, plays a critical role in brain development. Here, we performed a computational search using the non-palindromic T-box binding sequence, namely the non-palindromic T-element, to determine the putative downstream target genes of Tbr-1. More than 20 identified genes containing the non-palindromic T-element in the 5' regulatory region were found expressed in brain. Luciferase reporter assays using cultured hippocampal neurons showed that overexpression of Tbr-1 and CASK-enhanced promoter activities of some of these putative target genes, including NMDAR subunit 2b (NR2b), glycine transporter, interleukin 7 receptor (IL-7R) and OX-2. Among these genes, NR2b promoter responded strongest to overexpression of Tbr-1 and CASK. Deletion of the non-palindromic T-elements from NR2b promoter impaired the induction by Tbr-1 and CASK. We also examined expression of these target genes in Tbr-1 knockout mice, it was found that NR2b expression was consistently downregulated. Similarly, both RNA and protein expression levels of NMDAR subunit 1 (NR1), which also contains the non-palindromic T-elements in its 5' regulatory region, were reduced in Tbr-1 knockout mice. We suggest that Tbr-1/CASK protein complex regulates expression of these downstream target genes and thus modulates neuronal activity and function.