The elucidation of the mechanisms of neural development using glutamic acid decarboxylase knockout mice

点击翻译按钮获取中文摘要

1.gamma-Aminobutyric acid (GABA) is synthesized by glutamic acid decarboxylase (GAD). We have isolated and characterized the mouse GAD (mGAD-1 and mGAD-2) genes. The mGAD-1 gene spans over 45 kb in length and contains 19 exons including two 5'noncoding exons (exon 0A and exon 0B). On the other hand, the mGAD-2 gene spans more than 50 kb in length. We made an mGAD-2-targetting construct for generating knockout mice. The construct was electroporated into mouse ES cells and drug-resistant clones were selected. We select the clones that have targeted disruption of the mGAD-2 gene. The clones will be used to make the mGAD-2 knockout mice and these mice would be useful to elucidate the role of mGAD genes in neural development.2.To understand the molecular mechanisms underlying the regulation of mGAD-1 gene expression, we have isolated the 5'-flanking region of the mGAD-1 gene. Sequence analysis of the 5'-flanking region revealed the presence of a number of putative regulatory elements including NRSE,Krox-24, and Sp1. Using transgenic mice, we have examined the expression pattern conferred by a 10.2 kb fragment of the mGAD-1 gene (including 8.4 kb of 5'sequences) fused to the bacterial lacZ reporter gene. In two separate founder lines, the transgene expression was observed in neurons of particular brain regions such as the olfactory tubercle, cerebral cortex, and basal ganglia, where expression of GAD-1 gene has been reported to be abundant. These results suggest that the 10.2-kb DNA fragment of the mGAD-1 gene may be essential for its GABA ergic cell-specific expression in vivo.