Two Glutamate Decarboxylase Forms Corresponding to the Mammalian GAD65 and GAD67 are Expressed During Development of the Chick Telencephalon

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Abstract
The gamma-aminobutyric acid (GABA)-synthesizing enzyme glutamate decarboxylase (GAD) was studied during development of the chick telencephalon. By means of reverse-phase HPLC analysis, we showed that GABA indeed accumulates during embryogenesis, whereas the levels of glutamate, the substrate for GAD, are more or less unchanged up to later developmental stages. The enzyme activity increased approximately 25-fold from embryonic day 3 to embryonic day 17. Immunoblotting data revealed that two GAD proteins, of approximately 65 and 67 kDa, were present during the period investigated. Furthermore, Northern blot analysis with probes obtained from rat cDNA sequences, as well as a chicken-specific probe for GAD65 generated by means of reverse transcriptase-polymerase chain reaction (RT-PCR), strengthened the interpretation that the chick embryo expresses genes corresponding to GAD65 and GAD67. The rat probes recognized transcript sizes of 3.9 kb (GAD65) and 5.6 kb (GAD67), sizes which are different from those of the rat brain (Erlander et al., Neuron, 7, 91-100, 1991). Sequencing of the RT-PCR products revealed a high level of homology (82% at the nucleotide level) between the mammalian and chick GAD65 genes. Taken together, these findings suggest that the chick embryo expresses two GAD genes during embryogenesis. The functional properties of each gene product remain to be investigated.