Comparative distribution of GAD(65) and GAD(67) mRNAs and proteins in the rat spinal cord supports a differential regulation of these two glutamate decarboxylases in vivo

Gamma-aminobutyric acid (GABA) synthesis can result from the action of at least two glutamic acid decarboxylase (GAD) isoforms, GAD(65) and GAD(67) possibly involved in distinct mechanisms. We have made the hypothesis that GAD(65) may respond to short-term changes and is present in neurons with a phasic activity, while GAD(67) may rather provide GABA for the metabolic pool and for supporting tonic levels of synaptic transmission (Erlander et al.: Neuron 7:91-100, 1991; Feldblum et al.: J Neurosci Res 34:689-706, 1993). In the present work we have tested this hypothesis in the rat spinal cord where both types of activities have been identified. The correlation of GABA immunodetection with the distribution of GAD(65) and GAD(67) mRNAs and proteins has evinced in the dorsal horn a differential regulation of the two isoforms. In situ hybridization has revealed, in the dorsal horn, relatively higher levels of GAD(67) mRNA than of GAD(65), while immunodetection of the proteins demonstrated numerous punctate profiles with both GAD antisera. Reverse transcription-polymerase chain reaction (RT-PCR) data confirmed the abundance of the GAD(67) transcripts compared to GAD(65) in the rat spinal cord. In contrast, within the ventral horn, there was a greater number of GAD(67)-immunoreactive (IR) profiles mostly located around motoneurons. The paucity of GAD(65) immunoreactivity in the ventral horn cannot be related to a different accessibility of the antigens to the epitopes since on the same section a dense GAD(65) staining was detected in the dorsal horn. Hence, a number of biochemical and electrophysiological data support the concept of the involvement of glycine as the major inhibitory system within the ventral horn which may explain the low levels of GAD transcription in this region. The paucity of GAD(65) in the ventral horn may also reflect a functional difference, suggesting a predominance of GAD(67) in neurons under tonic activity. In the dorsal horn, where neurons with phasic and tonic firing patterns have been disclosed, GAD(65) may, in addition, provide GABA for responses to short-term changes. (C) 1995 Wiley-Liss, Inc.