Gamma-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian cortex. Extensive work in animals has shown that changes in GABA metabolism plays an important role in the original and spread of seizure activity. This project will center on the enzymatic control of flux through key enzymes of GABA synthesis (glutamic acid decarboxylase, GAD) and degradation (GABA-transaminase) in the cortex of rats treated with vigabatrin (gamma-vinyl-GABA), a highly specific GABA-transaminase inhibitor and one of the most promising new anti-epileptic drugs under clinical development. Our general hypothesis is that changes in the rate of cortical GABA synthesis following a prolonged increase in GABA concentration in vivo can be interpreted quantitatively in terms of GABA-induced changes in the isoform composition and activity of GAD. Also that inhibition of GABA metabolism interrupts carbon and nitrogen cycling between neurons and glia, altering glutamate and glutamine metabolism in vivo. We propose four specific aims: 1) Investigate the relationship between the in vivo rate of cortical GABA synthesis, GABA concentration and GAD activity (i.e., total GAD and major GAD isoforms) after acute and prolonged GABA elevation, 2) Determine the changes in the in vivo rate of cortical GABA synthesis and GAD activity during seizures and 3) in response to stimulation of the somatosensory cortex. 4) Determine the relationship between the changes in the rate of turnover of GABA measured in brain tissue in vivo and GABA in the extracellular fluid during acute and chronic GABA-transaminase inhibition. Using in vivo NMR Spectroscopy, Magnetic Resonance Imaging, in vivo microdialysis, immunologic and enzymatic assays in vitro, we will examine the role of GABa and other modulators in the control of GABA synthesis and the regulation of GABA levels in vivo. The uniqueness of this project derives from 1) the approach of combining in vivo and in vitro techniques, 2) the track record of the investigators in addressing questions using NMR, and 3) the fact that this project complements ongoing clinical investigations of GABA and glutamate metabolism in epilepsy and the effects of antiepileptic medications on the metabolic pathways of the human brain.
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