Neurotransmitter glutamate (GLU) is thought to derive primarily from the recycling of glutamine to GLU by phosphate-activated glutaminase (PAG). Previously, we reported that mice heterozygous for PAG (PAG-hets) show a 50% reduction in PAG activity, and are sensitized to the stimulant effects of MK-801 (SFN 2003). To examine the impact on GLUergic transmission, we used a monoclonal anti-GLU antibody (Glu2, Diasorin) to visualize GLU distribution; we found that the overall intensity of GLU staining was reduced to about 40%, p = 0.007 in PAG-het cortex. While there was some diminution in cell body labeling, neuropil staining was profoundly diminished, consistent with deficient neurotransmission. GLU microdialysis using the no-net flux method in the anterior cingulate cortex of awake PAG-hets showed that extracellular GLU levels did not differ between controls (0.73 ± 0.19 µM) and PAG-hets (0.70 ± 0.23 µM). Reverse-dialysis of L-trans-pyrrolidine-2,4-dicarboxylate (PDC) consistently increased GLU levels in wil...