We examined acute changes in MRS-visible glutamate and glutamine after stimulation with N-acetylcysteine (NAC), since NAC reportedly decreases synaptic glutamate via activation of inhibitory metabotropic glutamate receptors. In 10 healthy adults, NAC significantly reduced Glx in the basal ganglia and prefrontal cortex. In the basal ganglia, the changes in Glx were driven by changes in Gln, suggesting that Gln might represent a proxy marker for synaptic glutamate. In the frontal lobe, the MEGAPRESS edited spectra showed greater sensitivity to changes in Glx than short TE PRESS or the edit OFF subspectra. Acute compartmental shifts in glutamate are detectable with MRS.