According to preclinical studies, glutamate has been implicated in the pathogenesis of anxiety. In order to elucidate the role of glutamate in anxiety and panic in humans, brain glutamate+glutamine (Glx) levels were measured during CCK-4 induced panic using magnetic resonance spectroscopy (MRS). Eighteen healthy subjects underwent CCK-4 challenge. MR spectra were obtained from the anterior cingulate cortex (ACC) using a single voxel point resolved spectroscopy method (PRESS) and analyzed using LCModel. A combined fitting of glutamine and glutamate (Glx) was performed. Panic was assessed using the Acute Panic Inventory (API) and Panic Symptom Scale (PSS) scores. Moreover, hypothalamic-pituitary-adrenal (HPA) axis stimulation was monitored throughout the challenge. There was a significant panic response following CCK-4 as revealed by a marked increase in both panic scores (API: F[1,17]=149.41; p<0.0001; PSS: F[1,17]=88.03; p<0.0001) and a significant increase in heart rate (HR: F[1,17]=72.79; p<0.0001). MRS measures showed a significant increase of brain glutamate+glutamine/creatine (Glx/Cr) levels peaking at 5-10 minutes after challenge (F[1,17]=15.94; p=0.001). There was also a significant increase in CCK-4 related cortisol release (F[6,11]=8.68; p=0.002). Finally, significant positive correlations were found between baseline Glx/Cr and both APImax (r=.598; p=0.009) and the maximum heart rate during challenge HRmax (r=.519; p=0.027). Our results suggest that CCK-4 induced panic is accompanied by a significant glutamate increase in the bilateral ACC. The results add to the hypothesis of a disturbance of the inhibitory-excitatory equilibrium and suggest that apart from static alterations also rapid and dynamic neurochemical changes might be relevant for the neural control of panic attacks.Neuropsychopharmacology accepted article preview online, 5 March 2013; doi:10.1038/npp.2013.61.