The pathophysiological underpinnings of bipolar disorder are not fully understood. However, they may be due in part to changes in the phosphatidylinositol second messenger system (PI-cycle) generally, or changes in myo-inositol concentrations more specifically. Dextro-amphetamine has been used as a model for mania in several human studies as it causes similar subjective and physiological symptoms. We wanted to determine if dextro-amphetamine altered myo-inositol concentrations in vivo as it would clearly define a mechanism linking putative changes in the PI-cycle to the subjective psychological changes seen with dextro-amphetamine administration. Fifteen healthy human volunteers received a baseline scan, followed by second scan 75 min after receiving a 25 mg oral dose of dextro-amphetamine. Stimulated echo proton magnetic resonance spectroscopy (MRS) scans were preformed at 3.0 Tesla (T) in the dorsal medial prefrontal cortex (DMPFC). Metabolite data were adjusted for tissue composition and analyzed using LCModel. Twelve adult male rats were treated acutely with a 5-mg/kg intraperitoneal dose of dextro-amphetamine. After 1 h rats were decapitated and the brains were rapidly removed and frozen until dissection. Rat brains were dissected into frontal, temporal, and occipital cortical areas, as well as hippocampus. Tissue was analyzed using a Varian 18.8 T spectrometer. Metabolites were identified and quantified using Chenomx Profiler software. The main finding in the present study was that myo-inositol concentrations in the DMPFC of human volunteers and in the four rat brain regions were not altered by acute dextro-amphetamine. While it remains possible that the PI-cycle may be involved in the pathophysiology of bipolar disorder, it is not likely that the subjective and physiological of dextro-amphetamine are mediated, directly or indirectly, via alternations in myo-inositol concentrations.