The pathogenesis of morbidity associated with hyponatremia is postulated to be determined by the state of intracellular cerebral osmolytes. Previously inaccessible, these metabolites can now be quantitated by proton magnetic resonance spectroscopy. An in vivo quantitative assay of osmolytes was performed in 12 chronic hyponatremic patients (mean serum sodium 120 meq/liter) and 10 normal controls. Short echo time proton magnetic resonance spectroscopy of occipital gray and parietal white matter locations revealed dramatic reduction in the concentrations of several metabolites. In gray matter, myo-inositol was most profoundly reduced at 49% of control value. Choline containing compounds were reduced 36%, creatine/phosphocreatine 19%, and N-acetylaspartate 11% from controls. Similar changes were found in white matter. Recovery of osmolyte concentrations was demonstrated in four patients studied 8-14 wk later. These results are consistent with a reversible osmolyte reduction under hypoosmolar stress in the intact human brain and offer novel suggestions for treatment and monitoring of this common clinical event.