Proton magnetic resonance spectroscopy (MRS) signals from lipids in brain have been observed to increase after ischemic brain injury. However, neither the chemical identity nor the cellular location of these lipids has been established. The aim of the present study was to identify the origin of MRS lipid signals in rat brain after temporary (90 min) middle cerebral artery occlusion (MCAO). Fatty acyl proton signals were detected by short-echo one and two dimensional (1)H MRS in superfused brain slices from the infarcted hemisphere 1-5 days after MCAO. The intensities of these signals were strongly correlated with the amount of triacylglyceride and cholesterol ester in lipid extracts from the samples (r(2)=0.96, P<0.05) and were not correlated with the amount of free fatty acids in the tissue. Histological staining of tissue revealed the presence of neutral lipid droplets in infarcted regions. Dual labeling by immunohistochemistry demonstrated that these droplets were localized to microglia/macrophage (OX-42-labeled cells). These results strongly suggest that (1)H MRS lipid signals from brain after stroke arise from microglia/macrophage phagocytosis of cellular membranes.