Glutamine accumulation in rat kidney was investigated by giving large loads of gluta, mate and aspartate and measuring glutamine changes in seven completely defined portions of the nephron and in small patches from the thin limb area and from the papilla. Both glutamate and aspartate (which is partly coverted to glutamate) caused increases up to sixfold in glutamine which were mainly confined to the proximal straight tubule. Plasma glutamine is minimally affected. In contrast, large glutamine loads doubled plasma glutamine and caused about a doubling of glutamine concentrations in all parts of the nephron, but resulted in no greater change in the proximal straight tubule than elsewhere. Simultaneous inulin and glutamine measurements make it seem likely that the proximal convoluted tubule is a major site of glutamine reabsorption. The low glutamine concentrations in these cells (only 40% higher than that in plasma) indicate a very small gradient at the antiluminal surface. We suggest that the proximal straight tubule is the major site of glutamine synthesis but that it may not be an important site of glutamine breakdown.