Intracellular pH changes in rat brain tissue were investigated during low or high extracellular pH induced by acetazolamide or sodium bicarbonate, respectively. Intracellular pH was measured by 31P-MRS in the brain of spontaneously breathing rats under intraperitoneal sodium pentobarbital anaesthesia. Extracellular pH was calculated from the results of blood gas analysis. After intravenous injection of sodium bicarbonate (280 mg/kg), the extracellular pH rose significantly (p less than 0.05) from 7.47 +/- 0.06 to 7.82 +/- 0.15 (mean +/- SE). After administration of acetazolamide (50 mg/kg), the extracellular pH dropped significantly (p less than 0.05) from 7.45 +/- 0.02 to 7.34 +/- 0.03. Despite the changes in extracellular pH, the intracellular pH of rat brain did not change significantly under either condition. The following four factors are thought to contribute to the maintenance of intracellular pH in the normal brain: 1) production and consumption of H+ by brain metabolism, 2) physicochemical buffering, 3) transmembrane transport of H+ and its equivalent, 4) compensatory adaptation of circulatory factors. These mechanisms are not disturbed in the brain of rats that are breathing spontaneously, because the cerebral circulation and energy metabolism are preserved in the normal range.