This study evaluated the relative importance of perfusion and diffusion mechanisms in compartmental models of blood:tissue helium exchange in the brain. Helium has different physiochemical properties from previously studied gases and is a common diluent gas in underwater diving where decompression schedules are based on theoretical models of inert gas kinetics. Helium kinetics across the cerebrum were determined during and after 15 minutes of helium inhalation, at separate low and high steady states of cerebral blood flow in seven sheep under isoflurane anaesthesia. Helium concentrations in arterial and sagittal sinus venous blood were determined using gas chromatographic analysis and sagittal sinus blood flow was monitored continuously. Parameters and model selection criteria of various perfusion-limited or perfusion-diffusion compartmental models of the brain were estimated by simultaneous fitting of the models to the sagittal sinus helium concentrations for both blood flow states. Purely perfusion-limited models fitted the data poorly. Models that allowed a diffusion-limited exchange of helium between a perfusion-limited tissue compartment and an unperfused deep compartment provided better overall fit of the data and credible parameter estimates. Fit to the data was also improved by allowing counter current diffusion shunt of helium between arterial and venous blood. These results suggest a role of diffusion in blood:tissue helium equilibration in brain.