The presence and localisation of dehydroepiandrosterone sulphotransferase was assessed in adult rat brain. The activity of the enzyme was investigated and was found to be approximately four times less than that in rat liver. Rat brain dehydroepiandrosterone sulphotransferase activity was measured as 1.39 ± 0.04 nm/μg/min. The protein expression of dehydroepiandrosterone sulphotransferase was evaluated using Western blotting. The presence of a band at 35 kDa signified the presence of this enzyme in adult rat brain. Finally, the localisation of this enzyme was investigated by immunohistochemistry and by assessing activity in nine regions of brain. Immunohistochemistry showed positive staining in the hippocampus and the dendate gyrus, and in the large neurons of the midbrain, indicating the presence of dehydroepiandrosterone sulphotransferase. Western blotting showed the presence of a band in all nine regions, and enzyme activity was highest in the hippocampus, frontal cortex, thalamus and basal ganglia. These results imply that dehydroepiandrosterone and dehydroepiandrosterone sulphate (DHEAS) have a defined role in brain function and are produced in situ independently of the peripheral endocrine system.