The role of arylalkylamine N-acetyltransferase (AANAT) in neuronal functioning has been suggested based on biochemical assays; only scarce evidence indicates neuronal expression of the mRNA encoding for this enzyme that catalyzes the conversion of serotonin into N-acetylserotonin. Using a quantitative reverse transcriptase polymerase chain reaction (RT-PCR) assay with internal standards, and an in-situ RT-PCR hybridization assay we found evidence for the expression of AANAT in the rat brain. In the localization studies, the most prominent AANAT mRNA signal was found in the granule neurons of the hippocampus, the olfactory bulb, and the cerebellum, and in the gray matter of the spinal cord. Diurnal differences in AANAT mRNA content were observed in the pineal gland but not in the hippocampus; the content of AANAT mRNA was lower both in the pineal gland and the hippocampus of old (24 months) compared with young (2 months) rats. These data are consistent with the hypothesis that AANAT may play a physiological role in mammalian central nervous system neurons. Further studies are warranted into the possible functional significance of neuronal expression of AANAT mRNA.