Co-localization of glial fibrillary acidic protein (GFAP) and radioactivity was examined after intraventricular injection of [3H]5-HT in adult rat brains. Radioactivity localized over GFAP-positive astrocytes was seen, especially when image-enhancing techniques were applied to the data. Also slices prepared from astrogliotic hippocampi of rats pretreated with kainic acid showed a twofold increased uptake of [3H]5-HT compared to control slices. This indicates that the uptake of [3H]5-HT seen in primary astrocyte cultures also occurs for astrocytes in situ. Also, as with astrocyte cultures, only some of the GFAP(+) astrocytes in situ showed localization of radioactivity, supporting the concept of intraregional heterogeneity of astrocyte functions.