Using simultaneous autoradiography and immunofluorescence we have investigated the functional heterogeneity amongst oligodendrocytes and astrocytes in primary mouse central nervous system (CNS) culture as expressed by differences in their ability to accumulate gamma-[3H]aminobutyric acid [( 3H]GABA) and D-[3H]aspartate. We have used a range of specific antibodies that identify oligodendrocytes and astrocytes, from precursor to fully mature cells, to address the question of whether all neuroglial cells are capable of expressing this function. Our results showing that A2B5-, 03-, and galactocerebroside-positive cells became heavily labelled with these two neuroactive amino acids, whereas cells expressing the myelin proteins 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNP) and myelin basic protein (MBP) did not, demonstrate that this capacity is already present in oligodendrocytes at early developmental stages but may not extend to fully mature cells. Astrocytes in culture exhibited a large degree of variability with respect to their ability to transport GABA and D-aspartate. When grown in either serum-containing or serum-free hormone supplemented culture medium two morphologically distinct of glial fibrillary acidic protein (GFAP)-positive astrocyte were identified, process-bearing and epithelioid. Process-bearing cells became heavily labelled with the amino acids under both growth conditions, whereas, data showed that although epithelioid astrocytes were not, or only lightly, labelled with either amino acid in serum-containing cultures, when grown in serum-free culture medium they became more heavily labelled. Thus the expression, in culture, by epithelioid astrocytes, of one of the functions attributed to these cells is largely dependent on growth conditions.