Previous studies from this laboratory have shown that C-6 rat glioma cells (2B clone) exhibit specific phenotypic characteristics depending on passage in culture and that these populations respond differentially to addition of various exogenous compounds to the medium. Early passage (<25) C-6 glial cells express low glutamine synthetase activity (a marker for astrocytes) and with increasing cell passage (>70) C-6 glial cells express more astrocytic properties with respect to both glutamine synthetase (GS) and morphology. In this study, cells from both early (glioblastic) and late (astrocytic) passage were examined for their response to the phospholipid, platelet-activating factor (PAF). We found that PAF increased GS activity in early passage (glioblastic) cells and more importantly it increased GS activity in late passage cells, already committed to the astrocytic phenotype. Furthermore, cells from both passages failed to respond to addition of lyso-PAF, the non-biologically active analog of PAF, to the medium. By following the uptake of 3H-PAF into cells, we observed that >90% of the phospholipid was taken into the cells within the first hour of incubation. We compared the PAF effects with that of dibutyryl cyclic AMP (dBcAMP) and R020-1724, a phosphodiesterase inhibitor. Cells from the early passage responded to both dBcAMP and R020-1724 treatments with a significant increase in GS activity whereas cells from the late passage showed no significant change, confirming earlier reports from this laboratory. These findings indicate that the response of C-6 glioma cells to PAF (at least in the late passage) is not mediated via cyclic AMP. We suggest that in early passage cells PAF promotes expression of the astrocytic phenotype and in late passage cells PAF mediates a gliosis-type response.