Most previous studies on Schwann cell proliferation in vitro have used serum-containing media. This complicates the analysis of agents required for cell division since serum contains an ill-defined mixture of hormones and growth factors. Serum-free medium has therefore been used to analyse the response of Schwann cell to previously identified Schwann cell mitogens. Serum factors were not necessary for DNA synthesis in response to platelet-derived growth factor, basic fibroblast growth factor, or glial growth factor, provided they were used in combination with forskolin to elevate intracellular cAMP. Transforming growth factor beta 1, a Schwann cell mitogen in serum, was not mitogenic under these conditions. Neither the growth factors nor forskolin were effective when used alone. Growth control was analysed further using long-term cultured Schwann cells that had spontaneously immortalized. Measurements of endogenous cAMP levels in short- and long-term Schwann cells revealed that long-term cells had two to three times higher basal cAMP levels. As predicted by these findings, platelet-derived growth factor, basic fibroblast growth factor, and glial growth factor stimulated DNA synthesis in long-term cells without requiring costimulation by agents which elevate cAMP (while transforming growth factor beta 1 had no effect).(ABSTRACT TRUNCATED AT 250 WORDS)