An immortalized clonal cell line (AP-16) has been established from glial cultures obtained from neonatal mouse cerebra by multipassages under serum-free conditions. Immunofluorescent experiments showed that AP-16 cells expressed a marker for glial progenitors (A2B5) and did not express markers for oligodendrocytes (galactocerebroside) or mature astrocytes (glial fibrillary acidic protein: GFAP). Treatment with transforming growth factor-beta1 (TGF-beta1) or fetal calf serum (FCS) for 2 days induced AP-16 cells to differentiate into A2B5-negative, GFAP-positive, phenotypically mature astrocytes. AP-16 cells depended on epidermal growth factor for survival, and their growth was inhibited by FCS. These results indicate that AP-16 cells retained the properties of astrocyte progenitors. An enzyme-linked immunosorbent assay showed that AP-16 cells synthesized fibronectin and laminin, and that the expression of fibronectin was increased by TGF-beta1. AP-16 cells should be useful for studying the roles of TGF-beta1 in the differentiation of astrocyte progenitors.