Tumor necrosis factor alpha (TNF α) and lymphotoxin alpha (LT α) induce pleiotropic cellular effects through low-affinity 55 kDa type-1 receptors (TNFR1, CD120a) and high-affinity 75 kDa type-2 receptors (TNFR2, CD120b). Both cytokines have potent biological effects on glial cells and are strongly implicated in the pathology of central nervous system (CNS) demyelinating diseases. However, to date, neither constitutive nor cytokine-induced TNFR expression by glial cells have been definitively characterized. We therefore characterized TNF receptors at the molecular, protein, and functional levels in rat astrocytes, microglia, and oligodendrocytes. Northern blotting demonstrated that all three types of glia constitutively transcribed a single TNFR1 mRNA. IFN γ increased transcript levels in all three types of glia, but TNF α increased levels only in oligodendrocytes. Microglia constitutively transcribed three distinct TNFR2 mRNAs, levels of which were increased by either IFN γ or TNF α. In contrast, astrocytes and oligodendrocytes constitutively transcribed nearly undetectable levels of TNFR2 mRNAs, and levels were not affected by IFN γ, TNF α, or oligodendrocyte maturation. Immunocytochemical staining of glial cells corroborated Northern data by demonstrating that glia express a parallel pattern of TNFR proteins on their cell surfaces. In co-cultures of microglia plated atop irradiated astrocytes, human TNF α (which, on mouse cells, binds TNFR1 exclusively) induced microglial cell proliferation, whereas murine TNF α (which binds both TNFRs) did not. Collectively, the data show that microglia, a primary source of TNF α at CNS inflammatory sites, express both TNFR1 and TNFR2, whereas astrocytes and oligodendrocytes, whose embryological origin differs from that of microglia, predominantly express TNFR1. TNF α increases expression of TNFR1 by oligodendrocytes whereas it increases expression of TNFR2 by microglia. Microglia proliferation data suggest that signals transduced through TNFR2 directly or indirectly inhibit signals transduced through TNFR1. Different patterns of TNFR expression by glia at sites of CNS inflammation may be critical in determining whether TNF has activational, proliferative, or cytotoxic effects on these cells.