Primary neonatal murine astrocyte cultures were used to investigate the role of these glial cells in host defense of the central nervous system (CNS) against Toxoplasma gondii. For comparison, neonatal murine microglial cells were also studied. Microscopic analyses revealed that uptake of T. gondii into astrocytes was parasite-driven and was followed by uniform intracellular survival and multiplication of tachyzoites. Treatment of astrocytes with interferon (IFN)-γ and lipopolysaccharide (LPS) had no apparent effect on the survival or growth of T. gondii. Microglia, on the other hand, had both an intrinsic phagocytosis-associated anti toxoplasma activity and a nitric oxide-dependent inhibitory activity that was up-regulated by IFN-γ and LPS. The results of this study suggest that in contrast to microglial cells, astrocytes may provide a safe harbor within the CNS for T. gondii.