Following infection with Neospora caninum, BALB/c mice were shown to be resistant to an acute infection but developed a latent chronic infection. However, BALB/c background gamma interferon (IFN-)-deficient mice were sensitive to the acute infection. Since the immune response in IFN--deficient mice is scantly known, we examined the function of macrophages, major histocompatibility complex (MHC) class II expression, T-cell responses, and serum cytokine levels in the mice. All IFN--deficient mice died within 9 days of infection with N. caninum, whereas those treated with exogenous IFN- lived longer. Although N. caninum invaded various organs in both types of mice at the early stage of infection, the parasite was not detected in the brains of resistant hosts until 21 days postinfection (dpi). Peritoneal macrophages from IFN--deficient mice were activated by exogenous IFN- associated with inhibition of parasite growth and nitric oxide production as were those from BALB/c mice. IFN--deficient mice failed to increase MHC class II expression on macrophages. Moreover, BALB/c mice induced T-cell proliferation while IFN--deficient mice did not. However, in vivo treatment with exogenous IFN- induced up-regulated MHC class II expression in IFN--deficient mice. BALB/c mice treated with an antibody to CD4 showed an increase in morbidity and mortality after parasite infection. In serum, significant levels of IFN- and interleukin-4 (IL-4) were detected in resistant hosts, whereas IL-10 was detected in IFN--deficient mice. The levels of IL-12 in IFN--deficient mice were higher than those in BALB/c mice at 7 dpi. The present study indicates that early IFN- production has a crucial role in the activation of peritoneal macrophages for the induction of protective immune responses against N. caninum.