CD8(+) memory T cells are abundant and are activated in a near-synchronous manner by infection, thereby providing a unique opportunity to evaluate the coordinate functional and phenotypic changes that occur in vivo within hours of viral challenge. Using two disparate virus challenges of mice, we show that splenic CD8(+) memory T cells rapidly produced IFN-γ in vivo; however, within 18-24 h, IFN-γ synthesis was terminated and remained undetectable for ≥ 48 h. A similar on/off response was observed in CD8(+) memory T cells in the peritoneal cavity. Cessation of IFN-γ production in vivo occurred despite the continued presence of immunostimulatory viral Ag, indicating that the initial IFN-γ response had been actively downregulated and that the cells had been rendered refractory to subsequent in vivo Ag contact. Downregulation of IFN-γ synthesis was accompanied by the upregulation of inhibitory receptor expression on the T cells, and ex vivo analyses using synthetic peptides revealed a concurrent hierarchical loss of cytokine responsiveness (IL-2, then TNF, then IFN-γ) taking place during the first 24 h following Ag contact. Thus, within hours of virus challenge, CD8(+) memory T cells display the standard hallmarks of T cell exhaustion, a phenotype that previously was associated only with chronic diseases and that is generally viewed as a gradually developing and pathological change in T cell function. Our data suggest that, instead, the "exhaustion" phenotype is a rapid and normal physiological T cell response.