An impaired ability to regulate the activation of microglia by fractalkine (CX(3)CL1) leads to persistent neuroinflammation and behavioral alterations following lipopolysaccharide (LPS) challenge. While these responses are usually transient, LPS injection caused prolonged depressive-like behavior in fractalkine receptor deficient mice (CX(3)CR1(-/-)) that was associated with exaggerated microglial activation and induction of the tryptophan (TRP) degrading enzyme indoleamine 2,3-dioxygenase (IDO). IDO activation and subsequent generation of neuroactive kynurenine metabolites may have a pivotal role in the development of depression. Therefore, the purpose of this study was to determine the extent to which LPS-induced depressive-like behavior in CX(3)CR1(-/-) mice was dependent on IDO activation. CX(3)CR1(-/-) mice were implanted prior to LPS challenge with a slow release pellet of 1-methyl-tryptophan (1-MT), a competitive inhibitor of IDO. Here we show that the depressive-like behavior evident in CX(3)CR1(-/-) mice 72h after LPS injection was abrogated by inhibition of IDO. LPS also decreased body weight and locomotor activity in CX(3)CR1(-/-) mice, but these effects were independent of 1-MT. Consistent with the increased metabolism of TRP by IDO, the ratio of 3-hydroxykynurenine (3-HK) to TRP was increased in the brain 72h after LPS. Increased serotonin (5-HT) turnover was also evident in the brain. The LPS-associated increases in both 3-HK:TRP and 5-HIAA:5-HT ratios were prevented by the inhibition of IDO. Last, IDO blockade attenuated microglial activation in the prefrontal cortex and hippocampus 72h after LPS. Collectively these data indicate that LPS-induced IDO activation contributes to persistent microglial activation and depressive-like behavior in CX(3)CR1(-/-) mice.