There is growing appreciation that the resident glial cells of the CNS, in particular microglia and astrocytes, are important participants in the generation of inflammation during brain infection. However, the mechanisms by which these cell types perceive and respond to microbial pathogens is only recently becoming apparent with the discovery of highly conserved families of pattern recognition receptors. In the current study we demonstrate that microglia and astrocytes express members of the NOD-like family of intracellular receptors. We demonstrate that the NOD2 pathway represents a functional mechanism by which these cells can augment the responses mediated by other families of pattern recognition receptors, specifically members of the Toll-like receptor family. Furthermore, we show that the NOD2 pathway contributes to the generation of potentially damaging inflammation in response to N. meningitidis and B. burgdorferi, both in vitro and in vivo, and that the elimination of this pathway results in a decrease in the production of important inflammatory mediators. Our additional studies involving viral infection of microglia and astrocytes highlight the potential of these cell types to respond to a variety of microbial types. Additionally, this work also demonstrated the replication-dependent nature of glial responses to vesicular stomatitis virus, a model of rabies, which points to the use of replication-dependent intracellular receptors such a Rig-I. Taken as a whole, this study implicates intracellular pattern recognition receptors as potentially important contributors to the glial immune responses within the CNS.