Innate immunity relies on pattern recognition receptors to detect the presence of infectious pathogens. In the case of Gram-positive bacteria, binding of bacterial lipopeptides to TLR2 is currently regarded as an important mechanism. In the present study, we used the synthetic bacterial lipopeptide Pam3CysSK4, a selective TLR2 agonist, to induce meningeal inflammation in rodents. In a 6-h rat model, intrathecal application of Pam3CysSK4 caused influx of leukocytes into the cerebrospinal fluid (CSF) and induced a marked increase of regional cerebral blood flow and intracranial pressure. In wild-type mice, we observed CSF pleocytosis and an increased number of apoptotic neurons in the dentate gyrus 24 h after intrathecal challenge. Inflammation and associated neuronal loss were absent in TLR2 knockout mice. In purified neurons, cytotoxicity of Pam3CysSK4 itself was not observed. Exposure of microglia to Pam3CysSK4 induced neurotoxic properties in the supernatant of wild-type, but not TLR2-deficient microglia. We conclude that TLR2-mediated signaling is sufficient to induce the host-dependent key features of acute bacterial meningitis. Therefore, synthetic lipopeptides are a highly specific tool to study mechanisms of TLR2-driven neurodegeneration in vivo.