Prolonged exposure of microglia to lipopolysaccharide modifies the intracellular signaling pathways and selectively promotes prostaglandin E2 synthesis

During inflammatory or degenerative processes microglial cells are likely to be exposed to activating agents that persist in brain parenchyma for prolonged periods. As our knowledge on microglial activation is largely based on in vitro studies in which microglial cultures are activated by a single administration of pro-inflammatory stimuli, we investigated the effects of repeated endotoxin (LPS) challenges on microglial functional state. Primary rat microglial cultures were subjected to one, two or three consecutive LPS-stimulation and the production of tumor necrosis factor-alpha (TNF-alpha), nitric oxide (NO), prostaglandin E-2 (PGE(2)) and 15-deoxy-Delta(12,14)-PGJ(2) (15d-PGJ(2)) measured. The ability of microglial cells to produce NO, TNF-alpha and 15d-PGJ(2) upon the first LPS challenge rapidly declined after the second and the third stimulations, whereas PGE(2) synthesis remained constantly elevated. Accordingly, the expression of inducible NO synthase decreased whereas cyclooxygenase-2 and microsomal PGE synthase remained up-regulated. The signaling pathways evoked by single or multiple LPS-stimulation were also profoundly different, when considering the activation of the transcription factors nuclear factor-kappa B and CREB, and of the p38 MAPK. Our observations suggest that prolonged exposure to LPS, and likely other activating agents, induces in microglia a functional state clearly distinct from that triggered by acute stimulation. The progressive down-regulation of pro-inflammatory molecules and the sustained release of PGE(2) could have important implications for the resolution of brain inflammation.