TNF‐α/IFN‐γ‐induced iNOS expression increased by prostaglandin E2 in rat primary astrocytes via EP2‐evoked cAMP/PKA and intracellular calcium signaling

Abstract Astrocytes, the most abundant glia in the central nervous system (CNS), produce a large amount of prostaglandin E 2 (PGE 2 ) in response to proinflammatory mediators after CNS injury. However, it is unclear whether PGE 2 has a regulatory role in astrocytic activity under the inflamed condition. In the present work, we showed that PGE 2 increased inducible nitric oxide synthase (iNOS) production by tumor necrosis factor‐α and interferon‐γ (T/I) in astrocytes. Pharmacological and RNA interference approaches further indicated the involvement of the receptor EP2 in PGE 2 ‐induced iNOS upregulation in T/I‐treated astrocytes. Quantitative real‐time polymerase chain reaction and gel mobility shift assays also demonstrated that PGE 2 increased iNOS transcription through EP2‐induced cAMP/protein kinase A (PKA)‐dependent pathway. Consistently, the effect of EP2 was significantly attenuated by the PKA inhibitor KT‐5720 and partially suppressed by the inhibitor (SB203580) of p38 mitogen‐activated protein kinase (p38MAPK), which serves as one of the downstream components of the PKA‐dependent pathway. Interestingly, EP2‐mediated PKA signaling appeared to increase intracellular Ca 2+ release through inositol triphosphate (IP3) receptor activation, which might in turn stimulate protein kinase C (PKC) activation to promote iNOS production in T/I‐primed astrocytes. By analyzing the expression of astrocytic glial fibrillary acidic protein (GFAP), we found that PGE 2 alone only triggered the EP2‐induced cAMP/PKA/p38MAPK signaling pathway in astrocytes. Collectively, PGE 2 may enhance T/I‐induced astrocytic activation by augmenting iNOS/NO production through EP2‐mediated cross‐talk between cAMP/PKA and IP3/Ca 2+ signaling pathways. © 2006 Wiley‐Liss, Inc.