Increasing cAMP attenuates induction of inducible nitric-oxide synthase in rat primary astrocytes

Nitric oxide produced by inducible nitric-oxide synthase (iNOS) in different brain cells in response to various cytokines plays an important role in the pathophysiology of stroke and other neurodegenerative diseases. This study underlines the importance of cAMP in inhibiting the induction of NO production by lipopolysaccharide (LPS) and cytokines in rat primary astrocytes. Compounds (forskolin, 8-bromo-cAMP, and (Sp)-cAMP) that increase cAMP and activate protein kinase A (PKA) were found to inhibit LPS- and cytokine-mediated production of NO as well as the expression of iNOS, whereas compounds (H-89 and (Rp)-cAMP) that decrease cAMP and PKA activity stimulated the production of NO and the expression of iNOS in rat primary astrocytes. Forskolin, but not the inactive analogue 1,9-dideoxyforskolin, inhibited NO production and iNOS expression in a dose-dependent manner in astrocytes. The inhibition of LPS- and/or cytokine-induced NO production in rat C6 glial cells by forskolin suggest that similar to astrocytes, iNOS expression in C6 cells is also regulated by similar mechanisms. In contrast, in rat peritoneal macrophages the cAMP analogues stimulated the LPS- and cytokine-induced production of NO. In vitro, the PKA had no effect on iNOS activity in LPS-treated astrocytes or macrophages, suggesting that PKA modulates the intracellular signaling events associated with the induction of iNOS biogenesis rather than the post-translational modification of iNOS. The compounds which activate PKA activity, blocked the activation of NF-kappabeta in astrocytes but stimulated the activation of NF-kappabeta in macrophages. This differential regulation of NF-kappabeta activation in two different cell types (astrocytes and macrophages) by the same second messenger (cAMP) indicates that intracellular events or pathways in the activation of NF-kappabeta may be different. Moreover, this inhibition of iNOS expression in LPS- and cytokine-treated astrocytes by cAMP may be of therapeutic potential in NO-mediated cytotoxicity in neurodegenerative diseases.