Ouabain activates NFκB through an NMDA signaling pathway in cultured cerebellar cells

• Published in: Neuropharmacology 2013-10, Vol.73, p.327-336
• Main Authors: de Sá Lima, L., Kawamoto, E.M., Munhoz, C.D., Kinoshita, P.F., Orellana, A.M.M., Curi, R., Rossoni, L.V., Avellar, M.C.W., Scavone, C.
• Format: Article
• Language: English
• Subjects: Animals; Bdnf; Brain-derived neurotrophic factor; Brain-Derived Neurotrophic Factor - metabolism; Cerebellum - drug effects; Cerebellum - metabolism; Dizocilpine Maleate - pharmacology; Dose-Response Relationship, Drug; Enzyme Inhibitors - pharmacology; Excitatory Amino Acid Antagonists - pharmacology; Flavonoids - pharmacology; Gene Expression Regulation - drug effects; Il-1β; Interleukin-1beta; NF-kappa B - metabolism; NFκB; NMDA; OUA; Ouabain - antagonists & inhibitors; Ouabain - pharmacology; Polyenes - pharmacology; Polyunsaturated Alkamides - pharmacology; Primary Cell Culture; Pyrazoles - pharmacology; Pyrimidines - pharmacology; Rats; Receptors, N-Methyl-D-Aspartate - agonists; Receptors, N-Methyl-D-Aspartate - antagonists & inhibitors; Signal Transduction - drug effects; Time Factors; Tnf-α; Tumor Necrosis Factor-alpha - metabolism
• ISSN: 0028-3908; 1873-7064
• DOI: 10.1016/j.neuropharm.2013.06.006

Na,K-ATPase, an ion pump, has been shown to interact with other proteins in signaling complexes in cardiac myocytes, renal and glial cells, and several other cell types. Our previous in vivo studies indicated that intrahippocampal administration of ouabain (OUA), an inhibitor of Na,K-ATPase, induces NFκB activation, leading to an increase in mRNA levels of target genes of this transcription factor in the rat hippocampus. The present work investigated whether OUA can regulate NF-κB in primary cultured rat cerebellar cells. Cells were treated with different concentrations of OUA (1, 10 or 100 μM) for different periods of time (1, 2 and 4 h). OUA induced a time- and concentration-dependent activation of NFκB (peak of activation: 10 μM, 2 h), involving both p50/p65 and p50/p50 NFκB dimers. OUA (10 μM, 2 h) induced upregulation of tumor necrosis factor α (Tnf-α), interleukin-1β (Il-1β), and brain derived neurotrophic factor (Bdnf) mRNA levels. Both NFκB activation and gene expression activation induced by OUA (10 μM) were abolished when cells were pre-treated for 20 min with MK-801 (N-Methyl-D-Aspartate (NMDA) receptor antagonist), manumycin A (farnesyltransferase inhibitor), PP-1(Src-family tyrosine kinase inhibitor) and PD98059 (mitogen-activated protein kinase (MAPK) inhibitor). OUA (10 μM) alone or in the presence of MK-801, PP-1, PD98059 did not cause cell death or DNA fragmentation. These findings suggest that OUA activates NFκB by NMDA-Src-Ras-like protein through MAPK pathways in cultured cerebellar cells. This pathway may mediate an adaptive response in the central nervous system. •OUA induced a time- and dose-dependent activation of NFκB (p50/p65 and p50/p50).•OUA induced upregulation of Tnf-α, Il-1β, and Bdnf mRNA levels.•NFκB activation and gene expression by OUA were blocked by NMDA-Src-MAPK cascade.•OUA activates NFκB by NMDA-MAPK pathway in cultured cerebellar cells.