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Cytokines regulate numerous cell processes, including connexin expression and gap junctional coupling. In this study, we examined the effect of ciliary neurotrophic factor (CNTF) on connexin43 (Cx43) expression and intercellular coupling in astrocytes. Murine cortical astrocytes matured in vitro were treated with CNTF (20 ng/ml), soluble ciliary neurotrophic factor receptor alpha (CNTFRalpha) (200 ng/ml), or CNTF-CNTFRalpha. Although CNTF and CNTFRalpha alone had no effect on Cx43 expression, the heterodimer CNTF-CNTFRalpha significantly increased both Cx43 mRNA and protein levels. Cx43 immunostaining correlated with increased intercellular coupling as determined by dye transfer analysis. By using the pharmacological inhibitor alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide (AG490), the increase in Cx43 was found to be dependent on the Janus tyrosine kinase/signal transducer and activator of transcription (JAK/STAT) pathway. Immunocytochemical analysis revealed that CNTF-CNTFRalpha treatment produced nuclear localization of phosphorylated STAT3, whereas CNTF treatment alone did not. Transient transfection of constructs containing various sequences of the Cx43 promoter tagged to a LacZ reporter into ROS 17/2.8 cells confirmed that the promoter region between -838 to -1693 was deemed necessary for CNTF-CNTFRalpha to induce heightened expression. CNTF-CNTFRalpha did not alter Cx30 mRNA levels, suggesting selectivity of CNTF-CNTFRalpha for connexin signaling. Together in the presence of soluble receptor, CNTF activates the JAK/STAT pathway leading to enhanced Cx43 expression and intercellular coupling.

molecular biology of the cell

The Complex of Ciliary Neurotrophic Factor-Ciliary Neurotrophic Factor Receptor α Up-Regulates Connexin43 and Intercellular Coupling in Astrocytes via the Janus Tyrosine Kinase/Signal Transducer and Activator of Transcription Pathway