Mechanisms of interleukin-1β-induced Ca2+ signals in mouse cortical astrocytes:: roles of store- and receptor-operated Ca2+ entry

Many neurodegenerative disorders are accompanied by chronic glial activation, which is characterized by the abundant production of proinflammatory cytokines, such as IL-1 beta. IL-1 beta disrupts Ca2+ homeostasis and stimulates astrocyte reactivity. The mechanisms by which IL-1 beta induces Ca2+ dysregulation are not completely defined. Here, we examined how acute and chronic ( 24-48 h) treatment with IL-1 beta affect Ca2+ homeostasis in freshly dissociated and primary cultured mouse cortical astrocytes. Cytosolic free Ca2+ concentration ([Ca2+](cyt)) was measured with fura-2 using digital imaging. An acute application of 10 ng/ml IL-1 beta induced Ca2+ mobilization from intracellular stores and activated storeoperated Ca2+ entry (SOCE) and receptor-operated Ca2+ entry (ROCE) in both freshly dissociated and cultured actrocytes. Treatment of cultured astrocytes with IL-1 beta for 24 and 48 h elevated resting [Ca2+](cyt), decreased Ca2+ store content [associated with sarco(endo) plasmic reticulum Ca2+-ATPase 2b downregulation], and augmented ROCE. Based on evidence that receptor-operated, but not store-operated Ca2+ channels are Ba2+ permeable, Ba2+ entry was used to distinguish receptor-operated Ca2+ channels from store-operated Ca2+ channels. ROCE was activated by the diacylglycerol analog, 1-oleoyl-2-acetylsnglycerol (OAG). In the presence of extracellular Ba2+, OAG-induced elevations of cytosolic Ba2+ (fura-2 340-to-380-nm ratio) were significantly larger in astrocytes treated with IL-1 beta. These changes in IL-1 beta-treated astrocytes correlate with augmented expression of transient receptor potential cation channel ( TRPC) 6 protein, which likely mediates ROCE. Knockdown of the TRPC6 gene markedly reduced ROCE. The data suggest that IL-1 beta-induced dysregulation of Ca2+ homeostasis is the result of enhanced ROCE and TRPC6 expression. The disruption of Ca2+ homeostasis appears to be an upstream component in the cascade of IL-1 beta-activated pathways leading to neurodegeneration.