The propagation of intercellular calcium waves has been studied as a mechanism by which cooperative cell activity can be achieved. In astrocytes two distinct pathways have been demonstrated: and intercellular pathway mediated by gap junctions, and an extracellular pathway mediated by P2-type nucleotide receptors. Recently, we showed that activation of astrocyte with the cytokine IL-1beta profoundly altered these astrocyte-astrocyte communication pathways, inducing a rapid loss of gap junction connectivity but an enhancement of P2 nucleotide receptor signaling associated with a shift in the functional P2Y receptor subtype(s) expressed. Conversely, blockade of P2 nucleotide receptors inhibited the high level activation of nuclear factor kappaB (NF-kappaB) mediated by IL-1beta, resulting in a significant inhibition of Il-1beta-inudced expression of tumor necrosis factor alpha IL-6, and type 2 nitric oxide synthase (NOS2). These data have led us to propose that in astrocytes, cytokine and P2 nucleotide receptor signaling pathways intersect such that cytokine signaling alters P2 receptor expression and function, and autocrine/paracrine signaling via P2 receptors regulates cytokine response elements. To test this hypothesis three aims are proposed. In the first the effect of cytokines on P2 nucleotide receptor subtype expression and function in cultured human astrocytes will be examined. The second will address the effect of P2 receptor signaling on the IL1beta and TNFalpha signaling cascades. In the third we will address the in vivo relevance of these observations by studying two well-defined models of CNS inflammation in mice with targeted gene deletions of the P2Y1 and P2Y2 receptors. In the CNS events such as inflammation, trauma, and stress lead to the release of high levels of extracellular nucleotides such as ATP, as well as cytokines such as IL1beta and TNFalpha, and are frequently associated with significant long-term CNS dysfunction. The overall goal of this project is to determine how inflammatory events affect astrocyte coordination and information processing, with the hope that an understanding of the role that purinergic receptors play in this process may provide new targets for therapeutic intervention resulting in maintenance/reestablishment of the normal functioning of the CNS environment.
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