Roles of the P2X7 receptor in C6 astroglioma: in vitro and in vivo studies

The purinergic P2X7 receptor (P2X7R) is an ionotropic adenosine triphosphate(ATP) receptor which is closely linked with pathological conditions in the central nervous system (CNS). Gliomas are the most common primary brain tumors with presently no cures. The roles of the P2X7R in these diseases have not been previously studied and in this work, I have used the rat C6 glioma as an experimental model system to investigate expression and function of the P2X7R in vitro and in vivo. The in vitro study has examined expression of the P2X7R in C6 cells and the involvement of this receptor in mediating cell functional responses. C6 glioma cells were found to express the P2X7R at both mRNA and protein levels. The P2X7Ragonist, 2', 3 '-(benzoy1-4-benzoy1)-ATP (BzATP) induced an increase in intracellularCa2+ concentration, an effect which was largely inhibited by periodate-oxidized ATP(OxATP), an irreversible P2X7R antagonist. BzATP treatment of C6 cells also resulted in ethidium bromide dye uptake indicating pore formation was induced byP2X7R activation. Chronic exposure of C6 cells to BzATP showed up-regulation of several pro-inflammatory factors including the chemokines monocyte chemoattractant protein-1 (MCP-1) and interleukin-8 (IL-8) and the angiogenic factor vascular endothelial growth factor (VEGF) suggesting the P2X7R in C6 cells is involved in mediating inflammation in tumors. In addition, BzATP treatment was found to enhance wound-induced cell migration, an effect which was inhibited in the presence of OxATP, or another P2X7R antagonist, Brilliant Blue G (BBG). The in vivo study examined whether pharmacological modulation of P2X7R with BBG altered tumor growth. C6 glioma cells were implanted into the striatum of rat brain and in situ P2X7R expression was shown to be associated with glioma cells and resident microglia. Preliminary results have indicated that inhibition of P2X7R leads to a reduced volume of brain tumors formed by transplanted C6 cells. The overall results from this study demonstrate the novel finding that C6 glioma cells express functional P2X7R and suggest pharmacological modulation of theP2X7R could serve as an effective strategy to inhibit the development and progression of brain tumors.Medicine, Faculty ofAnesthesiology, Pharmacology and Therapeutics, Department ofGraduat