Characterization of the cellular and biochemical mechanisms underlying the deleterious effects of IL-1beta to hypoxic neuronal injury

Date of Completion

January 2007

Keywords

Biology, Neuroscience

Degree

Ph.D.

Abstract

Changes in interleukin-β (IL-β) levels and/or signaling have been implicated in the pathogenesis of cerebral ischemia; however, the mechanism(s) by which this occurs are unknown. Therefore, the overall goal of this thesis project was to investigate the cellular and biochemical pathway(s) by which this cytokine contributes to neuronal injury. To do so, a suitable in vitro model system was developed utilizing murine mixed neuronal/astrocyte cortical cell cultures. In this model, IL-1β pre-treatment—simulating endogenous production of IL-1β following cerebral ischemia—potentiated hypoxic neuronal injury, mimicking events in the ischemic penumbra. The injury process coincided with an increase in extracellular glutamate levels and NMDA receptor-mediated 45Ca2+ uptake, and was inhibited by glutamate receptor antagonism. Utilizing pharmacologic and genetic approaches we showed that the detrimental effects of IL-1β in vitro were dependent on signaling through the IL-1 receptor type I (IL-1RI), a finding that was further confirmed by an in vivo approach, which demonstrated that IL-1RI-deficient mice were less susceptible to ischemic and excitotoxic injury. Utilizing chimeric cultures from IL-1RI-deficient mice, we then specifically identified astrocytes as the cell type mediating the ill effects of IL-1β, since the increased glutamate accumulation, as well as the enhanced vulnerability to hypoxia that followed IL-1β treatment, was not observed in chimeric cultures consisting of wild-type neurons plated on top of IL-1RI-deficient astrocytes. Furthermore, we showed that IL-1β-mediated potentiation of hypoxic neuronal injury was inhibited in the presence of the cystine glutamate exchanger (system xc−)/mGluR1 antagonists but not by selective mGluR1 antagonists, nor were mGluR1-deficient cultures protected, indicating a causative role for system xc − in the injury process. Last, we demonstrated that this is due to a selective increase in system xc− velocity induced by astrocytic IL-1RI signaling, a change that becomes neurotoxic when energy deprivation results in reduced glutamate uptake via the sodium-dependent glutamate transporters (system XAG−). These data identify alterations in system xc− as a previously undescribed mechanism in the development and progression of cerebral ischemic injury and describe system xc− as a novel target for the development of neuroprotective treatments following cerebral ischemia. ^

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