System xc- Activity and Astrocytes are necessary for interleukin-1β-mediated hypoxic neuronal injury

The purpose of this study was to elucidate the cellular/biochemical pathway(s) by which interleukin-1 beta (IL- 1 beta) contributes to the pathogenesis of hypoxic-ischemic brain damage. In vivo, IL-1 receptor type I (IL-1RI)-deficient mice showed smaller infarcts and less neurological deficits than wild-type animals after a 90 min reversible middle cerebral artery occlusion. In vitro, IL-1 beta mediated an enhancement of hypoxic neuronal injury in murine cortical cultures that was lacking in cultures derived from IL-1RI null mutant animals and was blocked by the IL-1 receptor antagonist or an IL-1RI blocking antibody. This IL-1 beta- mediated potentiation of hypoxic neuronal injury was associated with an increase in both cellular cystine uptake ([cystine](i)) and extracellular glutamate levels ([ glutamate](e)) and was prevented by either ionotropic glutamate receptor antagonism or removal of L-cystine, suggesting a role for the cystine/glutamate antiporter (System x(c)(-)). Indeed, dual System x(c)(-)/metabotropic glutamate receptor subunit 1 ( mGluR1) antagonism but not selective mGluR1 antagonism prevented neuronal injury. Additionally, cultures derived from mGluR1-deficient mice exhibited the same potentiation in injury after treatment with IL-1 beta as wild-type cultures, an effect prevented by System x(c)(-)/mGluR1 antagonism. Finally, assessment of System x(c)(-) function and kinetics in IL-1 beta-treated cultures revealed an increase in velocity of cystine transport (V-max), in the absence of a change in affinity (Km). Neither the enhancement in [cystine](i), [glutamate](e), or neuronal injury were observed in chimeric cultures consisting of IL-1RI(+)/(+) neurons plated on top of IL-1RI(-/-) astrocytes, highlighting the importance of astrocyte-mediated alterations in System x(c)(-) as a novel contributor to the development and progression of hypoxic neuronal injury.