Erythropoietin (Epo) has been used for many years in neonates for the treatment of anemia of prematurity. Epo has also been proposed for treatment of neonatal brain injury, as mounting evidence suggests neuroprotective properties for Epo. However, Epo's neuroprotective mechanism of action is poorly understood. In this study we hypothesized that Epo may confer neuroprotection by enhancing cellular redox defense brought about by cellular glutathione (GSH). This was examined in cultures of differentiated cortical neural stem cells and using the B104 cell line as model systems. Our data shows that Epo causes a time- and dose-dependent increase in expression and activity of system Xc-, the transporter responsible for uptake of cystine for the production of glutathione. Cystine uptake increases 3-5 fold in differentiated neural stem cells and B104 cells treated with Epo. Exposure of cells to 100 μM kainate suppressed cellular GSH and caused excitotoxicity, but GSH levels and cell viability were completely restored by Epo in the continued presence of kainate. This rescue effect of Epo vanished if system Xc- was inhibited pharmacologically using S4-CPG in the presence of Epo leading to marked cell death of B104 cells and cultured mouse cortical neural stem cells. This could also be achieved using xCT siRNA to decrease xCT expression. This data suggests that system Xc- activity and protein expression are positively regulated by Epo directly explaining its neuroprotective effect. © 2010 Elsevier B.V. All rights reserved.