Janus kinases/STAT pathway mediates cellular responses to certain oxidative stress stimuli and cytokines. Here we examine the activation of Stat1 and Stat3 in rat astrocyte cultures and its involvement in cell death. H(2)O(2), interferon (INF)-gamma and interleukin (IL)-6 but not IL-10 caused cell death. Stat1 was phosphorylated on tyrosine (Tyr)-701 after exposure to H(2)O(2), INF-gamma or IL-6 but not IL-10. Tyr-705 pStat3 was observed after H(2)O(2), IL-6 and IL-10. Also, H(2)O(2) induced serine (Ser)-727 phosphorylation of Stat1 but not Stat3. The degree of Tyr-701 pStat1 by the different treatments positively correlated with the corresponding reduction of cell viability. AG490, a Jak2 inhibitor, prevented Tyr-701 but not Ser-727, Stat1 phosphorylation. Also, AG490 inhibited Tyr-705 Stat3 phosphorylation induced by H(2)O(2) and IL-6 but did not prevent that induced by IL-10. Furthermore, AG490 conferred strong protection against cell death induced by INF-gamma, IL-6 and H(2)O(2). These results suggest that Jak2/Stat1 activation mediates cell death induced by proinflammatory cytokines and peroxides. However, we found evidence suggesting that AG490 reduces oxidative stress induced by H(2)O(2), which further shows that H(2)O(2) and/or derived reactive oxygen species directly activate Jak2/Stat1, but masks the actual involvement of this pathway in H(2)O(2)-induced cell death.