Reactive oxygen species (ROS) and reactive nitrogen species (RNS) direct the activation of distinct signaling pathways that determine cell fate. In this study, the pathways activated and the mechanisms by which ROS and RNS control the viability of pancreatic β-cells were examined. Although both nitric oxide and hydrogen peroxide (H₂O₂) induce DNA damage, reduce cell viability, and activate AMPK, the mechanisms of AMPK activation and cell death induction differ between each reactive species. Nitric oxide activates the unfolded protein and heat shock responses and MAPK kinase signaling, whereas H₂O₂ stimulates p53 stabilization and poly(ADP-ribose) polymerase (PARP) activation but fails to induce the unfolded protein or heat shock responses or MAPK activation. The control of cell fate decisions is selective for the form of stress. H₂O₂-mediated reduction in β-cell viability is controlled by PARP, whereas cell death in response to nitric oxide is PARP independent but associated with the nuclear localization of GAPDH. These findings show that both ROS and RNS activate AMPK, induce DNA damage, and reduce cell viability; however, the pathways controlling the responses of β-cells are selective for the type of reactive species.