Brain ischemic tolerance is an endogenous neuroprotective mechanism, whereby an experience of non-lethal ischemic episode (preconditioning; PC) produces resilience to subsequent lethal ischemia. We previously showed that PC caused activation of astrocytes and a subsequent upregulation of P2X7 receptors, activation of which induced ischemic tolerance via upregulation of HIF-1α in astrocytes. P2X7 receptor requires relatively higher extracellular ATP concentrations (ATPo) for its activation. However, the PC-evoked increase in ATPo was not enough to activate P2X7 receptor. Here, we show that astrocytes have a unique mechanism of P2X7 receptor activation with lower ATPo, thereby leading to ischemic tolerance. It has been reported that NAD at lower ATPo could induce prolonged activation of P2X7 receptor via ART2-catalyzed ADP-ribosylation. Thus, we tested effect of NAD on astrocytic P2X7 receptors, and found that NAD increased HIF-1α in WT astrocytes but not in P2X7 receptor-deficient astrocytes in vitro. We also found that ART2 is selectively expressed and upregulated by PC in astrocytes. Taken together, our findings suggest that astrocytes could activate P2X7 receptors by their unique mechanisms, i.e., NAD/ART2/P2X7 signal pathways and induce ischemic tolerance.