A variety of pharmacological agents (i.e. agonists and inhibitors) were used in time course studies to examine the signal transduction pathway associated with the NaCl-induced upregulation of antioxidant enzyme activity was examined. NaCl stress was shown to initiate an increase in superoxide levels, and the more NaCl tolerant callus tissue showed significant increases in antioxidant enzyme activity. Studies with fluridone, an inhibitor of ABA synthesis, suggested that both ABA-dependent and ABA-independent signaling pathways are operative. Studies with calcium channel blockers, a calcium ionophore, and protein kinase inhibitors indicate that both the mobilization of calcium and the activation of protein kinases are involved in the signal transduction pathway. An inhibitor of phospholipase C blocked the NaCl-induced upregulation of antioxidant enzyme activity, while the addition of phosphatidylinositol, myoinositol 1,4,5-triphosphate (IP₃), and diacylgycerol resulted in an increase in antioxidant enzyme activity, suggesting that phosphoinositides also play a role in the signal transduction pathway. A tentative signal transduction pathway for the NaCl-induced upregulation of antioxidant enzyme activity is provided.