RANKL, a member of tumor necrosis factor (TNF) superfamily, regulates the differentiation, activation, and survival of osteoclasts through binding to its cognate receptor, RANK. RANK can interact with several TNF-receptor-associated factors (TRAFs) and activates signaling molecules including Akt, NF-κB, and MAPKs. Although the transient elevation of reactive oxygen species (ROS) by receptor activation has been shown to act as a cellular secondary messenger, the involvement of ROS in RANK signaling pathways has been not characterized. In this study, we found that RANKL stimulated ROS generation in osteoclasts. Pretreatment of osteoclasts with the antioxidants N-acetyl-l-cystein and glutathione reduced RANKL-induced Akt, NF-κB, and ERK activation. The reduced NF-κB activity by antioxidants was associated with decreased IKK activity and IκBα phosphorylation. In contrast, antioxidants did not prevent TNF-α-induced Akt and NF-κB activation. Pretreatment with antioxidants also significantly reduced RANKL-induced actin ring formation, required for bone resorbing activity, and osteoclast survival. Taken together, our results suggest that ROS act as mediators in RANKL-induced signaling pathways and cellular events.