Regulation of B cell receptor (BCR)-induced Ca2＋ signaling by CD40 co-stimulation was compared in long-term BCR-stimulated immature (WEHI-231) and mature (Bal-17) B cells. In response to long-term pre-stimulation of immature WEHI-231 cells to Ձ-IgM antibody (0.5∼48 hr), the initial transient decrease in BCR-induced [Ca2＋]i was followed by spontaneous recovery to control level within 24 hr. The recovery of Ca2＋ signaling in WEHI-231 cells was not due to restoration of internalized receptor but instead to an increase in the levels of PLCՃ2 and IP3R-3. CD40 co-stimulation of WEHI-231 cells prevented BCR-induced cell cycle arrest and apoptosis, and it strongly inhibited the recovery of BCR-induced Ca2＋ signaling. CD40 co-stimulation also enhanced BCR internalization and reduced expression of PLCՃ2 and IP3R-3. Pre-treatment of WEHI-231 cells with the antioxidant N-acetyl-L-cysteine (NAC) strongly inhibited CD40-mediated prevention of the recovery of Ca2＋ signaling. In contrast to immature WEHI-231 cells, identical long-term Ձ-IgM pre-stimulation of mature Bal-17 cells abolished the increase in BCR-induced [Ca2＋]i, regardless of CD40 co-stimulation. These results suggest that CD40-mediated signaling prevents antigen-induced cell cycle arrest and apoptosis of immature B cells through inhibition of sustained BCR-induced Ca2＋ signaling.