Volume-sensitive outwardly rectifying (VSOR) anion channels play a key role in a variety of essential cell functions including cell volume regulation, cell death induction and intercellular communications. We previously demonstrated that, in cultured mouse cortical astrocytes, VSOR channels are activated in response to an inflammatory mediator, bradykinin, even without an increase in cell volume. Here we report that this VSOR channel activation must be mediated firstly by 'nanodomains' of high [Ca2+]i generated at the sites of both Ca2+ release from intracellular Ca2+ stores and Ca2+ entry at the plasma membrane. Bradykinin elicited a [Ca2+]i rise, initially caused by Ca2+ release and then by Ca2+ entry. Suppression of the [Ca2+]i rise by removal of extracellular Ca2+ and by depletion of Ca2+ stores suppressed the VSOR channel activation in a graded manner. Quantitative RT-PCR and suppression of gene expression with small interfering RNAs indicated that Orai1, TRPC1 and TRPC3 channels are involved in the Ca2+ entry and especially the entry through TRPC1 channels is strongly involved in the bradykinin-induced activation of VSOR channels. Moreover, Ca2+-dependent protein kinases Cα and β were found to mediate the activation after the [Ca2+]i rise through inducing generation of reactive oxygen species. Intracellular application of a slow Ca2+ chelator, EGTA, at 10 mM or a fast chelator, BAPTA, at 1 mM, however, had little effect on the VSOR channel activation. Application of BAPTA at 10 mM suppressed significantly the activation to one-third. These suggest that the VSOR channel activation induced by bradykinin is regulated by Ca2+ in the vicinity of individual Ca2+ release and entry channels, providing a basis for local control of cell volume regulation and intercellular communications.