The present work sets out to investigate how Ca(2+) regulates the volume-sensitive taurine-release pathway in HeLa cells. Addition of Ca(2+)-mobilizing agonists at the time of exposure to hypotonic NaCl medium augments the swelling-induced taurine release and subsequently accelerates the inactivation of the release pathway. The accelerated inactivation is not observed in hypotonic Ca(2+)-free or high-K(+) media. Addition of Ca(2+)-mobilizing agonists also accelerates the regulatory volume decrease, which probably reflects activation of Ca(2+)-activated K(+) channels. The taurine release from control cells and cells exposed to Ca(2+) agonists is equally affected by changes in cell volume, application of DIDS and arachidonic acid, indicating that the volume-sensitive taurine leak pathway mediates the Ca(2+)-augmented taurine release. Exposure to Ca(2+)-mobilizing agonists prior to a hypotonic challenge also augments a subsequent swelling-induced taurine release even though the intracellular Ca(2+)-concentration has returned to the unstimulated level. The Ca(2+)-induced augmentation of the swelling-induced taurine release is abolished by inhibition of calmodulin, but unaffected by inhibition of calmodulin-dependent kinase II, myosin light chain kinase and calcineurin. The effect of Ca(2+)-mobilizing agonists is mimicked by protein kinase C (PKC) activation and abolished in the presence of the PKC inhibitor Gö6850 and following downregulation of phorbol ester-sensitive PKC isoforms. It is suggested that Ca(2+) regulates the volume-sensitive taurine-release pathway through activation of calmodulin and PKC isoforms belonging to the novel subclass (nPKC).