The effects of REV5901 on intracellular calcium signalling in freshly isolated bovine articular chondrocytes

REV5901 is an inhibitor of regulatory volume decrease (RVD) a mechanotransduction pathway regulating cell volume in response to hypotonicity, with protective properties upon chondrocyte trauma impact in situ. As the mechanism of action of REV5901 is unknown and changes in intracellular calcium ([Ca2+](j)) have been linked to REV5901-loading, we investigated the effects of REV5901 on a known calcium signalling pathway. Upon REV5901 loading, there was significant increase in [Ca2+](j) reaching 37.97 +/- 5.67%, above basal levels which was reduced to 27.86 +/- 3.15% in the presence of 2 mmol/l EGTA. In the presence of U73122 or neomycin there was a decrease in calcium with inhibition factors (IF) of 0.39 +/- 0.09 and 0.37 +/- 0.08, respectively, whereas rottlerin abolished the REV5901-induced [Ca2+](j) rise. The role of calcium channels in contributing to the REV5901-induced calcium rise was investigated whereby the calcium rise was inhibited in the absence of extracellular sodium and by the addition of Gd3+ and Ruthenium red. These data show a phospholipase C beta 3-dependent release of calcium from intracellular stores as well as a sodium calcium exchanger-mediated influx in response to REV5901 loading, suggesting a potential role for calcium signalling in mediating the action of REV5901 in chondrocytes.