Volume-sensitive chloride channels (ICl,vol) mediate doxorubicin-induced apoptosis through apoptotic volume decrease in cardiomyocytes

Apoptosis is associated with early changes in cell volume through a mechanism called apoptotic volume decrease (AVD). As volume-sensitive chloride channels (I-C1,I-vol) are known to play a key role in the regulation of cell volume, this study investigated the role of I-C1,I-vol and AVD in doxorubicin-induced apoptotic cell death in adult rabbit ventricular cardiomyocytes. Exposure of cardiomyocytes to 1 mum doxorubicin induced a rapid and significant reduction in cell volume of cardiomyocytes (average of 15%), i.e. AVD as well as increases in the early markers of apoptosis, annexin V labeling and caspase-3 activity. Doxorubicin also induced the activation of a current characterized as I-C1,I-vol on the basis of the external chloride sensitivity and pharmacological properties with the patch clamp technique. Doxorubicin-induced AVD and apoptosis were both abolished when cardiomyocytes were exposed to the I-C1,I-vol inhibitors 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) (0.1 mm) or indanyloxyacetic acid 94 (IAA-94) (10 mum). The crucial role of I-C1,I-vol during AVD and apoptosis was confirmed using C-2-ceramide, another pro-apoptotic compound. These results demonstrate that activation of I-C1,I-vol plays a major role in the mechanism leading to cell shrinkage and apoptosis-induced AVD by agents such as doxorubicin or C-2-ceramide in adult cardiomyocytes.