The plasma membrane Na + /Ca 2+ exchange inhibitor KB‐R7943 is also a potent inhibitor of the mitochondrial Ca 2+ uniporter

Background and purpose: The thiourea derivative KB‐R7943, originally developed as inhibitor of the plasma membrane Na + /Ca 2+ exchanger, has been shown to protect against myocardial ischemia‐reperfusion injury. We have studied here its effects on mitochondrial Ca 2+ fluxes. Experimental approach. [Ca 2+ ] in cytosol, mitochondria and endoplasmic reticulum (ER), and mitochondrial membrane potential were monitored using both luminescent (targeted aequorins) and fluorescent (fura‐2, tetramethylrhodamine ethyl ester) probes in HeLa cells. Key results: KB‐R7943 was also a potent inhibitor of the mitochondrial Ca 2+ uniporter (MCU). In permeabilized HeLa cells, KB‐R7943 inhibited mitochondrial Ca 2+ uptake with a Ki of 5.5±1.3 μM (mean±S.D.). In intact cells, 10μM KB‐R7943 reduced by 80% the mitochondrial [Ca 2+ ] peak induced by histamine. KB‐R7943 did not modify the mitochondrial membrane potential and had no effect on the mitochondrial Na + /Ca 2+ exchanger. KB‐R7943 inhibited histamine‐induced ER‐Ca 2+ release in intact cells, but not in cells loaded with a Ca 2+ ‐chelator to damp cytosolic [Ca 2+ ] changes. Therefore, inhibition of ER‐Ca 2+ ‐release by KB‐R7943 was probably due to the increased feedback Ca 2+ ‐inhibition of inositol 1,4,5‐trisphosphate receptors after MCU block. This mechanism also explains why KB‐R7943 reversibly blocked histamine‐induced cytosolic [Ca 2+ ] oscillations in the same range of concentrations required to inhibit MCU. Conclusions and Implications: Inhibition of MCU by KB‐R7943 may contribute to its cardioprotective activity by preventing mitochondrial Ca 2+ ‐overload during ischemia‐reperfusion. In addition, the effects of KB‐R7943 on Ca 2+ homeostasis provide new evidence for the role of mitochondria modulating Ca 2+ ‐release and regenerative Ca 2+ ‐oscillations. Search for permeable and selective MCU inhibitors may yield useful pharmacological tools in the future. British Journal of Pharmacology (2007) 151 , 647–654; doi: 10.1038/sj.bjp.0707260