The plasma membrane Na+/Ca2+ exchange inhibitor KB-R7943 is also a potent inhibitor of the mitochondrial Ca2+ uniporter

Background and purpose: The thiourea derivative KB-R7943, originally developed as inhibitor of the plasma membrane Na+/Ca2+ exchanger, has been shown to protect against myocardial ischemia-reperfusion injury. We have studied here its effects on mitochondrial Ca2+ fluxes. Experimental approach. [Ca2+] 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 Ca2+ uniporter (MCU). In permeabilized HeLa cells, KB-R7943 inhibited mitochondrial Ca2+ uptake with a Ki of 5.5±1.3 μM (mean±S.D.). In intact cells, 10μM KB-R7943 reduced by 80% the mitochondrial [Ca2+] peak induced by histamine. KB-R7943 did not modify the mitochondrial membrane potential and had no effect on the mitochondrial Na+/Ca2+ exchanger. KB-R7943 inhibited histamine-induced ER-Ca2+ release in intact cells, but not in cells loaded with a Ca2+-chelator to damp cytosolic [Ca2+] changes. Therefore, inhibition of ER-Ca2+-release by KB-R7943 was probably due to the increased feedback Ca2+-inhibition of inositol 1,4,5-trisphosphate receptors after MCU block. This mechanism also explains why KB-R7943 reversibly blocked histamine-induced cytosolic [Ca2+] 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 Ca2+-overload during ischemia-reperfusion. In addition, the effects of KB-R7943 on Ca2+ homeostasis provide new evidence for the role of mitochondria modulating Ca2+-release and regenerative Ca2+-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