Detection of Mitochondrial ATP-Sensitive Potassium Channels in Rat Cardiomyocytes

Detection of Mitochondrial ATP-Sensitive Potassium Channels in Rat Cardiomyocytes

Mitochondrial ATP-sensitive potassium (mitoK<SUB>ATP</SUB>) channels play a role in early and late ischemic preconditioning. Nevertheless, the subunit composition of mitoK<SUB>ATP</SUB> channels remains unclear. In this study, we investigated the subunit composition of mitoK<SUB>ATP</SUB> channels in mitochondria isolated from rat cardiac<FONT COLOR=RED><SPAN STYLE= font-size:11pt; > </SPAN>myocytes. Mitochondria were visualized using the red fluorescence probe, Mitrotracker Red, while mitoK<SUB>ATP</SUB> channels were visualized using the green fluorescence probe, glibenclamide-BODIPY. The immunofluorescence confocal microscopy revealed the presence of Kir6.1, Kir6.2 and SUR2 present in the cardiac mitochondria. Western blot analysis was carried to further investigate the nature of mitoK<SUB>ATP</SUB> channels. For SUR proteins, a 140-kDa immunoreactive band that corresponded to SUR2, but no SUR1 was detected. For Kir6.2, three bands (∼44, ∼46, and ∼30 kDa) were detected, and a specific ∼46-kDa immunoreactive band corresponding to Kir6.1 was also observed. These observations suggest that the subunits of mitoK<SUB>ATP</SUB> channels in rat myocytes include Kir6.1, Kir6.2, and a<FONT COLOR=RED><SPAN STYLE= font-size:11pt; > </SPAN>SUR2-related sulfonylurea-binding protein.