心筋におけるクレアチンキナーゼの新しい機能

抄録

Present study was designed to demonstrate intracellular radial gradients of myoglobin oxygen saturation and ATP in single cardiomyocytes with elevated oxygen demand. Intracellular ATP concentration was indirectly assessed from intracellular free Mg²⁺ concentration using a Mg²⁺ sensitive ratiometric fluorescent dye mag-fura-2 (MF2). A bright field fluorescence microscopy was newly manufactured for simultaneous measurements of myoglobin light absorption (spectrophotometry) and MF2 fluorescence (fluorometry). Uncoupler of oxidative phosphorylation (1 μM CCCP) was used to increase oxygen flux. While significant intracellular gradients of myoglobin oxygen saturation was demonstrated in CCCP treated cells at physiological extracellular Po₂, no significant heterogeneity was found in MF2 fluorescence. In contrast, in cardiomyocytes treated with 0.5 mM iodoacetamide (a blocker of creatine kinase) in addition to CCCP, gradients of MF2 fluorescence from the sarcolemma to the cell core (indicating radial gradients of ATP) were demonstrated. Such MF2 fluorescence gradients were not demonstrated when extracellular oxygen concentration was elevated to 20%. From these results, it is concluded that significant gradients of ATP may be produced in the isolated single cardiomyocyte when mitochondrial oxygen consumption is moderately elevated at physiological extracellular Po₂. However, such potential ATP heterogeneities are effectively buffered in the normal cardiomyocyte by ATP supplementations from creatine phosphate. Thus, present results propose a new function of creatine kinase in the myocardium. This study was supported by JSPS KAKEN #15390061. [J Physiol Sci. 2006;56 Suppl:S73]