Imaging of myocardial metabolic activity by mitochondrial NADH fluorescence in a beating rat heart

To visualize myocardial NADH fluorescence, which reflects mitochondrial oxygen metabolism, the authors developed a fluorescence CCD system. It consists of an optical lens system with a barrier filter for NADH fluorescence, an ultraviolet light (365 nm) source, and a CCD video camera. The authors applied this system to 5 normal and 5 streptozotocin-induced diabetic Wistar rats. Heart was isolated and perfused retrograde from the aorta. Oxygen (O₂) saturation of perfusate was altered from 95% to 0% and then returned to 95%. Fluorescence intensity of each heart became brighter with different time course spatially indicating heterogeneous mitochondrial hypoxia developing after 0% O₂ saturated perfusate. Reversing to 95% O₂ perfusate, NADH fluorescence distribution became gradually patchy (bright: more hypoxic, dark: less hypoxic), indicating heterogeneous recovery process of mitochondrial functions. The location of patchy area was persistent throughout repeated procedures and was heart-specific in both normal and diabetic rats. In diabetic rats, the maximum brightness was higher, mean-time constant of hypoxic transition shorter and that of normoxic transition longer than in normal rats. In conclusion, the authors´ fluorescence CCD microscope revealed development of patchy hypoxic area and normoxic recovery process of myocardial mitochondria in rats. The faster transition to and the slower recovery from mitochondrial hypoxic state with higher maximum brightness in the diabetic heart indicate higher vulnerability to hypoxia than in the normal heart