The long term of this research is to develop quantitative imaging methods for tissue bioenergetic studies. Towards this end it is proposed to construct and evaluate an instrument that will image the response of mitochondrial metabolites to ischemia and hypoxia in rabbit hearts in situ. Instrument operation is based on fluorescence emission from mitochondrial NADH in heart epicardium. These changes can be measured fluorometically and will be imaged at the surface of perfused and in situ rabbit hearts. In the research of Phase I, a video fluorometer will be constructed from a low light level video camera, frame grabber, and computer. Alterations in regional NADH concentrations will be analyzed from multiwavelength fluorescence and/or reflectance images. Procedures for compensating fluorescence emission for internal absorption due to blood volume changes will be developed to generate quantitative fluorescence images. The efficiency of the compensation will be tested with rabbit hearts in situ during procedures involving local ischemia or hypoxia. Proof of feasibility in Phase I will allow the range of fluorochromes to be expanded in Phase II to include pH, metal ion, and membrane potential sensitive dyes. The instrument will be of value for a wide range of investigators who require information concerning regional metabolic alterations in tissues or organs.
