Pathways of Succinate Accumulation and Adenine Nucleotide Depletion in Cardiac Ischemia

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Project Summary/Abstract The energy utilized by the heart for contractions is primarily made by aerobic respiration. This process consists of oxidizing carbon substrates to fuel ATP synthesis. An essential substrate is oxygen which acts as the final electron acceptor. Without oxygen, ATP is rapidly depleted from the myocardium. There are a number of scenarios where the heart is exposed to hypoxic or anoxic conditions. Ischemia occurs when blood flow becomes restricted preventing oxygen delivery to part of the muscle. This occurs in ischemic heart disease. If blood flow is not restored in time, it can lead to irreversible damage, termed myocardial infarction. Organ transplantation is another scenario where the heart is away from blood supply during transportation. The amount of time the muscle is without oxygen determines the health of the tissue for the recipient. Situations such as these that involve an ischemic environment induce metabolic changes that damage the myocardium during reperfusion when oxygen is restored, known as ischemia-reperfusion injury. One such change is succinate accumulation. During reperfusion, the immense amount of succinate is responsible for reactive oxygen species (ROS) production that puts oxidative stress on cardiomyocytes. Another metabolic change seen during ischemia is the depletion of adenine nucleotides. This group includes AMP, ADP, and ATP and are all essential energy carriers in the myocardium. The depletion of these molecules results in impaired energy metabolism in the heart. Little is known about the mechanisms behind these metabolic changes. The goal of this project is to uncover the primary pathways of succinate accumulation and adenine nucleotide depletion in ischemic myocardium. The working hypothesis is the combined actions of the tricarboxylic acid (TCA) cycle, malate-aspartate shuttle (MAS), and the purine nucleotide cycle (PNC) provide the substrate utilized for succinate production and adenine nucleotides entering the PNC are shuttled into purine degradation pathways. Experiments involving anoxic isolated heart mitochondrial, ischemic ex vivo hearts, and transgenic rats will be used jointly with computational models of myocardial metabolism to test this hypothesis. The metabolic states of these different systems will be quantified by metabolomics and enzyme inhibitors that will be used to assess primary pathways of accumulation and depletion. Computational models will help with experimental design, refining, and testing hypotheses. The data from this project will have applications for both ischemic heart disease, organ transplantation, treatments for ischemia-reperfusion injury.