The role of mitochondrially bound arginase in the regulation of urea synthesis -: Studies with [U-15N4]arginine, isolated mitochondria, and perfused rat liver

The main goal of the current study was to elucidate the role of mitochondrial arginine metabolism in the regulation of N-acetylglutamate and urea synthesis. We hypothesized that arginine catabolism via mitochondrially bound arginase augments ureagenesis by supplying ornithine for net synthesis of citrulline, glutamate, N-acetylglutamate, and aspartate. [U-N-15(4)] arginine was used as precursor and isolated mitochondria or liver perfusion as a model system to monitor arginine catabolism and the incorporation of N-15 into various intermediate metabolites of the urea cycle. The results indicate that similar to 8% of total mitochondrial arginase activity is located in the matrix, and 90% is located in the outer membrane. Experiments with isolated mitochondria showed that similar to 60-70% of external [U-N-15(4)] arginine catabolism was recovered as N-15-labeled ornithine, glutamate, N-acetylglutamate, citrulline, and aspartate. The production of N-15-labeled metabolites was time- and dose-dependent. During liver perfusion, urea containing one (Um+1) or two (Um+2) N-15 was generated from perfusate [U-N-15(4)] arginine. The output of Um+2 was between 3 and 8% of total urea, consistent with the percentage of activity of matrix arginase. Um+1 was formed following mitochondrial production of [N-15] glutamate from [alpha,delta-N-15(2)]ornithine and transamination of [N-15] glutamate to [N-15] aspartate. The latter is transported to cytosol and incorporated into argininosuccinate. Approximately 70, 75, 7, and 5% of hepatic ornithine, citrulline, N-acetylglutamate, and aspartate, respectively, were derived from perfusate [U-N-15(4)] arginine. The results substantiate the hypothesis that intramitochondrial arginase, presumably the arginase-II isozyme, may play an important role in the regulation of hepatic ureagenesis by furnishing ornithine for net synthesis of N-acetylglutamate, citrulline, and aspartate.