Rates of ammonia formation from six amino acids by hepatocytes and liver mitochondria were compared with the rate of ammonia excretion by individual fish. Glutamine and asparagine are the most ammoniagenic substrates for hepatocytes and glutamine and glutamate, for isolated liver mitochondria. The main site of ammonia release is the mitochondrion. Ammonia is formed mainly via deamidation or transdeamination. Relatively small amounts of aspartate are formed from glutamate and glutamine by the fish liver mitochondria. This apparent "uncoupling" of glutamate transamination may represent an adaptation of carnivorous fish to the utilization of amino acids as a major energy source. More than 90% of the ammonia formed from glutamate and glutamine by isolated mitochondria are elaborated into the medium. Exiting ammonia is accompanied by a proton, but the source of this proton appears not to be the electrogenic proton pump or the carboxylate of exiting glutamic acid.
To determine the potential site(s) of fatty acid synthesis and source(s) of reducing equivalents, the activities of the cytoplasmic NADPH producing enzymes--isocitrate dehydrogenase (IDH), malic enzyme (ME), glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH), and of aconitase, ATP-citrate lyase (CCE) and malate dehydrogenase (MDH) were measured in homogenates of liver, intestine, visceral fat, red muscle and white muscle of eels (Anguilla rostrata) fed beef liver or worms, or fasted for 2 to 6 months. There were no differences in enzyme activities between eels fed beef liver or fasted for 2 months. Eels fed worms had significantly greater G6PDH activity than fasted eels. Liver size and hepatosomatic index decreased in fasted eels, but lipid content per gram of liver or muscle increased. Based on the total activities of the NADPH producing enzymes, the liver appeared to be the primary organ for lipogenesis, although the intestine contained active lipogenic enzymes as well. In the liver, IDH had the lowest Km (NADP) and highest activity of the NADP-dehydrogenases. In the liver cytoplasm, the low activities of CCE and ME and the presence of an active aconitase, with a 20-fold greater affinity than CCE for citrate, suggest tha citrate cleavage is unimportant and that IDH is a major source of reducing equivalents. The source of carbon for fatty acid synthesis is discussed in relation to these conclusions.
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