The effect of diabetes and the redox potential on amino acid content and release by isolated rat hemidiaphragms.

The free amino acid content of diaphragm muscles of control and diabetic rats was studied 5 days after the injection of streptozotocin. Muscles were prepared for analysis either immediately after sacrifice or following incubation in balanced salt solution containing 5.5 mM glucose, with or without an electron acceptor, 0.02 mM methylene blue. Diaphragms of diabetic rats contained significantly more free taurine, glutamate, and branched chain amino acids than the controls at sacrifice, and significantly less glutamine, serine, asparagine, lysine, arginine, histidine, threonine, citrulline, and carnosine. Alanine decreased in plasma of diabetic rats but not in diaphragms before incubation. Hemidiaphragms of diabetic rats produced less alanine and more glutamate during incubation than controls. After incubation they contained less than half as much alanine and glutamine and twice as much glutamate than the controls, having released approximately 40% less alanine and 25% more glutamate into the medium than the controls. Glutamine release was not significantly different between the two groups. Methylene blue increased the free alanine content in the tissue water as well as alanine release by control and by diabetic muscles; the glutamate content of muscles decreased concomitantly. The effects of methylene blue were greater in the diabetic group. Branched chain amino acid release by diabetic muscles decreased during incubation with methylene blue. Muscles of diabetic rats contained more alpha-ketoglutarate than the controls after incubation with or without methylene blue. Methylene blue increased the alpha-ketoglutarate content of muscles and its release into the medium, the effect being greater in diabetics than in controls. Hemidiaphragms from diabetic rats released less pyruvate during incubation than controls, while lactate release by the two groups was not significantly different. Incubation with methylene blue caused a marked increase in pyruvate release by diabetic muscles, and a lesser stimulation in controls; lactate release increased in both groups. After incubation the lactate/pyruvate ratio in muscles was lower in the methylene blue treated group. The in vitro effect of 0.02 mM phenazine methosulfate on alanine production was similar to that of methylene blue. The data is compatible with the hypothesis that the NADH/NAD ratio may exert a restraining effect on alanine production and release by muscle. The progressive increase in this ratio may play a role in the eventual deceleration of gluconeogenesis during a prolonged fast and may restrain this process in uncompensated diabetes.