THE EFFECTS OF ADENINE NUCLEOTIDES ON PYRUVATE METABOLISM IN RAT LIVER

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Abstract
The effects of adenine nucleotides on pyruvate mebabolism by isolated liver cells and isolated mitochondria were investigated. The amount of pyruvate carboxylated was estimated by determining the tricarboxylic acid-cycle intermediates, glutamate and aspartate accumulating in the incubation medium. The extent of pyruvate oxidation was assessed by measuring oxygen uptake and the yield of C14 O2 from [1-C14] pyruvate and [2-C14] pyruvate. When catalytic amounts of adenine nucleotides (1-2mM) were added to suspensions of isolated liver cells incubated with pyruvate an ATP:ADP ratio greater than 6:1 was maintained. Both puruvate oxidation to acetyl-CoA and the oxidation of acetyl-CoA through the tricarboxylic acid cycle were stimulated but pyruvate carboxylation was not affected. The production of acetyl-CoA exceeded the capacity of the cells for the oxidation of acetyl-CoA and the excess was converted into ketone bodies. If a low ATP:ADP ratio was maintained in isolated cells or mitochondria by incubating them with dinitrophenol or hexokinase pyruvate carboxylation was grossly inhibited, oxygen uptake depressed and ketone-body formation stimulated. Measurement of oxaloacetate concentrations confirmed that under these conditions oxaloacetate was rate-limiting for the oxidation of acetyl-CoA via the tricarboxylic acid cycle. The inclusion in the incubation medium of fumarate (1.25 mM) completely prevented the ketogenic action of dinitrophenol or hexokinase. When ADP (5mM) was added to a suspension of isolated liver cells incubated with pyruvate an actual ADP concentration of about 1 mM was attained. This brought about effects on pyruvate metabolism similar to those obtained with dinitrophenol or hexokinase. These results support the concept that the relative concentrations of adenine nucleotides within the liver cell may play a role in governing the rates of pyruvate oxidation and carboxylation. In addition, they provide further evidence that the availability of oxaloacetate in the liver cell can play a key role in determining whether acetyl-CoA arising from pyruvate is oxidized through the tricarboxylic acid cycle or converted into ketone bodies.