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The metabolic effects of glucagon and glucagon plus insulin on the isolated rat livers perfused with 10 mM sodium L-lactate as substrate were studied. Glucagon stimulated gluconeogenesis, ketogenesis and ureogenesis at the concentration used of 2.1 nM. The addition of insulin to give a glucagon-to-insulin ratio of 0.2 reversed all the glucagon effects.The glucagon enhancement of gluconeogenesis was accompanied by a rise in the cytosolic and mitochondrial state of reduction of the NAD system and a fall in the [ATP]/[ADP] ratio. The analysis of the intermediary metabolite concentrations suggested, as possible sites of glucagon action, the steps between pyruvate and phosphoenolpyruvate as well as the reactions catalyzed by phosphofructokinase and/or fructose bisphosphatase. All the changes in metabolite contents were abolished when insulin was present.Glucagon increased the intramitochondrial concentration of all the metabolites, whose intracellular distribution was calculated. The finding of a significant rise in the calculated intramitochondrial concentration of oxaloacetate points to pyruvate carboxylation as an important site of glucagon interaction with the gluconeogenic pathway. A primary event in the glucagon action redistributing intracellular metabolites seems to be the mitochondrial entry of malate. The possibility is discussed that the changes in metabolite cellular distribution were brought about by the increased cellular state of reduction caused by the hormone.Since an early study, in which the role of glucagon on hepatic gluconeogenesis was postulated [l -41, a considerable amount of information has been accumulated regarding its mechanism of action. However, the efforts do not seem to have been succesful, since as yet no satisfactory explanation has been found to elucidate the mechanism leading to the enhancement of the hepatic gluconeogenic flux under the different situations in which the hormone is known to be active.Recent work has pointed to the activation of some metabolic event between pyruvate and phosphoenolpyruvate as the cause for the observed effect of glucagon stimulation of the gluconeogenic flux from subEnzymes. Aconitate hydratase (EC 4.2

The effect of nucleotide energy levels in vivo on the different steps of protein synthesis has been studied. Hepatic anoxia was induced by interrupting the blood portal-vein flow. At 5 min of anoxia ATP fell to 59 % of the control values and the amino acid incorporation into protein was inhibited by more than 70%. This strong inhibition was not paralleled by polyribosomal breakdown. On the contrary, when fasted rats were used, at 5 min of anoxia the ribosomal state of aggregation was found to increase. Longer periods of anoxia resulted in a further decrease in triphosphonucleoside content and polyribosomal breakdown. Based on these results and other reports from the literature it is concluded that the K, for the GTP of the peptide-chain-elongation mechanism must be higher than the K,,, of the initiation step. This finding implies that variations of nucleotide levels in vivo within the physiological range may control protein synthesis at the elongation step without apparent changes in the polyribosomal profiles.Most of the cellular functions are somehow related to the energy level of the cell, since the phosphorylation state of the adenine nucleotides is known to play a central role in metabolic regulation [l-31. On this subject there are in the literature several reports pointing to a clear relationship between energy level of the cell and rates of protein synthesis. It was found how anoxia inhibited the incorporation of amino acids into the proteins of several isolated rat tissues [4-61, of several tissues of the intact animals [7] and of isolated cell suspensions [8]. Not only anoxia but also the perturbation of the energetic level of the cell by other means results in an altered rate of protein synthesis [9, lo]. Nevertheless, it is not clear yet which of the steps involved in polypeptide synthesis, initiation, elongation or both, are under this type of control [ll -141. The problem has been approached in this work by studying the ribosomal state of aggregation as reflected by the polyribosomal profiles. The rationale of this approach lies in the fact that under normal conditions initiation and elongation are both synchronized in such a way that any perturbation on only one of them should bring about variations in the ribosomal pattern of aggregation. For instance, it is knowrn that the use of inhibitors of peptide chain

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