Douglas A. Hems

Hepatic action of vasopressin: Lack of a role for adenosine-3′,5′-cyclic monophosphate

Vasopressin (anti-diuretic hormone) can stimulate glycogen breakdown and gluconeogenesis in the rat liver, at concentrations (0. I 1 .O ng/ml) which can occur in the intact rat [l] . This is a third major systemic action of vasopressin in rats; its concentrationdependence [I] resembles that of the pressor action, rather than the anti-diuretic effect. The hepatic action of vasopressin therefore has significance in the intact animal, in conditions where plasma vasopressin levels are high, such as haemorrhagic shock [2] , and warrants further investigation. The question arises of the mechanism of the hepatic action of vasopressin. A simple theory would be that adenosine cyclic-3’,5’-monophosphate (cyclic AMP) is implicated. To investigate this possibility, concentrations of cyclic AMP have been measured in the liver of intact rats, and in the medium during perfusion, in conditions where glucogon and adrenalin produced a rise in concentration of this nucleotide [3-51. In this report, it is shown that vasopressin does not produce an increase in cyclic AMP in the liver or perfusate, in any of the conditions tested. Thus the hepatic effect of vasopressin provides a clearcut example of a short-term hormonal effect on mammalian metabolic processes, which is not mediated by cyclic AMP.

Role of extracellular calcium in the action of vasopressin on hepatic glycogenolysis

Vasopressin (anti-diuretic hormone) has been shown to stimulate glycogen breakdown in perfused rat and mouse liver preparations [1-4] at concentrations which can occur in the intact animal (i.e. 0.1-1.0 mU/ml) especially during haemorrhagic shock [5]. In vivo, vasopressin (0.3 units) can produce hyperglycaemia, and this effect is more potent than that of glucagon [6]. It is clear that vasopressin does not act via cyclicAMP [3,7] and it is therefore of interest to consider an alternative mechanism. Such a mechanism could be via cation-dependent events, as have been implicated in many types of cellular response to stimuli, including hormonal effects on hepatic gluconeogenesis, [8-11 ]. In the investigation reported here, hepatocyte suspensions have been employed to demonstrate that vasopressin acts on this population of liver cells, and also to characterise the dependence on extracellular calcium and potassium of vasopressin action. The data suggest that vasopressin action is markedly dependent on extracellular Ca 2÷, to a greater extent than is the action of glucagon or adrenalin. Results obtained with the perfused liver are also presented which support this conclusion.

Metabolic actions of vasopressin, glucagon and adrenalin in the intact rat

Metabolic effects of vasopressin, glucagan and adrenalin were compared, in intact rats, especially in regard to time courses of effects. Hyperglycaemia was transient in response to vasopressin, prolonged following adrenalin, and, suprisingly, was not discernible after glucagon, except in response to a very large dose. Vasopressin decreased and adrenalin increased, the plasma free fatty acid concentration; both hormones decreased the triacylglycerol level. Muscle glycogen concentrations, measured in heart, diaphragm and skeletal muscle, exhibited small changes, with complex time courses, following hormone administration. Vasopressin brought about a rapid but transient activation of heaptic glycogen phosphorylase which resembled that due to adrenalin. The activation by glucagon of phosphorylase was greater and more prolonged, despite the absence of hyperglycaemia. In response to vasopressin, there was in increase in plasma insulin. Incorporation of 14C from [14C]glucose into glycogen or fatty acids was not influenced by vasopressin. Taken together, these results may be explained by rapid metabolic action of vasopressin on hepatic glycogenolysis, whereas adrenalin has multiple prolonged actions.

The hepatic clearance of circulating native and asialo carcinoembryonic antigen by the rat

Abstract Native human carcinoembryonic antigen has a circulating half life of less than 5 min when injected intravenously into rats. The intact rat accumulated carcinoembryonic antigen almost exclusively in the liver. Trace amounts were found in spleen and lung. The half life of the native glycoprotein in a rat liver perfusion system was 28 min while that of the asialo glycoprotein was 11 min. In both cases the time course for removal of glycoprotein from the perfusate was first order. After perfusion (90 min), about 10% of the glycoprotein was found in the bile. The rapid uptake of the native glycoprotein suggests that the recognition of terminal galactose may not be the only factor involved in determining the hepatic assimilation of glycoproteins.

Control mechanisms in the acceleration of hepatic glycogen degradation during hypoxia

Hepatic glycogen metabolism in aerobic and hypoxic conditions has been assessed with respect to glycogenolysis, phosphorylase alpha activity and nucleotide content. Insulin did not inhibit glycogen breakdown nor stimulate lipogenesis in the aerobic perfused liver. Partial ischaemia induced glycogen breakdown, release of glucose and changes in nucleotide content in the perfused liver. Phosphorylase alpha content increased within 2 min in response to total ischaemia, in vivo and in the perfused liver. This change was paralleled by an increase in hepatic AMP. Glycogen synthase alpha activity decreased, as did the hepatic content of both cyclic AMP and cyclic GMP.

Hormonal Control of Intermediary Metabolism in Obese Hyperglycemic Mice: II. Levels of Plasma Free Fatty Acid and Immunoreactive Insulin and Liver Glycogen

In mice with genetic obesity, the plasma concentration of total free fatty acid (FFA) was not significantly raised, and no abnormality was detected in the response of the plasma level of FFA to noradrenaline. The plasma concentration of immunoreactive insulin (IRI) in the fed state was higher in obese than lean mice, even if glucose was given to lean mice. The content of liver glycogen was consistently higher in obese mice. Following deprivation of food, plasma concentration of IRI decreased in obese mice, and that of FFA rose at the same rate as in lean mice. In obese mice which had been fasted for twenty-four hours or fed on a restricted quantity of food, the plasma level of IRI remained higher than in lean mice. On oral administration of glucose to fasted mice, the decline in plasma FFA concentration, compared to that in animals which received saline, was similar in obese and lean mice. In response to glucose, the plasma IRI concentration increased rapidly in obese mice. These results suggest that there is no primary impairment of adipose tissue triglyceride breakdown (“lipolysis”) in genetically obese mice, and that the hypersecretion of insulin in response to ingestion of food may more closely reflect their primary disorder.

Inhibition by parathyroid hormone of glycogen synthesis in the perfused rat liver

Parathyroid hormone can exert effects on metabolic processes; thus it may stimulate renal gluconeogenesis [l-3] or adipose tissue lipolysis [4-61. The renal content of cyclic adenosine 3’5’ monophosphate (cyclic AMP) is increased in response to parathyroid hormone, [ 1,7] . Hormones which act in the above fashion on kidney or fat, might be expected, by analogy with e.g. adrenalin and glucagon, to stimulate hepatic glycogen breakdown. Therefore this possibility was tested in the perfused rat liver. Rather than following glucose output, glycogen metabolism was tested directly, to exclude effects on gluconeogenesis; thus net glycogen synthesis was measured, as previously described [9,10]. This process exhibits greater sensitivity to another ‘catabolic’ hormone, viz. vasopressin, than does glycogen breakdown in livers from fed rats [lo] , which was another reason for selecting glycogen synthesis for study. The results presented here demonstrate that parathyroid hormone can inhibit hepatic net glycogen synthesis. During the progress of these experiments, reports appeared of stimulation by parathyroid hormone of glucose output in hepatocytes [ 1 l] and of cyclic AMP formation by the liver [ 11,121. The present work complements these studies, in documenting the hormone concentration-dependence of the action on hepatic glycogen metabolism, in the perfused liver, i.e. in a preparation where hormone responses may be reasonably presumed to correspond closely to events in vivo . Male Sprague-Dawley rats (200 g) were starved for 48 h from 10.00 h. Net glycogen synthesis was measured by a sequential-biopsy procedure in livers perfused with 50 ml saline containing albumin, red cells, glucose and gluconeogenic precursors, as described previously [9,10]. Parathyroid hormone (Lot 74/286) was very kindly provided by Dr J. A. Parsons (National Institute of Medical Research, Mill Hill, London, NW7, UK), and Dr J. R. L. O’Riordan (Middlesex Hospital, London W.l., UK). This material contained, per ampoule, 170 gg highly purified hormone, and 259 mg mannitol. For use, it was dissolved in about 1 ml 1% (w/v) bovine serum albumin (‘Crystalline’, Sigma Ltd) which had been heated at 56°C (pH 7.0) for 2 h. When the entire ampoule of hormone was not used during a day’s perfusions, the solution (pH 4.0) was rapidly frozen (using liquid nitrogen) in small aliquots, being used subsequently after only one thaw.

Hepatic metabolism in normal and genetically obese mice

Abstract Aspects of hepatic metabolism have been investigated in normal and genetically obese ( ob/ob ) mice. The formation of glucose and lipids was studied with 14 C-labelled glycerol, lactate, or alanine. Glucose was the major product of these precursors. In obese mice, the concentration and turnover of glycerol in blood were increased, compared to lean mice. However, maximal rates of glycerol metabolism, and the glycerol tolerance curve, were not altered in ob/ob mice. Increased glycerol turnover in blood of ob/ob mice is thus a result of enhanced breakdown of extra-hepatic triglyceride. This inference was confirmed by the increased turnover of plasma free fatty acid in ob/ob mice, which was present despite the decreased capacity for ketogenesis in the perfused liver, and the absence of hyperketonaemia. The rates of formation of blood glucose and liver lipid from a trace dose of [ 14 C] lactate were increased in ob/ob mice compared to lean mice No such difference was observed with [ 14 C] alanine. There were no marked differences in blood amino acid concentrations between lean and obese mice. In general, results suggested that gluconeogenesis was not markedly increased in ob/ob mice, although there was moderate enhancement of glucose or glycogen formation from lactate or glycerol. The hepatic content of intermediary metabolites was measured in mice. Levels of glucose 6-phosphate, glycerol 1-phosphate and citrate were somewhat diminished in the liver of ob/ob mice, compared to lean mice. The redox state of the free NADP couple was more oxidised in ob/ob mouse liver, which exhibits enhanced lipogenesis.

Metabolism of adrenaline in the isolated perfused liver of the rat

Abstract The liver has a high concentration of the enzymes necessary for the metabolism of catecholamines. Experiments with the perfused liver were designed to investigate the rate of hepatic uptake of adrenaline from blood and its subsequent metabolism. Adrenaline was rapidly removed from the perfusion medium, but did not accumulate in the liver. Metabolites of adrenaline, however, appeared in the liver and were soon found at a concentration exceeding that of the adrenaline in the medium. The concentration of adrenaline metabolites was directly proportional to the duration of the perfusion and the concentration of adrenaline in the medium. The time course of accumulation of adrenaline metabolites resembled that of their subsequent disappearance from the liver in “washout” experiments (half-time about 35 min). The accumulation of adrenaline metabolites in the liver and the removal of adrenaline from the perfusion medium were markedly reduced by the presence of cortieosterone (10 μg/ml in the medium.

Actions of vasopressin-related peptides on glycogen metabolism in the perfused rat liver

Abstract Both [8-lysine] vasopressin and oxyloein inhibited glycogen accumulation in the perfused liver from starved rats, at concentrations of 30–1000 ng/ml and 20–900 ng/ml respectively. [1-Deamino-8- d -arginine] vasopressin caused glycogenolysis in the perfused liver from fed rats over the concentration range 5–100 ng/ml. These effects resemble those previously reported for [8-arginine] vasopressin: they are discussed with reference to the potency of action of vasopressin-related peptides on the liver.