Dallas G. Clark

Effects of fatty acid oxidation on glucose utilization by isolated hepatocytes

We have studied the inhibitory action of long‐ and short‐chain fatty acids on hepatic glucose utilization in hepatocytes isolated from fasted rats. The rates of hepatic glucose phosphorylation and glycolysis were determined from the tritiated products of [2‐3H] and [6‐3H]glucose metabolism, respectively. The difference between these was taken as an estimate of the ‘cycling’ between glucose and glucose‐6‐phosphate. In the presence of 40 mM glucose this cycling was estimated at 0.68 μ;molmin/g wet wt. Glucose phosphorylation was unaffected during palmitate and hexanoate oxidation to ketone bodies but glycolysis was inhibited. The rate of glucose cycling was increased during this phase to 1.25 μmol/min > g. Following the complete metabolism of the fatty acids, glycolysis was reinstated and cycling rates returned to control levels. Hepatic glucose cycling appears to be an important component of the glucose/fatty acid cycle.

The Glycogen Storage Disease (gsd/gsd) Rat

Publisher Summary This chapter discusses the glycogen storage disease (gsd/gsd) rat. The glycogen storage diseases (GSDs) or glycogenoses are a group of genetically determined conditions that are characterized by a deficiency of one of the enzymes concerned with the degradation or synthesis of cellular glycogen; the defect may result in the presence of abnormal concentrations of this polysaccharide in one or more organs or tissues and the accumulation of glycogen having an unusual molecular structure. Defects of glycogen metabolism in humans were first reported in the 1920s; however, the fact that these diseases were caused by specific enzyme deficiencies was not appreciated until 30 years later. At the present time, at least 12 different forms of human GSD are recognized; in 10 of these, the deficient enzyme has been identified. There are many biochemical, genetic, and morphological similarities between the condition present in the gsd/gsd rat and human GSD Type IX. In both, the primary deficiency is a lack of hepatic phosphorylase b kinase that is transmitted as a recessive trait. The gsd/gsd rat model, moreover, has the added advantage that the disease is present in every member of an otherwise healthy, normally reproducing colony of common laboratory animals.

Energy expenditure and nutrient intake in long-distance runners

Abstract Apparent daily energy expenditure and nutrient intake of twelve male and eleven female athletes who run more than 70 km/wk were assessed using seven day diet and activity diaries. The mean daily energy expenditure (males 4055 kcal, females 2935 kcal) exceeded the mean daily energy intake (males 3485 kcal, females 2100 kcal) by ≈15% and ≈40% in the males and females respectively. There were no significant differences between the sexes with respect to the proportions of energy obtained from each of the major nutrients; carbohydrate provided 55.0% of total daily energy, protein 13.5%, fat 28.0% and alcohol 3.5%. These results indicate that these long distance runners have adopted a low fat, moderate to high carbohydrate diet which is quite similar to that recommeded for athletes.

A glycogen storage disease in rats. Morphological and biochemical investigations.

SummaryLiver and heart from a substrain of the NZR/Gd rat in which there is an inherited deficiency of liver phosphorylaseb kinase was examined by light and electron microscopy and compared to material from a related, but normal substrain.Hepatic tissue differed markedly from that of control animals. Hepatocytes contained more than twice as much free glycogen and visible lipid. Glycogen particles had an abnormal appearance and some glycogen was sequestered within large, membrane-bound vesicles. Hepatocyte lysosomes were increased by a third and mean cell volume by more than half. Lobular architecture was distorted by the presence of enlarged, irregularly-shaped hepatocytes. Free glycogen was present in the space of Disse and sinusoids and within lysosomes in Kupffer cells. There were increased amounts of collagen in the space of Disse. The changes resemble those described in human glycogen storage disease IXa.A study of hepatic tissue from fasted rats showed that affected animals have an impaired ability to mobilise their liver glycogen stores. An increase in visible lipid also occurred in affected, fasted animals.Cardiac tissue appeared to be normal.

Adrenergic regulation of pyruvate kinase and gluconeogenesis in hepatocytes from phosphorylase kinase-deficient (gsdgsd) rats

Abstract Hepatocytes were prepared from a strain of rats deficient in hepatic phosphorylase b kinase and were used to assess the role of this enzyme in the adrenergic regulation of pyruvate kinase and gluconeogenesis. Epinephrine (10 μM) stimulated glucose output and gluconeogenesis from 1.8 mM lactate but did not significantly affect the concentration of hepatocyte glycogen. In addition epinephrine treatment led to an inhibition of pyruvate kinase. The stimulation of gluconeogenesis and the inhibition of pyruvate kinase by epinephrine were blocked by both α- and β-antagonists: similar effects with epinephrine were observed in cells from control animals. It is concluded that mechanisms for the adrenergic regulation of pyruvate kinase and gluconeogenesis are similar in hepatocytes from both phosphorylase kinase-deficient and normal rats.

Some effects of different extracellular proteins on oxygen consumption and heat production in isolated rat hepatocytes

When hepatocytes prepared from 24-h-fasted rats were washed, suspended and incubated in Krebs-Henseleit bicarbonate-buffered saline, the endogenous rates of O2 consumption and heat production were 2.13 +/- 0.13 mumol/min per g wet wt. and 1.00 +/- 0.05 J/min per g wet wt. respectively. The inclusion of 2.5% (w/v) defatted and dialysed bovine serum albumin in either the cell suspension (washing) buffer or the cell incubation buffer produced a 20-25% increase in O2 consumption and heat production: these rates were increased by an additional 20-25% when the albumin (2.5%) was present in both the cell suspension and the cell incubation buffers. There was an inverse relationship between the increases in O2 consumption and heat production and the leakage of lactate dehydrogenase from the isolated hepatocytes: the inclusion of purified bovine serum albumin decreased lactate dehydrogenase leakage from 40% to 15% of total enzyme content. The calorimetric-respirometric ratios for hepatocytes incubated both in the absence (-461 +/- 19 kJ/mol O2) and presence (-477 +/- 8 kJ/mol O2) of the purified protein are very similar to the theoretical, thermochemically derived oxycaloric equivalents.

Inhibition of the substrate cycle glucose: Glucose 6-phosphate by physiological concentrations of fructose in perfued rat liver

Summary Livers from fed rats were perfused with whole rat blood containing glucose labelled uniformly with 14 C and specifically with 3 H at position 2. Infusion of fructose at a concentration of 1.8 μmol/ml of blood significantly depressed perfusate glucose concentrations and increased the apparent utilization of both radioactive glucoses. The substrate cycle glucose:glucose 6-phosphate, as assessed by comparison of the apparent rates of utilization of [U- 14 C] and [2- 3 H] glucose and the actual rates, determined from glucose specific radioactivities during the perfusion, was almost completely inhibited by fructose.