Dataset Information

Interactions of insulin, glucagon and dexamethasone in controlling the activity of glycerol phosphate acyltransferase and the activity and subcellular distribution of phosphatidate phosphohydrolase in cultured rat hepatocytes.

ABSTRACT: Rat hepatocytes were incubated in monolayer culture for 8 h. Glucagon (10nM) increased the total phosphatidate phosphohydrolase activity by 1.7-fold. This effect was abolished by adding cycloheximide, actinomycin D or 500 pM-insulin to the incubations. The glucagon-induced increase was synergistic with that produced by an optimum concentration of 100 nM-dexamethasone. Theophylline (1mM) potentiated the effect of glucagon, but it did not affect the dexamethasone-induced increase in the phosphohydrolase activity. The relative proportion of the phosphohydrolase activity associated with membranes was decreased by glucagon when 0.15 mM-oleate was added 15 min before the end of the incubations to translocate the phosphohydrolase from the cytosol. This glucagon effect was not seen at 0.5 mM-oleate. Since glucagon also increased the total phosphohydrolase activity, the membrane-associated activity was maintained at 0.15 mM-oleate and was increased at 0.5 mM-oleate. This activity at both oleate concentrations was also increased in incubations that contained dexamethasone, particularly in the presence of glucagon. Insulin increased the relative proportion of phosphatidate phosphohydrolase that was associated with membranes at 0.15 mM-oleate, but not at 0.5 mM-oleate. It also decreased the absolute phosphohydrolase activity on the membranes at both oleate concentrations in incubations that also contained glucagon and dexamethasone. None of the hormonal combinations significantly altered the total glycerol phosphate acyltransferase activity. However, glucagon significantly increased the microsomal activities, and insulin had the opposite effect. Glucagon also decreased the mitochondrial acyltransferase activity. There was a highly significant correlation between the total phosphatidate phosphohydrolase activity and the synthesis of neutral lipids from glycerol phosphate and 0.5 mM-oleate in homogenates of cells from all of the hormonal combinations. Phosphatidate phosphohydrolase activity is increased in the long term by glucocorticoids and also by glucagon through cyclic AMP. In the short term, glucagon increases the concentration of fatty acid required to translocate the cytosolic reservoir of activity to the membranes on which phosphatidate is synthesized. Insulin opposes the combined actions of glucagon and glucocorticoids. The long-term events explain the large increases in the phosphohydrolase activity that occur in vivo in a variety of stress conditions. The expression of this activity depends on increases in the net availability of fatty acids and their CoA esters in the liver.

SUBMITTER: Pittner RA

PROVIDER: S-EPMC1152646 | biostudies-other | 1985 Sep

REPOSITORIES: biostudies-other

ACCESS DATA

Similar Datasets

Project description:The subcellular distribution of Mg2+-dependent phosphatidate phosphohydrolase in rat adipocytes between a soluble and a membrane-bound fraction was measured by using both centrifugal fractionation and a novel Millipore-filtration method. The relative proportion of the phosphohydrolase associated with the particulate fraction was increased on incubation of cells with noradrenaline or palmitate. Insulin on its own decreased the proportion of the phosphohydrolase that was particulate and abolished the effect of noradrenaline, but not that of palmitate. The effect of noradrenaline on phosphohydrolase distribution was rapid, the effect being maximal within 10 min. Noradrenaline exerted this effect with a similar concentration-dependence to its lipolytic effect. Inclusion of albumin in homogenization buffers decreased the proportion of the phosphohydrolase that was particulate, but did not abolish the effect of noradrenaline. There was limited correlation between the proportion of the phosphohydrolase that was particulate and the measured rate of triacylglycerol synthesis in adipocytes incubated under a variety of conditions. Starvation, streptozotocin-diabetes and hypothyroidism decreased the specific activities of the phosphohydrolase and glycerolphosphate acyltransferase in homogenates from epididymal fat-pads. Restoration of these activities in the diabetic state was seen after administration of insulin over 2 days or, in the short term, within 2 h after a single administration of insulin. Administration of thyroxine over 3 days caused restoration of these activities in the hypothyroid state. Starvation and diabetes increased the proportion of the phosphohydrolase found in the microsomal fraction. This change was not seen when albumin was present in homogenization buffers. The possible role of fatty acids as regulators of the intracellular translocation of the phosphohydrolase, together with the role of this enzyme in the regulation of triacylglycerol synthesis in adipose tissue, is discussed.

Project description:Activities of phosphatidate phosphohydrolase and palmitoyl-CoA hydrolase were determined in cardiac subcellular fractions prepared from rabbits which has received tri-iodothyronine and from hamsters with hereditary cardiomyopathy (strain BIO 14.6). 1. Both mitochondrial and microsomal fractions of hyperthyroid rabbit hearts produced 4-5 times as much diacylglycerol 3-phosphate from glycerol 3-phosphate and palmitate as did those of euthyroid hearts. 2. Phosphatidate phosphohydrolase, measured with phosphatidate emulsion, was activated by 1mm-Mg(2+) in all but the mitochondrial fraction of euthyroid rabbit hearts. The activation was more pronounced in subcellular fractions isolated from hyperthyroid hearts, so that the measured activities were significantly increased above those of the controls. The highest activity was found in the microsomal and lysosomal fractions. 3. In the absence of Mg(2+) during incubation, the difference in phosphohydrolase activities between eu- and hyper-thyroid states was not significant. 4. The phosphohydrolase of subcellular fractions of control hamsters did not respond to addition of 0.5-8.0mm-Mg(2+). The enzyme from cardiomyopathic hearts was slightly inhibited by this bivalent cation and therefore significant increases in activity were observed only in the absence of Mg(2+) from the assay system. 5. The rate of reaction by soluble phosphatidate phosphohydrolase was similar regardless of the nature of the substrate. Both when microsomal-bound phosphatidate was used as the substrate and when phosphatidate suspension was used, the activity of soluble enzyme was lower than that of the microsomal and lysosomal enzymes measured with phosphatidate suspension; this was especially so when the assay was carried out in the absence of Mg(2+). Neither tri-iodothyronine nor cardiomyopathy influenced the soluble phosphohydrolase activity in the two species. 6. Neither tri-iodothyronine nor cardiomyopathy significantly changed palmitoyl-CoA hydrolase activities in subcellular fractions. 7. Microsomal diacylglycerol acyltransferase and myocardial triacylglycerol content were also unchanged in the hyperthyroid state.

Dataset Information

Interactions of insulin, glucagon and dexamethasone in controlling the activity of glycerol phosphate acyltransferase and the activity and subcellular distribution of phosphatidate phosphohydrolase in cultured rat hepatocytes.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets