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Effect of adrenaline on 32 P incorporation into rat fat-cell phospholipids.

ABSTRACT: 1. The phospholipid composition of fat-cells prepared from rat epididymal fat-pad was determined. 2. The incorporation of [(32)P]P(i) into the phospholipids of fat-cells incubated in glucose-free medium and the effect of adrenaline and of alpha- and beta-adrenergic blocking agents, were studied. 3. Incorporation of [(32)P]P(i) into fat-cell phospholipid increased with time; incubation with adrenaline resulted in increased incorporation that was related to the concentration of adrenaline. 4. The pattern of incorporation of [(32)P]P(i) into the individual phospholipids of fat-cells after incubation for 1h was determined; adrenaline (5.4mum) resulted in increased incorporation into phosphatidylcholine. 5. Incubation of fat-cells with propranolol (34mum) and adrenaline (5.4mum) resulted in abolition of adrenaline-stimulated lipolysis; there was a decrease in the specific radioactivity of phosphatidylcholine and an increase in the specific radioactivity of phosphatidylethanolamine, phosphatidic acid, phosphatidylinositol and cardiolipin compared with cells incubated with adrenaline alone. 6. Incubation of fat-cells with phenoxybenzamine (0.1mm) and adrenaline (5.4mum) resulted in stimulation of lipolysis, and in diminished specific radioactivities of phosphatidylcholine, phosphatidic acid, phosphatidylinositol, phosphatidylglycerol and choline plasmalogen compared with cells stimulated with adrenaline alone.

SUBMITTER: Stein JM

PROVIDER: S-EPMC1173805 | biostudies-other | 1972 Jul

REPOSITORIES: biostudies-other

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Project description:1. Glyceride biosynthesis from glycerol phosphate and [1-(14)C]palmitate was studied in liver homogenates of rats that were fed ad libitum or starved for 36-40hr. The changes in enzyme activity were related to total DNA content or total liver homogenate as these were found to be equivalent and to be the most meaningful parameters. 2. In liver homogenates from fed rats, labelled palmitate was incorporated mainly into phosphatidate (58% of the total incorporation into lipids), diglycerides (25%) and triglycerides (16%), whereas monoglycerides, cholesterol esters and phospholipids other than phosphatidate were labelled only to a small extent. Addition of particle-free supernatant to full homogenates increased the total incorporation of palmitate by 45% and the pattern of incorporation altered to 53% incorporated into triglycerides, 24% into diglycerides and 17% into phosphatidate. This result suggested that, in liver homogenates, phosphatidate phosphohydrolase (EC 3.1.3.4) may be rate-limiting in the biosynthesis of glycerides via the glycerol phosphate pathway. 3. Upon starvation, the amount of palmitate incorporated per liver into total phospholipids plus glycerides was decreased to between 68% and 75% of that observed with fed animals. In homogenates from fed animals 41-44% of the labelled phospholipids plus glycerides was in glycerides; this value increased to between 63% and 75% with starved rats. Of the palmitate incorporated into total phospholipids, between 85% and 86% was found in phosphatidate, independent of the nutritional state of the animal. The ratio of palmitate incorporated into triglycerides/diglycerides rose from 0.7, obtained with fed rats, to 1.0 with starved animals. 4. These results indicate that starvation caused a decrease in the activity (per total liver) of acyl-CoA-glycerol phosphate acyltransferase(s) (EC 2.3.1.15) and an increase in the activity of acyl-CoA-diglyceride acyltransferase (EC 2.3.1.20). The largest change, however, seemed to be related to the increased activity of the phosphatidate phosphohydrolase in the particle-free supernatant. 5. The latter enzyme was assayed in the particle-free supernatant with membrane-bound phosphatidate as substrate. In starvation, the activity per total liver was increased to between 130% and 190% and the specific activity to between 180% and 320% of the values for fed rats.

Project description:1. Dose-dependent effects of adrenaline on PDHa activity were investigated with both incubated rat epidiymal fat-pads and isolated adipocytes. 2. Adrenaline (10nM- 5 micrometer) decreased PDHa activity in fat-pads incubated with 5 mM-[U-14C]glucose + insulin (20 munits/ml). Changes in [U-14C]glucose incorporation into fatty acids in these tissues correlated only loosely with changes in PDHa activity. There was a good inverse relationship between adrenaline-induced changes in PDHa activity and increases in lipolysis (glycerol release). 3. Adrenaline (10nM - 0.5 micrometer) decreased PDHa activity in fat-pads incubated with 5 mM-[U-14C]pyruvate + insulin (20 munits/ml), whereas 1 micrometer- and 5 micrometer-adrenaline slightly increased PDHa activity. All concentrations of adrenaline tested decreased [U-14C]pyruvate incorporation into fatty acids. Between 10nM- and 0.5 micrometer-adrenaline percentage decreases in PDHa activity paralleled decreases in faty acid synthesis. 4. Effects of adrenaline on PDHa activity and fatty acid synthesis in fat-pads incubated with 5mM-[U-14C]pyruvate + insulin (20 munits/ml) could not be mimicked by addition of albumin-bound palmitate. 5. The response of PDHa activity to adrenaline (0.1 nM - 1 micrometer) in isolated adipocytes differed with the carbohydrate substrate used in the incubations. With 5 mM-glucose + insulin (20 munits/ml), PDHa activity was significantly increased by 10 nM-adrenaline, but not by 1 micrometer-adrenaline, the response to adrenaline being biphasic. There was some correlation between PDHa activity and accumulation of non-esterified fatty acids. With 5 mM-glucose alone adrenaline (0.1 nM - 1 micrometer) had no effect on PDHa activity even though lipolysis was increased by adrenaline (0.1 micrometer - 1 micrometer). With 5mM-fructose in the presence and absence of insulin, lipolytic doses of adrenaline decreased PDHa activity. No tested concentrations of adrenaline increased PDHa with this substrate. 6. In the presence of 5 mM-fructose, palmitate was significantly more effective than adrenaline with respect to the maximum decrease in PDHa activity that could be elicited. 4. The relationship of changes in PDHa activity to changes in lipogenesis and the likelihood of adrenaline-induced changes in PDHa activity being secondary to changes in non-esterified fatty acid metabolism are discussed.

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Effect of adrenaline on 32 P incorporation into rat fat-cell phospholipids.

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