Phospholipid composition and [14C]glycerol incorporation into glycerolipids of toad retina and brain

The phospholipid composition as well as the in vivo [14C]glycerol uptake in lipids was found to be similar in the toad brain and retina. The choroid lipid labeling was markedly different. An in vitro time-course study of [14C]glycerol incorporation in toad retina lipids disclosed that under the conditions of these experiments: (1) retina is able to rapidly synthesize phosphatidic acid from the radioactive precursor; (2) the sequence phosphatidic acid-diacylglycerol-triacylglycerol operates; (3) a high rate of phosphatidylinositol de novo biosynthesis takes place; (4) phosphoglycerides of choline and of ethanolamine are also heavily labeled after a lag period; (5) in vivo labeling profiles resembled those obtained in vitro mainly regarding phosphatidylinositol biosynthesis; and (6) the presence of glycerol kinase in the CNS is suggested.     

1-14C]Arachidonic acid incorporation into glycerolipids and prostaglandin synthesis in peritoneal macrophages: effect of chloramphenicol

Peritoneal macrophages from normal mice were labelled with [1-14C]arachidonic acid after 2 h culture. The uptake of arachidonic acid into cellular lipids was rapid, time-dependent and can be represented within the limit of the studied times by a parabolic regression. Indomethacin decreased the kinetics of uptake; this inhibition is dose-dependent. Chloramphenicol had no effect on macrophage [1-14C]arachidonic acid uptake. After 3 h, the radioactivity was recovered in phosphatidylcholine (38.6%), phosphatidylserine-phosphatidylinositol (8.5%), phosphatidylethanolamine (22.1%), diacylglycerol (2.9%), triacyglycerol (2%) and cholesteryl ester (11.8%). Chloramphenicol and indomethacin inhibited the labelling of phospholipids and stimulated the labelling of neutral lipids and cholesteryl ester. Studies on arachidonic acid release from glycerolipids of prelabelled 2-h cultured macrophages showed that phosphatidylcholine and phosphatidylserine-phosphatidylinositol are the major source of arachidonic acid in prostaglandin synthesis in macrophages stimulated or not by zymosan. Chloramphenicol inhibited release of fatty acid from phosphatidylcholine and phosphatidylserine-phosphatidylinositol; indomethacin had no effect. Both drugs inhibited prostaglandin synthesis in stimulated or non-stimulated macrophages. In the culture medium, indomethacin increased the release of free arachidonic acid by stimulated macrophages. Possible explanations for the mechanisms underlying these effects are presented. It is concluded that indomethacin and chloramphenicol exert profound effects on the metabolism of phospholipids and its zymosan activation. Chloramphenicol appears to impair prostaglandin synthesis through several mechanisms and especially through phospholipase inhibition.     

Effects of postdecapitation ischemia on the metabolism of [14C]arachidonic acid and [14C]palmitic acid in the mouse brain

The effect of postdecapitation ischemia on the labeling of the free fatty acid pool and their incorporation in lipids was examined during the first 10 min after decapitation in mouse brain that had been injected intracerebrally with either [1-14C]arachidonic acid or [1-14C]palmitic acid. One min after decapitation, animals injected with labeled arachidonic acid exhibited a greatly reduced incorporation of label in brain phospholipids, diglycerides, and triglycerides. When radioactive palmitic acid was used, brain lipids exhibited considerably less inhibition of label. However, a similar degree of inhibition was observed 10 min after decapitation with both fatty acids. At this time, free arachidonic acid had decreased 84% as compared to the 24% decrease observed in the controls, and about 77% of the free palmitic acid remained in the free fatty acid fraction as compared with 30% in the controls. This decreased labeling may reflect ATP shortage that affects the fatty acid activation-reacylation reactions or the enzymes involved. Alternatively, the enhanced endogenous free arachidonic acid may compete with the radiolabeled arachidonic acid resulting in an inhibition of lipid labeling. Inhibition of label may have been greater in radiolabeled arachidonic acid than palmitic because of the larger accumulation of the former endogenous fatty acid during early ischemia.     

The effect of methyl palmoxirate on incorporation of [U-14C]palmitate into rat brain

We examined the dose response, time course and reversibility of the effect of methyl 2-tetradecylglycidate (McN-3716, methyl palmoxirate or MEP), an inhibitor of -oxidation of fatty acids, on incorporation of radiolabeled palmitic acid ([U-¹⁴C]PA) from plasma into brain lipids of awake rats. MEP (0.1, 1 and 10 mg/kg) or vehicle was administered intravenously from 10 min to 72 hr prior to infusion of [U-¹⁴C]PA. Two hr pretreatment with MEP (0.1 to 10 mg/kg) increased brain organic radioactivity 1.2 to 1.8 fold and decreased brain aqueous radioactivity by 1.2 to 3.0 fold when compared to control values. At 10 mg/kg, MEP significantly increased brain organic fraction from 40% in controls to 85%, 30 min to 6 hr pretreatment, and resulted in a redistribution of the radiolabeled fatty acid toward triacylglycerol. MEP changed the lipid/aqueous brain ratio of incorporated [U-¹⁴C]PA from 0.67 to 5.7. The incorporation rate coefficient, k^*, was significantly increased by MEP (10 mg/kg) at 2 hr (31%), 4 hr (59%) and 6 hr (34%). All effects were reversed by 72 hr, consistent with a half-life of 2 days for carnitine palmitoyl transferase I. These results indicate that intravenous MEP may be used with [1-¹¹C]palmitic acid for studying brain lipid metabolism in vivo by positron emission tomography, as it significantly reduces the large unincorporated aqueous fraction that would result in high background radioactivity.     

Diffusion of intracerebrally injected [1-14C]arachidonic acid and [2-3H]Glycerol in the mouse brain

[2-³H]Glycerol and [1-¹⁴C]arachidonic acid were injected into the region of the frontal horn of the left ventricle of mice and were distributed rapidly throughout the brain. After 10 sec, most of the radioactive fatty acid was found in the hemisphere near the injection site; after 10 min, it was recovered in similar proportions in the cerebellum and brain stem. [2-³H]Glycerol showed a heterogeneous distribution, with most of the label remaining in the left hemisphere even after 10 min. On a fresh weight basis, cerebrum, cerebellum, and brain stem were found to contain similar amounts of labeled glycerol. However, the amount of [1-¹⁴C]arachidonate in cerebrum was only 50% of that recovered from cerebellum or brain stem. Brain ischemia or a single electroconvulsive shock reduced the spread of the label, producing an accumulation of radioactivity in the injected hemisphere, except for an increase in [2-³H]glycerol in the brain stem during ischemia. Despite the significant decrease in available precursor in the cerebellum and brain stem after electroshock, the amount of label incorporated into lipids was not altered in these areas and only slightly diminished in the cerebrum.     

The incorporation of [14C]palmitic acid into the lipids of mouse brain in vivo

Abstract— The half-life of free [¹⁴C]palmitic acid injected intracerebrally into C57BL/10J mice was less than 5 min. The rapid disappearance of radioactivity as palmitic acid was accompanied by increases in the radioactivity of the phosphatidic acids and the diacyl-glycerols. The peak specific radioactivity of the diacylglycerols occurred at about 6-8 min after injection. The triacylglycerols, phosphatidyl ethanolamines and phosphatidyl cholines exhibited increasing amounts of radioactivity during the first 40 min. At 160 min after injection, the distribution of radioactivity was similar to the pattern observed at 12 h. The biosynthetic pathway through the phosphatidic acids and the diacylglycerols to triacylglycerols, phosphatidyl ethanolamines and phosphatidyl cholines is apparently the major pathway in vivo for the esterification of free fatty acids in the brain.     

In vitro age-dependent incorporation of [1-14C]acetate into lipid subclasses in rat ventral prostate

The [1-14C]acetate incorporation into different lipid subclasses by the rat prostate gland was lineal between 20 and 80 mg of wet tissue. The in vitro [1-14C]acetate incorporation into lipid subclasses was a development-dependent process. The highest values of [1-14C]acetate incorporation into triacylglycerols, free cholesterol and esterified cholesterol were observed at puberty, but radioactivity incorporation into phospholipids was similar in both prepuberty and puberty, then decreasing in maturity. The relationship between triacylglycerols, free cholesterol and esterified cholesterol with respect to total lipids was about 12, 10 and 3.5%, respectively, values being maintained during the animal development. The in vitro [1-14C]acetate incorporation into lipid subclasses in castrated rats decreased considerably as compared with normal rats.     

The kinetics of incorporation in vivo of [14C]acetate and [14C]carbon dioxide into the fatty acids of glycerolipids in developing leaves

1. The patterns of incorporation of (14)C into glycerolipid fatty acids of developing maize leaf lamina from supplied [1-(14)C]acetate and from (14)CO(2) during steady-state photosynthesis were similar. Oleate of phosphatidylcholine and palmitate of phosphatidylglycerol attained linear rates of labelling more rapidly than did other fatty acids, particularly the linoleate and linolenate of monogalactosyl diacylglycerol. 2. After the transfer of lamina from labelled to unlabelled acetate, there was a decrease in labelled oleate and linoleate of phosphatidylcholine and a concomitant increase in the amount of radioactivity in the linoleate and linolenate of monogalactosyl diacylglycerol. 3. The rapidly labelled phospholipids, phosphatidylcholine and phosphatidylglycerol, were shown by differential and sucrose-density-gradient centrifugation to be associated with different organelles, the former being mainly in a low-density membrane fraction, probably microsomal, and the latter mainly in chloroplasts. 4. During a 48h period after supplying spinach leaves with [(14)C]acetate, radioactivity was lost from the oleate of phosphatidylcholine present in fractions sedimented at 12000g and 105000g, and accumulated in the linolenate of monogalactosyl diacylglycerol of the chloroplast. 5. It is proposed that the phosphatidylcholine of some non-plastid membranes is intimately involved in the process of oleate desaturation and that this lipid serves as a donor of unsaturated C(18) fatty acids to other lipids, principally monogalactosyl diacylglycerol, of the chloroplasts.     

[14C]Formate and [8-14C]Adenine incorporation into purine nucleotides of ischaemic rat liver after re-establishment of blood flow

• 1.1. After restoration of blood flow the incorporation rate of [¹⁴C]formate and [8-¹⁴C]adenine into purine nucleotides of ischaemic rat liver was investigated.
• 2.2. The incorporation rate of [¹⁴C]formate into adenine nucleotides was nearly 5-fold greater in post-ischaemic livers than in controls at 120 min point following blood restoration.
• 3.3. No difference in the rate of [8-¹⁴C]adenine incorporation into adenine nucleotides of post-ischaemic and control rat liver lobes was found.
• 4.4. After re-establishment of blood circulation a predominant contribution of the de novo biosynthetic pathway in the recovery of purine nucleotides of the ischaemic rat liver is emphasized.

     

Measles-virus-persistent infection in BGM cells. Modification of the incorporation of [3H]arachidonic acid and [14C]stearic acid into lipids.

In BGM cells chronically infected with measles virus, although the composition of the phospholipids is unaltered, the fatty acid composition is modified. Uninfected, lytic and persistently infected cells were labelled with [3H]arachidonic acid and [14C]stearic acid and their metabolic fate analysed. No difference in the total incorporation was observed in the different systems. However, the [14C]stearic acid and [3H]arachidonic acid were incorporated up to 2-fold and 13-fold respectively greater into the neutral lipid of persistently infected compared with that of uninfected cells. Both radioactive fatty acids were specifically accumulated in the triacylglycerol and non-esterified fatty acids fractions. Lytically infected cells were similar to uninfected cells. Although there was no significant difference in the incorporation of radioactivity into the total phospholipid in either system, there was a large decrease in [3H]arachidonic acid incorporated into phosphatidylethanolamine and to a lesser extent phosphatidylcholine and phosphatidylinositol in persistently infected cells. [14C]Stearic acid incorporation was also reduced in phosphatidylcholine and phosphatidylethanolamine fractions of persistently infected cells.     

Effects of dietary nonspecific nitrogen on [14C]glutamate and [14C]proline incorporation into bone proteins in chicks

The incorporation of [14C]glutamic acid into EDTA-soluble and -insoluble calvaria protein in vitro and [14C]proline into EDTA-insoluble femur protein in vivo was determined in chicks fed inadequate and adequate levels of nonspecific nitrogen (glutamic acid). In each instance, the amount of amino acid incorporated into bone protein was reduced by the low level of nonspecific nitrogen. It was concluded that the high incidence of leg abnormalities observed in chicks fed purified diets containing adequate levels of indispensable amino acids but lacking in total nitrogen might be associated with an inability to form bone matrix protein.     

Incorporation of [14C] arachidonic acid into ovine conceptus and endometrial lipids.

The purpose of this study was to quantitate conceptus and endometrial incorporation of [14C]arachidonic acid (AA) into individual neutral and polar lipids. Endometrium and conceptuses from pregnant ewes and endometrium from nonbred ewes were collected 14 and 16 d after onset of estrus (d 0). Tissues were incubated for 8 h at 37 degrees C in medium containing 1 microCi of [14C]AA. Thin-layer chromatographic procedures were used to separate 12 lipids. Radioactivity was measured in each lipid, and the amount (ng) of [14C]AA incorporated into each lipid was calculated. Conceptuses and endometrium incorporated more [14C]AA into triacylglycerols than into any other lipid. Day and tissue type affected differentially (i.e., day X tissue interaction) the incorporation of [14C]AA into several lipids; d-14 conceptuses incorporated [14C]AA more actively than did any other day-tissue combination. Results indicate that triacylglycerols may be an important reservoir for conceptus and endometrial AA. The remarkable ability of d-14 conceptuses to incorporate [14C]AA into various lipids may be important for their accelerated elongation and active prostaglandin synthetic system.