Glycerol incorporation into brain lipids in rat pups born to ethanol-intoxicated dams

The incorporation of labeled glycerol into glycerolipid of rat brain was influenced by the age of the animal (2-month-oldvs. newborn); indeed, 12 min after the administration, diglyceride was the most heavily labeled glycerolipid in the newborn brain, whereas the labeling of glycerophospholipid was highest in the adult. Various amounts of ethanol (0% to 36% of total energy intake) were administered to pregnant female rats, and the brains of their pups were examined for the ability to incorporate labeled glycerol into glycerolipid. The radioactivity incorporated into lipid diminished with increasing the amounts of alcohol consumed. The labeling pattern of lipid classes was also influenced; indeed, the radioactivity of diglyceride decreased markedly, whereas that of triglyceride and glycerophospholipid was affected to a lower degree. The distribution of radioactivity among different phospholipids also varied with age; on a percent basis, phosphatidylcholine was labeled less and phosphatidylinositol was labeled more in the newborn than in the adult. Ethanol influenced the pattern of glycerophospholipid labeling, increasing the radioactivity of phosphatidylserine and decreasing that of phosphatidylinositol.     

Valproic acid and bicuculline affect the formation of glycerolipid in rat brain

To ascertain the effects of bicuculline and of sodium valproate on the incorporation of glycerol into rat brain lipid, rats were divided into 5 groups: (a) controls; (b) treated with sodium valproate (400 mg/kg body wt); (c) treated with bicuculline (12.5 μmol/kg body wt); (d) treated with sodium valproate as in (b) + bicuculline as in (c); and (e) treated with bicuculline (25 μmol/kg body wt). Only rats of group (c) had seizures, which lasted until the end of the experiment. Each animal received 20 μCi of [2-³H]glycerol by intraventricular route and was sacrificed 12 min afterwards. Hippocampi and cerebella were taken and lipid extracted and separated by chromatography.

The type of treatment influenced very much the fate of injected, labeled glycerol. Indeed, total recovered radioactivity increased following either convulsions or the administration of valproate, whereas both treatments decreased the amount of radioactivity incorporated into lipid. These effects were more evident in cerebella than in hippocampi.

The distribution of radioactivity among lipid classes (diglyceride, triglyceride and total phospholipid) was also affected by seizures, which decreased the labeling ratio phospholipid/neutral lipid. The distribution of radioactivity among phospholipid classes was influenced by bicuculline (both at convulsant and non-convulsant doses) and these effects were sometimes antagonized by valproate. We conclude that some effects of bicuculline are exerted through the systemic modifications due to seizures and that other effects are probably connected to neuronal hyperfiring. The data reported in this paper are consistent with both mechanisms of action proposed for valproate, i.e. increased membrane permeability and modifications of GABAergic systems.     

[3H]Arachidonic acid metabolism in rat brain minces: Effects of nucleotide triphosphates,CDPcholine and CMP

Rat brain minces were used to investigate the effects of nucleotides on the metabolism of arachidonic acid in nerve tissue. Brain free fatty acids, neutral lipids and phospholipids, were radiolabeled in vivo following intracerebral injection of [³H]arachidonic acid. Minces were prepared from the radiolabeled cerebra and were incubated in a modified Krebs-Ringer buffer with and without various nucleotides. The incubation-induced accumulation of unesterified [³H]arachidonate was reduced in the presence of CDPcholine, ATP, CTP, GTP, and UTP. These nucleotides inhibited choline and inositol glycerophospholipid hydrolysis. They also reduced the amount of labeled diglycerides. However, CDPethanolamine had no effect on arachidonic acid metabolism in the mince preparation and CMP appeared to stimulate further hydrolysis of choline glycerophospholipids, resulting in increased accumulation of [³H]arachidonic acid and labeled diglycerides. We suggest that the production of unesterified [³H]arachidonate and labeled diglycerides is due to the involvement of more than one catabolic reaction, since the high energy nucleotides had similar effects on fatty acid accumulation, but different effects on phospholipid labeling.     

Analysis of phospholipid species in human blood using normal-phase liquid chromatography coupled with electrospray ionization ion-trap tandem mass spectrometry

A narrow-bore normal-phase high-performance liquid chromatography (HPLC) method was developed for separation of phospholipid classes in human blood. The separation was obtained using an HPLC diol column and a gradient of chloroform and methanol with 0.1% formic acid, titrated to pH 5.3 with ammonia and added 0.05% triethylamine. The HPLC system was coupled on-line with an electrospray ionisation ion-trap mass spectrometer. Chromatographic baseline separation was obtained between phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, lyso-phosphatidylcholine, phosphatidylinositol and phosphatidylserine, eluting in that order. The total run time was 30 min. Plasmalogen phosphatidylethanolamine and sphingomyelin, which both are substances with structural similarities to the glycerophospholipids, had similar retention time as phosphatidylethanolamine, but were well separated from the other glycerophospholipid classes. The species from each class were identified using MS² or MS³, which forms characteristic lyso-fragments. The combination of lyso-fragment mass, molecular ion and chromatographic retention time was used to identify each species, including 20 species of phosphatidylglycerol. The mass spectra obtained for the phospholipid classes are presented. Using this system 17 disaturated phospholipid species not earlier described to be present in blood were identified. The limit of detection varied between different phospholipid classes and was in the range 0.1–5 ng of injected substance.     

Labeling of lipids of retina subcellular fractions by [1-14C]eicosatetraenoate (20:4(n-6)) docosapentaenoate (22:5(n-3)) and docosahexaenoate (22:6(n-3))

(1-14C)-labeled (n-6) eicosatetraenoate, (n-3) docosapentaenoate and (n-3) docosahexaenoate (20:4, 22:5 and 22:6, respectively) are efficiently taken up and actively esterified into the lipids of bovine retina after 2 h incubation. Photoreceptor membranes, mitochondria, microsomes and postmicrosomal supernatants, which display significant differences in phospholipid and fatty acid compositions, are isolated after such incubations to study the labeling of lipids. The lipid classes preferentially labeled with the acids (1) largely differ among and within subcellular fractions, while (2) some common features in the treatment of the three polyenes are observed in each fraction. In all of them, the three acids are actively incorporated in phosphatidylcholine; ethanolamine glycerophospholipid, phosphatidylserine (PS) and phosphatidylinositol (PI) are highly labeled with 22:6, 22:5 and 20:4 respectively; within ethanolamine glycerophospholipid, the three label phosphatidylethanolamine in preference to plasmenylethanolamine. Most of the 14C esterified in mitochondria is in phospholipids. The endoplasmic reticulum produces in addition highly labeled triacylglycerols, also found in cytosol. High levels of 14C-labeled diacylglycerols are observed exclusively in photoreceptor membranes, where the specific radioactivity of PI is very high. The total amounts of 14C incorporated (1) are in general similar within a given fraction for the three polyenes, but (2) largely differ among fractions. The labeling of the highly unsaturated phospholipids of photoreceptor membranes is the lowest, while the postmicrosomal supernatant (whose lipids are relatively the poorest in polyenoic fatty acids) contains most of the labeled lipids isolated from retinas under these conditions. The results indicate that polyunsaturated species of retina phospholipids undergo an active synthesis and turnover, as well as an intense intracellular traffic among membranes.     

Composition and fatty acid content of rat ventral prostate phospholipids

The major phospholipids of rat ventral prostate have been separated and examined using thin-layer chromatography, gas chromatography and mass spectrometry. The main phospholipid classes were choline and ethanolamine glycerophospholipids, accounting for 77.9% of total lipid phosphorus. The prostate also contained small amounts of serine glycerophospholipids and sphingomyelin. The relative proportions of fatty acids in the different phospholipid classes were also determined. Arachidonic acid in prostatic phospholipids is contributed primarily by ethanolamine glycerophospholipids. This fraction contained 65-69 mol% plasmalogens, whereas choline and serine glycerophospholipid fractions contained less than 5 mol% plasmalogens. Ethanolamine, choline and serine plasmalogens contained mainly vinyl ethers of palmitic and stearic aldehydes. Ethanolamine plasmalogens also contained the vinyl ether of oleic aldehyde.     

Characterizations of Phospholipids from Paramecium tetraurelia Cells and Cilia*,†

Phospholipids from Paramecium tetraurelia strains 51s and d,95 cultures and isolated cilia were characterized. The following classes of phospholipids were identified in whole cell lipids: the 1-alkyl-2–acylglyceryl and the 1,2–diacylglyceryl forms of phosphonylethanolamine, phosphorylethanolamine. and phosphorylcholine; cardiolipin: ceramide aminoethylphosphonate and 5 other sphingolipids: phosphatidylserine; phosphatidylinositol; and lyso derivatives of the major glycerophospholipids. Cilia lipids were rich in ether lipids, phosphonolipids. and sphingolipids. Net lipid biosynthesis did not occur, as determined by the weight of lipids extracted from culture medium compared with the weight of lipids extracted from culture medium and ciliates after 7 days of growth. Total lipids/cell decreased with culture age. changes in the neutral lipid fraction accounting for the major decrease. Phospholipid class distributions changed with culture age—the glyceryl phosphorylethanolamine and choline content of cells decreased, while the glyceryl ohosphonylethanolamine content remained relatively constant: hence, the ratio of phosphonolipids to total phospholipids increased. All fatty acids observed in total lipids from cells and cilia were also present in the glycerophospholipids. Total lipids from cilia contained a greater percent of polyunsaturated fatty acid than those of whole cells. Whole cells and cilia glyceryl phosphonolipids contained up to 93% eicosatetraenoic plus eicosapentaenoic fatty acids. The enrichment of phosphonolipids in cilia accounted for most of the polyunsaturated fatty acid enrichment observed in cilia total lipids. The fatty acid composition of all major whole cell glycerophospholipid classes changed dramatically with culture age, while only small changes occurred in cilia glycerophospholipid fatty acids.     

INCORPORATION AND METABOLISM OF THE DIETARY TRANS-UNSATURATED FATTY ACID,ELAIDIC ACID,BY DEVELOPING RAT BRAIN1

Trans-unsaturated fatty acids, geometrical isomers of naturally occurring cis-acids, are dietary components and are incorporated into complex lipids of many tissues. There is little information about incorporation into brain and effects on CNS functions. In our experiments, mixtures of [l-¹⁴C]-elaidic acid and [9,10-³H]oleic were injected intragastrically into a total of 34 rats at 6, 12 and 16 days of age. Animals were killed 4, 8, 24, 48 and 96 h after administration and brain and liver lipids analyzed. With all ages examined, about 0.02–0.22% of the administered radioactivity from each fatty acid was found in brain lipids with incorporation increasing with time after administration. Phospholipids accounted for 60–85% of the total label from both fatty acids; of this phospholipid label, 40–50%, of the ¹⁴C was in unaltered irans-monoene. Up to 22% of the total ¹⁴C label recovered from brain was in cholesterol. By contrast to brain, labeling of liver lipid was much greater and was highest at 4 h after administration; there was proportionally less ¹⁴C or ³H label in palmitate and cholesterol compared to brain. Thus, intact trans-fatty acid, elaidic acid, was incorporated into developing brain, but at slower rates than into liver. These studies establish that the developing central nervous system does not exclude dietary trans-acids.     

Rapid β-oxidation of eicosapentaenoic acid in mouse brain: An in situ study

Analyses of brain phospholipid fatty acid profiles reveal a selective deficiency and enrichment in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), respectively. In order to account for this difference in brain fatty acid levels, we hypothesized that EPA is more rapidly β-oxidized upon its entry into the brain. Wild-type C57BL/6 mice were perfused with either ¹⁴C-EPA or ¹⁴C-DHA via in situ cerebral perfusion for 40 s, followed by a bicarbonate buffer to wash out the residual radiolabeled polyunsaturated fatty acid (PUFA) in the capillaries. ¹⁴C-PUFA-perfused brains were extracted for chemical analyses of neutral lipid and phospholipid fatty acids. Based on the radioactivity in aqueous, total lipid, neutral lipid and phospholipid fractions, volume of distribution (V_D, μl/g) was calculated. The V_D between ¹⁴C-EPA- and ¹⁴C-DHA-perfused samples was not statistically different for total lipid, neutral lipids or total phospholipids. However, the V_D of ¹⁴C-EPA in the aqueous fraction was 2.5 times higher than that of ¹⁴C-DHA (p=0.025), suggesting a more extensive β-oxidation than DHA. Furthermore, radiolabeled palmitoleic acid, a fatty acid that can be synthesized de novo, was detected in brain phospholipids from ¹⁴C-EPA but not from ¹⁴C-DHA-perfused mice suggesting that β-oxidation products of EPA were recycled into endogenous fatty acid biosynthetic pathways. These findings suggest that low levels of EPA in brain phospholipids compared to DHA may be the result of its rapid β-oxidation upon uptake by the brain.     

Schistosoma mansoni: synthesis and release of phospholipids, lysophospholipids, and neutral lipids by schistosomula

Lipids in the two surface membranes of Schistosoma mansoni may play an important role in the parasite's defense against host immunity. In particular, lysophosphatidylcholine lyses erythrocytes attached to the parasite and alters the lateral mobilities of their membrane proteins and lipids (Golan et al. 1986). Here, we have studied the incorporation of radiolabeled precursors into the major lipid classes of schistosomula as well as into lipids released by schistosomula into the medium. Radiolabeled polar head groups (choline and ethanolamine) and fatty acid precursors (palmitate and oleate) were linearly incorporated into parasite phospholipids. Fatty acids were differentially incorporated into the various phospholipid classes, principally into phosphatidylcholine and, to a lesser extent, into phosphatidylethanolamine, lysophosphatidylcholine, and phosphatidylserine. The major neutral lipid class labeled, triglycerides, had a decrease in specific activity with time after pulse labeling and the specific activity of the phospholipids increased with time. Thus, triglycerides may provide acyl chains for phospholipid synthesis. Choline was incorporated into phosphatidylcholine and lysophosphatidylcholine, and ethanolamine into phosphatidylethanolamine and lysophosphatidylethanolamine. No evidence was found for phospholipid methylation or demethylation in schistosomula. Labeled lipids were linearly and selectively released into the medium. Triglycerides were released at the highest rate with measurable quantities of phosphatidylcholine, lysophosphatidylcholine, and phosphatidylethanolamine also observed. Monopalmitoylphosphatidylcholine was the only lysophosphatidylcholine present in the medium as demonstrated by reverse-phase chromatography of released choline-labeled lysophosphatidylcholine. These studies demonstrate that schistosomula synthesize phospholipids and neutral lipids and release some of them into the culture medium. In particular, they release a single molecular species of a potent biologically active molecule, monopalmitoylphosphatidylcholine, that may play a role in the parasite's evasion of the immune response.     

Carbon tetrachloride target lipids in the rat liver microsomes. Effect of cystamine administration on their pattern of labeling by 14CCl4

¹⁴C from ¹⁴CCl₄ irreversibly binds to lipids from the smooth (SER) and rough (RER) endoplasmic reticulum. Most of the label is associated with the phospholipid fraction (> 95%). Prior cystamine administration decreased the extent of that binding but does not change its pattern of distribution. About the half of the label in phospholipids is in the phosphatidylcholine fraction; the other half is distributed similarly among lysophosphatidylcholine, sphingomyelin and phosphatidylethanolamine, while only a very minor fraction is associated with diphosphatidyl glycerol. No differences were found in the pattern of labeling of phospholipids in SER and RER.     

The glyceryl ethers of Paramecium phospholipids and phosphonolipids

Two glyceryl ethers, 1-O-hexadecyl glycerol and 1-O-cis-octadec-11-enyl glycerol, chimyl and paramecyl alcohol, respectively, were quantified in total phospholipids and five glycerophospholipid classes from cells and cilia of the ciliated protozoon, Paramecium tetraurelia. The ether content of 2-aminoethyl phosphonoglycerolipid was 85-90 mole %. Concentrations of ethers were greatest in the ethanolamine phosphonolipids greater than phosphatidylcholines greater than phosphatidylserines greater than phosphatidylethanolamines greater than phosphatidylinositols. The glyceryl ether concentrations in total cellular phospholipids increased with culture age in P. tetraurelia and P. multimicronucleatum cells. The glyceryl ether concentrations in the phospholipids of P. tetraurelia cilia remained constant from mid log to stationary phase of culture growth. Paramecium tetraurelia phospholipid glyceryl ether concentrations were made greater by supplementation of cultures with chimyl alcohol.     

Highly unsaturated phospholipid molecular species of rat erythrocyte membranes: selective incorporation of arachidonic acid into phosphoglycerides containing polyunsaturation in both acyl chains

This study describes for the first time the complete molecular species composition and turnover of [3H]arachidonic acid in various glycerophospholipid classes of rat erythrocytes, a model system that has been extensively used to investigate numerous membrane phenomena. Quantitative analysis of the individual molecular species of the choline, ethanolamine, serine, and inositol glycerophospholipid classes was possible by preparing their diradylglycerobenzoate derivatives that can be quantitated by on-line uv detection in conjunction with high-performance liquid chromatography; turnover of the molecular species containing arachidonate was evaluated in erythrocytes labeled with [3H]arachidonic acid. A unique observation was the significant amounts of 22:6-20:4, 20:4-20:4, and 18:2-20:4 species observed in the diacyl fractions of phosphatidylethanolamine and phosphatidylserine. Moreover, the analysis of the specific radioactivities of individual phospholipid species from erythrocytes incubated with [3H]arachidonic acid demonstrated a selective incorporation of arachidonic acid into the most highly unsaturated molecular species in all of the phospholipid classes examined. Although the 22:6-20:4, 20:4-20:4, and 18:2-20:4 species represented only 4.5% of the total mass of the diacyl phosphoglycerides, these species accounted for a major portion (37%) of the arachidonic acid incorporated into the phospholipids. These results demonstrate the existence of unique populations of phospholipid molecules in rat erythrocytes with a high degree of unsaturation that exhibit a very rapid metabolic turnover rate.     

The transport of cytidine into rat brain in vivo,and its conversion into cytidine metabolites

Double-labeled cytidine, with a³H/¹⁴C isotope ratio of 20.00, has been intraventricularly injected into the brain of young rats, and its fate followed up to 90 min from administration together with the time-course of labeling. The injected nucleoside enters the brain as an intact molecule and is immediately utilized without prior degradation. Cytidine is actively converted into uridine and CMP, the latter being then transformed by a stepwise mechanism into CDP and CTP, and finally into CDP-choline and CDP-ethanolamine. The results indicate that administered cytidine represents a compound likely to enter metabolic events, which lead to CDP-choline and CDP-ethanolamine synthesis, and presumably to phospholipid production.     

Effects of γ-aminobutyric acid on 33Pi incorporation into rat brain phospholipids in vivo

The effects of γ-aminobutyric acid (GABA), bicuculline and strychnine on the incorporation $\text{in vivo}$ of ³³Pi into phospholipids of rat brain were studied at 10 and 30 minutes after intracisternal injection of the radionuclide. GABA inhibited labeling of phospholipids in the three brain regions studied at both times. Bicuculline by itself had no significant effect on ³³Pi incorporation, but totally blocked the inhibitory effect of GABA in all three brain regions. Strychnine by itself inhibited phospholipid labeling in the brain stem and forebrain, had no significant effect on GABA inhibition of ³³Pi incorporation in the cerebellum and forebrain, and partially blocked the GABA effect in the brain stem. GABA inhibited ³³Pi incorporation into phosphatidic acid, phosphatidylinositol, phosphatidyl choline and phosphatidyl ethanolamine but had no effect on phosphatidyl serine. The data suggest that the inhibitory effects of GABA on CNS phospholipid labeling are mediated specifically through GABA receptor sites.     

Mass spectrometric analysis reveals changes in phospholipid, neutral sphingolipid and sulfatide molecular species in progressive epilepsy with mental retardation, EPMR, brain: a case study

Progressive epilepsy with mental retardation, EPMR, belongs to a group of inherited neurodegenerative disorders, the neuronal ceroid lipofuscinoses. The CLN8 gene that underlies EPMR encodes a novel transmembrane protein that localizes to the endoplasmic reticulum (ER) and ER-Golgi intermediate compartment. Recently, CLN8 was linked to a large eukaryotic protein family of TLC (TRAM, Lag1, CLN8) domain homologues with postulated functions in lipid synthesis, transport or sensing. By using liquid chromatography/mass spectrometry we analysed molecular species of major phosholipid and simple sphingolipid classes from cerebral samples of two EPMR patients representing a progressive and advanced state of the disease. The progressive state brain showed reduced levels of ceramide, galactosyl- and lactosylceramide and sulfatide as well as a decrease in long fatty acyl chain containing molecular species within these classes. Among glycerophospholipid classes, an increase in species containing polyunsaturated acyl chains was detected especially in phosphatidylserines and phosphatidylethanolamines. By contrast, saturated and monounsaturated species were overrepresented among phosphatidylserine, phosphatidylethanolamine and phosphatidylinositol classes in the advanced state sample. The observed changes in brain sphingo- and phospholipid molecular profiles may result in altered membrane stability, lipid peroxidation, vesicular trafficking or neurotransmission and thus may contribute to the progression of the molecular pathogenesis of EPMR.     

Daily rhythms of glycerophospholipid synthesis in fibroblast cultures involve differential enzyme contributions

Circadian clocks regulate the temporal organization of several biochemical processes, including lipid metabolism, and their disruption leads to severe metabolic disorders. Immortalized cell lines acting as circadian clocks display daily variations in [³²P]phospholipid labeling; however, the regulation of glycerophospholipid (GPL) synthesis by internal clocks remains unknown. Here we found that arrested NIH 3T3 cells synchronized with a 2 h-serum shock exhibited temporal oscillations in a) the labeling of total [³H] GPLs, with lowest levels around 28 and 56 h, and b) the activity of GPL-synthesizing and GPL-remodeling enzymes, such as phosphatidate phosphohydrolase 1 (PAP-1) and lysophospholipid acyltransferases (LPLAT), respectively, with antiphase profiles. In addition, we investigated the temporal regulation of phosphatidylcholine (PC) biosynthesis. PC is mainly synthesized through the Kennedy pathway with choline kinase (ChoK) and CTP:phosphocholine cytidylyltranferase (CCT) as key regulatory enzymes. We observed that the PC labeling exhibited daily changes, with the lowest levels every ∼28 h, that were accompanied by brief increases in CCT activity and the oscillation in ChoK mRNA expression and activity. Results demonstrate that the metabolisms of GPLs and particularly of PC in synchronized fibroblasts are subject to a complex temporal control involving concerted changes in the expression and/or activities of specific synthesizing enzymes.     

Lipid Compositions of Different Regions of the Human Brain During Aging

The neutral and phospholipid compositions of various regions of the human brain were analyzed using autopsy material covering the life period between 33 and 92 years of age. The protein content was also measured and, on a weight basis, this content is unchanged in the cerebellum, pons, and medulla oblongata, whereas in the 90-year-old group it decreases in the hippocampus, gray matter, and nucleus caudatus. In white matter, the protein content decreases continuously with age. The phospholipid composition is characteristic of the region investigated, but remains unchanged during aging. The total phospholipid content exhibits only a 5-10% decrease in the oldest age group. The content of dolichol and its polyisoprenoid pattern are also characteristic of the region analyzed. Between 33 and 92 years of age, the amount of dolichol in all portions of the brain increases three- to fourfold, but the isoprenoid pattern remains constant. The level of dolichyl-P varies between different regions, but only a moderate increase is seen with age. Ubiquinone content is highest in the nucleus caudatus, gray matter, and hippocampus, and in all areas this content is decreased to a great extent in the oldest age groups. All regions of the human brain are rich in cholesterol, but alterations in the amount of this lipid are highly variable during aging, ranging from no change to a 40% decrease.     

The Glyceryl Ethers of Paramecium Phospholipids and Phosphonolipids1

Two glyceryl ethers, 1-O-hexadecyl glycerol and 1-O-cis-octadec-11-enyl glycerol, chimyl and paramecyl alcohol respectively, were quantified in total phospholipids and five glycerophospholipid classes from cells and cilia of the ciliated protozoon Parammecium tetraurelia. The ether content of 2-aminoethyl phosphonoglycerolipid was 85–90 mole %. Concentrations of ethers were greatest in the ethanolamine phosphonolipids > phosphatidylcholines > phosphatidylserines > phosphatidylethanolamines > phosphatidylinositols. The glyceryl ether concentrations in total cellular phospholipids increased with culture age in P. tetraurelia and P. multimteronucleatum cells. The glyceryl ether concentrations in the phospholipids of P. tetraurelia cilia remained constant from mid log to stationary phase of culture growth. Paramecium tetraurelia phospholipid glyceryl ether concentrations were made greater by supplementation of cultures with chimyl alcohol.     

Arachidonic acid channelling in the phospholipid fractions and subcellular compartments of cultured myocardial cells.

Arachidonic acid (AA) channeling in cultured heart cells was studied following pulse labelling for 1 h. AA was shown to be esterified immediately and equally distributed between the neutral lipids and phospholipids. A rapid constant flow to various phospholipid classes occurred thereafter, while the AA oxidation was only between 12%. The subcellular distribution of AA was studied by nitrogen cavitation followed by fractionation on 6.7% percoll in sucrose-EDTA. After 1 h pulse labeling and 2 h post-pulse incubation, most of the radioactivity was found in the sarcolemmal fraction with a much smaller amount in the mitochondrial fraction.