Triglyceride Uptake in Muscles in Rats

Exogenous lipid is assimilated with different priorities in adipose tissue regions and varies in the fasting and fed conditions. The quantitative role of uptake of lipid in muscle has not been evaluated. In order to examine the uptake in other than adipose tissues, U¹⁴C-oleic acid in sesame oil was administered orally to conscious rats, and lipid label measured after different times in serum, heart, liver, mesenteric, retroperitoneal, inguinal and epididymal fat pads, as well as in red and white parts of gastrocnemius, extensor digitorum longus and soleus muscles. Lipid uptake in total adipose tissue was calculated from dissected adipose tissues plus lipids extracted from the eviscerated, skinned carcass. Lipid uptake in total muscle tissue was estimated from label in dissected muscles plus that in the carcass, assuming similar intracellular lipid contents and radioactivity as that averaged from dissected muscles. Lipid uptake in the liver was calculated from directly extracted lipid. Four hours after lipid administration to fed rats lipid radioactivity in heart and serum was minimal and had essentially disappeared at 8 hours. Liver label declined rapidly from peak values at or before 4 hours. Adipose tissue radioactivity increased gradually up to 16 hours and then decreased. Label in muscles was highest at 4 hours in the red gastrocnemius, and then decreased, while the other muscles showed a constant radioactivity over the observation period (24 hours). Radioactivity expressed per unit muscle mass seemed to be proportional to the oxidative capacity of muscles. In comparisons between fed and fasted rats at 16 hours, when adipose tissue label peaked, liver, individual muscles and carcass did not show any significant differences while adipose tissue label was fivefold higher in fed than fasted rats. The distribution of total measured lipid radioactivity between total adipose tissue, total muscle tissue and liver in fed rats at this time-point was 76. 8, 14. 4 and 8. 8% respectively, and in the fasted state 26. 4, 51. 6 and 22. 0%. These estimations suggest that lipid uptake in the fed state is dominated by adipose tissue, while in the fasted state the lipid uptake is higher in muscles than adipose tissues. It was concluded that uptake of absorbed, exogenous triglyceride in muscle is of significance, particularly in the fasted state. This lipid has a half life of several days. It is suggested that this lipid is oxidized in situ, contributing with a hidden fraction to lipid energy needs, or partially transferred to adipose tissue. Lipid uptake in muscle probably constitutes a significant fraction of assimilated exogenous lipid, particularly in the fasting state.     

Direct evidence of the role of propionate in choline synthesis in rats

Wistar rats were injected with 2-14C-propionate in a dose of 30 mu Ci/100 g bw, 2 h after food intake. Two hours after isotope injection the rats were decapitated to determine specific radioactivity (SR) in liver and brain lipids, in liver phosphatidylcholine (PC) and its structural components. The label was incorporated in liver lipids in a far greater amount. In liver PC, SR appeared the highest in glycerin and less higher in the fraction of higher fatty acids. The least amount of the label from 2-14C-propionate was incorporated in choline. The fact of the label incorporation in choline was recorded for the first time.     

Acetoin metabolism in animal tissues

The acetoin-synthesizing activity has been studied in the skeletal muscles, brain, liver and spleen homogenates (numbered as the activity decreases). The acetoin-synthesizing activity drastically increases in case of the acetaldehyde excess and alcohol intoxication. The acetaldehyde concentrations of above 1.10(-3) M inhibit the liver pyruvate dehydrogenase activity and increase the non-oxidative transformation of pyruvate. Acetoin is rapidly metabolized in the organism eliminating from blood 10 minutes after its injection. Acetoin is an effective precursor in the biosynthesis of lipids.     

Adaptation of adenosylmethionine metabolism and methionine recycling to variations in dietary methionine in the rat

Weanling rats were fed a casein-based diet supplemented to give dietary methionine (Met) concentrations of 0.41, 0.61, and 1.50%. After 2 weeks of feeding, the rats received intraperitoneally 800 nCi of 2-14C-labeled and/or methyl-3H-labeled L-Met. The animals were killed 20 min, 1 hr, or 2 hr after the isotope injection and the specific radioactivity of adenosylmethionine (AdoMet) as well as the total acid-soluble radioactivity was analyzed in the liver and skeletal muscle. Met concentrations of the liver and skeletal muscle were increased 20-fold by the diet containing 1.50% of Met. In the liver, but not in skeletal muscle, accumulation of AdoMet closely followed changes in Met concentration. Within 2 hr after intraperitoneal injection, the rate of disappearance of 3H label from the acid-soluble fraction was slow in both tissues; increasing in the liver and decreasing in skeletal muscle with increasing dietary Met concentration. At the same time, disappearance of 14C label was slow in both tissues in the rats fed the toxic Met diet, and also in the liver of the rats fed the Met-deficient diet. Decline of the specific radioactivity of the AdoMet pool with respect to 3H label was similar to that of 14C label in the skeletal muscle at all dietary Met concentrations. In the liver, the rate of disappearance of 14C label from the AdoMet pool was markedly increased and that of the 3H label slightly decreased with increasing dietary Met supply. Met deprivation resulted in rapid disappearance of 3H label from the hepatic AdoMet pool, whereas the disappearance of the 14C label was very slow. The results indicate that hepatic Met recycling is very effective with deficient or adequate dietary Met concentrations. In skeletal muscle, the capacity to catabolize extra Met is very limited and continuous flow of Met to liver takes place. Unlike in the liver, in skeletal muscle the transsulfuration route is not adaptable to changes in Met supply and plays a minor role in Met catabolism. The approach used to determine the efficacy and adaptation of methionine salvage pathways by following simultaneously the decline of the specific radioactivities of the methyl group and the methionyl carbon chain of AdoMet following intraperitoneal injection of double-labeled Met has several advantages over that used in literature reports. It offers a reliable means of observing these metabolic pathways in whole animals without disruption of metabolite fluxes.     

Preferential In Vivo Incorporation of [3H]Arachidonic Acid from Blood into Rat Brain Synaptosomal Fractions Before and After Cholinergic Stimulation

Abstract: Awake adult male rats were infused intravenously with [³H]arachidonic acid for 5 min, with or without prior administration of an M1 cholinergic agonist, arecoline (15 mg/kg i.p.). Methylatropine was also administered (4 mg/kg s.c.) to control and arecoline-treated animals. At 15 min postinfusion, the animals were killed, brains were removed and frozen, and subcellular fractions were obtained from homogenates of whole brain. Total radioactivity and radioactivity in various lipid classes were determined for each fraction following normalization for exposure by use of a unidirectional incorporation coefficient, k ^⋆_brain. In control animals, incorporation was greatest in synaptosomal and microsomal fractions, accounting for 50 and 30% of total label incorporated into membrane lipids, respectively. Arecoline increased incorporation in these two fractions by up to 400% but did not increase incorporation into the myelin, mitochondrial, or cytosolic fractions. Of the incorporated radioactivity, 50–80% was in phospholipid in microsomal and synaptosomal fractions, indicating that phospholipid is the major lipid affected by cholinergic stimulation. These results demonstrate that plasma [³H]arachidonic acid is preferentially incorporated into phospholipids of synaptosomal and microsomal fractions of rat brain. Cholinergic stimulation increases incorporation into these fractions, likely by activation of phospholipase A₂ and/or C in association with acyltransferase activity. Thus, intravenously infused radiolabeled arachidonic acid can be used to examine synapse-mediated changes in brain phospholipid metabolism in vivo.     

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.     

Ether lipid metabolism. Incorporation of O-hexadecyl ethanediol into rat brain lipids.

1-O-[1'-14C]Hexadecyl ethanediol was administered intracerebrally to myelinating rat brain, and incorporation of radioactivity into brain lipids was followed over a 48-h period: (1) O-Hexadecyl ethanediol was metabolized primarily through oxidative ether bond cleavage, and much of the label was recovered in phospholipid acyl groups. (2) Substantial amounts of radioactivity were also found in choline and ethanolamine phospholipids having an O-hexadecyloxyethyl glycerol backbone. This means that alkyl ethanediol was used in glycerol ether biosynthesis as are long-chain primary alcohols. (3) Acidic hydrolysis of the ethanolamine glycerophosphatide fraction yielded also labeled hexadecanol which may indicate desaturation of 1-O-hexadecyloxyethyl 2-acyl glycerophosphoryl ethanolamine to the plasmalogen analogue. (4) Small amounts of the substrate were oxidized to O-hexadecyl glycolic acid and incorporated into the phospholipids. The substrate did not serve as precursor of O-hexadecyl ethanediol phosphorylcholine or phosphorylethanolamine in the brain.     

The dynamics of [3H]-cholesterol incorporation into the liver and aortic tissue of guinea pigs on long-term ethanol administration]

The effect of ethanol administration to guinea pigs (4 g/kg, per os) on the dynamics of [3H]-cholesterol incorporation into the liver and aorta tissues was studied for 3 months. It has been discovered that specific radioactivity of the control animals linearly increased during 24 hours in the blood serum. Ethanol reduced it as compared with the control only 0.5 h after a label has been introduced. Cholesterol renovation in the liver remained unchanged under the prolonged effect of ethanol. In the aorta the ethanol effect was characterized by a decrease of [3H]-cholesterol specific radioactivity 0.5 h after its administration. However, in this case the ratio of aorta/blood serum radioactivity increased. A day after the labelled cholesterol administration to alcoholized animals the radioactivity calculated per 1 mg of cholesterol and per unit of tissue weight and referred to the blood serum radioactivity was lower as compared to the control level.     

The effect of orally administered secondary autoxidation products of linoleic acid on the activity of detoxifying enzymes in the rat liver

Radioactive secondary autoxidation products of linoleic acid were administered orally to rats and the incorporation of radioactive substances into lipids was investigated in the liver. The radioactive substances were significantly incorporated into hepatic mitochondrial and microsomal lipids 12 h after the administration. 80% of the radioactivity in mitochondria was detected in neutral lipids. The radioactivity in microsomal neutral lipids significantly decreased and the activity in phospholipids increased 12 h after the administration. On the other hand, contents of lipid peroxide and thiobarbituric acid reactive substances in liver were significantly increased by 40% at 15 h after the administration of the secondary autoxidation products. Activity of marker enzymes used for an indication of the hepatic injury was also elevated. Glutathione peroxidase activity increased 3-fold and catalase activity increased 1.5-fold. Activity of mitochondrial NAD-dependent aldehyde dehydrogenase, however, was decreased by 50%. It seems likely that the secondary autoxidation products orally administered are detoxified in the hepatic mitochondria, metabolized to neutral lipids, and further metabolized to phospholipids in microsomes, while as the incorporated secondary autoxidation products induces hepatic injury by lipid peroxidation.     

Kinetics of four 11C-labelled enkephalin peptides in the brain, pituitary and plasma of rhesus monkeys

The kinetics of four 11C-labelled enkephalin peptides: Tyr-Gly-Gly-Phe-Met (Met-enkephalin), Tyr-D-Met-Gly-Phe-Pro-NH2 [D-Met2,Pro5)-enkephalinamide), Tyr-D-Ala-Gly-Phe-Met-NH2 (DALA) and Tyr-D-Ala-D-Ala-Phe-Met-NH2 (TAAFM) all labelled at the methyl group of methionine was studied in the Rhesus monkey. After intravenous administration, the regional kinetics in the head, lungs, liver and kidneys were followed by means of positron emission tomography (PET). The total radioactivity in blood and urine was measured and the composition of 11C-labelled peptide fragments in plasma in vivo and in vitro was analysed by liquid chromatography. With PET, an increased radioactivity was observed in the brain and pituitary over the 60-90 min investigation period after i.v. injection of the peptides. The highest radioactivities were noted for Met-enkephalin, followed by DALA and D-Met2, Pro5-enkephalinamide, while very low radioactivities were found for TAAFM. The uptake of Met-enkephalin- and DALA-derived radioactivity was of the same order as has previously been shown for morphine in the brain and considerably higher than that of D-Met2,Pro5-enkephalinamide and TAAFM, respectively. A large fraction of the brain radioactivity derived from Met-enkephalin and DALA probably emanated from [11C]methionine as indicated by plasma and urine analysis. Met-Enkephalin was rapidly eliminated from plasma in vitro with an half-life of less than two minutes, whereas DALA was stable suggesting clearance by other tissues than plasma. In conclusion, both Met-enkephalin and DALA, were rapidly hydrolyzed in vivo to [11C]methionine. [11C]Methionine was probably taken up in the brain, as the radioactivity increased with time in different brain regions as measured with PET.D-Met2,Pro5-Enkephalinamide and TAAFM were virtually stable in vivo and at least part of the radioactivity observed in the brain may have represented the intact peptide.     

Incorporation of Glycerol and Ethanolamine into Glycerophospholipid in Rat Brain Areas During Bicuculline-Induced Convulsive Seizures

The effect of bicuculline-induced convulsive seizures on lipid metabolism has been studied in four brain areas (cerebellum, cerebral cortex, hippocampus, and brainstem) using [2-3H]glycerol and [1,2-14C]ethanolamine as radioactive lipid precursors administered simultaneously with bicuculline. Twelve minutes after the administration, the uptake of radioactivity depended both on brain area and treatment, being generally higher in convulsing rats. The uptake of glycerol was influenced to a larger extent than that of ethanolamine and increased during convulsions, but its incorporation into lipids did not. In contrast, the amount of ethanolamine incorporated into lipids increased during bicuculline-induced seizures. The difference in behavior of glycerol and of ethanolamine is also indicated by the decrease of the 3H/14C ratio of phosphatidyl-ethanolamine in various brain areas during convulsions. It is, therefore, evident that the metabolism of the two precursors is affected differently by seizures.     

Spinal Morphine Administration Reduces the Fatty Acid Contents in Spinal Cord and Brain by Increasing Oxidative Stress

It is well known that oxidative stress damages bimolecules such as DNA and lipids. No study is available on the morphine-induced oxidative damage and fatty acids changes in brain and spinal tissues. The aim of this work was to determine the effects of morphine on the concentrations and compositions of fatty acid in spinal cord segments and brain tissues in rabbits as well as lipid peroxidation (LP) and glutathione (GSH) levels in cortex brain. Twelve New Zealand albino rabbits were used and they were randomly assigned to two groups of 6 rabbits each. First group used as control although morphine administrated to rats in second group. Cortex brain and (cervical, thoracic, lumbar) samples were taken. The fatty acids between n:18.0 and 21.0 were present in spinal cord sections and n:10 fatty acids in control animals were present in the brain tissues. Compared to n:20.0–24.0 fatty acids in spinal cord sections and 8.0 fatty acids in the brain tissues of drug administered animals. The concentration and composition of the fatty acid methyl esters in spinal cord and brain tissues was decreased by morphine treatments. LP levels in the cortex brain were increased although GSH levels were decreased by the morphine administration. In conclusion, unsaturated fatty acids contents in brain and spinal cord sections and GSH were reduced by administrating spinal morphine although oxidative stress as LP increased. The inhibition oxidative damage may be a useful strategy for the development of a new protection for morphine administration as well as opiate abuse.     

Generation of Arachidonic Acid and Diacylglycerol Second Messengers from Polyphosphoinositides in Ischemic Fetal Brain

Intracerebral administration of [3H]arachidonic acid ([3H]ArA) into 19-20-day-old rat embryos, resulted in a rapid incorporation of label into brain lipids. One hour after injection, 55.6 +/- 8.2, 18.0 +/- 3.4, and 13.7 +/- 1.3% of the total radioactivity was associated with phosphatidylcholine, phosphatidylinositol, and phosphatidylethanolamine, respectively. Approximately 10% of radioactivity was found acylated in neutral lipids of which free ArA comprised only 1.5 +/- 0.2% of the total radioactivity. Complete restriction of the maternal-fetal circulation for < or = 40 min did not affect the rate of [3H]ArA incorporation (t1/2 = 2 min) into fetal brain lipids, suggesting an effective acylation mechanism that proceeds irrespective of the impaired blood flow. After a short restriction period (5 min), the radioactivity in diacylglycerol was elevated by 50%. After a longer restriction period (20 min), the radioactivity in the free fatty acid and diacylglycerol fractions increased to values of 130 and 87%, respectively. Polyphosphoinositides prelabeled with either [3H]ArA or 32P were rapidly degraded after 5 min of ischemia. After 20 min, the decrease in phosphatidylinositol-4-phosphate and phosphatidylinositol-4,5-bisphosphate radioactivity was 47 and 70%, respectively. Double labeling of phospholipids with [14C]palmitic acid and [3H]ArA indicated a preferential loss of [3H]ArA within the polyphosphoinositide species after 20 min, but not after 5 min of ischemia. The specific activity of [14C]palmitate remained unchanged. The current data suggest phospholipase C-mediated diacylglycerol formation at the beginning of the insult followed by a phospholipase A2-mediated ArA liberation at a later time, both enzymes presumably acting preferentially on polyphosphoinositide species.     

Regulation of phospholipid metabolism in differentiating cells from rat brain cerebral hemipheres in culture. II. Incorporation of [U-14C]ethanolamine into 1-alkenyl,2-acyl-and 1,2 diacyl-ethanolamine phosphoglycerides.

Cultured dissociated cells from rat embryo cerebral hemisphere incorporate [3H]-and [U-14C]ethanolamine into cellular lipids. Nearly all radioactivity in the lipid fractions is incorporated into 1,2-diacylethanolamine phosphoglycerides and 1-alkenyl,2-acylethanolamine phosphoglycerides (plasmalogen). Kinetic data suggest that the rate of labeling of both ethanolamine phospholipids from the phosphorylethanolamine is similar. A relative increase of the plasmalogen labeling is observed when free ethanolamine is continually present in the medium. The rate of incorporation of label from ethanolamine and phosphorylethanolamine into lipids was measured using a double label technique. Based upon these studies, an independent labeling pattern of the ethanolamine moiety of plasmalogens is suggested. A relative delay for the incorporation of label in plasmalogens could be explained by the presence of a variety of cell types which may differ in their capacity for phospholipid biosynthesis. The rate of incorporation of phosphorylethanolamine into the phosphatidylethanolamine was not affected by the presence of high concentrations of either choline or serine.     

The effect of ionizing irradiation on the lipid biosynthesis by thymus and liver

New Zealand rabbits, fasted for 12 hours, were subjected to 500 rads of whole-body irradiation. Analysis of thymus lipids, at various time intervals following irradiation, showed a threefold increase of triglycerides at 24 hours. Fatty acid composition of the 600 X g supernatant was not affected at 24 hours after irradiation. Lipid biosynthesis from acetate-1-14C by the thymus homogenates was increased to a small extent at 4 hours following irradiation, while the radioactivity distribution into fatty acids was not considerably affected. Contrary to the above findings, fatty acid synthesis from acetate-l-14C by the liver preparations showed a decreased incorporation between the fourth and twelfth hour following irradiation. Counting of the radioactivity of the separated fatty acids suggested that the system for synthesis of short-chain fatty acids was impaired as early as 4 hours following irradiation.     

Glucose uptake in vivo in skeletal muscles of insulin-injected chicks

Glucose uptake across the plasma membrane in animal cells plays a crucial role in whole-body glucose homeostasis. Insulin-stimulated glucose transport activity in vivo in several tissues was estimated using the 2-deoxy-D-[1-(3)H]glucose ([(3)H]2DG) uptake determination method. A tracer dose of [(3)H]2DG was injected intravenously into 8-day-old chicks (Gallus gallus) administered simultaneously or previously with porcine insulin (40 microg/kg BW). After 10 or 20 min, several major tissues, including skeletal and cardiac muscle, were sampled and their 2-deoxy-D-[1-(3)H]glucose 6-phosphate content analyzed. Plasma glucose concentration and [(3)H]2DG radioactivity were lowered by insulin within 20 min of [(3)H]2DG administration, while the plasma [(3)H]2DG/glucose ratio was not significantly different between chicks injected with insulin and their control counterparts. A marked uptake of 2DG was observed in cardiac tissue and brain, followed by kidney and skeletal muscles. In skeletal muscles, insulin increased the 2DG uptake in soleus, extensor digitorum longus and pectoralis superficialis muscles. On the other hand, no significant increases in insulin-induced 2DG uptake were detected in cardiac muscle or adipose tissue compared to controls. The results show that glucose transport across the plasma membrane in vivo in most skeletal muscles tested, but not cardiac muscle, was increased by insulin administration to chicks. These findings suggest that an insulin-responsive glucose transport mechanism is present in chickens, even though they intrinsically lack GLUT4 homologous gene, the insulin-responsive glucose transporter in mammals.     

Metabolism of [1-(14)C]arachidonic acid in ruminant tissues during the pre- and postnatal periods of development under in vitro conditions]

After in vitro incubation of liver, skeletal muscle and adipose tissues from the fetuses and adult cattle as well as of placenta tissue with [1-14C]arachidonic acid, about 90% of the radioactive label was found in the lipids, 10%-in the prostaglandins, and 0.1%-in 14CO2. Arachidonic acid was utilized for the synthesis of lipids and prostaglandins in the majority of fetal tissues in a much greater degree, whereas that in energy-linked process--in a smaller degree compared with adult cattle tissues. [1-14C]arachidonic acid metabolism in the placenta and liver of the given species proceeds much more intensely than that in the skeletal muscles and adipose tissue. In tissues of adult animals [1-14C]arachidonic acid is predominantly utilized for the synthesis of phospholipids, whereas that in fetal tissues is utilized for the synthesis of phospholipids, cholesterol, esters and triglycerides.     

Axonal transport in nigro-neostriatal neurons during morphine tolerance development and abstinence in rats.

Axonal transport of [³H]protein in the nigro-neostriatal pathway in rats was examined during acute and chronic morphine administration and during morphine abstinence. Two days after a microinjection of [³H]lysine into the left substantia nigra zona compacta, more than 95% of the radioactivity present in the rat forebrain was protein-bound. Examination of frozen frontal brain sections revealed that 80–90% of the labelled protein of the injected side was located in brain areas traversed by the nigro-neostriatal pathway. As a positive control, intranigrally administered colchicine reduced the amount of [³H]protein transported after 5 days to the nucleus caudatus-putamen (neostriatum) to approx 18-26% of control. In animals rendered morphine-dependent by subcutaneous implantation of tablets containing 75 mg of morphine base, 27–86% more radioactivity accumulated in the neostriatum at 3, 4 and 5 days after [³H]lysine injection. In contrast, 23–48% less radioactivity was recovered in the neostriatal areas of animals withdrawing from morphine 24 h after [³H]lysine. Gel electrophoresis of soluble and particulate [³H]protein fractions from neostriatal tissues indicated that the gel patterns of radioactivity were not altered by chronic morphine administration. Neither morphine administration nor morphine abstinence altered the rate or amount of [³H]lysine incorporation into protein of the substantia nigra. These data demonstrate that chronic morphine administration was accompanied by a generalized increase in the amount of labelled protein transported to the neostriatum but the procedure was not sufficiently sensitive to detect a minor qualitative alteration of any particular protein(s). Furthermore, these data suggest that either the capacity or the rate of nigro-neostriatal protein transport may be increased during chronic morphine administration in the rat.     

THE EFFECT OF MORPHINE ON THE TRANSPORT AND METABOLISM OF INTRACISTERNALLY-INJECTED LEUCINE IN THE RAT

—Intracisternally injected l or d-[¹⁴C]leucine was retained longer in the brains of morphine-treated rats than in saline-injected control animals. This resulted in higher levels of the labelled leucine and of labelled metabolites of the l-isomer in free pools of brain tissue. However, the absolute levels of brain amino acids and the relative distribution of radioactivity among l-leucine metabolites in brain were unaffected by treatment with morphine, indicating that no disturbance of leucine oxidation through the citric acid cycle was produced by the drug. The inhibition of protein synthesis caused by acute administration of morphine was calculated to be greater than previously reported since morphine treatment increased the specific radioactivity of the free pool of leucine in brain following the intracisternal injection of the labelled amino acid. Possible mechanisms responsible for these morphine effects are discussed.     

Role of peroxisomal fatty acyl-CoA beta-oxidation in phospholipid biosynthesis

We have already reported that peroxisomal beta-oxidation has an anabolic function, supplying acetyl-CoA for bile acid biosynthesis [H. Hayashi and A. Miwa, 1989, Arch. Biochem. Biophys. 274, 582-589]. The anabolic significance of peroxisomal beta-oxidation was further investigated in the present study by using clofibrate, a peroxisome proliferator, as an experimental tool. Clofibrate suppressed 3-hydroxymethylglutaryl-CoA reductase activity (the key enzyme of cholesterol synthesis) and enhanced fatty acyl-CoA oxidase activity (the rate-limiting enzyme of beta-oxidation). Rats were fed a chow containing 0.25% clofibrate for 2 weeks, and then a bile duct fistula was implanted. [1-14C]lignoceric acid, which is degraded exclusively by peroxisomal FAOS, was injected into the rats 24 h after the operation. By this time, the secondary bile acids and pooled cholesterol which would normally be secreted into the bile are considered to have been exhausted from the liver. Clofibrate significantly decreased the incorporations of radioactivity into biliary bile acid (40% of the control) and cholesterol (50%), but did not affect biliary lipid contents. [14C]Acetyl-CoA formed by peroxisomal beta-oxidation of [1-14C]lignoceric acid was preferentially utilized for syntheses of long-chain fatty acids and phospholipids rather than synthesis of cholesterol or triglyceride. The radioactivities incorporated into the former two lipids were increased 2-fold over the control by administration of clofibrate, while the incorporation into triglyceride was decreased to approximately half. In particular, the incorporation into phosphatidylethanolamine was increased as much as 3.5-fold over the control. The contents of these lipids in the liver were not affected by clofibrate. The results suggest that peroxisomal beta-oxidation plays an important role in the biosynthesis of functional lipids such as phospholipids (this work), in addition to bile acids and cholesterol (previous report) by supplying acetyl-CoA.