In vitro substrate utilization for lipid synthesis in liver explants from hyperthyroid chickens

• 1.1. Indian River male broiler chickens growing from 7 to 28 days of age were fed diets containing 12, 18, 24 and 30% protein + 0 or 1 mg triiodothyronine (T₃)/kg of diet to study energetic costs of lipogenesis and the use of various substrates for in vitro lipogenesis.
• 2.2. De novo lipid and CO₂ production were determined in the presence of [1-¹⁴C]pyruvate, [2-¹⁴q]pyruvate, [3-¹⁴C]pyruvate, [2-¹⁴C]acetate and [U-¹⁴C]alanine.
• 3.3. Oxygen consumption was determined in mitochondrial preparations to estimate the energetic costs in expiants synthesizing lipid.
• 4.4. Radiolabeled CO₂ derived from [1-¹⁴C]pyruvate was used as an estimate of coenzyme A availability in liver expiants. Lipids derived from [2-¹⁴C]pyruvate, [2-¹⁴C]acetate and [U-¹⁴C]alanine estimate relative substrate efficiency.
• 5.5. Labeled CO₂ production from [1-¹⁴C]pyruvate was greatest in that group fed a 12% protein diet and least in the group fed a 30% protein diet.
• 6.6. In addition, T₃ increased CO₂ production from [1-¹⁴C]pyruvate.
• 7.7. The production of ¹⁴CO₂ from the second carbon of pyruvate or acetate was increased by T₃.
• 8.8. The low-protein diet (12% protein) increased (P <0.05) lipogenesis.
• 9.9. Adding T₃ to the diets decreased carbon flux into lipid from all substrates, but increased CO₂ production from all substrates without changing stage 3 and 4 respiration rates in mitochondrial preparations.
• 10.10. These observations imply that coenzyme A availability may have regulated de novo lipogenesis in the present study.
• 11.11. It was also concluded that previously noted effects of T₃ on intermediary metabolism may involve metabolic pathways that do not involve changes in mitochondrial function.

     

The pathway of the biosynthesis of non-methylene-interrupted dienoic fatty acids in molluscs

• 1.1. The metabolic fate of 1-¹⁴C-acetate administered to the marine bivalve mollusc Mytilus edulis was investigated.
• 2.2. The active incorporation of the label in 20:2 non-methylene-interrupted dienoic (NMID) fatty acids was found.
• 3.3. Acetate incorporation patterns and specific radioactivity of mussel acids suggest that 22:2Δ7,13 and 22:2/gD7,15 arose by C₂ elongation of 20:2Δ5,11 and 20:2Δ5,13 respectively.
• 4.4. The proposed pathway of NMID fatty acid biosynthesis in molluscs is discussed.

     

Effect of pH on metabolism of the glutamine carbon skeleton by renal cortical mitochondria

• 1.1. To determine the effect of altered acid-base homeostasis on the intramitochondrial metabolism of the glutamine carbon skeleton ¹⁴CO₂ production from [U-¹⁴C]glutamine by isolated rat renal cortical mitochondria was measured.
• 2.2. Mitochondria from rats with chronic metabolic acidosis either showed no change or diminished ¹⁴CO₂ production in comparison with pair fed controls.
• 3.3. By contrast, when the pH of the medium incubating mitochondria from normal rats was manipulated (pH 7.0, 7.4, 7.7), ¹⁴CO₂ production was clearly altered, but the direction and magnitude of the change depended on the glutamine concentration used (0.5 or 10.0 mM).
• 4.4. Mitochondria produced significant quantities of ¹⁴CO₂ when [1,4 ¹⁴C]succinate was used as substrate, indicating that ¹⁴CO₂ production from glutamine does not originate solely from the decarboxylation of α KG.
• 5.5. Thus chronic acidosis and pH, per se, affect intramitochondrial glutamine carbon skeleton metabolism in different fashions, but the specific mechanism cannot be elucidated using ¹⁴CO₂ production from [U-¹⁴C]glutamine.
• 6.6. Additional studies directly quantitating the metabolic products of glutamine have confirmed these findings and more precisely defined the sites of metabolic alteration.

     

The effect of temperature on the acid-base status of the blood of the hatching quail (Coturnix c. japonica)

• 1.1. The ambient temperature of embryos of pipped eggs was reduced from 38 to 28°C for a period of 45 min.
• 2.2. The blood P_CO2 was lower and the blood more alkaline at 28°C than at 38°C.
• 3.3. At 28°C plasma [HCO₃⁻] ] was lower than predicted from the blood buffer line determined in vitro.
• 4.4. The plasma concentrations of strong ions and lactate were the same at both temperatures.
• 5.5. After the ambient temperature had been returned to 38°C for a period of 45 min, blood pH was more acidic than before cooling, but there was no difference in blood P_CO2.
• 6.6. The plasma [HCO₃⁻] was the same as that at 28°C and plasma [K⁺] was higher than before cooling.
• 7.7. The results arc discussed in relation to the factors affecting blood pH in embryos at this stage of development.

     

Fetal lung metabolism: Response to maternal fasting

• 1.1. Fetal lung metabolic response to maternal fasting late in gestation was investigated.
• 2.2. Maternal fasting 4 days before term was associated with low fetal plasma glucose and insulin levels but increased levels of fetal plasma glucagon, glycerol, lactate and fatty acids.
• 3.3. Fetuses from fasted mothers showed a significant decrease in body weight (30%), lung weight (30%) and lung glycogen (46%), but no change in lung protein, phospholipid or total lung DNA, suggesting that lung size is affected more than maturation.
• 4.4. Fetal lung slices incubated in vitro showed that lactate oxidation to CO₂ equalled that of glucose in control fetal lungs and was unaffected by maternal fasting, while glucose oxidation was depressed (23%).
• 5.5. Maternal fasting significantly decreased in vitro incorporation of [U-¹⁴C]-glucose, [U-¹⁴C]lactate and [1-¹⁴C]palmitate into lung phospholipids.
• 6.6. Fetal lungs from fasted mothers showed increased conversion of lactate to glucose, indicating gluconeogenic potential by fetal lung.
• 7.7. These studies show that plasma lactate serves as an important energy fuel and substrate for lipid synthesis for the fetal lung, and maternal fasting markedly alters fetal lung metabolism.

     

Respiration in the eastern water dragon,Physignathus lesueurii (agamidae)

• 1.1. Some aspects of the gas exchange system of a diving lizard, Physignathus lesuewii were studied.
• 2.2. Breathing patterns were analysed.
• 3.3. Breathing rate increases logarithmically with temperature and Q₁₀ = 1.8. LogBR = −0.237 + 0.0256 T.
• 4.4. Gas tensions in lung air and arterial and venous blood were measured. Arterial pH declines with increasing temperature.
• 5.5. Temperature has a marked effect on oxygen affinity of the blood (ΔH = −10.1 kcal mol⁻). A Bohr effect was also noted.
• 6.6. CO₂ equilibrium curves were drawn.
• 7.7. The results are considered with a view to anticipating the efficiency of the gas exchange system of this species under conditions of variable temperature and during diving.

     

Bioelectrical potentials across the mantle epithelium of Achatina fulica

• 1.1. The shell side of the mantle of Achatina fulica is several millivolts positive to the blood side in vitro.
• 2.2. The electrical potential does not depend on Na⁺, Ca²⁺, Mg²⁺, K⁺ or HCO₃⁻ but requires the presence of chloride on the shell side.
• 3.3. The potential difference and short-circuit current ranged from 3.0 to 30.0 mV and 15.0 to 75 μA/cm² with averages at 10m V and 50 μA/cm² respectively.
• 4.4. The electrical gradient is reduced by 2,4-dinitrophenol, thiocyanate and furosemide but not by ouabain, CO₂ or acetozolamide.
• 5.5. It is suggested that the nature and mechanism of electrogenesis in Achatina parallels that of the Helix mantle.

     

Metabolic utilization of leucine in a fat and a lean line of chicken

• 1.1. In vivo incorporation into body lipids and breast muscle proteins from l-[U-¹⁴C]leucine was studied in genetically lean or fat male chickens, fed or starved, 1 or 24 hr after intraperitoneal injection.
• 2.2. Lipogensis and portein synthesis from labelled leucine were significantly higher in fat chickens than in lean birds, particularly in those in the fed state.
• 3.3. Radioactivity in the free amino acid pool was greater in fat birds irrespective of the nutritional state.
• 4.4. However, utilization of injected l-[U-¹⁴C]leucine for lipogenesis was no more than 2%.

     

Mechanism of pepsin-catalyzed aminotranspeptidation reactions

• 1.1. The tetrapeptide Ala₂-Nph₂ (where Nph = p-nitrophenylalanyl) is treated by porcine pepsin to study the mechanism of aminotranspeptidation reactions.
• 2.2. The major initial product is Ala₂-Nph and the major transpeptidation products are Nph₂ and Nph₃ accompanied by some Nph, a little Nph₄, Ala₂-Nph₃ and Ala₂-Nph₄.
• 3.3. Oligomers of Nph greater than tetramers are formed near the end of the reaction.
• 4.4. In presence of [³H]Nph, no incorporation of Nph into the transpeptidation products is observed.
• 5.5. ¹⁸O-Iabeling shows extensive incorporation of ¹⁸O atoms from [¹⁸O]water in the carbonyl oxygens of Nph residues.

     

Influence of dietary sources of fat on lipid synthesis in mink (Mustela vison) mammary tissue

• 1.1. The fatty acid composition of the triglyceride fraction of mink milk sampled during mid-lactation (day 28 post partum) from two nursing mink was compared to that of plasma samples and to the fatty acid composition of the feed rations used.
• 2.2. Chemical analysis of the triglyceride composition of mink milk demonstrated only minute concentrations of fatty acids with a chain length below C₁₄.
• 3.3. The saturated C_16:0- and C_18:0-unit fatty acids in mink milk made up for 24–40% of the total amount of fatty acids extracted, the remainder being represented by mono and polyunsaturated long-chain (C₁₆-C₂₄) fatty acids.
• 4.4. Preliminary in vitro experiments proved the incorporation of¹⁴C-labelled glucose, acetate or palmitate into triacylglycerols in cultures of mink mammary tissue to be linear for at least 2 hr.
• 5.5. The in vitro capacity for de novo fatty acid synthesis in mink mammary tissue using 14C-labelled glucose or acetate was low, i.e. ranging from 0.096–0.109 nmol/g (fresh tissue)/min, and amounted to only about 5% of that obtained in the case of [¹⁴C]palmitic acid incubation.
• 6.6. Following ¹⁴C-labeIled acetic or palmitic acid incubation of mink mammary tissue neither desaturation nor chain elongation was observed.
• 7.7. In response to long-term feeding on rations with two different sources of animal fat (F = fish oil or L = lard) the influence of compositional changes in dietary neutral lipids on the fatty acid composition of the lipids of mink milk is discussed.

     

Anabolic and catabolic pathways of pyrimidine metabolism in rat liver

• 1.1. Use of pyrimidine nucleotide precursors labelled in various positions on the ring shows that in rat liver, pyrimidine nucleotides formed from orotate follow anabolic pathways almost exclusively, whereas trace quantities of uridine are mostly degraded to β-alanine and its metabolites.
• 2.2. Annomalies in the ratios of [¹⁴C] and [³H] in various common nucleotide products of orotate and uridine can be accounted for on the basis of metabolic compartmentation and recycling of CO₂.

     

Intramitochondrial reductive carboxylation of 2-oxoglutarate in adipose tissue and its contribution to fatty acid synthesis

• 1.1. The reductive carboxylation of 2-oxoglutarate was found to proceed in mitochondria of rat epididymal fat pads and rabbit perirenal adipose tissue at a rate similar to that in liver mitochondria.
• 2.2. In rat fat pads the incorporation of ¹⁴C from [5-¹⁴C]2-oxoglutarate into fatty acids via the carboxylation was suppressed by butylmalonate by 30%.
• 3.3. 2-Oxoglutarate and glutamate stimulated the incorporation into fatty acids of ¹⁴C from [2-¹⁴C]acetate in rat fat pads with the simultaneous reduction of tissue NADP. These effects persisted after inhibition of succinate dehydrogenase by malonate.
• 4.4. It is concluded that in adipose tissue 2-oxoglutarate carboxylation proceeds in both the cytoplasm and mitochondria. Therefore, it can supply carbon atoms as well as NADPH for fatty acid synthesis.

     

Light-induced transient increase of the activity of topoisomerase I in plasmodia of Physarum polycephalum

• 1.1. Activity of topoisomerase I and incorporation of [³H]uridine and [¹⁴C]thymidine were monitored during light-induced sporulation of the slime mold Physarum polycephalun.
• 2.2. A 4-fold transient increase of topoisomerase I activity but not of [³H]uridine or [¹⁴C]thymidine incorporation was observed after 42 hr of illumination with 6 hr impulses.
• 3.3. The activity of topoisomerase I did not increase in the absence of light impulses. However, ca 5-fold increase of the activity was observed in dark when 100 μ M dibutyryl-cAMP was administered 12 hr before harvesting of plasmodia.
• 4.4. Fluorodeoxyuridine and cycloheximide administered 36 hr after starting of the illumination cancelled the increase of the activity of topoisomerase I.
• 5.5. After 7 days of the illumination, when fruiting bodies appeared, the activity of topoisomerase I dropped to about 15% of the initial value.

     

Respiratory properties of blood and hemoglobin solutions from the piranha

• 1. Respiratory properties of piranha blood are distinguished from those of other fish primarily by the high CO₂ buffering capacity (ΔHCO₃/⁻ΔpH= 19.6mmol/l for oxygenated blood and 39.1 mmol/l for deoxygenated blood).
• 2. The concentration of nucleoside triphosphates (NTP) and the half-saturation tension (P₅₀) of whole blood were found to be inversely related to body size.
• 3. The higherP₅₀ in smaller fish, analogous to values obtained in previous studies involving interspecies comparisons, could be adaptive to a higher weight-specific metabolic rate.
• 4. Both ATP and guanosine triphosphate (GTP) lowered the oxygen affinity of purified hemoglobin solutions, accounting for the size-dependent correlation ofP₅₀ and NTP concentration in whole blood.
• 5. While similar in concentration in red cells, GTP is more potent than ATP as an allosteric modifier of hemoglobin function.

     

Sulphate conjugation is the main route of pentachlorophenol metabolism in Daphnia magna

• 1.1. Daphnia magna were exposed for 24 hr to ¹⁴C-labelled pentachlorophenol (PCP) at an initial concentration of 20μg/l in the incubation water. Occurrence of free PCP and its metabolites were measured both from the animals and the water.
• 2.2. Hydrophilic metabolites excreted into water were analysed, after acid or enzymatic hydrolyses, with a liquid-liquid extraction and TLC.
• 3.3. PCP was metabolized and excreted, perhaps solely, via the sulphate conjugation. The average excretion rate, 2.65nmol/g/hr, accounted for 35% of the absorption rate measured at the start of exposure.
• 4.4. Neonate daphnids had an equal ability to metabolize PCP as the older animals. Bioconcentration in young animals was, however, only 23% of that in adult ones.
• 5.5. Effect of naturally humic water on metabolization and excretion of PCP was negligible.

     

Phosphoinositides and inositol phosphates in Discopyge tschudii electrocyte membranes

• 1.1. The metabolism of inositol (Ins)-containing phospholipids and inositol phosphates has been studied by following the incorporation and distribution of myo-[³ H]Ins in metabolically active electrocytes from the electric ray Discopyge tschudii.
• 2.2. The apparent initial rate of myo-[³H]Ins incorporation into total phosphoinositides was ca 8.2 fmol/mg protein/hr. Phosphatidylinositol (Ptdlns) displayed the highest levels of labelling. Lithium inhibited this incorporation probably by limiting the recycling of myo-[³H]Ins from [³H]Ins-monophosphate.
• 3.3. The formation of water-soluble products of phosphoinositides between 7 and 24 hr was 4.1 ± 0.2, 0.4 ± 0.2 and 3.0 ± 1.0 fmol/μmmol total lipid phosphorus for myo-[³H]InsP, -InsP₂ and Ins-P₃ respectively.
• 4.4. Lithium ions are shown to modulate phosphoinositide synthesis and Ins-phosphate accumulation. Ins-mono-, bis- and tris-phosphate production was enhanced 5-, 3- and 2-fold by Li +.
• 5.5. The above results suggest the participation of a C-type phospholipase and of Li-sensitive phosphatases in the modulation of phosphoinositide metabolism in the electrocyte.

     

Effect of lindane on phosphatidylinositol synthesis by cerebral cortex after acute and subchronic treatment

• 1.1. The incorporation of myo-[2-³H]inositol into phosphatidylinositols was unmodified in brain cortex miniprisms from convulsant rats.
• 2.2. However, the incorporation had increased by 300–400% in non convulsant rats which had received the same amount of lindane at a lower concentration.
• 3.3. This result suggests that phosphatidylinositols are implicated in the convulsion syndrome.
• 4.4. Experiments with lindane added in vitro were performed with both subchronically lindane intoxicated and untreated rats.
• 5.5. The results show an interesting lack of parallelism.
• 6.6. This might indicate the development of some resistance to the effects of lindane, possibly as the result of complex compensatory changes in inositol lipid biosynthesis.

     

Biosynthesis of lipid-linked oligosaccharides in embryonic liver. Glucosylation of mannose containing dolichyl diphosphate oligosaccharide

• 1.1. A maximum rate of dolichyl phosphate [¹⁴C]glucose synthesis from 55-day embryos was achieved at 16nM concentration of exogenous dolichyl phosphate and exceeded about 3 times that without addition of dolichyl phosphate.
• 2.2. The highest values of [¹⁴C]glucose incorporation from UDP-[¹⁴C]glucose into dolichyl phosphate [¹⁴C]glucose, dolichyl diphosphate [¹⁴C]Glc-oligosaccharides and proteins were reached at 5 min time point of incubation of liver microsomes both from embryos and sows.
• 3.3. The radioactive incorporation into proteins was about 7-fold higher in liver microsomes from sows compared to that from embryos, probably due to the greater content of acceptor proteins in microsomes from sows.
• 4.4. The enzymatic transfer of Glc₃-oligosaccharide from a lipid carrier to endogenous protein acceptor in microsomes from pig embryonic and adult livers was considerably faster than the removal of glucose residues during the initial stages of processing of protein-bound oligosaccharides.
• 5.5. One labelled compound was discovered in the Chcl₃-Ch₃Oh-H₂O (1:1:0.3, by vol) extract after incubation of liver microsomes from embryos and sows with UDP-[¹⁴C]glucose. On the basis of its mobility on the chromatogram it appears to be GlcNAc₂Man₉Glc₃.

     

An investigation of gluconeogenesis in marine teleosts,and the effect of long-term exercise on hepatic gluconeogenesis

• 1.1. Gluconeogenesis from ¹⁴C-substrates was measured in liver slices from marine teleosts.
• 2.2. 0.56 to 2.78 μmols of lactate were incorporated/g wet wt/hr with the higher rates corresponding to the active species.
• 3.3. Snapper (Chrysophrys auratus) and bream (Acanthopagrus butcheri) exercised continuously for 14 days showed a substantial increase in the incorporation of lactate; snapper confined for 4 months showed a significant decrease in the incorporation of lactate, compared to freshly caught individuals.
• 4.4. Pyruvate carboxylase and fructose 1,6-diphosphatase were found in red muscle. Some phosphoenolpyruvate carboxykinase may also be present. The three enzymes were present in liver. Possible roles for the enzymes in teleost muscle are discussed.