scholarly journals The metabolism of phospholipids in mouse brain slices

1966 ◽  
Vol 101 (3) ◽  
pp. 674-679 ◽  
Author(s):  
PA Clayton ◽  
CE Rowe
Keyword(s):  
Phosphatidic Acid ◽  
Mouse Brain ◽  
Grey Matter ◽  
Microsomal Fraction ◽  
Brain Slices ◽  
Mitochondrial Fraction ◽  
Subcellular Fractions ◽  
The Other ◽  
Metabolic Inhibitors ◽  
Radioactive Labelling

1. Slices of mouse brain grey matter were incubated with [(32)P]phosphate and [1-(14)C]acetate. Doubly labelled phospholipids were extracted from subcellular fractions prepared from the slices in a mixture of metabolic inhibitors, under conditions where there was negligible change in radioactive labelling during the preparation. Two tissue fractions were studied in detail; one contained a high proportion of mitochondria and the other was mainly microsomal. 2. In all tissue fractions the highest incorporations of both [(32)P]phosphate and [1-(14)C]acetate occurred into phosphatidylcholine. 3. After incubation for 1hr., the (32)P/(14)C ratios for phosphatidylcholine, phosphatidylethanolamine and phosphatidic acid in the mitochondrial fraction were similar to those in the microsomal fraction. 4. The (32)P/(14)C ratios were similar in phosphatidylcholine and phosphatidylethanolamine and much lower than those in phosphatidic acid and phosphatidylinositol.

Comparative studies of CDP-diacylglycerol synthase in rat liver mitochondria and microsomes

1993 ◽  
Vol 71 (3-4) ◽  
pp. 183-189 ◽  
Author(s):  
Amy Y. P. Mok ◽  
Gordon E. McDougall ◽  
William C. McMurray
Keyword(s):  
Phosphatidic Acid ◽  
Rat Liver ◽  
Mitochondrial Fraction ◽  
Liver Mitochondria ◽  
Subcellular Fractions ◽  
Rat Liver Mitochondria ◽  
Mitochondrial Enzyme ◽  
Egg Lecithin ◽  
Microsomal Membranes ◽  
Concentration Curves

CDP-diacylglycerol for polyglycerophosphatide biogenesis can be synthesized within rat liver mitochondria. Contamination by microsomal membranes cannot account for the CDP-diacylglycerol synthesis found in the mitochondria. Phosphatidic acid from egg lecithin was the best substrate for the synthesis of CDP-diacylglycerol in both subcellular fractions. Concentration curves for CTP and Mg2+ differed for the two subcellular fractions. Microsomal CDP-diacylglycerol synthase was specifically stimulated by the nucleotide GTP; this stimulatory effect by GTP was not observed in the mitochondrial fraction. By comparison, the microsomal enzyme was more sensitive towards sulfhydryl inhibitors than the mitochondrial enzyme. The enzymes could be solubilized from the membrane fractions using 3-[(cholamidopropyl)dimethylammonio]-1-propanesulfonate, and the detergent-soluble activity could be partially restored by addition of phospholipids. Based on the differences in properties, it was concluded that there are two distinct enzyme localizations for CDP-diacylglycerol synthesis in mitochondria and microsomes from rat liver.Key words: CDP-diacylglycerol, synthase, phosphatidic acid, mitochondria, microsomes, solubilization.

scholarly journals Biogenesis of mitochondria. Phospholipid synthesis in vitro by yeast mitochondrial and microsomal fractions

1974 ◽  
Vol 144 (2) ◽  
pp. 265-275 ◽  
Author(s):  
G S Cobon ◽  
P D Crowfoot ◽  
A W Linnane
Keyword(s):  
Phosphatidic Acid ◽  
Microsomal Fraction ◽  
De Novo ◽  
Specific Activity ◽  
Neutral Lipids ◽  
Mitochondrial Fraction ◽  
Biogenesis Of Mitochondria ◽  
Glycerolipid Synthesis ◽  
Phosphatidylcholine Synthesis

The ability in vitro of yeast mitochondrial and microsomal fractions to synthesize lipid de novo was measured. The major phospholipids synthesized from sn-[2-3H]glycerol 3-phosphate by the two microsomal fractions were phosphatidylserine, phosphatidylinositol and phosphatidic acid. The mitochondrial fraction, which had a higher specific activity for total glycerolipid synthesis, synthesized phosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylserine and phosphatidic acid, together with smaller amounts of neutral lipids and diphosphatidylglycerol. Phosphatidylcholine synthesis from both S-adenosyl[Me-14C]methionine and CDP-[Me-14C]choline appeared to be localized in the microsomal fraction.

scholarly journals Subcellular distribution of cytochrome c in rat liver. Methods for its extraction and purification

1967 ◽  
Vol 105 (2) ◽  
pp. 427-442 ◽  
Author(s):  
N. F. González-Cadavid ◽  
P. N. Campbell
Keyword(s):  
Cytochrome C ◽  
Rat Liver ◽  
Subcellular Distribution ◽  
Microsomal Fraction ◽  
Gradient Centrifugation ◽  
Ammonium Phosphate ◽  
Mitochondrial Fraction ◽  
Subcellular Fractions ◽  
Nuclear Fraction ◽  
Extraction And Purification

1. A method for the extraction and purification of cytochrome c from rat liver is described. The method depends on multiple chromatography on Amberlite IRC-50 with elution with ammonium phosphate buffers of differing ionic composition and pH, interspersed with gel filtration with Sephadex G-25. Conditions leading to denaturation are avoided and the product is chromatographically pure. 2. The method may be used for the quantitative analysis of cytochrome c either in unfractionated liver or in subcellular fractions. 3. Two pools of cytochrome c were detected, one extractable at pH4·0 with distilled water and the other extracted from the residues of the first extraction with 0·15m-sodium chloride. 4. For subcellular distribution studies the liver was homogenized in 0·3m-sucrose and a nuclear fraction (washed thoroughly to remove trapped mitochondria), a mitochondrial fraction, a heavy microsomal fraction, a standard microsomal fraction and the cell sap were isolated. The mitochondrial fraction was subfractionated further by density-gradient centrifugation. Each fraction was analysed for protein, RNA, DNA, succinate–neotetrazolium oxidoreductase and glucose 6-phosphatase. 5. A total of 123μg. of cytochrome c was obtained/g. wet wt. of rat liver. 6. Values for the percentage subcellular distribution of cytochrome c are: nuclear fraction, 24·4; mitochondrial fraction, 57·2; heavy microsomal fraction, 5·2; standard microsomal fraction, 10·6; cell sap, 2·7. 7. Three out of the eight mitochondrial subfractions separated by gradient centrifugation contained 76% of the cytochrome c and 85% of the succinate–neotetrazolium oxidoreductase present in the mitochondrial fraction. 8. In unfractionated liver 94% of the cytochrome c was extracted at pH4·0 with water whereas in most of the subcellular fractions the corresponding value was approx. 75–80%.

Phosphatidic acid synthesis in the heart. 1. Effect of age and species difference in the mitochondrial and microsomal synthesis

1977 ◽  
Vol 55 (4) ◽  
pp. 308-314 ◽  
Author(s):  
K. J. Kako ◽  
G. Zaror-Behrens ◽  
S. D. Peckett
Keyword(s):  
Bovine Serum Albumin ◽  
Phosphatidic Acid ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Mitochondrial Fraction ◽  
Subcellular Fractions ◽  
Newborn Rats ◽  
Adult Rats ◽  
Rat Hearts ◽  
Weanling Rats

Rates of syntheses of monoacyl- and diacyl-glycerol 3-P (phosphate) were determined in the mitochondrial and microsomal fractions prepared from hearts of rats and rabbits, to compare characteristics of the acylation reactions by the two subcellular fractions. The assays were carried out with the subcellular fractions prepared from (i) hearts of hyperthyroid animals, and (ii) hearts of newborn and weanling rats. In addition, the effect of an addition of bovine serum albumin in the assay system was examined. (1) Administration of thyroid hormones increased the acyltransferase activity in rabbit hearts but not that in rat hearts. (2) Mitochondrial and microsomal fractions of hearts of newborn rats acylated glycerol 3-P at a rate 1.3–4 times greater than those of adult rats. The rate of acylation by the mitochondrial fraction of weanling rats was also high, but the rate of microsomal acylation was slightly lower than that of adult rats. By contrast, in newborn rats, diacylglycerol 3-P formation by the liver microsomes was not greater than that of the adults, although its formation by the newborn liver mitochondria was greater. (3) The accumulation of monoacylglycerol 3-P during the assay was accelerated by the addition of increasing amounts of bovine serum albumin. Therefore, the monoacylglycerol 3-P formation was less than 10% of the diacylglycerol 3-P formation in the assay containing no albumin and with the rat subcellular fractions, whereas nearly four times more monoacyl- than diacyl-glycerol 3-P was synthesized in the presence of 20 mg albumin. (4) The ratio of monoacyl- to diacyl-glycerol 3-P formation by the mitochondrial fraction was greater than that of microsomal fraction at any concentration of albumin in both rats and rabbits. At an equal albumin concentration in the assay, relatively more diacyl- than monoacyl-glycerol 3-P was formed in the mitochondrial fraction of newborn rat hearts as compared with adult hearts. (5) In conclusion, our data concerning the age and species differences in acyltransferase activities support a view that the mitochondrial fraction of both rat and rabbit hearts, in addition to the microsomal enzymes, is capable of catalyzing the de novo synthesis of phosphatidic acid.

scholarly journals The distribution of molecular species of phosphatidylinositol in ox brain and its subcellular fractions

1972 ◽  
Vol 128 (3) ◽  
pp. 587-595 ◽  
Author(s):  
M. G. Luthra ◽  
A. Sheltawy
Keyword(s):  
White Matter ◽  
Molecular Species ◽  
Regional Differences ◽  
Grey Matter ◽  
Subcellular Fractions ◽  
The Other ◽  
Cerebral Hemispheres ◽  
Molecular Species Composition ◽  
Its Regions ◽  
Post Mitochondrial Supernatant

1. The phosphatidylinositol content of white and grey matter of ox cerebral hemispheres did not differ. The phosphatidylinositol from grey matter was slightly enriched in palmitic acid and arachidonic acid, and that from white matter was enriched in eicosatrienoic (C20:3) acid. These regional differences were apparently due to the greater content of myelin in the white matter, since the same tendencies were observed when combined myelinic and non-myelinic subcellular fractions prepared from the cerebral hemispheres were compared. 2. Purified phosphatidylinositol was converted into its triacetylated methylated derivative and resolved to its molecular species by t.l.c. on AgNO3-impregnated silica gel. The tetraenoic molecular species was predominant in phosphatidylinositol from ox cerebral hemispheres, and this feature characterized all the phosphatidylinositol samples extracted from its regions or subcellular fractions. The grey matter was more enriched in the tetraenoic species and the white matter in the trienoic species. 3. The molecular-species composition of phosphatidylinositol from the subcellular fractions of ox cerebral hemispheres was studied. The trienoic species constituted nearly one-fifth of the phosphatidylinositol from two myelinic fractions. ‘Large myelin’ was more enriched in this species than was ‘small myelin’. Both fractions also contained greater concentrations of the dienoic species than the non-myelinic subcellular fractions. The latter fractions, one containing nuclei and the other nerve endings plus mitochondria, were enriched in the monoenoic and tetraenoic species of phosphatidylinositol. The post-mitochondrial supernatant exhibited a pattern of distribution of phosphatidylinositol species intermediate between the myelinic and non-myelinic fractions.

Ca2+fluxes in developingTrichoderma viridemycelium

2000 ◽  
Vol 46 (4) ◽  
pp. 312-324 ◽  
Author(s):  
Martin ŠŠimkovič ◽  
Svetlana Kryšštofová ◽  
L'udovít Varečka
Keyword(s):  
Life Cycle ◽  
Energy Metabolism ◽  
Microsomal Fraction ◽  
Trichoderma Viride ◽  
Mitochondrial Fraction ◽  
The Other ◽  
Submerged Cultivation ◽  
Transport Systems ◽  
Mycelium Growth ◽  
X Ray

The properties of both Ca2+influx and efflux in the mycelium during the life cycle of Trichoderma viride were studied by means of45Ca2+and by X-ray fluorescence spectroscopy measurements. The properties of the45Ca2+influx and effluxes indicate that they are mediated by different transport systems. The Ca2+influx could be mediated by an electrogenic Ca2+/nH+antiport, or by an Ca2+uniport system. Both Ca2+influx and efflux were stimulated by the uncouplers (and the treatment leading to the suppression of energy metabolism) and by azalomycin F, an antifungal agent. Salicylate stimulated the Ca2+efflux, but inhibited the Ca2+influx. In the isolated preparation of crude vacuolar/mitochondrial fraction, salicylate induced the Ca2+release, as did A23187. Azalomycin F moderately released Ca2+from the microsomal fraction. On the other hand, uncouplers did not release Ca2+from the isolated organelles, but inhibited to a different extent the ATP-dependent and -independent Ca2+influx. The results could be explained in terms of the capacitative Ca2+influx mechanism. The rate of45Ca2+influx, or of the40Ca2+content, was maximal after about 30 h of submerged cultivation, and then decreased. The results show that loading of internal Ca2+stores occurs in the early stages of the development of mycelium only, and the Ca2+influx mechanism is developmentally down-regulated, being almost nonexistent during its later stages. In older mycelium, growth seems to be autonomous of the extracellular Ca2+until the onset of conidiation.Key words: Trichoderma viride, development, Ca2+influx, Ca2+efflux, salicylate, uncoupler, azalomycin F.

Calcium uptake by subcellular fractions of human umbilical artery

1976 ◽  
Vol 231 (4) ◽  
pp. 1074-1081 ◽  
Author(s):  
RI Clyman ◽  
VC Manganiello ◽  
CJ Lovell-Smith ◽  
M Vaughan
Keyword(s):  
Calcium Uptake ◽  
Umbilical Artery ◽  
Microsomal Fraction ◽  
Heat Stability ◽  
Mitochondrial Fraction ◽  
Subcellular Fractions ◽  
Atp Concentration ◽  
Mitochondrial Fractions ◽  
Human Umbilical Artery ◽  
Arterial Patency

Two different mechanisms for the active accumulation of Ca2+ by subcellular fractions of human umbilical artery are described. One, located in the mitochondrial fraction, was induced by exogenous ATP or respiratory substrates (ADP and succinate) and was inhibited by azide. The other, located in the microsomal fraction, was induced by ATP and potentiated by oxalate, but not inhibited by azide. Increasing ATP concentrations up to 4-5 mM increased microsomal Ca2+ accumulation, whereas increasing ATP concentration above 2-3 mM caused inhibition of mitochondrial Ca2+ uptake. Although changing pH from 7.4 to 7.2 had no effect on mitochondrial Ca2+ accumulation, it doubled microsomal uptake. Neither adenosine 3',5'-monophosphate nor guanosine 3',5'-monophosphate in the presence or absence of protein kinase and kinase modulator affected Ca2+ uptake by or phosphorylation of the subcellular fractions. Partially purified protein kinases from umbilical and beef skeletal muscle contained a component(s) distinguishable from the kinase on the basis of its heat stability that enhanced ATP-induced Ca2+ uptake by mitochondrial fractions from the umbilical artery. It is suggested that alterations in Ca2+ sequestration induced by changes in ATP concentration and intracellular pH in mitochondrial and microsomal fractions, respectively, could play a role in the control of arterial patency and closure with changes in PO2.

Enhancement of the Release Reaction Not Accompanied with Enhanced Phosphatidylinositol Turnover in the Reserpinized Rabbit Blood Platelets

1983 ◽  
Vol 50 (02) ◽  
pp. 595-600 ◽  
Author(s):  
Y Watanabe ◽  
M Soda ◽  
N Fukamachi ◽  
B Kobayashi
Keyword(s):  
Phosphatidic Acid ◽  
Blood Platelets ◽  
Specific Protein ◽  
The Other ◽  
Release Reaction ◽  
Rabbit Blood ◽  
Phosphatidylinositol Turnover ◽  
Activated State ◽  
32P Incorporation ◽  
Platelet Release

SummaryThrombin-induced platelet release reaction examined with secretion of calcium and N-acetylglucosaminidase was significantly enhanced in the platelets from reserpine-treated rabbits as compared with the control. On the other hand, 32P-incorporation into phosphatidic acid was suppressed in the reserpinized platelets in activated state. Thrombin induced phosphatidylinositol (PI)- breakdown, which was examined by decreases in radioactivity and content of PI, and an increase in diacylglycerol, was not enhanced in the reserpinized platelets as compared with the control. The phosphorylation of the specific protein coupled to thrombin- induced platelet PI-breakdown was not stimulated in the reserpinized platelets as compared with the control. In contrast to PI, PC-degradation by thrombin was significantly stimulated in the reserpinized platelets. Possible existence of pathway(s) other than that associated with an enhancement of Pl-tumover is conceivable as a mechanism involved in platelet release reaction.

scholarly journals Quantitative measurement of reactive oxygen species in ex vivo mouse brain slices

2021 ◽  
Vol 2 (1) ◽  
pp. 100332
Author(s):  
Chirag Vasavda ◽  
Solomon H. Snyder ◽  
Bindu D. Paul
Keyword(s):  
Reactive Oxygen Species ◽  
Mouse Brain ◽  
Quantitative Measurement ◽  
Ex Vivo ◽  
Brain Slices ◽  
Reactive Oxygen ◽  
Oxygen Species

scholarly journals Patch-clamp and multi-electrode array electrophysiological analysis in acute mouse brain slices

2021 ◽  
Vol 2 (2) ◽  
pp. 100442
Author(s):  
Kevin M. Manz ◽  
Justin K. Siemann ◽  
Douglas G. McMahon ◽  
Brad A. Grueter
Keyword(s):  
Patch Clamp ◽  
Mouse Brain ◽  
Brain Slices ◽  
Electrode Array ◽  
Electrophysiological Analysis ◽  
Multi Electrode Array
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