scholarly journals Disialoganglioside GDla of rat brain subcellular particles during development

1978 ◽  
Vol 174 (2) ◽  
pp. 655-657 ◽  
Author(s):  
H K M Yusuf ◽  
J W T Dickerson
Keyword(s):  
Rat Brain ◽  
Microsomal Fraction ◽  
Subcellular Fractionation ◽  
Dendritic Arborization ◽  
Total Ganglioside ◽  
Rat Cerebrum

The increase observed in the amount of the disialoganglioside GDlof the rat cerebrum during development between 21 and 81 days of age accounted for nearly 40% of the overall increase in total ganglioside in the tissue during the same period. Subcellular fractionation showed the microsomal fraction to contribute by far the most towards this increase in Cerebral ganglioside GDla. It is suggested that microsomal ganglioside GDla may serve as a marker for dendritic arborization in the rat cerebrum.

Secretagogue-induced changes in subcellular Ca2+ distribution in isolated pancreatic acini

1981 ◽  
Vol 240 (2) ◽  
pp. G130-G140
Author(s):  
R. L. Dormer ◽  
J. A. Williams
Keyword(s):  
Atomic Absorption Spectrometry ◽  
High Speed ◽  
Microsomal Fraction ◽  
Absorption Spectrometry ◽  
Subcellular Fractionation ◽  
Differential Centrifugation ◽  
Zymogen Granules ◽  
Pancreatic Acini ◽  
Induced Changes ◽  
Stimulation Of

In a prior study, we demonstrated that pancreatic secretagogues increased both the uptake into and washout of 45Ca2+ from isolated mouse pancreatic acini. The net result of these processes was an initial fall in total acinar cell Ca2+ content. In the present study, we have employed subcellular fractionation of acini under conditions that minimized posthomogenization redistribution of Ca2+ in order to localize those organelles involved in intracellular Ca2+ fluxes. Homogenization and differential centrifugation of acini, preloaded with 45Ca2+ and subjected to a period of washout, showed that carbachol induced an increased loss of 45Ca2+ from all fractions isolated. The high-speed microsomal fraction lost 45Ca2+ to a greater extent than did whole acini; measurement of total Ca2+ by atomic absorption spectrometry showed a net loss of Ca2+ from this fraction. Purification of the lower-speed fractions indicated that carbachol increased 45Ca2+ exchange with both zymogen granules and mitochondria, but net Ca2+ levels in these organelles were unchanged. It was concluded that stimulation of pancreatic acini by carbachol results in the release of calcium from a microsomal compartment leading to a rise in cytoplasmic Ca2+, increased exchange with granule and mitochondrial Ca2+, and increased efflux of Ca2+ from the cell.

Redistribution of rat brain esterases during subcellular fractionation

1969 ◽  
Vol 183 (2) ◽  
pp. 253-264 ◽  
Author(s):  
A.H. Koeppen ◽  
K.D. Barron ◽  
J. Bernsohn
Keyword(s):  
Rat Brain ◽  
Subcellular Fractionation

scholarly journals The biosynthesis of sulphogalactosyldiacylglycerol of rat brain in vitro

1977 ◽  
Vol 166 (3) ◽  
pp. 429-435 ◽  
Author(s):  
G. Subba Rao ◽  
Leonard N. Norcia ◽  
Joanne Pieringer ◽  
Ronald A. Pieringer
Keyword(s):  
Enzyme Activity ◽  
Rat Brain ◽  
Enzyme Preparation ◽  
Microsomal Fraction ◽  
Inhibitory Effect ◽  
Subcellular Fractions ◽  
Triton X ◽  
Derivatives Of

Triton X-100 extracts of rat brain microsomal fraction catalyse the formation of sulphogalactosyldiacylglycerol from galactosyldiacylglycerol and adenosine 3′-phosphate 5′-sulphatophosphate. Of the various subcellular fractions of brain assayed, the microsomal fraction contained most (79%) of the adenosine 3′-phosphate 5′-sulphatophosphate–galactosyldiacylglycerol sulphotransferase activity. The enzyme activity was stimulated by Triton X-100 and showed linearity with increasing time, concentrations of enzyme and added substrates. ATP and KF prolonged the linearity of the activity with time, but ATP had an overall inhibitory effect on the sulphotransferase. Both ATP and KF inhibit the degradation of adenosine 3′-phosphate 5′-sulphatophosphate, which probably causes the increased linearity of the sulphotransferase reaction with time. The enzyme preparation did not catalyse the transfer of sulphate from adenosine 3′-phosphate 5′-sulphatophosphate to either cholesterol or galabiosyldiacylglycerol (galactosylgalactosyldiacylglycerol). Significant differences between the formation of sulphogalactosyldiacylglycerol and cerebroside sulphate catalysed by the same enzyme preparation were noted. ATP and Mg2+ strongly inhibit the formation of sulphogalactosyldiacylglycerol but equally strongly stimulate the synthesis of cerebroside sulphate. The apparent Km for galactosyldiacylglycerol is 200μm, and that for cerebroside is 45μm. Galactosyldiacylglycerol and cerebroside are mutually inhibitory toward the synthesis of sulphated derivatives of each. These data do not necessarily lead to the conclusion that two sulphotransferases are present, but they do indicate a possible means of controlling the synthesis of these two sulpholipids.

scholarly journals Activation of a peroxisome-proliferating catabolite of cholic acid to its CoA ester

1993 ◽  
Vol 296 (1) ◽  
pp. 265-270 ◽  
Author(s):  
T Nishimaki-Mogami ◽  
A Takahashi ◽  
Y Hayashi
Keyword(s):  
Cholic Acid ◽  
Rat Liver ◽  
Microsomal Fraction ◽  
Liver Microsomes ◽  
Subcellular Fractionation ◽  
Fatty Acyl ◽  
Peroxisome Proliferator ◽  
Rat Liver Microsomes ◽  
Triton X ◽  
Long Chain

We have shown that a microbial cholic acid catabolite (4R)-4-(2,3,4,6,6a beta,7,8,9,9a alpha,9b beta-decahydro-6a beta-methyl-3-oxo- 1H-cyclopenta[f]quinolin-7 beta-yl)valeric acid (DCQVA), is a potent peroxisome proliferator. In this paper a possible key stage in DCQVA metabolism, the activation of DCQVA to its CoA ester, has been investigated in rat liver microsomes and particulate fractions. The microsomal reaction was dependent on CoA, ATP, DCQVA (0.2-1 mM) and protein content. The reaction was decreased by storage at 4 degrees C, preincubation of microsomes at 37 degrees C for 5 min, or inclusion of Triton X-100 in the reaction mixture. Such treatments also enhanced generation of long-chain fatty acyl-CoAs, as determined by h.p.l.c. analysis. The same effect was caused by exposing the microsomes to phospholipase A2, suggesting that endogenous fatty acids may compete with DCQVA for esterification with CoA. Subcellular fractionation of rat liver demonstrated that the activity of DCQVA-CoA synthesis was localized predominantly in the microsomal fraction, in contrast to long-chain fatty acyl-CoA synthetase, which was distributed among all particulate fractions. Administration of clofibrate of rats did not affect the distribution of DCQVA-CoA synthesis activity. In contrast to a 2-fold induction of long-chain fatty acyl-CoA synthetase by clofibrate treatment, the activity of DCQVA-CoA synthesis in the microsomal fraction decreased by 80%. These results suggest that DCQVA is activated by an enzyme distinct from long-chain fatty acyl-CoA synthetase. The resulting perturbation of fatty acid metabolism may be involved in the mechanism whereby DCQVA causes peroxisome proliferation.

A Rapidly Sedimenting Fraction of Rat Liver Endoplasmic Reticulum

1973 ◽  
Vol 13 (2) ◽  
pp. 447-459 ◽  
Author(s):  
J. A. LEWIS ◽  
J. R. TATA
Keyword(s):  
Endoplasmic Reticulum ◽  
Rat Liver ◽  
Microsomal Fraction ◽  
Close Association ◽  
Balance Sheet ◽  
Subcellular Fractionation ◽  
Electron Microscopic ◽  
Microsomal Preparation ◽  
Low Speed ◽  
Microscopic Studies

Balance-sheet experiments carried out to account for the distribution of endoplasmic reticulum fragments during subcellular fractionation of rat liver showed that a large proportion of these fragments are present in the pellets of low-speed centrifugation. Using glucose-6-phosphatase and RNA as markers we found that approximately 50% of the fragments of endoplasmic reticulum sedimented in the pellet of a 640-g centrifugation, 10% in that of a 6000-g centrifugation and 35% in the pellet of a 105000-g centrifugation. Starvation of the animals before use did not alter this distribution, nor did the use of more vigorous homogenization conditions. We have developed a procedure for removing nuclei and erythrocytes from the material sedimenting at 640g to give a fraction (rapidly sedimenting ER fraction or RS-ER) similar to the standard microsomal preparation. Centrifugation of this RS-ER fraction over 1.3 M sucrose yields subfractions of high and low RNA content analogous to the rough and smooth microsomal fractions. Electron-microscopic studies showed that, whereas the rough microsomal fraction consisted of ribosome-studded vesicles of varying size and content density, the rough RS-ER fraction contained a mixture of mitochondria and double lamellar membranes with ribosomes attached. These double lamellar membranes closely resemble the endoplasmic reticulum of intact rat liver. The double lamellar membranes are frequently observed grouped in stacks and in close association with the mitochondria. The significance of the association between endoplasmic reticulum and mitochondria of the RS-ER fraction and the relation between it and the standard microsomal preparation are discussed.

scholarly journals The Early Onset Dystonia Protein TorsinA Interacts with Kinesin Light Chain 1

2004 ◽  
Vol 279 (19) ◽  
pp. 19882-19892 ◽  
Author(s):  
Christoph Kamm ◽  
Heather Boston ◽  
Jeffrey Hewett ◽  
Jeremy Wilbur ◽  
David P. Corey ◽  
...  
Keyword(s):  
Rat Brain ◽  
Light Chain ◽  
Cortical Neurons ◽  
Early Onset ◽  
Subcellular Fractionation ◽  
Wild Type ◽  
Anterograde Transport ◽  
Glutamic Acid Residue ◽  
Carboxyl Terminal

Early onset dystonia is a movement disorder caused by loss of a glutamic acid residue (Glu302/303) in the carboxyl-terminal portion of the AAA+protein, torsinA. We identified the light chain subunit (KLC1) of kinesin-I as an interacting partner for torsinA, with binding occurring between the tetratricopeptide repeat domain of KLC1 and the carboxyl-terminal region of torsinA. Coimmunoprecipitation analysis demonstrated that wild-type torsinA and kinesin-I form a complexin vivo. In cultured cortical neurons, both proteins co-localized along processes with enrichment at growth cones. Wild-type torsinA expressed in CAD cells co-localized with endogenous KLC1 at the distal end of processes, whereas mutant torsinA remained confined to the cell body. Subcellular fractionation of adult rat brain revealed torsinA and KLC associated with cofractionating membranes, and both proteins were co-immunoprecipitated after cross-linking cytoplasmically oriented proteins on isolated rat brain membranes. These studies suggest that wild-type torsinA undergoes anterograde transport along microtubules mediated by kinesin and may act as a molecular chaperone regulating kinesin activity and/or cargo binding.

Saturable binding of [3H]phenytoin to rat brain membrane fraction

1981 ◽  
Vol 59 (4) ◽  
pp. 402-407 ◽  
Author(s):  
W. M. Burnham ◽  
L. Spero ◽  
M. M. Okazaki ◽  
B. K. Madras
Keyword(s):  
Rat Brain ◽  
Binding Sites ◽  
Membrane Fraction ◽  
Proteolytic Enzymes ◽  
Subcellular Fractionation ◽  
Brain Membrane ◽  
High Affinity ◽  
Number Of Binding Sites ◽  
Very High ◽  
Maximal Capacity

Preliminary studies indicate that [3H]phenytoin binds in a saturable and reversible fashion to at least two distinct sites in the membrane fraction of whole rat brain. One of these displays a high affinity (Kd = 6 nM) and a low maximal capacity (Bmax = 10 pmol/g protein). The other has a low affinity (Kd = 4.8 μM) and is estimated to have a very high maximal capacity. Phenytoin binding is reduced if the membrane fraction is preincubated with proteolytic enzymes and subcellular fractionation studies indicate that the P2 fraction has the largest number of binding sites. Competition experiments fail to reveal significant binding interactions with putative neurotransmitters or with other drugs except the hydantoins and anticonvulsant barbiturates. Although it is premature to speculate on the clinical significance of these findings, it is encouraging to note that the low affinity site has a Kd very similar to the therapeutic levels of phenytoin found in cerebrospinal fluid and that there seems to be some relationship between binding potency and anticonvulsant potency within the hydantoin series.

Depolymerization of polyadenylic acid by subcellular fractions of rat brain and liver

1969 ◽  
Vol 47 (3) ◽  
pp. 283-289 ◽  
Author(s):  
M. R. V. Murthy ◽  
A. D. Bharucha ◽  
C. Raynaud-Jammet
Keyword(s):  
Rat Brain ◽  
Microsomal Fraction ◽  
Soluble Fraction ◽  
Total Activity ◽  
Relative Increase ◽  
Nuclear Fraction ◽  
Polyadenylic Acid ◽  
Rat Brain And Liver ◽  
Liver Homogenates ◽  
Maximum Increment

Rat brain and liver homogenates depolymerized polyadenylic acid when added to a reaction mixture containing this polynucleotide. The activity in the homogenate declined progressively with the age of the tissues. This was reflected in a parallel reduction in the activity of the soluble fraction. In brain, the activity in the nuclear fraction also declined in the adult to half the level of the newborn. In contrast, liver nuclei had approximately the same activity at all stages of growth.With advancement in age, an increasingly greater proportion of the total activity of the tissues was contained in the nuclear fraction, while at the same time the proportion of activity in the soluble fraction decreased. The proportion of activity contained in the mitochondrial–microsomal fraction also increased with growth in brain, with the maximum increment in activity occurring after 8 weeks of age. In liver, there was actually a decrease of activity in this fraction during the same period. At all ages, the mitochondrial–microsomal fraction of brain contained a higher proportion of activity and the nuclear fraction of brain contained a lower proportion of activity compared to corresponding fractions of liver. The presence of polyadenylic acid degrading activity in these fractions and its relative increase with age may indicate a changing emphasis in the pattern of RNA metabolism during growth; for example, a higher rate of RNA synthesis in the young and a higher rate of RNA turnover in the adult.When the soluble fraction of rat brain was dialyzed, the polyadenylic acid degrading activity of this fraction was stimulated by the addition of inorganic orthophosphate. Brain and liver homogenates also mediated an ADP – inorganic phosphate exchange reaction which was highest in the newborn and decreased rapidly with age. These observations indicate that at least a part of the polyadenylic acid degrading activity in brain and liver extracts may be due to phosphorolytic action.

scholarly journals ADENOSINE TRIPHOSPHATASE ACTIVITY IN RAT BRAIN FOLLOWING DIFFERENTIAL FIXATION WITH FORMALDEHYDE, GLUTARALDEHYDE, AND HYDROXYADIPALDEHYDE

1965 ◽  
Vol 13 (3) ◽  
pp. 191-205 ◽  
Author(s):  
RICHARD M. TORACK
Keyword(s):  
Electron Microscopy ◽  
Enzymatic Activity ◽  
Rat Brain ◽  
Adenosine Triphosphatase ◽  
Physiological Activity ◽  
Formalin Fixation ◽  
Adenosine Triphosphatase Activity ◽  
Chemical Inhibition ◽  
Rat Cerebrum ◽  
The Brain

Differential fixation of rat brain has been described using formaldehyde, glutaraldehyde and hydroxyadipaldehyde by perfusion or prolonged immersion. Fixation by prolonged immersion appears preferable since it produces a similar result with greater simplicity. Distribution of reaction product resulting from adenosine triphosphate hydrolysis in these fixed brains appears different and characteristic for each of these fixatives when they are used in this manner. More adenosine triphosphatase activity in rat brain was observed following formalin fixation than after fixation with either glutaraldehyde on hydroxyadipaldehyde; in this respect formalin fixation is recommended for over-all study of adenosine triphosphatase activity in the brain. The use of glutaraldehyde and hydroxyadipaldehyde seems indicated when study of enzymes surviving these fixatives is desired. Beside varying inactivation by different aldehyde fixatives, adenosine triphosphatase activity of rat brain has been characterized by distinct substrate preference and by chemical inhibition. Specificity of adenosine triphosphatase localization in rat cerebrum by electron microscopy seems enhanced by such differential fixation since differences in enzymatic activity not previously apparent can he recognized in closely related structures such as components of the blood-brain barrier. Since adenosine triphosphatase activity of astroglia in corpus callosum as well as in subpial and subependymal glial networks is glutaraldehyde resistant, an enzymatic similarity perhaps related to their physiological activity is indicated in these cells. Astroglia of cortex evince enzyme activity that survives only formalin fixation, suggesting a different function for cortical astrocytes. The enzymatic activity of oligodendrocytes appears to be a diphosphatase inactivated by glutaraldehyde and hydroxyadipaldehyde, and in this is strikingly similar to diphosphatase of Golgi apparatus.

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