Direct Phosphorylation Effects of Epinephrine on the Plasma Membranes of Intact Rat Fat Cells

1981 ◽  
Vol 2 (1) ◽  
pp. 55-66 ◽  
Author(s):  
Ellen S. Kang ◽  
Ronald E. Gates
Keyword(s):  
Plasma Membranes ◽  
Fat Cells ◽  
Rat Fat Cells

scholarly journals A rapid immunological procedure for the isolation of hormonally sensitive rat fat-cell plasma membrane

1976 ◽  
Vol 154 (1) ◽  
pp. 11-21 ◽  
Author(s):  
J P Luzio ◽  
A C Newby ◽  
C N Hales
Keyword(s):  
Plasma Membrane ◽  
Adenylate Cyclase ◽  
Plasma Membranes ◽  
Membrane Preparation ◽  
Fat Cells ◽  
Fat Cell ◽  
Cell Plasma ◽  
Rat Fat Cells ◽  
Plasma Membrane Preparation ◽  
Isolated Rat

1. A rapid method for the isolation of hormonally sensitive rat fat-cell plasma membranes was developed by using immunological techniques. 2. Rabbit anti-(rat erythrocyte) sera were raised and shown to cross-react with isolated rat fat-cells. 3. Isolated rat fat-cells were coated with rabbit anti-(rat erythrocyte) antibodies, homogenized and the homogenate made to react with an immunoadsorbent prepared by covalently coupling donkey anti-(rabbit globulin) antibodies to aminocellulose. Uptake of plasma membrane on to the immunoadsorbent was monitored by assaying the enzymes adenylate cyclase and 5′-nucleotidase and an immunological marker consisting of a 125I-labelled anti-(immunoglobulin G)-anti-cell antibody complex bound to the cells before fractionation. Contamination of the plasma-membrane preparation by other subcellular fractions was also investigated. 4. By using this technique, a method was developed allowing 25-40% recovery of plasma membrane from fat-cell homogenates within 30 min of homogenization. 5. Adenylate cyclase in the isolated plasma-membrane preparation was stimulated by 5 μm-adrenaline.

scholarly journals Coypu insulin. Primary structure, conformation and biological properties of a hystricomorph rodent insulin

1986 ◽  
Vol 238 (2) ◽  
pp. 345-351 ◽  
Author(s):  
M Bajaj ◽  
T L Blundell ◽  
R Horuk ◽  
J E Pitts ◽  
S P Wood ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Model Building ◽  
Spatial Arrangement ◽  
Biological Properties ◽  
Bovine Insulin ◽  
Fat Cells ◽  
C Terminus ◽  
Biological Potency ◽  
A Chain ◽  
Rat Fat Cells

Insulin from a hystricomorph rodent, coypu (Myocaster coypus), was isolated and purified to near homogeneity. Like the other insulins that have been characterized in this Suborder of Rodentia, coypu insulin also exhibits a very low (3%) biological potency, relative to pig insulin, on lipogenesis in isolated rat fat-cells. The receptor-binding affinity is significantly higher (5-8%) in rat fat-cells, in rat liver plasma membranes and in pig liver cells, indicating that the efficacy of coypu insulin on receptors is about 2-fold lower than that of pig insulin. The primary structures of the oxidized A- and B-chains were determined, and our sequence analysis confirms a previous report [Smith (1972) Diabetes 21, Suppl. 2, 457-460] that the C-terminus of the A-chain is extended by a single residue (i.e. aspartate-A22), in contrast with most other insulin sequences, which terminate at residue A21. In spite of a large number of amino acid substitutions (relative to mammalian insulins), computer-graphics model-building studies suggest a similar spatial arrangement for coypu insulin to that for pig insulin. The substitution of the zinc-co-ordinating site (B10-His----Gln) along with various substitutions on the intermolecular surfaces involved in the formation of higher aggregates are consistent with the observation that this insulin is predominantly ‘monomeric’ in nature. The c.d. spectrum of coypu insulin is relatively similar to those of casiragua insulin and of bovine insulin at low concentration.

Growth hormone regulates cytosolic free calcium in rat fat cells by maintaining L-type calcium channels

1996 ◽  
Vol 270 (5) ◽  
pp. C1478-C1484 ◽  
Author(s):  
S. Gaur ◽  
H. Yamaguchi ◽  
H. M. Goodman
Keyword(s):  
Growth Hormone ◽  
Steady State ◽  
Plasma Membranes ◽  
Free Calcium ◽  
Fat Cells ◽  
Cytosolic Free Calcium ◽  
Fura 2 ◽  
Rat Adipocytes ◽  
Rat Fat Cells ◽  
Ca2 Channels

In freshly isolated individual rat adipocytes, cytosolic free Ca2+ concentration ([Ca2+]i) as measured with fura 2 slowly declined during incubation but was sustained, or even somewhat increased, by brief treatment with growth hormone (GH) at the beginning of a 3-h incubation period. GH-treated adipocytes were more permeable to Ca2+ than GH-deprived cells as indicated, using Mn2- as a surrogate and monitoring influx by the rate of quenching of fura 2 fluorescence. Blockage of Ca2- channels with 100 nM nimodipine lowered [Ca2+]i in GH-treated cells to the level seen in GH-deprived cells. Increases in [Ca2+]i or the rate of Mn2+ entry were twofold greater in GH-treated than in GH-deprived cells when extracellular K+ was increased to 30 mM. Similarly, the Ca2+ channel agonist BAY K 5552 or the diacylglycerol analogue 1,2-dioctanoyl-sn-glycerol increased [Ca2+]i more in GH-treated than in GH-deprived adipocytes. Ca(2+)-ATPase activity was two times higher in plasma membranes isolated from GH-treated than from GH-deprived cells. Continued synthesis of Ca(2+)-ATPase may depend on [Ca2+]i, since the effects of GH on [Ca2+]i and Ca(2+)-ATPase were blocked by a cycloheximide or verapamil. We suggest that voltage-sensitive L-type Ca2+ channels regulate steady-state [Ca2+]i in rat adipocytes and that GH maintains the number or functional integrity of these channels.

scholarly journals Use of a novel rapid preparation of fat-cell plasma membranes employing Percoll to investigate the effects of insulin and adrenaline on membrane protein phosphorylation within intact fat-cells

1980 ◽  
Vol 192 (2) ◽  
pp. 457-467 ◽  
Author(s):  
G J Belsham ◽  
R M Denton ◽  
M J A Tanner
Keyword(s):  
Plasma Membranes ◽  
Microsomal Fraction ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Sodium Dodecyl ◽  
Fat Cells ◽  
Fat Cell ◽  
Rapid Preparation ◽  
Cell Plasma ◽  
Rat Fat Cells

1. A rapid method was developed for the preparation of plasma membranes from either isolated rat fat-cells or intact epididymal fat-pads with the use of density-gradient centrifugation in the presence of Percoll. On the basis of 5′-nucleotidase activity, the yield of plasma membranes was about 50% and purification over 10-fold. Activities of marker enzymes indicated that contamination by mitochondria and microsomal fraction was small. 2. Incorporation of 32Pi into proteins associated with plasma membranes within isolated fat-cells was investigated. Four major bands of labelled phosphoproteins were separated by sodium dodecyl sulphate/polyacrylamide-slab-gel electrophoresis; the apparent subunit mol.wts. were 67 000, 61 000, 26 000 and 20 000. None of these phosphoprotein bands corresponded to periodate/Schiff-staining glycoproteins. The extent of phosphorylation of the 61 000 mol.wt phosphoprotein band was increased by about 30 and 60% after exposure of fat-cells for 15 min to insulin or adrenaline respectively.

Lipoprotein lipase secretion from isolated rat fat cells of different size

Life Sciences ◽  
1986 ◽  
Vol 39 (22) ◽  
pp. 2111-2119 ◽  
Author(s):  
Susan K. Fried ◽  
Mario DiGirolamo
Keyword(s):  
Lipoprotein Lipase ◽  
Fat Cells ◽  
Rat Fat Cells ◽  
Isolated Rat
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