scholarly journals The effect of insulin on plasma-membrane and mitochondrial-membrane potentials in isolated fat-cells

1981 ◽  
Vol 196 (1) ◽  
pp. 133-147 ◽  
Author(s):  
R J Davis ◽  
M D Brand ◽  
B R Martin
Keyword(s):  
Plasma Membrane ◽  
Mitochondrial Membrane ◽  
Weak Acid ◽  
Membrane Potentials ◽  
Ph Gradient ◽  
Protonmotive Force ◽  
Fat Cells ◽  
Fat Cell ◽  
Intact Cells ◽  
Insulin Effect

1. A recently developed technique for the measurement of plasma-membrane and mitochondrial-membrane potentials in intact cells by using the distribution of 86Rb+ and [3H]methyltriphenylphosphonium+ has enabled us to characterize a novel insulin effect on fat-cell mitochondria. For control cells the plasma-membrane and mitochondrial-membrane potentials were 75 mV and 152 mV respectively. Insulin (10 mu units/ml) caused a 9 mV hyperpolarization of the plasma membrane and a 19 mV depolarization of the mitochondrial membrane. 2. The insulin-dependent mitochondrial depolarization was observed at physiological insulin concentrations (10 mu units/ml) and was apparent when the cells metabolized a wide variety of substrates. 3. Evidence from the uptake of the weak acid 5,5-dimethyloxazolidine-2,4-dione by fat-cells was interpreted as indicating that the mitochondrial pH gradient was increased by insulin. 4. Insulin alters the balance between the electrical and pH-gradient components that form the mitochondrial protonmotive force. A model is proposed.

scholarly journals The properties and extracellular location of 5′-nucleotidase of the rat fat-cell plasma membrane

1975 ◽  
Vol 146 (3) ◽  
pp. 625-633 ◽  
Author(s):  
A C Newby ◽  
J P Luzio ◽  
C N Hales
Keyword(s):  
Plasma Membrane ◽  
Fat Cells ◽  
Isolated Fat Cells ◽  
Fat Cell ◽  
Cell Plasma Membrane ◽  
Intact Cells ◽  
Low Concentrations ◽  
Cell Plasma ◽  
Atp Analogues ◽  
Extracellular Location

1. A phosphohydrolase specific for 5′-nucleotides was characterized by using a particulate fraction from isolated fat-cells. 2. The activity of intact cells towards 5′-AMP was studied. 3. The activity in either situation had the same KM for AMP (45 muM) and was inhibited by low concentrations of ATP (less than 50 muM), but less potently by the ATP analogues AMP-P(CH2)P(adenylyl (β γ-methylene)diphosphonate) and AMP-P)NH)P (adenylylimidodiphosphate). 4. Homogenization of intact fat-cells caused no increase in activity and at least 85% of the activity was recovered in the particulate preparation. 5. The preparation of fat-cells used in this work was not freely permeable to AMP. 6. The ability of intact fat-cells to hydrolyse AMP implies that 5′-nucleotidase is an ectoenzyme in fat-cells. 7. Concentrations of ATP 100 times lower than intracellular concentrations inhibit the enzyme when added extracellularly to intact fat-cells, implying that this effect is also medicated at the extracellular face of the membrane. 8. Antibodies raised to whole liver cells and whole fat-cells inhibit 5′-nucleotidase in intact cells. 9. Incubation of intact fat-cells with adrenaline (1 mug/ml) or insulin (50 mui.u./ml) failed to alter the KM or Vmax. of the enzyme.

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.

Proton compartmentation in rat renal cortical tubules

1989 ◽  
Vol 256 (6) ◽  
pp. F986-F993 ◽  
Author(s):  
A. C. Schoolwerth ◽  
F. A. Gesek ◽  
R. M. Culpepper
Keyword(s):  
Cell Membrane ◽  
Mitochondrial Membrane ◽  
Weak Acid ◽  
Ph Gradient ◽  
Inner Mitochondrial Membrane ◽  
Mitochondrial Content ◽  
Medium Ph ◽  
Intracellular Compartments ◽  
Alpha Ketoglutarate Dehydrogenase ◽  
The Relationship

To study the control of renal ammoniagenesis, a technique was developed to estimate simultaneously intracellular (pHi) and intramitochondrial (pHm) pH in suspensions of rat renal cortical tubules. pHi was estimated with the fluorescent probe 2',7'biscarboxyethyl-5(6)-carboxy-fluorescein (BCECF). The intracellular distribution of the weak acid 5,5-dimethyloxazolidine-2,4-dione (DMO) allowed calculation of pHm with the use of values of pHi obtained with BCECF and tubule mitochondrial content. At medium pH (pHe) 7.4, pHi was 7.08 +/- 0.02. Over the pHe range 7.0-7.7, pHi was linearly related to pHe, but the pH gradient across the cell membrane decreased as pHe was lowered. No difference in the relationship between pHe and pHi was obtained when tubules were incubated in the presence of a nonbicarbonate or bicarbonate-buffered medium. Changes in pHe with bicarbonate-buffered media resulted in identical pHi values, whether the changes were induced by altered bicarbonate or CO2 content. At pHe 7.4, pHm was 7.78 +/- 0.6 in bicarbonate-buffered medium but was higher (0.2-0.3 pH units) when tubules were bathed in nonbicarbonate-buffered medium. pHm was linearly related to pHi in either buffer. The pH gradient across the inner mitochondrial membrane was also positively correlated with pHe. The present studies indicate the suitability of the techniques for estimating pHi and pHm simultaneously in suspensions of rat renal cortical tubules. Parallel changes occur in both intracellular compartments when pHe is altered. pHm, which is approximately 0.7 pH units greater than pHi, decreases in acute acidosis. This decrease may be important in stimulating renal ammoniagenesis, possibly by activation of alpha-ketoglutarate dehydrogenase.

scholarly journals Energy transduction in intact synaptosomes. Influence of plasma-membrane depolarization on the respiration and membrane potential of internal mitochondria determined in situ

1980 ◽  
Vol 186 (1) ◽  
pp. 21-33 ◽  
Author(s):  
I D Scott ◽  
D G Nicholls
Keyword(s):  
Plasma Membrane ◽  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Significant Proportion ◽  
Membrane Potentials ◽  
Simultaneous Estimation ◽  
Anaerobic Glycolysis ◽  
Mitochondrial Potential ◽  
Plasma Membrane Potential

A method is described, based on the differential accumulation of Rb+ and methyltriphenylphosphonium, for the simultaneous estimation of the membrane potentials across the plasma membrane of isolated nerve endings (synaptosomes), and across the inner membrane of mitochondria within the synaptosomal cytoplasm. These determinations, together with measurements of respiratory rates, and ATP and phosphocreatine concentrations, are used to define the bioenergetic behaviour of isolated synaptosomes under a variety of conditions. Under control conditions, in the presence of glucose, the plasma and mitochondrial membrane potentials are respectively 45 and 148mV. Addition of a proton translocator induces a 5-fold increase in respiration, and abolishes the mitochondrial membrane potential. The addition of rotenone to inhibit respiration does not affect the plasma membrane potential, and only lowers the mitochondrial membrane potential to 128mV. Evidence is presented that ATP synthesis by anaerobic glycolysis is sufficient under these conditions to maintain ATP-dependent processes, including the reversal of the mitochondrial ATP synthetase. Addition of oligomycin under non-respiring conditions leads to a complete collapse of the mitochondrial potential. Even under control conditions the plasma membrane (Na+ + K+)-dependent ATPase is responsible for a significant proportion of the synaptosomal ATP turnover. Veratridine greatly increases respiration, and depolarizes the plasma membrane, but only slightly lowers the mitochondrial membrane potential. High K+ and ouabain also lower the plasma membrane potential without decreasing the mitochondrial membrane potential. In non-respiring synaptosomes, anaerobic glycolysis is incapable of maintaining cytosolic ATP during the increased turnover induced by veratridine, and the mitochondrial membrane potential collapses. It is concluded that the internal mitochondria must be considered in any study of synaptosomal transport.

Anti-Trypanosoma cruzi activity of costic acid isolated from Nectandra barbellata (Lauraceae) is associated with alterations in plasma membrane electric and mitochondrial membrane potentials

2020 ◽  
Vol 95 ◽  
pp. 103510 ◽  
Author(s):  
Vinicius S. Londero ◽  
Thais A. Costa-Silva ◽  
Andre G. Tempone ◽  
Gislene M. Namiyama ◽  
Fernanda Thevenard ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Trypanosoma Cruzi ◽  
Mitochondrial Membrane ◽  
Membrane Potentials

Water content of rat adipose tissue and isolated adipocytes in relation to cell size

1976 ◽  
Vol 231 (5) ◽  
pp. 1568-1572 ◽  
Author(s):  
M DiGirolamo ◽  
JL Owens
Keyword(s):  
Adipose Tissue ◽  
Water Content ◽  
Cell Volume ◽  
Cell Size ◽  
Intracellular Water ◽  
Male Rats ◽  
Fat Cells ◽  
Fat Cell ◽  
Tissue Water ◽  
Adipose Mass

Epididymal adipose tissue composition and adipocyte water content were studied in male rats during growth and development of spontaneous obesity. The data show that a highly significant positive correlation exists between fat-cell volume and intracellular water space (IWS) (r=.967, P less than .001). Intracellular water, expressed as picoliters per fat cell, varied from 1.5-2 in small fat cells (mean vol, 30-50 pl) to 9-10 in large cells (800-1,000 pl). When expressed as percent of fat-cell volume, IWS varied from 5-7% in the small fat cells to 1-1.3% in the large ones. Total adipose tissue water continued to increase with increasing adipose mass. Similarly, total adipocyte water increased with enlarging cell size and tissue mass. The contribution of total adipocyte water (as contrasted to that of nonadipocyte water) to total tissue water, however, was found to be limited (less than 23%) and to decline progressively with adipose mass expansion.

PROTEIN KINASE ACTIVITY ASSOCIATED WITH THE FAT CELL PLASMA MEMBRANE

1981 ◽  
Vol 9 (2) ◽  
pp. 232P-232P
Author(s):  
G. J. Belsham ◽  
R. W. Brownsey ◽  
R. M. Denton
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase ◽  
Kinase Activity ◽  
Protein Kinase Activity ◽  
Fat Cell ◽  
Cell Plasma Membrane ◽  
Cell Plasma
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