Rates of efflux and intracellular concentrations of potassium, sodium and chloride ions in isolated fat-cells from the rat

1. The metabolism of K+, Na+ and Cl− has been investigated in isolated fat-cells prepared from the epididymal adipose tissue of rats. 2. Methods are described for measuring the intracellular water space, the rates of loss of intracellular 42K+, 22Na+ and 36Cl− and the intracellular concentrations of K+, Na+ and Cl− in isolated fat-cells. 3. The intracellular water space, measured as the [3H]water space minus the [carboxylic acid−14C]inulin space, was 3·93±0·38μl./100mg. cell dry wt. 4. The first-order rate constants for radioisotope effluxes from isolated fat-cells were 0·029min.−1 for 42K+, 0·245min.−1 for 22Na+ and 0·158min.−1 for 36Cl−. 5. The intracellular concentrations of K+, Na+ and Cl− were 146m-equiv./l., 18·6±2·9m-equiv./l. and 43±2·4m-equiv./l. respectively. 6. The total intracellular K+ content of isolated fat-cells was determined by atomic-absorption spectrophotometry to confirm the value obtained from the radioisotope-efflux data. 7. The ion effluxes from isolated fat-cells were: K+, 1·5pmoles/cm.2/sec., Na+, 1·6pmoles/cm.2/sec., and Cl−, 2·4pmoles/cm.2/sec. 8. The membrane potential of isolated fat-cells calculated from the Cl− distribution ratio was −28·7mv.

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Distribution of nonelectrolytes in muscle

American Journal of Physiology-Legacy Content ◽

10.1152/ajplegacy.1961.200.4.651 ◽

1961 ◽

Vol 200 (4) ◽

pp. 651-655 ◽

Cited By ~ 22

Author(s):

Emil Bozler

Keyword(s):

Small Molecules ◽

Muscle Fibers ◽

Intracellular Water ◽

Stomach Muscle ◽

First Order ◽

First Order Kinetics ◽

Inulin Space ◽

Muscle Water ◽

Osmotic Effects ◽

Water Space

Uptake and rate of efflux of some nonelectrolytes was studied in stomach muscle and sartorius of the frog. After complete equilibration the concentration in muscle water approximated that of the medium for glycerol, but was higher for urea and thiourea. Erythritol space averaged 50%. d-Arabinose and sucrose distributed themselves in a smaller volume, which, however, was always greater than that of inulin. Efflux of sugars did not follow first-order kinetics and became very slow after a few hours. This was true also for muscles washed in formalin. In glycerol-extracted muscle fibers, sucrose space was nearly as large as water space, but inulin space was significantly lower. It is concluded that sugars and some other nonelectrolytes penetrate into the fibers but are excluded from part of the intracellular water and, therefore, produce osmotic effects. The results support the hypothesis that most of the fiber water is contained in segregated or narrow spaces, hindering diffusion of all except very small molecules.

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Factors affecting the permeability of isolated fat-cells from the rat to [42K]potassium and [36Cl]chloride ions

Biochemical Journal ◽

10.1042/bj1170615 ◽

1970 ◽

Vol 117 (3) ◽

pp. 615-621 ◽

Cited By ~ 25

Author(s):

M. C. Perry ◽

C. N. Hales

Keyword(s):

Initial Phase ◽

Chloride Ions ◽

Fat Cells ◽

Adrenocorticotrophic Hormone ◽

Isolated Fat Cells ◽

Free Medium ◽

Fat Cell ◽

Factors Affecting ◽

Exchange Diffusion ◽

High K

1. The effluxes of 42K+ and 36Cl− from isolated fat-cells from the rat were studied under a variety of conditions known to affect the metabolism of the cells. 2. 42K+ efflux from isolated fat cells was increased in a Na+-free–high-K+ medium and decreased in a K+-free medium. The existence of K+ exchange diffusion across the fat-cell membrane is suggested. 3. 36Cl− efflux from isolated fat-cells was decreased when the Cl− component of the wash medium was replaced by acetate. The basal 36Cl− efflux is suggested to be partly by Cl− exchange diffusion and partly in company with a univalent cation. 4. A variety of lipolytic stimuli, adrenaline, adrenocorticotrophic hormone, N-6,O-2′-dibutyryladenosine cyclic 3′:5′-monophosphate and theophylline, increased 42K+ efflux from isolated fat-cells. The adrenaline stimulation was biphasic; an initial, rapid and transient increase in 42K+ loss from the fat-cells was followed by a slower, more prolonged, increase in 42K+ efflux. The initial phase was inhibited by phentolamine but not by propranolol. 5. Insulin increased 42K+ efflux only after preincubation with the cells.

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Effect of nutrition on subcellular localization of rat fat-cell lipoprotein lipase

Biochemical Journal ◽

10.1042/bj1720239 ◽

1978 ◽

Vol 172 (2) ◽

pp. 239-245 ◽

Cited By ~ 17

Author(s):

A Vanhove ◽

C Wolf ◽

M Breton ◽

M C Glangeaud

Keyword(s):

Adipose Tissue ◽

Lipoprotein Lipase ◽

Plasma Membranes ◽

Total Activity ◽

Normal Diet ◽

Epididymal Adipose Tissue ◽

Fat Cells ◽

Isolated Fat Cells ◽

Fat Cell ◽

Intact Tissue

This study supports the possibility for multiple subcellular forms of lipoprotein lipase. 1. The total activity of lipoprotein lipase per g of intact epididymal adipose tissue from fed rats is much higher than that from starved rats. 2. The isolated fat-cells of fed and of starved rats have lipoprotein lipase of almost the same activity per g of fat-pads. The isolated fat-cells of starved rats have a much higher proportion of total activity per g of the intact tissue than do those of fed rats. 3. Under the conditions of homogenization used, only a small proportion of the total activity per g of intact tissue from fed rats was associated with the fat layer which floated to the top of the homogenate during low-speed centrifugation. The different proportions of the specific enzyme activity found in each subcellular fraction are described. 4. Lipoprotein lipase from plasma membranes and microsomal fractions from starved and fed rats was purified by affinity chromatography. 5. The total activity of microsomal lipoprotein lipase per g of intact adipose tissue is enhanced by a normal diet. 6. In intact epididymal adipose tissue from fed rats, the activity per g of tissue of lipoprotein lipase of plasma membranes is much higher than that in the same fraction from starved rats. By contrast, the activities per g of tissue in plasma membranes obtained from starved or from fed rats by collagenase treatment were similar.

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STUDIES IN CONGENITAL GENERALIZED LIPODYSTROPHY

Acta Endocrinologica ◽

10.1530/acta.0.0790720 ◽

1975 ◽

Vol 79 (4) ◽

pp. 720-728 ◽

Cited By ~ 4

Author(s):

Oddmund Søvik ◽

Svein Oseid

Keyword(s):

Plasma Insulin ◽

Dilution Effect ◽

Insulin Level ◽

Tolerance Test ◽

High Plasma ◽

Epididymal Adipose Tissue ◽

List Type ◽

Fat Cells ◽

Isolated Fat Cells ◽

Congenital Generalized Lipodystrophy

ABSTRACT Suppressible and non-suppressible insulin-like activities (ILA) of plasma from 3 patients with congenital generalized lipodystrophy have been studied, employing isolated fat cells from rat epididymal adipose tissue. In all 3 patients the fasting ILA was markedly increased compared with the normal controls. In one of the patients (I. T.) total ILA rose to about 800 μU per ml during an iv glucose tolerance test. The observed total ILA was in all cases (controls included) equal to or slightly higher than the previously determined immunoreactive plasma insulin (IRI). The exact determination of ILA was, however, hampered by a dilution effect, which was present even at high plasma dilutions. In 2 of the patients addition of insulin antiserum inhibited plasma ILA by about 50 %. In the third patient (I. T.), who exhibited the highest insulin level, at least 85 % of the activity was suppressed by insulin antibodies. The levels of non-suppressible ILA were higher than in the controls in terms of μU per ml, but lower than in the controls when related to total ILA. These findings strongly support our previous conclusion that the elevated plasma insulin seen in congenital generalized lipodystrophy is mainly due to true pancreatic insulin. Since the effect of plasma insulin on isolated fat cells were freely expressed, i. e. suppressible ILA was equal to or slightly lower than IRI, the presence of a circulating insulin antagonist in this disease may be excluded.

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Clearing-factor lipase in adipose tissue. Distinction of different states of the enzyme and the possible role of the fat cell in the maintenance of tissue activity

Biochemical Journal ◽

10.1042/bj1120203 ◽

1969 ◽

Vol 112 (2) ◽

pp. 203-209 ◽

Cited By ~ 52

Author(s):

V. J. Cunningham ◽

D S Robinson

Keyword(s):

Adipose Tissue ◽

Lipase Activity ◽

Total Activity ◽

Epididymal Adipose Tissue ◽

Fat Cells ◽

Isolated Fat Cells ◽

Fat Cell ◽

Prolonged Incubation ◽

Intact Tissue

1. Incubation of intact epididymal adipose tissue from fed rats at 37° in an albumin solution at pH7·4 in vitro results in rapid loss of clearing-factor lipase activity until a low activity, stable to prolonged incubation, is attained. The clearing-factor lipase activity of intact tissue from starved rats, which is initially much less than that of tissue from fed rats, is mainly stable to incubation at 37°. 2. Much of the clearing-factor lipase activity of intact epididymal adipose tissue from fed rats is inactivated by collagenase. The enzyme activity of intact tissue from starved rats is not inactivated by collagenase. 3. The clearing-factor lipase activity of fat cells isolated from the epididymal adipose tissue of fed rats is stable to prolonged incubation at 37°. It represents only a small proportion of the total activity of the intact tissue. In starved rats, the isolated fat cells contain a much higher proportion of the activity of the intact tissue. Their activity is also stable at 37°. 4. Incubation of isolated fat cells in a serum-based medium leads to a progressive rise in clearing-factor lipase activity. Actinomycin increases the extent of this rise in activity. No rise in clearing-factor lipase activity occurs when stromal-vascular cells isolated from epididymal adipose tissue are incubated in the medium. 5. The findings indicate that less than 20% of the activity of intact adipose tissue from fed rats is retained when fat cells are isolated from the tissue by collagenase treatment. The activity that is lost could be that which normally functions in the uptake of triglyceride fatty acids by the tissue.

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The clearing-factor lipase activity of isolated fat-cells

Biochemical Journal ◽

10.1042/bj1460481 ◽

1975 ◽

Vol 146 (2) ◽

pp. 481-488 ◽

Cited By ~ 34

Author(s):

A Cryer ◽

P Davies ◽

E R Williams ◽

D S Robinson

Keyword(s):

Enzyme Activity ◽

Adipose Tissue ◽

Lipase Activity ◽

Incubation Medium ◽

Cell Activity ◽

Molecular Forms ◽

Epididymal Adipose Tissue ◽

Fat Cells ◽

Isolated Fat Cells ◽

Fat Cell

1. When fat-cells are isolated from the epididymal adipose tissue of 24h-starved rats and incubated at 25 degrees C in the presence of dialysed serum, glucose, insulin, amino acids and heparin, the total clearing-factor lipase acitivity of the incubation system increases progressively over a period of several hours. 2. All of the increase in activity is accounted for by the appearance of enzyme in the appearance of enzyme in the incubation medium and the fat-cell activity does not change significantly. Cycloheximids, at a concentration that prevents protein synthesis, does not affect the appearance of enzyme in the incubation medium, but the fat-cell enzyme activity is decreased in its presence. 3. The magnitude of the increase in total clearing factor lipase activity is unaffected by the omission of heparin from the medium. However, less enzyme is extracted in tis absence and the fat-cell activity increases. Cycloheximide again only affects the rise in cell activity and does not alter the activity in the incubation medium. 4. When serum in the incubation medium is replaced by casein, the distribution of enzyme between the cells and the medium is changed, but the magnitudes of the increases in total enzyme activity are similar. 5. These characteristics of the clearing-factor lipase response of isolated fat-cells differ in several respects from those observed earlier with intact adipose tissue from 24h-starved rats (Robinson & Wing, 1971; Cryer et al., 1973). The differences could be due, in part, to changes in the relative amounts of two different molecular forms of the enzyme that occur during the isolation of the fat-cells.

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