Roles of cAMP and free fatty acids on the activity of the hexose monophosphate shunt

Summary Basal glucose utilization by isolated rat adipocytes have been found to be increased ten times in the presence of certain preparations of albumin. In these conditions the effects of several adrenergic agonists and related compounds on glucose oxidation, lipolysis and triacylglycerol synthesis in isolated fat cells have been studied. Oxidation of D(1-¹⁴C) glucose in rat adipocytes was almost completely inhibited by norepinephrine and isoproterenol when added to incubated fat cells. Agents able to modify intracellular AMP cyclic levels by different mechanisms display a similar ability to imitate the effect of lipolytic agents. The inhibition of glucose oxidation due to norepinephrine and isoproterenol is partially reverted by propanolol. Under the same conditions in which norepinephrine and isoproterenol markedly reduced glucose conversion to ¹⁴CO₂, they stimulated lipolysis and triacylglycerol synthesis and in this case propanolol also reverted those actions. However, in these experimental conditions, norepinephrine and isoproterenol did not raise CAMP levels 10 min after hormone addition.It is concluded from these data that glucose oxidation through hexose monophosphate shunt, activation of lipolysis and triacylglycerol synthesis in isolated rat fat cells by lipolytic agents occurs by a mechanism(s) that depend(s) on intracellular free fatty acids levels.     

Effects of a new hypoglycemic agent A-4166 on glucose metabolism in rat adipocytes and muscle tissues

The effects of the oral administration of a non-sulfonylurea hypoglycemic agent, the phenylalanine derivative A-4166, on serum insulin and glucose levels and glucose metabolism in isolated rat adipocytes and slices of muscle tissues were studied. An increase in serum insulin and a decrease in glucose levels were observed 30 minutes after A-4166 administration to rats fed basal or high fat diet. No changes in basal glucose transport in isolated fat cells were observed after the administration of A-4166. The effect of in vitro added insulin was, however, stronger in rats fed basal diet and treated with A-4166. An elevation of the membrane glucose transporter GLUT 4 was observed in rats treated with A-4166. An increase of basal lipogenesis, measured by incorporation of radiocarbon labeled glucose into lipids, was noted in adipocytes from rats fed high fat diet. The addition of insulin was followed by stimulation of lipogenesis in rats fed basal diet, however, this hormone had no effect in rats fed high fat diet. The administration of A-4166 did not affect the basal or insulin stimulated lipogenesis. Basal glucose oxidation in the diaphragm was not influenced by high fat diet or by A-4166 treatment. In the soleus muscle, basal glucose oxidation was decreased in rats fed high fat diet, and treatment with A-4166 increased the glucose oxidation up to values observed in the control basal diet fed rats. These results indicate that the administration of A-4166 can affect glucose metabolism in muscle tissue and the sensitivity of adipocytes to insulin.     

Effects of insulin and norepinephrine on glucose transport and metabolism in rat brown adipocytes. Potentiation by insulin of norepinephrine-induced glucose oxidation

Glucose is an important fuel for rat brown adipose tissue in vivo and its utilization is highly sensitive to insulin. In this study, the different glucose metabolic pathways and their regulation by insulin and norepinephrine were examined in isolated rat brown adipocytes, using [6-14C]glucose as a tracer. Glucose utilization was stimulated for insulin concentrations in the range of 40-1000 microU/ml. Furthermore, the addition of adenosine deaminase (200 mU/ml) or adenosine (10 microM) did not alter insulin sensitivity of glucose metabolism. The major effect of insulin (1 mU/ml) was a respective 7-fold and 5-fold stimulation of lipogenesis and lactate synthesis, whereas glucose oxidation remained very low. The 5-fold stimulation of total glucose metabolism by 1 mU/ml of insulin was accompanied by an 8-fold increase in glucose transport. In the presence of norepinephrine (8 microM), total glucose metabolism was increased 2-fold. This was linked to a 7-fold increase of glucose oxidation, whereas lipogenesis was greatly inhibited (by 72%). In addition, norepinephrine alone did not modify glucose transport. The addition of insulin to adipocytes incubated with norepinephrine, induced a potentiation of glucose oxidation, while lipogenesis remained very low. In conclusion, in the presence of insulin and norepinephrine glucose is a oxidative substrate for brown adipose tissue. However the quantitative importance of glucose as oxidative fuel remains to be determined.     

Phosphorylation of zymogen granule membrane proteins in intact rat pancreatic acinar cells

The comparative effects of insulin and ethanolamine on 14CO2 production and lipid synthesis from [U-14C]-D-glucose in isolated rat adipocytes were studied. Ethanolamine (10 mM) increased 14CO2 production (glucose oxidation) about 5-fold and lipogenesis about 3-fold as compared to the control. Ethanolamine was more efficient than 25 microU/ml insulin regarding both parameters, but it was less efficient than 200 microU/ml insulin in glucose oxidation, and equally potent in lipogenesis. The combination of ethanolamine and insulin was more active than insulin alone. The mechanisms of ethanolamine action include facilitation of glucose transport and increase of pyruvate dehydrogenase activity.     

Inositol glycan mimics the action of insulin on glucose utilization in rat adipocytes

Some of the actions of insulin may be mediated by the intracellular generation of an inositol phosphate glycan that modulates the activities of certain metabolic enzymes. The actions of this molecule were evaluated on glucose utilization in intact rat adipocytes. The inositol glycan led to the dose-dependent stimulation of glucose oxidation and lipogenesis. The extent of stimulation was similar to that observed for insulin. The stimulation of lipogenesis was seen only at high concentrations of glucose, suggesting regulation of processes distal to glucose uptake. The effects of the inositol glycan on intact adipocytes were specifically attenuated with inositol monophosphate in a dose dependent manner. These results further support a role for this substance as a second messenger for some of the actions of insulin, and indicate that the cellular uptake of the inositol glycan may occur by a specific transport system.     

The role of glutathione in rat adipocyte pentose phosphate cycle activity

An assay for reduced and oxidized glutathione was adapted to isolated rat epididymal adipocytes in order to correlate pentose phosphate cycle activity and glutathione metabolism. In collagenase-digested adipocytes the [GSH/GSSG] molar ratio was in excess of 100. Cells incubated for 1 hr with low glucose concentrations (0.28–0.55 mm) had higher GSH contents (3.2 μg/10⁶ cells) than in the absence of glucose (2.3 μg/10⁶ cells). The glutathione oxidant diamide caused a dose-related decrease in intracellular GSH, an increase in GSSG released into the medium, but no detectable change in the low intracellular GSSG content. The intracellular content of GSH and amount of GSSG released into the medium were therefore taken to reflect the glutathione status of the adipocytes most closely. Addition of H₂O₂ to a concentration of 60 μm to adipocytes caused to decline within 5 min in GSH content, which was less severe and more rapid to recover in the presence of 1.1 mm glucose, suggesting that the concomitant stimulation of glucose C-1 oxidation induced by the peroxide in the presence of glucose provided NADPH for regeneration of GSH. Further evidence for tight coupling between adipocyte [GSH/GSSG] ratios and pentose phosphate cycle activity was that (i) lowering intracellular GSH to 35–60% of control values by agents as diverse in action as t-butyl hydroperoxide, diamide, or the sulfhydryl blocker N-ethylmaleimide resulted in optimal stimulation of glucose C-1 oxidation and fractional pentose phosphate cycle activity, and (ii) incubating adipocytes directly with 2.5 mm GSSG resulted in a slight increase in glucose C-1 oxidation and when 0.5 mm NADP⁺ was also added a synergistic effect on pentose phosphate cycle activity was found. On the other hand, electron acceptors such as methylene blue did not lower cellular GSH content, but did stimulate the pentose phosphate cycle, confirming a site of action independent of glutathione metabolism. The results show that (i) glucose metabolism by the pentose phosphate cycle contributes to regeneration of GSH and that (ii) glutathione metabolism either directly or via coupled changes in [NADPH/NADP⁺] ratios may play a significant role in short-term control of the pentose phosphate cycle.     

Independent control of selected insulin-sensitive cell membrane and intracellular functions — the linkage of cell membrane and intracellular events controlled by insulin

Summary The effects of pH, oxidation reduction compounds and trypsin on insulin binding, hexose transport, and activation of glycogen synthase were studied utilizing rat adipocytes. In this paper the effect of pH is examined; while in the subsequent two papers the effects of glutathione and trypsin are examined. Increase in pH from 6 to 8.5 increased labelled glucose oxidation, 2-deoxyglucose transport as well as labelled insulin binding to the receptor. Enhanced insulin binding was due to an increased rate of association k₊₁ with no effect the rate of dissociation k₋₁ resulting in a decreased equilibrium dissociation constant K_D. Glycogen synthase activity was unaffected by increase in pH when adipocytes were incubated with or without glucose. Insulin in contrast to pH was effective in increasing the activity of glycogen synthase. With 2-deoxyglucose, % glycogen synthaseI activity was increased by an increase in pH. Glycogen synthase activity was thus stimulated by insulin by the direct mechanism, previously termed mechanism 1, involving the formation of a chemical mediator, and clearly distinguishable from the activation of hexose transport, previously termed mechanism 2(1). Increase in labelled glucose oxidation and in 2-deoxyglucose transport with increased pH, as well as insulin stimulation, was abolished by preincubation with trypsin, or cytochalasin B; suggesting that trypsin-sensitive and cytochalasin B-binding protein(s) presumably in the plasma membrane are involved in these effects of pH. Since increase in pH alone activates cell membrane-mediated hexose transport and insulin receptor binding under conditions where glycogen synthase is not activated, increase in pH acts presumably by a non-mediator mechanism. Insulin acts at the membrane to enhance further the effects of increased pH, via a mediator mechanism.     

Metabolic shift from lipogenesis to glycogenesis in the last instar larval fat body of the silkworm,Bombyx mori

When [¹⁴C]glucose was injected into the last instar larvae of the silkworm, Bombyx mori, the label was incorporated into various tissues at varying degrees depending on the developmental stages. Fat body exhibited high incorporation rates throughout the feeding periods. Silk glands became active in incorporation but midgut decreased toward larval maturation. The pulse labeling experiment clearly demonstrated that the metabolic shift from lipogenesis to glycogenesis occurred in fat body at the middle of the last instar; a predominant incorporation was found in lipids when [¹⁴C]glucose was injected at the early stage, while at the late stage glycogen synthesis became most active. Incorporation into fat body proteins was not a major factor throughout the instar. Extirpation of silk glands enhanced incorporation into glycogen and proteins at the late stage but did not affect lipid synthesis. Long-term chase showed that fat body lipids and proteins synthesized at the early stage were totally carried over into the pupal fat body, while much glycogen produced at the late stage was used during the larval-pupal transformation with the remainder carried over into the pupa.From these results the metabolic shift from lipogenesis to glycogenesis in fat body is discussed in relation to the storage function of the fat body for pupal metamorphosis.     

DEVELOPMENT OF LIPOGENESIS IN RAT BRAIN CORTEX: THE DIFFERENTIAL INCORPORATION OF GLUCOSE AND ACETATE INTO BRAIN LIPIDS IN VITRO

—The oxidation to CO₂ and the incorporation of [U-¹⁴C]glucose and [U-¹⁴C]acetate into lipids by cortex slices from rat brain during the postnatal period were investigated. The oxidation of [U-¹⁴C]glucose was low in 2-day-old rat brain, and increased by about two-fold during the 2nd and 3rd postnatal weeks. The oxidation of [U-¹⁴C]acetate was increased markedly in the second postnatal week, but decreased to rates observed in 2-day-old rat brain at the time of weaning. Both labeled substrates were readily incorporated into non-saponifiable lipids and fatty acids by brain slices from 2-day-old rat. Their rates of incorporation and the days on which maximum rates occurred were different, however, maximum incorporation of [U-¹⁴C]glucose and [U-¹⁴]acetate into lipid fractions being observed on about the 7th and 12th postanatal days, respectively. The metabolic compartmentation in the utilization of these substrates for lipogenesis is suggested. The activities of glucose-6-phosphate dehydrogenase, cytosolic NADP-malate dehydrogenase, cytosolic NADP-isocitrate dehydrogenase, ATP-citrate lyase and acetyl CoA carboxylase were measured in rat brain during the postnatal period. All enzymes followed somewhat different courses of development; the activity of acetyl CoA carboxylase was, however, the lowest among other key enzymes in the biosynthetic pathway, and its developmental pattern paralleled closely the fatty acid synthesis from [U-¹⁴C]glucose. It is suggested that acetyl CoA carboxylase is a rate-limiting step in the synthesis de novo of fatty acids in developing rat brain.     

Quercetin selectively inhibits insulin receptor function in vitro and the bioresponses of insulin and insulinomimetic agents in rat adipocytes.

We report here that quercetin, a naturally occurring bioflavonoid, is an effective blocker of insulin receptor tyrosine kinase-catalyzed phosphorylation of exogenous substrate. The ID50 was estimated to be 2 +/- 0.2 microM in cell-free experiments, using a partially purified insulin receptor and a random copolymer of glutamic acid and tyrosine as a substrate. Insulin-stimulated autophosphorylation of the receptor itself was not blocked by quercetin (up to 500 microM). In intact rat adipocytes, quercetin inhibited insulin-stimulating effects on glucose transport, oxidation, and its incorporation into lipids. Inhibition of lipogenesis (50%) occurred at 47 +/- 4 microM, whereas full inhibition was evident at 110 +/- 10 microM quercetin. In contrast, the effect of insulin in inhibiting lipolysis remained unaltered in quercetin-treated adipocytes. The inhibitor was devoid of general adverse cell affects. Basal activities and the ability of lipolytic agents to stimulate lipolysis were not affected. Inhibition by quercetin enabled us to evaluate which insulinomimetic agents are dependent on tyrosine phosphorylation of endogenous substrates for stimulating glucose metabolism. Quercetin blocked lipogenesis mediated by insulin, wheat germ agglutinin, and concanavalin A. The lipogenic effect of Zn2+ and Mn2+ was partially blocked, whereas that of vanadate was not affected at all.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of insulinomimetic antibodies against rat adipocyte membranes by hybridoma cells

SJL mice were injected intraperitoneally with adipocyte plasma membranes or with intrinsic membrane proteins obtained by extraction of plasma membranes with dimethylmaleic anhydride. Three days after the boost injection, the spleens were removed and fused with NS-1, a thioguanine-resistant myeloma cell line derived from P₃X63 Ag⁸ (Balb/c). Following selection for hybrids with hypoxanthine, aminopterin, and thymidine, medium of the hybrid cells was tested for its ability to bind to the plasma membrane of the adipocyte and to stimulate the oxidation of D-(1-¹⁴C) glucose to ¹⁴CO₂. Approximately 40% of the wells containing hybridomas derived from splenocytes of SJL mice immunized with plasma membranes produced immunoglobulin that bound to adipocyte plasma membranes. About 30% of these mimicked the ability of insulin to stimulate the oxidation of D-(1-¹⁴C) glucose to ¹⁴CO₂ in adipocytes. Media from 51% of the wells containing hybridomas derived from splenocytes of SJL mice immunized with intrinsic membrane proteins produced immunoglobulin that bound to the plasma membrane and 48% of those stimulated glucose oxidation. The bioactivity of the hybrid cell media could be blocked by adsorption with intrinsic membrane proteins or by the removal of immunoglobulins using formalin-fixed Staphylococcus aureus. The hybrids generated in this study can be divided into three categories: (1) hybrids that secrete antibodies that can bind to plasma membranes and mimic insulin action of glucose transport; (2) hybrids that secrete antibodies that bind to plasma membranes but do not stimulate the oxidation of D-(1-¹⁴C) glucose to ¹⁴CO₂; and (3) hybrids that produce no antimembrane antibodies. The data suggest that interaction of immunoglobulins with specific membrane proteins is essential in mimicking the action of insulin on glucose transport and oxidation in the rat adipocyte.     

Mechanism of insulin resistance in the post receptor events in sheep: 3-O-methylglucose transport in ovine adipocytes

A severe resistance to the stimulatory action of insulin on glucose metabolism has been shown in ruminant adipose tissue or isolated adipocytes as compared to that of rats. To elucidate the mechanism of insulin resistance in ruminants, we measured the stimulatory effect of insulin on 3-O-methylgulose transport and on intracellular glucose metabolism in isolated adipocytes from sheep and rats. At a glucose concentration (0.1 mM) where transport is thought to be rate-limiting for metabolism, lipogenesis from [U-14C]glucose by ovine adipocytes was markedly less than by rat adipocytes in both the basal state and at all insulin concentrations. The responsiveness to insulin assessed by percent increase above basal was reduced to about 15% of that in rat adipocytes, but the insulin sensitivity was similar, because the insulin concentration giving half-maximal stimulation, ED50, did not differ significantly between ovine and rat adipocytes. The maximal insulin-stimulated 3-O-methylglucose transport in ovine adipocytes per cell was less than 20% of that in rat adipocytes, with a significant lowering in basal rates of transport. However, when data was expressed per 3-O-methylglucose equilibrium space no significant differences were found between ovine and rat in the basal transport rates, but a lowered ability of insulin to stimulate glucose transport was still seen in ovine adipocytes. The dose-response curve for glucose transport was slightly shifted to the right in ovine adipocytes compared to rat adipocytes, indicating a small decrease in insulin sensitivity. The decrease in glucose transport was due to 60% reduction in the maximum velocity in the insulin--stimulated state, with no change in the Km.     

Changes in prostaglandin E1 stimulation of glucose oxidation in rat adipocytes during maturation and aging

Prostaglandin E₁ stimulates glucose oxidation in isolated rat adipocytes in a time and concentration dependent manner. Maximal stimulation requires 2 hours exposure to prostaglandin, although effects can be detected by 0.5 hours or earlier. In contrast to prostaglandin E₁, prostaglandin F₂α has essentially no effect on glucose oxidation. Maximal stimulation by prostaglandin E₁ at all ages tested occurs at concentrations of 10^?5 ? 10^?4M. Stimulation is greatest in cells of mature (10–12 month old) animals at 81 ± 9% above basal levels of glucose oxidation. This is to reduced to 48 ± 8% in cells of senescent (23–26 month old) animals, and at 23 ± 18% in cells of young (2–3 month old) rats is not significantly different from basal oxidation in most animals. These results are consistent with data for adipocytes and other cell types indicating that responsiveness to certain hormones is altered during maturation and aging.     

Map4k4 suppresses Srebp-1 and adipocyte lipogenesis independent of JNK signaling

Adipose tissue lipogenesis is paradoxically impaired in human obesity, promoting ectopic triglyceride (TG) deposition, lipotoxicity, and insulin resistance. We previously identified mitogen-activated protein kinase kinase kinase kinase 4 (Map4k4), a sterile 20 protein kinase reported to be upstream of c-Jun NH₂-terminal kinase (JNK) signaling, as a novel negative regulator of insulin-stimulated glucose transport in adipocytes. Using full-genome microarray analysis we uncovered a novel role for Map4k4 as a suppressor of lipid synthesis. We further report here the surprising finding that Map4k4 suppresses adipocyte lipogenesis independently of JNK. Thus, while Map4k4 silencing in adipocytes enhances the expression of lipogenic enzymes, concomitant with increased conversion of ¹⁴C-glucose and ¹⁴C-acetate into TGs and fatty acids, JNK1 and JNK2 depletion causes the opposite effects. Furthermore, high expression of Map4k4 fails to activate endogenous JNK, while Map4k4 depletion does not attenuate JNK activation by tumor necrosis factor α. Map4k4 silencing in cultured adipocytes elevates both the total protein expression and cleavage of sterol-regulated element binding protein-1 (Srebp-1) in a rapamycin-sensitive manner, consistent with Map4k4 signaling via mechanistic target of rapamycin complex 1 (mTORC1). We show Map4k4 depletion requires Srebp-1 upregulation to increase lipogenesis and further show that Map4k4 promotes AMP-protein kinase (AMPK) signaling and the phosphorylation of mTORC1 binding partner raptor (Ser792) to inhibit mTORC1. Our results indicate that Map4k4 inhibits adipose lipogenesis by suppression of Srebp-1 in an AMPK- and mTOR-dependent but JNK-independent mechanism.     

Regulation of glucose transport in isolated adipocytes by levamisole.

When isolated rat adipocytes were incubated with increasing concentrations of levamisole (0.5-5 mM), basal glucose oxidation decreased by almost 50% and insulin-stimulated glucose oxidation decreased by 90%. The decrease in glucose oxidation correlated with an inhibition of glucose transport, since levamisole at 5.0 mM decreased basal 3-O-methylglucose transport by 60% and insulin-stimulated transport by 80%. Diamide-stimulated glucose transport was also inhibited approximately 80% by 5.0 mM levamisole. Levamisole at concentrations up to 5.0 mM had no effect on phosphofructokinase activity. The present results suggest that levamisole inhibits glucose utilization by inhibiting glucose transport in a concentration-dependent manner.     

Effects of zinc supplementation on the plasma glucose level and insulin activity in genetically obese (ob/ob) mice

The effects of zinc supplementation (20 mM ZnCl₂ from the drinking water for eight weeks) on plasma glucose and insulin levels, as well as its in vitro effect on lipogenesis and lipolysis in adipocytes were studied in genetically obese (ob/ob) mice and their lean controls (+/?). Zinc supplementation reduced the fasting plasma glucose levels in both obese and lean mice by 21 and 25%, respectively (p < 0.05). Fasting plasma insulin levels were significantly decreased by 42% in obese mice after zinc treatment. In obese mice, zinc supplementation also attenuated the glycemic response by 34% after the glucose load. The insulin-like effect of zinc on lipogenesis in adipocytes was significantly increased by 80% in lean mice. However, the increment of 74% on lipogenesis in obese mice was observed only when the zinc plus insulin treatment was given. This study reveals that zinc supplementation alleviated the hyperglycemia of ob/ob mice, which may be related to its effect on the enhancement of insulin activity.     

Adenosine modulation of fat cell responsiveness to insulin and oxytocin

We have investigated the effects of adenosine on the stimulation of glucose oxidation and lipogenesis by oxytocin and insulin in rat epididymal adipocytes. The addition of adenosine deaminase (1 U/ml) to the assay medium reduced the maximal oxytocin response (glucose oxidation and lipogenesis) to between 25 and 50% of the maximum response in control cells. The maximal response to insulin was not appreciably affected under these conditions. The addition of adenosine (10 or 30 microM) increased the cell sensitivity to oxytocin by elevating the maximum rate of oxytocin-stimulated glucose metabolism. Adenosine also increased the cell sensitivity to insulin by decreasing its ED50. A change in ED50, however, was observed only when control or adenosine-treated cells were compared to adenosine deaminase-treated cells; but not when control and adenosine-treated cells were compared. On its own, adenosine also caused an appreciable increase in both glucose oxidation and lipogenesis (ED50 approximately equal to 3 microM adenosine). The difference in the effect of adenosine on oxytocin action, compared with the effect on insulin action, points to differences in the mechanisms by which insulin and oxytocin stimulate glucose metabolism in adipocytes.     

Comparison of the effects of insulin and H2O2 on adipocyte glucose transport

The abilities of insulin and the insulin mimickers spermine and H₂O₂ to stimulate 3-O-methyl glucose transport in isolated rat ft cells were stuided in an attempt to determine possible common mechanisms of action. All three agents caused a seven- to 12-fold stimulation of initial rates of glucose transport with insulin being the most effective agent. Insulin and spermine displayed similar time courses for the onset of their stimulation of transport; an initial lag before any effect was seen and then a gradual rise until the full effect was reached. The time course of H₂O₂ activation of glucose transport was different since stimulation was seen at the earliest time point tested and then gradually rose to the maximal effect. Trypsinization of cells removed insulin receptors and rendered the cells insensitive to insulin but not to spermine or H₂O₂. However, trypsinization did alter the time course of H₂O₂ action, causing an initial lag phase to appear and a general slowing of the activation kinetics. Pretreatment of cells with 2,4-dinitrophenol to lower ATP levels prevented the stimulatory effects of insulin and the mimickers. All three of the agents revealed a similar temperature dependency for stimulated glucose transport, resulting in linear Arrhenius plots with activation energies of 10.2–11.4 kcal/mole. These results show that (1) H₂O₂ does not act directly on the glucose transport system of rat adipocytes and (2) insulin and H₂O most likely act through a common energy-dependent biochemical pathway to stimulate glucose transport, but H₂O₂ enters the stimulus-response sequence distal to the initial steps in insulin action.     

Effect of Carbohydrate Overfeeding on Whole Body and Adipose Tissue Metabolism in Humans

Objective: To evaluate the effect of a 4‐day carbohydrate overfeeding on whole body net de novo lipogenesis and on markers of de novo lipogenesis in subcutaneous adipose tissue of healthy lean humans. Research Methods and Procedures: Nine healthy lean volunteers (five men and four women) were studied after 4 days of either isocaloric feeding or carbohydrate overfeeding. On each occasion, they underwent a metabolic study during which their energy expenditure and net substrate oxidation rates (indirect calorimetry), and the fractional activity of the pentose‐phosphate pathway in subcutaneous adipose tissue (subcutaneous microdialysis with 1, 6¹³C₂, 6, 6²H₂ glucose) were assessed before and after administration of glucose. Adipose tissue biopsies were obtained at the end of the experiments to monitor mRNAs of key lipogenic enzymes. Results: Carbohydrate overfeeding increased basal and postglucose energy expenditure and net carbohydrate oxidation. Whole body net de novo lipogenesis after glucose loading was markedly increased at the expense of glycogen synthesis. Carbohydrate overfeeding also increased mRNA levels for the key lipogenic enzymes sterol regulatory element‐binding protein‐1c, acetyl‐CoA carboxylase, and fatty acid synthase. The fractional activity of adipose tissue pentose‐phosphate pathway was 17% to 22% and was not altered by carbohydrate overfeeding. Discussion: Carbohydrate overfeeding markedly increased net de novo lipogenesis at the expense of glycogen synthesis. An increase in mRNAs coding for key lipogenic enzymes suggests that de novo lipogenesis occurred, at least in part, in adipose tissue. The pentose‐phosphate pathway is active in adipose tissue of healthy humans, consistent with an active role of this tissue in de novo lipogenesis.     

Development of insulin sensitivity in white adipose tissue during the suckling-weaning transition in the rat. Involvement of glucose transport and lipogenesis.

The changes of insulin responsiveness of white adipose tissue during the suckling-weaning transition in the rat were investigated in vitro on isolated adipocytes. Insulin binding, glucose transport and glucose metabolism in adipocytes from suckling rats and from rats weaned on to a high-carbohydrate (HC) or a high-fat (HF) diet were compared. Despite similar insulin binding, insulin-stimulated glucose transport rate is lower in adipocytes from suckling rats and HF-weaned rats than in adipocytes from HC-weaned rats. Moreover, whereas insulin markedly stimulates glucose metabolism in adipocytes from HC-weaned rats, glucose metabolism is totally unresponsive to insulin in adipocytes from suckling and HF-weaned rats. This insulin resistance is associated with a very low rate of lipogenesis and low activities of acetyl-CoA carboxylase, fatty acid synthase and pyruvate dehydrogenase.