Enhancement of leptin receptor signaling by SOCS3 deficiency induces development of gastric tumors in mice.

Leptin acts on its receptor (ObR) in the hypothalamus to inhibit food intake and energy expenditure. Leptin and ObR are also expressed in the gastrointestinal tract; however, the physiological significance of leptin signaling in the gut remains uncertain. Suppressor of cytokine signaling 3 (SOCS3) is a key negative feedback regulator of ObR-mediated signaling in the hypothalamus. We now show that gastrointestinal epithelial cell-specific SOCS3 conditional knockout (T3b-SOCS3 cKO) mice developed gastric tumors by enhancing leptin production and the ObRb/signal transducer and activator of transcription 3 (STAT3) signaling pathway. All T3b-SOCS3 cKO mice developed tumors in the stomach but not in the bowels by 2 months of age, even though the SOCS3 deletion occurred in both the epithelium of stomach and bowels. The tumors developed in the absence of the inflammatory response and all cKO mice died within 6 months. These tumors displayed pathology and molecular alterations, such as an increase in MUC2 (Mucin 2, oligomeric mucus/gel-forming) and TFF3 (trefoil factor 3), resembling human intestinal-type gastric tumors. Administration of antileptin antibody to T3b-SOCS3 cKO mice reduced hyperplasia of gastric mucosa, which is the step of the initiation of gastric tumor. These data suggest that SOCS3 is an antigastric tumor gene that suppresses leptin overexpression and ObRb/STAT3 hyperactivation, supporting the hypothesis that the leptin/ObRb/STAT3 axis accelerates tumorigenesis and that it may represent a new therapeutic target for the treatment of gastric cancer.

Ceramide and adenosine 5'-monophosphate-activated protein kinase are two novel regulators of 11beta-hydroxysteroid dehydrogenase type 1 expression and activity in cultured preadipocytes.

Increased activity of intracellular glucocorticoid reactivating enzyme, 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) in obese adipose tissue contributes to adipose dysfunction. As recent studies have highlighted a potential role of preadipocytes in adipose dysfunction, we tested the hypothesis that a variety of metabolic stress mediated by ceramide or AMP-activated protein kinase (AMPK) would regulate 11beta-HSD1 in preadipocytes. The present study is the first to show that 1) expression of 11beta-HSD1 in 3T3-L1 preadipocytes was robustly induced when cells were treated with cell-permeable ceramide analogue C(2) ceramide, bacterial sphingomyelinase, and sphingosine 1-phosphate, 2) 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR)-induced activation of AMPK augmented the expression and enzyme activity of 11beta-HSD1, and 3) these results were reproduced in human preadipocytes. We demonstrate for the first time that C(2) ceramide and AICAR markedly induced the expression of CCAAT/enhancer-binding protein (C/EBP) beta and its binding to 11beta-HSD1 promoter. Transient knockdown of C/EBPbeta protein by small interfering RNA markedly attenuated the expression of 11beta-HSD1 induced by C(2) ceramide or AICAR. The present study provides novel evidence that ceramide- and AMPK-mediated signaling pathways augment the expression and activity of 11beta-HSD1 in preadipocytes by way of C/EBPbeta, thereby highlighting a novel, metabolic stress-related regulation of 11beta-HSD1 in a cell-specific manner.

Role of AMP-activated protein kinase in leptin-induced fatty acid oxidation in muscle.

Leptin regulates energy homoeostasis through central and peripheral mechanisms. Initial steps in leptin action include signalling through a cytokine-like receptor which activates the JAK/STAT pathway. We investigated whether the metabolic effects of leptin in muscle could be mediated by the AMP-activated protein kinase (AMP kinase). Through studies involving leptin injection intrahypothalamically or intravenously, as well as incubation of soleus muscle or cultured muscle cells with leptin, we determined that leptin stimulates fatty acid oxidation in skeletal muscle by activating AMP kinase. Leptin exerts this effect directly at the level of muscle and also through the hypothalamic sympathetic nervous system, specifically engaging alpha-adrenergic receptors in muscle. This represents a novel and important pathway mediating leptin's metabolic actions.

Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase.

Adiponectin (Ad) is a hormone secreted by adipocytes that regulates energy homeostasis and glucose and lipid metabolism. However, the signaling pathways that mediate the metabolic effects of Ad remain poorly identified. Here we show that phosphorylation and activation of the 5'-AMP-activated protein kinase (AMPK) are stimulated with globular and full-length Ad in skeletal muscle and only with full-length Ad in the liver. In parallel with its activation of AMPK, Ad stimulates phosphorylation of acetyl coenzyme A carboxylase (ACC), fatty-acid oxidation, glucose uptake and lactate production in myocytes, phosphorylation of ACC and reduction of molecules involved in gluconeogenesis in the liver, and reduction of glucose levels in vivo. Blocking AMPK activation by dominant-negative mutant inhibits each of these effects, indicating that stimulation of glucose utilization and fatty-acid oxidation by Ad occurs through activation of AMPK. Our data may provide a novel paradigm that an adipocyte-derived antidiabetic hormone, Ad, activates AMPK, thereby directly regulating glucose metabolism and insulin sensitivity in vitro and in vivo.

ATP-sensitive K+ channels in the hypothalamus are essential for the maintenance of glucose homeostasis.

Glucose-responsive (GR) neurons in the hypothalamus are thought to be critical in glucose homeostasis, but it is not known how they function in this context. Kir6.2 is the pore-forming subunit of K(ATP) channels in many cell types, including pancreatic beta-cells and heart. Here we show the complete absence of both functional ATP-sensitive K+ (K(ATP)) channels and glucose responsiveness in the neurons of the ventromedial hypothalamus (VMH) in Kir6.2-/- mice. Although pancreatic alpha-cells were functional in Kir6.2-/-, the mice exhibited a severe defect in glucagon secretion in response to systemic hypoglycemia. In addition, they showed a complete loss of glucagon secretion, together with reduced food intake in response to neuroglycopenia. Thus, our results demonstrate that KATP channels are important in glucose sensing in VMH GR neurons, and are essential for the maintenance of glucose homeostasis.

Involvement of bradykinin and nitric oxide in leptin-mediated glucose uptake in skeletal muscle.

Regulation of glucose metabolism in peripheral tissues by leptin has been highlighted recently, although its mechanism is unclear. In this study, we postulated that bradykinin and nitric oxide (NO) are involved in the effect of leptin-mediated glucose uptake in peripheral tissues and examined these possibilities. Injection of leptin (200 pg/mouse) into the ventromedial hypothalamus-enhanced glucose uptake in skeletal muscle and brown adipose tissue, but not in white adipose tissue. Treatment with Hoe140 (0.1 mg/kg), bradykinin B2 receptor antagonist, or L-NAME (N:(G)-nitro-L-arginine methyl ester) (30 mg/kg), nitric oxide synthase inhibitor, did not influence the basal level of glucose uptake in skeletal muscle and the adipose tissue, whereas Hoe140 and L-NAME inhibited leptin-mediated glucose uptake in skeletal muscles, but had no effect in adipose tissue. However, Hoe140 and L-NAME did not inhibit insulin (1.0 U/kg)-mediated glucose uptake in all tissues examined. Taken together, these results suggest that leptin enhances bradykinin and/or the NO system, which contributes at least partially to the enhanced glucose uptake in skeletal muscles.

Aggravation of chemically-induced injury in perfused rat liver by extracellular ATP.

The effects of purinergic receptor agonists on acute liver damage and hemodynamics were studied using chemically-induced liver injury. Rat livers were perfused in situ 24 h after treatment with D-galactosamine (800 mg/kg, i.p.). In these livers, infusion of ATP (50 microM) into the portal vein caused a rapid increase in the leakage of LDH and AST from perfused liver in a dose dependent manner, accompanied with flow reduction. The similar but less effective responses were also observed by the infusion of ADP. Infusion of adenosine, a P1-receptor agonist, induced only minimal changes of liver damage and flow rate. The ATP-induced changes were almost completely suppressed by P2-receptor antagonist, suramin, but not affected by P1-receptor antagonist, 8-phenyltheophylline. Pretreatment of rats with gadolinium chloride, which depletes Kupffer cells, did not inhibit the potentiation of liver damage caused by ATP, whereas hemodynamic effects of ATP were significantly attenuated by gadolinium. These results indicate that extracellular ATP aggravates acute liver injury mediated by P2-type purinergic receptors.

Interferon-gamma induces AT(2) receptor expression in fibroblasts by Jak/STAT pathway and interferon regulatory factor-1.

The expression of angiotensin II type 2 (AT(2)) receptor is closely associated with cell growth, differentiation, and/or injury. We examined the effect of interferon (IFN)-gamma on AT(2) receptor expression in mouse fibroblast R3T3 cells and demonstrated that IFN-gamma treatment increased the expression of AT(2) receptor mRNA as well as its binding. Interferon regulatory factor (IRF)-1 was induced in mouse fibroblast R3T3 cells after IFN-gamma stimulation, and electrophoretic mobility shift assay showed an increase in IRF-1 binding with the IRF-specific binding sequence in the AT(2) receptor gene promoter region after IFN-gamma stimulation. The IRF-1 gene promoter contains an IFN-gamma-activated sequence (GAS) motif for possible binding of signal transducer(s) and activator(s) of transcription (STAT). Indeed, in R3T3 cells, IFN-gamma treatment resulted in rapid activation of Janus kinase (Jak) 1, Jak2, and STAT1 via tyrosine phosphorylation. Electrophoretic mobility shift assay with the GAS probe revealed increased STAT1 binding to the IRF-1 gene promoter in response to IFN-gamma stimulation. Transfection of GAS-binding oligonucleotides inhibited the effect of IFN-gamma on IRF-1 production, resulting in the AT(2) receptor trans-activation. Taken together, our data show that IFN-gamma upregulates AT(2) receptor expression in R3T3 cells via the activation of the intracellular Jak/STAT pathway and production of IRF-1.

Role of the sympathetic nervous system and insulin in enhancing glucose uptake in peripheral tissues after intrahypothalamic injection of leptin in rats.

Our previous study demonstrated that microinjection of leptin into the ventromedial hypothalamus (VMH) dramatically increased glucose uptake in the heart, brown adipose tissue (BAT), and skeletal muscles, but not in white adipose tissue (WAT) in conscious unrestrained rats, as assessed in vivo by the 2-[3H]deoxyglucose method. Here we examined the role of the sympathetic nervous system and insulin in enhanced glucose uptake by tissues after hypothalamic leptin injection. Pretreatment with guanethidine significantly suppressed the increased glucose uptake by the tissues in response to leptin injected into the VMH, whereas bilateral adrenal demedullation had no significant effect. Treatment with propranolol but not phenoxybenzamine also decreased significantly enhanced glucose uptake by the tissues. We further examined the interaction of the effects of hypothalamic leptin and insulin administered peripherally by clamping the glucose concentrations at a constant level. When leptin was injected into the VMH and a maximal dose of insulin was administered intravenously, the rates of glucose uptake by the heart, BAT, and skeletal muscles, but not by WAT, markedly increased beyond the values reached by insulin stimulation alone. Surgical sympathetic denervation of BAT abolished the enhancement of glucose uptake in this tissue, decreasing to the level stimulated by insulin alone. These results appear to indicate that leptin in the hypothalamus enhances glucose uptake in certain peripheral tissues through mediation of a beta-adrenergic mechanism for the sympathetic nerves innervating the tissues and that central leptin and peripheral insulin have a synergistic role in augmenting tissue glucose uptake.

Cross talk between angiotensin II type 1 and type 2 receptors: cellular mechanism of angiotensin type 2 receptor-mediated cell growth inhibition.

Angiotensin (Ang) II plays an important role in regulating cardiovascular hemodynamics as well as cardiovascular structure. At least two distinct receptor subtypes of Ang II have been defined on the basis of their differential pharmacological and biochemical properties, and designated as Ang II type 1 (AT1) receptor and type 2 (AT2) receptor. Most of the known effects of Ang II in adult tissues are attributable to the AT1 receptor. Recent cloning of the AT2 receptor has revealed a variety of new physiological effects of Ang II. AT1 and AT2 receptors belong to the seven-transmembrane receptor family. However, the function and signaling mechanism of these receptor subtypes are quite different. These receptors seem to exert opposite effects in terms of cardiovascular hemodynamics and cell growth. Growth inhibitory effects of AT2 receptors are unique in that this receptor activates a variety of phosphatases and cross talks with the signaling of other seven-transmembrane, G protein-coupled receptors, as well as other classes of growth factor receptors. We will review recent concepts of the molecular and cellular mechanisms of AT2 receptor action in this article.

L-Glutamate and insulin enhance glycogen synthesis in cultured astrocytes from the rat brain through different intracellular mechanisms.

The effects of L-glutamate and insulin on glycogen synthesis in astrocytes were examined. L-Glutamate and insulin both stimulated glycogen synthesis in primary cultures of rat astrocytes in a dose-dependent manner, as measured by the incorporation of 14C from [14C]glucose into glycogen. D-Aspartate also increased the incorporation of 14C into glycogen. When insulin and L-glutamate were added together, the glycogen synthesis as well as glycogen content of the cells was additively increased. Wortmannin, an inhibitor of phosphatidylinositol 3-kinase, had little effect on glycogen synthesis induced by L-glutamate, whereas it suppressed the insulin-induced glycogen synthesis. These results suggest that the insulin- and L-glutamate-induced glycogen syntheses are mediated by different intracellular mechanisms. In fact, insulin stimulated the conversion of glycogen synthase b to glycogen synthase a, which was suppressed by wortmannin. L-Glutamate and D-aspartate, however, did not increase the level of glycogen synthase a activity. By contrast, L-glutamate increased 2-deoxy-D-[3H]glucose uptake by the astrocytes, whereas insulin did not affect the uptake. These results suggest that insulin stimulates glycogen synthesis in astrocytes by activating glycogen synthase, which is dependent on a wortmannin-sensitive signaling pathway. L-Glutamate, however, enhances the glucose uptake, which contributes to the increase in glycogen synthesis in the cells.

Microinjection of leptin into the ventromedial hypothalamus increases glucose uptake in peripheral tissues in rats.

We studied the effects of microinjection of leptin into the ventromedial hypothalamus (VMH) and lateral hypothalamus (LH) on glucose uptake in peripheral tissues in unanesthetized rats. The rate of glucose uptake was assessed in vivo by 2-[3H]deoxyglucose incorporation. Single injection of leptin into VMH increased glucose uptake in brown adipose tissue (BAT), heart, skeletal muscles, and spleen but not in white adipose tissue or skin. On the other hand, microinjection of leptin into LH had little effect on glucose uptake in those tissues. The plasma concentrations of glucose and insulin were unaltered by intrahypothalamic injection of leptin into either VMH or LH. Among skeletal muscles, the increase in glucose uptake induced by intrahypothalamic injection of leptin was greater in the soleus than in the extensor digitorum longus. Likewise, the increased glucose uptake in the gastrocnemius in response to leptin was more prominent in the red part than in the white part of the tissue. When surgical sympathetic denervation of the interscapular BAT was performed, the enhanced glucose uptake by BAT in response to intrahypothalamic leptin was completely suppressed. These findings suggest that intrahypothalamic injection of leptin preferentially increases glucose uptake by some peripheral tissues through activation of the VMH-sympathetic (or its neighboring medial hypothalamus-sympathetic) nervous system, thereby contributing to the maintenance of energy balance.

Effects of noradrenaline on the cell-surface glucose transporters in cultured brown adipocytes: novel mechanism for selective activation of GLUT1 glucose transporters.

Glucose transport into rat brown adipocytes has been shown to be stimulated directly by the sympathetic neurotransmitter, noradrenaline, without a significant increase in the protein content of either GLUT1 or GLUT4 glucose transporter in the plasma membrane [Shimizu, Kielar, Minokoshi and Shimazu (1996) Biochem. J. 314, 485-490]. In the present study, we labelled the exofacial glucose-binding sites of GLUT1 and GLUT4 with a membrane-impermeant photoaffinity reagent, 2-N-[4-(1-azitrifluoroethyl)benzoyl]-[2-3H]1,3-bis- (D-mannos-4-yloxy)-2-propylamine (ATB-[3H]BMPA), to determine which isoform is responsible for the noradrenaline-induced increase in glucose transport into intact brown adipocytes in culture. Insulin stimulated the rate of hexose transport by increasing ATB-[3H]BMPA-labelled cell-surface GLUT4. In contrast, the noradrenaline-induced increase in glucose transport was not accompanied by an increased ATB-[3H]BMPA labelling of GLUT4, nor with an increased amount of GLUT4 in the plasma membrane fraction as assessed by Western blotting, indicating that noradrenaline does not promote the translocation of GLUT4. However, noradrenaline induced an increase in photoaffinity labelling of cell-surface GLUT1 without an apparent increase in the immunoreactive GLUT1 protein in the plasma membrane. This is suggestive of an increased affinity of GLUT1 for the ligand. In fact, the Ki value of non-radioactive ATB-BMPA for 2-deoxy-D-glucose uptake was significantly decreased after treatment of the cells with noradrenaline. The increased photoaffinity labelling of GLUT1 and increased glucose transport caused by noradrenaline were inhibited by a cAMP antagonist, cAMP-S Rp-isomer. These results demonstrate that noradrenaline stimulates glucose transport in brown adipocytes by enhancing the functional activity of GLUT1 through a cAMP-dependent mechanism.

The beta3-adrenergic agonist BRL37344 increases glucose transport into L6 myocytes through a mechanism different from that of insulin.

In the present study, we examined the effects of BRL37344, a selective beta3-adrenergic agonist, on glucose transport into L6 myocytes and the results were compared with the effects of insulin. Insulin increased 2-deoxyglucose (2-DG) uptake in a dose-dependent manner, with maximal stimulation at 10(-7)M. BRL37344 ranging from 10(-7) to 10(-5)M also enhanced 2-DG uptake in the absence of insulin. The effects of insulin and BRL37344 were completely additive, suggesting that these two agents enhance glucose uptake by L6 myocytes through different mechanisms. In fact, BRL37344 apparently did not increase tyrosine phosphorylation of cellular proteins in L6 myocytes, whereas insulin stimulated tyrosine phosphorylation of 180-190 and 95 kDa proteins. Furthermore, BRL37344-induced increase in glucose transport was not blocked by wortmannin, an inhibitor of phosphatidylinositol 3-kinase, whereas the insulin-induced effect was completely abolished. When L6 myocytes were incubated with insulin, the content of GLUT4 in the plasma membrane was increased. However, BRL37344 did not affect the GLUT4 content in the plasma membrane. BRL37344 did not increase the Vmax value for glucose uptake but decreased the Km value, although insulin increased the Vmax value. These results suggest that BRL37344 enhances glucose transport into L6 myocytes through a signaling pathway different from that of insulin and that the mechanism does not involve the translocation of GLUT4, but may be due to an increase in the intrinsic activity of GLUT present in the plasma membrane.

Activation of mitogen-activated protein kinase by norepinephrine in brown adipocytes from rats.

We have investigated the adrenergic control of mitogen-activated protein kinase (MAPK) activity in brown adipocytes. Cold exposure in rats led to an activation of MAPK in brown adipose tissue, as determined by the gel mobility shift assay and in-gel kinase assay. In contrast, no activation was seen after surgical sympathetic denervation of the tissue. The neurotransmitter, norepinephrine (NE), directly activated MAPK of brown adipocytes in primary cultures in the absence of insulin and serum. NE-induced activation of MAPK was mimicked by beta-adrenergic agonists, including a beta 3-agonist, BRL37344. Activation of MAPK also was observed by an alpha-agonist, phenylephrine, the extent of which being much lower than that by beta-agonists. The effect of NE was attenuated by the beta-adrenergic antagonist, propranolol. Dibutyryl cAMP also mimicked the effect of NE. The phorbol ester, phorbol-12-myristate, 13-acetate(PMA), could induce activation of MAPK, but pretreatment of the cultured cells with PMA to down-regulate protein kinase C did not abolish the ability of NE in activating MAPK. Furthermore, a selective inhibitor of phosphatidylinositol-3 kinase, wortmannin, did not inhibit the effect of NE, whereas insulin-induced activation of MAPK was totally suppressed. These results demonstrate that NE activates MAPK directly in brown adipocytes and that the effect of NE is not mediated by PMA-sensitive protein kinase C or wortmannin-sensitive phosphatidylinositol-3 kinase but rather is likely to be dependent on beta-receptor-mediated increase in cAMP with a minor contribution of alpha-receptor-mediated signals.

Noradrenaline increases glucose transport into brown adipocytes in culture by a mechanism different from that of insulin.

Glucose uptake into brown adipose tissue has been shown to be enhanced directly by noradrenaline (norepinephrine) released from sympathetic nerves. In this study we characterized the glucose transport system in cultured brown adipocytes, which responds to noradrenaline as well as insulin, and analysed the mechanism underlying the noradrenaline-induced increase in glucose transport. Insulin increased 2-deoxyglucose (dGlc) uptake progressively at concentrations from 10(-11) to 10(-6) M, with maximal stimulation at 10(-7) M. Noradrenaline concentrations ranging from 10(-8) to 10(-6) M also enhanced dGlc uptake, even in the absence of insulin. The effects of noradrenaline and insulin on dGlc uptake were additive. The stimulatory effect of noradrenaline was mimicked by the beta3-adrenergic agonist, BRL37344, at concentrations two orders lower than noradrenaline. Dibutyryl cyclic AMP also mimicked the stimulatory effect of noradrenaline, and the antagonist of cyclic AMP, cyclic AMP-S Rp-isomer, blocked the enhancement of glucose uptake due to noradrenaline. Furthermore Western blot analysis with an anti-phosphotyrosine antibody revealed that, in contrast with insulin, noradrenaline apparently does not stimulate intracellular phosphorylation of tyrosine, suggesting that the noradrenaline-induced increase in dGlc uptake depends on elevation of the intracellular cyclic AMP level and not on the signal chain common to insulin. When cells were incubated with insulin, the content of the muscle/adipocyte type of glucose transporter (GLUT4) in the plasma membrane increased, with a corresponding decrease in the amount in the microsomal membrane. In contrast, noradrenaline did not affect the subcellular distribution of GLUT4 or that of the HepG2/erythrocyte type of glucose transporter. Although insulin increased Vmax. and decreased the Km value for glucose uptake, the effect of noradrenaline was restricted to a pronounced decrease in Km. These results suggest that the mechanism by which noradrenaline stimulates glucose transport into brown adipocytes is not due to translocation of GLUT but is probably due to an increase in the intrinsic activity of GLUT, which is mediated by a cyclic AMP-dependent pathway.

Expression of beta3-adrenoceptor and stimulation of glucose transport by beta3-agonists in brown adipocyte primary culture.

Precursor cells of brown adipocytes were isolated from the interscapular brown fat of newborn rats and cultured on collagen-coated dishes. When confluent cells were treated with dexamethasone, mRNAs for muscle/adipocyte type of glucose transporter, hormone-sensitive lipase, and CCAAT/enhancer binding protein alpha were increased remarkably, confirming a predominant effect of dexamethasone on the terminal differentiation of the cultured cells. Effects of dexamethasone on the expression of three subtypes of beta-adrenoceptor were also examined. beta1- and beta2-adrenoceptor mRNAs remained constant regardless of dexamethasone-treatment, while beta3-adrenoceptor mRNA was present only in dexamethasone-treated differentiated cells. To assess the metabolic response mediated by beta3-adrenoceptor, glucose transport into the cells was estimated. Norepinephrine enhanced glucose transport in dexamethasone-treated differentiated cells, but not in undifferentiated cells. beta3-Adrenergic agonists mimicked completely the stimulatory effect of norepinephrine at concentrations lower by two orders of magnitude. These results suggest that the beta3-adrenoceptor is expressed during the course of differentiation in brown adipocytes and plays a significant role in the response of glucose transport to adrenergic stimulation.

Dexamethasone induces the GLUT4 glucose transporter, and responses of glucose transport to norepinephrine and insulin in primary cultures of brown adipocytes.

Glucose uptake into brown adipose tissue is enhanced directly by norepinephrine released from the sympathetic nerves. In the present study, we tried to establish culture conditions for brown adipocytes which are favorable for investigation of this unique glucose transport. Stromal-vascular cells isolated from the interscapular brown adipose tissue of newborn rats differentiated into brown adipocytes expressing the uncoupling protein, when the cells were maintained on collagen-coated dishes. These cells, however, did not show an increase in 2-[3H]deoxyglucose transport in response to insulin or norepinephrine, nor did they exhibit expression of the GLUT4 glucose transporter, whereas GLUT1 was present, as judged on Western blotting. Pre-treatment of confluent cells with dexamethasone induced a response of glucose transport to either insulin or norepinephrine, and the expression of GLUT4, together with notable accumulation of lipid droplets. The induction of GLUT4 expression by dexamethasone was dose-dependent and potentiated by insulin. These results indicate that treatment of cultured brown adipocytes with dexamethasone makes it feasible to analyze the mechanism underlying the enhancement of glucose transport induced by norepinephrine.

Regulatory mechanism of the ventromedial hypothalamus in enhancing glucose uptake in skeletal muscles.

To evaluate roles of the sympathetic nervous system in enhancing glucose uptake in skeletal muscles in response to electrical stimulation of the ventromedial hypothalamic nucleus (VMH), the effects of guanethidine treatment and adrenal demedullation on the response were examined in rats anesthetized and injected with muscle relaxant, pancuronium bromide. Pretreatment with guanethidine effectively suppressed the increase in the rate constant of glucose uptake, measured by the 2-deoxy-D-[3H]glucose method, in skeletal muscles upon VMH stimulation. However, bilateral adrenodemedullation had no significant effect. These results suggest that the VMH is intimately concerned in the regulation of glucose uptake in skeletal muscles through intermediation of the sympathetic nerves.

Effect of a beta 3-adrenergic agonist, BRL35135A, on glucose uptake in rat skeletal muscle in vivo and in vitro.

The effects of the beta 3-agonist, BRL35135A, on glucose uptake in the peripheral tissues of the rat, including skeletal muscle, were studied using the 2-[3H]deoxyglucose method in anaesthetized adult animals. Intravenous infusion of the beta 3-agonist dose-dependently increased the rate constant of glucose uptake in three types of skeletal muscle, brown adipose tissue, white adipose tissue, heart and diaphragm, but not in the brain, spleen or lung. Although infusion of the beta 3-agonist did not change the plasma concentration of glucose appreciably, it caused an increase in the plasma concentration of insulin when given at more than 25 micrograms/kg per h. To ascertain whether the effect of the beta 3-agonist on glucose uptake in skeletal muscle is mediated by insulin, glucose uptake into soleus muscle isolated from young rats was also measured in vitro using different concentrations of the beta 3-agonist. The beta 3-agonist BRL37344 (an active metabolite of BRL35135A) significantly increased glucose transport in a dose-dependent manner, with maximum stimulation at 100 pmol/l. These results demonstrate that glucose uptake in skeletal muscle can be enhanced independently of the action of insulin, probably through the mediation of beta 3-adrenoceptors present in the tissue.