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A study was conducted to examine the mechanism of the adipose tissue inducible nitric oxide synthase (iNOS) in endotoxemia. Male Sprague-dawley rats were injected with bacterial endotoxin lipopolysaccharide (LPS) to increase the iNOS messenger RNA and protein levels of the white and brown adipose tissues. Results suggest that LPS treatment has significantly increased iNOS activity and expression in both the white and brow adipose tissues.

The influence of nitric oxide on the insulin-dependent glucose uptake of skeletal muscle and adipose tissue is investigated using nitric oxide synthase (NOS) inhibition during hyperinsulinemic-euglycemic clamp studies in rats. NOS inhibition resulted in a significant reduction of the glucose uptake of several tissues and organs in vivo. No such effects were observed in vitro. These results are suggestive of the dependence of insulin-mediated glucose uptake on nitric oxide.

Electrical stimulation induces GLUT-4 translocation to the plasma membrane and transverse tubules of red gastrocnemius (RG) and white gastrocnemius (WG) muscles. Experiments were conducted testing the hypothesis that GLUT-4 translocation to the cell surface may underlie the responsiveness of type II muscle fibers. The greater translocation of GLUT-4 to transverse tubules of WG in contrast to RG may provide an explanation on both muscle types' responsiveness toward supramaximal electrical stimulation.

Mammalian skeletal muscles have been classified according to structure and function but not according to fiber types. Herein, skeletal muscles with insulin-induced sodium (Na+)-potassium (K+)-adenosine triphosphatase (ATPase) activity were classified according to muscle fiber type. Na+-K+-ATPase activity was detected in muscles composed mostly of oxidative (red) fibers but not in glycolytic (white) fibers. Insulin-induced translocation of Na+-K+-ATPase subunits to the plasma membrane was also observed in the red muscles, but not in white muscles.

A fractionation protocol that can separate plasma membranes, T tubules and internal membranes from skeletal muscle cells was developed. The muscles were minced, treated with phenylmethylsulfonylfluroide, homogenized and suspended in lithium bromide and tris(hydroxymethyl)aminomethane. These were then centrifuged and resuspended in magnesium sulfate and N-2 hydroxyethylpiperazine-N'-2-ethanesulfonic acid. An internal membrane fraction derived from this method was found to be a source of GLUT-4 glucose transporter. Insulin increases the presence of GLUT-4 in the T tubules and plasma membranes assayed through this method.

Experiments with alpah2-subunits or the Na+-K+-ATPase in the surface skeletal muscles of mammals indicate that an acute stimulation of insulin increases the number of immunolabeled alpha2-subunits in the plasma membrane. There is a gain in the range of 1.5 times to 3.7 times in the plasma membrane. The results provide morphological quantitative evidence that the alpha2-immuno reactive subunits of the Na+-K+-ATPase are present at both the mammalian skeletal muscle (intracellularly) and the plasma membrane.

The mechanism responsible for the specific decrease in BAT mitochondrial content in obese SHR/N-cp rats may be related to a marked insulin resistance and the defective activity of the sympathetic nervous system. This was inferred from experiments on mitochondrial thermogenic capacity in brown adipose tissue (BAT) in obese animals to test whether insulin resistance of these animals is specifically associated with a thermogenic defect in BAT. Extreme insulin resistance induces BAT metabolism alterations.

A study was conducted to examine the influence of norepinephrine and insulin on glucose movement in obese spontaneously hypertensive rats (O-SHR) brown adipocytes separated from obese nondiabetic Lister and Albany (O-LA) rats. The results revealed that significant metabolic imperfections exist in the brown adipose tissue or O-SHR but not in O-LA cells, that postreceptor imperfections are present in O-SHR adipocyte refractoriness to norepinephrine and that decreased mitochondrial content could indicate the main metabolic changes which explain the lowered influence of norepinephrine on glucose transport and thermogenesis.

Purified high-fat diets are commonly used to promote obesity and metabolic disturbances in animal models. However, most studies also use unpurified chow (low in fat and high in fiber content) as a "healthy" diet which can impact gut health. Here, we investigated the impact of feeding chow or purified diets varying in fat content on metabolic and intestinal functions in relation to temporal changes in gut microbiota composition. C57BL/6J male mice were fed either chow or purified low-fat (LF) or high-fat (HF) diet for 12 wk. LF-fed mice displayed similar glucose and insulin homeostasis and bile acid profile than chow-fed mice but did exhibit higher fasting insulinemia and impaired insulin clearance, which was associated with increased weight gain and fat mass. Compared with chow, feeding LF or HF diets increased hepatic steatosis and inflammation, and impaired intestinal integrity. Changes in gut microbiota preceded these effects, with both purified diets inducing rapid and dramatic decrease of bacterial diversity and short-chain fatty acid (SCFA) production. Importantly, branched SCFA levels increased only in HF-fed mice suggesting that this response is driven by excessive dietary fat rather than lack of fiber content. LF-fed mucus-associated microbial composition was also closer to that of chow-fed mice than that of HF-fed mice, suggesting the impact of fat content on this specific microbial community. We have identified distinct and overlapping gut microbiome and metabolic impairments caused by low fiber or high-fat contents in HF-fed animals, revealing their selective mechanisms underpinning the gastrointestinal and metabolic impacts in obesity. [ABSTRACT FROM AUTHOR]

The roles of sex and sex-hormones on the metabolic consequences of intermittent hypoxia (IH, a reliable model of sleep apnea) are unknown. We used intact male or female mice and ovariectomized (OVX) females treated with vehicle (Veh) or estradiol (E2) and exposed to normoxia (Nx) or IH (6% O2, 10 cycles/h, 12 h/day, 2 wk). Mice were then fasted for 6 h, and we measured fasting glucose and insulin levels and performed insulin or glucose tolerance tests (ITT or GTT). We also assessed liver concentrations of glycogen, triglycerides (TGs), and expression levels of genes involved in aerobic or anaerobic metabolism. In males, IH lowered fasting levels of glucose and insulin, slightly improved glucose tolerance, but altered glucose tolerance in females. In OVX-Veh females, IH reduced fasting glucose and insulin levels and strongly impaired glucose tolerance. E2 supplementation reversed these effects and improved homeostasis model assessment of b-cell function (HOMA-b), a marker of pancreatic glucose-induced insulin released. IH decreased liver TG concentration in males and slightly increased glycogen in OVX-Veh females. Liver expression of glycolytic (Ldha) and mitochondrial (citrate synthase, Pdha1) genes was reduced by IH in males and in OVX-Veh females, but not in intact or OVX-E2 females. We conclude that 1) IH reduced fasting levels of glycemia in males and in ovariectomized females. 2) IH improves glucose tolerance only in males. 3) In females IH decreased glucose tolerance, this effect was amplified by ovariectomy, and reversed by E2 supplementation. 4) During IH exposures, E2 supplementation appears to improve pancreatic b cells functions. [ABSTRACT FROM AUTHOR]

Prevention of skeletal muscle insulin resistance by dietary cod protein in high fat-fed rats. Am J Physiol Endocrinol Metab 281: E62-E71, 2001.--In the present study, we tested the hypothesis that fish protein may represent a key constituent of fish with glucoregulatory activity. Three groups of rats were fed a high-fat diet in which the protein source was casein, fish (cod) protein, or soy protein; these groups were compared with a group of chow-fed controls. High-fat feeding led to severe whole body and skeletal muscle insulin resistance in casein- or soy protein-fed rats, as assessed by the euglycemic clamp technique coupled with measurements of 2-deoxy-D-[[sup.3]H]glucose uptake rates by individual tissues. However, feeding cod protein fully prevented the development of insulin resistance in high fat-fed rats. These animals exhibited higher rates of insulin-mediated muscle glucose disposal that were comparable to those of chow-fed rats. The beneficial effects of cod protein occurred without any reductions in body weight gain, adipose tissue accretion, or expression of tumor necrosis factor-a in fat and muscle. Moreover, L6 myocytes exposed to cod protein-derived amino acids showed greater rates of insulin-stimulated glucose uptake compared with cells incubated with casein- or soy protein-derived amino acids. These data demonstrate that feeding cod protein prevents obesity-induced muscle insulin resistance in high fat-fed obese rats at least in part through a direct action of amino acids on insulin-stimulated glucose uptake in skeletal muscle cells. amino acids; high-fat diet; adipose tissue

Cytokines and endotoxin induce cytokine receptors in skeletal muscle. Am J Physiol Endocrinol Metab 279: E196-E205, 2000.--Proinflammatory cytokines are important factors in the regulation of diverse aspects of skeletal muscle function; however, the muscle cytokine receptors mediating these functions are uncharacterized. Binding kinetics (dissociation constant = 39 [+ or -] 4.7 x [10.sup.-9] M, maximal binding = 3.5 [+ or -] 0.23 x [10.sup.-12] mol/mg membrane protein) of muscle tumor necrosis factor (TNF) receptors were obtained. Skeletal muscle was found to express mRNAs encoding interleukin-1 type I and II receptors, interleukin-6 receptor (IL-6R), and interferon-y receptor by RT-PCR, but these receptors were below limits of detection of ligandbinding assay ([is greater than or equal to] -1 fmol binding sites/mg protein). Twenty-four hours after intraperitoneal administration of endotoxin to rats, TNF receptor type II (TNFRII) and IL-6R mRNA were increased in skeletal muscle (P [is less than] 0.05). In cultured L6 cells, the expression of mRNA encoding TNFRII and IL-6R receptors was induced by TNF-[Alpha], and all six cytokine receptor mRNA were induced by a mixture of TNF-[Alpha], IFN-[Gamma], and endotoxin (P [is less than] 0.05). This suggests that the low level of cytokine receptor expression is complemented by a capacity for receptor induction, providing a clear mechanism for amplification of cytokine responses at the muscle level. tumor necrosis factor-[Alpha]

Bonen, Arend, Dragana Miskovic, Mio Tonouchi, Kathleen Lemieux, Mariengela C. Wilson, Andre Marette, and Andrew P. Halestrap. Abundance and subcellular distribution of MCT1 and MCT4 in heart and fast-twitch skeletal muscles. Am J Physiol Endocrinol Metab 278: E1067-E1077, 2000.--The expression of two monocarboxylate transporters (MCTs) was examined in muscle and heart. MCT1 and MCT4 proteins are coexpressed in rat skeletal muscles, but only MCT1 is expressed in rat hearts. Among six rat fast-twitch muscles (red and white gastrocnemius, plantaris, extensor digitorum longus, red and white tibialis anterior) there was an inverse relationship between MCT1 and MCT4 (r = -0.94). MCT1 protein was correlated with MCT1 mRNA (r = 0.94). There was no relationship between MCT4 mRNA and MCT4 protein. MCT1 (r = -0.97) and MCT4 (r = 0.88) protein contents were correlated with percent fast-twitch glycolytic fiber. When normalized for their mRNAs, MCT1 but not MCT4 was still correlated with the percent fast-twitch glycolytic fiber composition of rat muscles (r = -0.98). MCT1 and MCT4 were also measured in plasma membranes (PM), triads (TR), T tubules (TT), sarcoplasmic reticulum (SR), and intracellular membranes (IM). There was an intracellular pool of MCT4 but not of MCT1. The MCT1 subcellular distribution was as follows: PM (100%) [is greater than] TR (31.6%) [is greater than] SR (15%) = TT (14%) [is greater than] IM (1.7%). The MCT4 subcellular distribution was considerably different [PM (100%) [is greater than] TR (66.5%) [is greater than] TT (36%) = SR (43%) [is greater than] IM (24%)]. These studies have shown that 1) the mechanisms regulating the expression of MCT1 (transcriptional and posttranscriptional) and MCT4 (posttranscriptional) are different and 2) differences in MCT1 and MCT4 expression among muscles, as well as in their subcellular locations, suggest that they may have different roles in muscle. lactate; transport; transporters; fast-twitch fibers; plasma membrane; T tubules, messenger ribonucleic acid; monocarboxylate transporter 1; monocarboxylate transporter 4

Effect of metformin on the vascular and glucose metabolic actions of insulin in hypertensive rats. Am J Physiol Gastrointest Liver Physiol 278: G682-G692, 2000.--We investigated the long-term effect of metformin treatment on blood pressure, insulin sensitivity, and vascular responses to insulin in conscious spontaneously hypertensive rats (SHR). The rats were instrumented with intravascular catheters and pulsed Doppler flow probes to measure blood pressure, heart rate, and blood flow. Insulin sensitivity was assessed by the euglycemic hyperinsulinemic clamp technique. Two groups of SHR received metformin (100 or 300 mg [multiply by] [kg.sup.-1] [multiply by] [day.sup.-1]) for 3 wk while another group of SHR and a group of Wistar Kyoto (WKY) rats were left untreated. We found that vasodilation of skeletal muscle and renal vasculatures by insulin is impaired in SHR. Moreover, a reduced insulin sensitivity was detected in vivo and in vitro in isolated soleus and extensor digitorum longus muscles from SHR compared with WKY rats. Three weeks of treatment with metformin improves the whole-body insulin-mediated glucose disposal in SHR but has no blood pressure-lowering effect and no influence on vascular responses to insulin (4 mU [multiply by] [kg.sup.-l] [multiply by ] [min.sup.-1]). An improvement in insulin-mediated glucose transport activity was detected in isolated muscles from metformin-treated SHR, but in the absence of insulin no changes in basal glucose transport activity were observed. It is suggested that part of the beneficial effect of metformin on insulin resistance results from a potentiation of the hormone-stimulating effect on glucose transport in peripheral tissues (mainly skeletal muscle). The results argue against a significant antihypertensive or vascular effect of metformin in SHR. insulin sensitivity; hypertension; blood flow; skeletal muscle

Lavigne, Charles, Andre Marette, and Helene Jacques. Cod and soy proteins compared with casein improve glucose tolerance and insulin sensitivity in rats. Am. J. Physiol. Endocrinol. Metab. 278: E491-E500, 2000.--The aim of the present study was to determine the effects of feeding various dietary proteins on insulin sensitivity and glucose tolerance in rats. Male Wistar rats were fed for 28 days with isoenergetic diets containing either casein, soy protein, or cod protein. Cod protein-fed and soy protein-fed rats had lower fasting plasma glucose and insulin concentrations compared with casein-fed animals. After intravenous glucose bolus, cod protein- and soy protein-fed rats induced lower incremental areas under glucose curves compared with casein-fed animals. Improved peripheral insulin sensitivity was confirmed by higher glucose disposal rates in cod protein-and soy protein-fed rats (15.2 [+ or -] 0.3 and 13.9 [+ or -] 06 mg [multiplied by] [kg.sup.-1] [multiplied by] [min.sup.-1], respectively) compared with casein-fed animals (6.5 [+ or -] 0.7 mg [multiplied by] [kg.sup.-1] [multiplied by] [min.sup.-1], P [is less than] 0.05). Moreover, test meal experiments revealed that, in the postprandial state, the lower plasma insulin concentrations in cod protein- and soy protein-fed animals could be also due to decreased pancreatic insulin release and increased hepatic insulin removal. In conclusion, the metabolic responses to three common dietary proteins indicate that cod and soy proteins, when compared with casein, improve fasting glucose tolerance and peripheral insulin sensitivity in rats. triglycerides; intravenous glucose tolerance test; test meal; hyperinsulinemic-euglycemic clamp.