Ghrelin infusion in humans induces acute insulin resistance and lipolysis independent of growth hormone signaling

OBJECTIVE—Ghrelin is a gut-derived peptide and an endogenous ligand for the growth hormone (GH) secretagogue receptor. Exogenous ghrelin stimulates the release of GH (potently) and adrenocorticotropic hormone (ACTH) (moderately). Ghrelin is also orexigenic, but its impact on substrate metabolism is controversial. We aimed to study direct effects of ghrelin on substrate metabolism and insulin sensitivity in human subjects.RESEARCH DESIGN AND METHODS—Six healthy men underwent ghrelin (5 pmol · kg⁻¹ · min⁻¹) and saline infusions in a double-blind, cross-over study to study GH signaling proteins in muscle. To circumvent effects of endogenous GH and ACTH, we performed a similar study in eight hypopituitary adults but replaced with GH and hydrocortisone. The methods included a hyperinsulinemic-euglycemic clamp, muscle biopsies, microdialysis, and indirect calorimetry.RESULTS—In healthy subjects, ghrelin-induced GH secretion translated into acute GH receptor signaling in muscle. In the absence of GH and cortisol secretion, ghrelin acutely decreased peripheral, but not hepatic, insulin sensitivity together with stimulation of lipolysis. These effects occurred without detectable suppression of AMP-activated protein kinase phosphorylation (an alleged second messenger for ghrelin) in skeletal muscle.CONCLUSIONS—Ghrelin infusion acutely induces lipolysis and insulin resistance independently of GH and cortisol. We hypothesize that the metabolic effects of ghrelin provide a means to partition glucose to glucose-dependent tissues during conditions of energy shortage.Ghrelin, an endogenous ligand for the growth hormone (GH) secretagogue receptor (GHS-R), stimulates GH and adrenocorticotropic hormone (ACTH) secretion (1) in addition to having orexigenic and gastrokinetic effects (2,3). The observation that GHS-R is located in peripheral tissues suggests that ghrelin may exert direct effects (4). The effects of ghrelin on substrate in humans are uncertain, but insulin resistance and stimulation of lipolysis have been reported (5–7). However, it remains difficult to segregate direct effects from effects related to GH and cortisol, and we have recently demonstrated that somatostatin infusion fails to sufficiently suppress ghrelin-induced GH and cortisol secretion (8). Hormonally replaced hypopituitary patients constitute a means for studying putative GH- and cortisol-independent effects of ghrelin in human subjects in vivo.We aimed to study potential direct effects of ghrelin on substrate metabolism and insulin sensitivity in the postabsorptive state. In one experiment in healthy adults, we assessed whether ghrelin-induced GH release translated into GH signaling in skeletal muscle, in the event of which the importance of abrogating indirect effects of ghrelin is obvious. Second, we studied the effects of ghrelin exposure on whole-body and regional substrate metabolism in the basal and insulin-stimulated state in hypopituitary patients on stable replacement with GH and hydrocortisone.     

Interleukin-6 induces cellular insulin resistance in hepatocytes

Interleukin (IL)-6 is one of several proinflammatory cytokines that have been associated with insulin resistance and type 2 diabetes. A two- to threefold elevation of circulating IL-6 has been observed in these conditions. Nonetheless, little evidence supports a direct role for IL-6 in mediating insulin resistance. Here, we present data that IL-6 can inhibit insulin receptor (IR) signal transduction and insulin action in both primary mouse hepatocytes and the human hepatocarcinoma cell line, HepG2. This inhibition depends on duration of IL-6 exposure, with a maximum effect at 1-1.5 h of pretreatment with IL-6 in both HepG2 cells and primary hepatocytes. The IL-6 effect is characterized by a decreased tyrosine phosphorylation of IR substrate (IRS)-1 and decreased association of the p85 subunit of phosphatidylinositol 3-kinase with IRS-1 in response to physiologic insulin levels. In addition, insulin-dependent activation of Akt, important in mediating insulin's downstream metabolic actions, is markedly inhibited by IL-6 treatment. Finally, a 1.5-h preincubation of primary hepatocytes with IL-6 inhibits insulin-induced glycogen synthesis by 75%. These data suggest that IL-6 plays a direct role in insulin resistance at the cellular level in both primary hepatocytes and HepG2 cell lines and may contribute to insulin resistance and type 2 diabetes.     

Overfeeding rapidly induces leptin and insulin resistance.

In common forms of obesity, hyperphagia, hyperinsulinemia, and hyperleptinemia coexist. Here, we demonstrate rapid induction of insulin and leptin resistance by short-term overfeeding. After 3 and 7 days on the assigned diet regimen, rats were tested for their biological responses to acute elevations in plasma insulin and leptin concentrations. Severe resistance to the metabolic effects of both leptin and insulin ensued after just 3 days of overfeeding. During the insulin clamp studies, glucose production was decreased by approximately 70% in control rats and 28-53% in overfed rats. Similarly, leptin infusion doubled the contribution of gluconeogenesis to glucose output in control rats but failed to modify gluconeogenesis in overfed animals. These findings demonstrate a paradoxical and rapid collapse of the leptin system in response to nutrient excess. This partial failure is tightly coupled with the onset of insulin resistance.     

Mechanism of amino acid-induced skeletal muscle insulin resistance in humans

Plasma concentrations of amino acids are frequently elevated in insulin-resistant states, and a protein-enriched diet can impair glucose metabolism. This study examined effects of short-term plasma amino acid (AA) elevation on whole-body glucose disposal and cellular insulin action in skeletal muscle. Seven healthy men were studied for 5.5 h during euglycemic (5.5 mmol/l), hyperinsulinemic (430 pmol/l), fasting glucagon (65 ng/l), and growth hormone (0.4 microg/l) somatostatin clamp tests in the presence of low (approximately 1.6 mmol/l) and increased (approximately 4.6 mmol/l) plasma AA concentrations. Glucose turnover was measured with D-[6,6-(2)H(2)]glucose. Intramuscular concentrations of glycogen and glucose-6-phosphate (G6P) were monitored using (13)C and (31)P nuclear magnetic resonance spectroscopy, respectively. A approximately 2.1-fold elevation of plasma AAs reduced whole-body glucose disposal by 25% (P < 0.01). Rates of muscle glycogen synthesis decreased by 64% (180--315 min, 24 plus minus 3; control, 67 plus minus 10 micromol center dot l(-1) center dot min(-1); P < 0.01), which was accompanied by a reduction in G6P starting at 130 min (DeltaG6P(260--300 min), 18 plus minus 19; control, 103 plus minus 33 micromol/l; P < 0.05). In conclusion, plasma amino acid elevation induces skeletal muscle insulin resistance in humans by inhibition of glucose transport/phosphorylation, resulting in marked reduction of glycogen synthesis.     

Acute vasoconstriction-induced insulin resistance in rat muscle in vivo

Insulin-mediated changes in blood flow are associated with altered blood flow distribution and increased capillary recruitment in skeletal muscle. Studies in perfused rat hindlimb have shown that muscle metabolism can be regulated by vasoactive agents that control blood flow distribution within the hindlimb. In the present study, the effects of a vasoconstrictive agent that has no direct effect on skeletal muscle metabolism but that alters perfusion distribution in rat hindlimb was investigated in vivo to determine its effects on insulin-mediated vascular action and glucose uptake. We measured the effects of alpha-methylserotonin (alpha-met5HT) on mean arterial blood pressure, heart rate, femoral blood flow, hindlimb vascular resistance, and glucose uptake in control and euglycemic insulin-clamped (10 mU x min(-1) x kg(-1)) anesthetized rats. Blood flow distribution within the hindlimb muscles was assessed by measuring the metabolism of 1-methylxanthine (1-MX), an exogenously added substrate for capillary xanthine oxidase. Alpha-met5HT (20 microg x min(-1) x kg(-1)) infusion alone increased mean arterial blood pressure by 25% and increased hindlimb vascular resistance but caused no change in femoral blood flow. These changes were associated with decreased hindlimb 1-MX metabolism indicating less capillary flow. Insulin infusion caused decreased hindlimb vascular resistance that was associated with increased femoral blood flow and 1-MX metabolism. Treatment with alpha-met5HT infusion commenced before insulin infusion prevented the increase in femoral blood flow and inhibited the stimulation of 1-MX metabolism. Alpha-met5HT infusion had no effect on hindlimb glucose uptake but markedly inhibited the insulin stimulation of glucose uptake (P < 0.05) and was associated with decreased glucose infusion rates to maintain euglycemia (P < 0.05). A significant correlation (P < 0.05) was observed between 1-MX metabolism and hindlimb glucose uptake but not between femoral blood flow and glucose uptake. The results indicate that in vivo, certain types of vasoconstriction in muscle such as elicited by 5HT2 agonists, which prevent normal insulin recruitment of capillary flow, cause impaired muscle glucose uptake. Moreover, if vasoconstriction of this kind results from stress-induced increase in sympathetic outflow, then this may provide a clue as to the link between hypertension and insulin resistance that is often observed in humans.     

Indinavir induces acute and reversible peripheral insulin resistance in rats

The use of HIV protease inhibitors (PIs) has been associated with several metabolic changes, including lipodystrophy, hyperlipidemia, and insulin resistance. The etiology of these adverse effects remains unknown. PIs have recently been found to cause acute and reversible inhibition of GLUT4 activity in vitro. To determine the acute in vivo effects of indinavir on whole-body glucose homeostasis, glucose tolerance tests were performed on PI-na?ve Wistar rats immediately after a single intravenous dose of indinavir. Glucose and insulin levels were significantly elevated in indinavir-treated versus control rats (P < 0.05) during the initial 30 min of the glucose tolerance test. Under euglycemic- hyperinsulinemic clamp conditions, indinavir treatment acutely reduced the glucose infusion rate required to maintain euglycemia by 18 and 49% at indinavir concentrations of 14 and 27 micromol/l, respectively. Muscle 2-deoxyglucose uptake was similarly reduced under these conditions. Restoration of insulin sensitivity was observed within 4 h after stopping the indinavir infusion. Indinavir did not alter the suppression of hepatic glucose output under hyperinsulinemic conditions. These data demonstrate that indinavir causes acute and reversible changes in whole-body glucose homeostasis in rats and support the contribution of GLUT4 inhibition to the development of insulin resistance in patients treated with PIs.     

Acute pain induces an instant increase in natural killer cell cytotoxicity in humans and this response is abolished by local anaesthesia

We have investigated the effect of pain without tissue injury on natural killer (NK) cell activity in peripheral blood in humans and the effect of local anaesthesia on the response. Ten subjects were investigated during two sessions. First, self-controlled painful electric stimulation was applied to abdominal skin for 30 min to an intensity of 8 on a visual analogue scale (0-10). Next, the electric intensity profile was reproduced during local anaesthesia (mepivacaine 10 mg ml-1 s.c. to a total dose of 2.5 mg kg-1). NK cell cytotoxicity was measured using a 4-h 51Cr-release assay against K562 target cells. NK cell activity increased from mean 22 (SEM 4)% (baseline) to 35 (6)% and 36 (5)% after 15 and 30 min of painful stimulation, respectively (P < 0.02). A simultaneous increase in the number of CD56+ cells in peripheral blood during pain was found. Stimulation after local anaesthesia did not change either NK cell activity or number. Parallel and significant increases in concentrations of plasma epinephrine and serum cortisol were observed. These changes were abolished by local anaesthesia. We conclude that acute severe pain without tissue injury markedly increased NK cell cytotoxicity. Local anaesthesia completely abolished this immunological and hormonal response.      

Acute insulin resistance in myocardial ischemia: causes and consequences

Diabetes mellitus is associated with increased risk for cardiovascular mortality because of multiple pathophysiologic mechanisms. Acute stress-induced hyper-glycemia during acute myocardial infarction has gained much attention, as blood glucose levels seem to be an independent risk factor for acute myocardial infarction-related death. Clinical studies that identify stress-induced hyperglycemia as a risk factor are reviewed and its causes are discussed. They can be summarized as the consequence of acute insulin resistance, which in its turn is caused by stress hormones and by proinflammatory cytokines. Hyperglycemia causes the release of proinflammatory cytokines, the induction of reactive radicals, alterations in cardiovascular substrate metabolism, and propagation of coagulation and apoptosis. These all have harmful effects during and after acute myocardial infarction. Recommendations are for strict glycemic control in hyperglycemic patients with acute myocardial infarction, although the target glucose level is still a subject of debate.     

Altered skeletal muscle lipase expression and activity contribute to insulin resistance in humans

OBJECTIVE

Insulin resistance is associated with elevated content of skeletal muscle lipids, including triacylglycerols (TAGs) and diacylglycerols (DAGs). DAGs are by-products of lipolysis consecutive to TAG hydrolysis by adipose triglyceride lipase (ATGL) and are subsequently hydrolyzed by hormone-sensitive lipase (HSL). We hypothesized that an imbalance of ATGL relative to HSL (expression or activity) may contribute to DAG accumulation and insulin resistance.

RESEARCH DESIGN AND METHODS

We first measured lipase expression in vastus lateralis biopsies of young lean (n = 9), young obese (n = 9), and obese-matched type 2 diabetic (n = 8) subjects. We next investigated in vitro in human primary myotubes the impact of altered lipase expression/activity on lipid content and insulin signaling.

RESULTS

Muscle ATGL protein was negatively associated with whole-body insulin sensitivity in our population (r = −0.55, P = 0.005), whereas muscle HSL protein was reduced in obese subjects. We next showed that adenovirus-mediated ATGL overexpression in human primary myotubes induced DAG and ceramide accumulation. ATGL overexpression reduced insulin-stimulated glycogen synthesis (−30%, P < 0.05) and disrupted insulin signaling at Ser1101 of the insulin receptor substrate-1 and downstream Akt activation at Ser473. These defects were fully rescued by nonselective protein kinase C inhibition or concomitant HSL overexpression to restore a proper lipolytic balance. We show that selective HSL inhibition induces DAG accumulation and insulin resistance.

CONCLUSIONS

Altogether, the data indicate that altered ATGL and HSL expression in skeletal muscle could promote DAG accumulation and disrupt insulin signaling and action. Targeting skeletal muscle lipases may constitute an interesting strategy to improve insulin sensitivity in obesity and type 2 diabetes.Skeletal muscle insulin resistance is a strong risk factor of type 2 diabetes and cardiovascular diseases (1,2). Dysfunctional adipose tissue can lead to lipid oversupply and increased flux of free fatty acids (FFAs) into skeletal muscle and is associated with the accumulation of intramyocellular triacylglycerols (IMTGs) (3–5). This chronic lipid overload in tissues can evolve to a state of lipotoxicity leading to cell dysfunction (6). The term “lipotoxicity” defines more generally in skeletal muscle a state of lipid overload (increased concentrations of long-chain acyl-CoA, diacylglycerols [DAGs], and ceramide) causing insulin resistance (7–12). DAGs have been shown to activate novel protein kinase C (PKC) isoforms, such as the novel PKCθ leading to insulin receptor substrate-1 (IRS-1) serine phosphorylation and impaired downstream insulin signaling (10–13). DAGs can be formed through multiple pathways but are also formed as intermediates during triacylglycerol (TAG) synthesis and hydrolysis (14). The control of lipolysis in skeletal muscle has mainly been attributed to hormone-sensitive lipase (HSL), which exhibits a 10-fold higher specific activity for DAG than TAG (15). HSL null mice display normal TAG hydrolase activity after an overnight fast and accumulate large amounts of DAG (16). Recently it was shown that adipose triglyceride lipase (ATGL) plays a major role in the regulation of cellular TAG stores in various tissues of the body, including heart and skeletal muscle (17,18). ATGL specifically drives the hydrolysis of TAG into DAG (18). ATGL-deficient mice are more insulin sensitive and glucose tolerant despite a threefold increase in TAG content in their skeletal muscle (17). The molecular mechanism underlying this phenotype remains unclear. In the current study, we hypothesized that an imbalance of ATGL relative to HSL could increase intracellular DAG concentrations and promote insulin resistance. To test this hypothesis, we first examined the relationship between muscle ATGL expression and whole-body insulin sensitivity in a wide range of subjects. We next manipulated the expression/activity of ATGL and HSL in vitro in cultured human primary skeletal muscle cells and evaluated its impact on lipid pools and insulin signaling.     

Combined effect of growth hormone and cortisol on late posthypoglycemic insulin resistance in humans

The occurrence and mechanisms for late (6.5- to 7.5-h) posthypoglycemic insulin resistance were studied with the euglycemic clamp in 19 healthy subjects. Comparisons were made with a control study with the same insulin infusion rate but where hypoglycemia was prevented by glucose infusion. Glucose production and utilization were studied with D-[3-3H] glucose infusions. Hypoglycemia induced marked insulin resistance shown by lower glucose infusion rates compared with the control study 3.1 +/- 0.3 vs. 6.0 +/- 0.7 mg.kg-1.min-1, P less than .001). This late posthypoglycemic insulin resistance was mainly due to a decreased insulin effect on glucose utilization. Infusion of propranolol did not prevent insulin resistance, whereas somatostatin partially prevented its appearance. Somatostatin plus metyrapone completely normalized posthypoglycemic insulin resistance. A positive correlation (r = .72, P less than .001) was found between initial insulin sensitivity and percent reduction of the insulin effect after hypoglycemia. Thus, hypoglycemia is followed by prolonged (6- to 8-h) insulin resistance. In contrast to early-phase (2- to 3-h) resistance, long-term resistance is not due to beta-adrenergic stimulation but to the combined effect of growth hormone and cortisol. This resistance is also more pronounced in subjects with initially high insulin sensitivity.     

Contribution of insulin resistance to catabolic effect of prednisone on leucine metabolism in humans

To determine the role insulin resistance may play in the catabolic effect of high-dose prednisone therapy, healthy volunteers were studied on four occasions with the hormone-clamp technique at two insulin infusion rates. Subjects were studied after 5 days of prednisone (60 mg/day) or no steroid treatment and were infused with somatostatin, glucagon, growth hormone, [3H]glucose, [14C]leucine, and insulin (0.1 or 0.2 mU.kg-1.min-1). At each rate of insulin infusion, the rate of leucine oxidation was increased (P less than .001) after steroid treatment. Leucine flux, an indicator of whole-body proteolysis, was similar in the presence or absence of steroid treatment (2.26 +/- 0.08 vs. 2.13 +/- 0.04 mumol.kg-1.min-1, respectively) at the lower rate of insulin infusion but was higher during steroid treatment (2.18 +/- 0.06 vs. 1.84 +/- 0.13 mumol.kg-1.min-1) at the 0.2-mU.kg-1.min-1 insulin infusion. Steroid pretreatment had no significant effect on the nonoxidative rates of leucine disappearance. These data provide strong evidence that the protein wasting associated with glucocorticosteroid therapy is in part the result of steroid-induced resistance to the antiproteolytic effect of insulin and an increase in the oxidation (and thus wasting) of one essential amino acid, leucine.     

Molecular mechanisms of insulin resistance in humans and their potential links with mitochondrial dysfunction

Recent studies using magnetic resonance spectroscopy have shown that decreased insulin-stimulated muscle glycogen synthesis due to a defect in insulin-stimulated glucose transport activity is a major factor in the pathogenesis of type 2 diabetes. The molecular mechanism underlying defective insulin-stimulated glucose transport activity can be attributed to increases in intramyocellular lipid metabolites such as fatty acyl CoAs and diacylglycerol, which in turn activate a serine/threonine kinase cascade, thus leading to defects in insulin signaling through Ser/Thr phosphorylation of insulin receptor substrate (IRS)-1. A similar mechanism is also observed in hepatic insulin resistance associated with nonalcoholic fatty liver, which is a common feature of type 2 diabetes, where increases in hepatocellular diacylglycerol content activate protein kinase C-epsilon, leading to reduced insulin-stimulated tyrosine phosphorylation of IRS-2. More recently, magnetic resonance spectroscopy studies in healthy lean elderly subjects and healthy lean insulin-resistant offspring of parents with type 2 diabetes have demonstrated that reduced mitochondrial function may predispose these individuals to intramyocellular lipid accumulation and insulin resistance. Further analysis has found that the reduction in mitochondrial function in the insulin-resistant offspring can be mostly attributed to reductions in mitochondrial density. By elucidating the cellular and molecular mechanisms responsible for insulin resistance, these studies provide potential new targets for the treatment and prevention of type 2 diabetes.     

Intracerebroventricular administration of neuropeptide Y induces hepatic insulin resistance via sympathetic innervation

OBJECTIVE—We recently showed that intracerebroventricular infusion of neuropeptide Y (NPY) hampers inhibition of endogenous glucose production (EGP) by insulin in mice. The downstream mechanisms responsible for these effects of NPY remain to be elucidated. Therefore, the aim of this study was to establish whether intracerebroventricular NPY administration modulates the suppressive action of insulin on EGP via hepatic sympathetic or parasympathetic innervation.RESEARCH DESIGN AND METHODS—The effects of a continuous intracerebroventricular infusion of NPY on glucose turnover were determined in rats during a hyperinsulinemic-euglycemic clamp. Either rats were sham operated, or the liver was sympathetically (hepatic sympathectomy) or parasympathetically (hepatic parasympathectomy) denervated.RESULTS—Sympathectomy or parasympathectomy did not affect the capacity of insulin to suppress EGP in intracerebroventricular vehicle–infused animals (50 ± 8 vs. 49 ± 6 vs. 55 ± 6%, in hepatic sympathectomy vs. hepatic parasympathectomy vs. sham, respectively). Intracerebroventricular infusion of NPY significantly hampered the suppression of EGP by insulin in sham-denervated animals (29 ± 9 vs. 55 ± 6% for NPY/sham vs. vehicle/sham, respectively, P = 0.038). Selective sympathetic denervation of the liver completely blocked the effect of intracerebroventricular NPY administration on insulin action to suppress EGP (NPY/hepatic sympathectomy, 57 ± 7%), whereas selective parasympathetic denervation had no effect (NPY/hepatic parasympathectomy, 29 ± 7%).CONCLUSIONS—Intracerebroventricular administration of NPY acutely induces insulin resistance of EGP via activation of sympathetic output to the liver.Since Claude Bernard first observed in the 1850s that puncture of the floor of the fourth cerebral ventricle elevates blood glucose levels, the fundamental role of the brain in the control of glucose metabolism has been firmly established (1,2). Intracerebroventricular infusion of insulin inhibits endogenous glucose production (EGP) (3), and downregulation of hypothalamic insulin receptors by antisense oligonucleotides precludes suppression of EGP by (circulating) insulin to a considerable extent (4). Thus, hypothalamic insulin signaling appears to play a role in the control of EGP. Insulin inhibits neuropeptide Y (NPY)-producing neurons in the arcuate nucleus of the hypothalamus (5). Intracerebroventricular administration of NPY hampers the capacity of insulin to suppress EGP (6), suggesting that silencing of arcuate NPY neurons may contribute to the inhibitory effect of intracerebroventricular insulin administration on EGP.The downstream mechanism responsible for the effects of hypothalamic NPY on hepatic fuel flux remains to be established. Arcuate NPY neurons project to the paraventricular nucleus (PVN) and various other hypothalamic nuclei (7). The hypothalamus is a major source of forebrain input into the sympathetic nervous system (8); it partakes in the control of cholinergic outflow to visceral organs (9,10), and it orchestrates the release of various pituitary hormones (11,12). Either of these neuroendocrine systems can impact on EGP (13–15).Here, we test the hypothesis that intracerebroventricular administration of NPY hampers the ability of insulin to suppress EGP via autonomic nervous inputs to the liver. To this end, rats whose livers were selectively stripped of sympathetic or parasympathetic nerves received intracerebroventricular infusion of NPY or vehicle. The capacity of insulin to inhibit EGP was quantified by hyperinsulinemic-euglycemic clamp.     

Activation of the hexosamine pathway by glucosamine in vivo induces insulin resistance of early postreceptor insulin signaling events in skeletal muscle.

To explore potential cellular mechanisms by which activation of the hexosamine pathway induces insulin resistance, we have evaluated insulin signaling in conscious fasted rats infused for 2-6 h with saline, insulin (18 mU x kg(-1) x min(-1)), or insulin and glucosamine (30 micromol x kg(-1) x min(-1)) under euglycemic conditions. Glucosamine infusion increased muscle UDP-N-acetylglucosamine concentrations 3.9- and 4.3-fold over saline- or insulin-infused animals, respectively (P < 0.001). Glucosamine induced significant insulin resistance to glucose uptake both at the level of the whole body and in rectus abdominis muscle, and it blunted the insulin-induced increase in muscle glycogen content. At a cellular level, these metabolic effects were paralleled by inhibition of postreceptor insulin signaling critical for glucose transport and glycogen storage, including a 45% reduction in insulin-stimulated insulin receptor substrate (IRS)-1 tyrosine phosphorylation (P = 0.02), a 44% decrease in IRS-1 association with the p85 regulatory subunit of phosphatidylinositol (PI) 3-kinase (P = 0.03), a 34% reduction in IRS-1-associated PI 3-kinase activity (P = 0.03), and a 51% reduction in insulin-stimulated glycogen synthase activity (P = 0.03). These alterations in postreceptor insulin signaling were time-dependent and paralleled closely the progressive inhibition of systemic glucose disposal from 2 to 6 h of glucosamine infusion. We also demonstrated that glucosamine infusion results in O-linked N-acetylglucosamine modification of IRS-1 and IRS-2. These data indicate that activation of the hexosamine pathway may directly modulate early postreceptor insulin signal transduction, perhaps via posttranslation modification of IRS proteins, and thus contribute to the insulin resistance induced by chronic hyperglycemia.     

Ultradian oscillations of insulin secretion in humans

Ultradian rhythmicity appears to be characteristic of several endocrine systems. As described for other hormones, insulin release is a multioscillatory process with rapid pulses of about 10 min and slower ultradian oscillations (50--120 min). The mechanisms underlying the ultradian circhoral oscillations of insulin secretion rate (ISR), which arise in part from a rhythmic amplification of the rapid pulses, are not fully understood. In humans, included in the same period range is the alternation of rapid eye movement (REM) and non-REM (NREM) sleep cycles and the associated opposite oscillations in sympathovagal balance. During sleep, the glucose and ISR oscillations were amplified by about 150%, but the REM-NREM sleep cycles did not entrain the glucose and ISR ultradian oscillations. Also, the latter were not related to either the ultradian oscillations in sympathoagal balance, as inferred from spectral analysis of cardiac R-R intervals, or the plasma fluctuations of glucagon-like peptide-1 (GLP-1), an incretin hormone known to potentiate glucose-stimulated insulin. Other rhythmic physiological processes are currently being examined in relation to ultradian insulin release.     

Development of muscle insulin resistance after liver insulin resistance in high-fat-fed rats

Muscle and hepatic insulin resistance are two major defects of non-insulin-dependent diabetes mellitus. Dietary factors may be important in the etiology of insulin resistance. We studied progressive changes in the development of high-fat-diet-induced insulin resistance in tissues of the adult male Wistar rat. In vivo insulin action was compared 3 days and 3 wk after isocaloric synthetic high-fat or high-starch feeding (59 and 10% cal as fat, respectively). Basal and insulin-stimulated glucose metabolism were assessed in the conscious 5- to 7-h fasted state with the euglycemic clamp (600 pM insulin) with a [3-3H]-glucose infusion. Fat feeding significantly reduced suppressibility of hepatic glucose output by insulin after both 3 days and 3 wk of diet (P less than 0.01). However, a significant impairment of insulin-mediated peripheral glucose disposal was only present after 3 wk of diet. Further in vivo [3H]-2-deoxyglucose uptake studies supported this finding and demonstrated adipose but not muscle insulin resistance after 3 days of high-fat feeding. Muscle triglyceride accumulation due to fat feeding was not significant at 3 days but had doubled by 3 wk in red muscle (P less than 0.001) compared with starch-fed controls. By 3 wk, high-fat-fed animals had developed significant glucose intolerance. We conclude that fat feeding induces insulin resistance in liver and adipose tissue before skeletal muscle with early metabolic changes favoring an oversupply of energy substrate to skeletal muscle relative to metabolic needs. This may generate later muscle insulin resistance.     

Pulsatile insulin secretion dictates systemic insulin delivery by regulating hepatic insulin extraction in humans

In health, insulin is secreted in discrete pulses into the portal vein, and the regulation of the rate of insulin secretion is accomplished by modulation of insulin pulse mass. Several lines of evidence suggest that the pattern of insulin delivery by the pancreas determines hepatic insulin clearance. In previous large animal studies, the amplitude of insulin pulses was related to the extent of insulin clearance. In humans (and in large animals), the amplitude of insulin oscillations is approximately 100-fold higher in the portal vein than in the systemic circulation, despite only a fivefold dilution, implying preferential hepatic extraction of insulin pulses. In the present study, by direct hepatic vein sampling in healthy humans, we sought to establish the extent of first-pass hepatic insulin extraction and to determine whether the pattern of insulin secretion (insulin pulse mass and amplitude) dictates the hepatic insulin clearance and thereby delivery of insulin to extrahepatic insulin-responsive tissues. Five nondiabetic subjects (two men and three women, mean age 32 years [range 25-39], BMI 24.9 kg/m(2) [21.2-27.1]) participated. Insulin and C-peptide delivery from the splanchnic bed was measured in basal overnight-fasted state and during a glucose infusion of 2 mg . kg(-1) . min(-1) by simultaneous sampling from the hepatic vein and an arterialized vein along with direct estimation of splanchnic blood flow. Fractional insulin extraction was calculated from the difference between the C-peptide and insulin delivery rates from the liver. The time patterns of insulin concentrations and hepatic insulin clearance were analyzed by deconvolution and Cluster analysis, respectively. Cross-correlation analysis was used to relate C-peptide secretion and insulin clearance. Glucose infusion increased peripheral glucose concentrations from 5.4 +/- 0.1 to 6.4 +/- 0.4 mmol/l (P < 0.05). Likewise, insulin and C-peptide concentrations increased during glucose infusion (P < 0.05). Hepatic insulin clearance increased with glucose infusion (1.06 +/- 0.18 vs. 2.55 +/- 0.38 pmol . kg(-1) . min(-1); P < 0.01), but fractional hepatic insulin clearance was stable (78.2 +/- 4.4 vs. 84 0. +/- 3.9%, respectively; P = 0.18). Insulin secretory-burst mass rose during glucose infusion (P < 0.05), whereas the interburst interval remained unchanged (4.4 +/- 0.2 vs. 4.5 +/- 0.3 min; P = 0.36). Cluster analysis identified an oscillatory pattern in insulin clearance, with peaks occurring approximately every 5 min. Cross-correlation analysis between prehepatic C-peptide secretion and hepatic insulin clearance demonstrated a significant positive association without detectable (<1 min) time lag. Insulin secretory-burst mass strongly predicted insulin clearance (r = 0.81, P = 0.0043). In conclusion, in humans, approximately 80% of insulin is extracted during the first liver passage. The liver rapidly responds to fluctuations in insulin secretion, preferentially extracting insulin delivered in pulses. The mass (and therefore amplitude) of insulin pulses traversing the liver is the predominant determinant of hepatic insulin clearance. Therefore, through this means, the pulse mass of insulin release dictates both hepatic (directly) as well as extra-hepatic (indirectly) insulin delivery. These findings emphasize the dual role of the liver and pancreas and their relationship mediated through magnitude of insulin pulse mass in regulating the quantity and pattern of systemic insulin delivery.