bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of the microdialysis technique in the dog fat pad
Authors: Stallknecht Bente , Madsen Joop , Galbo Henrik , Bülow Jens
Published: December 2017

In the present study the microdialysis technique was evaluated in an isolated autoperfused dog fat pad. Concentrations of glucose, lactate, and glycerol were measured in interstitial fluid by microdialysis and simultaneously in arterial and adipose venous plasma. Adipose tissue blood flow was measured by both133Xe washout and timed weighing of venous blood. Metabolite concentrations in adipose venous plasma calculated from interstitial and arterial metabolite concentrations and133Xe washout were positively correlated with measured venous concentrations (glucose: r = 0.95, lactate: r = 0.92, glycerol: r = 0.81). Calculated and measured venous plasma concentrations did not differ for either glucose or lactate, but for glycerol, calculated concentration was on average 76% of measured concentration. Metabolite exchanges (Fick’s principle) calculated from interstitial metabolite concentrations were positively correlated with measured exchanges only for lactate ( r = 0.69). In conclusion, metabolite concentrations in adipose venous plasma can be calculated from microdialysis measurements with greater accuracy for glucose and lactate than for glycerol. The precision, however, is too low to allow calculation of metabolite exchange when arteriovenous metabolite differences are low.
Stimulation by 2-deoxy-d-glucose tetraacetates of hormonal secretion from the perfused rat pancreas
Authors: Leclercq-Meyer Viviane , Kadiata Marcel M. , Malaisse Willy J.
Published: December 2017

The effects of α- and β-2-deoxy-d-glucose tetraacetate (1.7 and 8.5 mM) on insulin, somatostatin, and glucagon secretion from isolated rat pancreases perfused in the presence of 8.3 mM d-glucose were compared with those of unesterified 2-deoxy-d-glucose tested at the same two concentrations. The unesterified glucose analog caused, in a concentration-related manner, inhibition of glucose-induced insulin and somatostatin release and augmentation of glucagon secretion. The two anomers of 2-deoxy-d-glucose tetraacetate, however, increased the secretion rate of all three hormones; this effect was also related to the concentration of the esters. No obvious anomeric specificity of the secretory response to 2-deoxy-d-glucose tetraacetate was observed. These findings indicate that the insulinotropic action of hexose esters cannot be accounted for solely by the metabolic effect of their glucidic moieties. They suggest that the A, B, and D cells of the endocrine pancreas are each equipped with a receptor system responsible for the direct recognition of monosaccharide esters as secretagogues. They further support the view that a paracrine effect of insulin on glucagon-producing cells does not represent a major component in the regulation of their secretory activity.
Insulin receptor autophosphorylation in cultured myoblasts correlates to glucose disposal in Pima Indians
Authors: Youngren Jack F. , Goldfine Ira D. , Pratley Richard E.
Published: December 2017

In a previous study [Youngren, J. F., I. D. Goldfire, and R. E. Pratley. Am. J. Physiol. 273 ( Endocrinol. Metab. 36): E276–E283, 1997] of skeletal muscle biopsies from insulin-resistant, nondiabetic Pima Indians, we demonstrated that diminished insulin receptor (IR) autophosphorylation correlated with in vivo insulin resistance. In the present study, to determine whether decreased IR function is a primary trait of muscle, and not secondary to an altered in vivo environment, we cultured myoblasts from 17 nondiabetic Pima Indians in whom insulin-stimulated glucose disposal (M) was measured during hyperinsulinemic-euglycemic glucose clamps. Myoblast IR autophosphorylation was determined by a highly sensitive ELISA. IR autophosphorylation directly correlated with M ( r = 0.56, P = 0.02) and inversely correlated with the fasting plasma insulin ( r = −0.58, P < 0.05). The relationship between M and IR autophosphorylation remained significant after M was adjusted for the effects of percent body fat (partial r = 0.53, P < 0.04). The relationship between insulin resistance and the capacity for myoblast IR autophosphorylation in nondiabetic Pima Indians suggests that variations in IR-signaling capacity may be intrinsic characteristics of muscle that contribute to the genetic component determining insulin action in this population.
Parenteral amino acid intake alters the anabolic actions of insulin-like growth factor I in rats
Authors: Kee Anthony J. , Baxter Robert C. , Carlsson Anthony R. , Smith Ross C.
Published: December 2017

The anabolic properties of insulin-like growth factor (IGF) I are attenuated by oral diets that are low in protein. However, it is not known whether parenteral nutrition (PN) providing a low amino acid (AA) input will influence IGF-I action. With the use of a rat model, this study examined the interaction between AA input (1.27 and 0.62 g N ⋅ kg body wt−1⋅ 24 h−1, AA and ½AA groups, respectively) and recombinant human IGF-I (rhIGF-I, 2.5 mg ⋅ kg body wt−1⋅ 24 h−1) infusion on the composition of the carcass and organs and on plasma insulin, IGF-I, IGF-binding protein 1 (IGFBP-1), and acid-labile subunit (ALS) concentrations. Carcass protein deposition only occurred in the AA groups ( P < 0.003) and was not influenced by administration of rhIGF-I. However, visceral protein loss persisted in the AA group but was prevented by rhIGF-I infusion. The changes in water content of the carcass and the organs were generally in the expected proportion of normal lean tissue. The accumulation of lipid that follows the infusion of the AA-deficient PN was prevented by rhIGF-I infusion, which may indicate an improved energy utilization. Neither serum insulin nor ALS concentrations were influenced by the level of AA infusion but were reduced by rhIGF-I administration. However, plasma IGF-I levels were elevated by higher AA infusion and by IGF-I administration. Also, IGFBP-1 concentrations were reduced by the higher AA infusion and increased with rhIGF-I administration. Interestingly, there was a significant interaction effect between both of these influences. It is concluded that free IGF-I concentration, which may be regulated by IGFBP-1 through a direct effect of AAs on the liver, may have an important role in regulating anabolism in visceral and possibly skeletal tissue during PN.
The ς-ligand (+)-pentazocine depresses M current and enhances calcium conductances in frog melanotrophs
Authors: Soriani O. , le Foll F. , Galas L. , Roman F. , Vaudry H. , Cazin L.
Published: December 2017

Gramicidin-perforated patch clamp experiments and microfluorimetric measurements were performed to study the ionic mechanisms involved in the ς-receptor-mediated stimulation of frog ( Rana ridibunda) pituitary melanotrophs. The ς-ligand (+)-pentazocine (50 μM) depressed a sustained outward K+current. The kinetic properties of this K+component, investigated by analyzing tail currents, were reminiscent of those of the M current ( IM), with an activation threshold close to −60 mV, a −21-mV half-maximal activation potential, and two-component exponential deactivation kinetics at −90 mV. (+)-Pentazocine (20 μM) produced a 12-mV rightward shift of the activation curve and accelerated the deactivation rate of the tail current. It is also demonstrated that (+)-pentazocine (20 μM) reversibly increased both voltage-dependent calcium conductances and internal calcium level. Altogether, these results suggest that the ς-receptor-induced modulation of IMand calcium currents likely underlies the increase of intracellular [Ca2+].
Differentiation-dependent inhibition of proteolysis by norepinephrine in brown adipocytes
Authors: Desautels M. , Heal S.
Published: December 2017

The objective was to evaluate whether norepinephrine (NE) and other hormonal factors have direct effects on protein degradation in brown fat cells. NE inhibited proteolysis by 35–45% in mouse brown adipocytes differentiated in culture. Insulin also inhibited protein degradation but significantly less than NE, whereas glucagon and leptin had no effect. The inhibitory effect of NE was partially antagonized by propranolol but not by prazosin, and dose-response curves with BRL-37344 (a β3-agonist), isoproterenol (a β1/β2-agonist) and dobutamide (a β1-agonist) were consistent with the involvement of a β3-adrenergic receptor. Furthermore, forskolin mimicked the effects of NE, whereas additions of A-23187 or phorbol esters had no effect, alone or in combination with NE or forskolin. Thus inhibition of proteolysis by NE likely involves a β3-adrenergic receptor-mediated increase in cAMP. In contrast, NE, BRL-37344, and dobutamide had no effect on proteolysis in preadipocytes. Inhibition of proteolysis by NE was due at least in part to inhibition of autophagy. Thus inhibition of proteolysis by NE and insulin in mature brown adipocytes is likely an important process contributing to brown fat growth and atrophy under many physiological or pathological conditions.
Mobilization of GLUT-4 from intracellular vesicles by insulin and K+depolarization in cultured H9c2 myotubes
Authors: Yu Bo , Poirier Laurie A. , Nagy Laura E.
Published: December 2017

The insulin-responsive glucose transporter, GLUT-4, moves from an intracellular compartment to the cell surface in response to insulin and/or muscle contraction. Treatment of H9c2 myotubes with insulin significantly increased uptake of 2-deoxyglucose. Depolarization of the myotubes by increasing extracellular [K+], which mimics the initial phases of excitation-contraction coupling, also increased 2-deoxyglucose uptake. The K+- but not insulin-evoked increase was blocked by dantrolene, an inhibitor of Ca2+release from the sarcoplasmic reticulum. In contrast, wortmannin, an inhibitor of phosphatidylinositol 3-kinase, blocked insulin- but not K+-stimulated 2-deoxyglucose uptake. Increased glucose uptake in response to insulin or K+depolarization was associated with increased GLUT-4 in plasma membranes and depletion of a population of small intracellular GLUT-4-containing vesicles. Similarly, in H9c2 cells transfected with c-myc-tagged GLUT-4, translocation of c-myc GLUT-4 to the cell surface was increased after stimulation with insulin or K+depolarization. Taken together, these data demonstrate that insulin and K+depolarization increase glucose uptake by recruiting GLUT-4 from intracellular vesicles to the plasma membrane of H9c2 myotubes via distinct signaling mechanisms.
T3stimulates resting metabolism and UCP-2 and UCP-3 mRNA but not nonphosphorylating mitochondrial respiration in mice
Authors: Jekabsons Mika B. , Gregoire Francine M. , Schonfeld-Warden Nancy A. , Warden Craig H. , Horwitz B. A.
Published: December 2017

The molecular basis for variations in resting metabolic rate (RMR) within a species is unknown. One possibility is that variations in RMR occur because of variations in uncoupling protein 2 (UCP-2) and uncoupling protein 3 (UCP-3) expression, resulting in mitochondrial proton leak differences. We tested the hypothesis that UCP-2 and -3 mRNAs positively correlate with RMR and proton leak. We treated thyroidectomized and sham-operated mice with triiodothyronine (T3) or vehicle and measured RMR, liver, and skeletal muscle mitochondrial nonphosphorylating respiration and UCP-2 and -3 mRNAs. T3stimulated RMR and liver UCP-2 and gastrocnemius UCP-2 and -3 expression. Mitochondrial respiration was not affected by T3and did not correlate with UCP-2 and -3 mRNAs. Gastrocnemius UCP-2 and -3 expression did correlate with RMR. We conclude 1) T3did not influence intrinsic mitochondrial properties such as membrane structure and composition, and 2) variations in UCP-2 and -3 expression may partly explain variations in RMR. One possible explanation for these data is that T3stimulates the leak in vivo but not in vitro because a posttranslational regulator of UCP-2 and -3 is not retained in the mitochondrial fraction.
Glutamate transport asymmetry and metabolism in the functioning kidney
Authors: Schuldt Scott , Carter Patsy , Welbourne Tomas
Published: December 2017

Renal glutamate extraction in vivo shows a preference for the uptake ofd-glutamate on the antiluminal and l-glutamate on the luminal tubule surface. To characterize this functional asymmetry, we isolated rat kidneys and perfused them with an artificial plasma solution containing either d- orl-glutamate alone or in combination with the system [Formula: see text]specific transport inhibitor,d-aspartate. To confirm that removal of glutamate represented transport into tubule cells, we monitored products formed as the result of intracellular metabolism and related these to the uptake process. Perfusion withd-glutamate alone resulted in a removal rate that equaled or exceeded thel-glutamate removal rate, with uptake predominantly across the antiluminal surface;l-glutamate uptake occurred nearly equally across both luminal and antiluminal surfaces. Thus the preferential uptake ofd-glutamate at the antiluminal and l-glutamate at the luminal surface confirms the transport asymmetry observed in vivo. Equimolard-aspartate concentration blocked most of the antiluminald-glutamate uptake and a significant portion of the luminall-glutamate uptake, consistent with system [Formula: see text] activity at both sites. d-Glutamate uptake was associated with 5-oxo-d-proline production, whereas l-glutamate uptake supported both glutamine and 5-oxo-l-proline formation;d-aspartate reduced production of both 5-oxoproline and glutamine. The presence of system[Formula: see text] activity on both the luminal and antiluminal tubule surfaces, exhibiting different reactivity towardl- andd-glutamate suggests that functional asymmetry may reflect two different[Formula: see text] transporter subtypes.
Metabolic control analysis of insulin-stimulated glucose disposal in rat skeletal muscle
Authors: Jucker Beat M. , Barucci Nicole , Shulman Gerald I.
Published: December 2017

Metabolic control analysis was used to calculate the distributed control of insulin-stimulated skeletal muscle glucose disposal in awake rats. Three separate hyperinsulinemic infusion protocols were performed: 1) protocol I was a euglycemic (∼6 mM)-hyperinsulinemic (10 mU ⋅ kg−1⋅ min−1) clamp, 2) protocol II was a hyperglycemic (∼11 mM)-hyperinsulinemic (10 mU ⋅ kg−1⋅ min−1) clamp, and 3) protocol III was a euglycemic (∼6 mM)-hyperinsulinemic (10 mU ⋅ kg−1⋅ min−1)-lipid/heparin (increased plasma free fatty acid) clamp. [1-13C]glucose was administered in all three protocols for a 3-h period, during which time [1-13C]glucose label incorporation into [1-13C]glycogen, [3-13C]lactate, and [3-13C]alanine was detected in the hindlimb of awake rats via13C-NMR. Combined steady-state and kinetic data were used to calculate rates of glycogen synthesis and glycolysis. Additionally, glucose 6-phosphate (G-6- P) was measured in the hindlimb muscles with the use of in vivo31P-NMR during the three infusion protocols. The clamped glucose infusion rates were 31.6 ± 2.9, 49.7 ± 1.0, and 24.0 ± 1.5 mg ⋅ kg−1⋅ min−1at 120 min in protocols I– III, respectively. Rates of glycolysis were 62.1 ± 10.3, 71.6 ± 11.8, and 19.5 ± 3.6 nmol ⋅ g−1⋅ min−1and rates of glycogen synthesis were 125 ± 15, 224 ± 23, and 104 ± 17 nmol ⋅ g−1⋅ min−1in protocols I– III, respectively. Insulin-stimulated G-6- Pconcentrations were 217 ± 8, 265 ± 12, and 251 ± 9 nmol/g in protocols I– III, respectively. A top-down approach to metabolic control analysis was used to calculate the distributed control among glucose transport/phosphorylation [GLUT-4/hexokinase (HK)], glycogen synthesis, and glycolysis from the metabolic flux and G-6- P data. The calculated values for the control coefficients ( C) of these three metabolic steps ([Formula: see text]= 0.55 ± 0.10,[Formula: see text]= 0.30 ± 0.06, and[Formula: see text] = 0.15 ± 0.02; where J is glucose disposal flux, and glycogen syn is glycogen synthesis) indicate that there is shared control of glucose disposal and that glucose transport/phosphorylation is responsible for the majority of control of insulin-stimulated glucose disposal in skeletal muscle.
Lack of skeletal muscle uncoupling protein 2 and 3 mRNA induction during fasting in type-2 diabetic subjects
Authors: Vidal Hubert , Langin Dominique , Andreelli Fabrizio , Millet Laurence , Larrouy Dominique , Laville Martine
Published: December 2017

Skeletal muscle uncoupling protein 2 and 3 (UCP-2 and UCP-3) mRNA levels are increased during calorie restriction in lean and nondiabetic obese subjects. In this work, we have investigated the effect of a 5-day hypocaloric diet (1,045 kJ/day) on UCP-2 and UCP-3 gene expression in the skeletal muscle of type-2 diabetic obese patients. Before the diet, UCP-2 and UCP-3 mRNA levels were more abundant in diabetic than in nondiabetic subjects. The long (UCP-3L) and short (UCP-3S) forms of UCP-3 transcripts were expressed at similar levels in nondiabetic subjects, but UCP-3S transcripts were twofold more abundant than UCP-3Ltranscripts in the muscle of diabetic patients. Calorie restriction induced a two- to threefold increase in UCP-2 and UCP-3 mRNA levels in nondiabetic patients. No change was observed in type-2 diabetic patients. Variations in plasma nonesterified fatty acid level were positively correlated with changes in skeletal muscle UCP-3L( r = 0.6, P < 0.05) and adipose tissue hormone-sensitive lipase ( r = 0.9, P < 0.001) mRNA levels. Lack of increase in plasma nonesterified fatty acid level and in hormone-sensitive lipase upregulation in diabetic patients during the diet strengthens the hypothesis that fatty acids are associated with the upregulation of uncoupling proteins during calorie restriction.
Regulation of skeletal muscle glycogen phosphorylase and PDH during maximal intermittent exercise
Authors: Parolin Michelle L. , Chesley Alan , Matsos Mark P. , Spriet Lawrence L. , Jones Norman L. , Heigenhauser George J. F.
Published: December 2017

The time course for the activation of glycogen phosphorylase (Phos) and pyruvate dehydrogenase (PDH) and their allosteric regulators was determined in human skeletal muscle during repeated bouts of maximal exercise. Six subjects completed three 30-s bouts of maximal isokinetic cycling separated by 4-min recovery periods. Muscle biopsies were taken at rest and at 6, 15, and 30 s of exercise during bouts 1 and 3. Phos was rapidly activated within the first 6 s of bout 1 from 12% at rest to 47% at 6 s. The activation of PDH increased from 14% at rest to 48% at 6 s and 95% at 15 s of bout 1. Phos reverted back to basal values at the end of the first bout, whereas PDH remained fully activated. In contrast, in the third bout, PDH was 42% at rest and was activated more rapidly and was nearly completely activated by 6 s, whereas Phos remained at basal levels (range 14–20%). Lactate accumulation was marked in the first bout and increased progressively from 2.7 to 76.1 mmol/kg dry wt with no further increase in bout 3. Glycogen utilization was also marked in the first bout and was negligible in bout 3. The rapid activation of Phos and slower activation of PDH in bout 1was probably due to Ca2+release from the sarcoplasmic reticulum. Lactate accumulation appeared to be due to an imbalance of the relative activities of Phos and PDH. The increase in H+concentration may have served to reduce pyruvate production by inhibiting Phos transformation and may have simultaneously activated PDH in the third bout such that there was a better matching between pyruvate production and oxidation and minimal lactate accumulation. As each bout progressed and with successive bouts, there was a decreasing ability to stimulate substrate phosphorylation through phosphocreatine hydrolysis and glycolysis and a shift toward greater reliance on oxidative phosphorylation.
Fetal-to-maternal transfer of 3,3′,5-triiodothyronine sulfate and its metabolite in sheep
Authors: Wu Sing-Yung , Polk Daniel H. , Huang Wen-Sheng , Fisher Delbert A.
Published: December 2017

Earlier studies have shown that sulfoconjugation is a major pathway of thyroid hormone metabolism in fetal mammals. To assess the placental transfer of sulfoconjugates in the pregnant sheep model, we measured 3,3′,5-triiodothyronine (T3) sulfate (T3S), 3,3′-diiodothyronine sulfate (T2S), and T3 concentrations in fetal serum and in maternal serum and urine after T3S infusion to the fetus ( n = 5) or the ewe ( n = 6). Maternal infusion of T3S did not increase fetal serum T2S, T3S, or T3 concentrations. In contrast, fetal infusion of T3S produced significant increases in maternal serum T2S and T3S but not T3 concentrations. Fetal T3S infusion also increased maternal urine excretion of T3S. However, the 4-h cumulative maternal urinary excretion of T2S and T3S after fetal T3S infusion was less than the excretion observed after fetal infusion of equimolar amounts of T3 in our previous study. It is concluded that fetal serum T2S and T3S can be transferred to maternal compartments. However, compared with T3, these sulfoconjugates may be less readily transferred.
High nocturnal body temperatures and disturbed sleep in women with primary dysmenorrhea
Authors: Baker Fiona C. , Driver Helen S. , Rogers Geoffrey G. , Paiker Janice , Mitchell Duncan
Published: December 2017

Primary dysmenorrhea is characterized by painful uterine cramps, near and during menstruation, that have an impact on personal life and productivity. The effect on sleep of this recurring pain has not been established. We compared sleep, nocturnal body temperatures, and hormone profiles during the menstrual cycle of 10 young women who suffered from primary dysmenorrhea, without any menstrual-associated mood disturbances, and 8 women who had normal menstrual cycles. Dysmenorrheic pain significantly decreased subjective sleep quality, sleep efficiency, and rapid eye movement (REM) sleep but not slow wave sleep (SWS), compared with pain-free phases of the menstrual cycle and compared with the controls. Even before menstruation, in the absence of pain, the women with dysmenorrhea had different sleep patterns, nocturnal body temperatures, and hormone levels compared with the controls. In the mid-follicular, mid-luteal, and menstrual phases, the dysmenorrheics had elevated morning estrogen concentrations, higher mean in-bed temperatures, and less REM sleep compared with the controls, as well as higher luteal phase prolactin levels. Both groups of women had less REM sleep when their body temperatures were high during the luteal and menstrual phases, implying that REM sleep is sensitive to elevated body temperatures. We have shown that dysmenorrhea is not only a disorder of menstruation but is manifest throughout the menstrual cycle. Furthermore, dysmenorrheic pain disturbs sleep, which may exacerbate the effect of the pain on daytime functioning.
Effect of high-altitude acclimation on NEFA turnover and lipid utilization during exercise in rats
Authors: McClelland Grant B. , Hochachka Peter W. , Weber Jean-Michel
Published: December 2017

Relative exercise intensity (or %maximum O2consumption,V˙o2 max) controls fuel selection at sea level (SL) and after high-altitude acclimation (HA) in rats. In this context we used indirect calorimetry, [1-14C]palmitate infusions, and muscle triacylglycerol (TAG) measurements to determine 1) total lipid oxidation, 2) the relationship between circulatory nonesterified fatty acid (NEFA) flux and concentration, and 3) muscle TAG depletion after exercise in HA-acclimated rats. Aerobic capacity is decreased in trained rats after 10 wk of acclimation. Both SL and HA showed the same relative use of lipids at 60% [62 ± 5% (HA) and 61 ± 3% (SL) of O2consumption (V˙o2)] and 80% [46 ± 6% (HA) and 47 ± 5% (SL) ofV˙o2] of their respective V˙o2 max. At 60% V˙o2 max, plasma [NEFA] were higher in HA, but rate of appearance was essentially the same in both groups (at 30 min, 38 ± 9 vs. 49 ± 6 μmol ⋅ kg−1⋅ min−1in HA and SL, respectively). At this intensity SL showed no significant decrease in muscle TAG, but in HA it decreased by 64% in soleus and by 90% in red gastrocnemius. We conclude that 1) the relative contributions of total lipid are the same in SL and HA, contrary to differences in [NEFA], because the relationship between flux rate and [NEFA] is modified after acclimation, and 2) muscle TAG may play a more important role at HA.
The adipoinsular axis: effects of leptin on pancreatic β-cells
Authors: Kieffer Timothy J. , Habener Joel F.
Published: December 2017

The prevalence of obesity and related diabetes mellitus is increasing worldwide. Here we review evidence for the existence of an adipoinsular axis, a dual hormonal feedback loop involving the hormones insulin and leptin produced by pancreatic β-cells and adipose tissue, respectively. Insulin is adipogenic, increases body fat mass, and stimulates the production and secretion of leptin, the satiety hormone that acts centrally to reduce food intake and increase energy expenditure. Leptin in turn suppresses insulin secretion by both central actions and direct actions on β-cells. Because plasma levels of leptin are directly proportional to body fat mass, an increase of adiposity increases plasma leptin, thereby curtailing insulin production and further increasing fat mass. We propose that the adipoinsular axis is designed to maintain nutrient balance and that dysregulation of this axis may contribute to obesity and the development of hyperinsulinemia associated with diabetes.
Glucosamine-induced insulin resistance in 3T3-L1 adipocytes
Authors: Heart Emma , Choi Woo S. , Sung Chin K.
Published: December 2017

To study molecular mechanisms for glucosamine-induced insulin resistance, we induced complete and reversible insulin resistance in 3T3-L1 adipocytes with glucosamine in a dose- and time-dependent manner (maximal effects at 50 mM glucosamine after 6 h). In these cells, glucosamine impaired insulin-stimulated GLUT-4 translocation. Glucosamine (6 h) did not affect insulin-stimulated tyrosine phosphorylation of the insulin receptor and insulin receptor substrate-1 and -2 and weakly, if at all, impaired insulin stimulation of phosphatidylinositol 3-kinase. Glucosamine, however, severely impaired insulin stimulation of Akt. Inhibition of insulin-stimulated glucose transport was correlated with that of Akt activity. In these cells, glucosamine also inhibited insulin stimulation of p70 S6 kinase. Glucosamine did not alter basal glucose transport and insulin stimulation of GLUT-1 translocation and mitogen-activated protein kinase. In summary, glucosamine induced complete and reversible insulin resistance in 3T3-L1 adipocytes. This insulin resistance was accompanied by impaired insulin stimulation of GLUT-4 translocation and Akt activity, without significant impairment of upstream molecules in insulin-signaling pathway.
Estimation of doubly labeled water energy expenditure with confidence intervals
Authors: Jones Richard H. , Sonko Bakary J. , Miller Leland V. , Thureen Patti J. , Fennessey Paul V.
Published: December 2017

Bivariate regression is used to estimate energy expenditure from doubly labeled water data. Two straight lines are fitted to the logarithms of the enrichments of oxygen-18 and deuterium simultaneously as a bivariate regression, so that the correlations between the oxygen and deuterium regression coefficients can be estimated. Maximum likelihood methods are used to extend bivariate regression to unbalanced situations caused by missing observations and to include replicate laboratory determination from the same urine samples, even if one of the replicates is missing. Use of maximum likelihood allows the determination of a confidence interval for the energy expenditure based on the log likelihood surface rather than use of the propagation of variance methods for nonlinear transformations. The model is extended to include the subject's deviations from the two lines as a bivariate continuous-time first-order autoregression to allow for serial correlation in the observations. The analysis of data from two subjects, one without apparent serial correlation and one with serial correlation, is presented.
Hormone-independent activation of EGP during hypoglycemia is absent in type 1 diabetes mellitus
Authors: Mevorach Michèle , Kaplan Jonathan , Chang Chee Jen , Rossetti Luciano , Shamoon Harry
Published: December 2017

It has been suggested that insulin-induced suppression of endogenous glucose production (EGP) may be counteracted independently of increased epinephrine (Epi) or glucagon during moderate hypoglycemia. We examined EGP in nondiabetic ( n = 12) and type 1 diabetic (DM1, n = 8) subjects while lowering plasma glucose (PG) from clamped euglycemia (5.6 mmol/l) to values just above the threshold for Epi and glucagon secretion (3.9 mmol/l). Individualized doses of insulin were infused to maintain euglycemia during pancreatic clamps by use of somatostatin (250 μg/h), glucagon (1.0 ng ⋅ kg−1⋅ min−1), and growth hormone (GH) (3.0 ng ⋅ kg−1⋅ min−1) infusions without need for exogenons glucose. Then, to achieve physiological hyperinsulinemia (HIns), insulin infusions were fixed at 20% above the rate previously determined for each subject. In nondiabetic subjects, PG was reduced from 5.4 ± 0.1 mmol/l to 3.9 ± 0.1 mmol/l in the experimental protocol, whereas it was held constant (5.3 ± 0.2 mmol/l and 5.5 mmol/l) in control studies. In the latter, EGP (estimated by [3-3H]glucose) fell to values 40% of basal ( P < 0.01). In contrast, in the experimental protocol, at comparable HIns but with PG at 3.9 ± 0.1 mmol/l, EGP was activated to values about twofold higher than in the euglycemic control ( P < 0.01). In DM1 subjects, EGP failed to increase in the face of HIns and PG = 3.9 ± 0.1 mmol/l. The decrease from basal EGP in DM1 subjects (4.4 ± 1.0 μmol ⋅ kg−1⋅ min−1) was nearly twofold that in nondiabetics (2.5 ± 0.8 μmol ⋅ kg−1⋅ min−1, P < 0.02). When PG was lowered further to frank hypoglycemia (∼3.1 mmol/l), the failure of EGP activation in DM1 subjects was even more profound but associated with a 50% lower plasma Epi response ( P < 0.02) compared with nondiabetics. We conclude that glucagon- or epinephrine-independent activation of EGP may accompany other counterregulatory mechanisms during mild hypoglycemia in humans and is impaired or absent in DM1.
Contribution of changes in the chloride driving force to the fading ofIGABAin frog melanotrophs
Authors: le Foll Frank , Soriani Olivier , Vaudry Hubert , Cazin Lionel
Published: December 2017

Chloride redistribution during type A γ-aminobutyric acid (GABAA) currents ( IGABA) has been investigated in cultured frog pituitary melanotrophs with imposed intracellular chloride concentration ([Cl−]i) in the whole cell configuration or with unaltered [Cl−]iusing the gramicidin-perforated patch approach. Prolonged GABA exposures elicited reproducible decaying currents. The decay of IGABAwas associated with both a transient fall of conductance ( gGABA) and shift of current reversal potential ( EGABA). The shift of EGABAappeared to be time and driving force dependent. In the gramicidin-perforated patch configuration, repeated GABA exposures induced currents that gradually vanished. The fading of IGABAwas due to persistent shifts of EGABAas a result of gGABArecovering from one GABA application to another. In cells alternatively clamped at potentials closely flanking resting potential and submitted to a train of brief GABA pulses, a reversal of IGABAwas observed after 150 s recording. It is demonstrated that, in intact frog melanotrophs, shifts of EGABAcombine with genuine receptor desensitization to depress IGABA. These findings strongly suggest that shifts of EGABAmay act as a negative feedback, reducing the bioelectrical and secretory responses induced by an intense release of GABA in vivo.
Catabolism of arginine and ornithine in the perfused rat liver: effect of dietary protein and of glucagon
Authors: O'Sullivan Dan , Brosnan John T. , Brosnan Margaret E.
Published: December 2017

The rates of oxidation of arginine and ornithine that occurred through a reaction pathway involving the enzyme ornithine aminotransferase (EC 2.6.1.13 ) were determined using14C-labeled amino acids in the isolated nonrecirculating perfused rat liver. At physiological concentrations of these amino acids, their catabolism is subject to chronic regulation by the level of protein consumed in the diet.14CO2production from [U-14C]ornithine (0.1 mM) and from [U-14C]arginine (0.2 mM) was increased about fourfold in livers from rats fed 60% casein diets for 3–4 days. The catabolism of arginine in the perfused rat liver, but not that of ornithine, is subject to acute regulation by glucagon (10−7M), which stimulated arginine catabolism by ∼40%. Dibutyryl cAMP (0.1 mM) activated arginine catabolism to a similar extent. In retrograde perfusions, glucagon caused a twofold increase in the rate of arginine catabolism, suggesting an effect of glucagon on arginase in the perivenous cells.
Regulation of glycogen phosphorylase and PDH during exercise in human skeletal muscle during hypoxia
Authors: Parolin Michelle L. , Spriet Lawrence L. , Hultman Eric , Hollidge-Horvat Melanie G. , Jones Norman L. , Heigenhauser George J. F.
Published: December 2017

The present study examined the acute effects of hypoxia on the regulation of skeletal muscle metabolism at rest and during 15 min of submaximal exercise. Subjects exercised on two occasions for 15 min at 55% of their normoxic maximal oxygen uptake while breathing 11% O2(hypoxia) or room air (normoxia). Muscle biopsies were taken at rest and after 1 and 15 min of exercise. At rest, no effects on muscle metabolism were observed in response to hypoxia. In the 1st min of exercise, glycogenolysis was significantly greater in hypoxia compared with normoxia. This small difference in glycogenolysis was associated with a tendency toward a greater concentration of substrate, free Pi, in hypoxia compared with normoxia. Pyruvate dehydrogenase activity (PDHa) was lower in hypoxia at 1 min compared with normoxia, resulting in a reduced rate of pyruvate oxidation and a greater lactate accumulation. During the last 14 min of exercise, glycogenolysis was greater in hypoxia despite a lower mole fraction of phosphorylase a. The greater glycogenolytic rate was maintained posttransformationally through significantly higher free [AMP] and [Pi]. At the end of exercise, PDHawas greater in hypoxia compared with normoxia, contributing to a greater rate of pyruvate oxidation. Because of the higher glycogenolytic rate in hypoxia, the rate of pyruvate production continued to exceed the rate of pyruvate oxidation, resulting in significant lactate accumulation in hypoxia compared with no further lactate accumulation in normoxia. Hence, the elevated lactate production associated with hypoxia at the same absolute workload could in part be explained by the effects of hypoxia on the activities of the rate-limiting enzymes, phosphorylase and PDH, which regulate the rates of pyruvate production and pyruvate oxidation, respectively.
Combined intraportal infusion of acetylcholine and adrenergic blockers augments net hepatic glucose uptake
Authors: Shiota Masakazu , Jackson Patricia , Galassetti Pietro , Scott Melanie , Neal Doss W. , Cherrington Alan D.
Published: December 2017

Portal glucose delivery in the conscious dog augments net hepatic glucose uptake (NHGU). To investigate the possible role of altered autonomic nervous activity in the effect of portal glucose delivery, the effects of adrenergic blockade and acetylcholine (ACh) on hepatic glucose metabolism were examined in 42-h-fasted conscious dogs. Each study consisted of an equilibration (−120 to −20 min), a control (−20 to 0 min), and a hyperglycemic-hyperinsulinemic period (0 to 300 min). During the last period, somatostatin (0.8 μg ⋅ kg−1⋅ min−1) was infused along with intraportal insulin (1.2 mU ⋅ kg−1⋅ min−1) and glucagon (0.5 ng ⋅ kg−1⋅ min−1). Hepatic sinusoidal insulin was four times basal (73 ± 7 μU/ml) and glucagon was basal (55 ± 7 pg/ml). Glucose was infused peripherally (0–300 min) to create hyperglycemia (220 mg/dl). In test protocol, phentolamine and propranolol were infused intraportally at 0.2 μg and 0.1 μg ⋅ kg−1⋅ min−1from 120 min on. ACh was infused intraportally at 3 μg ⋅ kg−1⋅ min−1from 210 min on. In control protocol, saline was given in place of the blockers and ACh. Hyperglycemia-hyperinsulinemia switched the net hepatic glucose balance (mg ⋅ kg−1⋅ min−1) from output (2.1 ± 0.3 and 1.1 ± 0.2) to uptake (2.8 ± 0.9 and 2.6 ± 0.6) and lactate balance (μmol ⋅ kg−1⋅ min−1) from uptake (7.5 ± 2.2 and 6.7 ± 1.6) to output (3.7 ± 2.6 and 3.9 ± 1.6) by 120 min in the control and test protocols, respectively. Therefter, in the control protocol, NHGU tended to increase slightly (3.0 ± 0.6 mg ⋅ kg−1⋅ min−1by 300 min). In the test protocol, adrenergic blockade did not alter NHGU, but ACh infusion increased it to 4.4 ± 0.6 and 4.6 ± 0.6 mg ⋅ kg−1⋅ min−1by 220 and 300 min, respectively. These data are consistent with the hypothesis that alterations in nerve activity contribute to the increase in NHGU seen after portal glucose delivery.
Tolbutamide and diazoxide modulate phospholipase C-linked Ca2+signaling and insulin secretion in β-cells
Authors: Schöfl Christof , Börger Julia , Mader Thilo , Waring Mark , von zur Mühlen Alexander , Brabant Georg
Published: December 2017

Arginine vasopressin (AVP), bombesin, and ACh increase cytosolic free Ca2+and potentiate glucose-induced insulin release by activating receptors linked to phospholipase C (PLC). We examined whether tolbutamide and diazoxide, which close or open ATP-sensitive K+channels (KATPchannels), respectively, interact with PLC-linked Ca2+signals in HIT-T15 and mouse β-cells and with PLC-linked insulin secretion from HIT-T15 cells. In the presence of glucose, the PLC-linked Ca2+signals were enhanced by tolbutamide (3–300 μM) and inhibited by diazoxide (10–100 μM). The effects of tolbutamide and diazoxide on PLC-linked Ca2+signaling were mimicked by BAY K 8644 and nifedipine, an activator and inhibitor of L-type voltage-sensitive Ca2+channels, respectively. Neither tolbutamide nor diazoxide affected PLC-linked mobilization of internal Ca2+or store-operated Ca2+influx through non-L-type Ca2+channels. In the absence of glucose, PLC-linked Ca2+signals were diminished or abolished; this effect could be partly antagonized by tolbutamide. In the presence of glucose, tolbutamide potentiated and diazoxide inhibited AVP- or bombesin-induced insulin secretion from HIT-T15 cells. Nifedipine (10 μM) blocked both the potentiating and inhibitory actions of tolbutamide and diazoxide on AVP-induced insulin release, respectively. In glucose-free medium, AVP-induced insulin release was reduced but was again potentiated by tolbutamide, whereas diazoxide caused no further inhibition. Thus tolbutamide and diazoxide regulate both PLC-linked Ca2+signaling and insulin secretion from pancreatic β-cells by modulating KATPchannels, thereby determining voltage-sensitive Ca2+influx.
Mechanism of muscle glycogen autoregulation in humans
Authors: Laurent Didier , Hundal Ripudaman S. , Dresner Alan , Price Thomas B. , Vogel Suzanne M. , Petersen Kitt Falk , Shulman Gerald I.
Published: December 2017

To examine the mechanism by which muscle glycogen limits its own synthesis, muscle glycogen and glucose 6-phosphate (G-6- P) concentrations were measured in seven healthy volunteers during a euglycemic (∼5.5 mM)-hyperinsulinemic (∼450 pM) clamp using13C/31P nuclear magnetic resonance spectroscopy before and after a muscle glycogen loading protocol. Rates of glycogen synthase ( Vsyn) and phosphorylase ( Vphos) flux were estimated during a [1-13C]glucose (pulse)-unlabeled glucose (chase) infusion. The muscle glycogen loading protocol resulted in a 65% increase in muscle glycogen content that was associated with a twofold increase in fasting plasma lactate concentrations ( P < 0.05 vs. basal) and an ∼30% decrease in plasma free fatty acid concentrations ( P < 0.001 vs. basal). Muscle glycogen loading resulted in an ∼30% decrease in the insulin-stimulated rate of net muscle glycogen synthesis ( P < 0.05 vs. basal), which was associated with a twofold increase in intramuscular G-6- P concentration ( P < 0.05 vs. basal). Muscle glycogen loading also resulted in an ∼30% increase in whole body glucose oxidation rates ( P < 0.05 vs. basal), whereas there was no effect on insulin-stimulated rates of whole body glucose uptake (∼10.5 mg ⋅ kg body wt−1⋅ min−1for both clamps) or glycogen turnover ( Vsyn/ Vphoswas ∼23% for both clamps). In conclusion, these data are consistent with the hypothesis that glycogen limits its own synthesis through feedback inhibition of glycogen synthase activity, as reflected by an accumulation of intramuscular G-6- P, which is then shunted into aerobic and anaerobic glycolysis.
Islet amyloid polypeptide (amylin)-deficient mice develop a more severe form of alloxan-induced diabetes
Authors: Mulder Hindrik , Gebre-Medhin Samuel , Betsholtz Christer , Sundler Frank , Ahrén Bo
Published: December 2017

To examine whether islet amyloid polypeptide (IAPP), other than through amyloid formation, may be of importance in diabetes pathogenesis, IAPP-deficient mice (IAPP−/−) were challenged with alloxan ( day 0). Diabetes in IAPP−/−mice was more severe at day 35, indicated by greater weight loss; glucose levels were higher in alloxan-treated IAPP−/−mice, whereas insulin levels were lower, indicating a greater impairment of islet function. Accordingly, glucose levels upon intravenous glucose challenges at days 7 and 35 were consistently higher in alloxan-treated IAPP−/−mice. At day 35, insulin mRNA expression, but not β-cell mass, was lower in untreated IAPP−/−mice. Yet, upon alloxan administration, β-cell mass and numbers of β-cell-containing islets were significantly more reduced in IAPP−/−mice. Furthermore, they displayed exaggerated β-cell dysfunction, because in their remaining β-cells, insulin mRNA expression was significantly more impaired and the localization of glucose transporter-2 was perturbed. Thus the lack of IAPP has allowed exaggerated β-cell cytotoxic actions of alloxan, suggesting that there may be beneficial features of IAPP actions in situations of β-cell damage.
Kinetics of l-[1-13C]leucine when ingested with free amino acids, unlabeled or intrinsically labeled casein
Authors: Metges Cornelia C. , El-Khoury Antoine E. , Selvaraj Ambalini B. , Tsay Rita H. , Atkinson Alan , Regan Meredith M. , Bequette Brian J. , Young Vernon R.
Published: December 2017

In two groups of five adults, each adapted to two different dietary regimens for 6 days, the metabolic fate of dietary [1-13C]leucine was examined when ingested either together with a mixture of free amino acids simulating casein (extrinsically labeled; condition A), along with the intact casein (extrinsically labeled; condition B), or bound to casein (intrinsically labeled; condition C). Fed state leucine oxidation (Ox), nonoxidative leucine disposal (NOLD), protein breakdown, and splanchnic uptake have been compared using an 8-h oral [1-13C]leucine and intravenous [2H3]leucine tracer protocol while giving eight equal hourly mixed meals. Lower leucine Ox, increased NOLD, and net protein synthesis were found with condition Ccompared with condition A (19.3 vs. 24.9; 77 vs. 55.8; 18.9 vs. 12.3 μmol ⋅ kg−1 ⋅ 30 min−1; P < 0.05). Ox and NOLD did not differ between conditions B and C. Splanchnic leucine uptake calculated from [1-13C]- and [2H3]leucine plasma enrichments was between 24 and 35%. These findings indicate that the form in which leucine is consumed affects its immediate metabolic fate and retention by the body; the implications of these findings for the tracer balance technique and estimation of amino acid requirements are discussed.
Regulation of MAP kinase pathway activity in vivo in human skeletal muscle
Authors: Osman Abdullah A. , Pendergrass Merri , Koval Janice , Maezono Katsumi , Cusi Kenneth , Pratipanawatr Thongchai , Mandarino Lawrence J.
Published: December 2017

Insulin and exercise potently stimulate glucose metabolism and gene transcription in vivo in skeletal muscle. A single bout of exercise increases the rate of insulin-stimulated glucose uptake and metabolism in skeletal muscle in the postexercise period. The nature of the intracellular signaling mechanisms that control responses to exercise is not known. In mammalian tissues, numerous reports have established the existence of the mitogen-activated protein (MAP) kinase signaling pathway that is activated by a variety of growth factors and hormones. This study was undertaken to determine how a single bout of exercise and physiological hyperinsulinemia activate the MAP kinase pathway. The euglycemic-hyperinsulinemic clamp and cycle ergometer exercise techniques combined with percutaneous muscle biopsies were used to answer this question. In healthy subjects, within 30 min, insulin significantly increased MAP kinase [isoforms p42MAPKand p44MAPK(ERK1 and ERK2)] phosphorylation (141 ± 2%, P < 0.05) and activity (177 ± 5%, P < 0.05), and the activity of its upstream activator MEK1 (161 ± 16%, P < 0.05). Insulin also increased the activity of the MAP kinase downstream substrate, the p90 ribosomal S6 kinase 2 (RSK2) almost twofold (198 ± 45%, P < 0.05). In contrast, a single 30-min bout of moderate-intensity exercise had no effect on the MAP kinase pathway activation from MEK to RSK2 in muscle of healthy subjects. However, 60 min of exercise did increase extracellular signal-related kinase activity. Therefore, despite similar effects on glucose metabolism after 30 min, insulin and exercise regulate the MAP kinase pathway differently. Insulin more rapidly activates the MAP kinase pathway.
Hepatic glucose metabolism during intraduodenal glucose infusion: impact of infection
Authors: McGuinness Owen P. , Ejiofor Joseph , Lacy D. Brooks , Schrom Nancy
Published: December 2017

We previously reported that infection decreases hepatic glucose uptake when glucose is given as a constant peripheral glucose infusion (8 mg · kg−1· min−1). This impairment persisted despite greater hyperinsulinemia in the infected group. In a normal setting, hepatic glucose uptake can be further enhanced if glucose is given gastrointestinally. Thus the aim of this study was to determine whether hepatic glucose uptake is impaired during an infection when glucose is given gastrointestinally. Thirty-six hours before study, a sham (SH, n = 7) or Escherichia coli-containing (2 × 109organisms/kg; INF; n = 7) fibrin clot was placed in the peritoneal cavity of chronically catheterized dogs. After the 36 h, a glucose bolus (150 mg/kg) followed by a continuous infusion (8 mg · kg−1· min−1) of glucose was given intraduodenally to conscious dogs for 240 min. Tracer ([3-3H]glucose and [U-14C]glucose) and arterial-venous difference techniques were used to assess hepatic and intestinal glucose metabolism. Infection increased hepatic blood flow (35 ± 5 vs. 47 ± 3 ml · kg−1· min−1; SH vs. INF) and basal glucose rate of appearance (2.1 ± 0.2 vs. 3.3 ± 0.1 mg · kg−1· min−1). Arterial insulin concentrations increased similarly in SH and INF during the last hour of glucose infusion (38 ± 8 vs. 46 ± 20 μU/ml), and arterial glucagon concentrations fell (62 ± 14 to 30 ± 3 vs. 624 ± 191 to 208 ± 97 pg/ml). Net intestinal glucose absorption was decreased in INF, attenuating the increase in blood glucose caused by the glucose load. Despite this, net hepatic glucose uptake (1.6 ± 0.8 vs. 2.4 ± 0.9 mg · kg−1· min−1; SH vs. INF) and consequently tracer-determined glycogen synthesis (1.3 ± 0.3 vs. 1.0 ± 0.3 mg · kg−1· min−1) were similar between groups. In summary, infection impairs net glucose absorption, but not net hepatic glucose uptake or glycogen deposition, when glucose is given intraduodenally.