Jucker B M, Rennings A J, Cline G W, Shulman G I
Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06520-8020, USA.
J Biol Chem. 1997 Apr 18;272(16):10464-73. doi: 10.1074/jbc.272.16.10464.
The effects of increased plasma free fatty acids (FFA) on insulin-dependent whole body glucose disposal, skeletal muscle glycolysis, glycogen synthesis, pyruvate versus FFA/ketone oxidation, and glucose 6-phosphate (Glu-6-P) were investigated in the awake rat. A control group (glycerol-infused) and high plasma FFA group (Liposyn-infused) were clamped at euglycemia (approximately 6 mM)-hyperinsulinemia (10 milliunits/kg/min) throughout the experiment (180-240 min). In the initial experiment, 13C NMR was used to observe [1-13C]glucose incorporation into [1-13C]glycogen in the rat hindlimb for glycogen synthesis calculations and into [3-13C]lactate and [3-13C]alanine for glycolytic flux calculations. These experiments were followed by 31P NMR measurements of Glu-6-P changes under identical conditions of the initial experiment. Plasma FFA concentrations were 2.25 +/- 0.36 and 0.20 +/- 0.03 mM in the high plasma FFA and control groups respectively (p < 0.0005). Glucose infusion rates (Ginf) decreased significantly in the Liposyn-infused rats (29.5 +/- 0.7 and 27.2 +/- 1.2 mg/kg/min for control and high plasma FFA group, respectively, at 15 min to 30.7 +/- 2.3 and 17.7 +/- 1.3 mg/kg/min, respectively, at the end of the experiment, p < 0.002). Glycogen synthesis rates were 163 +/- 32 and 104 +/- 17 nmol/g/min, and glycolytic rates were 57.9 +/- 8.0 and 19. 5 +/- 3.6 nmol/g/min (p < 0.002) in the control and high plasma FFA groups, respectively. The relative flux of pyruvate versus free fatty acids and ketones entering the tricarboxylic acid cycle was greater in the control (57 +/- 9%) versus high plasma FFA group (25 +/- 4%) (p < 0.005) as assessed by [4-13C]glutamate/[3-13C]lactate steady state isotopic enrichment measurements. Finally, Glu-6-P concentrations increased by 29.8 +/- 7.0 and 52.8 +/- 12.3% (p < 0. 05) in the control and high plasma FFA groups, respectively, above their basal concentrations by 180 min. In conclusion, we have demonstrated the ability to use in vivo NMR to elucidate the metabolic fate of glucose within skeletal muscle of an awake rat during a euglycemic-hyperinsulinemic clamp and increased levels of plasma FFA. These data suggest that increased concentrations of plasma FFA inhibit insulin-stimulated muscle glucose metabolism in the rat through inhibition of glycolysis.
在清醒大鼠中研究了血浆游离脂肪酸（FFA）增加对胰岛素依赖的全身葡萄糖处置、骨骼肌糖酵解、糖原合成、丙酮酸与FFA/酮氧化以及葡萄糖6-磷酸（Glu-6-P）的影响。在整个实验（180 - 240分钟）期间，将对照组（输注甘油）和高血浆FFA组（输注Liposyn）钳定在正常血糖（约6 mM）-高胰岛素血症（10毫单位/千克/分钟）状态。在最初的实验中，使用13C NMR观察大鼠后肢中[1-13C]葡萄糖掺入[1-13C]糖原以进行糖原合成计算，以及掺入[3-13C]乳酸和[3-13C]丙氨酸以进行糖酵解通量计算。这些实验之后，在与最初实验相同条件下进行31P NMR测量Glu-6-P的变化。高血浆FFA组和对照组的血浆FFA浓度分别为2.25±0.36和0.20±0.03 mM（p < 0.0005）。输注Liposyn的大鼠的葡萄糖输注速率（Ginf）显著降低（对照组在15分钟时为29.5±0.7毫克/千克/分钟，高血浆FFA组为27.2±1.2毫克/千克/分钟，实验结束时分别为30.7±2.3毫克/千克/分钟和17.7±1.3毫克/千克/分钟，p < 0.002）。对照组和高血浆FFA组的糖原合成速率分别为163±32和104±17纳摩尔/克/分钟，糖酵解速率分别为57.9±8.0和19.5±3.6纳摩尔/克/分钟（p < 0.002）。通过[4-13C]谷氨酸/[3-13C]乳酸稳态同位素富集测量评估，进入三羧酸循环的丙酮酸与游离脂肪酸和酮的相对通量在对照组（57±9%）高于高血浆FFA组（25±4%）（p < 0.005）。最后，在180分钟时，对照组和高血浆FFA组的Glu-6-P浓度分别比其基础浓度增加了29.8±7.0%和52.8±12.3%（p < 0.05）。总之，我们已经证明能够使用体内NMR来阐明清醒大鼠在正常血糖-高胰岛素钳夹和血浆FFA水平升高期间骨骼肌内葡萄糖的代谢命运。这些数据表明，血浆FFA浓度增加通过抑制糖酵解来抑制大鼠胰岛素刺激的肌肉葡萄糖代谢。
