Merry T L, Dywer R M, Bradley E A, Rattigan S, McConell G K
Department of Physiology, The University of Melbourne, Parkville, Victoria 3010, Australia.
J Appl Physiol (1985). 2010 May;108(5):1275-83. doi: 10.1152/japplphysiol.01335.2009. Epub 2010 Mar 4.
There is evidence that reactive oxygen species (ROS) contribute to the regulation of skeletal muscle glucose uptake during highly fatiguing ex vivo contraction conditions via AMP-activated protein kinase (AMPK). In this study we investigated the role of ROS in the regulation of glucose uptake and AMPK signaling during low-moderate intensity in situ hindlimb muscle contractions in rats, which is a more physiological protocol and preparation. Male hooded Wistar rats were anesthetized, and then N-acetylcysteine (NAC) was infused into the epigastric artery (125 mg.kg(-1).h(-1)) of one hindlimb (contracted leg) for 15 min before this leg was electrically stimulated (0.1-ms impulse at 2 Hz and 35 V) to contract at a low-moderate intensity for 15 min. The contralateral leg did not receive stimulation or local NAC infusion (rest leg). NAC infusion increased (P<0.05) plasma cysteine and cystine (by approximately 360- and 1.4-fold, respectively) and muscle cysteine (by 1.5-fold, P=0.001). Although contraction did not significantly alter muscle tyrosine nitration, reduced (GSH) or oxidized glutathione (GSSG) content, S-glutathionylation of protein bands at approximately 250 and 150 kDa was increased (P<0.05) approximately 1.7-fold by contraction, and this increase was prevented by NAC. Contraction increased (P<0.05) skeletal muscle glucose uptake 20-fold, AMPK phosphorylation 6-fold, ACCbeta phosphorylation 10-fold, and p38 MAPK phosphorylation 60-fold, and the muscle fatigued by approximately 30% during contraction and NAC infusion had no significant effect on any of these responses. This was despite NAC preventing increases in S-glutathionylation with contraction. In conclusion, unlike during highly fatiguing ex vivo contractions, local NAC infusion during in situ low-moderate intensity hindlimb contractions in rats, a more physiological preparation, does not attenuate increases in skeletal muscle glucose uptake or AMPK signaling.
有证据表明，在高度疲劳的离体收缩条件下，活性氧（ROS）通过AMP激活的蛋白激酶（AMPK）参与调节骨骼肌葡萄糖摄取。在本研究中，我们调查了ROS在大鼠低-中等强度原位后肢肌肉收缩过程中对葡萄糖摄取和AMPK信号传导的调节作用，这是一种更接近生理状态的实验方案和准备工作。雄性带帽Wistar大鼠麻醉后，在一侧后肢（收缩腿）的腹主动脉中注入N-乙酰半胱氨酸（NAC，125 mg·kg⁻¹·h⁻¹）15分钟，然后对该腿进行电刺激（2 Hz和35 V的0.1-ms脉冲），使其以低-中等强度收缩15分钟。对侧腿不接受刺激或局部NAC注入（静止腿）。注入NAC使血浆半胱氨酸和胱氨酸增加（P<０.０５，分别增加约360倍和1.4倍），肌肉半胱氨酸增加（1.5倍，P = 0.001）。虽然收缩未显著改变肌肉酪氨酸硝化、还原型谷胱甘肽（GSH）或氧化型谷胱甘肽（GSSG）含量，但收缩使约250和150 kDa蛋白条带的S-谷胱甘肽化增加（P<0.05）约１.７倍，而NAC可阻止这种增加。收缩使骨骼肌葡萄糖摄取增加（P<0.05）20倍、AMPK磷酸化增加6倍、ACCβ磷酸化增加10倍、p38 MAPK磷酸化增加60倍，并且在收缩和NAC注入期间肌肉疲劳约3０％，但对这些反应均无显著影响。尽管NAC可阻止收缩引起的S-谷胱甘肽化增加，但仍出现上述结果。总之，与高度疲劳的离体收缩不同，在大鼠原位低-中等强度后肢收缩（一种更接近生理状态的准备工作）过程中局部注入NAC，并不会减弱骨骼肌葡萄糖摄取或AMPK信号传导的增加。
