Bihler I, McNevin S R, Sawh P C
Biochim Biophys Acta. 1985 Jan 18;844(1):9-18. doi: 10.1016/0167-4889(85)90227-7.
Cardiac myocytes were isolated from adult rat ventricles by a method which preserves their functional integrity, including long survival in physiological concentrations of Ca2+. Sarcolemmal glucose transport was assessed by measuring linear initial uptake rates of the nonmetabolized glucose analog 3-O-methyl-D-glucose. Transport was saturable and showed competition by D-glucose and other features of chemical and stereo-selectivity. Transport was stimulated by insulin in a dose-dependent manner, resulting in an almost 5-fold increase in Vmax, with little change in Km. Stimulation of 3-methylglucose transport by insulin was largely Ca2+-dependent. Omission of Ca2+ from the incubation medium caused a minor rise in basal 3-methylglucose uptake but the insulin-stimulated rise in Vmax was only 30%. The Ca2+ antagonist D600 also antagonized stimulation of hexose transport by insulin. In all the above respects, 3-methylglucose transport in myocytes is identical to that in intact heart muscle. In addition, the decrease in insulin response by Ca2+ omission was partially reversed by subsequent return to a Ca2+-containing medium. ATP levels remained stable in the absence of Ca2+, showing that the Ca2+ dependence did not reflect nonspecific cell damage.
采用一种能保持其功能完整性的方法，从成年大鼠心室中分离出心肌细胞，包括在生理浓度的Ca2+中长时间存活。通过测量非代谢性葡萄糖类似物3 - O - 甲基 - D - 葡萄糖的线性初始摄取率来评估肌膜葡萄糖转运。转运具有饱和性，并表现出对D - 葡萄糖的竞争性以及化学和立体选择性的其他特征。转运受到胰岛素的剂量依赖性刺激，导致Vmax几乎增加5倍，而Km变化很小。胰岛素对3 - 甲基葡萄糖转运的刺激在很大程度上依赖于Ca2+。从孵育培养基中去除Ca2+会导致基础3 - 甲基葡萄糖摄取略有增加，但胰岛素刺激的Vmax增加仅为30%。Ca2+拮抗剂D600也能拮抗胰岛素对己糖转运的刺激。在上述所有方面，心肌细胞中3 - 甲基葡萄糖的转运与完整心肌中的转运相同。此外，通过随后恢复到含Ca2+的培养基中，因去除Ca2+而导致的胰岛素反应降低部分得到逆转。在没有Ca2+的情况下，ATP水平保持稳定，表明Ca2+依赖性并不反映非特异性细胞损伤。
