Nakazaki Mitsuhiro, Kakei Masafumi, Ishihara Hisamitsu, Koriyama Nobuyuki, Hashiguchi Hiroshi, Aso Katsumi, Fukudome Michiyo, Oka Yoshitomo, Yada Toshihiko, Tei Chuwa
First Department of Internal Medicine, Faculty of Medicine, Kagoshima University, Kagoshima 890-8520, Japan.
J Physiol. 2002 May 1;540(Pt 3):781-9. doi: 10.1113/jphysiol.2001.013048.
Insulin-secreting MIN6 cells overexpressing uncoupling protein-1 (UCP1) were studied regarding insulin secretion in response to various secretagogues. Overexpression of UCP1 prevented an increase of cytosolic ATP levels induced by glucose. In contrast, glucose utilization was not affected, nor was glycerol phosphate flux. The UCP1-expressing cells showed an inability to increase cytosolic Ca(2+) concentration (Ca(2+)) in response to glucose or alpha ketoisocaproate and this resulted in less insulin secretion, whereas initial reduction in Ca(2+) occurring upon either nutrient addition was not affected. Moreover, the effectiveness of tolbutamide on Ca(2+) increase was reduced and the dose-response relations for insulin secretion induced by the agent was shifted toward the right in the UCP1-expressing cells. The resting membrane potential of the UCP1-expressing cells was significantly hyperpolarized by 6.2 mV compared with control cells. In the perforated and conventional whole-cell patch-clamp configurations, the conductance density of ATP-sensitive K(+) (K(ATP)) channels of the UCP1-expressing cells was 6-fold and 1.7-fold greater than that of the control cells, respectively. The sensitivity of K(ATP) channels for tolbutamide was not different between two groups, indicating that in intact cells more than 6-fold higher concentrations of tolbutamide were required to reduce the K(ATP) channel currents of UCP1-expressing cells to the same levels as of the control cells. The current density of the voltage-dependent Ca(2+) channels was not influenced. In conclusion, UCP1-expressing cells showed a refractoriness to respond to tolbutamide as well as nutrients. An upregulated activity of K(ATP) channels was associated with unresponsiveness to the agent in the cells with impaired mitochondrial function.
研究了过表达解偶联蛋白-1（UCP1）的胰岛素分泌MIN6细胞对各种促分泌剂的胰岛素分泌情况。UCP1的过表达阻止了葡萄糖诱导的胞质ATP水平升高。相比之下，葡萄糖利用未受影响，磷酸甘油通量也未受影响。表达UCP1的细胞在响应葡萄糖或α-酮异己酸时无法增加胞质Ca(2+)浓度（Ca(2+)），这导致胰岛素分泌减少，而在添加任何一种营养物质时最初发生的Ca(2+)降低不受影响。此外，在表达UCP1的细胞中，甲苯磺丁脲对Ca(2+)升高的有效性降低，该药物诱导的胰岛素分泌的剂量反应关系向右移动。与对照细胞相比，表达UCP1的细胞的静息膜电位显著超极化6.2 mV。在穿孔和传统全细胞膜片钳配置中，表达UCP1的细胞的ATP敏感性钾（K(ATP)）通道的电导密度分别比对照细胞高6倍和1.7倍。两组之间K(ATP)通道对甲苯磺丁脲的敏感性没有差异，这表明在完整细胞中，需要超过6倍浓度的甲苯磺丁脲才能将表达UCP1的细胞的K(ATP)通道电流降低到与对照细胞相同的水平。电压依赖性Ca(2+)通道的电流密度不受影响。总之，表达UCP1的细胞对甲苯磺丁脲以及营养物质表现出不应性。K(ATP)通道活性上调与线粒体功能受损细胞对该药物无反应有关。
