Long chain fatty acid receptor function using visualization of mitochondrial function

在这项研究中，我们通过可视化线粒体功能和细胞内钙离子浓度来评估长链脂肪酸受体的功能。线粒体的pH值被认为是反映其功能活性的指标。然而，目前还没有一个实用的系统来定量和连续地测量线粒体的pH值。近年来，用GFP法成功地计算了有机质的pH值，但仍不是直接定量计算。在这里，我们开发了一种新的系统，用两个GFP之间的荧光比率来评估活细胞的线粒体pH。该pH探针系统能较好地评价较宽的pH范围。应用葡萄糖、甲苯丁胺、精氨酸、钾等胰岛素刺激物质，测定线粒体pH值。在培养的β细胞中，葡萄糖和钾可使线粒体pH立即下降，精氨酸和甲苯丁胺可使线粒体pH缓慢下降。β-细胞线粒体pH值的基础水平通过在培养介质中的曝气而增加。然而，GPR40激动剂能迅速降低线粒体的pH。这一结果表明，GPR40的信号转导途径不同于其他信号转导途径。此外，我们通过在STC-1细胞中的钙显影来评估GPR120的激动剂。

In this study, we evaluate the long chain fatty acid receptor function using visualization of mitochondrial function and intracellular Ca^<2+> concentration. Mitochondrial pH is known to reflect its functional viability. However, there has been no useful system to measure the quantitative and serial mitochondrial pH. Recently, a pH of organum has been successfully calculated by using GFP, but still it is not a direct quantitative calculation. Here, we developed the new system to evaluate mitochondrial pH of living cell with fluorescence ratio between two GFP. This pH probe system could evaluate the wide and preferable pH range. We applied insulin-stimulating substance, such as glucose, tolbutamide, arginine and potassium, and measure mitochondrial pH. In the cultured β-cells, the mitochondrial pH decreased immediately by glucose and potassium, and leisurely decreased by arginine and tolbutamide. A basal level of the mitochondrial pH of β-cells has increased by aeration in a cultured medium. However, GPR40 agonist quickly decreases the mitochondrial pH. This result suggested that the signal transduction of GPR40 is differ from the others. Additionally, we evaluate the agonist of GPR120 using Ca2+ visualization in STC-1 cell.