Abstract：
Mitochondrial membrane potential (MMP) plays a crucial role in the function of cells and organelles, involving various cellular physiological processes, including energy production, formation of reactive oxygen species (ROS), unfolded protein stress, and cell survival. Currently, there is a lack of genetically encoded fluorescence indicators (GEVIs) for MMP. In our screening of various GEVIs for their potential monitoring MMP, the Accelerated Sensor of Action Potentials (ASAP) demonstrated optimal performance in targeting mitochondria and sensitivity to depolarization in multiple cell types. However, mitochondrial ASAPs also displayed sensitivity to ROS in cardiomyocytes. Therefore, two ASAP mutants resistant to ROS were generated. A double mutant ASAP3-ST exhibited the highest voltage sensitivity but weaker fluorescence. Overall, four GEVIs capable of targeting mitochondria were obtained and named mitochondrial potential indicators 1-4 (MPI-1-4). In vivo, fiber photometry experiments utilizing MPI-2 revealed a mitochondrial depolarization during isoflurane-induced narcosis in the M2 cortex.
摘要：
线粒体膜电位（MMP）在细胞和细胞器的功能中起着至关重要的作用，涉及各种细胞生理过程，包括能源生产，活性氧（ROS）的形成，未折叠的蛋白质应激，细胞存活。目前,缺乏基因编码的MMP荧光指标(GEVIs)。在我们筛选各种GEVIs的潜在监测MMP时，加速动作电位传感器（ASAP）在靶向线粒体和多种细胞类型对去极化的敏感性方面表现最佳。然而,线粒体ASAP也显示出对心肌细胞中ROS的敏感性。因此,产生两个抗ROS的ASAP突变体。双突变体ASAP3-ST表现出最高的电压灵敏度，但荧光较弱。总的来说,获得了四个能够靶向线粒体的GEVIs，并将其命名为线粒体电位指标1-4（MPI-1-4）。在体内,利用MPI-2进行的纤维光度法实验显示，异氟烷诱导的M2皮层麻醉过程中线粒体去极化。
