Shortly before the experiments, we carefully dissociated cells with a 1% trypsin/EDTA solution, and an aliquot of the suspension containing cell clumps was rapidly placed in a recording chamber adherently attached to the working stage of a DM-IL inverted microscope (Leica; Highrise Instrument, Taichung, Taiwan). The electrodes which were used to record were fabricated from Kimax-51® capillaries with 1.5–1.8 mm in diameter (Kimble® 34500-99; Merck, Taipei, Taiwan) by using a PC-10 vertical puller (Narishige; Taiwan Instrument, Tainan, Taiwan), and their tips were then fire-polished with MF-83 microforge (Narishige). When the electrodes were filled up with different internal solutions described above, their resistance was measured to range between 3 and 5 MΩ, for the purpose of making good GΩ-seal formation. We performed patch clamp recordings in cell-attached, inside-out or whole-cell configuration by using either an RK-400 (Bio-Logic, Claix, France) or an Axopatch-200B amplifier (Molecular Devices; Bestgen Biotech, New Taipei City, Taiwan), as elaborated elsewhere [29 (link),31 (link),52 (link),59 (link)]. Whole-cell current recordings were established by rupturing the patch of membrane isolated with GW sealing by the patch pipette, then bringing the cell interior into contact with the pipette interior.
Free full text: Click here
