Real-time impedance of hTM monolayers was measured using the electric cell-substrate impedance sensing (ECIS) Zθ system (Applied BioPhysics), as described (Wang et al., 2016 (link)). In brief, cells were seeded on 8W10E+ arrays coated with rat collagen before seeding. Once a stable monolayer was established, the cells were exposed to test agents (TREK-1 activators and inhibitors) or control media, and the resistance was measured in each well for 1 h at 10-s intervals in response to a 4-kHz AC frequency (40 electrodes per well). Resistance for each well was normalized to the baseline resistance before the addition of agonist/antagonist to account for baseline differences in the electrodes. Treatment wells were normalized to the control resistance at each time point to determine change in resistance relative to control.