Whole-cell Patch Clamp Measurements Protocol

Whole-cell patch clamp measurements were performed as described earlier (Badheka et al., 2015 (link)). Measurements were carried out on YFP positive cells, in an extracellular solution containing (in mM) 137 NaCl, 5 KCl, 1 MgCl₂, 10 HEPES and 10 glucose, pH 7.4. The intracellular solution contained (in mM) 135 Cs-Metanesulfonate, 1 MgCl₂, 10 HEPES, 5 EGTA, 4 NaATP (pH 7.25). Patch clamp pipettes were prepared from borosilicate glass capillaries (Sutter Instruments) using a P-97 pipette puller (Sutter Instrument) and had a resistance of 4–6 MΩ. In all experiments after formation of gigaohm-resistance seals, the whole-cell configuration was established and currents were recorded using a ramp protocol from −100 mV to +100 mV over 500 ms preceded by a −100 mV step for 200 ms; the holding potential was −60 mV, and this protocol was applied once every 2 s. The currents were measured with an Axopatch 200B amplifier, filtered at 5 kHz, and digitized through the Digidata 1440A interface. In all experiments, cells that had a passive leak current more than 100 pA were discarded. Data were collected and analyzed with the PClamp10.6 (Clampex) acquisition software (Molecular Devices, Sunnyvale, CA), and further analyzed and plotted with Origin 8.0 (Microcal Software Inc, Northampton, MA, USA).