Ca2 ionophore 2 eth 129

Double-barreled Ca²⁺-selective microelectrodes were prepared from thin-walled borosilicate with 1.2 and 1.5 mm outside diameter (OD) (PB150F-4, World Precision Instruments, Sarasota, FL, USA). The ion-selective barrel (1.5-mm OD) was silanized with dimethyldichlorosilane vapor and then backfilled with the Ca²⁺ ionophore II (ETH 129) Sigma-Aldrich in Saint Louis, MO, USA. The remaining portion of the ion-selective barrel was backfilled with pCa7, as previously described in [10 (link)]. The resting membrane potential (RMP) barrel (1.2-mm OD) was backfilled with 3 M KCl before the measurements. The RPM- and Ca²⁺-specific potentials were acquired at a frequency of 1000 Hz using AxoGraph software (version 4.6; Axon Instruments, San Jose, CA, USA), then and stored on a computer for further analysis. Before and after each measurement, individual calibration of the Ca²⁺-selective microelectrodes was performed by following the previously described protocol [10 (link)]. If the calibration curves obtained before and after the measurement differed by more than 3 mV, the data from that microelectrode were excluded from further analysis [10 (link)].

Double-barreled Ca²⁺-selective microelectrodes were prepared and individually calibrated as described previously (Eltit et al., 2013 (link)). Ca²⁺ ionophore II (ETH 129; Fluka Sigma–Aldrich, St. Louis, MO, USA) was used to backfill the Ca²⁺-selective microelectrode. Resting membrane potential (Vm) and Ca²⁺ potentials were recorded via a high impedance amplifier (FD-223-WPI, Sarasota, FL, USA), as described previously (Lopez et al., 2018b (link)). After obtaining measurements of resting [Ca²⁺]_r, all microelectrodes were recalibrated. If the initial and final calibration curves did not agree within 3 mV, data from that microelectrode were discarded.