2.6. Short-Circuit Current Measurement

Short-circuit current (I_sc) measurement of polarized HNE was performed as described [19,27]. HNE were treated with CFTR modulators or 0.2% DMSO (vehicle control) for 24 h at 37 °C in serum-free PneumaCult-ALI medium (StemCell Technologies). Snapwell filters were mounted in Ussing chambers (Physiologic Instruments) in Krebs-bicarbonate Ringer (KBR) buffer (140 mM Na⁺, 120 mM Cl⁻, 5.2 mM K⁺, 25 mM HCO₃⁻, 2.4 mM HPO₄, 0.4 mM H₂PO₄, 1.2 mM Ca²⁺, 1.2 mM Mg²⁺, 5 mM glucose, pH 7.4), which was mixed by bubbling with carbogen (95% O₂ and 5% CO₂). To generate a basolateral-to-apical chloride gradient NaCl was replaced with 115 mM Na⁺ gluconate in the apical buffer and the I_sc was determined in the presence of 100 μM amiloride. The transepithelial voltage was clamped at 0 mV (VCC MC8 multichannel voltage/current clamp, Physiologic Instruments) after compensating for voltage offsets and current and resistance were recorded at 37 °C with the Acquire and Analyze package (Physiologic Instruments).