The effect of NOC-5, ZAP, and ODQ on the Ca²⁺ signaling induced by 5-HT in airway SMCs. (A) A fluorescence confocal image of part of an airway in a lung slice showing the epithelial cells (EPCs) lining the airway lumen and the underlying SMCs. (B, top trace) Ca²⁺ changes in a small region (∼7 × 7 pixels, indicated by the white square in A) within an SMC plotted as a ratio (F/F₀) with respect to time showing the effect of 10 µM NOC-5 on the Ca²⁺ oscillations induced by 0.1 µM 5-HT. In the bottom part of B, a line scan analysis from the longitudinal axis of the SMC (indicated by the dashed line in A) shows the propagation of the Ca²⁺ oscillations along the SMC as Ca²⁺ waves (near vertical white lines). The position of both the ROI and the line scan was adjusted to track the same area inside the SMC during airway contraction. NOC-5 transiently blocked the Ca²⁺ oscillations, which resumed with a lower frequency. (C and D) Ca²⁺ traces and line scan showing the effect of 10 µM ZAP and 5 µM ODQ on the Ca²⁺ oscillations induced by 0.1 µM 5-HT and subsequent inhibition by 10 µM NOC-5 in single SMCs. ZAP or ODQ alone had no effect on the Ca²⁺ oscillations induced by 5-HT. However, in the presence of ZAP, NOC-5 further decreased the frequency and increased the amplitude and duration of the Ca²⁺ waves (line scan: lines brighter and wider, respectively). In contrast, ODQ blocked the inhibitory action of NOC-5 on the Ca²⁺ oscillations. Representative data from at least seven different slices from three mice are shown. A movie of the effect of NOC-5 on 5-HT–induced Ca²⁺ signaling in airway SMCs is shown in Video 2.