(A) Confocal image (left) of alveolar epithelium and SiglecF-positive AMs loaded with the cytosol-localizing dye, calcein. Dashed squares enclose 2 AMs subjected to photobleaching. High-magnification images show calcein fluorescence in a hyperinflation-responsive AM (#1, upper panel) and nonresponsive AM (#2, lower panel) before and after bleach at indicated time points. Replicated 8 times in 4 lungs. (B) Plot shows increase in cytosolic Ca²⁺ in AMs after hyperinflation against efficiency of gap junctional communication (GJC) quantified in terms of fluorescence recovery after photobleaching (FRAP). Data are for 27 AMs from 4 lungs. Line drawn by linear regression. (C) Confocal images show calcein green–loaded sessile AMs identified by SiglecF and CD11c labeling. Pseudocolor high-power images of sessile AMs are shown in prebleach and 0- and 60-minute after bleach periods. White dashed circles indicate photobleached regions. Bars show group data quantifications of calcein fluorescence within sessile AMs at indicated time points. n = 3 and 4 lungs, respectively, for Cx43^FL and AM-Cx43^KO groups. Cx43^FL, floxed littermate control. AM-Cx43^KO, CD11cCre-Cx43^fl/fl. (D) Tracings in different colors show Ca²⁺ responses in AMs from indicated mice. Bars show percentage of AMs per field of view that responded to hyperinflation by increasing Ca²⁺ in CD11cCre-Cx43^fl/fl mice (AM-Cx43^KO) and littermate controls (Cx43^FL). Data analyses. Group data are mean ± SEM. P values were calculated by paired (C) or unpaired (D) t test.