(A) Schematic of tamoxifen administration to generate Ascl1-mutant mice. (B) Representative images of control (Ascl1^creERT2/+ Rosa26^tdTomato/+) and mutant (Ascl1^cre/fl Rosa26^tdTomato/+) lung sections stained with anti-CGRP and anti-TUJ1 antibodies to characterize neuropeptide expression and innervation of tdTomato-lineaged PNECs, respectively. (C and D) Quantification of PNEC numbers based on CGRP staining or Ascl1 lineage at indicated ages. For this and all cell number quantifications that follow, each dot represents the average value across multiple sections representing the full range of depths from 1 lung (20–25 sections per lung, 10–15 sections per trachea), unless otherwise indicated. (E) Representative images of control and mutant trachea sections of Ascl1-lineaged cells. (F) Quantification of Ascl1-lineaged cells in the trachea. (G) Relative transcript level of neuroendocrine markers as assayed by qRT-PCR in P16 whole lungs. (H) Representative images of lung sections stained with anti–cleaved caspase-3 to label apoptotic cells (white arrowheads) in Ascl1-lineaged cells. (I) Close-up of boxed area from mutant lung (P21) from H. One-way ANOVA was used for C and D (n = 3–4 for each group). Student’s t test was used for F (n = 3 for each group). * for P < 0.05, ** for P < 0.01, *** for P < 0.001, and **** for P < 0.0001. Error bars represent mean ± SD. Scale bar size 100 μm for B and E; 50 μm for H and I. TUJ1, beta III tubulin; qRT, quantitative reverse transcription.