Lack of Piezo1 expression reduces differentiation in vivo. (A) Fluorescent ISH of Piezo1 mRNA in the E10.5 mouse brain. Representative ISH images of WT embryo sections for the forebrain, midbrain, and hindbrain regions shown in top three panels, and Piezo1 KO mutant (midbrain) shown in the bottom panel. Piezo1 mRNA signal shown in yellow, TUJ1 antibody staining of neurons shown in magenta, and Hoechst-stained nuclei in cyan. The white dashed lines demarcate the neuroepithelium, and the red solid box highlights a blood vessel; ap, apical/ventricular border. Images are representative of n = 3 embryos for WT and n = 2 embryos for Piezo1 KO. (B) Representative WT and Piezo1 KO midbrain regions from embryo sections immunostained with NESTIN (cyan), TUJ1 (yellow), and SOX2 (magenta). The white dashed lines demarcate the neuroepithelial borders. (C) Gardner-Altman estimation plots of the area of TUJ1+ regions normalized to the total area of the neuroepithelium for WT and Piezo1 KO embryos from images as in B. Data are from n = 6 embryos (26 unique fields of view) for WT and n = 5 embryos (27 unique fields of view) for Piezo1 KO from five independent experiments (P value calculated by two-sample t-test, Cohen’s d = −1.32). See also Video 1 and Fig. S2.