Fig. 5. Intrinsic Sema3A cell-autonomously modulates the Nrp1 level at the growth cone surface. (A) In situ hybridization of Nrp1 mRNA (NP1) on spinal transverse sections from Sema3AiresEGFP and EGFP embryos. Brackets indicate the electroporated side. Scale bar: 100 µm. (B,C) Nrp1 and NgCAM immunostaining of EGFP⁺ and EGFP^- growth cones of Sema3AiresEGFP and ctrlEGFP explants, with membrane permeabilization (B), and without permeabilization (C). Scale bars: 10 µm. (D-G) Histograms showing fluorescence intensities per unit of surface at the growth cone. Fluorescence intensities were normalized to EGFP⁺ conditions in D-F. Fluorescence intensities of pShScramble-EGFP⁺ and pShSema3A-EGFP⁺ growth cones were normalized to the pShScramble-EGFP^- or pShSema3A-EGFP^- conditions, respectively (G). ^**, P<0.01; ^***, P<0.001 with Mann-Whitney test. The number of growth cones analyzed is indicated for each condition. (H) Nrp1 and NgCAM immunostaining of EGFP⁺ and EGFP^- growth cones of pShScramble-EGFP and pShSema3A-EGFP explants without permeabilization.