Fig. 6. Non cell-autonomous effects of motoneuronal Sema3A in conditions of forced overexpression suggests that intrinsic Sema3A acts via secretion. (A) Myc immunostaining on transverse sections of Sema3AiresEGFP HH25 embryo showing that Myc-tagged Sema3A is targeted to the defasciculated spinal nerve (arrowhead). (B) Histogram showing the percentage (±s.d.) of EGFP⁺ or EGFP^- collapsed growth cones following Sema3A and control supernatant exposure, of Sema3AiresEGFP, or control ventral spinal cord explants. ^**, P<0.01; ^***, P<0.001 with Student's t-test; n.s.: non significant. The total number of axons from three independent experiments is indicated on the bars. (C,D) Histograms showing Nrp1 (NP1) and NgCAM immunofluorescence intensities per unit of surface at the growth cone in conditions of increased level of Sema3A overexpression. Immunolabeling of cell surface Nrp1 and NgCAM was performed on non-permeabilized explants. Fluorescence intensities were normalized to EGFP⁺ conditions. ^**, P<0.01; ^***, P<0.001 with Mann-Whitney test. The number of axons examined is indicated on the bars.