Fig. 6. Slits guide VNO axons by chemorepulsion. E14.5 VNO explants were placed in a collagen-matrix next to COS cell aggregates transfected with Slit1 (B), Slit2 (C), Slit3 (D) or mock transfected (control, A). Lipofection was used to transfect 6 cm² dishes with a total amount of 0.75 µg Slit2 DNA and 1.5 µg DNA for Slit1 and Slit3. After 3 days in culture, outgrowth of VNO axons was recorded and quantified (E). (A) Axon-outgrowth from VNO explants was radial and not influenced by mock-transfected COS cells (n=33 explants). (B-D) Slit1-Slit3 caused chemorepulsion of VNO axons. (E) The repulsive effect of Slit1-Slit3 on VNO axons was quantified by measuring the length of the neurite front in the proximal (P) and distal quadrants (D) of the VNO explant relative to the cell aggregate. In controls, a P/D ratio of 1 (0.82±0.18) indicates almost no repulsive activity of mock-transfected cell aggregates. By contrast, Slit1-Slit3 shift the P/D ratio to 0.25±0.13, 0.1±0.11 and 0.25±0.18, respectively, indicating a strong Slit-mediated chemorepulsion of VNO axons. Scale bar: 100 µm.