The mean turning angle evoked by VEGF164 in the presence this website of control IgG was 16.8° ± 2.4° (n = 9), but 0.0° ± 2.6° (n = 10) in the presence of the function-blocking anti-NRP1 antibody (p < 0.001). VEGF164 therefore signals through NRP1 to attract the growth cones of presumptive contralateral RGC axons. Based on these findings,

together with the expression pattern of VEGF164 and NRP1 and the loss-of-function phenotypes of the corresponding mouse mutants in vivo, we conclude that VEGF164 signals to NRP1-expressing RGC growth cones to promote axon crossing at the chiasmatic midline. Nerves and blood vessels ramify through tissues in strikingly similar patterns and develop during embryogenesis under the control of similar cellular and molecular mechanisms (reviewed by Ruiz de Almodovar et al., 2009 and Adams and Eichmann, 2010). Thus, classical axon guidance cues of the ephrin, netrin,

and SLIT families affect the growth of blood vessels. Conversely, it has been hypothesized that the main selleckchem vascular growth factor VEGF-A is important for axon growth and guidance, either in its own right or by competing with SEMA3A for NRP1 binding (reviewed by Carmeliet, 2003 and Ruiz de Almodovar et al., 2009). However, evidence is still lacking that VEGF-A controls axon guidance in vivo. By demonstrating that VEGF164 is expressed at the optic chiasm midline, is essential for RGC axon guidance and fasciculation in vivo, and promotes RGC axon outgrowth and attractive growth cone turning, we provide

evidence that VEGF-A is a physiological axon guidance cue (Figures 8A and 8B). We found that loss of VEGF164 or its receptor, NRP1, perturbs axon crossing at the optic chiasm in a similar manner in vivo, causing optic tract defasciculation and increasing ipsilateral projection. Because VEGF and NRP1 are well known for their essential roles in blood vessel growth (Kawasaki et al., 1999, Ruhrberg et al., 2002 and Gerhardt et al., 2004), we used endothelial-specific NRP1 mutants to exclude the possibility that loss of VEGF164 signaling inhibits contralateral axon growth indirectly by disrupting L-NAME HCl the brain vasculature. These mutants suffer blood vessel defects similar to those seen in full NRP1 knockouts (Gu et al., 2003), but do not display defects in midline crossing of contralateral RGC axons. VEGF164/NRP1 signaling therefore controls axon crossing at the optic chiasm independently of its role in blood vessels. Instead, our results support a model in which VEGF164 signals through NRP1 in RGC growth cones to regulate axon pathfinding directly (Figure 8B). Thus, we found that NRP1 is expressed strongly by contralateral RGC axons throughout the period of optic chiasm development, and that VEGF164 is a powerful chemoattractant for growth cones from presumptive contralateral RGC axons that acts in a NRP1-dependent fashion.