Both IgCAMs and the actin cytoskeleton play critical roles in neuronal growth cone motility and guidance. However, it is unclear how IgCAM receptors transduce signals from the plasma membrane to induce actin remodeling. Previous studies have shown that local clustering and immobilization of apCAM, the Aplysia homolog of NCAM, induces Src kinase activity and F-actin polymerization in the peripheral domain of cultured Aplysia bag cell growth cones. Therefore, we wanted to test whether the Src kinase substrate and actin regulator cortactin could be a molecular link between Src activity and actin assembly during apCAM-mediated growth cone guidance. Here, we cloned Aplysia cortactin and showed that it is abundant in the nervous system. Immunostaining of growth cones revealed a strong colocalization of cortactin with F-actin in filopodial bundles and at the leading edge of lamellipodia. Perturbation of the cytoskeleton indicated that cortactin distribution largely depends on actin filaments. Furthermore, active Src colocalized with cortactin in regions of actin assembly, including leading edge and filopodia tips. Finally, we observed that cortactin, like F-actin, localizes to apCAM adhesion sites mediating growth cone guidance. Altogether, these data suggest that cortactin is a mediator of IgCAM-triggered actin assembly involved in growth cone motility and guidance. © 2008 Wiley-Liss, Inc.