The adhesive interactions of nerve growth cones stabilize elongating nerve fibers and mediate transmembrane signaling to regulate growth cone behaviors. We used interference reflection microscopy and immunocytochemistry to examine the dynamics and composition of substratum contacts that growth cones of chick sensory neurons make with extracellular adhesive glycoproteins, fibronectin and laminin. Interference reflection microscopy indicated that sensory neuronal growth cones on fibronectin-treated substrata, but not on laminin, make contacts that have the appearance and immobility of fibroblastic focal contacts. Interference reflection microscopy and subsequent immunocytochemical staining showed that beta 1 integrin and phosphotyrosine residues were concentrated at growth cone sites that resemble focal contacts. Two other components of focal contacts, paxillin and zyxin, were also co-localized with concentrated phosphotyrosine residues at sites that resemble focal contacts. Such staining patterns were not observed on laminin-treated substrata. Growth cone migration on fibronectin-treated substrata was inhibited by herbimycin A, a tyrosine kinase inhibitor. We conclude that sensory neuronal growth cones distinguish fibronectin from laminin by making contacts with distinct organization and regulation of cytoskeletal components at the adhesive sites. This finding suggests that growth cone interactions with different adhesive molecules lead to distinctive transmembrane organization and signaling to regulate nerve fiber elongation.