Prior studies in our laboratory have demonstrated an association of specific gap junction proteins with intramembranous bone formation in the avian mandible. The purpose of the present study was to extend these observations by determining if there was a relationship between the expression of one of the gap junction proteins examined previously (connexin43) and the expression of specific cell adhesion (CAM) and/or substrate adhesion (SAM) molecules [i.e. NCAM, A-CAM (N-cadherin) and tenascin (tenascin-C)] that have previously been shown to be associated with bone formation. Immunohistochemical localization of connexin43, tenascin, NCAM and N-cadherin was performed on serial sections of mandibles of chick embryos from 6 to 12 days of incubation. Analysis of adjacent serial sections revealed that the NCAM and tenascin immunostaining that appeared initially on the lateral aspect of Meckel's cartilage preceded the overt expression of trabecular bone. At subsequent stages, NCAM and tenascin staining gradually overlapped the region of connexin43 expression. In contrast, the expression of N-cadherin was found to colocalize with that of connexin43 from the first appearance of connexin43 expression. Most significantly, although the domains of NCAM and tenascin expression were initially separate from that of connexin43, bone formation originated only in the region where these domains intersected. These findings suggest that, of the CAMs and SAMs examined, N-cadherin appears to be associated with the establishment of cell contacts responsible for the presence and/or maintenance of connexin43-mediated gap junctional communication, while tenascin and NCAM appear to be associated, in a more specific manner, with processes that accompany the overt expression of the osteogenic phenotype.