Osteocytes embedded in the matrix of bone are thought to be mechanosensory cells involved in the regulation of both the mass and structure of bone formation. The osteocyte is thought to signal the osteoclast to induce bone resorption and the osteoblast to begin bone formation. Coordination of these remodeling processes is mediated, in part, by cell-to-cell gap junction-mediated intercellular communication in transmitting mechanical signals to the other bone cells essential for bone formation and remodeling. The objective of this application is to understand the role that gap junction channels play in regulating the signals stimulated by mechanical stress in osteocytes. The central hypothesis of the project is that gap junction-mediated intercellular communication is one of the major pathways that mediate and coordinate the signals generated by mechanical loading. This hypothesis has been formulated on the basis of strong preliminary findings, which suggest that gap junctions are essential for mediating the osteocyte cell responses to mechanical stress, and that intercellular communication mediated by gap junctions between osteocytes is stimulated in the presence of the applied strain. The central hypothesis to be tested and the objective of the application will be accomplished by pursuing four specific aims: 1). Confirm the essential role that gap junctions play in osteocyte responses to mechanical stress induced by fluid flow; 2). Identify and characterize the connexin(s) responsible for formation of functional gap junction channels in osteocytes; 3). Determine if fluid flow is more effective than mechanical stretch in regulating gap junction function in osteocytes, and 4). Determine if prostaglandin E2 regulates osteocyte gap function response to shear stress induced by fluid flow. The proposed research is innovative, because osteocyte-like cell lines will be employed as the principal model system with complementary experiments using primary osteocyte cultures. Moreover, this study combines comprehensive biochemical, molecular, genetic and functional approaches with unique mechanical engineering applications. It is our expectation that our experimental findings will have a major impact on our understanding on how signals generated by mechanical strain are coordinated between the osteocytes and other cellular elements of the bone micro-environment. The outcomes will be significant because this new knowledge will contribute to broaden our understanding of how mechanical signals are transduced and modulated in osteocytes. Furthermore, this research activity should make a contribution to the general strategies for the prevention/treatment of bone diseases such as osteoporosis and to new ideas and potential molecular targets for drug development and discovery.