Myoblast fusion is critical for skeletal muscle formation and growth. The molecular mechanisms of myoblast fusion are not completely understood. Cell fusion is an ordered process of migration, cell-cell contact and adhesion. Thereafter, the plasma membranes align and membrane fusion occurs. Extensive cytoskeletal reorganization occurs before and after cell fusion. Several proteins involved in cytoskeletal arrangements are required for myoblast fusion in Drosophila and mammalian cells. Dynamic cytoskeletal assemblies regulating cell migration and fusion are dependent upon the localized hydrolysis of ATP. Phosphocreatine can serve to replenish ATP at sites of high ATP consumption. The generation of phosphocreatine is dependent upon the enzyme creatine kinase (CK). Roles for CK have been demonstrated in cytoskeletal rearrangements and cell motility in various cell types. In this study we hypothesized that myoblast fusion is a high-energy process requiring creatine. We show that adding exogenous creatine to cultures of differentiating primary muscle cells enhanced myoblast fusion with myotubes in a creatine-kinase dependent manner. Pretreating proliferating myoblast cultures with creatine also enhanced myoblast fusion during later stages of myogenesis. The fusion promoting effects of creatine were dependent on actin, as latrunculin blocked the effects of creatine. These results suggest that intracellular creatine acts as a temporally regulated energy source providing energy during cytoskeletal rearrangements in myoblast fusion.