In vitro studies have shown that the f-actin cytoskeleton in osteoblasts exhibits an adaptive response after being subjected to mechanical stimuli. We hypothesize that transient disruption and subsequent re-polymerization of the cytoskeleton is responsible for this adaptive response. Therefore, in this study, we determine the occurrence of transient disruption of the f-actin fiber bundles and their subsequent reassembly after in vitro equi-axial stretching in MLO-A5 cells and after in vivo forearm compression loading in mice. In vitro studies are conducted to determine f-actin disruption after varying mechanical stimulus parameters that are known to affect bone formation. Results indicate that the f-actin cytoskeleton is disrupted in vitro as a function of applied mechanical stimulus parameters and that the f-actin bundles reassemble after loading induced disruption within 3 minutes after cessation of loading. The disruption of the f-actin cytoskeleton depends on the magnitude of stretch, the numbers of loading cycles, frequency, the insertion of rest between loading cycles and extracellular calcium. In vivo studies also demonstrate disruption of the f-actin cytoskeleton in cells embedded in the bone matrix immediately after mechanical loading. These studies suggest that adaptation of the f-actin fiber bundles of the cytoskeleton in response to applied loads occurs by disruption and subsequent re-polymerization.