Objective: Hydrostatic pressure is crucial component of cell environment and fundamental physical quantity, also it is the main factor of both cell integrity and function. Pressure variation disorder, beyond physiological limits, may lead to pathological states. In this study, we examined the effect of hydrostatic pressure on apoptosis induction, viability, morphology, adhesion potency to substrate and migration of differentiated PC-12 cells. Materials and Methods: PC-12 as a neuronal cell line maintained in RPMI 1640 culture medium supplemented with 10% fetal bovine serum. Staurosporine was used for differentiating of mitotic PC-12 cells to post mitotic and differentiated neuronal cells. Exclusion Dye was used for viability assay, total neurite length of each cell as well as morphometry. TUNEL staining was also performed for apoptosis detection, adhesion potency of cells to substrate and evaluation of cell migration. Results: Hydrostatic pressure, over physiological limits, induced apoptosis in differentiated PC-12 cells. It changed cell viability gradually and reduction happened significantly after 24 hours (p<0.05). In compare to the control group, hydrostatic pressure reduced total neurite length, adhesion potency to substrate and migration of cells in the examined group (p<0.05). Conclusion: Hydrostatic pressure induced apoptosis in differentiated PC-12 cells as a result of inappropriate interaction between cells and substrate. We propose that apoptosis in differentiated PC-12 cells may be an anoikis causing to lose the attachment to the substrate.