The preferential association of cholesterol and sphingolipids within plasma membranes forms an organized compartment termed lipid rafts. Addition of caveolin proteins to this lipid milieu induces the formation of specialized invaginated plasma membrane structures called caveolae. Both lipid rafts and caveolae are purported to function in vesicular transport and cell signaling. We, and others, have shown that disassembly of rafts/caveolae through depletion of plasma membrane cholesterol mitigates mechanotransduction processes in endothelial cells. Since osteoblasts are subjected to fluid-mechanical forces, we hypothesize that cholesterol-rich plasma membrane microdomains also serve the mechanotransduction process in this cell type. Cultured human fetal osteoblasts were subjected to either sustained hydrostatic pressure or laminar shear stress using a pressure column or parallel plate apparatus, respectively. We found that sustained hydrostatic pressure induced protein tyrosine phosphorylation, activation of Erk 1/2, and enhanced c-fos expression in both a time- and magnitude-dependent manner. Similar responses were observed in cells subjected to laminar shear stress. Both sustained hydrostatic pressure- and shear stress- induced signaling was significantly reduced in osteoblasts pre-exposed to either filipin or methyl--cyclodextrin. These mechanotransduction responses were restored upon reconstitution of lipid rafts/caveolae suggesting that cholesterol-rich plasma membrane microdomains participate in the mechanotransduction process in osteoblasts. In addition, mechanical force induced phospho-proteins were localized within caveolin-enriched membrane fractions. These data support the concept that lipid rafts and caveolae serve a general function as cell surface mechanotransduction sites within the plasma membrane.