Regulation of mechanical interactions between fibroblasts and the substratum by stretch-activated Ca2+ entry

Ca2+ ions have long been implicated in regulating various aspects of cell movements. We found that stretching forces applied through flexible substrata induced increases in both intracellular Ca2+ concentration and traction forces of NIH3T3 fibroblasts. Conversely, application of gadolinium, an inhibitor of stretch-activated ion channels, or removal of extracellular free Ca2+ caused inhibition of traction forces. Gadolinium treatment also inhibited cell migration without affecting the spread morphology or protrusive activities. Local application of gadolinium to the trailing region had no detectable effect on the overall traction forces, while local application to the leading edge caused a global inhibition of traction forces and cell migration, suggesting that stretch-activated channels function primarily at the leading edge. Immunofluorescence microscopy indicated that gadolinium caused a pronounced decrease in vinculin and phosphotyrosine concentrations at focal adhesions. Our observations suggest that stretch-activated Ca2+ entry in the frontal region regulates the organization of focal adhesions and the output of mechanical forces. This mechanism probably plays an important role in sustaining cell migration and in mediating active and passive responses to mechanical signals.