Integrin mediated mechano-transduction in vascular smooth muscle cells (VSMC) plays an important role in the physiological control of tissue blood flow and vascular resistance. To test whether force applied to specific ECM-integrin interactions could induce myogenic-like mechanical activity at focal adhesion sites, we used atomic force microscopy (AFM) to apply controlled forces to specific ECM adhesion sites on arteriolar VSMC. The tip of AFM probes were fused with a borosilicate bead (2~5 µm) coated with fibronectin (FN), collagen type-I (CNI), laminin (LN) or vitronectin (VN). ECM-coated beads induced clustering of ₅ and ₃ integrins and actin filaments at sites of bead-cell contact indicative of focal adhesion formation. Step increases of an upward (z-axis) pulling force (800~1600 pN) applied to the bead-cell contact site for FN specific focal adhesions induced a myogenic-like, force-generating response from the VSMC resulting in a counteracting downward pull by the cell. This micromechanical event was blocked by cytochalasin D, but was enhanced by jasplakinolide. Function-blocking antibodies to ₅₁ and _v3 integrins also blocked the micromechanical cell event in a concentration-dependent manner. Similar pulling experiments with CNI, VN, or LN failed to induce the myogenic-like micromechanical events. Collectively, these results demonstrate that mechanical force applied to integrin-FN adhesion sites induces an actin-dependent myogenic-like, micromechanical event. Focal adhesions formed by different ECM proteins exhibit different mechanical characteristics, and FN appears of particular relevance in its ability to strongly attach to VSMC and to induce myogenic-like, force-generating reaction from sites of focal adhesion in response to externally applied forces.