Plasma membrane protrusion is fundamental to cell motility, but its regulation by the extracellular environment is not well elucidated. We have quantified lamellipodial protrusion dynamics in human vascular smooth muscle cells exposed to fibroblast growth factor 2 (FGF-2) and type I collagen, two distinct ligands presented to vascular cells during arterial remodeling. Video microscopy revealed that FGF-2 stimulated a modest increase in lamellipodial protrusion rate that peaked within 15 min. This response was associated with immediate but transient activation of Rac1 and was inhibited in cells infected with retrovirus containing cDNA encoding dominant-negative Rac1. A 1-h exposure to FGF-2 also set up a second phase of more striking lamellipodial protrusion evident at 24-36 h. This delayed response was most pronounced when cells were on type 1 collagen and was associated with FGF-2-induced expression of collagenase-1 that localized to the edge of protruding lamellipodia. Moreover, late membrane protrusion was inhibited when cells were on collagenase-resistant type I collagen, implicating degraded collagen as a mediator. For cells on collagen, the immediate activation of Rac1 by FGF-2 was followed by a sustained wave of Rac1 activation that was inhibited when cleavage of the collagen triple helix was prevented and also by blockade of alpha(v)beta(3) integrin. We conclude that lamellipodial protrusion in smooth muscle cells can be regulated by waves of Rac1 activation, corresponding to the sequential presentation of FGF-2 and remodeled collagen. The findings thus reveal a previously unrecognized level of coordination among extracellular input that enables cells to maintain protrusive activity over prolonged periods.