Unactivated platelets contain about 69% G actin and less than 10% of the contractile proteins in a cytoskeletal core resistant to extraction with 1% Triton X-100. Activation by thrombin leads, within 1 min, to the formation of pseudopodia and contractile gels, accompanied by the reduction of the G-actin content to about 22% and the development of cytoskeletal cores containing 70%-80% of the total actin and 60%-80% of the total myosin and actin-binding protein. Inhibition of pseudopodal formation by pretreatment with cytochalasin B before thrombin activation results in the loss of most of the actin-binding protein and about one third of the actin from the cytoskeletal core. Myosin incorporation and contractile gel formation are unaltered by this treatment. Conversely, activation with phorbol 12-myristate 13-acetate leads to pseudopodal but not contractile gel formation, with cytoskeletal cores containing mostly actin and actin-binding protein. These results demonstrate that there are separable cytoskeletal assembly processes in platelets for pseudopodal and contractile gel formation.