The distribution of actin and myosin II was studied by immunofluorescence microscopy in Dictyostelium discoideum cells that had been stimulated with a chemoattractant, cAMP. The amounts of actin but not of myosin II increased in the cortical region of cells after 5-10 seconds of stimulation by cAMP at 21°C (the first peak). After a transient decrease in the amount of actin in the cortical region, the amounts of both actin and myosin II increased in the cortical region after 25-35 seconds of stimulation (the second peak). The amounts of myosin II decreased in the cortical region thereafter but actin became localized in the pseudopods. The stimulated cells extended many short projections that contained actin at the time of the first peak and the cell bodies contracted at the time of the second peak. Foci in the cortical actin network decreased in number at 20-30 sec and recovered thereafter. A quantitative examination using a fluorocytometer revealed that the amounts of actin filaments in the cells increased at both peaks. The calcium ionophore A23187 and Ca²⁺ ion induced the reorganization of actin and myosin II to a certain extent. Furthermore, Ca²⁺ ions inhibited the return of myosin II to the endoplasm in CAMP-stimulated cells after preincubation with A23187. Thus, Ca²⁺ ions may play a dual role, being involved in the regulation of both the initiation and the termination of the reorganization of cytoskeletons. The cAMP-stimulated responses of cytoskeletons were not inhibited by EGTA, La³⁺ ions, or ruthenium red but were inhibited by preincubation with BAPTA-AM. These observations suggest that increase of Ca²⁺ ions from intracellular stores may play a crucial role in the response of cytoskeletons.