The inhibitory effect of cytochalasin B on contraction of smooth muscle cells isolated from guinea-pig taenia coli was investigated. Cytochalasin B (10-70 microM) inhibited the high K+ (70 mM)-induced contraction in a dose-dependent manner, and the maximum and the half-maximum effects were obtained at 50 and 15 microM, respectively. Cytochalasin B (70 microM) decreased ATPase activity in skinned guinea-pig taenia coli. However, cytochalasin B (50 microM) had no significant effect on the voltage-dependent Ca2+ currents, the passive membrane properties or the membrane potential. Cytochalasin B also had no effect on the phosphorylation of 20 kDa myosin light chain induced by high K+ and cytosolic Ca2+ levels. These results suggest that the inhibition of contraction by cytochalasin B may be due to its effects on actin of microfilaments and contractile filaments of guinea-pig taenia coli smooth muscle cells.