Fertilized hen eggs were treated externally with 2,4-dichlorophenoxyacetic butyl ester (2,4-D b.e.) (3.1 mg/egg) before the start of the incubation. Actomyosin and sarcoplasmic reticulum adenosine triphosphatase (ATPase) activities from leg and complexus muscles of chicks hatched from treated eggs were measured. No significant variations were detected in the ATPase activities of actomyosin, but the sarcoplasmic reticulum Mg2(+)-activated ATPase and Ca2+, Mg2(+)-activated ATPase were inhibited 50 and 38% respectively. 45Ca2+ uptake into soleus muscle was increased by the 2,4-D b.e. treatment. The compartmental analysis of 45Ca2+ uptake kinetics showed increases in Ca2+ fluxes in sarcolemma and mitochondria and in the mitochondrial calcium pool. Isolated soleus muscles were treated with 2,4-dichlorophenoxyacetic acid (2,4-D) or 2,4-D b.e. [14C]2,4-D reached it highest level in these muscles after 1 hr of treatment. The in vitro treatment with 2,4-D or 2,4-D b.e. increased 45Ca2+ uptake into the muscles. 2,4-D b.e. produced greater alterations than 2,4-D. The compartmental analysis of the 45Ca2+ uptake kinetics also showed increases of the mitochondrial Ca2+ pool and Ca2+ fluxes through sarcolemma and mitochondria. These results led to a hypothesis based on Ca2+ permeability alterations for explaining the myopathic actions of these phenoxyherbicides.