The present experiments were performed to study the question whether intraarterially infused acetylcholine changes in local tissue perfusion of the skeletal muscle. In experiments on cats the triceps surae muscles were isolated. While acetylcholine (0.5 mug/kg X min) was infused intraarterially on one side, the opposite side was taken as control. 4 minutes after the start of the infusion non-radioactive microspheres of 15 mum diameter were injected into the ascending aorta. Subsequently the muscles were freeze-sectioned into 1.0 mm thick slices and cleared by sodium hydroxide and glycerine. The concentration of microspheres per mm3 was measured in representative cross sections of the muscle. Higher concentrations of microspheres were observed in the proximal than in the distal slices of the acetylcholine-treated side. The distribution of microspheres was found to be different in the segments as well: higher values were found in the central than in peripheral segments. The untreated side showed a qualitatively similar state. The differences in the distribution pattern, however, were smaller than in the acetylcholine-treated muscles. It is concluded that the increased inhomogeneity of the regional perfusion observed in this study during vasodilation corresponds to the results reporting a changed nutritive tissue perfusion during an acetylcholine treatment.