The radioactive microsphere technique was applied to determine simultaneous cardiac output and flow distribution in the rat. Left ventricular injections of large numbers of microspheres were given, without significant adverse effects, allowing determination of flow to organs and tissues with low perfusion rates. In order to determine coronary blood flow it was necessary to excise the inner lining of the left ventricle, thus eliminating activity from deposits of microspheres. Cardiac output determination showed less variation with the sampling catheter in the abdominal aorta than in the femoral artery. It is concluded that the microsphere method can be conveniently used for hemodynamic studies in the rat, and that the abdominal aorta is the preferred site for the placement of the reference catheter in the rat.