The interactions of blood flow, A-V O2 difference (AVDO2), and A-V shunting were measured in normal hindlimbs of nine anesthetized dogs. An aorto-iliac nonpulsatile perfusion pump was used to change femoral artery blood flow from zero (collateral flow only) to twice its baseline level. Femoral AVDO2 was measured by in-line spectrophotometric O2 analysis. A-V shunting was measured with radio-labeled microspheres. Systemic hemodynamic parameters and temperature remained constant during the experiments. Despite changes in femoral mean arterial pressure (160 to 54 mm Hg) and AVDO2 (1.8 to 8.2 ml O/2/dl) that occurred with femoral blood flow reduction, peripheral A-V shunting remained constant at 4.1-5.5%. Alpha-adrenergic ablation (sympathectomy) was used to increase A-V shunting (up to 20%) during part of this experiment. When hindlimb blood flow was normal or increased, autoregulation of O2 extraction maintained constant hindlimb O2 consumption, despite sympathectomy-induced changes in A-V shunting. Subnormal femoral artery blood flow reduced hindlimb O2 consumption, and in this setting the increased A-V shunting further decreased femoral AVDO2 and O2 consumption. Since AVDO2 is dependent upon both blood flow and the variable efficiency of cellular O2 extraction, it cannot be used as an accurate indicator of A-V shunting. Direct microsphere techniques should be applied to A-V shunt measurements in clinical settings where A-V shunting is suspected.