Background We compared the contractile responses to endothelin-1 (ET-1) with and without the inhibition of ET-A receptors and protein kinase C-alpha (PKC-α) in the human peripheral microvasculature of diabetic and case-matched, nondiabetic patients. Methods Chest wall skeletal muscle was harvested from patients with and without diabetics undergoing cardiac surgery. Peripheral arterioles (90–180 μm in diameter) were dissected from the harvested tissue. Microvascular constriction was assessed by videomicroscopy in response to ET-1 with and without an endothelin-A (ET-A) receptor antagonist, an endothelin B (ET-B) antagonist, or a PKC-α inhibitor. Results ET-1 induced a dose-dependent contractile response of skeletal muscle arterioles from diabetic and nondiabetic patients. The contractile response of diabetic arterioles from both prebypass and postbypass to ET-1 (10−9 mol/L) was decreased compared with those of nondiabetic patients ( P < .05). The contractile responses of microvessels of both diabetics and nondiabetics to ET-1 were inhibited in the presence of either ET-A receptor antagonist BQ123 (10−7 mol/L) or the PKC-α inhibitor safingol (2 × 10−5 mol/L, P < .05, respectively). In contrast, the ET-1–induced vasoconstriction was not affected by the administration of the ET-B receptor antagonist BQ788 (10−7 mol/L). There were no differences in skeletal muscle levels of the ET-A and ET-B receptors between diabetic and nondiabetic groups. Conclusion Diabetic patients demonstrated a decreased contractile response to ET-1 in human peripheral microvasculature. The contractile response of diabetic vessels to ET-1 occurs via activation of ET-A receptors and PKC-α. These results provide novel mechanisms of ET-1–induced contraction in vasomotor dysfunction in patients with diabetes.