Nerve blood flow (NBF) is reduced in experimental diabetic neuropathy (EDN), but the mechanism of its reduction is uncertain. We tested the hypothesis that reduced NBF might be due to alterations of nitric oxide synthase (NOS) and endothelin of microvascular endothelial cells of sciatic nerve. We evaluated epineurial arteriolar vasoreactivity in response to superfused test agents. NBF was measured using microelectrode H2 polarography. Vasoconstrictor responses to endothelin-1 (ET-1; 10(-6), 10(-7), 10(-8), 10(-9), 10(-10) M) showed dose-response curves with similar EC50 values, indicating no change in potency. We applied the NOS inhibitor NG-nitro-L-arginine and observed reduced inhibition of NBF in EDN, correctable with insulin treatment and also with infused L-arginine. We conclude that vasoreactivity is disturbed in EDN, and is due to a combination of an impairment of NOS activity with reduced NO and increased endothelin effect (normal receptor sensitivity and increased plasma values) in EDN. Hyperglycemia is likely to be the mechanism of NOS inhibition since insulin treatment reversed this abnormality.