The purpose of the study was to examine endothelium‐dependent and ‐independent vasodilation of conduit and resistance vessels in sickle cell anemia (HbSS). We measured the effects of intra‐arterial infusion of methacholine, sodium nitroprusside, and N^G‐monomethyl‐L‐arginine (L‐NMMA) on forearm blood flow using venous occlusion plethysmography in eight patients with HbSS and 11 HbAA controls. We assessed vasodilation of the conduit brachial artery using ultrasound in 17 patients with HbSS and 41 nonanemic controls. Forearm blood flow response to methacholine was similar in HbSS and HbAA, while the response to sodium nitroprusside was greater in those with HbSS. Nitric oxide synthase inhibition with L‐NMMA attenuated methacholine‐induced forearm blood flow to a lesser extent in HbSS compared to HbAA, suggesting diminished nitric oxide (NO) contribution to endothelium‐dependent vasodilation. Flow‐mediated and nitroglycerin‐induced dilation were impaired in HbSS compared to controls and were not affected by the antioxidant vitamin C or the precursor substrate for NO synthesis L‐arginine. NO bioactivity is reduced but differs in peripheral conduit and resistance vessels in HbSS. Resistance vessels have preserved response to exogenous NO donors but have diminished contribution of NO to endothelium‐dependent vasodilation. Conduit vessels demonstrate impaired vasodilation to endogenous and exogenous NO. Am. J. Hematol. 74:104–111, 2003. © 2003 Wiley‐Liss, Inc.