The objective of this study was to determine whether endothelium-derived nitric oxide (NO) production is reduced at the macrocirculatory and microcirculatory levels during sepsis. To examine this, rats were subjected to sepsis by cecal ligation and puncture (CLP). At 5 h after CLP (i.e., midpoint of hyperdynamic sepsis) or sham operation, the aorta and superior mesenteric artery were isolated. Responses to an endothelium-dependent vasodilator, acetylcholine (ACh), and an endothelium-independent vasodilator, nitroglycerin (NTG), were determined. In additional studies, the small intestine was isolated 5 or 20 h (hypodynamic sepsis) after CLP. Responses to ACh and NTG were determined in the isolated intestine. The results indicate that endothelium-dependent relaxation in both the aorta and superior mesenteric artery was depressed at 5 h after CLP. In contrast, there was no significant difference in the relaxation induced by NTG. Moreover, ACh- induced vascular relaxation in the isolated small intestine decreased at 5 and 20 h post-CLP without any significant alterations in NTG-induced relaxation. Since studies have shown that ACh-induced relaxation in the aorta is reduced at 20 h after CLP, it could be concluded that endothelium-derived NO release is depressed during hyperdynamic and hypodynamic stages of sepsis, not only in large arteries, but also at the microcirculatory level.