Our objective was to test the hypothesis that acute exposure of human skin vasculature to nicotine may have deleterious effects on endothelial function. Vasoconstriction and vasorelaxation in isolated perfused human skin flaps (~8 × 18 cm) derived from dermolipectomy specimens were assessed by studying changes in skin perfusion pressure measured by a pressure transducer, and skin perfusion was assessed by a dermofluorometry technique (n = 4 or 5). It was observed that nicotine (10⁷ M) amplified (P < 0.05) the norepinephrine (NE)-induced concentration-dependent (10⁷-10⁵ M) increase in skin vasoconstriction compared with the control. This amplification effect of nicotine in NE-induced skin vasoconstriction was not blocked by the nicotine-receptor antagonist hexamethonium (10⁶ M) or the cyclooxygenase inhibitor indomethacin (10⁵ M). It was also observed that ACh and nitroglycerin (NTG) elicited a concentration-dependent (10⁸-10⁵ M) vasorelaxation in skin flaps preconstricted with 8 × 10⁷ M of NE. The vasorelaxation induced by ACh was attenuated (P < 0.05) in the presence of nicotine (10⁷ M) compared with the control. However, skin vasorelaxation induced by NTG was not affected by nicotine (10⁷ M). ACh and NTG are known to induce endothelium-dependent and -independent vasorelaxation, respectively. The present findings were interpreted to indicate that acute exposure of human skin vasculature to nicotine was associated with 1) amplification of NE-induced skin vasoconstriction and 2) impairment of endothelium-dependent skin vasorelaxation. Cyclooxygenase products and nicotine receptors blocked by hexamethonium were not involved in the amplification of NE-induced skin vasoconstriction by nicotine. These findings may provide further insight into the pathogenesis of skin vasospasm in skin flap surgery and skin ischemic disease associated with cigarette smoking or use of smokeless tobacco.