The effects of exchange transfusion (ET) with a modified hemoglobin (pyridoxalated hemoglobin polyoxyethylene conjugate, or PHP) on endothelium-dependent relaxation of microvasculature were examined in rat mesenteric arterioles using image-splitter television microscopy. To examine the endothelium-dependent relaxation, we applied acetylcholine (ACh) to arterioles preconstricted with norepinephrine (NE) and measured changes in arteriolar diameter. Topical application of NE (6 x 10(-8) to 6 x 10(-6)M) decreased the diameter of the arterioles dose-dependently to a mean of 45% (SE +/- 4%, n = 6) of the control. Topical application of ACh (7 x 10(-8) to 7 x 10(-6)M) increased the diameter of the arterioles preconstricted with NE in a dose-dependent manner and almost fully reversed the NE-induced decrease in diameter. ET with 6% PHP solution (30 ml/kg) induced an increase in the mean arterial blood pressure (MABP) from 117 +/- 5 to 132 +/- 4 mm Hg (n = 6, p < 0.05), with a concomitant decrease in diameter of arterioles, from 22.6 +/- 2.5 to 19.8 +/- 2.7 microns (p < 0.05). The MABP and arteriolar diameter gradually returned to the control level within 30 min after ET. Arteriolar changes in diameter in response to NE and ACh examined 30 min after ET were similar to those of the control. ET with 6% stroma-free hemoglobin (SFH; 30 ml/kg) did not alter arteriolar changes in diameter in response to NE and ACh. Results suggest that circulating PHP and SFH moieties do not interfere with endothelium-derived relaxing factor(s) induced by ACh in rat mesenteric microvasculatures in situ.