AIM: To determine the influence of acute hyperbaric oxygenation on the flow-induced dilation in healthy Sprague-Dawley rats by a method of measuring flow-induced dilation. MATERIALS AND METHODS: This study included 11 healthy Sprague-Dawley rats divided into two groups – control, CTRL (N=6) and A_HBOT group (N=5) which was exposed to acute hyperbaric oxygenation in hyperbaric oxygen chamber to 100% oxygen on 2 bar for 2 hours. After that procedure, rats were anesthetized with ketamine (75 mg/kg) and midazolam (0,5 mg/kg) and then decapitated. Middle cerebral arteries were isolated from rats’ brains, and then the arteries were incubated in the pressure myograph system 60 minutes under the pressure of Δ80 mmHg (P1 is Δ80mmHg, P2 is Δ0 mmHg) due to basal diameter estimation. After incubation, arteries were exposed to flow which is obtained by simultaneous changes of in-flow and out-flow pressure with pressure gradients Δ10, Δ20, Δ40, Δ60, Δ100 mmHg. After basal measurements of the arteries, flow-induced dilation is measured in presence of acetylcholine (ACh) and sodium nitroprusside (SNP). At the end of the protocol, PSS solution which was creating pressure gradient with its flow was replaced with PSS Ca-free solution, for the purpose of measuring maximum diameter of arteries. RESULTS: Flow-induced dilation of middle cerebral arteries has significantly decreased in the group treated with acute hyperbaric oxygen therapy, in comparison to healthy untreated animals. The dilation response to acetylcholine (Ach) induced dilation (AChID) of the middle cerebral artery, which is an endothelium-dependent vasodilation, is less sensitive in treated group than in control group. Such data suggests reduced sensitivity of the middle cerebral artery in treated animals after the application of acute hyperbaric oxygenation on endothelial-dependent vasodilation, which indicates an important role of endothelium in mechanism of flow-induced dilation. The blood vessel response to the endothelium-independent nitrogen oxide donor (SNP) was conserved and no differences were found between the two groups. CONCLUSION: Acute hyperbaric oxygenation distorts flow-induced dilation by increasing the level of oxidative stress.