Objectives: To determine the role of the Tff3 peptides in the endothelium-dependent vascular reactivity of the isolated blood vessels and to clarify the specific mechanisms of Tff3 peptide affecting the flow-induced vasodilation in Tff3−/−/C57BL/6N gene knockout mice and wild-type mice (C57BL/6N). Methods: Ten weeks old healthy male specimens were fed with standard (0,4% NaCl) and high-salt (4% NaCl) commercial rodent chow. They took food and water (or water with diluted TEMPOL, 1 mM) ad libitum over a period of 7 days. Carotid artery reactivity to gradual increase of the flow was measured in the presence or without the NO synthase (NOS) inhibitors, Nω-nitro-L-arginine methyl ester (L-NAME); cyclooxygenase inhibitors, indomethacin (INDO); the CYP450 epoxidase selective inhibitor MS-PPOH; agonists and antagonists of calcium-sensitive potassium channels (KCa2+, NS-1619; iberiotoxin); ATP-sensitive potassium channels (KATP, cromakalim; glibenclamide) and TRPV4 (GSK1016790A; RN-1734). The levels of nitric oxide and superoxide were determined using in situ fluorescence technique. The activity of the antioxidant enzyme GPx in the serum was measured in line with the standardised spectrophotometric protocol. The gene expression of antioxidant enzymes (Cu/Zn SOD, GPx1 and CAT) was determined using the quantitative real-time PCR method. Results: The high-salt diet (HS) impaired the flow-induced response of the wild type (WT) mice carotid artery. The recovered flow-induced dilation was observed under in vivo conditions in the presence of the superoxide “scavenger” TEMPOL in WT HS mice. The increased vascular oxidative stress was confirmed with the estimated levels of endothelial superoxide/ROS in carotid artery. The dilation in TFF3 LS, WT LS i WT HS groups was equally dependent on the levels of NO, the cyclooxygenases and EET metabolites, while the dilation in the TFF3 HS group was COX-dependent. In WT LS group, the TRPV4, Ca2+-activated K+ channels and KATP channels have a FID role, while in the TFF3 LS group the Ca2+-activated K+ channels and KATP channels have a significant role. In the TFF3 HS group, the agonists of TRPV4 and KATP channels increased the FID, while in the WT HS group the FID was increased by the agonists of the TRPV4 i KCa2+ channels. The increased vascular oxidative stress was confirmed with the estimated levels of endothelial superoxide/ROS in carotid artery. WT mice have a substantially lower catalase gene expression. The HS diet resulted in the substantially reduced GPx activity in the WT HS group. Conclusion: Due to the increased oxidative stress, the HS diet substantially decreases the vasodilation in the WT mice, whereas it does not affect the response of TFF3 knockout mice. The knockout of the Tff3 gene decreases the vasodilation compared to WT mice. The HS diet and the Tff3 gene knockout alter the FID mechanisms and ion channel involvement in FID.