L nio

Second‐order mesenteric arteries (MA) were cleaned from surrounding fat tissue and mounted on glass micropipettes by keeping the same orientation of the flow in a pressure myograph chamber (DanisMyoTechnology, Aarhus, Denmark) as previously described.²² (link) Briefly, vessels were perfused with oxygenated (95% O₂ and 5% CO₂) Krebs solution (mmol/L: NaCl 118, KCl 4.7, MgCl₂ 1.2, KH₂PO₄ 1.2, CaCl₂ 2.5, NaHCO₃ 25, and glucose 10.1) at 37°C, and maintained at 80 mm Hg for 30 minutes, before preconstriction with phenylephrine (1×10⁻⁶ mol/L) followed by a cumulative concentration‐response curve of acetylcholine (1×10⁻¹⁰ to 3×10⁻⁵ mol/L) to assess the integrity of the endothelium. Vessels with acetylcholine‐induced vasodilation lower than 70% were excluded. Concentration‐response curves of phenylephrine (1×10⁻⁹ to 3×10⁻⁵ mol/L) and S1P (1×10⁻¹² to 3×10⁻⁹ mol/L) were performed. Flow‐dependent vasodilation and myogenic tone were also assessed as previously described.²² (link) MA were incubated with 100 µmol/L of N5‐(1‐iminoethyl)‐l‐ornithine dihydrochloride (L‐NIO, Tocris, 15 minutes), nonspecific inhibitor of eNOS⁴¹ (link) followed by vasodilation in response to acetylcholine to assess basal and stimulated NO production.