Vascular Tissue Preparation for Ex Vivo Studies

Main conduit renal and third-order mesenteric arteries were microdissected and cleaned of adherent fat in ice-cold Krebs solution, as previously described.^{24 (link)} Arteries were incubated in DMEM/F-12 culture medium (Sigma Aldrich, Dorset, United Kingdom) supplemented with 1% penicillin-streptomycin (Sigma Aldrich, Dorset, United Kingdom) in a 37°C incubator with 5% CO₂ for 16 hours (for myo-inositol/raffinose treatments) or 48 hours (for gene-silencing experiments). In double knockdown experiments, arteries were incubated in the morpholinos transfection mix for 32 hours and then incubated in medium containing either vehicle or raffinose plus myo-inositol for 16 hours.