Indomethacin

Salts used in preparation of aCSF were purchased from Fisher Scientific (UK). Indomethacin, aminoguanidine, n-acetyl-L-cysteine, uric acid, Coomassie Brilliant Blue, and agents used in the preparation of the patch pipette solution, apart from QX-314 (Tocris UK) and MK801, were purchased from Sigma (UK). Ketamine was supplied by Fort Dodge Animal Health Ltd (Southampton, UK). Drugs were stored as frozen, concentrated stock solutions in distilled water and applied by dilution in the aCSF perfusion immediately before use. The following drugs were supplied by Tocris UK or Ascent Scientific UK: DCS, ACBC (1-aminocyclobutane-1-carboxylic acid), DCKA (5,7-dichlorokynurenic acid), 2-AP5 (D,L-2-amino-5-phosphonopentanoic acid), MK801, D-serine, TTX, NBQX (6-nitro-7 sulphamoylbenzo[f]quinoxalone-2,3-dione disodium), bicuculline methiodide and picrotoxin.

The compounds and suppliers used in the experiments described throughout the manuscript were: sodium pentobarbital (Dolethal^®; Vetoquinol, Madrid, Spain); heparin sodium from Rovi (Madrid, Spain); atropine sulphate (Scharlau, Barcelona, Spain); indomethacin (Acofarma, Barcelona, Spain); alloxan monohydrate, 1-PBG, BRL-54443 maleate salt, glibenclamide, NA bitartrate, and 5-HT hydrochloride (Merck Life Sciences S.L.U., Madrid, Spain); 5-CT, 8-OH-DPAT, α-methyl-5-HT, AS-19, cisapride, CGS-12066B, L-694,247, LY310762 hydrochloride, ODQ, and SB 699551 (Tocris Bioscience, Bristol, UK).
All the compounds were dissolved in saline at the time of experimentation, except AS-19 (dissolved in EtOH 5%), cisapride (0.01 M HCl), and both indomethacin and glibenclamide (dissolved in a mixture of PEN). None of the vehicles alter either RPP or SBP.

Atropine, Au-MUDA-AT, carbachol, potassium chloride (Mettler Toledo, Greifensee, Switzerland) were dissolved in aqueous stock solutions; forskolin (Tocris, Bristol, UK) and indomethacin were dissolved in ethanol. If not indicated otherwise, drugs were from Sigma (Steinheim, Germany).

PSS for vessel dissection, isolation and preparation contained (in mM) the following: KCl 6.0, NaCl 112, NaHCO₃ 1.18, MgSO₄ 1.18, KH₂PO₄ 1.18, CaCl₂ 1.8 and glucose 10. The pH of the PSS was adjusted to and maintained at 7.4 by continuous flow of normal air into the PSS. The amount of 60 mM K⁺-PSS (60K) was prepared by equimolar replacement of NaCl with KCl in the PSS. Empagliflozin was purchased from Ambeed Inc. (Arlington Heights, IL, USA) and used in the concentration range of 0.001–100 µM. Phenylephrine (PE) and 4-aminopyridine (4-AP) were purchased from Sigma-Aldrich (St. Louis, MO, USA) and used at a final concentration of 1 µM and 1 mM, respectively. Indomethacin (10 µM), DPO-1 (1 µM), Linopirdine (10 µM), Psora-4 (100 nM), Glibenclamide (10 µM), paxilline (10 µM), ODQ (10 µM), KT 5823 (1 µM), L-NNA (10 µM), SNP (10 µM) and acetylcholine (ACh, 1 µM)) were purchased from Tocris (Minneapolis, MN, USA). 4-AP, SNP and PE were dissolved in distilled water. Empagliflozin, DPO-1, Linopirdine, Psora-4, Indomethacin, Glibenclamide, paxilline, ODQ, KT5823, L-NNA and ACh were dissolved in dimethyl sulfoxide (DMSO). The final DMSO concentration (<0.1%) in the myograph chamber had no significant effect on arterial contractility.