5 aminoimidazole 4 carboxamide 1 β d ribofuranoside aicar

The antibodies used were: mouse monoclonal anti-Kv1.4 (1:50, clone K13/31, UC Davis/NIH NeuroMab Facility), goat polyclonal anti-Kv7.1 (1:100), Alexa Fluor®488-conjugated donkey anti-mouse IgG (1:200) and Alexa Fluor®555-conjugated donkey anti-goat IgG (1:500). Alexa Flour®647 phalloidin (1:200) and 4′,6-diamidino-2-phenylindole (DAPI) was used to stain actin filaments and nuclei, respectively. All fluorescent antibodies and probes were purchased from Thermo Fischer Scientific.
The activators and inhibitors used in this study were as follows: 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR, Sigma-Aldrich, 0.5 mM), 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranosyl-5′-monophosphate (ZMP, Sigma-Aldrich), 2-O-Tetradecanoylphorbol 13-acetate (PMA, Sigma-Aldrich, 100 nM), 52-(4-Morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002, Sigma-Aldrich, 10 µM), 2-Cyclopentyl-4-(5-phenyl-1H-pyrrolo[2,3-b]pyridin-3-yl-benzoic acid (GSK650394, Tocris Bioscience, 1 µM).

5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR, an AMPK activator) and compound C (an AMPK inhibitor) were purchased from Sigma-Aldrich (St. Louis, MO, USA).

Cucurbitacin B (CuB, 100 μM), glucose (10%), denatonium benzoate (10 mM), 2-APB (10 μM), U73122 (10 μM), U73343 (10 μM), rolipram (50 μM), STO-690 (100 μM), Dorsomorphin (50 μM), forskolin (5 μM), and AICAR (5-Aminoimidazole-4-carboxamide-1-β-D-ribofuranoside, 1–5 mM) were purchased from Sigma-Aldrich (Sigma-Aldrich, MO, United States). Gallein (10 μM) was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, United States). Lactisole (10 μM) was purchased from Endeavour Speciality Chemicals (Daventry, United Kingdom). Metformin, Diabex tab was purchased from Daewoong Pharmaceutical Co., Ltd. (Gyeonggi-do, South Korea). PBS was purchased from Corning (NY, United States) Bovine serum albumin (BSA) was purchased from RMBIO (MT, United States). Exendin 9-39 was purchased from Abcam (Cambridge, United Kingdom).

The mPGES-1 inhibitor, AF3485 N-[9-(2-hydroxyethyl)-9H-carbazol-3yl]-2-(trifluoromethyl) benzamide (MW 398.38), was synthesized in Angelini S.p.A. (Rome, Italy). Lipopolysaccharide (LPS, derived from Escherichia Coli 026:B6), the irreversible peroxisome proliferator-activated receptor (PPAR)γ antagonist, GW9662, the complete Freund's adjuvant (CFA), the proteasome inhibitor MG132 (carbobenzoxy-Leu-Leu-leucinal), the cell-permeable AMP-activated protein kinase (AMPK) inhibitor, Compound C [ComC; {6-[4-(2-piperidin-1-ylethoxy) phenyl]-3-pyridin-4-ylpyrazolo [1,5-a] pyrimidine}], and cell-permeable activator of AMPK, AICAR (5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The selective COX-2 inhibitors, celecoxib was purchased from CCS.Chem.Co., Ltd (Zhejiang, China) while L-745337, was kindly provided by Merck Frosst Canada. The compounds were used in different experimental conditions, as described in detail below.