Nor binaltorphimine dihydrochloride

The A1 adenosine receptor (A1AR) agonist n6-cyclohexyladenosine (CHA, 1mM, Sigma Aldrich); the NMDA receptor antagonist (2R)-amino-5-phosphonopentanoate (AP5, 5 mM, Tocris); the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione disodium salt hydrate (CNQX, 5 mM, Tocris); the peptidase inhibitor (2S,3R)-3-Amino-2-hydroxy-5-methylhexanoyl-Val-Val-Asp hydrochloride hydrate (Amastatin, 1 mM, Sigma Aldrich); Dynorphin A (100 μM, Tocris); and the κ-opioid receptor antagonist nor-Binaltorphimine dihydrochloride (nor-BNI, 27 μM intraparenchymal or 1.6 mM ICV, Tocris) were dissolved in isotonic saline.

The following drugs were used: (i) acetylsalicylic acid (ASA), apamin, atropine, bradykinin, caffeine, capsaicin, capsazepine (CAPZ), charybdotoxin, glibenclamide, haloperidol, l-glutamic acid, phorbol 12-myristate 13-acetate (PMA), pindolol, tetraethylammonium chloride, and yohimbine were purchased from Sigma-Aldrich (St. Louis, MO, USA); (ii) naltrindole hydrochloride, nor-binaltorphimine dihydrochloride and β-funaltrexamine hydrochloride were purchased from Tocris Bioscience (Ellisville, Missouri, USA); and (iii) acetic acid, dimethyl sulfoxide (DMSO), and methanol were purchased from Fisher Scientific (England). bradykinin, capsaicin, l-glutamic acid, and PMA were dissolved in physiological saline (0.9% (w/v) NaCl), while ASA, MECN, and CAPZ were dissolved in distilled water containing 10% DMSO (v/v). The vehicle used alone had no effects per se on the nociceptive responses in mice. All drugs, chemicals, and MECN solutions were administered in 10 mL/kg volumes and were freshly prepared just before being used.

Stock solutions of a broad-spectrum opioid antagonist (naloxone hydrochloride (Tocris, Bristol, UK)), a μ-receptor antagonist (naloxonazine dihydrochloride (Tocris)), a δ-receptor antagonist (naltrindole hydrochloride (Tocris)) and a κ-receptor antagonist (nor-binaltorphimine dihydrochloride (Tocris)) were dissolved in pyrogen-free saline (PFS). Pilocarpine (1 mg/μl, Sigma-Aldrich, St. Louis, MO, USA) was also dissolved in PFS. The stock solutions were stored at 4 °C until use. Our previous results and others have indicated that the appropriate microinjection dosage for naloxonazine, naltrindole and nor-binaltorphimine to selectively block μ-, δ- and κ-opioid receptors, respectively, without interaction with other opioid receptor subtypes, is within 20 μg [12 , 13 , 32 (link), 33 (link)]. In the current study, naloxone, naloxonazine, naltrindole and nor-binaltorphimine were microinjected at a dose of 10 μg/μl, which according to our previous studies efficiently exhibits pharmacological blockade [12 , 13 ]. The total volume for each injection was 1 μl and the duration of injection was 3 to 5 min. Our previous study has demonstrated that microinjection of 1 μl solution into the CeA does not cause CeA lesion [34 (link)].