Pnu282987

The following drugs were used: PNU-282987, Tocris Bioscience (Bristol, UK); DMSO, LLC Tula Pharmaceutical Factory (Tula, Russia); saline, REACHIM (Old Kupavna, Moscow Region, Russia).

Cells were grown on laminin-covered glass placed on 24-well plates under the same conditions described in Section 2.4. To conduct the electrophysiological recording of the α7 nAChR-mediated macroscopic currents, a whole-cell patch clamp was set up as follows. Cells attached to the glass were transferred to a bath filled with extracellular electrophysiology solution (140 mM NaCl, 2 mM CaCl₂, 2.8 mM KCl, 4 mM MgCl₂, 20 mM HEPES, 10 mM glucose, pH 7.4). A microelectrode pipette was filled with intracellular buffer solution (140 mm CsCl, 6 mm CaCl₂, 2 mm MgCl₂, 2 mm MgATP, 0.4 mm NaGTP, 10 mm HEPES/CsOH, 20 mm BAPTA/KOH). A microelectrode was attached to the cell membrane until a resistance of at least 1 GOhm was reached. After establishing the gigaseal, the cell membrane was ruptured using suction and the recordings were created. A typical experiment consisted of 1 s cell wash with control extracellular buffer, then the bath solution was changed to an extracellular solution with 1, 10, or 100 μM of α7 nAChR-selective agonist PNU-282987 (Tocris, Bristol, UK) supplemented with 10 μM of α7 nAChR-selective positive allosteric modulator PNU-120596 (Tocris) for 5 s followed by 5 s wash-out with control extracellular buffer.

The agonist PNU282987 and antagonist MLA were purchased from Tocris Bioscience (Bristol, UK), and LPS was purchased from Sigma–Aldrich (St. Louis, MO, USA). The [A10L]PnIA linear crude peptide was synthesized by Bankpeptide Biological Technology Co., Ltd. (Hefei, China) and purified in our laboratory.
A CCK-8 kit, DMEM-H medium, Penicillin/Streptomycin Solution, and PBS buffer were purchased from Sangon Biotech (Shanghai, China). FBS was purchased from Procell (Wuhan, China). A FastPure Cell/Tissue Total RNA Isolation Kit, HiScript II Q Select RT SuperMix for qPCR (+gDNA wiper), 2 × Taq Plus Master Mix Ⅱ, and ChamQ Universal SYBR qPCR Master Mix were obtained from Vazyme (Nanjing, China). A TNF-α ELISA kit, IL-6 ELISA kit, and IL-1β ELISA kit were purchased from Thermo Scientific (Waltham, MA, USA).

The groups VAChT-KD^HOM + LPS + PNU and WT + LPS + PNU received PNU 282987 treatment (Tocris, Bristol, UK), intraperitoneal, an agonist the α7 nAChR, in the dose of 10 mg/kg per animal, six hours after LPS instillation [16 (link)]