Il 1β

Synaptoneurosomes were prepared from mouse hippocampus as described17 (link) and western blot analysis was performed as previously27 (link). Hippocampal proteins were separated by SDS-PAGE and immunoblotted using the following primary antibodies: P2X7R (1:200, Cat n: APR-004, Alomone labs), IL-1β (1:2000, Cat n: ab9772. Abcam), synaptophysin (1:1000, Cat n: S5768. Sigma-Aldrich), α-tubulin (1:2000, Cat n: T6199. Sigma-Aldrich) and β-actin (1:2000, Cat n: A5441. Sigma-Aldrich). Membranes were then incubated with horseradish peroxidase-conjugated secondary antibodies (Jackson ImmunoResearch, Plymouth, PA, USA) and bands visualized using Supersignal West Pico Chemiluminescence Substrate (Pierce, Rockford, IL, USA). Images were captured using a Fuji-Film LAS-300 (Fuji, Sheffield, UK), and densitometry was performed using AlphaEaseFC4.0 gel-scanning integrated optical density software (Alpha Innotech, San Leandaro, CA, USA).

TNF-α and IL-1β were obtained from Alomone (Jerusalem, Israel); Fasudil, Y-27632, ethidium (Etd⁺) bromide and lanthanum (La³⁺) chloride were obtained from Sigma-Aldrich (St. Louis, MO, USA). TBARS assay kit was obtained from Cayman Chemicals (Ann Arbor, MI, USA), and CellTiter96^® Non-Radioactive Cell Proliferation Assay was obtained from Promega (Madison, WI, USA). The mimetic peptides gap19 (KQIEIKKFK, intracellular loop domain of Cx43) and TAT-L2 (YGRKKRRQRRRDGANVDMHLKQIEIKKFKYGIEEHGK, second intracellular loop domain of Cx43) were obtained from GenScript (Piscataway Township, NJ, USA) [25 (link)]. The monoclonal anti-α-tubulin antibody was purchased from Sigma-Aldrich (St. Louis, MO, USA). The polyclonal anti-phosphorylated-MYPT1 (Thr696) antibody was obtained from Merck Millipore (Darmstadt, Germany). The monoclonal anti-MYPT1 antibody was purchased from BD Transduction Laboratories (San José, CA, USA). The monoclonal anti-unphosphorylated Cx43 antibody was obtained from Invitrogen (Carlsbad, CA, USA). Anti-mouse and anti-rabbit secondary antibodies conjugated to horseradish peroxidase were from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA).