Mitofusin 1

To acquire specimens for immunoblot, the mice were perfused with cold PBS and one side of the hippocampus was dissected immediately (Wang et al., 2021b (link)). Hippocampal tissue was homogenized at 4°C in RIPA buffer (containing protease and phosphatase inhibitors) and followed by sonication (10 s × three times with a 10-s interval). Ten micrograms of protein was loaded for immunoblot as previously described (Wang et al., 2021b (link)). GAPDH was used as a loading control for total lysate immunoblots. Primary antibodies used were as follows: p-DRP(Ser616) (Thermo Fisher; #PA5-64821, 1:500), DRP1 (Cell Signaling, #8570, AB_10950498, 1:1000), mitofusin-1 (Abcam, #104274, 1:500), mitofusin-2 (Cell Signaling, #9482, AB_2716838, 1:1000), p-MFF (Ser146) (Cell Signaling, #49281, 1:1000), MFF (Cell Signaling, #84580, AB_2728769, 1:1000), Tom20 (Cell Signaling, #42406, AB_2687663, 1:1000), VDAC1 (Santa Cruz, #sc-390996, 1:50), OXPHOS (Abcam, #ab110413, AB_2629281, 1:2000), and GAPDH (Cell Signaling #2118s; AB_561053, 1:1,000) overnight at 4°C followed by incubation with an IR-dye-labeled secondary antibody for 1 hour and measured with LiCor Odyssey followed by densitometric analysis.

High glucose Dulbecco’s modified Eagle’s medium (DMEM) and heat-inactivated fetal bovine serum (FBS) were from GIBCO (Grant Island, NY, USA). Trypsin, uridine, penicillin, streptomycin and Tween-20 were from Sigma Aldrich (St. Louis, MO, USA).
All other chemicals were of analytical grade
The anti-Drp1(Western Blotting (WB): 1:2500) and anti-Opa1 (WB: 1:2500) were obtained from BD Transduction Laboratories (San Diego, CA, USA), Mitofusin 1 (WB 1:2500) was from Abcam (Cambridge, UK), Mitofusin 2 (WB 1:2500), β-actin (WB: 1:50 000) and Fis1 (WB 1:1000) from Sigma Aldrich (St. Luis, MO, USA). Alexa Fluor 488 Phalloidin and DAPI were purchased from Invitrogen (Eugene, OR, USA).
Secondary antibodies labelled with horseradish peroxidase (WB: 1:5000) were from Santa Cruz Biotechnology (Santa Cruz, CA, USA), and antibody used for immunofluorescence staining was purchased from Jackson ImmunoResearch Europe, Ltd. (Newmarket, UK) and Invitrogen (Eugene, OR, USA). IRDye 680 and IRDye 800 Secondary Antibodies (1:5000) were from Li-Cor Biosciences (Bad Homburg, Germany).