Journal: Clinical and Translational Medicine
Article Title: Low‐glucose‐sensitive TRPC6 dysfunction drives hypoglycemia‐induced cognitive impairment in diabetes
doi: 10.1002/ctm2.205
Figure Lengend Snippet: RH specifically represses hippocampal TRPC6 channel in diabetic mice. (A) Relative TRPC6 mRNA expression (n = 6) in PC12 cells cultured in medium with normal glucose concentration (NG, 5.6 mM) or medium with high glucose concentration (HG, 25 mM). NG+LG, cells cultured in NG medium and received three times of low glucose stimulation (12 hours per times, LG, 3.0 mM); HG+LG, cells cultured in HG medium and received three times of low glucose stimulation (12 hours per times, LG, 3.0 mM). (B) Representative Western blots of TRPC6 protein in PC12 cells in four groups. GAPDH served as a loading control. Quantitative data were shown on the right (n = 4). (C) Relative mRNA expression levels of TRPC6 in hippocampus from nondiabetic (ND) mice, ND mice with recurrent moderate hypoglycemia (ND‐RH), mice with STZ‐induced type 1 diabetes mellitus (DM), and DM mice with RH (DM‐RH) (n = 6 samples from six mice). (D) Representative Western blots of TRPC6 in hippocampus. GAPDH served as a loading control. Quantitative results are shown on the right (n = 3 tissues from three mice). TPRC6 −/− , TRPC6 global knockout mice. (E) Relative mRNA expression levels of TRPC1, TRPC3, TRPC4, TRPC5, and TRPC7 in hippocampus (n = 6–8 samples from four mice). * P < .05, ** P < .01, *** P < .001. Statistical significance was assessed using a one‐way ANOVA or Kruskal‐Wallis test
Article Snippet: The primary antibodies were used, including anti‐TRPC1 (ACC‐010, 120 kDa), anti‐TRPC3 (ACC‐016, 100 kDa), anti‐TRPC4 (ACC‐018, 97 kDa), anti‐TRPC5 (ACC‐020, 100 kDa), anti‐TRPC6 antibody (ACC‐017, 110 kDa), anti‐TRPC7 antibody (ACC‐066, 95 kDa), anti‐TRPV1 (ACC‐030, 90 kDa), and anti‐TRPV4 (ACC‐034, 100 kDa) from Alomone; anti‐LKB1 (PA5‐96062, 48 kDa), anti‐phospho‐CaMMK2 (PA5‐105225, 65 kDa), and anti‐phospho‐DRP1 (S616, PA5‐64821, predicted to react with Ser585 in rats and Ser600 in mice, 83 kDa) antibodies from Invitrogen; anti‐CaMKK2 (ab135979, 65 kDa), anti‐Drp1 (ab184247, 83 kDa), anti‐phospho‐DRP1 (S637, ab193216, predicted to react with Ser656 in rats and Ser616 in mice, 83 kDa), anti‐PSD95 (ab238135, 80 kDa), anti‐Synapsin I (ab254349, 74 kDa), and anti‐SYP (ab32127, 34 kDa) antibodies from Abcam; anti‐AMPKα (2532S, 62 kDa) and anti‐phospho‐AMPKα (Thr172, 2535S, 62 kDa) antibodies from CST, followed by incubation with the secondary antibodies (ZSGB‐BIO).
Techniques: Expressing, Cell Culture, Concentration Assay, Western Blot, Knock-Out