Fig. 4

HFD promotes synaptic IR in wild-type and heterozygous TBA2.1 mice. Representative immunoblots of total protein extracts from hippocampus (a) and cortex (e) of wild-type (+/+) or heterozygous TBA2.1 (Tg) mice fed with a RD or HFD. Membranes were probed with antibodies detecting pAKT (Ser473), AKT, pIRS1 (Ser612) and IRS1 as well as β-actin as a loading control. Scatter dot plots of the normalized pAKT (b, f), IRS1 (c, g) and pIRS (d, h) in hippocampus (n = 10) and cortex (n = 10). i Endogenous PI3Kγ was immunoprecipitated from hippocampal lysates of wild-type and heterozygous TBA2.1 mice fed with a RD or HFD. The amount of PI(3,4,5)P₃ produced by the enzyme coupled to the beads was determined by ELISA. The inhibitor AS-605240 was added to the wild-type/RD control group and used as a negative control (n = 6). j, k Immunoblot analysis showing no differences in the amounts of immunoprecipitated PI3Kγ between wild-type or heterozygous TBA2.1 mice fed with a RD or HFD (n = 6). ***P < 0.001, **P < 0.01, *P < 0.05 versus control by two-way ANOVA followed by Bonferroni’s post-hoc test. n.s. = non-significant. Two-tailed Student’s t-test was done to compare +/+ RD versus +/+ RD treated with AS-605240 in i and k. Data are presented as the mean ± SEM