Figure 4. Bin1 deletion in cardiomyocytes impairs GLUT4 translocation and glucose utilization, weakening systemic glucose response to insulin in mice.

(A) Representative GLUT4 confocal images in isolated cardiomyocytes from WT and Bin1 HT mice with or without prior AAV9-cBIN1 rescue. Cardiomyocytes were treated with 0 or 10 nM insulin for 20 minutes before collection. The images of the boxed areas with the corresponding fluorescence intensity profiles are included in the bottom panels. Scale bar: 10 μm. (B) Quantification of GLUT4 peak power density at t-tubules (n = 30–47 cells from 3 hearts per group). (C) 2DG uptake following insulin (0 and 10 nM) stimulation in cardiomyocytes isolated from each group (n = 10–12 repeats from 3 animals per group). (D) ¹³C enrichment of extracellular lactate and intracellular glycolysis and TCA cycle intermediates from cardiomyocytes isolated from each group (n = 3–5). Schematic for metabolic pathway of ¹³C₆-glucose is included. Pyr., pyruvate; Lac., lactate; Glc, glucose; F6P, fructose-6-phophate; F1,6BP, fructose-1,6-biphosphate; DHAP, dihydroxyacetone phosphate; 3PG, 3-phosphoglyceric acid; PEP, phosphoenolpyruvate; Cit., citrate; α-KG, alpha-ketoglutarate; Succ., succinate; Fum., fumarate; Mal, malate. (E) Percent of peak blood glucose during iGTT in each group (n = 4). (F) Percent of baseline blood glucose during iITT in each group (n = 4). Data are presented as mean ± SEM. Two-way ANOVA followed by Tukey’s (3 treatment groups, B–F) or Bonferroni’s (2 insulin doses, B and C) test is used. *, **, *** indicates P < 0.05, 0.01, 0.001, respectively, for comparison versus WT + PBS (B–F); ^†, ^††, ^††† indicates P < 0.05, 0.01, 0.001, respectively, for comparison between Bin1 HT + PBS and Bin1 HT + cBIN1 (B–F); and ^#, ^### indicates P < 0.05, 0.001, respectively, for comparison of 10 nM versus 0 nM insulin within each group (B and C).