Figure 6.

The relative impact of 5-aminoimidazole-4-carboxamide riboside (AICAR) on AMPK activation in the old mouse heart. A: representative Western blots for phosphorylated AMPK (pAMPK) and total AMPK in the cardiac lysate obtained from 21-mo-old female and male mice. Hearts were excised 3 h after a single injection with AICAR (0.166 mg/g body wt) or saline. Each blot contains a sex-unmatched reference: S, saline; A, AICAR, for the sex-unmatched lanes. Diagrams represent pAMPK/AMPK ratio (B) and pAMPK level (C); each circle or triangle depicts a unique replicate (n = 3 samples/group). Relative normalization of the two separated blots was ensured by using the sex-unmatched references loaded on each blot. P value was calculated using a one-way ANOVA test, followed by a post hoc study to determine the significant differences among the groups. P values were then adjusted for multiple parameters. D: impact of higher dose of AICAR (0.5 mg/g body wt) on old male heart function. Effect of AICAR treatment on the LAV changes was determined by repeated-measures (RM) one-way ANOVA test/each group (n = per group). Overall significance between two groups along the time of treatment was assessed using RM two-way ANOVA test. E: representative Western blot for pAMPK and total AMPK in lysates of cardiac fibroblasts (from male and female old mice) preincubated with 0.5 mM AICAR (+) or DMSO (−) for 30 min. F: diagram representing the change in pAMPK/AMPK ratio in cardiac fibroblasts (n = 5–7 samples/group). P value was calculated using an one-way ANOVA test, followed by a post hoc study with Tukey correction to determine the significant differences among the groups. P values were adjusted for multiple parameters.