Activation of AMPK leads to translocation of TFEB to the nucleus and increased expression of flcn and fnip2 in zebrafish. A) Embryos at 3 dpf were treated with 991 10 µM or vehicle (DMSO) for 16 h. Tissue lysates (20 µg) were resolved by SDS-PAGE and Western blot analysis was performed using the indicated antibodies. B) Tg(actc1b:tfeb-ZsGreen);Tg(actc1b:nls-mCherry) embryos at 3 dpf were treated with 991 10 µM or vehicle (DMSO) for 24 h. Embryos were mounted live in water containing 0.016% tricaine and imaged with ImageXpress confocal system at original magnification, ×20, the scale bar corresponds to 50 µm (n = 12). C) Schematic summary of the acute exercise study design using WT and prkaa1−/−;prkaa2−/− zebrafish. D) Immediately after the 3-h training session, muscle samples were collected from the zebrafish. Tissue lysates (20 µg) were resolved by SDS-PAGE and Western blot analysis was performed using the indicated antibodies. E) Relative mRNA transcript levels of flcn and fnip2 were determined by qPCR. Data are shown as box-and-whisker plots (minimum to maximum) of values normalized to control WT (n = 6). Two-way ANOVA with the factors of genetic background and exercise, plus the interaction between these 2 factors, was performed. The graph shows the significance of the interaction. *P < 0.05, ****P < 0.0001.
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