Fig 2: 2-Deoxy glucose impairs cardiomyocyte proliferation.(a) Experimental design for the 2-DG injections to inhibit glycolysis in injured zebrafish hearts. (b) Confocal image of injured zebrafish hearts at seven dpi either injected with PBS or 2-DG stained for Mef2c (green) and PCNA (magenta). Zoom-in images of the borderzone are shown below overview pictures for both PBS (cyan box) and 2-DG (orange box). Arrowheads indicate nuclei positive for Mef2c and PCNA. Scale bar indicates 200 µm (overview) or 20 µm (zoom-in). (c) Quantification of the proliferating cardiomyocytes (double Mef2c/PCNA positive) in the border zone of PBS and 2-DG treated hearts represented as the proportion of proliferating cardiomyocytes compared to the average percentage in the PBS injected group. Each dot represents a single heart (three sections per heart analyzed). Hearts were pooled from two separate experiments. Error bars represent mean ± standard deviation. ***, p<0,0001.

Fig 3: Cardiac slices retain characteristics of the regenerating heart. (A) Schematic representation of workflow. (B) Immunolabeling on 5 dpi whole hearts, 5 dpi cardiac slices after 1 day culture and 5 dpi cardiac slices after 3 day culture from a Tg(nppa:mCitrine) transgenic line. Trabecular cardiomyocytes are marked with mCitrine (green) and embryonic myosin is marked with N2.261 (magenta). Scale bars: 200 µm (overview); 50 µm (zoom). (C) Immunolabeling on 5 dpi whole hearts, 5 dpi cardiac slices after 1 day culture and 5 dpi cardiac slices after 3 day culture from a Tg(nppa:mCitrine) transgenic line. Cardiomyocyte nuclei are marked with MEF2 (green) and proliferating nuclei are marked with PCNA (magenta). Scale bars: 200 µm (overview) or 10 µm (zoom). (D) Quantification of MEF2+/PCNA+ cells in the border zone of injured hearts/cardiac slices and remote zone of uninjured hearts/cardiac slices. Each dot represents a result from one heart or cardiac slice. Data are mean±s.d. *P<0.05, ***P<0.001, ****P<0.0001.