Fig. 4
Monitoring NADH/NAD+ dynamics by SoNar in zebrafish embryos at 24 or 48 h post-fertilization. In vivo fluorescence imaging A-B and quantification C of zebrafish larvae (A, 24 hpf; B, 48 hpf) expressing SoNar or iNapC in response to 5 μM rotenone (mitochondria complex I inhibitor) indicating regions of interest (white dashed line). Highly viable wild-type AB zebrafish embryos were collected at one or two-cell stage and 1 nL sensor protein in HEPES buffer was injected into the animal pole. Embryos were collected and placed in a Petri dish with egg water (60 μg/mL Sea salts) at 28 °C. Larvae was anesthetized with 0.6 mM tricaine in E3 medium (5 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl2, 0.33 mM MgSO4) and then imaged via a Leica TCS SP8 SMD confocal laser scanning microscope with a HCX Plan APO CS 10/0.40 NA or HC Plan FLUOTAR 5/0.15 NA dry objective. Zebrafish larvae in 48 hpf were dechorionated under the stereomicroscope using fine tweezers. All fluorescence images were pseudocolored by F405/F488 (R405/488). Scale bar, 100 μm A or 200 μm B. The handling procedures were approved by the Institutional Animal Care and Use Committee of Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. For A-C, adapted from the reference (Zou et al. 2018)