NADH FLIM reveals cellular bioenergetics and mitochondrial morphology during Drosophila convergent extension.

Mitochondria are dynamic organelles that can fragment or fuse to support different bioenergetic demands and cellular processes, although the role of mitochondrial fission and fusion during embryonic development is not well understood. Fluorescence lifetime imaging (FLIM) of the mitochondrial cofactor NADH can be used to visualize mitochondrial networks and infer aspects of cellular bioenergetics in a label-free manner. We used NADH FLIM to test whether germband cells undergo changes in cellular metabolism during Drosophila convergent extension (CE)--a process in which hundreds of epithelial cells undergo coordinated intercalation to elongate the embryo. Contrary to our expectations, we did not observe significant changes in NADH lifetime or mitochondrial topology during CE, suggesting that germband cells do not need to alter their baseline metabolism to fuel intercalation. Inhibiting mitochondrial fission led to hyper-fused basal networks and robustly increased NADH lifetime, whereas inhibiting fusion led to hyperfragmented apical networks and decreased NADH lifetime. Inhibiting either mitochondrial fission or fusion increased cell intercalation errors during CE, suggesting that a precise network topology is required for proper tissue elongation. These defects could be rescued by knocking down ROS scavengers, suggesting that one of the roles of mitochondria during CE is to create a ROS-rich environment to support cell motility. This study demonstrates the utility of NADH FLIM for visualizing mitochondria and characterizing bioenergetics during development in live embryos, and this technique should be broadly applicable to many other systems.