Decrease in laser ablation threshold for epithelial tissue microsurgery in a living Drosophila embryo during dorsal closure.

In this work, we use a two-photon fluorescence microscope for combined imaging and laser tissue ablation of a living Drosophila Melanogaster embryo. By using tightly focused near-infrared femtosecond pulses at MHz repetition rate and of sub-nanojoule energy we are able to produce microsurgery on the epithelial tissue within a Drosophila embryo at the final stages of its embryonic development. Ablation was performed on labelled and unlabelled embryos during and after dorsal closure. We observed that ablation of GFP-labelled tissue required lower energy deposition than unlabelled tissue ensuring that the tissue ablation is mediated by multiphoton absorption of Green Fluorescent Protein (GFP). In addition, the energy deposition to produce ablation is further decreased during dorsal closure. These results show the presence of additional tensile forces on the tissue during dorsal closure. Furthermore, an increased activity of actin near the laser wounds was observed as the tissue heals.