Optical Clearing and Light Sheet Microscopy Imaging of Amphioxus

0555020 - ÚMG 2022 RIV CH eng J - Journal Article
Macháčová, Simona - Chmelová, Helena - Vávrová, Anna - Kozmik, Zbyněk - Kozmiková, Iryna
Optical Clearing and Light Sheet Microscopy Imaging of Amphioxus.
Frontiers in Cell and Developmental Biology. Roč. 9, July (2021), č. článku 702986. ISSN 2296-634X. E-ISSN 2296-634X
R&D Projects: GA ČR GA20-25377S
Institutional support: RVO:68378050
Keywords : amphioxus * light sheet microscopy * clearing technique * whole mount immunohistochemistry * photoreceptor * acetylated tubulin * melanopsin
OECD category: Cell biology
Impact factor: 6.081, year: 2021
Method of publishing: Open access
https://www.frontiersin.org/articles/10.3389/fcell.2021.702986/full

Cephalochordates (amphioxi or lancelets) are representatives of the most basally divergent group of the chordate phylum. Studies of amphioxus development and anatomy hence provide a key insight into vertebrate evolution. More widespread use of amphioxus in the evo-devo field would be greatly facilitated by expanding the methodological toolbox available in this model system. For example, evo-devo research on amphioxus requires deep understanding of animal anatomy. Although conventional confocal microscopy can visualize transparent amphioxus embryos and early larvae, the imaging of later developmental stages is problematic because of the size and opaqueness of the animal. Here, we show that light sheet microscopy combined with tissue clearing methods enables exploration of large amphioxus specimens while keeping the surface and the internal structures intact. We took advantage of the phenomenon of autofluorescence of amphioxus larva to highlight anatomical details. In order to investigate molecular markers at the single-cell level, we performed antibody-based immunodetection of melanopsin and acetylated-alpha-tubulin to label rhabdomeric photoreceptors and the neuronal scaffold. Our approach that combines light sheet microscopy with the clearing protocol, autofluorescence properties of amphioxus, and antibody immunodetection allows visualizing anatomical structures and even individual cells in the 3D space of the entire animal body.
Permanent Link: http://hdl.handle.net/11104/0329638