Zebrafish as a Model to Study Chemokine Function

Zebrafish have emerged as a powerful model organism to study embryo morphogenesis. Due to their optical clarity, they areuniquely suited for time-lapse imaging studies, providing insights into the dynamic processes underlying tissue formationand cell migration. These studies have been tremendously facilitated by the availability of transgenic zebrafish lines, labellingdistinct embryonic structures, individual cells, or even subcellular structures, such as the nucleus. Zebrafish studies haverevealed that the migration of several different cell types in the embryo is controlled by chemokines, small vertebrate-specificproteins. Here, we report methods to analyze the expression pattern of a given chemokine and its receptor in transgenic zebrafishusing fluorescent in situ hybridization in combination with an anti-green fluorescent protein (GFP) antibody staining. Wefurthermore illustrate how to image migrating cell populations using time-lapse microscopy in double-transgenic embryos. Weshow how to investigate cell number and direction of migration by using a nuclear-localized GFP. The combination of this transgenewith a membrane-targeted red fluorescent protein allows for the simultaneous determination of changes in cell shape, suchas the formation of filopodial extensions. We exemplify this by describing how a mutation in the chemokine receptor
cxcr4a
affects endothelial cell migration and blood vessel formation. Finally, we provide a method to perform fluorescent angiographyto monitor blood vessel perfusion in chemokine receptor mutants.