<i>Ex vivo</i> imaging reveals neutrophil behaviors in the adult zebrafish heart

Current approaches to investigate cells in mature internal organs of vertebrate model organisms focus primarily on cell morphology, numbers, and spatial distribution using static images from fixed samples, failing to provide information on cell behavior. Here, we focus on the robust dynamics of neutrophils and analyze their motility and morphodynamic (i.e., time-resolved shape changes) behaviors in adult zebrafish hearts ex vivo. To modulate neutrophil activity, we exposed cryoinjured heart explants to dexamethasone (DEX) or lipopolysaccharide (LPS), anti- and pro-inflammatory drugs, respectively. While DEX halted neutrophil motility and morphodynamics, LPS drastically increased them, emphasizing the suitability of this experimental setup to modulate and investigate cell behavior. Detailed analyses identified parameters predictive of cell activity and confirmed that temporal changes in morphological parameters, including two-dimensional measures, are stronger indicators of cell behavior than their corresponding mean values. Profiling neutrophil behavior during cardiac regeneration further revealed temporal variation of behavioral states and substantial heterogeneity at each time point. Altogether, we introduce the use of a cardiac explant system to analyze cell behaviors over time, and present it as an additional platform for compound screening.