Non-apoptotic pioneer neutrophils initiate an endogenous swarming response in a zebrafish tissue injury model

Neutrophils are rapidly recruited to inflammatory sites where they are able to coordinate their migration to form clusters, a process termed neutrophil swarming. The factors which initiate neutrophil swarming are not understood, requiring the development of new in vivo models. Using transgenic zebrafish larvae to study neutrophil migration, we demonstrate that neutrophil swarming is conserved in zebrafish immunity, sharing essential features with mammalian systems. We identified that one pioneer neutrophil was sufficient to induce neutrophil swarming after adopting a distinctive morphology at the wound site, followed by the coordinated migration of neutrophils to form a swarm. Using a FRET reporter of neutrophil apoptosis, we demonstrate that pioneer neutrophils do not undergo caspase-3 mediated apoptosis prior to swarming. These data provide some of the first evidence of endogenous neutrophil migration patterns prior to swarming and demonstrate that the zebrafish can be used to dissect the mechanisms modulating neutrophil swarm initiation.