In vivo single-cell RNA metabolic labeling resolves early transcriptional responders in the regenerating zebrafish heart

The adult zebrafish heart can regenerate after injury, but the earliest gene expression changes that trigger this process remain poorly understood. Here we show that in vivo single-cell RNA metabolic labeling, which marks newly made RNA in individual cells, can capture rapid responses in the adult zebrafish heart after injury. Within the first 6 h, we detect activation of innate immune programs, including Toll-like receptor signaling, in a subset of macrophage-like immune cells. Analysis of a larger single-cell dataset indicates that neutrophils also contribute to this early response. Guided by these data, we show that macrophage-specific inhibition of the Toll-like receptor adaptor MyD88 reduces the pro-inflammatory macrophage response at the injury site and improves early hallmarks of regeneration. Our work establishes RNA metabolic labeling as a useful approach for measuring acute responses in vivo at single-cell resolution and identifies early immune-cell activation as a tunable component of heart regeneration.