Direct In Vivo Manipulation and Imaging of Calcium Transients in Neutrophils Identify a Critical Role for Leading-Edge Calcium Flux

• Published in: Cell reports (Cambridge) Vol. 13; no. 10; pp. 2107 - 2117
• Main Authors: Beerman, Rebecca W., Matty, Molly A., Au, Gina G., Looger, Loren L., Choudhury, Kingshuk Roy, Keller, Philipp J., Tobin, David M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 15.12.2015; Elsevier
• Subjects: Animals; Animals, Genetically Modified; Calcium - metabolism; Calcium Signaling - physiology; Chemotaxis - physiology; Microscopy, Fluorescence - methods; Neutrophils - metabolism; Rats; TRPV Cation Channels - genetics; TRPV Cation Channels - metabolism; Zebrafish
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2015.11.010

Calcium signaling has long been associated with key events of immunity, including chemotaxis, phagocytosis, and activation. However, imaging and manipulation of calcium flux in motile immune cells in live animals remain challenging. Using light-sheet microscopy for in vivo calcium imaging in zebrafish, we observe characteristic patterns of calcium flux triggered by distinct events, including phagocytosis of pathogenic bacteria and migration of neutrophils toward inflammatory stimuli. In contrast to findings from ex vivo studies, we observe enriched calcium influx at the leading edge of migrating neutrophils. To directly manipulate calcium dynamics in vivo, we have developed transgenic lines with cell-specific expression of the mammalian TRPV1 channel, enabling ligand-gated, reversible, and spatiotemporal control of calcium influx. We find that controlled calcium influx can function to help define the neutrophil’s leading edge. Cell-specific TRPV1 expression may have broad utility for precise control of calcium dynamics in other immune cell types and organisms. [Display omitted] •Light-sheet microscopy in zebrafish reveals neutrophil calcium dynamics•Migrating zebrafish neutrophils display enriched leading-edge calcium flux•Mammalian TRPV1 allows drug-inducible, cell-specific regulation of calcium•Leading-edge calcium signals can be instructive in setting neutrophil direction In vivo visualization of subcellular calcium dynamics in leukocytes has been challenging. Beerman et al. deploy light-sheet microscopy to identify leading-edge calcium enrichment in migrating zebrafish neutrophils. Calcium manipulation using cell-specific expression of a mammalian ion channel or pharmacological inhibition implicates leading-edge calcium in directional control of migrating neutrophils.