title: Direct In Vivo Manipulation and Imaging of Calcium Transients in Neutrophils Identify a Critical Role for Leading-Edge Calcium Flux.
abstract: 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.
authors.affiliation: Department of Molecular Genetics and Microbiology, Duke University, Durham, NC 27710, USA.
authors.forename: Rebecca W
authors.initials: RW
authors.lastname: Beerman
authors.affiliation: Department of Molecular Genetics and Microbiology, Duke University, Durham, NC 27710, USA; University Program in Genetics and Genomics, Duke University, Durham, NC 27710, USA.
authors.forename: Molly A
authors.initials: MA
authors.lastname: Matty
authors.affiliation: Department of Molecular Genetics and Microbiology, Duke University, Durham, NC 27710, USA.
authors.forename: Gina G
authors.initials: GG
authors.lastname: Au
authors.affiliation: Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, VA 20147, USA.
authors.forename: Loren L
authors.initials: LL
authors.lastname: Looger
authors.affiliation: Department of Biostatistics and Bioinformatics, Duke University, Durham, NC 27710, USA.
authors.forename: Kingshuk Roy
authors.initials: KR
authors.lastname: Choudhury
authors.affiliation: Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, VA 20147, USA.
authors.forename: Philipp J
authors.initials: PJ
authors.lastname: Keller
authors.affiliation: Department of Molecular Genetics and Microbiology, Duke University, Durham, NC 27710, USA; Department of Immunology, Duke University, Durham, NC 27710, USA. Electronic address: david.tobin@duke.edu.
authors.forename: David M
authors.initials: DM
authors.lastname: Tobin
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002633:Chemotaxis / Q000502:physiology; D008856:Microscopy, Fluorescence / Q000379:methods*; D009504:Neutrophils / Q000378:metabolism*; D051381:Rats; D050916:TRPV Cation Channels / Q000235:genetics / Q000378:metabolism*; D015027:Zebrafish
chemical_list: D050916:TRPV Cation Channels; D002118:Calcium