Tracking dynamic gap junctional coupling in live cells by local photoactivation and fluorescence imaging

Song Yang, Wen Hong Li

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Intercellular communication through gap junction channels is crucial for maintaining cell homeostasis and synchronizing physiological functions of tissues and organs. In this chapter, we present a noninvasive fluorescence imaging assay termed LAMP (local activation of a molecular fluorescent probe) that consists of the following steps: loading cells with a caged and cell permeable coumarin probe (NPE-HCCC2/AM), locally photolyzing the caged coumarin in one or a subpopulation of coupled cells, monitoring cell–cell dye transfer by digital fluorescence microscopy, and post-acquisition analysis to quantify the rate of junction dye transfer using Fick’s equation. The LAMP assay can be conveniently carried out in fully intact cells to assess the extent and degree of cell coupling, and is compatible with other fluorophores emitting at different wavelengths to allow multicolor imaging. Moreover, by carrying out multiple photo-activations in a coupled cell pair, LAMP assay can track changes in cell coupling strength between coupled cells, hence providing a powerful method for investigating the regulation of junctional coupling by cellular biochemical changes.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages181-191
Number of pages11
Volume1437
DOIs
StatePublished - 2016

Publication series

NameMethods in Molecular Biology
Volume1437
ISSN (Print)10643745

Keywords

  • Caged coumarin
  • Caged dye
  • Gap junction communication
  • Infrared-LAMP assay
  • LAMP assay
  • NPE-HCCC2/AM
  • Photoactivation

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Yang, S., & Li, W. H. (2016). Tracking dynamic gap junctional coupling in live cells by local photoactivation and fluorescence imaging. In Methods in Molecular Biology (Vol. 1437, pp. 181-191). (Methods in Molecular Biology; Vol. 1437). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-3664-9_13