Measuring Ca²⁺ sparks in cardiac myocytes

Niall Macquaide, Virginie Bito, Karin R. Sipido*

*Corresponding author for this work

Research output: Contribution to journalArticle

Abstract

This protocol describes the measurement of Ca(2+) sparks in intact myocytes by using a Ca(2+)-sensitive dye and imaging using laser scanning confocal microscopy. It takes advantage of spontaneous Ca(2+)-release events-sparks-using them as a measure of the activity of ryanodine receptors (RyRs). Two methodologies are described: One requires that cardiomyocytes be stimulated, preferably under voltage clamp by depolarizing pulses, until steady-state is reached, and then stimulation is stopped and Ca(2+) sparks are recorded. The second requires that cells be permeabilized and bathed in a solution to load the cell with Ca(2+) sufficient to elicit Ca(2+) sparks, but not Ca(2+) waves. These are then analyzed offline to quantify spark frequency and morphology. The advantages and disadvantages of each approach are discussed.

Original languageEnglish
Pages (from-to)490-7
Number of pages8
JournalCold Spring Harbor protocols
Volume2015
Issue number5
DOIs
Publication statusPublished - 1 May 2015

Keywords

  • Calcium/analysis
  • Cytological Techniques/methods
  • Fluorescent Dyes/metabolism
  • Microscopy, Confocal/methods
  • Myocytes, Cardiac/chemistry
  • Patch-Clamp Techniques/methods
  • Staining and Labeling/methods

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