High-speed digital imaging of cytosolic Ca2+ and contraction in single cardiomyocytes

B. O'Rourke, D. K. Reibel, A. P. Thomas

Research output: Contribution to journalArticlepeer-review

Abstract

A charge-coupled device (CCD) camera, with the capacity for simultaneous spatially resolved photon counting and rapid frame transfer, was utilized for high-speed digital image collection from an inverted epifluorescence microscope. The unique properties of the CCD detector were applied to an analysis of cell shortening and the Ca2+ transient from fluorescence images of fura-loaded cardiomyocytes. On electrical stimulation of the cell, a series of sequential subimages was collected and used to create images of Ca2+ within the cell during contraction. The high photosensitivity of the camera, combined with a detector-based frame storage technique, permitted collection of fluorescence images 10 ms apart. This rate of image collection was sufficient to resolve the rapid events of contraction, e.g., the upstroke of the Ca2+ transient (< 40 ms) and the time to peak shortening (< 80 ms). The technique was used to examine the effects of β-adrenoceptor activation, fura-2 load, and stimulus frequency on cytosolic Ca2+ transients and contractions of single cardiomyocytes. β-Adrenoceptor stimulation resulted in pronounced increases in peak Ca2+, maximal rates of rise and decay of Ca2+, extent of shortening, and maximal velocities of shortening and relaxation. Raising the intracellular load of fura-2 had little effect on the rising phase of Ca2+ or the extent of shortening but extended the duration of the Ca2+ transient and contraction. In related experiments utilizing differential-interference contrast microscopy, the same technique was applied to visualize sarcomere dynamics in contracting cells. This newly developed technique is a versatile tool for analyzing the Ca2+ transient and mechanical events in studies of excitation-contraction coupling in cardiomyocytes.

Original languageEnglish (US)
Pages (from-to)H230-H242
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume259
Issue number1 28-1
DOIs
StatePublished - 1990
Externally publishedYes

Keywords

  • Isolated cardiomyocytes
  • charge-coupled device
  • contractility
  • fura-2
  • isoproterenol
  • β-adrenergic receptors

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

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