Who has the beat? Imaging subcellular calcium concentrations over rapid time scales.

Research Areas:

Cell biology, Cardiovascular biology, Calcium dynamics

Imaging Needs:

Sensitivity to infrared wavelengths

Imaging System:
  • Zeiss Axiovert 100 inverted microscope
  • 63× oil immersion lens
  • Sutter Instrument, LB-LS/Q17 fluorexcence excitation source
  • 175W Xenon lamp
  • Liquid light guide (LLG, 2 meters, 3 mm diameter)
  • Zeiss filter set 38 HE (470/40 nm; 525/50 nm)
  • Fluo-4AM
  • Hamamatsu ImagEM C9100-12 CCD camera
    • 100 frames/second
    • 8.192 mm square sensor
    • 512 × 512 pixels resolution
Imaging cellular events in real time

Find out how Fang Huang, Jeorg Bewersdorf and colleagues use the sCMOS technology in the ORCA-Flash4.0 camera to achieve video-rate imaging at nanometer scales. Read now.

THE QUESTION

What physiological changes in the cell contribute to the beat-to-beat variability in heart rate?

The human heart rate exhibits a certain amount of variability, with the length of time between each beat governed by a complex array of physiological interactions. Loss of this variability is associated with aging and disease.

Both the autonomic nervous system and cardiac pacemaker cells—sinoatrial node cells (SANC)—contribute to heart rate variability. The existing paradigm views the variability contributed by SANC as stemming from membrane ion channels. However, focus is now shifting to include the role of Ca++ release into the cytoplasm from the sarcoplasmic reticulum.

THE BARRIERS

In the absence of action potentials, the sarcoplasmic reticulum from SANC spontaneously genearates local calcium releases (LCRs). Do these LCRs also affect beat-to-beat cycle length in an individual SANC? Addressing this question requires the ability to rapidly acquire images and to precisely localize fluorescence—and the corresponding Ca++ levels—within the cell.

THE SOLUTION

Beat-to-Beat Variation in Periodicity of Local Calcium Releases Contributes to Intrinsic Variations of Spontaneous Cycle Length in Isolated Single Sinoatrial Node Cells
Oliver Monfredi, Larissa A. Maltseva, Harold A. Spurgeon, Mark R. Boyett, Edward G. Lakatta, Victor A. Maltsev
PLoS One. 2013; 8(6): e67247. PMCID: PMC3695077.

Monfredi, et al,1 developed a semi-automated image analysis method to detect LCRs in a given spontaneous cycle using a custom-made image analysis program (developed by Victor Maltsev), and measured Ca++ levels using Fluo-4 and Hamamatsu’s ImagEM C9100-12 EM-CCD camera. Through quantitatively precise localization and detection of fluorescence, Monfredi, et al,1 were able to show that the average period of LCRs is linked to overall beat-to-beat variability in the cycle length of SANC.

THE POSSIBILITIES

Monfredi, et al,1 relied on precise localization and fast frame rates of Hamamatsu’s ImagEM camera to measure and localize subcellular Ca++ concentrations. Imagine what’s possible with the even faster capabilities and wider fields-of-view of Hamamatsu’s ORCA-Flash4.0 technology. Learn more, read Exciting Advances Push the Limits of Visualization.

References

  1. Monfredi, et al. Beat-to-Beat Variation in Periodicity of Local Calcium Releases Contributes to Intrinsic Variations of Spontaneous Cycle Length in Isolated Single Sinoatrial Node Cells. PLoS One. 2013; 8(6): e67247. PMCID: PMC3695077.
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