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The Journal of Physiological Sciences

, Volume 61, Issue 2, pp 141–149 | Cite as

The beginning of the calcium transient in rat embryonic heart

  • Takeshi KobayashiEmail author
  • Sachiko Maeda
  • Nobutoshi Ichise
  • Tatsuya Sato
  • Takehito Iwase
  • Sumihiko Seki
  • Yoichi Yamada
  • Noritsugu Tohse
Original Paper

Abstract

Although many researchers have tried to observe the beginning of the heartbeat, no study has shown the beginning of the calcium transient. Here, we evaluate the beginning of the calcium transient in the Wistar rat heart. We first tried to reveal when the heart of the Wistar rat begins to contract because no previous study has evaluated the beginning of the heartbeat in Wistar rats. Observation of embryos transferred to a small incubator mounted on a microscope revealed that the heart primordium, the so-called cardiac crescent, began to contract at embryonic day 9.99–10.13. Observation of embryos loaded with fluo-3 AM revealed that the beginning of the calcium transient precedes the initiation of contraction which precedes the appearance of the linear heart tube. Nifedipine (1 μM), but not ryanodine (1 μM), abolished the calcium transients. These results indicate that calcium transients in the early embryonic period involve exclusively calcium entry through L-type calcium channels in contrast to the situation in mature hearts. This study provides the first demonstration of the relationship between morphological changes in the heart primordium and the beginning of the calcium transient and contraction.

Keywords

Cardiogenesis Developmental biology Excitation-contraction coupling Calcium transient 

Notes

Acknowledgments

We thank Michael. P. Walsh, University of Calgary, for help with writing this manuscript. This work was supported by KAKENHI (22700427), Grant-in-Aid for Young Scientists (B), and by a program for developing the support system for upgrading education and research.

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

Legend for Supplementary Movie 1 Representative movie showing the beginning of contraction of the rat heart primordium, corresponding to the data marked with the asterisk in Fig. 1A. Upper panel shows the time course. The filled circle represents the second when an embryo was removed from a pregnant rat. The thin line represents the period when the heart primordium did not contract. The bold line represents the period when the heart primordium contracted. Panels a to d show the movement of the whole heart primordium corresponding to the period a to d in the time course of upper panel. Panels a′ to d′ show scale-up movies corresponding to the blue rectangles in panels a to d. (MOV 9306 kb)

Legend for Supplementary Movie 2 Representative movie showing the beginning of the calcium transient and contraction of the rat heart primordium loaded with fluo-3 AM, corresponding to Fig. 2. Upper panel shows the time course. Left edge of the thin line indicates the start time of observation. The thin line represents the period when the heart primordium did not contract. The white bar represents the period when the calcium transient was observed but contraction was not observed. The black bar represents the period when the heart primordium contracted with calcium transients. Panels a to d show the calcium transient corresponding to the period a to d in the time course of the upper panel. Panels e to i show the movement of the heart primordium corresponding to the period e to i in the time course of the upper panel. To allow the contraction to be clearly recognized, panels e′ to i′ show scaled-up and filtered movies corresponding to the area surrounded by the rectangle in each panel e to i. The emphasis of the edge of the contraction area was achieved with ImageJ. The line in panels e′ to i′ corresponds to line 1 in Fig. 2F. Panel b shows the first calcium transients. The calcium transient had a small amplitude at first, which increased gradually. Panels g and g’ show the first contractions of the heart primordium. Along the line in panel g’, tissue motion was recognized from the middle of the movie. The direction of the first contractions was toward cranial. (MOV 10376 kb)

Legend for Supplementary Movie 3 Representative movie showing the effect of the L-type calcium channel blocker, nifedipine, on the beginning of the calcium transient, corresponding to Fig. 4. Upper panel shows the time course. Left edge of the thin line indicates the start time of observation. The thin line represents the period when the heart primordium did not contract. The white bar represents the period when the calcium transient was observed and the contraction was not observed. The black bar represents the period when the heart primordium contracted with calcium transients. The hatched box represents the period when the nifedipine (1 µM) was applied. Panels a to d show the calcium transient corresponding to the period a to d in the time course of the upper panel. Panel a shows the first calcium transients. The calcium transient was not recognized in panel c because of the application of nifedipine (1 µM). (MOV 10473 kb)

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Copyright information

© The Physiological Society of Japan and Springer 2011

Authors and Affiliations

  • Takeshi Kobayashi
    • 1
    Email author
  • Sachiko Maeda
    • 1
  • Nobutoshi Ichise
    • 1
  • Tatsuya Sato
    • 1
  • Takehito Iwase
    • 1
  • Sumihiko Seki
    • 1
  • Yoichi Yamada
    • 1
  • Noritsugu Tohse
    • 1
  1. 1.Department of Cellular Physiology and Signal TransductionSapporo Medical University School of MedicineSapporoJapan

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