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Updated: Jun 12, 2026
An Antegrade Perfusion Method for Cardiomyocyte Isolation from Mice
Published on: May 19, 2021
Fetal and neonatal development of Ca2+ transients and functional sarcoplasmic reticulum in beating mouse hearts
Yoichi Kawamura1, Takahiro Ishiwata, Mari Takizawa
1Department of Pediatrics, National Defense Medical College, Tokorozawa, Japan.
Background:
It is generally accepted that Ca(2+)-induced Ca(2+) release is not the predominant mechanism during embryonic stages. Most studies have been conducted either on primary cultures or acutely isolated cells, in which an apparent reduction of ryanodine receptor density and alterations in the cell shape have been reported. The aim of the present study was to investigate developmental changes in Ca(2+) transients using whole hearts of mouse embryos and neonates.
Methods And Results:
Fluo-3 fluorescence signals from stimulated whole hearts were detected using a photomultiplier and stored as Ca(2+) transients. The upstroke and decay of Ca(2+) transients became more rapid from the late embryonic stages to the neonatal stage. After thapsigargin application (an inhibitor of the sarcoplasmic Ca(2+)-ATPase [SERCA]), time to 50% relaxation (T(50)) of Ca(2+) transients was significantly prolonged. There were no significant changes in T(50) after Ru360 application (an inhibitor of mitochondrial Ca(2+) uniporter). The rate of increase in the amplitude of Ca(2+) transients after caffeine application became larger during developmental stages.
Conclusions:
Ca(2+) homeostasis developmentally changes from a slow rise and decay of Ca(2+) transients to rapid kinetics after the mid-embryonic stage. SERCA began to contribute significantly to Ca(2+) homeostasis at early embryonic stages and sarcoplasmic reticulum Ca(2+) contents increased from embryonic to neonatal stages, whereas mitochondrial Ca(2+) uptake did not contribute to Ca(2+) transients on a beat-to-beat basis.
Insights
Cardiac calcium (Ca2+) handling matures during development. Sarcoplasmic reticulum Ca2+-ATPase (SERCA) significantly contributes to Ca2+ homeostasis in embryonic hearts, with increasing SR Ca2+ content from embryonic to neonatal stages.
Area of Science:
- Cardiovascular Physiology
- Developmental Biology
- Cellular Calcium Signaling
Background:
- Ca(2+)-induced Ca(2+) release is not the primary mechanism in embryonic development.
- Previous studies on isolated cells reported reduced ryanodine receptor density and altered cell shape.
- Investigating Ca(2+) transients in whole embryonic and neonatal mouse hearts is crucial.
Purpose of the Study:
- To investigate developmental changes in Ca(2+) transients in whole mouse hearts.
- To understand the role of SERCA and mitochondrial Ca(2+) uptake in embryonic cardiac function.
Main Methods:
- Detection of Ca(2+) transients using Fluo-3 fluorescence and photomultiplier in stimulated whole hearts.
- Application of thapsigargin (SERCA inhibitor) and Ru360 (mitochondrial Ca(2+) uniporter inhibitor).
- Analysis of Ca(2+) transient kinetics, including upstroke, decay, and relaxation time (T(50)).
Main Results:
- Ca(2+) transient upstroke and decay accelerated from late embryonic to neonatal stages.
- Thapsigargin significantly prolonged T(50), indicating SERCA's role in relaxation.
- Ru360 did not significantly alter T(50), suggesting minimal mitochondrial Ca(2+) contribution to beat-to-beat transients.
- Caffeine-induced Ca(2+) transient amplitude increase was larger in later developmental stages.
Conclusions:
- Cardiac Ca(2+) homeostasis shifts from slow to rapid kinetics post-mid-embryonic stage.
- SERCA becomes a significant contributor to Ca(2+) homeostasis early in embryonic development.
- Sarcoplasmic reticulum Ca(2+) content increases during development.
- Mitochondrial Ca(2+) uptake does not play a significant role in beat-to-beat Ca(2+) transients.
