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Updated: May 3, 2026
Isolating Myofibrils from Skeletal Muscle Biopsies and Determining Contractile Function with a Nano-Newton Resolution Force Transducer
Published on: May 7, 2020
Phase-targeting rapid cryofixation of the beating heart and histological analysis unveil contractile state-dependent
Shoko Tamura1, Kentaro Mochizuki2, Yasuaki Kumamoto3,4
1Department of Pathology and Cell Regulation, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto, 602-8566, Japan.
Insights
Cardiac phase-targeting cryofixation reveals sarcomere length changes during heartbeats. Sarcomere length (SL) is shorter in systole and longer in diastole, with some variations observed within the myocardium.
Area of Science:
- Cardiology
- Cellular Biology
- Biophysics
Background:
- The heart functions as a syncytium of cardiomyocytes.
- Coordinated contractions and relaxations are essential for cardiac function.
- Understanding sarcomere length (SL) dynamics is crucial for cardiac physiology.
Purpose of the Study:
- To investigate phase-dependent differences in myocyte sarcomere arrangements during the cardiac cycle.
- To analyze spatial inhomogeneity of sarcomere length (SL) within the myocardium.
- To develop a novel cryofixation technique for studying cardiac structures.
Main Methods:
- Cardiac phase-targeting rapid cryofixation of Langendorff-perfused rat hearts.
- α-actinin immunohistochemistry to visualize sarcomere length (SL).
- Generation of myocardial heatmaps to visualize SL distributions.
Main Results:
- Sarcomere length (SL) was significantly shorter during systole than diastole.
- Heatmaps revealed spatially inhomogeneous SL distributions within and among myocytes.
- Pharmacological relaxation attenuated SL inhomogeneity; ventricular fibrillation induced inhomogeneous SLs.
Conclusions:
- Cardiac phase-targeting cryofixation effectively visualizes dynamic sarcomere length (SL) changes.
- The study highlights significant spatial inhomogeneity in sarcomere length (SL) during the cardiac cycle.
- This cryofixation strategy offers new insights into spatiotemporal sarcomere structure and cardiac function.
Abstract:
The heart is a functional syncytium consisting of numerous cardiomyocytes that repetitively exhibit coordinated contractions/relaxations. However, the extent to which myocyte sarcomere arrangements in the heart differ across beats is unknown. To examine this, we conducted cardiac phase-targeting rapid cryofixation of Langendorff-perfused rat hearts. We adjusted the timepoint of cryogen exposure to the electrically paced heart and observed phase-dependent differences in the sarcomere length (SL) of subepicardial myocytes by α-actinin immunohistochemistry, namely a significantly shorter SL during systole than during diastole. We detected spatially inhomogeneous SL distributions by generating a heatmap of the myocardium. For peak systole the SL heatmap exhibited nearly uniform SL shortening within and among the individual myocytes with some myocardia exhibiting nonuniform SLs. During diastole, the heart showed predominant SL elongation, which was also accompanied by patchy distributions of locally short-SL regions, reflecting inhomogeneous SLs. This SL inhomogeneity was attenuated by pharmacological relaxation by 2,3-butanedione monoxime. The heatmap of the rapidly-frozen heart during ventricular fibrillation also revealed inhomogeneous SLs within and among individual myocytes. Overall, cardiac phase-targeting cryofixation unveiled in-depth behaviors on SL in the heart. Our cryofixation strategy will open a new horizon to clarify precise spatiotemporal changes in sarcomere structures and understand cardiac functions.
