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Cell Biology JoVE (Open Access) Citable · DOI

In Vitro Assessment of Cardiac Function Using Skinned Cardiomyocytes

DOI: 10.3791/60427-v
What you'll learn
  • ✓ Extract and permeabilize skinned cardiomyocytes from cardiac tissue biopsies
  • ✓ Measure active, passive, and calcium-sensitive contractile force in isolated myocytes
  • ✓ Assess myofilament function using mechanical and biochemical modulation techniques
Protocol

This protocol aims to describe step-by-step the technique of extraction and assessment of cardiac function using skinned cardiomyocytes. This methodology allows measurement and acutemodulation of myofilament function using small frozen biopsies that can be collected from different cardiac locations, from mice to men.

Difficulty
advanced
Total time
~4–6 hours per tissue sample (extraction, permeabilization, selection, and force measurements)
Model organism
Mouse; human cardiac tissue biopsies
Biosafety
BSL-1

Steps

1
Introduce skinned cardiomyocyte methodology and rationale

Overview of the skinned cardiomyocyte technique for direct measurement of myofilament contractile properties and acute modulation using small frozen biopsies from various cardiac regions.

▶ 00:04
2
Extract and permeabilize cardiac myocyte fibers

Dissociate frozen cardiac tissue biopsies and chemically permeabilize cell membranes to allow direct access to contractile apparatus while preserving myofilament structure.

▶ 01:00
3
Select and mechanically attach skinned myocytes

Identify suitable individual myocytes, attach them to force transducers via micropipette gluing, and position for mechanical testing.

▶ 02:46
4
Measure active, passive, and calcium sensitivity forces

Perform isometric force measurements under varying calcium concentrations and passive stretch conditions to assess myofilament contractile function and calcium responsiveness.

▶ 04:03
5
Analyze representative skinned cardiomyocyte data

Present and interpret typical force–calcium relationships and contractile parameters from assessed myocytes.

▶ 05:58
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