Cardiomyocyte sarcomere length variability: Membrane fluorescence versus second harmonic generation myosin imaging

Area of Science:

• Cardiovascular Biology
• Cellular Biophysics
• Microscopy Techniques

Background:

• Sarcomere length (SL) critically influences coordinated myocyte contraction.
• Accurate assessment of individual SL properties is vital for understanding cardiac function.
• Current optical methods like confocal fluorescence microscopy visualize Z-disks but not central sarcomeric structures.

Purpose of the Study:

• To compare averaged SL and its variability in isolated rat cardiomyocytes using ANEPPS fluorescence and SHG microscopy.
• To determine the optimal imaging technique for precise SL variability evaluation.

Main Methods:

• Isolated relaxed rat cardiomyocytes were imaged using both ANEPPS fluorescence and SHG microscopy.
• Sarcomere length and its variability were quantified using established metrics.
• Comparison of SL measurements obtained from both imaging modalities.

Main Results:

• Both ANEPPS and SHG microscopy yielded similar average (median) SL values.
• Sarcomere length variability was found to be two times smaller when measured by SHG microscopy compared to ANEPPS.
• SHG microscopy visualizes A-band structures directly, avoiding interference seen with Z-disk-focused methods.

Conclusions:

• Optical methods with comparable spatial resolution provide valid average SL estimations.
• SHG microscopy is proposed as the gold standard for evaluating SL variability due to its direct visualization of A-bands.
• SHG microscopy offers a prospective tool for mapping sarcomere variability in vitro and in vivo, aiding in understanding its role in myocardial function.