Mechanical measurements using permeabilized myocardial strips
Frozen tissue sections were thawed in ice-cold skinning solution and
dissected into thin strips (~180 µm in diameter and 700
µm long). These were skinned overnight at 4°C, transferred to storage
solution, and stored at –20°C for 0-5 days. On the day of experiments,
strips were mounted between a motor (P841.40, Physik Instrumente,
Auburn, MA) and a strain gauge (AE801, Kronex, Walnut Creek, CA),
lowered into a 30 µL droplet of relaxing solution (with or without
mavacamten) and stretched to a sarcomere length of 1.9 or 2.3 µm as
measured by digital Fourier transform analysis (IonOptix Corp, Milton,
MA). Solutions were maintained at physiological temperature (37°C)
throughout each experiment, using 2-3 strips at each condition for each
heart.
Strips were activated in solutions (with or without mavacamten) at pCa
values ranging from 8.0 to 4.5 to measure the steady-state, isometric
force-pCa relationship. Force values were normalized to the
cross-sectional area of each preparation and reported as stress values
with units of kN m‑2. Stress-pCa curves from each
strip were fit to a 4-parameter Hill equation using MATAB (version
9.0.4, Mathworks, Natick, MA):