A game of load and calcium determines muscle relaxation

Muscle contraction is like rowing a boat, where myosin motors (oars) pull on the thin actin filaments (the water) to generate movement. This is a process triggered when calcium levels rise and cause changes in the thin filament, allowing myosin motors to connect . The myosin motors on the thick filament also switch from "off" (inactive) to "on" (active) conformation, so they can pull on actin and create force and shortening. The thick filament can sense how much force is needed and adjust how many motors to turn on.

On the other hand, muscle relaxation has not been as widely studied as muscle activation. This is a critical component to muscle mechanics, especially in fast processes like eye movement. The ability from muscles to quickly activate and relax is impaired in diseases such as Parkinson’s and dystonia or in old age, so a better understanding of this process is needed.

“Just lowering calcium doesn’t rapidly relax the muscles, so we decided to focus on other factors like load, which could play a role in how muscles switch off”, explains Cameron Hill, researcher at King’s College London and corresponding author of the publication.

With this aim, they came to the ESRF’s ID02 beamline to use time-resolved small-angle X-ray diffraction to determine how muscle thin and thick filaments switch off when calcium or muscle load decreases. “At ID02 we can use a series of different camera lengths to visualize what happens at the sarcomere level and within it. The unique ability of ID02 is that we can characterise this at 31 metres and at 3 metres from the same sample, which is currently impossible to do elsewhere”, explains Hill.

Thick and thin filament activation states and myosin motor conformations with respect to force (T/T0). Load is removed by rapid shortening of the muscle at high [Ca2+]i in the grey-shaded region, resulting in partial recovery of the OFF states of the thick and thin filaments (green box). Load remains high whilst calcium is rapidly removed after the final electrical stimulation at 120ms in the blue-shaded region where attached myosin motors keep both the thick and thin filaments ON (blue box).

The results showed that that reducing load from maximal when the calcium is high  switches off both the thick and thin filaments. When load is high but calcium decreases after activation, the thick filaments initially remain fully on, although the number of myosin motors bound to actin decreases and the force per attached motor increases. That initial slow phase of relaxation finishes abruptly when the sarcomeres lose tension. This triggers a redistribution of sarcomere lengths, which leads to the rapid completion of mechanical relaxation.

“It is the first time structural changes in both the thick and thin filaments have been characterized during relaxation of a mammalian muscle with this temporal and spatial resolution”, says Hill. Whilst this research is fundamental, the new findings can contribute to can contribute to the development of new treatments for diseases that affect muscle function’.

Reference:

Hill, C. PNAS, March 12, 2025. 122 (11) e2416324122

https://doi.org/10.1073/pnas.2416324122

Text by Montserrat Capellas Espuny

Top image: This 2D X-ray diffraction pattern is collected at 3.2m. The beam is much smaller than the muscle (beam = ~140um horizontally, ~40um vertically; mouse EDL = ~3m horizontally, 12mm vertically), so it is visualising the myosin-containing thick filament and actin-containing thin filament structures within the sarcomeres. Because the thick and thin filaments have repeating helical structures within them, the collected diffraction pattern give rise to these strongly diffracting reflections that look like yellow-white strips along the meridian, and off-meridional “layer lines” which look like horizontal strips of intensity that radiate outwards from the meridian. The bright spots along the equator come from the hexagonal lattice arrangement of the thick and thin filaments.