Oscillatory work and the step that generates force in single myofibrils from rabbit psoas

The elementary molecular step that generates force by cross-bridges (CBs) in active muscles has been under intense investigation in the field of muscle biophysics. It is known that an increase in the phosphate (P_i) concentration diminishes isometric force in active fibers, indicating a tight coupling between the force generation step and the P_i release step. The question asked here is whether the force generation occurs before P_i release or after release. We investigated the effect of P_i on oscillatory work production in single myofibrils and found that P_i-attached state(s) to CBs is essential for its production. Oscillatory work is the mechanism that allows an insect to fly by beating its wings, and it also has been observed in skeletal and cardiac muscle fibers, implying that it is an essential feature of all striated muscle types. With our studies, oscillatory work disappears in the absence of P_i in experiments using myofibrils. This suggests that force is generated during a transition between steps of oscillatory work production, and that the states involved in force production must have P_i attached. With sinusoidal analysis, we obtained the kinetic constants around the P_i release steps, established a CB scheme, and evaluated force generated (and supported) by each CB state. Our results demonstrate that force is generated before P_i is released, and the same force is maintained after P_i is released. Stretch activation and/or delayed tension can also be explained with this CB scheme and forms the basis of force generation and oscillatory work production.