Elastic Properties of Connecting Filaments Along the Sarcomere

Abstract

The elasticity of the connecting filament—the filament that anchors the thick filament to the Z-line—has been investigated using rigor release, freeze-break and immunolabelling techniques. When relaxed insect flight muscle was stretched and then allowed to go into rigor, then released, the recoil forces of the connecting filaments caused sarcomeres to shorten. Thin filaments, prevented from sliding by rigor links, were found crumpled against the Z-line. Thus, rigor release experiments demonstrate the spring-like nature of the connecting filaments in insect flight muscle.

In vertebrate skeletal muscle, however, the same protocol did not result in sarcomere shortening. Absence of shortening was due to either smaller stiffness of connecting filaments and/or higher stiffness of the thin filaments relative to insect flight muscle. The spring-like nature of the connecting filament was confirmed with the freeze break technique. When the frozen sarcomeres were broken along the A-I junction, the broken connecting filaments retracted to the Ni-line level, independently of the thin filaments, demonstrating the basic elastic nature of these filaments.

To study the elastic properties of the connecting filaments along the sarcomere, the muscle was labelled with monoclonal antibodies against a titin epitope near the Ni-line, and another very near the A-I junction in the I-band. Before labelling, fibers were pre-stretched to varying extents. Based on filament retraction and epitope translation with stretch, we could conclude: (1) the A-band domain of the connecting filament is ordinarily bound to the thick filaments; (2) at higher degrees of stretch, connecting filaments become free of the thick filaments, and the freed segments are intrinsically elastic; (3) between the A-I junction and the N₁-line, connecting filaments behave independently of thin filaments; between N₁- and Z-lines, however, they are firmly associated with the thin filaments.