Interaction between titin and thin filaments in intact cardiac muscle.

A 'freeze break' technique and immunoelectron microscopy were used to study the elastic properties of cardiac titin filaments. Small bundles consisting of a few fibres from freshly prepared dog papillary muscle were quickly frozen and broken under liquid nitrogen to fracture sarcomeres in planes perpendicular to the filament axes. Breaks occurred at each of several regions along the sarcomeres. The still-frozen specimens were thawed during fixation to allow elastic filaments to retract. The broken muscle segments were then treated with monoclonal titin antibody 9D10 which labelled a unique epitope in the I-band. In sarcomeres broken at the A-I junction, the titin filaments reacted toward the Z-line, independently of the thin filaments. The retracted epitopes did not reach the Z-line; retraction stopped at the N1-line level. In sarcomeres broken near the Z-line, the titin filaments retracted in the opposite direction, to the tip of the thick filaments. When the break occurred in the A-band, by contrast, the titin-epitope position was unaffected. On the basis of these results, and despite the reported interaction of titin and actin in vitro, it appears that cardiac titin molecules form elastic filaments that are functionally independent of the thin filaments. Near the Z-line, however, the titin filaments seem to associate firmly with the thin filaments.