NONUNIFORM ELASTICITY OF TITIN IN CARDIAC MYOCYTES - A STUDY USING IMMUNOELECTRON MICROSCOPY AND CELLULAR MECHANICS

Titin (also known as connectin) is a muscle-specific giant protein fou nd inside the sarcomere, spanning from the Z-line to the M-line. The I -band segment of titin is considered to function as a molecular spring that develops tension when sarcomeres are stretched (passive tension) . Recent studies on skeletal muscle indicate that it is not the entire I-band segment of titin that behaves as a spring; some sections are i nelastic and do not take part in the development of passive tension, T o better understand the mechanism of passive tension development in th e heart, where passive tension plays an essential role in the pumping function, we investigated titin's elastic segment in cardiac myocytes using structural and mechanical techniques, Single cardiac myocytes we re stretched by various amounts and then immunolabeled and processed f or electron microscopy in the stretched state. Monoclonal antibodies t hat recognize different titin epitopes were used, and the locations of the titin epitopes in the sarcomere were studied as a function of sar comere length, We found that only a small region of the I-band segment of titin is elastic; its contour length is estimated at similar to 75 nm, which is only similar to 40% of the total I-band segment of titin , Passive tension measurements indicated that the fundamental determin ant of how much passive tension the heart develops is the strain of ti tin's elastic segment. Furthermore, we found evidence that in sarcomer es that are slack (length, similar to 1.85 mu m) the elastic titin seg ment is highly folded on top of itself, Based on the data, we propose a two-stage mechanism of passive tension development in the heart, in which, between sarcomere lengths of similar to 1.85 mu m and similar t o 2.0 mu m, titin's elastic segment straightens and, at lengths longer than similar to 2.0 mu m, the molecular domains that make up titin's elastic segment unravel, Sarcomere shortening to lengths below slack ( similar to 1.85 mu m) also results in straightening of the elastic tit in segment, giving rise to a force that opposes shortening and that te nds to bring sarcomeres back to their slack length.