The precise binding of the antibody to the Z-line was confirmed with the fluorescently labeled secondary antibody localized at regular intervals between adjacent A-bands in myofibrils (Fig. within the myofibril, and the sarcomere yielding occurred in clusters composed of several adjacent sarcomeres. The labeling of Z-line with anti–actinin antibody significantly suppressed the clustered sarcomere yielding. These results strongly suggest that the contractile system of muscle possesses the mechanism of structure-based inter-sarcomere coordination. Keywords: myofibril, sarcomere yielding, Z-line, partial activation The periodic architecture of striated muscle relies on the series connection of the basic contractile units, called sarcomeres, which are connected through the Z-line to form a myofibril. Each sarcomere is composed of a bipolar array of myofilaments, i.e., the thick (myosin) and thin (actin) filaments. The cyclic interaction of myosin with actin coupled to ATP hydrolysis produces force and motion along the long axis of the myofibril (1, 2). Such characteristics of striated muscle imply that individual sarcomeres are structurally and mechanically interconnected and interact with each other. Tension and length responses of sarcomeres to the external perturbations have revealed various mechanochemical properties of striated muscle, particularly those relating to the mechanism of force generation (3C6) MK-4305 (Suvorexant) and to MK-4305 (Suvorexant) the attachment/detachment kinetics of cross-bridges (7C9). On the other hand, MK-4305 (Suvorexant) the interaction between sarcomeres remains unclear, primarily because the widely used technique for measuring sarcomere response is laser light diffraction, where the individual sarcomere dynamics are obscured by the ensemble averaging of thousands of sarcomeres connected in parallel and in series in muscle fibers. Recent developments in microscopic analysis using skeletal and cardiac myofibrils revealed various dynamic properties of individual sarcomeres (or even half-sarcomeres) upon force generation and relaxation of striated muscle (10C14). One of the particular characteristics found by this approach is the spontaneous oscillation of sarcomeres, which is observed at partial activating conditions. The myofilament-generated oscillation was first reported by Fabiato and Fabiato in skinned cardiac cells at fixed concentrations of free Ca2+, suggesting the existence of Ca2+-independent regulatory mechanism in sarcomeres (15). One decade later, we found that the steady periodic oscillation, named SPOC (SPontaneous Oscillatory Contraction), can be well reproduced at partial activation by adding exogenous ADP and inorganic phosphate (Pi) to the relaxing solution (in the presence of ATP and the absence of Ca2+) (16), allowing the quantitative analysis of sarcomere behavior. During SPOC, each sarcomere repeats slow-shortening and rapid-lengthening phases with a period of a few seconds, lasting from minutes to hours. In addition, the lengthening phase propagates to adjacent sarcomeres along the long axis of myofibrils (SPOC wave) at a faster rate than the diffusion of chemical components. These observations strongly suggest that FLB7527 the cooperative behavior of sarcomeres is not attributable to artificial non-uniform activation, but is an intrinsic property of striated muscle. Moreover, such dynamic properties of sarcomeres show a possible link to physiological functions. For example, the period of SPOC strongly correlates with that of heart beat in various animal species (17), and the relaxation of single sarcomeres propagates along the myofibril after the rapid removal of Ca2+ (12). These reports imply that the cooperative behavior of individual sarcomeres, which is concealed by the ensemble averaging, plays a significant role for the efficient work and motion in muscle. Hence, the analysis at the single sarcomere level must be indispensable for fully understanding the regulatory mechanism that striated muscle possesses. In this study, we investigated the length-response of individual sarcomeres to externally applied load, using mechanical manipulation of skeletal myofibrils with microneedles. The characteristic aspects of this study are: 1) The behavior of individual sarcomeres was observed under a phase-contrast microscope simultaneously with measuring tension response (18). 2) The myofibrils were activated, in almost all cases, by adding MgADP to the relaxing conditions (19), which results in the regulation of sarcomeric activity by MK-4305 (Suvorexant) strong-binding (or ADP-bound) cross-bridges independently of Ca2+. 3) The mechanical.