Pairs of frog semitendinosus muscles were stimulated tetanically for 2 s at 0 degree C and, after 0.75 s of isometric contraction, were released at constant velocity for 0.16 s. The distance shortened was 0.3 micron per sarcomere. The extra heat (shortening heat) associated with the release was determined by comparison with isometric control tetani. The mean sarcomere length at the start of stimulation was varied between 2.25 and 3.75 microns. At the greater initial lengths there was considerable resting tension and heat was absorbed during releases of unstimulated muscles. This thermoelastic effect was also present during stimulation, so shortening heat values were calculated from the difference between the heat production of the stimulated and unstimulated muscle for releases over the same length range and at the same velocity. Laser diffraction was used to measure mean sarcomere length in the part of the muscle from which heat was recorded. At the greater initial lengths these central sarcomeres slowly elongated by a small amount during nominally isometric tetani. When a release was given during a tetanus the central sarcomere shortening was slightly less than that in a release of the unstimulated muscle with the same change of muscle length. In the period of a tetanus after the release had ended the rate of central sarcomere elongation was greater than that in an isometric tetanus. Shortening heat production had a large early component, during and just after the release, then continued to increase slowly up to the end of the tetanus. It is likely that sarcomere length redistribution along the muscle is responsible for the apparent slow phase, but has little effect on the early component. Shortening heat (determined as the early component) decreased linearly with increasing muscle length in the range studied. The intercept on the length axis was 3.78 +/- 0.08 micron per sarcomere (mean +/- S.E. of mean, n = 7, based on sarcomere length before stimulation). The corresponding intercept for the isometric tension before the release was 3.78 +/- 0.03 micron per sarcomere. The isometric heat rate at this time also decreased with increasing muscle length, but at 3.75 microns per sarcomere was still 50.0 +/- 2.0% of its maximum value. The results suggest that shortening heat, like isometric tension, is produced by the interaction of thick and thin filaments in contracting muscle.