Single fibers, 25-40 microm wide and 0.5-0.7 mm long, were isolated from the flexor digitorum brevis muscle of the mouse. Force and movement were recorded (21-27 degrees C) from the fiber as a whole and, in certain experiments, from a short marked segment that was held at constant length by feedback control. The maximum tetanic force, 368+/-57 kN/m2 (N = 10), was not significantly different from that recorded in frog muscle fibers at equal temperature. However, the rising phase of the tetanus was considerably slower in the mammalian fibers, 202+/-20 ms (N = 17) being required to reach 90% of maximum tetanic force as compared with 59+/-4 ms (N = 20) in the frog muscle fibers. Similar to the situation in frog muscle fibers, the force-velocity relation exhibited two distinct curvatures located on either side of a breakpoint near 80% of the isometric force. Maximum speed of shortening was 4.0+/-0.3 fiber lengths s(-1) (N = 6). The relationship between tetanic force and sarcomere length was studied between 1.5 and 4.0 microm sarcomere spacings, based on length-clamp recordings that were free of 'tension creep'. There was a flat maximum (plateau) of the length-tension relation between approximately 2.0 and 2.4 microm sarcomere lengths. The descending limb of the length-tension relation (linear regression) intersected the length axis (zero force) at 3.88 microm and reached maximum force at 2.40 microm sarcomere length. The slope of the descending limb is compatible with a thick filament length of 1.63 microm and an average thin filament length of 1.10 microm. These values accord well with recent electron microscope measurements of myofilament length in mammalian muscle.