To determine the optimal rhythm of distraction and level of osteotomy for maximal muscle preservation during limb lengthening, histologic and ultrastructural analysis of the peroneus tertius muscle was performed in 46 goats. Thirty-nine animals with 20% or 30% tibial lengthening at the rate of 0.75 mm per day at each osteotomy level were divided in two groups. The first group included 15 animals undergoing tibial middiaphyseal lengthening with automated distractor producing one (n = 5), four (n = 5), or 720 (n = 5) increments per day. The second group included 24 animals in which lengthening was performed at the proximal (n = 4), middle (n = 6), distal (n = 6), or double (n = 8) level of the tibia with distraction rhythm of three times per day. The remaining seven animals served as controls. Histomorphologic features of the lengthened muscles were characterized by a polymorphous appearance with foci of necrosis, atrophy, and sclerosis scattered among apparently normal muscle fibers. The intensity and dispersion of degenerative changes were in direct proportion to the amount of lengthening but in reverse proportion to the frequency of distraction. Tibial lengthening at the diaphysis caused intense fibroblast proliferation and remarkable muscle sclerosis, whereas metaphyseal distraction was associated with a predominance of muscle tissue regeneration over atrophy and sclerosis. Bifocal lengthening and a more fractionated rhythm of distraction were associated with enhanced preservation of existing muscle fibers and greater regenerative activity of the muscle.