Hyaluronic acid (HA) is a component of the extracellular matrix (ECM) in most vertebrate tissues and is thought to play a significant role during development, wound healing, and regeneration. In vitro studies have shown that HA enhances muscle progenitor cell recruitment and inhibits premature myotube fusion, implicating a role for this glycosaminoglycan in functional repair. However, the spatiotemporal distribution of HA during muscle growth and repair was unknown. We hypothesized that inducing hypertrophy via synergist ablation would increase the expression of HA and the HA synthases (HAS1-HAS3). We found that HA and HAS1-HAS3 were significantly upregulated within the plantaris muscle in response to Achilles tenectomy. HA concentration significantly increased 2.8-fold after 2 days but decreased towards levels comparable to age-matched controls by 14 days. Using immunohistochemistry, we found the colocalization of HAS1-HAS3 with macrophages, blood vessel epithelia, and fibroblasts varied in response to time and/or tenectomy. At the level of gene expression, only HAS1 and HAS2 significantly increased with respect to both time and tenectomy. The profiles of additional genes that influence ECM composition during muscle repair, tenascin-C, type I collagen, the HA-degrading hyaluronidases (Hyal) and matrix metalloproteinases (MMP) were also investigated. Hyal1 and Hyal2 were highly expressed in skeletal muscle but did not change after tenectomy; however, indicators of hypertrophy, MMP-2 and MMP-14, were significantly upregulated from 2 to 14 days. These results indicate that HA levels dynamically change in response to a hypertrophic stimulus and various cells may participate in this mechanism of skeletal muscle adaptation.