High-intensity eccentric-concentric contractions of the jaw-closing muscles induce muscle soreness, fatigue and functional impairment of the jaw, resembling the symptoms of myalgia, according to the Diagnostic Criteria for Temporomandibular Disorders (DC/TMD). However, it is claimed that repetition of similar exercises can minimise these detrimental effects. This study aimed to evaluate the response of jaw-closing muscles following two series of intense eccentric-concentric exercises of the masticatory muscles in healthy subjects. Twelve pain-free participants underwent 2 sessions of intense eccentric-concentric jaw exercises, with 1-week interval in between. Each session of jaw exercises comprises 6 sets of 5-minute-long bouts of concentric-eccentric contractions. Self-reported muscle fatigue and pain, maximum mouth opening without pain (MMO), pain pressure thresholds (PPTs) of temporalis and masseter muscles and maximum voluntary bite force (MVBF) were recorded before, immediately after, 24 and 48hours after each bout of exercises. ANOVA for repeated measurements was used to analyse the data. During session 2, muscle pain and fatigue were statistically significantly decreased (P<.05) as compared to session 1. Furthermore, statistically significant increases of MVBF (P<.005), MMO (P<.005) and PPTs (P<.005) were found at session 2 as compared to session 1. Within the limitations of the study, is can be concluded that the repetition of eccentric-concentric jaw-closing exercises results in signs of muscle training. Future studies can elucidate whether this motor training might be useful for the treatment of myalgia.