This study aimed to examine peripheral fatigue and the resultant damage to the masseter muscle due to prolonged low-frequency stimulation. Thirty male rats were divided into S1, S2, S4, Dantr and Sham groups. The left masseters were used as experimental muscles. A pair of stimulation electrodes was placed on the left masseter. A stimulating session included rectangular electric pulses of 18 Hz (5 mA, ∼18 V, 0.7 ms) for 2 h with a 3 min rest period between sessions. One session was given to the S1 group, two sessions to the S2 group and four sessions to the S4 group. Four sessions were given to the Dantr group with administration of dantrolene to determine any artifacts of the electrical current. No electric stimulation was given to both side masseters in the Sham group or to the control (right) masseters in the other groups. In each session, jaw-closing force increased to a peak within 1 min and attenuated to the steady force. The peak force decreased as the session advanced in each group. Both side masseters were dissected after the stimulations and examined histologically. The experimental masseter was significantly heavier than that of the controls in the S1, S2 and S4 groups, and the muscle fibres showed irregularity of size and shape with enlargement of interstitial space and infiltration of mononuclear cells into the fibres. However, no such histological change was observed in the Dantr and Sham groups. It was confirmed that fatigue and damage to muscle fibres could be induced in masticatory muscles by prolonged low-frequency stimulation. © 2005 Elsevier Ltd. All rights reserved.