Healing process following controlled lesion of the round window membrane in guinea pigs, as a model of round window membrane rupture in human, was studied by light microscopy, transmission and scanning electron microscopy. The middle ear of the guinea pigs was aseptically opened by a post-auricular incision to expose the round window membrane without touching the auditory ossicles and tympanic membrane . Then, a slit perforation, 0.5-0.8mm in length, was made with a micro knife in the same direction as those of the membrane fibers. The following healing process could be divided into three stages based on complete closing times of the layers consisting the membrane. The first stage was about a week, after the lesion and until the complete closing of the middle layer. In this stage, the opening of the lesion persisted and around this opening moderate hypertrophy of the outer and middle layers, 0.2mm in width and showing a snake head-like profile in section, was observed. The inner layer partially peeled off from the middle layer. The second stage continued about one or two weeks until the complete closing of the outer layer. In this stage, the healing portion presented a shallow and elongated crater-like appearance because of outer layer cells growing on the newly extended middle layer and leakage of the endolymph was still noted. The last stage was from about three weeks to three months after the operation, when the lesion completely healed. The crater-like swelling of membrane was still observed in the early time of this stage, but it gradually disappeared. Recovery of the inner layer was the latest and the old inner layer floating in the perilymph was progressively replaced by a newly formed network beneath it. The endolymph leakage continued even after the middle layer had closed but ceased when the outer layer closed. These finidings suggest that complete closure of both of the middle and outer layers is required to cease the endolymph leakage.