The ovary of the mare has a unique structure which differs totally from that of other mammals. However, because of its relatively large size, conventional histological techniques were unsuitable for the observation of the internal structure of the whole ovary. Three-dimensional internal structure microscopy (3D-ISM) consists of a cryotome-CCD camera-laser disc recorder-PC-based control system coupled with a graphic workstation. The internal structure of the ovary is observed by processing over more than 1,000 stored images of serially sliced surfaces of each frozen equine ovary. The 3D reconstruction was done using the full-coloured, volume-rendering method. The relationship between the localization of medulla, cortex and ovulation fossa was clarified. The ovulation fossa is localized in the centre of the ovary and is surrounded by a broad ovarian cortex. A trace of ovulation was observed only at the ovulation fossa. Medulla are localized in narrow peripheral areas. The phenomenon of the competition to occupy the cortical area ahead of the ovulation fossa by developing secondary follicles was visualized. Spatial localization of various sized follicles was identified from 3D-reconstructed images. In this study, it has been clarified that application or this novel computerized technique can clarify the anatomical arrangements of the equine ovary and the complex mechanism of equine follicular development.