In the ciliated protozoan, Blepharisma japonicum, the pink-colored pigment (blepharismin) contained in the pigment granules is believed to be the photoreceptor pigment responsible for the step-up photophobic response. When the cells partially bleached by extrusion of the pigment granules caused by cold shock were subsequently cultured under illuminated conditions, the pigment-less granules regenerated and the cells were further bleached (pigment content below 0.5%). The photosensitivity of such colorless cells disappeared completely. In contrast, the blepharismin pigment regenerated gradually when such colorless cells were transferred to darkness. The photosensitivity of the cells also recovered with regeneration of the pigment. We found that blepharismin pigment was not photobleached in the absence of O2. The step-up photophobic response was also completely repressed in the absence of O2. These results strongly confirm that blepharismin is a photoreceptor pigment mediating photobehavior of Blepharisma and that O2 is required for the early step in the phototransduction of the light-excited pigment.