PURPOSE. Environmental light plays an important role in the process of emmetropization. This study investigated how the retina integrates wavelength and temporal signals to regulate eye emmetropization. METHODS. Guinea pigs (n ¼ 220) were randomly divided into 11 groups (n ¼ 20/group) that received different environmental lighting (12:12 light cycle) for 8 weeks: white, green, or blue light at steady, 0.5 or 20 Hz. White-steady group was repeated for each wavelength. Refraction, axial length, and corneal curvature were measured using streak retinoscopy, Ascan ultrasonography, and keratometry, respectively, every 2 weeks. RESULTS. (1) In white light, the white-0.5 Hz group was more myopic than the white-steady group or the white-20 Hz group (both P < 0.0001), with a longer axial length (both P < 0.0001). White-20 Hz did not significantly differ from white-steady. (2) At low temporal frequencies (0 and 0.5 Hz), green-steady (P ¼ 0.0008) and green-0.5 Hz (P < 0.0001), were more myopic than the white-steady group, with longer axial lengths (both P < 0.0001). No significant difference was found between green-0.5 Hz and green-steady. Blue-steady and blue-0.5 Hz were more hyperopic than white-steady (both P < 0.0001), with shorter axial lengths (both P < 0.0001). Blue-0.5 Hz showed no significant difference from blue-steady. (3) At high temporal frequencies (20 Hz), green-20 Hz, was more hyperopic than green-steady or green-0.5 Hz (both P < 0.0001) and had a shorter axial length (both P < 0.0001). Green-20 Hz showed a 1.10 D hyperopic shift compared to green-steady. Blue-20 Hz was less hyperopic than blue-steady (P < 0.0001) or blue-0.5 Hz (P ¼ 0.0012), with a longer axial length (both P < 0.0001). Blue-20 Hz showed a 1.18 D myopic shift compared to blue-steady. CONCLUSIONS. Eyes use both wavelength and temporal frequency of light to regulate emmetropization. Their interactions provide different cues to control eye growth. At low temporal frequencies, the eye can use wavelength defocus to guide eye growth. This signal is weakened at high temporal frequencies.