Purpose: The purpose of this study is to investigate the relationship between macular and peripapillary retinal nerve fiber layer thickness and amblyopia. Methods: A total of 56 pediatric patients with hyperopic anisometropic amblyopia were evaluated in this study. As the control group, we used the patients’ fellow eyes. A comprehensive eye examination was performed, including best-corrected visual acuity with Snellen charts (converted to logMAR for analysis), slit-lamp examination, fundus examination, cover and, cover-uncover testing, and ocular motility testing. The macular thickness, retinal nerve fiber layer thickness, axial length and optic disc area of both eyes were measured. Results: The mean age was 7.25±1.89 years. For the amblyopic and fellow eyes, the mean best-corrected visual acuity was 0.33± 0.20 logMAR and 0.0± 0.0 logMAR, respectively. Macular center thicknesses were 225,55±18,86 µm and 215,7±12,96 µm in amblyopic and fellow eyes, respectively. Macular 6mm ring thicknesses were 281,32±33,51 µm and 256,07±28,91 µm in amblyopic and fellow eyes, respectively. RNFLT were 108,39±11,59 µm and 104,61±8,43 µm in amblyopic and fellow eyes, respectively There was a statistically significant difference in the macular center thickness, in macular 6-mm ring area thickness and in RNFLT (p<0.05). There was a positive correlation between visual acuity of amblyopic eyes and macular thickness and RNFLT. Conclusions: This study explored objectively measured retinal changes in amblyopia and found a slightly thicker central macular region and 6-mm ring area thicknesses and RNFLT in amblyopic compared with normal eyes.