Background: Optic nerve diffusion imaging is a useful investigational tool of optic nerve microstructure, but is limited by eye-movement-induced optic nerve movement and artifacts from surrounding cerebrospinal fluid, fat, bone and air. Attempts at improving patient cooperation, thus voluntarily limiting eye movement during a standard diffusion imagingsequence, are usually futile. The aim of this study was to establish the impact of optic nerve movement on clinical diffusion parameters of the optic nerve. Method: Twenty-nine healthy volunteers with intact vision and intact conjugate gaze were recruited and subjected to magnetic resonance diffusion-weighted imaging (DWI) and diffusion-tensor imaging (DTI) of the optic nerves. Twenty right eyes had nerve tracking done using single-shot echo-planar imaging at 20 time points over 3 minutes. Optic nerve movement measurements were correlated with diffusion parameters of apparent diffusion coefficient (ADC), mean diffusivity (MD), fractional anisotropy (FA) and anisotropic index(AI) using Spearman’s rank correlation. Results: No significant correlations were noted between optic nerve movement parameters and ADC in the axial plane and MD of the optic nerve. Low to moderate negative correlations were noted between optic nerve movement parameters and AI and FA and positive correlation with ADC in the radial plane. Conclusion: Optic nerve movement documented during the timespan of standard diffusion sequences (DWI and DTI) has a negative effect on the anisotropic diffusion parameters of the optic nerve. With greater eye movement, optic nerve diffusion appears less anisotropic owing to greater radial diffusion.