Although a few automated hippocampal subfield segmentation methods have been developed, the effects of amnestic mild cognitive impairment (aMCI) on the hippocampal subfield volumes on magnetic resonance imaging (MRI) are not clear. The aim of this study was to investigate the hippocampal subfield volume changes and their relationships with various neuropsychological tests in aMCI using an automated hippocampal subfield segmentation technique. Forty-five subjects with aMCI and 49 group-matched healthy control subjects underwent 3-tesla MRI scanning, and hippocampal subfield volumes were measured and compared. Additionally, we explored the correlation pattern between hippocampal subfield volumes and the Korean version of the Consortium to Establish a Registry for Alzheimer’s Disease (CERAD-K) neuropsychological test scores in aMCI subjects. Subjects with aMCI exhibited significant hippocampal volume reductions in the presubiculum, subiculum and cornu ammonis 2–3 areas compared with healthy subjects. In addition, we also found significant positive correlations between presubiculum and subicular area volumes and the CERAD-K verbal and visuospatial delayed recall scores in aMCI. This study was the first to explore the relationships between hip-pocampal subfield volumes and various types of cognitive performances in aMCI. These structural changes might be at the core of the underlying neurobiological mechanisms of hippocampal dysfunction in aMCI.