Participants with mild cognitive impairment (MCI) have a higher likelihood of developing Alzheimer's disease (AD) compared to those without MCI, and functional magnetic resonance neuroimaging (fMRI) used with MCI participants may prove to be an important tool in identifying early biomarkers for AD. We tested the hypothesis that functional connectivity differences exist between older adults with and without MCI using resting-state fMRI. Data were collected on over 200 participants of the Rush Memory and Aging Project, a community-based, clinical-pathological cohort study of aging. From the cohort, 40 participants were identified as having MCI, and were compared to 40 demographically matched participants without cognitive impairment. MCI participants showed lesser functional connectivity between the posterior cingulate cortex and right and left orbital frontal, right middle frontal, left putamen, right caudate, left superior temporal, and right posterior cingulate regions; and greater connectivity with right inferior frontal, left fusiform, left rectal, and left precentral regions. Furthermore, in an alternate sample of 113, connectivity values in regions of difference correlated with episodic memory and processing speed. Results suggest functional connectivity values in regions of difference are associated with cognitive function and may reflect the presence of AD pathology and increased risk of developing clinical AD.