Background: Hemodialysis (HD) leads to cognitive impairment; however, the pathophysiology of maintenance HD remains unclear. This study aimed to investigate the longitudinal alterations in gray matter volume (GMV) and cerebral blood flow (CBF) in patients on follow-up HD compared with baseline HD, examine the alterations in functional connectivity (FC) by defining co-changed brain regions as seed points, and investigate the correlation between the co-changed brain regions and neuropsychological test scores. Methods: Twenty-seven patients with HD and 30 healthy controls were enrolled in this study. All participants underwent high-resolution T1-weighted imaging, arterial spin labeling, and functional MR imaging to measure GMV, CBF, and FC. The patients on HD were assessed at baseline and 3 years subsequently. Results: The right and left medial superior frontal gyrus (SFGmed.L) exhibited significantly lower GMV and CBF in patients on follow-up HD compared with patients on baseline HD and lower FC between the SFGmed.L and left middle temporal gyrus (MTG.L). Decreased FC between the SFGmed.L and MTG.L was positively correlated with neuropsychological test scores in the follow-up HD group. Conclusions: Reduced GMV and CBF may result in decreased FC between the SFGmed.L and MTG.L, which may be associated with cognitive impairment in patients on maintenance HD. Our findings provide unique insights into the pathological mechanisms of patients on maintenance HD with cognitive impairment.