BACKGROUND: Glymphatic functioning is implicated in cognitive and affective functioning. Given that rumination, a major risk factor for major depressive disorder (MDD), is a cognitive process regulating information processing, knowledge of the neurobiological mechanisms underpinning the relationship among glymphatic functioning, rumination, and depression would offer significant insight into the precipitating and maintenance mechanisms of MDD. METHODS: This study recruited 53 MDD patients and 47 matched healthy controls (HCs). Diffusion Tensor Image Analysis along the Perivascular Space (DTI-ALPS) index was computed as a proxy of the glymphatic functioning. Rumination and depressive severity were evaluated using Ruminative Response Scale (RRS) and Hamilton Depression Rating Scale (HAMD), respectively. Static and dynamic functional connectivity (FC/dFC) analyses were performed, and associations with neurotransmitter maps were explored. RESULTS: MDD patients showed reduced glymphatic function compared to HCs, with lower glymphatic function correlating with more severe depression and rumination. Rumination mediated the glymphatic-depression relationship. Furthermore, overlapping static FC involving default mode and subcortical networks linked the glymphatic functioning and rumination. Edge-centric dynamic FC analysis showed reduced State 3 occurrence and heightened rumination, further mediating the glymphatic-rumination relationship in HCs. Both FC biomarkers spatially correlated with various neurotransmitter maps (e.g. dopamine). CONCLUSIONS: Glymphatic dysfunction may exacerbate depression by disrupting brain networks and neurotransmitter balance, trapping individuals in maladaptive rumination. Enhancing glymphatic flow (e.g. via physical exercise) could restore neurobiological health, breaking the maladaptive cycles. This highlights glymphatic functioning as a potential therapeutic target bridging neurobiology, cognition, and depression severity.