Vitamin D deficiency (VDD) is a worldwide problem, especially among the elderly and epidemiological studies have found a link between VDD, cognitive decline and depression. However, it is unclear whether VDD directly drives the development of depression and cognitive decline or whether it is secondary to co-occurring pathology.In this study, we sought to determine whether long-term VDD causes memory impairments and depressive-like behavior alone or in combination with chronic stress and to elucidate co-occurring dysfunctions in neuronal and glial plasticity in the dentate gyrus (DG) of the hippocampus.4-month-old C57/Bl6J mice were assigned to a vitamin D-free diet for 6 months to induce long-term VDD, followed by 8 weeks of unpredictable chronic mild stress (UCMS).UCMS induced depressive-like behavior, memory impairments and alterations in grooming behavior. VDD induced impairments in episodic-like and contextual fear memory and reduced grooming behavior, but not depressive-like behavior. Interestingly, VDD did not pose as an additional risk factor for cognitive dysfunctions and depressive-like behavior in combination with UCMS. In fact, in VDD mice UCMS improved self-care behavior. In the hippocampus, VDD and UCMS independently reduced total cell number and neurogenesis in the DG. VDD increased apoptosis and induced glial reactivity in the DG, but these effects were not amplified by UCMS. Our findings indicate while long-term VDD in adulthood caused cell loss and altered neuronal and glial plasticity in the DG, UCMS did not exacerbate these effects and could contribute to mechanisms regulating altered stress response.