Adult hippocampal neurogenesis is considered as the progression from neural stem cell to mature cells in subventricular (SVZ) and subgranular zone (SGZ) of dentate gyrus (DG) of the hippocampus. Neurogenesis is essential for antidepressants efficacy and, amongst many factors, it is influenced by the fluctuations in estradiol levels across the estrous cycle, pregnancy and aging. In this study SGZ cell proliferation, survival, differentiation and Tail Suspension Test (TST), as a depressive-like behavioural paradigm, have been evaluated in naïve female as well as in ovariectomized (OVX) mice. Both groups were treated with chronic (21-days) melatonin, already known in improving a depression-like state by stimulating neurogenesis and synaptic plasticity. To correlate neuronal development with neuronal plasticity, dendritic spine density has been analyzed. The number of bromodeoxyuridine (BrdU) and doublecortin (DCX) immunoreactive cells of OVX mice was unchanged compared to that one of naïve, while melatonin administration increased BrdU+ and DCX+ cells in both groups. As concerns the TST, estrogen deprivation did not affect the immobility time in naive and OVX mice, contrarily to melatonin. Moreover, in OVX mice, depletion of estrogens impairs the spine maturation in granule cells of DG, while melatonin increases this number. Our data demonstrate that SGZ proliferation is not dependent on endogenous levels of estradiol and show that melatonin stimulates cell formation in the hippocampus in an estrogen-independent manner. Melatonin may be thus useful in improving brain functions acting on birth, survival and differentiation of new neurons in the hippocampus, stimulating maturation of spines and exerting an antidepressant-like action under estrogen-deprived conditions.