To investigate whether melatonin protects neurons from apoptosis, we used amyloid beta-peptide 25–35 (Aβ25–35) to induce apoptosis in cultured hippocampal neurons, and monitored the apoptotic activity of the neurons with or without melatonin treatment. Present study shows that melatonin at concentrations of 1 × 10−6 and 1 × 10−5 mol/L prevents neuronal morphological changes induced by apoptosis, as characterized by the shrunken and rounded morphology caused by condensed chromatin and fragmented DNA. Melatonin further exhibited the ability to inhibit apoptotic internucleosomal DNA degradation. Moreover, flow cytometric analysis of cell cycle demonstrated that melatonin abolished the Aβ25–35-induced apoptotic peak. Our results suggest that melatonin may play an important role to protect neurons from Aβ25–35-induced apoptosis.