Adult neurogenesis, the differentiation of new neurons from neural stem cells (NSCs), occurs primarily within the subventricular zone (SVZ) and the hippocampal dentate gyrus (DG). Caffeine, the most widely used psychostimulant, is a potent non-selective adenosine receptor antagonist. Though the adenosinergic system has been shown to regulate adult brain plasticity, caffeine impact on postnatal neurogenesis and synaptogenesis remains elusive. Thus, the main goal of this work was to dissect caffeine effects on the differentiation and maturation of postnatal NSCs into neurons.Our results indicate an effect of caffeine in the regulation of neurogenesis in vitro, with caffeine (125 µM) inducing a significant increase in cell proliferation in SVZ-derived neurospheres at DIV1, while in DG-derived neurospheres an increase in the number of mature neurons was observed at DIV7. No significant alterations in cell survival were observed in both niches in response to caffeine exposure. Regarding synaptogenesis, in SVZ-derived new-born neurons, caffeine decreased the number of inhibitory synapses at DIV14. This structural alteration of GABAergic synapses was accompanied by a functional alteration of synapses. Calcium imaging revealed an excitatory action of GABAergic synapses in SVZ-derived new-born neurons, which was significantly diminished by caffeine. In DG-derived neurospheres, caffeine also reduced GABAergic synaptogenesis at DIV7, whilst it increased glutamatergic synaptogenesis at DIV14. Therefore, caffeine has opposite effect on GABAergic and glutamatergic synaptogenesis.In sum, this study sheds light on caffeine influence on postnatal neurogenesis and synaptogenesis, giving novel insights about its effects on brain plasticity.