Neural stem cells (NSCs) are found in discrete regions of the mammalian brain. During adulthood, NSCs can be a source of new neurons and oligodendrocytes in neurological disorders. However, these newborn cells are not sufficient to overcome the neurological deficits involved by neural loss. Therefore, the identification of novel mechanisms responsible for modulating NSC fate represent a major key issue for future brain repair strategies. Several studies suggest that mitochondria have an important role in regulating NSC differentiation and lineage determination. However, the molecular mechanisms involved in this regulation remain unknown. Hence, our work aims to dissect how mitochondria biogenesis and dynamics can modulate the NSC differentiation into neurons or oligodendrocytes. For this, NSCs were obtained by isolating subventricular zone (SVZ) and dentate gyrus (DG) cells from P1-3 mouse models. Seeding density, culture conditions and number of passages were determined. Moreover, the multipotency of SVZ/DG-derived NSPCs, obtained from different passages, was also accessed. Additionally, expression of proteins involved in mitochondrial biogenesis and fusion/fission appear altered during NSPC differentiation, while mitochondrial network revealed different morphologies in cells from different lineages. The results obtained will provide novel findings concerning the role of mitochondrial dynamics in NSC fate.