Stable and continuous supply of essential biomolecules is critical to mimic in vivo microenvironments wherein spontaneous generation of various cell types occurs. Here, we report a new platform that enables highly efficient neuronal cell generation of neural stem cells using single metal-organic framework (MOF) nanoparticle-embedded nanopit arrays (SMENA). By optimizing the physical parameters of homogeneous periodic nanopatterns, each nanopit can confine single nMOFs (UiO-67) that are specifically designed for long-term storage and release of retinoic acid (RA). The SMENA platform successfully inhibited physical interaction with cells, which contributed to remarkable stability of the nMOF (RA⊂UiO-67) structure without inducing nanoparticle-mediated toxicity issues. Owing to the continuous and long-term supply of RA, the neural stem cells showed enhanced mRNA expressions of various neurogenesis-related activities. The developed SMENA platform can be applied to other stem cell sources and differentiation lineages and is therefore useful for various stem cell-based regenerative therapies.