Human brain organoids offer a unique opportunity to understand developmental milestones, but we still have limited knowledge about the complex molecular mechanisms involved in long-term processes of maturation and maintenance of the tissue. In this study, we profiled brain cortical organoids cultured for periods from 180 to over 1000 days using single-cell RNA-sequencing, electron microscopy and functional assays.Our analysis revealed various neuronal populations with specific subclusters that gradually decreased in complexity over time, while astrocyte populations became more prominent. We hypothesized that spontaneous activity, and the establishment of functional synapses might play a crucial role in maintaining neuronal identity over time. To investigate this, we transferred younger organoids to a culture medium that promotes synaptic maturation, and evaluated morphology and synapse formation using electron microscopy. We found increased synaptic markers and a rescue of specific neuronal populations, such as callosal projection neurons and corticofugal projection neurons. We also performed functional essays to ensure that the organoids were electrically active. Using extracellular single-unit recordings with a multielectrode array (MEA), we detected network bursts and action potentials with features that were changing over the developmental trajectory.In parallel, we explored the potential of cortical progenitors after several months of culturing: we discovered populations of committed cortical progenitors that have unexpected plasticity and can be receptive to environmental triggers.Our findings provide insight into the developmental capabilities of long-term organoid cultures exploring fundamental processes that might contribute to the maintenance of human neuronal identity and progenitor plasticity.