Faravelli, I., Antón-Bolaños, N. et al. (2025).
bioRxiv (Cold Spring Harbor Laboratory).
Abstract
The human brain develops and matures over an exceptionally prolonged period of time that spans nearly two decades of life. Processes that govern species-specific aspects of human postnatal brain development are difficult to study in animal models. While human brain organoids offer a promising in vitro model, they have thus far been shown to largely mimic early stages of brain development. Here, we developed human brain organoids for an unprecedented 5 years in culture, optimizing growth conditions able to extend excitatory neuron viability beyond previously-known limits. Using module scores of maturation-associated genes derived from a time course of endogenous human brain maturation, we show that brain organoids transcriptionally age with cell type-specificity through these many years in culture. Whole-genome methylation profiling reveals that the predicted epigenomic age of organoids sampled between 3 months and 5 years correlates precisely with time spent in vitro, and parallels epigenomic aging in vivo. Notably, we show that in chimeric organoids generated by mixing neural progenitors derived from “old” organoids with progenitors from “young” organoids, old progenitors rapidly produce late neuronal fates, skipping the production of earlier neuronal progeny that are instead produced by their young counterparts in the same co-cultures. The data indicate that human brain organoids can mature and record the passage of time over many years in culture. Progenitors that age in organoids retain a memory of the time spent in culture reflected in their ability to execute age-appropriate, late developmental programs.
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This is pretty wild. This study demonstrates that human brain organoids can be cultured for an unprecedented five years, during which they don't just survive but actively mature, recording the passage of time through coordinated transcriptional and epigenetic programs that parallel the slow development of the endogenous human brain. The researchers developed an "Activity-Permissive Medium" (APM) that significantly enhanced neuronal survival, synaptic activity, and structural complexity over long periods. Crucially, they showed that neural progenitor cells within these aged organoids retain a "memory" of their developmental time. When old progenitors were mixed with young ones in chimeric organoids, the old cells skipped early developmental steps and rapidly generated late-born neuronal types (like callosal projection neurons), indicating they have an internal clock that dictates their fate potential based on their age.