An international team of researchers has maintained brain organoids, three-dimensional models of human brain tissue, for longer than ever before. For over five years, the tissue continued to mature in ways that closely resemble human brain development. The findings have been published in Nature. By extending the lifespan of these organoids, the researchers create new opportunities to investigate neurodevelopment, model brain disorders and test potential drugs.
The human brain continues to develop until the age of twenty. To study this prolonged process, researchers have traditionally relied on donated human brain tissue and animal models. Both approaches have generated major insights, but each has its limitations: donated tissue provides only snapshots of brain development, and animal brains differ from the human brain in cell-type composition and timing of development.
Researchers therefore increasingly use brain organoids, models of human brain tissue derived from stem cells, to study human brain development. "These models allow us to track development over time and examine how different brain cell types emerge," says Noelia Antón-Bolaños , Assistant Professor at UMC Utrecht. So far, however, most studies have focused on the earliest stages of development. This is because researchers could not maintain brain organoids in culture for extended periods. Antón-Bolaños and her colleagues therefore investigated how far human brain organoids can continue to mature.
Sustaining development
The researchers faced one key challenge: keeping the brain organoids healthy for several years. Standard culture conditions did not adequately support neuronal activity over extended periods. "During human brain development, neurons display spontaneous activity," Antón-Bolaños explains. "By adapting the composition of the culture medium, we supported that activity, kept the neurons active, and maintained the neuronal populations for much longer." Dedicated researchers also took care of the cultures in an isolated environment throughout the entire study.
Using this approach, the team maintained organoids for over five years. At defined time points, the researchers profiled the cell types present, gene-expression patterns, epigenetic changes, and neuronal activity.
Keeping track of time
The researchers then examined whether the organoids simply stayed alive or continued to develop. Several lines of evidence pointed to the same conclusion. Different brain cell types appeared in the same order as during human brain development, neurons formed increasingly complex connections, and genes became active or inactive at the expected times.
Some of the strongest evidence came from small chemical modifications to DNA. "In the human brain, these epigenetic changes accumulate according to a characteristic developmental pattern," Antón-Bolaños explains. "We observed the same pattern in the brain organoids." After approximately one year, the organoids even displayed features that normally emerge only after birth. "The cells are outside the body, yet they still follow approximately the same developmental timeline as we do—and even more closely than we had anticipated."
Retaining memory
The team also found that mature cells retained a memory of developmental time. "When we dissociated an older organoid and allowed the cells to grow again, they produced the cell types associated with a late developmental stage," Antón-Bolaños says."Yet when we combined older cells with younger cells, the older cells regained the ability to produce neurons—but only the types associated with later stages of development."
This is striking because the human brain stops producing neurons relatively early and later switches to producing glial cells. "If we can identify the signals that reactivate neuron production in older cells, it could provide new ways to study neurodegenerative disorders."
A platform for studying brain disorders
Because the organoids continue to mature over years, researchers can examine a much larger portion of the human developmental trajectory. This is particularly relevant for conditions such as autism spectrum disorder and schizophrenia , in which altered neurodevelopment plays an important role. "These brain organoids are also highly reproducible," Antón-Bolaños says. "This is essential for using them as disease models and may eventually support drug-testing studies in organoids."
The organoids are still developing in the laboratory. The researchers now want to determine how environmental cues, such as stimulation with light, improve further maturation. The field also aims to improve features that remain incomplete, including vascularization and the layered organization of the cerebral cortex.
"We now know that these models have the capacity to continue developing for years," Antón-Bolaños says. "The next step is to understand how to provide optimal conditions for that capacity to unfold. That will bring us closer to more faithful models of the human brain."
About this research
This research was conducted at Harvard University in the laboratory of Paola Arlotta. The study was led by Noelia Antón-Bolaños and Irene Faravelli.
