During fetal development, the human brain generates an excess of neurons, most of which are eliminated shortly before and after birth. New research from Harvard Medical School and Boston Children's Hospital suggests that this is a critical quality-control mechanism.
A study published Oct 28 in Nature shows that neurons with multiple copy number variants — abnormal duplications or deletions of segments of DNA — are surprisingly widespread in the developing brain. Copy number variants can impact brain function and may contribute to neuropsychiatric conditions, including autism, schizophrenia, epilepsy, and intellectual disability.
But the research also shows that the vast majority of these flawed neurons are selectively eliminated, likely because the genomic alterations disrupt their ability to form synapses.
"Between 13 and 46 percent of neurons in the developing brain are riddled with copy number variants, but there seems to be a mechanism by which these abnormal neurons are eliminated around the time of birth," said co-senior author Peter Park , professor of biomedical informatics in the Blavatnik Institute at HMS.
The findings reveal an important process in the developing brain and could lead to a better understanding of autism and other disorders in which neural development goes awry.
"The brain is selecting for genomic integrity," said co-first author Diane Shao , HMS instructor of neurology at Boston Children's. "If that quality control system can be disrupted, that could be part of what goes wrong in neurodevelopmental disorders like autism. Understanding the mechanism could eventually point us toward what to look for, and maybe what to protect."
Competition as quality control
To understand the prevalence of copy number variants within developing human brains, the researchers needed to sequence the genomes of thousands of individual neurons — a prohibitively expensive process.
The researchers designed a new, lower-cost methodology that allowed them to lightly sequence the genomes of single neurons, akin to taking a low-resolution photo. They used this method on 1,526 neurons from 16 individuals, then validated their results with deeper sequencing of 831 neurons from 12 individuals. All the samples came from tissue banks funded by the National Institutes of Health and other organizations.
Co-first author Yifan Zhao, who was a PhD student in Park's lab at the time, developed computational methods to interpret the data and identify copy number variants within each genome.
"We've long known that copy number changes are important in brain disorders, but we haven't had a good way to study them in individual neurons at scale," Park said. "Now we can see these changes one neuron at a time, map where in the genome they occur, and compare how common they are from before birth to old age."
The researchers found that an average of 25.9 percent (and in some cases nearly half) of neurons in the prenatal cortex had widespread copy number variants, but this rapidly decreased to an average of 6.2 percent after birth.
"We were totally unprepared for how common these were in early development," said co-senior author Christopher Walsh , HMS Bullard Professor of Pediatrics and Neurology at Boston Children's Hospital. "But it makes sense in the context of what we know about the formation of excess neurons and in the context of the IVF literature." Over the last decade, in vitro fertilization clinics have found that early embryos have very high rates of cells with abnormal genomes, but the vast majority of them develop into healthy children, Walsh said.
The researchers also confirmed earlier findings that flawed neurons continue to be eliminated throughout adulthood, albeit at a much slower rate.
The team then took a closer look at how abnormal neurons were being eliminated, sequencing both the DNA and the RNA from each of 44,861 individual cells to identify possible pathways involved in the process. They found that neurons with widespread copy number variants also had abnormal expression of genes involved in the creation and maintenance of synapses — connections with other neurons.
Previous research has shown that neurons compete for synapses and those that fail to make these critical connections during development typically die off. The findings suggest that neurons with too many copy number variants can't compete as effectively.
"This competition for synapses has been described for years, but without the benefit of a genomic understanding," said Walsh, who is also an investigator of the Howard Hughes Medical Institute. "We didn't realize that this struggle for survival — this Darwinian competition — was nature's quality control mechanism in the brain."
Diversity and dysregulation
While the most abnormal neurons seem to be eliminated early on in development, the research shows that some variation persists into adulthood even in healthy brains. The researchers don't yet know whether this diversity is simply an accidental byproduct of development or contributes something useful, but it is one of many threads they are intending to pull.
Currently, they are looking into whether this neural elimination process is different in developing brains of those with autism and other neurodevelopmental disorders. Autism has already been linked to dysregulated synaptic pruning (the trimming of weak or unused neural connections).
Walsh and his colleagues are also exploring whether this process plays a role in the death of neurons that occurs in adult-onset neurodegenerative disorders, such as Alzheimer's disease.
"We think that a lot of different diseases may show abnormalities in this process," Walsh said. "It's opened up a whole new universe of things to think about."
Authorship, funding, disclosures
Additional authors of this work include Joseph Brew, Yahia Adla, Urmi Ghosh, Vinayak V. Viswanadham, Lovelace J. Luquette, Sijing Zhao, Andrea J. Kriz, Xuyu Qian, Takumi Taketomi, Shannon Coy, Sandro Santagata , Michael Miller, and Fuminori Tsuruta.
This work was supported by the National Institutes of Health (grants K08-NS128074, R01-NS032457, R01-HG012573, and R01-CA269805), the Templeton Foundation, the SMaHT consortium (UH3-NS132144), the Program in Genetics and Genomics PhD Training (grant T32GM141745), the Howard Hughes Medical Institute Jane Coffin Childs Fellowship, the HMS Program in Neuroscience Award, Ludwig Cancer Research and the Ludwig Center at Harvard, and the Brigham and Women's Hospital President's Scholars Award. Christopher A. Walsh is an Investigator of the Howard Hughes Medical Institute.
Park and Walsh are members of the scientific advisory board for Bioskryb Genomics, Inc. Walsh is also a member of the scientific advisory board for Mosaica Therapeutics and an advisor to Maze Therapeutics.
