Newly Discovered Sex Differences in the Expression of Thousands of Genes in the Cerebral Cortex

A new study, based on RNA sequencing of single nuclei, could pave the way towards a better understanding of why certain neurological conditions, including migraine, are more common in one sex versus the other.

Many brain-based diseases, including neurodegenerative, psychiatric, and neurodevelopmental disorders, show significant sex differences in who is most at risk of developing them. While these epidemiological differences are quite clear, why they exist in the first place is less so. That certainly applies to migraine, as researchers still seek a better grasp of why this condition is roughly 2 to 3 times more common in women compared to men.

But now, an important new study advances understanding in this area of research by profiling, in 6 regions of the human cerebral cortex, the expression of genes in women versus men, relying on a technique called single-nucleus RNA-sequencing (snRNA-seq). Using tissue from postmortem brain donors, investigators based at the US National Institutes of Health in Bethesda, US, sequenced the RNA of more than a million single nuclei, discovering that more than 3,000 genes were differentially expressed between the sexes.

Notably, while sex chromosome genes showed the largest differences in expression between males and females, most of the differentially expressed genes were autosomal genes, which are genes not on the sex chromosomes.

Another key finding was that more genes than expected escaped X-chromosome inactivation, a process in which cells in females show reduced expression of genes on one of their two X chromosomes. Knowing which genes escape this process could explain why certain neurological disorders affect one sex over the other.

“I think this is important work and a step toward better understanding sex differences in brain function,” wrote William Renthal, Brigham and Women’s Hospital, and Harvard Medical School, Boston, US, in an email to Migraine Science Collaborative. “While additional donor samples could further strengthen their findings, the methods they use and initial findings are rigorous and exciting,” according to Renthal, a headache and pain investigator who uses snRNA-seq in his own research but was not involved with the new work.

While the study did not focus on migraine per se, it has obvious relevance to it.

“Migraine is one of the clearest examples of a neurologic disease with strong sex differences, particularly after puberty, and the paper itself cites migraine among conditions more common in females,” said Renthal. “This paper shows that sex can alter the molecular context in which genetic risk acts, and that this context differs by cell type and brain region. So, while it doesn’t study migraine specifically, it contributes to a foundation of knowledge about gene expression variation between males and females, and may help explain how similar genetic risk for migraine between males and females can lead to different rates of migraine.”

The study, and a related perspective, appeared in the April 16, 2026, issue of the journal Science.

Different scales, complementary approaches
To learn more about the nature and scope of sex differences in gene expression in the brain, co-corresponding authors Alex DeCasien and Armin Raznahan, of the US National Institute of Mental Health in Bethesda, wanted to overcome a hurdle in previous research of this kind.

“At the time we started this study, most work on sex differences in gene activity (expression) in the brain had looked at only a few brain regions – typically with brain samples that mixed up lots of different cells. We wanted to be able to catch if differences in brain gene expression varied between brain regions and cell types,” they wrote in a joint email to Migraine Science Collaborative.

To do so, they applied snRNA-seq, a technique that can measure gene expression in individual cell types, to 169 cortical samples obtained from 15 female and 15 male postmortem brain donors, all adults from 26 to 78 years of age. The samples came from 6 cortical regions, 4 of which showed reproducible sex differences in gray matter volume, and 2 of which did not, in previous structural magnetic resonance imaging studies. Choosing which brain regions to study based on that imaging evidence, the investigators believe, is one of the most novel contributions of their study – and one that complements the snRNA-seq approach.

“[W]e picked the brain regions for cellular analysis using an independent measure of sex differences from neuroimaging. RNA sequencing and neuroimaging represent two very different spatial scales of brain measurement – so our study design also let us ask if there were particular cellular features (micron scale) that tracked with sex differences in the average volume of a brain region (mm [millimeter] scale),” said DeCasien and Raznahan.

All in all, the study dataset would include roughly 1.2 million nuclei from excitatory neurons, inhibitory neurons, and glia (a non-neuronal cell type) and other cells, with the researchers focusing on 19 subclasses of these cell types for their analysis.

Consistent with previous studies, there were no sex differences in cell type proportions across or within any of the brain regions examined in the study. This finding suggested that differences in gray matter volume that exist among males and females stem more from differences in cellular morphology – the shape, size, and other features of cells.

An X surprise
Also in line with previous work, sex accounted for less than 1% of the overall variance in gene expression, with genes on the sex chromosomes showing the largest differences in expression between males and females. However, the researchers identified more than 3,000 distinct genes with sex-biased expression, most of which, it turned out, were autosomal genes. When looking specifically at where the sex differences were consistent across all brain regions and cell types, the researchers found that 133 genes met this criteria, with 119 of those being autosomal genes.

“This is notable because few autosomal genes with sex-biased expression have been reported previously in human or mouse cortex,” wrote Jessica Tollkuhn, Cold Spring Harbor Laboratory, US, and S. Marc Breedlove, Michigan State University, East Lansing, US, in the accompanying perspective.

There was another surprise related to the sex chromosomes: Many more genes than expected escaped from X-chromosome inactivation. This inactivation is an important process, referred to as a gene dosage compensation mechanism; without it, females would have twice the expression levels of genes on the X chromosome.

“Because one of the two X chromosomes in every XX female cell is ‘silenced’ (inactivated), the few genes on that chromosome that manage to ‘escape’ the inactivation represent a major aspect of sex differences in gene expression (because these X genes can only be expressed from one chromosome in males, but may also be partly expressed from the second inactivated X in females),” the authors explained to Migraine Science Collaborative. “There is some evidence from prior work that X-inactivation and escape may vary between different tissues and cell types – but there had never been a detailed cell type survey in the brain. Our study takes a first look at this, and reveals several brain-specific escape genes.”

The payoff of knowing which genes escape X-inactivation could be a better understanding of sex biases in the prevalence of certain brain disorders, as these genes may contribute to the pathophysiological processes underlying disease.

“I think [X-inactivation escape] is potentially really important for understanding neurological afflictions that affect women more often than they affect men,” including pain and migraine, said Ted Price, a pain researcher at the University of Texas at Dallas, US. Price, who performs snRNA-seq analysis from human dorsal root ganglia in his own research using tissue from organ donors, was not involved in the new work.

Seeking patterns, biological processes, and cellular components
Zeroing in on the autosomal genes, the researchers next wanted to understand whether there were patterns to the sex differences in gene expression, based on brain region, on cell type, or on both.

Thirteen distinct patterns, which the researchers called signatures or clusters, emerged. Some clusters were region specific. For instance, there was a cluster in the fusiform gyrus, where both neurons and glial cells showed sex differences in gene expression in that particular area of the cortex. Of note, the fusiform gyrus, which is the cortical region associated with the largest sex differences in brain volume, also showed the largest differences in gene expression of all the cortical areas included in the study.

Other patterns emerged based on cell type. For example, there was a cluster comprising astrocytes, a type of glial cell, across several brain regions. And then there were patterns according to both cell type and brain region. For instance, there were clusters comprising inhibitory neurons, excitatory neurons, or both, in the caudal insula and retrosplenial cortex.

The researchers also wondered about which biological processes the differentially expressed autosomal genes within each cluster play a role in – and there were many, ranging from immunity to cell signaling to metabolism. There was also a wide range of cellular components associated with the differentially expressed genes, including synapses, membranes, and the extracellular matrix, to name a few.

“By pinpointing these sexually differentiated processes, the data provide a treasure trove for the discovery of biomarkers of and/or therapeutic targets for differential disease risk in men and women,” wrote Tollkuhn and Breedlove in their perspective.

Additional findings indicated that sex hormones regulated the sex-biased autosomal genes – estrogen and progesterone in the case of female-biased genes, and androgens for male-biased genes – as did transcription factors linked to the X chromosome.

Finally, many of the sex-biased genes were ones that a previous genome-wide association study (GWAS) had identified as being associated with the risk for conditions more common in one sex than another, especially for certain neurodevelopmental diseases.

Sex differences in the human brain at cell type resolution. Credit: DeCasien et al. Science. 2026 Apr 16;392(6795):eaea9063.

A resource for the field – and an appreciation of tissue donors
As for future studies, the investigators are following up in a number of areas. One is how sex differences in gene expression might vary based on factors such as age or exposure to different environmental factors.

Another is to prove their findings that hormones, and genes on the sex chromosomes, drive the sex differences in autosomal gene expression. The researchers are also interested in understanding whether gender plays a role in the sex differences. Finally, they’re seeking a better grasp of the association of sex-biased gene expression with the biological processes and risk for disease that their current study highlighted.

The investigators also hope the study spurs efforts by other researchers to better understand sex differences in gene expression. That would include those working in the migraine field.

“We hope that the detailed lists we’ve published on sex-biased gene expression in the human brain can serve as a rich database for other scientists to cross-reference with their own findings. For example, scientists looking at risk genes for migraine, or genes that differ in expression as a function of migraine status or risk factors, can now ask, do these genes show sex differences in expression within any specific cell types or brain regions? That might help narrow down mechanisms for sex differences in migraine,” said DeCasien and Raznahan.

“Which sex-biased expression differences actually affect brain function and migraine susceptibility? This paper is a cellular map that can be used to start to answer this question,” said Renthal, along similar lines.

Finally, while everyone who spoke to Migraine Science Collaborative agreed that a larger number of donor samples are needed for future studies, the enormous contribution of people who have donated tissue thus far for research is already being felt. This is something that drew particular attention from Price, who said he has come to greatly value the participation of donors in the research he does to better understand pain.

“It’s amazing how widely recognized it is among people that this is a really meaningful gift to humanity – to be able to give your precious tissues to turn it into understanding that can hopefully change what we know about the brain and the nervous system, and hopefully also lead to big advances in treating neurological disorders,” he said.

Neil Andrews is a science journalist and executive editor of the Migraine Science Collaborative.

Image credit: 123RF Stock Photo/crystallight

References
Sex effects on gene expression across the human cerebral cortex at cell type resolution.
DeCasien et al.
Science. 2026 Apr 16;392(6795):eaea9063.

Express yourself.
Tollkuhn J, Breedlove SM.
Science. 2026 Apr 16;392(6795):251-252.