Unraveling the complexities of the prefrontal cortex in humans and non-human primates
Sir Charles Sherrington famously referred to the brain as an enchanted loom. More than 80 years later, that metaphor rings truer than ever: like a loom, the brain weaves together a pattern of heterogeneous cell types, anatomy, and tissue organization into a tapestry greater than the sum of its parts.
One notable part is the granular dorsolateral prefrontal cortex (dlPFC), a brain region highly involved in emotional processing, higher cognition, and neuropsychiatric disease (1,2). The dlPFC is also an evolutionary specialization of primates and, as much as we share with our closest evolutionary cousins, this begs the question: what is it about this brain region that makes us similar? More importantly, what makes us different?
Recently, Ma et al. tackled these questions by using single nuclei RNA-seq (snRNA-seq) and single nuclei ATAC-seq (snATAC-seq) to better understand the complex weave of primate species–specific cellular and anatomical diversity (3).
Common patterns and divergent threads in the dlPFC
Using the Chromium Single Cell 3’ Gene Expression assay, researchers performed snRNA-seq on dlPFC tissue from humans and three other primates (chimpanzees, rhesus macaques, and common marmosets) and characterized 114 distinct subtypes of cells. While 109 subtypes were detected in each of the four species, five of these exhibited species specificity:
- An excitatory neuronal subtype (absent in marmosets)
- An inhibitory neuronal subtype (present only in marmosets)
- An astroglial subtype (seen only in humans and chimpanzees)
- A microglial subtype (seen only in humans and chimpanzees)
- A human-specific microglial subtype (not detected in any other species)
After validating their findings with other species-specific snRNA-seq datasets, the researchers chose to focus on the human-specific microglial subtype. Intriguingly, they noted this subtype was similar to a previously reported population involved in immune homeostasis that was notably distinct from disease-associated microglial subtypes.
Turning their attention to the 19 genes that were both 1) enriched in microglia in humans and 2) enriched specifically in microglia versus another immune cell type (macrophages), the group identified FOXP2–a transcription factor linked to multiple neuropsychiatric disorders–as a target of interest. The researchers highlighted this as interesting given that, while some FOXP2-linked diseases were associated with microglial activation and neuroinflammation, prior studies had shown FOXP2 expression only in neurons.
Tugging on the thread of microglial FOXP2
Researchers next validated their findings of FOXP2 expression in both neurons and microglia in human dlPFC by demonstrating immunofluorescent colocalization of FOXP2 with neuronal (NeuN) and microglial (IBA1) markers.
Interestingly, while other snRNA-seq and microglial bulk RNA-seq data confirmed human- and microglia-specific FOXP2 expression, the group discovered it was enriched in an excitatory neuronal subtype—the small excitatory neurons found in cortical layers 3 to 5 (L3-5) that are responsible for the aforementioned “granular” appearance of the primate dlPFC. Lending weight to the notion that this is a primate evolutionary specialization, this enrichment was specific to primates (not observed in 30 non-primate mammals) and to areas involved in higher cognitive functions.
FOXP2 and the transcriptional weave of the dlPFC
Since FOXP2 is a transcription factor, the researchers used Chromium Single Cell Multiome ATAC + Gene Expression to construct possible regulatory networks by simultaneously assaying gene expression and chromatin accessibility in human dlPFC.
Using this multiomic dataset, they characterized both potential upstream regulators of FOXP2 and its prospective downstream targets. Consistent with the role of FOXP2 in neurological disorders, DSCAM—a Down’s syndrome–associated gene and predicted target of FOXP2—was specifically expressed in human microglia only. Similarly, multiple FOXP2 targets restricted to neurons in layers 3 to 5 exhibited primate-specific expression compared to mice, consistent with FOXP2 playing a major role in the coordination of these networks.
Following the thread to its conclusion
It’s an often-quoted statistic that we share roughly 99% of our DNA with chimpanzees and bonobos, but our overwhelming genetic and neuroanatomical similarities belie our stark cognitive differences. In this work, Ma et al. demonstrated just a few of the ways that, in spite of the broad cellular similarities humans have in common with our closest cousins, marked transcriptional differences can arise due to subtle shifts in species-specific cellular diversity.
While we touched on a lot of their findings in this blog, this was only a small subset of the truly rich dataset Ma et al. generated. Read their article to discover what else they found, or take a deeper dive into the single cell and multiomic tools they used—and learn how they can help you unravel your own research questions.
- Preuss TM and Wise SP. Evolution of prefrontal cortex. Neuropsychopharmacology 47: 3–19 (2022). doi: 10.1038/s41386-021-01076-5
- Lewis DA and Mirnics K. Transcriptomic alterations in schizophrenia: disturbing the functional architecture of the dorsolateral prefrontal cortex. Progress in Brain Research 158: 141–152 (2006). doi:10.1016/S0079-6123(06)58007-0
- Ma S, et al. Molecular and cellular evolution of the primate dorsolateral prefrontal cortex. Science 377 (2022). doi: 10.1126/science.abo7257