Astrocyte layers in the mammalian cerebral cortex revealed by a single-cell in situ transcriptomic map

Abstract

While the cerebral cortex is organized into six excitatory neuronal layers, it is unclear whether glial cells show distinct layering. Here, we developed a high-content pipeline, the Large-area Spatial Transcriptomic (LaST) map, which can quantify singlecell gene expression in situ. Screening 46 candidate genes for astrocyte diversity across the mouse cortex, we identified superficial, mid, and deep astrocyte identities in gradient layer patterns that were distinct from those of neurons. Astrocyte layer features, established in early postnatal cortex, mostly persisted in adult mouse and human cortex. Single cell RNA sequencing and spatial reconstruction analysis further confirmed the presence of astrocyte layers in the adult cortex. Satb2 and Reeler mutations that shifted neuronal post-mitotic development were sufficient to alter glial layering, indicating an instructive role for neuronal cues. Finally, astrocyte layer patterns diverged between mouse cortical regions. These findings indicate that excitatory neurons and astrocytes are organized into distinct lineage-associated laminae.

Identification of region-specific astrocyte subtypes at single cell resolution

“…The robustness of our data is demonstrated by consistency with a study from Bayraktar et al 41 , which uses large-scale ISH-based mapping to study gene expression in somatosensory cortex of P14 mice (see Supplementary Fig. 19).…”

“…Our results distinguish multiple astrocyte subtypes at the transcriptomic level. However, the picture is substantially more complicated than for neurons, with little clear evidence for distinct cellular hierarchies in clustering, most likely due to multiple axes of heterogeneity being present 4,5,20,41 . However, although astrocyte subtypes share common genes associated with core functions, the transcriptomic differences between the subtypes are still sufficiently large to suggest distinct specializations in known astrocyte functions 13,42 .…”

Molecular Logic of Cellular Diversification in the Mammalian Cerebral Cortex

“…The inferred tree also uncovered previously unappreciated expression patterns of genes traditionally considered lineage-restricted, which we validated by Slide-seq. For example, Rorb, a marker of layer 4 stellate neurons, and Fezf2, a selector gene for SCPNs, were also expressed in the astroglia lineage, consistent with recent reports that Fezf2 is expressed in astrocytes (Bayraktar, Bartels et al 2020) (Supplementary information Figure 5B). Pcp4, a marker for CFuPN (Arlotta, Molyneaux et al 2005), was expressed in migratory neurons of both the CFuPN and CPN lineages, which we confirmed by presence of Pcp4 signal in the intermediate zone using Slide-seq (Supplementary information Figure 5A).…”

No Longer Underappreciated: The Emerging Concept of Astrocyte Heterogeneity in Neuroscience

“…It is also required for expression of the potassium channel Kir4.1 in astrocytes of the ventral horn of the spinal cord [47]. Furthermore, genetic manipulations affecting cortical layering, for example neuronal deletion of Dab1 [16] or Satb2 [48], also influence astrocyte positioning.…”