Although the cerebral cortex is organized into six excitatory neuronal layers, it is unclear whether glial cells show distinct layering. In the present study, we developed a high-content pipeline, the large-area spatial transcriptomic (LaST) map, which can quantify single-cell 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 the 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.
For more details, please refer to the paper (doi:10.1038/s41593-020-0602-1) by Dr Omer Ali Bayraktar.
Fig. 1 in the paper: LaST map pipeline for mapping cortical neuronal subtypes in situ.
a, Design of automated spatial transcriptomic pipeline. b, High-resolution imaging of large tissue areas. Shown are ×40 z-projection images of Rorb+ L4 neurons in the P14 mouse barrel cortex. c, Automated mapping of layer neuron marker expression and layer neuron subtypes in the mouse barrel cortex. Automatically identified single neurons are plotted as solid circles and colored according to expression (middle panels) or subtype classification (right panels). d, Identification of neuronal subtypes based on unbiased classification of single-cell-level smFISH data; t-SNE and hierarchical clustering of 46,888 cortical neurons yielded 10 subpopulations. Top: violin plots show single-cell expression profiles of clusters; highest RNA spot counts per cell are shown on the left. Middle: histograms showing total number of cells per cluster. Bottom: spatial distribution of clusters across cortical areas and five normalized cortical depth bins, shown as the percentage of total neurons in given area/depth bin (bottom). e–h, Single-cell mapping of cortical neuron subtypes. e, Low-magnification images of P14 hemisections from four different anatomical levels. f, Broad cortical areas included in the analysis. g, Maps of 10 major neuronal populations. h, Spatial distribution of area-restricted L5 RorbhighCux2midBcl11blow neurons (n = 1 mouse, 10 tissue sections independently imaged). Scale bars, 10 μm (b), 100 μm (c) and 1 mm (e). A, auditory; M, motor; PT, parietal; S–A, anterior somatosensory; S–BF, somatosensory barrel; S–L, somatosensory lateral; S–M, medial somatosensory; V, visual.