Abstract
The neocortex consists of a vast number of diverse neurons that form distinct layers and intricate circuits at the single-cell resolution to support complex brain functions1. Diverse cell-surface molecules are thought to be key for defining neuronal identity, and they mediate interneuronal interactions for structural and functional organization2,3,4,5,6. However, the precise mechanisms that control the fine neuronal organization of the neocortex remain largely unclear. Here, by integrating in-depth single-cell RNA-sequencing analysis, progenitor lineage labelling and mosaic functional analysis, we report that the diverse yet patterned expression of clustered protocadherins (cPCDHs)—the largest subgroup of the cadherin superfamily of cell-adhesion molecules7—regulates the precise spatial arrangement and synaptic connectivity of excitatory neurons in the mouse neocortex. The expression of cPcdh genes in individual neocortical excitatory neurons is diverse yet exhibits distinct composition patterns linked to their developmental origin and spatial positioning. A reduction in functional cPCDH expression causes a lateral clustering of clonally related excitatory neurons originating from the same neural progenitor and a significant increase in synaptic connectivity. By contrast, overexpression of a single cPCDH isoform leads to a lateral dispersion of clonally related excitatory neurons and a considerable decrease in synaptic connectivity. These results suggest that patterned cPCDH expression biases fine spatial and functional organization of individual neocortical excitatory neurons in the mammalian brain.
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Data availability
The scRNA-seq matrix of cPcdh isoform expression is provided in Supplementary Table 4. Source data are provided with this paper.
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No code or model was generated in this study.
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Acknowledgements
We thank members of the Shi laboratory and H. Jiang for discussions and input; and J. L. Lefebvre and J. A. Weiner for the Pcdhgfcon3 and Pcdhgc3 mouse lines. This work was supported by grants from the Ministry of Science and Technology of China (2021ZD0202300), the National Natural Science Foundation of China (32021002 to S.-H.S. and 31630039 to Q.W.), Beijing Outstanding Young Scientist Program (BJJWZYJH01201910003012), Beijing Municipal Science & Technology Commission (Z20111000530000 and Z211100003321001), the Chinese Institute for Brain Research (Beijing) and the Simons Foundation (SFARI GC232866) (to S.-H.S.) and the Postdoctoral Program of the Tsinghua Centre for Life Sciences (to S.L.).
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X.L. and S.-H.S. conceived the project. X.L. performed the majority of the experiments and data analysis. S.L., B.L. and S.H. performed electrophysiological recordings and analysis. X.G. and S.L. helped with the scRNA-seq experiments. J.L., X.-Y.Y., X.G., S.Z. and J. Yang performed the scRNA-seq data analysis. Y.L. helped with the immunostaining. X.Z. and J. Yan helped with the mouse genotyping. A.L.J. advised on the project. H.S. helped with isoform detection and sequencing design. X.L., Q.W. and S.-H.S. wrote the paper with input from all of the other authors.
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Extended data figures and tables
Extended Data Fig. 1 MADM-labelling of individual excitatory neuron clones in the neocortex and quality check of cell aspiration and extraction.
a, Schematic diagram of MADM labelling strategy. b, c, Representative 3D reconstruction images of symmetric (b) or asymmetric (c) excitatory neuron clones induced at E12 and analysed at P21. Different coloured dots represent the cell bodies of labelled neurons. The x-/y-/z-axes indicate the spatial orientation of the clone with the x-axis parallel to the brain pial surface and the y-axis perpendicular to the pial surface. Coloured lines indicate the layer boundaries. Scale bars, 100 μm. d, e, Representative fluorescence images of individual clones in live brain slices subjected to glass pipette aspiration and extraction shown in Fig. 1band c. Scale bars, 50 μm. f—h, Quantifications of amplified cDNAs from negative controls (f, g) or individual extracted neurons (h) in the same experiment setting. i, Quantification of the cDNA library of individual extracted neurons. Data are representative of at least three independent experiments.
Extended Data Fig. 2 Quality check of the scRNA-seq analysis.
a—c, Quality control of individual extracted neurons after sequencing. Neurons with a low total detected gene number (n = 14), or a high mitochondrial gene (n = 2), or ERCC (n = 7) percentage (orange) were excluded from further analysis. Each symbol represents a neuron. d, e, Quantification of the number of the total detected genes (d) and cPcdh isoforms (e) in our clone dataset (n = 188). Each symbol represents a neuron. f, Histogram of the number of the detected cPcdh isoforms per neuron in our clone dataset (n = 188). g, h, Quantification of the number of the total detected gene (g) and cPcdh isoforms (h) in the previously published randomly collected neocortical excitatory neuron scRNA-seq dataset from the Allen Institute42 (n = 4,152). Each symbol represents a neuron. i, Histogram of the number of the detected cPcdh isoforms in the previously published randomly collected neocortical excitatory neuron scRNA-seq dataset from the Allen Institute42 (n = 4,152). j, Quantification of the number of the total detected cPcdh isoforms in clonally related excitatory neurons from different neocortical regions (motor cortex/MO, n = 32; somatosensory cortex/SS, n = 116; visual cortex/VIS, n = 40). k, Pearson correlation analysis between the average expression level of each cPcdh isoform in clonally related excitatory neurons of SS (n = 116) and VIS (n = 40). Each dot represents a cPcdh isoform. The grey bar indicates the 95% confidence interval. A similar display is used in subsequent panels (l–n). l, Pearson correlation analysis between the average expression level of each cPcdh isoform in clonally related excitatory neurons of SS (n = 116) and MO (n = 32). m, Pearson correlation analysis between the average expression level of each cPcdh isoform in clonally related excitatory neurons of MO (n = 32) and VIS (n = 40). n, Pearson correlation analysis between the average expression level of each cPcdh isoform in randomly collected excitatory neurons of MO (n = 3,893) and VIS (n = 7,347) in the previously published scRNA-seq datasets from the Allen Institute54. o, Frequency distribution of the mean similarities of cPcdh isoform expression pattern in the non-clonal dataset (n = 1,000 trails, grey bars), and the mean similarity of the clonal dataset (red line) from 32 clones. The non-clonal dataset was generated by a random permutation of the clonal dataset and statistics were performed using the permutation test (see Methods). p, Quantification of the pairwise similarity of cPcdh isoform expression for clonally related neocortical excitatory neurons in the same or different layers per clone (same layers, n = 31; different layers, n = 20). q, Quantification of the pairwise similarity of cPcdh isoform expression for neurons in the same or different layers of the randomly collected 150 neocortical excitatory neurons from the previously published Allen Institute dataset42 (same layers, n = 600; different layers, n = 400). r, Quantification of the pairwise similarity of cPcdh isoform expression for randomly collected excitatory neurons across different layers from the previously published Allen Institute dataset42 (L2/3-L2/3, n = 200; L2/3-L5, n = 200; L2/3-L6, n = 200). The n numbers indicate neurons (a–n), clones (p), and neuron pairs (q, r). Data are representative of at least three independent experiments. One-way ANOVA test without adjusted P value (j); Two-tailed Pearson correlation analysis (k–n); One-tailed permutation test (o); Two-tailed unpaired Student’s t-test (p–r). Box plots as in Fig. 1.
Extended Data Fig. 3 Combinatorial expression of cPcdh isoforms in individual neocortical excitatory neurons in the clone dataset.
Heatmap of cPcdh transcripts in 188 individual neocortical excitatory neurons from 32 clones. The existence of individual cPcdh isoforms is indicated by the red or green coloured boxes, the colour reflects fluorescence labelling, and the expression level is indicated by the colour gradient. Note that the majority of neurons express multiple cPcdh isoforms from three Pcdh clusters (a, b, and g) and the C-type Pcdh isoforms (ac2 and gc5).
Extended Data Fig. 4 Combinatorial expression of cPcdh isoforms in individual excitatory neurons of the motor cortex in the previously published Allen Institute dataset.
Heatmap of cPcdh transcripts in 200 individual excitatory neurons in the motor cortex randomly selected from the published Allen Institute dataset42. The existence of individual cPcdh isoforms is indicated by the blue coloured boxes and the expression level is indicated by the colour gradient. Note that the majority of neurons express multiple cPcdh isoforms from three Pcdh clusters (a, b, and g) and the C-type Pcdh isoforms (ac2 and gc5), similar to the neurons in the clone dataset.
Extended Data Fig. 5 Confirmation of layer identities for individual neocortical excitatory neurons in the clone dataset.
a, Uniform manifold approximation and projection (UMAP) plot for alignment of individual neurons in our clone dataset (L2/3, n = 95; L4, n = 26; L5, n = 41; L6, n = 26) to the previously published neocortical excitatory neuron scRNA-seq dataset from the Allen Institute57 (n = 9,100). Large dots with black outlines represent individual clonally related excitatory neuron in our dataset and small dots with no outline represent individual excitatory neurons in the reference Allen Institute dataset. Different colours reflect different subtypes of neocortical excitatory neurons. The n numbers indicate neurons. b, Sankey plot for the result of transferring subtype labels from the reference dataset (predicted) to our clone dataset (raw) based on scRNA-seq. Different colours reflect different subtypes of neocortical excitatory neurons. The mapped cell numbers are shown in the graph inset. c, No significant difference in the percentage of cells mapped to the corresponding reference neuron subtypes/layers between our dataset and a previously published dataset using a similar method45. The mapped cell numbers are shown in the bar graph. Two-tailed paired Student’s t-test was used for statistical analysis.
Extended Data Fig. 6 PCDHγ removal causes a lateral clustering of sister and cousin excitatory neurons in the neocortex.
a, b, Representative 3D reconstruction images of P21 WT (left) and Pcdhg cKO (right) symmetric (a) and asymmetric (b) excitatory neuron clones labelled by MADM. Coloured lines indicate the layer boundaries and coloured dots represent the cell bodies of labelled neurons. The x-/y-/z-axes indicate the spatial orientation of the clone with the x-axis parallel to the brain pial surface and the y-axis perpendicular to the pial surface. Scale bars: 100 μm. c—f, Quantification of the pairwise (c, d) and maximal (e, f) lateral and radial distances between sister neurons in individual WT and Pcdhg cKO clones (WT, n = 90 clones; Pcdhg cKO, n = 90 clones). g—j, Quantification of the pairwise (g, h) and maximal (i, j) lateral and radial distances between cousin neurons in individual WT and Pcdhg cKO clones (WT, n = 68 clones; Pcdhg cKO, n = 74 clones). Data are representative of four independent experiments. Two-tailed unpaired Student’s t-test was used for statistical analysis. Box plots as in Fig. 1.
Extended Data Fig. 7 PCDHγ removal does not affect the overall layer formation, the excitatory neuron density, or the dendritic morphology.
a, Quantification of the number of neurons in individual WT and Pcdhg cKO clones (WT, n = 158 clones; Pcdhg cKO, n = 164 clones). b, Representative confocal images of P35 WT (left) and Pcdhg cKO (Emx1-Cre;Pcdhgfcon3/fcon3) (right) neocortices stained for CUX1 (red), a superficial layer excitatory neuron marker, and CTIP2 (green), a deep layer excitatory neuron marker, and counter-stained with DAPI (blue). Note no change in either layer formation or neuronal density in the Pcdhg cKO neocortex compared with the WT control. Scale bars: 100 μm (left) and 20 μm (right). c, d, Quantification of the numbers of CUX1+ (c) and CTIP2+ (d) cells in the 10,000 μm2 rectangle area (WT, n = 4 brains; Pcdhg cKO, n = 3 brains). e, Representative reconstructed dendritic morphologies of WT and Pcdhg cKO excitatory neurons in different layers. Scale bar, 50 μm. f—i, Quantification of the neurite length (f), branch number (g), planar angle (h), or local angle (i) of WT and Pcdhg cKO neurons in different layers (L2/3: WT, n = 71; Pcdhg cKO, n = 55; L4: WT, n = 16; Pcdhg cKO, n = 24; L5: WT, n = 33; Pcdhg cKO, n = 46; L6: WT, n = 23; Pcdhg cKO, n = 40). The n numbers indicate neurons. j, Confocal images of representative E16 Doublecortin (Dcx) promoter driven eGFP expressing (green) cortices electroporated at E14 and stained for DCX (red) and DAPI (blue). The arrow points to the example cell. Note that eGFP-expressing cells are mostly located in the intermediate zone and positive for DCX. Scale bars: 100 µm (left) and 5 µm (right). k, Schematic diagram of in utero intraventricular injection of low-titre retroviruses containing Dcx promoter driven eGFP (green, Ctrl) and Cre/tdTomato (red, Pcdhg cKO) into the Pcdhgfcon3 mice at E12. l, Representative 3D reconstruction images of the control and Pcdhg cKO clones. Note that the Pcdhg cKO clone is more laterally clustered than the control clone. Scale bar, 100 μm. m, Quantification of the number of neurons in individual control and Pcdhg cKO clones (Ctrl, n = 67 clones from ~305 brain slices/5 brains; Pcdhg cKO, n = 78 clones from ~315 brain slices/5 brains). n—q, Quantification of the pairwise (n, o) and maximal (p, q) lateral and radial distances between neurons in individual control and Pcdhg cKO clones (Ctrl, n = 67 clones; Pcdhg cKO, n = 78 clones). Data are representative of four (a), three (b—d, j—q), or ten (e—i) independent experiments. Two-tailed unpaired Student’s t-test was used for statistical test. Data are presented as mean ± SEM (c, d). Box plots as in Fig. 1.
Extended Data Fig. 8 PCDHγ removal causes a lateral clustering of excitatory neuron clones during embryonic and neonatal neocortical development.
a, Representative 3D reconstruction images of E15 WT (left) and Pcdhg cKO (right) clones labelled by MADM at E12. Coloured lines indicate the layer boundaries and coloured dots represent the cell bodies of labelled cells. The x-/y-/z-axes indicate the spatial orientation of the clone with the x-axis parallel to the brain pial surface and the y-axis perpendicular to the pial surface. Similar symbols and displays are used in subsequent panels. Scale bar, 100 μm. b—e, Quantification of the pairwise (b, c) and maximal (d, e) lateral and radial distances between cells in individual E15 WT and Pcdhg cKO clones (WT, n = 54 clones from ~160 brain slices/4 brains; Pcdhg cKO, n = 62 clones from ~150 brain slices/4 brains). f, Representative 3D reconstruction images of E18 WT (left) and Pcdhg cKO (right) clones labelled by MADM at E12. Scale bar, 100 μm. g—j, Quantification of the pairwise (g, h) and maximal (i, j) lateral and radial distances between cells in individual E18 WT and Pcdhg cKO clones (WT, n = 60 clones from ~140 brain slices/4 brains; Pcdhg cKO, n = 76 clones from ~135 brain slices/4 brains). k, l, Representative 3D reconstruction images of P7 WT (left) and Pcdhg cKO (right) symmetric (k) and asymmetric (l) clones labelled by MADM at E12. Coloured lines indicate the layer boundaries and coloured dots represent the cell bodies of labelled neurons. Note that the Pcdhg cKO clones are more laterally clustered than the WT clones. Scale bars, 100 μm. m—p, Quantification of the pairwise (m, n) and maximal (o, p) lateral and radial distances between neurons in individual P7 WT and Pcdhg cKO clones (WT, n = 66 clones from ~204 brain slices/4 brains; Pcdhg cKO, n = 71 clones from ~215 brain slices/4 brains). Data are representative of three independent experiments. Two-tailed unpaired Student’s t-test was used for statistical test. Box plots as in Fig. 1.
Extended Data Fig. 9 PCDHγC3 overexpression leads to a lateral dispersion of sister and cousin excitatory neurons.
a, b, Representative 3D reconstruction images of P21 WT (left) and Pcdhgc3 OE (right) symmetric (a) and asymmetric (b) excitatory neuron clones labelled by MADM. Coloured lines indicate the layer boundaries and coloured dots represent the cell bodies of labelled neurons. The x-/y-/z-axes indicate the spatial orientation of the clone with the x-axis parallel to the brain pial surface and the y-axis perpendicular to the pial surface. Scale bars: 100 μm. c–f, Quantification of the pairwise (c, d) and maximal (e, f) lateral and radial distances between sister neurons in individual WT and Pcdhgc3 overexpressing clones (WT, n = 99 clones; Pcdhgc3 OE, n = 68 clones). g—j, Quantification of the pairwise (g, h) and maximal (i, j) lateral and radial distances between cousin neurons in individual WT and Pcdhgc3 OE clones (WT, n = 79 clones; Pcdhgc3 OE, n = 86 clones). Data are representative of four independent experiments. Two-tailed unpaired Student’s t-test was used for statistical test. Box plots as in Fig. 1.
Extended Data Fig. 10 PCDHγC3 overexpression does not affect the number of clonally related excitatory neurons or the dendritic morphology.
a, Quantification of the number of neurons in individual WT and Pcdhgc3 OE clones in the neocortex (WT, n = 178 clones; Pcdhgc3 OE, n = 154 clones). b, Representative reconstructed dendritic morphologies of WT and Pcdhgc3 OE excitatory neurons in different layers. Scale bar, 50 μm. c–f, Quantification of the neurite length (c), branch number (d), planar angle (e), or local angle (f) of WT and Pcdhgc3 OE neurons in different layers (L2/3: WT, n = 58; Pcdhgc3 OE, n = 55; L4: WT, n = 17; Pcdhgc3 OE, n = 29; L5: WT, n = 34; Pcdhgc3 OE, n = 30; L6: WT, n = 23; Pcdhgc3 OE, n = 31). The n numbers indicate neurons. g, Schematic diagram of in utero intraventricular injection of low-titre retroviruses with Doublecortin (Dcx) promoter driven eGFP (green, Ctrl) and Cre/tdTomato (red, Pcdhgc3 OE) into the Pcdhgc3/c3 mice at E12. h, Representative 3D reconstruction images of the control and Pcdhgc3 OE clones. Note that the Pcdhgc3 OE clones are more laterally dispersed than the control clones. Scale bar, 100 μm. i, Quantification of the number of neurons in individual control and Pcdhgc3 OE clones (Ctrl, n = 62 clones from ~255 brain slices/4 brains; Pcdhgc3 OE, n = 66 clones from ~260 brain slices/4 brains). j–m, Quantification of the pairwise (j, k) and maximal (l, m) lateral and radial distances between neurons in individual control and Pcdhgc3 OE clones (Ctrl, n = 62 clones; Pcdhgc3 OE, n = 66 clones). Data are representative of four (a), ten (b—f), or three (g—m) independent experiments. Two-tailed unpaired Student’s t-test was used for statistical test. Box plots as in Fig. 1.
Extended Data Fig. 11 PCDHγC3 overexpression leads to a lateral dispersion during early neocortical development.
a, Representative 3D reconstruction images of E15 WT (left) and Pcdhgc3 OE (right) clones labelled by MADM at E12. Coloured lines indicate the layer boundaries and coloured dots represent the cell bodies of labelled cells. The x-/y-/z-axes indicate the spatial orientation of the clone with the x-axis parallel to the brain pial surface and the y-axis perpendicular to the pial surface. Similar symbols and displays are used in subsequent panels. Scale bar, 100 μm. b—e, Quantification of the pairwise (b, c) and maximal (d, e) lateral and radial distances between cells in individual E15 WT and Pcdhgc3 OE clones (WT, n = 60 clones from ~180 brain slices/5 brains; Pcdhgc3 OE, n = 67 clones from ~160 brain slices/4 brains). f, Representative 3D reconstruction images of E18 WT (left) and Pcdhgc3 OE (right) clones labelled by MADM at E12. Scale bar, 100 μm. g—j, Quantification of the pairwise (g, h) and maximal (i, j) lateral and radial distances between cells in individual E18 WT and Pcdhgc3 OE clones (WT, n = 53 clones from ~106 brain slices/3 brains; Pcdhgc3 OE, n = 39 clones from ~100 brain slices/3 brains). k, l, Representative 3D reconstruction images of P7 WT (left) and Pcdhgc3 OE (right) symmetric (k) and asymmetric (l) clones labelled by MADM at E12. Coloured lines indicate the layer boundaries and coloured dots represent the cell bodies of labelled neurons. Note that the Pcdhgc3 OE clones are more laterally dispersed than the WT clones. Scale bars, 100 μm. m—p, Quantification of the pairwise (m, n) and maximal (o, p) lateral and radial distances between neurons in individual P7 WT and Pcdhgc3 OE clones (WT, n = 62 clones from ~160 brain slices/3 brains; Pcdhgc3 OE, n = 61 clones from ~156 brain slices/3 brains). Data are representative of three independent experiments. Two-tailed unpaired Student’s t-test was used for statistical test. Box plots as in Fig. 1.
Extended Data Fig. 12 PCDHγ removal does not affect the basic membrane properties of neocortical excitatory neurons.
a, Quantification of the inter-soma distance between clonally related WT and Pcdhg cKO excitatory neurons and nearby non-clonally related excitatory neurons (clonally related: WT, n = 415; Pcdhg cKO, n = 330; non-clonally related: WT, n = 157; Pcdhg cKO, n = 141). b, Summary of the rate of chemical synaptic connections between the clonally related WT and Pcdhg cKO excitatory neurons and nearby non-clonally related excitatory neurons at P14–20 (WT clone & non-clone, n = 337 from ~53 brain slices/15 brains; Pcdhg cKO clone & non-clone, n = 318 from ~52 brain slices/17 brains) and P21–35 (WT clone & non-clone, n = 235 from ~46 brain slices/16 brains; Pcdhg cKO clone & non-clone, n = 225 from ~40 brain slices/12 brains). The specific numbers of recorded pairs are shown in the bar graphs. c, Representative sample traces of the responses of excitatory neurons to somatic current injections in the WT (left) and Pcdhg cKO (right) neocortices. Scale bars: 50 mV and 200 msec. d—f, Summary of the resting membrane potential (RMP) (d, WT, n = 45; Pcdhg cKO, n = 48), input resistance (e, WT, n = 44; Pcdhg cKO, n = 48), and maximal firing rate (f, WT, n = 44; Pcdhg cKO, n = 47) of WT and Pcdhg cKO excitatory neurons. g, Quantification of the inter-soma distance between WT and Pcdhg cKO sister (left) or cousin (right) excitatory neurons (sister: WT, n = 216; Pcdhg cKO, n = 177; cousin: WT, n = 199; Pcdhg cKO, n = 153). h, i, Summary of the rate of synaptic connections between WT and Pcdhg cKO sister or cousin neuronal pairs with regard to the angular orientation of their cell bodies relative to the pia. Each symbol represents a neuronal pair. The numbers of recorded pairs and the rates of synaptic connections are shown in the graphs. The n numbers indicate neuron pairs (a, b, g–i) or neurons (d—f). Data are representative of at least 20–30 independent experiments. Two-tailed unpaired Student’s t-test (a, d–g); Two-tailed χ2 test (b, h, i). Box plots as in Fig. 1.
Extended Data Fig. 13 PCDHγC3 overexpression does not affect the basic membrane properties of neocortical excitatory neurons.
a, Quantification of the inter-soma distance between clonally related WT and Pcdhgc3 OE excitatory neurons and nearby non-clonally related excitatory neurons (clonally related: WT, n = 318; Pcdhgc3 OE, n = 324; non-clonally related: WT, n = 128; Pcdhgc3 OE, n = 123). b, Summary of the frequency of chemical synaptic connections between the clonally related WT and Pcdhgc3 OE excitatory neurons and nearby non-clonal excitatory neurons in P14–20 (WT clone & non-clone, n = 242 from ~44 brain slices/13 brains; Pcdhgc3 OE clone & non-clone, n = 273 from ~53 brain slices/18 brains) and P21–35 (WT clone & non-clone, n = 204 from ~31 brain slices/11 brains; Pcdhgc3 OE clone & non-clone, n = 204 from ~32 brain slices/11 brains). The specific numbers of recorded pairs are shown in the bar graphs. c, Representative sample traces of the responses of excitatory neurons to somatic current injections in the WT (left) and Pcdhgc3 OE (right) neocortices. Scale bars: 50 mV and 200 msec. d—f, Summary of the resting membrane potential (RMP) (d, WT, n = 45; Pcdhgc3 OE, n = 54), input resistance (e, WT, n = 44; Pcdhgc3 OE, n = 62), and maximal firing rate (f, WT, n = 42; Pcdhgc3 OE, n = 62) of WT and Pcdhgc3 OE excitatory neurons. g, Quantification of the inter-soma distance between WT and Pcdhgc3 OE sister (left) or cousin (right) excitatory neurons (sister: WT, n = 167; Pcdhgc3 OE, n = 215; cousin: WT, n = 151; Pcdhgc3 OE, n = 109). h, i, Summary of the rate of synaptic connections between WT and Pcdhgc3 OE sister or cousin neuronal pairs with regard to the angular orientation of their cell bodies relative to the pia. Each symbol represents a neuronal pair. The numbers of recorded pairs and the rates of synaptic connections are shown in the graphs. The n numbers indicate neuron pairs (a, b, g-i) or neurons (d—f). Data are representative of at least 20–30 independent experiments. Two-tailed unpaired Student’s t-test (a, d—g); Two-tailed χ2 test (b, h, i). Box plots as in Fig. 1.
Supplementary information
Supplementary Table 1
A summary of statistical tests (P values) for Extended Data Figs. 12h,i and 13h,i.
Supplementary Table 2
Mouse information used in individual experiments.
Supplementary Table 3
A summary of the numbers of brains, brain slices/sections, clones or neurons in individual experiments.
Supplementary Table 4
The scRNA-seq matrix of cPcdh isoform expression in individual clonally related neocortical excitatory neurons.
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Lv, X., Li, S., Li, J. et al. Patterned cPCDH expression regulates the fine organization of the neocortex. Nature 612, 503–511 (2022). https://doi.org/10.1038/s41586-022-05495-2
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DOI: https://doi.org/10.1038/s41586-022-05495-2
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