Article Figures & Data
Figures
- Figure 1.
EGFP-expressing cells located in the molecular layers of the CXCR4-EGFP mouse are not immunoreactive for astrocytic or microglial markers. A, Low-magnification images of the hippocampal dentate gyrus of the CXCR4-EGFP mouse. Left, EGFP expression (green). Middle, GFAP immunoreactivity (red). Right, Previous images superimposed with additional DAPI counterstaining (blue). SLM, stratum lacunosum-moleculare; ML, molecular layer; GCL, granule cell layer; Hil, hilus; SP, stratum pyramidale. B, Higher-magnification view of the region in proximity to the HF (dotted line); left, middle, and right as in A. Notice the total lack of co-localization between the EGFP and GFAP signals. Filled arrowheads indicate EGFP-expressing somata, whereas empty arrowheads mark the position of GFPA-labeled cells. C, D, Identical organization as in A, B, respectively, but immunoreactivity is shown for the microglial marker IBA1. Notice that EGFP-positive cells are not IBA1-immunoreactive.
- Figure 2.
EGFP expression in the neurogenic niche of the CXCR4-EGFP mouse. A, Low-magnification images of the hippocampal dentate gyrus showing EGFP expression (left, green), DCX immunoreactivity (middle, red), and the two signals superimposed with DAPI (blue) counterstain (right). SLM, stratum lacunosum-moleculare; ML, molecular layer; GCL, granule cell layer; Hil, hilus; SP, stratum pyramidale. B, Higher magnification reveals EGFP expression (green, left) in DCX-negative (blue, middle left), GFAP-positive (red, middle right) cells with the soma in the GCL and processes extending to the inner molecular layer (IML) as well as in DCX-positive, GFAP-negative cells, in the subgranular zone (SGZ). The right panel shows all the previous signals superimposed. Dotted lines mark the border between different layers. The boxes in the right panel are shown in more detail in C, D, for EGFP expression (left, green), GFAP/DCX immunoreactivity (middle, red and blue, respectively), and with the signal superimposed and additional DAPI counterstaining (white, right). Filled arrowheads mark the position of the soma and processes of the cells of interest for reference. Asterisk marks EGFP-negative, DCX-positive cell.
- Figure 3.
Overview of the distribution of EGFP-positive and GAD67-immunoreactive neurons in the CXCR4-EGFP mouse (P14). A, left, Low-magnification fluorescence image of the hippocampus. Notice the numerous EGFP-labeled neurons localized in the molecular layers on either side of the HF. Middle, Immunoreactivity for GAD67 in the same section. Notice the more scattered distribution of labeled cells within all hippocampal layers. Right, Overlap of the previous images with nuclear DAPI staining. Specific layers and areas are indicated as follows: SO, stratum oriens; SP, stratum pyramidale; SL, stratum lucidum; SLM, stratum lacunosum-moleculare; HF, hippocampal fissure; ML, molecular layer; GCL, granule cell layer; Hil, hilus; CA3, cornu ammonis 3; CA1, cornu ammonis 1; Sub, subiculum. Dashed lines indicate the borders between the CA3 and CA1 subfields and of the subiculum. Hippocampal molecular layers (MLs) are highlighted in red. Circles indicate the position of EGFP-positive cells (green/white) and GAD67-positive cells (blue/white). B, Same experimental setup as in A, but observed at higher magnification. The dotted line indicates the HF. Notice the lack of overlap between the EGFP and GAD67 signals. Arrowheads indicate EGFP-positive Cajal–Retzius cells (green) and GAD67-positive interneuron (blue). Insets show the neurons indicated by red arrowheads at higher magnification.
- Figure 4.
Spatiotemporal distribution of EGFP-labeled Cajal–Retzius cells versus GAD67-immunoreactive interneurons of the molecular layers. A, Summary plot showing the normalized location of Cajal–Retzius cells and GABAergic interneurons of the hippocampal molecular layers. Data from n = 3 animals (P14), n = 690 Cajal–Retzius cells and n = 713 interneurons. Schematic lines indicate the granule cell layer, HF, and borders of the infra-pyramidal blade of the dentate gyrus and subicular complex. B, left, Fractional density plot of Cajal–Retzius cells (green) and 5% maximum density area for interneurons (dotted red contour). Middle, Similar to the left panel, but with fractional density of interneurons (blue) and the 5% maximum density area for Cajal–Retzius cells limited by the dotted red line. Notice the more restricted area occupied by EGFP-labeled Cajal–Retzius cells compared to GAD67-immunoreactive interneurons. Right, Left and middle panels superimposed. C, Distances of Cajal–Retzius and GABAergic interneurons from the HF. Left, Microscopic field with measurements of the shortest distances between the soma of Cajal–Retzius cells and interneurons (green and blue lines and circles, respectively) from the HF (white dotted line). Middle, Summary scatterplot for measurements from Cajal–Retzius cells (CR, green dots) and GABAergic interneurons (IN, blue dots). HF, hippocampal fissure. The abscissa indicates the relative position of the studied cell from the hippocampal pole (0) to the end of the HF (1). ML/DG, molecular layer of the dentate gyrus; SLM, stratum lacunosum-moleculare. Right (left panel), Relative distributions of the distances to the HF for the two neuronal populations (green, Cajal–Retzius cells; blue, interneurons). Notice the clear grouping of Cajal–Retzius cells in the vicinity of the HF compared to a more homogeneous distribution in the case of interneurons. Right (right panel), Summary box charts indicating the values of the distances from the fissure calculated separately for Cajal–Retzius cells and interneurons of stratum lacunosum-moleculare (top) and of the molecular layer of the dentate gyrus (bottom). Notice, in both cases, the shorter distances of Cajal–Retzius cells. D, Cajal–Retzius cell are positioned in a non-random fashion. Left, Measurement of nearest neighbor distances between Cajal–Retzius cells and interneurons. The image shows examples for a Cajal–Retzius cell (green dot) and an interneuron (blue dot). The nearest neighbor (Cajal–Retzius cell to Cajal–Retzius cell and interneuron to interneuron) is identified in both cases by a white dot. The distance to the nearest neighbor is marked by the thick tapered lines, whereas distances to other cells are shown by faint dotted lines (green for Cajal–Retzius cells and blue for interneurons). The white dashed line indicates the HF. Middle, Summary graph for Cajal–Retzius cells (green dots) and interneurons (blue dots). The abscissa indicates the relative position from the hippocampal pole (0) to the end of the CA1 region (1). Right (left panel), Distribution histograms of the data shown in the scatterplot. Right (right panel), Summary box charts for nearest neighbor distances between Cajal–Retzius cells (green box), interneurons (blue box). Notice the much shorter distances for Cajal–Retzius cells. E, left, Comparison of the linear densities of Cajal–Retzius cells and interneurons at different postnatal stages (P14, top panel; P60, bottom panel). Cajal–Retzius cells and interneurons are counted in green and blue, respectively. Middle, Summary boxplot for linear densities measured at P7, P14, P30, and P60. Notice the larger density of Cajal–Retzius cells progressively approaching the values measured for interneurons with brain maturation. Boxes have been slightly shifted to the left and right of the actual time points (P7, P14, P30, and P60) to avoid their superimposition. The connecting dotted lines are aligned to the actual time points. Right, Summary graph of the ratio of the densities of Cajal–Retzius cells over interneurons. Notice that at early developmental stages the density of Cajal–Retzius cells is more than double the one of the entire population of GAD67-positive cells in the same molecular layers. n.s., p > 0.05, ***p < 0.001.
- Figure 5.
Comparison of biometric morphologic parameters between Cajal–Retzius cells and interneurons. A, Regions of interest (soma and nucleus: white dotted lines) and markers in individual Cajal–Retzius cells (three leftmost panels: EGFP, DAPI, and superimposed) and B, interneurons (three rightmost panels: GAD67, DAPI, and superimposed). C–H, Comparisons of the distributions for the measured parameters (top graphs) in Cajal–Retzius cells and interneurons (green and blue, respectively) and their summary box charts (bottom insets). Notice the overall smaller size of Cajal–Retzius cells compared to interneurons; (C) nuclear (left) and somatic (right) perimeter, (D) nuclear (left) and somatic (right) area, (E) nuclear (left) and somatic (right) minimal Feret diameter (Fmin), (F) nuclear (left) and somatic (right) maximal Feret diameter (Fmax), (G) nuclear (left) and somatic (right) aspect ratios, (H) nuclear/somatic areas ratio, (I) principal component analysis of all the measured parameters. Notice the clear separation in clusters between Cajal–Retzius cells (green circles) and interneurons (blue circles). *p < 0.05, ***p < 0.001.
- Figure 6.
Reelin expression in the hippocampal molecular layers is not specific for Cajal–Retzius cells. A, Low-magnification images of EGFP fluorescence (top left, green), reelin immunoreactivity (top right, red) and GAD67 immunoreactivity (bottom left, blue), and of all the signals superimposed (bottom right). Notice the presence of reelin-immunoreactive cells in all hippocampal layers. B, Higher-magnification view reveals the presence of reelin both in EGFP-positive Cajal–Retzius cells and GAD67-expressing interneurons. EGFP expression (top left), reelin immunoreactivity (top right), GAD67 immunoreactivity (bottom left), and merged signals with DAPI (white) nuclear counterstaining (bottom right). Solid arrowheads indicate Cajal–Retzius cells; outlined arrowheads indicate GAD67-positive interneurons. C, left, Summary plot of the relative fraction of reelin-expressing neurons that are positive for EGFP, GAD67, or neither EGFP nor GAD67 (nnor). Right, Regional subdivision of the data in the left panel for different regions of interest. 1, CA3 subfield; 2, CA1 subfield; 3, subiculum. n.s., p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001.
- Figure 7.
The nuclear transcription factor COUP-TFII is expressed both in Cajal–Retzius cells and interneurons of the molecular layers. A, Low-magnification overview of the hippocampus tested for EGFP expression (top left, green), COUP-TFII immunoreactivity (top right, red), GAD67 immunoreactivity (bottom right, blue), and of all the signals superimposed (bottom right). B, Cells of the molecular layers identified by their EGFP expression as Cajal–Retzius cells (top left, green) or GAD67-expressing interneurons (bottom left, blue) express COUP-TFII (top right, red). Bottom right, All signals superimposed with DAPI (white) nuclear counterstaining. Solid arrowheads indicate Cajal–Retzius cells; outlined arrowheads indicate GAD67-positive interneurons. C, Left, Overall graph of the relative fraction of COUP-TFII-immunopositive cells that express EFGP, GAD67, or neither EGFP nor GAD67 (nnor). Right, Regional subdivision of the data in the left panel for different regions of interest. 1, CA3 subfield; 2, CA1 subfield; 3, subiculum. Notice that in any case, the largest fraction of COUP-TFII labeled neurons identifies Cajal–Retzius cells.
- Figure 8.
Immunoreactivity for the calcium binding protein calretinin in the molecular layers of the hippocampus is predominantly due to its expression by Cajal–Retzius cells. A, Low-magnification overview of the hippocampal molecular layers, tested for EGFP fluorescence (top left, green), calretinin immunoreactivity (top right, red), GAD67 immunoreactivity (bottom left, blue), and of the merged signals (bottom right). B, High magnification of molecular layer neurons identified as Cajal–Retzius cells by their EGFP expression (top left, green) or GABAergic interneurons by GAD67 immunoreactivity (bottom left, blue). Cajal–Retzius cells and GAD67-positive interneurons show calretinin immunoreactivity (top right, red). All signals superimposed with DAPI (white) nuclear counterstaining (bottom right). Solid arrowheads indicate Cajal–Retzius cells, outlined arrowheads indicate GAD67-positive interneurons. C, left, Summary plot of the relative fraction of calretinin-positive cells that express EGFP, GAD67, or neither EGFP nor GAD67 (nnor). Right, Regional subdivision of the data in the left panel for different regions of interest. 1, CA3 subfield; 2, CA1 subfield; 3, subiculum. Notice that the vast majority of calretinin-positive cells express EGFP but not GAD67.
- Figure 9.
The tumor protein p73 is specifically expressed by Cajal–Retzius cells. A, Low-magnification overview of the hippocampal molecular layers, tested for EGFP expression (top left, green), p73 immunoreactivity (top right, red), GAD67 immunoreactivity (bottom left, blue), and of all the merged signals (bottom right). B, High magnification of molecular layer neurons identified as Cajal–Retzius cells by their EGFP expression (top left, green) or GABAergic interneurons by GAD67 immunoreactivity (bottom left, blue). Only Cajal–Retzius cells show p73 immunoreactivity (top right, red). Bottom right, All signals superimposed with DAPI (white) nuclear counterstaining. Solid arrowheads indicate Cajal–Retzius cells, outlined arrowheads indicate GAD67-positive interneurons. C, left, Summary plot of the relative fraction of p73-positive cells that express EGFP, GAD67, or neither EGFP nor GAD67 (nnor). Right, Regional subdivision of the data in the left panel for different regions of interest. 1, CA3 subfield; 2, CA1 subfield; 3, subiculum. Notice that p73 was never found in GAD67-positive cells, irrespective of the region examined.
- Figure 10.
High specificity of p73 staining for hippocampal Cajal–Retzius cells in the hippocampal formation. A, Low-magnification images of the hippocampus of the CXCR4-EGFP mouse with NeuN staining (top left, blue), EGFP-expression (middle left, green), and p73 immunoreactivity (bottom left, red). Please refer to the online figure to see p73 staining in this panel as it is difficult to see in the non-enlarged printed version. Right, All channels merged with boxes (1, 2, 3, 4, 5, 6, and 7) indicating regions of interest (ROIs) enlarged in the insets with labeled with the same numbers. NC, neocortex; EC, entorhinal cortex; paraS, parasubiculum; preS, presubiculum; Sub, subiculum; CA1, cornu ammonis subfield 1; CA2, cornu ammonis subfield 2; CA3, cornu ammonis subfield 3; DG, dentate gyrus; GCL, granule cell layer; ML, molecular layer; Hil, hilus; V, ventricle. Insets 1–6 show corresponding ROI at higher magnifications for EGFP expression (top), p73 immunoreactivity (middle), and signals superimposed including NeuN (bottom). Notice that p73 staining is specifically confined to EGFP-expressing cells of the molecular layers (SLM, stratum lacunosum moleculare; OML, outer molecular layer) and is absent in the other layers and regions considered. GCL, granule cell layer; IML, inner molecular layer; SP, stratum pyramidale; SR, stratum radiatum. Inset 7, EGFP positivity (left, green) and p73 labeling (middle, red) are both present in ependymal cells surrounding the ventricle (right, signals superimposed plus DAPI counterstain). Filled arrowheads indicate EGFP-positive cilia.
- Figure 11.
Post hoc anatomic reconstruction and basic electrophysiological properties of Cajal–Retzius cells, GABAergic interneurons, and semilunar cells. A, Neurolucida reconstructions of biocytin-filled neurons (somato-dendritic domains in red and axons in green, Cajal–Retzius cell; blue, interneuron; and orange, semilunar granule cell), respectively. Notice the distinct morphologies of the three cell types. B, top panel, Oblique contrast illumination image of a hippocampal Cajal–Retzius cell soma in an acute brain slice (left) and its respective firing pattern (right, current step: +40 pA/–60 pA, 1 s). Middle panel, Same, but for a GABAergic interneuron (current step: +280 pA/–100 pA, 1 s). Bottom panel, Dentate gyrus semilunar cell (current step: +120 pA/–100 pA, 1 s). C, Summary box plots of the basic electrical properties of Cajal–Retzius cells (CR), GABAergic interneurons (IN), and semilunar cells (SL). Left panel, Membrane input resistance (Rm). Middle panel, Membrane potential (Vm). Right panel, EPSC/IPSC ratio. Notice the null ratio of Cajal–Retzius cells as a result of the complete absence of EPSCs. D, Spontaneous synaptic currents. Top panel, Cajal–Retzius cells. The inset to the right shows the recording at higher temporal magnification of (time window marked by red arrowheads). Notice the absence of events at –60 mV. Middle and bottom panels, Same experiments, but for a GABAergic interneuron and a dentate gyrus semilunar cell, respectively. Notice the presence of synaptic events both at +10 mV (IPSCs) and at –60 mV (EPSCs).
- Figure 12.
Dendritic and axonal morphology distinguish Cajal–Retzius cells from GABAergic interneurons and semilunar cells. A, left panel, Micrograph of a typical Cajal–Retzius cell revealed by a DAB reaction. Dashed line represents the HF. Notice the typical bipolar morphology, with a single dendrite emerging from one pole (left) and the axon from the opposite pole (right) of the soma. Right panel: examples of three Cajal–Retzius cells, reconstructed with a Neurolucida system. Somato-dendritic domains shown in red, axons in green. B, Fractional density plot of dendrites (left panel, red), axons (middle panel, green), and axo-dendritic overlay (right, high-contrast LUT). Cells (n = 16) were aligned at soma (white circle). Grid size is 25 × 25 µm2. C, Same experiments as in A but for a GABAergic interneuron. Somato-dendritic domain shown in red, axons shown in blue. Notice the multipolar dendritic configuration and heterogeneity of GABAergic interneurons. The axonal arborization mainly surrounds the somato dendritic domain. D, Same experiment as in B but for (n = 17) GABAergic interneurons. E, Same experiments as in A but for a semilunar cell. Somato-dendritic domain shown in red, axons shown in blue. Notice the semilunar-like dendritic configuration directed toward the HF and stereotypical appearance of these neurons. Axonal arborizations show few segments within the molecular layers and penetrate the granule cell layer (gray) to project to CA3. F, Same experiment as in B but for (n = 6) semilunar cells. G, Sholl analysis of the dendritic domains of all the cell types (same n as in B, D, and F, respectively) with a 25 µm circular increment (from soma). Lines show the means at every increment, transparent area illustrate the SD. H, Box plots of the total dendritic length (left), number of dendrites emerging from cell (middle), and number dendrite endings (right). Notice the overall sparse dendritic configuration of Cajal–Retzius cells.
Tables
- Table 1.
Developmental profile of the relative proportions of EGFP-expressing and GAD67-expressing neurons of the hippocampal molecular layers in the CXCR4-EGFP mouse
Measurement Age P7 P14 P30 P60 EGFP-positive 1541 1381 1079 870 GAD67-positive 652 1114 870 1041 EGFP/GAD67-positive 1 1 0 0 Notice that EGFP- and GAD67-labeled neurons remain two distinct populations throughout P7–P60. Results obtained from three mice per age.
- Table 2.
Summary table of the distances and densities of Cajal–Retzius cells and interneurons measured in mice of different developmental stages (P7–P60, n = 3 animals for every age)
Age Measurement Unit P7 P14 P30 P60 Cell type Distance to HF
ML - dentate gyrusµm 20.29 ± 1.02 23.45 ± 0.88 21.46 ± 1.06 26.00 ± 1.03 CR 74.82 ± 4.71 72.54 ± 2.99 72.52 ± 2.77 87.06 ± 3.3 IN 27% 32% 30% 30% CR/IN ratio 219/120 641/300 522/223 435/278 #CR/#IN Distance to HF
SLM - cornu ammonisµm 33.79 ± 1.89 34.49 ± 1.48 23.17 ± 1.26 26.43 ± 1.14 CR 113.06 ± 5.49 129.71 ± 3.02 130.04 ± 3.67 112.26 ± 3.33 IN 30% 27% 18% 24% CR/IN ratio 245/223 514/633 393/492 337/659 #CR/#IN Nearest neighbor
distanceµm 15.33 ± 0.46 18.71 ± 0.39 19.81 ± 0.43 22.80 ± 0.62 CR 38.81 ± 1.24 39.61 ± 0.62 39.61 ± 0.68 39.99 ± 0.79 IN 40% 47% 50% 57% CR/IN ratio 464/343 1155/933 945/715 772/937 #CR/#IN Density at HF Cells/100 µm 17.33 ± 0.63 12.27 ± 1.02 8.64 ± 0.79 6.74 ± 0.53 CR 7.62 ± 0.33 9.89 ± 0.5 + 6 7.34 ± 0.68 8.05 ± 0.55 IN 227% 124% 118% 84% CR/IN ratio 770/326 1381/1114 1079/870 870/1041 #CR/#IN Compare to results illustrated in Figure 2. CR: EGFP-identified Cajal–Retzius cells, in: GAD67-immunoreactive interneurons. #CR and #IN indicate the number of the different cell types used for the analysis.
- Table 3.
Summary table of the values for the various morphometric parameters measured as in Figure 3 for mice of different ages (P7–P60, n = 3 animals for every age)
Age Measurement Unit P7 P14 P30 P60 Cell type Nucleus Soma Nucleus Soma Nucleus Soma Nucleus Soma Perimeter µm 28.77 ± 1.59 40.25 ± 1.62 25.45 ± 0.49 34.47 ± 0.83 24.12 ± 0.56 32.54 ± 0.71 23.92 ± 0.41 31.72 ± 0.79 CR 36.06 ± 1.16 49.74 ± 1.55 34.83 ± 0.33 48.53 ± 0.83 33.64 ± 0.91 47.74 ± 0.54 31.12 ± 0.65 42.56 ± 1.03 IN 79.8% 80.9% 73.1% 71.0% 71.7% 68.2% 76.9% 76.9% CR/IN ratio Area µm² 53.08 ± 3.05 85.08 ± 5.84 46.32 ± 1.43 71.58 ± 2.38 42.12 ± 2.18 68.36 ± 3.01 38.31 ± 1.08 55.26 ± 1.93 CR 94.74 ± 4.38 157.8 ± 8.71 93.04 ± 1.87 159.37 ± 4.64 94.03 ± 1.99 121.32 ± 2.84 70.63 ± 2.59 116.42 ± 4.11 IN 56.0% 53.9% 49.8% 44.9% 44.8% 56.4% 54.2% 47.5% CR/IN ratio Feret max µm 10.31 ± 0.22 16.91 ± 0.67 9.31 ± 0.18 13.93 ± 0.42 10.01 ± 0.32 14.27 ± 0.55 9.41 ± 0.27 13.21 ± 0.46 CR 13.08 ± 0.42 18.5 ± 0.64 12.26 ± 0.12 18.05 ± 0.38 12.08 ± 0.37 16.21 ± 0.41 11.54 ± 0.35 16.54 ± 0.57 IN 78.8% 91.4% 75.9% 77.2% 82.9% 88.0% 81.5% 79.9% CR/IN ratio Feret min µm 6.55 ± 0.31 7.32 ± 0.34 6.42 ± 0.14 7.21 ± 0.13 6.21 ± 0.12 7.24 ± 0.21 5.31 ± 0.17 6.07 ± 0.19 CR 9.34 ± 0.26 12.03 ± 0.45 9.78 ± 0.15 12.19 ± 0.25 9.11 ± 0.18 11.41 ± 0.22 10.03 ± 0.25 7.99 ± 0.19 IN 78.8% 91.4% 75.9% 77.2% 82.9% 88.0% 81.5% 79.9% CR/IN ratio Aspect ratio - 1.65 ± 0.09 2.36 ± 0.08 1.47 ± 0.04 1.95 ± 0.06 1.81 ± 0.09 2.08 ± 0.04 1.83 ± 0.1 2.25 ± 0.12 CR 1.41 ± 0.04 1.57 ± 0.07 1.26 ± 0.02 1.5 ± 0.04 1.32 ± 0.02 1.55 ± 0.07 1.46 ± 0.06 1.67 ± 0.07 IN 117.0% 150.3% 116.7% 130.0% 137.1% 134.2% 125.3% 134.7% CR/IN ratio n 23/20 31/34 32/31 25/25 #CR/#IN CR: EGFP-identified Cajal–Retzius cells, in: GAD67-immunoreactive interneurons. #CR and #IN indicate the number of the different cell types used for the analysis.
Movies
- Movie 1.
Developmental profile of Cajal–Retzius cells identified by p73 immunoreactivity and reelin-positive GABAergic interneurons (p73-negative) in a model of a horizontal brain slice. Density plot (scaled to minimal/maximal density for each time point) was calculated based on measurements of P7, P14, P30, and P60 animals (n = 3 each, n = 6 per animal). Data points were calculated with linear extrapolation for P2–P7 and with linear interpolation for P7–P14, P14–P30, and P30–P60. Notice the contrast between the fading Cajal–Retzius cells (indicated by red arrowhead) and the persistent GABAergic interneurons in the neocortex (NC).
Extended Data Figure 4-1
Nearest neighbor distances of Cajal–Retzius cells and GABAergic interneurons compared to a simulated a random equal distribution (R.E.D.). A, Plot of Cajal–Retzius cells (n = 461), interneurons (n = 511), and R.E.D. cells (n = 500), in a normalized model of the hippocampus. B, Plot of the linear cell density of Cajal–Retzius cells (n = 45 slices) and interneurons (n = 33 slices) against their average nearest neighbor distance (per slice). The R.E.D. perfectly follows an allometric fitted function (red line, y = 111.83×−0.54; R 2 = 1), based on a simulation with n = 5× values (25 iterations each). Notice that the GABAergic interneurons are closely associated with the function of the R.E.D. suggesting their equal distribution in the hippocampal molecular layers. The distribution of Cajal–Retzius cells is shifted downwards, suggesting a clustered distribution. Selection of data points shown in A have a black outline. Download Figure 4-1, TIF file.
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