The Cellular and Molecular Landscapes of the Developing Human Central Nervous System

Abstract

The human CNS follows a pattern of development typical of all mammals, but certain neurodevelopmental features are highly derived. Building the human CNS requires the precise orchestration and coordination of myriad molecular and cellular processes across a staggering array of cell types and over a long period of time. Dysregulation of these processes affects the structure and function of the CNS and can lead to neurological or psychiatric disorders. Recent technological advances and increased focus on human neurodevelopment have enabled a more comprehensive characterization of the human CNS and its development in both health and disease. The aim of this review is to highlight recent advancements in our understanding of the molecular and cellular landscapes of the developing human CNS, with focus on the cerebral neocortex, and the insights these findings provide into human neural evolution, function, and dysfunction.

Keywords: brain development; developmental milestones; evolution; genomics; lateralization; neurodevelopmental disorders; regulatory elements; species differences; transcription factor.

Figure 1. Timeline of Key Human Neurodevelopmental Processes and Functional Milestones

The figure provides a summary of some key cellular processes in the developing prefrontal cortex and functional milestones. Illustrations in the top panel show the gross anatomical features of the developing and adult CNS, with prenatal brain features magnified. The second panel, which is duplicated at the bottom of the figure, provides a timeline of human development and the associated periods (designed by Kang et al., 2011), and age in postconceptional days (pcd), postconceptional weeks (pcw) and postnatal years (y). The schematic below details the approximate timing and sequence of key cellular processes and developmental milestones. Bars indicate the peak developmental period in which each feature is acquired; dotted lines indicate that feature acquisition occurs at these ages, though to a relatively minor degree; arrows indicate that the feature is present throughout life. Relevant references pertaining to each process or milestone are provided in the rightmost column: a, (Gould et al., 1990; Malik et al., 2013); b, (Bystron et al., 2006; Meyer, 2007; Workman et al., 2013); c, (Choi and Lapham, 1978; deAzevedo et al., 2003; Kang et al., 2011); d, (Kang et al., 2011; Yeung et al., 2014); e, (Huttenlocher, 1979; Kwan et al., 2012; Molliver et al., 1973; Petanjek et al., 2011); f, (Miller et al., 2012; Yakovlev and Lecours, 1967); g, (Huttenlocher, 1979; Petanjek et al., 2011); h, (Kostovic and Rakic, 1990), i, (Kwan et al., 2012; Kwan et al., 2008); j, (Aldama, 1930; Brodmann, 1909); k, (Humphrey and Hooker, 1959); l, (Eswaran et al., 2007); m, (Bellieni and Buonocore, 2012); n, (Polishuk et al., 1975); o, (Clowry, 2007; de Vries et al., 1985; Ianniruberto and Tajani, 1981; Johnson and Blasco, 1997; Van Dongen and Goudie, 1980); p, (W. H. O. Multicentre Growth Reference Study Group, 2006); q, (McManus et al., 1988; Ramsay, 1980); r, (Dosman et al., 2012; Gerber et al., 2010; Johnson and Newport, 1989); s, (Johnson and Newport, 1989); t, (Zahn-Waxler et al., 1992); u, (Meltzoff and Moore, 1977); v, (Amsterdam, 1972; Butterworth, 1990); w, (Harris, 2000); x, (Dumontheil, 2014); y, (Rajan et al., 2014); z, (Catts et al., 2013; Heaton et al., 1993).

Figure 2. Psychiatric and Neurological disorders Have Discrete Ages of Onset

The bars indicate the age range that each disorder commonly effects, with less frequent ages of diagnosis denoted as dotted lines. The light gray shading corresponds to adolescence. Note that the age of diagnosis is highly variable between brain disorders. Many psychiatric disorders emerge in adolescence and early adulthood. This variability is indicative of dysregulation of tightly controlled developmental processes and highlights the necessity of defining the spatiotemporal molecular and cellular processes in healthy and diseased human CNS. Based in part on data from (Kessler et al., 2007; Lee et al., 2014)

Figure 3. Global Spatio-Temporal Dynamics of the Human Brain Transcriptome

(A) Multidimensional scaling (MDS) plot of global transcriptional differences across regions and time. The most pronounced differences (approximately two-thirds) occur during prenatal development (periods 1 to 7). By contrast, over four decades of adulthood (periods 13–15), less than 1% of genes are differentially expressed. Each dot represents a sample and is colored according to period as defined by Kang et al., 2011; the dotted line indicates birth. (B) MDS plot in (A) colored by brain regions. The most prominent differences were between cerebellar cortex (CBC) and forebrain regions. Abbreviations: NCX: neocortex; HIP: hippocampus; AMY: amygdala; STR: striatum; MD: mediodorsal nucleus of thalamus. (C) Weighted gene co-expression analysis identifies modules of co-expressed genes associated with distinct spatio-temporal expression patterns and biological processes in human brain development. Module M20 from Kang et al., 2011 shown here comprises genes that are downregulated simultaneously in different regions, with the exception of postnatal CBC, as the brain matures. M20 is enriched for genes encoding transcriptions factors involved in neurogenesis and pan-neuronal differentiation. (D) Module M2 shown here comprises genes that are upregulated simultaneously, peaking first in childhood, in all regions, with the exception of postnatal CBC, as the brain matures. This module was enriched for genes associated with neuronal maturation processes like synaptic transmission, ion-transport, and calcium signaling. C and D were adapted with permission from Kang et al., 2011.

Figure 4. Divergent Organization and Transcritonal Profiles of Human and Mouse Neocortical Stem/Progentior Cells

(A) Schematic representations of the mid-fetal human (left; 20 pcw) and mouse (right; 18.5 pcd) brain and neocortex (blue). The smaller image of the mouse brain indicates the approximate size difference between human and mouse brains and neocrtex (blue) at approximately equivalent age. The zoomed image of the mouse brain highlights differences in brain size and the position along the rostral caudal axis (dotted line) of the illustrations in (B). Adapted with permission from Gulden and Sestan, 2014. (B) A schematic of the cellular composition of the human (left) and mouse (right) fetal forebrain wall detailing differences in neurogenic processes and stem/progenitor types between species. In mouse, apical or inner RG (aRG and iRG) cells (dark green) divide asymetrically to produce both a daughter intermediate progenitor cell (IPC) that subsequently divides symmetrically in the overlying SVZ (light green) and an excitatory projection neuron (PN) that migrates along RG fibers to the CP (blue). In human, the SVZ is greatly expanded and contains a large population of asymetrically dividing basal or outer RG (bRG and oRG) cells, which enables the production of greater numbers of neurons. (C) A plot of the top 5,000 genes found to be enriched in RG cells by Lui and colleagues (Lui et al., 2014) are plotted. Ordinate values reflect the specificity of a gene within the neocortex to human RG cells while values along the abscissa represent expression differences between human and mouse RG. Red lines and green lines denote one- and two- standard deviations from the mean differential expression score. The vast majority of RG-enriched genes are similarly expressed in mouse and human. However, a considerable number of genes with high differential expression between species are observed, including a number of genes with high expression in human but not mouse RG cells. Among these genes, PDGFD was shown to increase the proliferative capacity of the neural progenitors. Adapted with permission from Figure 2d and Expanded Data Table 3 in Lui et al., 2014. (D) A heatmap showing differential gene expression within discrete zones (layers) of the fetal neocortical wall at 21 pcw. These differences underlie the distinct cell types, cellular processes, and stages of maturation in each zone. Abbreviations: SG, subpial granular layer; MZ, marginal zone; CPo and CPi, outer and inner cortical plate; SP, subplate zone; IZ, intermediate zone; SZo and SZi, outer and inner subventricular zones; VZ, ventricular zone. (E) A Nissl stain on the left delineates each fetal neocortical zone. Notable genes enriched in each are shown by in situ hybridization in the panels to the right, confirming findings of zone enriched expression identified by microarrays. The red arrows mark bands of enriched expression of calbindin2 (CALB2) and zic family member 1 (ZIC1) in the MZ and SG, respectively. Images in D and E were adapted with permission from Miller et al., 2014.

Figure 5. Transcriptional Differences Among Human Neocortical Areas are Temporally Regulated

(A) Unsupervised hierarchical clustering of the 11 neocortical regions/areas profiled by Pletikos et al., 2014, based on the transcriptome of each area from the period of fetal development throughout adulthood, showing relative transcriptional differences. Abbreviations: OFC, orbital prefrontal cortex; DFC, dorsal prefrontal cortex; VFC, ventral prefrontal cortex; MFC, medial prefrontal cortex; M1C, primary motor cortex; S1C, primary somatosensory cortex; IPC, posterior inferior parietal cortex; A1C, primary auditory cortex; STC, superior temporal cortex; ITC, inferior temporal cortex; V1C, primary visual cortex. (B) Boxplots of subsampling permutations show the number of expressed (blue) and differentially expressed (red) genes among neocortical areas across fetal development (periods 3–7), infancy (periods 8 and 9), childhood (periods 10 and 11), adolescence (period 12), and adulthood (periods 13–15). Note that the total number of genes expressed decreases over development. However, the number of differentially expressed genes observed over development exhibits a temporal hourglass pattern with the highest number in fetal development, a marked decline in infancy and childhood, and an increase in adolescence through adulthood. (C) A 3D heatmap showing the number (post hoc Tukey test) of genes with differential expression between any two neocortical areas, demonstrating that the hourglass pattern of inter-areal differential expression persists in all neocortical areas, but is most prominent in MFC and V1C. (D–G) Examples of gene co-expression modules (M) with a temporally regulated gradient-like expression pattern in the fetal neocortex (see circles with colored scale overlying each area). Modules M91, M100, M2, and M118 show frontal (D), medial fronto-occipital (E), posterior perisylvian (F) and middle perisylvian (G) enriched expression, respectively. Fp, frontal pole; Tp, temporal pole; Op, occipital pole. (H–L) Radar charts showing shared and divergent expression gradients in human (blue) versus Rhesus macaque (green) fetal neocortex of specific intramodular hub genes (CLMP [M91], C13ORF38 [M80], WNT7B [M6], and NR2F2 [M13]). A–K were adapted with permission from Pletikos, et al. 2014.

Figure 6. Shared and Divergent Expression Patterns in the Fetal Human and Mouse Neocortex

(A) CBLN2 is enriched throughout the CP of the mid-fetal human prefrontal cortex (PFC), whereas in mouse, at a comparable period of development, expression is enriched in the upper layers of frontal cortex. (B) NPY has highly divergent expression along the rostral caudal axis in human versus mouse mid-fetal neocortex. In human, but not mouse, NPY is enriched in the mid-fetal occipito-temporal CP. The expression of NPY in sparsely distributed interneurons of the CP and SP zone is conserved. Human data were adapted with permission from Johnson et al., 2009. Mouse in situ hybridization images were obtained from the Allen Developing Mouse Brain Atlas (http://developingmouse.brain-map.org; (Thompson et al., 2014). Abbreviations are the same as in Figure 5.

Figure 7. Evolutionary Changes in a Cis-Regulatory Element Active in Neocortical Stem/Progenitor Cells

(A) Schematic demonstrating expansion of transcriptional start sites over evolution in noncoding exon 1 of the Gpr56 gene (2 in zebra fish, 5 in mouse, and 17 in human), a gene that promotes proliferation of neural progenitor cells. (B) A different expression pattern was observed after driving expression of lacZ (blue staining) with either the mouse or human variant of the promoter of one of the transcription start sites (E1m). The mouse element was able to recapitulate the full extent of mouse Gpr56 expression, but expression driven from the human element was restricted to a rostro-lateral band. (C) Consistent with a role in driving rostra-lateral expression specifically, a 15 base pair deletion in the E1m element eliminated expression of GFP from the rostro-lateral forebrain in transgenic mice harboring an allele of Gfp driven by the E1m element. (D) Deletion of this element was observed in patients with perisylvian polymicrogyria in accordance with a role in driving expression of GPR56 in the lateral neocortex (light green). In these patients, this mutation likely led to malformations of the perisylvian neocortex important for language among other functions. A–D were adapted with permission from (Bae et al., 2014).