Molecular mechanisms regulating neurogenesis in the developing mouse cerebral cortex

TLDR: Differences at the transcriptional level between direct neurogenic and largely non-neurogenic radial glial cells are revealed, supporting their intrinsic lineage differences.

Abstract: Radial glial cells are a widespread non-neuronal cell type in the developing central nervous system (CNS) of all vertebrates. In the cortex, distinct subsets of radial glial cells coexist that are either multipotent or specified towards the generation of neurons or glial cells (Malatesta et al., 2000). Radial glial cells in the cerebral cortex are also the source of a second type of neurogenic progenitors, called basal progenitors. However, whether the generation of basal progenitors occurs in a stochastic manner or whether a specific lineage of radial glial cells is specified towards the generation of these progenitors has not been previously known. To identify functionally distinct lineages of cortical radial glial cells, I developed a new strategy using fluorescence-activated cell sorting (FACS) to isolate them and study their progeny. I isolated radial glial cells by FACS from a transgenic mouse line in which green fluorescent protein (GFP) expression is under the control of the human GFAP promoter. Strikingly, GFP intensity was correlated with cell fate. Selective enrichment of cells with a higher GFP intensity separated a largely non-neurogenic from a neurogenic (low GFP-intensity) subsets of radial glial cells. Notable differences on the progeny of these distinct sets of radial glia were found. The neurogenic radial glial cells subset generated neurons directly and those that are largely non-neurogenic also gave rise to a small proportion of Tbr2-positive basal progenitors that are then neurogenic. Thus, this last subset comprises an indirect neurogenic population of radial glial cells present in the developing cortex. Microarray analysis of these distinct sets of radial glial cells revealed profound differences in their gene expression. Genes related to gliogenesis, proliferation and cell-cycle regulation were expressed at higher levels in the largely non-neurogenic set of radial glia while genes related to neurogenesis, cell adhesion, neurotransmitter secretion and axon guidance were expressed mostly in the neurogenic subset. Moreover, the set of genes expressed at higher levels in the neurogenic radial glia was down-regulated at later stages (cortical radial glia at E18). Thus, this analysis reveals differences at the transcriptional level between direct neurogenic and largely non-neurogenic radial glial cells, supporting their intrinsic lineage differences. The functional analysis of a key fate determinant for neurogenesis from radial glia discovered in this transcriptome analysis will also be presented. This gene is the transcription factor AP2γ which was expressed at significantly higher levels in radial glial cells generating basal progenitors. AP2γ is restricted to the ventricular zone (VZ)/subventricular zone (SVZ) regions of the developing cerebral cortex in the entire nervous system and is also highly expressed in primate and human cortical progenitors. Its genetic deletion within the mouse cerebral cortex results in the molecular misspecification of basal progenitors with decreased levels of Tbr2 and Math3 expression, as well as their overproliferation associated with increased cell death specifically in the occipital cortex. This causes a reduction in upper layer neuron generation with intriguing functional defects in visual acuity. Gain-of-function studies also revealed the important role of AP2γ for the adequate specification and development of basal progenitors in the cerebral cortex, while apical progenitors were not affected by the loss- and gain-of-function of this transcription factor. Thus, I show for the first time the prospective isolation of distinct radial glia subtypes in the mouse developing cortex demonstrated at the molecular and functional level.