Bulk and Mosaic Deletions ofEgfrReveal Regionally Defined Gliogenesis in the Developing Mouse Forebrain

Abstract

The Epidermal growth factor receptor (EGFR) plays a role in cell proliferation and differentiation in healthy development and in tumors, however its requirement for brain development remains unclear. Here we used a conditional mouse allele for Egfr to examine its contributions to perinatal forebrain development at the tissue level. Subtractive bulk ventral and dorsal deletions of Egfr revealed that ventral but not dorsal telencephalic EGFR expression is critical for forebrain homeostasis and the presence of transiently EGFR-dependent glial lineages including robust depletion of oligodendrocytes that last the mouse lifetime. Sparse deletion using Mosaic Analysis with Double Markers (MADM) surprisingly reveals a regional requirement for EGFR in dorsal, but not ventral glial lineages including both astrocytes and oligodendrocytes. These findings lead to a model in which EGFR-independent ventral glial progenitors compensate for the absence of EGFR-dependent dorsal glia in the forebrain, thus masking its precise role in the bulk model.

COMBINe: Automated Detection and Classification of Neurons and Astrocytes in Tissue Cleared Mouse Brains

“…Analysis of neurons and astrocytes in the Emx:MADM:F/F forebrain hemispheres using COMBINe confirmed a phenotype analyzed in our recent publication in rostral-caudal differences in EGFR-dependent and EGFR-independent astrocyte production (Zhang et al, 2022). It is important to note that all cells in the Emx:MADM:F/F forebrain are Egfr-null regardless of their MADM color.…”

“…One dataset was obtained from MADM forebrains in which all labeled cells are wildtype ( Emx:MADM: +/+), while the two others were from MADM forebrains with heterozygous ( Emx:MADM:F /+) or homozygous ( Emx:MADM:F/F ) deletions of Egfr . The F /+ and F/F MADM genotypes were phenotypically distinct with robust disruptions in production of glia, while largely preserving the neuronal production that developmentally occurs earlier than gliogenesis (Zhang et al, 2022).…”

“…For completeness, glia labeled using Emx:MADM forebrains consisted of oligodendrocytes and astrocytes. However, the morphology and size of mature astrocytes was highly distinguishable, whereas oligodendrocytes were small and difficult to detect, and their numbers were small compared to neurons and astrocytes (Zhang et al, 2022). Thus, we trained our platform to distinguish between neurons and astrocytes.…”

“…We used a conditional mouse allele for the Epidermal growth factor receptor ( Egfr ) in combination with MADM alleles. Using established Emx1 cre line to restrict recombination to the dorsal telencephalon (cerebral cortices and the hippocampal formation), three genotypes were obtained: Emx:MADM: +/+, Emx:MADM:F /+, and Emx:MADM:F/F (Zhang et al, 2022). We have recently reported on variable effects on specification and differentiation of MADM glia in cerebral cortices of these mice, which has led to the developmental model that EGFR-dependent and EGFR-independent perinatal gliogenesis regulate forebrain development cooperatively (Zhang et al, 2022).…”