A critical role for dorsal progenitors in cortical myelination

Abstract

Much controversy regarding the anatomical sources of oligodendrocytes in the spinal cord and hindbrain has been resolved. However, the relative contribution of dorsal and ventral progenitors to myelination of the cortex is still a subject of debate. To assess the contribution of dorsal progenitors to cortical myelination, we ablated the basic helix-loop-helix transcription factor Olig2 in the developing dorsal telencephalon. In Olig2-ablated cortices, myelination is arrested at the progenitor stage. Under these conditions, ventrally derived oligodendrocytes migrate dorsally into the Olig2-ablated territory but cannot fully compensate for myelination deficits observed at postnatal stages. Thus, spatially restricted ablation of Olig2 function unmasks a contribution of dorsal progenitors to cortical myelination that is much greater than hitherto appreciated.

Figure 1.

Conditional ablation of Olig2 in cortical progenitors. A, Olig2 conditional knock-out mice are generated through homologous recombination in embryonic stem cells with the Olig2-coding region flanked by two loxP sites. B, Generation of Olig2-ablated alleles by Cre-mediated loxP site recombination preceded by Flpase-mediated FRT site recombination of the neomycin selection cassette in the targeted genome. C, Olig2 mutant alleles were confirmed by Southern blot with a 5′ external probe (wild-type and mutant fragment sizes; 5 and 7 kb, respectively). D, Olig2 mutant alleles are confirmed by PCR analysis [wt +/+, 100 bp; Olig2 mutant (c/+ and c/c), 150 bp]. E, Coronal sections of hGFAPCre forebrain at E14.5 were immunostained with a Cre antibody. Red and white arrows indicate the subventricular zone of the dorsal telencephalon and the ganglionic eminence, respectively. F, β-Galactosidase activity in the forebrain of hGFAPCre;Rosa26 mice at E14.5. Red and green arrows indicate the dorsal and ventral of forebrain, respectively. G–N, Expression of Olig2 and PdgfαR mRNAs (arrows) in the forebrain of Olig2c/-;hGFAP^Cre (Cko^G) and control mice (ctrl, Olig2c/+;hGFAP^Cre) was analyzed by in situ hybridization at E14.5 and E16.5 as indicated.

Figure 2.

Oligodendrocyte myelination deficit in the Olig2-ablated cortex. A–N, Expression of oligodendroglial markers in coronal cortical sections from control and Olig2-ablated (Cko^G) mice at neonatal P7 (A, B), perinatal P14 (C, D, G–N), and adult stage P79 (E, F) was analyzed by in situ hybridization with antisense probes Olig2, Mbp, Plp/dm20, or PdgfaR, as well as O4 immunostaining as indicated by arrows. Arrowhead in H indicates residual Olig2+ cells in the cerebral white matter. O, P, Electron micrographs of cross-sections of the corpus callosum from control (O) and Cko^G (P) animals at P21. Red and blue arrows indicate myelin sheaths and dysmyelinated axons, respectively. Q–R, Expression of Mbp mRNA in the ventral forebrain of Olig2 mutant (Cko^G) mice at P14 (Q) and adult P79 (R). Arrows and arrowheads indicate Mbp expression in the cortex and the ventral forebrain, respectively. Scale bar: P, 500 nm.

Figure 3.

Contribution of dorsal and ventral progenitors to cortical myelination in Olig2Cko;Foxg1^Cre mice. A–L, In situ hybridization labeling of Mbp (A–F) and Plp (I–L) in the forebrain of control (A–C) and Olig2Cko;Foxg1^Cre (Cko^F) (D–K) mice at P12, P18, and P28 as indicated. Arrows in D–F and I–K indicate the dorsal regions of the cortex. Blue arrows in G (dorsal) and H (ventral) indicate Olig2 labeling in the Olig2 mutant forebrain at P5. M–P, Mbp expression is shown in the ventral forebrain of control (M, N) and Olig2 mutant (O, P) mice. The square areas in M and O are shown at a larger magnification in N and P, respectively. Arrows in M and O indicate the dorsal cortical regions.

Figure 4.

Cortical myelination deficit in Olig2Cko;Emx1^Cre mice. A, β-Galactosidase activity was examined in coronal brain sections of Emx1-Cre;Rosa26 reporter mice at P14. The red arrow indicates the cortex-restricted β-galactosidase expression. The green arrowhead indicates the ventral forebrain region such as the striatum. B–F, In situ hybridization labeling of Olig2 (B, C) and Mbp (D–F) is shown in the forebrain of control and Olig2Cko;Emx1^Cre (Cko^E) mice at P14. Arrows in B–E indicate Olig2 and Mbp expression in the cortex, respectively. Arrow in D indicates the formation of Mbp+ oligodendrocytes in the ventral forebrain of Olig2 mutants. Magnifications: A, 10×; B–E, 100×; D, 40×