Figure 1.
Schematic and expression profile of Plp-Cre transgene.
A. Schematic representation of the Plp-Cre transgene construct. The mouse Plp promoter is used to drive expression of the Cre cDNA. The initiating ATG within exon 1 has been mutated to prevent initiation of translation within the promoter region. B. Summary of Plp-Cre transgenic mice generated. C. RT-PCR analysis of the tissue expression profile of the Plp-Cre transgene for both founder lines. At P28, both lines adhered to a strictly CNS-specific expression pattern, with transcripts detected in the spinal cord, cerebellum and cortex. At P4, the transgene transcript in line F633 was detected solely in CNS tissues, whereas line F627 displayed a more widespread expression pattern, with transcripts also detected in kidney, liver and TA muscle. SC = spinal cord, Cer = cerebellum, Cor = cortex, Spln = spleen, Kid = kidney, Liv = liver, TA = tibialis anterior muscle. Actin mRNA amplification was used as control.
Figure 2.
Cre-recombinase is predominantly expressed in oligodendrocytes of P28 Plp-Cre mouse spinal cord.
A–I. Ventral white matter regions were double-stained with antibodies specific to CC-1 as a marker for mature oligodendrocytes (green), Cre (red) and counterstained with DAPI. Cre was not detected in WT spinal cord sections (B). Sections from transgenic mice of both F633 (D–F) and F627 (G–I) lines exhibited a similar pattern of Cre expression. High levels of Cre recombinase protein was detected in the CC1-positive oligodendrocytes. J. Ventral white matter region stained for GFAP (green) as a marker for astrocytes, Cre (red) and counterstained with DAPI. A small number of Cre-positive cells co-labeled with GFAP (arrow). K. Ventral white matter region stained for Ng2 (green) as a marker for OPCs, Cre (red) and counterstained with DAPI. A small number of Cre-positive cells co-labeled with Ng2 (arrow). L. Ventral grey matter region stained for NeuN (green) as a neuronal marker, Cre (red) and counterstained with DAPI. There were no NeuN positive cells that co-labeled with Cre. Scale bar = 20 µm.
Figure 3.
Cre-recombinase is predominantly expressed in oligodendrocytes of P28 Plp-Cre mouse cerebellum.
A–I. Cerebellar sections were double-stained with antibodies specific to CC-1 as a marker for mature oligodendrocytes (green), Cre (red) and counterstained with DAPI. Cre was not detected in WT cerebellar sections (B). Sections from transgenic mice of both F633 (D–F) and F627 (G–H) lines exhibited a similar pattern of Cre expression. High levels of Cre recombinase protein was detected in the CC1-positive oligodendrocytes. J. Cerebellar section stained for GFAP (green) as a marker for astrocytes, Cre (red) and counterstained with DAPI. A small number of weakly stained Cre-positive cells co-labeled with GFAP (arrow). K. Representative example within the molecular layer of cerebellar section stained for Ng2 (green) as a marker for OPCs, Cre (red) and counterstained with DAPI. A small number of weakly stained Cre-positive cells co-labeled with Ng2 (arrow). L. Representative example within granular layer of cerebellar section stained for NeuN (green) as a neuronal marker, Cre (red) and counterstained with DAPI. There were no NeuN positive cells co-labeled with Cre. GL = granular layer, WMT = white matter tract. Scale bar = 20 µm.
Table 1.
Distribution of Cre-positive cell types in P28 Plp-Cre CNS tissues.
Figure 4.
Cre-recombinase is expressed both in and outside the oligodendrocyte lineage in spinal cords of P4 Plp-Cre mice.
A–L. Ventral spinal cord sections from WT and Plp-Cre line 627 mice were double-stained with antibodies specific to Cre (red) and either CC-1 (green) as a marker for mature oligodendrocytes (A–B, G–H), Ng2 (green) as a marker for OPCs (C–D, I–J), or GFAP (green) as a marker for astrocytes (E–F, K–L), and counterstained with DAPI. Cre was not detected in WT sections (A–F). As expected, many Cre-positive cells were CC-1-positive, however, a large percentage also co-stained for Ng2 and GFAP. Examples of Cre and glial marker co-labeling are denoted by arrows. WM = white matter, GM = grey matter. Scale bar = 20 µm.
Figure 5.
Cre-recombinase is expressed both in and outside the oligodendrocyte lineage in the cerebellum of P4 Plp-Cre mice.
A–D, G–J. Deep cerebellar white matter regions from WT and Plp-Cre line 627 mice were double-stained with antibodies specific to Cre (red) and either CC-1 (green) as a marker for mature oligodendrocytes (A–B, G–H), or Ng2 (green) as a marker for OPCs (C–D, I–J) and counterstained with DAPI. Cre was not detected in WT sections (A–D). A large percentage of Cre-positive cells within the region co-stained for CC-1 and Ng2. E–F, K–L. Cerebellar sections of the developing granular layer from WT and Plp-Cre line 627 mice were double-stained with antibodies specific to Cre (red), the neuronal marker NeuN (green) and counterstained with DAPI. Cre was not detected in WT sections (E–F). Many Cre-positive cells co-stained for NeuN. Examples of Cre-positive cells co-labeling with neuronal or glial markers are denoted by arrows. Scale bar = 20 µm.
Table 2.
Distribution of Cre-positive cell types in P4 Plp-Cre CNS tissues.
Figure 6.
β-galactosidase expression in Plp-CreERt;ROSA26LacZ mice given tamoxifen at either at P16 or P4.
A. β-galactosidase expression in Plp-CreERt;ROSA26LacZ mice given tamoxifen at P16 and examined by whole mount X-gal staining at P60. In the brain, β-galactosidase activity is predominantly observed in white matter tracts. In the spinal cord, β-galactosidase activity is detected in the dorsal and ventral white matter tracts, but is absent from the ventral and dorsal roots. Finally, β-galactosidase activity is apparent throughout the length of the optic nerve, but not in the sciatic nerve. B. β-galactosidase expression in Plp-CreERt;ROSA26LacZ mice given tamoxifen at P4 and examined by whole mount X-gal staining at P28. In the brain, β-galactosidase activity is detectable outside of large white matter tracts. In the spinal cord, β-galactosidase activity is detectable in the dorsal and ventral white matter tracts, and is also present in the ventral and dorsal roots. Finally, β-galactosidase is expressed throughout the length of the optic nerve, albeit at a reduced level, and the sciatic nerve is now positive for β-galactosidase activity. CC = corpus callosum, PCL = Purkinje cell layer, IGL = internal granular layer, VR = ventral root, DR = dorsal root. Scale bars = 1 mm in A and B Brain, 500 µm in B Spinal Cord.
Figure 7.
β-galactosidase expression outside of the oligodendrocyte lineage in CNS tissues of Plp-Cre;ROSA26LacZ mice at E10.5 and E15.5. A.
β-galactosidase expression in Plp-Cre;ROSA26LacZ embryos at E10.5. Shown are representative examples of E10.5 embryos from lines F627 and F633 after whole mount X-gal staining. Both lines show β-galactosidase expression throughout the developing brain and spinal cord. B. β-galactosidase expression in Plp-Cre;ROSA26LacZ mice at E15.5. Shown is an example of whole mount staining of tissues from line F633. β-galactosidase activity was detected throughout both the brain, and ventral and dorsal spinal cord. Scale bars = 2 mm in A and B Brain and Spinal Cord, 1 mm in A and B Optic nerve and Sciatic nerve.
Figure 8.
EGFP expression occurs outside of the oligodendrocyte lineage in CNS tissue of P28 Plp-Cre;mT/mG mice.
A. Representative example of mT/mG reporter expression in the absence of Cre recombinase in sections of the cerebellum from non-transgenic mice. Only tomato red expression is detected and EGFP expression is absent. B. EGFP expression in cerebellum of Plp-Cre;mT/mG mice. EGFP is expressed throughout the cerebellum, including the granular, Purkinje and molecular layers. Non-recombined cells, marked by tomato red expression, are limited to the vasculature. C. EGFP expression in forebrain region of Plp-Cre;mT/mG mice. EGFP is detected throughout the region. Non-recombined cells, marked by tomato red expression, are limited predominantly to vasculature. D. EGFP expression in spinal cord of Plp-Cre;mT/mG mice. EGFP is detected throughout the spinal cord including gray and white matter zones. Non-recombined cells, marked by tomato red expression, are limited to vasculature. Scale bar = 50 µm.