To characterize the DsRed-fluorescent cells in the EGL, we stained for several astroglial markers

To characterize the DsRed-fluorescent cells in the EGL, we stained for several astroglial markers. m. NIHMS211743-supplement-02.tif (6.7M) GUID:?74FDBE85-4238-42E5-BFE1-830F49216922 03: Supplementary Figure 3. Characterization of Pax6 expression in CGE;CAG-EGFP; promoter, we BI-4464 found 2 populations of DsRed+ cells in the EGL in the first postnatal week defined by bright and faint DsRed fluorescent signal. Bright DsRed+ cells have a protein expression profile and electrophysiological characteristics typical of astrocytes, but faint DsRed+ cells in the EGL and internal granule cell layer (IGL) express markers and physiological properties of immature neurons. To determine if these BI-4464 astroglial cells gave rise to GCPs, we genetically tagged them with EGFP or gal reporter genes at postnatal day (P)3-P5 using a promoter driven inducible recombinase. We found that promoter+ cells in the EGL are proliferative and express glial and neural stem cell markers. In addition, immature granule cells (GCs) en route to the IGL at P12 as well as GCs in the mature cerebellum, 30 days after recombination, express the reporter protein, suggesting that promoter+ cells in the EGL generate a subset of granule cells. The identification of glial cells which function BI-4464 as neuronal progenitor cells profoundly impacts our understanding of cellular plasticity in the developing cerebellum. from a knock-in animals where the gene is replaced with a LacZ reporter the EGL does not form. However, a small population of positive cells are generated from the rhombic lip and migrate tangentially over the BI-4464 surface of the cerebellum. These cells are retained postnatally and express the astroglial marker GFAP (Jensen et al., 2004) Reinforcing these findings and the notion that not all cells of the EGL are (Okano-Uchida et al., 2004). However, the generation of mature cerebellar GCs from postnatal promoter, and give rise to juvenile and adult NSC/NPCs of the subventricular zone and dentate gyrus, in which both the mouse and human promoters are expressed (Casper and McCarthy, 2006; Doetsch et al., 1999; Ganat et al., 2006; Garcia et al., 2004; Seri et al., 2001). In this paper, we examine whether Rabbit polyclonal to ZNF167 the EGL contains neural progenitor cells with glial characteristics that generate GCs. We used several lines of transgenic mice expressing reporter genes under the human promoter (recombinase (CreErT2), were then used to permanently mark cells with promoter activity with reporter genes and study their fate at specific time points in cerebellar development. Our results indicate that in a temporally restricted window, limited to the first two postnatal weeks of cerebellar development. RESULTS The EGL contains GFAP-expressing cells with characteristics BI-4464 of neural progenitors In our experiments, we used transgenic lines driven by the to identify, characterize and fate map astroglial cells in the cerebellum of neonatal mice. The promoter is expressed in radial glia, earlier than the mouse promoter, which is not highly expressed at embryonic stages (see GENSAT database at: http://www.gensat.org/imagenavigatorfileselection.jsp?gene.id=507&mouseage.id=8). However, after birth, both promoters largely identify the same cell types. The mouse promoter is highly expressed both by Bergmann glia and by other glial cells throughout the cerebellum (http://www.gensat.org/imagenavigatorfileselection.jsp?gene.id=507&mouseage.id=9). Consistently, in mice (data not shown). At P0-P2, DsRed-fluorescent cells were found throughout the EGL (Fig 1 ACD). By P4 the majority of bright DsRed-fluorescent cells were in the inner EGL and at both time points contacted the pial surface with a single process (Fig. 1 ECH and see Supplementary Fig. 1). We also found DsRed-fluorescent cells in other regions of the cerebellum, including the IGL and PL (Bergmann glia), as expected (Fig. 1). To characterize the DsRed-fluorescent cells in the EGL, we stained for several astroglial markers. Blbp, a marker of radial glia and immature astrocytes (Ganat et al., 2002; Malatesta et al., 2003), stained the majority of DsRed-fluorescent cells in the EGL at P0-P2 (Fig. 1 ACD). As expected, the classical astrocytic marker GFAP was not expressed in P0-P2 cerebellum (data not shown). By P4, DsRed-fluorescent cells in the EGL stained positive for GFAP (Fig. 1 ECH). Open in a separate window Figure 1 Cells expressing immature glial markers and promoter activity are present in the cerebellar EGLIn promoter-expressing radial glial cells in the embryonic rhombic lip are progenitors for GCs (Schuller et al., 2008; Spassky et al., 2008), we stained for known markers of NSCs such as nestin, Sox2 and Mushahi1 (Ellis et al., 2004; Imura et al., 2006; Kaneko et al., 2000; Zappone et al., 2000). We found that DsRed-fluorescent cells stained for.