Temporally regulated expression of Cre recombinase in neural stem cells

Abstract

The use of mouse gene targeting to study molecules important in neural development is oftentimes impaired by early embryonic lethality. In order to address later roles for such molecules, specifically in neural stem cells, we generated transgenic mice that express both the tetracycline-inducible molecule rtTA-M2 and GFP under the control of the neural precursor specific form of nestin. Developmental analysis of these mice demonstrates that GFP expression is exclusive to the neural tube. Adult expression of GFP is seen only in known areas of adult neurogenesis, namely, the subventricular zone and the dentate gyrus. When crossed with a second transgenic mouse (TetOp-Cre) that expresses the Cre recombinase under the control of the tetracycline responsive promotor, we demonstrate temporal induction of Cre in bigenic animals exposed to doxycycline. We further demonstrate the feasibility of this approach by using the ROSA-26 reporter mouse to mediate recombination in neural precursor cells.

Dynamic Contribution of Nestin-Expressing Stem Cells to Adult Neurogenesis

“…These data are in agreement with links between nestin expression and a specific lineage in many non-neuronal and oncogenic stem cells (Wiese et al, 2004). However, they are in contrast to the multilineage role for nestin in the embryo (Beech et al, 2004;Yu et al, 2005;Carlen et al, 2006;Imayoshi et al, 2006;Kuo et al, 2006;Burns et al, 2007), most notably in our nestin-CreER T2 /R26R-YFP mouse (Battiste et al, 2007). This underscores the importance of the permissive neurogenic microenvironment in determining the ultimate phenotypic fate of the progeny of nestinexpressing cells.…”

“…To address this challenge in studying neurogenesis, we and others created conditional and inducible nestin-driven transgenic mice (Beech et al, 2004;Yu et al, 2005;Carlen et al, 2006;Imayoshi et al, 2006;Kuo et al, 2006;Burns et al, 2007). Whereas many of these mouse models are highly effective in labeling and tracking the multipotent lineage of nestinrecombined cells in the embryo, only three of these models have demonstrated efficient labeling within the adult SVZ (Yu et al, 2005;Carlen et al, 2006;Burns et al, 2007), and none have labeled the large pool of adult SGZ stem and progenitor cells.…”

“…Whereas many of these mouse models are highly effective in labeling and tracking the multipotent lineage of nestinrecombined cells in the embryo, only three of these models have demonstrated efficient labeling within the adult SVZ (Yu et al, 2005;Carlen et al, 2006;Burns et al, 2007), and none have labeled the large pool of adult SGZ stem and progenitor cells.…”

Bax limits adult neural stem cell persistence through caspase and IP3 receptor activation

“…51 Bax-deficient mice were crossed to nestin-GFP expressing transgenic mice and maintained as heterozygotes for the nestin-GFP transgene and genotyped also according to a previously published protocol. 22 BrdU was injected as previously described where six WT and six Bax-deficient mice were injected intraperitoneally with 50 mg/gm bodyweight daily for 7 days and then killed after 3 weeks. 12 For flow cytometry, the SVZ from four Bax-deficient/nestin-GFP and four littermate controls that only contained the nestin-GFP transgene were analyzed by digesting the SVZ tissue in DMEM with a mixture of 0.1% neural protease, 0.01% papain and 0.01% DNAse (all from Invitrogen) at 371C for 30 min.…”

“…To definitively determine that the neural precursor cell number was increased in the Baxdeficient SVZ, we crossed Bax-deficient mice to transgenic mice that express GFP under the control of the neural precursor specific form of the nestin promoter. 22 The adult Bax-deficient SVZ shows a 50% increase in the number of GFP-expressing cells as assayed by Fluorescence Activated Cell Sorting (FACS) analysis (Figure 1h). Thus, ablation of Bax function causes increased numbers of neural precursors in the adult mouse subventricular zone.…”