Figure 2

CreER^T2 fusion protein expression driven by the regulatory elements of the CaMKIIα gene allows targeting of specific regions in the adult brain upon tamoxifen treatment. (A, B) β-galactosidase staining of brain slices revealed that animals harboring a ROSA26 Cre reporter allele and the CaMKCreER^T2 transgene (R26R/CaMKCreER^T2, two copies) show low Cre activity in absence of the ligand only within the hippocampus (Hc) (A), but strong staining within the cortex (Co) and hippocampus (Hc) upon tamoxifen treatment (B). (D-F) Analysis of β-galactosidase positive neurons using cryosections of the hippocampus of R26R/CaMKCreER^T2 mice revealed sparse recombination in the granular neurons of the dentate gyrus (DG) and in hippocampal neurons of the CA1 region in the absence of tamoxifen. (C) β-galactosidase staining of a brain slice isolated from an animal harboring a ROSA26 Cre reporter allele and the constitutive CaMKCre transgene (R26R/CaMKCre) revealed recombination in the hippocampus and cortex as well as recombination in olfactory bulbs (Ob), striatum (St) and cerebellum (Ce). (G-I) Recombination of a conditional GR allele achieved using the CaMKCreER^T2 transgene (two copies) plus tamoxifen treatment (2 × 1 mg tamoxifen for five consecutive days) is comparable to recombination obtained using the constitutive CaMKCre transgene. Recombination levels detected in distinct brain regions (Co, Hc, Ht = hypothalamus, Ce) of mice heterozygous for the conditional GR allele (GR^flox) and either the constitutive (GR^flox/wt/CaMKCre) (I) or the inducible (GR^flox/wt/CaMKCreER^T2, tamoxifen-treated) (G) Cre transgene (two copies) were determined using Southern blot analysis. The Southern blot strategy is depicted in (H), black boxes represent exons three and four of the GR gene, the arrowheads the loxP sites and S marks SacI recognition sites. The probe detects in case of the GR^flox and the GR^wt allele a DNA fragment of similar size and in case of the GR^null allele a smaller DNA fragment. The bands were quantified by phosphoimager and the percentage of recombination for each region is indicated. Animals for Cre reporter and Southern blot analysis were treated with 1 mg tamoxifen or vehicle twice a day for five consecutive days and sacrificed ten days after the last injection.