Thalamocortical input to layer 4 carries the major ascending sensory information to the mammalian sensory cortex and is crucial for the function and plasticity of sensory cortical areas. Here we report identification of a Six3-cre transgene that is selectively expressed in layer 4 of sensory cortical areas but not in the thalamus. In the mature somatosensory cortex Cre recombinase expressed from the transgene is able to mediate gene deletion in the overwhelming majority of layer 4 neurons, including GABAergic interneurons. The gene deletion in layer 4 mainly occurs during the first postnatal week. This cre transgene therefore provides a useful tool for examining the role of proteins expressed in layer 4 neurons.The mouse Six3 gene is a vertebrate homologue of Drosophila sine oculis and is expressed in the anterior-most neural plate and the developing eye in embryos (Oliver et al., 1995). Its promoter and regulatory elements have been used to generate several Six3-cre transgenic mouse lines (Furuta et al., 2000). Many of these cre transgenes are able to mediate deletion of a floxed gene in the hypothalamus and the retina (Furuta et al., 2000;Liu et al., 2007). We found that one of the reported lines, #69 (termed Six3-cre#69 here), was also able to mediate gene deletion in layer 4 of sensory cortical areas.To monitor the activity of Cre recombinase in the brain of Six3-cre#69 mice, we crossed Six3-cre#69 mice to Rosa26 (R26R) mice (Soriano, 1999) to obtain R26R/+;Six3-cre#69 double heterozygous mice. Once the R26R allele is recombined by Cre recombinase, it starts to express β-galactosidase that produces blue color in X-gal staining. To determine where Six3-cre#69 mediates recombination of the R26R locus in the brain, we performed X-gal staining on coronal brain sections from either adult double heterozygous mice or adult R26R/+ mice. In the absence of a cre transgene, no β-galactosidase activity was detected in R26R/+ brain sections (Fig. 1l). In R26R/+;Six3-cre#69 brain sections, we did detect β-galactosidase activity in the auditory cortex ( Fig. 1g-i), the somatosensory cortex (Fig. 1a-g), and the visual cortex ( Fig. 1g-j), which indicates that Six3-cre#69 is expressed in the sensory cortical areas. There were many more Cre-expressing cells in the somatosensory cortex than in the other two sensory cortical areas. In addition, the Cre activity was also present in some ventral forebrain structures, including the bed nucleus of the stria terminalis, the medial preoptic nucleus, and the anterior hypothalamus ( Fig. 1b-f). No Cre activity was observed in the midbrain, the hindbrain, or the cerebellum (Fig. 1). In the sensory cortical areas, X-gal staining seems to be specifically localized to layer 4. The recombination pattern was preserved in all six adult R26R/+;Six3-cre#69 mice we analyzed.
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NIH-PA Author ManuscriptTo confirm that the Cre activity is restricted to layer 4 in sensory cortical areas, we performed immunohistochemistry with an antibody to Neu...