Expression of Lhx6 in the adult and developing mouse retina
Introduction
The LIM homeodomain family of transcription factors plays a role in many processes involved in interneuron migration and development (Bourgouin et al., 1992; Ericson et al., 1992; Taira et al., 1995; Hobert and Westphal, 2000). Lhx6, a member of this family, is expressed in the medial ganglionic eminence (MGE) of the ventral telencephalon where the vast majority of cortical interneurons are generated (Grigoriou et al., 1998; Kimura et al., 1999; Lavdas et al., 1999). Lhx6 is required for tangential migration of GABAergic interneuron progenitors, and for their correct distribution in the cortical layers of postnatal animals (Alifragis et al., 2004).
The expression of Lhx6 begins embryonically in retinal progenitors at the embryonic stage (E13.5), and is expressed in cells in the ganglion cells layer (GCL), as well as in the outer portion of the inner nuclear layer (INL) (Balasubramanian et al., 2014). To confirm the expression of Lhx6 during retinal development in mice, the Lhx6-GFP knock-in allele was used to activate constitutive expression of a GFP reporter in Lhx6 expressing cells. By performing double labeling experiments using GFP and retinal markers in the mouse retina at P56, we found that GFP + Lhx6 lineage cells were predominantly expressed in amacrine cells (ACs) and retinal ganglion cells (RGCs) in the adult retina. Retinal sections at P5, P6, P8, P15, P21, and P28 were stained with GFP to validate the initial time of expression and location of Lhx6 in the developing mouse retina. To determine the neuronal cell types that express Lhx6, double labeling experiments were performed using GFP and various retinal cell markers in the differentiating retina at P7 and P15. Our findings showed that Lhx6 may contribute to cell-fate determination or maturation of retinal cells not only in RGCs but also in subsets of ACs.
Section snippets
Animals
The generation of Lhx6-GFP knock-in mice was as described elsewhere. In brief, a cDNA-encoding enhanced GFP was targeted to the locus encoding Lhx6 using homologous recombination. Mice with a C57BL⁄6 background were used to generate a heterozygous progeny, which are referred to as Lhx6-GFP mice (May et al., 2008). The day of birth was designated as P0. Animal experiments were performed in accordance with the institutional guidelines that were drafted by the Laboratory Animal Center, Hangzhou
Lhx6 was predominantly expressed in ACs and RGCs in the adult retina
Previously, we have shown that the expression of Lhx6 mRNA was observed in cells in the GCL and cells in the INL postnatally (Balasubramanian et al., 2014). Therefore, we used Lhx6-GFP knock-in mice to investigate the expression of Lhx6 in the adult retina. Co-labeling experiments using GFP and Lhx6 were performed to assess the mosaicism of GFP expression in medial ganglionic eminence (MGE). As shown in Fig. S1 (A-C), GFP was strongly expressed in Lhx6+ cells. To identify the cell types
Conclusion
In summary, our data showed that Lhx6 was expressed in the Brn3a + RGCs, ChAT + ACs, and Isl1+ ACs in the mouse retina at P56 (Fig. 1). These results demonstrated for the first time the precise cell types that co-localize with Lhx6 in the adult mouse retina. Lhx6 is initially expressed in the developing retina at P6 (Fig. 2A and B). Lhx6 was only expressed in the cells in the INL at P8 (Fig. 2C). However, Lhx6 was expressed in both INL and GCL cells from P15 to P28 (Fig. 2D–F). In addition,
Author contributions
Conceived and designed experiments: DH LG. Performed experiments: RG DZ PL XD. Analyzed the data: DH. Wrote the manuscript: DH LG.
Conflicts of interest
The authors have declared that no competing interests exist.
Acknowledgement
This study was supported by Zhejiang Natural Science Foundation No.2012C13023-1.
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