FoxP2 is a parvocellular-specific transcription factor in the visual thalamus of monkeys and ferrets

Abstract

Although the parallel visual pathways are a fundamental basis of visual processing, our knowledge of their molecular properties is still limited. Here, we uncovered a parvocellular-specific molecule in the dorsal lateral geniculate nucleus (dLGN) of higher mammals. We found that FoxP2 transcription factor was specifically expressed in X cells of the adult ferret dLGN. Interestingly, FoxP2 was also specifically expressed in parvocellular layers 3-6 of the dLGN of adult old world monkeys, providing new evidence for a homology between X cells in the ferret dLGN and parvocellular cells in the monkey dLGN. Furthermore, this expression pattern was established as early as gestation day 140 in the embryonic monkey dLGN, suggesting that parvocellular specification has already occurred when the cytoarchitectonic dLGN layers are formed. Our results should help in gaining a fundamental understanding of the development, evolution, and function of the parallel visual pathways, which are especially prominent in higher mammals.

Keywords: FoxP2; X cells; ferret; monkey; parvocellular.

Figure 1.

Characterization of anti-FOXP2 antibodies. (A) Immunohistochemical localization of Foxp2 protein in the mouse cerebral cortex at P13. Two independent anti-FOXP2 antibodies showed Foxp2 expression primarily in layer 6 and weakly in layer 5. Cortical layers are indicated with numbers. (B) Localization of Foxp2 mRNA and Foxp2 protein in the mouse cerebral cortex at P10. The merged image shows that Foxp2 mRNA-positive cells (pseudo-colored in green) and Foxp2 protein-positive cells (orange) are completely overlap. Cortical layers are indicated with numbers. (C) High magnification images of B. (D) FoxP2 immunoreactivities in the adult ferret visual cortex. Consistent with the results obtained using mice, 2 independent anti-FOXP2 antibodies predominantly recognized layer 6 and weakly recognized layer 5 in the ferret primary visual cortex at P101. SMI-32–positive cells (arrowheads) indicate the location of layer 5. Cortical layers are indicated with numbers in merged images. Coronal sections. Experiments were repeated at least 3 times in different animals and gave consistent results. Scale bars: 100 μm (A, B, D) and 25 μm (C).

Figure 2.

FoxP2 expression in the adult ferret dLGN. (A) Double staining with FoxP2 immunohistochemistry and Zic2 in situ hybridization using horizontal sections of the adult ferret dLGN. While Zic2-positive cells were detected throughout the entire dLGN (dashed line), FoxP2-positive cells were mostly restricted to the inner dLGN (double arrow). (B) FoxP2-positive cells were restricted to the A/A1 layers of the ferret dLGN. LGN layers were labeled by injecting CTBs into the eyes. Horizontal sections were stained with anti-FOXP2 antibody. (C) High magnification images of areas in B (boxes). (D) Double immunostaining with 2 independent anti-FOXP2 antibodies. Both antibodies resulted in the same expression patterns. (E) Distribution patterns of FoxP2-positive cells across the adult ferret dLGN. The serial horizontal sections from CTB-injected animals were stained with anti-FOXP2 antibody. Experiments were repeated at least 3 times in different animals and gave consistent results. Scale bars: 200 μm (A, B, E), 50 μm (C), and 25 μm (D).

Figure 3.

Characterization of FoxP2-positive cells in the A/A1 layer at the cellular level. (A) Double immunostaining with anti-FOXP2 antibody plus either anti-NeuN or anti-HuC/D antibody. FoxP2 was expressed in a subset of NeuN- or HuC/D-positive neurons (arrows). (B) Double immunostaining with anti-FOXP2 antibody plus either anti-GFAP or anti-APC antibody. GFAP-positive astrocytes (arrowheads) and APC-positive oligodendrocytes (arrowheads) did not express FoxP2. (C) Double staining with in situ hybridization and FoxP2 immunohistochemistry. FoxP2 expression was almost entirely limited to VGLUT2-positive cells (arrows). (D) Double immunostaining for FoxP2 plus either Cat-301 or SMI-32. Cat-301–positive or SMI-32–positive Y cells did not express FoxP2 (arrowheads). (E) Histogram of soma area sizes of Cat-301–positive cells and FoxP2-positive cells. FoxP2-positive cells were significantly smaller than Cat-301–positive cells (mean ± SD; *P< 0.01, Mann–Whitney's U test). (F) Double staining for fluorescent Nissl plus either Cat-301 or FoxP2. Soma area sizes were measured using Nissl staining patterns. Cat-301–positive cells were larger than FoxP2-positive cells (arrowheads). Horizontal sections. Experiments were repeated at least 3 times in different animals and gave consistent results. Scale bars: 20 μm (A–D) and 50 μm (F).

Figure 4.

Parvocellular-specific expression of FoxP2 in the adult monkey dLGN. (A) dLGN layers visualized with Nissl staining. (B) Immunostaining for FoxP2 within the red box in A. FoxP2 is specifically expressed in the parvocellular layers. LGN layers are indicated with numbers. (C) High magnification images. FoxP2 was strongly expressed in parvocellular layers 3–6, but was absent in magnocellular layers 1–2 and in the koniocellular layers (layer S and interlaminar zones). (D) Distinct distribution patterns of FoxP2-, Cat-301–, and calbindin-D28–positive cells. Sections were stained with fluorescent Nissl plus either anti-FoxP2, Cat-301, or anti-calbindin–D28 antibody. FoxP2-positive cells were specifically located in the parvocellular layers (arrowheads) and were rarely found in the magnocellular and koniocellular layers. Faint FoxP2 signals were observed around small vessels (open arrowheads). Cat-301–immunoreactivity was detected in layers 1 and 2, and calbindin-D28–immunoreactivity was found in the layer S and interlaminar zones. LGN layers are indicated with numbers and arrows. Experiments were repeated at least 3 times in different animals and gave consistent results. Scale bars: 500 μm (A), 250 μm (B), and 100 μm (C, D).

Figure 5.

FoxP2 expression in the monkey and ferret dLGN during development. (A) FoxP2 expression in the monkey dLGN on GD140. FoxP2 was specifically expressed in the parvocellular layers. High magnification images in the white box of the upper panel are shown in the lower panels. (B) FoxP2 expression in the dLGN (arrows) on GD85. FoxP2 immunoreactivity was stronger in the dorsal portion of the dLGN, although not as clearly so as on GD140. Frontal sections. (C) Double immunostaining for FoxP2 and HuC/D using the P1 ferret dLGN. The area within the dashed lines indicates the dLGN. Whereas HuC/D-positive cells are distributed throughout the dLGN, FoxP2-positive cells are prominent in the inner dLGN. (D) High magnification images of the inner dLGN at P14. FoxP2-positive cells were a subset of HuC/D-positive neurons. Horizontal sections. Scale bars: 1 mm (A, upper), 250 μm (A, lower), 500 μm (B), 200 μm (C), and 20 μm (D).