Geniculocortical input drives genetic distinctions between primary and higher-order visual areas - PubMed (original) (raw)

Geniculocortical input drives genetic distinctions between primary and higher-order visual areas

Shen-Ju Chou et al. Science. 2013.

Abstract

Studies of area patterning of the neocortex have focused on primary areas, concluding that the primary visual area, V1, is specified by transcription factors (TFs) expressed by progenitors. Mechanisms that determine higher-order visual areas (V(HO)) and distinguish them from V1 are unknown. We demonstrated a requirement for thalamocortical axon (TCA) input by genetically deleting geniculocortical TCAs and showed that they drive differentiation of patterned gene expression that distinguishes V1 and V(HO). Our findings suggest a multistage process for area patterning: TFs expressed by progenitors specify an occipital visual cortical field that differentiates into V1 and V(HO); this latter phase requires geniculocortical TCA input to the nascent V1 that determines genetic distinctions between V1 and V(HO) for all layers and ultimately determines their area-specific functional properties.

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Figures

Fig. 1

Fig. 1. Selective deletion of geniculocortical TCA projection to V1 in RORα-IRES-Cre floxed COUP-TF1 cKO mice occurs early postnatally

(A) Geniculocortical projection to V1 is selectively absent in P7 cKO mice. 5-HT immunostaining on tangential sections through layer 4 of P7 WT (COUP-TF1fl/+; RORαCre/+) and cKO (COUP-TF1fl/fl; RORαCre/+) flattened cortices is shown. Rostral is at left and medial at the top. 5-HT staining reveals TCA input from principal sensory thalamic nuclei to primary sensory areas: dLG to V1, VP to S1, and MG to the primary auditory area (A). The 5-HT–negative region surrounding V1 is composed of VHO. In P7 cKO mice, 5-HT immuno-stained geniculocortical TCA input to V1 is absent. 5-HT staining of S1 appeared modestly diminished in cKO as compared to WT mice probably because COUP-TF1 may influence TCA input from VP, which expresses Cre (fig. S1) and exhibits COUP-TF1 deletion (fig. S2), but VP is less affected than dLG by COUP-TF1 deletion (figs. S2 and S4 to S6). (B) Time course of the deletion of geniculocortical TCA projection from dLG to V1 revealed in WT (COUP-TF1fl/+; RORαCre/+; Ai14) and cKO (COUP-TF1fl/fl; RORαCre/+; Ai14) mice crossed to the Ai14 line with the Cre-inducible axon reporter tdTomato. Sagittal sections from E16.5, P1, and P3 WT and cKO cortices, showing TCAs labeled by tdTomato reporter activated by Cre expressed in dLG and VP (anterior to the left, dorsal at the top) are shown. At E16.5, labeled TCAs (arrows) are densely packed in the subplate underlying the cortical plate of nascent V1, with the TCA projection being indistinguishable between WT and cKO mice. At P1, the geniculocortical TCA projection is beginning to invade the overlying CP of V1 in WT mice but is retarded in the cKO mice. By P3, the geniculocortical TCA projection is densely terminating in V1 of WT mice but is virtually absent from V1 in cKO mice. Abbreviations are as follows: 4, layer 4; ic, internal capsule; arrowheads approximate the anterior (A) – posterior (P) extent of nascent V1. Scale bars, 0.5 mm in (A) and 0.2 mm in (B).

Fig. 2

Fig. 2. Differentiation of complementary gene markers that distinguish V1 from VHO requires the postnatal influence of geniculocortical TCA projection

(A) 5-HT immunostained tangential sections of flattened cortices to reveal TCA input and RORβ and Igfbp4 in situ hybridization (ISH) from WT (COUP-TF1fl/+; RORαCre/+) and cKO (COUP-TF1fl/fl; RORαCre/+) mice; the field shown is indicated in the schematic. 5-HT and RORβ labeling were performed on sections through layer 4 of P7 cortices; Igfbp4 labeling was done on sections through layers 2/3 of P14 cortices. In WT mice, RORβ is strongly expressed in V1 and low in VHO, whereas Igfbp4 expression is low in V1 and high in VHO. The lack of TCA input in cKO V1 (*) is accompanied by a loss in differential expression patterns of RORβ and Igfbp4 that distinguish V1 from VHO in WT mice. Scale bar, 1 mm. (B) RORβ expression at P7 from collapsing all tangential sections through layer 4. The rectangles are 100 μm wide and 2700 μm long and indicate the area used for pixel intensity measurements shown in (C) and (D). (C) Plot of the normalized pixel intensity (mean ± SEM) in the rectangular field shown in (B). Each bin is 100 μm wide and 20 μm long and encompasses expression throughout layer 4 (z axis). The expression of RORβ in barrel B1 in the posterior medial barrel subfield of S1 is set at 100; other data are normalized to it. In WT (black, n = 4) mice, the RORβ expression is high in S1 and V1 and low in VHO, whereas in cKO mice (red, n = 4), expression is flattened across the occipital visual cortical field due to up-regulation in VHO and down-regulation in V1. (D) Bar graph of the statistical analysis performed with data from (C). Significant difference in RORβ expression intensity between VHO (white) and V1 (black) was observed in WT mice (*P < 0.0001), but not in cKO mice (n.s., not significant).

Fig. 3

Fig. 3. V1 and VHO delineated by complementary gene expression patterns are replaced with a uniform occipital visual cortical field after deletion of geniculo-cortical TCA input

Photos show the posterior part of P7 WT (COUP-TF1fl/+; RORαCre/+) and cKO (COUP-TF1fl/fl; RORαCre/+) cortices, as in the schematic at top left, processed using whole-mount in situ hybridization (WMISH) for Igfbp5, Cad8, and Lmo4. Igfbp5 expression specifically delineates V1; Cad8 and Lmo4 show lower expression in V1 and higher expression in VHO and delineate both. These differential expression patterns in WT mice are lost in the cKO cortex and replaced with a uniform expression field that encompasses the entire occipital visual cortical field (marked with *). Scale bar, 0.5 mm.

Fig. 4

Fig. 4. Geniculocortical input drives differential patterning of genes and proteins that contribute to functional distinctions between V1 and VHO in the adult cortex

Immunostaining for m2AChR and the SMI-32 epitope (a nonphosphorylated epitope on medium and heavy chains of neurofilaments that distinguishes functionally different forms) and ISH with an m2AChR probe on flattened tangential (A) and sagittal sections (B) of WT (COUP-TF1fl/+; RORαCre/+) and cKO (COUP-TF1fl/fl; RORαCre/+) cortices from adult (3–month-old) mice is shown. Rostral is to the left. In WT mice, m2AChR is highly expressed in layer 4 of V1, and SMI-32 is highly expressed in layers 3 and 5 of V1; both are expressed at low or nondetectable levels in VHO of WT mice. In the cKO occipital cortex, m2AChR and SMI-32 are greatly down-regulated in V1 and modestly up-regulated in VHO, resulting in uniform expression across the occipital visual cortical field (marked with *) posterior to S1 that would normally differentiate into V1 and VHO. Scale bars, 0.5 mm.

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