Expression of classic cadherins and delta-protocadherins in the developing ferret retina

Abstract

Background: Cadherins are a superfamily of calcium-dependent adhesion molecules that play multiple roles in morphogenesis, including proliferation, migration, differentiation and cell-cell recognition. The subgroups of classic cadherins and delta-protocadherins are involved in processes of neural development, such as neurite outgrowth, pathfinding, target recognition, synaptogenesis as well as synaptic plasticity. We mapped the expression of 7 classic cadherins (CDH4, CDH6, CDH7, CDH8, CDH11, CDH14, CDH20) and 8 delta-protocadherins (PCDH1, PCDH7, PCDH8, PCDH9, PCDH10, PCDH11, PCDH17, PCDH18) at representative stages of retinal development and in the mature retina of the ferret by in situ hybridization.

Results: All cadherins investigated by us are expressed differentially by restricted populations of retinal cells during specific periods of the ferret retinogenesis. For example, during embryonic development, some cadherins are exclusively expressed in the outer, proliferative zone of the neuroblast layer, whereas other cadherins mark the prospective ganglion cell layer or cells in the prospective inner nuclear layer. These expression patterns anticipate histogenetic changes that become visible in Nissl or nuclear stainings at later stages. In parallel to the ongoing development of retinal circuits, cadherin expression becomes restricted to specific subpopulations of retinal cell types, especially of ganglion cells, which express most of the investigated cadherins until adulthood. A comparison to previous results in chicken and mouse reveals overall conserved expression patterns of some cadherins but also species differences.

Conclusions: The spatiotemporally restricted expression patterns of 7 classic cadherins and 8 delta-protocadherins indicate that cadherins provide a combinatorial adhesive code that specifies developing retinal cell populations and intraretinal as well as retinofugal neural circuits in the developing ferret retina.

Figure 1

Histology and differential expression of cadherins in central regions of the developing ferret retina. Expression of cadherin-4 (CDH4; B), cadherin-6 (CDH6; C), cadherin-7 (CDH7; D), cadherin-8 (CDH8; E) and cadherin-11 (CDH11; F) was mapped with cRNA probes at embryonic day 23 (E23), E30, E38, postnatal day 2 (P2), P13, P25, and at the mature stage (P60). For neuroanatomical orientation, Nissl-stained sections (thionin) are shown in A. The different layers of the retina are indicated on the right side of each panel. Large arrows point at large ganglion cells (> 15 μm in largest diameter) in the ganglion cell layer (GCL). Small arrows point at single, large cells of the inner nuclear layer (INL) adjacent to the inner plexiform layer (IPL). Large arrowheads point at negative inner layers of the neuroblast layer (NBL). Small arrowheads point at putative horizontal cells adjacent to the outer plexiform layer (OPL) or putative developing horizontal cells in the NBL, respectively. The outer nuclear layer (ONL) is shown up to the outer limiting membrane and does not include the outer photoreceptor segments after P13. The pigment epithelium next to the neuroblast layer (NBL) is indicated by an asterisk. Scale bar in F, 50 μm, applies to all panels.

Figure 2

Differential expression of cadherins in central regions of the ferret retina during development. Expression of cadherin-14 (CDH14; A), cadherin-20 (CDH20; B), protocadherin-1 (PCDH1; C), protocadherin-7 (PCDH7; D), protocadherin-8 (PCDH11; E) and protocadherin-9 (PCDH9; F) was mapped with cRNA probes at embryonic day 23 (E23), E30, E38, postnatal day 2 (P2), P13, P25 and at the mature stage (P60). For neuroanatomical orientation, the different layers of the retina are indicated on the right side of each panel. Large arrows point at large ganglion cells (> 15 μm in largest diameter), whereas small arrows point at several cells of the ganglion cell layer (GCL) in A (CDH14). Large arrowheads point at negative inner layers of the neuroblast layer (NBL). Small arrowheads point at putative horizontal cells adjacent to the outer plexiform layer (OPL) or putative developing horizontal cells in the NBL, respectively. The outer nuclear layer (ONL) is shown up to the outer limiting membrane and does not include the outer photoreceptor segments after P13. The pigment epithelium next to the NBL is indicated by an asterisk. Areas boxed in C and F are shown at a higher magnification in Figure 4B and G, respectively. Other abbreviations: INL, inner nuclear layer; IPL, inner plexiform layer. Scale bar in F, 50 μm, applies to all panels.

Figure 3

Differential expression of cadherins in central regions of the ferret retina during development. Expression of protocadherin-10 (PCDH10; A), protocadherin-11 (PCDH11; B), protocadherin-17 (PCDH17; C) and protocadherin-18 (PCDH18; D) was mapped with cRNA probes at embryonic day 23 (E23), E30, E38, postnatal day 2 (P2), P13, P25 and at the mature stage (P60). For neuroanatomical orientation, the different layers of the retina are indicated on the right side of each panel. Large arrows point at large ganglion cells (> 15 μm in largest diameter), whereas small arrows point at several cells in the ganglion cell layer (GCL) in D (PCDH18). Large arrowheads point at amacrine cells at the inner margin of the inner nuclear layer (INL), next to the inner plexiform layer (IPL). Small arrowheads point at putative horizontal cells adjacent to the outer plexiform layer (OPL). The outer nuclear layer (ONL) is shown up to the outer limiting membrane and does not include the outer photoreceptor segments after P13. The pigment epithelium next to the neuroblast layer (NBL) is indicated by an asterisk. Scale bar in F, 50 μm, applies to all panels.

Figure 4

Cell type-specific expression of cadherins revealed by in situ hybridization (dark purple precipitate). A, B: Expression of protocadherin-7 (PCDH7; A) and protocadherin-1 (PCDH1; B) in the ganglion cell layer (GCL) at postnatal day 60 (P60). Note the positive large ganglion cells (large arrowhead in A, 19 μm diameter; large arrowhead in B, 26 μm; small arrowhead in B, 20 μm), a negative large ganglion cell (large arrow in B, 25 μm), and a positive, putative small ganglion cell (small arrow in A, 11 μm). The elongated cell in the nerve fiber layer (NFL, small arrow in B) probably represents an astrocyte. Panel B shows the area boxed in Fig. 2C. C-E: Putative displaced ganglion cells adjacent to the inner plexiform layer (IPL; arrows) express cadherin-6 at P13 (CDH6; C), protocadherin-10 at P25 (PCDH10; D), and protocadherin-9 at P60 (PCDH9; E). Cells are identified as ganglion cells based on their large somata (13 μm, 15 μm, and 14 μm, respectively) and nuclei (10 μm, 11 μm, and 12 μm, respectively) and the large gaps occupied by them between neighboring amacrine cells. F, G: Cadherin-20 expression by putative Müller glia at P13 (CDH20, arrowhead in F) and PCDH9 expression by putative bipolar cells (arrowhead in G). The small arrow marks a positive amacrine cell. Panel G shows the area boxed in Fig. 2F. H: A large horizontal cell expressing protocadherin-1 at P60 (PCDH1; arrow). I: Photoreceptor cells throughout most of the outer nuclear layer (ONL) express cadherin-8 at P60 (CDH8; arrowhead). Cell nuclei are shown in light pink color (A, B, F-I) or light blue color (Hoechst nuclear counterstain; C-E). The different layers of the retina are indicated on the right-hand side of each row of panels. Other abbreviations: INL, inner nuclear layer; OPL, outer plexiform layer. Scale bars are 20 μm.