The Molecular Logic of Synaptic Specificity in the Retinocollicular Pathway

Vision is vital for many animals to interact with the world around them. Visual perception commences within the retina, where various types of retinal ganglion cells (RGCs) extract visual information from photoreceptor cells and transfer it to the central nervous system (Hahn et al., 2023). Among the primary targets of RGCs is the superior colliculus (SC), a prominent sensorimotor hub that integrates multisensory inputs and drives reflexive behaviors and higher cognitive functions (Cang et al., 2018). Of particular significance within the SC is its superficial layer (sSC) that receives input from most RGCs and exhibits diverse molecular cell types and visual responses (Liu et al., 2023). Over the course of five decades, investigations into retino-colliculus (retinocollicular) circuitry at a mesoscopic scale have established its value as a model for studying the molecular mechanisms of circuitry assembly (Cang et al., 2018).

Investigation of retinocollicular pathways has classically focused on retinotopic maps and layer-specific connectivity. Visual space in the sSC is represented as a two-dimensional retinotopic map due to topographic retinocollicular circuitry (Cang and Feldheim, 2013). Specifically, the nasal-temporal (N-T) axis of the visual field (azimuth) is mirrored along the anterior-posterior (A-P) axis of the SC, while the dorsal-ventral (D-V) axis (elevation) is represented along the medial-lateral (M-L) axis of the SC. The sSC's precise retinotopic organization is important for aligning sensory inputs from the visual, somatosensory, auditory, and motor cortices in the deeper layer of SC. The alignment of these sensory and motor maps enables the SC to respond to sensory stimuli occurring at the same spatial location and to initiate appropriate motor responses. During brain development, the retinotopic map is thought to establish initially from molecular cues and then be refined by retinal spontaneous activity (Cang et al., 2018).

More recent work has revealed that the specificity of retinocollicular projections goes much deeper than the level of retinotopic maps, to the level of individual cell types (Tsai et al., 2022). Studies have revealed that different types of RGCs arborize within distinct sublaminae of the mouse SC (Hong et al., 2011), consistent with the laminar organization of SC functions such as direction selectivity and orientation selectivity (Wang et al., 2010; Cang et al., 2018). Moreover, some subsets of RGCs selectively sends axons to collicular neurons that project to the parabigeminal nucleus or/and the pulvinar nucleus of the thalamus (Reinhard et al., 2019); the functional characteristics of these subsets of RGCs correlate well with the visual responses of these nuclei (Reinhard et al., 2019). Tracing and transcriptomic studies have also led to the identification of eight distinct molecular types of collicular neurons that receive inputs from RGCs (Tsai et al., 2022), and in-depth examination has revealed cell type-specific retinocollicular circuits. For example, αRGCs show distinct connectivity with SC wide-field neurons (NPWFs) that express the glycoprotein nephronectin, but such connectivity has not been observed with On–Off direction-selective ganglion cells (ooDSGCs; Reinhard et al., 2019; Tsai et al., 2022).

How do the retinal ganglion cell types recognize and connect to their target collicular cell types? A recent study has shed light on one particular mechanism: an interaction between the cell adhesion molecules Integrin α8β1 and Nephronectin (Npnt) that guides αRGCs sublaminar choice to selectively connect with NPWFs in the SO sublayer of sSC (Tsai et al., 2022). Other cell adhesion molecules may mediate specific partner identification by other RGCs. One class of cell adhesion molecules known for roles in synaptic specificity is the Cadherin (Cdh) family of proteins. Cdhs have been shown to promote target-specific axonal arborization and synaptic recognition within inner retina circuits (Duan et al., 2018). However, the role of Cdhs in wiring retinocollicular circuits during neural development remains unclear.

In a recent article in The Journal of Neuroscience, Matcham et al. (2023) investigated the involvement of Cdhs in the development of retinocollicular circuits. The authors first examined the expression pattern of Cdhs in the SC. They detected Cdh4, Cdh8, and Cdh13 expression in the sSC and established transgenic mouse lines to visualize the cells expressing these Cdhs. Comparing the morphology and laminar distribution of Cdh-positive cells with known sSC morphological cell types revealed that WFs express Cdh13. In situ hybridization confirmed the expression of Cdh13 in WFs and demonstrated the lack of expression in other morphological cell types of excitatory neurons, including Narrow Field cells (NFs) and stellate cells. This was further confirmed by analyzing single-cell RNA sequencing (scRNA-seq) clustering of retinorecipient SC cells, which revealed that Cdh13 is selectively expressed in NPWFs.

Next, Matcham and colleagues elucidated the role of Cdh13 in wiring a subset of RGCs to Wide Field cells (WFs) of the sSC. To investigate the necessity of Cdh13 for retinocollicular connectivity, Matcham et al. (2023) knocked out Cdh13 specifically from WFs using transgenic mouse lines. This manipulation resulted in a reduction in mature spine density on WF dendrites, most strikingly in the apical region nearest to the SC surface. Knockdown of Cdh13 with short hairpin RNA (shRNA) also significantly reduced spine density, particularly of mature spines, on WFs of adult mice. However, the knockdown did not affect NF spine density, suggesting a cell type-specific role for Cdh13 in influencing dendritic spine density. Since germline knock-out and adult knockdown of Cdh13 both result in dendritic spine loss, Cdh13 likely plays an important role in retinocollicular connectivity.

Immunohistochemical labeling of Cdh13 along with cell type-specific RGC markers identified a subtype of Cdh13-positive RGCs that were distinct from αRGC, ooDSGC, ipRGCs, or F-RGCs. Given that Cdh13 operates homophilically to regulate synaptic specificity, Matcham et al. (2023) sought to determine whether the expression of presynaptic Cdh13 in RGCs was necessary for maintaining mature spines in the SC. Knocking down Cdh13 expression with shRNA specifically in RGCs caused a significant loss of mature spines in postsynaptic WFs, while the axonal arborization of RGCs within SC laminae remained unaffected. These findings provide compelling evidence that Cdh13 facilitates retinocollicular connectivity on WFs and suggest Cdh13 promotes synapse formation by a specific type of RGCs.

These findings revealed the role of Cdh13–Cdh13 signaling in establishing cell type-specific retinocollicular circuits. The unique expression of Cdh13 within a specific subpopulation confines the role to a subset of RGCs and WFs, thereby providing cell-type specificity. Notably, previous research has revealed that the interaction between Integrin α8β1 and Npnt mediates selective wiring from αRGCs to the same collicular population (Tsai et al., 2022). But unlike deletion of Cdh13 in the retina, knockdown of Integrin α8β1 in αRGCs or Npnt in NPWFs caused mistargeting of RGC axons. These differences suggest that RGC subtypes may use different wiring rules to recognize postsynaptic collicular neurons during retinocollicular synaptogenesis. Further exploration is warranted to uncover the mechanisms instructing the specific connectivity of other retinocollicular cells.

Investigating circuitry assembly at the cell-type level may offer insights into the wiring principles underlying brain function. For example, WFs, the most extensively studied population within the SC (Cang et al., 2018), are predominantly situated in the SO sublayer, with extensive dendritic arbors extending to the surface of the SC, where they receive inputs from ten types of RGCs (Cang et al., 2018; Reinhard et al., 2019). Despite their large receptive fields, WFs exhibit a preference for detecting small moving objects. However, the specific contributions of various RGC types with diverse functional properties to the visual tuning of WFs remain largely unexplored. Given that the removal of specific cell adhesion molecules such as Cdh13, Integrin α8β1, or Npnt disrupts connections between specific cell types, it is reasonable to anticipate such manipulations have distinct functional and behavioral consequences on WFs. By using high-throughput scRNA-seq to identify cell adhesion molecules enriched within specific cell types (Tsai et al., 2022; Matcham et al., 2023), and then knocking out these proteins, we may gain a better understanding not only of how RGCs recognize and connect with their synaptic partners in the SC but also how these specific connections contribute to the overall function of SC cells.