Synaptic Convergence Patterns onto Retinal Ganglion Cells Are Preserved despite Topographic Variation in Pre- and Postsynaptic Territories

Abstract

Sensory processing can be tuned by a neuron's integration area, the types of inputs, and the proportion and number of connections with those inputs. Integration areas often vary topographically to sample space differentially across regions. Here, we highlight two visual circuits in which topographic changes in the postsynaptic retinal ganglion cell (RGC) dendritic territories and their presynaptic bipolar cell (BC) axonal territories are either matched or unmatched. Despite this difference, in both circuits, the proportion of inputs from each BC type, i.e., synaptic convergence between specific BCs and RGCs, remained constant across varying dendritic territory sizes. Furthermore, synapse density between BCs and RGCs was invariant across topography. Our results demonstrate a wiring design, likely engaging homotypic axonal tiling of BCs, that ensures consistency in synaptic convergence between specific BC types onto their target RGCs while enabling independent regulation of pre- and postsynaptic territory sizes and synapse number between cell pairs.

Figure 1.. Dendritic Morphology and PSD95 Puncta Density of OFF-S and OFF-T RGCs across the Retina

(A and B) Single-cell morphology of OFF-S ganglion cells within the (A) nasal (N) or the (B) temporal (T) retina labeled by biolistic transfection. (C) Average dendritic field size at different retinal locations; p = 0.012. (D) Dendritic diameter as a function of percent distance along the N-T axis. Data collected in total from 12 biolistic transfected retinas and 4 dye-injected retinas are shown. (E and F) Dendritic morphology for OFF-T RGCs in the (E) dorsal (D) or (F) ventral (V) retinas. (G) Average dendritic diameters of OFF-T RGCs at different retinal locations; p = 0.007. (H) Dendritic diameter as a function of percent distance along the D-V axis.(D and H) Regression line fitted to the pooled data. Data collected in total from 17 biolistic transfected retinas and 19 dye-injected retinas are shown. (I–L) Whole-mount views of an OFF-S (I) and OFF-T (K) RGC biolistically labeled by BFP and PSD95-RFP and the respective distributions of PSD95 puncta across the arbors (J and L). Magnified views of a stretch of dendrite (boxed regions) are shown in the lower panels of (I) and (K). PSD95-RFP puncta identified for quantification shown by gray dots (J and L, lower panels). (M and O) Average linear density of PSD95 puncta for OFF-S (M) and OFF-T (O) at different retinal locations. (N and P) Scatterplots show linear density of PSD95 puncta as a function of dendritic field size of OFF-S (N) and OFF-T (P). Linear regression: OFF-S: β = 6.890e–07, p = 0.348, r² = 0.04; OFF-T: b = −1.615e–06, p = 0.025, r² = 0.14. * indicates p < 0.05 in (C) and (G). Error bars are SEM. See also Figure S4.

Figure 2.. Variations in BC Axonal Size with Retinal Location

(A–D) BC axonal terminal sizes were determined using transgenic lines in which a single BC type expresses fluorescent protein (A, T1, Vsx1-cerulean2; C, T3a,Vsx1-cerulean1; D, T4, 5HTR2a-EGFP) or estimated from their Voronoi domains based on their axon stalk positions (B, T2, white dots). (E–H) Axonal sizes at four retinal locations are shown here for T1 (E), T2 (F), T3a (G), and T4 (H) bipolar cells. D, dorso-nasal; DT, dorso-temporal; VN, ventro-nasal; VT, ventro-temporal. **p < 0.001; ***p < 0.0001. Numbers indicate number of cells from 2–5 retinas. Error bars are SEM. See also Figures S1 and S4.

Figure 3.. OFF-S and OFF-T Alpha RGCs Are Differentially Connected to 4 Types of OFF BCs

(A and B) Maps of PSD95-FP apposed (yellow dots) or not apposed (magenta dots) to axon terminals of each BC type are shown for OFF-S (A) or OFF-T alpha RGCs (B). Magnified views of an example of PSD95 (yellow) puncta on the RGC dendrite apposed (solid arrowhead, 1) or not apposed (open arrowhead, 2) to axon terminals of the BC are shown on the left. Because labeling of T1 and T4 BCs was not complete, we only determined the synaptic contacts on the dendritic segments of the RGCs within an area where BCs were labeled. Grey segments of the dendrites indicate regions where there were no labeled BCs. (C) Quantification (mean ± SEM) showing the pro-portion of the total number of synapses on the dendritic arbor of OFF-S (left) and OFF-T (right) RGCs contributed by each BC type at different retinal locations. Number of cells in each region is provided. See also Figures S2, S3, and S4.

Figure 4.. Ribbon Densities of 4 OFF BC Types Are Unchanged across the Retina

(A) Maximum intensity projections of confocal image stacks of isolated axon terminals and ribbons (Ribeye-ribeye-tagRFP) of four types of BCs at dorso-nasal (DN) and ventro-temporal (VT) locations. (B) Quantification of ribbon density (mean ± SEM) per axon terminal at two retinal locations. Numbers indicate number of cells from 2–4 retinas. (C) Scatterplot of ribbon number as a function of axon terminal volume. Solid lines represent linear regression lines. T1: β= 0.5209, p = 0.039, r² = 0.2515; T2:β = 0.3761, p = 0.007, r² = 0.4671; T3a: p = 0.534; b = 0.4256, p = 0.0112, r² = 0.4078; T4: β= 0.3136, p < 0.001, r² = 0.6865. See also Figure S4.