Abstract
Purpose: :
Detection of motion direction is a visual processing task which occurs already in the retina. The responsible "direction selective" (DS) circuit includes several subtypes of ganglion cells (DSGCs) and a certain class of amacrine cells, named "starburst" (SACs.) Inhibition from SACs onto DSGCs is key in generating retinal DS signals. Here we report on the nature of the GABA receptors mediating this inhibition.
Methods: :
Rabbit retinae were dye-injected to label individual ON-OFF DSGCs and SACs, then immunostained against GABAA receptor subunits α1 and α2 (GABA-R α1 & α2.) Retinae of transgenic mice expressing GFP-tagged gephyrin protein in random GCs (Thy1-gephyrin-GFP) were also stained with these antibodies, and putative ON-OFF DSGCs were analyzed for postsynaptically anchored receptors.
Results: :
GABA-R α2 aggregates in two denser bands that overlap with staining against ChAT, a marker for SACs. Indeed, GABA-R α2 immunoreactive puncta are aligned with SAC varicosities, which represent the cell’s output structures. Varicosities along the distal dendrites of individually labeled ON SACs are almost always associated with GABA-R α2 (90%, n = 245, 5 cells), though not their proximal dendrites (2 cells.) GABA-R α2 can be found also on DSGCs dendrites (20-45 puncta/ 100 µm, 1 cell) and many gephyrin puncta along DSGC dendrites co-localize with GABA-R α2, both in the ON (58%, n = 517, 3 cells) and OFF sublayers (67%, n = 634, 3 cells.) All counts are significantly higher than chance, as assessed by quantifying the random signal overlap when one channel is manipulated spatially. In contrast, GABA-R α1 can be only occasionally seen on elements of the DS circuitry, at incidences equal to chance (SAC varicosities: 38%, n = 181, 5 cells; postsynaptic gephyrin puncta on ON-OFF DSGCs: ON - 14%, n = 361, 3 cells, OFF - 16%, n = 327, 3 cells.)
Conclusions: :
We performed a careful quantitative analysis of the distribution of two GABA receptors in the DS circuitry. We clearly show that GABA-R α2, but not GABA-R α1, is associated with SAC output structures and expressed at postsynaptic sites on the ON and OFF dendrites of DSGCs. We postulate that GABA-R α2 is the critical receptor mediating DS inhibition in the retina.
Keywords: retinal connections, networks, circuitry • inhibitory receptors • amacrine cells