GABA A Receptors Containing the a2 Subunit Are Critical for Direction-Selective Inhibition in the Retina Olivia Nicola Auferkorte 1 *, Tom Baden 2,3 , Sanjeev Kumar Kaushalya 4 , Nawal Zabouri 1 , Uwe Rudolph 5,6 , Silke Haverkamp 1 , Thomas Euler 2,3,7 1 Neuroanatomy, Max-Planck-Institute for Brain Research, Frankfurt/M., Germany, 2 Bernstein Center for Computational Neuroscience (BCCN), University of Tu ¨ bingen, Tu ¨ bingen, Germany, 3 Werner Reichardt Centre for Integrative Neuroscience (CIN), University of Tu ¨ bingen, Tu ¨ bingen, Germany, 4 Department of Biomedical Optics, Max- Planck-Institute for Medical Research, Heidelberg, Germany, 5 Laboratory of Genetic Neuropharmacology, McLean Hospital, Belmont, Massachusetts, United States of America, 6 Department of Psychiatry, Harvard Medical School, Boston, Massachusetts, United States of America, 7 Institute for Ophthalmic Research, University Hospital Tu ¨ bingen, Tu ¨ bingen, Germany Abstract Far from being a simple sensor, the retina actively participates in processing visual signals. One of the best understood aspects of this processing is the detection of motion direction. Direction-selective (DS) retinal circuits include several subtypes of ganglion cells (GCs) and inhibitory interneurons, such as starburst amacrine cells (SACs). Recent studies demonstrated a surprising complexity in the arrangement of synapses in the DS circuit, i.e. between SACs and DS ganglion cells. Thus, to fully understand retinal DS mechanisms, detailed knowledge of all synaptic elements involved, particularly the nature and localization of neurotransmitter receptors, is needed. Since inhibition from SACs onto DSGCs is crucial for generating retinal direction selectivity, we investigate here the nature of the GABA receptors mediating this interaction. We found that in the inner plexiform layer (IPL) of mouse and rabbit retina, GABA A receptor subunit a2 (GABA A R a2) aggregated in synaptic clusters along two bands overlapping the dendritic plexuses of both ON and OFF SACs. On distal dendrites of individually labeled SACs in rabbit, GABA A R a2 was aligned with the majority of varicosities, the cell’s output structures, and found postsynaptically on DSGC dendrites, both in the ON and OFF portion of the IPL. In GABA A R a2 knock-out (KO) mice, light responses of retinal GCs recorded with two-photon calcium imaging revealed a significant impairment of DS responses compared to their wild-type littermates. We observed a dramatic drop in the proportion of cells exhibiting DS phenotype in both the ON and ON-OFF populations, which strongly supports our anatomical findings that a2-containing GABA A Rs are critical for mediating retinal DS inhibition. Our study reveals for the first time, to the best of our knowledge, the precise functional localization of a specific receptor subunit in the retinal DS circuit. Citation: Auferkorte ON, Baden T, Kaushalya SK, Zabouri N, Rudolph U, et al. (2012) GABA A Receptors Containing the a2 Subunit Are Critical for Direction- Selective Inhibition in the Retina. PLoS ONE 7(4): e35109. doi:10.1371/journal.pone.0035109 Editor: Steven Barnes, Dalhousie University, Canada Received December 5, 2011; Accepted March 13, 2012; Published April 10, 2012 Copyright: ß 2012 Auferkorte et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was funded by Deutsche Forschungsgemeinschaft (DFG) (HA 5277/3-1, EXC307) and the Max Planck Society. This study is part of the research program of the Bernstein Center for Computational Neuroscience, Tu ¨ bingen, funded by the German Federal Ministry of Education and Research (BMBF; FKZ: 01GQ1002). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: Olivia.Auferkorte@brain.mpg.de Introduction One of many neural computations performed by the retina is direction selectivity (DS), with certain types of retinal ganglion cells (RGCs) tuned to specific directions of image motion. Direction-selective ganglion cells (DSGCs) were first systemati- cally studied in the rabbit several decades ago [1]. They robustly fire when presented with a light stimulus moving in a particular (‘‘preferred’’) direction, but are silent when the stimulus moves in the opposite (‘‘null’’) direction. Since then, three functionally distinct types of DSGCs have been identified: the originally described ON-OFF type in the rabbit [1,2] makes up the most numerous population of DSGCs, has a bistratified morphology [3,4] and detects brighter-than-background (‘‘ON’’) as well as darker-than-background (‘‘OFF’’) moving stimuli [5]. The other ‘‘classical’’ DSGC, the ON type [2], has monostratified dendrites [6,7] and only responds to brighter-than-background stimuli. The ON and ON-OFF DSGCs can further be distinguished by their velocity tuning – the ON type prefers slower stimuli – and the fact that ON DSGCs detect global movement whereas ON-OFF DSGCs are specialized for local motion [5,8]. Furthermore, ON and ON-OFF DSGCs comprise different functional subtypes [5,6,9,10,11,12,13]. Recently, in the mouse retina another GC was described that exhibits a DS phenotype: an OFF cell with a strongly asymmetrical wedge-shaped dendritic tree that is pointing ventrally [14]. It responds preferentially to motion from the soma to the dendritic tips, suggesting that the underlying DS mechanism makes use of the cell’s highly asymmetric morphol- ogy. In contrast, most classical DSGCs have rather symmetrical dendritic arbors that do not necessarily correlate with the cell’s preferred motion direction (but see [15]). These DGSCs rely mainly on spatially asymmetric inhibition from GABAergic interneurons, the starburst amacrine cells (SACs; for review see [16,17,18]), which generate local DS signals within their dendrites [19,20,21]. However, classical DSGCs may contain a subpopulation with more pronounced asymmetrical dendritic arbors, shown to contribute to DS responses, at least for lower motion velocities [15]. PLoS ONE | www.plosone.org 1 April 2012 | Volume 7 | Issue 4 | e35109