DOI: 10.5061/dryad.47d7wm3kx
title: An ON-type direction selective ganglion cell in primate retina

description: To maintain a stable and clear image of the world, our eyes reflexively
 follow the direction in which a visual scene is moving. Such gaze
 stabilisation mechanisms reduce image blur as we move in the environment.
 In non-primate mammals, this behaviour is initiated by retinal output
 neurons called ON-type direction-selective ganglion cells (ON-DSGCs),
 which detect the direction of image motion and transmit signals to
 brainstem nuclei that drive compensatory eye movements. However, in
 primates, ON-DSGCs have not yet been identified in the retina, raising the
 possibility that this reflex is mediated by cortical visual areas. Here,
 we mined single-cell RNA transcriptomic data from primate retina to
 identify a candidate ON-DSGC. We then combined two-photon calcium imaging,
 molecular identification, and morphological analysis, to reveal a
 population of ON-DSGCs in the macaque retina. The morphology, molecular
 signature, and GABAergic mechanisms that underlie direction selectivity in
 primate ON-DSGCs are highly conserved with lower mammals. We further show
 that the human retina contains a homologous cell type.  The
 presence of ON-DSGCs in primates highlights the need to examine the
 contribution of subcortical retinal mechanisms to normal and aberrant gaze
 stabilisation in the developing and mature visual system. 
description: # An ON-type direction selective ganglion cell in primate retina *** This
 dataset contains the raw data from the paper: "An ON-type direction
 selective ganglion cell in primate retina". The data are from the
 following experiments: * Immunohistochemistry (IMHC): Pieces of macaque
 retina were stained for BNC2, RBPMS, and FOXP2 to analyze the density and
 proportion of pRGC10s and pRGC16s (Figure 1d-e), and their mosaic
 properties (Extended Data Fig. 1). IMHC was performed after calcium
 imaging to determine the molecular identity of recorded cells (macaque;
 Figure 2, mouse; Extended Data Fig. 5.). Candidate human On-DSGCs were
 labeled with BNC2 and RPBMS (Figure 4). Additional antibody validation was
 also performed (Extended Data Fig. 7). * Calcium imaging: Calcium
 responses to light stimuli were recorded from the ganglion cell layer in
 ex vivo macaque (Figure 2, Figure 3e-h) and mouse retinas (Extended Data
 Fig. 5). Stimuli include a bar drifting in 8 directions (2.24 degrees/s
 for macaque; 8 degrees/s for mouse). A subset of macaque direction
 selective cells were centred in the scan field and probed with bright bars
 moving in the preferred and null directions at a range of velocities
 (0.57-9 degrees/s) (Extended Data Fig. 4). One centred cell was also
 stimulated with bright spots of increasing diameter (25-750 μm) (Extended
 Data Fig. 4). * DiI filling: Iontophoretically filling cells with DiI in
 macaque tissue that was pre-labelled with BNC2. We then analysed their
 morphology (Figure 1f), stratification, and relationship with starburst
 amacrine cell processes (Figure 3a-d). * In situ hybridization (RNAscope):
 Used to validate single cell RNA sequencing data that suggests pRGC10 may
 contain multiple subtypes. One putative pRGC10 subtype shows expression of
 *FSTL4* (Extended Data Fig. 2d-e). See the manuscript methods for further
 details on experimental design and analyses. See the readme.html files
 included in each figure's folder for more information on the
 datasets. ## Description of the data and file structure The dataset is
 organized in folders according to Figure or Extended Data Figure (EDF)
 number. A "readme" .html file in each .zip file contains
 additional information on how the data are organized. The content of each
 zip file is summarized below: **Figure\_1 - BNC2 is a putative marker of
 macaque ON-DSGCs** Raw .czi confocal image files (supporting Fig. 1d-e)  
 **Figure\_2 - pRGC10 cells are direction-selective** Igor .pxp files
 containing raw calcium imaging data from macaque retina Raw .tif image
 files showing 2P calcium imaging scan fields registered to IMHC . zip
 files containing Image J ROIsets delineating cells in each scan field  
 **Figure\_3 - GABAergic inhibition is required for directional tuning of
 primate ON-DSGCs** Raw .czi and .tif image files of cells used for
 stratification analysis (supporting Fig. 3d)   **Figure\_4 - A candidate
 ON-DSGC in human retina** Raw .czi image files (supporting Fig. 4b)  
 **EDF\_1 - Macaque pRGC10 mosaic properties suggest the presence of
 multiple mosaics** Raw/processed image files (.czi & .tif) and
 Image J .roi files (supporting EDF 1b-f).   **EDF\_2 - pRGC10 cells are
 comprised of molecular sub-clusters** Raw .czi image files supporting
 RNAscope analysis (EDF 2d-e)   **EDF\_3 - BNC2+ RGCs are concentrated on
 the horizontal midline** Raw .zvi image files   **EDF\_4. ON-DSGCs show ON
 responses to spot and bar stimuli and are tuned to slower velocities**
 Readme file only pointing to location of relevant data files. **EDF\_5 -
 Identification of mouse ON-DSGCs with calcium imaging and *post-hoc*
 immunostaining** Igor .pxp files containing raw calcium imaging data from
 mouse retina Raw .tif image files showing 2P calcium imaging scan fields
 registered to IMHC . zip files containing Image J ROIsets delineating
 cells in each scan field   **EDF\_6 - Post-hoc molecular identification of
 a macaque ON-DSGC treated with gabazine.** Readme file with details of
 .tif image files showing 2P calcium imaging scan fields registered to IMHC
   **EDF\_7 - Specificity of BNC2 antibodies.** Raw .czi image files
 supporting EDF 7a-b, d-f   **EDF\_8 - BNC2+ cells with bistratified
 morphology** Raw .czi confocal image files ## Usage notes * Images are
 .czi or .tif files, which can both be read in
 [FIJI/ImageJ](https://imagej.net/software/fiji/downloads). * Calcium
 imaging responses are in a .pxp file which can be read by [Igor
 Pro](https://www.wavemetrics.com/products/igorpro) * The
 "readme" files are .html and can be opened with any web browser.
 ## Other data Publicly available single cell RNA sequencing data: *
 [GEO:GSE118480](https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE118480) (macaque) * [GEO:GSE148077](https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE148077) (human) * [GEO:GSE137400](https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE137400) (mouse)