Molecular classification of zebrafish retinal ganglion cells links genes to behavior

Retinal ganglion cells (RGCs) form an array of feature detectors, which transmit visual information through segregated channels to central brain regions. Characterizing RGC diversity is required to understand the logic of this functional parcellation. Using single- cell transcriptomics, we systematically classified RGCs in adult and larval zebrafish, thereby identifying marker genes for >30 stable and several transient cell types, cell type-specific markers, and their maturational changes. We used this catalog to engineer transgenic driverreporter lines, enabling experimental genetic access to specific RGC types. Strikingly, expression of a few transcription factors often predicts dendrite morphological features, in particular and axonal projections to specific tectal layers and extratectal targets. Moreover, iIn vivo calcium imaging revealed that molecularly defined RGCs exhibit highly specific functional tuningphysiological properties. Finally, chemogenetic ablation of eomesa+ RGCs, which comprise three melanopsin-expressing types with a specific projections to thalamus, pretectum and one deep tectal layerpattern, selectively impaired a visual behavior, phototaxis. Our study establishes a framework and provides a resource for systematically studying the functional architecture of the visual system. RGCs were labeled using transgenic Tg(isl2b:tagRFP) zebrafish that express RFP in all RGCs. Retinas from larval (5 dpf) and adult (4-6 months) fish were dissected and sorted using FACS. For the larval dataset, 200 manually dissected retinas were dissociated in one experiment and single cell profiles were collected across 3 replicates. For the adult dataset, about 20 retinas per batch were dissected and dissociated and droplet RNA sequencing was performed collecting a total of 15 replicates across 5 experiments. The cDNA libraries were sequenced on the Illumina HiSeq 2500 to a depth of ~30,000 reads per cell.