Retinal ganglion cell topography and spatial resolving power in the pajama cardinalfish Sphaeramia nematoptera (Bleeker, 1856).

We studied the topography of retinal ganglion cells (GCs) and estimated spatial resolving power (SRP) in the pajama cardinalfish Sphaeramia nematoptera (Bleeker, 1856), a relatively small brightly colored fish inhabiting coral reefs and lagoons in the Western Pacific. S. nematoptera is an active night predator feeding on near-bottom animal plankton and benthos. DAPI staining was used to label nuclei of GCs and non-GCs in the inner plexiform and ganglion cell layers. Non-GCs were distinguished from GCs in Nissl-stained retinal wholemounts based on cell size, shape, and staining intensity. The proportion of displaced amacrine cells (DACs) varied from 15.46 ± 1.12 (visual streak [VS]) to 17.99 ± 1.06% (dorsal periphery) (mean ± S.E.M., N = 5); the respective proportions of glial cells were 6.61 ± 0.84 and 5.89 ± 0.76%. Thus, 76%-78% of cells in the ganglion cell layer and inner plexiform layer were GCs. The minimum spatial coverage of GCs (3600-4600 cells/mm2) was detected in the dorsal and ventral periphery. It gradually increased toward the central retina to form a moderate VS. The maximum GC density (11,400-12,400 cells/mm2) was registered in the central portion of the VS. No pronounced concentric retinal specializations were found. The total number of GCs ranged within 595.2-635.9 × 103. The anatomical spatial resolving power was minimum in the ventral periphery (4.91-5.53 cpd) and maximum in the central portion of the VS (8.47-9.07 cpd). The respective minimum separable angles were 0.18-0.20° and 0.11-0.12°. The relatively high spatial resolving power and presence of the VS in the pajama cardinalfish are in line with its highly visual behavior.

Regulation of retinal amacrine cell generation by miR-216b and Foxn3.

The mammalian retina contains a complex mixture of different types of neurons. We find that microRNA miR-216b is preferentially expressed in postmitotic retinal amacrine cells in the mouse retina, and expression of miR-216a/b and miR-217 in retina depend in part on Ptf1a, a transcription factor required for amacrine cell differentiation. Surprisingly, ectopic expression of miR-216b directed the formation of additional amacrine cells and reduced bipolar neurons in the developing retina. We identify the Foxn3 mRNA as a retinal target of miR-216b by Argonaute PAR-CLIP and reporter analysis. Inhibition of Foxn3, a transcription factor, in the postnatal developing retina by RNAi increased the formation of amacrine cells and reduced bipolar cell formation. Foxn3 disruption by CRISPR in embryonic retinal explants also increased amacrine cell formation, whereas Foxn3 overexpression inhibited amacrine cell formation prior to Ptf1a expression. Co-expression of Foxn3 partially reversed the effects of ectopic miR-216b on retinal cell formation. Our results identify Foxn3 as a novel regulator of interneuron formation in the developing retina and suggest that miR-216b likely regulates Foxn3 and other genes in amacrine cells.

Regional Variation of Gap Junctional Connections in the Mammalian Inner Retina.

The retinas of many species show regional specialisations that are evident in the differences in the processing of visual input from different parts of the visual field. Regional specialisation is thought to reflect an adaptation to the natural visual environment, optical constraints, and lifestyle of the species. Yet, little is known about regional differences in synaptic circuitry. Here, we were interested in the topographical distribution of connexin-36 (Cx36), the major constituent of electrical synapses in the retina. We compared the retinas of mice, rats, and cats to include species with different patterns of regional specialisations in the analysis. First, we used the density of Prox1-immunoreactive amacrine cells as a marker of any regional specialisation, with higher cell density signifying more central regions. Double-labelling experiments showed that Prox1 is expressed in AII amacrine cells in all three species. Interestingly, large Cx36 plaques were attached to about 8-10% of Prox1-positive amacrine cell somata, suggesting the strong electrical coupling of pairs or small clusters of cell bodies. When analysing the regional changes in the volumetric density of Cx36-immunoreactive plaques, we found a tight correlation with the density of Prox1-expressing amacrine cells in the ON, but not in the OFF sublamina in all three species. The results suggest that the relative contribution of electrical synapses to the ON- and OFF-pathways of the retina changes with retinal location, which may contribute to functional ON/OFF asymmetries across the visual field.

Feedback from retinal ganglion cells to the inner retina.

Retinal ganglion cells (RGCs) are thought to be strictly postsynaptic within the retina. They carry visual signals from the eye to the brain, but do not make chemical synapses onto other retinal neurons. Nevertheless, they form gap junctions with other RGCs and amacrine cells, providing possibilities for RGC signals to feed back into the inner retina. Here we identified such feedback circuitry in the salamander and mouse retinas. First, using biologically inspired circuit models, we found mutual inhibition among RGCs of the same type. We then experimentally determined that this effect is mediated by gap junctions with amacrine cells. Finally, we found that this negative feedback lowers RGC visual response gain without affecting feature selectivity. The principal neurons of the retina therefore participate in a recurrent circuit much as those in other brain areas, not being a mere collector of retinal signals, but are actively involved in visual computations.

Amacrine Cells Forming Gap Junctions With Intrinsically Photosensitive Retinal Ganglion Cells: ipRGC Types, Neuromodulator Contents, and Connexin Isoform.

Purpose: Intrinsically photosensitive retinal ganglion cells (ipRGCs) signal not only centrally to non-image-forming visual centers of the brain but also intraretinally to amacrine interneurons through gap junction electrical coupling, potentially modulating image-forming retinal processing. We aimed to determine (1) which ipRGC types couple with amacrine cells, (2) the neuromodulator contents of ipRGC-coupled amacrine cells, and (3) whether connexin36 (Cx36) contributes to ipRGC-amacrine coupling. Methods: Gap junction-permeable Neurobiotin tracer was injected into green fluorescent protein (GFP)-labeled ipRGCs in Opn4Cre/+; Z/EG mice to stain coupled amacrine cells, and immunohistochemistry was performed to reveal the neuromodulator contents of the Neurobiotin-stained amacrine cells. We also created Opn4Cre/+; Cx36flox/flox; Z/EG mice to knock out Cx36 in GFP-labeled ipRGCs and looked for changes in the number of ipRGC-coupled amacrine cells. Results: Seventy-three percent of ipRGCs, including all six types (M1-M6), were tracer-coupled with amacrine somas 5.7 to 16.5 µm in diameter but not with ganglion cells. Ninety-two percent of the ipRGC-coupled somas were in the ganglion cell layer and the rest in the inner nuclear layer. Some ipRGC-coupled amacrine cells were found to accumulate serotonin or to contain nitric oxide synthase or neuropeptide Y. Knocking out Cx36 in M2 and M4 dramatically reduced the number of coupled somas. Conclusions: Heterologous gap junction coupling with amacrine cells is widespread across mouse ipRGC types. ipRGC-coupled amacrine cells probably comprise multiple morphologic types and use multiple neuromodulators, suggesting that gap junctional ipRGC-to-amacrine signaling likely exerts diverse modulatory effects on retinal physiology. ipRGC-amacrine coupling is mediated partly, but not solely, by Cx36.

Jmjd3 Plays Pivotal Roles in the Proper Development of Early-Born Retinal Lineages: Amacrine, Horizontal, and Retinal Ganglion Cells.

Purpose: Trimethylation of histone H3 at lysine 27 (H3K27me3) is a critical mediator of transcriptional gene repression, and Jmjd3 and Utx are the demethylases specific to H3K27me3. Using an in vitro retinal explant culture system, we previously revealed the role of Jmjd3 in the development of rod bipolar cells; however, the roles of Jmjd3 in the development of early-born retinal cells are unknown due to limitations concerning the use of retinal explant culture systems. In this study, we investigated the roles of Jmjd3 in the development of early-born retinal cells. Methods: We examined retina-specific conditional Jmjd3 knockout (Jmjd3-cKO) mice using immunohistochemistry and quantitative reverse transcription PCR and JMJD3 binding to a target locus by chromatin immunoprecipitation analysis. Results: We observed reductions in amacrine cells (ACs) and horizontal cells (HCs), as well as lowered expression levels of several transcription factors involved in the development of ACs and HCs in the Jmjd3-cKO mouse retina. JMJD3 bound the promoter regions of these transcription factors. Notably, an elevated number of retinal ganglion cells (RGCs) was observed at embryonic stages, whereas RGCs were moderately reduced at later postnatal stages in the Jmjd3-cKO retina. We also observed reduced expression of Eomes, which is required for the maintenance of RGCs, as well as lower H3K27me3 level and lower JMJD3 binding in the promoter region of Eomes in RGC-enriched cells. Conclusions: The results indicated that Jmjd3 has critical roles in the development of early-born retinal subtypes, and suggested biphasic roles of Jmjd3 in RGC production and maintenance.

TRPC5 regulates axonal outgrowth in developing retinal ganglion cells.

The TRPC5 ion channel is activated upon depletion of intracellular calcium stores, as well as by various stimuli such as nitric oxide (NO), membrane stretch, and cold temperatures. TRPC5 is abundantly expressed in the central nervous system where it has important neuronal functions. In the chick retina, TRPC5 expression was shown to be restricted to amacrine cells (ACs) and Müller glial cells, although its expression was also observed in the ganglion cell layer (GCL) in displaced ACs, as determined by their characteristic cell morphology. However, it is possible that this expression analysis alone might be insufficient to fully understand the expression of TRPC5 in retinal ganglion cells (RGCs). Hence, we analyzed TRPC5 expression by in situ hybridization and immunostaining in the developing mouse retina, and for the first time identified that developing and mature RGCs strongly express TRPC5. The expression begins at E14.5, and is restricted to ACs and RGCs. It was reported that TRPC5 negatively regulates axonal outgrowth in hippocampal neurons. We thus hypothesized that TRPC5 might have similar functions in RGCs since they extend very long axons toward the brain, and this characteristic significantly differs from other retinal cell types. To elucidate its possible involvement in axonal outgrowth, we inhibited TRPC5 activity in developing RGCs which significantly increased RGC axon length. In contrast, overexpression of TRPC5 inhibited axonal outgrowth in developing RGCs. These results indicate that TRPC5 is an important negative regulator of RGC axonal outgrowth. Since TRPC5 is a mechanosensor, it might function to sense abnormal intraocular pressure changes, and could contribute to the death of RGCs in diseases such as glaucoma. In this case, excessive Ca2+ entry through TRPC5 might induce dendritic and axonal remodeling, which could lead to cell death, as our findings clearly indicate that TRPC5 is an important regulator of neurite remodeling.

Interactions between Dpr11 and DIP-γ control selection of amacrine neurons in Drosophila color vision circuits.

Drosophila R7 UV photoreceptors (PRs) are divided into yellow (y) and pale (p) subtypes. yR7 PRs express the Dpr11 cell surface protein and are presynaptic to Dm8 amacrine neurons (yDm8) that express Dpr11's binding partner DIP-γ, while pR7 PRs synapse onto DIP-γ-negative pDm8. Dpr11 and DIP-γ expression patterns define 'yellow' and 'pale' color vision circuits. We examined Dm8 neurons in these circuits by electron microscopic reconstruction and expansion microscopy. DIP-γ and dpr11 mutations affect the morphologies of yDm8 distal ('home column') dendrites. yDm8 neurons are generated in excess during development and compete for presynaptic yR7 PRs, and interactions between Dpr11 and DIP-γ are required for yDm8 survival. These interactions also allow yDm8 neurons to select yR7 PRs as their appropriate home column partners. yDm8 and pDm8 neurons do not normally compete for survival signals or R7 partners, but can be forced to do so by manipulation of R7 subtype fate.

AMIGO2 Scales Dendrite Arbors in the Retina.

The size of dendrite arbors shapes their function and differs vastly between neuron types. The signals that control dendritic arbor size remain obscure. Here, we find that in the retina, starburst amacrine cells (SACs) and rod bipolar cells (RBCs) express the homophilic cell-surface protein AMIGO2. In Amigo2 knockout (KO) mice, SAC and RBC dendrites expand while arbors of other retinal neurons remain stable. SAC dendrites are divided into a central input region and a peripheral output region that provides asymmetric inhibition to direction-selective ganglion cells (DSGCs). Input and output compartments scale precisely with increased arbor size in Amigo2 KO mice, and SAC dendrites maintain asymmetric connectivity with DSGCs. Increased coverage of SAC dendrites is accompanied by increased direction selectivity of DSGCs without changes to other ganglion cells. Our results identify AMIGO2 as a cell-type-specific dendritic scaling factor and link dendrite size and coverage to visual feature detection.

Photopharmacologic Vision Restoration Reduces Pathological Rhythmic Field Potentials in Blind Mouse Retina.

Photopharmacology has yielded compounds that have potential to restore impaired visual responses resulting from outer retinal degeneration diseases such as retinitis pigmentosa. Here we evaluate two photoswitchable azobenzene ion channel blockers, DAQ and DAA for vision restoration. DAQ exerts its effect primarily on RGCs, whereas DAA induces light-dependent spiking primarily through amacrine cell activation. Degeneration-induced local field potentials remain a major challenge common to all vision restoration approaches. These 5-10 Hz rhythmic potentials increase the background firing rate of retinal ganglion cells (RGCs) and overlay the stimulated response, thereby reducing signal-to-noise ratio. Along with the bipolar cell-selective photoswitch DAD and second-generation RGC-targeting photoswitch PhENAQ, we investigated the effects of DAA and DAQ on rhythmic local field potentials (LFPs) occurring in the degenerating retina. We found that photoswitches targeting neurons upstream of RGCs, DAA (amacrine cells) and DAD (bipolar cells) suppress the frequency of LFPs, while DAQ and PhENAQ (RGCs) had negligible effects on frequency or spectral power of LFPs. Taken together, these results demonstrate remarkable diversity of cell-type specificity of photoswitchable channel blockers in the retina and suggest that specific compounds may counter rhythmic LFPs to produce superior signal-to-noise characteristics in vision restoration.

Topography of Neurons in the Rod Pathway of Human Retina.

Purpose: The objective of this study was to map the distribution and density of the three major components of the classical scotopic "night vision" pathway (rods, rod bipolar, and AII amacrine cells) in postmortem human retinas. Methods: Four postmortem donor eyes (male and female, aged 44-56 years) were used to cut vertical sections through the temporal horizontal meridian. The sections were processed for immunohistochemistry and imaged using high-resolution multichannel confocal microscopy. Rods, rod bipolar, and AII amacrine cells were counted along the temporal horizontal meridian. Two additional retinas were used for intracellular injections. Results: Rod peak density is close to 150,000 cells/mm2 at 4 to 5 mm (15° to 20°) eccentricity, declining to below 70,000 cells/mm2 in peripheral retina. Rod bipolar density is lower but follows a similar distribution with peak density near 10,000 cells/mm2 between 2 and 4 mm (7° to 15°) eccentricity declining to below 4000 cells/mm2 in peripheral retina. The peak density of AII amacrine cells (near 4000 cells/mm2) is located close to the fovea, at 0.5- to 2 mm-eccentricity (2° to 7°) and declines to below 1000 cells/mm2 in the periphery. Thus, convergence between rods and AII cells increases from central to peripheral retina. Conclusions: Comparison with human psychophysics and ganglion cell density indicates that the spatial resolution of scotopic vision is limited by the AII mosaic at eccentricities below 15° and by the midget ganglion cell mosaic at eccentricities above 15°.

The circadian clock gene Bmal1 is required to control the timing of retinal neurogenesis and lamination of Müller glia in the mouse retina.

A single pool of multipotent retinal progenitor cells give rise to the diverse cell types within the mammalian retina. Such cellular diversity is due to precise control of various cellular processes like cell specification, proliferation, differentiation, and maturation. Circadian clock genes can control the expression of key regulators of cell cycle progression and therefore can synchronize the cell cycle state of a heterogeneous population of cells. Here we show that the protein encoded by the circadian clock gene brain and muscle arnt-like protein-1 (Bmal1) is expressed in the embryonic retina and is required to regulate the timing of cell cycle exit. Accordingly, loss of Bmal1 during retinal neurogenesis results in increased S-phase entry and delayed cell cycle exit. Disruption in cell cycle kinetics affects the timely generation of the appropriate neuronal population thus leading to an overall decrease in the number of retinal ganglion cells, amacrine cells, and an increase in the number of the late-born type II cone bipolar cells as well as the Müller glia. Additionally, the mislocalized Müller cells are observed in the photoreceptor layer in the Bmal1 conditional mutants. These changes affect the functional integrity of the visual circuitry as we report a significant delay in visual evoked potential implicit time in the retina-specific Bmal1 null animals. Our results demonstrate that Bmal1 is required to maintain the balance between the neural and glial cells in the embryonic retina by coordinating the timing of cell cycle entry and exit. Thus, Bmal1 plays an essential role during retinal neurogenesis affecting both development and function of the mature retina.-Sawant, O. B., Jidigam, V. K., Fuller, R. D., Zucaro, O. F., Kpegba, C., Yu, M., Peachey, N. S., Rao, S. The circadian clock gene Bmal1 is required to control the timing of retinal neurogenesis and lamination of Müller glia in the mouse retina.

Development of ON and OFF cholinergic amacrine cells in the human fetal retina.

Choline acetyltransferase (ChAT) expressing retinal amacrine cells are present across vertebrates. These interneurons play important roles in the development of retinal projections to the brain and in motion detection, specifically in generating direction-selective responses to moving stimuli. ChAT amacrine cells typically comprise two spatially segregated populations that form circuits in the 'ON' or 'OFF' synaptic layers of the inner retina. This stereotypic arrangement is also found across the adult human retina, with the notable exception that ChAT expression is evident in the ON but not OFF layer of the fovea, a region specialized for high-acuity vision. We thus investigated whether the human fovea exhibits a developmental path for ON and OFF ChAT cells that is retinal location-specific. Our analysis shows that at each retinal location, human ON and OFF ChAT cells differentiate, form their separate synaptic layers, and establish non-random mosaics at about the same time. However, unlike in the adult fovea, ChAT immunostaining is initially robust in both ON and OFF populations, up until at least mid-gestation. ChAT expression in the OFF layer in the fovea is therefore significantly reduced after mid-gestation. OFF ChAT cells in the human fovea and in the retinal periphery thus follow distinct maturational paths.

Multiple cell types form the VIP amacrine cell population.

Amacrine cells are a heterogeneous group of interneurons that form microcircuits with bipolar, amacrine and ganglion cells to process visual information in the inner retina. This study has characterized the morphology, neurochemistry and major cell types of a VIP-ires-Cre amacrine cell population. VIP-tdTomato and -Confetti (Brainbow2.1) mouse lines were generated by crossing a VIP-ires-Cre line with either a Cre-dependent tdTomato or Brainbow2.1 reporter line. Retinal sections and whole-mounts were evaluated by quantitative, immunohistochemical, and intracellular labeling approaches. The majority of tdTomato and Confetti fluorescent cell bodies were in the inner nuclear layer (INL) and a few cell bodies were in the ganglion cell layer (GCL). Fluorescent processes ramified in strata 1, 3, 4, and 5 of the inner plexiform layer (IPL). All tdTomato fluorescent cells expressed syntaxin 1A and GABA-immunoreactivity indicating they were amacrine cells. The average VIP-tdTomato fluorescent cell density in the INL and GCL was 535 and 24 cells/mm2 , respectively. TdTomato fluorescent cells in the INL and GCL contained VIP-immunoreactivity. The VIP-ires-Cre amacrine cell types were identified in VIP-Brainbow2.1 retinas or by intracellular labeling in VIP-tdTomato retinas. VIP-1 amacrine cells are bistratified, wide-field cells that ramify in strata 1, 4, and 5, VIP-2A and 2B amacrine cells are medium-field cells that mainly ramify in strata 3 and 4, and VIP-3 displaced amacrine cells are medium-field cells that ramify in strata 4 and 5 of the IPL. VIP-ires-Cre amacrine cells form a neuropeptide-expressing cell population with multiple cell types, which are likely to have distinct roles in visual processing.

The organization of tyrosine hydroxylase-immunopositive cells in the sparrow retina.

The purpose of this study was to identify tyrosine hydroxylase-immunopositive (TH+) cells in the sparrow retina using immunocytochemistry and quantitative analysis. All TH+ cells were conventional amacrine cells. Based on dendritic morphology, at least two types were observed. The first type had a single thick primary process that descended from the cell body and many densely beaded processes in substrata (s) 1, less beaded processes in s3, and spiny processes in s4/5 of the inner plexiform layer. The dendrites of the second type appeared similar in each layer, but it displayed several primary processes that spread laterally away from the soma before descending to the inner plexiform layer. The average density of TH+ cells was 37.48 ± 1.97 cells/mm2 (mean ± standard deviation; n = 4), and the estimated total number of TH+ cells was 3,061.25 ± 192.79. The highest and lowest densities of TH+ cells were located in the central dorsotemporal retina and periphery of the ventronasal retina, respectively. TH+ cells did not express calbindin-D28 K, calretinin, or parvalbumin. These results suggest that all TH+ cells in specific amacrine cell subpopulations are involved in retinal information processing in both the ON and OFF sublaminae in sparrow retina.

Rod-cone crossover connectome of mammalian bipolar cells.

The basis of cross-suppression between rod and cone channels has long been an enigma. Using rabbit retinal connectome RC1, we show that all cone bipolar cell (BC) classes inhibit rod BCs via amacrine cell (AC) motifs (C1-6); that all cone BC classes are themselves inhibited by AC motifs (R1-5, R25) driven by rod BCs. A sparse symmetric AC motif (CR) is presynaptic and postsynaptic to both rod and cone BCs. ON cone BCs of all classes drive inhibition of rod BCs via motif C1 wide-field GABAergic ACs (γACs) and motif C2 narrow field glycinergic ON ACs (GACs). Each rod BC receives ≈10 crossover AC synapses and each ON cone BC can target ≈10 or more rod BCs via separate AC processes. OFF cone BCs mediate monosynaptic inhibition of rod BCs via motif C3 driven by OFF γACs and GACs and disynaptic inhibition via motifs C4 and C5 driven by OFF wide-field γACs and narrow-field GACs, respectively. Motifs C4 and C5 form halos of 60-100 inhibitory synapses on proximal dendrites of AI γACs. Rod BCs inhibit surrounding arrays of cone BCs through AII GAC networks that access ON and OFF cone BC patches via motifs R1, R2, R4, R5 and a unique ON AC motif R3 that collects rod BC inputs and targets ON cone BCs. Crossover synapses for motifs C1, C4, C5, and R3 are 3-4× larger than typical feedback synapses, which may be a signature for synaptic winner-take-all switches. J. Comp. Neurol. 527:87-116, 2019. © 2016 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc.

AII amacrine cells in the primate fovea contribute to photopic vision.

The AII amacrine cell is known as a key interneuron in the scotopic (night-vision) pathway in the retina. Under scotopic conditions, rod signals are transmitted via rod bipolar cells to AII amacrine cells, which split the rod signal into the OFF (via glycinergic synapses) and the ON pathway (via gap junctions). But the AII amacrine cell also has a "day job": at high light levels when cones are active, AII connections with ON cone bipolar cells provide crossover inhibition to extend the response range of OFF cone bipolar cells. The question whether AII cells contribute to crossover inhibition in primate fovea (where rods and rod bipolar cells are rare or absent) has not been answered. Here, immunohistochemistry and three-dimensional reconstruction show that calretinin positive cells in the fovea of macaque monkeys and humans have AII morphology and connect to cone bipolar cells. The pattern of AII connections to cone bipolar cells is quantitatively similar to that of AII cells outside the fovea. Our results support the view that in mammalian retina AII cells first evolved to serve cone circuits, then later were co-opted to process scotopic signals subsequent to the evolution of rod bipolar cells.

Light-dependent pathways for dopaminergic amacrine cell development and function.

Retinal dopamine is a critical modulator of high acuity, light-adapted vision and photoreceptor coupling in the retina. Dopaminergic amacrine cells (DACs) serve as the sole source of retinal dopamine, and dopamine release in the retina follows a circadian rhythm and is modulated by light exposure. However, the retinal circuits through which light influences the development and function of DACs are still unknown. Intrinsically photosensitive retinal ganglion cells (ipRGCs) have emerged as a prime target for influencing retinal dopamine levels because they costratify with DACs in the inner plexiform layer and signal to them in a retrograde manner. Surprisingly, using genetic mouse models lacking specific phototransduction pathways, we find that while light influences the total number of DACs and retinal dopamine levels, this effect does not require ipRGCs. Instead, we find that the rod pathway is a critical modulator of both DAC number and retinal dopamine levels.

A Self-Regulating Gap Junction Network of Amacrine Cells Controls Nitric Oxide Release in the Retina.

Neuromodulators regulate circuits throughout the nervous system, and revealing the cell types and stimulus conditions controlling their release is vital to understanding their function. The effects of the neuromodulator nitric oxide (NO) have been studied in many circuits, including in the vertebrate retina, where it regulates synaptic release, gap junction coupling, and blood vessel dilation, but little is known about the cells that release NO. We show that a single type of amacrine cell (AC) controls NO release in the inner retina, and we report its light responses, electrical properties, and calcium dynamics. We discover that this AC forms a dense gap junction network and that the strength of electrical coupling in the network is regulated by light through NO. A model of the network offers insights into the biophysical specializations leading to auto-regulation of NO release within the network.

Notch signalling patterns retinal composition by regulating atoh7 during post-embryonic growth.

Patterning of a continuously growing naive field in the context of a life-long growing organ such as the teleost eye is of high functional relevance. Intrinsic and extrinsic signals have been proposed to regulate lineage specification in progenitors that exit the stem cell niche in the ciliary marginal zone (CMZ). The proper cell-type composition arising from those progenitors is a prerequisite for retinal function. Our findings in the teleost medaka (Oryzias latipes) uncover that the Notch-Atoh7 axis continuously patterns the CMZ. The complement of cell types originating from the two juxtaposed progenitors marked by Notch or Atoh7 activity contains all constituents of a retinal column. Modulation of Notch signalling specifically in Atoh7-expressing cells demonstrates the crucial role of this axis in generating the correct cell-type proportions. After transiently blocking Notch signalling, retinal patterning and differentiation is re-initiated de novo Taken together, our data show that Notch activity in the CMZ continuously structures the growing retina by juxtaposing Notch and Atoh7 progenitors that give rise to distinct complementary lineages, revealing coupling of de novo patterning and cell-type specification in the respective lineages.