OFF Cone Bipolar Cells Research Articles

High temporal frequency light response in mouse retina requires FAT3 signaling in bipolar cells.

Vision is initiated by the reception of light by photoreceptors and subsequent processing via downstream retinal neurons. Proper cellular organization depends on the multi-functional tissue polarity protein FAT3, which is required for amacrine cell connectivity and retinal lamination. Here we investigated the retinal function of Fat3 mutant mice and found decreases in physiological and perceptual responses to high frequency flashes. These defects did not correlate with abnormal amacrine cell wiring, pointing instead to a role in bipolar cell subtypes that also express FAT3. The role of FAT3 in the response to high temporal frequency flashes depends upon its ability to transduce an intracellular signal. Mechanistically, FAT3 binds to the synaptic protein PTPσ, intracellularly, and is required to localize GRIK1 to OFF-cone bipolar cell synapses with cone photoreceptors. These findings expand the repertoire of FAT3's functions and reveal its importance in bipolar cells for high frequency light response.

Cross-species scRNA-seq reveals the cellular landscape of retina and early alterations in type 2 diabetes mice

Single-cell RNA sequencing (scRNA-seq) analysis have provided an unprecedented resolution for the studies on diabetic retinopathy (DR). However, the early changes in the retina in diabetes remain unclear. A total of 8 human and mouse scRNA-seq datasets, containing 276,402 cells were analyzed individually to comprehensively delineate the retinal cell atlas. The neural retinas were isolated from the type 2 diabetes (T2D) and control mice, and scRNA-seq analysis was conducted to evaluate the early effects of diabetes on the retina. Bipolar cell (BC) heterogeneity were identified. We found some stable BCs across multiple datasets, and explored their biological functions. A new RBC subtype (Car8_RBC) in the mouse retina was validated using the multi-color immunohistochemistry. AC149090.1 was significantly upregulated in the rod cells, ON cone BCs (CBCs), OFF CBCs, and RBCs in T2D mice. Additionally, the interneurons, especially BCs, were the most vulnerable cells to diabetes by integrating scRNA-seq and genome-wide association studies (GWAS) analyses. In conclusion, this study delineated a cross-species retinal cell atlas and uncovered the early pathological alterations in the retina of T2D mice.

Using optogenetics to dissect rod inputs to OFF ganglion cells in the mouse retina.

Light responses of rod photoreceptor cells traverse the retina through three pathways. The primary pathway involves synapses from rods to ON-type rod bipolar cells with OFF signals reaching retinal ganglion cells (RGCs) via sign-inverting glycinergic synapses. Secondly, rod signals can enter cones through gap junctions. Finally, rods can synapse directly onto cone OFF bipolar cells. To analyze these pathways, we obtained whole cell recordings from OFF-type α RGCs in mouse retinas while expressing channelrhodopsin-2 in rods and/or cones. Optogenetic stimulation of rods or cones evoked large fast currents in OFF RGCs. Blocking the primary rod pathway with L-AP4 and/or strychnine reduced rod-driven optogenetic currents in OFF RGCs by ~1/3. Blocking kainate receptors of OFF cone bipolar cells suppressed both rod- and cone-driven optogenetic currents in OFF RGCs. Inhibiting gap junctions between rods and cones with mecloflenamic acid or quinpirole reduced rod-driven responses in OFF RGCs. Eliminating the exocytotic Ca2+ sensor, synaptotagmin 1 (Syt1), from cones abolished cone-driven optogenetic responses in RGCs. Rod-driven currents were not significantly reduced after isolating the secondary pathway by eliminating Syt1 and synaptotagmin 7 (Syt7) to block synaptic release from rods. Eliminating Syt1 from both rods and cones abolished responses to optogenetic stimulation. In Cx36 KO retinas lacking rod-cone gap junctions, optogenetic activation of rods evoked small and slow responses in most OFF RGCs suggesting rod signals reached them through an indirect pathway. Two OFF cells showed faster responses consistent with more direct input from cone OFF bipolar cells. These data show that the secondary rod pathway supports robust inputs into OFF α RGCs and suggests the tertiary pathway recruits both direct and indirect inputs.

Impact of Retinal Degeneration on Response of ON and OFF Cone Bipolar Cells to Electrical Stimulation.

In retinal degenerative diseases, such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD), the photoreceptors become stressed and start to degenerate in the early stages of the disease. Retinal prosthetic devices have been developed to restore vision in patients by applying electrical stimulation to the surviving retinal cells. However, these devices provide limited visual perception as the therapeutic interventions are generally considered in the later stages of the disease when only inner retinal layer cells are left. A potential treatment option for retinal degenerative diseases in the early stages can be stimulating bipolar cells, which receive presynaptic signals from photoreceptors. In this work, we constructed computational models of healthy and degenerated (both ON and OFF-type) cone bipolar cells (CBCs) with realistic morphologies extracted from connectomes of the healthy and early-stage degenerated rabbit retina. We examined these cells' membrane potential and axon terminal calcium current differences when subjected to electrical stimulation. In addition, we investigated how differently healthy and degenerated cells behave with respect to various stimulation parameters, including pulse duration and cells' distance from the stimulating electrode. The results suggested that regardless of the position of the OFF CBCs in the retina model, there is not a significant difference between the membrane potential of healthy and degenerate cells when electrically stimulated. However, the healthy ON CBC axon terminal membrane potential rising time-constant is shorter (0.29 ± 0.03 ms) than the degenerated cells (0.8 ± 0.07 ms). Moreover, the ionic calcium channels at the axon terminals of the cells have a higher concentration and higher current in degenerated cells (32.24 ± 6.12 pA) than the healthy cells (13.64 ± 2.88 pA) independently of the cell's position.

Glycine Release Is Potentiated by cAMP via EPAC2 and Ca2+ Stores in a Retinal Interneuron.

Neuromodulation via the intracellular second messenger cAMP is ubiquitous at presynaptic nerve terminals. This modulation of synaptic transmission allows exocytosis to adapt to stimulus levels and reliably encode information. The AII amacrine cell (AII-AC) is a central hub for signal processing in the mammalian retina. The main apical dendrite of the AII-AC is connected to several lobular appendages that release glycine onto OFF cone bipolar cells and ganglion cells. However, the influence of cAMP on glycine release is not well understood. Using membrane capacitance measurements from mouse AII-ACs to directly measure exocytosis, we observe that intracellular dialysis of 1 mm cAMP enhances exocytosis without affecting the L-type Ca2+ current. Responses to depolarizing pulses of various durations show that the size of the readily releasable pool of vesicles nearly doubles with cAMP, while paired-pulse depression experiments suggest that release probability does not change. Specific agonists and antagonists for exchange protein activated by cAMP 2 (EPAC2) revealed that the cAMP-induced enhancement of exocytosis requires EPAC2 activation. Furthermore, intact Ca2+ stores were also necessary for the cAMP potentiation of exocytosis. Postsynaptic recordings from OFF cone bipolar cells showed that increasing cAMP with forskolin potentiated the frequency of glycinergic spontaneous IPSCs. We propose that cAMP elevations in the AII-AC lead to a robust enhancement of glycine release through an EPAC2 and Ca2+ store signaling pathway. Our results thus contribute to a better understanding of how AII-AC crossover inhibitory circuits adapt to changes in ambient luminance.SIGNIFICANCE STATEMENT The mammalian retina operates over a wide dynamic range of light intensities and contrast levels. To optimize the signal-to-noise ratio of processed visual information, both excitatory and inhibitory synapses within the retina must modulate their gain in synaptic transmission to adapt to different levels of ambient light. Here we show that increases of cAMP concentration within AII amacrine cells produce enhanced exocytosis from these glycinergic interneurons. Therefore, we propose that light-sensitive neuromodulators may change the output of glycine release from AII amacrine cells. This novel mechanism may fine-tune the amount of tonic and phasic synaptic inhibition received by bipolar cell terminals and, consequently, the spiking patterns that ganglion cells send to the upstream visual areas of the brain.

Rod and Cone Connections With Bipolar Cells in the Rabbit Retina.

Rod and cone pathways are segregated in the first stage of the retina: cones synapse with both ON- and OFF-cone bipolar cells while rods contact only rod bipolar cells. However, there is an exception to this specific wiring in that rods also contact certain OFF cone bipolar cells, providing a tertiary rod pathway. Recently, it has been proposed that there is even more crossover between rod and cone pathways. Physiological recordings suggested that rod bipolar cells receive input from cones, and ON cone bipolar cells can receive input from rods, in addition to the established pathways. To image their rod and cone contacts, we have dye-filled individual rod bipolar cells in the rabbit retina. We report that approximately half the rod bipolar cells receive one or two cone contacts. Dye-filling AII amacrine cells, combined with subtractive labeling, revealed most of the ON cone bipolar cells to which they were coupled, including the occasional blue cone bipolar cell, identified by its contacts with blue cones. Imaging the AII-coupled ON cone bipolar dendrites in this way showed that they contact cones exclusively. We conclude that there is some limited cone input to rod bipolar cells, but we could find no evidence for rod contacts with ON cone bipolar cells. The tertiary rod OFF pathway operates via direct contacts between rods and OFF cone bipolar cells. In contrast, our results do not support the presence of a tertiary rod ON pathway in the rabbit retina.

Differential Contribution of Gap Junctions to the Membrane Properties of ON- and OFF-Bipolar Cells of the Rat Retina

Gap junctions are ubiquitous within the retina, but in general, it remains to be determined whether gap junction coupling between specific cell types is sufficiently strong to mediate functionally relevant coupling via electrical synapses. From ultrastructural, tracer coupling and immunolabeling studies, there is clear evidence for gap junctions between cone bipolar cells, but it is not known if these gap junctions function as electrical synapses. Here, using whole-cell voltage-clamp recording in rat (male and female) retinal slices, we investigated whether the gap junctions of bipolar cells make a measurable contribution to the membrane properties of these cells. We measured the input resistance (RN) of bipolar cells before and after applying meclofenamic acid (MFA) to block gap junctions. In the presence of MFA, RN of ON-cone bipolar cells displayed a clear increase, paralleled by block of the electrical coupling between these cells and AII amacrine cells in recordings of coupled cell pairs. For OFF-cone and rod bipolar cells, RN did not increase in the presence of MFA. The results for rod bipolar cells are consistent with the lack of gap junctions in these cells. However, for OFF-cone bipolar cells, our results suggest that the morphologically identified gap junctions between these cells do not support a junctional conductance that is sufficient to mediate effective electrical coupling. Instead, these junctions might play a role in chemical and/or metabolic coupling between subcellular compartments.

Rod Photoreceptors Signal Fast Changes in Daylight Levels Using a Cx36-Independent Retinal Pathway in Mouse.

Temporal contrast detected by rod photoreceptors is channeled into multiple retinal rod pathways that ultimately connect to cone photoreceptor pathways via Cx36 gap junctions or via chemical synapses. However, we do not yet understand how the different rod pathways contribute to the perception of temporal contrast (changes in luminance with time) at mesopic light levels, where both rods and cones actively respond to light. Here, we use a forced-choice, operant behavior assay to investigate rod-driven, temporal contrast sensitivity (TCS) in mice of either sex. Transgenic mice with desensitized cones (GNAT2 cpfl3 line) were used to identify rod contributions to TCS in mesopic lights. We found that at low mesopic lights (400 photons/s/μm2 at the retina), control and GNAT2 cpfl3 mice had similar TCS. Surprisingly, at upper mesopic lights (8000 photons/s/μm2), GNAT2 cpfl3 mice exhibited a relative reduction in TCS to low (<12 Hz) while maintaining normal TCS to high (12-36 Hz) temporal frequencies. The rod-driven responses to high temporal frequencies developed gradually over time (>30 min). Furthermore, the TCS of GNAT2 cpfl3 and GNAT2 cpfl3 ::Cx36-/- mice matched closely, indicating that transmission of high-frequency signals (1) does not require the rod-cone Cx36 gap junctions as has been proposed in the past; and (2) a Cx36-independent rod pathway(s) (e.g., direct rod to OFF cone bipolar cell synapses and/or glycinergic synapses from AII amacrine cells to OFF ganglion cells) is sufficient for fast, mesopic rod-driven vision. These findings extend our understanding of the link between visual circuits and perception in mouse.SIGNIFICANCE STATEMENT The contributions of specific retinal pathways to visual perception are not well understood. We found that the temporal processing properties of rod-driven vision in mice change significantly with light level. In dim lights, rods relay relatively slow temporal variations. However, in daylight conditions, rod pathways exhibit high sensitivity to fast but not to slow temporal variations, whereas cone-driven responses supplement the loss in rod-driven sensitivity to slow temporal variations. Our findings highlight the dynamic interplay of rod- and cone-driven vision as light levels rise from night to daytime levels. Furthermore, the fast, rod-driven signals do not require the rod-to-cone Cx36 gap junctions as proposed in the past, but rather, can be relayed by alternative Cx36-independent rod pathways.

Corrigendum: Gap Junctions in A8 Amacrine Cells Are Made of Connexin36 but Are Differently Regulated Than Gap Junctions in AII Amacrine Cells.

[This corrects the article DOI: 10.3389/fnmol.2019.00099.].

Gap Junctions in A8 Amacrine Cells Are Made of Connexin36 but Are Differently Regulated Than Gap Junctions in AII Amacrine Cells.

In the mammalian retina, amacrine cells represent the most diverse cell class and are involved in the spatio-temporal processing of visual signals in the inner plexiform layer. They are connected to bipolar, other amacrine and ganglion cells, forming complex networks via electrical and chemical synapses. The small-field A8 amacrine cell was shown to receive non-selective glutamatergic input from OFF and ON cone bipolar cells at its bistratified dendrites in sublamina 1 and 4 of the inner plexiform layer. Interestingly, it was also shown to form electrical synapses with ON cone bipolar cells, thus resembling the rod pathway-specific AII amacrine cell. In contrast to the AII cell, however, the electrical synapses of A8 cells are poorly understood. Therefore, we made use of the Ier5-GFP mouse line, in which A8 cells are labeled by GFP, to study the gap junction composition and frequency in A8 cells. We found that A8 cells form <20 gap junctions per cell and these gap junctions consist of connexin36. Connexin36 is present at both OFF and ON dendrites of A8 cells, preferentially connecting A8 cells to type 1 OFF and type 6 and 7 ON bipolar cells and presumably other amacrine cells. Additionally, we show that the OFF dendrites of A8 cells co-stratify with the processes of dopaminergic amacrine cells from which they may receive GABAergic input via GABAA receptor subunit α3. As we found A8 cells to express dopamine receptor D1 (but not D2), we also tested whether A8 cell coupling is modulated by D1 receptor agonists and antagonists as was shown for the coupling of AII cells. However, this was not the case. In summary, our data suggests that A8 coupling is differently regulated than AII cells and may even be independent of ambient light levels and serve signal facilitation rather than providing a separate neuronal pathway.

Dopamine D1 receptor activation reduces local inner retinal inhibition to light-adapted levels.

During adaptation from dim to bright environments, changes in retinal signaling are mediated, in part, by dopamine. Dopamine is released with light and can modulate retinal receptive fields, neuronal coupling, inhibitory receptors, and rod pathway inhibition. However, it is unclear how dopamine affects inner retinal inhibition to cone bipolar cells, which relay visual information from photoreceptors to ganglion cells and are important signal processing sites. We tested the hypothesis that dopamine (D)1 receptor activation is sufficient to elicit light-adapted inhibitory changes. Local light-evoked inhibition and spontaneous activity were measured from OFF cone bipolar cells in dark-adapted mouse retinas while stimulating D1 receptors, which are located on bipolar, horizontal, and inhibitory amacrine cells. The D1 agonist SKF38393 reduced local inhibitory light-evoked response magnitude and increased response transience, which mimicked changes measured with light adaptation. D1-mediated reductions in local inhibition were more pronounced for glycinergic than GABAergic inputs, comparable with light adaptation. The effects of D1 receptors on light-evoked input were similar to the effects on spontaneous input. D1 receptor activation primarily decreased glycinergic spontaneous current frequency, similar to light adaptation, suggesting mainly a presynaptic amacrine cell site of action. These results expand the role of dopamine to include signal modulation of cone bipolar cell local inhibition. In this role, D1 receptor activation, acting primarily through glycinergic amacrine cells, may be an important mechanism for the light-adapted reduction in OFF bipolar cell inhibition since the actions are similar and dopamine is released during light adaptation. NEW & NOTEWORTHY Retinal adaptation to different luminance conditions requires the adjustment of local circuits for accurate signaling of visual scenes. Understanding mechanisms behind luminance adaptation at different retinal levels is important for understanding how the retina functions in a dynamic environment. In the mouse, we show that dopamine pathways reduce inner retinal inhibition similar to increased background luminance, suggesting the two are linked and highlighting a possible mechanism for light adaptation at an early retinal processing center.

Contribution of Bipolar Cells of Cone ON and OFF Pathways to Electroretinograms Elicited by Ultraviolet and Middle Wavelength Stimuli

ABSTRACTPurpose: To determine the contribution of the ON and OFF cone bipolar cell pathways to the electroretinograms (ERGs) elicited by ultraviolet (UV) and middle wavelength light in mice.Materials and Methods: The experiments were performed on 8- to 10-week-old C57BL/6J mice. The ERGs elicited by single-flash and flickering UV light stimuli were compared to those elicited by green light stimuli under photopic conditions. Pharmacological agents were used to selectively block the ON and OFF pathways contributing to the ERGs. Saline was used as a control. The flicker ERGs elicited by UV light were compared to the ERGs elicited by green light after the injection of the pharmacological agents to determine the contribution of the cone ON and OFF pathways to the ERGs.Results: The photopic single-flash and flicker ERGs were more sensitive to the UV light stimuli than to those elicited by green light stimuli. The flicker ERG responses elicited by both UV and green light stimuli at stimulus frequencies lower than 15-Hz decreased after L-2-amino-4-phosphobutyric acid was injected. The ERGs elicited by UV light at 30-Hz and by green light at frequencies lower than 15-Hz decreased significantly after the intravitreal injection of cis-2, 3-piperidine-dicarboxylic acid. An analysis of the ON and OFF components of the flicker ERGs showed that there might be pharmacological differences between the UV light-sensitive responses and the green light-sensitive responses.Conclusions: These results suggest that the UV light-sensitive cones connect to both the ON and OFF bipolar cells differently than that of the green light-sensitive cones.

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.

Synaptic Transfer between Rod and Cone Pathways Mediated by AII Amacrine Cells in the Mouse Retina

To understand computation in a neural circuit requires a complete synaptic connectivity map and a thorough grasp of the information-processing tasks performed by the circuit. Here, we dissect a microcircuit in the mouse retina in which scotopic visual information (i.e., single photon events, luminance, contrast) is encoded by rod bipolar cells (RBCs) and distributed to parallel ON and OFF cone bipolar cell (CBC) circuits via the AII amacrine cell, an inhibitory interneuron. Serial block-face electron microscopy (SBEM) reconstructions indicate that AIIs preferentially connect to one OFF CBC subtype (CBC2); paired whole-cell patch-clamp recordings demonstrate that, depending on the level of network activation, AIIs transmit distinct components of synaptic input from single RBCs to downstream ON and OFF CBCs. These findings highlight specific synaptic and circuit-level features that allow intermediate neurons (e.g., AIIs) within a microcircuit to filter and propagate information to downstream neurons.

The morphological characterization of orientation-biased displaced large-field ganglion cells in the central part of goldfish retina.

The vertebrate retina has about 30 subtypes of ganglion cells. Each ganglion cell receives synaptic inputs from specific types of bipolar and amacrine cells ramifying at the same depth of the inner plexiform layer (IPL), each of which is thought to process a specific aspect of visual information. Here, we identified one type of displaced ganglion cell in the goldfish retina which had a large and elongated dendritic field. As a population, all of these ganglion cells were oriented in the horizontal axis and perpendicular to the dorsal-ventral axis of the goldfish eye in the central part of retina. This ganglion cell has previously been classified as Type 1.2. However, the circuit elements which synapse with this ganglion cell are not yet characterized. We found that this displaced ganglion cell was directly tracer-coupled only with homologous ganglion cells at sites containing Cx35/36 puncta. We further illustrated that the processes of dopaminergic neurons often terminated next to intersections between processes of ganglion cells, close to where dopamine D1 receptors were localized. Finally, we showed that Mb1 ON bipolar cells had ribbon synapses in the axonal processes passing through the IPL and made ectopic synapses with this displaced ganglion cell that stratified into stratum 1 of the IPL. These results suggest that the displaced ganglion cell may synapse with both Mb1 cells using ectopic ribbon synapses and OFF cone bipolar cells with regular ribbon synapses in the IPL to function in both scotopic and photopic light conditions.

AMPA receptors at ribbon synapses in the mammalian retina: kinetic models and molecular identity.

In chemical synapses, neurotransmitter molecules released from presynaptic vesicles activate populations of postsynaptic receptors that vary in functional properties depending on their subunit composition. Differential expression and localization of specific receptor subunits are thought to play fundamental roles in signal processing, but our understanding of how that expression is adapted to the signal processing in individual synapses and microcircuits is limited. At ribbon synapses, glutamate release is independent of action potentials and characterized by a high and rapidly changing rate of release. Adequately translating such presynaptic signals into postsynaptic electrical signals poses a considerable challenge for the receptor channels in these synapses. Here, we investigated the functional properties of AMPA receptors of AII amacrine cells in rat retina that receive input at spatially segregated ribbon synapses from OFF-cone and rod bipolar cells. Using patch-clamp recording from outside-out patches, we measured the concentration dependence of response amplitude and steady-state desensitization, the single-channel conductance and the maximum open probability. The GluA4 subunit seems critical for the functional properties of AMPA receptors in AII amacrines and immunocytochemical labeling suggested that GluA4 is located at synapses made by both OFF-cone bipolar cells and rod bipolar cells. Finally, we used a series of experimental observables to develop kinetic models for AII amacrine AMPA receptors and subsequently used the models to explore the behavior of the receptors and responses generated by glutamate concentration profiles mimicking those occurring in synapses. These models will facilitate future in silico modeling of synaptic signaling and processing in AII amacrine cells.

How do horizontal cells 'talk' to cone photoreceptors? Different levels of complexity at the cone-horizontal cell synapse.

The first synapse of the retina plays a fundamental role in the visual system. Due to its importance, it is critical that it encodes information from the outside world with the greatest accuracy and precision possible. Cone photoreceptor axon terminals contain many individual synaptic sites, each represented by a presynaptic structure called a 'ribbon'. These synapses are both highly sophisticated and conserved. Each ribbon relays the light signal to one ON cone bipolar cell and several OFF cone bipolar cells, while two dendritic processes from a GABAergic interneuron, the horizontal cell, modulate the cone output via parallel feedback mechanisms. The presence of these three partners within a single synapse has raised numerous questions, and its anatomical and functional complexity is still only partially understood. However, the understanding of this synapse has recently evolved, as a consequence of progress in understanding dendritic signal processing and its role in facilitating global versus local signalling. Indeed, for the downstream retinal network, dendritic processing in horizontal cells may be essential, as they must support important functional operations such as contrast enhancement, which requires spatial averaging of the photoreceptor array, while at the same time preserving accurate spatial information. Here, we review recent progress made towards a better understanding of the cone synapse, with an emphasis on horizontal cell function, and discuss why such complexity might be necessary for early visual processing.

Connectomics of synaptic microcircuits: lessons from the outer retina.

Photoreceptors form a sophisticated synaptic complex with bipolar and horizontal cells, transmitting the signals generated by the phototransduction cascade to downstream retinal circuitry. The cone photoreceptor synapse shows several characteristic anatomical connectivity motifs that shape signal transfer: typically, ON-cone bipolar cells receive photoreceptor input through invaginating synapses; OFF-cone bipolar cells form basal synapses with photoreceptors. Both ON- and OFF-cone bipolar cells are believed to sample from all cone photoreceptors within their dendritic span. Electron microscopy and immunolabelling studies have established the robustness of these motifs, but have been limited by trade-offs in sample size and spatial resolution, respectively, constraining precise quantitative investigation to a few individual cells. 3D-serial electron microscopy overcomes these limitations and has permitted complete sets of neurons to be reconstructed over a comparatively large section of retinal tissue. Although the published mouse dataset lacks labels for synaptic structures, the characteristic anatomical motifs at the photoreceptor synapse can be exploited to identify putative synaptic contacts, which has enabled the development of a quantitative description of outer retinal connectivity. This revealed unexpected exceptions to classical motifs, including substantial interaction between rod and cone pathways at the photoreceptor synapse, sparse photoreceptor sampling and atypical contacts. Here, we summarize what was learned from this study in a more general context: we consider both the implications and limitations of the study and identify promising avenues for future research.

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.

Amyloid Precursor-Like Protein 2 deletion-induced retinal synaptopathy related to congenital stationary night blindness: structural, functional and molecular characteristics.

BackgroundAmyloid precursor protein knockout mice (APP-KO) have impaired differentiation of amacrine and horizontal cells. APP is part of a gene family and its paralogue amyloid precursor-like protein 2 (APLP2) has both shared as well as distinct expression patterns to APP, including in the retina. Given the impact of APP in the retina we investigated how APLP2 expression affected the retina using APLP2 knockout mice (APLP2-KO).ResultsUsing histology, morphometric analysis with noninvasive imaging technique and electron microscopy, we showed that APLP2-KO retina displayed abnormal formation of the outer synaptic layer, accompanied with greatly impaired photoreceptor ribbon synapses in adults. Moreover, APLP2-KO displayed a significant decease in ON-bipolar, rod bipolar and type 2 OFF-cone bipolar cells (36, 21 and 63 %, respectively). Reduction of the number of bipolar cells was accompanied with disrupted dendrites, reduced expression of metabotropic glutamate receptor 6 at the dendritic tips and alteration of axon terminals in the OFF laminae of the inner plexiform layer. In contrast, the APP-KO photoreceptor ribbon synapses and bipolar cells were intact. The APLP2-KO retina displayed numerous phenotypic similarities with the congenital stationary night blindness, a non-progressive retinal degeneration disease characterized by the loss of night vision. The pathological phenotypes in the APLP2-KO mouse correlated to altered transcription of genes involved in pre- and postsynatic structure/function, including CACNA1F, GRM6, TRMP1 and Gα0, and a normal scotopic a-wave electroretinogram amplitude, markedly reduced scotopic electroretinogram b-wave and modestly reduced photopic cone response. This confirmed the impaired function of the photoreceptor ribbon synapses and retinal bipolar cells, as is also observed in congenital stationary night blindness. Since congenital stationary night blindness present at birth, we extended our analysis to retinal differentiation and showed impaired differentiation of different bipolar cell subtypes and an altered temporal sequence of development from OFF to ON laminae in the inner plexiform layer. This was associated with the altered expression patterns of bipolar cell generation and differentiation factors, including MATH3, CHX10, VSX1 and OTX2.ConclusionsThese findings demonstrate that APLP2 couples retina development and synaptic genes and present the first evidence that APLP2 expression may be linked to synaptic disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s13041-016-0245-z) contains supplementary material, which is available to authorized users.