Neural Circuits Underlying Multifeature Extraction in the Retina.

Classic ON-OFF direction-selective ganglion cells (DSGCs) that encode the four cardinal directions were recently shown to also be orientation-selective. To clarify the mechanisms underlying orientation selectivity, we employed a variety of electrophysiological, optogenetic, and gene knock-out strategies to test the relative contributions of glutamate, GABA, and acetylcholine (ACh) input that are known to drive DSGCs, in male and female mouse retinas. Extracellular spike recordings revealed that DSGCs respond preferentially to either vertical or horizontal bars, those that are perpendicular to their preferred-null motion axes. By contrast, the glutamate input to all four DSGC types measured using whole-cell patch-clamp techniques was found to be tuned along the vertical axis. Tuned glutamatergic excitation was heavily reliant on type 5A bipolar cells, which appear to be electrically coupled via connexin 36 containing gap junctions to the vertically oriented processes of wide-field amacrine cells. Vertically tuned inputs are transformed by the GABAergic/cholinergic "starburst" amacrine cells (SACs), which are critical components of the direction-selective circuit, into distinct patterns of inhibition and excitation. Feed-forward SAC inhibition appears to "veto" preferred orientation glutamate excitation in dorsal/ventral (but not nasal/temporal) coding DSGCs "flipping" their orientation tuning by 90° and accounts for the apparent mismatch between glutamate input tuning and the DSGC's spiking response. Together, these results reveal how two distinct synaptic motifs interact to generate complex feature selectivity, shedding light on the intricate circuitry that underlies visual processing in the retina.

An uncommon neuronal class conveys visual signals from rods and cones to retinal ganglion cells.

Neurons in the central nervous system (CNS) are distinguished by the neurotransmitter types they release, their synaptic connections, morphology, and genetic profiles. To fully understand how the CNS works, it is critical to identify all neuronal classes and reveal their synaptic connections. The retina has been extensively used to study neuronal development and circuit formation. Here, we describe a previously unidentified interneuron in mammalian retina. This interneuron shares some morphological, physiological, and molecular features with retinal bipolar cells, such as receiving input from photoreceptors and relaying visual signals to retinal ganglion cells. It also shares some features with amacrine cells (ACs), particularly Aii-ACs, such as their neurite morphology in the inner plexiform layer, the expression of some AC-specific markers, and possibly the release of the inhibitory neurotransmitter glycine. Thus, we unveil an uncommon interneuron, which may play an atypical role in vision.

The Variety of Mechanosensitive Ion Channels in Retinal Neurons.

Alterations in intraocular and external pressure critically involve the pathogenesis of glaucoma, traumatic retinal injury (TRI), and other retinal disorders, and retinal neurons have been reported to express multiple mechanical-sensitive channels (MSCs) in recent decades. However, the role of MSCs in visual functions and pressure-related retinal conditions has been unclear. This review will focus on the variety and functional significance of the MSCs permeable to K+, Na+, and Ca2+, primarily including the big potassium channel (BK); the two-pore domain potassium channels TRAAK and TREK; Piezo; the epithelial sodium channel (ENaC); and the transient receptor potential channels vanilloid TRPV1, TRPV2, and TRPV4 in retinal photoreceptors, bipolar cells, horizontal cells, amacrine cells, and ganglion cells. Most MSCs do not directly mediate visual signals in vertebrate retinas. On the other hand, some studies have shown that MSCs can open in physiological conditions and regulate the activities of retinal neurons. While these data reasonably predict the crossing of visual and mechanical signals, how retinal light pathways deal with endogenous and exogenous mechanical stimulation is uncertain.

Cell numbers, cell ratios, and developmental plasticity in the rod pathway of the mouse retina.

The precise specification of cellular fate is thought to ensure the production of the correct number of neurons within a population. Programmed cell death may be an additional mechanism controlling cell number, believed to refine the proper ratio of pre- to post-synaptic neurons for a given species. Here, we consider the size of three different neuronal populations in the rod pathway of the mouse retina: rod photoreceptors, rod bipolar cells, and AII amacrine cells. Across a collection of 28 different strains of mice, large variation in the numbers of all three cell types is present. The variation in their numbers is not correlated, so that the ratio of rods to rod bipolar cells, as well as rod bipolar cells to AII amacrine cells, varies as well. Establishing connectivity between such variable pre- and post-synaptic populations relies upon plasticity that modulates process outgrowth and morphological differentiation, which we explore experimentally for both rod bipolar and AII amacrine cells in a mouse retina with elevated numbers of each cell type. While both rod bipolar dendritic and axonal arbors, along with AII lobular arbors, modulate their areal size in relation to local homotypic cell densities, the dendritic appendages of the AII amacrine cells do not. Rather, these processes exhibit a different form of plasticity, regulating the branching density of their overlapping arbors. Each form of plasticity should ensure uniformity in retinal coverage in the presence of the independent specification of afferent and target cell number.

Recovery of dopaminergic amacrine cells after strobe light stimulation in the developing rat retina.

Concerns regarding the impact of strobe light on human health and life have recently been raised. Sources of strobe light include visual display terminals, light-emitting diodes, and computer monitors. Strobe light exposure leads to visual discomfort, headaches, and poor visual performance and affects the number of dopaminergic amacrine cells (DACs) in the developing retina, as well as retinal dopamine levels in animals. DACs serve as the sole source of retinal dopamine, and dopamine release from the retina is activated by light exposure following a circadian rhythm. Using a Sprague-Dawley rat model, this study sought to determine whether changes in DACs caused by strobe light are recoverable after ceasing strobe light exposure during retinal development. From eye opening (postnatal 2 weeks), rats in the control group were reared under normal light (an unflickering 150 lux incandescent lamp with a 12 h light/dark cycle), whereas those in the experimental group (i.e., strobe-recovery group) were reared under strobe light (2 Hz for 12 h/day) exposure for 2 weeks. After postnatal week 4, normal light was provided to all animals to observe the reversibility of the effect of strobe light. Immunohistochemistry and immunoblot analysis for the rate limiting enzyme for dopamine synthesis, tyrosine hydroxylase (TH), as well as high-pressure liquid chromatography for measuring dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) were performed at postnatal weeks 4, 6, 8, and 10. The number of type I and type II TH-immunoreactive (TH-IR) cells across the entire retina was counted to evaluate whether changes in DACs induced by strobe light could recover after ceasing strobe light exposure. The number of type I TH-IR cells slightly decreased but remained at a constant level in the control group. In contrast, the number of type I TH-IR cells rapidly decreased up to postnatal week 6, but then increased after postnatal week 8 in the strobe-recovery group. Subsequently, the number of type I TH-IR cells eventually reached a number similar to that in the control group. In addition, the number of intermediate-sized TH-IR cells were increased at postnatal weeks 8 and 10 and the dopamine level was decreased at postnatal week 8 in the strobe-recovery group. However, the levels of DOPAC and TH proteins did not differ between the two groups. This suggests that changes in DACs caused by strobe light are reversible and that type II TH-IR cells may play a key role in this recovery.

Loss of optic nerve oligodendrocytes during maturation alters retinal organization.

The myelin sheath facilitates signal conduction along axons in white matter tracts, and when disrupted, can result in significant functional deficits. Demyelination, observed in diseases like multiple sclerosis and optic neuritis, are associated with neural degeneration, however the extent of this damage on upstream circuitry is not well understood. Here we use the MBP-iCP9 mouse model to induce selective oligodendrocyte ablation in the optic nerve at P14 via a chemical inducer of dimerization (CID), resulting in partial demyelination of retinal ganglion cell (RGC) axons with minimal inflammation after two weeks. Oligodendrocyte loss reduced axon diameter and altered compound action potential waveforms, blocking conduction in the slowest-conducting axon populations. Demyelination resulted in disruptions to the normal composition of the retina, including reduced density of RBPMS+, Brn3a+, and OFF-transient RGCs, thinning of the IPL, and reduced density of displaced amacrine cells. The INL and ONL were unaffected by oligodendrocyte loss, suggesting that demyelination-induced deficits in this model are specific to the IPL and GCL. These results show that a partial demyelination of a subpopulation of RGC axons disrupts optic nerve function and affects the organization of the retinal network. This study highlights the significance of myelination in maintaining upstream neural connectivity and provides support for targeting neuronal degeneration in treatments of demyelinating diseases.

A retinal circuit for the suppressed-by-contrast receptive field of a polyaxonal amacrine cell.

Amacrine cells are a diverse population of interneurons in the retina that play a critical role in extracting complex features of the visual world and shaping the receptive fields of retinal output neurons (ganglion cells). While much of the computational power of amacrine cells is believed to arise from the immense mutual interactions among amacrine cells themselves, the intricate circuitry and functions of amacrine-amacrine interactions are poorly understood in general. Here we report a specific interamacrine pathway from a small-field, glutamate-glycine dual-transmitter amacrine cell (vGluT3) to a wide-field polyaxonal amacrine cell (PAS4/5). Distal tips of vGluT3 cell dendrites made selective glycinergic (but not glutamatergic) synapses onto PAS4/5 dendrites to provide a center-inhibitory, surround-disinhibitory drive that helps PAS4/5 cells build a suppressed-by-contrast (sbc) receptive field, which is a unique and fundamental trigger feature previously found only in a small population of ganglion cells. The finding of this trigger feature in a circuit upstream to ganglion cells suggests that the sbc form of visual computation occurs more widely in the retina than previously believed and shapes visual processing in multiple downstream circuits in multiple ways. We also identified two different subpopulations of PAS4/5 cells based on their differential connectivity with vGluT3 cells and their distinct receptive-field and luminance-encoding characteristics. Moreover, our results revealed a form of crosstalk between small-field and large-field amacrine cell circuits, which provides a mechanism for feature-specific local (<150 µm) control of global (>1 mm) retinal activity.

Photoreceptive Ganglion Cells Drive Circuits for Local Inhibition in the Mouse Retina.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) exhibit melanopsin-dependent light responses that persist in the absence of rod and cone photoreceptor-mediated input. In addition to signaling anterogradely to the brain, ipRGCs signal retrogradely to intraretinal circuitry via gap junction-mediated electrical synapses with amacrine cells (ACs). However, the targets and functions of these intraretinal signals remain largely unknown. Here, in mice of both sexes, we identify circuitry that enables M5 ipRGCs to locally inhibit retinal neurons via electrical synapses with a nonspiking GABAergic AC. During pharmacological blockade of rod- and cone-mediated input, whole-cell recordings of corticotropin-releasing hormone-expressing (CRH+) ACs reveal persistent visual responses that require both melanopsin expression and gap junctions. In the developing retina, ipRGC-mediated input to CRH+ ACs is weak or absent before eye opening, indicating a primary role for this input in the mature retina (i.e., in parallel with rod- and cone-mediated input). Among several ipRGC types, only M5 ipRGCs exhibit consistent anatomical and physiological coupling to CRH+ ACs. Optogenetic stimulation of local CRH+ ACs directly drives IPSCs in M4 and M5, but not M1-M3, ipRGCs. CRH+ ACs also inhibit M2 ipRGC-coupled spiking ACs, demonstrating direct interaction between discrete networks of ipRGC-coupled interneurons. Together, these results demonstrate a functional role for electrical synapses in translating ipRGC activity into feedforward and feedback inhibition of local retinal circuits.SIGNIFICANCE STATEMENT Melanopsin directly generates light responses in intrinsically photosensitive retinal ganglion cells (ipRGCs). Through gap junction-mediated electrical synapses with retinal interneurons, these uniquely photoreceptive RGCs may also influence the activity and output of neuronal circuits within the retina. Here, we identified and studied an electrical synaptic circuit that, in principle, could couple ipRGC activity to the chemical output of an identified retinal interneuron. Specifically, we found that M5 ipRGCs form electrical synapses with corticotropin-releasing hormone-expressing amacrine cells, which locally release GABA to inhibit specific RGC types. Thus, ipRGCs are poised to influence the output of diverse retinal circuits via electrical synapses with interneurons.

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.

Zfhx3 modulates retinal sensitivity and circadian responses to light.

Mutations in transcription factors often exhibit pleiotropic effects related to their complex expression patterns and multiple regulatory targets. One such mutation in the zinc finger homeobox 3 (ZFHX3) transcription factor, short circuit (Sci, Zfhx3Sci/+ ), is associated with significant circadian deficits in mice. However, given evidence of its retinal expression, we set out to establish the effects of the mutation on retinal function using molecular, cellular, behavioral and electrophysiological measures. Immunohistochemistry confirms the expression of ZFHX3 in multiple retinal cell types, including GABAergic amacrine cells and retinal ganglion cells including intrinsically photosensitive retinal ganglion cells (ipRGCs). Zfhx3Sci/+ mutants display reduced light responsiveness in locomotor activity and circadian entrainment, relatively normal electroretinogram and optomotor responses but exhibit an unexpected pupillary reflex phenotype with markedly increased sensitivity. Furthermore, multiple electrode array recordings of Zfhx3Sci/+ retina show an increased sensitivity of ipRGC light responses.

Selective glycinergic input from vGluT3 amacrine cells confers a suppressed-by-contrast trigger feature in a subtype of M1 ipRGCs in the mouse retina.

KEY POINTS: M1 intrinsically photosensitive retinal ganglion cells (ipRGCs) are known to encode absolute light intensity (irradiance) for non-image-forming visual functions (subconscious vision), such as circadian photoentrainment and the pupillary light reflex. It remains unclear how M1 cells respond to relative light intensity (contrast) and patterned visual signals. The present study identified a special form of contrast sensitivity (suppressed-by-contrast) in M1 cells, suggesting a role of patterned visual signals in regulating non-image-forming vision and a potential role of M1 ipRGCs in encoding image-forming visual cues. The study also uncovered a synaptic mechanism and a retinal circuit mediated by vesicular glutamate transporter 3 (vGluT3) amacrine cells that underlie the suppressed-by-contrast response of M1 cells. M1 ipRGC subtypes (M1a and M1b) were revealed that are distinguishable based on synaptic connectivity with vGluT3 amacrine cells, receptive field properties, intrinsic photo sensitivity and membrane excitability, and morphological features, suggesting a division of visual tasks among discrete M1 subpopulations. ABSTRACT: The M1 type ipRGC (intrinsically photosensitive retinal ganglion cell) is known to encode ambient light signals for non-image-forming visual functions such as circadian photo-entrainment and the pupillary light reflex. Here, we report that a subpopulation of M1 cells (M1a) in the mouse retina possess the suppressed-by-contrast (sbc) trigger feature that is a receptive field property previously found only in ganglion cells mediating image-forming vision. Using optogenetics and the dual patch clamp technique, we found that vesicular glutamate transporter 3 (vGluT3) (vGluT3) amacrine cells make glycinergic, but not glutamatergic, synapses specifically onto M1a cells. The spatiotemporal and pharmacological properties of visually evoked responses of M1a cells closely matched the receptive field characteristics of vGluT3 cells, suggesting a major role of the vGluT3 amacrine cell input in shaping the sbc trigger feature of M1a cells. We found that the other subpopulation of M1 cells (M1b), which did not receive a direct vGluT3 cell input, lacked the sbc trigger feature, being distinctively different from M1a cells in intrinsic photo responses, membrane excitability, receptive-field characteristics and morphological features. Together, the results reveal a retinal circuit that uses the sbc trigger feature to regulate irradiance coding and potentially send image-forming cues to non-image-forming visual centres in the brain.

Structure and function of the gap junctional network of photoreceptive ganglion cells.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) signal not only anterogradely to drive behavioral responses, but also retrogradely to some amacrine interneurons to modulate retinal physiology. We previously found that all displaced amacrine cells with spiking, tonic excitatory photoresponses receive gap-junction input from ipRGCs, but the connectivity patterns and functional roles of ipRGC-amacrine coupling remained largely unknown. Here, we injected PoPro1 fluorescent tracer into all six types of mouse ipRGCs to identify coupled amacrine cells, and analyzed the latter's morphological and electrophysiological properties. We also examined how genetically disrupting ipRGC-amacrine coupling affected ipRGC photoresponses. Results showed that ipRGCs couple with not just ON- and ON/OFF-stratified amacrine cells in the ganglion-cell layer as previously reported, but also OFF-stratified amacrine cells in both ganglion-cell and inner nuclear layers. M1- and M3-type ipRGCs couple mainly with ON/OFF-stratified amacrine cells, whereas the other ipRGC types couple almost exclusively with ON-stratified ones. ipRGCs transmit melanopsin-based light responses to at least 93% of the coupled amacrine cells. Some of the ON-stratifying ipRGC-coupled amacrine cells exhibit transient hyperpolarizing light responses. We detected bidirectional electrical transmission between an ipRGC and a coupled amacrine cell, although transmission was asymmetric for this particular cell pair, favoring the ipRGC-to-amacrine direction. We also observed electrical transmission between two amacrine cells coupled to the same ipRGC. In both scenarios of coupling, the coupled cells often spiked synchronously. While ipRGC-amacrine coupling somewhat reduces the peak firing rates of ipRGCs' intrinsic melanopsin-based photoresponses, it renders these responses more sustained and longer-lasting. In summary, ipRGCs' gap junctional network involves more amacrine cell types and plays more roles than previously appreciated.

Functional connexin35 increased in the myopic chicken retina.

Our previous research showed that increased phosphorylation of connexin (Cx)36 indicated extended  coupling of AII amacrine cells (ACs) in the rod-dominant mouse myopic retina. This research will determine whether phosphorylation at serine 276 of Cx35-containing gap junctions increased in the myopic chicken, whose retina is cone-dominant. Refractive errors and ocular biometric dimensions of 7-days-old chickens were determined following 12 h and 7 days induction of myopia by a -10D lens. The expression pattern and size of Cx35-positive plaques were examined in the early (12 h) and compensated stages (7 days) of lens-induced myopia (LIM). At the same time, phosphorylation at serine 276 (functional assay) of Cx35 in strata 5 (S5) of the inner plexiform layer was investigated. The axial length of the 7 days LIM eyes was significantly longer than that of non-LIM controls (P < 0.05). Anti-phospho-Ser276 (Ser276-P)-labeled plaques were significantly increased in LIM retinas at both 12 h and 7 days. The density of Ser276-P of Cx35 was observed to increase after 12 h LIM. In the meanwhile, the areas of existing Cx35 plaques did not change. As there was more phosphorylation of connexin35 at Ser276 at both the early and late stages (12 h) and 7 days of LIM chicken retinal activity, the coupling with ACs could be increased in myopia development of the cone-dominated chicken retina.

Visual Disfunction due to the Selective Effect of Glutamate Agonists on Retinal Cells.

One of the causes of nervous system degeneration is an excess of glutamate released upon several diseases. Glutamate analogs, like N-methyl-DL-aspartate (NMDA) and kainic acid (KA), have been shown to induce experimental retinal neurotoxicity. Previous results have shown that NMDA/KA neurotoxicity induces significant changes in the full field electroretinogram response, a thinning on the inner retinal layers, and retinal ganglion cell death. However, not all types of retinal neurons experience the same degree of injury in response to the excitotoxic stimulus. The goal of the present work is to address the effect of intraocular injection of different doses of NMDA/KA on the structure and function of several types of retinal cells and their functionality. To globally analyze the effect of glutamate receptor activation in the retina after the intraocular injection of excitotoxic agents, a combination of histological, electrophysiological, and functional tools has been employed to assess the changes in the retinal structure and function. Retinal excitotoxicity caused by the intraocular injection of a mixture of NMDA/KA causes a harmful effect characterized by a great loss of bipolar, amacrine, and retinal ganglion cells, as well as the degeneration of the inner retina. This process leads to a loss of retinal cell functionality characterized by an impairment of light sensitivity and visual acuity, with a strong effect on the retinal OFF pathway. The structural and functional injury suffered by the retina suggests the importance of the glutamate receptors expressed by different types of retinal cells. The effect of glutamate agonists on the OFF pathway represents one of the main findings of the study, as the evaluation of the retinal lesions caused by excitotoxicity could be specifically explored using tests that evaluate the OFF pathway.

The effects of early diabetes on inner retinal neurons.

Diabetic retinopathy is now well understood as a neurovascular disease. Significant deficits early in diabetes are found in the inner retina that consists of bipolar cells that receive inputs from rod and cone photoreceptors, ganglion cells that receive inputs from bipolar cells, and amacrine cells that modulate these connections. These functional deficits can be measured in vivo in diabetic humans and animal models using the electroretinogram (ERG) and behavioral visual testing. Early effects of diabetes on both the human and animal model ERGs are changes to the oscillatory potentials that suggest dysfunctional communication between amacrine cells and bipolar cells as well as ERG measures that suggest ganglion cell dysfunction. These are coupled with changes in contrast sensitivity that suggest inner retinal changes. Mechanistic in vitro neuronal studies have suggested that these inner retinal changes are due to decreased inhibition in the retina, potentially due to decreased gamma aminobutyric acid (GABA) release, increased glutamate release, and increased excitation of retinal ganglion cells. Inner retinal deficits in dopamine levels have also been observed that can be reversed to limit inner retinal damage. Inner retinal targets present a promising new avenue for therapies for early-stage diabetic eye disease.

Inhibitory components of retinal bipolar cell receptive fields are differentially modulated by dopamine D1 receptors.

During adaptation to an increase in environmental luminance, retinal signaling adjustments are mediated by the neuromodulator dopamine. Retinal dopamine is released with light and can affect center-surround receptive fields, the coupling state between neurons, and inhibitory pathways through inhibitory receptors and neurotransmitter release. While the inhibitory receptive field surround of bipolar cells becomes narrower and weaker during light adaptation, it is unknown how dopamine affects bipolar cell surrounds. If dopamine and light have similar effects, it would suggest that dopamine could be a mechanism for light-adapted changes. We tested the hypothesis that dopamine D1 receptor activation is sufficient to elicit the magnitude of light-adapted reductions in inhibitory bipolar cell surrounds. Surrounds were measured from OFF bipolar cells in dark-adapted mouse retinas while stimulating D1 receptors, which are located on bipolar, horizontal, and inhibitory amacrine cells. The D1 agonist SKF-38393 narrowed and weakened OFF bipolar cell inhibitory receptive fields but not to the same extent as with light adaptation. However, the receptive field surround reductions differed between the glycinergic and GABAergic components of the receptive field. GABAergic inhibitory strength was reduced only at the edges of the surround, while glycinergic inhibitory strength was reduced across the whole receptive field. These results expand the role of retinal dopamine to include modulation of bipolar cell receptive field surrounds. Additionally, our results suggest that D1 receptor pathways may be a mechanism for the light-adapted weakening of glycinergic surround inputs and the furthest wide-field GABAergic inputs to bipolar cells. However, remaining differences between light-adapted and D1 receptor-activated inhibition demonstrate that non-D1 receptor mechanisms are necessary to elicit the full effect of light adaptation on inhibitory surrounds.

Mobile zinc increases rapidly in the retina after optic nerve injury and regulates ganglion cell survival and optic nerve regeneration.

Retinal ganglion cells (RGCs), the projection neurons of the eye, cannot regenerate their axons once the optic nerve has been injured and soon begin to die. Whereas RGC death and regenerative failure are widely viewed as being cell-autonomous or influenced by various types of glia, we report here that the dysregulation of mobile zinc (Zn2+) in retinal interneurons is a primary factor. Within an hour after the optic nerve is injured, Zn2+ increases several-fold in retinal amacrine cell processes and continues to rise over the first day, then transfers slowly to RGCs via vesicular release. Zn2+ accumulation in amacrine cell processes involves the Zn2+ transporter protein ZnT-3, and deletion of slc30a3, the gene encoding ZnT-3, promotes RGC survival and axon regeneration. Intravitreal injection of Zn2+ chelators enables many RGCs to survive for months after nerve injury and regenerate axons, and enhances the prosurvival and regenerative effects of deleting the gene for phosphatase and tensin homolog (pten). Importantly, the therapeutic window for Zn2+ chelation extends for several days after nerve injury. These results show that retinal Zn2+ dysregulation is a major factor limiting the survival and regenerative capacity of injured RGCs, and point to Zn2+ chelation as a strategy to promote long-term RGC protection and enhance axon regeneration.

Local synaptic integration enables ON-OFF asymmetric and layer-specific visual information processing in vGluT3 amacrine cell dendrites.

A basic scheme of neuronal organization in the mammalian retina is the segregation of ON and OFF pathways in the inner plexiform layer (IPL), where glutamate is released from ON and OFF bipolar cell terminals in separate inner (ON) and outer (OFF) sublayers in response to light intensity increments and decrements, respectively. However, recent studies have found that vGluT3-expressing glutamatergic amacrine cells (GACs) generate ON-OFF somatic responses and release glutamate onto both ON and OFF ganglion cell types, raising the possibility of crossover excitation in violation of the canonical ON-OFF segregation scheme. To test this possibility, we recorded light-evoked Ca2+ responses from dendrites of individual GACs infected with GCaMP6s in mouse. Under two-photon imaging, a single GAC generated rectified local dendritic responses, showing ON-dominant responses in ON sublayers and OFF-dominant responses in OFF sublayers. This unexpected ON-OFF segregation within a small-field amacrine cell arose from local synaptic processing, mediated predominantly by synaptic inhibition. Multiple forms of synaptic inhibition compartmentalized the GAC dendritic tree and endowed all dendritic varicosities with a small-center, strong-surround receptive field, which varied in receptive field size and degree of ON-OFF asymmetry with IPL depth. The results reveal a form of short-range dendritic autonomy that enables a small-field, dual-transmitter amacrine cell to process diverse dendritic functions in a stratification level- and postsynaptic target-specific manner, while preserving the fundamental ON-OFF segregation scheme for parallel visual processing and high spatial resolution for small object motion and uniformity detection.

Dopamine-Dependent Sensitization of Rod Bipolar Cells by GABA Is Conveyed through Wide-Field Amacrine Cells.

The vertebrate retina has the remarkable ability to support visual function under conditions of limited illumination, including the processing of signals evoked by single photons. Dim-light vision is regulated by several adaptive mechanisms. The mechanism explored in this study is responsible for increasing the light sensitivity and operational range of rod bipolar cells, the retinal neurons operating immediately downstream of rod photoreceptors. This sensitization is achieved through the sustained dopamine-dependent GABA release from other retinal neurons. Our goals were to identify the cell type responsible for the GABA release and the site of its modulation by dopamine. Previous studies have suggested the involvement of amacrine and/or horizontal cells. We now demonstrate, using mice of both sexes, that horizontal cells do not participate in this mechanism. Instead, sustained GABA input is provided by a subpopulation of wide-field amacrine cells, which stimulate the GABAC receptors at rod bipolar cell axons. We also found that dopamine does not act directly on either of these cells. Rather, it suppresses inhibition imposed on these wide-field cells by another subpopulation of upstream GABAergic amacrine cells, thereby sustaining the GABAC receptor activation required for rod bipolar cell sensitization.SIGNIFICANCE STATEMENT The vertebrate retina has an exquisite ability to adjust information processing to ever-changing conditions of ambient illumination, from bright sunlight to single-photon counting under dim starlight. Operation under each of these functional regimes requires an engagement of specific adaptation mechanisms. Here, we describe a mechanism optimizing the performance of the dim-light channel of vision, which consists of sensitizing rod bipolar cells by a sustained GABAergic input originating from a population of wide-field amacrine cells. Wide-field amacrine cells span large segments of the retina, making them uniquely equipped to normalize and optimize response sensitivity across distant receptive fields and preclude any bias toward local light-intensity fluctuations.

Convergence and Divergence of CRH Amacrine Cells in Mouse Retinal Circuitry.

Inhibitory interneurons sculpt the outputs of excitatory circuits to expand the dynamic range of information processing. In mammalian retina, >30 types of amacrine cells provide lateral inhibition to vertical, excitatory bipolar cell circuits, but functional roles for only a few amacrine cells are well established. Here, we elucidate the function of corticotropin-releasing hormone (CRH)-expressing amacrine cells labeled in Cre-transgenic mice of either sex. CRH cells costratify with the ON alpha ganglion cell, a neuron highly sensitive to positive contrast. Electrophysiological and optogenetic analyses demonstrate that two CRH types (CRH-1 and CRH-3) make GABAergic synapses with ON alpha cells. CRH-1 cells signal via graded membrane potential changes, whereas CRH-3 cells fire action potentials. Both types show sustained ON-type responses to positive contrast over a range of stimulus conditions. Optogenetic control of transmission at CRH-1 synapses demonstrates that these synapses are tuned to low temporal frequencies, maintaining GABA release during fast hyperpolarizations during brief periods of negative contrast. CRH amacrine cell output is suppressed by prolonged negative contrast, when ON alpha ganglion cells continue to receive inhibitory input from converging OFF-pathway amacrine cells; the converging ON- and OFF-pathway inhibition balances tonic excitatory drive to ON alpha cells. Previously, it was demonstrated that CRH-1 cells inhibit firing by suppressed-by-contrast (SbC) ganglion cells during positive contrast. Therefore, divergent outputs of CRH-1 cells inhibit two ganglion cell types with opposite responses to positive contrast. The opposing responses of ON alpha and SbC ganglion cells are explained by differing excitation/inhibition balance in the two circuits.SIGNIFICANCE STATEMENT A goal of neuroscience research is to explain the function of neural circuits at the level of specific cell types. Here, we studied the function of specific types of inhibitory interneurons, corticotropin-releasing hormone (CRH) amacrine cells, in the mouse retina. Genetic tools were used to identify and manipulate CRH cells, which make GABAergic synapses with a well studied ganglion cell type, the ON alpha cell. CRH cells converge with other types of amacrine cells to tonically inhibit ON alpha cells and balance their high level of excitation. CRH cells diverge to different types of ganglion cell, the unique properties of which depend on their balance of excitation and inhibition.