Vertebrate Retina Research Articles

Formulation and Implementation of Nonlinear Integral Equations to Model Neural Dynamics Within the Vertebrate Retina

Existing computational models of the retina often compromise between the biophysical accuracy and a hardware-adaptable methodology of implementation. When compared to the current modes of vision restoration, algorithmic models often contain a greater correlation between stimuli and the affected neural network, but lack physical hardware practicality. Thus, if the present processing methods are adapted to complement very-large-scale circuit design techniques, it is anticipated that it will engender a more feasible approach to the physical construction of the artificial retina. The computational model presented in this research serves to provide a fast and accurate predictive model of the retina, a deeper understanding of neural responses to visual stimulation, and an architecture that can realistically be transformed into a hardware device. Traditionally, implicit (or semi-implicit) ordinary differential equations (OES) have been used for optimal speed and accuracy. We present a novel approach that requires the effective integration of different dynamical time scales within a unified framework of neural responses, where the rod, cone, amacrine, bipolar, and ganglion cells correspond to the implemented pathways. Furthermore, we show that adopting numerical integration can both accelerate retinal pathway simulations by more than 50% when compared with traditional ODE solvers in some cases, and prove to be a more realizable solution for the hardware implementation of predictive retinal models.

K+ Channels of Müller Glial Cells in Retinal Disorders.

Müller cell is the major type of glial cell in the vertebrate retina. Müller cells express various types of K+ channels, such as inwardly rectifying K+ (Kir) channels, big conductance Ca2+-activated K+ (BKCa) channels, delayed rectifier K+ channels (KDR), and transient A-type K+ channels. These K+ channels play important roles in maintaining physiological functions of Müller cells. Under some retinal pathological conditions, the changed expression and functions of K+ channels may contribute to retinal pathogenesis. In this article, we reviewed the physiological properties of K+ channels in retinal Müller cells and the functional changes of these channels in retinal disorders.

Plasticity of Retinal Gap Junctions: Roles in Synaptic Physiology and Disease.

Electrical synaptic transmission via gap junctions underlies direct and rapid neuronal communication in the central nervous system. The diversity of functional roles played by electrical synapses is perhaps best exemplified in the vertebrate retina, in which gap junctions are expressed by each of the five major neuronal types. These junctions are highly plastic; they are dynamically regulated by ambient illumination and circadian rhythms acting through light-activated neuromodulators. The networks formed by electrically coupled neurons provide plastic, reconfigurable circuits positioned to play key and diverse roles in the transmission and processing of visual information at every retinal level. Recent work indicates gap junctions also play a role in the progressive cell death and aberrant activity seen in various pathological conditions of the retina. Gap junctions thus form potential targets for novel neuroprotective therapies in the treatment of neurodegenerative retinal diseases such as glaucoma and ischemic retinopathies.

MicroRNAs in Retinal Development.

The small RNA regulatory molecules called microRNAs (miRNAs) play key roles in the development of most organisms. The expression of many different miRNAs has been described in the developing and mature vertebrate retina. The ability of miRNAs to regulate a diversity of messenger RNA targets allows them to have effects on many different developmental processes, but the functions of only a few miRNAs have been documented to date. Developmental transitions between cell states appear to be particularly sensitive to miRNA loss of function, as evidenced by specific miRNA knockdowns or from global perturbations in miRNA levels (e.g., Dicer deletion). However, we are still in only the very early stages of understanding the range of cellular functions miRNAs control during development.

Protective role of carotenoids in the visual cycle.

Exposure to light and accumulation of aberrant visual cycle by-products causes stress in the retina. The physical and chemical properties of carotenoids may provide protection against such scenario. These pigments exist in retinas of many vertebrates, including humans. However, the absence of carotenoids in mice, the preferred ophthalmologic animal model, hindered molecular and biochemical examination of the pigments' role in vision. We established a mouse model that accumulates significant amounts of carotenoids in the retina due to inactivating mutations in the Isx and Bco2 genes. We introduced a robust light damage protocol for the mouse retina using green (532 nm) and blue (405 nm) low-energy lasers. We observed that blue but not green laser light treatment triggered the formation of aberrant retinaldehyde isomers in the retina. The production of these visual cycle by-products was accompanied by morphologic damage in inferior parts of the mouse retina. Zeaxanthin supplementation of mice shielded retinoids from these photochemical modifications. These pigments also reduced the extent of the damage to the retina after the blue laser light insult. Thus, our study discovered a novel role of carotenoids in the visual cycle and indicated that vertebrates accumulate carotenoids to shield photoreceptors from short-wavelength light-induced damage.-Widjaja-Adhi, M. A. K., Ramkumar, S., von Lintig, J. Protective role of carotenoids in the visual cycle.

Electrophysiological Analysis of Retinal Oscillation in Normal and Degenerated Retina

The vertebrate retina is one of the most sophisticated parts of the nervous system. It comprises five classes of neurons and one glial type cell. During development, but prior to a vertebrate's eyes opening, retinal circuits are refined by endogenous neural activity. Characteristic patterns of activity, including oscillatory activity, occur in the normal retina, whereas distinctive alternative patterns occur in abnormal retinas. In this paper, we first describe the electrophysiological and spike sorting methods used to study retinal oscillations. Next, we describe the mechanisms and functions of oscillation in the normal retina. Finally, we characterize the distinctive oscillations and abnormal spontaneous activities in the degenerative retina.

Characterization of a morphogenetic furrow specific Gal4 driver in the developing Drosophila eye.

The ability to express a gene of interest in a spatio-temporal manner using Gal4-UAS system has allowed the use of Drosophila model to study various biological phenomenon. During Drosophila eye development, a synchronous wave of differentiation called Morphogenetic furrow (MF) initiates at the posterior margin resulting in differentiation of retinal neurons. This synchronous differentiation is also observed in the differentiating retina of vertebrates. Since MF is highly dynamic, it can serve as an excellent model to study patterning and differentiation. However, there are not any Gal4 drivers available to observe the gain- of- function or loss- of- function of a gene specifically along the dynamic MF. The decapentaplegic (dpp) gene encodes a secreted protein of the transforming growth factor-beta (TGF-beta) superfamily that expresses at the posterior margin and then moves with the MF. However, unlike the MF associated pattern of dpp gene expression, the targeted dpp-Gal4 driver expression is restricted to the posterior margin of the developing eye disc. We screened GMR lines harboring regulatory regions of dpp fused with Gal4 coding region to identify MF specific enhancer of dpp using a GFP reporter gene. We employed immuno-histochemical approaches to detect gene expression. The rationale was that GFP reporter expression will correspond to the dpp expression domain in the developing eye. We identified two new dpp-Gal4 lines, viz., GMR17E04-Gal4 and GMR18D08-Gal4 that carry sequences from first intron region of dpp gene. GMR17E04-Gal4 drives expression along the MF during development and later in the entire pupal retina whereas GMR18D08-Gal4 drives expression of GFP transgene in the entire developing eye disc, which later drives expression only in the ventral half of the pupal retina. Thus, GMR18D08-Gal4 will serve as a new reagent for targeting gene expression in the ventral half of the pupal retina. We compared misexpression phenotypes of Wg, a negative regulator of eye development, using GMR17E04-Gal4, GMR18D08-Gal4 with existing dpp-Gal4 driver. The eye phenotypes generated by using our newly identified MF specific driver are not similar to the ones generated by existing dpp-Gal4 driver. It suggests that misexpression studies along MF needs revisiting using the new Gal4 drivers generated in our studies.

Localization of cholecystokinin in the zebrafish retina from larval to adult stage

The peptide hormone cholecistokinin (CCK) plays a key role in the central and peripheral nervous system. It is known to be involved in the digestive physiology and in the regulation of food intake. Moreover, the CCK expression has also been detected in the retina of different vertebrates, including fish, although its biological activity in this tissue remains to be elucidated. In literature no data are yet available about the CCK-immunoreactivity in the zebrafish retina during development. Therefore, the aim of the study was to investigate the distribution of sulfated cholecystokinin octapeptide (CCK8-S) as a well preserved form during evolution in the zebrafish retina from 3days post hatching (dph) until adult stage, using immunohistochemistry in order to elucidate the potential role of this protein in the development and maintenance of normal retinal homeostasis. The cellular distribution of CCK in the retina was similar from 3 dph to 40days post fertilization (dpf) when immunoreactivity was found in the photoreceptors layer, in the outer plexiform layer, in the inner plexiform layer and, to a lesser extent, in the ganglion cell layer (GCL). Immunohistochemical localization at 50 dpf as well as in the adult stage was observed in a subpopulation of amacrine cells in the proximal inner nuclear layer, in the inner plexiform layer, in displaced amacrine cells and in retinal ganglion cells in the GCL. Our results demonstrate for the first time the occurrence of CCK in the zebrafish retina from larval to adult stage with a different pattern of distribution, suggesting different roles of CCK during retinal cells maturation.

Circadian Regulation of Mitochondrial Dynamics in Retinal Photoreceptors.

Energy expenditure and metabolism in the vertebrate retina are under circadian control, as we previously reported that the overall retinal ATP content and various signaling molecules related to metabolism display daily or circadian rhythms. Changes in the fission and fusion process of mitochondria, the major organelles producing ATP, in retinal photoreceptors are largely dependent on light exposure, but whether mitochondrial dynamics in photoreceptors and retinal neurons are under circadian control is not clear. Herein, we investigated the possible roles of circadian oscillators in regulating mitochondrial dynamics, mitophagy, and redox states in the chicken retina and mammalian photoreceptors. After entrainment to 12:12-h light-dark (LD) cycles for several days followed by free-running in constant darkness (DD), chicken embryonic retinas and cone-derived 661W cells were collected in either LD or DD at 6 different zeitgeber time (ZT) or circadian time (CT) points. The protein expression of mitochondrial dynamin-related protein 1 (DRP1), mitofusin 2 (MFN2), and PTEN-induced putative kinase 1 (PINK1) displayed daily rhythms, but only DRP1 was under circadian control in the chicken retinas and cultured 661W cells. In addition, cultured chicken retinal cells responded to acute oxidative stress differently from 661W cells. Using pMitoTimer as a mitochondrial redox indicator, we found that the mitochondrial redox states were more affected by light exposure than regulated by circadian oscillators. Thus, this study demonstrates that the influence of cyclic lights might outweigh the circadian regulation of complex mitochondrial dynamics in light-sensing retinal cells.

Parallel opsin switches in multiple cone types of the starry flounder retina: tuning visual pigment composition for a demersal life style

Variable expression of visual pigment proteins (opsins) in cone photoreceptors of the vertebrate retina is a primary determinant of vision plasticity. Switches in opsin expression or variable co-expression of opsins within differentiated cones have been documented for a few rodents and fishes, but the extent of photoreceptor types affected and potential functional significance are largely unknown. Here, we show that both single and double cones in the retina of a flatfish, the starry flounder (Platichthys stellatus), undergo visual pigment changes through opsin switches or variable opsin co-expression. As the post-metamorphic juvenile (i.e., the young asymmetric flatfish with both eyes on one side of the body) grows from ~5 g to ~196 g, some single cones and one member of unequal double cones switched from a visual pigment with maximum wavelength of absorbance, λmax, at shorter wavelengths (437 nm and 527 nm) to one with longer λmax (456 nm and 545 nm, respectively) whereas other cones had intermediate visual pigments (λmax at 445 nm or 536 nm) suggesting co-expression of two opsins. The shift toward longer wavelength absorbing visual pigments was in line with maximizing sensitivity to the restricted light spectrum at greater depths and achromatic detection of overhead targets.

Lrit1, a Retinal Transmembrane Protein, Regulates Selective Synapse Formation in Cone Photoreceptor Cells and Visual Acuity

In the vertebrate retina, cone photoreceptors play crucial roles in photopic vision by transmitting light-evoked signals to ON- and/or OFF-bipolar cells. However, the mechanisms underlying selective synapse formation in the cone photoreceptor pathway remain poorly understood. Here, we found that Lrit1, a leucine-rich transmembrane protein, localizes to the photoreceptor synaptic terminal and regulates the synaptic connection between cone photoreceptors and cone ON-bipolar cells. Lrit1-deficient retinas exhibit an aberrant morphology of cone photoreceptor pedicles, as well as an impairment of signal transmission from cone photoreceptors to cone ON-bipolar cells. Furthermore, we demonstrated that Lrit1 interacts with Frmpd2, a photoreceptor scaffold protein, and with mGluR6, an ON-bipolar cell-specific glutamate receptor. Additionally, Lrit1-null mice showed visual acuity impairments in their optokinetic responses. These results suggest that the Frmpd2-Lrit1-mGluR6 axis regulates selective synapse formation in cone photoreceptors and is essential for normal visual function.

Activation of retinal glial (Müller) cells by extracellular ATP induces pronounced increases in extracellular H+ flux.

Small alterations in extracellular acidity are potentially important modulators of neuronal signaling within the vertebrate retina. Here we report a novel extracellular acidification mechanism mediated by glial cells in the retina. Using self-referencing H+-selective microelectrodes to measure extracellular H+ fluxes, we show that activation of retinal Müller (glial) cells of the tiger salamander by micromolar concentrations of extracellular ATP induces a pronounced extracellular H+ flux independent of bicarbonate transport. ADP, UTP and the non-hydrolyzable analog ATPγs at micromolar concentrations were also potent stimulators of extracellular H+ fluxes, but adenosine was not. The extracellular H+ fluxes induced by ATP were mimicked by the P2Y1 agonist MRS 2365 and were significantly reduced by the P2 receptor blockers suramin and PPADS, suggesting activation of P2Y receptors. Bath-applied ATP induced an intracellular rise in calcium in Müller cells; both the calcium rise and the extracellular H+ fluxes were significantly attenuated when calcium re-loading into the endoplasmic reticulum was inhibited by thapsigargin and when the PLC-IP3 signaling pathway was disrupted with 2-APB and U73122. The anion transport inhibitor DIDS also markedly reduced the ATP-induced increase in H+ flux while SITS had no effect. ATP-induced H+ fluxes were also observed from Müller cells isolated from human, rat, monkey, skate and lamprey retinae, suggesting a highly evolutionarily conserved mechanism of potential general importance. Extracellular ATP also induced significant increases in extracellular H+ flux at the level of both the outer and inner plexiform layers in retinal slices of tiger salamander which was significantly reduced by suramin and PPADS. We suggest that the novel H+ flux mediated by ATP-activation of Müller cells and of other glia as well may be a key mechanism modulating neuronal signaling in the vertebrate retina and throughout the brain.

Amphioxus, motion detection, and the evolutionary origin of the vertebrate retinotectal map

The axonal projection from the retina to the optic tectum maps visual information isomorphically from one to the other and serves as a model for the development of sensory maps more generally in the vertebrate brain. How or why this connection evolved is not known, nor why the midbrain is so important to the processing of visual information. Amphioxus is potentially informative here because its eye homolog, the frontal eye, also has a neural connection to a region of the brain now known to be homologous with the caudal diencephalon and midbrain. The frontal eye has only a one-dimensional receptor array, but simple alterations to the pattern and plane of cell division would have been sufficient to generate a structure more like the vertebrate retina. Accounting for the retinotectal map poses more of a problem. The hypothesis developed here is that this is best explained as a consequence of a prior association between the roof of the anterior nerve cord and an array of rhabdomeric photoreceptors, homologous with the Joseph cells of amphioxus, that were used by the common ancestor of amphioxus and vertebrates for detecting moving shadows. Hence, a rudimentary tectal map could have been present before the evolution of image-forming eyes and been coopted by them secondarily. Assuming the orientation of this map was fixed from the start relative to the external world, its retinal counterpart would have had to adjust to this to accommodate the image reversal that accompanies the conversion of a flat receptor array to a camera-type eye. Exploring this hypothesis further will require more information than is currently available on the Joseph cells, especially as to where and how their neural output is processed.

Novel Animal Model of Crumbs-Dependent Progressive Retinal Degeneration That Targets Specific Cone Subtypes.

PurposeHuman Crb1 is implicated in some forms of retinal degeneration, suggesting a role in photoreceptor maintenance. Multiple Crumbs (Crb) polarity genes are expressed in vertebrate retina, although their functional roles are not well understood. To gain further insight into Crb and photoreceptor maintenance, we compared retinal cell densities between wild-type and Tg(RH2-2:Crb2b-sfEX/RH2-2:GFP)pt108b transgenic zebrafish, in which the extracellular domain of Crb2b-short form (Crb2b-sfEX) is expressed in the retina as a secreted protein, which disrupts the planar organization of RGB cones (red, green, and blue) by interfering with Crb2a/2b-based cone-cone adhesion.MethodsWe used standard morphometric techniques to assess age-related changes in retinal cell densities in adult zebrafish (3 to 27 months old), and to assess effects of the Crb2b-sfEX transgene on retinal structure and photoreceptor densities. Linear cell densities were measured in all retinal layers in radial sections with JB4-Feulgen histology. Planar (surface) densities of cones were determined in retinal flat-mounts. Cell counts from wild-type and pt108b transgenic fish were compared with both a “photoreceptor maintenance index” and statistical analysis of cell counts.ResultsAge-related changes in retinal cell linear densities and cone photoreceptor planar densities in wild-type adult zebrafish provided a baseline for analysis. Expression of Crb2b-sfEX caused progressive and selective degeneration of RGB cones, but had no effect on ultraviolet-sensitive (UV) cones, and increased numbers of rod photoreceptors.ConclusionsThese differential responses of RGB cones, UV cones, and rods to sustained exposure to Crb2b-sfEX suggest that Crb-based photoreceptor maintenance mechanisms are highly selective.

Neuronal Migration and Lamination in the Vertebrate Retina.

In the retina, like in most other brain regions, developing neurons are arranged into distinct layers giving the mature tissue its stratified appearance. This process needs to be highly controlled and orchestrated, as neuronal layering defects lead to impaired retinal function. To achieve successful neuronal layering and lamination in the retina and beyond, three main developmental steps need to be executed: First, the correct type of neuron has to be generated at a precise developmental time. Second, as most retinal neurons are born away from the position at which they later function, newborn neurons have to move to their final layer within the developing tissue, a process also termed neuronal lamination. Third, these neurons need to connect to their correct synaptic partners. Here, we discuss neuronal migration and lamination in the vertebrate retina and summarize our knowledge on these aspects of retinal development. We give an overview of how lamination emerges and discuss the different modes of neuronal translocation that occur during retinogenesis and what we know about the cell biological machineries driving them. In addition, retinal mosaics and their importance for correct retinal function are examined. We close by stating the open questions and future directions in this exciting field.

Simulator Development and Analysis for Signal Flow Pathway in Vertebrate Retina

Simulator Development and Analysis for Signal Flow Pathway in Vertebrate Retina

Molecular Landscapes of Embryonic and Post-Embryonic Neurogenesis in the Vertebrate Retina

Zebrafish retinas, undergoing life-long neurogenesis, provide an integrating model to resolve molecular landscapes of embryonic and post-embryonic neurogenesis. By single-cell RNAseq of zebrafish embryonic retinal progenitors and post-embryonic ciliary marginal zone (CMZ) progenitors, we define 7 distinguished states, 4 independent states (S1/S3/S5/S7) separated by 3 transitional states (S2/S4/S6), that develop over time and reveal aspects of lineage evolution. Genes that define earlier states are more critical for making retinas and post-embryonic CMZ, while genes that define later states are more critical for specifying individual cell fates. Remarkably, transitional and independent states expressed different cell-cycle phase genes, suggesting state transition relies on cell cycle. Intriguingly, similar analysis CMZ cells revealed similar set of states, suggesting embryonic and post-embryonic retinogensis conform to the same molecular program, although some interesting differences in rod genesis. Thus, our findings exemplify molecular landscapes of embryonic and post-embryonic neurogenesis in a vertebrate CNS tissue.

TRPV4 Does Not Regulate the Distal Retinal Light Response.

The transient receptor potential vanilloid isoform 4 (TRPV4) functions as polymodal transducer of swelling, heat, stretch, and lipid metabolites, is widely expressed across sensory tissues, and has been implicated in pressure sensing in vertebrate retinas. Although TRPV4 knockout mice exhibit a variety of mechanosensory, nociceptive, and thermo- and osmoregulatory phenotypes, it is not known whether the transmission of light-induced signals in the eye is affected by the loss of TRPV4. We utilized field potentials, a measure of rod and cone signaling, to determine whether TRPV4 impacts on the generation and/or transmission of the photoreceptor light response and neurotransmission. Luminance intensity-response relationships were acquired in anesthetized wild-type and TRPV4-/- mice and evaluated for peak amplitude and implicit time under scotopic and photopic conditions. We found that the morphology of the outer retina is unaffected by the ablation of the Trpv4 gene. Calcium imaging of dissociated Müller glia showed that selective TRPV4 stimulation induces oscillatory calcium signals in adjacent rods. However, no differences in scotopic or photopic light-evoked signaling in the distal retina were observed in TRPV4-/- eyes, suggesting that TRPV4 signaling in healthy Müller cells does not modulate the transmission of light-evoked signals at rod and cone synapses.

The inverted retina and the evolution of vertebrates: an evo-devo perspective

The inverted retina is a basic characteristic of the vertebrate eye. This implies that vertebrates must have a common ancestor with an inverted retina. Of the two groups of chordates, cephalochordates have an inverted retina and urochordates a direct retina. Surprisingly, recent genetics studies favor urochordates as the closest ancestor to vertebrates. The evolution of increasingly complex organs such as the eye implies not only tissular but also structural modifications at the organ level. How these configurational modifications give rise to a functional eye at any step is still subject to debate and speculation. Here we propose an orderly sequence of phylogenetic events that closely follows the sequence of developmental eye formation in extant vertebrates. The progressive structural complexity has been clearly recorded during vertebrate development at the period of organogenesis. Matching the chain of increasing eye complexity in Mollusca that leads to the bicameral eye of the octopus and the developmental sequence in vertebrates, we delineate the parallel evolution of the two-chambered eye of vertebrates starting with an early ectodermal eye. This sequence allows for some interesting predictions regarding the eyes of not preserved intermediary species. The clue to understanding the inverted retina of vertebrates and the similarity between the sequence followed by Mollusca and chordates is the notion that the eye in both cases is an ectodermal structure, in contrast to an exclusively (de novo) neuroectodermal origin in the eye of vertebrates. This analysis places cephalochordates as the closest branch to vertebrates contrary to urochordates, claimed as a closer branch by some researchers that base their proposals in a genetic analysis.

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.