New treatment for amblyopia based on rules of synaptic plasticity: a randomized clinical trial.

Amblyopia resulting from early deprivation of vision or defocus in one eye reflects an imbalance of input from the eyes to the visual cortex. We tested the hypothesis that asynchronous stimulation of the two eyes might induce synaptic plasticity and rebalance input. Experiments on normal adults showed that repetitive brief exposure of grating stimuli, with the onset of each stimulus delayed by 8.3 ms in one eye, results in a shift in perceptual eye dominance. Clinical studies (Clinical trial registration number: ChiCTR2100049130), using popular 3D movies with similar asynchrony between the two eyes (amblyopic eye stimulated first) to treat anisometropic amblyopia, established that just 10.5 h of conditioning over <3 weeks produced improvement that met criteria for successful treatment. The benefits of asynchronous conditioning accumulate over 20-30 45 min sessions, and are maintained for at least 2 years. Finally, we demonstrate that asynchronous binocular treatment alone is more effective than patching only. This novel treatment is popular with children and is some 50 times more efficient than patching alone.

Intraocular Amphiregulin antibody and axial elongation in nonhuman primates.

Purpose: To examine the effect of intraocularly applied amphiregulin antibody on physiological axial elongation in young nonhuman primates. Methods: The experimental study included six male 12-months-old macaque nonhuman primates (body weight:2.46 ± 0.25kg;range:2.20-2.90kg). In the experimental group (n=3 animals), three intravitreal injections of amphiregulin antibody (100µg/50µl) were applied to the left eyes at intervals of 4-6 weeks, and injections of phosphate buffered solution (50µl) were applied to the right eyes. Three other animals were assigned to a blank control group. Results: During the study period of 23.6 weeks, axial length in the experimental group did not change in the left eyes (18.91 ± 0.37mm to 18.94 ± 0.67mm;P=0.90), while it linearly increased in the right eyes (18.87 ± 0.38mm to 19.24 ± 0.53mm;P=0.056) and in the control group (left eyes:19.15 ± 0.22mm to 19.48 ± 0.22mm;P=0.009; right eyes:19.17 ± 0.15 mm to 19.46 ± 0.23 mm;P=0.024). The interocular difference in axial elongation increased in the experimental group from -0.11 ± 0.12mm at 4 weeks after baseline to -0.34 ± 0.15mm at the study end, while in the control group, the interocular side difference did not change significantly (from 0.01 ± 0.10 mm to 0.03 ± 0.08 mm;P=0.38). The difference in the interocular difference in axial elongation between the two groups was significant at 8 weeks (P=0.01), 15 weeks (P=0.007), and at study end (P=0.02). The interocular difference in axial length correlated with the interocular difference in vitreous cavity length (standardized regression coefficient beta:0.85;P<0.001). The interocular axial length difference was inversely associated with the interocular refractive error difference (beta:-0.49;P<0.001). Conclusions: Intraocularly applied amphiregulin antibody (100µg) reduced the physiological ocular axial elongation in juvenile nonhuman primates.

A Visual Circuit Related to Habenula Underlies the Antidepressive Effects of Light Therapy.

Light plays a pivotal role in the regulation of affective behaviors. However, the precise circuits that mediate the impact of light on depressive-like behaviors are not well understood. Here, we show that light influences depressive-like behaviors through a disynaptic circuit linking the retina and the lateral habenula (LHb). Specifically, M4-type melanopsin-expressing retinal ganglion cells (RGCs) innervate GABA neurons in the thalamic ventral lateral geniculate nucleus and intergeniculate leaflet (vLGN/IGL), which in turn inhibit CaMKIIα neurons in the LHb. Specific activation of vLGN/IGL-projecting RGCs, activation of LHb-projecting vLGN/IGL neurons, or inhibition of postsynaptic LHb neurons is sufficient to decrease the depressive-like behaviors evoked by long-term exposure to aversive stimuli or chronic social defeat stress. Furthermore, we demonstrate that the antidepressive effects of light therapy require activation of the retina-vLGN/IGL-LHb pathway. These results reveal a dedicated retina-vLGN/IGL-LHb circuit that regulates depressive-like behaviors and provide a potential mechanistic explanation for light treatment of depression.

Morphological properties of medial amygdala-projecting retinal ganglion cells in the Mongolian gerbil.

The amygdala is a limbic structure that is involved in many brain functions, including emotion, learning and memory. It has been reported that melanopsin-expressing retinal ganglion cells (ipRGCs) innervate the medial amygdala (MeA). However, whether conventional RGCs (cRGCs) project to the MeA remains unknown. The goal of this study was to determine if cRGCs project to the MeA and to determine the morphological properties of MeA-projecting RGCs (MeA-RGCs). Retrogradely labeled RGCs in whole-mount retinas were intracellularly injected to reveal their dendritic morphologies. Immunohistochemical staining was performed to selectively label ipRGCs (MeA-ipRGCs) and cRGCs (MeA-cRGCs). The results showed that 95.7% of the retrogradely labeled cells were cRGCs and that the rest were ipRGCs. Specifically, MeA-cRGCs consist of two morphological types. The majority of them exhibit small but dense dendritic fields and diffuse ramification patterns as previously reported in RGB2 (95%), while the rest exhibit small but sparse dendritic branching patterns resembling those of RGB3 cells (5%). MeA-ipRGCs consist of M1 and M2 subtypes. The MeA-RGCs showed an even retinal distribution patterns. The soma and dendritic field sizes of the MeA-RGCs did not vary with eccentricity. In conclusion, the present results suggest that MeA-RGCs are structurally heterogeneous. These direct RGCs that input to the MeA could be important for regulating amygdala functions.

High glucose levels impact visual response properties of retinal ganglion cells in C57 mice-An in vitro physiological study.

This study investigated visual response properties of retinal ganglion cells (RGCs) under high glucose levels. Extracellular single-unit responses of RGCs from mouse retinas were recorded. And the eyecup was prepared as a flat mount in a recording chamber and superfused with Ames medium. The averaged RF size of the ON RGCs (34.1±2.9, n=14) was significantly smaller than the OFF RGCs under the HG (49.3±0.3, n=12) (P<0.0001) conditions. The same reduction pattern was also observed in the osmotic control group (HM) between ON and OFF RGCs (P<0.0001). The averaged luminance threshold (LT) of ON RGCs increased significantly under HG or HM (HG: P<0.0001; HM: P<0.0002). OFF RGCs exhibited a similar response pattern under the same conditions (HG: P<0.01; HM: P<0.0002). The averaged contrast gain of ON cells was significantly lower than that of OFF cells with the HM treatment (P<0.015, unpaired Student's t test). The averaged contrast gain of ON cells was significantly higher than OFF cells with the HG treatment (P<0.0001). The present results suggest that HG reduced receptive field center size, suppressed luminance threshold, and attenuated contrast gain of RGCs. The impact of HG on ON and OFF RGCs may be mediated via different mechanisms.

Morphological evaluation of retinal ganglion cells expressing the L132C/T159C ChR2 mutant transgene in young adult cynomolgus monkeys.

To characterize recombinant AAV2 (rAAV2)-mediated expression of L132C/T159C ChR2 mutant in retinal ganglion cells (RGCs) of young adult cynomolgus monkeys. rAAV2 vectors carrying a fusion construct of the ChR2 mutant and GFP (ChR2-GFP) were delivered to the vitreous chamber by intravitreal injection. Expression patterns of the ChR2 mutant in RGCs were examined by immunohistochemical methods three months after injection. The RNA-binding protein with multiple splicing (RBPMS) was used as an RGC specific marker to differentiate RGCs from other retinal neurons and non-neuronal cells. The numbers of RBPMS+ and GFP+ double-labeled RGCs in the central foveal varied with the eccentricity. The expression peaked within 100 µm from the edge of the foveola and drastically decreased to a single superficial RGC layer approximately 300 µm from the edge. On average, the ratio of the double-labeled RGCs versus RBPMS+ RGCs approached 0.32±0.15 (n=14 fields) at the central foveal region (0.1 to 0.53 mm). We observed that the ratio reached 0.78±0.16 (n=21 fields) at peripheral retinal locations (eccentricity >7 mm). This investigation demonstrates that RBPMS could serve as a valuable RGC specific marker for future investigations in this field.

A retinoraphe projection regulates serotonergic activity and looming-evoked defensive behaviour.

Animals promote their survival by avoiding rapidly approaching objects that indicate threats. In mice, looming-evoked defensive responses are triggered by the superior colliculus (SC) which receives direct retinal inputs. However, the specific neural circuits that begin in the retina and mediate this important behaviour remain unclear. Here we identify a subset of retinal ganglion cells (RGCs) that controls mouse looming-evoked defensive responses through axonal collaterals to the dorsal raphe nucleus (DRN) and SC. Looming signals transmitted by DRN-projecting RGCs activate DRN GABAergic neurons that in turn inhibit serotoninergic neurons. Moreover, activation of DRN serotoninergic neurons reduces looming-evoked defensive behaviours. Thus, a dedicated population of RGCs signals rapidly approaching visual threats and their input to the DRN controls a serotonergic self-gating mechanism that regulates innate defensive responses. Our study provides new insights into how the DRN and SC work in concert to extract and translate visual threats into defensive behavioural responses.

Effect of alpha lipoic acid on retinal ganglion cell survival in an optic nerve crush model.

PURPOSE: This study was conducted to determine whether alpha lipoic acid (ALA) promotes the survival of retinal ganglion cells (RGCs) in a rat model of optic nerve crush (ONC) injury and to investigate the neuroprotective mechanisms of ALA in the retina in this ONC injury model. METHODS: Adult male Sprague-Dawley rats (180-220 g) were subjected to ONC injury surgery. ALA (63 mg/kg) was injected intravenously 1 day before or after the ONC injury. Animals were euthanized after 10 days, and the number of ganglion cells positive for RNA-binding protein with multiple splicing (Rbpms), which is an RGC marker, were counted on the whole mount retinas. In addition, immunofluorescence and immunoblotting were performed to examine the localization and levels of erythropoietin receptor (EPOR) and neurotrophin-4/5 (NT4/5) in the retinas in all experimental groups. To determine whether the EPO/EPOR signaling pathway was involved in the ALA antioxidant pathway, the rats were subjected to ruxolitinib (INCB018424, 0.25 mg/kg, bid, intraperitoneal, i.p.) treatment after the animals were injected intravenously with ALA 1 day before ONC injury. RESULTS: The average number of Rbpms-positive cells/mm2 in the control group (sham-operated group), the ONC group, the ALA-ONC group, and the ONC-ALA group retinas was 2219±28, 418±8, 848±22, and 613±18/mm2, respectively. The ALA-ONC and ONC-ALA groups showed a statistically significantly increased RGC survival rate compared to the ONC group. There were statistical differences in the RGC survival rates between the ALA-ONC (39%) and ONC-ALA groups (28%; p<0.05). Immunofluorescent labeling showed that EPOR and NT4/5 expression was significant in the retinal ganglion cell layer (GCL). At the same time, western blot analysis revealed that ALA induced upregulation of EPOR protein and NT4/5 protein expression in the retina after ONC injury. However, INCB018424 reversed the protective effects of ALA on the ONC retinas. CONCLUSIONS: ALA has neuroprotective effects on RGCs after ONC injury. Moreover, prophylactic administration of ALA may have a stronger neuroprotective effect against ONC-induced damage. Based on these data, we also conclude that the endogenous EPO/EPOR signaling pathway may contribute to the protective effects of ALA in the retina after ONC injury.

Protective Effect of ALA in Crushed Optic Nerve Cat Retinal Ganglion Cells Using a New Marker RBPMS.

In this study we first sought to determine whether RNA-binding protein with multiple splicing (RBPMS) can serve as a specific marker for cat retina ganglion cells (RGCs) using retrograde labeling and immunohistochemistry staining. RBPM was then used as an RGC marker to study RGC survival after optic nerve crush (ONC) and alpha-lipoic acid (ALA) treatment in cats. ALA treatment yielded a peak density of RBPMS-alpha cells within the peak isodensity zone (>60/mm2) which did not differ from ONC retinas. The area within the zone was significantly enlarged (control: 2.3%, ONC: 0.06%, ONC+ALA: 0.1%). As for the 10-21/mm2 zone, ALA treatment resulted in a significant increase in area (control: 34.5%, ONC: 12.1%, ONC+ALA: 35.9%). ALA can alleviate crush-induced RGC injury.

ON and OFF retinal ganglion cells differentially regulate serotonergic and GABAergic activity in the dorsal raphe nucleus.

The dorsal raphe nucleus (DRN), the major source of serotonergic input to the forebrain, receives excitatory input from the retina that can modulate serotonin levels and depressive-like behavior. In the Mongolian gerbil, retinal ganglion cells (RGCs) with alpha-like morphological and Y-like physiological properties innervate the DRN with ON DRN-projecting RGCs out numbering OFF DRN-projecting RGCs. The DRN neurons targeted by ON and OFF RGCs are unknown. To explore retino-raphe anatomical organization, retinal afferents labeled with Cholera toxin B were examined for association with the postsynaptic protein PSD-95. Synaptic associations between retinal afferents and DRN serotonergic and GABAergic neurons were observed. To explore retino-raphe functional organization, light-evoked c-fos expression was examined. Light significantly increased the number of DRN serotonergic and GABAergic cells expressing c-Fos. When ON RGCs were rendered silent while enhancing the firing rate of OFF RGCs, c-Fos expression was greatly increased in DRN serotonergic neurons suggesting that OFF DRN-projecting RGCs predominately activate serotonergic neurons whereas ON DRN-projecting RGCs mainly target GABAergic neurons. Direct glutamatergic retinal input to DRN 5-HT neurons contributes to the complex excitatory drive regulating these cells. Light, via the retinoraphe pathway can modify DRN 5-HT neuron activity which may play a role in modulating affective behavior.

Effects of Tauroursodeoxycholic Acid and Alpha-Lipoic-Acid on the Visual Response Properties of Cat Retinal Ganglion Cells: An In Vitro Study.

PURPOSE: To investigate the effects of tauroursodeoxycholic acid (TUDCA) and alpha-lipoic-acid (ALA) on the visual response properties of cat retinal ganglion cells (RGCs) in wholemount retinas. METHODS: Young adult cats were divided into three groups: control, ALA, and TUDCA. In vitro single-unit extracellular recordings were performed on wholemount retinas to objectively evaluate the visual response properties of RGCs prior and post to antioxidant treatment. The visual responses properties of RGCs, including receptive field size, luminance threshold, and contrast sensitivity, were collected online and analyzed off-line with Axon Pclamp9. RESULTS: Most of the RF sizes were larger than those plotted prior to the 60 minutes dark adaptation. The luminance threshold was elevated in the control group (no treatment) but reduced post ALA treatment and significantly reduced post TUDCA treatment. The contrast threshold was significantly elevated in the control group (no treatment) and clearly elevated post ALA treatment but effectively sustained post TUCDA treatment. CONCLUSIONS: Retinal neurocircuitry deteriorates in wholemount retinas, resulting in abnormal visual response properties in RGCs. Alpha-lipoic-acid and TUDCA exerted beneficial neuroprotective effects by activating the antioxidant pathway, partially restoring the functionality of retinal neurocircuitry and significantly improving the visual response properties of RGCs. However, TUDCA appears to be more effective than ALA in reducing irradiance thresholds and improving contrast sensitivity.

Dorsal raphe nucleus projecting retinal ganglion cells: Why Y cells?

Retinal ganglion Y (alpha) cells are found in retinas ranging from frogs to mice to primates. The highly conserved nature of the large, fast conducting retinal Y cell is a testament to its fundamental task, although precisely what this task is remained ill-defined. The recent discovery that Y-alpha retinal ganglion cells send axon collaterals to the serotonergic dorsal raphe nucleus (DRN) in addition to the lateral geniculate nucleus (LGN), medial interlaminar nucleus (MIN), pretectum and the superior colliculus (SC) has offered new insights into the important survival tasks performed by these cells with highly branched axons. We propose that in addition to its role in visual perception, the Y-alpha retinal ganglion cell provides concurrent signals via axon collaterals to the DRN, the major source of serotonergic afferents to the forebrain, to dramatically inhibit 5-HT activity during orientation or alerting/escape responses, which dis-facilitates ongoing tonic motor activity while dis-inhibiting sensory information processing throughout the visual system. The new data provide a fresh view of these evolutionarily old retinal ganglion cells.

The Dorsal Raphe Nucleus Receives Afferents From Alpha-Like Retinal Ganglion Cells and Intrinsically Photosensitive Retinal Ganglion Cells in the Rat.

PURPOSE: A retinal projection into the dorsal raphe nucleus (DRN), namely, the retino-raphe projection, exists in many species. The rat is one of several species in which a retino-raphe projection has been described; however, the retinal ganglion cell (RGC) types that contribute to this pathway are unknown. METHODS: Retrograde tracing via cholera toxin subunit B (CTB) was used to reveal DRN-projecting RGCs in rats, combined with intracellular injection in vitro, melanopsin immunostaining in whole-mounted retinas, and serotonin immunostaining to define the DRN. We modified methods of CTB injection into DRN used previously in order to avoid possible contamination with other retinorecipient regions, particularly the superior colliculus (SC). RESULTS: The majority of DRN-projecting RGCs showed alpha-like morphology, and some CTB-positive RGCs were colabeled with melanopsin. Approximately 80% of the total population of CTB-labeled DRN-projecting RGCs was alpha-like cells including ON alpha cells and OFF alpha cells; these alpha-like cells were melanopsin immunonegative. Approximately 10% of the remaining DRN-projecting RGCs were melanopsin immunopositive, in which the M1 subtype of intrinsically photosensitive retinal ganglion cell (ipRGC) provided the dominant projection of ipRGCs into DRN, with only few non-M1 ipRGCs involved. The DRN-projecting ipRGCs could be retrogradely labeled following tracer injection into all rostrocaudal aspects of the DRN. CONCLUSIONS: Both conventional RGCs with alpha-like morphology and melanopsin-expressing ipRGCs project into the rat DRN. Approximately 10% of DRN-projecting RGCs were colabeled with melanopsin, and the majority of these were the M1 subtype of ipRGCs. An ipRGC component of the retino-raphe projection may contribute to a sustained light-mediated modulation of DRN serotonin release.

Sulforaphane protects rodent retinas against ischemia-reperfusion injury through the activation of the Nrf2/HO-1 antioxidant pathway.

Retinal ischemia-reperfusion (I/R) injury induces oxidative stress, leukocyte infiltration, and neuronal cell death. Sulforaphane (SF), which can be obtained in cruciferous vegetables such as broccoli, exerts protective effects in response to oxidative stress in various tissues. These effects can be initiated through nuclear factor E2-related factor 2 (Nrf2)-mediated induction of heme oxygenase-1 (HO-1). This investigation was designed to elucidate the neural protective mechanisms of SF in the retinal I/R rat model. Animals were intraperitoneally (i.p.) injected with SF (12.5 mg/kg) or vehicle (corn oil) once a day for 7 consecutive days. Then, retinal I/R was made by elevating the intraocular pressure (IOP) to 130 mmHg for 1 h. To determine if HO-1 was involved in the Nrf2 antioxidant pathway, rats were subjected to protoporphyrin IX zinc (II) (ZnPP, 30 mg/kg, i.p.) treatments at 24 h before retinal ischemia. The neuroprotective effects of SF were assessed by determining the morphology of the retina, counting the infiltrating inflammatory cells and the surviving retinal ganglion cells (RGCs) and amacrine cells, and measuring apoptosis in the retinal layers. The expression of Nrf2 and HO-1 was studied by immunofluorescence analysis and western blotting. I/R induced a marked increase of ROS generation, caused pronounced inflammation, increased the apoptosis of RGCs and amacrine cells and caused the thinning of the inner retinal layer (IRL), and these effects were diminished or abolished by SF pretreatment. Meanwhile, SF pretreatment significantly elevated the nuclear accumulation of Nrf2 and the level of HO-1 expression in the I/R retinas; however, ZnPP reversed the protective effects of SF on I/R retinas. Together, we offer direct evidence that SF had protective effects on I/R retinas, which could be attributed, at least in part, to the activation of the Nrf2/HO-1 antioxidant pathway.

Dendritic morphology of caudal periaqueductal gray projecting retinal ganglion cells in Mongolian gerbil (Meriones unguiculatus).

In this study we investigated the morphological features of the caudal periaqueductal gray (cPAG)-projecting retinal ganglion cells (RGCs) in Mongolian gerbils using retrograde labeling, in vitro intracellular injection, confocal microscopy and three-dimensional reconstruction approaches. cPAG-projecting RGCs exhibit small somata (10-17 µm) and irregular dendritic fields (201-298 µm). Sizes of somata and dendritic fields do not show obvious variation at different distance from the optic disk (eccentricity). Dendrites are moderately branched. Morphological analysis (n = 23) reveals that cPAG-projecting RGCs ramified in sublamina a and b in the inner plexiform layer. These cells exhibit different stratification patterns based on the thickness of dendritic bands in sublaminas a and b: majority of analyzed cells (16 out of 23) have two bands of arborizations share similar thickness. The rest of analyzed cells (7 out of 23) exhibit thinner band in sublamina a than in sublamina b. Together, the present study suggests that cPAG of Mongolian gerbil could receive direct retinal inputs from two types of bistratified RGCs. Furthermore, a small subset of melanopsin-expressing RGCs (total 41 in 6 animals) is shown to innervate the rostral PAG (rPAG). Functional characteristics of these non-visual center projecting RGCs remain to be determined.

Activation of the Nrf2/HO-1 antioxidant pathway contributes to the protective effects of Lycium barbarum polysaccharides in the rodent retina after ischemia-reperfusion-induced damage.

Lycium barbarum polysaccharides (LBP), extracts from the wolfberries, are protective to retina after ischemia-reperfusion (I/R). The antioxidant response element (ARE)-mediated antioxidant pathway plays an important role in maintaining the redox status of the retina. Heme oxygenase-1 (HO-1), combined with potent AREs in its promoter, is a highly effective therapeutic target for the protection against neurodegenerative diseases, including I/R-induced retinal damage. The aim of our present study was to investigate whether the protective effect of LBP after I/R damage was mediated via activation of the Nrf2/HO-1-antioxidant pathway in the retina. Retinal I/R was induced by an increase in intraocular pressure to 130 mm Hg for 60 minutes. Prior to the induction of ischemia, rats were orally treated with either vehicle (PBS) or LBP (1 mg/kg) once a day for 1 week. For specific experiments, zinc protoporphyrin (ZnPP, 20 mg/kg), an HO-1 inhibitor, was intraperitoneally administered at 24 h prior to ischemia. The protective effects of LBP were evaluated by quantifying ganglion cell and amacrine cell survival, and by measuring cell apoptosis in the retinal layers. In addition, HO-1 expression was examined using Western blotting and immunofluorescence analyses. Cytosolic and nuclear Nrf2 was measured using immunofluorescent staining. LBP treatment significantly increased Nrf2 nuclear accumulation and HO-1 expression in the retina after I/R injury. Increased apoptosis and a decrease in the number of viable cells were observed in the ganglion cell layer (GCL) and inner nuclear layer (INL) in the I/R retina, which were reversed by LBP treatment. The HO-1 inhibitor, ZnPP, diminished the LBP treatment-induced protective effects in the retina after I/R. Taken together, these results suggested that LBP partially exerted its beneficial neuroprotective effects via the activation of Nrf2 and an increase in HO-1 protein expression.

Melanopsin-expressing retinal ganglion cell loss and behavioral analysis in the Thy1-CFP-DBA/2J mouse model of glaucoma.

In this study, the role of melanopsin-expressing retinal ganglion cells (mRGCs) in the glaucoma-induced depressive behavioral response pattern was investigated. The CFP-D2 transgenic glaucoma animal model from five age groups was used in this study. Immunohistochemical labeling, quantitative analysis of mRGC morphology, open field test (OFT), and statistical analysis were used. In comparison with C57 BL/6 mice, the age-matched CFP-D2 mice had significantly elevated intraocular pressure (IOP). We observed parallel morphological changes in the retina, including a reduction in the density of cyan fluorescent protein-(CFP) expressing cells (cells mm(-2) at 2 months of age, 1309±26; 14 months, 878±30, P<0.001), mRGCs (2 months, 48±3; 14 months, 19±4, P<0.001), Brn3b-expressing RGCs (2 months, 1283±80; 14 months, 950±31, P <0.001), Brn-3b expressing mRGCs (5 months, 50.17%±5.5%; 14 months, 12.61%±3.8%, P<0.001), and reduction in the dendritic field size of mRGCs (mm(2) at 2 months, 0.077±0.015; 14 months, 0.065±0.015, P<0.05). CFP-D2 mice had hyperactive locomotor activity patterns based on OFT findings of the total distance traveled, number of entries into the center, and time spent in the center of the testing apparatus. The glaucoma induced hyperactive response pattern could be associated with dysfunctional mRGCs, most likely Brn-3b-positive mRGCs in CFP-D2 mice.

Functional evaluation of iodoacetic acid induced photoreceptor degeneration in the cat.

Iodoacetic acid (IAA) has been applied to different species to acutely induce photoreceptor degeneration. The purpose of the present study was to use this toxin to thoroughly eliminate photoreceptors and induce complete blindness in the cat. IAA was delivered by single ear vein injection (20 mg kg(-1)). Six months after the IAA treatment, functional evaluations including pupillary light reflex (PLR), electroretinogram (ERG), visual behavior tests were performed. Morphological examinations were carried out after the functional evaluation. The present result shows that, six months after the IAA application, animals lost visual functions and became completely blind. High dose IAA application via ear vein delivery created an acute and reliable complete photoreceptor degeneration model in the cat. This model can be applied to genetic and cellular therapies for visual function restoration.

Roles for redox signaling by NADPH oxidase in hyperglycemia-induced heme oxygenase-1 expression in the diabetic retina.

PURPOSE: The antioxidant response element (ARE)-mediated antioxidant pathway has an important role in maintaining the redox status of the retina. The expression of ARE-mediated antioxidants, such as heme oxygenase-1 (HO-1), remains unclear in the db/db mice. We evaluated the expression of HO-1 in the retinas of db/db mice and investigated a possible role for NADPH oxidase. METHODS: Fresh retinas were harvested from 8-, 12-, and 20-week db/db or db/m mice. Reactive oxygen species were detected by dihydroethidium. The expression levels of HO-1, Nox2, and Nox4 were evaluated by immunohistochemistry and Western blotting. In vitro retina explants culture was used to assess the role of NADPH oxidase in high glucose-induced HO-1 expression. RESULTS: The expression of HO-1 was increased in the retinas of 8-week db/db mice, while it was decreased in 20-week db/db mice compared to age-matched controls. Similarly, the activation of Nox4 was increased in the retinas at 8 weeks and returned to basal levels at 20 weeks in db/db mice compared to age-matched controls. The activation of Nox2 was increased in the retinas of 8-, 12-, and 20-week db/db mice compared to age-matched controls. The NADPH oxidase inhibitors apocynin and DPI significantly blocked the HO-1 expression that was induced by high glucose levels in cultured retina explants. CONCLUSIONS: The expression patterns of HO-1, Nox2, Nox4 in db/db mouse retinas, and the suppressive effects of NADPH oxidase inhibitors on the expression of HO-1 induced by high glucose levels in cultured retina explants suggest that the expression of HO-1 is, at least partially, mediated by NADPH oxidase in this diabetic animal model.