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Inherited retinal degeneration (RD) comprises a heterogeneous group of retinopathies that rank among the main causes of blindness. Tauroursodeoxycholic acid (TUDCA) is taurine conjugate hydrophilic bile acid that demonstrates profound protective effects against a series of neurodegenerative diseases related to oxidative stress. This study sought to evaluate the TUDCA induced effects of on a pharmacologically induced RD animal model by electroretinogram (ERG) examination, behavior tests, morphological analysis and immunochemistry assay. Massive photoreceptor degeneration in mice retina was induced by an intraperitoneal administration of N-methyl-N-nitrosourea(MNU). Subcutaneous delivery of TUDCA inhibits effectively the photoreceptor loss and visual impairments in the MNU administered mice. In the retinal flat-mounts of TUDCA treated mice, the cone photoreceptors were efficiently preserved. Furthermore, the multi-electrodes array (MEA) was used to detect the firing activities of retinal ganglion cells within the inner retinal circuits. TUDCA therapy could restrain the spontaneous firing response, enhance the light induced firing response, and preserve the basic configurations of ON-OFF signal pathway in degenerative retinas. Our MEA assay provided an example to evaluate the potency of pharmacological compounds on retinal plasticity. TUDCA affords these protective effects by modulating apoptosis and alleviating oxidative stress in the degenerative retina. In conclusion, TUDCA therapy can ameliorate the photoreceptor degeneration and rectify the abnormities in visual signal transmission. These findings suggest that TUDCA might act as a potential medication for these retinopathies with progressive photoreceptor degeneration., (Copyright © 2019 Tangdu Hospital, Fourth Military Medical University. Published by Elsevier Masson SAS.. All rights reserved.)

We assessed the neuroprotective effects of Lycium barbarum Polysaccharides (LBP) on photoreceptor degeneration and the mechanisms involved in oxidative stress in light-exposed mouse retinas. Mice were given a gavage of LBP (150 mg/kg or 300 mg/kg) or phosphate buffered saline (PBS) for 7 days before exposure to light (5000 lx for 24 h). We found that LBP significantly improved the electroretinography (ERG) amplitudes of the a- and b-waves that had been attenuated by light exposure. In addition, changes caused by light exposure including photoreceptor cell loss, nuclear condensation, an increased number of mitochondria vacuoles, outer membrane disc swelling and cristae fractures were distinctly ameliorated by LBP. LBP treatment also significantly prevented the generation of reactive oxygen species (ROS) compared with PBS treatment. The levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and thioredoxin reductase (TrxR1) mRNA were decreased in PBS-treated mice compared with controls but increased remarkably in LBP-treated mice. The mRNA levels of the DNA repair gene Poly (ADP-ribose) polymerase (PARP14) was increased in PBS-treated mice but decreased significantly in the LBP-treated mice. Our findings indicate that pretreatment with LBP effectively protected photoreceptor cells against light-induced retinal damage probably through the up-regulation of the antioxidative genes Nrf2 and TrxR1, the elimination of oxygen free radicals, and the subsequent reduction in the mitochondrial reaction to oxidative stress and enhancement in antioxidant capacity. In addition, the decreased level of PARP14 mRNA in LBP-treated mice also indicated a protective effect of LBP on delaying photoreceptor in the light-damaged retina., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Purpose: Retinitis pigmentosa (RP) is a group of inherited diseases characterized by the death of rod photoreceptors, followed by the death of cone photoreceptors, progressively leading to partial or complete blindness. Currently no specific treatment is available for RP patients. Sulforaphane (SFN) has been confirmed to be an effective antioxidant in the treatment of many diseases. In this study, we tested the therapeutic effects of SFN against photoreceptor degeneration in Pde6b rd10 mice., Methods: rd10 mice and C57/BL6 wild-type (WT) mice were treated with SFN and saline, respectively, from P6 to P20. Electroretinography (ERG), terminal deoxynucleotidyl transferase dUTP nick end labeling and western blot were tested, respectively, at P21 for the analysis of retinal function, retinal cell apoptosis or death and the protein express of GRP78/BiP (TUNEL) as a marker of endoplasmic reticulum (ER) stress., Results: Compared with the saline group, the SFN-treated group showed significantly higher ERG a-wave and b-wave amplitudes, less photoreceptor death, and the downregulation of GRP78/BiP., Conclusions: Our data showed that SFN ameliorated the retinal degeneration of rd10 mice, which is possibly related to the downregulation of GRP78 expression.

Background: To characterize the effects of intravitreal injections of iodoacetic acid (IAA) in comparison to its systemic application as a measure to induce unilateral photoreceptor degeneration., Methods: Seven-week-old C57BL/6 J mice received either intravitreal injections of IAA or systemic treatment (intraperitoneal vs intravenous) and were observed in the following 5 weeks using ERG, OCT, and histology., Results: Systemic treatment with IAA induced high toxic effects and a high mortality in contrast to the intravitreal injection. Intraperitoneal application had no effect on the retina. Intravenous application of 2 × 30 mg/kg BW IAA (time between injections 3.5 h) resulted in an extinction of the ERG and a thinning of the retina, in particular of the outer nuclear layer (ONL) indicating photoreceptor degeneration. Animals receiving intravitreal injections developed cataracts already at low concentrations (up to 100% at 0.25 mg/kg BW). Higher intravitreal IAA doses led to extinguished ERGs. In histology, a thinning of the entire retina was observed that was most prominent in the inner part of the retina., Conclusions: In contrast to intraperitoneal administration, intravenous application of IAA led to a selective photoreceptor degeneration. After intravitreal injection, dense cataracts were already observed at concentrations lower than those needed to induce changes in the ERG. ERG results must be interpreted carefully. A thinning of all retinal layers rather than a specific outer retinal degeneration was observed upon intravitreal injection. IAA is not a useful model to induce outer retinal degeneration in mice.

Purpose: To develop and characterize a feline model of retinal degeneration induced by intravitreal injection of adenosine triphosphate (ATP)., Methods: Nineteen normally sighted adult cats received 100 μL intravitreal injections of ATP with a final concentration of 11, 22, or 55 mM at the retina. Four animals were euthanized 30 hours after injection and retinal sections examined for apoptosis using a TUNEL cell death assay. In the remaining animals, structural and functional changes were characterized over a 3-month period using a combination of electroretinography (ERG) and optical coherence tomography (OCT)., Results: Using a TUNEL cell death assay, we detected widespread photoreceptor death 30 hours after injection with 55 mM intravitreal ATP. All concentrations of ATP caused loss of retinal function and gross changes in retinal structure within 2 weeks of injection. Intravitreal injection of ATP led to a rapid loss of rod photoreceptor function and a gradual loss of cone photoreceptor function within 3 months. Outer nuclear layer thickness was globally reduced by 3 months, with the inner nuclear layer including the retinal nerve fiber layer remaining intact. Structural abnormalities were observed, including focal retinal detachment with evidence of both intravitreal and intraretinal inflammation in some eyes., Conclusions: Development of an ATP-induced feline model of retinal degeneration provides a rapid and effective large-eyed animal model for research into vision restoration., (Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.)

Purpose: We investigated how the N-methyl-DL-aspartic acid (NMDA) receptor contributes to generating oscillatory potentials (OPs) of the electroretinogram (ERG) in the Royal College of Surgeons (RCS) rat., Methods: Scotopic ERGs were recorded from dystrophic and wild-type congenic (WT) RCS rats (n = 20 of each) at 25, 30, 35, and 40 days of age. The stimulus intensity was increased from -2.82 to 0.71 log cd-s/m(2) to obtain intensity-response function. NMDA was injected into the vitreous cavity of the right eyes. The left eyes were injected with saline as controls. The P3 obtained by a-wave fitting was digitally subtracted from the scotopic ERG to isolate the P2. For the OPs, the P2 was digitally filtered between 65 and 500 Hz. The amplitudes of OP1, OP2, OP3, and OP4 were then measured and summed and designated as ΣOPs. The implicit times of OP1, OP2, and OP3 were also measured. The frequency spectra of the OPs were analyzed using fast Fourier transform (FFT)., Results: The maximum ERG a- and b-waves as well as ΣOPs amplitudes reduced with age in dystrophic rats. Compared with intravitreal saline injection, administration of NMDA decreased ΣOPs amplitudes from 30 days of age in dystrophic rats, while it did not attenuate ΣOPs amplitudes in WT rats. The implicit times of the OPs of the maximum ERG were prolonged by NMDA injections in WT and dystrophic rats. NMDA/saline ratios of ΣOPs amplitudes area under the FFT curves were significantly lower in dystrophic rats from 30 days of age than that in WT rats., Conclusion: In the early stage of photoreceptor degeneration, intravitreal NMDA injection attenuated OPs amplitudes in dystrophic rats. This indicates that NMDA receptors play a significant role in generating OPs amplitudes with advancing photoreceptor degeneration.

Anthocyanins are known to have antioxidant effects and thus may play an important role in preventing various degenerative diseases. In this study, we examined the effect of anthocyanins extracted from the seed coat of black soybean on an animal model of retinal degeneration (RD), a leading cause of photoreceptor cell death resulting in blindness. RD was induced in rats by an intraperitoneal injection of N-methyl-N-nitrosourea (MNU) (50mg/kg), a DNA-methylating agent that causes photoreceptor damage. Anthocyanins extracted from black soybean seed coat (50mg/kg) were daily administered, orally, for 1, 2, and 4 weeks after MNU injection. Electroretinographic (ERG) recordings and morphological analyses were performed. In control rats with MNU-induced retinal damage, the ERG recordings showed a gradual significant time-dependent reduction in both a- and b-wave amplitudes compared with those of normal animals. In the MNU-induced RD rats given anthocyanins for 4 weeks, ERG responses were significantly increased compared with untreated RD rats, more apparently in scotopic stimulation than in the photopic condition. However, in the MNU-injected rats given anthocyanins for 1 and 2 weeks, the increase in ERG responses was not significant. Morphologically, the outer nuclear layer, where photoreceptors reside, was well preserved in the anthocyanin-treated rat retinas throughout the experimental period. In addition, retinal injury, evaluated by immunolabeling with an antibody against glial fibrillary acidic protein, was markedly reduced in anthocyanin-treated retinas. These results demonstrate that anthocyanins extracted from black soybean seeds can protect retinal neurons from MNU-induced structural and functional damages, suggesting that anthocyanins from black soybean seed coat may be used as a useful supplement to modulate RD., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Purpose: To determine how the different stages of retinal processing change after photoreceptor degeneration in rabbits carrying the Pro347Leu rhodopsin mutation (Tg rabbits)., Methods: Cone electroretinograms (ERGs) were elicited by 150-ms duration stimuli from 13 Tg rabbits at 12 and 24 weeks of age. The ERG recordings were made before and after an intravitreal injection of tetrodotoxin citrate (TTX) plus N-methyl-dl-aspartic acid (NMDA), with the addition of 2-amino-4-phosphonobutyric acid (APB) and then cis-2,3-piperidine-dicarboxylic acid (PDA) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Digital subtraction of the ERG after the injection from the ERG before the injection was used to extract the components that were blocked by these drugs. Thirteen age-matched, wild-type (WT) rabbits were studied with the same protocol., Results: In Tg rabbits, the cone ERGs elicited by intermediate intensities had a depolarizing pattern. At 12 weeks of age, the photoreceptor and OFF-bipolar/horizontal cell responses reflected in the ERG in the Tg rabbits did not differ significantly from those in the WT rabbits. The ON-bipolar cells and the third-order neuronal responses recorded after pharmacologic blockade were significantly enhanced in the Tg rabbits compared with those recorded in the WT rabbits. At 24 weeks of age, the ERG waveforms representing the photoreceptors and OFF-bipolar/horizontal cell responses were significantly decreased, but those representing the ON-bipolar cell and third-order neuronal responses were still preserved in the Tg rabbits., Conclusions: A depolarizing pattern of the cone ERG responses was seen in Pro347Leu Tg rabbits. The enhancement or preservation of the ON-bipolar cell response in the ERGs contributed to shaping the waveform in the Tg rabbits. In this model, the functional alterations in the ON-pathway took place before the deterioration of cone photoreceptor function.

Purpose: To determine whether photoreceptor degeneration in transgenic (Tg) rabbits carrying the Pro347Leu rhodopsin mutation alters the neural activity of the middle and inner retinal neurons., Methods: Multifocal electroretinograms (mfERGs) were recorded from eight 12-week-old Tg rabbits both before and after intravitreal injection of the following: tetrodotoxin citrate (TTX), N-methyl-DL: -aspartic acid (NMDA), 2-amino-4-phosphonobutyric acid (APB), and cis-2,3-piperidine-dicarboxylic acid (PDA). Digital subtraction of the mfERGs recorded after the drugs were administered from those recorded before was used to extract the components that were eliminated by these drugs. Eight agematched, wild-type (WT) rabbits were studied with the same protocol., Results: There was no reduction in the amplitude of the cone photoreceptor response of the mfERGs in Tg rabbits. Both the first positive and the first negative waves of the ON-bipolar cell responses were significantly larger in the Tg than in the WT rabbits. Late negative waves of the ON-bipolar cell response were recorded only in the WT rabbits. The first negative wave of the inner retinal responses was larger in the Tg than in the Wt rabbits. The late positive waves were seen mainly in the WT rabbits., Conclusions: The ON-bipolar cell and inner retinal responses were altered at the early stage of photoreceptor degeneration in Tg rabbits despite the preservation of the cone photoreceptor responses.

Purpose: To study the neuroprotective properties of low-dose, sustained-release intravitreous fluocinolone acetonide (FA) in transgenic S334ter-4 rats., Methods: S334ter-4 rats aged 4 weeks were divided into four groups: 0.5 microg/d FA-loaded intravitreous drug delivery implant (IDDI); 0.2 microg/d FA-loaded IDDI; inactive IDDI; and unoperated controls. Electroretinography (ERG) was performed before surgery and every 2 weeks after surgery for 8 weeks. When the rats were 12 weeks of age, outer nuclear layer (ONL) and inner nuclear layer (INL) thicknesses were measured. Microglial cell counts were obtained from retinal wholemounts labeled for Iba-1., Results: At the end of the study, unoperated and inactive IDDI-implanted rats demonstrated 50% to 60% reductions in ERG amplitudes compared with those recorded at 4 weeks (P < 0.001 for both groups). FA 0.2-microg/d animals demonstrated 15% amplitude attenuation, while FA 0.5-microg/d animals showed 30% reduction. ONL thickness in FA 0.2-microg/d-treated eyes was 25.8% +/- 2.3% higher than in control group eyes (P < 0.001) and 30.0% +/- 2.1% higher than in inactive IDDI-implanted eyes (P < 0.001). In FA 0.5-microg/d-treated eyes, ONL thickness was 22.4% +/- 2.8% higher than in control group eyes (P < 0.001) and 22.3% +/- 3.7% higher than in inactive IDDI-implanted eyes (P < 0.01). No statistically significant difference was observed between the two control groups. No statistically significant difference between the two FA-treated groups was found. FA-treated groups demonstrated significantly fewer activated microglial cells than control groups., Conclusions: Chronic intravitreous infusion of FA preserves ONL cell morphology and ERG a- and b-wave amplitudes and reduces retinal neuroinflammation in S334ter rats. Based on these findings, the synthetic corticosteroid FA may promise a therapeutic role in patients with retinal degeneration.

Purpose: Mice lacking retinal pigment epithelium-specific 65-kDa protein (RPE65) develop retinopathy and blindness resembling Leber congenital amaurosis. Effects of 9-cis-retinyl acetate (9-cis-R-Ac) on visual function and retinopathy progression were tested in Rpe65(-/-) mice., Methods: Young C57Bl/6 mice were given 9-cis-R-Ac in each of four different oil-based vehicle solutions by gastric gavage to identify the vehicle most suitable for drug delivery by measuring retinoid levels in plasma. Then doses of 9-cis-R-Ac ranging from 1 to 100 mg/kg were administered to 5- to 12-week-old Rpe65(-/-) mice by different treatment regimens, including single doses and either intermittent or daily doses for various periods up to 8 weeks. Retinoid effects on visual function were evaluated by electroretinography, retinoid analyses, histologic methods, and vision-dependent behavioral testing., Results: Soybean oil vehicle provided the highest 9-cis-R-Ac metabolite levels in plasma. Single doses of 9-cis-R-Ac (6.25-50 mg/kg) provided significant dose-dependent improvement in electroretinographic responses. Well-tolerated daily doses (1-12.5 mg/kg) for 2 weeks induced remarkable improvement of retinal function. Significant dose-dependent improvement of electroretinographic responses was observed 6 days after administration of 9-cis-R-Ac daily for 3 days at 1 to 12.5 mg/kg. Mice given either daily or intermittent 9-cis-R-Ac treatment at 1 and 4 mg/kg and evaluated 8 weeks later displayed dose-dependent improvement of retinal function and morphology, whereas retinal function deteriorated in control animals. Treated mice also performed better than control animals in vision-dependent behavioral tests., Conclusions: Treatment with 9-cis-R-Ac improves visual function and preserves retinal morphology in Rpe65(-/-) mice.

Purpose: In nonocular systems, activation of opioid receptors has been shown to ameliorate tissue damage induced by ischemic stress. The current study was an investigation of whether opioid receptors activated by endogenous or exogenous agonists can ameliorate ischemic retinal injury., Methods: In an investigation of whether endogenous opioid receptor-activation reduces ischemic injury, the effects of the opioid antagonist naloxone (3 mg/kg; IP) on retinal neuroprotection induced by ischemic preconditioning (IPC) were evaluated. Whether exogenous opioid administration can reduce ischemic retinal injury was determined by pretreating rats with morphine (0.01-10 mg/kg) before injury. Morphometric and electroretinogram (ERG) analyses were used to assess the differences in retinal structure and function. The expression of opioid receptor subtypes was evaluated by Western blot and immunohistochemical analyses., Results: In control animals, 7 days after ischemic retinal injury, ERG a- and b-wave amplitudes were significantly reduced (23% and 41%, respectively). In addition, degeneration of the inner retina resulted in a 34% reduction in overall retina thickness. In animals receiving IPC before ischemic injury, ERG wave forms and retinal morphology were preserved. Pretreatment with naloxone reversed both the functional and structural retinal protection induced by IPC. In animals treated with morphine 24-hours before ischemic injury, ERG waveforms were preserved in a dose-dependent fashion (ED(50) = 0.18 mg/kg), and this protective response was reversed by naloxone pretreatment. Immunohistochemical and Western blot data demonstrated that the delta-, kappa-, and mu-opioid receptor subtypes are expressed in the retina., Conclusions: These data provide evidence that activation of one (or more) opioid receptor(s) facilitates the development of IPC within the retina and can reduce ischemic retina injury.

This study investigated contributions from the retinal On and Off pathways, and the spiking and nonspiking activity of neurons in those pathways to the pattern ERG of the mouse. Light-adapted pattern and ganzfeld ERGs were recorded from anesthetized C57BL/6 mice 3-4 months of age. Recordings were made before and after intravitreal injections of PDA (cis-2,3-piperidine-dicarboxylic acid) to block transmission to hyperpolarizing 2nd order and all 3rd order neurons, TTX (tetrodotoxin) to block Na(+)-dependent spiking, APB (2-amino-4-phosphonobutyric acid) to block synapses between photoreceptors and ON-bipolar cells, and APB + TTX and PDA + TTX cocktails. The pattern stimuli consisted of 0.05 cy/deg gratings reversing in contrast at 1 Hz, presented at various contrasts (50-90%) and a rod saturating mean luminance. For flash ERGs, brief green ganzfeld flashes were presented on a rod-suppressing green background. Recordings were made 39-42 days after unilateral optic nerve crush (ONC) in a subset of animals in which ganglion cell degeneration was subsequently confirmed in retinal sections. Pattern ERGs were similar in waveform for all contrasts, with a positive wave (P1) peak for 90% contrast around 60 ms on average and maximum trough for a negative wave (N2) around 132 ms after each contrast reversal; amplitudes were greatest for 90% contrast which became the standard stimulus. ONC eliminated or nearly eliminated the pattern ERG but did not affect the major waves of the flash ERG. PDA and TTX both delayed P1 and N2 waves of the pattern ERG, and reduced their amplitudes, with effects of PDA on N2 greater than those of TTX. In the flash ERG, PDA reduced a-wave amplitudes, removed OPs but hardly affected b-wave amplitudes. In contrast, TTX reduced b-wave amplitudes substantially, as previously observed in rat. APB removed P1 of the pattern ERG, but left a negative wave of similar timing and amplitude to N2. In the flash ERG, APB removed the b-wave, producing a negative ERG. Addition of TTX to the APB injection removed most of N2 of the pattern ERG, while other waves of the pattern and flash ERG resembled those after APB alone. Addition of TTX to the PDA injection had little effect on the pattern ERG beyond that of PDA alone, but it prolonged the b-wave of the flash ERG. In conclusion, this study confirmed that a selective lesion of ganglion cells will practically eliminate the pattern ERG. The study also showed that P1 of the mouse pattern ERG is dominated by contributions, mainly spiking, from ON pathway neurons, whereas N2 reflects substantial spiking activity from the OFF pathway as well as nonspiking contributions from both pathways.

Purpose: To investigate the intraocular effect of rhodamine 6G (R6G) on retinal structures and function in an in vivo rat model and to develop an in vivo method for accurate evaluation of new dyes for intraocular surgery., Methods: R6G in physiologic saline solution (PSS) was injected into the vitreous of adult Brown Norway rats at concentrations of 0.0002%, 0.002%, 0.02%, 0.2%, and 0.5%. Control animals received only PSS. Retinal toxicity was assessed by retinal ganglion cell (RGC) counts, light microscopy 7 days later, photopic electroretinography (ERG), and measurement of scotopic sensitivity and recovery of dark adaptation 48 hours and 7 days after intravitreous injection., Results: R6G at concentrations of 0.2% and 0.5% led to a dose-dependent loss of RGC. The most significant loss occurred at 0.5%. Lower concentrations (0.0002%, 0.002%, and 0.02%) produced no statistically significant retinal ganglion cell loss. Analysis of the eyes by light microscopy showed no structural changes in the central retina, although injections of 0.5% R6G were followed by impressive degenerative changes adjacent to the injection sites. ERGs showed no effects of the highest R6G concentration on rods, kinetics of rhodopsin recovery after bleaching, or cone-driven responses., Conclusions: R6G can be safely injected in doses of up to 0.02% in rats, but has a toxic effect on retinal ganglion cells at higher concentrations. Accumulation of R6G may be a problem at higher concentrations, particularly at the injection site.

In most retinal degenerations in humans and in animal models, photoreceptor cells die by apoptosis. Although the biochemical features are similar in all apoptotic cells, different molecular events lead the cell to death. In the present study we used a rat model of inherited retinal degeneration, the RCS rats, to investigate the involvement of the proteases, caspases and/or calpains, in photoreceptor apoptosis. In the first experiments, rats were untreated or injected intravitreally at post natal day 27 (P27) with the large broad spectrum caspase inhibitor, ZVAD, the calpain inhibitor, MuhPhe, or with the vehicle, DMSO. Retinal status was evaluated at P35 and P42 by electroretinography, morphometry and apoptotic nuclei detection. DMSO and MuhPhe had no effect on RCS retinas as evidenced by equivalent loss of function and equivalent number of apoptotic cells than in untreated group. ZVAD transiently reduced apoptotic cells and preserved photoreceptor function at P35 but not at P42. These results suggest that caspases but not calpains are involved in retinal degeneration in the RCS. In the second experiments, RCS rats were injected twice at P27 and P35 with ZVAD or DMSO. Although ZVAD-treated retinas were preserved at P35 compared to the DMSO controls, the second injection of ZVAD did not extend the preserving effect to P42. Moreover, a single injection of ZVAD at P35 had no preserving effect at P42. All these data taken together suggest that caspases do not play a pivotal role after P35. In a fourth set of experiments, we used specific caspase inhibitors to elucidate which caspase was activated. The caspase-1/4 inhibitor (YVAD) or the caspase-3/7 inhibitor (DEVD) were injected intravitreally at P27 and retinal status was evaluated at P35 and P42. Electroretinograms and apoptotic nuclei detection demonstrated that YVAD and DEVD preserved photoreceptors at P35 but not at P42. These results suggest that both caspase-1/4 and caspase-3/7 play a major role in the apoptotic pathway between P27 and P35 in retinal degeneration of RCS rats. In this study, we show that 1/ the photoreceptor apoptotic process in the RCS rat involves caspases but not calpains, and 2/ the retinal degeneration seems to be composed of different phases involving different molecular players. Indeed, we have demonstrated that caspases are playing a major role at P35, but not at P42.

To investigate the effect of nilvadipine, a calcium channel blocker, upon the retina of retinal degeneration slow (rds) mouse, nilvadipine was intraperitoneally injected into heterozygous rds mice for up to 200 days. The effect of nilvadipine was evaluated by electroretinography (ERG), light and electron microscopies, DNA microarray, quantitative reverse transcriptase polymerase chain reaction (RT-PCR), and western-blot analysis. After nilvadipine treatment, both a- and b-waves of ERG were significantly higher than in the control group (p<0.01). Although there was no difference in histological findings by light microscopy between the nilvadipine treated group and control group, apparent preservation of photoreceptor disc was demonstrated by electron microscopy in the treated group. Rhodopsin level was also increased in the treated group comparing to the control group. The DNA microarray analysis detected increased expression of genes encoding proteins which function in protein synthesis, growth factors and neurotrophic factor like ciliary neurotrophic factor (CNTF) and fibroblast growth factors (FGFs22 and 13). Decreased expression of genes coding for proteins related to proteolysis, apoptosis and growth factor (FGF18) was also demonstrated. Increased expression of CNTF, FGF22 and FGF13 and decreased expression of FGF18 were confirmed by both quantitative RT-PCR and western-blot analysis. In addition, FGF2 was constitutively expressed in both treated and control groups. Since CNTF has been known to retard retinal degeneration by rds mouse or other models of inherited retinal degeneration, it is possible that nilvadipine has a photoreceptor survival effect on rds retinal degeneration partly by enhancing expression of endogenous CNTF in the retina.

Purpose: To investigate how the third-order neuronal response contributes to shaping the electroretinogram (ERG) in the Royal College of Surgeons (RCS) rat., Methods: Full-field ERGs were recorded from dystrophic RCS rats (n = 30) at 4, 6, 8, 10, 12, or 14 weeks of age in response to different stimulus intensities (maximum intensity, 0.84 log cd-s/m(2)). N-methyl-DL: -aspartic acid (NMDA, 5 mM) was injected into the vitreous cavity of the right eyes to eliminate the third-order neuronal response. The left eyes received the vehicle and served as controls. The third-order neuronal response was isolated by digitally subtracting waveforms of the NMDA-injected eyes from those of the control eyes., Results: The ERG a- and b-waves deteriorated with the age of the rat. The third-order neuronal response was preserved to a greater degree than the b-wave despite progression of photoreceptor degeneration. Intravitreal injection of NMDA attenuated the a-wave and enhanced the b-wave across the stimulus range from low to middle intensities. This tendency became more pronounced with advancing rat age. In aged dystrophic RCS rats this phenomenon was seen even at maximum intensity. The difference between NMDA-injected and vehicle-injected eyes was larger for the threshold than for the maximum amplitude at each examined time point (P < 0.001). Intravitreal injection of NMDA decreased implicit times of the a- and b-waves after the rats reached 8 weeks of age (P < 0.005 for the a-wave)., Conclusion: With advancing photoreceptor degeneration, the third-order neuronal response made a greater contribution to shaping the a- and b-waves in dystrophic RCS rats., (Copyright (c) Japanese Ophthalmological Society 2007.)

Purpose: In this study, the hypothesis that increased intraocular levels of iron cause oxidative damage to the retina was tested., Methods: Adult C57BL/6 mice were given an intravitreous injection of saline or 0.10, 0.25, or 0.50 mM FeSO(4). Scotopic electroretinograms (ERGs) were performed 3, 7, and 14 days after injection, and photopic ERGs were performed on day 14. Hydroethidine was used to identify superoxide radicals and lipid peroxidation was visualized by staining for hydroxynonenal (HNE). Retinal cell death was evaluated by TUNEL and measurement of inner nuclear layer (INL) and outer nuclear layer (ONL) thickness. Levels of rhodopsin and cone-opsin mRNA were measured by quantitative real time RT-PCR. Cone density was assessed by peanut agglutinin staining and confocal microscopy., Results: Compared with retinas in saline-injected eyes, retinas from eyes injected with FeSO(4) showed greater fluorescence after intravenous injection of hydroethidine due to superoxide radicals in photoreceptors, greater photoreceptor staining for HNE, a marker of lipid peroxidation, and increased expression of Heme oxygenase 1, an indicator of oxidative stress. ERG b-wave amplitudes were reduced (photopic > scotopic) in FeSO(4)-injected eyes compared with those in saline-injected eyes. Numerous TUNEL-stained nuclei were seen along the outer border of the ONL, the location of cone cell nuclei, at 1 and 2 days after injection of FeSO(4). In FeSO(4)-injected eyes, the thickness of the ONL, but not the INL, was significantly reduced, and 17 days after injection, there were 3.8- and 2.6-fold reductions in the mRNAs for M-cone and S-cone opsin, respectively, whereas there was no significant difference in rhodopsin mRNA. Confocal microscopy of peanut agglutinin-stained sections showed dose-dependent FeSO(4)-induced cone drop out., Conclusions: Increased intraocular levels of FeSO(4) cause oxidative damage to photoreceptors with greater damage to cones than rods. This finding suggests that the oxidative defense system of cones differs from that of rods and other retinal cells, and that cones are more susceptible to damage from the type of oxidative stress imposed by iron.

Purpose: Neurotrophic factors and neurotrophins are well-known to have neuroprotective efficacy against retinal injury. The aim of this experiment is to investigate the signal transduction pathway of ciliary neurotrophic factor (CNTF) on the upregulation of viability of retinal primary culture and retinal protection against constant light damage in vivo. CNTF is known to enhance the viability of retinal culture and provide protection under constant light exposure conditions, but little is known about how the signal transduction pathways of CNTF affect retina function., Methods: Primary retinal cultures were prepared from 7-day-old Wistar rats. Brain-derived neurotrophic factor (BDNF) (0.1, 1, 10 ng/ml), CNTF (0.1, 1, 10 ng/ml), PD98059 (10, 100, 1000 nM), or LY294002 (10, 100, 1000 nM) was added to these cultures at the time of cell preparation. After 3 days, the percentage of cells surviving was assessed using alamarBlue. For the in vivo experiment, inhibitors for the MAPKK (PD98059, 10 microg/eye) or PI3K (LY294002, 10 microg/eye) pathways were injected into the vitreous together with CNTF (1 microg/eye) 2 days before constant light exposure. Electroretinogram (ERG) analysis was performed to investigate which pathway was used by CNTF., Results: CNTF at 1, 10, or 100 ng/ml enhanced cell viability in retinal cultures. The cell-survival activity of CNTF was blocked by 10 ng/ml LY294002 (Dunnet's test, p < 0.05). In vivo, the neuroprotective activity of CNTF in constant-light conditions was attenuated by 10 microg/eye LY294002 (Dunnet's test, p < 0.05)., Conclusions: These data suggest that CNTF promotes cell survival via the PI3K signaling pathway in vitro and in vivo.

Intraocular injections of ciliary neurotrophic factor (CNTF) or intraocular adenovirus-mediated CNTF gene transfer have been reported to inhibit retinal alterations in inherited retinal degeneration in mice strains. To investigate whether or not CNTF administered by eye drops prevents retinal degeneration in streptozotocin (STZ)-induced diabetic rats, recombinant CNTF was administered to eyes of diabetic rats (n=20) twice daily for 1 month after the onset of diabetes. The b-wave amplitude of electroretinogram in CNTF-administered diabetic rats was significantly larger than that of diabetic rats, and approached that of the controls. Atrophy of the inner plexiform layer, and cavity formation in the pigment epithelium, which were observed in diabetic rats, were prevented in CNTF-administered diabetic rats. These results indicate that CNTF administration by eye drops prevents retinal degeneration in STZ-induced diabetic rats.

In the retinas of Royal College of Surgeons (RCS) rats light induces an increase in distal extracellular potassium irrespective of the age, between days 19-24 and days 29-35 postpartum, but by days 29-35 the ERG b-wave has become reduced. The synaptic blocker 2-amino-4-phosphonobutyric acid (APB) causes the abolition of both the b-wave and the potassium increase at any age. MgCl2 greatly reduces the b-wave at all ages and abolishes the potassium increase in older rats, but in younger rats the potassium increase is enlarged. Since this increase occurs in the absence of the b-wave it is unlikely that the on-bipolar cells are the only sources of the b-wave. Because the NMDA receptor blocker ketamine reduces the b-wave, third order neurons, which possess NMDA receptors, could contribute to the b-wave.

When preparing isolated rabbit retinas we found in some animals fundi which were not uniformly dark but had abnormal areas of red coloration. The in situ electroretinograms (ERG) of 82 rabbits recorded after 1 h of dark adaptation were checked for abnormalities indicative of a degenerative disorder. The ERGs of eight rabbits with small dark adapted b-waves (< or = 250 microV) were re-recorded and their b-waves found to decline with time. The greatest reduction, in three rabbits, was > or = 150 microV over 2.5 years. After 1 year, however, the light adapted b-waves were similar to those of rabbits with normal dark adapted b-waves. The majority of the progeny of these rabbits also had small b-waves, which became still smaller in 2 years. Ultrastructural studies of two rabbit retinas of the first generation showed pathological changes of the pigment epithelium (Wrigstad, Hanitzsch & Nilsson, Ultrastructural and electrophysiological studies of the retina and the retinal pigment epithelium in rabbits with low b-wave amplitudes, in preparation). Evidently there is an inheritable defect in the pigment epithelium which first impairs the rod pathway.

When pure, synthetic lipid hydroperoxides (LHP) were injected into the vitreous body of albino rabbits, electrical activity was decreased in all components of the electroretinogram (ERG) in a progressive and time-related manner. In the early phase morphological changes occurred at the interface between the photoreceptor outer segments and the retinal pigment epithelium (RPE) in response to LHP. These findings commenced within a few hours after injection and continued during the following 2-3 weeks, when the ERG was completely extinguished. Two days after injection, the rod outer segments (ROS) were swollen and damage of the RPE apical villi was observed. This initial event was followed by additional changes in the RPE which embraced swelling, accumulation of residual bodies, complete loss of ROS and enlargement and disruption of Bruch's membrane. The precursors of residual bodies appeared to result from focal, peroxidative damage to ROS discs which apparently rendered these materials undegradable by the RPE. As ROS degeneration continued, the RPE showed hypertrophy and modification. These studies provided evidence for a sequential destruction of the neural retina and RPE during oxidative damage involving lipid peroxidation. The mechanism appeared to differ from that produced by other toxic compounds or those which resulted from vitamin E or A deficiency. This new model system is thought to be useful in 1) explaining differences in susceptibility of inner ROS disks versus other membranes, 2) determining how the RPE metabolizes abnormal ROS, 3) studying RPE reactivity following trauma and/or retinal detachment, and 4) determining factors which produce degeneration of Bruch's membrane.

Clofazamine is an iminophenazine dye with antimycobacterial activity which has recently been used to treat mycobacterium avium complex infections in patients with acquired immunodeficiency syndrome. The authors present the second report of a presumed clofazamine-induced bull's-eye maculopathy and generalized retinal degeneration in a patient with AIDS. The importance of closely following patients on clofazamine, especially those with AIDS who may be particularly susceptible to developing this toxicity, is stressed.

Tunicamycin (TM), an inhibitor of dolichylphosphate-mediated protein glycosylation, was injected intravitreally into the eyes of diurnal rodents with cone-dominated retinas. Injection of 1 microgram of the B2 isomer led to a progressive degeneration of the photoreceptor outer segments and disruption of the RPE-photoreceptor interface that took place over a 10-day period. Cone outer segments were shortened by postinjection day 6 and virtually absent by day 9. The microvilli that normally protrude from the apical surface of the retinal pigment epithelium were replaced by a fringe of shortened processes. The other retinal layers showed no morphological evidence of disruption. Retinal sensitivity, as measured by electroretinographic b-wave threshold, showed a significant and progressive decline over the 10-day course of the experiment that paralleled the disruption of retinal morphology. These results suggest that TM leads to similar morphological and electrophysiological effects on rod and cone photoreceptors.