Scotopic Electroretinogram Research Articles

Iodate induced toxic retinopathy: a case report

BackgroundIodine deficiency is a common preventable cause of intellectual and developmental disabilities. Iodine in the form of potassium iodate is added to the common salt under the National Iodine Deficiency Disorder Control Program in India. Overdose of iodate can lead to retinal toxicity.Case presentationWe hereby present a case of a 34 year old male patient who presented to us 10 years following iodate ingestion. There was widespread outer retinal atrophy, foveal atrophy and sub-retinal fibrosis noted on fundus evaluation. The fundus fluorescein angiogram was suggestive of window defects while the scotopic electroretinogram showed diminished amplitude pointing towards a grave prognosis.ConclusionsExcessive ingestion of potassium iodate can lead to outer retinal atrophy due to its toxicity to the retinal pigment epithelium and photo-receptors. The degree of damage is dependent on the ingested dose.

Laquinimod protects the optic nerve and retina in an experimental autoimmune encephalomyelitis model

BackgroundThe oral immunomodulatory agent laquinimod is currently evaluated for multiple sclerosis (MS) treatment. Phase II and III studies demonstrated a reduction of degenerative processes. In addition to anti-inflammatory effects, laquinimod might have neuroprotective properties, but its impact on the visual system, which is often affected by MS, is unknown. The aim of our study was to investigate potential protective effects of laquinimod on the optic nerve and retina in an experimental autoimmune encephalomyelitis (EAE) model.MethodsWe induced EAE in C57/BL6 mice via MOG35–55 immunization. Animals were divided into an untreated EAE group, three EAE groups receiving laquinimod (1, 5, or 25 mg/kg daily), starting the day post-immunization, and a non-immunized control group. Thirty days post-immunization, scotopic electroretinograms were carried out, and mice were sacrificed for histopathology (HE, LFB), immunohistochemistry (MBP, Iba1, Tmem119, F4/80, GFAP, vimentin, Brn-3a, cleaved caspase 3) of the optic nerve and retina, and retinal qRT-PCR analyses (Brn-3a, Iba1, Tmem119, AMWAP, CD68, GFAP). To evaluate the effect of a therapeutic approach, EAE animals were treated with 25 mg/kg laquinimod from day 16 when 60% of the animals had developed clinical signs of EAE.ResultsLaquinimod reduced neurological EAE symptoms and improved the neuronal electrical output of the inner nuclear layer compared to untreated EAE mice. Furthermore, cellular infiltration, especially recruited phagocytes, and demyelination in the optic nerve were reduced. Microglia were diminished in optic nerve and retina. Retinal macroglial signal was reduced under treatment, whereas in the optic nerve macroglia were not affected. Additionally, laquinimod preserved retinal ganglion cells and reduced apoptosis. A later treatment with laquinimod in a therapeutic approach led to a reduction of clinical signs and to an improved b-wave amplitude. However, no changes in cellular infiltration and demyelination of the optic nerves were observed. Also, the number of retinal ganglion cells remained unaltered.ConclusionFrom our study, we deduce neuroprotective and anti-inflammatory effects of laquinimod on the optic nerve and retina in EAE mice, when animals were treated before any clinical signs were noted. Given the fact that the visual system is frequently affected by MS, the agent might be an interesting subject of further neuro-ophthalmic investigations.

Improving the quality of electroretinogram recordings using active electrodes

The aim of this study was to compare the quality of electroretinogram (ERG) recordings using a custom built active electrode with attached amplifier versus a standard (passive) ERG electrode. Scotopic and photopic ERG responses were recorded from five adult albino rabbits using a custom built active electrode on one eye and a passive electrode on the other. For the active electrode, the ERG-jet electrode (Universo S.A., La Chaux-De-Fonds, Switzerland) was used as the transducer with the cable cut short and soldered directly to the input of a customized amplifier. The passive electrode was a standard ERG jet electrode. The signal to noise ratio and reproducibility of ERGs were compared. The noise was significantly lower in the active electrode compared to the passive electrode (p = 0.009) resulting in signals being recorded at lower stimulation strengths with the active electrode. The scotopic a-wave was significantly larger in the active electrode at all supra-threshold stimulation intensities (p < 0.05) and the scotopic b-wave amplitudes were also higher in the active electrode at all supra-threshold stimulation intensities but was only statistically significant between −3.25 and −1 log cd.s.m−2 (p < 0.05). The photopic a- and b-wave amplitudes were also higher in the active electrode and statistically significant between −0.75 and 0.48 log cd.s.m−2 for the a-wave and −1.25 to −1 log cd.s.m−2 for the b-wave (p < 0.05). The intra-observer repeatability, inter-sessions reproducibility and reliability of the signals were better in the active electrode as evidenced by lower coefficient of variation (CV) and coefficient of repeatability (CR) with high intra-class correlation coefficient (ICC) of the a- and b-wave parameters of the active electrode. These findings suggest that the custom built active ERG electrode produces less noise than the passive electrode, allowing responses to be recorded at lower stimulation strengths. It produces greater signal amplitudes and improved reproducibility and is therefore a better device for investigating retinal function.

Tissue inhibitor of metalloproteinases 1 enhances rod survival in the rd1 mouse retina.

Retinitis pigmentosa (RP), an inherited retinal degenerative disease, is characterized by a progressive loss of rod photoreceptors followed by loss of cone photoreceptors. Previously, when tissue inhibitor of metalloproteinase 1 (TIMP1), a key extracellular matrix (ECM) regulator that binds to and inhibits activation of Matrix metallopeptidase 9 (MMP9) was intravitreal injected into eyes of a transgenic rhodopsin rat model of RP, S334ter-line3, we discovered cone outer segments are partially protected. In parallel, we reported that a specific MMP9 and MMP2 inhibitor, SB-3CT, interferes with mechanisms leading to rod photoreceptor cell death in an MMP9 dependent manner. Here, we extend our initial rat studies to examine the potential of TIMP1 as a treatment in retinal degeneration by investigating neuroprotective effects in a classic mouse retinal degeneration model, rdPde6b-/- (rd1). The results clearly demonstrate that intravitreal injections of TIMP1 produce extended protection to delay rod photoreceptor cell death. The mean total number of rods in whole-mount retinas was significantly greater in TIMP-treated rd1 retinas (postnatal (P) 30, P35 (P<0.0001) and P45 (P<0.05) than in saline-treated rd1 retinas. In contrast, SB-3CT did not delay rod cell death, leading us to further investigate alternative pathways that do not involve MMPs. In addition to inducing phosphorylated ERK1/2, TIMP1 significantly reduces BAX activity and delays attenuation of the outer nuclear layer (ONL). Physiological responses using scotopic electroretinograms (ERG) reveal b-wave amplitudes from TIMP1-treated retinas are significantly greater than from saline-treated rd1 retinas (P<0.05). In later degenerative stages of rd1 retinas, photopic b-wave amplitudes from TIMP1-treated rd1 retinas are significantly larger than from saline-treated rd1 retinas (P<0.05). Our findings demonstrate that TIMP1 delays photoreceptor cell death. Furthermore, this study provides new insights into how TIMP1 works in the mouse animal model of RP.

Loss of XBP1 accelerates age-related decline in retinal function and neurodegeneration

BackgroundAging is the strongest risk factor for neurodegenerative diseases and extended age results in neuronal degeneration and functional decline in the visual system. Among many contributing factors to age-related deterioration of neurons is an insufficient activation of the Unfolded Protein Response (UPR) in the endoplasmic reticulum (ER) in response to cellular stress. X-box binding protein 1 (XBP1) is a major component of the UPR and is essential for maintaining protein homeostasis and reducing cellular stresses. Herein, we investigate the role of XBP1 in maintaining morphological and functional integrity in retinal neurons during adulthood and the early stages of aging.MethodsThe basal and induced levels of XBP1 activation in the retina were measured in young adult and aged mice. Conditional knockout (cKO) of XBP1 in retinal neurons was achieved by crossing XBP1 floxed mice with a retina specific Cre-recombinase line (Chx10-Cre). Retinal morphology, neuronal populations including photoreceptors, bipolar cells, and retinal ganglion cells (RGCs), synaptic structure, and microglial activation were examined with immunohistochemistry and staining of retinal sections. Retinal function was evaluated with light-adapted (photopic) and dark adapted (scotopic) electroretinograms. Retinal mitochondrial function and metabolism was assessed by Seahorse XFe24 Extracellular Flux Analyzer.ResultsThe retinas of aged wild type (WT) mice display a significantly reduced basal level of Xbp1s and compromised activation of ER stress response. In XBP1 cKO mice, significant structural degeneration of the retina, evidenced by thinning of retinal layers and a loss of RGCs, and functional defects indicated by diminished photopic and scotopic ERG b-waves are observed at the age of 12–14 months. Furthermore, discontinuous and disorganized synaptic laminae, colocalized with activated microglia, in the inner plexiform layer is found in the XBP1 cKO retinas. In addition, cKO mice demonstrate a significant increase in ectopic synapses between bipolar cells and photoreceptors, which is strikingly similar to WT mice at 20–24 months of age. These changes are associated with defective retinal glycolysis while mitochondrial respiratory function appears normal in the cKO retina.ConclusionsXBP1 cKO mice at 12–14 months of age show significant structural, functional, and metabolic deficits that closely resemble WT mice twice that age. Our findings suggest that the absence of XBP1, a critical component of the UPR, accelerates age-related retinal neurodegeneration.

Α-Melanocyte-Stimulating Hormone Protects Early Diabetic Retina from Blood-Retinal Barrier Breakdown and Vascular Leakage via MC4R

Background/Aims: Blood-retinal barrier (BRB) breakdown and vascular leakage is the leading cause of blindness of diabetic retinopathy (DR). Hyperglycemia-induced oxidative stress and inflammation are primary pathogenic factors of this severe DR complication. An effective interventional modality against the pathogenic factors during early DR is needed to curb BRB breakdown and vascular leakage. This study sought to examine the protective effects of α-Melanocyte-stimulating hormone (α-MSH) on early diabetic retina against vascular hyperpermeability, electrophysiological dysfunction, and morphological deterioration in a rat model of diabetes and probe the mechanisms underlying the α-MSH’s anti-hyperpermeability in both rodent retinas and simian retinal vascular endothelial cells (RF6A). Methods: Sprague Dawley rats were injected through tail vein with streptozotocin to induce diabetes. The rats were intravitreally injected with α-MSH or saline at Week 1 and 3 after hyperglycemia. In another 2 weeks, Evans blue assay, transmission electron microscopy, electroretinogram (ERG), and hematoxylin and eosin (H&E) staining were performed to examine the protective effects of α-MSH in diabetic retinas. The expression of pro-inflammatory factors and tight junction at mRNA and protein levels in retinas was analyzed. Finally, the α-MSH’s anti-hyperpermeability was confirmed in a high glucose (HG)-treated RF6A cell monolayer transwell culture by transendothelial electrical resistance (TEER) measurement and a fluorescein isothiocyanate-Dextran assay. Universal or specific melanocortin receptor (MCR) blockers were also employed to elucidate the MCR subtype mediating α-MSH’s protection. Results: Evans blue assay showed that BRB breakdown and vascular leakage was detected, and rescued by α-MSH both qualitatively and quantitatively in early diabetic retinas; electron microscopy revealed substantially improved retinal and choroidal vessel ultrastructures in α-MSH-treated diabetic retinas; scotopic ERG suggested partial rescue of functional defects by α-MSH in diabetic retinas; and H&E staining revealed significantly increased thickness of all layers in α-MSH-treated diabetic retinas. Mechanistically, α-MSH corrected aberrant transcript and protein expression of pro-inflammatory factor and tight junction genes in the diseased retinas; moreover, it prevented abnormal changes in TEER and permeability in HG-stimulated RF6A cells, and this anti-hyperpermeability was abolished by a universal MCR blocker or an antagonist specific to MC4R. Conclusions: This study showed previously undescribed protective effects of α-MSH on inhibiting BRB breakdown and vascular leakage, improving electrophysiological functions and morphology in early diabetic retinas, which may be due to its down-regulating pro-inflammatory factors and augmenting tight junctions. α-MSH acts predominantly on MC4R to antagonize hyperpermeability in retinal microvessel endothelial cells.

Morphological analyses of the retinal photoreceptor cells in the nocturnally adapted owl monkeys.

Owl monkeys are the only one species possessing the nocturnal lifestyles among the simian monkeys. Their eyes and retinas have been interested associating with the nocturnal adaptation. We examined the cellular specificity and electroretinogram (ERG) reactivity in the retina of the owl monkeys by comparison with the squirrel monkeys, taxonomically close-species and expressing diurnal behavior. Owl monkeys did not have clear structure of the foveal pit by the funduscope, whereas the retinal wholemount specimens indicated a small-condensed spot of the ganglion cells. There were abundant numbers of the rod photoreceptor cells in owl monkeys than those of the squirrel monkeys. However, the owl monkeys’ retina did not possess superiority for rod cell-reactivity in the scotopic ERG responses. Scanning electron microscopic observation revealed that the rod cells in owl monkeys’ retina had very small-sized inner and outer segments as compared with squirrel monkeys. Owl monkeys showed typical nocturnal traits such as rod-cell dominance. However, the individual photoreceptor cells seemed to be functionally weak for visual capacity, caused from the morphological immaturity at the inner and outer segments.

Effects of Prenatal Alcohol Exposure on the Visual System of Monkeys Measured at Different Stages of Development

Fetal alcohol spectrum disorder (FASD) is a developmental disease characterized by behavioral problems and physical defects including malformations of the eye and associated optical defects. How these malformations affect retinal functioning is not well known, although animal models have suggested that scotopic vision is particularly deficient. Age is also known to affect scotopic vision. Here, we determined the combined effects of age and fetal alcohol exposure (FAE) on retinal function using full-field electroretinograms (ERGs) in monkeys (Chlorocebus sabaeus). ERGs were recorded in monkeys aged 3- to 12-years old, at multiple flash intensities under scotopic and photopic conditions, and functions were fit to the amplitudes of the a- and b-waves. We found that both age and alcohol exposure affected ERGs. In photopic ERGs, amplitudes increased with age, and were higher in FAEs than controls, for data related to the OFF- and ON-pathways. In scotopic ERGs, amplitudes were decreased in young FAE compared with age-matched controls but only for the rod-dominated responses, while at brighter flashes, alcohol exposure led to an increase in the amplitude of the a- and b-waves. The ERGs from the FAE animals closely resembled the data from the older sucrose-control monkeys. This suggests that the FAE monkey retina ages more quickly than the control monkeys. This large sample of nonhuman primates, with carefully monitored ethanol exposure, demonstrates the critical interplay between age and alcohol when assessing the integrity of the retina. We suggest that ERGs might be an important adjunct to diagnosing human FASD.

Electroretinographic Assessment of Inner Retinal Signaling in the Isolated and Superfused Murine Retina

ABSTRACTPurpose: Longer-lasting electroretinographic recordings of the isolated murine retina were initially achieved by modification of a phosphate-buffered nutrient solution originally developed for the bovine retina. During experiments with a more sensitive mouse retina, apparent model-specific limitations were addressed and improvements were analyzed for their contribution to an optimized full electroretinogram (ERG).Material and methods: Retinas were isolated from dark-adapted mice, transferred to a recording chamber and superfused with different solutions. Scotopic and photopic ERGs were recorded with white flashes every 3 minutes. The phosphate buffer (Sickel-medium) originally used was replaced by a carbonate-based system (Ames-medium), the pH of which was adjusted to 7.7–7.8. Moreover, addition of 0.1 mM BaCl2 was investigated to reduce b-wave contamination by the slow PIII component typically present in the murine ERG.Results: B-wave amplitudes were increased by the pH-shift (pH 7.4 to pH 7.7) from 22.9 ± 1.9 µV to 37.5 ± 2.5 µV. Improved b-wave responses were also achieved by adding small amounts of Ba2+ (100 µM), which selectively suppressed slow PIII components, thereby unmasking more of the true b-wave amplitude (100.0% with vs. 22.2 ± 10.7% without Ba2+). Ames medium lacking amino acids and vitamins was unable to maintain retinal signaling, as evident in a reversible decrease of the b-wave to 31.8 ± 3.9% of its amplitude in complete Ames medium.Conclusions: Our findings provide optimized conditions for ex vivo ERGs from the murine retina and suggest that careful application of Ba2+ supports reliable isolation of b-wave responses in mice. Under our recording conditions, murine retinas show reproducible ERGs for up to six hours.

Neuroprotective effect of bilberry extract in a murine model of photo-stressed retina.

Excessive exposure to light promotes degenerative and blinding retinal diseases such as age-related macular degeneration and retinitis pigmentosa. However, the underlying mechanisms of photo-induced retinal degeneration are not fully understood, and a generalizable preventive intervention has not been proposed. Bilberry extract is an antioxidant-rich supplement that ameliorates ocular symptoms. However, its effects on photo-stressed retinas have not been clarified. In this study, we examined the neuroprotective effects of bilberry extract against photo-stress in murine retinas. Light-induced visual function impairment recorded by scotopic and phototopic electroretinograms showing respective rod and cone photoreceptor function was attenuated by oral administration of bilberry extract through a stomach tube in Balb/c mice (750 mg/kg body weight). Bilberry extract also suppressed photo-induced apoptosis in the photoreceptor cell layer and shortening of the outer segments of rod and cone photoreceptors. Levels of photo-induced reactive oxygen species (ROS), oxidative and endoplasmic reticulum (ER) stress markers, as measured by real-time reverse transcriptase polymerase chain reaction, were reduced by bilberry extract treatment. Reduction of ROS by N-acetyl-L-cysteine, a well-known antioxidant also suppressed ER stress. Immunohistochemical analysis of activating transcription factor 4 expression showed the presence of ER stress in the retina, and at least in part, in Müller glial cells. The photo-induced disruption of tight junctions in the retinal pigment epithelium was also attenuated by bilberry extract, repressing an oxidative stress marker, although ER stress markers were not repressed. Our results suggest that bilberry extract attenuates photo-induced apoptosis and visual dysfunction most likely, and at least in part, through ROS reduction, and subsequent ER stress attenuation in the retina. This study can help understand the mechanisms of photo-stress and contribute to developing a new, potentially useful therapeutic approach using bilberry extract for preventing retinal photo-damage.

Test-retest reliability of scotopic full-field electroretinograms in rabbits.

To explore test-retest reliability of standard scotopic full-field ERG measurements in New Zealand White rabbits. TheERG is widely used for testing of retinal integrity after any ocular treatment. We here present detailed stimulus-response dependencies for single healthy and untreated animals, concentrating on test-retest reproducibility. Five New Zealand White rabbits (aged 8-10weeks, weight about 2.0-2.5kg) underwent binocular ERG measurements after intramuscular anesthesia and pharmaceutical pupillary dilatation at a baseline day and 10days later. Eleven scotopic flash strengths (0.0001-10cds/m2) were presented. Variability was quantified via the 95% limits of agreement (LOA). The a-waves displayed the typical monotonic sigmoid amplitude increase with flash strength, and the b-waves peaked at 0.01cds/m2, followed by a marked dip at 0.1-0.3cds/m2. LOA of both waves went through a maximum in the dip region. LOA divided by mean amplitudes (relative LOA) was fairly flat over flash strength, around 20% beyond the dip. Intraindividual interocular variability was markedly lower, around 10%. Scotopic ERGresponses in rabbits display a region of high variability at 0.1-0.3cds/m2; beyond that region the amplitude-LOA is 20%, the interocular LOA being half that value. The use of intraindividual control eyes for testing any toxicity of ocular agents thus appears markedly more sensitive. As a rule of thumb, we found the relative 95% LOA as 33% between individuals, 20% across sessions and 10% between eyes.

Retinal A2A and A3 adenosine receptors modulate the components of the rat electroretinogram.

Adenosine is a neuromodulator present in various areas of the central nervous system, including the retina. Adenosine may serve a neuroprotective role in the retina, based on electroretinogram (ERG) recordings from the rat retina. Our purpose was to assess the role of A2A and A3 adenosine receptors in the generation and modulation of the rat ERG. The flash ERG was recorded with corneal electrodes from Sprague Dawley rats. Agonists and antagonists for A2A and A3 receptors, and adenosine were injected (5 µl) into the vitreous. The effects on the components of the single flash scotopic and photopic ERGs were examined, and ERG flicker. Adenosine (0.5 mM) increased the mean amplitudes of the scotopic ERG a-waves (68 ± 8 to 97 ± 14 µV, P = 0.042), and b-waves (236 ± 38 µV to 305 ± 42 µV). A2A agonist CGS21680 (2 mM) reduced the mean amplitude of the ERG b-wave, from 298 ± 21 µV in response to the brightest stimulus to 212 ± 19 µV (P = 0.005), and mean scotopic oscillatory potentials (OPs) from 100 ± 9 µV to 47 ± 11 µV (P = 0.023). ZM241385 [4 mM], an A2A antagonist, decreased the scotopic b-wave of the ERG. A3 agonist 2-CI-IB-MECA (0.5 mM) increased the a-wave, while decreasing the scotopic and photopic ERG b-waves, and the scotopic OPs. A3 antagonist VUF5574 (1 mM) increased the mean amplitude of the scotopic a-wave (66 ± 8 to 140 ± 29 µV, P = 0.046) and b-wave (224 ± 20 to 312 ± 39 µV, P = 0.0037). No significant effects on ERG flicker were found. We conclude that retinal neurons containing A2A and/or A3 adenosine receptors contribute to the generation of the ERG a- and b-waves and OPs.

Spermidine Oxidation-Mediated Degeneration of Retinal Pigment Epithelium in Rats.

Retinal pigment epithelium (RPE) degeneration is a crucial event in dry age-related macular degeneration and gyrate atrophy. The polyamine spermidine has been shown to induce RPE cell death in vitro. The present study aimed to establish a novel in vivo model of spermidine-induced RPE degeneration and to determine whether spermidine-induced RPE cell death involves oxidative mechanisms. In this study, spermidine caused ARPE-19 cell death in a concentration-dependent manner. This effect was prevented by removal of serum from the culture medium or treatment with amine oxidase inhibitors, N-acetylcysteine (NAC), or aldehyde dehydrogenase (ALDH). Intravitreal injection of spermidine into rats significantly increased the permeability of the blood-retinal barrier and decreased the amplitudes of scotopic electroretinogram a- and b-waves. Histological analysis revealed that spermidine induced vacuolation, atrophy, and dropout of RPE cells, leading to the disruption of photoreceptor outer segments. Simultaneous intravitreal administration of NAC and ALDH with spermidine prominently inhibited the functional and morphological changes induced by spermidine. In conclusion, this study demonstrated that the intravitreal administration of spermidine induced RPE cell dysfunction and death followed by photoreceptor degeneration in rats. These effects of spermidine are thought to be mediated by oxidative stress and a toxic aldehyde generated during spermidine oxidation.

Chemogenetic Activation of ipRGCs Drives Changes in Dark-Adapted (Scotopic) Electroretinogram.

PurposeThe purpose of this study was to investigate the impact of activating melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs) on dark-adapted (scotopic) electroretinograms (ERG).MethodsWe used mice (Opn4Cre/+) expressing cre recombinase in melanopsin-expressing cells for a targeted gene delivery of a chemogenetic Gq-coupled receptor, hM3Dq, to ipRGCs. Intraperitoneal injection of clozapine N-oxide (CNO) at 5 mg/kg was used for acute activation of hM3Dq and thus excitation of ipRGCs in darkness. Dark-adapted flash ERGs were recorded across a 9-fold range of irradiances from hM3Dq Opn4Cre/+ and control Opn4Cre/+ mice before and after intraperitoneal injection of CNO. A- and b-wave amplitudes and implicit times and oscillatory potentials (OPs) were analyzed. Paired-flash stimuli were used to isolate cone-driven responses.ResultsClozapine N-oxide application suppressed a- and b-wave amplitudes of the dark-adapted ERG across the flash intensity range in hM3Dq Opn4Cre/+ mice compared to control mice. Examination of the normalized irradiance-response functions revealed a shift in b-wave but not a-wave sensitivity. No changes in a- and b-wave implicit times were detected. Total OP amplitudes were also reduced in hM3Dq Opn4Cre/+ mice compared to controls following CNO administration. The paired-flash method revealed reduction in both the first (rods and cones) and second (cones only) flash response.ConclusionsAcute and selective activation of ipRGCs modulates the amplitude of both a- and b-waves of the scotopic ERG, indicating that the influence of this ganglion cell class on the retinal physiology extends to the photoreceptors as well as their downstream pathways.

Cell surgery and growth factors in dry age-related macular degeneration: visual prognosis and morphological study.

BackgroundThe aim of this research was to study the overall restoration effect on residual retinal cells through surgically grafted autologous cells onto the surrounding tissue, choroid and retina in order to produce a constant secretion of growth factors (GFs) in dry age-related macular degeneration (AMD) patients.Results6 months after surgery, several values were statistically significant in the group with higher RTA. Also patient compliance analysis (PCA) in relation to functional change perception appeared to be very good.MethodsThirty-six eyes of 25 patients (range 64-84 years of age) affected by dry AMD were included in study, and divided in two groups by spectral domain-optical coherence tomography (SD-OCT): group A with retinal thickness average (RTA) less than 250 microns (μm) and group B with RTA equal to or more than 250 μm. Adipocytes, adipose-derived stem cells from the stromal-vascular fraction, and platelets from platelet-rich plasma were implanted in the suprachoroidal space. Particularly, the following parameters were evaluated: best corrected visual acuity (BCVA) for far and near distance, retinal thickness maps, scotopic and photopic electroretinogram (ERG), and microperimetry (MY). All statistical analyses were performed with STATA 14.0 (Collage Station, Texas, USA).ConclusionsThe available set of GFs allowed biological retinal neuroenhancement. After 6 months it improved visual performance (VP), but the increase was better if RTA recorded by OCT was higher, probably in relation to the presence of areas with greater cellularity.

Ciliopathy-associated IQCB1/NPHP5 protein is required for mouse photoreceptor outer segment formation.

Null mutations in the human IQCB1/NPHP5 (nephrocystin-5) gene that encodes NPHP5 are the most frequent cause of Senior-Løken syndrome, a ciliopathy that is characterized by Leber congenital amaurosis and nephronophthisis. We generated germline Nphp5-knockout mice by placing a β-Geo gene trap in intron 4, thereby truncating NPHP5 at Leu87 and removing all known functional domains. At eye opening, Nphp5-/- mice exhibited absence of scotopic and photopic electroretinogram responses, a phenotype that resembles Leber congenital amaurosis. Outer segment transmembrane protein accumulation in Nphp5-/- endoplasmic reticulum was evident as early as postnatal day (P)6. EGFP-CETN2, a centrosome and transition zone marker, identified basal bodies in Nphp5-/- photoreceptors, but without fully developed transition zones. Ultrastructure of P6 and 10 Nphp5-/- photoreceptors revealed aberrant transition zones of reduced diameter. Nphp5-/- photoreceptor degeneration was complete at 1 mo of age but was delayed significantly in Nphp5-/-;Nrl-/- (cone only) retina. Nphp5-/- mouse embryonic fibroblast developed normal cilia, and Nphp5-/- kidney histology at 1 yr of age showed no significant pathology. Results establish that nephrocystin-5 is essential for photoreceptor outer segment formation but is dispensable for kidney and mouse embryonic fibroblast ciliary formation.-Ronquillo, C. C., Hanke-Gogokhia, C., Revelo, M. P., Frederick, J. M., Jiang, L., Baehr, W. Ciliopathy-associated IQCB1/NPHP5 protein is required for mouse photoreceptor outer segment formation.

Brimonidine Enhances the Electrophysiological Response of Retinal Ganglion Cells through the Trk-MAPK/ERK and PI3K Pathways in Axotomized Eyes

ABSTRACTPurpose: To investigate changes in retinal ganglion cell (RGC) activity by measuring the positive scotopic threshold response (pSTR) of the electroretinogram (ERG) in axotomized eyes after brimonidine injection.Methods: In 50 adult Sprague-Dawley rats, the left eye was axotomized and injected with phosphate buffered saline (PBS) or brimonidine and the contralateral right eye was left untreated. Scotopic ERGs were recorded simultaneously from both eyes on days 1, 2, 3, 7, and 10 after the intravitreal injection, and the amplitude of the a- and b-waves and the pSTR were measured. Surviving RGCs in the flat-mounted retinas were counted 10 days after axotomy. In addition to brimonidine, K252a (an inhibitor of tyrosine kinase phosphorylation of the Trk receptors), U0126 (a MAPK/ERK kinase inhibitor), and LY294002 (phosphoinositide 3-kinases [PI3Ks]) were also injected intravitreally into the left eye, and ERGs were recorded using the same protocol.Results: The pSTR amplitude increased significantly in the axotomized eyes with brimonidine, to 122.9 ± 5.0%, 161.8 ± 8.3%, and 133.6 ± 8.1% on days 1, 2, and 3 (P < 0.01), respectively, compared to the axotomized eyes treated with PBS (control). The increased pSTR amplitude returned to normal (103.6 ± 6.7%) on day 7, although there were a greater number of surviving RGCs in the treatment groups than in the controls. The intravitreal injection of K252a, U0126, or LY294002 significantly attenuated the increase in pSTR induced by intravitreal brimonidine (P < 0.01).Conclusion: Intravitreal brimonidine enhanced the survival and electrophysiological activity of the RGCs in rats. The mechanism of this electrophysiological change may involve activation of the Trk-MAPK/ERK and Trk-PI3K signals.

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

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

Characterizing the Retinal Function of Psammomys obesus: A Diurnal Rodent Model to Study Human Retinal Function

ABSTRACTPurpose: To compare the retinal function of a diurnal murid rodent, Psammomys obesus, with that of Wistar albino rat and human subjects.Materials and methods: Adult Psammomys obesus were captured and transferred to the animal facilities where they were maintained at 25°C with standard light/dark cycles and natural halophilic plants, rich in water and mineral salts. Standard full-field photopic and scotopic electroretinograms were obtained.Results: The right eye of all animals displayed well detectable and reproducible scotopic and photopic electroretinogram (ERG) responses. Results were compared with those obtained from human subjects and Wistar rats. ERG measurement showed that the amplitudes of scotopic responses in Psammomys obesus are quite similar to those of human subjects. The amplitude of the photopic a-wave was comparable to that of humans and six times higher than that of the albino rat. The amplitudes of photopic b-wave, photopic oscillatory potentials (OPs), and 30 Hz flicker were all markedly larger in Psammomys obesus compared to those obtained from human subjects and Wistar rats. Furthermore, like the human photopic ERG, the photopic ERG of Psammomys obesus also includes prominent post b-wave components (i.e. i- and d-waves) while the ERG of Wistar rats does not.Conclusions: Our results suggest that the retinal function of Psammomys obesus, especially the cone-mediated function, shares several features with that of human subjects. We believe that Psammomys obesus represents an interesting alternative to study the structure and function of the normal and diseased retina in a human-like rodent model of retinal function.

Protective effects of hydrogen-rich saline against N-methyl-N-nitrosourea-induced photoreceptor degeneration

The N-methyl-N-nitrosourea (MNU)-treated rat is typically used as an animal model of chemically-induced retinitis pigmentosa (RP). Reactive oxygen species (ROS) have been recognized as the crucial contributor to the retinal photoreceptor apoptosis seen in MNU-treated rats. In the present study, we explored the therapeutic effects of hydrogen-rich saline (HRS), a selective ROS scavenger, on MNU-induced photoreceptor degeneration. Intraperitoneal (IP) administration of HRS ameliorated MNU-induced photoreceptor degeneration in terms of morphology and function: Sharply decreased thickness of the retinal outer nuclear layer (ONL) and flattened photopic and scotopic electroretinogram (ERG) waveforms, typically seen in response to MNU treatment, were substantially rescued in rats cotreated with MNU and HRS (MNU + HRS). Moreover, the terminal deoxyuridine triphosphate nick-end labeling (TUNEL) assay revealed a smaller number of apoptotic photoreceptors in the MNU + HRS group compared that in the MNU group. Compared to MNU-treated rats, retinal malondialdehyde (MDA) content in MNU + HRS rats significantly decreased while superoxide dismutase (SOD) activity significantly increased. Morphological and multi-electrode array (MEA) analyses revealed more efficient preservation of the architecture and field potential waveforms in particularly the peripheral regions of the retinas within the MNU + HRS group, compared to that in the MNU group. However, this enhanced protection of structure and function in the peripheral retina is unlikely the result of site-dependent variation in the efficacy of HRS; rather, it is most likely due to reduced susceptibility of peripheral photoreceptors to MNU-induced degeneration. Inner retinal neuron function in the MNU + HRS rats was better preserved, with fewer apoptotic photoreceptors in the ONL. Collectively, these results support the rationale for future clinical evaluation of HRS as a therapeutic agent for human RP.