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The Electroretinogram: ERG
Abstract
As early as 1865, Holmgren found that a light stimulus could cause a change in the electrical potential of the amphibian eye. Shortly afterwards, similar findings were reported by Dewar from Scotland. He showed that light illumination through the pupil, which had previously been covered, caused a slight movement of a galvanometer, suggestive of a positive electrical change in the cornea relative to the back of the eye (1). This light-induced electrical activity of the eye was called the electroretinogram. Now, the electroretinogram response is commonly abbreviated to the ERG. Gotch (2) was the first to report that the response of the eye to a light flash consisted of two waves; first the cornea became negative and then a positive wave of larger amplitude appeared. Later, Einthoven and Jolly (3) separated the ERG response into three waves. The first wave to appear immediately after turning on a light stimulus was negative on the cornea. It was followed by a positive wave and a final slower wave that was also positive. Einthoven and Jolly (3) suggested that the light stimulus triggered a chain of reactions leading to the formation of products A, B, and C, and that every electrical wave indicated a change in a "relevant" product. These authors' work was the foundation for the form of analysis of the ERG used to the present day. The waves are called a-, b- and c-waves. An additional corneal-positive wave, that is more rarely recorded at the termination of the light flash, is called the d-wave.
... Another evidence for a threshold mechanism in the neural tissue, which is considerably different from a single spike threshold, appears in many in-vivo electrophysiological studies that involved stimuli with increasing intensities. As representative examples,we list here the studies by Ebert and Koch (1997), Frot et al. (2007), and Perlman (2007). These studies demonstrate that the electrophysiological response considerably changes after crossing a certain level of stimulus intensity. ...
... Interestingly, after crossing the threshold intensity, the electrophysiological response waveform includes a negative potential component that has an opposite polarity comparing to neural spiking response, which suggests that non-neuron cells may be involved. Figure 14a shows the threshold behaviour of the negative wave as measured in retinal studies by Perlman (2007), and Figure 14b shows a closer look on the negative electrical wave from the same study. Figures 14a-c show similar responses that were obtained by Ebert and Koch (1997) in a study that examined leg reflex movement in rats, in response to a loud auditory stimulus (startle response). ...
... Figures 14a-c show similar responses that were obtained by Ebert and Koch (1997) in a study that examined leg reflex movement in rats, in response to a loud auditory stimulus (startle response). The reflex movement appeared only when stimuli intensities were above 110 dB and they were preceded by a sharp negative EEG wave similar to the one reported by Perlman (2007). The latencies of the responses is typically several tens of milliseconds, and the total duration of the negative peak followed by the positive peak lasts less than two hundred milliseconds. ...
... Scotopic a-wave amplitude measures primarily the function of rod cells and scotopic b-wave amplitude the function of neurons that are post-synaptic to the rod cells, i.e. mostly bipolar cell function. Cone cell function is measured by photopic bwave amplitude and flicker ERG after rod cells are temporarily inactivated [26]. When a single dose of CeNPs was injected intravitreally before 3 weeks of age, we observed increased rod cell function up to 34 days post injection (dpi; Figs 1 and 2). ...
... Scotopic b-wave amplitude reflects the function of neurons in the inner retina predominately bipolar neurons which are post-synaptic to rod photoreceptor cells. Photopic b-wave amplitude and flicker ERG reflect the function of cone cells [26]. Each data point represented the average from at least three individual rats or six eyes. ...
Purpose:
Inorganic catalytic nanoceria or cerium oxide nanoparticles (CeNPs) are bona fide antioxidants that possess regenerative radical scavenging activities in vitro. Previously, we demonstrated that CeNPs had neuroprotective and anti-angiogenic properties in rodent retinal degeneration and neovascularization models. However, the cellular mechanisms and duration of the catalytic activity of CeNPs in preventing photoreceptor cell loss are still unknown. In this study, we sought to answer these questions using the P23H-1 rat, an autosomal dominant retinitis pigmentosa (adRP) model.
Methods:
A single dose of either saline or CeNPs was delivered intravitreally into the eyes of P23H-1 rats at 2-3 weeks of age. Retinal functions were examined at 3 to 7 weeks post injection. We quantified retinal proteins by Western blot analyses and counted the number of apoptotic (TUNEL+) profiles in the outer nuclear layer (ONL) of retinal sections. We measured free 8-isoprostanes to quantify lipid peroxidation in retinal tissues.
Results:
We observed increased rod and cone cell functions up to three weeks post injection. Apoptotic cells were reduced by 46%, 56%, 21%, and 24% at 3, 7, 14, 21 days, respectively, after CeNPs injection compared to saline. Additionally, reduction of lipid peroxidation in the retinas of CeNPs-treated vs saline-treated animals was detected 14 days post injection.
Conclusions:
We validated that CeNPs were effective in delaying loss of photoreceptor cell function in an adRP rat model. This represents the fourth rodent retinal disease model that shows delay in disease progression after a single application of CeNPs. We further demonstrated that CeNPs slowed the rate of photoreceptor cell death. We deduced that the catalytic activity of CeNPs in vivo in this rat model to be undiminished for at least 7 days and then declined over the next 14 days after CeNPs administration.
... Under dark-adapted conditions, rod activity is usually normal or close to normal [29,31], although a decrease in scotopic responses has been reported in some patients [32]. To date, ffERG is a common, non-invasive tool widely used in visual electrophysiology for its ability to objectively measure activity originating from the retina [33]. By manipulating the type of light stimulus and its intensity, it is possible to measure different components of the retina, such as oscillatory potentials (OPs), in addition to the typical a-and b-waves that reflect the hyperpolarization of the photoreceptors [34], and the depolarization of the bipolar cells [35] and Müller cells [36], respectively. ...
Purpose
The aim of this exploratory study is to investigate the role of S-cones in oscillatory potentials (OPs) generation by individuals with blue-cone monochromacy (BCM), retaining S-cones, and achromatopsia (ACHM), lacking cone functions.
Methods
This retrospective study analyzed data from 39 ACHM patients, 20 BCM patients, and 26 controls. Central foveal thickness was obtained using spectral-domain optical coherence tomography, while amplitude and implicit time (IT) of a- and b-waves were extracted from the ISCEV Standard dark-adapted 3 cd.s.m⁻² full-field ERG (ffERG). Time–frequency analysis of the same measurement enabled the extraction of OPs, providing insights into the dynamic characteristics of the recorded signal.
Results
Both ACHM and BCM groups showed a significant reduction (p < .00001) of a- and b-wave amplitudes and ITs as well as the power of the OPs compared to the control groups. The comparison between ACHM and BCM didn’t show any statistically significant differences in the electrophysiological parameters. The analysis of covariance revealed significantly reduced central foveal thickness in the BCM group compared to ACHM and controls (p < .00001), and in ACHM compared to controls (p < .00001), after age correction and Tukey post-hoc analysis.
Conclusions
S-cones do not significantly influence OPs, and the decline in OPs' power is not solely due to a reduced a-wave. This suggests a complex non-linear network influenced by photoreceptor inputs. Morphological changes don’t correlate directly with functional alterations, prompting further exploration of OPs’ function and physiological role.
... Electroretinography, ERG, for review see [7,8], measures the electrical responses of the retina evoked by light stimulation from the eye surface. It is a summed activity of multiple retinal cell populations and consists of overlapping positive and negative potentials that originate from different stages of retinal processing. ...
The purpose of the study is to demonstrate coherent optical tomography and electroretinography techniques adopted from the human clinical practice to assess the morphology and function of the mouse retina in a high-throughput phenotyping environment. We present the normal range of wild-type C57Bl/6NCrl retinal parameters in six age groups between 10 and 100 weeks as well as examples of mild and severe pathologies resulting from knocking out a single protein-coding gene. We also show example data obtained by more detailed analysis or additional methods useful in eye research, for example, the angiography of a superficial and deep vascular complex. We discuss the feasibility of these techniques in conditions demanding a high-throughput approach such as the systemic phenotyping carried out by the International Mouse Phenotyping Consortium.
... It has previously been reported that degenerative changes in the retina are early events in the development of DR 43 . Therefore, we tested retinal function by electroretinography (ERG), which allows for effective assessment of photoreceptor activity (a-wave), Müller and bipolar cell activity (b-wave), and RPE function (c-wave) 44 . With prolonged diabetes (4 months after induction) the amplitudes of dark-adapted scotopic a-and b-waves (Fig. 1a-c), as well as the lightadapted photopic b-wave were reduced in Akt2 fl/fl mice compared to nondiabetic controls at high flash levels (Fig. 1d, e). ...
The retinal pigment epithelium (RPE) plays an important role in the development of diabetic retinopathy (DR), a leading cause of blindness worldwide. Here we set out to explore the role of Akt2 signaling—integral to both RPE homeostasis and glucose metabolism—to DR. Using human tissue and genetically manipulated mice (including RPE-specific conditional knockout (cKO) and knock-in (KI) mice), we investigate whether Akts in the RPE influences DR in models of diabetic eye disease. We found that Akt1 and Akt2 activities were reciprocally regulated in the RPE of DR donor tissue and diabetic mice. Akt2 cKO attenuated diabetes-induced retinal abnormalities through a compensatory upregulation of phospho-Akt1 leading to an inhibition of vascular injury, inflammatory cytokine release, and infiltration of immune cells mediated by the GSK3β/NF-κB signaling pathway; overexpression of Akt2 has no effect. We propose that targeting Akt1 activity in the RPE may be a novel therapy for treating DR.
... That this is the case is also suggested by the toxic changes evident in the RPE cells themselves. Given the extensive damage done to the outer retina, it is not surprising that rescue of Kcnj13 expression in the RPE is not accompanied by complete recovery of the a-and b-wave amplitudes, but does show recovery of the c-wave, which is generated by the RPE, at least in rodents (Perlman et al., 1995). ...
Purpose: We constructed and characterized knockout and conditional knockout mice for KCNJ13, encoding the inwardly rectifying K ⁺ channel of the Kir superfamily Kir7.1, mutations in which cause both Snowflake Vitreoretinal Degeneration (SVD) and Retinitis pigmentosa (RP) to further elucidate the pathology of this disease and to develop a potential model system for gene therapy trials.
Methods: A Kcnj13 knockout mouse line was constructed by inserting a gene trap cassette expressing beta-galactosidase flanked by FRT sites in intron 1 with LoxP sites flanking exon two and converted to a conditional knockout by FLP recombination followed by crossing with C57BL/6J mice having Cre driven by the VMD2 promoter. Lentiviral replacement of Kcnj13 was driven by the EF1a or VMD2 promoters.
Results: Blue-Gal expression is evident in E12.5 brain ventricular choroid plexus, lens, neural retina layer, and anterior RPE. In the adult eye expression is seen in the ciliary body, RPE and choroid. Adult conditional Kcnj13 ko mice show loss of photoreceptors in the outer nuclear layer, inner nuclear layer thinning with loss of bipolar cells, and thinning and disruption of the outer plexiform layer, correlating with Cre expression in the overlying RPE which, although preserved, shows morphological disruption. Fundoscopy and OCT show signs of retinal degeneration consistent with the histology, and photopic and scotopic ERGs are decreased in amplitude or extinguished. Lentiviral based replacement of Kcnj13 resulted in increased ERG c- but not a- or b- wave amplitudes.
Conclusion: Ocular KCNJ13 expression starts in the choroid, lens, ciliary body, and anterior retina, while later expression centers on the RPE with no/lower expression in the neuroretina. Although KCNJ13 expression is not required for survival of the RPE, it is necessary for RPE maintenance of the photoreceptors, and loss of the photoreceptor, outer plexiform, and outer nuclear layers occur in adult KCNJ13 cKO mice, concomitant with decreased amplitude and eventual extinguishing of the ERG and signs of retinitis pigmentosa on fundoscopy and OCT. Kcnj13 replacement resulting in recovery of the ERG c- but not a- and b-waves is consistent with the degree of photoreceptor degeneration seen on histology.
... Eleven of these reported use of visual evoked potentials (VEPs), 21,22,29,33,46,48,51,57,59,65,67 five articles used electroretinography (ERG) 33,43,44,46,57 and seven used electroencephalography (EEG). 21,22,33,34,41,46,51 Each of these electrophysiology techniques measure different functions; ERGs assess the functional integrity of the retina, 93 VEPs are used to determine the integrity of the visual pathways from the macula to the visual cortex 94 and EEG is used in the diagnosis of neurological disease and to monitor brain activity. 95 A score of 2 was assigned to articles that reported use of VEP, as this provides relevant information on the function of the primary visual pathway. ...
Purpose
Cerebral visual impairment (CVI) is the leading cause of childhood visual impairment in the developed world. Despite this, there are no agreed clinical guidelines for the investigation and diagnosis of the condition. Before development of such guidelines can commence, it is important to recognise which approaches are currently employed. This systematic review evaluated the literature to identify which methods of assessment are currently used to investigate and diagnose childhood CVI.
Methods
Medline, Embase, CINAHL, Scopus and the Cochrane Library databases were systematically searched in January 2020 using defined search terms. Articles were included if they: (i) were research papers, conference abstracts or research protocols published in peer‐reviewed scientific journals, or relevant textbooks; (ii) included a clinical investigation of CVI in children; (iii) provided an explanation or criteria to diagnose CVI and (iv) were specifically investigating cerebral/cortical visual impairment. Methods used to a) assess and b) diagnose CVI were extracted from included articles. ‘Assessment scores’ were assigned for each method employed by researchers to investigate and diagnose CVI to quantify and compare approaches between articles. A quality grading was also applied to each article.
Results
Of 6454 identified articles, 45 met the inclusion criteria. From these, 10 categories of assessment utilised within included articles were identified: (1) Medical history, (2) Vision assessment/ophthalmologic examination, (3) Neuroimaging, (4) Visual behaviour and direct observation, (5) Structured history‐taking, (6) Visual perception tests, (7) Ocular movement and posture assessment, (8) Intelligence/IQ assessment, (9) Clinical electrophysiology and (10) Neurodevelopmental tests. In terms of diagnostic criteria, the most commonly reported approach was one of exclusion, i.e., CVI was diagnosed when visual dysfunction could not be attributed to abnormalities detected in the anterior visual pathway.
Conclusion
There is a lack of common practice in the approaches used by clinicians to investigate and diagnose CVI in children. At present, a ‘diagnosis of exclusion’ remains the most common means to diagnose CVI. Development of clinical guidelines for assessment and diagnosis are necessary to ensure consistency in the diagnosis of CVI and the timely implementation of support to alleviate the impact of CVI on the child’s daily living.
... We analysed the amplitudes of main ERG componentsa-wave and b-wave-in the responses to blue and red brief flashes. These components are assumed to represent the function of photoreceptors and bipolar cells, respectively [23]. During the experimental protocol, we recorded the responses to flashes with increasing intensity (a discretization of 0.5 orders of magnitude in photons mm −2 ) for each of the prepared retinal quadrants ( figure 3). ...
Migratory birds are known to be sensitive to external magnetic field (MF). Much indirect evidence suggests that the avian magnetic compass is localized in the retina. Previously, we showed that changes in the MF direction could modulate retinal responses in pigeons. In the present study, we performed similar experiments using the traditional model animal to study the magnetic compass, European robins. The photoresponses of isolated retina were recorded using ex vivo electroretinography (ERG). Blue- and red-light stimuli were applied under an MF with the natural intensity and two MF directions, when the angle between the plane of the retina and the field lines was 0° and 90°, respectively. The results were separately analysed for four quadrants of the retina. A comparison of the amplitudes of the a- and b-waves of the ERG responses to blue stimuli under the two MF directions revealed a small but significant difference in a- but not b-waves, and in only one (nasal) quadrant of the retina. The amplitudes of both the a- and b-waves of the ERG responses to red stimuli did not show significant effects of the MF direction. Thus, changes in the external MF modulate the European robin retinal responses to blue flashes, but not to red flashes. This result is in a good agreement with behavioural data showing the successful orientation of birds in an MF under blue, but not under red illumination.
... If the computer failed to calculate the proper point, experimenters would manually measure the results. Since b wave represents the transduction of extracellular currents and is considered to be the major component of the human ERG recording as used in clinical and experimental analysis of retinal function (Perlman, 1995), we typically focus on the amplitude of b wave in the present study. ...
Retinal degenerative diseases (RDDs) are the leading causes of blindness and currently lack effective treatment. Cytotherapy has become a promising strategy for RDDs. The transplantation of olfactory ensheathing cells (OECs) or neural stem cells (NSCs) has recently been applied for the experimental treatment of RDDs. However, the long-term outcomes of single-cell transplantation are poor. The combined transplantation of multiple types of cells might achieve better effects. In the present study, OECs [containing olfactory nerve fibroblasts (ONFs)] and NSCs were cotransplanted into the subretinal space of Royal College of Surgeons (RCS) rats. Using electroretinogram (ERG), immunofluorescence, Western blot, and in vitro Transwell system, the differences in the electrophysiological and morphological changes of single and combined transplantation as well as the underlying mechanisms were explored at 4, 8, and 12 weeks postoperation. In addition, using the Transwell system, the influence of OECs on the stemness of NSCs was discovered. Results showed that, compared to the single transplantation of OECs or NSCs, the combined transplantation of OECs and NSCs produced greater improvements in b-wave amplitudes in ERGs and the thickness of the outer nuclear layer at all three time points. More endogenous stem cells were found within the retina after combined transplantation. Glial fibrillary acidic protein (GFAP) expression decreased significantly when NSCs were cotransplanted with OECs. Both the vertical and horizontal migration of grafted cells were enhanced in the combined transplantation group. Meanwhile, the stemness of NSCs was also better maintained after coculture with OECs. Taken together, the results suggested that the combined transplantation of NSCs and OECs enhanced the improvement in retinal protection in RCS rats, providing a new strategy to treat RDDs in the future.
... Electroretinogram is one of the most efficient electrophysiological techniques for detecting retinal function (Perlman, 1995;Meigen, 2015). We explored the effect of IRF3 deficiency on retinal function by ERG. ...
Age-related changes in visual function and retina structure are very common in aged animals, but the underlying mechanisms of these changes remain unclear. Here we report that the expression of interferon regulatory factor 3 (IRF3), a critical immune regulatory factor, is dramatically down-regulated in mouse retinas during aging. To address the role of IRF3 in the retina, we examined the structure and function of retinas in young (3-4 months) and old (22-24 months) Irf3-/- mice in comparison to age-matched wildtype (WT) mice. We found that IRF3 deletion resulted in impaired electroretinogram (ERG) responses and decreased retinal thickness in both young and old mice. In addition, numerous synapses of the outer plexiform layer (OPL) were found obviously extending into outer nuclear layer (ONL) in Irf3-/- mice, along with a reduction of the average synapse density in the OPL. These changes suggest that IRF3 deletion may accelerate retinal senescence. In support of this hypothesis, a number of classic senescence-associated markers were found in remarkably elevated level in Irf3-/- retina, including p53, p16INK4a, inositol-requiring enzyme 1α (IREα), p-H2A.X and promyelocytic leukemia protein (PML). Overall, our results indicate that maintenance normal IRF3 levels is necessary for retinal structure and function and suggest that IRF3 is an important regulator of retinal senescence.
... Scholl et al demonstrated significantly phase-advanced cone-driven ERG responses in BVMD patients, 19 in accordance with our findings of faster pupillary responses in BVMD patients. Furthermore, faster ERG responses are recorded using incremental light intensities, 21 which may suggest that loss of pigment and RPE cells in BVMD may allow more light to reach the photoreceptors which may induce faster responses to relatively increase the sensation of light. Specifically, measurement of LMCV in a test point at the center of the VF (which correlates with the primary location of BVMD pathology) demonstrated high sensitivity and specificity in discriminating between BVMD patients and controls. ...
Purpose
To determine the pupil response of Best vitelliform macular dystrophy (BVMD) patients for focal blue and red light stimuli presented at 76 test points in a 16.2° visual field (VF) using a chromatic pupilloperimeter.
Methods
An observational study was conducted in 16 participants: 7 BVMD patients with a heterozygous BEST1 mutation and 9 similar-aged controls. All participants were tested for best-corrected visual acuity, chromatic pupilloperimetry and Humphrey perimetry. Percentage of pupil contraction (PPC), maximal pupil contraction velocity (MCV) and latency of MCV (LMCV) were determined.
Results
The mean PPC and MCV recorded in BVMD patients in response to red stimuli were lower by >2 standard errors (SEs) from the mean of controls in 47% and 43% of VF test points, respectively. The mean PPC and MCV recorded in the patients in response to blue stimuli were lower by >2 SEs from the mean of controls in 36% and 24% of VF test points, respectively. The patients’ mean and median MCV recorded in response to red light correlated with their Humphrey mean deviation score (r=−0.714, P=0.071 and r=−0.821, P=0.023, respectively) and visual acuity (r=0.709, P=0.074 and r=0.655, P=0.111, respectively). A substantially shorter mean LMCV was recorded in BVMD patients compared to controls in 54% and 93% of VF test points in response to red and blue light, respectively. Receiver operating characteristic analysis for LMCV in response to red light identified a test point at the center of the VF with high diagnostic accuracy (area under the curve of 0.94).
Conclusion
Chromatic pupilloperimetry may potentially be used for objective noninvasive assessment of rod and cone cell function in different locations of the retina in BVMD patients.
... The features observed with day 21 chick retina include an early receptor potential (ERP) which is a 2 ms biphasic waveform with a latency of <1 µs after light stimulation. The instantaneous ERP response which represents the dipole changes [28] due to the conformational changes in the receptors (VRsvisual rhodopsins) that follow photon absorption, prominently can be observed in the sensor recordings. ...
Quantitative assessment involving noncontact electrical methods without the need for microscopy can enable a remote user to monitor the cell growth and development at different stages. In this line of pursuit, the utility of ultrahigh impedance, air‐suspended capacitively coupled electric potential sensors to monitor electrically active cells is examined. Standard cell lines and chick retinal explants on a variety of optoelectronically active polymer substrates are utilized to demonstrate the utility of this sensor placed in proximity without physical contact. Burst firing in developing chick retina in the early stage when photoreceptors are not developed can be observed via the photoexcitation of optoelectronic substrates using this noncontact sensing method.
... Other aspects of the electrophysiology techniques may also limit estimates of spatial resolving power. For example, as both species have a horizontal visual streak of higher resolution, behavioural measures may better reflect peak resolving power, since PERG responses are a summed response of all cells across the retina (Granit 1933;Perlman 1995). In addition, contrast sensitivity estimates based on eyecup preparations were lower than those from anaesthetised sharks. ...
In most animals, vision plays an important role in detecting prey, predators and conspecifics. The effectiveness of vision in assessing cues such as motion and shape is influenced by the ability of the visual system to detect changes in contrast in both space and time. Understanding the role vision plays in shark behaviour has been limited by a lack of knowledge about their temporal resolution, contrast sensitivity and spatial resolution. In this study, an electrophysiological approach was used to compare these measures across five species of sharks: Chiloscyllium punctatum, Heterodontus portusjacksoni, Hemiscyllium ocellatum, Mustelus mustelus and Haploblepharus edwardsii. All shark species were highly sensitive to brightness contrast and were able to detect contrast differences as low as 1.6%. Temporal resolution of flickering stimuli ranged from 28 to 44 Hz. Species that inhabit brighter environments were found to have higher temporal resolution. Spatial resolving power was estimated in C. punctatum, H. portusjacksoni and H. ocellatum and ranged from 0.10 to 0.35 cycles per degree, which is relatively low compared to other vertebrates. These results suggest that sharks have retinal adaptations that enhance contrast sensitivity at the expense of temporal and spatial resolution, which is beneficial for vision in dimly lit and/or low contrast aquatic environments.
... There is a general consensus that the neuronal generator of the b-wave is primarily the depolarizing (ON) bipolar cells, while the d-wave depends mainly on the activity of hyperpolarizing (OFF) bipolar cells with minor contribution of the photoreceptor response at stimulus offset and activity of proximal retinal neurons (see reviews [ 10,11 ]). Thus, the effects of histamine on the ERG band d-wave could be related to an altered activity of the ON and OFF bipolar cells. ...
The role of histamine in visual information processing in the retina is largely unknown. In this study we investigated the effects of histamine and the selective H1 receptor agonist TFMH on the b-wave (ON response) and d-wave (OFF response) of dark adapted frog electroretinogram (ERG). We obtained that 5 μM histamine enhanced the amplitude of both the ON and OFF responses and shortened their implicit time. The selective H1 receptor agonist TFMH (5 μM) also enhanced the amplitude of both ERG waves. Its effect upon the OFF response was expressed in the same degree as that of histamine, while its effect upon the ON response was less pronounced. Our results demonstrate that histamine has a significant role in visual information processing through the retinal ON and OFF channels and that a part of its action is mediated by H1 receptors.
... *P < 0.05, **P < 0.01, Student's t-test. [54]. Thus, we speculated that the possible reasons for increased ERG a-or b-wave amplitudes were come from the de-differentiation of Müller cells which gradually acquired a neural fate and established a robust integration with second order neurons, and formed efficient visual functional units. ...
Retinal regeneration and repair are severely impeded in higher mammalian
animals. Although Müller cells can be activated and show some characteristics of
progenitor cells when injured or under pathological conditions, they quickly form
gliosis scars. Unfortunately, the basic mechanisms that impede retinal regeneration
remain unknown. We studied retinas from Royal College of Surgeon (RCS) rats and
found that let-7 family molecules, let-7e and let-7i, were significantly overexpressed in
Müller cells of degenerative retinas. It demonstrated that down-regulation of the RNA
binding protein Lin28B was one of the key factors leading to the overexpression of let-
7e and let-7i. Lin28B ectopic expression in the Müller cells suppressed overexpression
of let-7e and let-7i, stimulated and mobilized Müller glia de-differentiation,
proliferation, promoted neuronal commitment, and inhibited glial fate acquisition of
de-differentiated Müller cells. ERG recordings revealed that the amplitudes of a-wave
and b-wave were improved significantly after Lin28B was delivered into the subretinal
space of RCS rats. In summary, down-regulation of Lin28B as well as up-regulation
of let-7e and let-7i may be the main factors that impede Müller cell de-differentiation
and proliferation in the retina of RCS rats.
... In fact, the b-wave is not a direct reflection of photoreceptors but its amplitude correlates directly with the activity of bipolar cells [86]. However, bipolar cells are post-synaptic to [87], and activated by, the photoreceptors. In the ERG examination, light is the only stimulation and the photoreceptors are the only cells of our body which respond to light stimulation. ...
Introduction
Degenerative retinal diseases like age-related macular degeneration (AMD) are the leading cause of blindness. Cell transplantation showed promising therapeutic effect for such diseases, and embryonic stem cell (ESC) is one of the sources of such donor cells. Here, we aimed to generate retinal progenitor cells (RPCs) from rat ESCs (rESCs) and to test their therapeutic effects in rat model.
Methods
The rESCs (DA8-16) were cultured in N2B27 medium with 2i, and differentiated to two types of RPCs following the SFEBq method with modifications. For rESC-RPC1, the cells were switched to adherent culture at D10, while for rESC-RPC2, the suspension culture was maintained to D14. Both RPCs were harvested at D16. Primary RPCs were obtained from P1 SD rats, and some of them were labeled with EGFP by infection with lentivirus. To generate Rax::EGFP knock-in rESC lines, TALENs were engineered to facilitate homologous recombination in rESCs, which were cotransfected with the targeting vector and TALEN vectors. The differentiated cells were analyzed with live image, immunofluorescence staining, flow cytometric analysis, gene expression microarray, etc. RCS rats were used to mimic the degeneration of retina and test the therapeutic effects of subretinally transplanted donor cells. The structure and function of retina were examined.
Results
We established two protocols through which two types of rESC-derived RPCs were obtained and both contained committed retina lineage cells and some neural progenitor cells (NPCs). These rESC-derived RPCs survived in the host retinas of RCS rats and protected the retinal structure and function in early stage following the transplantation. However, the glia enriched rESC-RPC1 obtained through early and longer adherent culture only increased the b-wave amplitude at 4 weeks, while the longer suspension culture gave rise to evidently neuronal differentiation in rESC-RPC2 which significantly improved the visual function of RCS rats.
Conclusions
We have successfully differentiated rESCs to glia enriched RPCs and retinal neuron enriched RPCs in vitro. The retinal neuron enriched rESC-RPC2 protected the structure and function of retina in rats with genetic retinal degeneration and could be a candidate cell source for treating some degenerative retinal diseases in human trials.
... As shown in Fig. 14(c), if a pure photoreceptor response is recorded, i.e., postphotoreceptor neurons are blocked, the a-wave may need more time to recover back to the baseline, which results in a longer time to reach a peak compared with the a-wave of a full-field ERG. [138][139][140][141] From this perspective, if we assume the rising-phase IOSs originate from retinal photoreceptors, the measured time to peak of the IOS should be longer than that of the a-wave of a full-field ERG, which is consistent with the experimental results in Fig. 13. ...
Intrinsic optical signal (IOS) imaging promises to be a noninvasive method for high-resolution examination of retinal physiology, which can advance the study and diagnosis of eye diseases. While specialized optical instruments are desirable for functional IOS imaging of retinal physiology, in depth understanding of multiple IOS sources in the complex retinal neural network is essential for optimizing instrument designs. We provide a brief overview of IOS studies and relationships in rod outer segment suspensions, isolated retinas, and intact eyes. Recent developments of line-scan confocal and functional optical coherence tomography (OCT) instruments have allowed in vivo IOS mapping of photoreceptor physiology. Further improvements of the line-scan confocal and functional OCT systems may provide a feasible solution to pursue functional IOS mapping of human photoreceptors. Some interesting IOSs have already been detected in inner retinal layers, but better development of the IOS instruments and software algorithms is required to achieve optimal physiological assessment of inner retinal neurons. © 2015 Society of Photo-Optical Instrumentation Engineers (SPIE).
... The effects of ON channel blockade on the OFF channel activity in distal retina have been investigated in two ways: 1) by recording the activity of individual OFF bipolar cells during retinal APB treatment and 2) by recording the electroretinographic (ERG) OFF response (d-wave) during the same treatment. It is known that the activity of large populations of ON and OFF bipolar cells is reflected in the b-and d-waves of the diffuse ERG [reviews: 119,120]. ...
In the vertebrate retina, visual signals are segregated into parallel ON and OFF pathways, which
provide information for light increments and decrements. The segregation is first evident at the level of the
ON and OFF bipolar cells and it apparently remains as signals propagate to higher brain visual centers. A
fundamental question in visual neuroscience is how these two parallel pathways function: are they
independent from each other or do they interact somehow? In the latter case, what kinds of mechanisms are
involved and what are the consequences from this cross-talk? This review summarizes current knowledge
about the types of interactions between the ON and OFF channels in nonmammalian and mammalian retina. Data
concerning the ON-OFF interactions in distal retina revealed by recording of single bipolar cell activity and
electroretinographic ON (b-wave) and OFF (d-wave) responses are presented. Special emphasis is put on the ON-OFF
interactions in proximal retina and their dependence on the state of light adaptation in mammalian retina. The involvement
of the GABAergic and glycinergic systems in the ON-OFF crosstalk is also discussed.
... We recorded local ERG simultaneously with ganglion cell activity in order to give an insight into the retinal site of picrotoxin action. It is known that the ERG b-wave (ON response) and d-wave (OFF response) depend mainly on the activity of the ON and OFF bipolar cells in distal retina (Reviews: Perlman 1995; Frishman 2006 ). We have demonstrated that picrotoxin increases the amplitude of both the band d-waves in all cases (irrespective of its effect on ganglion cell activity) (Fig. 4), indicating that the diverse effects of the blocker on the ganglion cells are due to its action in the proximal, but not distal retina. ...
Ganglion cells are the output retinal neurons that convey visual information to the brain. There are ~20 different types of ganglion cells, each encoding a specific aspect of the visual scene as spatial and temporal contrast, orientation, direction of movement, presence of looming stimuli; etc. Ganglion cell functioning depends on the intrinsic properties of ganglion cell's membrane as well as on the excitatory and inhibitory inputs that these cells receive from other retinal neurons. GABA is one of the most abundant inhibitory neurotransmitters in the retina. How it modulates the activity of different types of ganglion cells and what is its significance in extracting the basic features from visual scene are questions with fundamental importance in visual neuroscience. The present review summarizes current data concerning the types of membrane receptors that mediate GABA action in proximal retina; the effects of GABA and its antagonists on the ganglion cell light-evoked postsynaptic potentials and spike discharges; the action of GABAergic agents on centre-surround organization of the receptive fields and feature related ganglion cell activity. Special emphasis is put on the GABA action regarding the ON-OFF and sustained-transient ganglion cell dichotomy in both nonmammalian and mammalian retina.
... These components are usually used for assessment of the retinal ON and OFF channel activity. The ERG b-and d-waves are thought to depend mainly on the activity of ON and OFF bipolar cells, respectively ( [212][213][214][215][216]; reviews: [217,218]). The role played by GABA in overall function of the distal retina can be easily investigated by application of exogenous GABA or blocking the retinal GABAergic neurotransmission and following up the changes of the ERG waves. ...
In the vertebrate retina, visual signals are segregated into parallel ON and OFF pathways, which provide information for light increments and decrements. The segregation is first evident at the level of the ON and OFF bipolar cells in distal retina. The activity
of large populations of ON and OFF bipolar cells is reflected in the b- and d-waves of the diffuse electroretinogram (ERG). The role of gamma-aminobutyric acid (GABA), acting through ionotropic GABA receptors in shaping the ON and OFF responses in distal retina, is a matter of debate. This review summarized current knowledge about the types of the GABAergic neurons and ionotropic GABA receptors in the retina as well as the effects of GABA and specific GABAA and GABAC receptor antagonists on the activity of the ON and OFF bipolar cells in both nonmammalian and mammalian retina. Special emphasis is put on the effects on b- and d-waves of the ERG as a useful tool for assessment of the overall function of distal retinal ON and OFF channels. The role of GABAergic system in establishing the ON-OFF asymmetry concerning the time course and absolute and relative sensitivity of the ERG responses under different conditions of light adaptation in amphibian retina is also discussed.
... To move our probe toward future clinical usage, physiological evidence of potential probe toxicity was assayed by electroretinogram (ERG). Full-field ERG has been used in both humans and lower animals as an indicator of retinal function and ERG changes can provide evidence of retinal injury or toxicity, particularly to the outer retina [25,26]. Under conditions of dark adaptation, the A-wave is generally associated with rod photoreceptor activity while the B-wave is associated with a combination of Muller and bipolar cell layer activity [27]. ...
Peptide probes for imaging retinal ganglion cell (RGC) apoptosis consist of a cell-penetrating peptide targeting moiety and a fluorophore-quencher pair flanking an effector caspase consensus sequence. Using ex vivo fluorescence imaging, we previously validated the capacity of these probes to identify apoptotic RGCs in cell culture and in an in vivo rat model of N-methyl- D-aspartate (NMDA)-induced neurotoxicity. Herein, using TcapQ488, a new probe designed and synthesized for compatibility with clinically-relevant imaging instruments, and real time imaging of a live rat RGC degeneration model, we fully characterized time- and dose-dependent probe activation, signal-to-noise ratios, and probe safety profiles in vivo. Adult rats received intravitreal injections of four NMDA concentrations followed by varying TcapQ488 doses. Fluorescence fundus imaging was performed sequentially in vivo using a confocal scanning laser ophthalmoscope and individual RGCs displaying activated probe were counted and analyzed. Rats also underwent electroretinography following intravitreal injection of probe. In vivo fluorescence fundus imaging revealed distinct single-cell probe activation as an indicator of RGC apoptosis induced by intravitreal NMDA injection that corresponded to the identical cells observed in retinal flat mounts of the same eye. Peak activation of probe in vivo was detected 12 hours post probe injection. Detectable fluorescent RGCs increased with increasing NMDA concentration; sensitivity of detection generally increased with increasing TcapQ488 dose until saturating at 0.387 nmol. Electroretinography following intravitreal injections of TcapQ488 showed no significant difference compared with control injections. We optimized the signal-to-noise ratio of a caspase-activatable cell penetrating peptide probe for quantitative non-invasive detection of RGC apoptosis in vivo. Full characterization of probe performance in this setting creates an important in vivo imaging standard for functional evaluation of future probe analogues and provides a basis for extending this strategy into glaucoma-specific animal models.
Purpose
The loss of Dicer in Müller glia (MG) results in severe photoreceptor degeneration, as it occurs in retinitis pigmentosa or age-related macular degeneration; however, the sequence of events leading to this severe degenerative state is unknown. The aim of this study was to conduct a chronological functional and structural characterization of the pathological events in MG-specific Dicer-conditional knockout (cKO) mice in vivo and histologically.
Methods
To delete Dicer and mature microRNAs (miRNAs) in MG, two conditional Dicer1 knockout mouse strains (Rlbp-CreER:tdTomato:Dicer-cKOMG and Glast-CreER:tdTomato:Dicer-cKOMG) were created. Optical coherence tomography (OCT), electroretinograms (ERGs), and histological analyses were conducted to investigate structural and functional changes up to 6 months after Dicer deletion.
Results
Dicer/miRNA loss in MG leads to (1) impairments of the area spanning from the external limiting membrane (ELM) to the retinal pigment epithelium (RPE), (2) cone photoreceptor dysfunction, and (3) retinal remodeling and functional loss of the inner retina at 1, 3, and 6 months after Dicer loss, respectively, in both of the knockout mouse strains. Furthermore, in the Rlbp-CreER:tdTomato:Dicer-cKOMG strain, rod photoreceptor impairment was found 4 months after Dicer depletion (4) accompanied by alteration of RPE integrity (5).
Conclusions
MG Dicer loss in the adult mouse retina impacts cone function prior to any measurable changes in rod function, suggesting a pivotal role for MG Dicer and miRNAs in supporting cone health. A partially impaired RPE, however, seems to accelerate rod degeneration and overall degenerative events.
Purpose
To explore the effect of cobalt toxicity on vision.
Methods
A total of 103 wild-type (WT) mice were injected with cobalt chloride by two routes in different concentrations: single intravenous (IV) high or low doses (total, n = 43); or daily repeated intraperitoneal (IP) high (three days) or low (28 days, 56 days) dose, and low-dose cobalt with added minocycline (56 days) (total, n = 60); 10 WT mice served as a control group. An additional group of 17 immunodeficient NOD scid gamma (NSG) mice were injected IV or IP with cobalt, and 10 NSG mice served as control. Cobalt levels were measured in blood, urine, and tears by particle-induced X-ray emission (PIXE). Macroscopic, immunohistochemical, electroretinography (ERG), and molecular studies were done.
Results
PIXE revealed cobalt elimination from the blood by two hours, with increased levels in urine but under the detection limit in tears. In the retina, ERG recordings showed decreased b-wave amplitude. Apoptosis mainly involved the inner retina, with inner retinal inflammatory reaction in both WT and less in the NSG mice. In the optic nerves, an increased microglial and astrocytic activation was noted.
Conclusions
This study demonstrated functional visual impairment with extensive inflammatory reaction secondary to cobalt toxicity in mice.
Age-related macular degeneration (AMD) is a major cause of blindness that affects people over 60. While aging is the prominent factor in AMD, studies have reported a higher prevalence of AMD in women compared to age-matched men. Higher levels of the innate immune response’s effector proteins complement factor B and factor I were also found in females compared to males in intermediate AMD. However, the mechanisms underlying these differences remain elusive. Peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) is a key regulator of mitochondrial biogenesis and metabolic pathways. Previously, we showed that Pgc-1α repression and high-fat diet induce drastic AMD-like phenotypes in mice. Our recent data revealed that Pgc-1α repression alone can also induce retinal pigment epithelium (RPE) and retinal dysfunction in mice, and its inhibition in vitro results in lipid droplet accumulation in human RPE. Whether sex is a contributing factor in these phenotypes remains to be elucidated. Using electroretinography, we demonstrate that sex could influence RPE function during aging independent of Pgc-1α in wild-type (WT) mice. We further show that Pgc-1α repression exacerbates RPE and retinal dysfunction in females compared to aged-match male mice. Gene expression analyses revealed that Pgc-1α differentially regulates genes related to antioxidant enzymes and mitochondrial dynamics in males and females. RPE flat mounts immunolabeled with TOMM20 and DRP1 indicated a sex-dependent role for Pgc-1α in regulating mitochondrial fission. Analyses of mitochondrial network morphology suggested sex-dependent effects of Pgc-1α repression on mitochondrial dynamics. Together, our study demonstrates that inhibition of Pgc-1α induces a sex-dependent decline in RPE and retinal function in mice. These observations on the sex-dependent regulation of RPE and retinal function could offer novel insights into targeted therapeutic approaches for age-related RPE and retinal degeneration.
Many inherited retinal diseases target photoreceptors, which transduce light into a neural signal that is processed by the downstream visual system. As photoreceptors degenerate, physiological and morphological changes to retinal synapses and circuitry reduce sensitivity and increase noise, degrading visual signal fidelity. Here, we pharmacologically targeted the first synapse in the retina in an effort to reduce circuit noise without sacrificing visual sensitivity. We tested a strategy to partially replace the neurotransmitter lost when photoreceptors die with an agonist of receptors that ON bipolars cells use to detect glutamate released from photoreceptors. In rd10 mice, which express a photoreceptor mutation that causes retinitis pigmentosa (RP), we found that a low dose of the mGluR6 agonist L-2-amino-4-phosphonobutyric acid (L-AP4) reduced pathological noise induced by photoreceptor degeneration. After making in vivo electroretinogram recordings in rd10 mice to characterize the developmental time course of visual signal degeneration, we examined effects of L-AP4 on sensitivity and circuit noise by recording in vitro light-evoked responses from individual retinal ganglion cells (RGCs). L-AP4 decreased circuit noise evident in RGC recordings without significantly reducing response amplitudes, an effect that persisted over the entire time course of rod photoreceptor degeneration. Subsequent in vitro recordings from rod bipolar cells (RBCs) showed that RBCs are more depolarized in rd10 retinas, likely contributing to downstream circuit noise and reduced synaptic gain, both of which appear to be ameliorated by hyperpolarizing RBCs with L-AP4. These beneficial effects may reduce pathological circuit remodeling and preserve the efficacy of therapies designed to restore vision.
Significance Statement
Retinitis Pigmentosa (RP) is an inherited degenerative disease that affects more than two million people worldwide. RP patients first lose peripheral and low-light vision due to the progressive death of their highly sensitive rod photoreceptors. Photoreceptor degeneration induces pathological noise within the retinal circuit, leading to dramatic structural changes that may hamper therapies to restore visual sensitivity. We discovered a pharmacological treatment that reduces pathological activity in a mouse model of RP without diminishing signaling in surviving circuitry. Partially replacing the neurotransmitter lost when photoreceptors die reduced noise in the retinal circuit without eliminating light sensitivity. This approach could limit the impact of the disease on retinal neurons and preserve the efficacy of subsequent restorative therapies.
Purpose
Electroretinography (ERG) is used to assess retinal function in ophthalmology clinics and animal models of ocular disease; however, analyzing ERG waveforms can be a time-intensive process with interobserver variability. We developed ERGAssist, an automated approach, to perform non-subjective and repeatable feature identification (“marking”) of the ERG waveform.
Methods
The automated approach denoised the recorded waveforms and then located the b-wave after applying a lowpass filter. If an a-wave was present, the lowpass filter wave was also used to help locate the a-wave, which was considered the initial large negative response after the flash stimuli. Oscillatory potentials (OPs) were found using a bandpass filter on the denoised waveform. We used two cohorts. One was a Coherence cohort that consisted of ERGs with eight dark-adapted and three light-adapted stimuli in Brown Norway rats (−6 to 1.5 log cd·s/m²). The Verification cohort consisted of control and diabetic (DM) Long Evans rats. We examined retinal function using a five-step dark-adapted protocol (−3 to 1.9 log cd·s/m²).
Results
ERGAssist showed a strong correlation with manual markings of ERG features in our Coherence dataset, including the amplitudes (a-wave: r² = 0.99; b-wave: r² = 0.99; OP: r² = 0.92) and implicit times (a-wave: r² = 0.96; b-wave: r² = 0.90; OP: r² = 0.96). In the Verification cohort, both approaches detected differences between control and DM animals and found longer OP implicit times (P < 0.0001) in DM animals.
Conclusions
These results provide verification of ERGAssist to identify features of the full-field ERG.
Translational Relevance
This ERG analysis approach can increase the rigor of basic science studies designed to investigate retinal function using full-field ERG. To aid the community, we have developed an open-source graphical user interface (GUI) implementing the methods presented.
We present a detailed physiological model of the (human) retina that includes the biochemistry and electrophysiology of phototransduction, neuronal electrical coupling, and the spherical geometry of the eye. The model is a parabolic-elliptic system of partial differential equations based on the mathematical framework of the bi-domain equations, which we have generalized to account for multiple cell-types. We discretize in space with non-uniform finite differences and step through time with a custom adaptive time-stepper that employs a backward differentiation formula and an inexact Newton method. A refinement study confirms the accuracy and efficiency of our numerical method. Numerical simulations using the model compare favorably with experimental findings, such as desensitization to light stimuli and calcium buffering in photoreceptors. Other numerical simulations suggest an interplay between photoreceptor gap junctions and inner segment, but not outer segment, calcium concentration. Applications of this model and simulation include analysis of retinal calcium imaging experiments, the design of electroretinograms, the design of visual prosthetics, and studies of ephaptic coupling within the retina.
Introduction: The eye is an excellent target for gene therapy because of its anatomical features. Gene therapy to treat ocular disorders relies on efficient gene delivery and transgene expression in the target cells. The aim of this study was to compare the biodistribution and safety of two different AAV serotypes after intravitreal (IVT) and subretinal injections.
Methods: AAV2 (1 × 10 ¹² vg/mL) and AAV9 (5 × 10 ¹² vg/mL) vectors expressing an enhanced green fluorescent protein (EGFP) and an AAV9-empty (6 × 10 ¹¹ vg/mL) vector were injected intravitreally or subretinally into both eyes of adult C57Bl/OlaHsd mice. The biodistribution of the viral vectors in the eye and off-target tissues was studied using qPCR. GFP expression was studied from cryosections, and GFP transduction efficacy was verified using immunohistostaining for GFP. In addition, electroretinography (ERG) was used to assess the effect of vectors on retinal function.
Results: In addition to the eyes, viral vector copies were found in distant off-target tissues such as the liver, especially after AAV9-EGFP IVT and subretinal injections. AAV9-EGFP injections showed more GFP expression throughout the retina compared to AAV2-EGFP. AAV2-EGFP IVT showed transgene expression mainly in the ganglion cell layer, whereas subretinal injection showed GFP expression in the retinal pigment epithelium. In addition, GFP was expressed at a moderate level in the liver after both injection routes of AAV9 and in parts of the brain after all injection groups except AAV9-empty. Lowered a- and b-amplitude values were seen in ERG in both scotopic and photopic experiments after AAV9-EGFP subretinal injection compared to all other groups.
Discussion: This study shows that intraocular injection of AAV2 and AAV9 transduces retinal cells. Although the more efficient transduction of the retina, negative effect on the retinal function, and off-target transgene expression of AAV9 makes AAV2 a more suitable gene delivery vector to treat ocular disorders.
Purpose:
The purpose of this study was to evaluate the electrical responses in the retina and cortex of migraine patients with electrophysiological tests and compare with healthy controls.
Materials and methods:
This prospective study included 18 migraine patients with visual aura and 28 healthy controls. Pattern-reversal visual evoked potentials (VEP) and flash electroretinography (fERG) of migraine patients during the headache-free period were compared with healthy controls.
Results:
There were statistically significant differences in VEP results: P100 and N75 amplitudes increased significantly (P = 0.025 and P = 0.007 respectively) and P100 latency decreased significantly in migraine patients (P = 0.022). Furthermore, fERG scotopic combined cone and rod amplitude increased significantly in migraine patients (P = 0.01).
Conclusion:
Migraine brain displays abnormal visual evoked responses in between migraine attacks. In migraine eye, scotopic cone and rod response increased. The results of this study support the hyperexcitability of the retina and cortex in patients with migraine.
Brain visual dynamics encode rich functional and biological patterns of the neural system, and if decoded, are of great promise for many applications such as intention understanding, cognitive load quantization and neural disorder measurement. We here focus on the understanding of the brain visual dynamics for the Amyotrophic lateral sclerosis (ALS) population, and propose a novel system that allows these so-called 'lock-in' patients to 'speak' with their brain visual movements. More specifically, we propose an intelligent system to decode the eye bio-potential signal, Electrooculogram (EOG), thereby understanding the patients' intention. We first propose to leverage a deep learning framework for automatic feature learning and classification of the brain visual dynamics, aiming to translate the EOG to meaningful words. We afterwards design and develop an edge computing platform on the smart phone, which can execute the deep learning algorithm, visualize the brain visual dynamics, and demonstrate the edge inference results, all in real-time. Evaluated on 4,500 trials of brain visual movements performed by multiple users, our novel system has demonstrated a high eye-word recognition rate up to 90.47%. The system is demonstrated to be intelligent, effective and convenient for decoding brain visual dynamics for ALS patients. This research thus is expected to greatly advance the decoding and understanding of brain visual dynamics, by leveraging machine learning and edge computing innovations.
Achromatopsia (ACHM) is an inherited autosomal recessive disease lacking cone photoreceptors functions. In this study, we characterize the time-frequency representation of the full-field electroretinogram (ffERG) component oscillatory potentials (OPs), to investigate the connections between photoreceptors and the inner retinal network using ACHM as a model. Time-frequency characterization of OPs was extracted from 52 controls and 41 achromat individuals. The stimulation via ffERG was delivered under dark-adaptation (DA, 3.0 and 10.0 cd·s·m⁻²) to assess mixed rod-cone responses. The ffERG signal was subsequently analyzed using a continuous complex Morlet transform. Time-frequency maps of both DA conditions show the characterization of OPs, disclosing in both groups two distinct time-frequency windows (~70–100 Hz and >100 Hz) within 50 ms. Our main result indicates a significant cluster (p < 0.05) in both conditions of reduced relative power (dB) in ACHM people compared to controls, mainly at the time-frequency window >100 Hz. These results suggest that the strongly reduced but not absent activity of OPs above 100 Hz is mostly driven by cones and only in small part by rods. Thus, the lack of cone modulation of OPs gives important insights into interactions between photoreceptors and the inner retinal network and can be used as a biomarker for monitoring cone connection to the inner retina.
Blast-mediated traumatic brain injuries (bTBI) cause long-lasting physical, cognitive, and psychological disorders, including persistent visual impairment. No known therapies are currently utilized in humans to lessen the lingering and often serious symptoms. With TBI mortality decreasing due to advancements in medical and protective technologies, there is growing interest in understanding the pathology of visual dysfunction after bTBI. However, this is complicated by numerous variables, e.g., injury location, severity, and head and body shielding. This review summarizes the visual outcomes observed by various, current experimental rodent models of bTBI, and identifies data showing that bTBI activates inflammatory and apoptotic signaling leading to visual dysfunction. Pharmacologic treatments blocking inflammation and cell death pathways reported to alleviate visual deficits in post-bTBI animal models are discussed. Notably, techniques for assessing bTBI outcomes across exposure paradigms differed widely, so we urge future studies to compare multiple models of blast injury, to allow data to be directly compared.
Juvenile neuronal ceroid lipofuscinosis (JNCL, aka. juvenile Batten disease or CLN3 disease) is a lysosomal storage disease characterized by progressive blindness, seizures, cognitive and motor failures, and premature death. JNCL is caused by mutations in the Ceroid Lipofuscinosis, Neuronal 3 (CLN3) gene, whose function is unclear. Although traditionally considered a neurodegenerative disease, CLN3 disease displays eye-specific effects: JNCL often first presents as vision loss; and vision loss has also been reported in non-syndromic CLN3 disease. Here we described the roles of CLN3 protein in maintaining healthy retinal pigment epithelium (RPE) and normal vision. Using electroretinogram, fundoscopy and microscopy, we showed impaired visual function, retinal autofluorescent lesions, and RPE disintegration and metaplasia/hyperplasia in a Cln3 ~ 1 kb-deletion mouse model [1] on C57BL/6J backgroun. Utilizing a combination of biochemical analyses, RNA-Seq, Seahorse XF bioenergetic analysis, and Stable Isotope Resolved Metabolomics (SIRM), we further demonstrated that loss of CLN3 increased autophagic flux, suppressed mTORC1 and Akt activities, enhanced AMPK activity, and up-regulated gene expression of the autophagy-lysosomal system in RPE-1 cells, suggesting autophagy induction. This CLN3 deficiency induced autophagy induction coincided with decreased mitochondrial oxygen consumption, glycolysis, the tricarboxylic acid (TCA) cycle, and ATP production. We also report for the first time that loss of CLN3 led to glycogen accumulation despite of impaired glycogen synthesis. Our comprehensive analyses shed light on how loss of CLN3 affect autophagy and metabolism. This work suggests possible links among metabolic impairment, autophagy induction and lysosomal storage, as well as between RPE atrophy/degeneration and vision loss in JNCL.
Investigators have increasingly turned to studying the retina as a window into brain structure and function. In neuropsychiatric diseases, retinal cell structure as assessed by optical coherence tomography (OCT) and retinal cell function as assessed by various forms of electroretinography (ERG) indicate the presence of notable changes. In addition, many studies indicate significant correlations between retinal changes and clinical features such as cognitive decline, overall illness severity, and progression of illness. Here, we review retinal findings in psychiatric (schizophrenia, autism, mood disorders, attention deficit hyperactivity disorder, anorexia nervosa), and neurologic (multiple sclerosis, Parkinson's disease, Alzheimer's disease and mild cognitive impairment, Huntington's disease, traumatic brain injury) conditions, in terms of their potential as biomarkers of disease onset, progression, severity, and outcomes. Consistency and variability in findings across studies are highlighted, and implications for future research are discussed. Potential confounds and methodological issues central to studies of retinal structure and function in neuropsychiatry are also considered. The review concludes with discussions of: a) recent advances in retinal imaging and their potential applications for studying brain disorders; and b) the potential for applications of artificial intelligence to increasing the predictive validity of retinal data.
Studies in experimental ischemia models by permanent bilateral common carotid artery occlusion (BCCAO) have reported reduced retinal electrophysiological function, coupled with inner retinal degeneration and gliosis. In the current study, we tested the hypothesis that long-term (up to 14 days) BCCAO impairs oxygen delivery (DO2), which affects oxygen metabolism (MO2) and extraction fraction (OEF), electrophysiological function, morphology, and biochemical pathways. Twenty-one rats underwent BCCAO (N = 12) or sham surgery (N = 9) and were evaluated in separate groups after 3, 7, or 14 days. Electroretinography (ERG), optical coherence tomography, blood flow and vascular oxygen tension imaging, and morphological and biochemical evaluations were performed in both eyes. Reduced ERG b-wave amplitudes and delayed implicit times were reported at 3, 7, and 14 days following BCCAO. Total retinal blood flow, MO2, and DO2 were reduced in all BCCAO groups. OEF was increased in both 3- and 7-day groups, while no significant difference was observed in OEF at 14 days compared to the sham group. At 14 days following BCCAO, total and inner retinal layer thickness was reduced, while the outer nuclear layer thickness and gliosis were increased. There was an increase in nuclei containing fragmented DNA at 3 days following BCCAO. The compensatory elevation in OEF following BCCAO did not meet the tissue demand, resulting in the subsequent reduction of MO2. The associations between retinal MO2, DO2, and retinal function were shown to be significant in the sequelae of persistent ischemia. In sum, measurements of DO2, MO2, and OEF may become useful for characterizing salvageable tissue in vision-threatening pathologies.
Background:
Differences between people with schizophrenia and psychiatrically healthy controls have been consistently demonstrated on measures of retinal function such as electroretinography (ERG), and measures of retinal structure such as optical coherence tomography (OCT). Since our 2015 review of this literature, multiple new studies have been published using these techniques. At the same time, the accumulation of data has highlighted the "fault lines" in these fields, suggesting methodological considerations that need greater attention in future studies.
Methods:
We reviewed studies of ERG and OCT in schizophrenia, as well as data from studies whose findings are relevant to interpreting these papers, such as those on effects of the following on ERG and OCT data: comorbid medical conditions that are over-represented in schizophrenia, smoking, antipsychotic medication, substance abuse, sex and gender, obesity, attention, motivation, and influences of brain activity on retinal function.
Results:
Recent ERG and OCT studies continue to support the hypothesis of retinal structural and functional abnormalities in schizophrenia, and suggest that these are relevant to understanding broader aspects of pathophysiology, neurodevelopment, and neurodegeneration in this disorder. However, there are differences in findings which suggest that the effects of multiple variables on ERG and OCT data need further clarification.
Conclusions:
The retina, as the only component of the CNS that can be imaged directly in live humans, has potential to clarify important aspects of schizophrenia. With greater attention to specific methodological issues, the true potential of ERG and OCT as biomarkers for important clinical phenomena in schizophrenia should become apparent.
LEDs, which are rapidly replacing the traditional light sources for general illumination, present unique challenges with respect to flicker, due to their different electrical behavior in the circuits in which they are embedded. Since the present-day electrical systems are AC, there is potential for LED output to be time varying, because driver electronics are almost always employed and because driver and mains are not ideal. For flicker, time varying light is not an issue if it is not perceived by humans. The human perception aspect introduces enormous complexities, but large body of psychophysics research over many decades has shed much light on this. There now exist well known and recognized standards which provide for the measure and quantification of flicker. This paper offers a general technical perspective on flicker with discussions of each of these topics.
Purpose
To determine the electroretinographical and psychophysical parameters that can help to verify patients’ complaints of reduced night vision.
Methods
We tested 275 consecutive patients with normal appearing fundi, complaining of visual difficulties at night, using flash electroretinography (ERG) and dark adaptation (DA) test. Two ERG parameters were used to assess a scotopic retinal function: the amplitude of the response to dim blue flash (the rod response) and the b-wave ratio (measured/expected). Dark adaptation was measured with green- and red-light stimuli after exposure to a bright, bleaching light. The psychophysical parameter of night vision was defined as the threshold for detection of the blue-green stimulus that was measured after 40–45 min in complete darkness.
Results
Fifty-five patients were excluded from the analysis because of a discrepancy between the two ERG parameters in assessment of scotopic retinal function. The remaining 220 patients were divided into 4 groups: (1) normal ERG and normal DA, (2) subnormal ERG and subnormal DA, (3) normal ERG and subnormal DA and (4) subnormal ERG and normal DA. The ERG and DA tests supported the complaint of visual difficulties at night in 67 patients (group 2), while 34 patients were characterized as having normal scotopic visual function (group 1). The other 119 patients (groups 3 and 4) presented a diagnostic dilemma because one test (ERG or dark adaptation) showed normal scotopic function, while the other indicated subnormal scotopic function.
Conclusion
Our findings indicate that ERG is an essential, but not sufficient test for verifying patient’s complaint on visual difficulties in the dark. We suggest using both electroretinography and psychophysical dark adaptation to test patients complaining of reduced night vision.
Purpose:
To define the functional roles of Grk1 and Grk7 in zebrafish cones in vivo.
Methods:
Genome editing was used to generate grk7a and grk1b knockout zebrafish. Electroretinogram (ERG) analyses of the isolated cone mass receptor potential and the b-wave were performed in dark-adapted zebrafish using a paired flash paradigm to determine recovery of cone photoreceptors and the inner retina after an initial flash. In addition, psychophysical visual response was measured using the optokinetic response (OKR).
Results:
ERG analysis demonstrated that deletion of either Grk1b or Grk7a in zebrafish larvae resulted in modestly lower rates of recovery of the isolated cone mass receptor potential from an initial flash compared to wildtype larvae. On the other hand, grk1b-/- and grk7a-/- larvae exhibited a b-wave recovery that was similar to wildtype larvae. We evaluated the OKR and found that deletion of either Grk1b or Grk7a leads to a small decrease in temporal contrast sensitivity and alterations in visual acuity.
Conclusions:
For the first time, we demonstrate that Grk1b and Grk7a both contribute to visual function in larval zebrafish cones. Since the difference between wildtype and each knockout fish is modest, it appears that either GRK is sufficient for adequate cone visual function.
Photoreceptor ribbon synapses tonically release glutamate. To ensure efficient signal transmission and prevent glutamate toxicity, a highly efficient glutamate removal system provided by members of the SLC1 gene family is required. By using a combination of biophysical and in vivo studies, we elucidate the role of excitatory amino acid transporter 2 (EAAT2) proteins in synaptic glutamate homeostasis at the zebrafish photoreceptor synapse. The main glutamate sink is provided by the glial EAAT2a, reflected by reduced electroretinographic responses in EAAT2a-depleted larvae. EAAT2b is located on the tips of cone pedicles and contributes little to glutamate reuptake. However, this transporter displays both a large chloride conductance and leak current, being important in stabilizing the cone resting potential. This work demonstrates not only how proteins originating from the same gene family can complement each other’s expression profiles and biophysical properties, but also how presynaptic and glial transporters are coordinated to ensure efficient synaptic transmission at glutamatergic synapses of the central nervous system.
The electroretinogram (ERG) is a sensitive and noninvasive method for testing retinal function. In this protocol, we describe a method for performing ERGs in mice. Contact lenses on the mouse cornea measure the electrical response to a light stimulus of photoreceptors and downstream retinal cells, and the collected data are analyzed to evaluate retinal function.
Electroretinography is an important part of specialised ophthalmologic diagnostics in the horse. For the integration of ERG in the ophthalmologic examination, a standard equine ERG examination protocol was developed. The RETIport® ERG-system of Acri.Tec AG was used to define normal findings using 6 horses with healthy eyes which underwent 3 repeated examinations for a total of 18 examinations. The normal findings were taken after light- and dark adaptation and repeatability was tested. Every examination was taken in standing horses, sedated with detomidine. 4 parameters of the electroretinogram were taken: a-wave amplitude, a-latency time, b-wave amplitude, b-latency time. No significant differences in all 4 parameters were encountered amongst the 6 horses. Repeatability was excellent and variability low. The unity of the active electrode and the LED-Stimulator is the special feature of the RETIport device. Because of the inseparable unity of the Koojiman-Electrode and the LED-Stimulator, motion artefacts rarely occurred. The method proved to be easy, safe and reliable. Therefore, electroretinography in the standing sedated horse can well be integrated in the ophthalmologic examination using the RETIport system.
We report on detection results obtained in 20 experiments in which the presence of an external auditory stimulus had to be detected from observing electrophysiological multi-unit activity in the brain stem of rats. The performance of the optimal Gaussian-signal-in-Gaussian-noise (model-based) detector is compared to that of the energy detector which is widely used in electrophysiology as well as in many other disciplines with similar signal characteristics. It is shown that the optimal model based detector is indeed superior, but the performance gap in favor of the optimal detector is substantial mainly in very low probabilities of false alarm errors. The performance of the energy detector is close to optimum in moderate and high probabilities of false alarm error. Furthermore, the energy detector is shown to be more resilient to isolated and short, yet intense disturbances. We discuss a conjecture inspired by the model underlying the optimal detector and the empirical results, that the neural tissue itself executes a modified energy detection scheme, and we review experimental results from the literature that allegedly support this conjecture.
The electroretinogram (ERG) is a noninvasive electrophysiological method for determining retinal function. Through the placement of an electrode on the surface of the cornea, electrical activity generated in response to light can be measured and used to assess the activity of retinal cells in vivo. This manuscript describes the use of the ERG to measure visual function in zebrafish. Zebrafish have long been utilized as a model for vertebrate development due to the ease of gene suppression by morpholino oligonucleotides and pharmacological manipulation. At 5-10 dpf, only cones are functional in the larval retina. Therefore, the zebrafish, unlike other animals, is a powerful model system for the study of cone visual function in vivo. This protocol uses standard anesthesia, micromanipulation and stereomicroscopy protocols that are common in laboratories that perform zebrafish research. The outlined methods make use of standard electrophysiology equipment and a low light camera to guide the placement of the recording microelectrode onto the larval cornea. Finally, we demonstrate how a commercially available ERG stimulator/recorder originally designed for use with mice can easily be adapted for use with zebrafish. ERG of larval zebrafish provides an excellent method of assaying cone visual function in animals that have been modified by morpholino oligonucleotide injection as well as newer genome engineering techniques such as Zinc Finger Nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9, all of which have greatly increased the efficiency and efficacy of gene targeting in zebrafish. In addition, we take advantage of the ability of pharmacological agents to penetrate zebrafish larvae to evaluate the molecular components that contribute to the photoresponse. This protocol outlines a setup that can be modified and used by researchers with various experimental goals.
Dopamine is the most abundant catecholamine in the vertebrate retina. Despite the description of retinal dopaminergic cells three decades ago, many aspects of their function in the retina remain unclear. There is no consensus among the authors about the stimulus conditions for dopamine release (darkness, steady or flickering light) as well as about its action upon the various types of retinal cells. Many contradictory results exist concerning the dopamine effect on the gross electrical activity of the retina [reflected in electroretinogram (ERG)] and the receptors involved in its action. This review summarized current knowledge about the types of the dopaminergic neurons and receptors in the retina as well as the effects of dopamine receptor agonists and antagonists on the light responses of photoreceptors, horizontal and bipolar cells in both nonmammalian and mammalian retina. Special focus of interest concerns their effects upon the diffuse ERG as a useful tool for assessment of the overall function of the distal retina. An attempt is made to reveal some differences between the dopamine actions upon the activity of the ON versus OFF channel in the distal retina. The author has included her own results demonstrating such differences.
Mutations in the human CACNA1F gene cause incomplete congenital stationary night blindness type 2 (CSNB2), a non-progressive, clinically heterogeneous retinal disorder. However, the molecular mechanisms underlying CSNB2 have not been fully explored. Here, we describe the positional cloning of a blind zebrafish mutant, wait until dark (wud), which encodes a zebrafish homolog of human CACNA1F. We identified two zebrafish cacna1f paralogs and showed that the cacna1fa transcript (the gene mutated in wud) is expressed exclusively in the photoreceptor layer. We demonstrated that Cacna1fa localizes at the photoreceptor synapse and is absent from wud mutants. Electroretinograms revealed abnormal cone photoreceptor responses from wud mutants, indicating a defect in synaptic transmission. Although there are no obvious morphological differences, we found that wud mutants lacked synaptic ribbons and that wud is essential for the development of synaptic ribbons. We found that Ribeye, the most prominent synaptic ribbon protein, was less abundant and mislocalized in adult wud mutants. In addition to cloning wud, we identified synaptojanin 1 (synj1) as the defective gene in slacker (slak), a blind mutant with floating synaptic ribbons. We determined that Cacna1fa was expressed in slak photoreceptors and that Synj1 was initially expressed wud photoreceptors, but was absent by 5 days postfertilization. Collectively, our data demonstrate that Cacna1fa is essential for cone photoreceptor function and synaptic ribbon formation and reveal a previously unknown yet critical role of L-VDCCs in the expression and/or distribution of synaptic ribbon proteins, providing a new model to study the clinical variability in human CSNB2 patients.
Retinal ganglion cells (RGCs) are specialized projection neurons that relay an immense amount of visual information from the retina to the brain. RGC signal inputs are collected by dendrites and output is distributed from the cell body via very thin (0.5-1 μm) and long (∼50 mm) axons. The RGC cell body is larger than other retinal neurons, but is still only a very small fraction (one ten thousandths) of the length and total surface area of the axon. The total distance traversed by RGCs extends from the retina, starting from synapses with bipolar and amacrine cells, to the brain, to synapses with neurons in the lateral geniculate nucleus.
To summarize current technique, indications, and pitfalls of electrophysiologic testing used in ophthalmology.
Visual evoked potentials (VEPs) may be useful as an objective measurement of refractive error in complicated patients. VEP P100 latency was found superior to color vision and visual field in early stages of hydroxychloroquine maculopathy. VEP results can be predictive of visual recovery in traumatic optic neuropathy. Multifocal electroretinogram (ERG) or VEP can provide an objective assessment of visual field defects not yet present on automated perimetry in patients with glaucomatous and nonglaucomatous optic neuropathies. In patients with intraocular lymphoma, reduced amplitudes of all ERG components can be recorded, with the b-wave amplitude being most significantly affected.
Various visual electrophysiologic tests are useful to the ophthalmologist, each with different indications. The flash ERG is most useful in diffuse retinal disorders, whereas the multifocal ERG is superior in localized retinal disease. VEPs can be valuable in diagnosing optic neuropathies, nonorganic visual loss, and assessing visual function in infants or children.
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