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Purpose: To determine the utility of ophthalmology evaluation, dark-adapted threshold, and full-field electroretinogram for early detection of Usher syndrome in young patients with bilateral sensorineural hearing loss. Methods: We identified 39 patients with secure genetic diagnoses of Usher Syndrome. Visual acuity, spherical equivalent, fundus appearance, dark-adapted threshold, and full-field electroretinogram results were summarized and compared to those in a group of healthy controls with normal hearing. In those Usher patients with repeated measures, regression analysis was done to evaluate for change in visual acuity and dark-adapted threshold with age. Spherical equivalent and full-field electroretinogram responses from dark- and light-adapted eyes were evaluated as a function of age. Results: The majority of initial visual acuity and spherical equivalent results were within normal limits for age. Visual acuity and dark-adapted threshold worsened significantly with age in Usher type 1 but not in Usher type 2. At initial test, full-field electroretinogram responses from dark- and light-adapted eyes were abnormal in 53% of patients. Remarkably, nearly half of our patients (17% of Usher type 1 and 30% of Usher type 2) would have been missed by tests of retinal function alone if evaluated before age 10. Conclusions: Although there is an association of abnormal dark-adapted threshold and full-field electroretinogram at young ages in Usher patients, it appears that a small but important proportion of patients would not be detected by tests of retinal function alone. Thus, genetic testing is needed to secure a diagnosis of Usher syndrome.

To investigate the effect of mild cataract and its surgery on the ISCEV standard electroretinogram (ERG) by comparing pre- and postoperative ERGs elicited from fully dilated eyes. Twenty-two patients participated. Each eye had cataract of grade 2 according to Emery-Little classification. None had complications during and after phacoemulsification and intraocular lens implantation. According to the ISCEV standard, pre- and 1-week postoperative full-field ERGs, dark adapted (DA) 0.01, DA 3, DA 3 oscillatory potentials (OPs), DA 10, and light adapted (LA) 3, and LA 3 flicker, were elicited from fully dilated eyes using skin electrodes. Photopic negative response (PhNR) 1 amplitude was measured from the baseline to the trough before the i-wave in the LA 3 ERG waveform. Statistical analysis was performed using SigmaPlot version 11.0 (Systat Software, Inc., San Jose, CA, USA). Pre- and postoperative data were compared using a paired t-test. Non-normally distributed data were evaluated using the Wilcoxon signed rank test. The pre- and 1-week postoperative amplitudes in each component were as follows: DA 0.01: 48.4 ± 19.5 μV and 57.1 ± 20.0 μV (p = 0.002), a-wave of DA3: 51.5 ± 14.8 μV and 58.3 ± 17.3 μV (p = 0.003), b-wave of DA3: 88.3 ± 27.5 μV and 101.5 ± 29.7 μV (p = 0.003), DA 3 ΣOPs (sum of DA 3 OP1, OP2, and OP3 amplitude): 30.7 ± 16.3 μV and 37.1 ± 21.9 μV (p

Photosystem I (PSI) is a major player in the light reactions of photosynthesis. In higher plants, it consists of a core complex and four external antennae, Lhca1–4 forming the PSI–light-harvesting complex I (LHCI) supercomplex. The protein and pigment composition as well as the spectroscopic properties of this complex are considered to be identical in different higher plant species. In addition to the four Lhca, a pool of mobile LHCII increases the antenna size of PSI under most light conditions. In this work, we have first investigated purified PSI complexes and then PSI in vivo upon long-term dark-adaptation of four well-studied plant species: Arabidopsis thaliana, Zea mays, Nicotiana tabacum and Hordeum vulgare. By performing time-resolved fluorescence measurements, we show that LHCII is associated with PSI also in a dark-adapted state in all the plant species investigated. The number of LHCII subunits per PSI is plant-dependent, varying between one and three. Furthermore, we show that the spectroscopic properties of PSI–LHCI supercomplexes differ in different plants.

To study whether rod- and cone-driven electroretinogram (ERG) responses are altered in myopia patients. Dark- and light-adapted ERGs were recorded from 57 myopic eyes of 32 patients aged 22–30 and 19 emmetropic eyes of 10 age-matched normal subjects. The myopic eyes were divided into 3 groups according to spherical equivalent (SE) of manifest refraction: 18 low myopia eyes (≤ − 3.00 diopter (D), 23 moderate myopia eyes (− 3.25 to − 6.00 D), and 16 high myopia eyes (> − 6.25 D). The amplitudes of the dark- and light-adapted ERG a- and b-waves, as well as the frequency spectra of the cone-driven and rod-driven oscillatory potentials (OPs), were analyzed by fast Fourier transform. The peak frequency, implicit time, and total power of the OPs were determined. The axial length was measured with an IOL Master. The ERG parameters including those of the cone- and rod-driven OPs were compared among three groups. The amplitudes of the a-wave and b-wave of the dark-adapted ERGs were increased with refractive power (P

Forensic practitioners are recommended to dark adapt their eyes prior to conducting evidential searches in the dark. The dark adaptation process remains poorly standardised across the discipline, with little quantified regarding the benefits of such preparative steps. Herein, we report the findings of a study that recruited 50 participants to assess the effectiveness of the Crime-lite Eye™, a darkness adaptation device developed to assist forensic practitioners both in the laboratory and in field. Participants were tasked with searching for the fluorescent signatures left by reaction of 1,8-diazafluoren-9-one (DFO) with amino acids, in a manner akin to the fluorogenic fingerprint treatment of porous evidence. Using an Epson Stylus Photo R265 inkjet printer, ink cartridges were filled with alanine solutions of various concentrations, allowing different motifs to be printed onto copy paper and subsequently developed using DFO. Participants searched for this ‘evidence’ both with and without dark adapted vision. On average, participants were able to locate and correctly recognise 16% more evidence once dark adapted using the Crime-lite Eye™. The increase in evidence located by participants once dark adapted suggests that crime scene officers should be dark adapting in order to visualise as much as possible. The time taken to dark adapt, 10 min on average during this study, is not excessively long, and should not significantly slow the investigation.

The magnetic compass of birds seems to be based on light-dependent radical-pair processes in the eyes. Cryptochromes are currently the only candidate proteins known in vertebrates that may serve as the primary radical-pair-based magnetoreceptor molecules. Previous immunohistochemical studies have suggested that cryptochrome 1a (Cry1a) is localised in the photoreceptor outer segments of the ultraviolet/violet (UV/V) cones, and it has been claimed that differences in Cry1a antibody staining intensities show that Cry1a is activated by light and that this should make Cry1a the most likely magnetoreceptive candidate molecule. Here, we present an independent study of Cry1a distribution within retinae of several bird species, ranging from non-migratory domestic chicken and rock pigeon to night-migratory passerines, using both the previously used antibody and two newly generated antibodies, one against the same epitope as the originally used antibody and one against a different epitope of Cry1a. We confirm the UV/V cone outer segment localisation of Cry1a in all the tested bird species. In some stainings, we found Cry1a immunoreactivity as a distinct punctate pattern throughout the whole length of the UV/V cone outer segments. These dots with a diameter of around 170 nm might suggest that many Cry1a molecules accumulate in distinct spots in the UV/V cone outer segments. However, we did not see any notable difference in Cry1a immunoreactivity between light- and dark-adapted retinae. We find no evidence whatsoever that a C-terminal antibody against Cry1a labels only a light-activated form of the Cry1a protein.

Purpose Functional studies of rods in age-related macular degeneration using the Medmont Dark-Adapted Chromatic Perimeter (DACP) have demonstrated impairments in scotopic sensitivities and dark adaptation (DA). We investigated the intersession repeatability of scotopic sensitivity and DA parameters including the rod intercept time recorded from the Medmont DACP. Methods Scotopic thresholds (14 test points) and DA using a 30% photobleach (eight test points) were measured on two separate days from participants 50 years of age or older with a range of age-related macular degeneration severity at loci superior and inferior to the fovea. Repeatability coefficients were calculated for prebleach scotopic sensitivity, and for DA parameters including rod intercept time. Results Twelve participants (mean age, 79.7 ± 8.1 years) repeated Medmont DACP testing within 50 days. Repeatability coefficients for prebleach scotopic sensitivity to long wavelength (red, 625 nm) and short wavelength (cyan, 505 nm) were 5.9 dB and 7.2 dB, respectively. The DA curve-derived repeatability coefficients for cone threshold was 3.9 dB, final threshold 5.3 dB, with an R value of 0.075 decades/min, rod intercept time 7.6 minutes, and RITslope 0.54 min/degree. Conclusions This study establishes repeatability coefficients for scotopic thresholds and multiple DA parameters obtained with the Medmont DACP in patients with age-related macular degeneration. These repeatability coefficients will serve as the basis for determining clinically meaningful change in rod function in future clinical trials. Translational relevance Measures of repeatability parameters of scotopic thresholds and DA are essential to the accurate interpretation of results in future studies and trials using these measures.

The full-field electroretinogram (ff-ERG) is a widely used clinical tool to evaluate generalized retinal function by recording electrical potentials generated by the cells in the retina in response to flash stimuli and requires mydriasis. The purpose of this study was to determine the intra-visit reliability and diagnostic capability of a handheld, mydriasis-free ERG, RETeval (LKC Technologies, Gaithersburg, MD, USA), in comparison with the standard clinical ff-ERG by measuring responses recommended by the International Society for Clinical Electrophysiology of Vision (ISCEV).This prospective, cross-sectional study included 35 patients recruited at the Hospital for Sick Children (median age = 17, range 11 months-69 years) who had undergone a clinical ff-ERG according to ISCEV standards. For RETeval (n = 35), pupils were undilated in most (n = 29) and sensor strip electrodes were placed under the inferior orbital rim. Stimulus settings on RETeval were equivalent to those used in the clinical ERG. Fifty-seven control participants (median age = 22, range 8-65 years) underwent undilated RETeval ERG to establish standard values for comparison. Patient waveform components with amplitudes 5th percentile, or implicit times 95th percentile of normal relative to control data were classified as abnormal for the RETeval system.The RETeval system demonstrated a high degree of within-visit reliability for amplitudes (ICC = 0.82) and moderate reliability for implicit times (ICC = 0.53). Cohen's Kappa analysis revealed a substantial level of agreement between the diagnostic capability of RETeval in comparison with clinical ff-ERG (k = 0.82), with a sensitivity and specificity of 1.00 and 0.82, respectively. Pearson's correlations for clinical ERG versus RETeval demonstrated a positive correlation for amplitudes across the rod (r = 0.65) and cone (r = 0.74) ERG waveforms. Bland-Altman plots showed no bias between the mean differences across all amplitude and implicit time parameters of the two systems.The present study demonstrated that RETeval is a reliable tool with reasonable accuracy in comparison with the clinical ERG. The portable nature of RETeval system enables its incorporation at resource-limited centers where the ff-ERG is not readily available. The avoidance of sedation and pupillary dilation are added advantages of RETeval ERG.

Purpose: The x-wave of the dark-adapted (DA) ERG to a red flash reflects DA cone function. This exploratory study of healthy adults aimed to investigate changes in the DA red ERG with flash strength and during dark adaptation to optimise visualisation and therefore quantification of the x-wave.\ud \ud Methods: The effect of altering red flash strength was investigated in four subjects by recording ERGs after 20 minutes dark adaptation to red flashes (0.2–2.0 cd s m-2) using skin electrodes and natural pupils. The effect of dark adaptation duration was investigated in 16 subjects during 20 minutes in the dark, by recording DA 1.5 red ERGs at 1, 2, 3, 4, 5, 10, 15 and 20 minutes.\ud \ud Results: For a dark adaption period of 20 minutes, the x-wave was more clearly visualised to weaker (< 0.6 cd s m-2) red flash strengths: to stronger flashes it became obscured by the b-wave. For red flashes of 1.5 cd s m-2, the x-wave was most prominent in ERGs recorded after 1–5 minutes of dark adaptation: with longer dark-adaptation, it was subsumed into the b-wave’s rising edge.\ud \ud Conclusions: This small study suggests that x-wave visibility in healthy subjects after 20 minutes dark adaptation is improved by using flashes weaker than around 0.6 cd s m-2; for flash strengths of 1.5 cd s m-2, x-wave visibility is enhanced by recording after only around 5 minutes of dark adaptation. No evidence was found that interim red flash ERGs affecting the dark-adapted state of the normal retina.

The International Society for Clinical Electrophysiology of Vision (ISCEV) standard for full-field electroretinography (ERG) describes a minimum procedure, but encourages more extensive testing. This ISCEV extended protocol describes an extension to the ERG standard, namely the dark-adapted (DA) red flash ERG. The DA red flash ERG can be incorporated conveniently within the ISCEV standard ERG protocol after a minimum of 20-min DA and recorded after the DA 0.01 ERG to a flash strength of 0.3 phot cd s m−2, eliciting a waveform with two positive peaks in healthy individuals. The first positive component is the cone-mediated x-wave with a peak at 30–50 ms; the second is a rod-mediated b-wave with a peak time of approximately 100 ms. Shorter DA times may be desirable to shorten the recording time or to alter the prominence of the early cone-mediated x-wave relative to the rod-mediated b-wave. The DA red flash ERG is used to aid the diagnosis of achromatopsia (rod monochromacy), cone dystrophy and other forms of cone system dysfunction, including “Bradyopsia” (RGS9/R9AP-retinopathy), when the DA red flash ERG x-wave is preserved in the absence of ISCEV standard LA ERGs. The DA red flash ERG can also help determine the origin of residual DA ERGs in cases of severe rod dysfunction, for example in disorders such as vitamin A deficiency, fundus albipunctatus (RDH5-retinopathy), Oguchi disease (SAG- or GRK1-retinopathy) and some rod-cone dystrophies. To shorter DA periods, the x-wave may be elicited without the following rod b-wave, shown to be helpful in abbreviated protocols for children.

Purpose Macular telangiectasia type 2 (MacTel) is a bilateral neurodegenerative disorder of the central macula. Previous findings indicated more functional impairment in low light conditions. We sought to further characterize retinal dysfunction using dark-adapted two-color fundus-controlled perimetry ("scotopic microperimetry"). Methods Participants of the MacTel Natural History Observation Registry study and age-matched healthy controls underwent retinal imaging including dual wavelength autofluorescence macular pigment optical density (MPOD) measurement. Retinal sensitivity was assessed with scotopic microperimetry using cyan (505 nm) and red (627 nm). Disease was graded into classes of MPOD loss (0 to 3). For perimetry analysis, the differences of the mean sensitivities (MacTel minus controls) were compared at each test location and the results were aggregated to global indices. Results Thirty-four eyes (19 patients, mean age 62.2 years) were compared with 25 eyes (25 controls, mean age 61.5 years). Both cyan and red sensitivity were lower in MacTel. This was more pronounced at one- and three-degree eccentricity. Eyes with MPOD class 0 did not exhibit a functional deficit. Class 1 had impaired cyan, but normal red sensitivity. Class 2 and 3 behaved similarly and had impaired cyan and red sensitivity with a relatively higher cyan impairment. Conclusions Rods might be compromised to a greater extent than cones. Linking to previous studies, our results might also hint toward (postreceptoral) dysfunction of the cone system in very early disease stages. Macular pigment loss and global perimetry indices seemed to reflect functional impairment and might be useful as adjunct measures for disease progression.

The International Society for Clinical Electrophysiology of Vision (ISCEV) standard for full-field electroretinography (ERG) describes a minimum protocol for clinical testing but encourages more extensive testing where appropriate. This ISCEV extended protocol describes an extension of the ISCEV full-field ERG standard, in which methods to record and evaluate the growth of the dark-adapted (DA) ERG b-wave with increasing stimulus energy are described. The flashes span a range that includes the weakest flash required to generate a reliable DA ERG b-wave and that required to generate a maximal b-wave amplitude. The DA ERG b-wave stimulus–response series (also known historically as the “intensity–response” or “luminance–response” series) can more comprehensively characterize generalized rod system function than the ISCEV standard ERG protocol and may be of diagnostic or prognostic value in disorders that cause generalized rod system dysfunction.

Purpose The purpose of this study was to characterize changes in the full-field flash electroretinogram (ERG) in association with psychophysical dark-adapted visual thresholds in patients with genetically characterized Duchenne muscular dystrophy (DMD) either lacking Dp427 (Up 30) or at least Dp260 in addition to Dp427 (Down 30). Methods Twenty-one patients with DMD and 27 age-similar controls participated in this study. Dark-adapted (0.01, 3.0, and 10 cd.s/m² flashes) and light-adapted (3.0 cd.s/m² flash) ERGs were recorded following International Society for Clinical Electrophysiology of Vision (ISCEV) standard protocols. Visual detection thresholds to 625-nm (cone function) and 527-nm (rod function) light-emitting diode (LED) flashes (2 degree diameter) were measured during a dark adaptation period after a 1-minute exposure to a bleaching light (3000 cd/m²). Initially, 8 minutes of interleaved 625-nm and 527-nm thresholds were measured. After an additional 5 minutes of dark-adaptation, a second set of threshold measurements to 527-nm stimuli was performed during the subsequent 6 minutes. Results Dark-adapted b-wave amplitude was significantly reduced to all strengths of flash and a-wave in response to the strong flash stimulus was delayed (15.6 vs. 14.7 ms, P < 0.05) in patients with Down 30 compared with controls. Dark-adapted cone thresholds did not differ among the groups (−2.0, −1.8, and −1.7 log cd/m² for Down 30, Up 30, and controls, respectively, P = 0.21). In contrast, dark-adapted rod thresholds were elevated (F(2,36) = 8.537, P = 0.001) in patients with Down 30 (mean = −3.2 ± 1.1 log cd/m²) relative to controls (mean = −4.2 ± 0.3 log cd/m²). Dark-adapted b-wave amplitudes were correlated with dark-adapted rod sensitivity in patients with DMD (Spearman Rho = 0.943, P = 0.005). The changes were much smaller or absent in patients with intact Dp260. Conclusions Dp260 is particularly required for normal rod-system function in dark adaptation.

Purpose To determine the intrasession test-retest reliability of mesopic and dark-adapted fundus-controlled perimetry in patients with intermediate age-related macular degeneration (iAMD). Methods We conducted a cross-sectional study with 23 iAMD patients (67.3 ± 8.2 years; range, 50-85; 78% female) and 24 healthy controls (61.3 ± 5.2 years; range, 50-71; 50% female) using a modified MAIA microperimeter. All patients underwent duplicate mesopic (achromatic stimuli, 400-800 nm) and dark-adapted (red stimuli, 627 nm) microperimetry, using a grid of 33 stimuli over 14° of the central retina. Main outcome measure was the intrasession test-retest reliability for pointwise sensitivity (PWS). Results PWS test-retest reliability was good among mesopic and dark-adapted testing in both patients and controls (coefficient of repeatability of 4.4, 4.52, 3.96, and 4.56 dB, respectively). Mean mesopic sensitivity in patients was 2.62 dB lower than in controls (P < 0.01); mean dark-adapted sensitivity was 2.49 dB lower than in controls (P < 0.01). Conclusions The modified MAIA device allows for reliable mesopic and dark-adapted microperimetry in iAMD patients. We found that iAMD is associated with both reduced mesopic and dark-adapted retinal sensitivity.

PURPOSE: To investigate retinal sensitivity in the junctional zone of geographic atrophy (GA) secondary to age-related macular degeneration using patient-tailored perimetry grids for mesopic and dark-adapted two-color fundus-controlled perimetry. METHODS: Twenty-five eyes with GA of 25 patients (prospective, natural-history Directional Spread in Geographic Atrophy study [DSGA; NCT02051998]) and 40 eyes of 40 normal subjects were included. Patient-tailored perimetry grids were generated using annotated fundus autofluorescence data. Customized software positioned test-points along iso-hulls surrounding the GA boundary at distances of 0.43°, 0.86°, 1.29°, 2.15°, and 3.01°. The grids were used for duplicate mesopic and dark-adapted two-color (cyan and red) fundus-controlled perimetry. Age-adjusted reference-data were obtained through regression analysis of normative data followed by spatial interpolation. RESULTS: The mean sensitivity loss for mesopic testing decreased with the distance to GA (-10.3 dB [0.43°], -8.2 dB [0.86°], -7.1 dB [1.29°], -6.8 dB [2.15°], and -6.6 dB [3.01°]; P < 0.01). Dark-adapted cyan sensitivity loss exceeded dark-adapted red sensitivity loss for all iso-hulls (-14.8 vs. -11.7 dB, -13.5 vs. -10.1 dB, -12.8 vs. -9.1 dB, -11.6 vs. -8.2 dB, -10.7 vs. -8.0 dB; P < 0.01). CONCLUSION: Patient-tailored fundus-controlled perimetry grids allowed for testing of retinal function in the junctional zone of GA with high spatial resolution. A distinct decrease in mesopic sensitivity loss between 0.43° (125 µm) and 1.29° (375 µm) was observed that leveled off at more distant test-points. In proximity to the GA boundary, the results indicate that rod exceeded cone dysfunction.

Our aim was to compare the electroretinographic (ERG) responses of two eyes obtained by consecutive unilateral recordings to those obtained by a simultaneous bilateral recording in sheep. Eight sheep underwent two full-field ERG recordings, using two recording strategies of the standard ISCEV protocol: consecutive unilateral recordings of one eye after the other, and simultaneous bilateral recording of both eyes. The order of recording strategy within an animal (unilateral/bilateral), eye recording sequence in the unilateral session (OD/OS), and amplifier channel assignment for each eye were all randomized. To test whether duration of dark adaptation and/or anesthesia affect the results, the ISCEV protocol was recorded bilaterally in six additional eyes following 38 min of patched dark adaptation, as was done for the second eye recorded in the consecutive unilateral recordings. The second recorded eye in the unilateral session had significantly higher scotopic b-wave amplitudes compared to the first recorded eye and to the bilaterally recorded eyes. A-wave amplitudes of the dark-adapted mixed rod–cone responses to a high-intensity flash were also significantly higher in the second eye compared to the first eye recorded unilaterally and to the bilaterally recorded eyes. Light-adapted responses were unaffected by the recording strategy. When the ISCEV protocol was recorded after 38 min of dark adaptation, the scotopic responses were higher than in the first eyes, and similar to those of the second eyes recorded unilaterally, suggesting that indeed the longer duration of anesthesia and dark adaptation are responsible for the increased scotopic responses of the second eye. Consecutive unilateral ERG recordings of two eyes result in higher amplitudes of the dark-adapted responses of the eye recorded second, compared to the eye recorded first and to bilaterally recorded eyes. The differences in scotopic responses can be attributed to different duration of dark adaptation and/or anesthesia of the two consecutively recorded eyes. Photopic responses are not affected. Therefore, simultaneous bilateral ERG responses should be recorded when possible, especially for evaluation of scotopic responses.

The ‘temporal rule’ of multisensory integration (MI) proposes that unisensory stimuli, and the neuronal responses they evoke, must fall within a window of integration. Ecological validity demands that MI should occur only for physically simultaneous events (which may give rise to non‐simultaneous neural activations), and spurious neural response simultaneities unrelated to environmental multisensory occurrences must somehow be rejected. Two experiments investigated the requirements of simultaneity for facilitative MI. Experiment 1 employed an reaction time (RT)/race model paradigm to measure audiovisual (AV) MI as a function of AV stimulus‐onset asynchrony (SOA) under fully dark adapted conditions for visual stimuli that were either rod‐ or cone‐isolating. Auditory stimulus intensity was constant. Despite a 155‐ms delay in mean RT to the scotopic vs. photopic stimulus, facilitative AV MI in both conditions occurred exclusively at an AV SOA of 0 ms. Thus, facilitative MI demands both physical and physiological simultaneity. Experiment 2 investigated the accuracy of simultaneity and temporal order judgements under the same stimulus conditions. Judgements of AV stimulus simultaneity or temporal order were significantly influenced by stimulus intensity, indicating different simultaneity requirements for these tasks. The possibility was considered that there are mechanisms by which the nervous system may take account of variations in response latency arising from changes in stimulus intensity in order to selectively integrate only those physiological simultaneities that arise from physical simultaneities. It was proposed that separate subsystems for AV MI exist that pertain to action and perception.

The characteristics of aquatic light fields are generally reflected in the visual systems of fishes inhabiting them. Therefore, research on light sensitivity of fish is useful to explain the correlation between the visual function and habitat, behavior and distribution of fish. Rockfish is an important species in coastal ecology and also one of the main species for culturing in Korea. To make a contribution on the maintenance of the fish resources and understanding the ecology of the rockfish, the visual spectral sensitivities of the dark-adapted rockfishes were measured in the range of visible light (405~660 nm) using a light source of light-emitting diodes (LEDs). In order to assess electrophysiological response of the fish, the ERGs (electroretinograms) of the dark-adapted rockfishes were recorded on a data logger (12 bits) and a laptop computer. Juvenile (n

To evaluate rod and cone contributions to the dark-adapted 15-Hz flicker electroretinogram (ERG) across a broad range of stimulus luminances by comparing rod-isolating (ERGR), cone-isolating (ERGC), and non-receptor-specific (ERGR+C) responses.Dark-adapted, full-field 15-Hz ERGs were obtained from four normally sighted subjects (ages 29-36 years) using a four-primary LED-based stimulating system. The primaries were either modulated sinusoidally in phase (ERGR+C) or were modulated in counter-phase to achieve rod isolation (ERGR) or cone isolation (ERGC) by means of triple silent substitution. Measurements were made for a broad range of luminances (-2.5 to 1.8 log scot. cd/m(2) in 0.2 log unit steps). Fourier analysis was used to obtain the amplitude and phase of the fundamental response component at each stimulus luminance.Stimulus luminance had different effects on response amplitudes and phases under the three paradigms. Specifically, ERGC amplitude and phase increased monotonically as luminance increased. The effects on ERGR+C and ERGR were complex: ERGR+C and ERGR amplitude was small and the phase decreased for low luminances, whereas amplitude and phase increased sharply at moderate luminances. For high luminances, ERGR+C amplitude and phase increased, whereas ERGR amplitude decreased and phase was approximately constant.At low luminances, the ERGR+C and ERGR functions can be attributed to interactions between two rod pathways. At high luminances, the functions can be accounted for by interactions between rod and cone pathways (ERGR+C) or rod insensitivity (ERGR). The ERGR paradigm minimizes cone intrusion, permitting assessment of rod function over a large range of luminance levels.

Purpose To determine the effective dynamic range (EDR), retest reliability, and number of discriminable steps (DS) for mesopic and dark-adapted two-color fundus-controlled perimetry (FCP) using the S-MAIA (Scotopic-Macular Integrity Assessment) "micro-perimeter." Methods In this prospective cross-sectional study, each of the 52 eyes of 52 subjects with various macular diseases (mean age 62.0 ± 16.9 years; range, 19.1-90.1 years) underwent duplicate mesopic (achromatic stimuli, 400-800 nm), dark-adapted cyan (505 nm), and dark-adapted red (627 nm) FCP using a grid of 61 stimuli covering 18° of the central retina. The EDR, the number of DS, and the retest reliability for point-wise sensitivity (PWS) were analyzed. The effects of fixation stability, sensitivity, and age on retest reliability were examined using mixed-effects models. Results The EDR was 10 to 30 dB with five DS for mesopic and 4 to 17 dB with four DS for dark-adapted cyan and red testing. PWS retest reliability was good among all three types of retinal sensitivity assessments (coefficient of repeatability ±5.79, ±4.72, and ±4.77 dB, respectively) and did not depend on fixation stability or age. PWS had no effect on retest variability in dark-adapted cyan and dark-adapted red testing but had a minor effect in mesopic testing. Conclusions Combined mesopic and dark-adapted two-color FCP allows for reliable topographic testing of cone and rod function in patients with various macular diseases with and without foveal fixation. Retest reliability is homogeneous across eccentricities and various degrees of scotoma depth, including zones at risk for disease progression. These reliability estimates can serve for the design of future clinical trials.

Author(s): Panorgias, Athanasios; Tillman, Megan; Sutter, Erich E; Moshiri, Ala; Gerth-Kahlert, Christina; Werner, John S | Abstract: PurposeTo investigate the topographic changes of the dark-adapted multifocal electroretinogram (mfERG) across adulthood in the central retina and compare the topography between macular versus extramacular, nasal versus temporal, and inferior versus superior retinal areas.MethodsSixty-five subjects (18-88 years) received a comprehensive dilated eye examination to ensure the health of their retina and were tested with a dark-adapted mfERG protocol using a 61-hexagon pattern. The lens absorption of each subject was also estimated using a heterochromatic flicker photometry (HFP) paradigm.ResultsThe response amplitude and latency of the dark-adapted mfERG showed a significant change with age, which was best described with a linear model. All the retinal areas examined demonstrated similar aging effects. The extramacular and temporal retina showed higher response amplitude and faster response latency when compared with the macular and nasal retinae, respectively. No difference was found in response amplitude and latency between the inferior and superior retina. The HFP results also showed a significant correlation with age, consistent with senescent increases in short wavelength absorption by the crystalline lens. However, the change in lens absorption did not exceed the magnitude of the change in response amplitude and latency.DiscussionOur results indicate that there is a decline in dark-adapted retinal activity as measured with the mfERG. These aging processes affect rods and rod-bipolar cells. Their decrease in response can be attributed to both optical and neural factors.

In higher plants, thylakoid membrane protein complexes show lateral heterogeneity in their distribution: photosystem (PS) II complexes are mostly located in grana stacks, whereas PSI and adenosine triphosphate (ATP) synthase are mostly found in the stroma-exposed thylakoids. However, recent research has revealed strong dynamics in distribution of photosystems and their light harvesting antenna along the thylakoid membrane. Here, the dark-adapted spinach (Spinacia oleracea L.) thylakoid network was mechanically fragmented and the composition of distinct PSII-related proteins in various thylakoid subdomains was analyzed in order to get more insights into the composition and localization of various PSII subcomplexes and auxiliary proteins during the PSII repair cycle. Most of the PSII subunits followed rather equal distribution with roughly 70% of the proteins located collectively in the grana thylakoids and grana margins; however, the low molecular mass subunits PsbW and PsbX as well as the PsbS proteins were found to be more exclusively located in grana thylakoids. The auxiliary proteins assisting in repair cycle of PSII were mostly located in stroma-exposed thylakoids, with the exception of THYLAKOID LUMEN PROTEIN OF 18.3 (TLP18.3), which was more evenly distributed between the grana and stroma thylakoids. The TL29 protein was present exclusively in grana thylakoids. Intriguingly, PROTON GRADIENT REGULATION5 (PGR5) was found to be distributed quite evenly between grana and stroma thylakoids, whereas PGR5-LIKE PHOTOSYNTHETIC PHENOTYPE1 (PGRL1) was highly enriched in the stroma thylakoids and practically missing from the grana cores. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: Keys to Produce Clean Energy.

Purpose To determine the retest variability of mesopic and two-color dark-adapted (DA) fundus-controlled perimetry (FCP), to evaluate the predictive value of patient reliability indices, and to analyze the extent of impairment of rod- and cone function in neovascular age-related macular degeneration (nAMD). Methods A total of 50 eyes of 50 patients with nAMD (mean age, 76.1 years) and 70 eyes of 70 age-similar normal subjects underwent multimodal imaging as well as mesopic and DA two-color perimetry using the S-MAIA device. A subset of patients (n = 28) underwent duplicate testing for retest reliability assessment. Mixed models were used for analysis of the hierarchical data. Results In eyes with nAMD, the coefficient of repeatability was (mean ± standard deviation [SD]) 5.99 ± 1.55 dB for mesopic, 6.14 ± 2.19 dB for DA cyan, and 6.06 ± 1.09 dB for DA red testing. “Patient reliability indices” explained 55%, 54.2%, and 64.2% of the variance in retest variability. The mean sensitivity loss was greater for DA cyan compared to DA red testing (cyan-red differences [mean ± SD] −2.63 ± 3.87 dB, P < 0.001). Conclusions The relatively greater degree of DA cyan versus DA red sensitivity loss indicates preferential rod vulnerability in nAMD, and qualifies rod function-based outcomes measures as potential sensitive and early markers of treatment response in nAMD. Translational Relevance The S-MAIA allows reliable testing of mesopic, DA cyan, and DA red sensitivity in patients with nAMD. Patient reliability indices may serve as eligibility criteria for clinical trials to identify patients with adequate retest reliability.

The most dominant feature of the electroretinogram, the b-wave, is thought to reflect ON-bipolar cell responses. However, a number of studies suggest that the b-wave is made up of several components. We consider the composition of the rat b-wave by subtracting corneal negative components obtained using intravitreal application of pharmacological agents to remove postreceptoral responses. By analyzing the intensity-response characteristic of the PII across a range of fixed times during and after a light step, we find that the rat isolated PII has 2 components. The first has fast rise and decay characteristics with a low sensitivity to light. GABAc-mediated inhibitory pathways enhance this transient-ON component to manifest increased and deceased sensitivity to light at shorter (

Walden-Coleman, A. E., Rajcan, I. and Earl, H. J. 2013. Dark-adapted leaf conductance, but not minimum leaf conductance, predicts water use efficiency of soybean (Glycine max L. Merr.). Can. J. Plant Sci. 93: 13–22. The conductance to water vapor of dark-adapted leaves (gdark) has been shown to be negatively correlated with whole-plant water use efficiency (WUE) in soybean, but the physiological basis of this relationship is unknown. It is also not clear how gdark compares with the minimum leaf conductance of wilted leaves (gmin), a trait that has been studied extensively across a broad range of species. We compared gdark to gmin of soybean leaves and found that gdark values were consistently much higher than gmin values measured on the same leaves. Also, across seven soybean varieties known to differ for WUE, gdark but not gmin was correlated with WUE. Thus, gdark and gmin should be considered distinct traits. We measured gdark at two different leaf positions, and found that gdark measured at the lower leaf position (two main stem nodes below the youngest fully expanded leaf) was best correlated with WUE. We then used this method to screen a selection of current commercial soybean varieties adapted to Ontario, Canada, for variation in gdark. The range in gdark among the commercial varieties was as broad as that measured previously among more diverse genotypes, suggesting that Ontario soybean varieties might also vary widely for WUE.

This paper reports the first electrolysis-based oxygen generation and transportation device for the treatment of retinal ischemia; containing an isolated biocompatible electrolyte and inductive power transfer for on-demand oxygen. A finite element model of the retina along with oxygen delivery data demonstrates the feasibility of the device. The estimated therapeutic window for the device was found to be 210 minutes for the light adapted retina, and 55 minutes for the dark adapted retina.

The aim of this research was to clarify and quantify the demands of the working environment and watch-keeping regime for large commercial ships in relation to dark adaptation. The night lookout task requires the identification of the relatively bright navigational lights of other ships against the dark background of the sky and sea. The probability of detection is determined by the ambient lighting conditions on the bridge and the dark adapted state of vision. Light levels were such that threshold sensitivity (after 15 minutes) was reduced by around 2 log units in comparison to complete darkness. This has implications for the effective range of navigational lights at sea as defined in regulations. The intensity and position of navigation lights on larger vessels is such that the sensitivity of the eye under typical bridge conditions is likely to be sufficient for their visibility to be acceptable. This may not be the case for less well lit small craft.

Chlorophyll fluorescence is a well established technique to rapidly and non-invasively determine photosynthesis parameters in plant leaves. It can be used in both laboratory and field settings, and frequently dark-adaptation of a leaf sample is called for. In the field, this can be accomplished on flat leaves using standard leaf clips supplied by instrument manufacturers. However, not all plant leaves are flat, many are cylindrical or otherwise three-dimensional in shape. The standard leaf clip does not close fully on three-dimensional leaves, therefore, does not allow the sample to be properly dark adapted in the field. A new leaf “wand” was developed that can be slipped over an entire cylindrical leaf or culm of rushes and sedges for both light- and dark-adapted measurements. This new leaf wand is compared to the standard leaf clip (DLC-8) using a Walz mini-PAM on Juncus roemerianus(Black needlerush). Results indicate that dark-adapted yield measurements are not significantly different between leaf clips, while light-adapted yields are higher with the leaf wand. The potential sources of difference in the optical path of the excitation light and fluorescence return are discussed and compared between leaf clips. Construction of specialized leaf wands should be considered for any leaves are not flat and therefore that do not fit the standard leaf clip for complete dark-adaptation under field conditions.

Purpose: To determine whether the pupillary light reflex (PLR) can serve as an indicator of photoreceptor function in patients with advanced typical retinitis pigmentosa (RP). Methods: Dark-adapted transient PLRs elicited by blue or white light over a luminance range of 4 log units were recorded from 27 eyes of 19 patients with advanced RP. Retinas were characterized according to fundus autofluorescence (AF) and dark-adapted perimetry. We qualitatively analyzed whether PLR thresholds were correlated with AF patterns or scotopic sensitivity. Quantitative analysis included correlations between relative pupillary constrictions (RPCs) elicited by blue light (≤–1 log cd/m2) and the area of abnormal ring or central AF, and between RPCs elicited by white light and perimetric mean sensitivity. Results: The PLRs of all patients showed varying degrees of threshold elevation and relative afferent pupil defects. We classified three types of abnormal fundus AF: abnormal ring AF, abnormal central AF, and fragmentary AF. PLR thresholds were largely consistent with the patterns of AF and scotopic sensitivity. Rod-mediated RPCs were not correlated with the area of the abnormal ring AF (p > 0.05), but were correlated with the area of abnormal central AF (p < 0.05). RPCs elicited with a white stimulus (–0.3 or 0.7 log cd/m2) were significantly correlated with the mean sensitivity of the dark-adapted perimetry. Conclusions: PLR testing is a powerful technique for assessing photoreceptor dysfunction. The high correlation with AF and dark-adapted perimetry suggests that the key to quantifying photoreceptor function using the transient PLR is to optimize the luminance of the stimulus.

This study characterizes differences in human ERGs based on ocular pigmentation. Light- and dark-adapted luminance-response (LR) series for a-, b- and i-waves and light-adapted oscillatory potentials (OPs) were recorded in 14 healthy volunteers (7 blue-eyed Caucasians; 7 brown-eyed Asians, aged 20–22 years). Amplitude interpolations were by logistic growth (Naka-Rushton), Gaussian or the combined ‘photopic hill’ functions. Implicit times (IT) for dark-adapted a- and b-waves, and for light-adapted a-, b- and i-waves were earlier in the blue-eyed group than in the brown-eyed group across all flash strengths (P < 0.05). For dark-adapted ERGs, saturated a-wave amplitude was larger for blue eyes (397 vs. 318 μV, P < 0.05) as was the a-wave to strong flash (10 cd·s/m2; 357 vs. 293 μV, P < 0.05) and the b-wave to ISCEV standard 0.01 (354 vs. 238 μV, P < 0.05). Light-adapted b-waves for midrange flash stimuli were much larger for the blue-eyed group (photopic hill, Gaussian peak: 155 vs. 82 μV, P < 0.001) with no difference in saturated amplitudes. Similarly, interpolated i-wave amplitudes were larger (48 vs. 18 μV, P < 0.01). For a light-adapted 2.6 stimulus, a- and b-waves were larger for the blue-eyed group (52 vs. 39 μV; 209 vs. 133 μV, P < 0.01) as were OP4 and OP5 (37.2 vs. 15.6 μV; 47.5 vs. 22.2 μV, P < 0.01), but OP1-OP3 did not differ. ERGs have shorter ITs in people with blue irides than in those with dark pigmentation. Amplitude differences are highly non-linear and substantially larger from eyes with light pigmentation for components thought to be associated with the OFF retinal pathways.

Based on a proposed model of the operation of the retina of a dark-adapted eye, the absolute sensitivity of the eye is calculated and is compared with that of a camera based on a cooled electron-multiplying CCD array. It is shown that the lowest observable brightnesses coincide in the two cases. The main factors that provide high visual sensitivity are indicated: a molecular light detector, a large detector area with a special noise-suppression system, and the absence of losses to scattering accompanying night vision.

The relatively large eye and pupil of the Piked Dogfish, Squalus acanthias, is visually arresting. However, knowledge of its basic visual characteristics lags far behind other areas in this generally well studied species. This study quantifies pupil dilation in a species that is naturally exposed to a broad range of light intensities and finds that the pupil area in the dark adapted state is 35.3% of the total eye area, an increase of 12.4% from the light adapted state. The anterior and posterior extents of the horizontal visual field are assessed and compared with both morphological and electrophysiological techniques and the results are integrated with the measured head yaw to derive the anterior convergence distance and blind area. The position of the eyes and the triangular, pointed snout of S. acanthias provides excellent anterior vision, which likely facilitates foraging upon its mobile prey.

Two EOG parameters - The Arden ratio (A) and an expression devised by Gliem (G) - from a sample of normal human subjects are studied. Their relations to the EOG potential and time parameters and to sex, age, pupillary diameter, degree of iris pigmentation, refractive error and axial length, ocular protrusion and interpupillary distance were assessed. Right eye and left eye distributions were congruent, although individual differences were sometimes appreciable, especially in the case of G. The two ratios were positively correlated and both showed a negative correlation to the interval between the dark trough and the light peak. Correlations to the EOG potential parameters, especially the light induced potential rise of the dark adapted eye, were also established. In the case of A, a higher level was disclosed in the male half of the sample. A negative age correlation, predominant in the female part of the sample, characterized both ratios. The two ratios were positively correlated to the pupil diameter, negatively correlated to the degree of refractive errror, and positively correlated to the degree of ocular protrusion. The consequences for the clinical EOG test are discussed, and it is concluded that the present EOG procedure is a qualitative rather than a quantitative test.

Purpose To determine the effects of a standardized intravenous dose of an α-2 agonist (Domitor®, Orion Pharma, distributed by Pfizer Animal Health, Exton, PA) on the electroretinogram (ERG) response in normal dogs. Methods Twenty-five normal dogs were used to collect ERG responses including a- and b-wave implicit times (IT) and amplitudes (AMP) before and after administration of medetomidine. Dogs were dark adapted for 20 min and ERGs were obtained using the HMsERG (RetVetCorp Inc., Columbia, MO). The QuickRetCheck protocol (Narfstrom) was employed to provide the following flash intensities: 10 mcd s/m2, 3 cd s/m2, and 10 cd s/m2. ERGs were repeated after 375 µg/m2 of medetomidine intravenously. Statistical analysis of the difference between the responses before and after medetomidine at all flash intensities was performed using a mixed effects model for anova. Results The P value for the effect of medetomidine on each of the ERG responses was

Glial Ca2+ excitability plays a key role in reciprocal neuron–glia communication. In the retina, neuron–glia signalling is expected to be maximal in the dark, but the glial Ca2+ signal characteristics under such conditions have not been evaluated. To address this question, we used bioluminescence imaging to monitor spontaneous Ca2+ changes under dark conditions selectively in Muller cells, the principal retinal glial cells. By combining this imaging approach with network analysis, we demonstrate that activity in Muller cells is organized in networks of coactive cells, involving 2–16 cells located distantly and/or in clusters. We also report that spontaneous activity of small networks (2–6 Muller cells) repeat over time, sometimes in the same sequential order, revealing specific temporal dynamics. In addition, we show that networks of coactive glial cells are inhibited by TTX, indicating that ganglion and/or amacrine neuronal cells probably regulate Muller cell network properties. These results represent the first demonstration that spontaneous activity in adult Muller cells is patterned into correlated networks that display repeated sequences of coactivations over time. Furthermore, our bioluminescence technique provides a novel tool to study the dynamic characteristics of glial Ca2+ events in the retina under dark conditions, which should greatly facilitate future investigations of retinal dark-adaptive processes.

The chromaticity of unique white viewed in illumination mode and under dark adapted conditions was investigated for 3 luminance levels (200, 1000 and 2000 cd/m²) using a unique white setting method. Unique white was found to encompass a rather large region in color space located slightly below the blackbody locus and centered around a CCT of 6600 K. Luminance level was found to have no significant effect on the mean unique white chromaticity. The high and low end points of the CIE class A and B white regions respectively under- and overestimate the chromaticity region perceived as white. Agreement along the Duv direction was quite good. However, another Duv related limit associated with white lighting (|Duv|≤5.4e-3) was found to be on the small side, especially for chromaticity values below the blackbody locus. The results for unique white viewed in illumination mode were compared to those reported for object mode presentation. Overall they were very comparable, although a statistical analysis does show a (just) significant effect of stimulus presentation mode for high (il)luminance levels. However, as no such effect could be established at the individual observer level, it was concluded that unique white chromaticity is essentially the same for both illumination and object mode stimulus presentation. ispartof: Optics Express vol:23 issue:10 pages:12488-12495 ispartof: location:United States status: published

Dopamine (DA) is an important modulator of light-adaptation in the vertebrate retina. A known source of DA in the retina is amacrine or interplexiform cells, but another potential source is the ret...

Part I: Historical Overview 1. The G Protein-Coupled Receptor Rhodopsin: A Historical Perspective Lukas Hofmann and Krzysztof Palczewski Part II: Rhodopsin Expression, Regeneration and Purification for Structural Studies 2. Rhodopsin Purification from Dark Adapted Bovine Retina Elise Blankenship and David T. Lodowski 3. Mammalian Expression, Purification, and Crystallization of Rhodopsin Variants Daniel Mattle, Ankita Singhal, Georg Schmid, Roger Dawson, and Jorg Standfuss 4. Imaging of Rhodopsin Crystals with Two-Photon Microscopy Grazyna Palczewska and David Salom Part III: Structure-function characterization of rhodopsin 5. Functional Stability of Rhodopsin in a Bicelle System: Evaluating G-Protein Activation by Rhodopsin in Bicelles Ali I. Kaya, T. M. Iverson, and Heidi E. Hamm 6. The Rhodopsin-Arrestin-1 Interaction in Bicelles Qiuyan Chen, Sergey A. Vishnivetskiy, Tiandi Zhuang, Min-Kyu Cho, Tarjani M. Thaker, Charles R. Sanders, Vsevolod V. Gurevich, and T. M. Iverson 7. Detection of Structural Waters and Their Role in Structural Dynamics of Rhodopsin Activation Liwen Wang and Mark Chance 8. Probing Conformational Changes in Rhodopsin Using Hydrogen-Deuterium Exchange Coupled to Mass Spectrometry Tivadar Orban and Yaroslav Tsybovsky 9. Analysis of Conformational Changes in Rhodopsin by Histidine Hydrogen-Deuterium Exchange David T. Lodowski and Masaru Miyagi 10. Investigation of Rhodopsin Dynamics in its Signaling State by Solid-State Deuterium NMR Spectroscopy Andrey V. Struts, Udeep Chawla, Suchithranga M.D.C. Perera, and Michael F. Brown 11. Sequential Structural Changes in Rhodopsin Occurring upon Photoactivation Naoki Kimata, Andreyah Pope, Dawood Rashid, Philip J. Reeves, and Steven O. Smith 12. Dynamic Single-Molecule Force Spectroscopy of Rhodopsin in Native Membranes Paul S.-H. Park and Daniel J. Muller Part IV: Rhodopsin Supramolecular Organization and its Complexes with Partner Proteins 13. High-Resolution Atomic Force Microscopy Imaging of Rhodopsin in Rod Outer Segment Disk Membranes Patrick D. Bosshart, Andreas Engel, and Dimitrios Fotiadis 14. Detection of Rhodopsin Dimerization in situ by PIE-FCCS, a Time-Resolved Fluorescence Spectroscopy Adam W. Smith 15. Oligomeric State of Rhodopsin within Rhodopsin-Transducin Complex Probed with Succinylated Concanavalin A Beata Jastrzebska 16. Quantification of Arrestin-Rhodopsin Binding Stoichiometry Ciara C.M. Lally and Martha E. Sommer 17. Rhodopsin Transient Complexes Investigated by Systems Biology Approaches Daniele Dell'Orco Part V: Rhodopsin and Photoreceptors 18. Three Dimensional Architecture of Murine Rod Cilium Revealed by Cryo-EM Theodore G. Wensel and Jared C. Gilliam 19. Monitoring of Rhodopsin Trafficking and Mistrafficking in Live Photoreceptors Kerrie H. Lodowski and Yoshikazu Imanishi 20. Measurements of Rhodopsin Diffusion within Signaling Membrane Microcompartments in Live Photoreceptors Mehdi Najafi and Peter D. Calvert Part VI: Treatment Strategies of Retinal Degenerative Disease 21. Kinetics of Rhodopsin's Chromophore Monitored in a Single Photoreceptor Leopold Adler IV, Nicholas P. Boyer, Chunhe Chen, and Yiannis Koutalos 22. Supplementation with Vitamin A Derivatives to Rescue Vision in Animal Models of Degenerative Retinal Diseases Lindsay Perusek, Akiko Maeda, and Tadao Maeda 23. Sustained Delivery of Retinoids to Prevent Photoreceptor Death Peter H. Tang and Rosalie K. Crouch 24. High-Throughput Screening Assays to Identify Small-Molecules Preventing Photoreceptor Degeneration Caused by the Rhodopsin P23H Mutation Yuanyuan Chen and Hong Tang 25. Gene Therapy to Rescue Retinal Degeneration Caused by Mutations in Rhodopsin Brian P. Rossmiller, Renee C. Ryals, and Alfred S. Lewin

Structural and biophysical studies of rhodopsin have long depended upon the ready availability of bovine retina from the meat-packing industry and the relative ease of obtaining homogenous preparations of rhodopsin in the quantities and purities necessary for such study. Herein we present a modular purification methodology employing a combination of several strategies, beginning with sucrose gradient isolation of rod outer segments (ROS) from bovine retina, detergent solubilization of ROS, selective extraction of rhodopsin starting from this detergent-solubilized ROS, and further purification via size-exclusion chromatography, resulting in a preparation of high-purity rhodopsin at high concentration suitable for crystallization or other biophysical study.

Dark-adapted rods in the area surrounding a luminance-modulated field can suppress flicker detection. However, the characteristics of the interaction between rods and each of the cone types are unclear. To address this issue, the effect that dark-adapted rods have on specific classes of receptoral and postreceptoral signals was determined by measuring the critical fusion frequencies (CFF) for receptoral L-, M-, and S-cone and postreceptoral luminance ([L+M+S] and [L+M+S+Rod]) and chromatic ([L/(L+M)]) signals in the presence of different levels of surrounding rod activity. Stimuli were generated with a two-channel photostimulator that has four primaries for a central field and four primaries for the surround, allowing independent control of rod and cone excitation. Measurements were made either with adaptation to the stimulus field after dark adaptation or during a brief period following light adaptation. The results show that dark-adapted rods maximally suppressed the CFF by ∼6 Hz for L-cone, M-cone, and luminance modulation. Dark-adapted rods, however, did not significantly alter the S-cone CFF. The [L/(L+M)] postreceptoral CFF was slightly suppressed at higher surround illuminances, that is, higher than surround luminances resulting in suppression for L-cone, M-cone, or luminance modulation. We conclude that rod-cone interactions in flicker detection occurred strongly in the magnocellular pathway.

SUMMARY The purpose of this study is to analyze the time-dependent molecular states of rhodopsin (Rho) phosphorylation in the specimens originating from eyeballs cryoimmobilized in situ in living animals. Whole eyeballs of living mice under various dark- and light-exposure conditions were quickly frozen using the in vivo cryotechnique with isopentane–propane cryogen cooled down in liquid nitrogen (196C). The frozen whole-mount eyeballs were freeze substituted in acetone containing paraformaldehyde and embedded in paraffin wax. Deparaffinized sections were immunostained with anti-phosphorylated 334 Ser Rho (PRho334) antibody. Immunoreactivity of P-Rho334 was specifically recognized in the outer segments of mouse retinas exposed to daylight. In the 12-h dark-adapted retinas, P-Rho334 immunoreactivity was completely eliminated. Moreover, in other retinas dark adapted for 12 or 36 hr and then exposed under the safety red light for 2 min, it was still barely recognized. Even in the eyeballs exposed to strong visible light for 10 sec, it was not detected. However, after 30, 60, and 180 sec of visible light exposure, P-Rho334 immunoreactivity was definitely recovered, similar to that under daylight condition. This is a new immunohistochemical approach to visualize the time-dependent Rho phosphorylation of living mice using the in vivo cryotechnique, in which changes could be detected within seconds following exposure to light. (J Histochem Cytochem 54:479–486, 2006)

Purpose: To determine the individual variability of the dark-adapted pupil diameter over 2 testing sessions using a standardized dark-adaptation protocol. Setting: Texas Tech University Health Sciences Center, Lubbock, Texas, USA. Methods: In this prospective observational cohort study, 40 volunteers with no history of ocular disease, surgery, or injury other than requirement for refractive correction were included. The right eye was tested. A standardized dark-adaptation protocol was used that controlled for accommodation and patient alertness. Infrared, still digital photographs were taken after 5 and 10 minutes of dark adaptation and analyzed independently by 2 investigators using digital-image software. Two test sessions were performed 1 to 7 days apart. Lifestyle factors such as sleep, diet, and exercise were not controlled. Results: The mean subject age was 31.5 years (range 20 to 49 years). There were 20 men; 27 subjects wore correction for myopia, and 13 wore no correction. The mean interval between test sessions was 2 days (range 1 to 7 days). The mean difference and 95% confidence intervals for pupil diameter difference between sessions were as follows: 5-minute readings, +0.032 mm (−0.030 to +0.094); 10-minute readings, −0.006 mm (−0.059 to +0.047); mean of 5- and 10-minute readings, +0.013 mm (−0.038 to +0.064). Using the paired t test, the pupil diameter did not differ significantly between sessions in 5-minute dark adaptation ( P = .2980), 10-minute dark adaptation ( P = .8263), or the mean ( P = .6049). Conclusion: Using a consistent dark-adaptation protocol that controlled for alertness, individuals aged 20 to 49 years showed no significant variation in dark-adapted pupil diameter when tested twice in 1 week.

The dark-adapted cone electroretinogram (ERG) is difficult to isolate because of unwanted rod intrusion. We compare dark-adapted cone estimates derived using three techniques. The first uses the cone response on a moderate rod saturating background to estimate the dark-adapted cone response. The second uses red and blue flashes to tease apart cone and rod responses (red-minus-blue technique, [Investigative Ophthalmology and Visual Science 31 (1990) 2283]). The third uses a bright flash to temporarily saturate rods, followed by a test flash that generates a putative cone-only response (2-flash technique [Investigative Ophthalmology and Visual Science 36 (1995) 1603]). By subtracting the cone estimates from ‘mixed’ ERG responses in the dark, rod isolated responses can be derived. The rod phototransduction parameters, derived using a computational model, are similar using the light-adapted and 2-flash cone estimates, but differ using the red-minus-blue estimates. The 2-flash cone estimate gives a cone waveform similar to the dark-adapted response of a patient with Oguchi stationary night blindness (a patient with no rod ERG responses and normal cone ERG responses). The growth of the cone response during light adaptation to steady backgrounds causes significant differences between the light-adapted and 2-flash cone waveforms at times beyond the first few milliseconds.

One of the purposes of this study was to examine whether b-wave measurements can be used in the evaluation of scotopic spectral sensitivity in Prussian carp measurements when the eyes were surgically deprived of cornea, lens, and most of the vitreous. Another goal was testing the new fitting procedure for A2-based photopigments. Using fitted amplitude-log intensity functions for threshold calculation, and two models for computer-assisted fitting of spectral sensitivity curves, no significant differences in λ max were found between rod photopigments and b-wave-based spectral sensitivity.

BACKGROUND: The retinal response to flickering stimuli (steady state ERG) recruits many retinal elements and is a sensitive indicator of early retinal dysfunction. This study reports the post-natal maturation of the steady state ERG response in guinea pigs. METHODS: The steady state ERG response to flickering stimuli (0.6 to 20 Hz) was recorded from dark adapted (more than 12 hrs) English Shorthair guinea pigs (n = 7) using flashes that produced rod and cone dominated responses. Temporal sensitivity functions and critical fusion frequencies (CFF) were derived over a range of ages from postnatal day (PND) 1 to 45. RESULTS: Guinea pig rod and cone temporal sensitivity functions show shape characteristics and CFF similar to humans. Furthermore, the post-natal development of the guinea pig temporal characteristics is also similar to that of humans - they are present at birth and mature rapidly post-natally. The time-course of CFF maturation is similar for rod and cone mediated responses. CONCLUSIONS: These data show that the temporal response and its maturation in the guinea pig retina is similar to that in humans. Therefore, we propose that the guinea pig is a particularly useful animal model to study retinal disease in early childhood.