Flicker Stimulation Research Articles

Audio-visual synchrony and spatial attention enhance processing of dynamic visual stimulation independently and in parallel: A frequency-tagging study

The neural processing of a visual stimulus can be facilitated by attending to its position or by a co-occurring auditory tone. Using frequency-tagging, we investigated whether facilitation by spatial attention and audio-visual synchrony rely on similar neural processes. Participants attended to one of two flickering Gabor patches (14.17 and 17 Hz) located in opposite lower visual fields. Gabor patches further “pulsed” (i.e. showed smooth spatial frequency variations) at distinct rates (3.14 and 3.63 Hz). Frequency-modulating an auditory stimulus at the pulse-rate of one of the visual stimuli established audio-visual synchrony. Flicker and pulsed stimulation elicited stimulus-locked rhythmic electrophysiological brain responses that allowed tracking the neural processing of simultaneously presented Gabor patches. These steady-state responses (SSRs) were quantified in the spectral domain to examine visual stimulus processing under conditions of synchronous vs. asynchronous tone presentation and when respective stimulus positions were attended vs. unattended. Strikingly, unique patterns of effects on pulse- and flicker driven SSRs indicated that spatial attention and audiovisual synchrony facilitated early visual processing in parallel and via different cortical processes. We found attention effects to resemble the classical top-down gain effect facilitating both, flicker and pulse-driven SSRs. Audio-visual synchrony, in turn, only amplified synchrony-producing stimulus aspects (i.e. pulse-driven SSRs) possibly highlighting the role of temporally co-occurring sights and sounds in bottom-up multisensory integration.

Study on the effects of brightness contrast on steady-state motion visual evoked potential.

Traditional steady - state visual evoked potential (SSVEP) using light flicker stimulation may easy cause visual fatigue with a consequent reduction of recognition accuracy. In the previous study, we proposed a steady - state motion visual evoked potential (SSMVEP) brain - computer interface (BCI) method. A black - white ring - shaped checkerboard was used as the visual stimulus paradigm in this study. The movement patterns of the checkerboard included contraction and expansion. Based on the signal-to-noise ratio (SNR), recognition accuracy and anti-fatigue properties, the effects of black-white brightness contrast on the brain response were investigated. Experimental results indicated that when the contrast ratio was the highest, it could obtain higher recognition accuracy and SNR, yet it was easy to cause visual fatigue. When the contrast was the lowest, the sensitivity of the eyes against flicker reduced to the lowest, and it could reduce visual fatigue. However the recognition accuracy and SNR were low. In contrast, the appropriate contrast could guarantee high SNR and recognition accuracy, and as much as possible to reduce the visual fatigue.

Dynamic retinal vessel analysis using a spatial light modulator for stimulation

Purpose : Dynamic vessel analysis (DVA) with the Retinal Vessel Analyzer (Imedos Systems UG) is the Gold standard to measure alterations in retinal vessel diameter in health and disease. The DVA guidelines provide flicker stimulation of 12.5 Hz in a 30° field of view [1] (fixed full-field). However, to investigate local effects in the regulation of retinal perfusion, e.g. neuro vascular coupling, spatially varying stimuli are required. We introduce and validate a spatial light modulator (SLM) based setup for DVA providing arbitrary stimuli in location and size. Methods : We studied 15 volunteers (8m, 7f, 24.7±1.1 yr) to investigate the repeatability of the novel setup and to compare the new setup with the Gold standard. The relative vaso-dilatation value was determined for both. We measured four primary vessels: one superior temporal artery and vein (STa/STv), one inferior temporal artery and vein (ITa/ITv). All subjects were free of ocular and systemic diseases. A high-power LED (515 nm) and a transmissive liquid crystal SLM were connected to a mydriatic fundus camera. The field of view was set to 30°. The SLM was controlled via standard graphics card and a customized software (60 Hz refresh rate / 15 Hz flicker frequency). Each volunteer was measured three times. One measurement with the Gold standard and two with the new setup, in a random order (10 min pause time). Repeatability and comparison of the two setups were analyzed using the Bland-Altmann method. To value the differences among the group means we performed the t-test. For groups that did not meet the normality assumption we employed the Mann-Whitney U test. Results : The repeatability analysis showed the following mean differences (MD), Limits of Agreement (LoA), both in %, and p-values. STa: MD=-0.78, LoA=-4.35/+2.79, p=0.134. ITa: MD=-0.67, LoA=-3.82/+2.48, p=0.131. STv: MD=-0.69, LoA=-4.30/+2.92, p=0.186. ITv: MD=-0.03, LoA=-3.00/+2.94, p=0.947. Comparing the two different setups the p-values were 0.545 (STa), 0.405 (ITa), 0.05 (STv), and 0.137 (ITv). Conclusions : We realized a working experimental setup and applied a new approach of Dynamic Vessel Analysis using a spatial light modulator. Validation was realized in comparison to the Gold standard. The agreement between the two different setups was predominantly good. Neither the repeatability analysis nor the comparison of the two setups showed significant differences.

Effect of Diffuse Luminance Flicker Light Stimulation on Total Retinal Blood Flow Assessed With Dual-Beam Bidirectional Doppler OCT.

We assess the increase in total retinal blood flow (TRBF) induced by flicker stimulation of the human retina in vivo and investigate the flicker induced hyperemia by means of a vascular flow model of the retinal circulation to study neurovascular coupling (NC). In six healthy subjects, TRBF was measured before and during stimulation with diffuse luminance flicker. Blood flow velocities in retinal vessels were measured via dual-beam bidirectional Doppler Fourier-domain optical coherence tomography (FD-OCT), retinal vessel diameters were assessed based on FD-OCT phase data. This allowed for the calculation of TRBF before and during visual stimulation. Additionally, a mathematical flow model for the retinal vasculature was adapted to study the implications of diameter variations on retinal perfusion. Measured and simulated perfusion was compared to draw conclusions on the diameter variations in different layers of the vascular tree. The measured mean baseline flow was 36.4 ± 6.5 μl/min while the mean flow during flicker stimulation was 53.4% ± 8.3 μl/min. The individual increase in TRBF during flicker stimulation ranged between 34% and 66%. The average increase in TRBF over all measured subjects was 47.6% ± 12.6%. Dual-beam bidirectional Doppler FD-OCT allowed quantifying NC in the human retina in vivo and may be a promising method for monitoring alterations in NC caused by various pathologies. The comparison of the measured data with the results obtained in the simulated vasculature indicates that the vasodilation induced by NC is more pronounced in smaller vessels.

Attenuated Fast Steady-State Visual Evoked Potentials During Human Sleep.

During sleep, external sensory events rarely elicit a behavioral response or affect perception. However, how sensory processing differs between wakefulness and sleep remains unclear. A major difficulty in this field stems from using brief auditory stimuli that often trigger nonspecific high-amplitude "K-complex" responses and complicate interpretation. To overcome this challenge, here we delivered periodic visual flicker stimulation across sleep and wakefulness while recording high-density electroencephalography (EEG) in humans. We found that onset responses can be separated from frequency-specific steady-state visual evoked potentials (SSVEPs) selectively observed over visual cortex. Sustained SSVEPs in response to fast (8/10 Hz) stimulation are substantially stronger in wakefulness than in both nonrapid eye movement (NREM) and REM sleep, whereas SSVEP responses to slow (3/5 Hz) stimulation are stronger in both NREM and REM sleep than in wakefulness. Despite wake-like spontaneous activity, responses in REM sleep were similar to those in NREM sleep and different than wakefulness, in accordance with perceptual disconnection during REM sleep. Finally, analysis of amplitude and phase in single trials revealed that stronger fast SSVEPs in wakefulness are driven by more consistent phase locking and increased induced power. These results suggest that the sleeping brain is unable to effectively synchronize large neuronal populations in response to rapid sensory stimulation.

Efficacy and Toxicity of Intravitreous Chemotherapy for Retinoblastoma: Four-Year Experience

To investigate the efficacy and toxicity of intravitreous melphalan for treatment of retinoblastoma, as a single agent or with concomitant topotecan. A total of 130 eyes of 120 patients with retinoblastoma receiving 630 intravitreous (melphalan, topotecan) or topotecan periocular injections. A total of 83 (64%) of these eyes were treated with concomitant ophthalmic artery chemosurgery (OAC). Retrospective cohort study. Indirect ophthalmoscopy and clinical imaging were used to evaluate clinical response. Ocular survival and disease-free survival were estimated using Kaplan-Meier methods in 130 eyes. Ocular toxicity was evaluated by clinical findings and electroretinography (ERG) on 244 evaluable injections in 63 patients using 30-Hz flicker responses. Analysis was performed using linear mixed effects models with a random intercept and slope for each patient and a fixed effect for number of injections, in addition to any other fixed effect of interest. Ocular survival, disease-free survival, ERG: peak-to-peak ERG amplitudes in response to 30-Hz photopic flicker stimulation. There were no disease- or treatment-related deaths, and no patient developed externalization of tumor or metastatic disease. Two-year Kaplan-Meier estimates of ocular survival and disease-free survival were 94.2% (95% confidence interval, 89.2-99.4) and 86.2% (95% confidence interval, 78.7-94.5), respectively. There was a significant association between the number of injections and diminished ERG responses, such thaton average each intravitreous melphalan injection was associated with a 5.3-μV decrease in ERG amplitude (P < 0.001). Concomitant intra-arterial chemotherapy (P= 0.01) and greater inherent ocular pigment also were significantly associated with a reduction in ERG (P= 0.045). Patient age and weight, new injection site location, addition of topotecan, concomitant focal treatment, and time interval between injections were not significantly associated with toxicity. Intravitreous melphalan is an effective treatment for vitreous seeding in retinoblastoma, resulting in high rates of ocular survival and disease-free survival. However, in this study, each injection of melphalan was associated, on average, with a decrement in ERG response. The findings suggest increased toxicity (1) when OAC is given within 1 week of the intravitreous injection and (2) in more deeply pigmented eyes.

Steady-State Motion Visual Evoked Potential (SSMVEP) Based on Equal Luminance Colored Enhancement.

Steady-state visual evoked potential (SSVEP) is one of the typical stimulation paradigms of brain-computer interface (BCI). It has become a research approach to improve the performance of human-computer interaction, because of its advantages including multiple objectives, less recording electrodes for electroencephalogram (EEG) signals, and strong anti-interference capacity. Traditional SSVEP using light flicker stimulation may cause visual fatigue with a consequent reduction of recognition accuracy. To avoid the negative impacts on the brain response caused by prolonged strong visual stimulation for SSVEP, steady-state motion visual evoked potential (SSMVEP) stimulation method was used in this study by an equal-luminance colored ring-shaped checkerboard paradigm. The movement patterns of the checkerboard included contraction and expansion, which produced less discomfort to subjects. Feature recognition algorithms based on power spectrum density (PSD) peak was used to identify the peak frequency on PSD in response to visual stimuli. Results demonstrated that the equal-luminance red-green stimulating paradigm within the low frequency spectrum (lower than 15 Hz) produced higher power of SSMVEP and recognition accuracy than black-white stimulating paradigm. PSD-based SSMVEP recognition accuracy was 88.15±6.56%. There was no statistical difference between canonical correlation analysis (CCA) (86.57±5.37%) and PSD on recognition accuracy. This study demonstrated that equal-luminance colored ring-shaped checkerboard visual stimulation evoked SSMVEP with better SNR on low frequency spectrum of power density and improved the interactive performance of BCI.

Nonlinear Origin of SSVEP Spectra-A Combined Experimental and Modeling Study.

Steady state visual evoked potentials (SSVEPs) are steady state oscillatory potentials elicited in the electroencephalogram (EEG) by flicker stimulation. The frequency of these responses maches the frequency of the stimulation and of its harmonics and subharmonics. In this study, we investigated the origin of the harmonic and subharmonic components of SSVEPs, which are not well understood. We applied both sine and square wave visual stimulation at 5 and 15 Hz to human subjects and analyzed the properties of the fundamental responses and harmonically related components. In order to interpret the results, we used the well-established neural mass model that consists of interacting populations of excitatory and inhibitory cortical neurons. In our study, this model provided a simple explanation for the origin of SSVEP spectra, and showed that their harmonic and subharmonic components are a natural consequence of the nonlinear properties of neuronal populations and the resonant properties of the modeled network. The model also predicted multiples of subharmonic responses, which were subsequently confirmed using experimental data.

Theoretical predictions of metabolic flow regulation in the retina

Purpose: This study uses a theoretical model to investigate the response of retinal blood flow to changes in tissue oxygen demand. The study is motivated by the need for a better understanding of metabolic flow regulation mechanisms in health and disease.Methods: A mathematical model is used to calculate retinal blood flow for different levels of tissue oxygen demand in the presence or absence of regulatory mechanisms. The model combines a compartmental view of the retinal vasculature and a Krogh cylinder description for oxygen delivery to retinal tissue.Results: The model predicts asymmetric behavior in response to changes in tissue oxygen demand. When all regulatory mechanisms are active, the model predicts a 6% decrease in perfusion when tissue oxygen demand is decreased by 50% and a 23% increase in perfusion when tissue oxygen demand is increased by 50%. In the absence of metabolic and carbon dioxide responses, the model predicts a constant level of blood flow that does not respond to changes in oxygen demand, suggesting the importance of these two response mechanisms. The model is not able to replicate the increase in oxygen venous saturation that has been observed in some flicker stimulation studies.Conclusions: The increase in blood flow predicted by the model due to an increase in oxygen demand is not in the same proportion as the change in blood flow observed with the same decrease in oxygen demand, suggesting that vascular regulatory mechanisms may respond differently to different levels of oxygen demand. These results might be useful for interpreting clinical and experimental findings in health and disease.

DYNAMIC AND STATIC VESSEL ANALYSIS IN PATIENTS WITH RETINITIS PIGMENTOSA: A Pilot Study of Vascular Diameters and Functionality.

To evaluate in vivo the vascular anatomy and functionality of early manifestation retinitis pigmentosa (RP) by means of a dynamic and static vessel analyzer. Fourteen patients with early RP and 14 normal subjects were consecutively enrolled in this observational, prospective study. Each patient underwent a complete ophthalmologic examination, including dynamic and static retinal vessel analysis using the Dynamic Vessel Analyzer. The patients with RP and the control group were well matched in age and sex. Patients with RP had a mean best-corrected visual acuity of 20/25 (range: 20/40-20/20). Dynamic vessel analysis performed in patients with RP showed an arterial and venous dilation during flicker stimulation of 5.28 ± 1.7% and 4.07 ± 1.78%, respectively. Only arterial dilation was statistically different compared with control subjects (3.33 ± 0.99%, P = 0.0062). Static retinal vessel analysis in patients with RP showed a decreased mean central retinal artery equivalent (P < 0.001) and central retinal vein equivalent (P < 0.001) compared with control subjects. By contrast, the arterial-to-venous ratio was similar in both groups (RP: 0.79 ± 0.11, control group 0.86 ± 0.04, P = 0.072). Our data confirm that retinal arterial and venous narrowing is present at an early stage in patients affected by RP. However, dynamic vessel analysis shows how the retina of patients with RP with no best-corrected visual acuity loss presents an augmented artery dilation response compared with normal subjects and retained neurovascular coupling.

Higher glucose level and systemic oxidative stress decrease the mean velocity index of the retinal artery during flickering light stimulation in type 1 diabetes.

AimTo determine whether higher glucose level and systemic oxidative stress decrease mean velocity (MV) index of the central retinal artery (CRA) during flickering light stimulation in type 1 diabetes (T1D).MethodsThe study was performed in the period from 2008 to 2015 at the University Eye Clinic in Ljubljana. 41 patients with T1D and 37 participants without diabetes were included. MV in the CRA was measured using Doppler ultrasound diagnostics in basal conditions and during 8 Hz flickering light irritation. The plasma levels of glucose, fructosamine, 8-hydroxy-2'-deoxyguanosine (8-OHdG), triglycerides, cholesterol, and low-density lipoprotein (LDL) were measured.ResultsPatients with T1D had significantly higher levels of blood glucose (P < 0.001), fructosamine (P < 0.001), and 8-OHdG (P < 0.001), but there were no significant differences in triglycerides (P = 0.108), cholesterol (P = 0.531), and LDL (P = 0.645) between the groups. Patients with T1D also had a significantly lower MV index in the CRA (1.11 ± 0.15 vs 1.24 ± 0.23; P = 0.010). In the T1D group, a significant negative correlation was found between the level of glucose (r = −0.58; P < 0.001), fructosamine (r = −0.46; P = 0.003), 8-OHdG (r = −0.48; P = 0.002) and the MV index in the CRA. At the same time, in this group fructosamine and 8-OHdG levels had a separate effect on the MV index (adjusted R2 = 0.38, P < 0.001).ConclusionHigher glucose levels, the medium-term glucose level, and systemic oxidative stress could importantly reduce retinal vasodilatation during flickering light irritation in patients with T1D.

A multi-command SSVEP-based BCI system based on single flickering frequency half-field steady-state visual stimulation

Steady-state visual evoked potentials (SSVEPs) are widely employed in brain-computer interface (BCI) applications, especially to control machines. However, the use of SSVEPs leads to eye fatigue and causes lower accuracy over the long term, particularly when multi-commands are required. Therefore, this paper proposes a half-field steady-state visual stimulation pattern and paradigm to increase the limited number of commands that can be achieved with existing SSVEP-based BCI methods. Following the theory of vision perception and existing half-field SSVEP-based BCI systems, the new stimulation pattern generates four commands using only one frequency flickering stimulus and has an average classification accuracy of approximately 75%. According to the proposed stimulus pattern, using only one frequency without requiring users to stare directly at the flickering stimulus allows users to easily focus on the system and experience less visual fatigue compared to existing systems. Furthermore, new half-field SSVEP-based BCI systems are proposed, incorporating our proposed feature extraction and decision-making algorithm. Extracting the signal from the occipital area and using a reference electrode position at the parietal area yielded better results compared to the central area. In addition, we recommend using an LED or LCD as the visual stimulus device (at the recommended size), which yielded comparable results to our proposed feature extraction and decision-making algorithm. Finally, an application of the proposed system is demonstrated for real-time television control.

Role of glial cells in regulating retinal blood flow during flicker‐induced hyperemia and systemic hyperoxia‐induced hyporemia in cats

SummaryBackgroundTo investigate the role of glial cells in the regulation of retinal circulation in response to flicker stimulation, which is related to neurovascular coupling, and systemic hyperoxia in the retina.MethodsUsing laser Doppler velocimetry, we measured the retinal blood flow (RBF) in first‐order retinal arterioles in anesthetized cats. After intravitreal injection of l‐2‐aminoadipic acid (LAA) as gliotoxic compound, we examined the changes in RBF in response to flicker stimulation of 16 Hz for 3 min and systemic hyperoxia induced by the inhalation of 100% oxygen for 10 min.ResultsThe RBF increased gradually and reached a maximal level after 2–3 min of flicker stimuli in the PBS group. In the LAA‐treated eyes, the increases in RBF during the flicker stimulation were significantly attenuated by one third of those in the control eyes. During hyperoxia, the decreases in RBF in response to hyperoxia were significantly attenuated in the LAA‐treated eyes compared with the control.ConclusionsThe current results suggest that the retinal glial cells play an important role in the regulation of retinal blood flow during Flicker‐Induced Hyperemia and Systemic Hyperoxia‐Induced Hyporemia.

Impact of combined hormonal contraceptives on vessels functionality.

To evaluate the dynamic and static retinal vascular functionality in young females using combined hormonal contraceptive (CHC). Thirty-eight consecutive young female subjects were enrolled in this study between January 2015 and December 2015. Subjects were divided in two groups: CHC group, defined as CHC use for ≥6months, and control group, defined as no current and prior CHC use. Participants underwent a dynamic and static retinal vessel analysis using the Dynamic Vessel Analyzer (DVA, Imedos, Jena, Germany). Seventeen subjects continuously took CHC for 54.6±29.3months, while 21 subjects belonged to control group. No difference was found between the CHC and control groups for age (p=0.1), smoking status (p=0.6), and systolic (p=0.3) and diastolic (p=0.1) blood pressure. With regard to dynamic analysis, women taking CHC exhibited a marked significant vasoconstriction following flicker stimulation in comparison with control group (-2.43±2.5 vs 0.63±2.1, respectively; p=0.0002). No significant difference was observed between groups for mean arterial (p=0.2) and venous dilatations (p=0.3), arteriovenous ratio (p=0.09), central retinal artery equivalent (p=0.4), and central retinal venous equivalent (p=0.5). CHC may affect vessel reactivity to flicker light by increasing arteries constriction. This may reflect systemic changes in vascular functionality in subjects using CHC. Moreover, CHC should be considered as a confounding bias in studies involving DVA.

Optical coherence tomography angiography of stimulus evoked hemodynamic responses in individual retinal layers.

Blood flow changes are highly related to neural activities in the retina. It has been reported that neural activity increases when flickering light stimulation of the retina is used. It is known that blood flow changes with flickering light stimulation can be altered in patients with vascular disease and that measurement of flicker-induced vasodilatation is an easily applied tool for monitoring functional microvascular alterations. However, details of distortions in retinal neurovascular coupling associated with major eye diseases are not well understood due to the limitation of existing techniques. In this study, flickering light stimulation was applied to mouse retinas to investigate stimulus evoked hemodynamic responses in individual retinal layers. A spectral domain optical coherence tomography (OCT) angiography imaging system was developed to provide dynamic mapping of hemodynamic responses in the ganglion cell layer, inner plexiform layer, outer plexiform layer and choroid layer before, during and after flickering light stimulation. Experimental results showed hemodynamic responses with different magnitudes and time courses in individual retinal layers. We anticipate that the dynamic OCT angiography of stimulus evoked hemodynamic responses can greatly foster the study of neurovascular coupling mechanisms in the retina, promising new biomarkers for retinal disease detection and diagnosis.

Factors Determining Flicker-Induced Retinal Vasodilation in Healthy Subjects.

The purpose of this study was to analyze factors determining retinal arterial and venous responses to stimulation with diffuse luminance flicker in healthy subjects. We retrospectively analyzed results obtained in 374 healthy subjects who had previously participated in clinical studies in our department. A total of 153 subjects underwent a protocol in which flicker stimulation was delivered through the fundus camera at 8 Hz (protocol 1), separating measurement and stimulation light depending on the wavelength, and 221 subjects underwent a protocol in which diffuse luminance flicker was delivered at 12.5 Hz with high modulation depth (protocol 2). We investigated whether sex, systemic blood pressure, baseline vessel size, blood plasma concentration of fasting glucose and hematocrit, and serum concentration of cholesterol, triglycerides, creatinine and C-reactive protein influenced the retinal vascular response to flicker stimulation. Flicker responses in arteries and veins were more pronounced in protocol 2 than in protocol 1 (P < 0.001, each). In both of the protocols the vascular response to stimulation with diffuse luminance flicker was larger in smaller vessels (P between 0.001 and 0.016). In protocol 2 the retinal arterial flicker response was negatively associated with cholesterol serum levels (P = 0.033); in protocol 1, only a tendency toward this effect was observed (P = 0.056). The present analysis indicates that retinal arterial and venous responses to stimulation with diffuse luminance flicker depend on the way the stimulation is delivered through the fundus camera. In addition, the flicker response varied with vessel size, that is, the smaller the vessel width, the larger the flicker response. Finally, our data indicate that, even within the normal range, higher cholesterol serum levels are associated with lower hyperemic flicker responses.

Preserved Pressure Autoregulation but Disturbed Cyclo-Oxygenase and Nitric Oxide Effects on Retinal Arterioles during Acute Hypoxia in Diabetic Patients without Retinopathy

Background: Acute hypoxia induces retinal vasodilatation, which depends on cyclooxygenase (COX) products and nitric oxide (NO) in vitro. However, it is unknown whether these mechanisms are active in diabetic patients and may contribute to the development of diabetic retinopathy. Methods: The Dynamic Vessel Analyzer was used to study the diameter regulation in retinal vessels during hypoxia in type 1 diabetic patients without retinopathy. The influence of NO and COX synthesis inhibition on the diameter of larger retinal vessels was studied during hypoxia, during isometric exercise and during flicker stimulation. Results: Increased arterial blood pressure during <smlcap>L</smlcap>-NMMA infusion and isometric exercise were paralleled by constriction of the retinal arterioles suggesting normal pressure autoregulation. Hypoxia significantly reduced the diameter responses during isometric exercise and during flicker stimulation as compared to normal persons. Conclusion: The findings support that changes in metabolic autoregulation develop before changes in pressure autoregulation in diabetic patients.

Wavelength-sensitivity of mouse retinal ganglion cells recorded by a high-density micro electrode array (MEA)

Event Abstract Back to Event Wavelength-sensitivity of mouse retinal ganglion cells recorded by a high-density micro electrode array (MEA) Florian Jetter1, Gabriel Bertotti2, Roland Thewes3 and Günther Zeck4* 1 NMI at the University Tuebingen, Neurochip Research, Germany 2 Technische Universität Berlin, Chair of Sensor and Actuator Systems, Germany 3 Technische Universität Berlin, Chair of Sensor and Actuator Systems, Germany 4 NMI at the University Tuebingen, Neurochip Research, Germany Motivation: High-density CMOS-MEAs can be used to simultaneously record the electrical spiking activity from hundreds of neurons [Bertotti et al., 2014]. Neuronal spiking can be induced by electrical stimulation [Eickenscheidt and Zeck 2014], by light stimulation applied to a light-sensitive retina [Zeck et al. 2011], or by light stimulation of optogenetically transfected neurons [Herrmann et al. 2014]. In this work, we investigate the wavelength-sensitivity of different mouse retinal ganglion cells and the intrinsic wavelength-sensitivity of the response of high-density CMOS-MEAs. Material and Methods: A CMOS-based high-density MEA comprising 4225 recording sites is used for recording the ganglion cell activity in C57/Bl6 mouse retina during flickering light stimulation (1 and 5 Hz, respectively) with different wavelength and of different light stimulus sizes. Light stimuli presented on a DMD (µ-matrix, Rapp Optoelectronic, Germany) are focused through a microscope objective on the retina. The DMD is illuminated by an LED system commonly used for optogenetic activation (pe-4000, coolLED, UK). Here the results for four stimulation wavelengths are presented (405, 470, 525, and 635nm) at intensities as high as 2 mW/mm² (470 nm). Recorded data is sampled at 25 kHz. Results: Light stimulation (stimulus area: 1 mm2) evokes spiking in the interfaced retina. Based on the stimulus polarity retinal ganglion cells are broadly classified in ON or OFF type, depending on whether they respond to light on- or offset, respectively. In Fig. 1A filtered recordings are shown of the measured extracellular voltage from two selected sensors during 1 Hz stimulation. Both, ON and OFF cell types respond to the three highest wavelengths used here. The sensor site recording the ON transient cell also detects activity from a second OFF cell with smaller amplitude. In Fig.1B we present the unfiltered extracellular voltage traces recorded by a third selected sensor site for all four wavelengths. Light onset leads to a measurable change of the current in the sensing transistor (cf. Bertotti et al., 2014) or more generally speaking of the sensed recording site response, reflected as a low-frequency change of signal back-converted into the voltage domain. However, this does not prevent the detection of light-induced spiking. We note, that the ganglion cell shown in Fig. 1B is not activated by the 635 nm light stimulus. Ongoing experiments investigate the sensitivity of retinal ganglion cells to chromatic stimulation and to stimuli presented at various intensities. The induced variations of the sensor signals are wavelength-dependent, with the highest change obtained for the lowest wavelength (405 nm) and undetectable changes for red light (635 nm). Conclusion: It is shown that recording of cellular spiking activity with CMOS-based MEAs is possible during optical stimulation. Ganglion cells in the mouse retina have different wavelength-sensitivities. The light-induced low-frequency drift of the sensor signals can be completely removed for the tested wavelengths and intensities using high-pass filtering.

Relation of retinal blood flow and retinal oxygen extraction during stimulation with diffuse luminance flicker.

Cerebral and retinal blood flow are dependent on local neuronal activity. Several studies quantified the increase in cerebral blood flow and oxygen consumption during activity. In the present study we investigated the relation between changes in retinal blood flow and oxygen extraction during stimulation with diffuse luminance flicker and the influence of breathing gas mixtures with different fractions of O2 (FiO2; 100% 15% and 12%). Twenty-four healthy subjects were included. Retinal blood flow was studied by combining measurement of vessel diameters using the Dynamic Vessel Analyser with measurements of blood velocity using laser Doppler velocimetry. Oxygen saturation was measured using spectroscopic reflectometry and oxygen extraction was calculated. Flicker stimulation increased retinal blood flow (57.7 ± 17.8%) and oxygen extraction (34.6 ± 24.1%; p < 0.001 each). During 100% oxygen breathing the response of retinal blood flow and oxygen extraction was increased (p < 0.01 each). By contrast, breathing gas mixtures with 12% and 15% FiO2 did not alter flicker–induced retinal haemodynamic changes. The present study indicates that at a comparable increase in blood flow the increase in oxygen extraction in the retina is larger than in the brain. During systemic hyperoxia the blood flow and oxygen extraction responses to neural stimulation are augmented. The underlying mechanism is unknown.

Role of Glial Cells in Regulating Retinal Blood Flow During Flicker-Induced Hyperemia in Cats

To investigate how glial cells participate in retinal circulation during flicker stimulation in cats. Using laser Doppler velocimetry, we measured the vessel diameter and blood velocity simultaneously and calculated the retinal blood flow (RBF) in feline first-order retinal arterioles. Twenty-four hours after intravitreal injections of L-2-aminoadipic acid (LAA), a gliotoxic compound, and the solvent of 0.01 N hydrochloric acid as a control, we examined the changes in RBF in response to 16-Hz flicker stimulation for 3 minutes. We also measured the changes in RBF 2 hours after intravitreal injection of Nω-propyl-L-arginine (L-NPA), a selective neuronal nitric oxide synthase inhibitor, in LAA-treated eyes. To evaluate the effects of LAA on retinal neuronal function, ERGs were monitored. Immunohistochemical examinations were performed. In LAA-treated eyes, histologic changes selectively occurred in retinal glial cells. There were no significant reductions in amplitude or elongation of implicit time in ERG after LAA injections compared with controls. In control eyes, the RBF gradually increased and reached the maximal level (53.5% ± 2.5% increase from baseline) after 2 to 3 minutes of flicker stimulation. In LAA-treated eyes, the increases in RBF during flicker stimulation were attenuated significantly compared with controls. In LAA-treated eyes 2 hours after injection of L-NPA, flicker-evoked increases in RBF decreased significantly compared with LAA-treated eyes. The current results suggested that increases in RBF in response to flicker stimulation were regulated partly by retinal glial cells.