Remote photonic sensing of action potential in mammalian nerve cells via histogram-based analysis of temporal spatial acoustic vibrations.

Label free and remote action potential detection in neurons can be of great importance in the neuroscience research field. This paper presents a novel label free imaging modality based on the detection of temporal vibrations of speckle patterns illuminating the sample. We demonstrated the feasibility of detecting action potentials originating from spontaneous and stimulated activity in cortical cell culture. The spatiotemporal vibrations of isolated cortical cells were extracted by illuminating the culture with a laser beam while the vibrations of the random back scattered secondary speckle patterns are captured by a camera. The postulated action potentials were estimated following correlation-based analysis on the captured vibrations, where the variance deviation of the signal from a Gaussian distribution is directly associated with the action potential events. The technique was validated in a series of experiments in which the optical signals were acquired concurrently with microelectrode array (MEA) recordings. Our results demonstrate the ability of detecting action potential events in mammalian cells remotely via extraction of acoustic vibrations.

A dichoptic presentation device and a method for measuring binocular temporal function in the visual system.

Recent studies highlight the importance of the temporal domain in visual processing. Critical Flicker Frequency (CFF), the frequency at which a flickering light is perceived as continuous, is a widely used measure for evaluating visual temporal processing. Another important issue to investigate is the cortical interactions arising between the flicker stimuli of both eyes. This paper presents a robust and reliable dichoptic tool for evaluating the CFF threshold in both eyes. This system is based on an analog output device used to independently drive two LEDs through a custom-written MATLAB code (using a laptop PC) for eliciting sinusoidal flickering stimuli and for psychophysically measuring the perceived CFF threshold. The luminance and phases of each LED are individually controlled, enabling the investigation of the effect of phase and luminance differences on binocular summation in subjects with different ocular pathologies. Experiments were designed to evaluate the CFF threshold through a psychophysical test, based on a discrimination task with a stimulus duration of 1 s, based on a temporal alternative forced-choice paradigm. The target stimulus temporal features were modulated using the staircase method. Subjects were requested to discriminate between a target stimulus (a flickering light at various frequencies) and a flickering light at a frequency of 120 Hz, which is significantly higher than the CFF in humans; therefore, it is perceived as constant. One of the main advantages of the introduced dichoptic presentation system is that it enables the visual temporal performance to be measured under both monocular and binocular conditions where phenomena such as temporal binocular summation (BS) can be evaluated. Moreover, the system offers great flexibility by introducing a stimulus phase shift, which enables studying how stimulus timing affects the temporal function at millisecond scale resolution. Our results confirm that no crosstalk exists between the eyes and that the system can reliably separate the stimuli presented to the eyes. Using this set-up, we observed the binocular summation of CFF for low target luminance levels. The CFF was significantly (p = 0.011) higher (5.2%) under binocular compared with monocular viewing conditions. More importantly, introducing an inter - ocular phase shift reduced the binocular CFF in normally sighted subjects. Finally, in amblyopic subjects the amblyopic eye showed a decrease of 3.9 Hz (15%) in CFF, compared with the fellow eye (p = 0.001). The ability to assess binocular temporal performance using a dichoptic display can shed light on visual temporal performance in general, and on binocular temporal summation processes in particular, both for subjects with normal binocular vision and for subjects with impaired binocular vision (e.g., amblyopic subjects). Furthermore, such a presentation set-up may facilitate the development of training paradigms aimed at improving binocular vision performance. In this paper we describe the system and methods in detail and provide all necessary computer code and other details that will enable an easy and quick adaptation of the method by scientists interested in studying the temporal resolution of the visual system in general, and in studying inter-ocular differences or interactions in particular.

Anti-VEGF-Aptamer Modified C-Dots-A Hybrid Nanocomposite for Topical Treatment of Ocular Vascular Disorders.

The vascular endothelial growth factor (VEGF) induces pathological angiogenetic ocular diseases. It is a scientific challenge to develop carriers for the controlled release of inhibitors for VEGF present in the back of the eye domain. Carbon dots (C-dots) functionalized with the VEGF aptamer are introduced and the hybrid nanoparticles are used for ocular nanomedicine. The C-dots are applied as effective carriers of the anti-VEGF aptamer across the cornea, yielding therapeutic levels upon topical administration. The hybrids show no toxicity for both in vitro and in vivo murine animal model, and further enable noninvasive intraocular concentration monitoring through the C-dots inherent fluorescence. In addition, the hybrid C-dots effectively inhibit VEGF-stimulated angiogenesis in choroidal blood vessels. This inhibition is comparable to two commercially available anti-VEGF drugs, bevacizumab and aflibercept. The hybrid aptamer-modified C-dots provide a versatile nanomaterial to treat age-related macular degeneration and diabetic retinopathy.

Interfacing the Cell with "Biomimetic Membrane Proteins".

Integral membrane proteins mediate a myriad of cellular processes and are the target of many therapeutic drugs. Enhancement and extension of the functional scope of membrane proteins can be realized by membrane incorporation of engineered nanoparticles designed for specific diagnostic and therapeutic applications. In contrast to hydrophobic insertion of small amphiphilic molecules, delivery and membrane incorporation of particles on the nanometric scale poses a crucial barrier for technological development. In this perspective, the transformative potential of biomimetic membrane proteins (BMPs), current state of the art, and the barriers that need to be overcome in order to advance the field are discussed.

An optimized protocol for generating labeled and transplantable photoreceptor precursors from human embryonic stem cells.

Cell replacement therapy is a promising approach for treatment of retinal degenerative diseases. Several protocols for the generation of photoreceptor precursors (PRP) from human embryonic stem cells (hESC) have been reported with variable efficiency. Herein, we show the advantages of use of size-controlled embryoid bodies in the ESC differentiation process using two differentiation protocols. We further explored cell-labeling methods for following the survival of PRP transplanted subretinally in rat eyes. Size-controlled embryoid bodies (EBs) generated using microwell dishes and non-size-controlled EBs generated using V-shaped 96-well plates were differentiated into PRP using two differentiation protocols. The differentiation protocols utilized two different combinations of growth factors. The first, Dkk1, Noggin, and IGF1, and the second protocol used IWR1e, SAG, and CHIR99021. Differentiation efficiency to PRP was analyzed by qPCR, immunocytochemistry, and fluorescence-assisted cell sorting (FACS). Size-controlled IWR1e yielded a significantly higher percent (86.4%) of PRP cells expressing CRX, compared with non-size-controlled IWR1e (51.4%, P = 0.026) or the size-controlled DKK1 protocol (70.5%, p = 0.007). In addition, the IWR1e differentiated cells exhibited a significantly higher fluorescence intensity of CRX immunostaining, compared with the DKK1 protocol, consistent with higher protein expression levels. The IWR1e cells exhibited higher maturation levels, as manifested by lower early neuronal marker PAX6 and pluripotency marker OCT4 levels compared with the DKK1 protocol. The expression of other late photoreceptor markers (NRL, recoverin) were similar among the differentiation groups. PRP cells were labeled by using hESC constitutively expressing EGFP or by AAV-GFP transduction. Finally, we transplanted the cells in the subretinal space of wild-type rats and monitored their survival over several weeks. The AAV2 serotype efficiently transduced the PRP cells, whereas other serotypes yielded low or no transduction. Following subretinal transplantation of GFP-labeled PRP, 63% of the cells were detected at 4 weeks post-transplantation. In conclusion, we show here that the IWR1e protocol using size-controlled EBs efficiently generated of PRP that could be labeled and followed in-vivo for weeks. The data from this study is an advance toward the goal of PRP transplantation therapy for retinal degenerative diseases.

Mechanisms of electrical vasoconstriction.

BACKGROUND: Electrical vasoconstriction is a promising approach to control blood pressure or restrict bleeding in non-compressible wounds. We explore the neural and non-neural pathways of electrical vasoconstriction in-vivo. METHODS: Charge-balanced, asymmetric pulses were delivered through a pair of metal disc electrodes. Vasoconstriction was assessed by measuring the diameter of rat saphenous vessels stimulated with low-voltage (20 V, 1 ms) and high-voltage (150 V, 10 µs) stimuli at 10 Hz for 5 min. Activation pathways were explored by topical application of a specific neural agonist (phenylephrine, alpha-1 receptor), a non-specific agonist (KCl) and neural inhibitors (phenoxybenzamine, 25 mg/ml; guanethidine, 1 mg/ml). Acute tissue damage was assessed with a membrane permeability (live-dead) fluorescent assay. The Joule heating in tissue was estimated using COMSOL Multiphysics modeling. RESULTS: During stimulation, arteries constricted to 41 ± 8% and 37 ± 6% of their pre-stimulus diameter with low- and high-voltage stimuli, while veins constricted to 80 ± 18% and 40 ± 11%, respectively. In arteries, despite similar extent of constriction, the recovery time was very different: about 30 s for low-voltage and 10 min for high-voltage stimuli. Neural inhibitors significantly reduced low-voltage arterial constriction, but did not affect high-voltage arterial or venous constriction, indicating that high-voltage stimuli activate non-neural vasoconstriction pathways. Adrenergic pathways predominantly controlled low-voltage arterial but not venous constriction, which may involve a purinergic pathway. Viability staining confirmed that stimuli were below the electroporation threshold. Modeling indicates that heating of the blood vessels during stimulation (< 0.2 °C) is too low to cause vasoconstriction. CONCLUSIONS: We demonstrate that low-voltage stimuli induce reversible vasoconstriction through neural pathways, while high-voltage stimuli activate non-neural pathways, likely in addition to neural stimulation. Different stimuli providing precise control over the extent of arterial and venous constriction as well as relaxation rate could be used to control bleeding, perfusion or blood pressure.

Subvisible retinal laser therapy: titration algorithm and tissue response.

PURPOSE: Laser therapy for diabetic macular edema and other retinal diseases has been used within a wide range of laser settings: from intense burns to nondamaging exposures. However, there has been no algorithm for laser dosimetry that could determine laser parameters yielding a predictable extent of tissue damage. This multimodal imaging and structural correlation study aimed to verify and calibrate a computational model-based titration algorithm for predictable laser dosimetry ranging from nondamaging to intense coagulative tissue effects. METHODS: Endpoint Management, an algorithm based on a computational model of retinal photothermal damage, was used to set laser parameters for various levels of tissue effect. The algorithm adjusts both power and pulse duration to vary the expected level of thermal damage at different percentages of a reference titration energy dose. Experimental verification was conducted in Dutch Belted rabbits using a PASCAL Streamline 577 laser system. Titration was performed by adjusting laser power to produce a barely visible lesion at 20 ms pulse duration, which is defined as the nominal (100%) energy level. Tissue effects were then determined for energy levels of 170, 120, 100, 75, 50, and 30% of the nominal energy at 1 hour and 3, 7, 30, and 60 days after treatment. In vivo imaging included fundus autofluorescence, fluorescein angiography, and spectral-domain optical coherence tomography. Morphologic changes in tissue were analyzed using light microscopy, as well as scanning and transmission electron microscopy. RESULTS: One hundred and seventy percent and 120% levels corresponded to moderate and light burns, respectively, with damage to retinal pigment epithelium, photoreceptors, and at highest settings, to the inner retina. 50% to 75% lesions were typically subvisible ophthalmoscopically but detectable with fluorescein angiography and optical coherence tomography. Histology in these lesions demonstrated some selective damage to retinal pigment epithelium and photoreceptors. The 30% to 50% lesions were invisible with in vivo multimodal imaging, and damage was limited primarily to retinal pigment epithelium, visible best with scanning electron microscopy. Over time, photoreceptors shifted into the coagulated zone, reestablishing normal retinal anatomy in lesions ≤100%, as seen in optical coherence tomography and light microscopy. Transmission electron microscopy at 2 months demonstrated restoration of synapses between shifted-in photoreceptors and bipolar cells in these lesions. Retinal pigment epithelium monolayer restored its continuity after 1 week in all lesions. No damage could be seen <30% level. CONCLUSION: A retinal laser dosimetry protocol based on the Endpoint Management algorithm provides reproducible changes in retinal morphology in animals with various levels of pigmentation. This algorithm opens doors to clinical trials of well-defined subvisible and nondestructive regimes of retinal therapy, especially important for treatment of macular disorders.

A novel GCaMP6f-RCS rat model for studying electrical stimulation in the degenerated retina.

Background: Retinal prostheses aim to restore vision by electrically stimulating the remaining viable retinal cells in Retinal Degeneration (RD) cases. Research in this field necessitates a comprehensive analysis of retinal ganglion cells' (RGCs) responses to assess the obtained visual acuity and quality. Here we present a novel animal model which facilitates the optical recording of RGCs activity in an RD rat. This model can significantly enhance the functional evaluation of vision restoration treatments. Methods: The development of the novel rat model is based on crossbreeding a retinal degenerated Royal College of Surgeons (RCS) rat with a transgenic line expressing the genetic calcium indicator GCaMP6f in the RGCs. Characterization of the model was achieved using Optical Coherence Tomography (OCT) imaging, histology, and electroretinography (ERG) at the ages of 4, 8, and 12 weeks. Additionally, optical recordings of RGCs function in response to ex-vivo subretinal electrical stimulations were performed. Results: Histological investigations confirmed the high expression of GCaMP6f in the RGCs and minimal expression in the inner nuclear layer (INL). OCT imaging and histological studies revealed the expected gradual retinal degeneration, as evident by the decrease in retinal thickness with age and the formation of subretinal debris. This degeneration was further confirmed by ERG recordings, which demonstrated a significant decrease in the b-wave amplitude throughout the degeneration process, culminating in its absence at 12 weeks in the GCaMP6f-RCS rat. Importantly, the feasibility of investigating subretinal stimulation was demonstrated, revealing a consistent increase in activation threshold throughout degeneration. Furthermore, an increase in the diameter of the activated area with increasing currents was observed. The spatial spread of the activation area in the GCaMP6f-RCS rat was found to be smaller and exhibited faster activation dynamics compared with the GCaMP6f-LE strain. Conclusion: This novel animal model offers an opportunity to deepen our understanding of prosthetically induced retinal responses, potentially leading to significant advancements in prosthetic interventions in visual impairments.

Perceptual learning based on a temporal stimulus enhances visual function in adult amblyopic subjects.

Studies have shown that Perceptual Learning (PL) can lead to enhancement of spatial visual functions in amblyopic subjects. Here we aimed to determine whether a simple flickering stimulus can be utilized in PL to enhance temporal function performance and whether enhancement will transfer to spatial functions in amblyopic subjects. Six adult amblyopic and six normally sighted subjects underwent an evaluation of their performance of baseline psychophysics spatial functions (Visual acuity (VA), contrast sensitivity (CS), temporal functions (critical fusion frequency (CFF) test), as well as a static and flickering stereopsis test, and an electrophysiological evaluation (VEP). The subjects then underwent 5 training sessions (on average, a total of 150 min over 2.5 weeks), which included a task similar to the CFF test using the method of constant stimuli. After completing the training sessions, subjects repeated the initial performance evaluation tasks. All amblyopic subjects showed improved temporal visual performance (CFF) in the amblyopic eye (on average, 17%, p << 0.01) following temporal PL. Generalization to spatial, spatio-temporal, and binocular tasks was also found: VA increased by 0.12 logMAR (p = 0.004), CS in backward masking significantly increased (by up to 19%, p = 0.003), and flickering stereopsis increased by 85 arcsec (p = 0.048). These results were further electrophysiologically manifested by an increase in VEP amplitude (by 43%, p = 0.03), increased Signal-to-Noise ratio (SNR) (by 39%, p = 0.024) to levels not different from normally sighted subjects, along with an improvement in inter-ocular delay (by 5.8 ms, p = 0.003). In contrast, no significant effect of training was found in the normally sighted group. These results highlight the potential of PL based on a temporal stimulus to improve the temporal and spatial visual performance in amblyopes. Future work is needed to optimize this method for clinical applications.

Temporal synchronization elicits enhancement of binocular vision functions.

Integration of information over the CNS is an important neural process that affects our ability to perceive and react to the environment. The visual system is required to continuously integrate information arriving from two different sources (the eyes) to create a coherent percept with high spatiotemporal precision. Although this neural integration of information is assumed to be critical for visual performance, it can be impaired under some pathological or developmental conditions. Here we took advantage of a unique developmental condition, amblyopia ("lazy eye"), which is characterized by an impaired temporal synchronization between the two eyes, to meticulously study the effect of synchronization on the integration of binocular visual information. We measured the eyes' asynchrony and compensated for it (with millisecond temporal resolution) by providing time-shifted stimuli to the eyes. We found that the re-synchronization of the ocular input elicited a significant improvement in visual functions, and binocular functions, such as binocular summation and stereopsis, were regained. This phenomenon was also evident in neurophysiological measures. Our results can shed light on other neural processing aspects and might also have translational relevance for the field of training, rehabilitation, and perceptual learning.

Optimizing the fabrication of a 3D high-resolution implant for neural stimulation.

BACKGROUND: Tissue-integrated micro-electronic devices for neural stimulation hold great potential in restoring the functionality of degenerated organs, specifically, retinal prostheses, which are aimed at vision restoration. The fabrication process of 3D polymer-metal devices with high resolution and a high aspect-ratio (AR) is very complex and faces many challenges that impair its functionality. APPROACH: Here we describe the optimization of the fabrication process of a bio-functionalized 3D high-resolution 1mm circular subretinal implant composed of SU-8 polymer integrated with dense gold microelectrodes (23µm pitch) passivated with 3D micro-well-like structures (20µm diameter, 3µm resolution). The main challenges were overcome by step-by-step planning and optimization while utilizing a two-step bi-layer lift-off process; bio-functionalization was carried out by N2 plasma treatment and the addition of a bio-adhesion molecule. MAIN RESULTS: In-vitro and in-vivo investigations, including SEM and FIB cross section examinations, revealed a good structural design, as well as a good long-term integration of the device in the rat sub-retinal space and cell migration into the wells. Moreover, the feasibility of subretinal neural stimulation using the fabricated device was demonstrated in-vitro by electrical activation of rat's retina. CONCLUSIONS: The reported process and optimization steps described here in detail can aid in designing and fabricating retinal prosthetic devices or similar neural implants.

Cortical responses to prosthetic retinal stimulation are significantly affected by the light-adaptive state of the surrounding normal retina.

Objective.Restoration of central vision loss in patients with age-related macular degeneration (AMD) by implanting a retinal prosthesis is associated with an intriguing situation wherein the central prosthetic vision co-exists with natural normal vision. Of major interest are the interactions between the prosthetic and natural vision. Here we studied the effect of the light-adaptive state of the normal retina on the electrical visual evoked potentials (VEPs) arising from the retinal prosthesis.Approach.We recorded electrical VEP elicited by prosthetic retinal stimulation in wild-type rats implanted with a 1 mm photovoltaic subretinal array. Cortical responses were recorded following overnight dark adaption and compared to those recorded following bleaching of the retina by light (520 nm) at various intensities and durations.Main results.Compared to dark-adapted responses, bleaching induced a 2-fold decrease in the prosthetic cortical response, which returned to the dark-adapted baseline within 30 min to several hours, depending on the degree of bleaching. This reduction was neither observed in Royal College of Surgeons (RCS) rats with a degenerated photoreceptor layer nor following intravitreal injection of a GABAa receptor blocker (bicuculine), suggesting the involvement of photoreceptors and a GABAa-mediated mechanism.Significance.These findings show a robust effect of the retinal light-adaptive state on the obtained prosthetic responses. If a similar effect is found in humans, this will have immediate implications on the design of prosthetic devices, where both natural and prosthetic vision co-exist, such as in AMD patients receiving a photovoltaic retinal implant. Similarly, standardization of the retinal light-adaptive state in prosthetic clinical trials should be considered.

Evaluation and Optimization of Methods for Generating High-Resolution Retinotopic Maps Using Visual Cortex Voltage-Sensitive Dye Imaging.

The localization and measurement of neuronal activity magnitude at high spatial and temporal resolution are essential for mapping and better understanding neuronal systems and mechanisms. One such example is the generation of retinotopic maps, which correlates localized retinal stimulation with the corresponding specific visual cortex responses. Here we evaluated and compared seven different methods for extracting and localizing cortical responses from voltage-sensitive dye imaging recordings, elicited by visual stimuli projected directly on the rat retina by a customized projection system. The performance of these methods was evaluated both qualitatively and quantitatively by means of two cluster separation metrics, namely, the (adjusted) Silhouette Index (SI) and the (adjusted) Davies-Bouldin Index (DBI). These metrics were validated using simulated data, which showed that Temporally Structured Component Analysis (TSCA) outperformed all other analysis methods for localizing cortical responses and generating high-resolution retinotopic maps. The analysis methods, as well as the use of cluster separation metrics proposed here, can facilitate future research aiming to localize specific activity at high resolution in the visual cortex or other brain areas.

Electrophysiologic Characterization of Developing Human Embryonic Stem Cell-Derived Photoreceptor Precursors.

Purpose: Photoreceptor precursor cells (PRPs) differentiated from human embryonic stem cells can serve as a source for cell replacement therapy aimed at vision restoration in patients suffering from degenerative diseases of the outer retina, such as retinitis pigmentosa and AMD. In this work, we studied the electrophysiologic maturation of PRPs throughout the differentiation process. Methods: Human embryonic stem cells were differentiated into PRPs and whole-cell recordings were performed for electrophysiologic characterization at days 0, 30, 60, and 90 along with quantitative PCR analysis to characterize the expression level of various ion channels, which shape the electrophysiologic response. Finally, to characterize the electrically induced calcium currents, we employed calcium imaging (rhod4) to visualize intracellular calcium dynamics in response to electrical activation. Results: Our results revealed an early and steady presence (approximately 100% of responsive cells) of the delayed potassium rectifier current. In contrast, the percentage of cells exhibiting voltage-gated sodium currents increased with maturation (from 0% to almost 90% of responsive cells at 90 days). Moreover, calcium imaging revealed the presence of voltage-gated calcium currents, which play a major role in vision formation. These results were further supported by quantitative PCR analysis, which revealed a significant and continuous (3- to 50-fold) increase in the expression of various voltage-gated channels concomitantly with the increase in the expression of the photoreceptor marker CRX. Conclusions: These results can shed light on the electrophysiologic maturation of neurons in general and PRP in particular and can form the basis for devising and optimizing cell replacement-based vision restoration strategies.

Cortical Interactions between Prosthetic and Natural Vision.

Outer retinal degenerative diseases, such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD), are among the leading causes of incurable blindness in the Western world [1]. Retinal prostheses have been shown to restore some useful vision by electrically stimulating the remaining retinal neurons [2]. In contrast to inherited retinal degenerative diseases (e.g., RP), typically leading to a complete loss of the visual field, in AMD patients the disease is localized to the macula, leaving the peripheral vision intact. Implanting a retinal prosthesis in the central macula in AMD patients [3, 4] leads to an intriguing situation where the patient's central retina is stimulated electrically, whereas the peripheral healthy retina responds to natural light stimulation. An important question is whether the visual cortex responds to these two concurrent stimuli similarly to the interaction between two adjacent natural light stimuli projected onto healthy retina. Here, we investigated the cortical interactions between prosthetic and natural vision based on visually evoked potentials (VEPs) recorded in rats implanted with photovoltaic subretinal implants. Using this model, where prosthetic and natural vision information are combined in the visual cortex, we observed striking similarities in the interactions of natural and prosthetic vision, including similar effect of background illumination, linear summation of non-patterned stimuli, and lateral inhibition with spatial patterns [5], which increased with target contrast. These results support the idea of combined prosthetic and natural vision in restoration of sight for AMD patients.

SEM/FIB Imaging for Studying Neural Interfaces.

Tissue and neural engineering for various regenerative therapies are rapidly growing fields. Of major interest is studying the complex interface between cells and various 3D structures by scanning electron microscopy with focused ion beam. Notwithstanding its unrivaled resolution, the optimal fixation, dehydration, and staining protocols of the samples while preserving the complex cell interface in its natural form, are highly challenging. The aim of this work was to compare and optimize staining and sample drying procedures in order to preserve the cells in their "life-like state" for studying the cell interface with either 3D well-like structures or gold-coated mushroom-shaped electrodes. The process involved chemical fixation using a combination of glutaraldehyde and formaldehyde, followed by gentle drying techniques in which we compared four methods: (critical point drying, hexamethyldisiloxane, repeats of osmium tetroxide-thiocarbohydrazide [OTOTO], and resin) in order to determine the method that best preserves the cell and cell interface morphology. Finally, to visualize the intracellular organelles and membrane, we compared the efficacy of four staining techniques: osmium tetroxide, osmium tetroxide and salts, osmium and uranyl acetate, and OTOTO. Experiments were performed on embryonic stem cell-derived photoreceptor precursors, neural cells, and a human retinal pigment epithelial cell line, which revealed that the optimal processing combination was resin drying and OTOTO staining, as manifested by preservation of cell morphology, the lowest percentage of cellular protrusion breakage as well as a high-quality image. The obtained results pave the way for better understanding the cell interface with various structures for enhancing various biomedical applications.

Carbon nanostructures as a scaffold for human embryonic stem cell differentiation toward photoreceptor precursors.

Carbon nanomaterials have been introduced as a scaffold for various biological applications due to their unique physical and electrical properties. Here we studied carbon nanotubes (CNTs) and carbon nanofibers (CNFs) as scaffold materials for the differentiation of human embryonic stem cells (hESCs) towards photoreceptor precursor cells (PRPs). We report on their cytoxicity, their effect on cell morphology, cell-surface interface and the differentiation process. To this end, hESCs were differentiated into PRPs on carbon nanofibers (CNFs), long horizontal CNTs (LHCNTs), vertically aligned CNTs (VACNTs) or glass (control) surfaces. The differentiated cells were investigated by immunohistochemistry, fluorescence imaging and electron microscopy. Our results revealed that the investigated nanomaterials were not cytotoxic to the cells during the differentiation process. The surface interface effect on the cells was apparent, affecting cell directionality, migration and morphology. Interestingly, cell fate was not dependent on the substrate type, as inferred from the similar dynamics of the loss of pluripotency and the comparable expression levels of the photoreceptor marker Crx for all investigated substrates. These results are important for better understanding the effect of nanomaterial surface interaction with differentiating neural cells in general, and for future use of these materials as scaffolds for differentiating photoreceptors for vision restoration in particular.

Refractive errors and schizophrenia.

BACKGROUND: Refractive errors (myopia, hyperopia and amblyopia), like schizophrenia, have a strong genetic cause, and dopamine has been proposed as a potential mediator in their pathophysiology. The present study explored the association between refractive errors in adolescence and schizophrenia, and the potential familiality of this association. METHODS: The Israeli Draft Board carries a mandatory standardized visual accuracy assessment. 678,674 males consecutively assessed by the Draft Board and found to be psychiatrically healthy at age 17 were followed for psychiatric hospitalization with schizophrenia using the Israeli National Psychiatric Hospitalization Case Registry. Sib-ships were also identified within the cohort. RESULTS: There was a negative association between refractive errors and later hospitalization for schizophrenia. Future male schizophrenia patients were two times less likely to have refractive errors compared with never-hospitalized individuals, controlling for intelligence, years of education and socioeconomic status [adjusted Hazard Ratio=.55; 95% confidence interval .35-.85]. The non-schizophrenic male siblings of schizophrenia patients also had lower prevalence of refractive errors compared to never-hospitalized individuals. CONCLUSIONS: Presence of refractive errors in adolescence is related to lower risk for schizophrenia. The familiality of this association suggests that refractive errors may be associated with the genetic liability to schizophrenia.

High-resolution VSDI retinotopic mapping via a DLP-based projection system.

High-resolution recording of visual cortex activity is an important tool for vision research. Using a customized digital mirror device (DMD) - based system equipped with retinal imaging, we projected visual stimuli directly on the rat retina and recorded cortical responses by voltage-sensitive dye imaging. We obtained robust cortical responses and generated high-resolution retinotopic maps at an unprecedented retinal resolution of 4.6 degrees in the field of view, while further distinguishing between normal and pathological retinal areas. This system is a useful tool for studying the cortical response to localized retinal stimulation and may shed light on various cortical plasticity processes.

Gold nanoparticles for multimodal high-resolution imaging of transplanted cells for retinal replacement therapy.

Aim: Longitudinal tracking of transplanted cells in clinical and experimental setups is crucial for evaluating the efficiency of retinal cell replacement therapies. Materials & methods: Gold nanoparticle-labeled photoreceptor precursors were transplanted in the vitreous and subretinal space of rats and were longitudinally tracked for over a month using optical coherence tomography, computed tomography and fluorescence fundus imaging. Results: This multimodal imaging approach enabled high-resolution long-term tracking and estimation of cell survival in the retina and vitreous, while displaying no toxic effects on the cells or the retina. Conclusion: These observations highlight the applicability of using gold nanoparticle for retinal cell tracking in existing experimental settings and its translational potential for providing more efficient retinal cell therapy in humans.