Evaluating the outcomes of pluripotent stem-cell-derived photoreceptor transplantation in retinal repair

\ua9 2025 The Author(s). The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.In recent decades, numerous research groups have focused on restoring visual function through the transplantation of stem cells into animal models of retinal neurodegeneration. Significant advancements in surgical techniques, the maturation of donor cells, and the production of cell suspensions, along with ensuring proper synaptic connectivity with the host environment, are key considerations for the potential implementation of this strategy in clinical practice. In this review, we summarize the latest progress in the transplantation of stem cell-derived photoreceptors, emphasizing the outcomes related to visual function observed in the used animal models. Additionally, we analyze the various methods of stem cell differentiation and the surgical techniques selected for transplanting these photoreceptor precursors. Finally, we report on functional assessments from recent studies to highlight the considerable potential of stem cell-derived photoreceptor transplants as a therapeutic approach for retinal degenerative diseases

Early-Stage Waves in the Retinal Network Emerge Close to a Critical State Transition between Local and Global Functional Connectivity

A novel, biophysically realistic model for early-stage, acetylcholine-mediated retinal waves is presented. In this model, neural excitability is regulated through a slow after-hyperpolarization (sAHP) operating on two different temporal scales. As a result, the simulated network exhibits competition between a desynchronizing effect of spontaneous, cell-intrinsic bursts, and the synchronizing effect of synaptic transmission during retinal waves. Cell-intrinsic bursts decouple the retinal network through activation of the sAHP current, and we show that the network is capable of operating at a transition point between purely local and global functional connectedness, which corresponds to a percolation phase transition. Multielectrode array recordings show that, at this point, the properties of retinal waves are reliably predicted by the model. These results indicate that early spontaneous activity in the developing retina is regulated according to a very specific principle, which maximizes randomness and variability in the resulting activity patterns

Segmentation of Retinal Ganglion Cells From Fluorescent Microscopy Imaging

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Systematic Comparison of Retinal Organoid Differentiation from Human Pluripotent Stem Cells Reveals Stage Specific, Cell Line, and Methodological Differences

A major goal in the stem cell field is to generate tissues that can be utilized as a universal tool for in vitro models of development and disease, drug development, or as a resource for patients suffering from disease or injury. Great efforts are being made to differentiate human pluripotent stem cells in vitro toward retinal tissue, which is akin to native human retina in its cytoarchitecture and function, yet the numerous existing retinal induction protocols remain variable in their efficiency and do not routinely produce morphologically or functionally mature photoreceptors. Herein, we determine the impact that the method of embryoid body (EB) formation and maintenance as well as cell line background has on retinal organoid differentiation from human embryonic stem cells and human induced pluripotent stem cells. Our data indicate that cell line-specific differences dominate the variables that underline the differentiation efficiency in the early stages of differentiation. In contrast, the EB generation method and maintenance conditions determine the later differentiation and maturation of retinal organoids. Of the latter, the mechanical method of EB generation under static conditions, accompanied by media supplementation with Y27632 for the first 48 hours of differentiation, results in the most consistent formation of laminated retinal neuroepithelium containing mature and electrophysiologically responsive photoreceptors. Collectively, our data provide substantive evidence for stage-specific differences in the ability to give rise to laminated retinae, which is determined by cell line-specific differences in the early stages of differentiation and EB generation/organoid maintenance methods at later stages

Refinement and Pattern Formation in Neural Circuits by the Interaction of Traveling Waves with Spike-Timing Dependent Plasticity

Traveling waves in the developing brain are a prominent source of highly correlated spiking activity that may instruct the refinement of neural circuits. A candidate mechanism for mediating such refinement is spike-timing dependent plasticity (STDP), which translates correlated activity patterns into changes in synaptic strength. To assess the potential of these phenomena to build useful structure in developing neural circuits, we examined the interaction of wave activity with STDP rules in simple, biologically plausible models of spiking neurons. We derive an expression for the synaptic strength dynamics showing that, by mapping the time dependence of STDP into spatial interactions, traveling waves can build periodic synaptic connectivity patterns into feedforward circuits with a broad class of experimentally observed STDP rules. The spatial scale of the connectivity patterns increases with wave speed and STDP time constants. We verify these results with simulations and demonstrate their robustness to likely sources of noise. We show how this pattern formation ability, which is analogous to solutions of reaction-diffusion systems that have been widely applied to biological pattern formation, can be harnessed to instruct the refinement of postsynaptic receptive fields. Our results hold for rich, complex wave patterns in two dimensions and over several orders of magnitude in wave speeds and STDP time constants, and they provide predictions that can be tested under existing experimental paradigms. Our model generalizes across brain areas and STDP rules, allowing broad application to the ubiquitous occurrence of traveling waves and to wave-like activity patterns induced by moving stimuli

A super-resolution approach for receptive fields estimation of neuronal ensembles

International audienceThe Spike Triggered Average (STA) is a classical technique to find a discrete approximation of the Receptive Fields (RFs) of sensory neurons [1], a required analysis in most experimental studies. One important parameter of the STA is the spatial resolution of the estimation, corresponding to the size of the blocks of the checkerboard stimulus images. In general, it is experimentally fixed to reach a compromise: If too small, neuronal responses might be too weak thus leading to RF with low Signal-to-Noise-Ratio; on the contrary, if too large, small RF will be lost, or not described with enough details, because of the coarse approximation. Other solutions were proposed consisting in starting from a small block size and updating it following the neuron response in a closed-loop to increase its response [2; 3; 4]. However, these solutions were designed for single cells and cannot be applied to simultaneous recordings of ensembles of neurons (since each RF has its own size and preferred stimulus). To solve this problem, we introduced a modified checkerboard stimulus where blocks are shifted randomly in space at fixed time steps. This idea is inspired from super-resolution techniques developed in image processing [4]. The main interest is that the block size can be large, enabling strong responses, while the resolution can be finer since it depends on the shift minimum size. In [5] was shown that the STA remains an unbiased RF estimator and, using simulated spike trains from an ensemble of Linear Nonlinear Poisson cascade neurons, it was predicted that this approach improves RF estimation over the neuron ensemble. Here, we test these predictions experimentally on the RFs estimation of 8460 ganglion cells from two mouse retinas, using recordings performed with a large scale high-density multielectrode array. To illustrate the main interest of the approach, in Figure 1 we show a representative example of STA for one neuron where RFs have been obtained using the three following stimuli (all presented during 15min, for one retina displayed at 10 Hz, for the other at 30 Hz): (A) standard checkerboard stimulus with block size of 160μm, (B) standard checkerboard stimulus with block size of 40μm, (C) checkerboard stimulus with block size of 160μm and arbitrary shifts of 40μm in x and y-directions. Results show spatial resolution can be improved in case (C), while nothing could be obtained in (B) by changing only the block size of the standard stimulus. At the population level, plot (D) shows the number of the RFs that could be recovered for each stimuli, using a decision criteria based of the RFs value distribution. Most of the RFs were mapped with both methods (A) and (C) (49.9%). However, the proposed case (C) allows to recover 51% of the mapped RFs at a resolution of 40μm, while in the classical case (A), 41% of the RFs could be found at a resolution of only 160μm. Thus, the method does improve the quality of the RF estimation and the amount of successfully mapped RFs in neural ensembles

CRX Expression in Pluripotent Stem Cell-Derived Photoreceptors Marks a Transplantable Subpopulation of Early Cones

Death of photoreceptors is a common cause of age-related and inherited retinal dystrophies, and thus their replenishment from renewable stem cell sources is a highly desirable therapeutic goal. Human pluripotent stem cells provide a useful cell source in view of their limitless self-renewal capacity and potential to not only differentiate into cells of the retina but also self-organize into tissue with structure akin to the human retina as part of three-dimensional retinal organoids. Photoreceptor precursors have been isolated from differentiating human pluripotent stem cells through application of cell surface markers or fluorescent reporter approaches and shown to have a similar transcriptome to fetal photoreceptors. In this study, we investigated the transcriptional profile of CRX-expressing photoreceptor precursors derived from human pluripotent stem cells and their engraftment capacity in an animal model of retinitis pigmentosa (Pde6brd1), which is characterized by rapid photoreceptor degeneration. Single cell RNA-Seq analysis revealed the presence of a dominant cell cluster comprising 72% of the cells, which displayed the hallmarks of early cone photoreceptor expression. When transplanted subretinally into the Pde6brd1 mice, the CRX + cells settled next to the inner nuclear layer and made connections with the inner neurons of the host retina, and approximately one-third of them expressed the pan cone marker, Arrestin 3, indicating further maturation upon integration into the host retina. Together, our data provide valuable molecular insights into the transcriptional profile of human pluripotent stem cells-derived CRX + photoreceptor precursors and indicate their usefulness as a source of transplantable cone photoreceptors. Stem Cells 2019;37:609–622

Unsupervised Spike Sorting for Large-Scale, High-Density Multielectrode Arrays

We present a method for automated spike sorting for recordings with high-density, large-scale multielectrode arrays. Exploiting the dense sampling of single neurons by multiple electrodes, an efficient, low-dimensional representation of detected spikes consisting of estimated spatial spike locations and dominant spike shape features is exploited for fast and reliable clustering into single units. Millions of events can be sorted in minutes, and the method is parallelized and scales better than quadratically with the number of detected spikes. Performance is demonstrated using recordings with a 4,096-channel array and validated using anatomical imaging, optogenetic stimulation, and model-based quality control. A comparison with semi-automated, shape-based spike sorting exposes significant limitations of conventional methods. Our approach demonstrates that it is feasible to reliably isolate the activity of up to thousands of neurons and that dense, multi-channel probes substantially aid reliable spike sorting

Refuting the challenges of the developmental shift of polarity of GABA actions: GABA more exciting than ever!

During brain development, there is a progressive reduction of intracellular chloride associated with a shift in GABA polarity: GABA depolarizes and occasionally excites immature neurons, subsequently hyperpolarizing them at later stages of development. This sequence, which has been observed in a wide range of animal species, brain structures and preparations, is thought to play an important role in activity-dependent formation and modulation of functional circuits. This sequence has also been considerably reinforced recently with new data pointing to an evolutionary preserved rule. In a recent 'Hypothesis and Theory Article', the excitatory action of GABA in early brain development is suggested to be "an experimental artefact" (Bregestovski and Bernard, 2012). The authors suggest that the excitatory action of GABA is due to an inadequate/insufficient energy supply in glucose-perfused slices and/or to the damage produced by the slicing procedure. However, these observations have been repeatedly contradicted by many groups and are inconsistent with a large body of evidence including the fact that the developmental shift is neither restricted to slices nor to rodents. We summarize the overwhelming evidence in support of both excitatory GABA during development, and the implications this has in developmental neurobiology. \ua9 2012 Ben-ari, Woodin, Sernagor, Cancedda, Vinay, Rivera,Legendre, Luhmann, Bordey, Wenner, Fukuda, Pol, Jean-luc and Cherubini