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Frontiers in Cellular Neuroscience 2024Mechanical sensitive channels expressed in mammalian retinas are effectors of elevated pressure stresses, but it is unclear how their activation affects visual function...
INTRODUCTION
Mechanical sensitive channels expressed in mammalian retinas are effectors of elevated pressure stresses, but it is unclear how their activation affects visual function in pressure-related retinal disorders.
METHODS
This study investigated the role of the transient potential channel vanilloid TRPV4 in photoreceptors and rod bipolar cells (RBCs) with immunohistochemistry, confocal microscopy, electroretinography (ERG), and patch-clamp techniques.
RESULTS
TRPV4 immunoreactivity (IR) was found in the outer segments of photoreceptors, dendrites and somas of PKCα-positive RBCs and other BCs, plexiform layers, and retinal ganglion cells (RGCs) in wild-type mice. TRPV4-IR was largely diminished in the retinas of homozygous TRPV4 transgenic mice. Genetically suppressing TRPV4 expression moderately but significantly enhanced the amplitude of ERG a- and b-waves evoked by scotopic and mesopic lights (0.55 to 200 Rh*rod s) and photopic lights (10-10 Rh*rod s) compared to wild-type mice in fully dark-adapted conditions. The implicit time evoked by dim lights (0.55 to 200 Rh*rod s) was significantly decreased for b-waves and elongated for a-waves in the transgenic mice. ERG b-wave evoked by dim lights is primarily mediated by RBCs, and under voltage-clamp conditions, the latency of the light-evoked cation current in RBCs of the transgenic mice was significantly shorter compared to wild-type mice. About 10% of the transgenic mice had one eye undeveloped, and the percentage was significantly higher than in wild-type mice.
CONCLUSIONS
The data indicates that TRPV4 involves ocular development and is expressed and active in outer retinal neurons, and interventions of TRPV4 can variably affect visual signals in rods, cones, RBCs, and cone ON BCs.
PubMed: 38903773
DOI: 10.3389/fncel.2024.1404929 -
BioRxiv : the Preprint Server For... May 2024The neurovascular unit (NVU), comprising vascular, glial and neural elements, supports the energetic demands of neural computation, but this aspect of the retina's...
The neurovascular unit (NVU), comprising vascular, glial and neural elements, supports the energetic demands of neural computation, but this aspect of the retina's trilaminar vessel network is poorly understood. Only the innermost vessel layer - the superficial vascular plexus (SVP) - is ensheathed by astrocytes, like brain capillaries, whereas glial ensheathment in other layers derives from radial Müller glia. Using serial electron microscopy reconstructions from mouse and primate retina, we find that Müller processes cover capillaries in a tessellating pattern, mirroring the tiled astrocytic endfeet wrapping brain capillaries. However, gaps in the Müller sheath, found mainly in the intermediate vascular plexus (IVP), permit different neuron types to contact pericytes and the endothelial cells directly. Pericyte somata are a favored target, often at spine-like structures with a reduced or absent vascular basement lamina. Focal application of adenosine triphosphate (ATP) to the vitreal surface evoked Ca signals in Müller sheaths in all three vascular layers. Pharmacological experiments confirmed that Müller sheaths express purinergic receptors that, when activated, trigger intracellular Ca signals that are amplified by IP -controlled intracellular Ca stores. When rod photoreceptors die in a mouse model of retinitis pigmentosa ( ), Müller sheaths dissociate from the deep vascular plexus (DVP) but are largely unchanged within the IVP or SVP. Thus, Müller glia interact with retinal vessels in a laminar, compartmentalized manner: glial sheathes are virtually complete in the SVP but fenestrated in the IVP, permitting direct neural-to-vascular contacts. In the DVP, the glial sheath is only modestly fenestrated and is vulnerable to photoreceptor degeneration.
PubMed: 38903067
DOI: 10.1101/2024.04.30.591885 -
Molecular Pharmaceutics Jun 2024Oxidative stress is pivotal in retinal disease progression, causing dysfunction in various retinal components. An effective antioxidant, such as probucol (PB), is vital...
Oxidative stress is pivotal in retinal disease progression, causing dysfunction in various retinal components. An effective antioxidant, such as probucol (PB), is vital to counteract oxidative stress and emerges as a potential candidate for treating retinal degeneration. However, the challenges associated with delivering lipophilic drugs such as PB to the posterior segment of the eye, specifically targeting photoreceptor cells, necessitate innovative solutions. This study uses formulation-based spray dry encapsulation technology to develop polymer-based PB-lithocholic acid (LCA) nanoparticles and assesses their efficacy in the 661W photoreceptor-like cell line. Incorporating LCA enhances nanoparticles' biological efficacy without compromising PB stability. studies demonstrate that PB-LCA nanoparticles prevent reactive oxygen species (ROS)-induced oxidative stress by improving cellular viability through the nuclear erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) pathway. These findings propose PB-LCA nanoparticles as a promising therapeutic strategy for oxidative stress-induced retinopathies.
PubMed: 38899552
DOI: 10.1021/acs.molpharmaceut.4c00269 -
Cell and Tissue Research Jun 2024The marine microturbellarian Macrostomum lignano (Platyhelminthes, Rhabditophora) is an emerging laboratory model used by a growing community of researchers because it...
The marine microturbellarian Macrostomum lignano (Platyhelminthes, Rhabditophora) is an emerging laboratory model used by a growing community of researchers because it is easy to cultivate, has a fully sequenced genome, and offers multiple molecular tools for its study. M. lignano has a compartmentalized brain that receives sensory information from receptors integrated in the epidermis. Receptors of the head, as well as accompanying glands and specialized epidermal cells, form a compound sensory structure called the frontal glandular complex. In this study, we used semi-serial transmission electron microscopy (TEM) to document the types, ultrastructure, and three-dimensional architecture of the cells of the frontal glandular complex. We distinguish a ventral compartment formed by clusters of type 1 (multiciliated) sensory receptors from a central domain where type 2 (collar) sensory receptors predominate. Six different types of glands (rhammite glands, mucoid glands, glands with aster-like and perimaculate granula, vacuolated glands, and buckle glands) are closely associated with type 1 sensory receptors. Endings of a seventh type of gland (rhabdite gland) define a dorsal domain of the frontal glandular complex. A pair of ciliary photoreceptors is closely associated with the base of the frontal glandular complex. Bundles of dendrites, connecting the receptor endings with their cell bodies which are located in the brain, form the (frontal) peripheral nerves. Nerve fibers show a varicose structure, with thick segments alternating with thin segments, and are devoid of a glial layer. This distinguishes platyhelminths from larger and/or more complex invertebrates whose nerves are embedded in prominent glial sheaths.
PubMed: 38898317
DOI: 10.1007/s00441-024-03901-x -
Nature Communications Jun 2024Phototransduction involves changes in concentration of ions and other solutes within photoreceptors and in subretinal space, which affect osmotic pressure and the...
Phototransduction involves changes in concentration of ions and other solutes within photoreceptors and in subretinal space, which affect osmotic pressure and the associated water flow. Corresponding expansion and contraction of cellular layers can be imaged using optoretinography (ORG), based on phase-resolved optical coherence tomography (OCT). Until now, ORG could reliably detect only photoisomerization and phototransduction in photoreceptors, primarily in cones under bright stimuli. Here, by employing a phase-restoring subpixel motion correction algorithm, which enables imaging of the nanometer-scale tissue dynamics during minute-long recordings, and unsupervised learning of spatiotemporal patterns, we discover optical signatures of the other retinal structures' response to visual stimuli. These include inner and outer segments of rod photoreceptors, retinal pigment epithelium, and subretinal space in general. The high sensitivity of our technique enables detection of the retinal responses to dim stimuli: down to 0.01% bleach level, corresponding to natural levels of scotopic illumination. We also demonstrate that with a single flash, the optoretinogram can map retinal responses across a 12° field of view, potentially replacing multifocal electroretinography. This technique expands the diagnostic capabilities and practical applicability of optoretinography, providing an alternative to electroretinography, while combining structural and functional retinal imaging in the same OCT machine.
Topics: Tomography, Optical Coherence; Animals; Retinal Pigment Epithelium; Retinal Rod Photoreceptor Cells; Retina; Light; Photic Stimulation; Algorithms; Male
PubMed: 38898002
DOI: 10.1038/s41467-024-49014-5 -
Experimental Eye Research Jun 2024This review examines the pivotal role of photoreceptor cells in ocular refraction development, focusing on dopamine (DA) as a key neurotransmitter. Contrary to the... (Review)
Review
This review examines the pivotal role of photoreceptor cells in ocular refraction development, focusing on dopamine (DA) as a key neurotransmitter. Contrary to the earlier view favoring cone cells, recent studies have highlighted the substantial contributions of both rod and cone cells to the visual signaling pathways that influence ocular refractive development. Notably, rod cells appeared to play a central role. Photoreceptor cells interact intricately with circadian rhythms, color vision pathways, and other neurotransmitters, all of which are crucial for the complex mechanisms driving the development of myopia. This review emphasizes that ocular refractive development results from a coordinated interplay between diverse cell types, signaling pathways, and neurotransmitters. This perspective has significant implications for unraveling the complex mechanisms underlying myopia and aiding in the development of more effective prevention and treatment strategies.
PubMed: 38897270
DOI: 10.1016/j.exer.2024.109976 -
Biomaterials Jun 2024Clinical results with photovoltaic subretinal prosthesis (PRIMA) demonstrated restoration of sight via electrical stimulation of the interneurons in degenerated retina,...
Clinical results with photovoltaic subretinal prosthesis (PRIMA) demonstrated restoration of sight via electrical stimulation of the interneurons in degenerated retina, with resolution matching the 100 μm pixel size. Since scaling the pixels below 75 μm in the current bipolar planar geometry will significantly limit the penetration depth of the electric field and increase stimulation threshold, we explore the possibility of using smaller pixels based on a novel 3-dimensional honeycomb-shaped design. We assessed the long-term biocompatibility and stability of these arrays in rats by investigating the anatomical integration of the retina with flat and 3D implants and response to electrical stimulation over lifetime - up to 32-36 weeks post-implantation in aged rats. With both flat and 3D implants, signals elicited in the visual cortex decreased after the day of implantation by more than 3-fold, and gradually recovered over the next 12-16 weeks. With 25 μm high honeycomb walls, the majority of bipolar cells migrate into the wells, while amacrine and ganglion cells remain above the cavities, which is essential for selective network-mediated stimulation of the retina. Retinal thickness and full-field stimulation threshold with 40 μm-wide honeycomb pixels were comparable to those with planar devices - 0.05 mW/mm with 10 ms pulses. However, fewer cells from the inner nuclear layer migrated into the 20 μm-wide wells, and stimulation threshold increased over 12-16 weeks, before stabilizing at about 0.08 mW/mm. Such threshold is still significantly lower than 1.8 mW/mm with a previous design of flat bipolar pixels, confirming the promise of the 3D honeycomb-based approach to high resolution subretinal prosthesis.
PubMed: 38897028
DOI: 10.1016/j.biomaterials.2024.122674 -
BioRxiv : the Preprint Server For... Jun 2024CLN3 Batten disease (also known as Juvenile Neuronal Ceroid Lipofuscinosis; JNCL) is a lysosomal storage disorder that typically initiates with retinal degeneration but...
PURPOSE
CLN3 Batten disease (also known as Juvenile Neuronal Ceroid Lipofuscinosis; JNCL) is a lysosomal storage disorder that typically initiates with retinal degeneration but is followed by seizure onset, motor decline and premature death. Patient-derived CLN3 disease iPSC-RPE cells show defective phagocytosis of photoreceptor outer segments (POSs). Because modifier genes are implicated in CLN3 disease, our goal here was to investigate a direct link between mutation and POS phagocytosis defect.
METHODS
Isogenic control and mutant stem cell lines were generated by CRISPR-Cas9-mediated biallelic deletion of exons 7 and 8. A transgenic ( ) Yucatan miniswine was also used to study the impact of mutation on POS phagocytosis. POS phagocytosis by cultured RPE cells was analyzed by Western blotting and immunohistochemistry. Electroretinogram, optical coherence tomography and histological analysis of and wild-type miniswine eyes were carried out at 6-, 36-, or 48-month age.
RESULTS
RPE ( RPE) displayed reduced POS binding and consequently decreased uptake of POS compared to isogenic control RPE cells. Furthermore, wild-type miniswine RPE cells phagocytosed POS less efficiently than wild-type POS. Consistent with decreased POS phagocytosis, lipofuscin/autofluorescence was decreased in miniswine RPE at 36 months-of-age and was followed by almost complete loss of photoreceptors at 48 months of age.
CONCLUSIONS
mutation (that affects up to 85% patients) affects both RPE and POSs and leads to photoreceptor cell loss in CLN3 disease. Furthermore, both primary RPE dysfunction and mutant POS independently contribute to impaired POS phagocytosis in CLN3 disease.
PubMed: 38895469
DOI: 10.1101/2024.06.09.597388 -
BioRxiv : the Preprint Server For... Jun 2024Rod photoreceptor formation in the postnatal mouse is a widely used model system for studying mammalian photoreceptor development. This experimental paradigm provides...
Rod photoreceptor formation in the postnatal mouse is a widely used model system for studying mammalian photoreceptor development. This experimental paradigm provides opportunities for both gain and loss-of-function studies which can be accomplished through in vivo plasmid delivery and electroporation. However, the cis-regulatory elements used to implement this approach have not been fully evaluated or optimized for the unique transcriptional environment of photoreceptors. Here we report that the use of a photoreceptor cis-regulatory element from the Crx gene in combination with broadly active promoter elements can increase the targeting of developing rod photoreceptors in the mouse. This can lead to greater reporter expression, as well as enhanced misexpression and loss-of-function phenotypes in these cells. This study also highlights the importance of identifying and testing relevant cis-regulatory elements when planning cell subtype specific experiments. The use of the specific hybrid elements in this study will provide a more efficacious gene delivery system to study mammalian photoreceptor formation.
PubMed: 38895286
DOI: 10.1101/2024.06.06.597220 -
International Journal of Molecular... Jun 2024Diabetic retinopathy (DR) is a very serious diabetes complication. Changes in the O-linked N-acetylglucosamine (O-GlcNAc) modification are associated with many diseases....
Diabetic retinopathy (DR) is a very serious diabetes complication. Changes in the O-linked N-acetylglucosamine (O-GlcNAc) modification are associated with many diseases. However, its role in DR is not fully understood. In this research, we explored the effect of O-GlcNAc modification regulation by activating AMP-activated protein kinase (AMPK) in DR, providing some evidence for clinical DR treatment in the future. Bioinformatics was used to make predictions from the database, which were validated using the serum samples of diabetic patients. As an in vivo model, diabetic mice were induced using streptozotocin (STZ) injection with/without an AMPK agonist (metformin) or an AMPK inhibitor (compound C) treatment. Electroretinogram (ERG) and H&E staining were used to evaluate the retinal functional and morphological changes. In vitro, 661 w cells were exposed to high-glucose conditions, with or without metformin treatment. Apoptosis was evaluated using TUNEL staining. The protein expression was detected using Western blot and immunofluorescence staining. The angiogenesis ability was detected using a tube formation assay. The levels of O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) in the serum changed in the DR patients in the clinic. In the diabetic mice, the ERG wave amplitude and retinal thickness decreased. In vitro, the apoptotic cell percentage and Bax expression were increased, and Bcl2 expression was decreased in the 661 w cells under high-glucose conditions. The O-GlcNAc modification was increased in DR. In addition, the expression of GFAT/TXNIP O-GlcNAc was also increased in the 661 w cells after the high-glucose treatment. Additionally, the Co-immunoprecipitation(CO-IP) results show that TXNIP interacted with the O-GlcNAc modification. However, AMPK activation ameliorated this effect. We also found that silencing the AMPKα1 subunit reversed this process. In addition, the conditioned medium of the 661 w cells may have affected the tube formation in vitro. Taken together, O-GlcNAc modification was increased in DR with photoreceptor cell degeneration and neovascularization; however, it was reversed after activating AMPK. The underlying mechanism is linked to the GFAT/TXNIP-O-GlcNAc modification signaling axis. Therefore, the AMPKα1 subunit plays a vital role in the process.
Topics: Diabetic Retinopathy; Animals; Mice; Acetylglucosamine; N-Acetylglucosaminyltransferases; Diabetes Mellitus, Experimental; Humans; AMP-Activated Protein Kinases; Male; Apoptosis; Metformin; beta-N-Acetylhexosaminidases; Retina; Mice, Inbred C57BL; Cell Line
PubMed: 38892474
DOI: 10.3390/ijms25116286