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1.
Acta Neuropathol Commun ; 12(1): 150, 2024 Sep 14.
Article in English | MEDLINE | ID: mdl-39300576

ABSTRACT

BACKGROUND: Glaucoma is a leading cause of blindness, affecting retinal ganglion cells (RGCs) and their axons. By 2040, it is likely to affect 110 million people. Neuroinflammation, specifically through the release of proinflammatory cytokines by M1 microglial cells, plays a crucial role in glaucoma progression. Indeed, in post-mortem human studies, pre-clinical models, and ex-vivo models, RGC degeneration has been consistently shown to be linked to inflammation in response to cell death and tissue damage. Recently, Rho kinase inhibitors (ROCKis) have emerged as potential therapies for neuroinflammatory and neurodegenerative diseases. This study aimed to investigate the potential effects of three ROCKis (Y-27632, Y-33075, and H-1152) on retinal ganglion cell (RGC) loss and retinal neuroinflammation using an ex-vivo retinal explant model. METHODS: Rat retinal explants underwent optic nerve axotomy and were treated with Y-27632, Y-33075, or H-1152. The neuroprotective effects on RGCs were evaluated using immunofluorescence and Brn3a-specific markers. Reactive glia and microglial activation were studied by GFAP, CD68, and Iba1 staining. Flow cytometry was used to quantify day ex-vivo 4 (DEV 4) microglial proliferation and M1 activation by measuring the number of CD11b+, CD68+, and CD11b+/CD68+ cells after treatment with control solvent or Y-33075. The modulation of gene expression was measured by RNA-seq analysis on control and Y-33075-treated explants and glial and pro-inflammatory cytokine gene expression was validated by RT-qPCR. RESULTS: Y-27632 and H-1152 did not significantly protect RGCs. By contrast, at DEV 4, 50 µM Y-33075 significantly increased RGC survival. Immunohistology showed a reduced number of Iba1+/CD68+ cells and limited astrogliosis with Y-33075 treatment. Flow cytometry confirmed lower CD11b+, CD68+, and CD11b+/CD68+ cell numbers in the Y-33075 group. RNA-seq showed Y-33075 inhibited the expression of M1 microglial markers (Tnfα, Il-1ß, Nos2) and glial markers (Gfap, Itgam, Cd68) and to reduce apoptosis, ferroptosis, inflammasome formation, complement activation, TLR pathway activation, and P2rx7 and Gpr84 gene expression. Conversely, Y-33075 upregulated RGC-specific markers, neurofilament formation, and neurotransmitter regulator expression, consistent with its neuroprotective effects. CONCLUSION: Y-33075 demonstrates marked neuroprotective and anti-inflammatory effects, surpassing the other tested ROCKis (Y-27632 and H-1152) in preventing RGC death and reducing microglial inflammatory responses. These findings highlight its potential as a therapeutic option for glaucoma.


Subject(s)
Neuroprotective Agents , Pyridines , Retinal Ganglion Cells , rho-Associated Kinases , Animals , Pyridines/pharmacology , rho-Associated Kinases/metabolism , rho-Associated Kinases/antagonists & inhibitors , Neuroprotective Agents/pharmacology , Rats , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Neuroinflammatory Diseases/drug therapy , Neuroinflammatory Diseases/metabolism , Retina/drug effects , Retina/pathology , Retina/metabolism , Amides/pharmacology , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/analogs & derivatives , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/pharmacology , Rats, Sprague-Dawley , Neuroprotection/drug effects , Neuroprotection/physiology , Microglia/drug effects , Microglia/metabolism , Microglia/pathology , Protein Kinase Inhibitors/pharmacology , Male , Optic Nerve Injuries/drug therapy , Optic Nerve Injuries/pathology , Optic Nerve Injuries/metabolism , Isoquinolines , Sulfonamides
2.
PLoS One ; 19(9): e0309400, 2024.
Article in English | MEDLINE | ID: mdl-39255314

ABSTRACT

Glaucoma is a blinding disease where the retinal ganglion cells and their axons degenerate. Degradation of axonal microtubules is thought to play a critical role in the pathogenesis, but the mechanism is unknown. Here we investigate whether microtubule disruption in glaucoma can be alleviated by metabolic rescue. The integrity of axonal microtubules and the morphology of the retinal nerve fibers were evaluated by second-harmonic generation microscopy in a mouse model of glaucoma, DBA/2J, which received a dietary supplement of nicotinamide (NAM) for reducing metabolic stress. It was compared with control DBA/2J, which did not receive NAM, and non-glaucomatous DBA/2J-Gpnmb+. We found that the morphology of the retinal nerve fibers, but not axonal microtubules, are significantly protected by NAM. The decoupling is analogous to microtubule deficit, a glaucoma pathology in which axonal microtubules exhibit rapid degradation compared to the morphology of the retinal nerve fibers. Understanding microtubule deficit could provide insights into the divergent responses to NAM. From co-registered images of second-harmonic generation and immunofluorescence, it was determined that microtubule deficit was not due to a shortage of tubulins. Furthermore, microtubule deficit colocalized with the sectors in which the retinal ganglion cells were disconnected from the brain, suggesting that microtubule disruption is associated with axonal transport deficit in glaucoma. Together, our data suggests significant role axonal microtubules play in glaucomatous degeneration, offering a new opportunity for neuroprotection.


Subject(s)
Disease Models, Animal , Glaucoma , Mice, Inbred DBA , Microtubules , Niacinamide , Retinal Ganglion Cells , Animals , Glaucoma/pathology , Glaucoma/metabolism , Glaucoma/drug therapy , Niacinamide/pharmacology , Niacinamide/therapeutic use , Mice , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Microtubules/drug effects , Microtubules/metabolism , Axons/drug effects , Axons/metabolism , Axons/pathology , Microscopy/methods , Nerve Fibers/drug effects , Nerve Fibers/pathology , Nerve Fibers/metabolism
3.
Int Ophthalmol ; 44(1): 370, 2024 Sep 06.
Article in English | MEDLINE | ID: mdl-39237823

ABSTRACT

PURPOSE: To assess structural (optical coherence tomography, fundus autofluorescence) and functional (contrast sensitivity and visual field) test results which were used for detecting early retinal changes in patients using oral hydroxychloroquine. METHODS: Patients using oral hydroxychloroquine for at least one year were divided into two groups according to the duration of drug use. Groups 1 and 2 consisted of patients with drug use for more than 5 years and 1-5 years, respectively. The drug-using groups were compared with the control group. The mean retinal nerve fiber layer (RNFL), central macular thickness (CMT), ganglion cell-inner plexiform layer (GC-IPL), static 10-2 visual field, fundus autofluorescence (FAF) imaging, and contrast sensitivity tests were performed and statistically compared between groups. RESULTS: Median and temporal quadrant RNFL thicknesses were found to be statistically significantly lower in the drug groups. In the drug groups, the GC-IPL sectoral and mean thicknesses were found to be statistically lower in all quadrants. Central macular thickness was also found to be similar in all three groups. There was no significant difference between the groups in visual field parameters. Macular FAF images were significantly higher in the drug users, but there was no significant difference between the three groups in foveal FAF images. Contrast sensitivity measurements were significantly lower in the drug groups than in the control group at all spatial frequencies except 6 and 18 cycles/degree. CONCLUSIONS: The combined use of structural and functional tests in patients using hydroxychloroquine provides useful information in detecting early retinal changes.


Subject(s)
Antirheumatic Agents , Contrast Sensitivity , Early Diagnosis , Fluorescein Angiography , Hydroxychloroquine , Macula Lutea , Retinal Diseases , Retinal Ganglion Cells , Tomography, Optical Coherence , Visual Fields , Humans , Hydroxychloroquine/adverse effects , Tomography, Optical Coherence/methods , Female , Male , Visual Fields/physiology , Visual Fields/drug effects , Middle Aged , Antirheumatic Agents/adverse effects , Retinal Diseases/chemically induced , Retinal Diseases/diagnosis , Macula Lutea/drug effects , Macula Lutea/pathology , Macula Lutea/diagnostic imaging , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/drug effects , Contrast Sensitivity/physiology , Contrast Sensitivity/drug effects , Fluorescein Angiography/methods , Adult , Nerve Fibers/pathology , Nerve Fibers/drug effects , Visual Acuity , Visual Field Tests/methods , Aged
4.
Neuromolecular Med ; 26(1): 37, 2024 Sep 12.
Article in English | MEDLINE | ID: mdl-39266914

ABSTRACT

As the primary connection between the eye and brain, the optic nerve plays a pivotal role in visual information transmission. Injuries to the optic nerve can occur for various reasons, including trauma, glaucoma, and neurodegenerative diseases. Retinal ganglion cells (RGCs), a type of neurons that extend axons through the optic nerve, can rapidly respond to injury and initiate cell death. Additionally, following optic nerve injury microglia, which serve as markers of neuroinflammation, transition from a resting state to an activated state. The phosphorylation of collapsin response mediator protein2 (CRMP2) in the semaphorin 3A (Sema3A) signalling pathway affects several processes, including axon guidance and neuron regeneration. In this study, we used an optic nerve crush (ONC) mouse model to investigate the effects of suppressing CRMP2 phosphorylation on microglia activation. We found that CRMP2 phosphorylation inhibitor suppressed RGCs loss and promoted neuronal regeneration following ONC. In addition, CRMP2 S522A mutant (CRMP2 KI) mice exhibited decreased microglial activation in both the retina and optic nerve following ONC. These results suggest that inhibiting the phosphorylation of CRMP2 can alleviate the loss of RGCs and microglial activation after optic nerve injury, providing insight into the development of treatments for optical neuropathies and neurodegenerative diseases.


Subject(s)
Intercellular Signaling Peptides and Proteins , Microglia , Nerve Regeneration , Nerve Tissue Proteins , Optic Nerve Injuries , Optic Nerve , Retina , Retinal Ganglion Cells , Semaphorin-3A , Animals , Optic Nerve Injuries/physiopathology , Optic Nerve Injuries/metabolism , Optic Nerve Injuries/drug therapy , Microglia/metabolism , Microglia/drug effects , Phosphorylation , Mice , Nerve Regeneration/drug effects , Intercellular Signaling Peptides and Proteins/genetics , Nerve Tissue Proteins/genetics , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/metabolism , Optic Nerve/metabolism , Semaphorin-3A/genetics , Semaphorin-3A/metabolism , Retina/drug effects , Retina/metabolism , Nerve Crush , Mice, Inbred C57BL , Male , Disease Models, Animal , Mice, Transgenic
5.
Acta Neuropathol Commun ; 12(1): 149, 2024 Sep 12.
Article in English | MEDLINE | ID: mdl-39267142

ABSTRACT

Motor neuron loss is well recognized in amyotrophic lateral sclerosis (ALS), but research on retinal ganglion cells (RGCs) is limited. Ocular symptoms are generally not considered classic ALS symptoms, although RGCs and spinal motor neurons share certain cell pathologies, including hallmark signs of glutamate neurotoxicity, which may be triggered by activation of extrasynaptic NMDA receptors (NMDARs). To explore potential novel strategies to prevent ALS-associated death of RGCs, we utilized inhibition of the TwinF interface, a new pharmacological principle that detoxifies extrasynaptic NMDARs by disrupting the NMDAR/TRPM4 death signaling complex. Using the ALS mouse model SOD1G93A, we found that the small molecule TwinF interface inhibitor FP802 prevents the loss of RGCs, improves pattern electroretinogram (pERG) performance, increases the retinal expression of Bdnf, and restores the retinal expression of the immediate early genes, Inhibin beta A and Npas4. Thus, FP802 not only prevents, as recently described, death of spinal motor neurons in SOD1G93A mice, but it also mitigates ALS-associated retinal damage. TwinF interface inhibitors have great potential for alleviating neuro-ophthalmologic symptoms in ALS patients and offer a promising new avenue for therapeutic intervention.


Subject(s)
Amyotrophic Lateral Sclerosis , Disease Models, Animal , Mice, Transgenic , Retinal Ganglion Cells , Animals , Amyotrophic Lateral Sclerosis/pathology , Amyotrophic Lateral Sclerosis/metabolism , Amyotrophic Lateral Sclerosis/drug therapy , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Mice , Electroretinography , Mice, Inbred C57BL , Neuroprotective Agents/pharmacology , Brain-Derived Neurotrophic Factor/metabolism , Humans , Basic Helix-Loop-Helix Transcription Factors/metabolism
6.
J Invest Surg ; 37(1): 2389379, 2024 Dec.
Article in English | MEDLINE | ID: mdl-39164010

ABSTRACT

OBJECTIVE: This investigation was to determine the relationship between changes in the expression levels of miR-134 and the E2F transcription factor 6 (E2F6) in mediating control of apoptosis in N-methyl-D-aspartate (NMDA)-induced glaucomatous mice. METHODS: Morphological and structural changes were quantitatively analyzed along with apoptosis in the retinal ganglion cell (RGC) layer, internal plexiform layer and RGCs. Glaucomatous RGCs were transfected, and cell viability and apoptosis were examined. The targeting relationship between miR-134 and E2F6 was analyzed, as well as their expression pattern. RESULTS: Intravitreal injection of NMDA induced a significant reduction in the number of RGCs and thinning of IPL thickness. miR-134 was highly expressed and E2F6 was lowly expressed in glaucoma mice. Suppression of miR-134 or E2F6 overexpression inhibited apoptosis in the glaucomatous RGCs and instead their proliferative activity. MiR-134 targeted inhibition of E2F6 expression. Suppressing rises in E2F6 expression reduced the interfering effect of miR-134 on glaucomatous RGC development. CONCLUSION: Depleting miR134 expression increases, in turn, E2F6 expression levels and in turn reduces glaucomatous RGC apoptosis expression.


Subject(s)
Apoptosis , E2F6 Transcription Factor , Glaucoma , MicroRNAs , N-Methylaspartate , Retinal Ganglion Cells , Animals , MicroRNAs/genetics , MicroRNAs/metabolism , Apoptosis/drug effects , Apoptosis/genetics , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/drug effects , Glaucoma/genetics , Glaucoma/pathology , Glaucoma/metabolism , Glaucoma/chemically induced , N-Methylaspartate/toxicity , Mice , E2F6 Transcription Factor/genetics , E2F6 Transcription Factor/metabolism , Mice, Inbred C57BL , Disease Models, Animal , Male , Intravitreal Injections , Cell Survival/drug effects
7.
Sci Adv ; 10(32): eadl5722, 2024 Aug 09.
Article in English | MEDLINE | ID: mdl-39110798

ABSTRACT

Dendrite pathology and synaptic loss result in neural circuit dysfunction, a common feature of neurodegenerative diseases. There is a lack of strategies that target dendritic and synaptic regeneration to promote neurorecovery. We show that daily human recombinant insulin eye drops stimulate retinal ganglion cell (RGC) dendrite and synapse regeneration during ocular hypertension, a risk factor to develop glaucoma. We demonstrate that the ribosomal protein p70S6 kinase (S6K) is essential for insulin-dependent dendritic regrowth. Furthermore, S6K phosphorylation of the stress-activated protein kinase-interacting protein 1 (SIN1), a link between the mammalian target of rapamycin complexes 1 and 2 (mTORC1/2), is required for insulin-induced dendritic regeneration. Using two-photon microscopy live retinal imaging, we show that insulin rescues single-RGC light-evoked calcium (Ca2+) dynamics. We further demonstrate that insulin enhances neuronal survival and retina-brain connectivity leading to improved optomotor reflex-elicited behaviors. Our data support that insulin is a compelling pro-regenerative strategy with potential clinical implications for the treatment and management of glaucoma.


Subject(s)
Glaucoma , Insulin , Retinal Ganglion Cells , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/drug effects , Glaucoma/drug therapy , Glaucoma/metabolism , Glaucoma/pathology , Insulin/metabolism , Insulin/pharmacology , Animals , Humans , Mice , Disease Models, Animal , Dendrites/metabolism , Dendrites/drug effects , Synapses/metabolism , Synapses/drug effects , Calcium/metabolism
8.
Invest Ophthalmol Vis Sci ; 65(10): 34, 2024 Aug 01.
Article in English | MEDLINE | ID: mdl-39186263

ABSTRACT

Purpose: In response to hypoxia, sympathetic fibers to the retina activate ß-adrenoceptors (ß-ARs) that play an important role in the regulation of vascular and neuronal functions. We investigated the role of ß3-AR using the mouse model of oxygen-induced retinopathy (OIR). Methods: Mouse pups were exposed to 75% oxygen at postnatal day 7 (PD7) followed by a return to room air at PD12. The ß3-AR preferential agonist BRL37344 was subcutaneously administered once daily at different times after the return to room air. At PD17, the OIR mice underwent flash and pattern electroretinogram. After sacrifice, retinal wholemounts were used for vessel staining or immunohistochemistry for astrocytes, Müller cells, or retinal ganglion cells (RGCs). In retinal homogenates, the levels of markers associated with neovascularization (NV), the blood-retinal barrier (BRB), or astrocytes were determined by western blot, and quantitative reverse-transcription polymerase chain reaction was used to assess ß3-AR messenger. Administration of the ß3-AR antagonist SR59230A was performed to verify BRL37344 selectivity. Results: ß3-AR expression is upregulated in response to hypoxia, but its increase is prevented by BRL37344, which counteracts NV by inhibiting the pro-angiogenic pathway, activating the anti-angiogenic pathway, recovering BRB-associated markers, triggering nitric oxide production, and favoring revascularization of the central retina through recovered density of astrocytes that ultimately counteracts NV in the midperiphery. Vasculature rescue prevents dysfunctional retinal activity and counteracts OIR-associated retinal ganglion cell loss. Conclusions: ß3-AR has emerged as a crucial intermediary in hypoxia-dependent NV, suggesting a role of ß3-AR agonists in the treatment of proliferative retinopathies.


Subject(s)
Adrenergic beta-3 Receptor Agonists , Disease Models, Animal , Electroretinography , Mice, Inbred C57BL , Oxygen , Receptors, Adrenergic, beta-3 , Retinal Neovascularization , Animals , Mice , Retinal Neovascularization/metabolism , Retinal Neovascularization/prevention & control , Retinal Neovascularization/pathology , Oxygen/toxicity , Adrenergic beta-3 Receptor Agonists/pharmacology , Receptors, Adrenergic, beta-3/metabolism , Animals, Newborn , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/metabolism , Ethanolamines/pharmacology , Retinal Vessels/drug effects , Retinopathy of Prematurity/metabolism , Retinopathy of Prematurity/drug therapy , Astrocytes/metabolism , Astrocytes/drug effects , Immunohistochemistry , Angiogenesis
9.
Sci Adv ; 10(35): eadk4062, 2024 Aug 30.
Article in English | MEDLINE | ID: mdl-39196935

ABSTRACT

The mammalian retina is considered an autonomous circuit, yet work dating back to Ramon y Cajal indicates that it receives inputs from the brain. How such inputs affect retinal processing has remained unknown. We confirmed brain-to-retina projections of histaminergic neurons from the mouse hypothalamus. Histamine application ex vivo altered the activity of various retinal ganglion cells (RGCs), including direction-selective RGCs that gained responses to high motion velocities. These results were reproduced in vivo with optic tract recordings where histaminergic retinopetal axons were activated chemogenetically. Such changes could improve vision of fast-moving objects (e.g., while running), which fits with the known increased activity of histaminergic neurons during arousal. An antihistamine drug reduced optomotor responses to high-speed moving stimuli in freely moving mice. In humans, the same antihistamine nonuniformly modulated visual sensitivity across the visual field, indicating an evolutionary conserved function of the histaminergic system. Our findings expose a previously unappreciated role for brain-to-retina projections in modulating retinal function.


Subject(s)
Histamine , Hypothalamus , Retina , Retinal Ganglion Cells , Animals , Histamine/pharmacology , Histamine/metabolism , Hypothalamus/metabolism , Hypothalamus/cytology , Hypothalamus/physiology , Mice , Retina/metabolism , Retina/physiology , Retina/drug effects , Retina/cytology , Retinal Ganglion Cells/physiology , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/metabolism , Neurons/metabolism , Neurons/physiology , Neurons/drug effects , Humans , Mice, Inbred C57BL , Visual Pathways/drug effects , Visual Pathways/physiology
10.
Pathol Res Pract ; 262: 155552, 2024 Oct.
Article in English | MEDLINE | ID: mdl-39180803

ABSTRACT

Glaucoma is a degenerative disease characterized by retinal ganglion cell (RGC) death and visual impairment caused by elevated intraocular pressure (IOP). Elevated IOP can activate microglia, which participate in ganglion cell injury. Based on the study of caveolin-1 (Cav-1) in glaucoma, we aimed to explore the effect and mechanism of Cav-1 on RGC apoptosis in mice with acute ocular hypertension (AOH). AOH mice were established, and Cav-1 was intravitreally injected. Retinal microglia and RGCs were isolated from neonatal mice. TUNEL staining, hematoxylin-eosin staining, immunohistochemistry, flow cytometry, PCR and western blotting were used to observe the effect of Cav-1 on RGCs and mouse retinas. The thickness of the whole retina and the inner retinal sublayer decreased significantly, retinal cell apoptosis increased after AOH injury, and Cav-1 treatment reversed the effect of AOH injury. In addition, Cav-1 treatment promoted the conversion of proinflammatory M1 microglia to anti-inflammatory M2 microglia. Microglia and RGCs were isolated from neonatal mice. Cav-1 protects RGCs from OGD/R-induced injury by changing the polarization status of retinal microglia in vitro. Further studies revealed that Cav-1 activated the Akt/PTEN signaling pathway and inhibited TLR4. Our study provides evidence that Cav-1 may be a promising therapeutic target for glaucoma.


Subject(s)
Caveolin 1 , Glaucoma , PTEN Phosphohydrolase , Proto-Oncogene Proteins c-akt , Retinal Ganglion Cells , Signal Transduction , Toll-Like Receptor 4 , Animals , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/drug effects , Caveolin 1/metabolism , Signal Transduction/physiology , PTEN Phosphohydrolase/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Mice , Glaucoma/metabolism , Glaucoma/pathology , Toll-Like Receptor 4/metabolism , Mice, Inbred C57BL , Apoptosis/drug effects , Microglia/metabolism , Microglia/pathology , Disease Models, Animal
11.
J Transl Med ; 22(1): 727, 2024 Aug 05.
Article in English | MEDLINE | ID: mdl-39103918

ABSTRACT

BACKGROUND: Vascular dysregulation is one of the major risk factors of glaucoma, and endothelin-1 (ET-1) may have a role in the pathogenesis of vascular-related glaucoma. Fruit extract from Lycium Barbarum (LB) exhibits anti-ageing and multitarget mechanisms in protecting retinal ganglion cells (RGC) in various animal models. To investigate the therapeutic efficacy of LB glycoproteins (LbGP) in ET-1 induced RGC degeneration, LbGP was applied under pre- and posttreatment conditions to an ET-1 mouse model. Retina structural and functional outcomes were characterised using clinical-based techniques. METHODS: Adult C57BL/6 mice were randomly allocated into four experimental groups, namely vehicle control (n = 9), LbGP-Pretreatment (n = 8), LbGP-Posttreatment (day 1) (n = 8) and LbGP-Posttreatment (day 5) (n = 7). Oral administration of LbGP 1 mg/Kg or PBS for vehicle control was given once daily. Pre- and posttreatment (day 1 or 5) were commenced at 1 week before and 1 or 5 days after intravitreal injections, respectively, and were continued until postinjection day 28. Effects of treatment on retinal structure and functions were evaluated using optical coherence tomography (OCT), doppler OCT and electroretinogram measurements at baseline, post-injection days 10 and 28. RGC survival was evaluated by using RBPMS immunostaining on retinal wholemounts. RESULTS: ET-1 injection in vehicle control induced transient reductions in arterial flow and retinal functions, leading to significant RNFL thinning and RGC loss at day 28. Although ET-1 induced a transient loss in blood flow or retinal functions in all LbGP groups, LbGP treatments facilitated better restoration of retinal flow and retinal functions as compared with the vehicle control. Also, all three LbGP treatment groups (i.e. pre- and posttreatments from days 1 or 5) significantly preserved thRNFL thickness and RGC densities. No significant difference in protective effects was observed among the three LbGP treatment groups. CONCLUSION: LbGP demonstrated neuroprotective effects in a mouse model of ET-1 induced RGC degeneration, with treatment applied either as a pretreatment, immediate or delayed posttreatment. LbGP treatment promoted a better restoration of retinal blood flow, and protected the RNFL, RGC density and retinal functions. This study showed the translational potential of LB as complementary treatment for glaucoma management.


Subject(s)
Endothelin-1 , Mice, Inbred C57BL , Neuroprotection , Retinal Ganglion Cells , Animals , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Endothelin-1/metabolism , Neuroprotection/drug effects , Electroretinography , Lycium/chemistry , Drugs, Chinese Herbal/pharmacology , Drugs, Chinese Herbal/therapeutic use , Neuroprotective Agents/pharmacology , Neuroprotective Agents/therapeutic use , Retinal Degeneration/drug therapy , Retinal Degeneration/pathology , Tomography, Optical Coherence , Male , Mice , Nerve Degeneration/pathology , Nerve Degeneration/drug therapy
12.
Exp Eye Res ; 246: 110017, 2024 Sep.
Article in English | MEDLINE | ID: mdl-39097072

ABSTRACT

Loss of retinal ganglion cells (RGCs) is central to the pathogenesis of optic neuropathies such as glaucoma. Increased RGC cAMP signaling is neuroprotective. We have shown that displacement of the cAMP-specific phosphodiesterase PDE4D3 from an RGC perinuclear compartment by expression of the modified PDE4D3 N-terminal peptide 4D3(E) increases perinuclear cAMP and protein kinase A activity in cultured neurons and in vivo RGC survival after optic nerve crush (ONC) injury. To explore mechanisms by which PDE4D3 displacement promotes neuroprotection, in this study mice intravitreally injected with an adeno-associated virus to express an mCherry-tagged 4D3(E) peptide were subjected to ONC injury and analyzed by single cell RNA-sequencing (scRNA-seq). 4D3(E)-mCherry expression was associated with an attenuation of injury-induced changes in gene expression, thereby supporting the hypothesis that enhanced perinuclear PKA signaling promotes neuroprotective RGC gene expression.


Subject(s)
Mice, Inbred C57BL , Optic Nerve Injuries , Retinal Ganglion Cells , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/drug effects , Animals , Mice , Optic Nerve Injuries/metabolism , Optic Nerve Injuries/drug therapy , Optic Nerve Injuries/genetics , Gene Expression Regulation , Disease Models, Animal , Cyclic Nucleotide Phosphodiesterases, Type 4/metabolism , Cyclic Nucleotide Phosphodiesterases, Type 4/genetics , Nerve Crush , Cell Survival , Intravitreal Injections , Signal Transduction , Cyclic AMP-Dependent Protein Kinases/metabolism , Male , Cells, Cultured
13.
PLoS One ; 19(8): e0308671, 2024.
Article in English | MEDLINE | ID: mdl-39116180

ABSTRACT

Glaucoma is a multifactorial optic neuropathy that primarily affecting retinal ganglion cells (RGC). Brimonidine is an intraocular pressure-lowering drug with reported neuroprotective properties. This study aimed to compare the neuroprotective effects of topical and intraperitoneal (IP) brimonidine on RGCs from different retinal segments in a murine optic nerve crush (ONC) model. METHODS: forty-one Balb/c mice underwent unilateral ONC and were divided into three study groups: fifteen animals received saline drops twice per day and two additional IP injections of saline; fourteen mice received brimonidine drops twice per day; and 12 mice received brimonidine eye drops twice per day and two additional IP brimonidine injections. Animals were sacrificed seven days post-ONC, and immunohistochemical staining of retinal whole mounts was performed using neuronal NeuN and GFAP staining. Microscopic pictures of the central, middle, and peripheral regions of the retina were taken. The density of the retinal cells was assessed. RESULTS: The total RGC density after ONC and RGC densities in all retinal eccentricities were significantly higher in the brimonidine eye drop and IP combination treatment group than in the saline drop + saline IP, and brimonidine drop treatment groups. CONCLUSIONS: brimonidine eye drops supplemented with IP brimonidine injections improved RGC survival in a preclinical model of ONC.


Subject(s)
Brimonidine Tartrate , Disease Models, Animal , Mice, Inbred BALB C , Nerve Crush , Neuroprotective Agents , Optic Nerve Injuries , Retinal Ganglion Cells , Animals , Brimonidine Tartrate/administration & dosage , Brimonidine Tartrate/pharmacology , Mice , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Neuroprotective Agents/pharmacology , Neuroprotective Agents/administration & dosage , Optic Nerve Injuries/drug therapy , Optic Nerve Injuries/pathology , Administration, Topical , Ophthalmic Solutions , Male , Injections, Intraperitoneal
14.
Sci Adv ; 10(31): eado0866, 2024 Aug 02.
Article in English | MEDLINE | ID: mdl-39093964

ABSTRACT

As part of the central nervous system, the optic nerve, composed of axons from retinal ganglion cells (RGCs), generally fails to regenerate on its own when injured in adult mammals. An innovative approach to promoting optic nerve regeneration involves manipulating the interactions between amacrine cells (ACs) and RGCs. Here, we identified a unique AC subtype, dopaminergic ACs (DACs), that responded early after optic nerve crush by down-regulating neuronal activity and reducing retinal dopamine (DA) release. Activating DACs or augmenting DA release with levodopa demonstrated neuroprotective effects and modestly enhanced axon regeneration. Within this context, we pinpointed the DA receptor D1 (DRD1) as a critical mediator of DAC-derived DA and showed that RGC-specific Drd1 overexpression effectively overcame subtype-specific barriers to regeneration. This strategy markedly boosted RGC survival and axon regeneration after crush and preserved vision in a glaucoma model. This study unveils the crucial role of DAC-derived DA signaling in optic nerve regeneration, holding promise for therapeutic insights into neural repair.


Subject(s)
Amacrine Cells , Dopamine , Nerve Regeneration , Optic Nerve , Retinal Ganglion Cells , Signal Transduction , Animals , Amacrine Cells/metabolism , Dopamine/metabolism , Nerve Regeneration/drug effects , Optic Nerve/metabolism , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/drug effects , Optic Nerve Injuries/metabolism , Optic Nerve Injuries/drug therapy , Optic Nerve Injuries/pathology , Mice , Axons/metabolism , Axons/physiology , Receptors, Dopamine D1/metabolism , Vision, Ocular/physiology , Disease Models, Animal
15.
Acta Neuropathol Commun ; 12(1): 137, 2024 Aug 23.
Article in English | MEDLINE | ID: mdl-39180087

ABSTRACT

A compromised capacity to maintain NAD pools is recognized as a key underlying pathophysiological feature of neurodegenerative diseases. NAD acts as a substrate in major cell functions including mitochondrial homeostasis, cell signalling, axonal transport, axon/Wallerian degeneration, and neuronal energy supply. Dendritic degeneration is an early marker of neuronal stress and precedes cell loss. However, little is known about dendritic structural preservation in pathologic environments and remodelling in mature neurons. Retinal ganglion cell dendritic atrophy is an early pathological feature in animal models of the disease and has been demonstrated in port-mortem human glaucoma samples. Here we report that a nicotinamide (a precursor to NAD through the NAD salvage pathway) enriched diet provides robust retinal ganglion cell dendritic protection and preserves dendritic structure in a rat model of experimental glaucoma. Metabolomic analysis of optic nerve samples from the same animals demonstrates that nicotinamide provides robust metabolic neuroprotection in glaucoma. Advances in our understanding of retinal ganglion cell metabolic profiles shed light on the energetic shift that triggers early neuronal changes in neurodegenerative diseases. As nicotinamide can improve visual function short term in existing glaucoma patients, we hypothesize that a portion of this visual recovery may be due to dendritic preservation in stressed, but not yet fully degenerated, retinal ganglion cells.


Subject(s)
Disease Models, Animal , Glaucoma , Neuroprotective Agents , Niacinamide , Retinal Ganglion Cells , Animals , Niacinamide/pharmacology , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Retinal Ganglion Cells/metabolism , Glaucoma/metabolism , Glaucoma/pathology , Neuroprotective Agents/pharmacology , Rats , Dose-Response Relationship, Drug , Male , Administration, Oral , Optic Nerve/drug effects , Optic Nerve/pathology , Optic Nerve/metabolism , Neuroprotection/drug effects , Neuroprotection/physiology , Dendrites/drug effects , Dendrites/pathology , Dendrites/metabolism , Vitamin B Complex/pharmacology , Vitamin B Complex/administration & dosage
16.
Biomed Pharmacother ; 178: 117270, 2024 Sep.
Article in English | MEDLINE | ID: mdl-39126773

ABSTRACT

The blood supply in the retina ensures photoreceptor function and maintains regular vision. Leber's hereditary optic neuropathy (LHON), caused by the mitochondrial DNA mutations that deteriorate complex I activity, is characterized by progressive vision loss. Although some reports indicated retinal vasculature abnormalities as one of the comorbidities in LHON, the paracrine influence of LHON-affected retinal ganglion cells (RGCs) on vascular endothelial cell physiology remains unclear. To address this, we established an in vitro model of mitochondrial complex I deficiency using induced pluripotent stem cell-derived RGCs (iPSC-RGCs) treated with a mitochondrial complex I inhibitor rotenone (Rot) to recapitulate LHON pathologies. The secretomes from Rot-treated iPSC-RGCs (Rot-iPSC-RGCs) were collected, and their treatment effect on human umbilical vein endothelial cells (HUVECs) was studied. Rot induced LHON-like characteristics in iPSC-RGCs, including decreased mitochondrial complex I activity and membrane potential, and increased mitochondrial reactive oxygen species (ROS) and apoptosis, leading to mitochondrial dysfunction. When HUVECs were exposed to conditioned media (CM) from Rot-iPSC-RGCs, the angiogenesis of HUVECs was suppressed compared to those treated with CM from control iPSC-RGCs (Ctrl-iPSC-RGCs). Angiogenesis-related proteins were altered in the secretomes from Rot-iPSC-RGC-derived CM, particularly angiopoietin, MMP-9, uPA, collagen XVIII, and VEGF were reduced. Notably, GeneMANIA analysis indicated that VEGFA emerged as the pivotal angiogenesis-related protein among the identified proteins secreted by health iPSC-RGCs but reduced in the secretomes from Rot-iPSC-RGCs. Quantitative real-time PCR and western blots confirmed the reduction of VEGFA at both transcription and translation levels, respectively. Our study reveals that Rot-iPSC-RGCs establish a microenvironment to diminish the angiogenic potential of vascular cells nearby, shedding light on the paracrine regulation of LHON-affected RGCs on retinal vasculature.


Subject(s)
Human Umbilical Vein Endothelial Cells , Induced Pluripotent Stem Cells , Optic Atrophy, Hereditary, Leber , Retinal Ganglion Cells , Humans , Optic Atrophy, Hereditary, Leber/metabolism , Optic Atrophy, Hereditary, Leber/pathology , Optic Atrophy, Hereditary, Leber/genetics , Induced Pluripotent Stem Cells/drug effects , Induced Pluripotent Stem Cells/metabolism , Human Umbilical Vein Endothelial Cells/metabolism , Human Umbilical Vein Endothelial Cells/drug effects , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/pathology , Mitochondria/drug effects , Mitochondria/metabolism , Phenotype , Reactive Oxygen Species/metabolism , Rotenone/pharmacology , Culture Media, Conditioned/pharmacology , Apoptosis/drug effects , Electron Transport Complex I/metabolism , Membrane Potential, Mitochondrial/drug effects , Neovascularization, Pathologic/metabolism , Angiogenesis
17.
Neuroscience ; 553: 145-159, 2024 Aug 16.
Article in English | MEDLINE | ID: mdl-38992567

ABSTRACT

Glutamate excitotoxicity is involved in retinal ganglion cell (RGC) death in various retinal degenerative diseases, including ischemia-reperfusion injury and glaucoma. Excitotoxic RGC death is caused by both direct damage to RGCs and indirect damage through neuroinflammation of retinal glial cells. Omidenepag (OMD), a novel E prostanoid receptor 2 (EP2) agonist, is a recently approved intraocular pressure-lowering drug. The second messenger of EP2 is cyclic adenosine monophosphate (cAMP), which activates protein kinase A (PKA) and exchange protein directly activated by cAMP (Epac). In this study, we investigated the neuroprotective effects of OMD on excitotoxic RGC death by focusing on differences in cAMP downstream signaling from the perspective of glia-neuron interactions. We established a glutamate excitotoxicity model in vitro and NMDA intravitreal injection model in vivo. In vitro, rat primary RGCs were used in an RGC survival rate assay. MG5 cells (mouse microglial cell line) and A1 cells (astrocyte cell line) were used for immunocytochemistry and Western blotting to evaluate the expressions of COX-1/2, PKA, Epac1/2, pCREB, cleaved caspase-3, inflammatory cytokines, and neurotrophic factors. Mouse retinal specimens underwent hematoxylin and eosin staining, flat-mounted retina examination, and immunohistochemistry. OMD significantly suppressed excitotoxic RGC death, cleaved caspase-3 expression, and activated glia both in vitro and in vivo. Moreover, it inhibited Epac1 and inflammatory cytokine expression and promoted COX-2, pCREB, and neurotrophic factor expression. OMD may have neuroprotective effects through inhibition of the Epac pathway and promotion of the COX-2-EP2-cAMP-PKA pathway by modulating glia-neuron interaction.


Subject(s)
Cyclic AMP-Dependent Protein Kinases , Cyclic AMP , Cyclooxygenase 2 , Neuroglia , Neuroprotective Agents , Retinal Ganglion Cells , Animals , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/metabolism , Neuroprotective Agents/pharmacology , Cyclic AMP-Dependent Protein Kinases/metabolism , Cyclooxygenase 2/metabolism , Cyclic AMP/metabolism , Mice , Neuroglia/drug effects , Neuroglia/metabolism , Receptors, Prostaglandin E, EP2 Subtype/metabolism , Receptors, Prostaglandin E, EP2 Subtype/antagonists & inhibitors , Receptors, Prostaglandin E, EP2 Subtype/agonists , Cell Death/drug effects , Cell Death/physiology , Signal Transduction/drug effects , Signal Transduction/physiology , Guanine Nucleotide Exchange Factors/metabolism , Rats, Sprague-Dawley , Rats , Glutamic Acid/metabolism , Glutamic Acid/toxicity , Mice, Inbred C57BL , Male , N-Methylaspartate/pharmacology , N-Methylaspartate/toxicity , Neurons/drug effects , Neurons/metabolism
18.
J Neurophysiol ; 132(2): 501-513, 2024 Aug 01.
Article in English | MEDLINE | ID: mdl-38958282

ABSTRACT

Neuromodulation in the retina is crucial for effective processing of retinal signal at different levels of illuminance. Intrinsically photosensitive retinal ganglion cells (ipRGCs), the neurons that drive nonimage-forming visual functions, express a variety of neuromodulatory receptors that tune intrinsic excitability as well as synaptic inputs. Past research has examined actions of neuromodulators on light responsiveness of ipRGCs, but less is known about how neuromodulation affects synaptic currents in ipRGCs. To better understand how neuromodulators affect synaptic processing in ipRGC, we examine actions of opioid and dopamine agonists have on inhibitory synaptic currents in ipRGCs. Although µ-opioid receptor (MOR) activation had no effect on γ-aminobutyric acid (GABA) currents, dopamine [via the D1-type dopamine receptor (D1R)]) amplified GABAergic currents in a subset of ipRGCs. Furthermore, this D1R-mediated facilitation of the GABA conductance in ipRGCs was mediated by a cAMP/PKA-dependent mechanism. Taken together, these findings reinforce the idea that dopamine's modulatory role in retinal adaptation affects both nonimage-forming and image-forming visual functions.NEW & NOTEWORTHY Neuromodulators such as dopamine are important regulators of retinal function. Here, we demonstrate that dopamine increases inhibitory inputs to intrinsically photosensitive retinal ganglion cells (ipRGCs), in addition to its previously established effect on intrinsic light responsiveness. This indicates that dopamine, in addition to its ability to intrinsically modulate ipRGC activity, can also affect synaptic inputs to ipRGCs, thereby tuning retina circuits involved in nonimage-forming visual functions.


Subject(s)
Dopamine , Receptors, GABA-A , Retinal Ganglion Cells , Animals , Retinal Ganglion Cells/physiology , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/metabolism , Dopamine/metabolism , Dopamine/pharmacology , Receptors, GABA-A/metabolism , Mice , Receptors, Dopamine D1/metabolism , Receptors, Dopamine D1/antagonists & inhibitors , Mice, Inbred C57BL , Receptors, Opioid, mu/metabolism , Male , Inhibitory Postsynaptic Potentials/physiology , Inhibitory Postsynaptic Potentials/drug effects , Female , Dopamine Agonists/pharmacology
19.
Neuroreport ; 35(13): 822-831, 2024 Sep 04.
Article in English | MEDLINE | ID: mdl-38973496

ABSTRACT

The aim of this study was to explore the neuroprotective effects of the P2X7 receptor antagonist A740003 on retinal ganglion cells (RGCs) in chronic intraocular hypertension (COH) experimental glaucoma mouse model. Bioinformatics was used to analyze the glaucoma-related genes. Western blot, real-time fluorescence quantitative PCR, and immunofluorescence staining techniques were employed to explore the mechanisms underlying the neuroprotective effects of A740003 on RGCs in COH retinas. Bioinformatic analysis revealed that oxidative stress, neuroinflammation, and cell apoptosis were highly related to the pathogenesis of glaucoma. In COH retinas, intraocular pressure elevation significantly increased the levels of translocator protein, a marker of microglial activation, which could be reversed by intravitreal preinjection of A740003. A740003 also suppressed the increased mRNA levels of proinflammatory cytokines interleukin (IL) 1ß and tumor necrosis factor α in COH retinas. In addition, although the mRNA levels of anti-inflammatory cytokine IL-4 and IL-10 were kept unchanged in COH retinas, administration of A740003 could increase their levels. The mRNA and protein levels of Bax and cleaved caspase-3 were increased in COH retinas, which could be partially reversed by A740003, while the levels of Bcl-2 kept unchanged in COH retinas with or without the injections of A740003. Furthermore, A740003 partially attenuated the reduction in the numbers of Brn-3a-positive RGCs in COH mice. A740003 could provide neuroprotective roles on RGCs by inhibiting the microglia activation, attenuating the retinal inflammatory response, reducing the apoptosis of RGCs, and enhancing the survival of RGCs in COH experimental glaucoma.


Subject(s)
Glaucoma , Mice, Inbred C57BL , Neuroprotective Agents , Purinergic P2X Receptor Antagonists , Retinal Ganglion Cells , Animals , Retinal Ganglion Cells/drug effects , Retinal Ganglion Cells/metabolism , Retinal Ganglion Cells/pathology , Glaucoma/drug therapy , Glaucoma/metabolism , Neuroprotective Agents/pharmacology , Mice , Purinergic P2X Receptor Antagonists/pharmacology , Disease Models, Animal , Male , Benzopyrans/pharmacology , Neuroprotection/drug effects , Apoptosis/drug effects , Receptors, Purinergic P2X7/metabolism , Receptors, Purinergic P2X7/drug effects , Ocular Hypertension/drug therapy , Ocular Hypertension/metabolism , Intraocular Pressure/drug effects , Carbazoles
20.
J Neurosci ; 44(35)2024 Aug 28.
Article in English | MEDLINE | ID: mdl-39060177

ABSTRACT

In retinitis pigmentosa (RP), rod and cone photoreceptors degenerate, depriving downstream neurons of light-sensitive input, leading to vision impairment or blindness. Although downstream neurons survive, some undergo morphological and physiological remodeling. Bipolar cells (BCs) link photoreceptors, which sense light, to retinal ganglion cells (RGCs), which send information to the brain. While photoreceptor loss disrupts input synapses to BCs, whether BC output synapses remodel has remained unknown. Here we report that synaptic output from BCs plummets in RP mouse models of both sexes owing to loss of voltage-gated Ca2+ channels. Remodeling reduces the reliability of synaptic output to repeated optogenetic stimuli, causing RGC firing to fail at high-stimulus frequencies. Fortunately, functional remodeling of BCs can be reversed by inhibiting the retinoic acid receptor (RAR). RAR inhibitors targeted to BCs present a new therapeutic opportunity for mitigating detrimental effects of remodeling on signals initiated either by surviving photoreceptors or by vision-restoring tools.


Subject(s)
Retinal Bipolar Cells , Synapses , Tretinoin , Animals , Retinal Bipolar Cells/drug effects , Retinal Bipolar Cells/physiology , Mice , Tretinoin/pharmacology , Male , Female , Synapses/drug effects , Synapses/physiology , Retinitis Pigmentosa/physiopathology , Retinitis Pigmentosa/genetics , Retinal Degeneration/physiopathology , Mice, Inbred C57BL , Mice, Transgenic , Retinal Ganglion Cells/physiology , Retinal Ganglion Cells/drug effects
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