Ophthalmology Commons^™

Demodex Blepharitis: A Comprehensive Review Of The Disease, Current Management, And Emerging Therapies, Michelle K. Rhee, Elizabeth Yeu, Melissa Barnett, Christopher J. Rapuano, Deepinder K Dhaliwal, Kelly K. Nichols, Paul Karpecki, Francis S. Mah, Arthur Chan, James Mun, Ian Benjamin Gaddie

Wills Eye Hospital Papers

Demodex blepharitis is a common disease of the eyelid, affecting approximately 25 million Americans. This article reviews what is known about the mechanisms and impact of Demodex blepharitis, risk factors, signs and symptoms, diagnostic techniques, current management options, and emerging treatments. Demodex mites contribute to blepharitis in several ways: direct mechanical damage, as a vector for bacteria, and by inducing hypersensitivity and inflammation. Risk factors for Demodex blepharitis include increasing age, rosacea, and diabetes. The costs, symptom burden, and psychosocial effects of Demodex blepharitis are considerable. The presence of collarettes is pathognomonic for Demodex blepharitis. Redness, dryness, discomfort, foreign body …

Extracellular-Vesicle-Based Therapeutics In Neuro-Ophthalmic Disorders, Hamed Massoumi, Sohil Amin, Mohammad Soleimani, Bita Momenaei, Mohammad Javad Ashraf, Victor H Guaiquil, Peiman Hematti, Mark I Rosenblatt, Ali R Djalilian, Elmira Jalilian

Wills Eye Hospital Papers

Extracellular vesicles (EVs) have been recognized as promising candidates for developing novel therapeutics for a wide range of pathologies, including ocular disorders, due to their ability to deliver a diverse array of bioactive molecules, including proteins, lipids, and nucleic acids, to recipient cells. Recent studies have shown that EVs derived from various cell types, including mesenchymal stromal cells (MSCs), retinal pigment epithelium cells, and endothelial cells, have therapeutic potential in ocular disorders, such as corneal injury and diabetic retinopathy. EVs exert their effects through various mechanisms, including promoting cell survival, reducing inflammation, and inducing tissue regeneration. Furthermore, EVs have shown …

Autophagy Requirements For Eye Lens Differentiation And Transparency, Lisa Brennan, M Joseph Costello, J Fielding Hejtmancik, A. Menko, S Amer Riazuddin, Alan Shiels, Marc Kantorow

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Recent evidence points to autophagy as an essential cellular requirement for achieving the mature structure, homeostasis, and transparency of the lens. Collective evidence from multiple laboratories using chick, mouse, primate, and human model systems provides evidence that classic autophagy structures, ranging from double-membrane autophagosomes to single-membrane autolysosomes, are found throughout the lens in both undifferentiated lens epithelial cells and maturing lens fiber cells. Recently, key autophagy signaling pathways have been identified to initiate critical steps in the lens differentiation program, including the elimination of organelles to form the core lens organelle-free zone. Other recent studies using ex vivo lens culture …

Tnf-Α And Nf-Κb Signaling Play A Critical Role In Cigarette Smoke-Induced Epithelial-Mesenchymal Transition Of Retinal Pigment Epithelial Cells In Proliferative Vitreoretinopathy, Victor Wang, Alison Heffer, Elisa Roztocil, Steven E Feldon, Richard T Libby, Collynn F Woeller, Ajay E Kuriyan

Wills Eye Hospital Papers

Proliferative vitreoretinopathy (PVR) is characterized by the growth and contraction of cellular membranes within the vitreous cavity and on both surfaces of the retina, resulting in recurrent retinal detachments and poor visual outcomes. Proinflammatory cytokines like tumor necrosis factor alpha (TNFα) have been associated with PVR and the epithelial-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells. Cigarette smoke is the only known modifiable risk factor for PVR, but the mechanisms are unclear. The purpose of this study was to examine the impact of cigarette smoke on the proinflammatory TNFα/NF-κB/Snail pathway in RPE cells to better understand the mechanisms through …

Cgas Drives Noncanonical-Inflammasome Activation In Age-Related Macular Degeneration, Nagaraj Kerur, Shinichi Fukuda, Daipayan Banerjee, Younghee Kim, Dongxu Fu, Ivana Apicella, Akhil Varshney, Reo Yasuma, Benjamin J. Fowler, Elmira Baghdasaryan, Kenneth M. Marion, Xiwen Huang, Tetsuhiro Yasuma, Yoshio Hirano, Vlad Serbulea, Meenakshi Ambati, Vidya L. Ambati, Yuji Kajiwara, Kameshwari Ambati, Shuichiro Hirahara, Ana Bastos-Carvalho, Yuichiro Ogura, Hiroko Terasaki, Tetsuro Oshika, Kyung Bo Kim, David R. Hinton, Norbert Leitinger, John C. Cambier, Joseph D. Buxbaum, M. Cristina Kenney, Bradley D. Gelfand, Jayakrishna Ambati

Ophthalmology and Visual Science Faculty Publications

Geographic atrophy is a blinding form of age-related macular degeneration characterized by retinal pigmented epithelium (RPE) death; the RPE also exhibits DICER1 deficiency, resultant accumulation of endogenous Alu-retroelement RNA, and NLRP3-inflammasome activation. How the inflammasome is activated in this untreatable disease is largely unknown. Here we demonstrate that RPE degeneration in human-cell-culture and mouse models is driven by a noncanonical-inflammasome pathway that activates caspase-4 (caspase-11 in mice) and caspase-1, and requires cyclic GMP-AMP synthase (cGAS)-dependent interferon-β production and gasdermin D-dependent interleukin-18 secretion. Decreased DICER1 levels or Alu-RNA accumulation triggers cytosolic escape of mitochondrial DNA, which engages cGAS. Moreover, caspase-4, gasdermin …

GabaB Receptor Attenuation Of GabaA Currents In Neurons Of The Mammalian Central Nervous System, Wen Shen, Changlong Nan, Peter T. Nelson, Harris Ripps, Malcolm M. Slaughter

Pathology and Laboratory Medicine Faculty Publications

Ionotropic receptors are tightly regulated by second messenger systems and are often present along with their metabotropic counterparts on a neuron's plasma membrane. This leads to the hypothesis that the two receptor subtypes can interact, and indeed this has been observed in excitatory glutamate and inhibitory GABA receptors. In both systems the metabotropic pathway augments the ionotropic receptor response. However, we have found that the metabotropic GABA_B receptor can suppress the ionotropic GABA_A receptor current, in both the in vitro mouse retina and in human amygdala membrane fractions. Expression of amygdala membrane microdomains in Xenopus oocytes by microtransplantation …

Naturally Occurring Deletion Mutants Of The Pig-Specific, Intestinal Crypt Epithelial Cell Protein Clca4b Without Apparent Phenotype., Stephanie Plog, Nikolai Klymiuk, Stefanie Binder, Matthew J. Van Hook, Wallace B. Thoreson, Achim D. Gruber, Lars Mundhenk

Journal Articles: Ophthalmology

The human CLCA4 (chloride channel regulator, calcium-activated) modulates the intestinal phenotype of cystic fibrosis (CF) patients via an as yet unknown pathway. With the generation of new porcine CF models, species-specific differences between human modifiers of CF and their porcine orthologs are considered critical for the translation of experimental data. Specifically, the porcine ortholog to the human CF modulator gene CLCA4 has recently been shown to be duplicated into two separate genes, CLCA4a and CLCA4b. Here, we characterize the duplication product, CLCA4b, in terms of its genomic structure, tissue and cellular expression patterns as well as its in vitro electrophysiological …

Ca2+ Diffusion Through Endoplasmic Reticulum Supports Elevated Intraterminal Ca2+ Levels Needed To Sustain Synaptic Release From Rods In Darkness., Minghui Chen, Matthew J. Van Hook, Wallace B. Thoreson

Journal Articles: Ophthalmology

UNLABELLED: In addition to vesicle release at synaptic ribbons, rod photoreceptors are capable of substantial slow release at non-ribbon release sites triggered by Ca(2+)-induced Ca(2+) release (CICR) from intracellular stores. To maintain CICR as rods remain depolarized in darkness, we hypothesized that Ca(2+) released into the cytoplasm from terminal endoplasmic reticulum (ER) can be replenished continuously by ions diffusing within the ER from the soma. We measured [Ca(2+)] changes in cytoplasm and ER of rods from Ambystoma tigrinum retina using various dyes. ER [Ca(2+)] changes were measured by loading ER with fluo-5N and then washing dye from the cytoplasm with …

Powerful Anti-Tumor And Anti-Angiogenic Activity Of A New Anti-Vascular Endothelial Growth Factor Receptor 1 Peptide In Colorectal Cancer Models, Valeria Cicatiello, Ivana Apicella, Laura Tudisco, Valeria Tarallo, Luigi Formisano, Annamaria Sandomenico, Younghee Kim, Ana Bastos-Carvalho, Augusto Orlandi, Jayakrishna Ambati, Menotti Ruvo, Roberto Bianco, Sandro De Falco

Ophthalmology and Visual Science Faculty Publications

To assess the therapeutic outcome of selective block of VEGFR1, we have evaluated the activity of a new specific antagonist of VEGFR1, named iVR1 (inhibitor of VEGFR1), in syngenic and xenograft colorectal cancer models, in an artificial model of metastatization, and in laser-induced choroid neovascularization. iVR1 inhibited tumor growth and neoangiogenesis in both models of colorectal cancer, with an extent similar to that of bevacizumab, a monoclonal antibody anti-VEGF-A. It potently inhibited VEGFR1 phosphorylation in vivo, determining a strong inhibition of the recruitment of monocyte-macrophages and of mural cells as confirmed, in vitro, by the ability to inhibit …

Swelling And Eicosanoid Metabolites Differentially Gate Trpv4 Channels In Retinal Neurons And Glia., Daniel A. Ryskamp, Andrew O. Jo, Amber M M. Frye, Felix Vazquez-Chona, Nanna Macaulay, Wallace B. Thoreson, David Križaj

Journal Articles: Ophthalmology

Activity-dependent shifts in ionic concentrations and water that accompany neuronal and glial activity can generate osmotic forces with biological consequences for brain physiology. Active regulation of osmotic gradients and cellular volume requires volume-sensitive ion channels. In the vertebrate retina, critical support to volume regulation is provided by Müller astroglia, but the identity of their osmosensor is unknown. Here, we identify TRPV4 channels as transducers of mouse Müller cell volume increases into physiological responses. Hypotonic stimuli induced sustained [Ca(2+)]i elevations that were inhibited by TRPV4 antagonists and absent in TRPV4(-/-) Müller cells. Glial TRPV4 signals were phospholipase A2- and cytochrome P450-dependent, …

Retinal Angiogenesis Suppression Through Small Molecule Activation Of P53, Sai H. Chavala, Younghee Kim, Laura Tudisco, Valeria Cicatiello, Till Milde, Nagaraj Kerur, Nidia Claros, Susan Yanni, Victor H. Guaiquil, William W. Hauswirth, John S. Penn, Shahin Rafii, Sandro De Falco, Thomas C. Lee, Jayakrishna Ambati

Ophthalmology and Visual Science Faculty Publications

Neovascular age-related macular degeneration is a leading cause of irreversible vision loss in the Western world. Cytokine-targeted therapies (such as anti-vascular endothelial growth factor) are effective in treating pathologic ocular angiogenesis, but have not led to a durable effect and often require indefinite treatment. Here, we show that Nutlin-3, a small molecule antagonist of the E3 ubiquitin protein ligase MDM2, inhibited angiogenesis in several model systems. We found that a functional p53 pathway was essential for Nutlin-3-mediated retinal antiangiogenesis and disruption of the p53 transcriptional network abolished the antiangiogenic activity of Nutlin-3. Nutlin-3 did not inhibit established, mature blood vessels …

Withaferin A Effectively Targets Soluble Vimentin In The Glaucoma Filtration Surgical Model Of Fibrosis, Paola Bargagna-Mohan, Sunil P. Deokule, Kyle G. Thompson, John Wizeman, Cidambi Srinivasan, Sunil Vooturi, Uday B. Kompella, Royce Mohan

Ophthalmology and Visual Science Faculty Publications

Withaferin A (WFA) is a natural product that binds to soluble forms of the type III intermediate filament (IF) vimentin. Currently, it is unknown under what pathophysiological contexts vimentin is druggable, as cytoskeltal vimentin-IFs are abundantly expressed. To investigate druggability of vimentin, we exploited rabbit Tenon's capsule fibroblast (RbTCF) cell cultures and the rabbit glaucoma filtration surgical (GFS) model of fibrosis. WFA potently caused G₀/G₁ cell cycle inhibition (IC₅₀ 25 nM) in RbTCFs, downregulating ubiquitin E3 ligase skp2 and inducing p27(Kip1) expression. Transforming growth factor (TGF)-ß-induced myofibroblast transformation caused development of cell spheroids with numerous elongated invadopodia, which WFA blocked …

Lateral Mobility Of L-Type Calcium Channels In Synaptic Terminals Of Retinal Bipolar Cells., Wallace B. Thoreson, Aaron J. Mercer, Karlene M. Cork, Robert J. Szalewski

Journal Articles: Ophthalmology

PURPOSE: Efficient and precise release of glutamate from retinal bipolar cells is ensured by the positioning of L-type Ca(2+) channels close to release sites at the base of the synaptic ribbon. We investigated whether Ca(2+) channels at bipolar cell ribbon synapses are fixed in position or capable of moving in the membrane.

METHODS: We tracked the movements of individual L-type Ca(2+) channels in bipolar cell terminals after labeling channels with quantum dots (QDs) attached to α(2)δ(4) accessory Ca(2+) channel subunits via intermediary antibodies.

RESULTS: We found that individual Ca(2+) channels moved within a confined domain of 0.13-0.15 μm(2) in bipolar …

Therapies For Neovascular Age-Related Macular Degeneration: Current Approaches And Pharmacologic Agents In Development., Mostafa Hanout, Daniel Ferraz, Mehreen Ansari, Natasha Maqsood, Saleema Kherani, Yasir J. Sepah, Nithya Rajagopalan, Mohamed Ibrahim, Diana V. Do, Quan Dong Nguyen

Journal Articles: Ophthalmology

As one of the leading causes of blindness, age-related macular degeneration (AMD) has remained at the epicenter of clinical research in ophthalmology. During the past decade, focus of researchers has ranged from understanding the role of vascular endothelial growth factor (VEGF) in the angiogenic cascades to developing new therapies for retinal vascular diseases. Anti-VEGF agents such as ranibizumab and aflibercept are becoming increasingly well-established therapies and have replaced earlier approaches such as laser photocoagulation or photodynamic therapy. Many other new therapeutic agents, which are in the early phase clinical trials, have shown promising results. The purpose of this paper is …

Rapid Synaptic Vesicle Endocytosis In Cone Photoreceptors Of Salamander Retina., Matthew J. Van Hook, Wallace B. Thoreson

Journal Articles: Ophthalmology

Following synaptic vesicle exocytosis, neurons retrieve the fused membrane by a process of endocytosis to provide a supply of vesicles for subsequent release and maintain the presynaptic active zone. Rod and cone photoreceptors use a specialized structure called the synaptic ribbon that enables them to sustain high rates of neurotransmitter release. They must also employ mechanisms of synaptic vesicle endocytosis capable of keeping up with release. While much is known about endocytosis at another retinal ribbon synapse, that of the goldfish Mb1 bipolar cell, less is known about endocytosis in photoreceptors. We used capacitance recording techniques to measure vesicle membrane …

Transcriptional Regulatory Network Analysis During Epithelial-Mesenchymal Transformation Of Retinal Pigment Epithelium., Craig H Pratt, Rajanikanth Vadigepalli, Praveen Chakravarthula, Gregory E Gonye, Nancy J Philp, Gerald B Grunwald

Rajanikanth Vadigepalli

PURPOSE: Phenotypic transformation of retinal pigment epithelial (RPE) cells contributes to the onset and progression of ocular proliferative disorders such as proliferative vitreoretinopathy (PVR). The formation of epiretinal membranes in PVR may involve an epithelial-mesenchymal transformation (EMT) of RPE cells as part of an aberrant wound healing response. While the underlying mechanism remains unclear, this likely involves changes in RPE cell gene expression under the control of specific transcription factors (TFs). Thus, the purpose of the present study was to identify TFs that may play a role in this process. METHODS: Regulatory regions of genes that are differentially regulated during …

Corneal Replication Is An Interferon Response-Independent Bottleneck For Virulence Of Herpes Simplex Virus 1 In The Absence Of Virion Host Shutoff, Tracy J. Pasieka, Vineet D. Menachery, Pamela C. Rosato, David A. Leib

Dartmouth Scholarship

Herpes simplex viruses lacking the virion host shutoff function (Δvhs) are avirulent and hypersensitive to type I and type II interferon (IFN). In this study, we demonstrate that even in the absence of IFN responses in AG129 (IFN-αβγR^−/−) mice, Δvhs remains highly attenuated via corneal infection but is fully virulent via intracranial infection. The data demonstrate that the interferon-independent inherent replication defect of Δvhs has a significant impact upon peripheral replication and neuroinvasion.

A Positive Feedback Synapse From Retinal Horizontal Cells To Cone Photoreceptors., Skyler L. Jackman, Norbert Babai, James J. Chambers, Wallace B. Thoreson, Richard H. Kramer

Journal Articles: Ophthalmology

Cone photoreceptors and horizontal cells (HCs) have a reciprocal synapse that underlies lateral inhibition and establishes the antagonistic center-surround organization of the visual system. Cones transmit to HCs through an excitatory synapse and HCs feed back to cones through an inhibitory synapse. Here we report that HCs also transmit to cone terminals a positive feedback signal that elevates intracellular Ca(2+) and accelerates neurotransmitter release. Positive and negative feedback are both initiated by AMPA receptors on HCs, but positive feedback appears to be mediated by a change in HC Ca(2+), whereas negative feedback is mediated by a change in HC membrane …

Quantal Amplitude At The Cone Ribbon Synapse Can Be Adjusted By Changes In Cytosolic Glutamate., Theodore M. Bartoletti, Wallace B. Thoreson

Journal Articles: Ophthalmology

PURPOSE: Vision is encoded at photoreceptor synapses by the number of released vesicles and size of the post-synaptic response. We hypothesized that elevating cytosolic glutamate could enhance quantal size by increasing glutamate in vesicles.

METHODS: We introduced glutamate (10-40 mM) into cone terminals through a patch pipette and recorded excitatory post-synaptic currents (EPSCs) from horizontal or OFF bipolar cells in the Ambystoma tigrinum retinal slice preparation.

RESULTS: Elevating cytosolic glutamate in cone terminals enhanced EPSCs as well as quantal miniature EPSCs (mEPSCs). Enhancement was prevented by inhibiting vesicular glutamate transport with 1S,3R-1-aminocyclopentane-1,3-dicarboxylate in the patch pipette. A low affinity glutamate …

Lateral Mobility Of Presynaptic L-Type Calcium Channels At Photoreceptor Ribbon Synapses., Aaron J. Mercer, Minghui Chen, Wallace B. Thoreson

Journal Articles: Ophthalmology

At most synapses, presynaptic Ca(2+) channels are positioned near vesicle release sites, and increasing this distance reduces synaptic strength. We examined the lateral membrane mobility of presynaptic L-type Ca(2+) channels at photoreceptor ribbon synapses of the tiger salamander (Ambystoma tigrinum) retina. Movements of individual Ca(2+) channels were tracked by coupling quantum dots to an antibody against the extracellular α(2)δ(4) Ca(2+) channel subunit. α(2)δ(4) antibodies labeled photoreceptor terminals and colocalized with antibodies to synaptic vesicle glycoprotein 2 and voltage-gated Ca(2+) channel 1.4 (Ca(V)1.4) α(1) subunits. The results show that Ca(2+) channels are dynamic and move within a confined region beneath the …

Calcium Regulates Vesicle Replenishment At The Cone Ribbon Synapse., Norbert Babai, Theodore M. Bartoletti, Wallace B. Thoreson

Journal Articles: Ophthalmology

Cones release glutamate-filled vesicles continuously in darkness, and changing illumination modulates this release. Because sustained release in darkness is governed by vesicle replenishment rates, we analyzed how cone membrane potential regulates replenishment. Synaptic release from cones was measured by recording postsynaptic currents in Ambystoma tigrinum horizontal or OFF bipolar cells evoked by depolarization of simultaneously voltage-clamped cones. We measured replenishment after attaining a steady state between vesicle release and replenishment using trains of test pulses. Increasing Ca(2+) currents (I(Ca)) by changing the test step from -30 to -10 mV increased replenishment. Lengthening -30 mV test pulses to match the Ca(2+) …

The Endogenous Soluble Vegf Receptor-2 Isoform Suppresses Lymph Node Metastasis In A Mouse Immunocompetent Mammary Cancer Model, Masa-Aki Shibata, Jayakrishna Ambati, Eiko Shibata, Romulo J. C. Albuquerque, Junji Morimoto, Yuko Ito, Yoshinori Otsuki

Ophthalmology and Visual Science Faculty Publications

BACKGROUND: Cancer metastasis contributes significantly to cancer mortality and is facilitated by lymphangiogenesis and angiogenesis. A new splicing variant, endogenous soluble vascular endothelial growth factor receptor-2 (esVEGFR-2) that we recently identified is an endogenous selective inhibitor of lymphangiogenesis. To evaluate the antimetastatic potential of esVEGFR-2, gene therapy with vector expressing esVEGFR-2 (pesVEGFR-2) or endostatin (pEndo) as a positive control was conducted on murine metastatic mammary cancer.

METHODS: Syngeneic inoculated metastatic mammary cancers received direct intratumoral injection of pesVEGFR-2, pEndo or pVec as control, once a week for six weeks. In vivo gene electrotransfer was performed on the tumors after each …

Endothelial Cell Capture Of Heparin-Binding Growth Factors Under Flow, Bing Zhao, Changjiang Zhang, Kimberly Forsten-Williams, Jun Zhang, Michael Fannon

Ophthalmology and Visual Science Faculty Publications

Circulation is an important delivery method for both natural and synthetic molecules, but microenvironment interactions, regulated by endothelial cells and critical to the molecule's fate, are difficult to interpret using traditional approaches. In this work, we analyzed and predicted growth factor capture under flow using computer modeling and a three-dimensional experimental approach that includes pertinent circulation characteristics such as pulsatile flow, competing binding interactions, and limited bioavailability. An understanding of the controlling features of this process was desired. The experimental module consisted of a bioreactor with synthetic endothelial-lined hollow fibers under flow. The physical design of the system was incorporated …

Calcium Homeostasis And Cone Signaling Are Regulated By Interactions Between Calcium Stores And Plasma Membrane Ion Channels., Tamas Szikra, Peter Barabas, Theodore M. Bartoletti, Wei Huang, Abram Akopian, Wallace B. Thoreson, David Krizaj

Journal Articles: Ophthalmology

Calcium is a messenger ion that controls all aspects of cone photoreceptor function, including synaptic release. The dynamic range of the cone output extends beyond the activation threshold for voltage-operated calcium entry, suggesting another calcium influx mechanism operates in cones hyperpolarized by light. We have used optical imaging and whole-cell voltage clamp to measure the contribution of store-operated Ca(2+) entry (SOCE) to Ca(2+) homeostasis and its role in regulation of neurotransmission at cone synapses. Mn(2+) quenching of Fura-2 revealed sustained divalent cation entry in hyperpolarized cones. Ca(2+) influx into cone inner segments was potentiated by hyperpolarization, facilitated by depletion of …

Accumulation Of Rhodopsin In Late Endosomes Triggers Photoreceptor Cell Degeneration, Yashodhan Chinchore, Amitavo Mitra, Patrick J. Dolph, Norbert Perrimon

Dartmouth Scholarship

Progressive retinal degeneration is the underlying feature of many human retinal dystrophies. Previous work using Drosophila as a model system and analysis of specific mutations in human rhodopsin have uncovered a connection between rhodopsin endocytosis and retinal degeneration. In these mutants, rhodopsin and its regulatory protein arrestin form stable complexes, and endocytosis of these complexes causes photoreceptor cell death. In this study we show that the internalized rhodopsin is not degraded in the lysosome but instead accumulates in the late endosomes. Using mutants that are defective in late endosome to lysosome trafficking, we were able to show that rhodopsin accumulates …

Feedback From Horizontal Cells To Rod Photoreceptors In Vertebrate Retina., Wallace B. Thoreson, Norbert Babai, Theodore M M. Bartoletti

Journal Articles: Ophthalmology

Retinal horizontal cells (HCs) provide negative feedback to cones, but, largely because annular illumination fails to evoke a depolarizing response in rods, it is widely believed that there is no feedback from HCs to rods. However, feedback from HCs to cones involves small changes in the calcium current (I(Ca)) that do not always generate detectable depolarizing responses. We therefore recorded I(Ca) directly from rods to test whether they were modulated by feedback from HCs. To circumvent problems presented by overlapping receptive fields of HCs and rods, we manipulated the membrane potential of voltage-clamped HCs while simultaneously recording from rods in …

Transcriptional Regulatory Network Analysis During Epithelial-Mesenchymal Transformation Of Retinal Pigment Epithelium., Craig H Pratt, Rajanikanth Vadigepalli, Praveen Chakravarthula, Gregory E Gonye, Nancy J Philp, Gerald B Grunwald

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

PURPOSE: Phenotypic transformation of retinal pigment epithelial (RPE) cells contributes to the onset and progression of ocular proliferative disorders such as proliferative vitreoretinopathy (PVR). The formation of epiretinal membranes in PVR may involve an epithelial-mesenchymal transformation (EMT) of RPE cells as part of an aberrant wound healing response. While the underlying mechanism remains unclear, this likely involves changes in RPE cell gene expression under the control of specific transcription factors (TFs). Thus, the purpose of the present study was to identify TFs that may play a role in this process.

METHODS: Regulatory regions of genes that are differentially regulated during …

Loss Of Sparc-Mediated Vegfr-1 Suppression After Injury Reveals A Novel Antiangiogenic Activity Of Vegf-A, Miho Nozaki, Eiji Sakurai, Brian J. Raisler, Judit Z. Baffi, Jassir Witta, Yuichiro Ogura, Rolf A. Brekken, E Helene Sage, Balamurali K. Ambati, Jayakrishna Ambati

Ophthalmology and Visual Science Faculty Publications

VEGF-A promotes angiogenesis in many tissues. Here we report that choroidal neovascularization (CNV) incited by injury was increased by excess VEGF-A before injury but was suppressed by VEGF-A after injury. This unorthodox antiangiogenic effect was mediated via VEGFR-1 activation and VEGFR-2 deactivation, the latter via Src homology domain 2-containing (SH2-containing) tyrosine phosphatase-1 (SHP-1). The VEGFR-1-specific ligand placental growth factor-1 (PlGF-1), but not VEGF-E, which selectively binds VEGFR-2, mimicked these responses. Excess VEGF-A increased CNV before injury because VEGFR-1 activation was silenced by secreted protein, acidic and rich in cysteine (SPARC). The transient decline of SPARC after injury revealed a temporal …

Paired-Pulse Depression At Photoreceptor Synapses., Katalin Rabl, Lucia Cadetti, Wallace B. Thoreson

Journal Articles: Ophthalmology

Synaptic depression produced by repetitive stimulation is likely to be particularly important in shaping responses of second-order retinal neurons at the tonically active photoreceptor synapse. We analyzed the time course and mechanisms of synaptic depression at rod and cone synapses using paired-pulse protocols involving two complementary measurements of exocytosis: (1) paired whole-cell recordings of the postsynaptic current (PSC) in second-order retinal neurons and (2) capacitance measurements of vesicular membrane fusion in rods and cones. PSCs in ON bipolar, OFF bipolar, and horizontal cells evoked by stimulation of either rods or cones recovered from paired-pulse depression (PPD) at rates similar to …

Kinetics Of Exocytosis Is Faster In Cones Than In Rods., Katalin Rabl, Lucia Cadetti, Wallace B. Thoreson

Journal Articles: Ophthalmology

Cone-driven responses of second-order retinal neurons are considerably faster than rod-driven responses. We examined whether differences in the kinetics of synaptic transmitter release from rods and cones may contribute to differences in postsynaptic response kinetics. Exocytosis from rods and cones was triggered by membrane depolarization and monitored in two ways: (1) by measuring EPSCs evoked in second-order neurons by depolarizing steps applied to presynaptic rods or cones during simultaneous paired whole-cell recordings or (2) by direct measurements of exocytotic increases in membrane capacitance. The kinetics of release was assessed by varying the length of the depolarizing test step. Both measures …