Anatomy and Neuroscience - Research Publications
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ItemRod and Cone Pathway Signalling Is Altered in the P2X7 Receptor Knock Out Mouse(PUBLIC LIBRARY SCIENCE, 2012-01-10)The P2X7 receptor (P2X7-R) is expressed in the retina and brain and has been implicated in neurodegenerative diseases. However, whether it is expressed by neurons and plays a role as a neurotransmitter receptor has been the subject of controversy. In this study, we first show that the novel vesicular transporter for ATP, VNUT, is expressed in the retina, verifying the presence of the molecular machinery for ATP to act as neurotransmitter at P2X7-Rs. Secondly we show the presence of P2X7-R mRNA and protein in the retina and cortex and absence of the full length variant 1 of the receptor in the P2X7-R knock out (P2X7-KO) mouse. The role of the P2X7-R in neuronal function of the retina was assessed by comparing the electroretinogram response of P2X7-KO with WT mice. The rod photoreceptor response was found to be similar, while both rod and cone pathway post-photoreceptor responses were significantly larger in P2X7-KO mice. This suggests that activation of P2X7-Rs modulates output of second order retinal neurons. In line with this finding, P2X7-Rs were found in the outer plexiform layer and on inner retinal cell classes, including horizontal, amacrine and ganglion cells. The receptor co-localized with conventional synapses in the IPL and was expressed on amacrine cells post-synaptic to rod bipolar ribbon synapses. In view of the changes in visual function in the P2X7-KO mouse and the immunocytochemical location of the receptor in the normal retina, it is likely the P2X7-R provides excitatory input to photoreceptor terminals or to inhibitory cells that shape both the rod and cone pathway response.
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ItemAmyloid Precursor Protein Is Required for Normal Function of the Rod and Cone Pathways in the Mouse Retina(PUBLIC LIBRARY SCIENCE, 2012-01-18)Amyloid precursor protein (APP) is a transmembrane glycoprotein frequently studied for its role in Alzheimer's disease. Our recent study in APP knockout (KO) mice identified an important role for APP in modulating normal neuronal development in the retina. However the role APP plays in the adult retina and whether it is required for vision is unknown. In this study we evaluated the role of APP in retinal function and morphology comparing adult wildtype (WT) and APP-KO mice. APP was expressed on neuronal cells of the inner retina, including horizontal, cone bipolar, amacrine and ganglion cells in WT mice. The function of the retina was assessed using the electroretinogram and although the rod photoreceptor responses were similar in APP-KO and WT mice, the post-photoreceptor, inner retinal responses of both the rod and cone pathways were reduced in APP-KO mice. These changes in inner retinal function did not translate to a substantial change in visual acuity as assessed using the optokinetic response or to changes in the gross cellular structure of the retina. These findings indicate that APP is not required for basic visual function, but that it is involved in modulating inner retinal circuitry.
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ItemMapping kainate activation of inner neurons in the rat retina(WILEY-BLACKWELL, 2013-08-01)Kainate receptors mediate fast, excitatory synaptic transmission for a range of inner neurons in the mammalian retina. However, allocation of functional kainate receptors to known cell types and their sensitivity remains unresolved. Using the cation channel probe 1-amino-4-guanidobutane agmatine (AGB), we investigated kainate sensitivity of neurochemically identified cell populations within the structurally intact rat retina. Most inner retinal neuron populations responded to kainate in a concentration-dependent manner. OFF cone bipolar cells demonstrated the highest sensitivity of all inner neurons to kainate. Immunocytochemical localization of AGB and macromolecular markers confirmed that type 2 bipolar cells were part of this kainate-sensitive population. The majority of amacrine (ACs) and ganglion cells (GCs) showed kainate responses with different sensitivities between major neurochemical classes (Ī³-aminobutyric acid [GABA]/glycine ACs > glycine ACs > GABA ACs; glutamate [Glu]/weakly GABA GCs > Glu GCs). Conventional and displaced cholinergic ACs were highly responsive to kainate, whereas dopaminergic ACs do not appear to express functional kainate receptors. These findings further contribute to our understanding of neuronal networks in complex multicellular tissues.
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ItemEarly remodeling of muller cells in the rd/rd mouse model of retinal dystrophy(WILEY, 2013-08-01)We studied the anatomical remodeling and gliosis of retinal MĆ¼ller cells in the rd/rd mouse model of photoreceptor degeneration. A computational calculation of glutamine synthetase immunoreactivity was developed so we could specifically quantify changes in MĆ¼ller cell anatomy between control mice (C57Bl/6) and the dystrophic strain. We found no change in the number of MĆ¼ller cell somata between mice strains, indicating no cell proliferation as a function of development and degeneration. The retinal area occupied by the total MĆ¼ller cell body (soma and processes) was significantly less in the rd/rd mouse retina compared with control mice. When only the outer retina was considered, we found rd/rd MĆ¼ller cell processes were dramatically reduced during the cone phase of photoreceptor degeneration. However, at older ages an increase in MĆ¼ller cell processes was seen. Conversely, glial fibrillary acidic protein (GFAP) expression showed a significant increase during cone degeneration followed by a reduction in older ages. MĆ¼ller cell electrophysiology, particularly K(+) currents and membrane potential, was similar between rd/rd and control MĆ¼ller cells during cone degeneration. Together, these results show that glial remodeling in the rd/rd retina follows separate phases-an initial conservative glial response involving the loss of MĆ¼ller cells processes, hyperexpression of GFAP, and preservation of normal electrophysiology followed by an active growth of MĆ¼ller cell processes, glial seal formation, and attenuation of GFAP expression after complete photoreceptor loss.
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ItemMapping cation entry in photoreceptors and inner retinal neurons during early degeneration in the P23H-3 rat retina(CAMBRIDGE UNIV PRESS, 2013-05)The proline-23-histidine line 3 (P23H-3) transgenic rat carries a human opsin gene mutation leading to progressive photoreceptor loss characteristic of human autosomal dominant retinitis pigmentosa. The aim of the present study was to evaluate neurochemical modifications in the P23H-3 retina as a function of development and degeneration. Specifically, we investigated the ion channel permeability of photoreceptors by tracking an organic cation, agmatine (1-amino-4-guanidobutane, AGB), which permeates through nonspecific cation channels. We also investigated the activity of ionotropic glutamate receptors in distinct populations of bipolar, amacrine, and ganglion cells using AGB tracking in combination with macromolecular markers. We found elevated cation channel permeation in photoreceptors as early as postnatal day 12 (P12) suggesting that AGB labeling is an early indicator of impending photoreceptor degeneration. However, bipolar, amacrine, or ganglion cells displayed normal responses secondary to ionotropic glutamate receptor activation even at P138 when about one half of the photoreceptor layer was lost and apoptosis and gliosis were observed. These results suggest that possible therapeutic windows as downstream neurons in inner retina appear to retain normal function with regard to AGB permeation when photoreceptors are significantly reduced but not lost.
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ItemNo Preview AvailableElectronic restoration of vision in those with photoreceptor degenerations(WILEY, 2012-09)Complete loss of vision is one of the most feared sequelae of retinal disease. Currently, there are few if any treatment options available to patients that may slow or prevent blindness in diseases caused by photoreceptor loss, such as retinitis pigmentosa and age-related macular degeneration. Electronic restoration of vision has emerged over recent years as a safe and viable option for those who have lost substantial numbers of photoreceptors and who are severely vision impaired. Indeed, there has been a dramatic increase in our understanding of what is required to restore vision using an electronic retinal prosthesis. Recent reports show that for some patients, restoration of vision to the point of reading large letters is possible. In this review, we examine the types of implants currently under investigation and the results these devices have achieved clinically. We then consider a range of engineering and biological factors that may need to be considered to improve the visual performance of newer-generation devices. With added research, it is hoped that the level of vision achieved with newer generation devices will steadily improve, resulting in enhanced quality of life for those with severe vision impairment.
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ItemNo Preview AvailableRetinal amino acid neurochemistry in health and disease(TAYLOR & FRANCIS LTD, 2013-05)Advances in basic retinal anatomy, genetics, biochemical pathways and neurochemistry have not only provided a better understanding of retinal function but have also allowed us to link basic science to retinal disease. The link with disease allowed measures to be developed that now provide an opportunity to intervene and slow down or even restore sight in previously 'untreatable' retinal diseases. One of the critical advances has been the understanding of the retinal amino acid neurotransmitters, related amino acids, their metabolites and functional receptors. This review provides an overview of amino acid localisation in the retina and examples of how retinal anatomy and amino acid neurochemistry directly links to understanding retinal disease. Also, the implications of retinal remodelling involving amino acid (glutamate) receptors are outlined in this review and insights are presented on how understanding of detrimental and beneficial retinal remodelling will provide better outcomes for patients using strategies for the preservation or restoration of vision. An internet-based database of retinal images of amino acid labelling patterns and other amino acid-related images in health and disease is located at http://www.aminoacidimmunoreactivity.com.
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ItemNo Preview AvailableA Naturally Occurring Mouse Model of Achromatopsia: Characterization of the Mutation in Cone Transducin and Subsequent Retinal Phenotype(ASSOC RESEARCH VISION OPHTHALMOLOGY INC, 2013-05)PURPOSE: This work investigates a novel, naturally occurring mouse model of achromatopsia. The specific missense mutation within the Gnat2 gene was identified and the subsequent retinal phenotype characterized. METHODS: The Gnat2 sequence was amplified using PCR from BALB/c and Gnat2(c.518A>G) retinae and the product sequenced. Retinal function was assessed at 3, 6, 9, and 12 months using the electroretinogram. Transducin and opsin expression were assessed at 3 and 12 months using immunohistochemistry and quantitative PCR. Retinal remodeling and MĆ¼ller cell gliosis were investigated using immunocytochemistry. RESULTS: An A to G missense mutation at position 518 of the Gnat2 gene was identified that resulted in an aspartic acid to glycine substitution. Gnat2(c.518A>G) animals showed no cone response, while the rod response was normal except for a decrease in the photoreceptor response at 12 months (a-wave, -14%). Gnat2(c.518A>G) retinal sections showed no transducin immunolabeling; however, protein was detected via Western blot. Gnat2 gene expression was only decreased at 12 months of age (-27%). There was reduced cone number at 12 months (-27%) and M-opsin showed evidence of mislocalization. Displaced photoreceptor terminals and altered horizontal cell, cone/rod bipolar cell morphology were evident at 3 months, becoming more extensive at 12 months with the emergence of MĆ¼ller cell gliosis. CONCLUSIONS: The Gnat2(c.518A>G) mouse contains a missense mutation that results in no cone function due to a misfolding of transducin. Cone photoreceptors also show signs of opsin mislocalization, retinal remodeling and degeneration. This naturally occurring model shows all the hallmark signs of achromatopsia.
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ItemNo Preview AvailableCcl2/Cx3cr1 Knockout Mice Have Inner Retinal Dysfunction but Are Not an Accelerated Model of AMD(ASSOC RESEARCH VISION OPHTHALMOLOGY INC, 2012-11)PURPOSE: The chemokine, Ccl2, and the fractalkine receptor, Cx3cr1, have both been implicated in the pathogenesis of age related macular degeneration (AMD), with mice lacking both genes exhibiting features of AMD by 3 months of age. However, recent reports indicate that this ascribed phenotype is due to the presence of a retinal degeneration mutation (crb1(rd8/rd8), rd8) on the background strain. Our aim was to characterize the retinal effects of lack of Ccl2 and Cx3cr1 (Ccl2(-/-)/Cx3cr1(EGFP/EGFP), CDKO-mice), in mice without the rd8 mutation. METHODS: Nine-month-old, CDKO and wildtype C57blk6J mice were investigated for retinal fundus appearance and histology. The function of the rod and cone pathways was assessed using the ERG. RESULTS: The CDKO mice did not develop lesions in the retinal fundus, and the ultrastructure of Bruch's membrane and the RPE were similar to that of C57blk6J mice. From the ERG, there was no change in the amplitude of the rod photoreceptor response, or in the rod or cone post-photoreceptor b-wave. However, the rod and cone ERG oscillatory potentials were significantly reduced in the CDKO animals, a phenotype apparent in Cx3cr1(EGFP/EGFP)- but not Ccl2(-/-)-founder lines. This correlated with aberrant amacrine cell morphology in the CDKO mice. In addition, MĆ¼ller cells were gliotic and microglial morphology subtly altered, indicative of retinal stress. CONCLUSIONS: These results suggest that in the absence of the rd8 mutation, the CDKO-mouse has a mild inner retinal phenotype characterized by altered amacrine cell function, but that it is not an accelerated model of AMD.
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ItemRelationship between the Magnitude of Intraocular Pressure during an Episode of Acute Elevation and Retinal Damage Four Weeks later in Rats(PUBLIC LIBRARY SCIENCE, 2013-07-29)PURPOSE: To determine relationship between the magnitude of intraocular pressure (IOP) during a fixed-duration episode of acute elevation and the loss of retinal function and structure 4 weeks later in rats. METHODS: Unilateral elevation of IOP (105 minutes) was achieved manometrically in adult Brown Norway rats (9 groups; nā=ā4 to 8 each, 10-100 mm Hg and sham control). Full-field ERGs were recorded simultaneously from treated and control eyes 4 weeks after IOP elevation. Scotopic ERG stimuli were white flashes (-6.04 to 2.72 log cd.s.m(-2)). Photopic ERGs were recorded (1.22 to 2.72 log cd.s.m(-2)) after 15 min of light adaptation (150 cd/m(2)). Relative amplitude (treated/control, %) of ERG components versus IOP was described with a cummulative normal function. Retinal ganglion cell (RGC) layer density was determined post mortem by histology. RESULTS: All ERG components failed to recover completely normal amplitudes by 4 weeks after the insult if IOP was 70 mmHg or greater during the episode. There was no ERG recovery at all if IOP was 100 mmHg. Outer retinal (photoreceptor) function demonstrated the least sensitivity to prior acute IOP elevation. ERG components reflecting inner retinal function were correlated with post mortem RGC layer density. CONCLUSIONS: Retinal function recovers after IOP normalization, such that it requires a level of acute IOP elevation approximately 10 mmHg higher to cause a pattern of permanent dysfunction similar to that observed during the acute event. There is a 'threshold' for permanent retinal functional loss in the rat at an IOP between 60 and 70 mmHg if sustained for 105 minutes or more.