The retina is a thin layer of tissue lining the back of the eye and is primarily responsible for sight in vertebrates. The neural retina has a distinct layered structure with three dense nuclear layers, separated by plexiform layers comprising of axons and dendrites, and a layer of photoreceptor segments. The retinal and choroidal vasculatures nourish the retina from either side, with an avascular layer comprised largely of photoreceptor cells. Diseases that directly affect the neural retina like retinal degeneration as well as those of vascular origin like diabetic retinopathy can lead to partial or total blindness. Early detection of these diseases can potentially pave the way for a timely intervention and improve patient prognosis. Current techniques of retinal imaging rely mainly on optical techniques, which have limited depth resolution and depend mainly on the clarity of visual pathway. Magnetic resonance imaging is a versatile tool that has long been used for anatomical and functional imaging in humans and animals, and can potentially be used for retinal imaging without the limitations of optical methods. The work reported in this thesis involves the development of high resolution magnetic resonance imaging techniques for anatomical and functional imaging of the retina in rats. The rats were anesthetized using isoflurane, mechanically ventilated and paralyzed using pancuronium bromide to reduce eye motion during retinal MRI. The retina was imaged using a small, single-turn surface coil placed directly over the eye. The several physiological parameters, like rectal temperature, fraction of inspired oxygen, end-tidal CO2, were continuously monitored in all rats. MRI parameters like T1, T2, and the apparent diffusion coefficient of water molecules were determined from the rat retina at high spatial resolution and found to be similar to those obtained from the brain at the same field strength. High-resolution MRI of the retina detected the three layers in wild-type rats, which were identified as the retinal vasculature, the avascular layer and the choroidal vasculature. Anatomical MRI performed 24 hours post intravitreal injection of MnCl2, an MRI contrast agent, revealed seven distinct layers within the retina. These layers were identified as the various nuclear and plexiform layers, the photoreceptor segment layer and the choroidal vasculature using Mn54Cl2 emulsion autoradiography. Blood-oxygenlevel dependent (BOLD) functional MRI (fMRI) revealed layer-specific vascular responses to hyperoxic and hypercapnic challenges. Relative blood volume of the retina calculated by using microcrystalline iron oxide nano-colloid, an intravascular contrast agent, revealed high blood-volume in the choroidal vasculature. Fractional changes to blood volume during systemic challenges revealed a higher degree of autoregulation in the retinal vasculature compared to the choroidal vasculature, corroborating the BOLD fMRI data. Finally, the retinal MRI techniques developed were applied to detect structural and vascular changes in a rat model of retinal dystrophy. We conclude that retinal MRI is a powerful investigative tool to resolve layer-specific structure and function in the retina and to probe for changes in retinal diseases. We expect the anatomical and functional retinal MRI techniques developed herein to contribute towards the early detection of diseases and longitudinal evaluation of treatment options without interference from overlying tissue or opacity of the visual pathway.

Patients with macular degeneration (MD) often fixate with a preferred retinal locus (PRL). Eye movements made while fixating with the PRL (in MD patients) has been observed to be maladaptive compared to those made while fixating with the fovea (normal sighted individuals). For example, in MD patients, PRL eye movements negatively affect fixation stability and re-fixation precision; consequently creating difficulty in reading and limits to their execution of other everyday activities. Abnormal eye movements from the PRL affect research on the physiological adaptations to MD. Specifically, previous research on cortical reorganization using functional magnetic resonance imaging (fMRI), indicates a critical need to accurately determine a MD patient's point of gaze in order to better infer existence of cortical reorganization. Unfortunately, standard MR compatible hardware eye-tracking systems do not work well with these patients. Their reduction in fixation stability often overwhelms the tracking algorithms used by these systems. This research investigates the use of an existing magnetic resonance imaging (MRI) based technique called Predictive Eye Estimation Regression (PEER) to determine the point of gaze of MD patients and thus control for fixation instability. PEER makes use of the fluctuations in the MR signal caused by eye movements to identify position of gaze. Engineering adaptations such as temporal resolution and brain coverage were applied to tailor PEER to MD patients. Also participants were evaluated on different fixation protocols and the results compared to that of the micro-perimeter MP-1 to test the efficacy of PEER. The fixation stability results obtained from PEER were similar to that obtained from the eye tracking results of the micro-perimeter MP-1. However, PEER's point of gaze estimations was different from the MP-1's in the fixation tests. The difference in this result cannot be concluded to be specific to PEER. In order to resolve this issue, advancements to PEER by the inclusion of an eye tracker in the scanner to run concurrently with PEER could provide more evidence of PEER's reliability. In addition, increasing the diversity of AMD patients in terms of the different scotoma types will help provide a better estimate of PEER flexibility and robustness.

The retina is a thin layer of tissue lining the back of the eye and is primarily responsible for sight in vertebrates. The neural retina has a distinct layered structure with three dense nuclear layers, separated by plexiform layers comprising of axons and dendrites, and a layer of photoreceptor segments. The retinal and choroidal vasculatures nourish the retina from either side, with an avascular layer comprised largely of photoreceptor cells. Diseases that directly affect the neural retina like retinal degeneration as well as those of vascular origin like diabetic retinopathy can lead to partial or total blindness. Early detection of these diseases can potentially pave the way for a timely intervention and improve patient prognosis. Current techniques of retinal imaging rely mainly on optical techniques, which have limited depth resolution and depend mainly on the clarity of visual pathway. Magnetic resonance imaging is a versatile tool that has long been used for anatomical and functional imaging in humans and animals, and can potentially be used for retinal imaging without the limitations of optical methods. The work reported in this thesis involves the development of high resolution magnetic resonance imaging techniques for anatomical and functional imaging of the retina in rats. The rats were anesthetized using isoflurane, mechanically ventilated and paralyzed using pancuronium bromide to reduce eye motion during retinal MRI. The retina was imaged using a small, single-turn surface coil placed directly over the eye. The several physiological parameters, like rectal temperature, fraction of inspired oxygen, end-tidal CO2, were continuously monitored in all rats. MRI parameters like T1, T2, and the apparent diffusion coefficient of water molecules were determined from the rat retina at high spatial resolution and found to be similar to those obtained from the brain at the same field strength. High-resolution MRI of the retina detected the three layers in wild-type rats, which were identified as the retinal vasculature, the avascular layer and the choroidal vasculature. Anatomical MRI performed 24 hours post intravitreal injection of MnCl2, an MRI contrast agent, revealed seven distinct layers within the retina. These layers were identified as the various nuclear and plexiform layers, the photoreceptor segment layer and the choroidal vasculature using Mn54Cl2emulsion autoradiography. Blood-oxygenlevel dependent (BOLD) functional MRI (fMRI) revealed layer-specific vascular responses to hyperoxic and hypercapnic challenges. Relative blood volume of the retina calculated by using microcrystalline iron oxide nano-colloid, an intravascular contrast agent, revealed a superfluous choroidal vasculature. Fractional changes to blood volume during systemic challenges revealed a higher degree of autoregulation in the retinal vasculature compared to the choroidal vasculature, corroborating the BOLD fMRI data. Finally, the retinal MRI techniques developed were applied to detect structural and vascular changes in a rat model of retinal dystrophy. We conclude that retinal MRI is a powerful investigative tool to resolve layerspecific structure and function in the retina and to probe for changes in retinal diseases. We expect the anatomical and functional retinal MRI techniques developed herein to contribute towards the early detection of diseases and longitudinal evaluation of treatment options without interference from overlying tissue or opacity of the visual pathway.

Sensory deprivation in one modality can enhance the development of the remaining modalities via mechanisms of synaptic plasticity. Mice of C3H strain suffers from RD1 retinal degeneration that leads to visual impairment at weaning age. Independently on the retinal degeneration there is also present olivocerebellar degeneration caused by Lurcher mutation. This neurodegenerative disorder causes motor deficits, increased CNS excitability as well as changes in synaptic plasticity. The aim of this study was to evaluate a role of whiskers in compensation of the visual deficit and to assess the influence of the olivocerebellar degeneration on this process. To differentiate contribution of the whiskers from other mechanisms that can take part in the compensation, we investigated the effect of both chronic and acute tactile deprivation. We focused on motor skills (rotarod, beam walking test), gait control (CatWalk system), spontaneous motor activity (open field) and the CNS excitability (audiogenic epilepsy). In the seeing mice without olivocerebellar degeneration, the removal of the whiskers had no effect. In the blind animals without olivocerebellar degeneration, chronic tactile deprivation caused changes in gait and impaired the performance in motor tests. Some other compensatory mechanisms were involved but the...

A Degeneração da Mácula Relacionada a Idade (DMRI) acomete pessoas com mais de 50 anos, comprometendo gravemente a visão. Desde 2005, têm-se sugerido uma correlação entre DMRI e o polimorfismo Y402H do Fator H (FH). Os mecanismos pelos quais a proteína FH participa da etiopatogenia dessa doença têm sido alvo de muitos estudos, desde então. Neste trabalho, investigamos a correlação entre esse polimorfismo e a expressão de proteínas da via alternativa e parâmetros do perfil lipídico de pacientes com DMRI. As concentrações de FH, Fator B, C3 e Proteína C-reativa foram semelhantes entre os grupos controle e paciente. As concentrações de Fator D e os autoanticorpos encontravam-se reduzidos nos pacientes; enquanto Fator I e os demais parâmetros do perfil lipídico estavam aumentados nesses pacientes. A variante Y402 aparentemente aderiu melhor à superfície das leptospiras (superfície ativadora da via alternativa) em relação à variante H402, mas não houve diferença entre as variantes em relação à ligação a células endoteliais (superfície não ativadora). / Age-related Macular Degeneration (AMD) affects people over 50 years, and severely prejudice the vision. Since 2005, it has been suggested a correlation between AMD and the Y402H polymorphism of Factor H (FH). After this, the mechanisms by which FH protein participates in the pathogenesis of this disease have been extensively studied. In this study, we investigated the correlation between this polymorphism and expression of proteins of the alternative pathway and lipid profile of patients with AMD. The concentrations of FH, Factor B, C3 and C-reactive protein were similar between the control and patient groups.Factor D concentrations and autoantibodies levels were reduced in patients, while Factor I concentrations and the levels of the other parameters of lipid profile were increased in these patients.Apparently, Y402 variant displays better adhesion to the surface of Leptospira (alternative pathway activating surface) than the H402 variant, but no difference between the variants of the linkage to endothelial cells (non-alternative pathway activating surface).

Sensory deprivation in one modality can enhance the development of the remaining modalities via mechanisms of synaptic plasticity. Mice of C3H strain suffers from RD1 retinal degeneration that leads to visual impairment at weaning age. Independently on the retinal degeneration there is also present olivocerebellar degeneration caused by Lurcher mutation. This neurodegenerative disorder causes motor deficits, increased CNS excitability as well as changes in synaptic plasticity. The aim of this study was to evaluate a role of whiskers in compensation of the visual deficit and to assess the influence of the olivocerebellar degeneration on this process. To differentiate contribution of the whiskers from other mechanisms that can take part in the compensation, we investigated the effect of both chronic and acute tactile deprivation. We focused on motor skills (rotarod, beam walking test), gait control (CatWalk system), spontaneous motor activity (open field) and the CNS excitability (audiogenic epilepsy). In the seeing mice without olivocerebellar degeneration, the removal of the whiskers had no effect. In the blind animals without olivocerebellar degeneration, chronic tactile deprivation caused changes in gait and impaired the performance in motor tests. Some other compensatory mechanisms were involved but the...

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Diss. (sammanfattning) Umeå : Umeå universitet, 2009. / Härtill 4 uppsatser. Även tryckt utgåva.

A non-experimental quantitative, descriptive and contextual study which sought to examine the level of awareness about Diabetic Retinopathy (DR), and how aware female diabetic patients were about retinal screening as a preventative measure to eye complications and blindness was conducted. The objective of the study was to explore and describe the variables related to the awareness level of female diabetic patients about Diabetic Retinopathy and diabetic retinal screening. A convenient sample of 149 respondents was obtained. A questionnaire was used to collect data. Data was analysed using the Statistical Package for Social Sciences (SPSS), 13.0 computer software program. Results evidenced a good level of awareness about DR. Recommendations based on the findings were made for consideration in clinical practice, education and research. / Health Studies / M.P.H.

La dégénérescence maculaire liée à l’âge ou DMLA est une pathologie oculaire qui touche près d’un million de personnes en France, et représente la première cause de cécité légale dans les pays industrialisés. C’est une affection multifactorielle (environnement, génétique), dans laquelle les stress inflammatoires, métaboliques et oxydants interviennent et aboutissent à la mort des photorécepteurs. L’apparition des drusen (dépôts de matériel extracellulaire contenant de l’amyloïde-β (Aβ)), entre les cellules de l’épithélium pigmentaire de la rétine (EPR) et la membrane de Brush, représente un facteur de risque de développement de la DMLA. De plus, le 4-hydroxynonenal (4-HNE) est un marqueur de stress oxydant dans la rétine de patients de différentes pathologies dégénératives comme la DLMA. L’identification des mécanismes moléculaires et cellulaires impliqués dans les dégénérescences rétiniennes la pathogenèse de la DMLA constitue un enjeu de santé publique, puisqu’elle permettrait de développer de nouvelles stratégies thérapeutiques anti-dégénératives.Le but de mon travail de thèse a été dans un premier temps de mieux comprendre le rôle de l’Aβ dans la dégénérescence rétinienne.Nous avons montré que l’Aβ induit une activation rapide des cellules microgliales, une gliose soutenue des cellules gliales de Müller (CGM), un œdème dans la rétine interne et une apoptose des photorécepteurs. La dégénérescence des photorécepteurs est en corrélation avec une activation soutenue de PERK, impliquée dans la voie pro-apoptotique de la réponse UPR. Par ailleurs la gliose des CGM est caractérisé par une délocalisation des canaux Kir4.1, une diminution de l’expression d’AQP4 et de la glutamine synthetase (GS), et une augmentation de l’expression des canaux Kir2.1 et du transporteur GLAST1, suggérant une dérégulation de l’homéostasie rétinienne contrôlée par ces protéines. Nous avons montré que l’inhibition de la réponse inflammatoire, par l’utilisation de l’indomethacine, un inhibiteur non stéroïdien de de la cyclooxygénase (COX) 2, réverse l’effet de l’Aβ sur l’expression des canaux Kir4.1 et sur GLAST1 mais pas celle de la GS et d’AQP4, suggérant un couplage partiel entre la gliose et la réponse inflammatoire dans notre modèle d’injection sous-rétinienne d’Aβ.Dans un deuxième temps, nous nous sommes intéressés au rôle du 4-HNE dans les CGM, un produit de peroxydation lipidique, qui est produit dans la rétine sous l’effet de l’Aβ. Nous avons observé qu’un stress oxydant unique et létal induit par le 4-HNE, entraîne la mort des CGM par apoptose dépendante de l’activation des caspases. L’utilisation d’antioxydants impliqués dans la régénération du glutathion (GSH), protège contre la mort des CGM. L’analyse du transcriptome des CGM soumises au 4-HNE a permis de mettre en évidence une réponse transcriptionnelle adaptative des CGM : une activation de la défense anti-oxydante, de la réponse UPR (unfolded protein response) au stress du réticulum endoplasmique, et un phénotype anti-inflammatoire. Par ailleurs, la surexpression de l’APP (amyloid protein precursor), dont l’expression du transcrit est augmentée sous l’effet du stress oxydant dans les CGM, protège ces cellules contre la mort induite par le 4-HNE. Cette protection est associée à une augmentation des capacités anti-oxydantes et à une activation de la voie de survie de la réponse UPR. L’ensemble de nos résultats montre un rôle de l’Aβ dans la dégénérescence des photorécepteurs et indique que le métabolisme de l’APP, ainsi que les voies de survie et pro-apoptotique de la réponse UPR pourraient constituer des cibles thérapeutiques contre la dégénérescence rétinienne induite par l’Aβ ou les stress oxydants. / Age related macular degeneration (AMD) is a leading cause of blindness in western countries and affects one million people in France. Multiple risk factors (genetics, environment) are involved in the pathogenesis of AMD. In addition, the AMD pathogenesis is strongly associated with chronic oxidative stress and inflammation that ultimately lead to photoreceptor death. AMD is characterized by the formation of drusen, extracellular deposits, including amyloid-β (Aβ), between the retinal pigmented epithelium and Bruch’s membrane. Moreover, 4-hydroxynonenal (4-HNE) is an oxidative stress marker of different retinal diseases including AMD. The determination of molecular and cell mechanisms involved in retinal degeneration and the pathogenesis of AMD is required in order to develop new therapeutic anti-degenerative approaches. The aim of our study was first to investigate the role of Aβ in retinal degeneration. We demonstrated that subretinal injection of Aβ induces an early activation of microglial cells, a sustained retinal Müller glial (RMG) cells gliosis, an oedema in the internal part of retina and photoreceptors apoptosis. The photoreceptors apoptosis was correlated with a sustained activation of PERK, a kinase implicated in the pro-apoptotic pathway of UPR (unfolded protein response). In addition, RMG gliosis has been characterized by a Kir4.1 channel redistribution, a down-regulation of AQP4 and glutamine synthetase (GS) expression, and an up-regulation of Kir2.1 channel and GLAST1 transporter expression, suggesting a dysregulation of the retinal homeostasis which is controlled by these proteins. The inhibition of the inflammatory response using indomethacin, a non-steroidal and non-specific cyclooxygenase (COX) 2 inhibitor, reversed Aβ-induced Kir4.1 channel redistribution and GLAST1 up-regulation but not GS and AQP4 down-regulation, suggesting a partial coupling between gliosis and inflammatory response in retinal degeneration after subretinal injection of Aβ in mice. The second part of our study aimed to investigate the effects on RMG cells of 4-HNE, a lipid peroxidation product that is up-regulated in retina after Aβ injection. We have shown that a single lethal oxidative stress using 4-HNE induces RMG cells apoptosis associated with caspase 3 and caspase 9 activation. Pre-treatment of RMG cells with anti-oxidative molecules involved in glutathione regeneration restored cell viability. Transcriptome analysis of RMG cells treated with 4-HNE showed an adaptive transcriptional response consisting in an activation of anti-oxidative stress cell defense, activation of UPR in response to endoplasmic reticulum stress and anti-inflammatory phenotype. APP (amyloid protein precursor) overexpression, which the transcript is up-regulated in RMG cells under oxidative stress, protects from 4-HNE-induced cell death. This protection is associated with an up-regulation of anti-oxidative cell defense and an activation of the pro-survival pathway of UPR. Our study pinpoints the role of Aβ in photoreceptors degeneration and suggests that targeting APP metabolism, pro and anti-apoptotic pathways of the UPR response may hel develop selective methods against retinal degeneration implicating Aβ and oxidative stress.