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

A deficiência de ferro é a carência nutricional mais frequente no mundo, uma vez que a população de crianças é uma das que mais sofre com essa condição. É sabido que o desenvolvimento cerebral é determinado não apenas por um plano genético, mas sim por uma forte interação de fatores genéticos e ambientais. Evidências emergentes sugerem que a estimulação precoce pode oferecer grande eficácia terapêutica, uma vez que o cérebro é notavelmente responsivo a essa interação com o ambiente. Dado que a estimulação tátil (TS) tem sido previamente demonstrada ser uma abordagem terapêutica eficaz e com potencial aplicação em seres humanos, o objetivo deste estudo foi verificar se a exposição à estimulação tátil desde o dia pós-natal (P) 1 até P32 durante 3 min/dia, poderia ser utilizada para prevenir alterações estruturais do nervo óptico de ratos mantidos com uma dieta deficiente em ferro durante o desenvolvimento pós-natal. Foram utilizados 72 ratos machos recém-nascidos (Wistar), sendo que as ratas-lactantes foram mantidas com dieta isocalórica com 35mg/Fe por kg de ração (Grupo ANTS) ou com 4mg/Fe por kg de ração (Grupo DNTS) durante todo o período de lactação e os filhotes receberam a dieta de suas respectivas ratas-lactantes após o desmame (P22-32). Metade dos filhotes de cada grupo foi submetida à TS diária (Grupo ATS e DTS), durante todo o período experimental (P01-32). Foram realizadas análises estrutural e ultraestrutural, em 3 diferentes idades, para avaliar a integridade tecidual e também a fim de determinar se as mudanças observadas na citoarquitetura do nervo óptico foram significativamente diferentes entre os grupos e idades. Verificou-se que os animais mantidos com dieta deficiente em ferro apresentaram baixo peso corporal a partir do desmame, revelando uma curva de crescimento menos acentuada. A baixa concentração de hemoglobina e hematócritos indicam que esses animais sofreram com anemia severa em todos os períodos estudados. A análise ultraestrutural qualitativa mostrou que a deficiência de ferro imposta durante o período crítico do desenvolvimento leva a sérios danos as fibras das células ganglionares da retina, com efeitos sobre o envoltório de mielina que frequentemente apresentou afrouxamento lamelar e em idades mais avançadas foram encontradas degenerações mielínicas e axonais. A análise ultraestrutural quantitativa mostrou que a dieta deficiente em ferro leva a um atraso no processo de mielinização que pode ser parcialmente revertido pelo tratamento com estimulação tátil. Além disso, fica claro que as fibras de menor diâmetro são mais sensíveis às lesões geradas pela deficiência de ferro e também ao tratamento, enquanto que as fibras de maior diâmetro são afetadas de forma desproporcional tanto pela deficiência de ferro quanto pela estimulação tátil. As lesões sugerem que a transmissão dos sinais elétricos pode estar prejudicada, interferindo com as funções normais do sistema visual. / Iron deficiency has a critical impact on maturational mechanisms of the brain and the damage related to neuroanatomical parameters is not satisfactorily reversed after iron replacement. However, emerging evidence suggest that enriched early experience may offer great therapeutic efficacy in cases of nutritional disorders postnatally, since the brain is remarkably responsive to its interaction with the environment. Given the fact that tactile stimulation (TS) treatment has been previously shown to be an effective therapeutic approach and with potential application to humans, here we ask whether exposure to TS treatment, from postnatal day (P) 1 to P32 for 3 min/day, could also be employed to prevent neuroanatomical changes in the optic nerve of rats maintained on an iron-deficient diet during brain development. It was verified that the animals maintained with iron deficient diet presented low weight from the weaning, revealing a lower growth curve. The low concentration of hemoglobin and hematocrits indicate that these animals suffered from severe anemia in all studied periods. The qualitative ultrastructural analysis showed that the iron deficiency imposed during the critical period of development leads to serious damage to the fibers from retinal ganglion cells, with effects on the myelin sheath that frequently presented lamellar loosening and myelin degenerations and axonal at more advanced age. Quantitative ultrastructural analysis has shown that the iron deficient diet leads to a delay in the myelination process that can be partially reversed by treatment with tactile stimulation. In addition, it is clear that the smaller diameter fibers are more sensitive to the iron deficiency and also to the treatment, whereas the larger fibers are disproportionately affected by both iron deficiency and tactile stimulation. The lesions suggest that the transmission of electrical signals may be impaired, interfering with the normal functions of the visual system.

Made available in DSpace on 2014-06-11T19:31:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-07-02Bitstream added on 2014-06-13T19:01:27Z : No. of bitstreams: 1 lachat_d_dr_jabo.pdf: 9336468 bytes, checksum: f5dea24e18ee6a7cb21848540c6d48af (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Diversos estudos mostraram que a ingestão de dieta com níveis inadequados de ferro pode causar, no sistema nervoso central (SNC) de ratos, alterações morfológicas, bioquímicas e comportamentais no animal. Esses estudos têm ainda indicado que animais deficientes em ferro apresentam redução no número de lamelas de mielina e prejuízos na aprendizagem. A deficiência de ferro é uma das mais comuns desordens nutricionais em pacientes pediátricos e adultos e atinge cerca de 2,5 a 5 bilhões de pessoas em todo mundo. A consequência mais explícita da deficiência de ferro é a anemia. O ferro está relacionado ao desenvolvimento de fibras nervosas mielínicas, as quais constituem mais de 80% do nervo óptico. Objetivou-se, na presente investigação, avaliar com o auxílio de microscopia eletrônica de transmissão, os possíveis efeitos da anemia ferropriva na estrutura do nervo óptico de ratos Wistar durante os períodos de lactação e pós-lactação. Os animais foram divididos em 2 grupos: Controle e Anêmico. Os anêmicos receberam uma dieta com 4 mg de ferro/Kg, e os controle, uma dieta com 35 mg de ferro/Kg. Avaliações do peso corpóreo, hemoglobina e hematócrito foram feitas para checar os efeitos da deficiência de ferro. Os animais foram anestesiados com cloridrato de quetamina IM (22 mg/Kg) e então sacrificados por perfusão transcardíaca com PBS 0,05M, pH 7,4, seguido da mistura fixadora paraformaldeído 2% e glutaraldeído 1% diluída em tampão fosfato. Um segmento do nervo óptico foi retirado e pós-fixado em solução de tetróxido de ósmio a 1% por duas horas a 4ºC, desidratado em acetona e incluído em araldite. Cortes ultra-finos com 60 nanômetros de espessura foram montados em grades de cobre, contrastados com acetato de uranila e citrato de chumbo, observados e fotografados ao microscópio eletrônico de transmissão para detalhada análise ultraestrutural... / Several studies showed that ingestion of diets with inadequate iron levels can cause morphological, biochemical and behavioral changes in the central nervous system (CNS) of rats. These studies have also shown that iron-deficient animals have reduced number of myelin lamellae and prejudice on learning. Iron deficiency is one of the most common nutritional disorders in pediatric patients and adults and affects about 2.5-5 billion people around the world. The most explicit result of iron deficiency is anemia. The iron is related to the development of myelinated nerve fibers, which constitute more than 80% of the optic nerve. The aim of this research is to evaluate, with transmission electronic microscopy, the possible effects of iron deficiency anemia in Wistar rats optic nerve structure during lactation and pos-lactation period. The animals were divided into 2 groups: Control and Anemic. The anemic group received 4 mg iron/Kg, the control group received 35 mg iron/Kg. Evaluation of body weight, hemoglobin and hematocrit were made to check the iron deficiency effects. The animals were anesthetized with ketamine 22 mg/Kg and then sacrificed by transcardiac perfusion with PBS 0.05 M, pH 7.4, followed by paraformaldehyde fixative mixture 2% and 1% glutaraldehyde. An optic nerve segment was removed and post-fixed in a solution of osmium tetroxide for two hours at 4°C, dehydrated in acetone and embedded in Araldite. Ultrathin sections with 60 nanometers thick were mounted on copper grids, contrasted with uranyl acetate and lead citrate, observed and photographed by transmission electronic microscope for detailed ultrastructural analysis of nerve fibers, blood vessels and glial cells. Both hematological and body weight were smaller in the anemic group. The ultrastructural analysis showed damaged myelinated and unmyelinated fibers, and glial cells of the anemic animals when compared with... (Complete abstract click electronic access below)

This thesis describes a series of investigations into the use of optic nerve head (ONH) imaging in primary open-angle glaucoma (POAG), and its relation to visual function. Accurate diagnosis is a key issue in POAG, particularly the difficult task of separating those with early disease from those healthy individuals who display signs of POAG. The purpose of this work is to improve diagnostic methods in glaucoma through use of ONH imaging and its relationship with visual field (VF) loss. First, the performance of a group of expert clinicians evaluating ONH photographs for glaucomatous damage was investigated. The results showed that even when their assessments are combined discrimination between eyes with and without POAG (based on VF loss) is far from perfect, highlighting the need for improvements in diagnosis. The possibility of combining structural and functional data to aid diagnosis was then considered. This requires VF loss and ONH damage to be strongly topographically related. The strength of this relationship was evaluated in 185 patients with POAG. 10,000 computer-generated maps between the ONH and VF were tested and the topographic relationship measured with each of these was compared to that using a published structure-based map. The weak topographic relationships found suggest that the application of these maps to individual patients is limited with current measures. The next chapter describes how a multispectral imaging (MSI, also called hyperspectral imaging) system was set-up for spatial evaluation of ONH oxygenation using a Beer-Lambert law model. Test-retest repeatability was tested and found to be acceptable for the purposes of the following studies. The MSI system was then used for an investigation of the relationship between ONH oxygenation and VF loss. 33 eyes of 18 patients underwent VF testing, MSI and HRT3 imaging. Superior-inferior asymmetries in VF sensitivity were compared to superior-inferior asymmetries in ONH oxygenation measured by MSI and in neuroretinal rim (NRR) area measured by HRT3. This way we take advantage of the typical progression of POAG and each eye acts as its own reference, negating the effect of a wide normal range and overlap between health and disease. This study found, for the first time, a strong association between ONH oxygenation and VF sensitivity. A re-analysis of the 33 ONH oxygenation maps was then performed to assess oxygenation only in the area of the NRR as defined by the HRT. Superior-inferior asymmetries in NRR oxygenation were then compared to superior-inferior asymmetries in VF loss, and the associations found were similarly strong. This study shows that MSI is capable of detecting areas of NRR deemed healthy tissue by structural imaging techniques, which are in fact poorly oxygenated and associated with VF defects. These findings show that NRR oxygenation measured by MSI is strongly related to VF loss. This important information complements existing technologies and may aid in the future diagnosis and management of patients with POAG.

Introdução: As doenças que envolvem a órbita representam um complexo problema cirúrgico, principalmente as localizadas no ápice orbitário, por onde passam estruturas críticas e um espaço pequeno. O uso do endoscópio por via endonasal para abordagem cirúrgica das lesões do ápice orbitário é uma técnica recente, com poucas citações na literatura. É necessário o estudo de referências anatômicas objetivas que tornem a cirurgia mais segura. Objetivo: Descrever os parâmetros anatômicos utilizados na abordagem cirúrgica endonasal endoscópica, assim como avaliar a concordância entre os hemicrânios do mesmo cadáver e as diferenças conforme o gênero. Casuística e métodos: Estudo anatômico em 30 cadáveres adultos, ambas as fossas nasais foram dissecadas (n=60 hemicrânios). Sob visibilização endoscópica endonasal, realizou-se a dissecção do ápice orbitário. Mensuramos a distância entre a crista etmoidal e o arco coanal para o forame óptico e para a fissura orbitária superior. Os resultados foram registrados na ficha de protocolo do estudo. Resultados: Foram dissecados 30 cadáveres, 60 hemicrânios ou lados. O sexo masculino foi mais prevalente, representando 63,3% dos cadáveres (19/30), enquanto o sexo feminino representou 36,7% (11/30). 43,3% dos cadáveres eram da raça branca (13/30), 20%, pardos (6/30), e 36,7%, negros (11/30). A correlação entre os valores conforme o lado nas seguintes aferições foi observada: Crista etmoidal - Forame óptico, (r=0,748, p=0.0001); Crista etmoidal - Fissura Orbitária Superior (r=0.785, p=0.0001), Arco coanal - Forame óptico (r=0,835, p=0.0001); Arco coanal - Fissura orbitária superior (r=0.820, p=0.0001). Foi obtido um Kappa de 0,444 na avaliação da concordância entre os lados em relação ao posicionamento da artéria etmoidal anterior no forame óptico. Conclusões: A sistematização da abordagem do ápice orbitário facilita seu acesso cirúrgico e a compreensão da anatomia. A crista etmoidal e o arco coanal se mostraram estruturas relevantes e com medidas constantes nos cadáveres estudados. Os valores do coeficiente de correlação de Spearman (r) foram maiores que 0,7, o que revela uma boa correlação entre as medidas dos hemicrânios do mesmo indivíduo. Ao analisarmos a concordância do posicionamento da artéria oftálmica entre os hemicrânios de um mesmo cadáver, podemos observar que a concordância foi moderada, o que representa assimetria e variação de localização da artéria. Ao compararmos as medidas aferidas entre os lados, observou-se que os valores são semelhantes e não houve diferença estatística das distâncias em nenhuma das referências anatômicas propostas para o estudo / Introduction: Diseases that affect the orbit pose a complex surgical challenge, particularly those involving the orbital apex, a small space through which critically important structures course. Endoscopic endonasal approaches to the surgical treatment of orbital apex lesions are a recent technique, with few citations in the literature. Research is still needed into objective anatomic landmarks that can improve surgical safety. Objective: To describe the anatomic landmarks used in endoscopic endonasal surgical approaches and assess agreement between placement of these landmarks in midsagittal sections of cadaver skulls and potential gender differences. Materials and methods: In this anatomic study, the nasal fossae of 30 adult cadavers were dissected (n=60 half-skulls). The orbital apex was dissected under endoscopic endonasal visualization. The distances between the ethmoidal crest and choanal arch to the optic foramen and to the superior orbital fissure were measured and recorded. Results: Overall, 30 cadavers were dissected for a total of 60 half-skulls or sides. The sample was predominantly male (63.3%, 19/30 cadavers); females accounted for the remaining 36.7% (11/30). Regarding skin color, 43.3% of cadavers were white (13/30), 20% were brown (6/30), and 36.7% were black (11/30). The following correlations between measurements according to side were observed: ethmoidal crest to optic foramen, r=0.748 (p=0.0001); ethmoidal crest to superior orbital fissure, r=0.785 (p=0.0001); choanal arch to optic foramen, r=0.835 (p=0.0001); choanal arch to superior orbital fissure, r=0.820 (p=0.0001). Analysis of the agreement of ophthalmic artery location within the optic foramen between skull halves revealed a kappa of 0.444. Conclusions: The approach systematization to the orbital apex will facilitate surgical access and improve understanding of the anatomy. In the cadavers studied in this sample, the ethmoidal crest and choanal arch were relevant structures and exhibited consistent measurements. Spearman correlation coefficients (r) were greater than 0.7, which is indicative of good correlation between measurements obtained in the skull halves of each cadaver. Analysis of the position of the ophthalmic artery in each skull half of the same cadaver revealed moderate agreement, which indicates asymmetry and variation in the location of this artery. Comparison of the measurements obtained in different sides showed similar values, with no statistically significant differences in the distances between any of the proposed anatomic landmarks

Le Syndrome de Wolfram (SW; OMIM #222300, prévalence 1-9 / 1000 000) est une maladie neurodégénérative, qui se présente avec un début juvénile, intégrant le diabète insipide, diabète sucré, l’atrophie optique (AO), et la surdité. AO est généralement son premier symptôme neurologique, commençant à l’âge de 11 ans et se terminant par la cécité 8 ans plus tard. Malheureusement, un modèle murin du SW approprié aux symptômes ophtalmologiques n'a pas encore été trouvé, donc la recherche de la thérapie pour sauver la vision en est à ces débuts. Dans cette thèse j’ai étudié l’atteinte visuelle de deux modèles de souris mutantes pour le SW et succès d’une approche de thérapie génique (TG) avec le gène humain WFS1.Premièrement, les souris Wfs1exon8del sont été examinées à 3 et 6 mois pour l’acuité visuelle (AV) et la sensibilité aux contrastes (SC) via changements dans le reflexe optomoteur (ROM), la fonction rétinienne neurale par électrorétinogramme (ERG), ainsi que la physiologie de l’œil par la fondoscopie et tomographie par cohérence optique (TCO). De plus, la proportion des cellules ganglionnaire de la rétine (CGRs) et la perte axonale dans le nerf optique (NO) à 7 mois ont été examinés avec marquage anti-Brn3a et microscopie électronique, respectivement. Il y avait une perte progressive de l’AV et la SC chez les souris KO à partir du 1 mois. Elle était accompagnée d'une pâleur du disque optique (DO), d'amincissement de la rétine ainsi que des lésions axonales. Par contre, il n’avait pas de perte des CGRs ni stress du réticulum endoplasmique dans la rétine. Brièvement, les souris KO présentent un phénotype ophtalmique du SW significatif et peuvent servir comme modèle.Deuxièmement, à la recherche d'un autre modèle du SW, les fonctions visuelles de la lignée Wfs1E864K de la souris ont été étudiées. Déjà à 1 mois, les souris Wfs1E864K/E864K avait une perte drastique de la fonction des CGRs, mais en gardant le nombre de cellules à un niveau normal. Ceci a été accompagné par un amincissement de la rétine et d’un sévère dommage du NO, comme montré par le TCO et la fondoscopie, respectivement. En conséquence, les souris Wfs1E864K/E864K, avec leur fort phénotype ophtalmique, pourraient servir comme modèle du SW classique.Enfin, pour enquêter sur les futures options de traitement contre le SW, les souris de la lignée Wfs1exon8del à 1 mois ont subi une TG intravitréenne avec AAV-2/2-CMV-WFS1. Les examens à 3 et 6 mois ont montré une amélioration de l’AV, ainsi que le sauvetage de la pâleur du DO et réduction des lésions axonales chez les souris KO. En outre, aucun effet indésirable lié à des injections TG n’ont été noté. Suivant cette idée, les souris Wfs1E864K/E864K ont également été soumis à la TG intravitréenne, délivrée à P14, mais sans succès.En conclusion, la lignée Wfs1exon8del de la souris est un modèle fiable du SW, y compris les aspects visuels. Je propose le modèle Wfs1E864K/E864K comme une alternative, en particulier pour enquêter sur la fonction de Wfs1 dans l'œil. Enfin, la GT intravitréenne avec WFS1 a un potentiel pour sauver partiellement le phénotype ophtalmique, ouvrant la voie vers le traitement pour les patients du SW / Wolfram Syndrome (WS; OMIM #222300, prevalence 1-9 / 1 000 000) has a juvenile onset and incorporates diabetes insipidus, diabetes mellitus, optic atrophy (OA), and deafness; leading to death in middle age. OA is its first neurological symptom, starting in adolescence and ending with blindness within 8 years. Unfortunately, a suitable WS mouse model comprising ophthalmologic symptoms has not yet been found, therefore the search for its treatment is delayed. In this thesis, I studied visual impairment in two WS mouse models along with a success of a gene therapy (GT) approach with the human WFS1 gene.Firstly, 3 and 6 months old Wfs1exon8del mice were examined for the visual acuity (VA) and contrast sensitivity via changes in the opto-motor reflex (OMR), the neural retinal function via electroretinogram (ERG), as well as the eye physiology via fundoscopy and optic coher-ence tomography (OCT). Also, the proportion of retinal ganglion cells (RGC) and the axonal loss at the age of 7 months were determined with anti-Brn3a immuno-labeling of retinal sections and electron microscopy of optic nerve (ON) sections, respectively. There was a progressive loss of VA and contrast sensitivity in Wfs1exon8del-/- mice, starting already at 1 month of age. It was accompanied by optic disc pallor, retinal thinning as well as axonal damage. However, there was no RGC loss and the endoplasmic reticulum (ER) stress in the retina was at a normal level. It suggested a presence of another cause for the reported degeneration in KO mice; in opposition to what was proposed in the literature. I brief, KO mice exhibit significant WS ophthalmic phenotype.Secondly, in search for another model, visual functions of Wfs1E864K mouse line were investigated. This line was originally a model of Wolfram-like Syndrome, characterized by dominant mutations in WFS1 leading to congenital progressive hearing impairment, diabetes mellitus and OA. Only homozygous mutants, however, showed expected visual impairment. Already at 1 month of age, Wfs1E864K/E864K mice had drastic loss of RGC function, albeit keeping the cell number at a normal level. This was accompanied by retinal thinning and a severe ON damage, as shown with OCT and fundoscopy, respectively. In contrast, the RGC function in Wfs1E864K/+ mice dropped slightly only at the age of 7 and 12 months, showing that the pathology of the E864K mutation-driven disease in mice is different than in humans. Therefore, Wfs1E864K/E864K mice, with their strong ophthalmic phenotype, could potentially serve as a model of the classical WS.Finally, to investigate future treatment options, 1 month old Wfs1exon8del+/+ (WT) and Wfs1exon8del-/- (KO) mice underwent a uni- and bi-lateral intravitreal gene therapy (GT) with AAV-2/2-CMV-WFS1. Exams at 3 and 6 months of age showed improved VA, as well as optic pallor and axonal damage rescue in KO mice. Also, no adverse effects related to either GT or sham injections were noted. Following this idea, the Wfs1E864K/E864K mice were also subjected to intravitreal GT, delivered at P14, but without success.In conclusion, Wfs1exon8del mouse line is a reliable model of WS, including the visual aspects. I propose the Wfs1E864K/E864K model as an alternative, especially to investigate Wfs1 function in the eye. Finally, the intravitreal AAV-driven GT with WFS1 has a potential to partially rescue the ophthalmic phenotype, paving the wave towards the treatment for WS patients.

Currently available methods for managing glaucoma, e.g. the planimetry on stereo disc photographs, involve a subjective component either by the patient or examiner. In addition, a few structures may overlap together on the essential 2-D images, which can decrease reproducibility. Spectral domain optical coherence tomography (SD-OCT) provides a 3-D, cross-sectional, microscale depiction of biological tissues. Given the wealth of volumetric information at microscale resolution available with SD-OCT, it is likely that better parameters can be obtained for measuring glaucoma changes that move beyond what is possible using fundus photography etc. The neural canal opening (NCO) is a 3-D single anatomic structure in SD-OCT volumes. It is proposed as a basis for a stable reference plane from which various optic nerve morphometric parameters can be derived. The overall aim of this Ph.D. project is to develop a framework to segment the 3-D NCO and its related structure retinal vessels using information from SD-OCT volumes and/or fundus photographs to aid the management of glaucoma changes. Based on the mutual positional relationship of the NCO and vessels, a multimodal 3-D scale-learning-based framework is developed to iteratively identify them in SD-OCT volumes by incorporating each other's pre-identified positional information. The algorithm first applies a 3-D wavelet-transform-learning-based layer segmentation and pre-segments the NCO using graph search. To aid a better NCO detection, the vessels are identified either using a SD-OCT segmentation approach incorporating the presegmented NCO positional information to the vessel classification or a multimodal approach combining the complementary features from SD-OCT volumes and fundus photographs (or a registered-fundus approach based on the original fundus vessel segmentation). The obtained vessel positional information is then used to help enhance the NCO segmentation by incorporating that to the cost function of graph search. Note that the 3-D wavelet transform via lifting scheme has been used to remove high frequency noises and extract texture properties in SD-OCT volumes etc. The graph search has been used for finding the optimal solution of 3-D multiple surfaces using edge and additionally regional information. In this work, the use of the 3-D wavelet-transform-learning-based cost function for the graph search is a further extension of the 3-D wavelet transform and graph search. The major contributions of this work include: 1) extending the 3-D graph theoretic segmentation to the use of 3-D scale-learning-based cost function, 2) developing a graph theoretic approach for segmenting the NCO in SD-OCT volumes, 3) developing a 3-D wavelet-transform-learning-based graph theoretic approach for segmenting the NCO in SD-OCT volumes by iteratively utilizing the pre-identified NCO and vessel positional information (from 4 or 5), 4) developing a vessel classification approach in SD-OCT volumes by incorporating the pre-segmented NCO positional information to the vessel classification to suppress the NCO false positives, and 5) developing a multimodal concurrent classification and a registered-fundus approach for better identifying vessels in SD-OCT volumes using additional fundus information.

En esta tesis estudiamos la población total de las células ganglionares de la retina (CGR) en rata y ratón, y desarrollamos un modelo experimental de hipertensión ocular mediante fotocoagulación láser. La población de CGR proyecta masivamente a los colículos superiores. Se observa una estría visual en la retina dorsal donde se encuentra las densidades más altas de CGR. La pequeña población de CGR ipsilateral se distribuye mayoritariamente en la periferia de la retina temporal. El aumento de la presión intraocular induce una compresión de los axones en la cabeza del nervio óptico que provoca una alteración del transporte axonal retrógrado, que induce una degeneración sectorial localizada y difusa de las CGR, preferentemente en la retina dorsal, así como de sus axones. La pérdida selectiva de las CGR en la capa de CGR, sugiere que la causa de la muerte de las CGR no se debe a una isquemia retiniana. / In this thesis we have studied the total population of retinal ganglion cells (RGCs) in rat and mouse, and developed an experimental model of ocular hypertension by laser photocoagulation. The RGC population projects massively to the superior colliculi. There is a visual streak in the dorsal retina where the highest densities of RGCs are found. The small population of ipsilateral RGCs is distributed mainly in the periphery of the temporal retina. The increase of intraocular pressure induces a compression of the axons at the optic nerve head that causes a disturbed retrograde axonal transport, inducing a localized, diffuse and sectorial degeneration of RGCs and their axons, preferably in the dorsal retina. The selective loss of RGCs in the RGC layer, suggests that the cause of the RGC loss is not due to retinal ischemia.

Purpose: To investigate species differences in the distribution and localization of alpha B-crystallin (ABC) in the normal retina and optic nerve head region, and to describe changes during outer and inner retina degeneration. Material and methods: Animals studied included mice, rats, cats, pigs, cows, and monkeys. Sections of the optic nerve and central retina were labeled with antibodies against ABC and glial fibrillary acidic protein (GFAP). Results: ABC was located in astrocytes and Muller cells with different intensities. During outer retina degeneration (dystrophic rat and Abyssinian cat), only late stages showed an increase in ABC in the retina and optic nerve head. Inner retina degeneration in the glaucoma mouse model showed no increase of ABC. In the monkey glaucoma model, only the innermost layer of the optic nerve head showed increased labeling for ABC. Conclusions: The distribution of ABC is species dependent and is (excluding the mouse) present in the nerve fiber layer of the retina and in the optic nerve head (localization of astrocytes). Chronic retinal degeneration does not necessarily lead to an over-expression of ABC. While in outer retinal degeneration induction was predominantly present in late stages, pressure-induced glaucoma led to a specific increase in ABC already in early stages indicating a local stress-response in this region.

[Truncated abstract] Developmentally, the rodent retinocollicular projection is often thought of as a homogenous projection of retinal ganglion cell (RGC) axons, however the extensive period of RGC neurogenesis and sequential arrival of their axons into central targets such as the superior colliulus (SC) suggests otherwise. RGC axons are already present in the developing SC at embryonic (E) day 16.5-17. RGCs born on E15 have innervated the SC by birth, whereas axons derived from RGCs that are born last (E19) do not grow into the SC until postnatal (P) days 4-6 (Dallimore et al., 2002). These observations may go someway to explaining why, after SC lesions in rats at P2, there is greater growth distal to the lesion site compared to lesions made at P6 (Tan and Harvey, 1997b). It may be that the post lesion growth is simply de novo growth of axons from late-born RGCs rather than regeneration of pre-existing, injured axons. Early and late cohorts of growing RGC axons presumably encounter different developmental terrains as they grow from retina to central targets, possibly resulting in differences in developmental milestones and growth potentials. There may also be differences in guidance cues, further suggesting that gene expression in early vs late born RGCs may differ. To examine differences between early (E15) and late (E19) born RGCs during development, the time-course and extent of programmed RGC death in normal rat pups, and RGC death following the removal of target-derived trophic factors, was assessed. ... On the other hand, LCM captured GCL analysed for gene expression at P0 and P7 revealed decreases in AKT, Math5, Notch1, c-jun, DCC, Arginase-1 mRNA levels and a considerable decrease in GAP-43 expression. It is not surprising to see differences in gene expression between whole eye and the more specific GCL samples, as the cells in all layers of the retina have very different functions and different developmental profiles. It is important to note decreases in mRNA expression in the GCL for a number of the genes analysed at P0 and P7, reflecting cessation of RGC death and completion of axonal growth into central visual targets. I also examined at the protein level expression of DCC, Arginase1, c-Jun and Bcl-2 at birth (P0) in BrdU labeled RGCs born on E15 or E19. When comparing the percentage of double labelled cells compared to the total number of cells expressing each protein, Bcl-2, c-Jun and Arg1 were expressed more in E15 RGCs (22.90%, 72.71%, and 16.44% respectively in E15 RGCs, compared with 0.52%, 13.17% and 3.59% in E19 RGCs). In contrast, DCC was expressed more at birth in E19 RGCs (18.05% in E19 RGCs compared with 9.23% in E15 RGCs). This shows there is clearly a difference in the expression of proteins in the two cohorts of RGCs, which is consistent with PCR data and with their growth state as their axons encounter the changes in the newborn brain. The overall findings of this research suggest that seemingly homogenous populations of neurons are quite different in their developmental profile and in their response to injury. This work may provide new ways of determining better strategies for CNS repair and the most effective way of targeting cells for regeneration and survival.