Thesis--Helsingfors University, 1938. / Includes bibliographical references (p. [71]-74).

Retinal degeneration is the progressive loss of neurons lining the posterior surface of the eye. Loss of a certain group of neurons called rod photoreceptors can occur as the result of genetic mutation. In humans, and in mammalian models of retinal degeneration, the death of these cells is permanent, and often followed by cone photoreceptor death, which leads to blindness. As a step towards understanding the implications of rod cell death in the retina, we generated transgenic X. laevis that expressed a novel form of caspase-9, with binding domains specific to the compound AP20187. We treated these transgenic animals with AP20187 and caused rod cell death by apoptosis in tadpoles and post metamorphic animals. Peak rod apoptosis occurred two days after drug exposure. We adapted an electroretinography apparatus, and protocols designed for mammals to measure functional changes in X. laevis rod and cone derived responses. We observed delayed secondary cone cell dysfunction after induced rod cell apoptosis, which was subsequently restored. These animals provide a simple and clinically relevant model of diseases like Retinitis pigmentosa, in which we will be able to probe in detail the mechanisms that govern cone cell dysfunction as a consequence of rod apoptosis. The unique ability of this species to recover from this insult will provide clues towards initiating similar recovery in humans. / Medicine, Faculty of / Graduate

Aims: The electroretinogram (ERG) can be used to independently assess the function of rod and cone photoreceptors within the human retina. The work in this thesis sought to investigate an alternative method of recording the ERG, using the silent substitution paradigm (Estevez and Spekreijse 1982). The aims are separated into two parts, firstly, the isolation and characterisation of the non-dark adapted rod photoreceptor response, and secondly, characterising the ERG response from L-, M- and S-cones. Methods: Rod, L-, M- and S-cone isolating as well as non-isolating sinusoidal flicker and transient square-wave stimuli were generated on a 4 primary LED ganzfeld stimulator to elicit ERGs from non-dark adapted participants with normal and compromised rod or cone function. Results: The results from the rod experiments showed that ERGs elicited by rod isolating silent substitution stimuli exhibit low-pass temporal frequency response characteristics with an upper response limit of 30Hz and saturate beyond 1000ph Td. Responses are optimal between 5 – 8 Hz and between 10-100 photopic Td. There is a significant correlation between the response amplitudes obtained with the silent substitution method and current standard clinical protocols. The results from the cone experiments showed that the L-, M- and S-cone stimulation produced ERGs with very different morphologies. L- and M-cone stimulation is of limited use as an objective measure of colour vision deficiency. Conclusion: Silent substitution provides an effective method for the isolation of human rod and cone photoreceptor function in subjects when stimuli are used within appropriate parameter ranges.

Hypothesis 1. MfERG is useful in the assessment of retinal dysfunction in patients following photodynamic therapy with verteporfin. / Hypothesis 2. MfERG is useful in the evaluation of retinal dysfunction in patients following safety-enhanced photodynamic therapy using half-dose verteporfin. / Hypothesis 3. MfERG is useful in the assessment of retinal dysfunction in patients receiving hydroxychloroquine therapy. / Hypothesis 4. MfERG findings correlate to certain extent with the visual field findings in patients receiving hydroxychloroquine therapy and mfERG might be more sensitive compared with 10-2 visual field testing in assessing retinal dysfunction associated with hydroxychloroquine therapy. / Hypothesis 5. MfERG is useful In the assessment of retinal dysfunction associated with intraoperative application of indocyaniue green for internal limiting membrane staining in epiretinal membrane surgery. / Multifocal electroretinography (mfERG) is an investigation which can provide objective assessment of retinal function, In contrast with full-field electroretinography which measures the mass electrical activity of the entire retina, mfERG allows simultaneous measurements of multiple retinal responses from the macula. / Several studies have demonstrated that mfERG might be useful in assessing retinal dysfunction caused by pharmacological agents and following laser therapy. This thesis aims to demonstrate the application of mfERG in the evaluation of retinal dysfunction associated with various ocular or systemic pharmacological agents. Three treatment modalities including photodynamic therapy with verteporfin, systemic use of hydroxychloroquine, and intraoperative application of indocyanine green dye were chosen for evaluation. These pharmacological agents were selected as they are associated with potential retinal dysfunction and are commonly encountered in the ophthalmic clinical practice. The thesis examines the following hypotheses: / Summary of studies arising from the thesis. Based on the findings from the above studies, it was demonstrated that mfERG can objectively evaluate the retinal dysfunction caused by a variety of ocular or systemic pharmacological agents. These included PDT with verteporfin, systemic therapy with hydroxychloroquine, as well as intraoperative application of ICG dye for ILM staining. The use of mfERG has enhanced the understanding of the underlying pathophysiology of drug-associated retinal toxicity. As mfERG becomes more widely available, its application will provide a valuable option for clinicians to assess toxic retinopathy objectively and enable safer administration of treatment to minimize potential retinal toxicity. (Abstract shortened by UMI.) / by Lai Yuk Yau, Timothy. / Source: Dissertation Abstracts International, Volume: 69-02, Section: B, page: 0951. / Thesis (M.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (p. 161-183). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / School code: 1307.

Thesis (M.S.)--University of Alabama at Birmingham, 2009. / Title from PDF title page (viewed on Sept. 3, 2009). Includes bibliographical references (p. 45-52).

PURPOSE. Recessive mutations in CLN7/MFSD8 usually cause variant late-infantile onset neuronal ceroid lipofuscinosis (vLINCL), a poorly understood neurodegenerative condition, though mutations may also cause nonsyndromic maculopathy. A series of 12 patients with nonsyndromic retinopathy due to novel CLN7/MFSD8 mutation combinations were investigated in this study. METHODS. Affected patients and their family members were recruited in ophthalmic clinics at each center where they were examined by retinal imaging and detailed electrophysiology. Whole exome or genome next generation sequencing was performed on genomic DNA from at least one affected family member. Immunofluorescence confocal microscopy of murine retina cross-sections were used to localize the protein. RESULTS. Compound heterozygous alleles were identified in six cases, one of which was always p.Glu336Gln. Such combinations resulted in isolated macular disease. Six further cases were homozygous for the variant p.Met454Thr, identified as a founder mutation of South Asian origin. Those patients had widespread generalized retinal disease, characterized by electroretinography as a rod-cone dystrophy with severe macular involvement. In addition, the photopic single flash electroretinograms demonstrated a reduced b- to a-wave amplitude ratio, suggesting dysfunction occurring after phototransduction. Immunohistology identified MFSD8 in the outer plexiform layer of the retina, a site rich in photoreceptor synapses. CONCLUSIONS. This study highlights a hierarchy of MFSD8 variant severity, predicting three consequences of mutation: (1) nonsyndromic localized maculopathy, (2) nonsyndromic widespread retinopathy, or (3) syndromic neurological disease. The data also shed light on the underlying pathogenesis by implicating the photoreceptor synaptic terminals as the major site of retinal disease.

Objetivo: Avaliar eletrofisiologicamente a função da retina do camundongo modelo de Alzheimer (3xTg-AD) comparando com seu controle (b6;129-PS1) em um estudo longitudinal com seis idades (2, 4, 6, 8, 10 e 12 meses). Métodos: Eletrorretinogramas (ERGs) foram registrados em 44 camundongos 3xTg-AD e em 23 controles, após administrada anestesia. Para o registro foi colocado um eletrodo de lente de contato sobre a córnea, um eletrodo de referência na cabeça e um terra na cauda. Em sessão de 30-40min de duração foram expostos ao seguinte protocolo de estimulação: 1) Adaptação ao escuro seguida de flashes nas intensidades: 0,003; 0,03; 0,3; 3 e 30 cd.s/m2; 2) Estimulação periódica (30 cd.s/m2) nas freqüências de 12, 18, e 30 Hz, sob luz de fundo (30 cd/m2). Resultados: Os ERGs mostraram dois tipos de respostas escotópicas tanto no grupo dos camundongos controles (b6;129- PS1) quanto nos modelos de Alzheimer. 13% dos camundongos controles e 72% dos modelos de AD apresentaram ERGs com potenciais oscilatórios presentes e tempo implícito da onda-b dentro da faixa esperada (45,31 ± 6,74 ms), enquanto no restante dos grupos, o ERG apresentou latência da onda-b muito aumentada (111,73 ± 22,56 ms) e potenciais oscilatórios ausentes. Devido a estes resultados, os grupos controle e experimental foram subdivididos em: b6;129 com OP, b6;129 sem OP; 3xTg-AD com OP e 3xTg-AD sem OP. Também foi incluído um grupo controle adicional constituído por 9 camundongos C57/B6. Comparando os cinco grupos, nenhuma diferença foi encontrada em relação à amplitude e à latência da onda-a. A amplitude da onda-b também foi semelhante para todos, ao contrário da latência para atingir o pico da onda-b dos grupos b6;129 sem OP e 3xTg-AD sem OP, que se apresentou duas vezes maior do que nos grupos com OP. As amplitudes dos cinco potenciais oscilatórios foram medidas individualmente e não mostraram diferenças entre os controles e os 3xTg-AD. Para o estímulo periódico, a amplitude do 1º harmônico dos grupos com OP mostrou clara diferença entre os grupos controle e o 3xTg-AD, tanto em 12 Hz como em 18 Hz. Os resultados dos dois grupos controle b6;129 e C57/B6 mantiveram-se muito próximos. Os grupos sem OP mantiveram-se sempre próximos a 10 V para as três freqüências de estimulação e mostraram atraso na diferença de fase média do 1º harmônico em 18 e 30 Hz, indicando maior lentidão de resposta, quando comparados aos primeiros. Conclusão: O camundongo 3xTg-AD e seu controle (b6;129) apresentam uma variante lenta e sem OPs do ERG escotópico em parte da população. Células bipolares, amácrinas e ganglionares podem estar alteradas nesses subgrupos (b6;129 sem OP e 3xTg-AD sem OP). Os grupos controle e 3xTg-AD com OPs diferiram quanto à amplitude de resposta à estimulação intermitente, diferença essa que implica em menor capacidade de processamento temporal para o modelo de AD. Sugerimos que as células bipolares de cones podem estar alteradas nos modelos de AD devido às amplitudes mais baixas dos 1os harmônicos desse grupo / Objective: To evaluate electrophysiologically the function of the retina of the Alzheimer model mouse (3xTg-AD) comparing it with its control (b6;129-PS1) in a longitudinal study at six ages (2, 4, 6, 8, 10 e 12 months) Methods: Electroretinograms (ERGs) were recorded in 44 anesthetized mice 3xTg-AD and in 23 controls, with a contact lens electrode placed on the cornea, a reference electrode on the head and a ground on the tail. During a 30-40min duration session the mice were exposed to the following stimulation protocol: 1) Scotopic response Dark adaptation followed by flashes at the following intensities: 0,003; 0,03; 0,3; 3 e 30 cd.s/m2; 2) Periodic stimulation (30 cd.s/m2) at the temporal frequencies of 12, 18, e 30 Hz, under background light (30 cd/m2). Results: The ERGs showed two types of scotopic responses, which ocurred in both the control mice (b6;129- PS1) and the Alzheimer´s models (3xTg-AD). 13% of the controls and 72% of the Alzheimer´s models mice presented ERGs with oscillatory potentials (OPs) and b-wave implicit times within the expected range (45,31 ± 6,74 ms), while for the other groups the ERG presented a very delayed b-wave latency (111,73 ± 22,56 ms) and absence of OPs. Given these results, the control and experimental groups were subdivided into: b6;129 with OPs, b6;129 without OPs; 3xTg-AD with OPs e 3xTg-AD without OPs. An additional control group with 9 mice C57/B6 was included. Comparing the five groups, no difference was found in a-wave amplitude and latency. The b-wave amplitude also did not differ among the groups, but the latency of the b-wave for the groups b6;129 without OPs and 3xTg-AD without OPs, was twice as long as in the groups with OPs. The amplitudes of the five OPs, measured individually, did not show differences between controls and 3xTg-AD groups. For the periodic stimulation the amplitude of the first harmonic of the Fourier transform of the groups with OPs showed a clear difference between the control and the 3xTg-AD groups, both for the 12 Hz and for the 18 Hz stimuli. The results of the two control groups (b6;129 and C57/B6) were very close. The groups without OPs had responses always close to 10 V for the three frequencies of stimulation and showed phase delay for the first harmonic, indicating response slowing, compared to the other groups. Conclusions: We found that a sub-group of both triple transgenic (3xTg-AD) and control mice (b6;129) manifest strikingly slow scotopic ERGs that lack OPs. We hypothesize that these response feature may reflect alterations in bipolar, amacrine and ganglion cells. The sub-group of triple transgenic and control mice that exhibited OPs differed in their response to flicker. Alzheimer model had significantly lower flicker-response amplitudes than the controls, suggesting impaired retinal temporal processing. We propose that the flicker results are consistent with alteration in cone bipolar cells in the Alzheimer model mice

O presente trabalho utilizou o eletrorretinograma (ERG) em coelhos (Oryctolagus cuniculus) para avaliar uma forma alternativa de aplicação do ácido micofenólico no tratamento da uveíte, uma doença ocular inflamatória. Uveíte é o termo utilizado para definir a inflamação da íris, coróide e corpo ciliar. O ERG é um registro de potenciais elétricos originados na retina em resposta à estimulação luminosa. O objetivo desse estudo é analisar alterações em funções da retina em olhos tratados com diferentes doses intravítreas de ácido micofenólico. Foram medidas amplitude e latência das ondas a e b do ERG para a função de intensidade-resposta e de resolução temporal. O micofenolato de mofetila (MMF) é um potente medicamento imunossupressor e vem sendo empregado com sucesso no tratamento de uveítes. O MMF é uma pró-droga, que é transformada no fígado em ácido micofenólico (MPA), o imunossupressor ativo. O uso de MMF causa efeitos colaterais e, para diminuir esses efeitos, este estudo analisa a injeção intravítea de MPA em coelhos. Foram utilizados um total de quarenta animais, divididos em seis grupos. Primeiramente, foram realizados exames eletrorretinográficos em olhos intactos, sem administração de nenhuma substância, para determinação dos parâmetros dos registros. Foram testados também os olhos injetados apenas com solução salina balanceada (SSB). Após essa etapa, foram feitos testes com olhos injetados apenas com o veículo (Polissorbato 80). Esse procedimento foi necessário para estabelecer se o veículo pode ser utilizado ou se apresenta algum grau de toxicidade para a retina. As doses usadas para avaliação da toxicidade da droga foram cinco: 5 g/ml, 50 g/ml, 200 g/ml, 1.000 g/ml e 10.000 g/ml. Os exames eletrorretinográficos foram realizados 7 e 30 dias após injeção intravítrea de MPA. Os resultados indicam que não houve alteração nos parâmetros de ERG nos olhos que receberam apenas o veículo. Também não acusaram indícios de toxicidade os olhos tratados com as doses de 5, 50 e 200 g/ml da droga. Apenas com as doses mais altas, de 1.000 e 10.000 g/ml, os olhos apresentaram diminuição de sensibilidade ao estímulo luminoso. Não é possível afirmar que as células da retina não foram danificadas pela injeção intravítrea de MPA, pois o ERG de campo total não detecta lesões pontuais. O pequeno número de olhos analisados e a necessidade de utilização de anestésicos podem levar a interpretações equivocadas dos resultados. As substâncias anestésicas podem afetar a neurotransmissão e, consequentemente, os resultados obtidos nos registros do ERG. As análises dos dados de amplitude e tempo implícito das ondas a e b foram muito consistentes e apresentam variações compatíveis com a literatura / This work uses the application of the electroretinogram (ERG) in rabbits (Oryctolagus cuniculus) in order to evaluate an alternative way of using the mycophenolic acid for treatment of uveitis, an inflammatory eye disease. Uveitis is an inflammation of the iris, choroid and ciliary body. The ERG is the record of electrical responses originated in the retina to luminous stimulation. The purpose of this study is to analyse changes in the retinal functions of eyes treated with distinct intravitreous doses of mycophnolic acid. A and b waves amplitude and implicit times for the intensity-response and temporal resolution functions. Mofetil mycophenolate (MMF) is a powerful immunossupressant drug successfully used for treating the uveitis. It is a pro-drug that is transformed in the liver into mycophenolic acid (MPA), the active immunossupressor. MMF causes collateral effects and, in order to reduce these effects, the intravitreous MPA injection in rabbits is analysed in this study. Forty rabbits were used, divided in 6 groups. First of all, electroretinografic exams in control eyes were made, without the application of any substance, for determining the recording parameters. Then were tested the eyes injected with only balanced saline solution (BSS). After that, tests were made in eyes injected only with the vehicle (Polissorbato 80). This procedure was necessary for finding out whether the vehicle may be used or is in some degree toxic for the retina. The doses for evaluation of the drug toxicity were five: 5 g/ml, 50 g/ml, 200 g/ml, 1.000 g/ml e 10.000 g/ml. By the examination of the distinct parameters of the ERG, several visual functions can be evaluated (inner and outer retina) and possible impairments in the temporal resolution, in the relationship luminous intensity and amplitude of the response, and in other functions. The results show there were no changes in the ERG parameters for the eyes where only the vehicle was applied. Also, no toxicity signs were detected in 5, 50 or 200 g/ml doses. Sensitivity to the luminous stimulus was only shown in eyes treated with higher doses, or, 1.000 and 10.000 g/ml. It is not possible to assert that the retina cells were not injured by the MPA injection, as the total field ERG does not detect spot lesions. The small quantity of tested eyes and the usage of anesthetics may lead to erroneous interpretations of the results. The anesthetic substances can affect the neurotransmission and consequently the results of ERG records. The analysis of the a and b wave amplitude and implicit time were very consistent and show variations consistent with the pertinent literature

Un des obstacles majeurs de la recherche en neurosciences est la difficulté d’accéder de manière directe au fonctionnement du cerveau afin de comprendre les mécanismes biologiques à l’origine des dysfonctionnements cérébraux dans les troubles psychiatriques. En tant qu’extension anatomique et développementale du système nerveux central, la rétine pourrait permettre d’offrir un accès indirect aux fonctions neurologiques cérébrales. Ainsi, l’investigation de la fonction rétinienne apporte l’unique opportunité d’étudier de manière objective un réseau neuronal complexe présentant des similarités avec celui du cerveau. Le cannabis est une substance neurotoxique identifiée comme modulant la transmission synaptique cérébrale par l’intermédiaire du système cannabinoïde mais les mécanismes précis à l’origine de ces anomalies sont peu connus. La première partie de ce travail consiste à présenter les bases neurobiologiques et les hypothèses physiopathologiques justifiant l’étude de la fonction rétinienne chez les usagers de cannabis, en se basant sur la présence du système cannabinoïde dans la rétine et son implication dans la régulation de la libération synaptique de neurotransmetteurs. La seconde partie discute l’intérêt de l’étude de la fonction rétinienne dans la recherche en psychiatrie avec des méthodes électrophysiologiques. Enfin, la dernière partie présente les dysfonctions rétiniennes présentes chez les usagers de cannabis, après un usage aigu ou régulier, évaluées par les techniques électrophysiologiques comme l’électrorétinogramme. Toutes ces données renforcent la pertinence de la rétine comme site d’investigation du cerveau et ouvrent éventuellement la perspective au développement de marqueurs fonctionnels / One of major obstacles in neuroscience research is the difficulty of directly accessing the brain function to understand the biological mechanisms underlying brain dysfunctions in psychiatric disorders. As an anatomical and developmental extension of the central nervous system, the retina could afford to offer an indirect access to brain neurological functions. Investigating the retinal function provides the unique opportunity to study in an objective way a complex neuronal network which shares similar properties with the brain. Cannabis is a neurotoxic substance identified as modulating brain synaptic transmission through the cannabinoid system, but the precise mechanisms underpinning these anomalies are poorly understood. The first part of this work is dedicated to present the neurobiological basis and pathophysiological hypotheses justifying the study of retinal function in cannabis users and is based on the presence of the cannabinoid system in the retina and its involvement in the regulation of synaptic neurotransmission. The second part discusses the interest of the study of retinal function with electrophysiological methods in psychiatric research. The last part presents the retinal dysfunctions detected in cannabis users, after acute or regular use, and assessed by electrophysiological techniques such as electroretinogram. All these data reinforce the relevance of the retina as a site of brain investigation and possibly open the prospect for the development of functional markers

Objetivos: Avaliar a capacidade do eletrorretinograma de padrão reverso (PERG) macular e multifocal (mf) de diferenciar pacientes com suspeita de glaucoma (SG) e glaucoma com defeito de campo hemianópico (GH) de controles, comparar a capacidade de discriminação do PERG e tomografia de coerência óptica (TCO) fourrier domain, e avaliar a relação entre as medidas do PERG e TCO. Métodos: Medidas do campo visual (CV) computadorizado, respostas do PERG transiente e modo estacionário e PERGmf foram obtidos dos SG (n=14, 24 olhos), GH (n=5, 7 olhos) e controles (n=19, 22 olhos). Os seguintes parâmetros de TCO foram investigados: camada de fibras nervosas da retina peripapilar (CFNRpp), espessura total da mácula e espessuras das camadas segmentadas da mácula. As medidas foram analizadas utilizando-se modelos lineares de efeito misto. Também foi avaliada a relação entre as medidas e a performance diagnóstica de cada tecnologia. Resultados: Comparado aos controles, a média do tempo de pico de P50 da resposta do PERG transiente estava reduzida nos SG e GH, enquanto que a fase, a amplitude do modo estacionário e respostas do PERGmf estavam anormais apenas no GH. A média das medidas da TCO de espessura macular e da CFNRpp nos SG e GH diferiram significativamente dos controles. Uma significativa relação foi observada entre o PERG e a maior parte dos dos parâmetros do CV central e TCO. A análise por regressão e componentes principais revelou que a TCO de nervo óptico e mácula, assim como o PERG transiente e PERGmf tiveram estatisticamente capacidade similar em discriminar os SG dos controles. Conclusões: Os parâmetros do PERG e da TCO podem estar anormais, com significativa relação entre as medidas, em uma porcentagem alta de olhos com SG com CV normal. Nossos achados sugerem que as duas tecnologias podem ser úteis e complementares na detecção precoce de glaucoma / Purpose: To evaluate the ability of macular and multifocal (mf) pattern electroretinogram (PERG) to differentiate glaucoma suspects (GS) and glaucoma with hemifield loss (GHL) from controls, to compare the discrimination ability of PERG and fourier-domain optical coherence tomography (fdOCT), and to assess the relationship between PERG and fdOCT measurements. Methods: Standard automated perimetry (SAP), steady-state and transient PERG responses and mfPERG measurements were obtained from GS (n=14, 24 eyes), GHL (n=5, 7 eyes) and controls (n=19, 22 eyes). The following fdOCT parameters were investigated: circumpapillary retinal nerve fiber layer (cpRNFL), full-thickness macula, and segmented macular layer thicknesses. Measurements were compared using mixed effects linear models. The relationships between measurements and the diagnostic performance of each technology were also assessed. Results: Compared to controls, average P50 peak time transient PERG responses were reduced in GS and GHL, whereas average phase and amplitude steady-state and mfPERG responses were abnormal only in GHL. The average fdOCT-measured cpRNFL and macular thickness measurements in GS and GHL differed significantly from controls. A significant relationship was found between PERG and most fdOCT or central SAP sensitivity parameters. Principal component regression analysis revealed that optic disc and macular OCT parameters, along with mfPERG and transient PERG parameters had statistically similar ability to discriminate GS from controls. Conclusions: PERG and OCT parameters may be abnormal, with significant correlations between measurements, in a high percentage of GS eyes with normal SAP. Our findings suggest that both technologies may be helpful and complementary in early glaucoma detection