Typescript (photocopy). / Digitized by Kansas Correctional Industries

Retinitis pigmentosa (RP) is a photoreceptor neurodegenerative disease. Patients with RP present with the loss of their peripheral visual field, and the disease will progress until there is a full loss of vision. Approximately 36,000 cases of simplex and familial RP worldwide are caused by a mutation in the rod-specific cyclic guanosine monophosphate phosphodiesterase (PDE6) complex. However, despite the need for treatment, mouse models with mutations in the alpha subunit of PDE6 have not been characterized beyond 1 month of age or used to test the pre-clinical efficacy of potential therapies for human patients with RP caused by mutations in PDE6A. We first proposed to establish the temporal progression of retinal degeneration in a mouse model with a mutation in the alpha subunit of PDE6: the Pde6anmf363 mouse. Next, we developed a surgical technique to enable us to deliver therapeutic treatments into the mouse retina. We then hypothesized that increasing PDE6a levels in the Pde6anmf363 mouse model, using an AAV2/8 gene therapy vector, could improve photoreceptor survival and retinal function when delivered before the onset of degeneration. Human RP patients typically will not visit an eye care professional until they have a loss of vision, therefore we further hypothesized that this gene therapy vector could improve photoreceptor survival and retinal function when delivered after the onset of degeneration, in a clinically relevant scenario. For each of these studies, we used histology, autofluorescence (AF) and infrared (IR) imaging to examine the appearance of the retinal cell layers and retinal pigment epithelium (RPE) that are affected in human RP patients. We also used electroretinograms (ERGs) to measure both photoreceptor-specific and global retinal visual function in the Pde6anmf363 mice. For our gene therapy experiments, we utilized a vector with the cell-type-specific rhodopsin (RHO) promoter: AAV2/8(Y733F)-Rho-Pde6a, to transduce Pde6anmf363 retinas after subretinal injection at either post-natal day (P) 5 or P21. We then monitored the effects of AAV2/8(Y733F)-Rho-Pde6a transduction over at least a quarter of the mouse lifespan. In the Pde6anmf363 mutant mouse model of RP, we found that by 2 months of age the number of photoreceptor cell nuclei is roughly halved in comparison to the 1 month time-point, and this degeneration continues until all photoreceptor cell nuclei have undergone degeneration by 4 months of age. Additionally, both loss of cone cell function and RPE atrophy are present by 5 months of age in these mice. After the development of a subretinal injection surgical procedure, we delivered the AAV2/8(Y733F)-Rho-Pde6a to the Pde6anmf363 mice at either P5 or P21. We found that a single injection enhanced survival of photoreceptors and improved retinal function. At 6 months of age, the treated eyes retained photoreceptor cell bodies, while there were no detectable photoreceptors remaining in the untreated eyes. More importantly, the treated eyes demonstrated functional visual responses even after the untreated eyes had lost all vision. Despite focal rescue of the retinal structure adjacent to the injection site, global functional rescue of the entire retina was observed. We have also determined that subretinal transduction of this rod-specific transgene at P21, when approximately half of the photoreceptor cells have undergone degeneration, has similar efficacy in rescuing cone cell function long-term as transduction before disease onset, at P5. Therefore, we concluded that the Pde6anmf363 mice mimic human RP caused by mutations in PDE6A. The establishment of the temporal and biochemical characteristics of photoreceptor neurodegeneration in the Pde6anmf363 mice allows for future studies to test therapeutic options using this animal model, since the progression of RP can be compared to the established time-course of degeneration. Additionally, the development of a standard method for performing subretinal injections allows for comparable results after this surgical technique is used to deliver gene therapy vectors into the perinatal mouse eye. Our gene therapy studies suggest that RP due to PDE6a deficiency in humans, in addition to PDE6b deficiency, is also likely to be treatable by gene therapy. Furthermore, AAV2/8(Y733F)-Rho-Pde6a is an effective gene therapy treatment that can be utilized in the clinical setting, in human patients who have lost portions of their peripheral visual field and are in the mid-stage of disease when they first present to an eye-care professional.

As we navigate the environment, images flow over our retinae and produce complex moving patterns including contraction and expansion. These in turn serve to inform us about our movement in the world, as well as about the positions and relative motions of the objects around us. The processes underlying the perception of radial motion were explored using psychophysics, eye tracking and functional magnetic resonance imaging (fMRI). The initial experiments studied how adaptation to radial motion affects discrimination using partially-coherent dot stimuli. Adaptation increased absolute detection thresholds in the adapted direction, but had no effect on the discrimination of higher pedestals. The change in sensitivity is consistent with a divisive inhibition mechanism, as well as with one based on proportion estimates. An orientation discrimination experiment adds support for the latter. Directed attention to one component of a motion-balanced transparently-moving stimulus impairs sensitivity to a level comparable to that following adaptation, although there were large inter-subject differences. A novel approach using relative velocity adjustments of transparently-moving dot fields was used to investigate the reference frame of motion adaptation, which is shown to be mainly retinotopic and diminished by gaze modulations. Finally, fMRI was used to probe the neural substrates of radial motion perception, including retinotopic mapping, localisation of MT/MST, and selectivity for contraction and expansion within the MT+ complex.

Glaucoma describes a group of optic neuropathies characterised by progressive irreversible loss of visual function. Within this thesis, a health economic model was constructed to map service provision from diagnosis considering two competing strategies: the current practice of annual visual field (VF) monitoring against the proposed guidelines of performing 6 VFs in the first two years. The constructed model found the proposed practice to be cost effective at a willingness to pay ceiling ratio of £30,000 per quality adjusted life year (QALY), identifying an incremental cost effectiveness ratio (ICER) of £21,679. The findings of the model however were potentially sensitive to the modelled infrastructure improvement costs required to undertake the proposed guidelines and a costing study to more accurately ascertain these costs was recommended. Following this study, statistical analysis of 473,252 VFs was undertaken to investigate trends in initial identification and progression rates whilst also narrowing their parameters within the health economic model. Consequently, the average level of glaucomatous vision loss at diagnosis was found to be improving by 0.11 dB per year on average whilst proportions of patients with ‘advanced’ loss at diagnosis fell significantly from 30% to 21%. Average progression rates were found to have fallen from -0.11 dB per year to -0.06 dB per year whilst average rates of loss in older eyes ( > 70 years) were found to progress faster than in younger eyes ( < 60 years). Furthermore, testing frequency was found not to vary by visual impairment risk factors. The constructed health economic model was subsequently updated to incorporate the more narrowly defined parameter distributions whilst also being re-specified to incorporate societal costs of visual impairment to count the true costs of the disease. This resulted in an improved ICER of £11,382. In conclusion, it is likely that implementing the proposed guidelines of 6 VFs in the first two years is more cost-effective than annual monitoring. This argument is further reinforced once societal costs are accounted for however a scoping study to examine the required costs of improving the glaucoma monitoring infrastructure is required.

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<em>Introduction:</em> Accommodation (change in ocular focus) and Vergence (change in ocular alignment) are two ocular motor systems that interact with each other to provide clear single binocular vision. While retinal blur drives accommodation as a reflex, retinal disparity changes accommodative position through the convergence-accommodation (or simply vergence-accommodation, VA) cross-link. Similarly, while retinal disparity primarily drives the vergence system, a change in retinal blur alters vergence through the accommodative-convergence (AC) cross-link. Although much information is known on the individual response dynamics of blur accommodation and disparity vergence, very little is known about the cross-linkages AC and VA. VA represents the unique situation where a stimulus to vergence (retinal disparity) drives a change in accommodation. When these dynamic measures are compared to those of vergence and blur accommodation a better understanding of the critical or rate limiting step within the system of vergence and accommodation can be determined. Accordingly, the purpose of this thesis was to determine the response dynamics of vergence driven accommodation (VA) and compare the response parameters to simultaneous measures of disparity vergence and blur driven accommodation. <br /><br /> <em>Methods:</em> A disparity stimulus generator (DSG) was modified to allow step stimulus demands of disparity to be created on a 0. 2 cpd non-accommodative difference of Gaussian target. Retinal disparity of different step amplitude demands were created as an ON / OFF paradigm and projected on a stereo monitor set at a distance of 1. 2m. Two experiments were conducted. The first experiment investigated the first order properties of VA in comparison to similar measures of blur driven accommodation (BA). The second study aimed at comparing the first order and second order dynamics of disparity vergence, VA and BA. <br /><br /> In the first experiment, stimulus measures of vergence, vergence-accommodation and BA were studied. Six normal young adult subjects participated in the study. Accommodation was measured continuously at 25Hz with the commercially available PowerRefractor (Multichannel systems, Germany). A Badal optical system was designed and accommodative response to step stimulus demands were measured. VA and BA measures obtained from the PowerRefractor were matched and plotted as main sequences (amplitude vs. peak velocity). Peak velocities between the two responses were compared using two-way analysis of variance (ANOVA) with Bonferroni post-tests. <br /><br /> In the second experiment, the response dynamics of vergence, vergence-accommodation, and blur accommodation were assessed and compared on 6 young adult subjects. Eye position was measured continuously by a stereo eye tracker at a sampling rate of 120Hz. A high speed photorefractor (sampling = 60Hz) was custom designed and synchronized with a stereo eye tracker to allow simultaneous measurement of vergence and VA. Monocular blur driven accommodation measures were also obtained with the Badal optometer and the high speed photorefractor (sampling = 75Hz). VA, BA and disparity vergence responses were analyzed and temporal parameters like latency, amplitude, duration, time to peak velocity, peak acceleration, duration of acceleration, and skewness were calculated. Main sequence plots (response amplitude vs. peak velocity) were generated and compared between disparity ON and disparity OFF. The dynamic measures of VA were compared to the measures of monocular blur driven accommodation. All comparisons were done using a two-way ANOVA with Bonferroni post-tests. <br /><br /> <em>Results:</em> <br /> <em>Study 1:</em> The results showed that response amplitude of VA during disparity ON and disparity OFF paradigms was linearly related to the peak velocity for an amplitude range of 0. 5 to 2. 5 D (Disparity ON: peak velocity of vergence-accommodation = 0. 812 * amplitude + 1. 564, R² = 0. 452, p<0. 0001 and Disparity OFF: peak velocity of vergence-accommodation = 1. 699* amplitude ? 0. 234, R² = 0. 86, p <0. 0001). The rate of change of peak velocity as a function of response magnitude was lower for VA during disparity ON compared to VA during disparity OFF. BA responses also showed amplitude dependent dynamic properties (Accommodation peak velocity = 1. 593 * amplitude - 0. 008, R² = 0. 84, p<0. 001; Dis-accommodation peak velocity = 1. 646 * amplitude - 0. 036, R² = 0. 77, p<0. 001). There was no statistical difference in the velocity of accommodation and dis-accommodation. <br /><br /> <em>Study 2:</em> When amplitudes were matched, disparity vergence response during disparity ON and disparity OFF had similar main sequence relationships. The mean values for the stimulus and response VA/V ratios were similar (0. 13±0. 05 D/&Delta; and 0. 15±0. 09 D/&Delta; respectively). All the temporal parameters of vergence-accommodation were similar during disparity ON and disparity OFF paradigms. When blur accommodation and vergence-accommodation measures were compared, all the first order and second order temporal parameters in the response were similar between the two systems. Also, disparity vergence exhibited significantly greater peak velocity and peak acceleration compared to two accommodation responses. The results also confirmed that the velocity of accommodation and dis-accommodation showed a statistically significant linear relationship as a function of amplitude for the range of amplitudes tested (Accommodation, y = 2. 55x + 0. 65, R² = 0. 55, p<0. 0001; Dis-accommodation, y = 2. 66x + 0. 50, R²=0. 65, p<0. 0001). <br /><br /> <em>Conclusions:</em> The dynamic properties of VA are amplitude dependent. Although initial results from study 1 suggested that VA may be slower during disparity ON, the results from study 2 using the high speed photorefractor and an improved analysis procedure showed that VA responses were equally fast between disparity ON (convergence) and disparity OFF (divergence). All temporal properties of VA were independent of vergence type (convergence/divergence). VA and BA have similar dynamic properties in humans suggesting that they may controlled by a common neural pathway or limited by the plant. Also, when compared to accommodation responses, disparity vergence exhibited greater velocities and accelerations reflecting the differences in the magnitude of neural innervation and plant mechanics between the two systems. The study also confirmed amplitude dependent response dynamics of blur driven accommodation and dis-accommodation.

<em>Introduction:</em> Accommodation (change in ocular focus) and Vergence (change in ocular alignment) are two ocular motor systems that interact with each other to provide clear single binocular vision. While retinal blur drives accommodation as a reflex, retinal disparity changes accommodative position through the convergence-accommodation (or simply vergence-accommodation, VA) cross-link. Similarly, while retinal disparity primarily drives the vergence system, a change in retinal blur alters vergence through the accommodative-convergence (AC) cross-link. Although much information is known on the individual response dynamics of blur accommodation and disparity vergence, very little is known about the cross-linkages AC and VA. VA represents the unique situation where a stimulus to vergence (retinal disparity) drives a change in accommodation. When these dynamic measures are compared to those of vergence and blur accommodation a better understanding of the critical or rate limiting step within the system of vergence and accommodation can be determined. Accordingly, the purpose of this thesis was to determine the response dynamics of vergence driven accommodation (VA) and compare the response parameters to simultaneous measures of disparity vergence and blur driven accommodation. <br /><br /> <em>Methods:</em> A disparity stimulus generator (DSG) was modified to allow step stimulus demands of disparity to be created on a 0. 2 cpd non-accommodative difference of Gaussian target. Retinal disparity of different step amplitude demands were created as an ON / OFF paradigm and projected on a stereo monitor set at a distance of 1. 2m. Two experiments were conducted. The first experiment investigated the first order properties of VA in comparison to similar measures of blur driven accommodation (BA). The second study aimed at comparing the first order and second order dynamics of disparity vergence, VA and BA. <br /><br /> In the first experiment, stimulus measures of vergence, vergence-accommodation and BA were studied. Six normal young adult subjects participated in the study. Accommodation was measured continuously at 25Hz with the commercially available PowerRefractor (Multichannel systems, Germany). A Badal optical system was designed and accommodative response to step stimulus demands were measured. VA and BA measures obtained from the PowerRefractor were matched and plotted as main sequences (amplitude vs. peak velocity). Peak velocities between the two responses were compared using two-way analysis of variance (ANOVA) with Bonferroni post-tests. <br /><br /> In the second experiment, the response dynamics of vergence, vergence-accommodation, and blur accommodation were assessed and compared on 6 young adult subjects. Eye position was measured continuously by a stereo eye tracker at a sampling rate of 120Hz. A high speed photorefractor (sampling = 60Hz) was custom designed and synchronized with a stereo eye tracker to allow simultaneous measurement of vergence and VA. Monocular blur driven accommodation measures were also obtained with the Badal optometer and the high speed photorefractor (sampling = 75Hz). VA, BA and disparity vergence responses were analyzed and temporal parameters like latency, amplitude, duration, time to peak velocity, peak acceleration, duration of acceleration, and skewness were calculated. Main sequence plots (response amplitude vs. peak velocity) were generated and compared between disparity ON and disparity OFF. The dynamic measures of VA were compared to the measures of monocular blur driven accommodation. All comparisons were done using a two-way ANOVA with Bonferroni post-tests. <br /><br /> <em>Results:</em> <br /> <em>Study 1:</em> The results showed that response amplitude of VA during disparity ON and disparity OFF paradigms was linearly related to the peak velocity for an amplitude range of 0. 5 to 2. 5 D (Disparity ON: peak velocity of vergence-accommodation = 0. 812 * amplitude + 1. 564, R² = 0. 452, p<0. 0001 and Disparity OFF: peak velocity of vergence-accommodation = 1. 699* amplitude ? 0. 234, R² = 0. 86, p <0. 0001). The rate of change of peak velocity as a function of response magnitude was lower for VA during disparity ON compared to VA during disparity OFF. BA responses also showed amplitude dependent dynamic properties (Accommodation peak velocity = 1. 593 * amplitude - 0. 008, R² = 0. 84, p<0. 001; Dis-accommodation peak velocity = 1. 646 * amplitude - 0. 036, R² = 0. 77, p<0. 001). There was no statistical difference in the velocity of accommodation and dis-accommodation. <br /><br /> <em>Study 2:</em> When amplitudes were matched, disparity vergence response during disparity ON and disparity OFF had similar main sequence relationships. The mean values for the stimulus and response VA/V ratios were similar (0. 13±0. 05 D/&Delta; and 0. 15±0. 09 D/&Delta; respectively). All the temporal parameters of vergence-accommodation were similar during disparity ON and disparity OFF paradigms. When blur accommodation and vergence-accommodation measures were compared, all the first order and second order temporal parameters in the response were similar between the two systems. Also, disparity vergence exhibited significantly greater peak velocity and peak acceleration compared to two accommodation responses. The results also confirmed that the velocity of accommodation and dis-accommodation showed a statistically significant linear relationship as a function of amplitude for the range of amplitudes tested (Accommodation, y = 2. 55x + 0. 65, R² = 0. 55, p<0. 0001; Dis-accommodation, y = 2. 66x + 0. 50, R²=0. 65, p<0. 0001). <br /><br /> <em>Conclusions:</em> The dynamic properties of VA are amplitude dependent. Although initial results from study 1 suggested that VA may be slower during disparity ON, the results from study 2 using the high speed photorefractor and an improved analysis procedure showed that VA responses were equally fast between disparity ON (convergence) and disparity OFF (divergence). All temporal properties of VA were independent of vergence type (convergence/divergence). VA and BA have similar dynamic properties in humans suggesting that they may controlled by a common neural pathway or limited by the plant. Also, when compared to accommodation responses, disparity vergence exhibited greater velocities and accelerations reflecting the differences in the magnitude of neural innervation and plant mechanics between the two systems. The study also confirmed amplitude dependent response dynamics of blur driven accommodation and dis-accommodation.

Thesis--Universität Erlangen-Nürnberg. / At head of title: Aus dem Seminar für Geschichte der Medizin der Universität Erlangen-Nürnberg. Vita. Includes index.