The goal of this project was to examine the visual system of the bonnethead shark (Sphyrna tiburo). The eyes of this shark are located at the extreme lateral ends of a broad, elongated cephalofoil. Better understanding of their visual system may aid in determining the adaptive benefits of their usual head shape. The proposed project examined one specific aspect of their visual system: the organization of retinal ganglion cells and identification of areas of increased resolution. Two experiments were conducted to realize these aims: (1) staining of retinal ganglion cells, to examine their distributional pattern, and (2) retrograde staining of retinal ganglion cells to determine morphology.

Doctor of Philosophy (PhD) / Background: The development of the blood supply in the primate retina has been extensively studied; however the relationship of the differentiating retina to the choroidal blood supply is less well known. The interaction of astrocytes and vascular endothelial cells promotes the development of the retinal vasculature from 14 weeks’ gestation (WG). Initially, astrocytes lead the developing capillaries from the optic nerve towards the macular area. However, neither astrocytes nor endothelial cells enter a prescribed avascular area, within which the fovea later forms. This may be attributed to expression of a factor that inhibits astrocyte and endothelial cell proliferation in the fovea. A factor found in the CNS that is already known to have these effects is transforming growth factor-β (TGF-β). Aims: This thesis investigated the relationship between retinal maturation and choroidal blood vessel supply and the possible role for TGF-β as an antiangiogenic factor in maintaining an avascular fovea during primate retinal development. Methods: Human eyes between 11 WG and 40 years were obtained with ethical approval from Prince of Wales Hospital and the NSW Lions Eye Bank and fixed and sectioned for histological procedures or prepared for polymerase chain reaction (PCR). Macaque eyes from foetal day (fd) 64 to postnatal 11years (p11y) were obtained from Bogor Agriculture University, Indonesia with the approval of the Ethics Committee of the University of Washington, Seattle, USA. Macaque eyes were also fixed and sectioned for immunohistochemistry and in situ hybridisation. RNA was extracted from human foetal retinas and used for RTPCR (Reverse Transcriptase PCR), QPCR (Quantitative PCR) and preparation of riboprobes. PCR products were analysed using both restriction digest and sequencing. RTPCR was used to identify TGF-β1, TGF-β2 and TGF-β3 in the developing human and in the developing and adult macaque retinas whilst QPCR was used to quantify the TGF-β isoforms in central compared to peripheral retina and in foetal compared to adult retina. In situ hybridisation was performed according to a standard protocol and visualised using Roche HNPP Fast Red detection set with designed riboprobes for TGF-β1, TGF-β2 and TGF-β3 (DIG RNA labelling kit). Some sections were counterstained with vimentin antibody. Immunohistochemistry was performed on human retina and choroid sections using antibodies to CD34 and Ki67 and on human and macaque retina using antibodies to synaptophysin, vimentin, GFAP, calbindin, S-opsin, RG-opsin, rhodopsin, TGF-β1, TGF-β2, TGF-β3 and their receptors TβRI and TβRII. Sections of the retina were imaged and analysed using either a Leica Confocal microscope and TCSNT software or Zeiss Confocal microscope and LSM 5 Pascal software. Data from the human retina and choroid sections corresponding to different regions (foveal, parafoveal nasal, parafoveal temporal, nasal and temporal) was collected to measure the number of Ki-67 immunolabelled mitotic endothelial cells and the area of CD34 immunolabelled choriocapillaris using Adobe Photoshop version 5.0.2, NIH software version 1.62 (measurement macros) and Excel. In the human and macaque sections the intensity of TGF-β protein and mRNA expression was captured from different regions of the retina (foveal, parafoveal nasal, parafoveal temporal, nasal, temporal, nasal to disc) to compile montages. Montages were then re-imported into LSM 5 Pascal software to measure the optical density across each montage along the ganglion cell layer, outer neuroblastic zone and photoreceptor layer collecting data in Excel for graphical representation. In addition to the montages, individual sections were assessed for co-localisation of TGF-β and TβR to various retinal cell types. Results: Analyses of choriocapillaris area and endothelial cell (EC) proliferation were able to demonstrate that the area of choriocapillaris endothelium is greater in the foveal region at all ages (14-18.5WG), that the rate of choriocapillaris EC proliferation declines dramatically over this same period and that the lowest rates of EC proliferation are at the incipient fovea. Most importantly these findings indicate that EC proliferation in the choriocapillaris does not appear to be promoted by increased metabolic activity in central retinal neurons which are more developed with higher oxygen and nutrient demands, which is the mechanism widely thought to regulate development of the retinal vasculature. PCR showed all TGF-β isoforms to be present in the human developing and adult retina. QPCR revealed that TGF-β2 was the most predominant isoform, followed by TGF-β3 with very small amounts of TGF-β1 seen. The isoforms were more abundant in developing rather than adult retina and in central rather than peripheral retina. Studies of the distribution of TGF-β protein and mRNA using immunohistochemistry and in situ hybridisation confirmed the low levels of TGF-β1 protein and mRNA observed in QPCR and demonstrated distinct centroperipheral gradients in the photoreceptor layer for TGF-β2 and TGF-β3. Relative high amounts of TGF-β in the fovea could affect vascular patterning due to TβRI seen in astrocytes which lead the blood vessels at the foveal rim at the level of the ganglion cell plexus. TGF-β2 and TGF-β3 expression is detected before formation of the foveal avascular zone (FAZ) at fd64 (~15WG) - fd73 (~17WG) with levels peaking in the foveal region at fd105 (~25WG) by the time the FAZ forms. Conclusions: This thesis has shown that EC proliferation in the choriocapillaris does not appear to be promoted by increased metabolic activity in central retinal neurons as reduced rates of EC proliferation in the ‘foveal’ chorioretinal location were observed at all ages studied between 14 and 18.5WG. The findings suggest that mechanisms regulating proliferation and growth of the choroidal vasculature are independent of differentiation in the neural retina and are therefore different to those governing the formation of the retinal vasculature. All TGF-β isoforms are expressed in developing and adult human and macaque retina with TGF-β2 being the predominant isoform. TGF-β isoforms are more abundant in central compared to peripheral retina and in developing compared to adult retina. Centro-peripheral gradients of TGF-β2 and TGF-β3 across the photoreceptor layer and TβRI on astrocytes support the presence of TGF-β in the fovea as an antiproliferative and antiangiogenic factor by helping to define the FAZ early in development, well before 23-25 WG in humans and before fd100 in macaques.

The purpose of this project is to develop a retinal imaging system suitable for routine examination or screening of mouse models that acquires fundus and Optical Coherence Tomography (OCT) images. The imaging system is composed of a digital camera with an objective for biomicroscopic examination of the fundus, an OCT interferometer, an OCT beam delivery system designed for the mouse eye, and a mouse positioning stage. The image acquisition is controlled with software that displays the fundus and OCT images in real-time, and allows the user to control the position of the OCT beam spot on the fundus image display. The system was used to image healthy mice and a mouse model of glaucoma. Fundus images and OCT scans were successfully acquired in both eyes of all mice with eyes that had clear optics. The study demonstrates the feasibility of acquiring simultaneous fundus and OCT images of the mouse retina, by a single operator, in a manner suitable for rapid evaluation of mouse models of retinal disease.

Calcineurin (CaN), a member of ser/thr protein phosphatase, was cloned from bovine retina. The peptide sequence of CaN A subunit is consisted of 511 amino acid residues. A 10 amino acid (A-T-V-E-A-I-E-A-D-E-A) deletion before the autoinhibitory domain was observed in bovine retina CaN A compared to bovine brain CaN A. The study on CaN activity and regulation demonstrated that different metal ions have different effects on its phosphatase activity. Ni2+ was found to be the strongest stimulator while Zn2+ was found to inhibit CaN phosphatase activity. Mn2+ was a relatively less effective stimulator compared to Ni2+. Fe2+ was also able to stimulate CaN phosphatase activity; in contrast, a previous study found Fe2+ slightly inhibited bovine brain CaN activity. The residues at 97-201 were found to be essential for bovine retina CaN A phosphatase activity. The residues at 407-456 also had an inhibitory effect on CaN A phosphatase activity in addition to the previously known auto inhibitory domain at 457-480. These observations suggest that bovine retina CaN A might possess some distinct structural characteristics compared to bovine brain CaN A.

Cell cycle exit (“birth”) of retinal progenitor cells (RPCs) is considered a watershed that is preceded by changing levels of cell cycle regulators, and followed rapidly by induction of a post M-phase differentiation cascade. Yet the actual dynamics of these events are largely unclear, thus whether mitosis separates pre- and post- birth differentiation cascades is unproven. We characterized the regulation of many division and differentiation markers relative to each other and final mitosis. Unexpectedly, classic “cell cycle” markers were present well beyond exit (e.g. Ki67, Pcna), early embryonic RPCs expressed “differentiation” markers that later labeled post-mitotic neurons exclusively (e.g. Brn3b, Tubb3, Ptf1a), and factors detected just after cell birth in the embryo were induced well beyond M-phase post-natally (e.g. Nrl, Crx). Thus, the dynamics of birth-associated events shift dramatically during development, even to either side of mitosis. Instead of mitosis behaving as a cog that activates post-exit differentation events we suggest that a common trigger induces both the exit and differentiation programs in RPCs, precisely coordinating their startpoints, but that each subsequent cascade unfolds independently. This model explains the convergence of birth and differentiation but also their temporal maliability. This view fits with our observation that in the absence of the Rb tumor suppressor, differentiation still initiates even without cell cycle exit. Finally, neoplastic transformation in the mouse retina requires loss of Rb and its relative p107, and emerging tumor features suggest an amacrine cell-of-origin. We studied Rb/p107 null clones, and noted two striking features. First, despite initial expansion of aberrantly dividing differentiating cells, apoptosis pruned clones precisely to wild type sizes. “Cell competition” maintains tissue size by selecting fitter over weaker progenitors; our data provide a unique example of competition among differentiating cells. Second, despite normal numbers of amacrine cells per Rb/p107 null clone, more clones contained amacrine cells and fewer had bipolar cells. Both this effect and ectopic division were E2f1-dependent. Thus, the oncogenic initiation event in mouse retinoblastoma triggers a very early fate switch, even before neoplastic transformation, broadening the possibilities for the cell-of-origin of retinoblastoma, and arguing that even very early stage tumors cannot be used to define cancer origin.

Cell cycle exit (“birth”) of retinal progenitor cells (RPCs) is considered a watershed that is preceded by changing levels of cell cycle regulators, and followed rapidly by induction of a post M-phase differentiation cascade. Yet the actual dynamics of these events are largely unclear, thus whether mitosis separates pre- and post- birth differentiation cascades is unproven. We characterized the regulation of many division and differentiation markers relative to each other and final mitosis. Unexpectedly, classic “cell cycle” markers were present well beyond exit (e.g. Ki67, Pcna), early embryonic RPCs expressed “differentiation” markers that later labeled post-mitotic neurons exclusively (e.g. Brn3b, Tubb3, Ptf1a), and factors detected just after cell birth in the embryo were induced well beyond M-phase post-natally (e.g. Nrl, Crx). Thus, the dynamics of birth-associated events shift dramatically during development, even to either side of mitosis. Instead of mitosis behaving as a cog that activates post-exit differentation events we suggest that a common trigger induces both the exit and differentiation programs in RPCs, precisely coordinating their startpoints, but that each subsequent cascade unfolds independently. This model explains the convergence of birth and differentiation but also their temporal maliability. This view fits with our observation that in the absence of the Rb tumor suppressor, differentiation still initiates even without cell cycle exit. Finally, neoplastic transformation in the mouse retina requires loss of Rb and its relative p107, and emerging tumor features suggest an amacrine cell-of-origin. We studied Rb/p107 null clones, and noted two striking features. First, despite initial expansion of aberrantly dividing differentiating cells, apoptosis pruned clones precisely to wild type sizes. “Cell competition” maintains tissue size by selecting fitter over weaker progenitors; our data provide a unique example of competition among differentiating cells. Second, despite normal numbers of amacrine cells per Rb/p107 null clone, more clones contained amacrine cells and fewer had bipolar cells. Both this effect and ectopic division were E2f1-dependent. Thus, the oncogenic initiation event in mouse retinoblastoma triggers a very early fate switch, even before neoplastic transformation, broadening the possibilities for the cell-of-origin of retinoblastoma, and arguing that even very early stage tumors cannot be used to define cancer origin.

Syfte: Syftet med studien var att, med hjälp av Optical Coherence Tomography (OCT), undersöka om det finns någon skillnad i central och perifer retinal tjocklek mellan olika ametropier. Metod: Studien omfattade 36 försökspersoner, som grupperades beroende på ametropi i en myop, emmetrop och hyperop grupp. Av de 36 försökspersonerna var det 15 myoper, 15 emmetroper och 6 hyperoper. En inledande mätning gjordes där försökspersonernas objektiva refraktion uppmättes med autorefraktor och därefter gjordes en avstämning i provbåge för att säkerställa refraktionen. Med OPKO Spectral OCT/SLO mättes retinas tjocklek både centralt och perifert på höger öga. För att analysera resultatet delades retina in i 15 olika zoner som jämfördes mellan de olika ametropierna. Resultat: Resultatet av studien visade en signifikant skillnad i foveal tjocklek mellan de olika ametropierna (p=0,03). Det var en siginifikant skillnad i retinal tjocklek mellan retinas zoner i alla tre ametropier (p<0,01), dock var det ingen signifikant skillnad i perifer retinal tjocklek mellan de tre olika ametropierna (p=0.07). Slutsats: Ingen skillnad i central och perifer retinal tjocklek kunde redovisas mellan de olika ametropierna. Då en tidigare studie har visat att den retinala tjockleken skiljer sig mellan olika ametropier kan resultatet av vår studie diskuteras då det i vår studie fanns brister som få antal personer, olika antal personer inom grupperna och en låg utbredning av synfel.

OBJETIVODemostrar la eficacia de la Vitrectomía vía Pars Plana sin indentación escleral , aire como taponador y sin decúbito prono en el manejo de roturas inferiores en el desprendimiento de la retina rhegmatógeno pseudofáquico primario.MétodosEstudio prospectivo no comparativo. Cincuenta y cinco ojos (55 pacientes) con desprendimiento de la retina rhegmatógeno primario pseudofáquico asociado a rotura(s) inferior(es) se les realizó Vitrectomía vía Pars Plana sin indentación escleral y aire como taponador. Los pacientes fueron divididos en dos grupos consecutivos; el primer grupo (15 ojos de 15 pacientes) realizaron 24 horas de posición en decúbito prono y el segundo grupo (40 ojos de 40 pacientes) no realizaron posicionamiento en el período postoperatorio. E período de seguimiento fue de 6 meses.ResultadosLa tasa de reaplicación anatómica inicial fue de 93,3% y 90% para los grupos con 24 horas de posición en decúbito prono y sin posicionamiento respectivamente. La tasa de reaplicación final (6 meses) fue alcanzada en el 100% de los casos. En el primer grupo la media de la mejor agudeza visual corregida preoperatoria fue 20/60 (rango 20/400 a 20/25) y la media postoperatoria fue 20/30 (rango 20/100 a 20/20). En un paciente la retina se redesprendió en la segunda semana por una rotura no detectada. Otro paciente desarrollo una membrana epirretiniana. En el segundo grupo la media de la mejor agudeza visual corregida preoperatoria fue 20/63 (rango de movimiento de manos a 20/20) y la media postoperatoria fue 20/30 (rango 20/60 a 20/20). Un paciente (2,5%) desarrollo proliferación vitrerretiniana en la sexta semana después de la cirugía. En 3 pacientes (7,5%) la retina se redesprendió presumiblemente debido a roturas no tratadas. Dos pacientes (5%) desarrollaron una membrana epirretiniana.Conclusiones La técnica de Vitrectomía vía Pars Plana sin indentación escleral y aire demostró ser efectiva en el Desprendimiento de la Retina Rhegmatógeno Pseudofáquico primario asociado a roturas inferiores con una baja tasa de complicaciones. Este estudio proporciona evidencia que dentro de las primeras horas del postopeatorio se crea una adherencia coriorretiniana suficiente para el desarrollo de una adhesión impermeable al paso de fluido, lo que nos permite reducir la necesidad del posicionamiento postoperatorio y el uso de aire como taponador. / ObjectiveTo determine the efficacy of a pars plane vitrectomy without scleral buckling while using air as a tamponade and no facedown positioning to manage inferior breaks in primary pseudophakic rhegmatogenous retinal detachment.MethodsProspective, noncomparative study was carried out. A total of 55 patients diagnosed with primary pseudophakic rhegmatogenous retinal detachment with inferior breaks were enrolled in two consecutive groups. Group 1: Fifteen patients (15 eyes) with 24 hours of prone positioning in the post operative period, and Group 2: Forty patients (40 eyes) with no facedown positioning during the post operative period. All 55 patients had a pars plana vitrectomy without scleral buckling and air as a tamponade performed to one of their eyes (55 eyes).ResultsThe anatomic reattachment rate was 93.3% for Group 1 and 90% for Group 2. The final reattachment rate was reached on 100% of the cases. For Group 1 the mean pre-operative best-corrected VA (BCVA) was 20/60 (range 20/400 to 20/25) and the mean post-operative best-corrected visual acuity was 20/30 (range 20/100 to 20/20). In one case the retina re-detached at the second week because of an undetected break. A postoperative epiretinal membrane was also observed in another patient. For Group 2 the mean pre-operative best-corrected VA (BCVA) was 20/63 (range, hand movements to 20/20) and the mean post-operative best-corrected visual acuity was 20/30 (range 20/60 to 20/20). One patient (2.5%) developed proliferative vitreoretinopathy at the sixth week after surgery. Three patients (7.5%) redetached, presumably due to untreated breaks. Two patients (5%) developed a macular epiretinal membrane that required surgery. Conclusions The surgical technique of pars plane vitrectomy without scleral buckling and air was effective (and with a low rate of complications) on the treatment of primary pseudophakic rhegmatogenous retinal detachment associated to inferior breaks. This study shows evidence that within the first hours of the post-operative period a chorioretinal adhesion gets generated that reduces the need for post-operative face-down positioning.

Propósito: Determinar cuáles son los principales factores que influyen en la impermeabilidad intraoperatoria de las roturas retinianas en el desprendimiento rhegmatógeno primario (DRR). Los objetivos secundarios son, valorar si es posible hacer impermeables las roturas al paso de fluido sin necesidad de taponamiento postoperatorio, determinar si el estado del cristalino influye sobre la técnica quirúrgica. Analizar las complicaciones intraoperatorias, postoperatorias y resultados anatómicos y funcionales.Material y métodos: Estudio prospectivo de 82 ojos de 82 pacientes con DRR. Los pacientes fueron intervenidos mediante Vitrectomía vía Pars Plana (VPP), las roturas se trataron mediante la disección exhaustiva del vítreo con ayuda de perfluorocarbono líquido, un "secado" intenso de la interfase retina-epitelio pigmentario y la realización intraoperatoria de láser bajo aire. Al final de la intervención se realizó intercambio por solución salina balanceada (BSS) y se estudió el comportamiento de las roturas tratadas. Se estudió el éxito anatómico y funcional al mes, tres y seis meses después de la intervención. Las medidas de asociación entre variables categóricas se evaluaron mediante el test exacto de Fisher. Los contrastes de hipótesis de igualdad de medias se han llevaron a cabo tras transformación del logaritmo de la inversa de las variables (LogMAR) aplicandotanto el test de t Student y ANOVA de comparación de medias.Resultados: El éxito anatómico primario fue del 84,1%. No influyeron en la reaplicación el tipo, número o localización de las roturas. Tampoco influyó en la tasa de reaplicación el estado del cristalino, el tipo de cirugía utilizada o el tipo de fotocoagulación. Tuvieron menos tasa de reaplicación los pacientes con proliferación vitreorretiniana en grado B pero no fue estadísticamente significativo. La agudeza visual media preoperatoria fue de 20/80, y la postoperatoria de 20/40. El 46,4% de los pacientes con mácula desprendida antes de la intervención, obtuvieron agudezas visuales superiores a 20/50. La tasa de proliferación vitreorretiniana postoperatoria fue del 2,4%. El porcentaje de complicaciones fue inferior a otras series de DRR.Conclusiones: Mediante la disección exhaustiva del vítreo alrededor de las roturas, un taponamiento intraocular de corta duración y la realización intraoperatoria de láser de diodo es posible realizar un sellado de las roturas retinianas, que las impermeabiliza al paso de fluido, sin necesidad de dejar un taponador tras la intervención. Los principales factores que han influido en la impermeabilidad de las roturas son la visualización y el tratamiento completo de éstas. No influyeron el número de cuadrantes afectos, las roturas múltiples, la existencia de roturas superiores a una hora, las roturas inferiores o el tiempo de evolución del desprendimiento. El estado del cristalino no influyó en el sellado de las roturas. El índice de complicaciones, los resultados anatómicos y funcionales han sido similares a otras técnicas quirúrgicas para el desprendimiento de retina / Purpose: To evaluate factors associated with intraoperative sealing of retinal breaks in the primary rhegmatogenous retinal detachment (RRD). Secondary objectives are evaluate the possibility of retinal breaks sealing without postoperative tamponade, influence of lens status in retinal detachment surgery, avaluation of intraoperative and postoperative complications, anatomic and functional outcomes.Methods: Prospective study of 82 eyes in 82 patients with RRD. Pars plana vitrectomy was performed in all patients, retinal breaks were treated with intensive vitreous peeling around the breaks, and with diode laser under air. At the end of the surgery saline balanced serum was introduced into the vitreous cavity and a meticulous study of retinal breaks was performed. We evaluate anatomic and functional outcomes in the first month, third and sixth months after surgery. Results: Primary anatomic success was 84,1%. Kind, number or placement of retinal breaks didn't influence the anatomic success, neither lens status nor surgical procedure. Patients with vitreo-retinal proliferation (PVR) type B had less rate of success than the others but there was no statistical difference. Mean preoperative visual acuity was 20/80 and postoperative was 20/40. The 46,4% of macula-off patients ended the study with visual acuity over 20/50. Postoperative PVR rate was 2,4%. Conclusions: With intensive vitreous peeling around the breaks, intraocular short tamponade, and intraoperative diode laser is possible to seal retinal breaks, without using a postoperative tamponade. Main factors in surgical success are visualization and treatment of all breaks, kind of RRD, multiple, larger or inferior breaks, evolution time or lens status didn't show any influence in success rate. Rate of complications, anatomic and functional outcomes was similar than other studies.

PURPOSE: Systemic hyperoxia has been induced using inspired gases in many studies to investigate vascular reactivity in the retinal vasculature. Technical limitations in the past resulted in inadequate control of systemic partial pressures of O2 and CO2, the latter of which tended to decrease secondary to induced hyperoxia. Recent development of a computerized gas delivery instrument has enabled the specific control of end-tidal CO2 (ETCO2) and fractional expired O2 (FeO2), independent of each other and of minute ventilation. The specific aims of each chapter are as follows: Chapter 3: To compare the magnitude and variability of the retinal vascular reactivity response to an isocapnic hyperoxic stimulus delivered using a manually-operated method to the newly developed computer-controlled gas sequencer. Chapter 4: To investigate the retinal hemodynamic response to incremental changes in hyperoxic stimuli during isocapnia. METHODS: Chapter 3: Ten young, healthy adults inhaled gases in a sequence of normoxic baseline, isocapnic hyperoxia, and normoxic recovery, using both gas delivery systems in random order. Chapter 4: Twelve healthy, young adults participated in a gas protocol consisting of 4 phases at varying fractional expired oxygen levels (FeO2): baseline (15%), hyperoxia I (40%), hyperoxia II (65%), and recovery (15%). End-tidal carbon dioxide (ETCO2) was maintained at an isocapnic level (~ 5%) throughout the experiment. In both Chapters 3 and 4, blood flow was derived from retinal arteriolar diameter and simultaneous blood velocity measurements of the superior temporal arteriole, acquired at 1-minute intervals during each of the phases of the gas protocol. RESULTS: Chapter 3: There was no interaction effect between the phases and gas delivery methods (p = 0.7718), but ETCO2 was significantly reduced during hyperoxia (p = 0.0002) for both methods. However, the magnitude of change in ETCO2 was physiologically insignificant i.e. <1%. The two systems differed in terms of FeO2 during hyperoxia, at a level of 85.27 ± 0.29% for the manual method, and 69.02 ± 2.84% for the computer method (p < 0.05). Despite this difference in oxygen concentrations, there was no difference in the vascular reactivity response for diameter (p = 0.7756), velocity (p = 0.1176), and flow (p = 0.1885) for equivalent gas phases between the two gas delivery systems. The inter-subject variability of retinal hemodynamic parameters was consistently lower using the computer-controlled gas sequencer. Chapter 4: Repeated measures ANOVA showed that there were significant influences of incremental changes in FeO2 on arteriolar diameter (p < 0.0001), blood velocity (p < 0.0001), and blood flow (p < 0.0001) in the retina. Paired t-tests of these retinal hemodynamic parameters during each phase in the gas sequence showed they were significantly different (p < 0.05) from each other, with the exception of baseline and recovery values. Incremental increases in FeO2 caused a linear decrease in group mean arteriolar diameter (R2 = 1, p = 0.002), group mean blood velocity (R2 = 0.9968, p = 0.04), and group mean blood flow (R2 = 0.9982, p= 0.03). CONCLUSIONS: Chapter 3: Inter-subject variability for virtually all retinal hemodynamic parameters was reduced using the computer-controlled method, presumably due to a higher degree of gas control. However, care needs to be exercised in the interprtetation of these results due to the relatively small sample size. A similar retinal hemodynamic response to isocapnic hyperoxia was induced using the two gas delivery systems, despite different levels of maximal FeO2. Chapter 4: Isocapnic hyperoxia elicits vasoconstriction and the reduction of retinal arteriolar blood flow in a dose-dependent manner over the range of FeO2 explored in this study.