Dry eye is a complex, multi-factorial disease that results in a compromised tear film and ocular surface. Clinicians and researchers alike have historically relied on an individual’s symptoms to diagnose and manage the condition, due to a lack of reliable objective methods for quantifying disease presence and severity. Of late, parameters such as tear film osmolarity and tear meniscus height have shown promise as valid methods for enumerating characteristics of the tear film that may aid the diagnosis of dry eye. Two new technologies have recently been introduced that can measure said parameters. The TearLab™ is a novel handheld nano-osmometer capable of measuring tear film osmolarity on samples as small as 50 nL. The device uses electrical conductance to measure osmolarity, and the small sample requirements purportedly allows the device to minimally disturb the natural state of the tear film. The RTVue-100 is a spectral-, or Fourier-domain optical coherence tomographer that has the ability to generate high resolution, cross-sectional images of the tear meniscus, and subsequently measure tear meniscus height. As little is published on the use of these technologies to evaluate the tear film, a series of studies was completed to determine their performance in both a normal and dry eye population.

Purpose: There is little agreement on the age at which visual acuity (VA) and contrast sensitivity (CS) become adult-like. The ultimate purpose of this thesis was to determine whether VA and CS are adult-like at the age of 6-8 or 9-12 years by using both objective and subjective methods in the same individuals. The objective method (sweep visually evoked potentials [sVEP]) has many parameters that may affect the measurement of VA or CS and previously these had not been studied systematically, especially in children. Therefore, a second purpose was to study the effects of these parameters on VA and contrast thresholds and to determine the parameters that give the most repeatable measurements and the greatest number of viable readings in children, to be compared to previous data obtained in adults. Methods: The effect of five criteria (C0-C4) for choosing the endpoint for the regression line fitting and three luminance levels (25, 50, and 100 cd/m2) on the sVEP VA and contrast thresholds (at 1 and 8 cpd) was investigated in six 6-8 year old children. Additionally, the effect of these parameters on the number of viable readings obtained from five active electrodes was investigated. C0 was derived from the sVEP software (PowerDiva), C1 used the best fit by eye to determine the range over which the regression line was fitted, C2 used the data point between signal peak and the last data point with an SNR ≥ 1, C3 was similar to C2 but was defined so that the threshold should be within the sweep range, and C4 was similar to C2 except that the SNR should not fall below one at any point within the range used for the regression line fitting. The effects of two electrode placements, three temporal frequencies (6, 7.5, and 10 Hz), sweep direction (low to high and high to low), presence or absence of a fixation target, three stimulus areas (6, 4, and 2° for VA and 15, 10, and 6° for contrast thresholds) and three sweep durations (10, 15, and 20 sec) on VA and contrast thresholds (at 1, 4, or 8 cpd) measured with sVEP were also investigated in six 6-8 year-old children and six adults with normal vision. Additionally, the effect of these parameters on the number of viable readings obtained from five active electrodes was investigated. The sVEP parameters that were found to give the best threshold measurements were employed in a cross sectional study of the development of VA and CS. In this study the objective sVEP technique and two psychophysical techniques were used. The psychophysical techniques were comprised of a two-alternative forced choice (2AFC) staircase for measuring VA and contrast thresholds and signal detection theory (SDT) for measuring contrast threshold. Crowded and uncrowded logMAR VA were also measured with a Bailey-Lovie logMAR chart. The study included three age groups (6-8, 9-12 year olds and adults). The criterion employed by each age group as indicated by the SDT was compared. Results: There was a significant effect of the criterion for choosing the endpoint for the regression line fitting (p < 0.05) on all the measures and a significant effect of luminance (p = 0.036) on contrast threshold at 1 cpd. Criterion C2 (in which the range for the regression line fit was defined to include all the data between the signal peak and the last data point [furthest from the peak] with an SNR ≥ 1) consistently gave more viable readings and better thresholds (i.e. higher VA and lower contrast thresholds) than the other criteria. Also C2 was the best criterion in terms of repeatability in children, and repeatability and validity in adults (Yadav et al., 2009). The luminance of 25 cd/m2 gave higher contrast thresholds than 50 or 100 cd/m2. There was a significant effect of temporal frequency on the number of viable readings for VA (p < 0.0001) and for contrast thresholds (p = 0.0001), with more viable readings at 7.5 Hz than at either 6 or 10 Hz. The adults gave more readings with the fixation target than without it (p = 0.04) for contrast threshold at 1 cpd. The smallest stimulus area used gave rise to fewer viable readings in both adults and children (p = 0.022 for VA and 0.0001 for contrast threshold). The other parameters (electrode placement, sweep direction and sweep duration) did not result in significant differences. There was a significant effect of age on crowded (p = 0.0001) and uncrowded (p < 0.0001) VA. The 6-8 year olds gave poorer VA than the 9-12 year olds or adults for both crowded and uncrowded VA. For the grating VA (sVEP and 2AFC staircase) there was a significant effect of age (p = 0.002). The 6-8 year olds had poorer VA than the 9-12 year olds or adults. For contrast threshold at 1 cpd, a significant effect of age was found for the 2AFC (p = 0.008) and SDT (p = 0.0003). The 6-8 year olds gave poorer contrast thresholds than adults with each procedure. For contrast thresholds at 8 cpd, there was a significant effect of age with the 2AFC staircase (p = 0.036). The 6-8 year olds gave poorer contrast thresholds than the 9-12 year olds. For SDT, there was a significant effect of age on criterion (p < 0.05), with adults being more likely to say “no” in the yes-no SDT procedure than both the 6-8 year olds and the 9-12 year olds for contrast threshold at 1 cpd. Adults were also more likely to say “no” than the 9-12 year olds for contrast thresholds at 8 cpd. Conclusions: This thesis has shown that VA and CS are not adult-like until the age of 9-12 years by these measures and that children do show differences in criterion compared to adults in psychophysical testing. This difference in criterion indicates the use of SDT or force-choice procedures to avoid this problem in any psychophysical developmental study. It has also shown that criterion for choosing the endpoint for the regression line fitting in the sVEP technique has the greatest effect on VA and contrast thresholds measurements and viable readings, while the other sVEP parameters have little effect on the thresholds.

Purpose To investigate ocular surface sensations, specifically ocular discomfort using psychophysical and clinical techniques. The measurement of discomfort on the ocular surface has been limited to the use of traditional rating scales until recently. This thesis focuses on the scaling of discomfort using a psychophysical approach and also investigates the less explored area of the influence of blur on ocular discomfort. The specific aims of each chapter are: Chapter 2: To evaluate the difference thresholds of the central cornea in lens and non-lens wearers. Chapter 3: To devise a novel scale for ocular discomfort, relating subjective estimation of discomfort arising from contact lens wear to discomfort produced by the pneumatic stimuli delivered by a modified Belmonte esthesiometer. Chapter 4: To evaluate the influence of blur on ocular comfort while systematically manipulating vision using habitual refractive correction, induced spatial and optical blur, and under the absence of visual structure. Chapter 5: To examine if subjects rate discomfort and intensity of suprathreshold pneumatic stimuli differently when viewing clear and defocused targets and to examine the suprathreshold scaling of stimuli under the same visual conditions. Methods Chapter 2: The mechanical sensitivity of the central cornea was determined in 12 lens wearers and 12 non-lens wearers using a modified Belmonte pneumatic esthesiometer. The mechanical threshold of the central cornea was first estimated using the method of limits. Then, a series of systematically increasing stimuli were presented, with the first stimuli being 25% less than the threshold. The subjects were asked to compare the intensity of each stimulus with the preceding one and report if any difference in intensity was detectable. The intensities at which the subjects perceived an increased intensity from the previous was recorded. The difference threshold (DL) was the differences between the stimulus intensities at which an increase was perceived and five DLs were measured for each subject. Weber’s constants that relate the size of the difference thresholds to the stimulus intensity were derived for each DL level and repeated measures ANOVA was used to compare the Weber’s constants in the lens and non-lens wearing groups. Chapter 3: Twenty seven participants were enrolled for this magnitude matching study. Soft (HEMA) contact lenses of eight different lens designs varying in base curve and diameter were fit on all participants. The study was conducted on two separate days with four lenses randomly assigned on each day. The assigned soft contact lens was placed on the chosen eye and the sensations were measured using a numerical rating scale. Following this, the subjects were asked to regulate the intensity of the pneumatic stimulus using the control dial in order to match the discomfort from the stimulus to the discomfort from contact lens wear. At the completion of magnitude matching, ratings of sensations were again recorded. Pearson product moment correlation was used to correlate the objective esthesiometer matches to the subjective ratings of discomfort reported by each participant. The method of least log squares was used to derive the power exponents as defined by Stevens’ power law and analyze the psychophysical functions. Repeated measures ANOVA was used to investigate the effect of lens sequence and session on ocular discomfort with contact lens wear. The impact of lens type and time on discomfort was studied using linear mixed modeling. Chapter 4: Twenty emmetropic subjects rated ocular comfort, vision and sensation attributes (burning, itching and warmth) under conditions of normal vision, spatial blur and dioptric defocus, each session lasting for five minutes. Subjects viewed digital targets projected from a distance of 3m, and ocular surface sensations, vision were rated using magnitude estimation. Dioptric defocus was produced using +6.00DS contact lenses and equivalent spatial blur was created by spatially blurring the targets. Clear target images were used during dioptric defocus and blurred images during spatial blur session. Comfort was also rated under the absence of visual structure in fifteen of the participants using a ganzfeld and black occluders. Repeated measures ANOVA was used to compare vision and comfort ratings between the different experimental conditions. Chapter 5: Twenty one participants were enrolled. Ocular discomfort was produced by delivering mechanical stimuli from a pneumatic esthesiometer, and participants were asked to rate the intensity of stimulus and the discomfort induced by it under clear and defocused visual conditions. Esthesiometry was performed on one eye while the fellow eye viewed either a clear or blurred 6/60 fixation target through a trial lens. For the clear visual condition, the trial lens contained +0.25DS over their distance refractive correction and for the defocused condition, an additional +4.00DS was used. Mechanical thresholds from the central cornea were estimated using ascending methods of limits and then stimuli that were 25%, 50%, 75% and 100 % above threshold were presented in random order. Participants rated intensity and discomfort of each stimulus using a 0-100 numerical scale where 0 indicated no sensation and 100 indicated highest imaginable intensity/discomfort. There were 3 sessions with clear visual conditions and 3 sessions with defocus, in random order. Results Chapter 2: The functions relating Weber’s constants to stimulus intensities were slightly different in lens and non-lens wearing groups, although the absolute thresholds were similar. Repeated measures ANOVA revealed a significant main effect of DL level on Weber’s constant (p<0.001), with the Weber’s fraction at the first DL being higher than the following DLs. A significant main effect of the group type was also observed, with the lens wearers showing higher Weber’s constants than the non-lens wearers (p=0.02) However, there was no interaction between DL level and lens wearing group on Weber’s constants (p=0.38). Chapter 3: The average and individual psychophysical functions appeared to follow Stevens’ power function, with mechanical and chemical stimuli giving rise to different power exponents. Examination of the individual transducer functions revealed that only about half of the subjects were able to match the contact lens sensations to the pneumatic stimulus discomfort, with both mechanical and chemical stimulation. The lens types did not have any impact (p=0.65) on the session or sequence in which the lens was presented, although an effect of session and sequence on discomfort was observed. The average discomfort ratings produced by the different lens types were similar. There appeared to be significant effects of time (p<0.001) on the reporting of discomfort with lens wear, with the discomfort upon lens insertion rated to be higher than after lenses settling. Chapter 4: Ratings of vision under spatial blur and dioptric defocus were significantly different (p<0.001) from normal vision condition. Vision with dioptric defocus was rated worse (p<0.001) than spatial blur. Significant differences in comfort were observed between normal vision and blur, including spatial blur (p=0.02) and dioptric defocus (p=0.001). However, there was no significant difference (p=0.99) in comfort between spatial blur and dioptric defocus. Comfort remained unchanged between normal vision, occluders and ganzfeld although vision was absent in the later two conditions. Chapter 5: There was no significant difference in mechanical thresholds under clear and defocused conditions with a paired t-test (p=0.66) and similar results were obtained with repeated measures ANOVA, with no significant difference in discomfort (p=0.10) and intensity (p=0.075) ratings between the two visual conditions. However, paired t-test between the derived exponents under clear and defocused conditions showed significant differences for discomfort (p=0.05) and no significant difference for the ratings of intensity (p=0.22). Comparison of exponents between discomfort and intensity showed a significant difference in both clear (p=0.02) and defocus conditions (p<0.001). Conclusions: Chapter 2: The differential sensitivity of the ocular surface can be successfully measured with a pneumatic esthesiometer and it appears that Weber’s law holds true for corneal nociceptive sensory processing. There are subtle differences in mechanical difference thresholds between lens and non-lens wearers suggesting the possibility of different neural activity levels in the two groups. Chapter 3: Subjective ratings of discomfort can be scaled by corneal esthesiometry in a selective group of people. In the subset of subjects with poorer correlations, perhaps the pneumatic mechanical stimulus was too localized and specific to match the complex sensations experienced while wearing contact lenses. However, there is also a group of subjects who are poor at making judgments about ocular comfort. Hence, the use of special sensory panels should be considered when ocular comfort is the primary outcome. Chapter 4: There does seem to be an association between clarity of vision and ocular comfort, although the pathways for pain and vision are perhaps exclusive. Interactions between vision and other senses have been reported, but a similar inter-sensory interaction between pain and vision is yet to be clearly demonstrated. The decreased comfort observed in this study might perhaps be due to nocebo or Hawthorne effects. Chapter 5: Suprathreshold scaling of pneumatic stimuli can vary with the viewing conditions, with defocus associated with higher exponents than clear visual conditions. However, the ratings of comfort appear to be similar under both the conditions. If defocus does affect comfort, it is subtle and does not affect the sensory components, but tiny effects through the affective aspect of pain can contribute to the differences in power exponents. The differences in the perception of comfort do not appear to be attributable to the differences in threshold or sensory intensity.

Purpose To investigate the impact of cigarrete smoking in a group of otherwise healthy young individuals on: 1) Retinal blood flow using Doppler based SD-OCT, 2) Retinal vascular reactivity using a gas sequencer to provoke hypercapnia via constant changes in PETCO2 (end-tidal partial pressure of CO2) and in PETO2 (end-tidal partial pressure of O2). Methods An automated gas flow controller was used to achieve normoxic hypercapnia in ten non-smokers (mean age 28.9 yrs, SD 4.58) and nine smokers (mean age 27.55 yrs, SD 4.77). Retinal blood flow measurements were obtained using Doppler OCT and cannon laser blood flowmeter (CLBF) during baseline, normoxic hypercapnia (15% increase in PETCO2 relative to homeostatic baseline) and post-hypercapnia in both the groups. Exhaled carbon monoxide level was measured in all subjects. Results In non-smokers, retinal arteriolar diameter, blood velocity and flow increased by +4.1% (SD 2.8, p<0.0001), +16.7% (SD 14.6, p=0.0004) and +29.6% (SD 12.5, p<0.0001) respectively, during normoxic hypercapnia; Similarly, the venous area, venous velocity and total retinal blood flow increased by 7% (SD 8.6, p=0.0418), 18.1% (SD 20.8, p=0.0068) and 26% (SD 22.9, p<0.0001) respectively. In smokers, normoxic hypercapnia resulted in a significant increase in velocity by 12.0% (SD 6.2, p=0.0019) and flow by 14.6% (SD 9.5, p=0.0029); though arteriolar diameter increased by 1.7% (SD 1.7, p=0.2616), the result was not statistically significant. Total retinal blood flow increased significantly by 19.3% (SD 18.4, p=0.002) in response to normoxic hypercapnia. However, there was no significant difference in venous area (p=0.3322) and venous velocity measurements (p=0.1185) during hypercapnia compared to baseline and recovery. Comparing smokers and non-smokers, only the percentage change in arteriolar diameter (p=0.0379) and flow (p=0.0101) was significantly different among the groups. Group mean PETCO2 was increased by 15.9% in the non-smoking group and by 15.7% in the smoking group, with a concomitant increase in PETO2 by approximately 1.5 to 2% in both groups. There was no significant difference in baseline PETCO2 level between smokers and non-smokers. Conclusions Retinal vascular reactivity in response to normoxic hypercapnia is significantly reduced in young healthy individuals who smoke compared to non-smokers. Further studies are needed to elucidate the exact reason behind the impaired retinal autoregulation to provocative stimuli in smokers.

No description available.

Saccadic eye movements, produced by the oculomotor system, are used to bring salient information in line with the high resolution fovea. It has been suggested that visual working memory, the cognitive system that temporarily stores and manipulates visual information (Baddeley & Hitch, 1974), is utilised by the oculomotor system in order to maintain saccade programmes across temporal delays (Belopolsky & Theeuwes, 2011). Saccadic eye movements have been found to deviate away from information stored in visual working memory (Theeuwes and colleagues, 2005, 2006). Saccadic deviation away from presented visual stimuli has been associated with top-down suppression (McSorley, Haggard, & Walker, 2006). This thesis examines the extent to which saccade trajectories are influenced by information held in visual working memory. Through a series of experiments behavioural memory data and saccade trajectory data were explored and evidence for visual working memory-oculomotor interaction was found. Other findings included specific interactions with the oculomotor system for the dorsal and ventral pathways as well as evidence for both bottom-up and top-down processing. Evidence of further oculomotor interaction with manual cognitive mechanisms was also illustrated, suggesting that visual working memory does not uniquely interact with the oculomotor system to preserve saccade programmes. The clinical and theoretical implications of this thesis are explored. It is proposed that the oculomotor system may interact with a variety of sensory systems to inform accurate and efficient visual processing.

The focus of this thesis is the improvement of acquisition and processing of Mueller matrix polarimetry using a confocal scanning laser ophthalmoscope (CSLO) and the application of Mueller matrix polarimetry to image the retina. Stepper motors were incorporated into a CSLO to semi-automate Mueller matrix polarimetry and were used in retinal image acquisition. Success rates of Fourier transform based edge detection filters, designed to improve the registration of retinal images, were compared. The acquired polarimetry images were used to reassess 2 image quality enhancement techniques, Mueller matrix reconstruction (MMR) and Stokes vector reconstruction (SVR), focusing on the role of auto-contrasting or normalization within the techniques and the degree to which auto-contrasting or normalization is responsible for image quality improvement of the resulting images. Mueller matrix polarimetry was also applied to find the retardance image of a malaria infected retinal blood vessel imaged in a confocal scanning laser microscope (CSLM) to visualize hemozoin within the vessel. Image quality enhancement techniques were also applied and image quality improvement was quantified for this blood vessel. The semi-automation of Mueller matrix polarimetry yielded a significant reduction in experimental acquisition time (80%) and a non-significant reduction in registration time (44%). A larger sample size would give higher power and this result might become significant. The reduction in registration time was most likely due to less movement of the eye, particularly in terms of decreased rotation seen between registered images. Fourier transform edge detection methods increased the success rate of registration from 73.9% to 92.3%. Assessment of the 2 MMR images (max entropy and max signal-to-noise ratio (SNR)) showed that comparison to the best CSLO images (not auto-contrasted) yielded significant average image quality improvements of 158% and 4% when quantified with entropy and SNR, respectively. When compared to best auto-contrasted CSLO images, significant image quality improvements were 11% and 5% for entropy and SNR, respectively. Images constructed from auto-contrasted input images were of significantly higher quality than images reconstructed from original images. Of the 2 other images assessed (modified degree of polarization (DOPM) and the first element of the Stokes vector (S0)), DOPM and S0 yielded significant average image quality improvements quantified by entropy except for the DOPM image of the RNFL. SNR was not improved significantly when either SVR image was compared to the best CSLO images. Compared to the best auto-contrasted CSLO images, neither DOPM nor S0 improved average image quality significantly. This result might change with a larger number of participants. When MMR were applied to images of malaria infected retinal slides, image quality was improved by 19.7% and 15.3% in terms of entropy and SNR, respectively, when compared to the best CSLO image. The DOPM image yielded image quality improvements of 8.6% and -24.3% and the S0 image gave improvements of 9.5% and 9.4% in entropy and SNR, respectively. Although percent increase in image quality was reduced when images were compared to initial auto-contrasted CSLO images, the final image quality was improved when auto-contrasting occurred prior to polarimetry calculations for max SNR and max entropy images. Quantitative values of retardance could not be found due to physical constraints in the CSLM that did not allow for characterization of its polarization properties and vibrational noise. Mueller matrix polarimetry used to find the retardance image of a malaria infected retina sample did yield visualization of hemozoin within the vessel but only qualitatively. In conclusion, improvements in the acquisition and registration of CSLO images were successful in leading to considerably shorter experimentation and processing times. In terms of polarimetric image quality improvement techniques, when compared to the best CSLO image. A large proportion of the improvement was in fact due to partially or completely stretching the pixel values across the dynamic range of the images within the algorithm of each technique. However, in general the image quality was still improved by the Mueller matrix reconstruction techniques using both entropy and SNR to generate the CSLO retinal images and the CSLM imaged malaria infected sample. In the malaria sample, retinal blood vessel visualization was also qualitatively improved. The images yielded from Mueller matrix polarimetry applied to a malaria infected retinal sample localized hemozoin within the blood vessel, but a quantitative image of the phase retardance could not be achieved.

Inherited retinal degenerative diseases can affect various components of the retina leading to blindness. Five different mutant strains of chicken have been studied extensively as potential models for inherited retinal degeneration. The Smoky Joe (SJ) chicken is a sixth genetically blind strain of White Leghorns that shows various degrees of blindness at hatch and by 8 weeks post-hatch, have complete blindness for those that are homozygous. The objective of this study was to characterize the retinal degeneration in these birds by histology, both during embryonic and post-hatch development, and to the retinal function using electroretinograms (ERG). For both embryonic and post-hatch development, a significantly lower number of cells were found in the retina of blind birds compared to sighted (both p<0.0001). The significant contributor to cell number decrease was the loss of amacrine cells located in the inner nuclear layer. Photoreceptors were also found to potentially decrease in number, but at a later stage. ERG recordings revealed decreases in amplitudes of b-waves and oscillatory potentials in blind birds, but not in sighted. Both histology and ERG findings support the idea that the inner retinal cells are affected. The results indicate that degeneration in the Smoky Joe retina occurs mostly within the inner nuclear layer affecting amacrine cells. This hampers the functional capacity of the retina, causing blindness.

Introduction: Oxygen delivery, or utilization, is a function of retinal blood flow and blood oxygen saturation. The retinal pigment epithelium (RPE), in particular, has been shown to have the highest levels of metabolic activity within the human body. Oxygen delivery is therefore of extreme importance to the maintenance of the health and integrity of the retina. Animal models presuppose that the oxygen tension in the retina is highest in the innermost layers at the level of the choriocapillaris, less in the photoreceptors and further decreases throughout the outer retinal structures. The choroid provides by far the largest component of the oxygen for consumption by the photoreceptors. A lack of oxygen stores in the inner retina therefore makes a constant supply crucial for its normal functioning. Blood flow dysfunction and subsequent hypoxia are both a feature in the pathogenesis of several major ocular diseases such as retinopathy of prematurity (ROP), age-related macular degeneration (ARMD), diabetic retinopathy (DR) and glaucoma. The development of methods to measure retinal blood flow and blood oxygen saturation is crucial to improve understanding of the patho-physiology of major ocular diseases. Purpose: The aims of this work were, firstly, to determine the least variable (range ± standard deviation) wavelength combination (610/548, 600/569 and 605/586) and subsequent ODR with the prototype HRC device. Secondly, using the ODR with the lowest measurement variability, we sought to quantify retinal blood SO2 in arterioles and venules and investigate the relationship between retinal blood SO2 and total retinal blood flow (TRBF) in response to stepwise changes in PETO2 in healthy participants. Retinal blood SO2 and TRBF were assessed using the IRIS HRC (Photon etc. Inc. Montreal, Canada) and the RTvue Doppler Fourier Domain OCT (Optovue Inc, Freemont, CA) instruments, respectively. Methods: Ten healthy participants between the ages of 23 and 37, with an average age of 28.3 years were evaluated in two descriptive cross-sectional studies. Two gas provocation protocols; hyperoxia (end-tidal oxygen; PETO2 of 100, 200, 300, 400mmHg) and hypoxia (PETO2 of 100, 80, 60, 50mmHg) were administered in a fixed sequential order. In each phase of gas provocation (via modulation of PETO2), retinal blood SO2 and TRBF measurements were acquired with the HRC and Doppler FD-OCT. The precise and repeated control of the partial end tidal pressures of oxygen (PETO2) and carbon dioxide (PETCO2) over the pre-determined phase duration, irrespective of the individuals’ respiratory rate, was made possible with the RespirAct (Thornhill Research Inc., Toronto, Canada); a sequential re-breathing gas delivery Results: In arterioles, the group range (±SD) of ODR values for baseline measurements (PETO2 of 100mmHg) was 0.169±0.061 for the 605/586 wavelength combination, 0.371±0.099 for the 600/569 wavelength combination and 0.340±0.104 for the 610/548 wavelength combination. In venules, the group range (±SD) of ODR values was 0.600±0.198 for the 605/586 wavelength combination, 0.569±0.169 for the 600/569 wavelength combination and 0.819±0.274 for the 610/548 wavelength combination. With the 605/586 combination at baseline 1 and 2 in arterioles, the group range (±SD) of ODR values was 0.607 ± 0.224 and 0.619 ± 0.158, respectively (p = 0.370), while in venules the group range (±SD) of ODR at baseline 1 and 2 was 0.289±0.750 and 0.284 ± 0.729, respectively (p = 0.714). For the 600/569 combination at baseline 1 and 2 in arterioles, the group range (±SD) of ODR values was 0.747±0.350 and 0.761±0.391, respectively (p = 0.424) while in venules the group range (±SD) of ODR at baseline 1 and 2 was 0.329±0.675 and 0.366±0.659, respectively (p = 0.372). For the 610/548 combination at baseline 1 and 2 in arterioles, the group range (±SD) of ODR values was 0.604±0.263 and 0.685±0.450, respectively (p = 0.056) while in venules, the group range (±SD) of ODR at baseline 1 and 2 was 0.292±0.746 and 0.285±1.009, respectively (p = 0.131). There was no statistical difference found between baseline ODR values (baseline 1 and 2) across all three wavelength combinations in both arterioles and venules. The mean retinal blood SO2 value at baseline in arterioles for 4 participants was 95.19% ± 31.04% and venules was 53.89% ± 17.24% (p = 0.115). There was a negative linear relationship between group retinal blood SO2 and TRBF values in the 10 participants studied, although the results of any of the 10 individuals did not show evidence of such a relationship using the described methodology. The Pearson’s correlation coefficient (r) between TRBF and SaO2 was r = -0.354 and p = 0.001 and between TRBF and SvO2 was r = - 0.295, p = 0.008 Conclusion: Of the three wavelength combinations investigated (605/586, 600/569 and 610/548), the 605/586 combination was shown to have the overall least variability. It would be unwise at this stage to adopt this wavelength combination for clinical usage, however, since it is presupposed that the 605/586 combination is also the most reliable combination to detect change in retinal blood SO2 i.e. lower variability of the 605/586 combination may be irrelevant if this combination proves to be insensitive to change in retinal blood SO2. The absolute mean ± SD retinal blood SO2 in the arterioles (SaO2) was 95.19% ± 31.04% and in the venules (SvO2) was 53.89% ± 17.24%. These values fell within the range expected and described in the literature. The magnitude of the difference between the SaO2 and SvO2 was also consistent with the literature. These findings were all appropriate for a low flow, high oxygen exchange vascular network typical of the inner retinal vascular system. Using group rather than individual data, TRBF was found in this study to relate inversely with SaO2 (r = -0.354 and p = 0.001) and SvO2 (r = – 0.295 and p=0.008), respectively. This relationship between TRBF and SaO2 and SvO2, was as expected based upon data derived primarily from animal models. This study is ground-breaking and unique, in that, it is the first study to concomitantly measure both retinal blood SO2 and TRBF in human participants. Individual data showed extensive variability and noise, thus limiting the strength of the association between TRBF and SaO2 and SvO2..

A previous study in chickens revealed that myosin light chain kinase (MLCK), f actin, and myosin are found on the crystalline lens. Their polygonal arrangement at the posterior surface resembles a muscle tissue, which suggests that these proteins may have a contractile role in accommodation. The ciliary muscle in chickens is skeletal in nature and, therefore, chickens were used to test the hypothesis that contractile microfilaments play a role in accommodation. Ciliary nerve-induced accommodation was measured in the presence of an MLCK inhibitor 1-(5-Iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-7). Eyes of 6-day old white Leghorn chickens (gallus gallus domesticus) were enucleated in Tyrode’s saline solution while keeping the ciliary nerve intact. One eye was treated with ML-7 and the other eye was treated with vehicle only. Three concentrations of ML-7 were used: 1 µM, 10 µM, and 100 µM. Two experiments were carried out, one including a (3×10 min) wash and one without. Focal lengths of the vehicle- and ML-7-treated eyes were measured before, during and after accommodation. Immunoblots were used to detect the amount of phosphorylated myosin with and without the inhibitor. Focal lengths for accommodation were shorter than those at rest (p<0.001). In the wash experiment, vehicle-treated eyes had higher accommodative amplitudes compared to ML-7-treated eyes for all three dosage groups. In the no-wash experiment, only the 1 µM group demonstrated the same trend as the wash experiment. For the 10 µM and 100 µM groups, ML-7-treated eyes had higher accommodative amplitudes compared to vehicle-treated eyes. Immunoblots revealed varying amounts of inhibition within pairs of eyes as well as between birds for both experiments. Results from this experiment indicate that ML-7 was not effective at determining whether contractile microfilaments found on the lens contribute to accommodation.