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Retinal vascular resistance is of interest in glaucoma research, as a potential link between retinal ganglion cell loss and observed phenomena including disrupted vascular autoregulation, altered biomechanical stiffness, and impaired neurovascular coupling. It can now be assessed in vivo, using laser speckle flowgraphy. However, continued progress in the field requires better understanding of its physiology. In this study, we test the hypothesis of homogeneity of vascular resistance indices between regions of the retina: specifically, between superior and inferior hemifields. The resistivity index (maximum flow minus minimum flow, as a proportion of the maximum) and pulsatility index (maximum minus minimum, as a proportion of the mean) were measured in major vessels within the optic nerve head, in the remaining tissue within the optic nerve head, and in peripapillary branch retinal arteries, separated in each case into superior and inferior quadrants. This was performed in 378 eyes of 189 participants with suspected, early or moderate glaucoma; and in 99 eyes of 50 participants without any ocular pathology. In the glaucoma cohort, the resistivity index was on average 9% higher superiorly than inferiorly in vessels within the optic nerve head; 8% higher superiorly in remaining tissue; and 8% higher superiorly in peripapillary vessels (all p < 0.001). The pulsatility index was on average 11% higher superiorly in all three locations (all p < 0.001). Average flow was slightly higher superiorly in major vessels in the nerve head, but higher inferiorly elsewhere. In the healthy control cohort, resistivity index was higher superiorly by 10% in vessels and 8% in tissue within the optic nerve head; pulsatility index was 12% and 10% higher superiorly respectively (all p < 0.001). The fact that these differences were similar between the two cohorts suggests that they are not caused by the disease process. However, it is notable that glaucomatous loss most frequently occurs first in the superior visual field, corresponding with the inferior retina. The finding that vascular resistance indices are consistently higher in the superior retina warrants further investigation, both for its causes and consequences.
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Purpose: It has been hypothesized that compromised ocular circulation in glaucoma may be concomitant of systemic changes. The purpose of this study is to test whether systemic blood flow pulse waveform patterns differ between individuals with glaucoma (GL), glaucoma suspects (GLS), and normal healthy controls (HC). Methods: The study included 35 bilateral GL, 67 bilateral GLS, 29 individuals with unilateral GL who were considered GLS in the other eye, and 44 healthy controls. Systemic pulsatile blood pressure waveforms were recorded using a finger cuff. A continuous 200 Hz plethysmography recording is made to obtain a pulse waveform. Waveform parameters were extracted using custom software from an average of eight pulse cycles. These were compared between GL, GLS, and HC groups on a per-eye basis, using generalized estimating equation models to account for intereye correlations; and plotted against disease severity by visual field linearized mean deviation (MDlin) and retinal nerve fiber layer thickness (RNFLT). Results: Averaged blood pressure was significantly lower in the HC group (mean ± standard deviation 91.7 ±11.7 mm Hg) than the GLS (102.4 ± 13.9) or GL (102.8 ± 13.7) groups, with P < 0.0001 (generalized estimating equation regression). Waveform parameters representing vascular resistance were higher in both GLS and GL groups than the HC group; and were correlated with RNFLT and MDlin (P ≤ 0.05). Conclusions: The shape of the systemic pulsatile waveform differs in individuals with GL/GLS suspects, compared to HC eyes. Blood pressure changes more rapidly in individuals with GL, which suggests higher arterial stiffness.
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Presión Sanguínea , Presión Intraocular , Hipertensión Ocular , Humanos , Masculino , Femenino , Persona de Mediana Edad , Presión Sanguínea/fisiología , Presión Intraocular/fisiología , Anciano , Hipertensión Ocular/fisiopatología , Campos Visuales/fisiología , Glaucoma/fisiopatología , Glaucoma/diagnóstico , Análisis de la Onda del Pulso , Frecuencia Cardíaca/fisiología , Adulto , Pletismografía , Fibras Nerviosas/patología , Células Ganglionares de la Retina/patologíaRESUMEN
Purpose: This study investigates the temporal relationship between blood flow changes and alterations in retinal nerve fiber layer thickness (RNFLT) and mean deviation (MD) in individuals with glaucoma. Methods: Blood flow, measured by mean blur rate in optic nerve head vessels (MBRv) and tissues (MBRt) using laser speckle flowgraphy (LSFG)-NAVI, was analyzed using structural equation models (SEMs). SEMs assessed whether the previous rate of one parameter predicted the current rate of the other parameter, adjusted for its own rate in the previous time interval. Data from 345 eyes of 174 participants were gathered from visits every six months. Results: Rates of change of both MBRv and MBRt were significantly predicted by their own rate in the previous time interval and by the rate of change of MD in the previous time interval (P < 0.001 and P = 0.043, respectively), but not by the rate of MD in the concurrent interval (P = 0.947 and P = 0.549), implying that changes in MD precede changes in blood flow. Rates of change of RNFLT were predicted by their own previous rate and the rate of change of MBRv and MBRt in either the previous interval (P = 0.002 and P = 0.008) or the concurrent interval (P = 0.001 and P = 0.018), suggesting that MBR may change before RNFLT. Conclusions: The evidence supports a temporal sequence where MD changes precede blood flow changes, which, in turn, may precede alterations in RNFLT.
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Glaucoma , Disco Óptico , Humanos , Campos Visuales , Retina , Fibras NerviosasRESUMEN
Purpose: To compare parameters of electroretinogram (ERG) responses for their ability to detect functional loss in early stages of nonhuman primate (NHP) experimental glaucoma (EG), including photopic negative responses (PhNR) to a standard brief red flash on a blue background (R/B) and 200-ms-long R/B and white-on-white (W/W) flashes, to W/W flicker stimuli (5-50 Hz), and to a dark-adapted intensity series. Methods: Light-adapted ERGs were recorded in 12 anesthetized monkeys with unilateral EG. Amplitudes and implicit times of the a-wave, b-wave, and d-wave were measured, as well as amplitudes of PhNRs and oscillatory potentials for flash onset and offset. Flicker ERGs were measured using peak-trough and fundamental frequency analyses. Dark-adapted ERG parameters were modeled by Naka-Rushton relationships. Results: Only PhNR amplitudes were significantly reduced in EG eyes compared to fellow control (FC) eyes. The d-wave implicit time was delayed in EG versus FC eyes only for the W/W long flash, but in all eyes it was 10 to 20 ms slower for R/B versus the W/W condition. Flicker ERGs were <0.5 ms delayed in EG versus FC overall, but amplitudes were affected only at 5 Hz. The brief R/B PhNR amplitude had the highest sensitivity to detect EG and strongest correlation to parameters of structural damage. Conclusions: The PhNR to the standard brief R/B stimulus was best for detecting and following early-stage functional loss in NHP EG. Translational Relevance: These results suggest that there would be no benefit in using longer duration flashes to separate onset and offset responses for clinical management of glaucoma.
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Glaucoma , Animales , Glaucoma/diagnóstico , OjoRESUMEN
Purpose: Tissue stiffening and alterations in retinal blood flow have both been suggested as causative mechanisms of glaucomatous damage. We tested the hypothesis that retinal blood vessels also stiffen, using laser speckle flowgraphy (LSFG) to characterize vascular resistance. Methods: In the longitudinal Portland Progression Project, 231 eyes of 124 subjects received LSFG scans of the optic nerve head (ONH) and automated perimetry every 6 months for six visits. Eyes were classified as either "glaucoma suspect" or "glaucoma" eyes based on the presence of functional loss on the first visit. Vascular resistance was quantified using the mean values of several instrument-defined parameterizations of the pulsatile waveform measured by LSFG, either in major vessels within the ONH (serving the retina) or in capillaries within ONH tissue, and age-adjusted using a separate group of 127 healthy eyes of 63 individuals. Parameters were compared against the severity and rate of change of functional loss using mean deviation (MD) over the six visits, within the two groups. Results: Among 118 "glaucoma suspect" eyes (average MD, -0.4 dB; rate, -0.45 dB/y), higher vascular resistance was related to faster functional loss, but not current severity of loss. Parameters measured in major vessels were stronger predictors of rate than parameters measured in tissue. Among 113 "glaucoma" eyes (average MD, -4.3 dB; rate, -0.53 dB/y), higher vascular resistance was related to more severe current loss but not rate of loss. Conclusions: Higher retinal vascular resistance and, by likely implication, stiffer retinal vessels were associated with more rapid functional loss in eyes without significant existing loss at baseline.
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Glaucoma , Hipertensión Ocular , Disco Óptico , Rigidez Vascular , Humanos , Presión Intraocular , Tomografía de Coherencia Óptica , Disco Óptico/irrigación sanguínea , Glaucoma/diagnóstico , Vasos RetinianosRESUMEN
Purpose: Assessment of glaucomatous damage in animal models is facilitated by rapid and accurate quantification of retinal ganglion cell (RGC) axonal loss and morphologic change. However, manual assessment is extremely time- and labor-intensive. Here, we developed AxoNet 2.0, an automated deep learning (DL) tool that (i) counts normal-appearing RGC axons and (ii) quantifies their morphometry from light micrographs. Methods: A DL algorithm was trained to segment the axoplasm and myelin sheath of normal-appearing axons using manually-annotated rat optic nerve (ON) cross-sectional micrographs. Performance was quantified by various metrics (e.g., soft-Dice coefficient between predicted and ground-truth segmentations). We also quantified axon counts, axon density, and axon size distributions between hypertensive and control eyes and compared to literature reports. Results: AxoNet 2.0 performed very well when compared to manual annotations of rat ON (R2 = 0.92 for automated vs. manual counts, soft-Dice coefficient = 0.81 ± 0.02, mean absolute percentage error in axonal morphometric outcomes < 15%). AxoNet 2.0 also showed promise for generalization, performing well on other animal models (R2 = 0.97 between automated versus manual counts for mice and 0.98 for non-human primates). As expected, the algorithm detected decreased in axon density in hypertensive rat eyes (P ⪠0.001) with preferential loss of large axons (P < 0.001). Conclusions: AxoNet 2.0 provides a fast and nonsubjective tool to quantify both RGC axon counts and morphological features, thus assisting with assessing axonal damage in animal models of glaucomatous optic neuropathy. Translational Relevance: This deep learning approach will increase rigor of basic science studies designed to investigate RGC axon protection and regeneration.
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Aprendizaje Profundo , Glaucoma , Ratas , Ratones , Animales , Células Ganglionares de la Retina/fisiología , Estudios Transversales , Modelos Animales de Enfermedad , Axones/fisiología , Glaucoma/diagnósticoRESUMEN
Purpose: Glia and their communication via connexin 43 (Cx43) gap junctions are known to mediate neurovascular coupling, a process driven by metabolic demand. However, it is unclear whether Cx43 mediated glial communication intermediates classical autoregulation. Here we used viral transfection and a glial fibrillary acidic protein (GFAP) promoter to downregulate glial Cx43 to evaluate its role in retinal vascular autoregulation to ocular perfusion pressure (OPP) reduction. Methods: Adult rats were intravitreally injected with the viral active construct or a control. Three weeks after the injection, eyes were imaged using confocal scanning laser ophthalmoscopy before and during a period of OPP decrease induced by blood draw to lower blood pressure or by manometric IOP elevation. Vessel diameter responses to the OPP decrease were compared between Cx43-downregulated and control-injected eyes. The extent of Cx43 downregulation was evaluated by Western blot and immunohistochemistry. Results: In control eyes, the OPP decrease induced dilatation of arterioles, but not venules. In Cx43-downregulated eyes, Cx43 expression in whole retina was decreased by approximately 40%. In these eyes, the resting diameter of the venules increased significantly, but there was no effect on arterioles. In Cx43-downregulated eyes, vasoreactivity evoked by blood pressure lowering was significantly compromised in both arterioles (P = 0.005) and venules (P = 0.001). Cx43 downregulation did not affect the arteriole responses to IOP elevation, whereas the responses of the venules showed a significantly greater decrease in diameter (P < 0.001). Conclusions: The downregulation of retinal Cx43 in GFAP-expressing cells compromises vasoreactivity of both arterioles and venules in response to an OPP decrease achieved via blood pressure lowering or IOP elevation. The results also suggest that Cx43-mediated glial communication actively regulates resting venular diameter.
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Presión Sanguínea/fisiología , Conexina 43/genética , Regulación de la Expresión Génica/fisiología , Proteína Ácida Fibrilar de la Glía/genética , Presión Intraocular/fisiología , Arteria Retiniana/fisiología , Vena Retiniana/fisiología , Animales , Western Blotting , Dependovirus/genética , Regulación hacia Abajo , Inmunohistoquímica , Masculino , Microscopía Confocal , Oftalmoscopía , Ratas , Ratas Endogámicas BN , Flujo Sanguíneo Regional , Retina/metabolismo , TransfecciónRESUMEN
In this work, we develop a robust, extensible tool to automatically and accurately count retinal ganglion cell axons in optic nerve (ON) tissue images from various animal models of glaucoma. We adapted deep learning to regress pixelwise axon count density estimates, which were then integrated over the image area to determine axon counts. The tool, termed AxoNet, was trained and evaluated using a dataset containing images of ON regions randomly selected from whole cross sections of both control and damaged rat ONs and manually annotated for axon count and location. This rat-trained network was then applied to a separate dataset of non-human primate (NHP) ON images. AxoNet was compared to two existing automated axon counting tools, AxonMaster and AxonJ, using both datasets. AxoNet outperformed the existing tools on both the rat and NHP ON datasets as judged by mean absolute error, R2 values when regressing automated vs. manual counts, and Bland-Altman analysis. AxoNet does not rely on hand-crafted image features for axon recognition and is robust to variations in the extent of ON tissue damage, image quality, and species of mammal. Therefore, AxoNet is not species-specific and can be extended to quantify additional ON characteristics in glaucoma and potentially other neurodegenerative diseases.
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Axones/fisiología , Biología Computacional/métodos , Aprendizaje Profundo , Modelos Biológicos , Nervio Óptico/fisiología , Células Ganglionares de la Retina/fisiología , Programas Informáticos , Algoritmos , Animales , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Femenino , Glaucoma/etiología , Glaucoma/metabolismo , Glaucoma/patología , Masculino , Nervio Óptico/patología , Enfermedades del Nervio Óptico/etiología , Enfermedades del Nervio Óptico/metabolismo , Enfermedades del Nervio Óptico/patología , Ratas , Reproducibilidad de los ResultadosRESUMEN
SIGNIFICANCE: The balance between oxygen and carbon dioxide sets the resting tone (or diameter) of retinal blood vessels. Eyes that are hypercapnic use up their "vasodilatory reserve" and therefore fail to respond adequately to changes in intraocular or blood pressure. PURPOSE: Retinal vessels are regulated by both myogenic and metabolic mechanisms. We considered whether alteration of metabolic status would modify the vascular response to ocular perfusion pressure (OPP) lowering in rat retina. METHODS: In pentobarbital anesthetized adult Brown-Norway rats, normocapnia or hypercapnia was achieved by artificially ventilating animals with air or 5% carbon dioxide in ~30% oxygen, respectively. Ocular perfusion pressure was gradually reduced to ~20 mmHg by either lowering blood pressure (slowly drawing blood from a femoral artery/vein) or manometrically increasing intraocular pressure under normocapnic or hypercapnic conditions. In all four groups (n = 7 eyes for each), a confocal scanning laser ophthalmoscope was used to acquire image sequences centered on the optic nerve throughout pressure modification. The diameter of arterioles and venules at various OPP levels was measured and expressed as percentage relative to their own baseline. The response of arterioles and venules to OPP lowering was compared between normocapnic and hypercapnic groups. RESULTS: Average arterial carbon dioxide partial pressures were 36.9 ± 2.6 mmHg in normocapnic and 64.1 ± 5.9 mmHg in hypercapnic (P < .001) animals. In the normocapnic groups, blood pressure lowering and intraocular pressure elevation resulted in significant vasodilation of both arterioles and venules (P < .0001). In the hypercapnic groups, OPP lowering-induced vasodilation was significantly attenuated compared with the corresponding normocapnic groups (P < .0001 for both, two-way analysis of variance). CONCLUSION: Hypercapnia significantly modified myogenic vascular autoregulation in response to OPP reduction.
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Presión Sanguínea/fisiología , Hipercapnia/fisiopatología , Presión Intraocular/fisiología , Músculo Liso Vascular/fisiopatología , Retina/fisiopatología , Vasos Retinianos/fisiopatología , Animales , Homeostasis , Humanos , Masculino , Oxígeno/sangre , Ratas , Ratas Endogámicas BN , Tonometría OcularRESUMEN
Purpose: Blood flow in the optic nerve head (ONH) is known to be reduced in eyes with advanced glaucoma. However, experimental results from non-human primates suggest an initial increase in ONH blood flow at the earliest stages of damage. This study assesses flow and pulsatile hemodynamics across a range of severities to test the hypothesis that this also occurs in human glaucoma. Methods: Laser speckle flowgraphy was used to measure average mean blur rate (MBRave) within ONH tissue (a correlate of capillary blood flow) and the pulsatile waveform in 93 eyes with functional loss and 74 glaucoma suspect/fellow eyes without functional loss. These were compared against results from 92 healthy control eyes. Parameters produced by the instrument's software were age-corrected, then compared between groups using generalized estimating equation models. Results: The mean MBRave in the control eyes was 12.5 units. In glaucoma suspect/fellow eyes, the mean was 16.4 units, higher with P < 0.0001. In eyes with functional loss, the mean was 13.8 units, lower than eyes without functional loss with P < 0.0001, although still higher than control eyes with P = 0.0096. Analysis of the pulsatile waveform suggested that the deceleration in flow as it approaches its maximum across the cardiac cycle was delayed in glaucoma. Conclusions: Blood flow within ONH capillaries was higher in glaucoma suspect eyes than in healthy controls. It was less elevated in eyes that had developed functional loss. The mechanisms causing these changes and their relation to concurrent changes in pulsatile hemodynamics remain under investigation.
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Velocidad del Flujo Sanguíneo/fisiología , Capilares/fisiopatología , Glaucoma de Ángulo Abierto/fisiopatología , Microcirculación/fisiología , Disco Óptico/irrigación sanguínea , Flujo Sanguíneo Regional/fisiología , Adulto , Anciano , Capilares/diagnóstico por imagen , Femenino , Angiografía con Fluoresceína , Fondo de Ojo , Glaucoma de Ángulo Abierto/diagnóstico , Humanos , Presión Intraocular , Flujometría por Láser-Doppler/métodos , Masculino , Persona de Mediana Edad , Pronóstico , Tomografía de Coherencia Óptica/métodosRESUMEN
PURPOSE: To compare diagnostic performance and structure-function correlations of multifocal electroretinogram (mfERG), full-field flash ERG (ff-ERG) photopic negative response (PhNR) and transient pattern-reversal ERG (PERG) in a non-human primate (NHP) model of experimental glaucoma (EG). METHODS: At baseline and after induction of chronic unilateral IOP elevation, 43 NHP had alternating weekly recordings of retinal nerve fiber layer thickness (RNFLT) by spectral domain OCT (Spectralis) and retinal function by mfERG (7F slow-sequence stimulus, VERIS), ff-ERG (red 0.42 log cd-s/m2 flashes on blue 30 scotopic cd/m2 background, LKC UTAS-E3000), and PERG (0.8° checks, 99% contrast, 100 cd/m2 mean, 5 reversals/s, VERIS). All NHP were followed at least until HRT-confirmed optic nerve head posterior deformation, most to later stages. mfERG responses were filtered into low- and high-frequency components (LFC, HFC, >75 Hz). Peak-to-trough amplitudes of LFC features (N1, P1, N2) and HFC RMS amplitudes were measured and ratios calculated for HFC:P1 and N2:P1. ff-ERG parameters included A-wave (at 10 ms), B-wave (trough-to-peak) and PhNR (baseline-to-trough) amplitudes as well as PhNR:B-wave ratio. PERG parameters included P50 and N95 amplitudes as well as N95:P50 ratio and N95 slope. Diagnostic performance of retinal function parameters was compared using the area under the receiver operating characteristic curve (A-ROC) to discriminate between EG and control eyes. Correlations to RNFLT were compared using Steiger's test. RESULTS: Study duration was 15 ± 8 months. At final follow-up, structural damage in EG eyes measured by RNFLT ranged from 9% above baseline (BL) to 58% below BL; 29/43 EG eyes (67%) and 0/43 of the fellow control eyes exhibited significant (>7%) loss of RNFLT from BL. Using raw parameter values, the largest A-ROC findings for mfERG were: HFC (0.82) and HFC:P1 (0.90); for ff-ERG: PhNR (0.90) and PhNR:B-wave (0.88) and for PERG: P50 (0.64) and N95 (0.61). A-ROC increased when data were expressed as % change from BL, but the pattern of results persisted. At 95% specificity, the diagnostic sensitivity of mfERG HFC:P1 ratio was best, followed by PhNR and PERG. The correlation to RNFLT was stronger for mfERG HFC (R = 0.65) than for PhNR (R = 0.59) or PERG N95 (R = 0.36), (p = 0.20, p = 0.0006, respectively). The PhNR flagged a few EG eyes at the final time point that had not been flagged by mfERG HFC or PERG. CONCLUSIONS: Diagnostic performance and structure-function correlation were strongest for mfERG HFC as compared with ff-ERG PhNR or PERG in NHP EG.
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Electrorretinografía/métodos , Glaucoma/diagnóstico , Glaucoma/fisiopatología , Animales , Modelos Animales de Enfermedad , Electrorretinografía/normas , Femenino , Glaucoma/patología , Macaca mulatta , Masculino , Fibras Nerviosas/patología , Fibras Nerviosas/fisiología , Disco Óptico/fisiopatología , Células Ganglionares de la Retina/patología , Células Ganglionares de la Retina/fisiología , Tomografía de Coherencia ÓpticaRESUMEN
Purpose: The purpose of this study was to test the hypothesis that retinal glial cells modify basal vessel diameter and pressure-initiated vascular regulation in rat retina. Methods: In rats, L-2-aminoadipic acid (LAA, 10 nM) was intravitreally injected to inhibit glial cell activity. Twenty-four hours following injection, retinal glial intracellular calcium (Ca2+) was labeled with the fluorescent calcium indicator Fluo-4/AM (F4, 1 mM). At 110 minutes after injection, intraocular pressure (IOP) was elevated from 20 to 50 mm Hg. Prior to and during IOP elevation, Ca2+ and retinal vessel diameter were assessed using a spectral-domain optical coherence tomography/confocal scanning laser ophthalmoscope. Dynamic changes in Ca2+ and diameter from IOP elevation were quantified. The response in LAA-treated eyes was compared with vehicle treated control eyes. Results: L-2-Aminoadipic acid treatment significantly reduced F4-positive cells in the retina (LAA, 16 ± 20 vs. control, 55 ± 37 cells/mm2; P = 0.02). Twenty-four hours following LAA treatment, basal venous diameter was increased from 38.9 ± 3.9 to 51.8 ± 6.4 µm (P < 0.0001, n = 20), whereas arterial diameter was unchanged (from 30.3 ± 3.5 to 30.7 ± 2.8 µm; P = 0.64). In response to IOP elevation, LAA-treated eyes showed a smaller increase in glial cell Ca2+ around both arteries and veins in comparison with control (P < 0.001 for both). There was also significantly greater IOP-induced vasoconstriction in both vessel types (P = 0.05 and P = 0.02, respectively; n = 6 each). Conclusions: The results suggest that glial cells can modulate basal retinal venous diameter and contribute to pressure-initiated vascular responses.
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Presión Intraocular/fisiología , Neuroglía/patología , Hipertensión Ocular/fisiopatología , Disco Óptico/irrigación sanguínea , Flujo Sanguíneo Regional/fisiología , Vasos Retinianos/fisiopatología , Tomografía de Coherencia Óptica/métodos , Vasoconstricción/fisiología , Animales , Modelos Animales de Enfermedad , Masculino , Hipertensión Ocular/patología , Oftalmoscopía , Ratas , Ratas Endogámicas BN , Células Ganglionares de la Retina/patología , Vasos Retinianos/patologíaRESUMEN
PURPOSE: We compare spectral-domain optical coherence tomography (SDOCT) measurements of minimum rim width (MRW), minimum rim area (MRA), and peripapillary retinal nerve fiber layer thickness (RNFLT) to complete orbital optic nerve axon counts in nonhuman primates (NHP) with unilateral experimental glaucoma (EG). METHODS: Biweekly SDOCT measurements of MRW, MRA, and RNFLT were acquired under manometric IOP control (10 mm Hg) in 51 NHP during baseline (mean ± SD, 5.0 ± 1.6 sessions) and after laser photocoagulation was applied to the trabecular meshwork of one eye to induce chronic IOP elevation. At the study endpoint (predefined for each NHP), 100% axon counts were obtained from each optic nerve. RESULTS: For SDOCT parameters at baseline, the correlation between the two eyes of each animal was strongest for RNFLT (R = 0.97) and MRW (R = 0.97), but lower for MRA (R = 0.85). At the final time point, average values in EG eyes relative to control eyes were: -22% for RNFLT, -38% for MRW, -36% for MRA, and -36% for optic nerve axons. The correlation with axon counts was strongest for RNFLT (R = 0.81), compared to MRW (R = 0.72, P = 0.001) or MRA (R = 0.70, P = 0.001). Diagnostic sensitivity was 75% for RNFLT, 90% for MRW, and 88% for MRA; all had 100% specificity. CONCLUSIONS: Peripapillary RNFLT was correlated more closely with total orbital optic nerve axon count than were the ONH parameters MRW or MRA. This is likely because glaucomatous deformation (beyond axon loss alone) has a greater influence on the ONH parameters MRW and MRA than on RNFLT.
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Axones/patología , Glaucoma/patología , Presión Intraocular/fisiología , Disco Óptico/patología , Células Ganglionares de la Retina/patología , Tomografía de Coherencia Óptica/métodos , Animales , Modelos Animales de Enfermedad , Femenino , Glaucoma/fisiopatología , Macaca mulatta , MasculinoRESUMEN
PURPOSE: To use optical coherence tomography (OCT) to test the hypothesis that optic nerve head (ONH) "cupping" in the monkey optic nerve transection (ONT) model does not include posterior laminar deformation. METHODS: Five monkeys (aged 5.5-7.8 years) underwent ONH and retinal nerve fiber layer (RNFL) OCT imaging five times at baseline and biweekly following unilateral ONT until euthanization at â¼40% RNFL loss. Retinal nerve fiber layer thickness (RNFLT) and minimum rim width (MRW) were calculated from each pre- and post-ONT imaging session. The anterior lamina cribrosa surface (ALCS) was delineated within baseline and pre-euthanasia data sets. Significant ONT versus control eye pre-euthanasia change in prelaminar tissue thickness (PLTT), MRW, RNFLT, and ALCS depth (ALCSD) was determined using a linear mixed-effects model. Eye-specific change in each parameter exceeded the 95% confidence interval constructed from baseline measurements. RESULTS: Animals were euthanized 49 to 51 days post ONT. Overall ONT eye change from baseline was significant for MRW (-26.2%, P = 0.0011), RNFLT (-43.8%, P < 0.0001), PLTT (-23.8%, P = 0.0013), and ALCSD (-20.8%, P = 0.033). All five ONT eyes demonstrated significant eye-specific decreases in MRW (-23.7% to -31.8%) and RNFLT (-39.6% to -49.7%). Four ONT eyes showed significant PLTT thinning (-23.0% to -28.2%). The ALCS was anteriorly displaced in three of the ONT eyes (-25.7% to -39.2%). No ONT eye demonstrated posterior laminar displacement. CONCLUSIONS: Seven weeks following surgical ONT in the monkey eye, ONH cupping involves prelaminar and rim tissue thinning without posterior deformation of the lamina cribrosa.
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Fibras Nerviosas/patología , Disco Óptico/patología , Enfermedades del Nervio Óptico/diagnóstico , Traumatismos del Nervio Óptico/diagnóstico , Células Ganglionares de la Retina/patología , Tomografía de Coherencia Óptica/métodos , Animales , Modelos Animales de Enfermedad , Presión Intraocular , Macaca mulatta , Masculino , Enfermedades del Nervio Óptico/etiología , Traumatismos del Nervio Óptico/complicacionesRESUMEN
PURPOSE: To evaluate structure and function of macular retinal layers in nonhuman primate (NHP) experimental glaucoma (EG). METHODS: Twenty-one NHP had longitudinal imaging of macular structure by SDOCT, 16 also had recordings of function by multifocal ERG. The average thickness over 15° was derived for seven individual SDOCT layers: macular nerve fiber layer (m-NFL), retinal ganglion cell layer (RGCL), inner plexiform layer (IPL), inner nuclear layer (INL), outer plexiform layer (OPL), outer nuclear layer+inner segments combined (ONL+IS), and outer segments (OS). Peripapillary RNFL thickness (ppRNFLT) was measured from a single circular B-scan with 12° diameter. Responses to a slow-sequence multifocal ERG (mfERG) stimulus (7F) were filtered (at 75 Hz) into low- and high-frequency components (LFC, HFC). RESULTS: At final follow-up, significant structural loss occurred only in EG eyes and only for ppRNFLT (-29 ± 23%), m-NFL (-17 ± 16%), RGCL (-22 ± 15%), and IPL (-19 ± 14%); though there was also a small increase in OPL (+6 ± 7%) and ONL+IS (4 ± 4%) and a similar tendency for INL. Macular structural loss was correlated with ppRNFLT only for the NFL, RGCL and IPL (R = 0.95, 0.93 and 0.95, respectively, P < 0.0001). Significant functional loss occurred only for HFC and N2 in EG eyes. Significant longitudinal structure-function correlations (P < 0.01) were observed only in EG eyes and only for mfERG HFC and N2: HFC was correlated with ppRNFLT (R = 0.69), macular NFL (R = 0.67), RGCL (R = 0.74), and IPL (R = 0.72); N2 was correlated with RGCL (R = 0.54) and IPL (R = 0.48). High-frequency components amplitude change was inversely correlated with outer retinal thickness change (= -0.66). CONCLUSIONS: Macular structural and functional losses are correlated and specific to ganglion cells over a wide range of EG severity. Outer retinal changes are likely due to inner retinal loss.
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Glaucoma/patología , Mácula Lútea/patología , Mácula Lútea/fisiopatología , Células Ganglionares de la Retina/patología , Tomografía de Coherencia Óptica/métodos , Animales , Modelos Animales de Enfermedad , Electrorretinografía , Femenino , Estudios de Seguimiento , Glaucoma/fisiopatología , Macaca mulatta , MasculinoRESUMEN
PURPOSE: We validated noninvasive Doppler-optical coherence tomography (OCT) blood flow measurements against the terminal microsphere method in a surgical induced optic nerve transection nonhuman primate model. METHODS: In 6 nonhuman primates, total retinal blood flow (TRBF) was measured with a custom-built dual-beam bidirectional Doppler Fourier Domain (FD)-OCT. Peripapillary retinal nerve fiber layer thickness (RNFLT) was measured by Spectralis spectral-domain (SD)-OCT. Measurements were performed every 10 to 15 days before and after unilateral optic nerve transection (ONT) until RNFLT was reduced by more than 40% from baseline. Before the animals were killed, TRBF was measured using the microsphere technique. RESULTS: A significant correlation between all arterial and venous Doppler OCT TRBF measurements was found in ONT and contralateral control eyes (both P < 0.01, n = 6). The Bland-Altman analysis showed a bias of 0.57 in the ONT group and 0.02 in the contralateral control group. Also, excellent agreement was observed between Doppler OCT and microsphere measurements (P < 0.01, r = 0.976, bias = 0.54). After ONT, TRBF and RNFLT decreased by -51% ± 42% and -44% ± 2% (n = 5), respectively. In the contralateral control eyes, TRBF and RNFLT were unchanged. CONCLUSIONS: Very good accordance was found between TRBF measurements, obtained with dual-beam bidirectional Doppler FD-OCT and the microsphere method. It also was possible to monitor changes over time in TRBF after ONT with Doppler OCT. These findings highlight the accuracy and potential of noninvasive Doppler OCT to provide valuable information for detecting early changes in ocular disease in future.
Asunto(s)
Velocidad del Flujo Sanguíneo/fisiología , Microesferas , Traumatismos del Nervio Óptico/fisiopatología , Flujo Sanguíneo Regional/fisiología , Retina/fisiopatología , Vasos Retinianos/fisiopatología , Tomografía de Coherencia Óptica/métodos , Animales , Modelos Animales de Enfermedad , Análisis de Fourier , Macaca mulatta , Masculino , Nervio Óptico/cirugía , Traumatismos del Nervio Óptico/patología , Retina/patología , Células Ganglionares de la Retina/patologíaRESUMEN
PURPOSE: To test the hypothesis that optic nerve head (ONH) blood flow (BF) and autoregulation compromise are consequences of optic nerve degeneration induced by surgical optic nerve transection (ONT). METHODS: In both eyes of five nonhuman primates, peripapillary retinal nerve fiber layer thickness (RNFLT) was measured by spectral-domain optical coherence tomography. Optic nerve head BF and dynamic autoregulation responses to a rapid manometric IOP increase (from 10-40 mm Hg) were measured by Laser Speckle Flowgraphy. The measurements were conducted every 10 to 15 days before and after unilateral ONT. Post-ONT measurements were repeated until RNFLT in the ONT eye was reduced by more than 40% of baseline value. RESULTS: After ONT, RNFLT, and ONH BF progressively declined over time (P < 0.0001 and P = 0.02, respectively). Longitudinal changes between the two were highly correlated (P < 0.0001). When data was grouped by test session, the first significant decreases for RNFLT and BF were found 13 ± 0.8 and 24 ± 3.2 days post ONT, respectively (P < 0.05, both). At the final time point (55 ± 0.5 days post ONT), RNFLT, and BF were reduced by 44% ± 2.0% and 38 ± 5.0% from baseline, respectively. Dynamic autoregulation analysis showed marginal increased response time in post-ONT eyes (P = 0.05). Control eyes showed no longitudinal changes for any parameter. CONCLUSIONS: The close association between RNFLT loss and ONH BF decrease following optic nerve degeneration demonstrated a clear cause and effect relationship. Increased BF response time appears to be a sign of dynamic autoregulation dysfunction in this ONT model.
Asunto(s)
Velocidad del Flujo Sanguíneo/fisiología , Homeostasis/fisiología , Atrofia Óptica/fisiopatología , Nervio Óptico/irrigación sanguínea , Flujo Sanguíneo Regional/fisiología , Animales , Modelos Animales de Enfermedad , Macaca mulatta , Masculino , Atrofia Óptica/patología , Tomografía de Coherencia Óptica/métodosRESUMEN
PURPOSE: To relate changes in retinal function and retinal nerve fiber layer (RNFL) retardance to loss of RNFL thickness and optic nerve axon counts in a nonhuman primate (NHP) model of experimental glaucoma (EG). METHODS: Bilateral longitudinal measurements of peripapillary RNFL thickness (spectral-domain optical coherence tomography, SDOCT; Spectralis), retardance (GDxVCC), and multifocal electroretinography (mfERG; VERIS) were performed in 39 NHP at baseline (BL; median, 5 recordings; range, 3-10) and weekly after induction of unilateral EG by laser photocoagulation of the trabecular meshwork. Multifocal ERG responses were high-pass filtered (>75 Hz) to measure high- and low-frequency component (HFC and LFC) amplitudes, including LFC features N1, P1, and N2. High-frequency component amplitudes are known to specifically reflect retinal ganglion cell (RGC) function. Complete (100%) axon counts of orbital optic nerves were obtained in 31/39 NHP. RESULTS: Postlaser follow-up was 10.4 ± 7.9 months; mean and peak IOP were 18 ± 5 and 41 ± 11 mm Hg in EG eyes, 11 ± 2 and 18 ± 6 mm Hg in control (CTL) eyes. At the final available time point, RNFL thickness had decreased from BL by 14 ± 14%, retardance by 20 ± 11%, and the mfERG HFC by 30 ± 17% (P < 0.0001 each). Longitudinal changes in retardance and HFC were linearly related to RNFL thickness change (R2 = 0.51, P < 0.0001 and R2 = 0.22, P = 0.002, respectively); LFC N2 was weakly related but N1 or P2 (N1: R2 = 0.07, P = 0.11; P1: R2 = 0.04, P = 0.24; N2: R2 = 0.13, P = 0.02). At zero change from BL for RNFL thickness (Y-intercept), retardance was reduced by 11% (95% confidence interval [CI]: -15.3% to -6.8%) and HFC by 21.5% (95% CI: -28.7% to -14.3%). Relative loss of RNFL thickness, retardance, and HFC (EG:CTL) were each related to axon loss (R2 = 0.66, P < 0.0001; R2 = 0.42, P < 0.0001; R2 = 0.42, P < 0.0001, respectively), but only retardance and HFC were significantly reduced at zero relative axon loss (Y-intercept; retardance: -9.4%, 95% CI: -15.5% to -3.4%; HFC: -10.9%, 95% CI: -18.6% to -3.2%; RNFL thickness: +1.8%, 95% CI: -4.9% to +5.4%). CONCLUSIONS: Retinal nerve fiber layer retardance and RGC function exhibit progressive loss from baseline before any loss of RNFL thickness or orbital optic nerve axons occurs in NHP EG. These in vivo measures might serve as potential biomarkers of early-stage glaucomatous damage preceding axon loss and RGC death.
Asunto(s)
Glaucoma/fisiopatología , Fibras Nerviosas/patología , Enfermedades del Nervio Óptico/fisiopatología , Células Ganglionares de la Retina/fisiología , Animales , Axones/patología , Modelos Animales de Enfermedad , Electrorretinografía , Femenino , Presión Intraocular , Macaca mulatta , Masculino , Disco Óptico/patología , Tomografía de Coherencia Óptica/métodosRESUMEN
Glaucoma is a multifactorial, neurodegenerative disease. The molecular mechanisms that underlie the pathophysiological changes in glaucomatous eyes, especially at the early stage of the disease, are poorly understood. Here, we report the findings from a quantitative proteomic analysis of retinas from experimental glaucoma (EG) eyes. An early stage of EG was modeled on unilateral eyes of five nonhuman primates (NHP) by laser treatment-induced elevation of intraocular pressure (IOP). Retinal proteins were extracted from individual EG eyes and their contralateral control eyes of the same animals, respectively, and analyzed by quantitative mass spectrometry (MS). As a result, a total, 475 retinal proteins were confidently identified and quantified. Results of bioinformatic analysis of proteins that showed an increase in the EG eyes suggested changes in apoptosis, DNA damage, immune response, cytoskeleton rearrangement and cell adhesion processes. Interestingly, hemoglobin subunit alpha (HBA) and Ras related C3 botulinum toxin substrate 1 (Rac1) were among the increased proteins. Results of molecular modeling of HBA- and Rac1-associated signaling network implicated the involvement of Mitogen-Activated Protein Kinase (MAPK) pathway in the EG, through which Rac1 may exert a regulatory role on HBA. This is the first observation of this potentially novel signaling network in the NHP retina and in EG. Results of Western blot analyses for Rac1, HBA and a selected MAPK pathway protein indicated synergistic changes in all three proteins in the EG eyes. Further, results of hierarchical cluster analysis of proteomes of control eyes revealed a clear age-proteome relationship, and such relationship appeared disrupted in the EG eyes. In conclusion, our results suggested an increased presence of a potentially novel signaling network at the early stage of glaucoma, and age might be one of the determinant factors in retinal proteomic characteristics under normal conditions.
RESUMEN
PURPOSE: To test the hypothesis that blood flow autoregulation in the optic nerve head has less reserve to maintain normal blood flow in the face of blood pressure-induced ocular perfusion pressure decrease than a similar magnitude intraocular pressure-induced ocular perfusion pressure decrease. MATERIALS AND METHODS: Twelve normal non-human primates were anesthetized by continuous intravenous infusion of pentobarbital. Optic nerve blood flow was monitored by laser speckle flowgraphy. In the first group of animals (n = 6), the experimental eye intraocular pressure was maintained at 10 mmHg using a saline reservoir connected to the anterior chamber. The blood pressure was gradually reduced by a slow injection of pentobarbital. In the second group (n = 6), the intraocular pressure was slowly increased from 10 mmHg to 50 mmHg by raising the reservoir. In both experimental groups, optic nerve head blood flow was measured continuously. The blood pressure and intraocular pressure were simultaneously recorded in all experiments. RESULTS: The optic nerve head blood flow showed significant difference between the two groups (p = 0.021, repeat measures analysis of variance). It declined significantly more in the blood pressure group compared to the intraocular pressure group when the ocular perfusion pressure was reduced to 35 mmHg (p < 0.045) and below. There was also a significant interaction between blood flow changes and the ocular perfusion pressure treatment (p = 0.004, adjusted Greenhouse & Geisser univariate test), indicating the gradually enlarged blood flow difference between the two groups was due to the ocular perfusion pressure decrease. CONCLUSIONS: The results show that optic nerve head blood flow is more susceptible to an ocular perfusion pressure decrease induced by lowering the blood pressure compared with that induced by increasing the intraocular pressure. This blood flow autoregulation capacity vulnerability to low blood pressure may provide experimental evidence related to the hemodynamic pathophysiology in glaucoma.