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BACKGROUND: Eye injuries comprise 10–13% of civilian improvised explosive device (IED) injuries. The bomb blast wave induces a normal and shear forces on the tissues, causing a large acute IOP elevation. This study calculated the biomechanical stresses and strains in the eye due to IED explosion via eye-specific fluid-structure interaction (FSI) models. METHODS: Blast occurred at 2, 3, and 4 m from the front and side of the victim and the weights of the IED were 1 and 2 kg. The ground was covered with the deformable soil to mimic the realistic IED explosion condition and reflect the blast wave. RESULTS: The IOP elevation of ~6,000–48,000 mmHg was observed in the eyes while the highest IOP was occurred with the IED weight and distance of 2 kg and 2 m (front) and the lowest was occurred with the IED weight and distance of 1 kg and 4 m (side). Our findings suggest the importance of the victim location and orientation concerning the blast wave when it comes to ocular injury assessment. IOP elevation of ~2900 and ~2700 mmHg were observed in ~1.6 ms after the blast for the IEDS weight of 2 kg and a victim distance of 2 m in front and side blasts, respectively, in consistence with the literature. Nonetheless, IOPs were considerably higher after ~1.6 ms due to the merging of the bomb blast wave and its reflection off the ground. CONCLUSIONS: The stresses and strains were highest for the frontal blast. Both side and frontal blasts caused higher stresses and strains at the rectus muscle insertions where the sclera is thinnest and prone to rupture. Blast angle has no considerable role in the resultant IOP. Front blast with a heavier IED resulted a higher stresses and deformations in the eye connective tissues compared to the side blast.

Glaucoma is among the leading causes of blindness worldwide that is characterized by irreversible damage to the retinal ganglion cell axons in the lamina cribrosa (LC) region of the optic nerve head (ONH), most often associated with elevated intraocular pressure (IOP). The LC is a porous, connective tissue structure that provides mechanical support to the axons as they exit the eye and the biomechanics of the LC microstructure likely play a crucial role in protecting the axons passing through it. There is a limited knowledge of the IOP-driven biomechanics of the LC microstructure, primarily due to its small size and the difficulty with imaging the LC both in vitro and in vivo. We present finite element (FE) models of three human eye posterior poles that include the LC microstructure and interspersed neural tissues (NT) composed of retinal axons that are constructed directly from segmented, binary images of the LC. These models were used to estimate the stresses and strains in the LC and NT for an acute IOP elevation from 0 to 45 mmHg and compared with identical models except that the LC was represented as a homogenized continuum material with either homogeneous isotropic neo-Hookean properties or heterogeneous properties derived from local connective tissue volume fraction (CTVF) and predominant LC beam orientation. Stresses and strains in the LC and NT microstructure were investigated, and results were compared against those from the models wherein the LC was represented as a homogenized continuum. The regionalized volumetric average stresses and strains showed that the microstructural model yielded similar patterns to our prior approach using an LC continuum representation with mapped LC CTVF/anisotropy, but the microstructural modeling approach allows analysis of the stresses and strains in the LC and NT separately. As expected, the LC beams carried most of the IOP load in the microstructural models but exhibited less strain, while the encapsulated NT exhibited lower stresses and much higher strains. Results also revealed that the continuum models underestimate the maximum strains in the LC beams and NT by a factor of 2-3. Microstructural modeling should provide greater insight into the biomechanical factors driving damage to the axons (NT) and LC connective tissue remodeling that occur in glaucoma. The methods presented are ideal for modeling any structure with a complex microstructure composed of different materials, such as trabecular bone, lung, and tissue engineering scaffolds such as decellularized LC. Matlab code for mesh generation from a segmented image stack of the microstructure is included as Supplemental Material. STATEMENT OF SIGNIFICANCE: Glaucoma is among the leading causes of blindness worldwide that is characterized by axon damage in the lamina cribrosa (LC) region of the eye. We present a new approach for finite element modeling the entire eye-specific 3D LC microstructure and the interspersed neural tissues, incorporated into an eye-specific posterior eye model that provides appropriate boundary and loading conditions. Results are presented for three human donor eyes, showing that prior modeling approaches underestimate the stresses and strains in the laminar microstructure. We constructed models from image stacks of the segmented microstructure (Matlab code included) using an approach that is ideal for modeling any structure with a complex microstructure composed of different materials, such as trabecular bone, lung, and tissue engineering scaffolds.

Background: Although the tissues comprising the ocular conventional outflow pathway have shown strong viscoelastic mechanical response to aqueous humor pressure dynamics, the viscoelastic mechanical properties of the trabecular meshwork (TM), juxtacanalicular connective tissue (JCT), and Schlemm’s canal (SC) inner wall are largely unknown. Methods: A quadrant of the anterior segment from two human donor eyes at low- and high-flow (LF and HF) outflow regions was pressurized and imaged using optical coherence tomography (OCT). A finite element (FE) model of the TM, the adjacent JCT, and the SC inner wall was constructed and viscoelastic beam elements were distributed in the extracellular matrix (ECM) of the TM and JCT to represent anisotropic collagen. An inverse FE-optimization algorithm was used to calculate the viscoelastic properties of the ECM/beam elements such that the TM/JCT/SC model and OCT imaging data best matched over time. Results: The ECM of the glaucoma tissues showed significantly larger time-dependent shear moduli compared to the heathy tissues. Significantly larger shear moduli were also observed in the LF regions of both the healthy and glaucoma eyes compared to the HF regions. Conclusions: The outflow tissues in both glaucoma eyes and HF regions are stiffer and less able to respond to dynamic IOP.

The purpose of this study was to assess ocular coat mechanical behavior using controlled ocular microvolumetric injections (MVI) of 15 µL of balanced salt solution (BSS) infused over 1 second into the anterior chamber (AC) via a syringe pump.Intraocular pressure (IOP) was continuously recorded at 200 Hz with a validated implantable IOP telemetry system in 7 eyes of 7 male rhesus macaques (nonhuman primates [NHPs]) during 5 MVIs in a series at native (3 trials), 15 and 20 mm Hg baseline IOPs, repeated in 2 to 5 sessions at least 2 weeks apart. Ocular rigidity coefficients (K) and ocular pulse volume (PV) were calculated for each trial. Data were averaged across sessions within eyes; PV was analyzed with a three-level nested ANOVA, and parameter relationships were analyzed with Pearson Correlation and linear regression.After MVI at native baseline IOP of 10.4 ± 1.6 mm Hg, IOP increased by 9.1 ± 2.8 mm Hg (∆IOP) at a 9.6 ± 2.7 mm Hg/s slope, ocular pulse amplitude (OPA) was 0.70 ± 0.13 mm Hg on average; the average K was 0.042 ± 0.010 µL-1 and average PV was 1.16 ± 0.43 µL. PV varied significantly between trials, days, and eyes (P ≤ 0.05). OPA was significantly correlated with K at native IOP: Pearson coefficients ranged from 0.71 to 0.83 (P ≤ 0.05) and R2 ranged from 0.50 to 0.69 (P ≤ 0.05) during the first trial.The MVI-driven ∆IOP and slope can be used to assess ocular coat mechanical behavior and measure ocular rigidity.Importantly, OPA at native IOP is correlated with ocular rigidity despite the significant variability in PV between heartbeats.

PURPOSE: Most studies on intraocular pressure (IOP) to monitor IOP “fluctuations” in glaucoma patients have been performed with snapshot tonometry techniques that obtain IOP measurements at single time points weeks to months apart. However, IOP telemetry has shown that IOP varies from second-to-second due to blinks, saccades, and systolic vascular filling. The purpose of this study was to characterize the cyclic pattern of baseline IOP and transient IOP fluctuations in 3 nonhuman primates (NHPs). METHODS: Bilateral IOP was measured using a proven implantable telemetry system and recorded 500 times per second, 24 hours a day, up to 451 continuous days in 3 male rhesus macaques aged 4 to 5 years old. The IOP transducers were calibrated every two weeks via anterior chamber cannulation manometry and all data were continuously corrected for signal drift via software, filtered for signal noise and dropout, and peaks and troughs were quantified and counted using a finite impulse response filter; waking hours were defined as 6:00–18:00 hours based on room light cycle. RESULTS: Fourier transform analyses of baseline IOP and the hourly mean frequency of transient IOP fluctuations > 0.6 mmHg, 0.6–5 mmHg and > 5 mmHg above baseline during waking hours exhibited an approximate 16- to 91-day cyclic pattern in all NHPs. There were no measured environmental or experimental factors associated with this cyclical pattern. CONCLUSIONS: While the importance of the cyclic pattern identified in IOP and its fluctuations is unknown at this time, it is plausible that this pattern is relevant to both homeostasis and pathophysiology of the ONH, corneoscleral shell, and aqueous outflow pathways.

The laminar region of the optic nerve head (ONH), thought to be the site of damage to the retinal ganglion cell axons in glaucoma, is continuously loaded on its anterior and posterior surfaces by dynamic intraocular pressure (IOP) and orbital cerebrospinal fluid pressure (CSFP), respectively. Thus, translaminar pressure (TLP; TLP = IOP-CSFP) has been proposed as a glaucoma risk factor.Three eye-specific finite element models of the posterior human eye were constructed, including full 3D microstructures of the load-bearing lamina cribrosa (LC) with interspersed laminar neural tissues (NTs), and heterogeneous, anisotropic, hyperelastic material formulations for the surrounding peripapillary sclera and adjacent pia. ONH biomechanical responses were simulated using three combinations of IOP and CSFP loadings consistent with posture change from sitting to supine.Results show that tensile, compressive, and shear stresses and strains in the ONH were higher in the supine position compared to the sitting position (P0.05). In addition, LC beams bear three to five times more TLP-driven stress than interspersed laminar NT, whereas laminar NT exhibit three to five times greater strain than supporting LC (P0.05). Compared with CSFP, IOP drove approximately four times greater stress and strain in the LC, NT, and peripapillary sclera, normalized per mm Hg pressure change. In addition, IOP drove approximately three-fold greater scleral canal expansion and anterior-posterior laminar deformation than CSFP per mm Hg (P0.05).Whereas TLP has been hypothesized to play a prominent role in ONH biomechanics, the IOP and CSFP effects are not equivalent, as IOP-driven stress, strain, and deformation play a more dominant role than CSFP effects.

The optic nerve head’s susceptibility to changes in the intraocular pressure profile is a function of both the acute and chronic response of its constituent tissues. This chapter looks at the characteristic changes to the optic nerve as a normal aging process and in glaucoma, in light of its functional anatomy and vascular supply.

The optimal approach for continuous measurement of intraocular pressure (IOP), including pressure transducer location and measurement frequency, is currently unknown. This study assessed the capability of extraocular (EO) and intraocular (IO) pressure transducers, using different IOP sampling rates and duty cycles, to characterize IOP dynamics. Transient IOP fluctuations were measured and quantified in 7 eyes of 4 male rhesus macaques (NHPs) using the Konigsberg EO system (continuous at 500 Hz), 12 eyes of 8 NHPs with the Stellar EO system and 16 eyes of 12 NHPs with the Stellar IO system (both measure at 200 Hz for 15 s of every 150 s period). IOP transducers were calibrated bi-weekly via anterior chamber manometry. Linear mixed effects models assessed the differences in the hourly transient IOP impulse, and transient IOP fluctuation frequency and magnitude between systems and transducer placements (EO versus IO). All systems measured 8000–12,000 and 5000–6500 transient IOP fluctuations per hour > 0.6 mmHg, representing 8–16% and 4–8% of the total IOP energy the eye must withstand during waking and sleeping hours, respectively. Differences between sampling frequency/duty cycle and transducer placement were statistically significant (p

Purpose Recent retrospective clinical studies and animal experiments have suggested that cerebrospinal fluid pressure (CSFP) is important in glaucoma, acting through the translaminar pressure (TLP = IOP - CSFP), which directly affects the optic nerve head. In this study, IOP and intracranial pressure (ICP; a surrogate of CSFP) were measured at various body positions to quantify the determinants of TLP. Methods We have developed an implantable wireless pressure telemetry system based on a small piezoelectric sensor with low temporal drift. Telemetry transducers were placed in the anterior chamber to measure IOP and in the brain parenchyma at eye height to measure ICP. IOP was calibrated against anterior cannulation manometry, and ICP/CSFP was calibrated against an intraparenchymal Codman ICP Express microsensor. We measured IOP, ICP, and TLP = IOP - ICP continuously at 200 Hz in three male nonhuman primates (NHPs) in three trials; pressures were then averaged for 30 seconds per body position. Relative change of IOP, ICP, and TLP from the supine (baseline) position to the seated, standing, and inverted positions were quantified. Results TLP changed significantly and instantaneously from the supine to seated (+14 mm Hg), supine to standing (+13 mm Hg) and supine to inverted (-12 mm Hg) positions (P < 0.05). There was no significant TLP change for supine to prone. ICP showed greater relative change than IOP. Conclusions TLP change due to body position change is driven more by ICP/CSFP than IOP. IOP, ICP, and TLP variability, coupled with telemetry, should allow us to test the hypotheses that IOP, ICP, or TLP fluctuations contribute independently to glaucoma onset or progression.

Purpose To characterize intraocular pressure (IOP) dynamics by identifying the sources of transient IOP fluctuations and quantifying the frequency, magnitude, associated cumulative IOP-related mechanical energy, and temporal distribution. Methods IOP was monitored at 500 Hz for periods of 16 to 451 days in nine normal eyes of six conscious, unrestrained nonhuman primates using a validated, fully implanted wireless telemetry system. IOP transducers were calibrated every two weeks via anterior chamber cannulation manometry. Analysis of time-synchronized, high-definition video was used to identify the sources of transient IOP fluctuations. Results The distribution of IOP in individual eyes is broad, and changes at multiple timescales, from second-to-second to day-to-day. Transient IOP fluctuations arise from blinks, saccades, and ocular pulse amplitude and were as high as 14 mm Hg (>100%) above momentary baseline. Transient IOP fluctuations occur ∼10,000 times per waking hour, with ∼2000 to 5000 fluctuations per hour greater than 5 mm Hg (∼40%) above baseline. Transient IOP fluctuations account for up to 17% (mean of 12%) of the total cumulative IOP-related mechanical energy that the eye must withstand during waking hours. Conclusions Transient IOP fluctuations occur frequently and comprise a large and significant portion of the total IOP loading in the eye and should, therefore, be considered in future studies of cell mechanotransduction, ocular biomechanics, and/or clinical outcomes where transient IOP fluctuations may be important. If IOP dynamics are similar in humans, clinical snapshot IOP measurements are insufficient to capture true IOP.

Glaucoma, a leading cause of blindness, is characterized by optic nerve damage related to intraocular pressure (IOP), but its full etiology is unknown. Researchers at UAB have devised a custom device to measure scleral strain continuously around the eye under fixed levels of IOP, which here is used to assess how strain varies around the posterior pole, with IOP, and across glaucoma risk factors such as age. The hypothesis is that scleral strain decreases with age, which could alter biomechanics of the optic nerve head and cause damage that could eventually lead to glaucoma. To evaluate this hypothesis, we adapted Bayesian Functional Mixed Models to model these complex data consisting of correlated functions on spherical scleral surface, with nonparametric age effects allowed to vary in magnitude and smoothness across the scleral surface, multi-level random effect functions to capture within-subject correlation, and functional growth curve terms to capture serial correlation across IOPs that can vary around the scleral surface. Our method yields fully Bayesian inference on the scleral surface or any aggregation or transformation thereof, and reveals interesting insights into the biomechanical etiology of glaucoma. The general modeling framework described is very flexible and applicable to many complex, high-dimensional functional data.

PURPOSE. To determine if in vivo strain response of the Optic Nerve Head (ONH) to IOP elevation visualized using Optical Coherence Tomography (OCT) video imaging and quantified using novel virtual extensometers was able to be provided repeatable measurements of tissue specific deformations. METHODS. The ONHs of 5 eyes from 5 non-human primates (NHPs) were imaged by Spectralis OCT. A vertical and a horizontal B-scan of the ONH were continuously recorded for 60 seconds at 9Hz (video imaging mode) during IOP elevation from 10 to 30 mmHg. Imaging was repeated over three imaging sessions. The 2D normal strain was computed by template-matching digital image correlation using virtual extensometers. ANOVA F-test (F) was used to compare inter-eye, inter-session, and inter-tissue variability for the prelaminar, Bruch’s membrane opening (BMO), lamina cribrosa (LC) and choroidal regions (against variance the error term). F-test of the ratio between inter-eye to inter-session variability was used to test for strain repeatability across imaging sessions (F(IS)). RESULTS. Variability of strain across imaging session (F=0.7263, p=0.4855) and scan orientation was not significant (F=1.053, p=0.3066). Inter session variability of strain was significantly lower than inter-eye variability (F(IS)=22.63, p=0.0428) and inter-tissue variability (F(IS)=99.33 p=0.00998). After IOP elevation, strain was highest in the choroid (−18.11%, p

Purpose of review This article presents, summarizes, and interprets the most recent advances in the study and understanding of the lamina cribrosa in glaucoma, in the context of previous work. Recent findings The lamina is an active living structure that responds to strain by changing morphology at the micro-scale and macro-scale in glaucoma. Changes in lamina cribrosa morphology in glaucoma include posteriorization of the laminar insertion into the sclera, increased cupping or depth of the lamina cribrosa, and the development of focal lamina cribrosa defects. These lamina cribrosa changes are associated with disk hemorrhages and visual field damage, and are detectable with clinical imaging techniques such as optical coherence tomography. Glaucomatous changes in the lamina cribrosa are thought to be driven by cellular processes mediated by focal cyclical mechanical strain. Strain is eye specific and mediated by intraocular pressure, cerebrospinal fluid pressure, scleral and lamina cribrosa morphology, and structural stiffness; deleterious lamina cribrosa strains can occur at all levels of mean intraocular pressure. Summary Laminar morphology is ever changing in health and disease, and recent studies have identified several promising morphological changes that are indicative of glaucoma susceptibility, onset, and progression.

Commercial finite element modeling packages do not have the tools necessary to effectively incorporate the complex anisotropic and heterogeneous material properties typical of the biological tissues of the eye. We propose a mesh-free approach to incorporate realistic material properties into finite element models of individual human eyes. The method is based on the idea that material parameters can be estimated or measured at so called control points, which are arbitrary and independent of the finite element mesh. The mesh-free approach approximates the heterogeneous material parameters at the Gauss points of each finite element while the boundary value problem is solved using the standard finite element method. The proposed method was applied to an eye-specific model a human posterior pole and optic nerve head. We demonstrate that the method can be used to effectively incorporate experimental measurements of the lamina cribrosa micro-structure into the eye-specific model. It was convenient to define characteristic material orientations at the anterior and posterior scleral surface based on the eye-specific geometry of each sclera. The mesh-free approach was effective in approximating these characteristic material directions with smooth transitions across the sclera. For the first time, the method enabled the incorporation of the complex collagen architecture of the peripapillary sclera into an eye-specific model including the recently discovered meridional fibers at the anterior surface and the depth dependent width of circumferential fibers around the scleral canal. The model results suggest that disregarding the meridional fiber region may lead to an underestimation of local strain concentrations in the retina. The proposed approach should simplify future studies that aim to investigate collagen remodeling in the sclera and optic nerve head or in other biological tissues with similar challenges.

PURPOSE: To compare the three-dimensional (3D) morphology of the deep load-bearing structures of the human optic nerve head (ONH) as revealed in vivo by spectral domain optical coherence tomography (SDOCT) with ex vivo quantitative 3D histology. METHODS: SDOCT imaging of the ONH was performed in six eyes from three brain-dead organ donors on life-support equipment awaiting organ procurement (in vivo conditions). Following organ procurement (ex vivo conditions), the eyes were enucleated and underwent a pars plana vitrectomy followed by pressurization to physiologic IOP and immersion fixation. Ex vivo ONH morphology was obtained from high-fidelity episcopic fluorescent 3D reconstruction. Morphologic parameters of the observed ONH canal geometry and peripapillary choroid, as well as the shape, visibility and depth of the lamina cribrosa were compared between ex vivo and in vivo measurements using custom software to align, scale, and manually delineate the different regions of the ONH. RESULTS: There was significant correspondence between in vivo and ex vivo measurements of the depth and shape of the lamina cribrosa, along with the size and shape of Bruch’s membrane opening (BMO) and anterior scleral canal opening (ASCO). Weaker correspondence was observed for choroidal thickness; as expected, a thinner choroid was seen ex vivo due to loss of blood volume upon enucleation (−79.9 %, p

Purpose This study quantified the thickness and depth of the lamina cribrosa (LC) and peripapillary scleral thickness in high-resolution three-dimensional (3D) fluorescent reconstructions of the optic nerve head (ONH) in eyes from donors of African (AD) and European descent (ED). Methods A total of 64 eyes (45 ED, 19 AD) from 51 normal donors were obtained within 6 hours of death and fixed at 10 mm Hg of pressure. The optic nerve head was trephined from the globe and digitally reconstructed at 1.5 × 1.5 × 1.5 μm voxel resolution with an automated episcopic fluorescence technique. The load-bearing ONH connective tissue surfaces were manually delineated in 3D using custom software. Results The lamina cribrosa and peripapillary sclera were significantly thinner in AD eyes adjusting for age and sex (LC was 24 ± 11 μm thinner; P = 0.0350; scleral was 56 ± 22 μm thinner; P = 0.0097). The lamina cribrosa was significantly thinner in females (23 ± 11 μm thinner; P = 0.0425). Age was not significantly associated with any morphologic parameter in the ED group. However, increasing age was associated with an increase in scleral thickness (1.3 μm/year, P = 0.0499) and an increase in LC depth (2.3 μm/year, P = 0.0035) in the AD group. The sclera was thickest in the superior and temporal regions while the LC was thinnest superiorly. Conclusions Substantial sectorial and racial differences in LC and scleral morphology were observed, as well as increasing LC depth and scleral thickness with age in the AD group. Results suggest greater age-related remodeling of the load-bearing ONH connective tissues in eyes from AD individuals that could explain, in part, the greater predilection to glaucomatous injury seen in aged AD populations.

Since the deformation of structures in the optic nerve head (ONH) is associated with glaucoma and other diseases of the optic nerve, measurement of this deformation is of current research interest. This paper considers the computation of cup depth, a measurement of the depth of the internal limiting membrane (ILM). The computation of cup depth requires a reference structure against which to measure the ILM, and the construction of two reference structures is discussed, one based on Bruch's membrane opening (BMO) and the other based on the anterior surface of the peripapillary sclera (AS). A main focus of the paper is the robust computation of mean cup depth, which requires a good sampling of a reconstruction of the ILM surface. To evaluate our algorithm, the construction of synthetic datasets is considered.

Purpose Recent retrospective clinical and animal studies suggest that cerebrospinal fluid pressure (CSFP) is important in glaucoma pathogenesis. Intraocular pressure (IOP) and CSFP are the driving components of translaminar pressure (TLP = IOP – CSFP), which acts across the lamina cribrosa (LC) thickness to create the translaminar pressure gradient (TLPG = TLP/LC thickness). Methods We developed an implantable wireless telemetry system based on a small piezoelectric sensor with low temporal drift. IOP, measured in the anterior chamber, and intracranial pressure (ICP), measured in the brain parenchyma (as a surrogate for CSFP) were measured at 200 Hz in three male rhesus macaques (nonhuman primates, NHPs) on a 10% duty cycle (15 seconds of every 150-second period). Three-dimensional LC thickness was autosegmented as the mean thickness of the visible hyperreflective band in 48 radial spectral-domain optical coherence tomography b-scans centered on the optic nerve head. Results Results indicated the rank order of IOP, ICP, TLP, and TLPG for waking, sleeping, and 24-hour periods averaged across all days. NHP 150110 had the highest IOP and ICP in all periods; however, it had the lowest TLPG in all periods due to its relatively thick LC. The other two NHPs showed similar shifts in the rank order of possible glaucoma risk factors. Conclusions IOP is the only modifiable and readily measurable pressure-based risk factor for glaucoma. However, other potential risk factors such as ICP, TLP, and TLPG, as well as their rank-order patterns, differed compared to IOP across subjects, demonstrating that a comprehensive view of relevant risk factors is warranted. Translational Relevance Future studies should consider including CSFP, TLP, and TLPG in addition to IOP as potential risk factors when assessing eye-specific glaucoma susceptibility.

Purpose To characterize ocular perfusion pressure (OPP) fluctuations with continuous telemetry over 24-hour periods across multiple days in nonhuman primates (NHPs) to test the hypotheses that OPP differs among NHPs and that the diurnal cycle of OPP is characterized by low OPP during sleep. Methods We have developed and validated two implantable radiotelemetry systems that allow continuous measurement of intraocular pressure (IOP), arterial blood pressure (BP), and OPP up to 500 Hz. OPP was measured unilaterally in 12 male NHPs for periods of 38 to 412 days. IOP transducers were calibrated directly via anterior chamber manometry, and OPP was calculated continuously as central retinal artery BP minus IOP. OPP data were corrected for signal drift between calibrations and averaged hourly. Results OPP varied widely among animals, with daily averages ranging from ∼47 to 65 mm Hg. In eight of 12 NHPs, OPP was significantly lower during sleep compared to waking hours. In three animals, the diurnal cycle was reversed and OPP was significantly higher during sleep (P < 0.05), and one NHP showed no diurnal cycle. Day-to-day OPP variability within NHPs was the largest source of overall OPP variability, even larger than the differences between NHPs. Average daily OPP showed an unexplained ∼32-day cyclic pattern in most NHPs. Conclusions Average OPP varied widely and exhibited differing diurnal cycles in NHPs, a finding that matches those of prior patient studies and indicates that OPP studies in the NHP model are appropriate. Infrequent snapshot measurements of either IOP or BP are insufficient to capture true IOP, BP, and OPP and their fluctuations.

Purpose To characterize relationships between intraocular pressure (IOP), mean arterial pressure (MAP), ocular perfusion pressure (OPP), IOP transient impulse, and IOP baseline impulse using continuous telemetry in nonhuman primates. Methods We used our validated implantable telemetry system to wirelessly record bilateral IOP and arterial BP at 500 Hz in 7 eyes of 4 male rhesus macaques, aged 4 to 5 years. IOP, MAP, OPP, IOP transient impulse, and IOP baseline impulse were averaged into 1-hour periods over 20 days for each NHP. IOP transient impulse was defined as the portion of total IOP due to transient IOP fluctuations

This nontechnical review is focused upon educating the reader on optic nerve head biomechanics in both aging and disease along two main themes: what is known about how mechanical forces and the resulting deformations are distributed in the posterior pole and ONH (biomechanics) and what is known about how the living system responds to those deformations (mechanobiology). We focus on how ONH responds to IOP elevations as a structural system, insofar as the acute mechanical response of the lamina cribrosa is confounded with the responses of the peripapillary sclera, prelaminar neural tissues, and retrolaminar optic nerve. We discuss the biomechanical basis for IOP-driven changes in connective tissues, blood flow, and cellular responses. We use glaucoma as the primary framework to present the important aspects of ONH biomechanics in aging and disease, as ONH biomechanics, aging, and the posterior pole extracellular matrix (ECM) are thought to be centrally involved in glaucoma susceptibility, onset and progression.

Purpose To study the effect and time course of body position changes on IOP in nonhuman primates. Methods We recorded continuous bilateral IOP measurements with a wireless telemetry implant in three rhesus macaques in seven different body positions. IOP measurements were acquired in the seated-upright, standing, prone, supine, right and left lateral decubitus positions (LDPs), and head-down inverted positions. Continuous IOP was recorded for 90 seconds in each position before returning to a supine reference position until IOP stabilized; measurements were averaged after IOP stabilized at each position. Results Head-down inversion increased IOP an average of 8.9 mm Hg, compared to the supine reference. In the LDP, IOP decreased an average of 0.5 mm Hg in the nondependent eye (i.e., the higher eye), while the fellow dependent (i.e., lower) eye increased an average of 0.5 mm Hg, compared to supine reference. Standing and seated positions decreased IOP 1.5 and 2.2 mm Hg, respectively, compared with supine reference. IOP changes occurred within 4 to 15 seconds of a body position change, and timing was affected by the speed at which body position was changed. Compared to the IOP in the supine position, the IOP in the inverted, prone, and seated positions was significantly different (P = 0.0313 for all). The IOP in the standing position was not statistically different from the IOP in the supine position (P = 0.094). In addition, the IOP was significantly different between the nondependent eye and the dependent eye in the LDPs compared to the supine position (P = 0.0313). Conclusions Body position has a significant effect on IOP and those changes persist over time.

Purpose To determine the magnitude of ocular hypotony and the length of recovery time to 6 and 10 mm Hg IOP following anterior chamber (AC) cannulation. Methods Bilateral IOP was recorded 500 times per second via telemetry immediately before, during, and immediately after AC cannulation with a 27-G needle in 10 different sessions at least 2 weeks apart in four male rhesus macaques (nonhuman primates; NHPs) aged 3- to 6-years old. Bilateral IOP was recorded continuously using a proven telemetry system while the NHPs were under general anesthesia during IOP transducer calibration experiments involving manometric control of IOP via AC cannulation, then continuously after the AC needles were removed until IOP recovered to precannulation levels. The change in IOP from baseline to AC cannulation was tested using the signed-rank test. The times necessary for IOP to recover to 6 and 10 mm Hg, respectively, were calculated. Results Average precannulation IOP was 11.5 mm Hg and significantly decreased to an average of 2.3 mm Hg immediately following AC needle removal (P = 0.0156). On average, IOP recovered from 2.3 to 6 and 10 mm Hg in 32.4 and 63.7 minutes, respectively. Recovery times of IOP were not affected by repeated AC cannulations every 2 weeks. Conclusions Generally, IOP recovers relatively quickly after repeated AC cannulation, and did not result in extended duration hypotony. It is important to consider hypotony in animal experiments and clinical procedures involving AC cannulation and paracentesis when consideration of IOP or its effects is important.

Purpose To determine the magnitude of IOP elevation associated with eye rubbing Study Design Experimental Study Subjects Three nonhuman primates (NHPs) Methods Three young adult male rhesus macaques were briefly anesthetized with ketamine and dexmedetomidine, and antibiotic ointment was placed in both eyes. The anesthetic was immediately reversed with atipamezole, and the animals quickly recovered. IOP was continuously recorded at 500 measurements per second during the experiment using a validated implantable wireless telemetry system; high-definition video was recorded while the NHP rubbed its eyes to remove the ointment, and rubbing events were marked in the IOP data. The experiment was repeated four times in each NHP, with numerous eye rubs recorded for each session, and these data were marked and the IOP elevation magnitudes were analyzed using NOTOCORD-hem software. Main Outcome Measures IOP elevation above baseline IOP due to eye rubbing Results IOP increased as much as 310 mmHg due to eye rubbing. The largest IOP elevations were associated with rubbing the eye and orbit with the back of the hand or wrist, rather than the fingers or knuckle. Eye rubs elicited mean IOP elevations of ∼80-150 mmHg above baseline for 3-4 seconds, with peak IOP elevations reaching 205-310 mmHg depending on the individual NHP and eye. Conclusions Rubbing the eyes causes momentary IOP elevations that average 109 mmHg above baseline IOP and can exceed 300 mmHg above baseline IOP in NHP eyes.

Purpose Recent retrospective clinical studies and animal experiments have suggested that cerebrospinal fluid pressure (CSFP) is important in glaucoma pathogenesis. Intraocular pressure (IOP) and CSFP are the driving components of the translaminar pressure (TLP), which directly effects the optic nerve head. This study measured the diurnal cycle of TLP using continuous wireless telemetry in nonhuman primates (NHPs), a common animal model of glaucoma. Methods We have developed an implantable wireless telemetry system based on a small piezoelectric pressure transducer with low drift. Unilateral IOP was measured in the anterior chamber of the eye, and intracranial pressure (ICP, a surrogate measure of CSFP) was measured in the brain parenchyma in four awake, behaving NHPs for periods of 22 to 281 days. IOP and ICP telemetry transducers were calibrated with direct pressure measurements in the eye (every 2 weeks) and brain (monthly). TLP was quantified in real time as IOP-ICP, and hourly means of IOP, ICP, and TLP were analyzed. Results Results show that mean ICP is significantly higher by an average of 4.8 ± 0.8 mmHg during sleeping hours in NHPs (P < 0.01). IOP showed a small but significant nocturnal elevation in two of four animals despite NHPs sleeping upright (P < 0.05). TLP was significantly lower during sleep (7.1 ± 0.6 mmHg; P < 0.01) than when the animals were awake and active (11.0 ± 0.9 mmHg), driven primarily by the large increase in ICP during sleep. Conclusions The 56% increase in TLP during waking hours in NHPs matches the increase in TLP due to postural change from supine to upright reported previously in humans.

Commercial finite element modeling packages do not have the tools necessary to effectively incorporate the complex anisotropic and heterogeneous material properties typical of the biological tissues of the eye. We propose a mesh-free approach to incorporate realistic material properties into finite element models of individual human eyes. The method is based on the idea that material parameters can be estimated or measured at so called control points, which are arbitrary and independent of the finite element mesh. The mesh-free approach approximates the heterogeneous material parameters at the Gauss points of each finite element while the boundary value problem is solved using the standard finite element method. The proposed method was applied to an eye-specific model a human posterior pole and optic nerve head. We demonstrate that the method can be used to effectively incorporate experimental measurements of the lamina cribrosa micro-structure into the eye-specific model. It was convenient to define characteristic material orientations at the anterior and posterior scleral surface based on the eye-specific geometry of each sclera. The mesh-free approach was effective in approximating these characteristic material directions with smooth transitions across the sclera. For the first time, the method enabled the incorporation of the complex collagen architecture of the peripapillary sclera into an eye-specific model including the recently discovered meridional fibers at the anterior surface and the depth dependent width of circumferential fibers around the scleral canal. The model results suggest that disregarding the meridional fiber region may lead to an underestimation of local strain concentrations in the retina. The proposed approach should simplify future studies that aim to investigate collagen remodeling in the sclera and optic nerve head or in other biological tissues with similar challenges.

PURPOSE To determine if laminar depth (LD) and prelaminar tissue volume (PTV) are associated with age and race in healthy human eyes. METHODS Optic nerve head images from enhanced depth imaging spectral-domain optical coherence tomography of 166 normal eyes from 84 subjects of African descent (AD) and European descent (ED) were manually delineated to identify the principal surfaces: internal limiting membrane, Bruch's membrane (BM), anterior sclera (AS), and anterior surface of the lamina cribrosa. These four surfaces defined the LD and PTV using Bruch's membrane opening (BMO) and AS for reference structures. Generalized estimating equations were used to evaluate whether the effect of age on each outcome was differential by race. RESULTS When age was analyzed as a continuous variable, the interaction term between age and race was statistically significant for mean LDBMO (P = 0.015) and mean LDAS (P = 0.0062) after adjusting for axial length and BMO area. For every 1-year increase in age, the LDAS was greater on average by 1.78 μm in AD subjects and less by 1.71 μm in ED subjects. Mean PTV was lower in the older subjects (1248 × 10(6) μm(3) AD, 881 × 10(6) μm(3) ED) compared to the younger subjects (1316 × 10(6) μm(3) AD, 1102 × 10(6) μm(3) ED) in both groups. CONCLUSIONS With increasing age, the LD changes differently across racial groups in normal subjects. The LD in ED subjects showed a significantly decreasing slope suggesting that the lamina moves anteriorly with age in this group.

PURPOSE We tested the hypothesis that there are age- and race-related differences in posterior scleral material properties, using eyes from human donors of European (20-90 years old, n = 40 eyes) and African (23-74 years old, n = 22 eyes) descent. METHODS Inflation tests on posterior scleral shells were performed while full-field, three-dimensional displacements were recorded using laser speckle interferometry. Scleral material properties were fit to each eye using a microstructure-based constitutive formulation that incorporates the collagen fibril crimp and the local anisotropic collagen architecture. The effects of age and race were estimated using Generalized Estimating Equations, while accounting for intradonor correlations. RESULTS The shear modulus significantly increased (P = 0.038) and collagen fibril crimp angle significantly decreased with age (P = 0.002). Donors of African descent exhibited a significantly higher shear modulus (P = 0.019) and showed evidence of a smaller collagen fibril crimp angle (P = 0.057) compared to donors of European descent. The in-plane strains in the peripapillary sclera were significantly lower with age (P < 0.015) and African ancestry (P < 0.015). CONCLUSIONS The age- and race-related differences in scleral material properties result in a loss of scleral compliance due to a higher shear stiffness and a lower level of stretch at which the collagen fibrils uncrimp. The loss of compliance should lead to larger high frequency IOP fluctuations and changes in the optic nerve head (ONH) biomechanical response in the elderly and in persons of African ancestry, and may contribute to the higher susceptibility to glaucoma in these at-risk populations.

Purpose To determine the relationship between peripapillary scleral strain change and cumulative differential IOP exposure in nonhuman primates (NHPs) with unilateral chronic ocular hypertension. Methods Posterior scleral shells from 6 bilaterally normal and 10 unilateral chronic ocular hypertension NHPs were pressurized from 5 to 45 mm Hg, and the resulting full-field, three-dimensional, scleral surface deformations were acquired using laser speckle interferometry. Scleral tensile strain (local tissue deformation) was calculated by analytical differentiation of the displacement field; zero strain was assumed at 5 mm Hg. Maximum principal strain was used to represent the scleral strain, and strains were averaged over a 15°-wide (∼3.6-mm) circumpapillary region adjacent to the ONH. The relative difference in mean strain was calculated between fellow eyes and compared with the differential cumulative IOP exposure within NHPs during the study period. The relationship between the relative difference in scleral strain and the differential cumulative IOP exposure in fellow eyes was assessed using an F test and quadratic regression model. Results Relative differential scleral tensile strain was significantly associated with differential cumulative IOP exposure in contralateral eyes in the chronic ocular hypertension NHPs, with the bilaterally normal NHPs showing no significant strain difference between fellow eyes. The sclera in the chronic ocular hypertension eyes was more compliant than in their fellow eyes at low levels of differential cumulative IOP exposure, but stiffer at larger differential IOPs (P < 0.0001). Conclusions These cross-sectional findings suggest that longitudinal IOP-induced changes in scleral mechanical behavior are dependent on the magnitude of differential cumulative IOP exposure.

Purpose To compare the effects of both injectable anesthesia (ketamine/dexmedetomidine versus ketamine/xylazine) and inhalant anesthesia (isoflurane) on IOP using continuous, bilateral IOP telemetry in nonhuman primates (NHP). Methods Bilateral IOP was recorded continuously using a proven implantable telemetry system in five different sessions at least 2 weeks apart in four male rhesus macaques under two conditions: ketamine (3 mg/kg) with dexmedetomidine (50 μg/kg) or ketamine with xylazine (0.5 mg/kg) for induction, both followed by isoflurane for maintenance. IOP transducers were calibrated via anterior chamber manometry. Bilateral IOP was averaged over 2 minutes after injectable anesthetic induction and again after isoflurane inhalant had stabilized the anesthetic plane, then compared to baseline IOP measurements acquired immediately prior to anesthesia (both before and after initial human contact). Results When compared to pre-contact baseline measurements, ketamine/dexmedetomidine injectable anesthesia lowers IOP by 1.5 mm Hg on average (P

To test the hypothesis that mechanical strain in the posterior human sclera is altered with age, 20 pairs of normal eyes from human donors aged 20 to 90 years old were inflation tested within 48-h postmortem. The intact posterior scleral shells were pressurized from 5 to 45 mmHg, while the full-field three-dimensional displacements of the scleral surface were measured using laser speckle interferometry. The full strain tensor of the outer scleral surface was calculated directly from the displacement field. Mean maximum principal (tensile) strain was computed for eight circumferential sectors (45° wide) within the peripapillary and mid-peripheral regions surrounding the optic nerve head (ONH). To estimate the age-related changes in scleral strain, results were fit using a functional mixed effects model that accounts for intradonor variability and spatial autocorrelation. Mechanical tensile strain in the peripapillary sclera is significantly higher than the strain in the sclera farther away from the ONH. Overall, strains in the peripapillary sclera decrease significantly with age. Sectorially, peripapillary scleral tensile strains in the nasal sectors are significantly higher than the temporal sectors at younger ages, but the sectorial strain pattern reverses with age, and the temporal sectors exhibited the highest tensile strains in the elderly. Overall, peripapillary scleral structural stiffness increases significantly with age. The sectorial pattern of peripapillary scleral strain reverses with age, which may predispose adjacent regions of the lamina cribrosa to biomechanical insult. The pattern and age-related changes in sectorial peripapillary scleral strain closely match those seen in disk hemorrhages and neuroretinal rim area measurement change rates reported in previous studies of normal human subjects.

The purpose of this study is to determine the effects of intraocular pressure (IOP) mean, maximum and variability on the rate of structural change in experimental glaucoma. Data were taken retrospectively from 59 non-human primates involved in ongoing studies of experimental glaucoma. IOP was measured by tonometry every 1-3 weeks, and these readings split into non-overlapping fixed-length windows. First, different characterizations of IOP variability were tested to find the one that was least correlated with the mean IOP within the same window. Next, the rates of change of the Mean Position of the Disc (MPD) from confocal scanning laser tomography, and Retinal Nerve Fiber Layer Thickness (RNFLT) from spectral domain ocular coherence tomography, were calculated over each window. Mixed effects models were formed to predict these rates based on the characterizations of IOP. Normalized root mean squared residual (RMSR) from the trend of IOP during windows of five IOP measurements provided a characterization of variability showing lowest correlation with mean IOP (r0.001). In univariate analyses, rate of change of MPD and RNFLT were predicted by mean IOP (p0.001 for both) and maximum IOP (p0.001 for both). IOP variability did not significantly predict change in MPD (p = 0.129) or RNFLT (p = 0.438). In bivariate models, maximum IOP was the most significant predictor of change. We conclude that normalized RMSR allows the effects of IOP variability to be assessed independently of mean IOP. Maximum IOP provided the best predictability of structural change, either causally or because it captures the contributions of both mean and variability.

Purpose Primates and rodents are used widely as animal models of glaucoma, but each has significant limitations. Researchers need additional animal models that closely resemble the relevant anatomy and pathologic features of the human disease to more quickly advance research. We validate a novel glaucoma animal model in tree shrews (Tupaia belangeri). Methods Experimental glaucoma was induced in adult tree shrews (n = 8) by injecting 50 μL of a 25 mg/mL ferromagnetic bead solution into the anterior chamber. Beads were directed into the iridocorneal angle with a magnet to impede aqueous outflow. Animals were followed for 3 months with weekly IOP measurements and biweekly spectral domain optical coherence tomography (SD-OCT) images of the optic nerve head. Histopathology of the optic nerve and optic nerve axon counts were completed at the end of the study. Results The 12-week average mean IOP was 22.7 ± 3.6 and 8.6 ± 2.9 mm Hg in the treated and control eyes, respectively. Longitudinal analysis showed significant retinal nerve fiber layer (RNFL) thinning throughout the study. Axon counts were significantly reduced (59.7%) in treated versus control eyes. SD-OCT imaging showed cupping and posterior displacement of the lamina cribrosa in glaucomatous eyes. RNFL thickness and optic nerve axon counts were reduced consistent with IOP elevation. Optic nerves demonstrated histopathology consistent with glaucomatous optic neuropathy. Conclusions Tree shrews with experimental glaucoma show key pathologic characteristics of the human disease. The tree shrew model of glaucoma has the potential to help researchers accelerate our understanding of glaucoma pathophysiology.

The optic nerve head (ONH) is of particular interest from a biomechanical perspective because it is a weak spot within an otherwise strong corneo-scleral envelope. The lamina cribrosa provides structural and functional support to the retinal ganglion cell axons as they pass from the relatively high-pressure environment in the eye to a low-pressure region in the retrobulbar cerebrospinal space. To protect the retinal ganglion cell axons within this unique environment, the lamina cribrosa in higher primates has developed into a complex structure composed of a three-dimensional network of flexible beams of connective tissue. The ONH is nourished by the short posterior ciliary arteries, which penetrate the immediate peripapillary sclera to feed capillaries contained within the laminar beams. This intrascleral and intralaminar vasculature is unique in that it is encased in load-bearing connective tissue, either within the scleral wall adjacent to the lamina cribrosa, or within the laminar beams themselves. Glaucoma is a multifactorial disease, and we believe that biomechanics not only determines the mechanical environment in the ONH, but also mediates IOP-related reductions in blood flow and cellular responses through various pathways. Our current understanding of the mechanical environment of the ONH is described, with particular emphasis on the influence of biomechanics in glaucoma.

Glaucoma is among the leading causes of blindness worldwide. The disease involves damage to the retinal ganglion cell axons that transmit visual information from the eye to the brain. Experimental evidence indicates that biomechanical mechanisms at different length scales are involved in pathophysiology of glaucoma, where chronic intraocular pressure (IOP) elevation at the organ level initiates axonal insult at the level of the lamina cribrosa. The lamina cribrosa consists of a porous collagen structure through which the axons of retinal ganglion cells (RGCs) pass on their path from the retina to the brain. The extent to which the structural mechanics of the lamina cribrosa contribute to the axonal insult remains unclear. In this book chapter, we give a short review of the present understanding of the structural mechanics of the lamina cribrosa and its role in glaucoma. The main aim is to present a first computationally coupled two-scale analysis of the lamina cribrosa that translates the IOP load at the macroscale to the mechanical insult of the axons within the mesostructure of the lamina cribrosa. The numerical results of two-scale analysis suggest that the collagen structures of the lamina cribrosa and its surrounding peripapillary sclera effectively provide mechanical support to the axons by protecting them from high tensile stresses even at elevated IOP levels. However, in-plane shear stresses in the axonal tissue may increase with increasing IOP at the posterior lamina insertion region and contribute to a mechanical insult of the RGC axons in glaucoma.

Deformation of the load-bearing tissues of the optic nerve head (ONH) due to IOP-generated stress is believed to play an important role in the pathogenesis of glaucomatous optic neuropathy.1–3 The peripapillary sclera (ppSc) provides mechanical boundary conditions of the ONH at the neural canal and transmits biomechanical strain directly to the lamina cribrosa (LC) at its insertion into the scleral canal wall. Hence, the IOP-related biomechanical response of the ppSc plays a crucial role in the regulation of biomechanical response and environment of the ONH and LC.4–6 Histomorphometric and computational modeling studies provide evidence that both the material properties and geometry of the ppSc, which are altered in glaucomatous eyes of both human patients and experimental nonhuman primates,7–11 significantly impact the scleral canal and LC deformations in response to IOP variation.12–16 These findings suggest that the material properties and geometry of the ppSc are determinants of the mechanical environment at the ONH. Mechanical strain induced by IOP not only alters the physical properties of load-bearing sclera, but activates the cells residing within the sclera that can in turn alter the material properties and geometry of the sclera through production of extracellular matrix (ECM) proteins and secretion of connective tissue remodeling factors, including matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs).17–19 In most mammalian species, fibroblasts are the primary cells found in sclera, although chondrocytes have been reported in the inner cartilaginous layer of sheep.20 In vivo assessments of scleral creep response in tree shrew eyes versus chick eyes suggest that specialized contractile fibroblasts, known as myofibroblasts, play an important role in defining the mechanical properties of sclera.21 Myofibroblasts are effector cells that drive connective tissue remodeling in the tissue injury–repair process.22 These cells are absent in most adult tissues under normal conditions, but can be differentiated from various progenitor cells, including fibroblasts, during tissue repair and remodeling processes.22 The sclera of adult human, monkey, tree shrew, and guinea pig has been found to contain an endogenous population of myofibroblasts.21,23,24 Although the origin and function of scleral myofibroblasts remain to be determined, it is thought that the presence of permanent scleral myofibroblasts might be a result of constant IOP-related strain, and that scleral myofibroblasts might work with collagen and other ECM components to resist the tensile effects of IOP on the eye. Myofibroblasts are activated fibroblasts that produce increased amounts of ECM proteins and ECM-regulating factors compared with their dormant progenitors.25 However, unlike fibroblasts, myofibroblasts develop a significant capacity for contractile force through expressing a variety of actomyosin components, including α smooth muscle actin (αSMA),26 making myofibroblasts capable of rapidly responding to imposed tissue stress, limiting expansion of the surrounding ECM. Because of the significant contractile potential, the number of myofibroblasts present in the ppSc would be expected to play a key role in biomechanical responses of the posterior sclera as well as the level of tensile stress applied on the ONH. The purpose of this study was to determine the effects of altered mechanical strain on human ppSc fibroblast-to-myofibroblast differentiation.

Mice are a common animal model for studies of aqueous humor dynamics and outflow physiology. The anatomy of the conventional outflow pathway in mice is similar to that of humans with a continuous Schlemm's canal and lamellated trabecular meshwork.1 Like primates, mice possess a ciliary muscle that forms tendinous connections to the elastic fiber net of the trabecular meshwork and the inner wall endothelium of Schlemm's canal.2 Compounds that affect outflow facility in humans, including pilocarpine,2,3 TGF-β2,4,5 latanoprost6–8 prostaglandin EP4 receptor agonist,9–11 and sphingosine 1-phosphate,11,12 similarly affect outflow facility in mice. Recently, mice have been used to validate novel compounds that increase outflow facility based on hits from screening assays of cellular contractility.13 Numerous investigators have measured outflow facility in mice.2,5,6,8,11,14–26 However, on account of the small dimensions of the mouse eye and the low flow rates involved, there is greater potential for physical factors to influence ocular perfusion measurements in mice compared with larger species. For example, evaporation from the surface of the eye, which is more pronounced in smaller eyes that have a larger surface to volume ratio, may lead to dehydration of the corneoscleral shell and artifactually increase the apparent outflow rate. Importantly, this effect would manifest as a pressure-independent outflow during perfusion. Posterior bowing of the iris, known as anterior chamber (AC) deepening, artificially increases outflow facility by applying traction to the trabecular meshwork.27,28 Anterior chamber deepening typically occurs during ocular perfusion via the AC, when the pressure in the AC exceeds that in the posterior chamber (PC). The pressure difference causes the iris–lens channel to collapse like a 1-way valve, preventing pressure equilibration across the iris. Anterior chamber deepening can be prevented by perfusion via the PC, by creating a fluidic shunt across the iris,29 or by iridectomy.30 Note that AC deepening is not synonymous with ‘AC depth,' which represents the distance between the posterior cornea and anterior lens. Because mice have a relatively large crystalline lens, it has been proposed that AC deepening may be negligible in mice,18 but this has not been specifically examined. To address these gaps in knowledge, this study examined the influence of hydration and AC deepening on pressure-dependent and pressure-independent outflow in enucleated mouse eyes. We also examined the effect of temperature that could account for up to 40% variation in apparent outflow facility due to changes in water viscosity between room and physiological temperature.