Outflow Pathway Research Articles

Insights into the practice of minimally invasive glaucoma surgery

Minimally invasive glaucoma surgeries (MIGS) offer an alternative approach, aiming to be both safe and effective. Defined by their ab interno technique, which avoids creating a bleb, MIGS procedures seek to reduce intraocular pressure (IOP) by enhancing the outflow pathways of aqueous humor. Techniques include stenting, incisions, and excisions of the trabecular meshwork, canaloplasty of Schlemm's canal, and drainage implants that enhance uveoscleral outflow to the subchoroidal space. Many of these methods show promise in reducing intraocular pressure and decreasing the need for medication. Nevertheless, with the increasing number of these procedures and devices on the market, it is difficult to keep track of the evidence supporting these methods. Although randomized controlled trials exist for many of the new MIGS procedures, the sample sizes are often small, and follow-up periods are limited. Additionally, there are no randomized controlled trials comparing different MIGS techniques or MIGS with traditional glaucoma surgery. These factors continue to make it challenging to select the most appropriate method for each patient. Therefore, it is essential to persist in conducting randomized controlled long-term studies to gain a deeper understanding of the applications of MIGS.

Differential impact of lymphatic outflow pathways on cerebrospinal fluid homeostasis.

Dysfunctional lymphatic drainage from the central nervous system (CNS) has been linked to neuroinflammatory and neurodegenerative disorders, but our understanding of the lymphatic contribution to CNS fluid autoregulation remains limited. Here, we studied forces that drive the outflow of the cerebrospinal fluid (CSF) into the deep and superficial cervical lymph nodes (dcLN and scLN) and tested how the blockade of lymphatic networks affects CNS fluid homeostasis. Outflow to the dcLN occurred spontaneously in the absence of lymphatic pumping and was coupled to intracranial pressure (ICP), whereas scLN drainage was driven by pumping. Impaired dcLN drainage led to elevated CSF outflow resistance and delayed CSF-to-blood efflux despite the recruitment of the nasal-to-scLN pathway. Fluid regulation was better compensated after scLN obstruction. The dcLN pathway exhibited steady, consistent drainage across conditions, while the nasal-to-scLN pathway was dynamically activated to mitigate perturbances. These findings highlight the complex physiology of CSF homeostasis and lay the groundwork for future studies aimed at assessing and modulating CNS lymphatic function.

Features of the condition of the perilimbal tissues of the eye during the surgical treatment of patients with open-angle glaucoma

primary open-angle glaucoma is a multifactorial chronic neurodegenerative disease characterized by acquired loss of retinal ganglion cells and subsequent optic nerve atrophy, and is often associated with increased intraocular pressure (IOP). As the leading cause of blindness and vision loss, glaucoma affects more than 70 million people worldwide (Tang et al., 2021), and unfortunately, there is no downward trend. Most currently available glaucoma treatments tend to target the formation of the aqueous humor or the uveal-scleral outflow pathway. They do not affect the primary outflow pathway, which is usually responsible for 70–90% of aqueous humor drainage into the circulatory system. However, dysfunction and blockage of this major pathway can lead to increased outflow resistance, increased IOP, and ultimately the development of glaucoma. Purpose: to investigate the condition of the perilimbal tissues of the eye during the surgical treatment of patients with open-angle glaucoma. Materials and methods: 22 patients with primary open-angle glaucoma who underwent combined treatment for glaucoma and age-related cataract were included in the examination and made up the main group of the examination. The comparison group, which consisted of 7 patients, included patients who had age-related cataracts and did not have glaucoma. The main group of patients was divided into two subgroups. In the first subgroup of the main group, the Tenon's sheath of the area where anti-glaucomatous intervention was performed was studied. Trabecula examination was performed in patients of the second subgroup of the main group. Results: on the histological preparations of the patients of the main group, unlike the comparison group, signs of a chronic inflammatory process were revealed: neovascularization with proliferation of endotheliocytes. It was established that the nuclei of cells of fibrous tissue are changed, namely, they are increased in size and more intensely colored. Significant swelling of the surrounding tissue was observed, which pathologically changed the very structure of the tissue. Pathological deposition of diffusely located pigment was noted extracellularly in tissue structures. Conclusions: The results obtained during the study of changes in the perilimbal tissues of the eye in patients with primary open-angle glaucoma will open new ways of personalized management of this category of patients, which will make it possible to formulate additional criteria for the diagnosis and treatment of glaucoma, aimed at overcoming the inflammatory process and prolonging the postoperative result.

Application progress of micropulse wave transscleral laser treatment in glaucoma

Glaucoma is the leading cause of blindness worldwide, and reducing intraocular pressure is crucial to delaying its progression. This review aims to explore the application progress of micropulse wave transscleral laser treatment (mTLT) in the treatment of glaucoma. This article introduces the evolution of ciliary body destructive surgical methods, and focuses on the mechanism of mTLT in the treatment of glaucoma, including reducing aqueous humor production, increasing the uveoscleral pathway and trabecular meshwork pathway for aqueous humor outflow. At the same time, the indications, contraindications, procedures and parameter settings of mTLT, its application effects in different types of glaucoma, possible complications, repeatability, and factors affecting the effect are discussed, and a comparison is made with the traditional TSCPC. However, the exact mechanism of mTLT in reducing intraocular pressure still needs to be further explored and clarified, and its related parameter settings still require a large number of clinical observations and studies.

Gender differences in canine anterior chamber dimensions: Potential implications for primary angle-closure glaucoma.

This study aimed to explore anatomical differences in the aqueous humor (AH) outflow pathway between male and female dogs using ultrasound biomicroscopy (UBM). Clinical data were collected from 30 eyes of 30 dogs (15 males: 10 right eyes, 5 left eyes; and 15 females: 7 right eyes, 8 left eyes) treated at the Veterinary Medical Teaching Hospital of Chungbuk National University, South Korea, between August 2018 and February 2024. The study conducted an in-depth UBM examination, specifically observing peripheral anterior chamber depth (ACD), geometric iridocorneal angle (ICA), and angle-opening distance (AOD) alongside measurements of ciliary cleft width (CCW), length (CCL), and area (CCA), and assessed the longitudinal fibers of ciliary muscle thickness (Lf-CMT) and the longitudinal and radial fibers of ciliary muscle thickness (LRf-CMT) for gender-based differences. The study identified a significant sex difference in peripheral ACD, with females displaying shallower depths compared to males, potentially clarifying the higher incidence of glaucoma in female dogs. No significant gender differences were found in ICA, AOD, CC parameters, or ciliary muscle parameters measurements. This research uncovered that peripheral ACD is significantly shallower in females than in males, potentially indicating a risk factor that could contribute to the development of primary angle-closure glaucoma.

Influence of perioperative therapy on filtration bleb formation and ophthalmotonus level after glaucoma surgery

Introduction. One limitation of glaucoma surgery is its reduced long-term effectiveness due to scarring of intraocular fluid outflow pathways. Appropriate perioperative therapy may extend the hypotensive effect by reducing scarring and improving the filtration bleb. Purpose: to evaluate the effect of perioperative use of Oku-Oku® on intraocular pressure (IOP) and filtration bleb condition in the early postoperative period following glaucoma surgery. Materials and methods. Thirty patients (30 eyes) who underwent non-penetrating glaucoma surgery (NPGS) were studied. The main group (15 eyes) received Oku-Oku® drops twice daily: the day before surgery, immediately before surgery, and for five days postoperatively. The control group (15 eyes) received no additional medication. Tonometry and assessment of conjunctival hyperemia were performed the day before surgery, and at 1 day, 2 weeks, and 1 month postoperatively. Filtration bleb characteristics were assessed using the Würzburg Clinical and Morphological Classification of Filtration Pads at the same time points, except for the preoperative assessment. Results. Glaucoma surgery significantly reduced IOP in both groups. After 1 month, the mean IOP in the main group was lower (15.1 ± 4.1 mm Hg) compared to the control group (17.9 ± 3.3 mm Hg). Conjunctival hyperemia was also lower in the main group (15.4 ± 2.7%) than in the control group (18.0 ± 3.1%). Filtration bleb parameters improved in both groups over time, with the greatest differences observed at 1 day and 2 weeks post-surgery. Conclusion. Perioperative use of Oku-Oku® reduced conjunctival hyperemia in the filtration bleb area during the first month after glaucoma surgery. This early improvement in the intraocular fluid deposition area may enhance the effectiveness of glaucoma surgery. Oku-Oku® can be used in comprehensive preparation for glaucoma surgery and other similar anterior segment procedures.

Anatomical insights from intraoperative OCT in microincisional trabeculectomy

The trabecular meshwork (TM) offers the maximum resistance to aqueous outflow that regulates the intraocular pressure. Newer ab interno procedures have opened up newer frontiers for understanding the functional relationship between the TM and Schlemm’s canal. This case provides newer anatomical insights from findings during intraoperative use of anterior segment optical coherence tomography during ab interno microincisional trabeculectomy. Growing evidence and intraoperative findings during ab interno trabeculectomy suggest that other parts of the TM and its intricate understanding of the physiology of the outflow pathway deserve more study.

Investigation of nitrogen conversion in ditch-treated greywater: Contributions of biochar and aeration

This study aimed to assess the contribution of different combinations of uninterrupted aeration and tidal flow modes, as well as shallow and deep biochar landfill methods, to nitrogen removal pathways in ditches. Compared to natural ditches, the addition of biochar and aeration resulted in higher water pollution removal rates, lower Greenhouse Gas (GHG) emissions, and reduced sediment leaching. The nitrogen mass balance revealed that Total Nitrogen removal efficiency of each ditch group was 25.33% for CA-C, 43.75% for DBT-C, 49.08% for UBT-C, 30.75% for DBA-C, and 45.16% for UBA-C. Compared to the control group (CA-C), the tidal flow group with deep biochar landfill (DBT-C)demonstrated the highest efficiency in total nitrogen removal. To elucidate the fate of nitrogen elements, potential transformation processes of unexplained nitrogen were explored. The primary outflow pathway of nitrogen pollution in ditches is through liquids. The nitrogen composition in the liquid of each ditch indicated that for shallow biochar configurations (UBT-C/UBA-C), denitrification might be the main factor limiting TN removal efficiency. In contrast, for deep biochar configurations (DBT-C/DBA-C), hydrolysis and nitrification of organic nitrogen were identified as limiting factors, with tidal flow alleviating these limitations. Overall, the combination of shallow biochar and tidal flow aeration in a ditch is the most effective method for removing rural greywater, providing valuable insights for the development of design parameters in greywater removal systems.

Groundwater flow system of the Abijata-Langano-Ziway lakes basin, Ethiopia

The Abijata-Langano-Ziway Lakes Basin (ALZLB) is situated in the Central part of the Main Ethiopian Rift. The availability and dynamics of groundwater in the Abijata-Langano-Ziway Lakes Basin (ALZLB) are primarily controlled by its geological and hydrogeological characteristics, shaped by volcanic-tectonic and sedimentary processes. The basin faces significant challenges, including drastic change in land use pattern, rapid population growth sustained by subsistence farming, over-extraction of water resources, and vulnerability to climate change and fragile ecosystems. These issues emphasize the urgent need for effective water resource management.To understand this complex system, a numerical groundwater flow model was employed to characterize the groundwater flow system within the ALZLB and examine its interaction with surface water bodies. The MODFLOW model translates the conceptual understanding of the basin's hydrogeology into a mathematical representation, allowing for numerical analysis. The model incorporates input parameters such as hydraulic conductivity and boundary conditions representing groundwater inflow and outflow. Steady-state numerical calculation was used to characterize the qualitative hydrogeological conceptual model into numerical representation and thereby describe the groundwater system.The calibrated model exhibited excellent agreement between simulated and observed groundwater levels. Statistical measures indicated a strong correlation (R2 = 0.98) and high efficiency (NSE = 0.97) in replicating the observed data. Additionally, the Mean Error (ME) of −8.3 m suggests minimal bias in the simulations. Further analysis of the histogram residuals revealed that a significant portion of the simulated values (65 % and 82 %) fell within ±20 m and ±30 m of the observed groundwater levels, respectively. This revealed the model's accuracy in capturing the groundwater system's behavior.The model identified groundwater recharge and constant head boundaries as the primary sources of groundwater inflow, contributing 445 million cubic meters per year (MCM/year) and 90 MCM/year, respectively. Conversely, constant head boundaries represented the most significant outflow pathway, with a simulated discharge of 519 MCM/year. The calibrated balance between inflow and outflow (discrepancy of −0.75 %) confirms that the model effectively simulates steady-state groundwater flow conditions. The calibrated model demonstrates the model's capability to accurately represent the basin's groundwater system.

Mitochondrial and Microtubule Defects in Exfoliation Glaucoma.

Exfoliation Syndrome (XFS) is an age-related systemic condition characterized by large aggregated fibrillar material deposition in the anterior eye tissues. This aggregate formation and deposition on the aqueous humor outflow pathway are significant risk factors for developing Exfoliation Glaucoma (XFG), a secondary open-angle glaucoma. XFG is a complex, multifactorial late-onset disease that shares common features of neurodegenerative diseases, such as altered cellular processes with increased protein aggregation, impaired protein degradation, and oxidative and cellular stress. XFG patients display decreased mitochondrial membrane potential and mitochondrial DNA deletions. Here, using Tenon Capsule Fibroblasts (TFs) from Normal (No Glaucoma) and XFG patients, we found that XFG TFs have impaired mitochondrial bioenergetics and increased reactive oxygen species (ROS) accumulation. These defects are associated with mitochondrial abnormalities as XFG TFs exhibit smaller mitochondria that contain dysmorphic cristae, with an increase in mitochondrial localization to lysosomes and slowed mitophagy flux. Mitochondrial dysfunction in the XFG TFs was associated with an increase in the dynamics of the microtubule cytoskeleton, decreased acetylated tubulin, and increased HDAC6 activity. Treatment of XFG TFs with a mitophagy inducer, Urolithin A, and a mitochondrial biogenesis inducer, NAD + precursor, Nicotinamide Ribose, improved mitochondrial bioenergetics and reduced ROS accumulation. Our results demonstrate abnormal mitochondria in XFG TFs and suggest that mitophagy inducers may represent a potential class of therapeutics for reversing mitochondrial dysfunction in XFG patients.

Physiological activation of liver X receptor provides protection against ocular inflammation in uveitic glaucoma

Virus-induced trabeculitis is considered a significant cause of uveitic glaucoma, being marked by a sudden increase in intraocular pressure and relatively mild inflammation in the anterior chamber of the eye. In previous proteome analyses of aqueous humor (AH) derived from Cytomegalovirus (CMV) uveitic glaucoma patients, we observed the liver X receptor (LXR) pathway to be among the most prominently activated canonical pathways. In the present study, we explored the role of the LXR pathway in the etiology of glaucoma in association with ocular inflammation. LXRα/β and ABCA1, the downstream targets of LXR, were distributed throughout the conventional AH outflow pathway of the human eye, and their increased levels in human trabecular meshwork cells in response to CMV infection and -lipopolysaccharide (LPS) treatment. Treatment with an LXR agonist (T091317) suppressed LPS-induced inflammation and this response was reversed under the deficiency of LXRα/LXRβ. Furthermore, in the rat endotoxin uveitis model, the LXR agonist significantly reduced infiltrating cells and expression of proinflammatory cytokines in the iris and retina. These results reveal upregulation of LXR-ABCA1 under inflammatory insult in the conventional AH outflow pathway, and activation of LXR exhibiting an anti-inflammatory effect, implying its essential physiological protective role in glaucoma associated with ocular inflammation.

Aqueous Humor Angiography.

Aqueous humor angiography, or aqueous angiography, is an anterior segment imaging technique capable of visualizing the conventional/trabecular aqueous humor outflow pathways. As a translational technique, it is applicable for in vivo imaging in living subjects and ex vivo imaging using whole-globe preparations or anterior segment perfusion setups. Excellent spatial and temporal resolution has enabled insights into the segmental distribution of aqueous humor outflow in physiological conditions as well as after trabecular bypass surgery. In this chapter, we thoroughly describe aqueous humor angiography in various experimental setups. The necessary materials for different settings and their application for in vivo and ex vivo experiments as well as notes on dye choice, dye sequences, and special considerations on performing aqueous humor angiography during surgery are presented.

Time-Lapse Live-Cell Imaging Using Fluorescent Protein Sensors in Outflow Pathway Cells Under Fluid Flow Conditions.

The role of shear stress in regulating aqueous humor (AH) outflow and intraocular pressure (IOP) in the trabecular meshwork (TM) and Schlemm's canal (SC) of the eye is an emerging field. Shear stress has been shown to activate mechanosensitive ion channels in TM cells and induce nitric oxide production in SC cells, which can affect outflow resistance and lower IOP. Live-cell imaging using fluorescent protein sensors has provided real-time data to investigate the physiological relationship between fluid flow and shear stress in the outflow pathway cells. The successful application of time-lapse live-cell imaging in primary cultured cells has led to the identification of key cellular and molecular mechanisms involved in regulating AH outflow and IOP, including the role of autophagy and primary cilia as mechanosensors. This chapter presents a detailed protocol for conducting time-lapse live-cell imaging under fluid flow conditions in the outflow pathway cells.

Evaluation of Cross-Linked Actin Networks (CLANs) in Human Trabecular Meshwork Cells and Tissues.

Elevated intraocular pressure (IOP) is a major risk factor for the development and progression of glaucoma, the leading cause of irreversible vision loss and blindness. An overall increase in resistance to aqueous humor outflow causes sustained elevation in IOP. Glaucomatous insults in the aqueous humor outflow pathway, including the trabecular meshwork (TM), precede such chronic physiological changes in IOP. These insults include ultrastructural changes with excessive extracellular matrix deposition and actin cytoskeletal reorganization that leads to pathological stiffening of the ocular tissues. One of the most common cytoskeletal changes associated with TM tissue stiffness in glaucoma is the increased prevalence of cross-linked actin networks (CLANs) in cells of the trabecular meshwork (TM) and lamina cribrosa (LC). In glaucomatous cells, rearrangement of linear actin stress fibers leads to formation of polygonal arrays within the cytoplasm, resembling a geodesic dome-like structure, that we identified as CLANs. In addition to increased amounts of CLANs in POAG TM cells and tissues, we also discovered that glucocorticoid (GC) and TGFβ2 signaling pathways associated with the development of ocular hypertension (OHT) and glaucoma also induced CLANs in the TM. Despite a clear association, we are yet to completely understand the mechanisms involved in CLAN formation and their direct relevance to disease pathology. In this chapter, we will describe methods to identify and characterize CLANs using fluorescent microscopy in primary TM cell cultures, ex vivo perfusion cultured human anterior segments, and in situ in human donor eyes.

Human Anterior Segment Perfusion Organ Culture.

The human anterior segment perfusion organ culture is an ex vivo model system for studying the human conventional outflow pathway with reference to pressure regulation. In this model, anterior segments dissected from human donor eyes can be fixed to a modified petri dish and perfused with media along with various study agents at the physiological flow rate of 2.5μL/min. The model mimics the one-way flow of aqueous humor in human eyes and can be used to evaluate the effects of various drugs on eye pressure in real time. Using this model, cells and tissues of the anterior segment can be maintained for up to 28days, enabling histological and molecular evaluations.

Green-Light-Triggered and Self-Calibrated Cascade Release of Nitric Oxide and Carbon Monoxide for Synergistic Glaucoma Therapy.

Glaucoma is an optic degenerative neuropathy that is driven by a vicious cycle of oxidative stress and mechanical stress. Current clinical treatments aim exclusively at alleviating mechanical stress by reducing the intraocular pressure (IOP). With the unattended oxidative stress, recurrence and deterioration of mechanical stress are inevitable. Nitric oxide (NO) and carbon monoxide (CO) are endogenous gaseous signaling molecules for vasodilation and anti-inflammation, respectively. Mounting evidence suggests an intricate interplay between NO and CO to mediate their biological roles, like how it takes two to dance a waltz. This leads to the concept of "gas waltz therapy" for glaucoma, in which NO is released to reduce IOP and stoichiometric CO is coreleased to suppress oxidative stress. CND570 is the first phototriggered cascade NO/CO donor, to the best of our knowledge. Notably, the release of NO/CO is accompanied by the concomitant release of a rhodamine dye whose bright fluorescence is harnessed as a convenient calibration mechanism of the gas release profile. CND570 exhibits excellent transcorneal permeability and reaches the target aqueous humor outflow pathway. Further, green-light irradiation triggers release of CO and NO in the eye tissue of glaucoma mice. NO and CO could promote the upregulation of soluble guanylate cyclase (sGC) in both in vitro and in vivo models. Notably, CND570 treatment significantly reduces the oxidative stress associated with glaucoma. NO/CO-based gas waltz therapy is a promising new avenue for glaucoma treatment.

Gene therapy for glaucoma: Targeting key mechanisms

Glaucoma is a group of optic neuropathies characterised by progressive retinal ganglion cell (RGC) degeneration and is the leading cause of irreversible blindness worldwide. Current treatments for glaucoma focus on reducing intraocular pressure (IOP) with topical medications. However, many patients do not achieve sufficient IOP reductions with such treatments. Patient compliance to dosing schedules also poses a significant challenge, further limiting their effectiveness. While surgical options exist for resistant cases, these are invasive and carry risks of complications. Thus, there is a critical need for better strategies to prevent irreversible vision loss in glaucoma. Gene therapy holds significant promise in this regard, offering potential long-term solutions by targeting the disease’s underlying causes at a molecular level. Gene therapy strategies for glaucoma primarily target the two key hallmarks of the disease: elevated IOP and RGC death. This review explores key mechanisms underlying these hallmarks and discusses the current state of gene therapies targeting them. In terms of IOP reduction, this review covers strategies aimed at enhancing extracellular matrix turnover in the conventional outflow pathway, targeting fibrosis, regulating aqueous humor production, and targeting myocilin for gene-specific therapy. Neuroprotective strategies explored include targeting neurotrophic factors and their receptors, reducing oxidative stress and mitochondrial dysfunction, and preventing Wallerian degeneration. This review also briefly highlights key research priorities for advancing gene therapies for glaucoma through the clinical pipeline, such as refining delivery vectors and improving transgene regulation. Addressing these priorities will be essential for translating advancements from preclinical models into effective clinical therapies for glaucoma.

Schlemm's canal-selective Tie2/TEK knockdown induces sustained ocular hypertension in adult mice

Deficient Angiopoietin-Tie2 signaling is linked to ocular hypertension in glaucoma. Receptor Tie2/TEK expression and signaling at Schlemm's canal (SC) is indispensable for canal integrity and homeostatic regulation of aqueous humor outflow (AHO) and intraocular pressure (IOP), as validated by conditional deletion of Tie2, its ligands (Angpt1, Angpt2 and Angpt3/4) or regulators (Tie1 and PTPRB/VE-PTP). However, these Tie2/TEK knockouts and conditional knockouts are global or endothelial, preventing separation of systemic and ocular vascular defects that impact retinal or renal integrity. To develop a more targeted model of ocular hypertension induced by selective knockdown of Tie2/TEK expressed in SC, we combined the use of viral vectors to target the canal, and two distinct gene-editing strategies to disrupt the Tie2 gene. Adeno-associated virus (AAV2) is known to transduce rodent SC when delivered into the anterior chamber by intracameral injection. First, delivery of Cre recombinase via AAV2.Cre into R26tdTomato/+ reporter mice confirmed preferential and stable transduction in SC endothelium. Next, to disrupt Tie2 expression in SC, we injected AAV2.Cre into homozygous floxed Tie2 (Tie2FL/FL) mice. This led to attenuated Tie2 protein expression along the SC inner wall, decreased SC area and reduced trabecular meshwork (TM) cellularity. Functionally, IOP was significantly and steadily elevated, whereas AHO facility was reduced. In contrast, hemizygous Tie2FL/+ mice responded to AAV2.Cre with inconsistent and low IOP elevation, corroborating the dose-dependency of ocular hypertension on Tie2 expression/activation. In a second model using CRISPR/SaCas9 genome editing, wild-type C57BL/6 J mice injected with AAV2.saCas9-sgTie2 showed similar selective SC transduction and comparable IOP elevation in course and magnitude to that induced by AAV2.Cre in Tie2FL/FL mice. Together, our findings, demonstrate that selective Tie2 knockdown in SC is a targeted strategy that reliably induces chronic ocular hypertension and reproduces glaucomatous damage to the conventional outflow pathway, providing novel models of SC-Tie2 signaling loss valuable for preclinical studies.

Effects of Genipin Crosslinking of Porcine Perilimbal Sclera on Mechanical Properties and Intraocular Pressure.

The mechanical properties of sclera play an important role in ocular functions, protection, and disease. Modulating the sclera's properties by exogenous crosslinking offers a way to expand the tissue's range of properties for study of the possible influences on the eye's behavior and diseases such as glaucoma and myopia. The focus of this work was to evaluate the effects of genipin crosslinking targeting the porcine perilimbal sclera (PLS) since the stiffness of this tissue was previously found in a number of studies to influence the eye's intraocular pressure (IOP). The work includes experiments on tensile test specimens and whole globes. The specimen tests showed decreased strain-rate dependence and increased relaxation stress due to the cross-linker. Whole globe perfusion experiments demonstrated that eyes treated with genipin in the perilimbal region had increased IOPs compared to the control globes. Migration of the cross-linker from the target tissue to other tissues is a confounding factor in whole globe, biomechanical measurements, with crosslinking. A novel quantitative genipin assay of the trabecular meshwork (TM) was developed to exclude globes where the TM was inadvertently crosslinked. The perfusion study, therefore, suggests that elevated stiffness of the PLS can significantly increase IOP apart from effects of the TM in the porcine eye. These results demonstrate the importance of PLS biomechanics in aqueous outflow regulation and support additional investigations into the distal outflow pathways as a key source of outflow resistance.

Robotic Visible-Light Optical Coherence Tomography Visualizes Segmental Schlemm's Canal Anatomy and Segmental Pilocarpine Response.

To use robotic visible-light OCT (vis-OCT) to study circumferential segmental Schlemm's canal (SC) anatomy in mice after topical pilocarpine administration. Anterior segment imaging was performed using a vis-OCT sample arm attached to a 6-degree-of-freedom robotic arm to maintain normal (perpendicular) laser illumination aimed at SC around the limbus. Sixteen mice were studied for repeatability testing and to study aqueous humor outflow (AHO) pathway response to topical drug. Pharmaceutical-grade pilocarpine (1%; n = 5) or control artificial tears (n = 9) were given, and vis-OCT imaging was performed before and 15 minutes after drug application. After SC segmentation, SC areas and volumes were measured circumferentially in control- and drug-treated eyes. Circumferential vis-OCT provided high-resolution imaging of the anterior segment and AHO pathways, including SC. Segmental SC anatomy was visualized with the average cross-sectional area greatest temporal (3971 ± 328 μm2) and the least nasal (2727 ± 218 μm2; p = 0.018). After pilocarpine administration, the iris became flatter, and SC became larger (pilocarpine: 26.8 ± 5.0% vs. control: 8.9 ± 4.6% volume increase; p = 0.030). However, the pilocarpine alteration was segmental as well, with a greater increase observed superior (pilocarpine: 31.6 ± 8.9% vs. control: 1.8 ± 5.7% volume increase; p = 0.023) and nasal (pilocarpine: 41.1 ± 15.3% vs. control: 13.9 ± 4.5% volume increase; p = 0.045). High-resolution circumferential non-invasive imaging using AS-OCT of AHO pathways is possible in living animals with robotic control. Segmental SC anatomy was seen at baseline and was consistent with the known segmental nature of trabecular AHO. Segmental SC anatomical response to a muscarinic agonist was seen as well. Segmental glaucoma drug response around the circumference of AHO pathways is a novel observation that may explain the variable patient response to glaucoma treatments.