This work is funded by National Health and Medical Research Council of Australia project grant (1046203), Australian Research Council Future Fellowship (FT130100338).

All experimental procedures were conducted according to the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes, set by the National Health and Medical Research Council in Australia. Ethics approval was obtained from the Howard Florey Institute Animal Ethics Committee (approval number 13-044-UM and 13-068-UM for rats and mice, respectively).
1. Intraocular Pressure Measurement in Conscious Rats
<ol>
	<li>Set the laboratory rebound tonometer to the rat setting. Swaddle the awake rat in a piece of soft cloth to calm the animal. Expose the head and neck. Gently hold the torso in one hand, with the animal&#39;s back resting against the investigator&#39;s chest. 
	NOTE: Topical anesthesia is not required.</li>
	<li>Use the other hand to bring the rebound tonometer near the rat&#39;s eye, so that the tip of the IOP probe is approximately 2 - 3 mm away from and perpendicular to the corneal apex. Use the right hand to measure IOP in the animal&#39;s right eye, and left hand for the left eye.</li>
	<li>Wait a few seconds for the rat to calm and press the measurement button once. Observe the tip of the IOP probe gently hit the corneal apex once; and hear the rebound tonometer beep once. 
	NOTE: A single beep of the tonometer confirms successful measurement, which can be read from the LCD screen. A double beep indicates a measurement error. Measurement errors can arise from factors such as inappropriate working distance between the probe and the cornea, an excessive tilt in the orientation of the tonometer, or the probe striking the eyelid or a non-central part of the cornea. Refer to the rebound tonometer manual from the manufacturer for further detail regarding measurement errors.</li>
	<li>Repeat step 1.3 ten times at an interval of 1 - 2 second, from these measurements derive an average IOP value for that time point. Reset the tonometer after the 5th reading.</li>
	<li>For serial monitoring, measure IOP at the same time of the day and under consistent lighting conditions to minimize variation due to the diurnal IOP cycle20,21.</li>
</ol>
2. Intraocular Pressure Measurement in Conscious Mice
<ol>
	<li>Set the rebound tonometer to the mouse setting according to manufacturer&#39;s instruction.</li>
	<li>To restrain the mouse by hand, place the mouse on a grill cage top and gently pull the tail backwards. 
	&#8203;NOTE: This will prompt the animal to grip onto the metal grill with its front legs and attempt to pull itself forward, which will slightly stretch its body.
	<ol>
		<li>Use the other hand to grasp the loose skin immediately behind the ears. Secure the lower body of the animal by holding the tail between the ring finger and middle finger (or between the little finger and your palm). 
		NOTE: Try not to grasp the skin too tight, to avoid suffocation and applying pressure on the eyes.</li>
	</ol>
	</li>
	<li>With the now free hand (initially holding the tail), bring the rebound tonometer near the mouse&#39;s eye, so that the tip of the IOP probe is approximately 2 - 3 mm from and perpendicular to the corneal apex. To measure the other eye, rotate the mouse so that the other eye is now in front of the tonometer.</li>
	<li>Wait for the mouse to calm and press the measurement button once. Observe the tip of the IOP probe gently hit the corneal apex; with a single beep confirming successful measurement. 
	NOTE: A double beep indicates a measurement error. It may help to have a second experimenter read and document the IOP readings whilst the first experimenter takes the measurements.</li>
	<li>Repeat step 2.4 to obtain ten successful readings to derive an IOP. Reset the tonometer after the 5th reading. Allow an interval of 1 - 2 seconds between readings.</li>
	<li>As per serial measurement in rats, measure mouse IOP at the same time of the day and under consistent lighting conditions.</li>
</ol>
3. Induction of Intraocular Pressure Elevation in Anesthetized Rats and Mice
<ol>
	<li>Clean the surgical bench with 0.5% chlorhexidine in 70% ethanol. Cover the bench with sterile drapes. Autoclave all surgical equipment beforehand. Ensure all experimenters wear appropriate personal protective equipment (surgical masks, gowns and sterilized gloves).</li>
	<li>To induce general anesthesia, place the animal in an induction chamber. Deliver 3 - 3.5% isoflurane with O2 at a flow rate of 3 L/min.
	<ol>
		<li>Maintain anesthesia with 1.5% isoflurane at 2 L/min delivered via a rodent face mask throughout the surgery. Ensure sufficient depth of anesthesia by the absence of a paw pinch reflex.</li>
		<li>Avoid respiratory depression by adjusting the flow rate when necessary to maintain the respiratory rate at approximately 60 breaths/min.</li>
	</ol>
	</li>
	<li>Randomly select one eye to induce ocular hypertension, with the contralateral eye to serve as an untreated control. Instill one drop of 0.5% proxymetacaine ophthalmic solution for topical anesthesia. To clean the ocular surface, rinse the eye with 3 mL of sterile normal saline.</li>
	<li>Cover the animal with a sterile, fenestrated surgical drape, exposing the eye to be sutured.</li>
	<li>Perform a purse-string suture on the bulbar conjunctiva around the globe. In rats, weave the 7/0 nylon suture parallel and 2 mm posterior to the limbus (Figure 1). In mice, place the 10/0 nylon suture at 1 mm posterior to the limbus.
	<ol>
		<li>Take care not to penetrate the sclera. A sudden pupillary dilation during the surgical procedure indicates the sclera has likely been penetrated.</li>
		<li>Anchor the suture on the conjunctiva using 5-6 anchor points in rats, and 4-5 anchor points in mice.</li>
		<li>Avoid direct compression on the major episcleral veins by threading the suture underneath the conjunctiva at the crossing of these veins. 
		NOTE: While we recommend avoiding compression of the major episcleral vein in rats, this is not routinely done in mice due to low visibility of these veins in mouse eyes. Even though the major veins are not directly compressed, it is likely that the smaller vessels in the episcleral vein plexus are under pressure, which may be a contributing factor to the sustained IOP elevation (please see Discussion for mechanism of IOP elevation).</li>
	</ol>
	</li>
	<li>Fasten the purse-string suture by tying a slipknot then followed by a second simple knot (Figure 1). To avoid an excessively high post-surgical IOP spike, have an assistant measure the IOP immediately before fastening the second knot.
	<ol>
		<li>If the IOP is found to be too high, adjust the slip knot by partially releasing the tension on one end of the suture (arrow in Figure 1A).</li>
		<li>After the desired IOP is achieved (ideally 30 - 60 mmHg in rats or 30 - 40 mmHg in mice), tie off the second knot while maintaining a continuous pulling force on that end of the suture (arrow in Figure 1A).</li>
		<li>After the second knot has been tightened, trim the ends of the suture to minimize any foreign body sensation. Monitor the animal during recover from general anesthesia. 
		NOTE: It is important to use the slipknot when tying the first knot to ensure adequate inward compression on the eye. After several weeks it is usually noted that the ends become embedded in the conjunctiva.</li>
	</ol>
	</li>
</ol>
4. Monitoring IOP
<ol>
	<li>Take the first IOP measurement at 2 minutes post-operatively under isoflurane anesthesia. Subsequently, monitor IOP when the rodent has regained consciousness as per the aforementioned steps 1 and 2. 
	NOTE: Monitor the IOP twice during the first day (2 minutes and 1 hour), daily in the first week and once or twice per week thereafter.</li>
</ol>
5. Assaying Retinal Structure and Function
<ol>
	<li>At the desired experimental end point (in this case after 8 weeks in rats and 12 weeks in mice), under general anesthesia using intraperitoneal injection with ketamine/xylazine, measure retinal function with the dark-adapted electroretinogram (ERG) as described in greater detail elsewhere15,16,17. 
	NOTE: We have found robust ganglion cell dysfunction, retinal nerve fibre layer thinning and ganglion cell loss for durations between 8-12 weeks. Others have successfully employed longer periods of IOP elevation14,15.</li>
	<li>Immediately after ERG measurement, measure the thickness of retinal nerve fiber layer (RNFL) and total retinal thickness using spectral domain optic coherence tomography (SD-OCT) 16,18.</li>
	<li>At the end of the longitudinal study, euthanize the animals under deep anesthesia.
	<ol>
		<li>Dissect the retina for histology18, for example immunostaining of whole-mount retina using a retinal ganglion cell (RGC) specific antibody such as RNA-binding protein with multiple splicing antibody (RBPMS) or brain-specific homeobox/POU domain protein 3A (Brn3a)16,19,22.</li>
	</ol>
	</li>
</ol>

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The authors have nothing to disclose.

The circumlimbal suture is a new model of chronic ocular hypertension. In addition to the studies from which the representative results are sourced16,18, this animal model has been utilized in a number of recent studies15,23,24,25,26. Comparison across these previous reports shows that the method produ...

Animal models provide an important platform for laboratory investigation of cellular processes underlying glaucoma pathogenesis, as well as to evaluate potential therapeutic interventions. Several inducible models have been developed to produce sustained intraocular pressure (IOP) elevation, the most important risk factor for glaucoma. Methods that have been applied to elevate IOP include: hypertonic saline injection in episcleral veins1, laser photocoagulation of the trabecular meshwork2 or of the limbal veins3, and intracameral injection of substances such as ghost red blood cells4, microbeads5,6 and viscoelastic agents7. Each approach has its advantages and limitations.
A good model for glaucoma should mimic the disease process, with minimal complication such as trauma, inflammation and media opacities. These complications are frequently associated with the procedures used to induce IOP elevation, and can confound interpretation of outcomes. For example, paracentesis of the anterior chamber, even when foreign substances are not introduced, has been shown to cause trauma and inflammation that is not representative of typical glaucomatous change8,9. In addition to the importance of avoiding inflammation, maintaining optical clarity facilitates in vivo imaging and electrophysiology to monitor disease progression. Although it is unclear to what extent these complications may affect disease investigations, it may be better to avoid penetrating the eye during model induction. The circumlimbal suture approach avoids penetration of the globe and facilitates in vivo longitudinal assessment of retinal structure and function. More importantly, this model differs from previous ones in its capacity to return IOP to baseline values by removal of the suture when required. IOP normalization may be useful for studying the cellular and molecular correlates of reversible and irreversible ganglion cell injury10,11,12,13,14.
This article focuses on the technique for model induction. Characterization of retinal injury&#160;induced by this model in rats and mice can be found in greater detail elsewhere15,16,17,18,19.

<table><tbody><tr><td>normal saline</td><td>Baxter International Inc</td><td>AHB1323</td><td>Maintain corneal hydration during surgery</td></tr><tr><td>Chlorhexadine 0.5%</td><td>Orion Laboratories</td><td>27411, 80085</td><td>Disinfection of surgical instrument</td></tr><tr><td>Isoflurane 99.9%</td><td>Abbott Australasia Pty Ltd</td><td>CAS 26675-46-7</td><td>Proprietory Name: Isoflo(TM) Inhalation anaaesthetic. Pharmaceutical-grade inhalation anesthetic mixed with oxygen gas for suture procedure</td></tr><tr><td>ocular lubricant</td><td>Alcon Laboratories&amp;nbsp;</td><td>1618611</td><td>Proprietory Name: Genteal, ocular lubricant to keep the other eye moist</td></tr><tr><td>Needle holder (microsurgery)</td><td>World Precision Instruments</td><td>555419NT</td><td>To hold needle during ocular surgery</td></tr><tr><td>Proxymetacaine 0.5%</td><td>Alcon Laboratories&amp;nbsp;</td><td>CAS 5875-06-9</td><td>Topical ocular analgesia</td></tr><tr><td>Scissors (microsurgery)</td><td>World Precision Instruments</td><td>501232</td><td>To cut excessive suture stump during ligation</td></tr><tr><td>Surgical drape</td><td>Vital Medical Supplies</td><td>GM29-612EE</td><td>Ensure sterile enviornment during surgery</td></tr><tr><td>Suture needle for rats (microsurgery)</td><td>Ninbo medical needles</td><td>151109</td><td>8-0 nylon suture attached with round needle, cutting edge 3/8, dual-needle, suture length 30cm</td></tr><tr><td>Suture needle for mice (microsurgery)</td><td>Ninbo medical needles</td><td>160905</td><td>10-0 nylon suture attached with round needle, cutting edge 3/8, dual-needle, suture length 30cm</td></tr><tr><td>Tweezers (microsurgery)</td><td>World Precision Instruments</td><td>500342</td><td>Manipulate tissues during ocular surgery</td></tr><tr><td>rebound tonometer</td><td>TONOLAB, iCare, Helsinki, Finland</td><td>TV02</td><td>for intraocular pressure monitoring</td></tr></tbody></table>

a model of glaucoma induced by circumlimbal suture in rats and mice

The circumlimbal suture is a technique for inducing experimental glaucoma in rodents by chronically elevating intraocular pressure (IOP), a well-known risk factor for glaucoma. This protocol demonstrates a step-by-step guide on this technique in Long Evans rats and C57BL/6 mice. Under general anesthesia, a "purse-string" suture is applied on the conjunctiva, around the equator and behind the limbus of the eye. The fellow eye serves as an untreated control. Over the duration of our study, which was a period of 8 weeks for rats and 12 weeks for mice, IOP remained elevated, as measured regularly by rebound tonometry in conscious animals without topical anesthesia. In both species, the sutured eyes showed electroretinogram features consistent with preferential inner retinal dysfunction. Optical coherence tomography showed selective thinning of the retinal nerve fiber layer. Histology of the rat retina in cross-section found reduced cell density in the ganglion cell layer, but no change in other cellular layers. Staining of flat-mounted mouse retinae with a ganglion cell specific marker (RBPMS) confirmed ganglion cell loss. The circumlimbal suture is a simple, minimally invasive and cost-effective way to induce ocular hypertension that leads to ganglion cell injury in both rats and mice.

The following results in rats18 and mice16 have been previously reported and are summarized here. The circumlimbal suture produced a similar pattern of IOP elevation in rats and mice (Figure 2). A brief IOP spike, up to 58.1 &#177; 2.7 mmHg in rats and 38.7 &#177; 2.2 mmHg in mice, was found immediately after the suture procedure. In rats, IOP magnitude gradually reduced over time to be 44 &#177; 6 mmHg and 32 &...

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A Model of Glaucoma Induced by Circumlimbal Suture in Rats and Mice

Chronic ocular hypertension is induced by applying a circumlimbal suture in rats and mice, leading to functional and structural deterioration of the retinal ganglion cells consistent with glaucoma.

The circumlimbal suture is a technique for inducing experimental glaucoma in rodents by chronically elevating intraocular pressure (IOP), a well-known ...

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10.3K Views.  University of Melbourne. Chronic ocular hypertension is induced by applying a circumlimbal suture in rats and mice, leading to functional and structural deterioration of the retinal ganglion cells consistent with glaucoma.

Video: A Model of Glaucoma Induced by Circumlimbal Suture in Rats and Mice

A Laser-induced Mouse Model of Chronic Ocular Hypertension to Characterize Visual Defects

Glaucoma, frequently associated with elevated intraocular pressure (IOP), is one of the leading causes of blindness. We sought to establish a mouse model of ocular hypertension to mimic human high-tension glaucoma. Here laser illumination is applied to the corneal limbus to photocoagulate the aqueous outflow, inducing angle closure. The changes of IOP are monitored using a rebound tonometer before and after the laser treatment. An optomotor behavioral test is used to measure corresponding changes in visual capacity. The representative result from one mouse which developed sustained IOP elevation after laser illumination is shown. A decreased visual acuity and contrast sensitivity is observed in this ocular hypertensive mouse. Together, our study introduces a valuable model system to investigate neuronal degeneration and the underlying molecular mechanisms in glaucomatous mice.

Chronic ocular hypertension is induced using laser photocoagulation of the trabecular meshwork in mouse eyes. The intraocular pressure (IOP) is elevated for several months after laser treatment. The decrease of visual acuity and contrast sensitivity of experimental animals are monitored using the optomotor test.

Glaucoma, frequently associated with elevated intraocular pressure (IOP), is one of the leading causes of blindness. We sought to establish a mouse model ...

Experimental Glaucoma Induced by Ocular Injection of Magnetic Microspheres

Progress in understanding the pathophysiology, and providing novel treatments for glaucoma is dependent on good animal models of the disease. We present here a protocol for elevating intraocular pressure (IOP) in the rat, by injecting magnetic microspheres into the anterior chamber of the eye. The use of magnetic particles allows the user to manipulate the beads into the iridocorneal angle, thus providing a very effective blockade of fluid outflow from the trabecular meshwork. This leads to long-lasting IOP rises, and eventually neuronal death in the ganglion cell layer (GCL) as well as optic nerve pathology, as seen in patients with the disease. This method is simple to perform, as it does not require machinery, specialist surgical skills, or many hours of practice to perfect. Furthermore, the pressure elevations are very robust, and reinjection of the magnetic microspheres is not usually required unlike in some other models using plastic beads. Additionally, we believe this method is suitable for adaptation for the mouse eye.

We present a method for inducing elevated intraocular pressure (IOP), by injecting magnetic microspheres into the rat eye, to model glaucoma. This leads to strong pressure rises, and extensive neuronal death. This protocol is easy to perform, does not require repeat injections, and produces stable long-lasting IOP rises.

Progress in understanding the pathophysiology, and providing novel treatments for glaucoma is dependent on good animal models of the disease. We present ...

A Magnetic Microbead Occlusion Model to Induce Ocular Hypertension-Dependent Glaucoma in Mice

The use of rodent models of glaucoma has been essential to understand the molecular mechanisms that underlie the pathophysiology of this multifactorial neurodegenerative disease. With the advent of numerous transgenic mouse lines, there is increasing interest in inducible murine models of ocular hypertension. Here, we present an occlusion model of glaucoma based on the injection of magnetic microbeads into the anterior chamber of the eye using a modified microneedle with a facetted bevel. The magnetic microbeads are attracted to the iridocorneal angle using a handheld magnet to block the drainage of aqueous humour from the anterior chamber. This disruption in aqueous dynamics results in a steady elevation of intraocular pressure, which subsequently leads to the loss of retinal ganglion cells, as observed in human glaucoma patients. The microbead occlusion model presented in this manuscript is simple compared to other inducible models of glaucoma and also highly effective and reproducible. Importantly, the modifications presented here minimize common issues that often arise in occlusion models. First, the use of a bevelled glass microneedle prevents backflow of microbeads and ensures that minimal damage occurs to the cornea during the injection, thus reducing injury-related effects. Second, the use of magnetic microbeads ensures the ability to attract most beads to the iridocorneal angle, effectively reducing the number of beads floating in the anterior chamber avoiding contact with other structures (e.g., iris, lens). Lastly, the use of a handheld magnet allows flexibility when handling the small mouse eye to efficiently direct the magnetic microbeads and ensure that there is little reflux of the microbeads from the eye when the microneedle is withdrawn. In summary, the microbead occlusion mouse model presented here is a powerful investigative tool to study neurodegenerative changes that occur during the onset and progression of glaucoma.

Here, we present a protocol to induce ocular hypertension in the murine eye that results in the loss of retinal ganglion cells as observed in glaucoma. Magnetic microbeads are injected into the anterior chamber and attracted to the iridocorneal angle using a magnet to block the outflow of aqueous humour.

The use of rodent models of glaucoma has been essential to understand the molecular mechanisms that underlie the pathophysiology of this multifactorial ...

Glaucoma-inducing Procedure in an In Vivo Rat Model and Whole-mount Retina Preparation

Glaucoma is a disease of the central nervous system affecting retinal ganglion cells (RGCs). RGC axons making up the optic nerve carry visual input to the brain for visual perception. Damage to RGCs and their axons leads to vision loss and/or blindness. Although the specific cause of glaucoma is unknown, the primary risk factor for the disease is an elevated intraocular pressure. Glaucoma-inducing procedures in animal models are a valuable tool to researchers studying the mechanism of RGC death. Such information can lead to the development of effective neuroprotective treatments that could aid in the prevention of vision loss. The protocol in this paper describes a method of inducing glaucoma - like conditions in an in vivo rat model where 50 &#181;l of 2 M hypertonic saline is injected into the episcleral venous plexus. Blanching of the vessels indicates successful injection. This procedure causes loss of RGCs to simulate glaucoma. One month following injection, animals are sacrificed and eyes are removed. Next, the cornea, lens, and vitreous are removed to make an eyecup. The retina is then peeled from the back of the eye and pinned onto sylgard dishes using cactus needles. At this point, neurons in the retina can be stained for analysis. Results from this lab show that approximately 25% of RGCs are lost within one month of the procedure when compared to internal controls. This procedure allows for quantitative analysis of retinal ganglion cell death in an in vivo rat glaucoma model.

Glaucoma-inducing Procedure in an In Vivo Rat Model and Whole-mount Retina Preparation

Glaucoma is characterized by damage to retinal ganglion cells. Inducing glaucoma in animal models can provide insight into the study of this disease. Here, we outline a procedure that induces loss of RGCs in an in vivo rat model and demonstrates the preparation of whole-mount retinas for analysis.

Glaucoma is a disease of the central nervous system affecting retinal ganglion cells (RGCs). RGC axons making up the optic nerve carry visual input to the ...

In vivo Structural Assessments of Ocular Disease in Rodent Models using Optical Coherence Tomography

Spectral-domain optical coherence tomography (SD-OCT) is useful for visualizing retinal and ocular structures in vivo. In research, SD-OCT is a valuable tool to evaluate and characterize changes in a variety of retinal and ocular disease and injury models. In light induced retinal degeneration models, SD-OCT can be used to track thinning of the photoreceptor layer over time. In glaucoma models, SD-OCT can be used to monitor decreased retinal nerve fiber layer and total retinal thickness and to observe optic nerve cupping after inducing ocular hypertension. In diabetic rodents, SD-OCT has helped researchers observe decreased total retinal thickness as well as decreased thickness of specific retinal layers, particularly the retinal nerve fiber layer with disease progression. In mouse models of myopia, SD-OCT can be used to evaluate axial parameters, such as axial length changes. Advantages of SD-OCT include in vivo imaging of ocular structures, the ability to quantitatively track changes in ocular dimensions over time, and its rapid scanning speed and high resolution. Here, we detail the methods of SD-OCT and show examples of its use in our laboratory in models of retinal degeneration, glaucoma, diabetic retinopathy, and myopia. Methods include anesthesia, SD-OCT imaging, and processing of the images for thickness measurements.

Here, we describe the use of spectral-domain optical coherence tomography (SD-OCT) to visualize retinal and ocular structures in vivo in models of retinal degeneration, glaucoma, diabetic retinopathy, and myopia.

Spectral-domain optical coherence tomography (SD-OCT) is useful for visualizing retinal and ocular structures in vivo. In research, SD-OCT is a valuable ...

Bioengineering

A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice

Elevated intraocular pressure (IOP) is a well-documented risk factor for glaucoma. Here we describe a novel, effective method for consistently inducing stable IOP elevation in mice that mimics the post-operative complication of using silicone oil (SO) as a tamponade agent in human vitreoretinal surgery. In this protocol, SO is injected into the anterior chamber of the mouse eye to block the pupil and prevent inflow of aqueous humor. The posterior chamber accumulates aqueous humor and this in turn increases the IOP of the posterior segment. A single SO injection produces reliable, sufficient, and stable IOP elevation, which induces significant glaucomatous neurodegeneration. This model is a true replicate of secondary glaucoma in the eye clinic. To further mimic the clinical setting, SO can be removed from the anterior chamber to reopen the drainage pathway and allow inflow of aqueous humor, which is drained through the trabecular meshwork (TM) at the angle of the anterior chamber. Because IOP quickly returns to normal, the model can be used to test the effect of lowering IOP on glaucomatous retinal ganglion cells. This method is straightforward, does not require special equipment or repeat procedures, closely simulates clinical situations, and may be applicable to diverse animal species. However, minor modifications may be required.

Here, we present a protocol to induce ocular hypertension and glaucomatous neurodegeneration in mouse eyes by intracameral injection of silicone oil and the procedure for silicone oil removal from the anterior chamber to return elevated intraocular pressure to normal.

Elevated intraocular pressure (IOP) is a well-documented risk factor for glaucoma. Here we describe a novel, effective method for consistently inducing ...

Full-Circle Cauterization of Limbal Vascular Plexus for Surgically Induced Glaucoma in Rodents

Glaucoma, the second leading cause of blindness worldwide, is a heterogeneous group of ocular disorders characterized by structural damage to the optic nerve and retinal ganglion cell (RGC) degeneration, resulting in visual dysfunction by interrupting the transmission of visual information from the eye to the brain. Elevated intraocular pressure is the most important risk factor; thus, several models of ocular hypertension have been developed in rodents by either genetic or experimental approaches to investigate the causes and effects of the disease. Among those, some limitations have been reported such as surgical invasiveness, inadequate functional assessment, requirement of extensive training, and highly variable extension of retinal damage. The present work characterizes a simple, low-cost, and efficient method to induce ocular hypertension in rodents, based on low-temperature, full-circle cauterization of the limbal vascular plexus, a major component of aqueous humor drainage. The new model provides a technically easy, noninvasive, and reproducible subacute ocular hypertension, associated with progressive RGC and optic nerve degeneration, and a unique post-operative clinical recovery rate that allows in vivo functional studies by both electrophysiological and behavioral methods.

The goal of this protocol is to characterize a novel model of glaucomatous neurodegeneration based on 360&#176; thermic cauterization of limbal vascular plexus, inducing subacute ocular hypertension.

Glaucoma, the second leading cause of blindness worldwide, is a heterogeneous group of ocular disorders characterized by structural damage to the optic ...

Neuroscience

Image-Guided Optical Coherence Tomography to Assess Structural Changes in Rodent Retinas

Ocular diseases, such as age-related macular degeneration, glaucoma, retinitis pigmentosa, and uveitis, are always accompanied by retinal structural changes. These diseases affecting the fundus always exhibit typical abnormalities in certain cell types in the retina, including photoreceptor cells, retinal ganglion cells, cells in the retinal blood vessels, and cells in the choroidal vascular cells. Noninvasive, highly efficient, and adaptable imaging techniques are required for both clinical practice and basic research. Image-guided optical coherence tomography (OCT) satisfies these requirements because it combines fundus photography and high-resolution OCT, providing an accurate diagnosis of tiny lesions as well as important changes in the retinal architecture. This study details the procedures of data collection and data analysis for image-guided OCT and demonstrates its application in rodent models of choroidal neovascularization (CNV), optic nerve crush (ONC), light-induced retinal degeneration, and experimental autoimmune uveitis (EAU). This technique helps researchers in the eye field to identify rodent retinal structural changes conveniently, reliably, and tractably.

The protocol presented here details the procedures of data collection and data analysis for image-guided optical coherence tomography (OCT) and demonstrates its application in multiple rodent models of ocular diseases.

Ocular diseases, such as age-related macular degeneration, glaucoma, retinitis pigmentosa, and uveitis, are always accompanied by retinal structural ...

Replicating Cataract Surgical Techniques in a Mouse Model Through LEC Extraction

Modeling Cataract Surgery in Mice

Cataract surgery (CS) is an effective treatment for cataracts, a major cause of visual disability worldwide. However, CS leads to ocular inflammation, and in the long term, it can result in posterior capsular opacification (PCO) and/or lens dislocation driven by the post-surgical overgrowth of lens epithelial cells (LECs) and their conversion to myofibroblasts and/or aberrant fiber cells. However, the molecular mechanisms by which CS results in inflammation and PCO are still obscure because most in vitro models do not recapitulate the wound healing response of LECs seen in vivo, while traditional animal models of cataract surgery, such as rabbits, do not allow the genetic manipulation of gene expression to test mechanisms. Recently, our laboratory and others have successfully used genetically modified mice to study the molecular mechanisms that drive the induction of proinflammatory signaling and LEC epithelial to mesenchymal transition, leading to new insight into PCO pathogenesis. Here, we report the established protocol for modeling cataract surgery in mice, which allows for robust transcriptional profiling of the response of LECs to lens fiber cell removal via RNAseq, the evaluation of protein expression by semi-quantitative immunofluorescence, and the use of modern mouse genetics tools to test the function of genes that are hypothesized to participate in the pathogenesis of acute sequelae like inflammation as well as the later conversion of LECs to myofibroblasts and/or aberrant lens fiber cells.

Author Spotlight: Unraveling the Molecular Mechanisms in PCO and Fibrosis Following Cataract Surgery

This method models cataract surgery in vivo by removing lens fiber cells from adult mice and leaving behind the capsular bag with attached lens epithelial cells (LECs). The injury response is then assessed at various times post-surgery using molecular and morphological criteria.

Cataract surgery (CS) is an effective treatment for cataracts, a major cause of visual disability worldwide. However, CS leads to ocular inflammation, and ...

Biology

 Induction of Elevated Intraocular Pressure in a Rat Model

<a href='https://app.jove.com/v/63442/full-circle-cauterization-limbal-vascular-plexus-for-surgically'>Source: Lani-Louzada, R., et al., Full-Circle Cauterization of Limbal Vascular Plexus for Surgically Induced Glaucoma in Rodents. J. Vis. Exp. (2022)</a>This video demonstrates the induction and monitoring of elevated intraocular pressure (IOP) in a rat model. The process involves cauterizing the limbal blood vessels, resulting in vessel occlusion and elevated IOP. The rat is then allowed to recover, and IOP is regularly monitored in both the experimental and control eyes for comparison.

Source: Lani-Louzada, R., et al., Full-Circle Cauterization of Limbal Vascular Plexus for Surgically Induced Glaucoma in Rodents. J. Vis. Exp. (2022)This ...

A Model of Glaucoma Induced by Circumlimbal Suture in Rats and Mice

Summary

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Chapters in this video

A Laser-induced Mouse Model of Chronic Ocular Hypertension to Characterize Visual Defects

Experimental Glaucoma Induced by Ocular Injection of Magnetic Microspheres

A Magnetic Microbead Occlusion Model to Induce Ocular Hypertension-Dependent Glaucoma in Mice

Glaucoma-inducing Procedure in an In Vivo Rat Model and Whole-mount Retina Preparation

In vivo Structural Assessments of Ocular Disease in Rodent Models using Optical Coherence Tomography

A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice

Full-Circle Cauterization of Limbal Vascular Plexus for Surgically Induced Glaucoma in Rodents

Image-Guided Optical Coherence Tomography to Assess Structural Changes in Rodent Retinas

Modeling Cataract Surgery in Mice

Induction of Elevated Intraocular Pressure in a Rat Model