The research reported in this publication was supported by the National Center for Advancing Translational Sciences of the National Institutes of Health under Award Numbers UL1TR002538 and KL2TR002539 and by Transformational Grant 19TPA34910194 from the American Heart Association.

This protocol is approved by the Institutional Animal Care and Use Committee (University of Utah IACUC Protocol Number 21-09021). Mature New Zealand White rabbits are obtained from commercial vendors.
1. Animal acquisition
<ol>
	<li>Acclimate the animals for the required duration after arrival according to institutional protocol, housing animals socially in a vivarium with standard chow diets. The acclimation period at our institution is 2 weeks.</li>
</ol>
2. Anesthesia and monitoring
<ol>
	<li>Induce general endotracheal anesthesia with intramuscular injection of buprenorphine (0.03 mg/kg) followed approximately 30 min later with a ketamine (25-35 mg/kg) and xylazine (3 mg/kg) intramuscular&#160;injection. Maintain anesthesia with 1%-5% isoflurane in oxygen administered via an endotracheal tube. During induction, use 100% FiO2, and then titrate down to the lowest FiO2 that maintains a 100% SpO2. 
	NOTE: Uninterrupted anesthesia is necessary to prevent movement by the animal so that the stroke will be the only disruption of the stroke-induction process. This also prevents spikes in BP that would result from agitation that could arise from inadequate anesthesia. Consistent oxygenation is also important to control for achieving comparable strokes. These measures are all accounted for in the representative results described below.</li>
	<li>Confirm adequate depth of anesthesia by applying noxious stimuli to the toe. Apply veterinary eye ointment on the eyes to prevent dryness.</li>
	<li>Monitor oxygen saturation by a pulse oximeter placed on the ear. Obtain intravenous access with an angiocatheter in an auricular vein. Ensure that it is secured with a suture or adhesive transparent film dressing.&#160;To mitigate vasospasm, place 0.25 inch of transdermal nitroglycerin on the inside of the ear after induction of anesthesia.</li>
	<li>Provide maintenance fluids with normal saline at a rate of 1 cc/kg/h. Place an esophageal temperature probe to monitor body temperature. Maintain normothermia (33-37 &#176;C) as needed with warming blankets placed under the animal.</li>
</ol>
3. Surgical preparation
<ol>
	<li>Place the rabbit in a supine position on a fluoroscopy-compatible operative table. Extend the head as it optimizes positioning for subsequent angiographic views. Rabbits have exquisitely sensitive arteries prone to vasospasm after instrumentation.&#160;</li>
	<li>Remove fur from both the inguinal regions using electric clippers. Next, palpate the bilateral femoral arterial pulses to confirm adequate clearance by trimming bilaterally. Prepare the skin with scrubs of chlorhexidine and alcohol, and then drape the skin in the usual sterile fashion.</li>
	<li>Administer local anesthesia by subcutaneously injecting 2 mL of 1% lidocaine in the bilateral inguinal regions. Make a 5 cm surgical incision with a number 10 blade at the site where lidocaine was injected. Use blunt dissection to expose the neurovascular bundle (Figure 1A). If needed, extend the incision to adequately expose an arterial segment large enough for access.</li>
	<li>Upon isolation of the neurovascular bundle, drip several drops of 1% lidocaine on the artery to prevent vasospasm. Separate the artery gently from the vein and the adjacent nerve using forceps. Identify the artery by the characteristic appearance of its muscular wall compared to the thin walls of the vein. The artery will have brighter blood, while the vein will contain darker blood.</li>
</ol>
4. Arterial access
<ol>
	<li>After the artery has been isolated, pass the right-angle forceps under the vessel. Grasp two vessel loops with the instrument and gently pass them under the artery. Situate one each at the upstream and downstream ends of the exposed vessel.</li>
	<li>Subject the artery to a gentle traction by pulling the vessel loops. At this point, inspect the vessel for any residual tissue, and remove it with gentle dissection (Figure 1B). This increases the chances of successful access.</li>
	<li>Use a 22 G angiocatheter for access. Slightly advance the catheter itself over the inner needle since this often sticks when completely seated and can dislodge the device during access attempts.</li>
	<li>After dissecting the vessel and preparing the angiocatheter, drip lidocaine on the vessel again. The artery will visibly dilate, which increases the chances of successful access and placement of a sheath using the Seldinger technique.</li>
	<li>Apply gentle traction to the downstream vessel loop to engorge the artery by reducing outflow. This also stabilizes the vessel for the access attempt. Slowly advance the needle of the angiocatheter into the middle of the exposed arterial segment (Figure 1C). When a flash of blood is seen in the angiocatheter and the chamber at its hub, advance the catheter over the needle into the arterial lumen.</li>
	<li>If the access attempt is unsuccessful, achieve hemostasis by applying traction on the upstream vessel loop. Flush the angiocatheter with saline and replace it on its introducer needle for additional attempts.</li>
	<li>When the angiocatheter is successfully placed in the vessel to its hub, advance a Cope microwire through the angiocatheter lumen and into the aorta (Figure 1D). Remove the angiocatheter over the wire and replace it with a 5 French slim hydrophilic sheath (Figure 1E).</li>
	<li>Confirm the return of the arterial blood through the sidearm tubing by opening the three-way valve. Flush the sheath with 0.9% saline and lock the valve closed during flushing.</li>
	<li>Secure the sheath hub to the adjacent skin with an additional 3-0 silk suture. Repeat this process for the contralateral femoral artery. For achieving higher efficiencies, two operators can simultaneously work while focusing on one artery each.</li>
</ol>
5. Cervicocerebral angiography and intracranial access
<ol>
	<li>Under fluoroscopic visualization, advance a 4 French glide catheter over a 0.035-inch glidewire inserted through the left femoral sheath. Position the tip of the glide catheter in the proximal left vertebral artery. Remove the wire and flush the catheter with heparinized 0.9% saline.</li>
	<li>Perform angiography by hand injecting the left vertebral artery with iodinated contrast under low magnification to visualize the entire head and neck (Figure 2A). Modulate the injection of the contrast solution by starting with a low-pressure injection that crescendos to visualize the entire vasculature. 
	NOTE: Sufficient injection to visualize the reflux down the right vertebral artery is needed, as this angiographic image will be used for roadmap guidance to select the right vertebral artery efficiently. Gentle injection is needed to minimize vasospasm or more profound injury. Additionally, excessive force or volume can induce transient motion from the animal even under deep anesthesia.</li>
	<li>For the left vertebral injection, inject 50% contrast diluted in normal saline with a gentle crescendo from a 3 cc syringe. Injecting 1-2 cc of the dilute contrast is typically sufficient. Determine the adequate amount of injection by checking the reflux down the right vertebral artery and into the right subclavian artery. During this injection, also note the posterior cerebral and superior cerebellar arteries, one of which will be the target to occlude with the microcatheter.</li>
	<li>Prepare a 2.4 French flow-directed microcatheter with a 0.010-inch microwire. Make a c-shape on the tip of the microwire. Under roadmap guidance, advance the microcatheter inside a 4 French glide catheter through the right femoral sheath and over the wire into the right vertebral artery. Due to the propensity for catheter-induced vasospasm, minimize the device manipulation time and the number of catheter attempts performed.</li>
	<li>Advance the microcatheter through the cervical segment of the right vertebral artery. To best pass the sharp turn from the V2 to V3 segment, advance the microcatheter alone while the microwire is back proximal to its tip. Leading with the microwire at this point will often cause selection of small side branches of the vertebral artery and can be the source of substantial vasospasm.</li>
	<li>After passing the sharp turn from V2 to V3, the microcatheter often passes easily to the proximal basilar artery. At this point, advance the microwire and select the desired posterior cerebral or superior cerebellar arteries. Microcatheter injections are not advised given the fragile nature of the intracranial arteries.</li>
	<li>Advance the microcatheter over the microwire into the target artery. Choose a proximal position as it is typically safest in the posterior to communicate due to its angulation at its origin. A deeper position is feasible in the superior cerebellar artery (Figure 2B).</li>
	<li>Repeat the angiogram by injecting the left vertebral artery catheter with high magnification over the head to confirm occlusion of the target artery (Figure 2B-C). For optimal imaging, inject full strength contrast in the 3 cc syringe. Typically, no more than 1 cc will be required for adequate opacification of all intracranial arteries.</li>
	<li>Gently remove the microwire from the microcatheter under fluoroscopic visualization to confirm a stable position. Place a stopcock on the hub of the microcatheter and close the stopcock to prevent blood loss from retrograde blood flow. Remove the left vertebral catheter to make the left femoral access sheath available.</li>
	<li>During the ensuing occlusion period, acquire intermittent fluoroscopic images to confirm a stable position of the occlusive microcatheter. Results of posterior cerebral artery occlusion periods ranging from 60-240 min have been published previously18.</li>
</ol>
6. Blood pressure measurement and modulation
<ol>
	<li>While one femoral access site is utilized for the occlusive intracranial microcatheter, use the contralateral sheath for BP measurement.</li>
	<li>Record continuous arterial BP readings with a 3 French gauge piezoresistive sensor, placed through a femoral sheath and advanced until the sensor tip is in the lower thoracic aorta. Connect this sensor to data acquisition hardware and visualize measured pressures with its associated software. Observe BP in the pressure visualization window. BP recordings can be exported into a spreadsheet for visualization in the statistics software.</li>
	<li>Alternatively, if mechanical manipulation of BP is desired using a balloon catheter, advance a 4 French 5 mm Fogarty balloon catheter through the available femoral sheath. Position the balloon in the infrarenal aorta. Use the 0.025-inch inner lumen for pressure tracing to continuously monitor BP upstream of the balloon and the 4 French diameter of the balloon for a second BP tracing line to be connected to the sheath for continuous monitoring of BP downstream of the balloon.</li>
</ol>
7. Euthanasia and tissue harvesting
<ol>
	<li>Remove the occlusive microcatheter after 3 h, and then continue arterial BP measurement and modulation for the additional desired period. A standard recovery period of 3 h is used for visualization of a completed infarct on subsequent histology.</li>
	<li>After completing the prescribed occlusion and recovery times, ensure that the animal is in a surgical plane of anesthesia and perform euthanasia (perfusion fixation with phosphate buffered solution,&#160;followed by decapitation after confirmation of absence of cardiac activity). Alternatively, perform perfusion fixation by infusing perfusate through a femoral sheath, and then transecting a jugular vein, the inferior vena cava, or the right atrium. 
	NOTE: Perfusion may be preferable for some postmortem analyses since gene expression or biomarker values may be affected by the solution. Both the techniques have been performed successfully by our group.</li>
	<li>In acute procedures with immediate harvesting of the brain, confirm euthanasia and decapitate the animal. Remove calvarium in a piecemeal fashion with rongeurs, starting at the occipital ridge and working anteriorly until the brain can be harvested intact. Place the brain in formalin or optimal cutting temperature solution and flash freeze, depending on the type of tissue analysis desired.</li>
</ol>

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MDA, GH, and MAJ are consultants for Certus Critical Care, Inc. MDA is a consultant for Johnson &amp; Johnson.

Substantial progress has been made in the management of IS, particularly considering advances in acute intervention and secondary prevention strategies. However, more work can be done to improve care for IS patients. Limited progress in other aspects of IS treatment, particularly in the realm of neuroprotection, likely results from the limitations in pathophysiological understanding of mechanistic processes at the tissue and molecular level. Impactful data from humans is unrealistic and likely impossible to acquire. In s...

Ischemic stroke (IS) is a leading cause of death and long-term disability worldwide, and its prevalence is projected to increase as society ages1. While substantial advances have been made in acute interventions and secondary prevention strategies, adjunctive neuroprotective treatments have not followed apace2,3,4,5,6,7. Further research is needed into stroke pathobiology because mechanisms by which therapies may or may not prove effective are poorly understood. This is largely due to the heterogeneous nature of the stroke patient population, many of whom have numerous comorbidities that confound analysis1. One driver of limitations in research is the absence of tissue-level data-the gold standard in biomedical research-due to the prohibitive morbidity of sampling tissue from the human central nervous system. Specifically, vascular tissue harvesting in a living human would cause a stroke, so vascular tissue is typically only obtained at autopsy, which is under-representative of the general population and skews toward more advanced disease in elderly patients with concomitant diagnoses.
In such cases, when sufficient human data cannot be utilized, animal models can bridge the data gaps. Large animal models of stroke are limited as most large animals used in research are ungulates having a rete mirabile that prevents direct endovascular access to the cerebral arteries8,9,10,11,12,13,14,15,16,17. Rabbits have a long history of use for the investigation of cardiovascular disease, including intracranial pathologies8,9,10,11,12,13,14,15,16,17. Rabbits present an ideal model for cerebrovascular diseases because they are large enough for endovascular catheterization and lack the rete mirabile that precludes intracranial access in other large mammals9,15,16,17. They have been previously utilized specifically for the investigation of IS through precise and well-controlled occlusion of an intracranial artery with a microcatheter18.
Blood pressure (BP) control, both through modulation of absolute BP or BP variability (BPV), the degree to which arterial BP fluctuates around a mean BP, is an emerging potential therapeutic target for IS patients after reports of worse outcomes in those with poorly controlled BP or BPV19,20,21,22. Mechanistic investigation into how changes lead to poor outcomes in IS patients is lacking. This is partly due to the difficulty in obtaining tissue-level data and performing well-controlled analyses in humans. To test interventions that modulate BP or BPV, animal models must be utilized to overcome these limitations. This report describes the successful pairing of a previously validated rabbit model of IS using controlled occlusion of the posterior cerebral artery in conjunction with continuous intra-arterial measurement of BP18. The method presented here improves on the previous approaches to stroke pathophysiology by applying a validated and reproducible stroke model to a system in which precise measurement and control of BP can be achieved. In this refined model, infarct burden can be assessed with post-procedural histopathologic staining of the harvested brain, which is also amenable to various stains and more advanced analyses such as spatial transcriptomics. Additionally, the occluded posterior circulation artery can also be chosen to be evaluated for morbidity analysis following survival procedures.

<table><tbody><tr><td>3-0 Silk Suture</td><td>Ethicon</td><td>A184H</td><td></td></tr><tr><td>Buprenorphine</td><td>Sigma-Aldrich</td><td>B9275</td><td></td></tr><tr><td>Catheter</td><td>Terumo</td><td>CG415</td><td>4F glide catheter</td></tr><tr><td>Endovascular Pressure Sensor</td><td>Millar</td><td>SPR-524</td><td></td></tr><tr><td>Euthasol</td><td>Virbac</td><td>PVS111</td><td></td></tr><tr><td>Guidewire</td><td>Terumo</td><td>GR1804</td><td></td></tr><tr><td>Iohexol</td><td>ThermoFisher</td><td>466651000</td><td>Iodinated Contrast</td></tr><tr><td>Ketamine</td><td>Biorbyt</td><td>orb61131</td><td></td></tr><tr><td>LabChart Software</td><td>ADInstruments</td><td></td><td></td></tr><tr><td>Lidocaine</td><td>Spectrum</td><td>LI102</td><td></td></tr><tr><td>Microcatheter</td><td>Medtronic</td><td>EV3 105-5056</td><td>Marathon Microcatheter</td></tr><tr><td>Microwire</td><td>Medtronic</td><td>EV3 103-0608</td><td>Mirage Microwire</td></tr><tr><td>PowerLab&amp;nbsp;</td><td>ADInstruments</td><td></td><td></td></tr><tr><td>Rabbit Brain 2mm Coronal Cutting Matrix</td><td>Ted Pella</td><td>15026</td><td></td></tr><tr><td>Saline</td><td>FisherScientific</td><td>23-535435</td><td></td></tr><tr><td>Sheath</td><td>Merit Medical</td><td>PSI-5F-11</td><td></td></tr><tr><td>Xylazine&amp;nbsp;</td><td>ThermoFisher</td><td>J61430.14</td><td></td></tr></tbody></table>

real-time monitoring and modulation of blood pressure in a rabbit model of ischemic stroke

Control of blood pressure, in terms of both absolute values and its variability, affects outcomes in ischemic stroke patients. However, it remains challenging to identify the mechanisms that lead to poor outcomes or evaluate measures by which these effects can be mitigated because of the prohibitive limitations inherent to human data. In such cases, animal models can be utilized to conduct rigorous and reproducible evaluations of diseases. Here we report refinement of a previously described model of ischemic stroke in rabbits that is augmented with continuous blood pressure recording to assess the impacts of modulation on blood pressure. Under general anesthesia, femoral arteries are exposed through surgical cutdowns to place arterial sheaths bilaterally. Under fluoroscopic visualization and roadmap guidance, a microcatheter is advanced into an artery of the posterior circulation of the brain. An angiogram is performed by injecting the contralateral vertebral artery to confirm occlusion of the target artery. With the occlusive catheter remaining in position for a fixed duration, blood pressure is continuously recorded to allow for tight titration of blood pressure manipulations, whether through mechanical or pharmacological means. At the completion of the occlusion interval, the microcatheter is removed, and the animal is maintained under general anesthesia for a prescribed length of reperfusion. For acute studies, the animal is then euthanized and decapitated. The brain is harvested and processed to measure the infarct volume under light microscopy and further assessed with various histopathological stains or spatial transcriptomic analysis. This protocol provides a reproducible model that can be utilized for more thorough preclinical studies on the effects of blood pressure parameters during ischemic stroke. It also facilitates effective preclinical evaluation of novel neuroprotective interventions that might improve care for ischemic stroke patients.

In the initial experiments with this model, our group successfully achieved the desired outcome of a posterior cerebral or superior cerebellar artery occlusion in 12 out of 14 animals (85.7%). For the experiment, seven males and seven females were studied. The mean animal weight was 3.6 kg (&#177; 0.46 kg). In the two animals in which success was not achieved, profound catheter-induced vasospasm precluded safe access to the intracranial circulation. In one rabbit, intracranial access could not be obtained due to occlusiv...

Watch this Scientific Journal Video about Real-Time Monitoring and Modulation of Blood Pressure in a Rabbit Model of Ischemic Stroke at JoVE.com

Real-Time Monitoring and Modulation of Blood Pressure in a Rabbit Model of Ischemic Stroke

Continuous arterial blood pressure recording allows the investigation of impacts of various hemodynamic parameters. This report demonstrates the application of continuous arterial blood pressure monitoring in a large animal model of ischemic stroke for determination of stroke pathophysiology, impact of different hemodynamic factors, and the assessment of novel treatment approaches.

Control of blood pressure, in terms of both absolute values and its variability, affects outcomes in ischemic stroke patients. However, it remains ...

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Medicine

1.1K Views.  University of Utah. Het begrijpen van de pathofysiologie van ischemische beroerte is beperkt vanwege de afwezigheid van hoogwaardige weefselgegevens bij mensen. Omdat dergelijke monsters onmogelijk te verkrijgen zijn, kunnen goed gecontroleerde diermodelstudies als surrogaat dienen. Het konijnenmodel dat in dit protocol wordt gebruikt, biedt consistente gegevens van hoge kwaliteit voor de ondervraging van ischemische beroerte.De hier beschreven techniek maakt nauwkeurige controle en modulatie van hemodyna...

Video: Real-Time Monitoring and Modulation of Blood Pressure in a Rabbit Model of Ischemic Stroke

Optimized System for Cerebral Perfusion Monitoring in the Rat Stroke Model of Intraluminal Middle Cerebral Artery Occlusion

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal models of acute ischemic stroke allows to confirm successful arterial occlusion and exclude subarachnoid hemorrhage. Cerebral perfusion monitoring can also be used to study intracranial collateral circulation, which is emerging as a powerful determinant of stroke outcome and a possible therapeutic target. Despite a recognized role of Laser Doppler perfusion monitoring as part of the current guidelines for experimental cerebral ischemia, a number of technical difficulties exist that limit its widespread use. One of the major issues is obtaining a secure and prolonged attachment of a deep-penetration Laser Doppler probe to the animal skull. In this video, we show our optimized system for cerebral perfusion monitoring during transient middle cerebral artery occlusion by intraluminal filament in the rat. We developed in-house a simple method to obtain a custom made holder for twin-fibre (deep-penetration) Laser Doppler probes, which allow multi-site monitoring if needed. A continuous and prolonged monitoring of cerebral perfusion could easily be obtained over the intact skull.

Cerebral perfusion monitoring has been demonstrated to improve accuracy in ischemic stroke models. Technical difficulties often limit the use of this essential tool for cerebrovascular research. In this video, an optimized system is shown to obtain a single or multi-site hemodynamic monitoring during intraluminal middle cerebral artery occlusion in rats.

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal ...

The Rabbit Blood-shunt Model for the Study of Acute and Late Sequelae of Subarachnoid Hemorrhage: Technical Aspects

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and controllable animal models that simulate both conditions are presently uncommon. Therefore, new models are needed in order to mimic human pathophysiological conditions resulting from SAH.
This report describes the technical nuances of a rabbit blood-shunt SAH model that enables control of intracerebral pressure (ICP). An extracorporeal shunt is placed between the arterial system and the subarachnoid space, which enables examiner-independent SAH in a closed cranium. Step-by-step procedural instructions and necessary equipment are described, as well as technical considerations to produce the model with minimal mortality and morbidity. Important details required for successful surgical creation of this robust, simple and consistent ICP-controlled SAH rabbit model are described.

The experimental intracranial pressure-controlled blood shunt subarachnoid hemorrhage (SAH) model in the rabbit combines the standard procedures &#8212; subclavian artery cannulation and transcutaneous cisterna magna puncture, which enables close mimicking of human pathophysiological conditions after SAH. We present step-by-step instructions and discuss key surgical points for successful experimental SAH creation.

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and ...

An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

Ischemic stroke leads to vasogenic cerebral edema and subsequent primary brain injury, which is mediated through destruction of the blood-brain barrier (BBB). Rats with induced ischemic stroke were established and used as in vivo models to investigate the functional integrity of the BBB. Spectrophotometric detection of Evans blue (EB) in the brain samples with ischemic injury could provide reliable justification for the research and development of novel therapeutic modalities. This method generates reproducible results, and is applicable in any laboratory without a need for special equipment. Here, we present a visualized and technical guideline on the detection of the extravasation of EB following induction of ischemic stroke in rats.

An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

The overall goal of this procedure is to provide a highly reproducible technique for in vivo assessment of the blood-brain barrier disruption in rat models of ischemic stroke.

Ischemic stroke leads to vasogenic cerebral edema and subsequent primary brain injury, which is mediated through destruction of the blood-brain barrier ...

Neuroscience

An Experimental Model of Diet-Induced Metabolic Syndrome in Rabbit: Methodological Considerations, Development, and Assessment

In recent years, obesity and metabolic syndrome (MetS) have become a growing problem for public health and clinical practice, given their increased prevalence due to the rise of sedentary lifestyles and unhealthy eating habits. Thanks to animal models, basic research can investigate the mechanisms underlying pathological processes such as MetS. Here, we describe the methods used to develop an experimental rabbit model of diet-induced MetS and its assessment. After a period of acclimation, animals are fed a high-fat (10% hydrogenated coconut oil and 5% lard), high-sucrose (15% sucrose dissolved in water) diet for 28 weeks. During this period, several experimental procedures were performed to evaluate the different components of MetS: morphological and blood pressure measurements, glucose tolerance determination, and the analysis of several plasma markers. At the end of the experimental period, animals developed central obesity, mild hypertension, pre-diabetes, and dyslipidemia with low HDL, high LDL, and an increase of triglyceride (TG) levels, thus reproducing the main components of human MetS. This chronic model allows new perspectives for understanding the underlying mechanisms in the progression of the disease, the detection of preclinical and clinical markers that allow the identification of patients at risk, or even the testing of new therapeutic approaches for the treatment of this complex pathology.

We describe methods to develop an experimental model of diet-induced metabolic syndrome (MetS) in rabbits using a high-fat, high-sucrose diet. Animals developed central obesity, mild hypertension, pre-diabetes, and dyslipidemia, thus reproducing the main components of human MetS. This chronic model will allow acquisition of knowledge underlying mechanisms of disease progression.

In recent years, obesity and metabolic syndrome (MetS) have become a growing problem for public health and clinical practice, given their increased ...

Simultaneous Electrocardiography Recording and Invasive Blood Pressure Measurement in Rats

For studies related to cardiovascular physiology or pathophysiology, blood pressure (BP) and electrocardiography are basic observational parameters. Research focusing on cardiovascular disease models, potential cardiovascular therapeutic targets or pharmaceutical agents requires assessment of systemic arterial pressure and heart rhythm changes. In situations where radio telemetry systems are not available or affordable, the technique of femoral artery cannulation is an alternative way to obtain intra-arterial pressure waveform recordings and systemic BP measurements. This technique is economical and can be performed with standard equipment in animal facilities. However, invasive arterial pressure recording requires cannulation of small arteries, which can be a challenging surgical skill. Here, we present step-by-step protocols for femoral artery cannulation procedures. Key procedures include the calibration of the data acquisition system, tissue dissection and femoral artery cannulation, and setup of the arterial cannulation system for pressure recording. Surface electrocardiography recording procedures are also included. We also present examples of BP recordings from normotensive and hypertensive rats. This protocol allows reliable direct recordings of systemic BP with simultaneous electrocardiography.

Here, we describe a setup for simultaneous recording of electrocardiography and intra-arterial blood pressure (BP) in experimental rats, which can be done with standard equipment in animal facilities and can be applied to physiological or pharmacological studies to investigate pathogenic or therapeutic mechanisms in cardiovascular medicine.

For studies related to cardiovascular physiology or pathophysiology, blood pressure (BP) and electrocardiography are basic observational parameters. ...

Intracranial Pressure Monitoring In Nontraumatic Intraventricular Hemorrhage Rodent Model

Survivors of intraventricular hemorrhage are often left with significant long-term memory impairment; thus, research utilizing intraventricular hemorrhage animal models is essential. In this study, we sought out ways to measure intracranial pressure, mean arterial pressure, and cerebral perfusion pressure during nontraumatic intraventricular hemorrhage in rats. The experimental design included three Sprague Dawley groups: sham, standard 200 &#181;l intraventricular hemorrhage, and vehicle control groups. By introducing an intraparenchymal fiberoptic pressure sensor, precise intracranial pressure measurements were obtained in all groups. Cerebral perfusion pressures were calculated with the knowledge of intracranial pressure and mean arterial pressure values. As expected, the intraventricular hemorrhage and vehicle control groups both experienced a rise in the intracranial pressure and subsequent decline in cerebral perfusion pressure during intraventricular injection of autologous blood and artificial cerebrospinal fluid, respectively. The addition of an intraparenchymal fiberoptic pressure sensor is beneficial in monitoring precise intracranial pressure changes.

Monitoring intracranial pressure in rodent&#160;models of nontraumatic intraventricular hemorrhage is not common in the current literature. Herein, we demonstrate a technique for measuring intracranial pressure, mean arterial pressure, and cerebral perfusion pressure during intraventricular hemorrhage in a rat&#160;animal model.

Survivors of intraventricular hemorrhage are often left with significant long-term memory impairment; thus, research utilizing intraventricular hemorrhage ...

Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits

Patients with ion channelopathies are at a high risk of developing seizures and fatal cardiac arrhythmias. There is a higher prevalence of heart disease and arrhythmias in people with epilepsy (i.e., epileptic heart.) Additionally, cardiac and autonomic disturbances have been reported surrounding seizures. 1:1,000 epilepsy patients/year die of sudden unexpected death in epilepsy (SUDEP). The mechanisms for SUDEP remain incompletely understood. Electroencephalograms (EEG) and electrocardiograms (ECG) are two techniques routinely used in the clinical setting to detect and study the substrates/triggers for seizures and arrhythmias. While many studies and descriptions of this methodology are in rodents, their cardiac electrical activity differs significantly from humans. This article provides a description of a non-invasive method for recording simultaneous video-EEG-ECG-oximetry-capnography in conscious rabbits. As cardiac electrical function is similar in rabbits and humans, rabbits provide an excellent model of translational diagnostic and therapeutic studies. In addition to outlining the methodology for data acquisition, we discuss the analytical approaches for examining neuro-cardiac electrical function and pathology in rabbits. This includes arrhythmia detection, spectral analysis of EEG and a seizure scale developed for restrained rabbits.

Using simultaneous video-EEG-ECG-oximetry-capnography, we developed a methodology to evaluate the susceptibility of rabbit models to develop provoked arrhythmias and seizures. This novel recording system establishes a platform to test the efficacy and safety of therapeutics and can capture the complex cascade of multi-system events that culminate in sudden death.

Patients with ion channelopathies are at a high risk of developing seizures and fatal cardiac arrhythmias. There is a higher prevalence of heart disease ...

Model of Ischemia and Reperfusion Injury in Rabbits

The protocol here provides a simple, highly replicable methodology to induce in situ acute regional myocardial ischemia in the rabbit for non-survival and survival experiments. New Zealand White adult rabbit is sedated with atropine, acepromazine, butorphanol, and isoflurane. The animal is intubated and placed on mechanical ventilation. An intravenous catheter is inserted into the marginal ear vein for the infusion of medications. The animal is pre-medicated with heparin, lidocaine, and lactated Ringer&#39;s solution. A carotid cut-down is performed to obtain arterial line access for blood pressure monitoring. Select physiologic and mechanical parameters are monitored and recorded by continuous real-time analysis.
With the animal sedated and fully anesthetized, either a fourth intercostal space small left thoracotomy (survival) or midline sternotomy (non-survival) is performed. The pericardium is opened, and the left anterior descending (LAD) artery is located.
A polypropylene suture is passed around the second or third diagonal branch of the LAD artery, and the polypropylene filament is threaded through a small vinyl tube, forming a snare. The animal is subjected to 30 min of regional ischemia, achieved by occluding the LAD by tightening the snare. Myocardial ischemia is confirmed visually by regional cyanosis of the epicardium. Following regional ischemia, the ligature is loosened, and the heart is allowed to re-perfuse.
For both survival and non-survival experiments, the myocardial function can be assessed via an echocardiography (ECHO) measurement of the fractional shortening. For non-survival studies, data from sonomicrometry collected using three digital piezoelectric ultrasonic probes implanted within the ischemic area and the left ventricle developed pressure (LVDP) using an apically inserted left ventricle (LV) catheter can be continuously acquired for evaluating the regional and global myocardial function, respectively.
For survival studies, the incision is closed, a left needle thoracentesis is performed for pleural air evacuation, and postoperative pain control is achieved.

The present study demonstrates a highly reproducible animal model of acute regional myocardial ischemia and reperfusion injury in rabbits using a left mini-thoracotomy for survival cases or a midline sternotomy for non-survival cases.

The protocol here provides a simple, highly replicable methodology to induce in situ acute regional myocardial ischemia in the rabbit for non-survival and ...

Periorbital Laser Doppler Flowmetry for MCAO-Induced Ischemic Stroke in Rodents

Periorbital Placement of a Laser Doppler Probe for Cerebral Blood Flow Monitoring Prior to Middle Cerebral Artery Occlusion in Rodent Models

Middle cerebral artery occlusion (MCAO) is the gold-standard method for preclinical modeling of ischemic stroke in rodents. However, successful occlusion is not guaranteed by even the most skilled surgical hands. Errors primarily occur when the filament is not placed at the correct depth and include instances of either no infarction or vessel perforation, which can cause death. Laser Doppler flowmetry (LDF) is a reliable technique that provides real-time feedback on regional cerebral blood flow (CBF) during the MCAO procedure. Here we demonstrate a rapid technique for periorbital placement of a laser Doppler probe for measurement of CBF in both mice and rats. Our rationale was to simplify LDF implementation, encouraging widespread usage for improved surgical reliability. The technique eliminates the need for skull thinning and specialized equipment, with placement at the periorbital region rather than dorsal placement, promoting efficiency and ease of adoption. The protocol described here encompasses presurgical preparations, periorbital Doppler probe placement, and post-operative care. Representative results include visual depictions of procedural elements along with representative LDF tracings illustrating successful MCAO surgeries, with instances of unsuccessful filament placement leading to complications. The protocol illustrates LDF in confirming proper filament placement and offers a simplified procedure compared to alternative methods.

Author Spotlight: Advancing Stroke Research with Periorbital Doppler Monitoring of Regional Cerebral Blood Flow in Rodent Models

A minimally invasive surgical procedure is shown here, which involves placing the laser Doppler probe onto the skull over the distal region of the middle cerebral artery (MCA), a periorbital location suitable for rats and mice, to assess blood flow during transient MCA occlusion.

Middle cerebral artery occlusion (MCAO) is the gold-standard method for preclinical modeling of ischemic stroke in rodents. However, successful occlusion ...

Multisystem Monitoring in Rabbit Model: A Method to Assess Neuro-Cardiac and Respiratory Parameters to Study the Oral Drug-induced Triggers for  Arrhythmia and Seizures

Source: Bosinski, C., et al. <a href="https://www.jove.com/v/62256">Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits.</a> J. Vis. Exp. (2021).
In this video, we describe simultaneous recording of video, EEG, ECG and respiratory cues in a rabbit model in response to orally administered drug to understand the sequence of events leading to seizure. The model can be used for testing potential drugs for their neuro-cardiac safety.

Source: Bosinski, C., et al. Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits. J. Vis. Exp. ...

Real-Time Monitoring and Modulation of Blood Pressure in a Rabbit Model of Ischemic Stroke

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