This work was supported by the National Natural Science Foundation of China (82173781 and 82373835), Postdoctoral research project (BKS212055), Science and Technology Innovation Project of Foshan Science and Technology Bureau (2320001007331), Guangdong Basic and Applied Basic Research Foundation (2019A1515010806), Key Field Projects (Intelligent Manufacturing) of General Universities in Guangdong Province (2020ZDZX2057), and the Scientific Research Projects (Characteristic Innovation) of General Universities in Guangdong Province (2019KTSCX195).

The experimental protocol was conducted in compliance with the Use of Laboratory Animals and Institutional Animal Care and Use Committee guidelines at Chengdu University of Traditional Chinese Medicine (Record number: 2019-DL-002). All animal research data have been documented following the ARRIVE (Animal Research: Reporting In Vivo Experiments) guidelines. Male Sprague Dawley (SD) rats weighing 250 g &#177; 20 g and aged 6-8 weeks were utilized for this study. The specifics regarding the animals, reagents, and equipment employed are listed in the Table of Materials.
1. Animal preparation
<ol>
	<li>Induce and maintain deep anesthesia in rats using Zoletil 50 (50mg/kg, IM) and Xylazine Hydrochloride (40mg/kg, IP).&#160;Ensure the body temperature is maintained at 37 &#177; 0.5 &#176;C using a rectal probe connected to a heating pad throughout the surgical procedure. Apply veterinary ointment to the rat&#39;s eyes to prevent them from drying out.</li>
	<li>Shave the hair from the head and neck of the rats. Use depilatory cream to remove fur from the head and neck, then wash off the cream with normal saline. Disinfect the skin at the surgical site by applying ethanol and povidone-iodine three times using sterile cotton balls.</li>
	<li>Make a 2 cm incision in the middle of the rat&#39;s head along the direction of the sagittal suture using a scalpel, and carefully remove the muscles covering the skull.</li>
	<li>Thin the skull on the ischemic side of the rat using a skull drill. Use normal saline to cool down and remove debris during the grinding process. Record the baseline blood flow of the rats using laser speckle contrast imaging (LSCI).</li>
	<li>Apply ethanol and povidone-iodine three times to the skin of the neck using sterile cotton balls. Make a 2 cm incision along the midline of the neck using a surgical blade. Use retractors to laterally pull back the skin and salivary glands, exposing the sternocleidomastoid and cervical muscles.</li>
	<li>Separate the sternocleidomastoid and cervical muscles to expose the carotid territory. Identify the vascular anatomy of the common carotid artery (CCA), the internal carotid artery (ICA), and the external carotid artery (ECA). Based on vascular anatomy, separate the CCA, as well as the CCA-derived ECA and ICA15. 
	NOTE: The anatomical structure of the rat indicates that the CCA region is mainly covered by sternocleidomastoid and cervical muscles, with the ICA and ECA derived from the CCA. After separation exposes the CCA, a Y-shaped fork can be seen along the CCA, representing the ICA and ECA. Be cautious not to damage the vagus nerve, which runs parallel to the CCA.</li>
</ol>
2. Occlusion of the MCA
<ol>
	<li>Tie an easily untangled knot over the external carotid artery (ECA) and common carotid artery (CCA) using a 3-0 silk thread to temporarily block blood flow. Position a vessel clip on the CCA approximately 0.5 cm from the first knot.
	<ol>
		<li>Dye the needle of a 5 mL syringe black. Create a small puncture in the CCA using the black-dyed needle and mark the pinholes in black.</li>
	</ol>
	</li>
	<li>Insert the monofilament nylon suture into the CCA through the black mark. Open the vascular clip and guide the nylon wire into the Internal Carotid Artery (ICA) until it halts with slight resistance. Tighten the second knot securely to keep the monofilament nylon suture in place inside the artery, preventing displacement from the blocking position.</li>
	<li>Note the time of ischemia at this point and measure the blood flow value on the ischemic side using LSCI. Administer drops of bupivacaine to the neck wound for analgesia and wound closure. Remove the anesthesia mask and allow the rat to recover. 
	NOTE: If the monofilament nylon suture encounters difficulty entering the ICA, retract it slightly and attempt insertion again.</li>
</ol>
3. Reperfusion of MCAO
<ol>
	<li>Following 90 min of ischemia, re-anesthetize the rats with isoflurane. Measure cerebral blood flow from the ischemic side using LSCI and ensure the monofilament nylon suture has not shifted.</li>
	<li>Untie the knots on the external carotid artery (ECA) to allow blood reperfusion. Release the knot on the common carotid artery (CCA), secure the monofilament suture, and withdraw the suture. Apply a vascular clip ahead of the fixed monofilament suture to prevent bleeding.</li>
	<li>Replace the first CCA knot with a vascular clip. Use tweezers to rotate the vessel incision sideways (Figure 1). Clamp the incision with tweezers and ligate it using 6-0 thread to repair the incision. Remove the vascular clip, check for leaks, and confirm complete reperfusion (Video 1). 
	NOTE: Perform ligation to repair the incision to avoid excessive vessel clamping, which could lead to CCA stenosis.</li>
	<li>Record cerebral cortical blood flow values using LSCI after incision ligation to confirm successful reperfusion. Close the neck incision with sutures and administer 1,00,000 units of penicillin and 2 mL of saline to prevent infection and dehydration.</li>
	<li>Maintain warmth until the rats regain consciousness. Offer soft food post-surgery and monitor the animals&#39; vital signs.</li>
</ol>
4. Evaluation of nerve function and cerebral ischemic injury
<ol>
	<li>Once the rats have fully awakened, their neural function will be assessed by researchers unaware of the animal grouping. Utilize the Longa score scale (Table 1) and the modified neurological severity score (mNSS) scale (Table 2) to evaluate the neurological function of all rats.</li>
	<li>After 24 h post-surgery, euthanize the rats under isoflurane-induced anesthesia (4% isoflurane at 4 L/min oxygen) via cervical dislocation16 (following institutionally approved protocols). Rinse the brain with ice water to remove any remaining blood.</li>
	<li>Divide the brain tissue into 5 sections and stain them with TTC. Capture images of both sides of the brain tissue slices, measure the ischemic area of the front and back sides using Image J software, and calculate the infarction rate using the following formula: 
	<img alt="Equation 1" src="/files/ftp_upload/67060/67060eq01.jpg" style="margin: auto;" /> 
	NOTE: S1 and S2 represent the infarct area of the proximal forebrain and proximal brainstem sides of the brain slices, respectively. The infarct area of each brain slice is corrected using Swanson&#39;s formula, where h represents the thickness of the brain slice. This formula, previously outlined by the author in prior research17, allows for a more accurate calculation of the infarct lesion size.</li>
	<li>Analyze the data using statistical and graphing software to assess the model&#39;s success rate and stability.</li>
	<li>Employ the immunofluorescence method to label cortical neurons in rats, further validating the neuronal damage caused by this method. After 24 h of ischemia-reperfusion in rats, cardiac perfusion using pre-cooled PBS was performed to obtain brain tissue.</li>
	<li>Embed and freeze the brain tissue into sections. Use neuronal antibodies such as the immature neuronal marker Doublecortin (DCX)18, neuronal nucleus marker Neuronal Nuclei (NeuN)18, and neuron dendritic marker Microtubule-associated protein-2 (MAP-2)19. Observe the expression of ischemic neurons using laser confocal microscopy.</li>
</ol>

Modified MCAO/R Model with Preserved ECA for Enhanced Reperfusion and Stability

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The middle cerebral artery occlusion (MCAO) model induced by a monofilament nylon suture is the most common method used for preparing MCAO models. This approach is widely adopted in preclinical studies and has gained recognition from many practitioners due to its simplicity, lack of need for craniotomy, minimal surgical trauma, and ability to achieve reperfusion.
There are two classical surgical techniques for intraluminal filament MCAO: the Koizumi method8 and the Long...

According to the World Health Organization, stroke has remained the second leading cause of death worldwide for the past decade, with a high incidence rate, high mortality, and high disability rate1,2. As the global population ages, the incidence of stroke is expected to increase in developing countries, potentially becoming the leading cause of premature death and disability in adults. Additionally, there is a trend for strokes to occur at a younger age3. The loss of the labor force after a stroke also places a heavy burden on families and society4. Therefore, the development of safe and effective treatments poses a major challenge in stroke research.
Animal models serve as crucial tools for studying the prevention and treatment of human diseases. The successful translation of stroke treatment strategies relies on the reproducibility and reliability of stroke animal models5,6. The middle cerebral artery (MCA) is a common site for clinical stroke, making the MCAO model the closest model to human ischemic stroke. The MCAO model, prepared using the suture method, has been favored by researchers due to advantages such as no craniotomy and easy control of ischemic time. It has been utilized in over 40% of neuroprotective experiments7. However, despite its numerous advantages, the operational details of this model remain a controversial topic for many researchers.
For the suture-induced middle cerebral artery occlusion (MCAO) model, reperfusion occurs by withdrawing the suture. Currently, two main methods are used for suture insertion: Koizumi's method8 and Longa's method9. In Koizumi's method, the suture enters the internal carotid artery (ICA) mainly through the common carotid artery (CCA) incision, while in Longa's method, it passes through the severed external carotid artery (ECA) into the ICA. During reperfusion, the Koizumi method requires permanently ligating the CCA incision and relies on the circle of Willis for reperfusion10. However, some studies suggest that effective reperfusion cannot be achieved solely through the compensatory supply of the circle of Willis after losing the CCA supply. Moreover, the circle of Willis exhibits high anatomical variability, especially in C57Bl/6 mice, increasing infarction variability and reducing experimental data reliability. Consequently, this method has been increasingly questioned by researchers11.
Longa's method involves inserting a suture through the severed ECA and then permanently ligating the internal carotid artery (ICA) once the suture is withdrawn. This preserves CCA patency, allowing blood perfusion up to 100% of baseline values. However, this method necessitates separating the external carotid artery and small arterial branches, cutting them off, or electrocoagulating them, making the procedure challenging. It also disrupts the brain's complete blood flow structure, which differs from the clinical patient's state12. Importantly, studies indicate that cutting or ligating the ECA can cause ischemic lesions in the muscles controlling chewing and swallowing, affecting animal diet and leading to postoperative animal death and severe sensory and motor damage in rats13,14.
Hence, a modified model preparation method is urgently needed to address these issues. This study introduces a modified MCAO modeling method that repairs the CCA insertion incision and achieves effective reperfusion. The procedure is simple, practical, and feasible, inducing significant neurological damage and replicable infarct lesions and providing valuable guidance for stroke researchers.

<table><tbody><tr><td>Animal anesthesia system</td><td>Rayward Life Technology Co., Ltd</td><td>R500IE</td><td></td></tr><tr><td>Animal temperature maintainer</td><td>Rayward Life Technology Co., Ltd</td><td>69020</td><td></td></tr><tr><td>Cy3 secondary antibody</td><td>Wuhan Saiweier Biotechnology Co., Ltd</td><td>GB21303</td><td></td></tr><tr><td>DAP1 antibody</td><td>Wuhan Saiweier Biotechnology Co., Ltd</td><td>G1012</td><td></td></tr><tr><td>DCX antibody</td><td>Wuhan Saiweier Biotechnology Co., Ltd</td><td>GB13434</td><td></td></tr><tr><td>Goat serum</td><td>Beyotime Biotechnology Co., LTD</td><td>C0265</td><td></td></tr><tr><td>GraphPad Prism</td><td>GraphPad Software</td><td>GraphPad Prism 8.0</td><td></td></tr><tr><td>ImageJ</td><td>National Institutes of Health</td><td>ImageJ software</td><td></td></tr><tr><td>Isofluran</td><td>Rayward Life Technology Co., Ltd</td><td>R510-22</td><td></td></tr><tr><td>Laser speckle blood flow imaging system</td><td>Rayward Life Technology Co., Ltd</td><td>PeriCam PSI NR</td><td></td></tr><tr><td>MAP-2 antibody</td><td>Wuhan Saiweier Biotechnology Co., Ltd</td><td>GB11128</td><td></td></tr><tr><td>Miniature hand-held skull drill</td><td>Rayward Life Technology Co., Ltd</td><td>87001</td><td></td></tr><tr><td>monofilament suture</td><td>Rayward Life Technology Co., Ltd</td><td>250-280g</td><td></td></tr><tr><td>NeuN antibody</td><td>Wuhan Saiweier Biotechnology Co., Ltd</td><td>GB11138</td><td></td></tr><tr><td>OCT embedding agent</td><td>BIOSHARP</td><td>BL557A</td><td></td></tr><tr><td>Penicillin sodium</td><td>Chengdu Kelong Chemical Co., Ltd.</td><td>17121709-2</td><td></td></tr><tr><td>Quick Antigen Retrieval Solution for Frozen Sections</td><td>Beyotime Biotechnology Co., LTD</td><td>P0090</td><td></td></tr><tr><td>SD rats</td><td>SPF ( Beijing ) Biotechnology Co.,Ltd.</td><td>250-280g</td><td></td></tr><tr><td>Triton X-100</td><td>Beyotime Biotechnology Co., LTD</td><td>ST795</td><td></td></tr><tr><td>TTC</td><td>Chengdu Kelong Chemical Co., Ltd.</td><td>2019030101</td><td></td></tr></tbody></table>

a modified model preparation for middle cerebral artery occlusion reperfusion

The middle cerebral artery occlusion reperfusion (MCAO/R) model is crucial for understanding the pathological mechanisms of stroke and for drug development.However, among the commonly used modeling methods, the Koizumi method often faces scrutiny due to its ligation of the common carotid artery (CCA) and its inability to achieve adequate reperfusion. Similarly, the Longa method has been criticized for disconnecting and ligating the external carotid artery (ECA). This study aims to introduce a modified model preparation method that preserves the integrity of the ECA, involves inserting a monofilament nylon suture through the CCA, repairing the ligated CCA incision, and maintaining reperfusion from the CCA. Reperfusion of blood flow was confirmed using laser speckle flow imaging. Evaluation methods such as the Longa scale, Modified Neurological Severity Score, triphenyltetrazolium chloride (TTC) staining, and immunofluorescence labeling of neurons demonstrated that this approach could induce stable ischemic nerve damage. This modified MCAO/R model protocol is simple and stable, providing valuable guidance for practitioners in the field of cerebral ischemia.

Laser speckle flow imaging demonstrated that prior to the occlusion of the monofilament nylon suture, there was abundant blood flow in the middle cerebral artery (MCA) area, and the baseline blood flow values of the rats were recorded. Following the occlusion of the MCA, the blood flow value on the ischemic side of the brain rapidly decreased. Before withdrawing the suture, the blood flow values on the ischemic side were rechecked to confirm whether the suture was occluding the MCA. The results indicated only a slight ch...

Author Spotlight: A Novel Approach to Cerebral Ischemia Modeling &#8211; Enhancing Reperfusion and Simplifying Procedure

This protocol describes the process of preparing for middle cerebral artery occlusion reperfusion via the common carotid artery.

A Modified Model Preparation for Middle Cerebral Artery Occlusion Reperfusion

The middle cerebral artery occlusion reperfusion (MCAO/R) model is crucial for understanding the pathological mechanisms of stroke and for drug ...

a-modified-model-preparation-for-middle-cerebral-artery-occlusion

Research

JoVE Journal

Medicine

817 Views.  Foshan University. This protocol describes the process of preparing for middle cerebral artery occlusion reperfusion via the common carotid artery.

Video: Author Spotlight: A Novel Approach to Cerebral Ischemia Modeling – Enhancing Reperfusion and Simplifying Procedure

Focal Cerebral Ischemia Model by Endovascular Suture Occlusion of the Middle Cerebral Artery in the Rat

Stroke is the leading cause of disability and the third leading cause of death in adults worldwide1. In human stroke, there exists a highly variable clinical state; in the development of animal models of focal ischemia, however, achieving reproducibility of experimentally induced infarct volume is essential. The rat is a widely used animal model for stroke due to its relatively low animal husbandry costs and to the similarity of its cranial circulation to that of humans2,3. In humans, the middle cerebral artery (MCA) is most commonly affected in stroke syndromes and multiple methods of MCA occlusion (MCAO) have been described to mimic this clinical syndrome in animal models. Because recanalization commonly occurs following an acute stroke in the human, reperfusion after a period of occlusion has been included in many of these models. In this video, we demonstrate the transient endovascular suture MCAO model in the spontaneously hypertensive rat (SHR). A filament with a silicon tip coating is placed intraluminally at the MCA origin for 60 minutes, followed by reperfusion. Note that the optimal occlusion period may vary in other rat strains, such as Wistar or Sprague-Dawley. Several behavioral indicators of stroke in the rat are shown. Focal ischemia is confirmed using T2-weighted magnetic resonance images and by staining brain sections with 2,3,5-triphenyltetrazolium chloride (TTC) 24 hours after MCAO.

Surgical induction of ischemic brain damage in the rat is a widely used model for stroke research. Here we demonstrate the induction of focal cerebral ischemia by occlusion of the middle cerebral artery. Visualization of the resulting infarct by histological staining and magnetic resonance imaging is also shown.

Stroke is the leading cause of disability and the third leading cause of death in adults worldwide1. In human stroke, there exists a highly variable ...

Neuroscience

Performing Permanent Distal Middle Cerebral with Common Carotid Artery Occlusion in Aged Rats to Study Cortical Ischemia with Sustained Disability

Stroke typically occurs in elderly people with a range of comorbidities including carotid (or other arterial) atherosclerosis, high blood pressure, obesity and diabetes. Accordingly, when evaluating therapies for stroke in animals, it is important to select a model with excellent face validity. Ischemic stroke accounts for 80% of all strokes, and the majority of these occur in the territory of the middle cerebral artery (MCA), often inducing infarcts that affect the sensorimotor cortex, causing persistent plegia or paresis on the contralateral side of the body. We demonstrate in this video a method for producing ischemic stroke in elderly rats, which causes sustained sensorimotor disability and substantial cortical infarcts. Specifically, we induce permanent distal middle cerebral artery occlusion (MCAO) in elderly female rats by using diathermy forceps to occlude a short segment of this artery. The carotid artery on the ipsilateral side to the lesion was then permanently occluded and the contralateral carotid artery was transiently occluded for 60 min. We measure the infarct size using structural T2-weighted magnetic resonance imaging (MRI) at 24 hr and 8 weeks after stroke. In this study, the mean infarct volume was 4.5% &#177; 2.0% (standard deviation) of the ipsilateral hemisphere at 24 hr (corrected for brain swelling using Gerriet&#8217;s equation, n = 5). This model is feasible and clinically relevant as it permits the induction of sustained sensorimotor deficits, which is important for the elucidation of pathophysiological mechanisms and novel treatments.

Here we present a protocol to produce permanent distal middle cerebral artery occlusion in elderly female rats with simultaneous occlusion of the carotid arteries to generate large cortical infarcts and sustained deficits. We show confirmation of the lesion size using structural MRI at 24 hr and 8 weeks after stroke.

Stroke typically occurs in elderly people with a range of comorbidities including carotid (or other arterial) atherosclerosis, high blood pressure, ...

Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ischemic in origin and occur in the territory of the middle cerebral artery (MCA). Currently the only Food and Drug Administration (FDA) approved drug for the treatment of this devastating disease is tissue plasminogen activator (tPA). However, tPA has a small therapeutic window for administration (3 - 6 hr), and is only effective in 4% of the patients who actually receive it. Current research focuses on understanding the pathophysiology of stroke in order to find potential therapeutic targets. Thus, reliable models are crucial, and the MCA occlusion (MCAo) model (also termed the intraluminal filament or suture model) is deemed to be the most clinically relevant surgical model of ischemic stroke, and is fairly non-invasive and easily reproducible. Typically the MCAo model is used with rodents, especially with mice due to all the genetic variations available for this species. Here we describe (and present in the video) how to successfully perform the MCAo model (with reperfusion) in mice to generate reliable and reproducible data.

In order to understand the pathophysiology of stroke, it is important to use reliable models. This paper will describe one of the most frequently used stroke models in mice, termed the middle cerebral artery occlusion (MCAo) model (also termed the intraluminal filament or suture model) with reperfusion.

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ...

Transient Middle Cerebral Artery Occlusion Model of Neonatal Stroke in P10 Rats

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature and mature brain. Stroke occurs commonly in the perinatal period in humans, and transient ischemia-reperfusion is the most common form of stroke in neonates. The reperfusion phase is a critical component of injury progression, which occurs over a period of days to weeks, and of the endogenous response to injury. This postnatal day 10 (p10) rat model of transient middle cerebral artery occlusion (tMCAO) creates a unilateral, non-hemorrhagic focal ischemia-reperfusion injury that can be utilized to study the mechanisms of focal injury and repair in the full-term-equivalent brain. The injury pattern that is produced by tMCAO is consistent and highly reproducible and can be confirmed with MRI or histological analyses. The severity of injury can be manipulated through changes in occlusion time and other methods that will be discussed.

Neonatal stroke is a significant cause of early brain injury requiring a translational model with consistent focal injury patterns and high reproducibility in order to enable study. This study describes the detailed surgical procedure for creating a non-hemorrhagic, unilateral focal ischemia-reperfusion injury in full-term-equivalent rodents.

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature ...

Middle Cerebral Artery Occlusion Allowing Reperfusion via Common Carotid Artery Repair in Mice

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue plasminogen activator (tPA) as an approved drug treatment to target ischemic strokes. Current research in the field of ischemic stroke focuses on better understanding the pathophysiology of stroke, to develop and investigate novel pharmaceutical targets. Reliable experimental stroke models are crucial for the progression of potential treatments. The middle cerebral artery occlusion (MCAO) model is clinically relevant and the most frequently used surgical model of ischemic stroke in rodents. However, the outcomes of this model, such as lesion volume, are associated with high levels of variability, particularly in mice. The alternative MCAO model described here allows the reperfusion of the common carotid artery (CCA) and the increased perfusion of the middle cerebral artery (MCA) territory, using a tissue pad with fibrinogen-based sealant to repair the vessel, and the improved welfare of the mice by avoiding external carotid artery (ECA) ligation. This reduces the reliance on the Circle of Willis, which is known to be highly anatomically variable in mice. Representative data show that using this alternative surgical approach decreases the variability in lesion volumes between the traditional MCAO approach and the alternative approach described here.

Intraluminal filament occlusion of the middle cerebral artery is the most frequently used in vivo model of experimental stroke in rodents. An alternative surgical approach to allow common carotid artery repair is performed here, which allows the reperfusion of the common carotid artery and a full reperfusion to the middle cerebral artery territory.

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue ...

Two-vessel Occlusion Mouse Model of Cerebral Ischemia-reperfusion

In this study, a middle cerebral artery (MCA) occlusion mouse model is employed to study cerebral ischemia-reperfusion. A reproducible and reliable mouse model is useful for investigating the pathophysiology of cerebral ischemia-reperfusion and determining potential therapeutic strategies for patients with stroke. Variations in the anatomy of the circle of Willis of C57BL/6 mice affects their infarct volume after cerebral-ischemia-induced injury. Studies have indicated that distal MCA occlusion (MCAO) can overcome this problem and result in a stable infarct size. In this study, we establish a two-vessel occlusion mouse model of cerebral ischemia-reperfusion through the interruption of the blood flow to the right MCA. We distally ligate the right MCA and right common carotid artery (CCA) and restore blood flow after a certain period of ischemia. This ischemia-reperfusion injury induces an infarct of stable size and a behavioral deficit. Peripheral immune cells infiltrate the ischemic brain within the 24 h infiltration period. Additionally, the neuronal loss in the cortical area is less for a longer reperfusion duration. Therefore, this two-vessel occlusion model is suitable for investigating the immune response and neuronal recovery during the reperfusion period after cerebral ischemia.

A mouse model of cerebral ischemia-reperfusion is established to investigate the pathophysiology of stroke. We distally ligate the right middle cerebral artery and right common carotid artery and restore blood flow after 10 or 40 min of ischemia.

In this study, a middle cerebral artery (MCA) occlusion mouse model is employed to study cerebral ischemia-reperfusion. A reproducible and reliable mouse ...

Modeling Stroke in Mice: Transient Middle Cerebral Artery Occlusion via the External Carotid Artery

Stroke is the third most common cause of mortality and the leading cause of acquired adult disability in developed countries. To date, therapeutic options are limited to a small proportion of stroke patients within the first hours after stroke. Novel therapeutic strategies are being extensively investigated, especially to prolong the therapeutic time window. These current investigations include the study of important pathophysiological pathways after stroke, such as post-stroke inflammation, angiogenesis, neuronal plasticity, and regeneration. Over the last decade, there has been increasing concern about the poor reproducibility of experimental results and scientific findings among independent research groups. To overcome the so-called &#8220;replication crisis&#8221;, detailed standardized models for all procedures are urgently needed. As an effort within the &#8220;ImmunoStroke&#8221; research consortium (https://immunostroke.de/), a standardized mouse model of transient middle cerebral artery occlusion (MCAo) is proposed. This model allows the complete restoration of the blood flow upon removal of the filament, simulating the therapeutic or spontaneous clot lysis that occurs in a large proportion of human strokes. The surgical procedure of this &#8220;filament&#8221; stroke model and tools for its functional analysis are demonstrated in the accompanying video.

Different models of middle cerebral artery occlusion (MCAo) are used in experimental stroke research. Here, an experimental stroke model of transient MCAo via the external carotid artery (ECA) is described, which aims to mimic human stroke, in which the cerebrovascular thrombus is removed due to spontaneous clot lysis or therapy.

Stroke is the third most common cause of mortality and the leading cause of acquired adult disability in developed countries. To date, therapeutic options ...

Establishing a Transcranial Middle Cerebral Artery Occlusion Mice Model

A Modified Transcranial Middle Cerebral Artery Occlusion Model to Study Stroke Outcomes in Aged Mice

In experimental stroke research, middle cerebral artery occlusion (MCAO) with an intraluminal filament is widely used to model ischemic stroke in mice. The filament MCAO model typically exhibits a massive cerebral infarction in C57Bl/6 mice that sometimes includes brain tissue in the territory supplied by the posterior cerebral artery, which is largely due to a high incidence of posterior communicating artery atresia. This phenomenon is considered a major contributor to the high mortality rate observed in C57Bl/6 mice during long-term stroke recovery after filament MCAO. Thus, many chronic stroke studies exploit distal MCAO models. However, these models usually produce infarction only in the cortex area, and consequently, the assessment of post-stroke neurologic deficits could be a challenge. This study has established a modified transcranial MCAO model in which the MCA at the trunk is partially occluded either permanently or transiently via a small cranial window. Since the occlusion location is relatively proximal to the origin of the MCA, this model generates brain damage in both the cortex and striatum. Extensive characterization of this model has demonstrated an excellent long-term survival rate, even in aged mice, as well as readily detectable neurologic deficits. Therefore, the MCAO mouse model described here represents a valuable tool for experimental stroke research.

Author Spotlight: Innovative Techniques and Future Directions in Stroke Research 

This protocol demonstrates a unique mouse stroke model with a medium-sized infarct and an excellent survival rate. This model allows preclinical stroke researchers to extend the ischemia duration, use aged mice, and assess long-term functional outcomes.

In experimental stroke research, middle cerebral artery occlusion (MCAO) with an intraluminal filament is widely used to model ischemic stroke in mice. ...

A Model of Transient Cerebral Ischemia by Occlusion of Middle Cerebral Artery

Transient Middle Cerebral Artery Occlusion Model of Stroke

Stroke stands as a major cause of death or chronic disability globally. Nevertheless, existing optimal treatments are limited to reperfusion therapies during the acute phase of ischemic stroke. To gain insights into stroke physiopathology and develop innovative therapeutic approaches, in vivo rodent models of stroke play a fundamental role. The availability of genetically modified animals has particularly propelled the use of mice as experimental stroke models.
In stroke patients, occlusion of the middle cerebral artery (MCA) is a common occurrence. Consequently, the most prevalent experimental model involves intraluminal occlusion of the MCA, a minimally invasive technique that doesn't require craniectomy. This procedure involves inserting a monofilament through the external carotid artery (ECA) and advancing it through the internal carotid artery (ICA) until it reaches the branching point of the MCA. After a 45 min arterial occlusion, the monofilament is removed to allow reperfusion. Throughout the process, cerebral blood flow is monitored to confirm the reduction during occlusion and subsequent recovery upon reperfusion. Neurological and tissue outcomes are evaluated using behavioral tests and magnetic resonance imaging (MRI) studies.

Author Spotlight: Assessing Ischemic Stroke Damage Through Middle Cerebral Artery Occlusion Model 

This protocol describes the model of transient focal cerebral ischemia in mice through intraluminal occlusion of the middle cerebral artery. Additionally, examples of outcome assessment are shown using magnetic resonance imaging and behavioral tests.

Stroke stands as a major cause of death or chronic disability globally. Nevertheless, existing optimal treatments are limited to reperfusion therapies ...

Middle Cerebral Artery Occlusion in Rats with Preserved Vascular Anatomy

A Rat Model of Middle Cerebral Artery Occlusion/Reperfusion Without Damaging the Anatomical Structure of Cerebral Vessels

Ischemic stroke stands as the primary cause of long-term disability and mortality among adults worldwide. Animal models of ischemic stroke have significantly contributed to our understanding of its pathological mechanisms and the development of potential treatments. Presently, there are two common methods involving filament (endovascular suture) techniques to induce animal models of cerebral ischemia. However, these methods have inherent limitations, such as reduced blood perfusion to the brain, damage to the external carotid artery system, impaired food and/or water intake, and sensory dysfunction of the face. This article introduces a new method for inducing a rat ischemic stroke model without compromising the cerebral vascular anatomy. In this study, the common carotid artery (CCA) of Sprague-Dawley rats was exposed, and an incision was made. A filament was then inserted through the incision into the internal carotid artery to occlude the middle cerebral artery. After 1.5 h of induced ischemia, the occluding filament was fully removed from both the internal carotid artery and the CCA. The incision in the CCA was subsequently sutured using 11-0 microsurgical sutures under a microscope (magnification 4x). Through the utilization of microsurgical techniques to repair the CCA, this study successfully developed a unique method to induce an ischemic stroke model in rats while preserving the anatomical integrity of cerebral blood vessels.

The current protocol describes a rat model of middle cerebral artery occlusion/reperfusion that preserves the anatomical structure of cerebral vessels without causing damage.