Name: Middle Cerebral Artery Occlusion (MCAO) to Induce Stroke in a Murine Model
Uploaded: 2023-05-05T14:50:00
Description: Watch this Scientific Journal Video about Middle Cerebral Artery Occlusion MCAO to Induce Stroke in a Murine Model at JoVE.com

middle cerebral artery occlusion (mcao) to induce stroke in a murine model

Establishing a Transcranial Middle Cerebral Artery Occlusion Mice Model

Nearly 800,000 people suffer a stroke in the US every year, and most of these strokes are ischemic in nature1. Timely restoration of the cerebral blood flow with tissue plasminogen activator (tPA) and/or thrombectomy is currently the most effective treatment for stroke patients; however, the full recovery of neurologic functions in the long term is rare2,3. Thus, searching for novel stroke therapy that targets functional improvement is an intense area of research that requires clinically relevant animal models of stroke. 
The most common ischemic stroke model in rodents uses intraluminal middle cerebral artery occlusion (MCAO) to induce stroke. In this model, initially developed by Zea Longa in 1989, a nylon filament is introduced into the internal carotid artery (ICA) to block the blood flow to the middle cerebral artery (MCA)4. However, this model has limitations. First, when the filament is inserted into the ICA, the blood flow to the posterior cerebral artery (PCA) could be partially blocked as well, especially in mice. Critically, the posterior communicating artery (PcomA), a small artery that connects anterior and posterior cerebral circulation, is frequently underdeveloped in some mouse strains, such as C57Bl/6, the strain predominantly used in experimental stroke research. This patency of the PcomA is believed to contribute to the variability in lesion size in mice after stroke5. Indeed, when blood flow to the PCA drops precipitously during MCAO, and the PcomA is unable to provide sufficient collateral blood flow, the stroke infarct can expand into the territory of the PCA. Moreover, in this model, a long duration of ischemia leads to a higher chance of mortality in mice. Consequently, a short MCAO duration of 30-60 min is typically used in mice. However, most stroke patients experience a few hours of ischemia before reperfusion treatment. Thus, a mouse stroke model with an extended duration of ischemia is of high clinical relevance. 
The overall goal of this procedure is to model ischemic stroke in mice that have a medium-sized infarct and an excellent survival rate. This transcranial MCAO model addresses critical attributes of clinical stroke, as prolonged ischemia can be performed, and aged mice tolerate this model well, allowing for the long-term assessment of functional recovery.

Author Spotlight: Innovative Techniques and Future Directions in Stroke Research 

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

Stroke affects nearly 800, 000 people annually in the US.Around 87%of this, ischemic, presenting as a severe medical condition. However, there are limited treatment options available especially during the chronic stroke phase. Our research aims to develop a recovery enhancing therapy to reduce disability and improve the quality of life for stroke patients.They widely use the filament, the middle cerebral artery occlusion model, as a drawback where the inserted filament can block a blood flow to the posterior cerebral artery. The C57BL/6 in mice often have an undisturbed posterior communicating artery leading to a high mortality in the later chronic stroke recovery phase. Our modified transcranial middle cerebral artery occlusion model partially across the MCA at the trunk generating brain damage in the cortex and the striatum via a small cranial window.This model achieves a high long-term survival rate, detectable neurological deficits, and works well even in aged mice. Our transcranial MCAO model can incorporate the critical attributes of clinical stroke, such as delayed reperfusion, neurological deficits, and aging factor. This model can be used to evaluate the therapeutical potential of new stroke treatments on long-term function recovery.

Watch this Scientific Journal Video about Middle Cerebral Artery Occlusion MCAO to Induce Stroke in a Murine Model at JoVE.com

Middle Cerebral Artery Occlusion (MCAO) to Induce Stroke in a Murine Model  

To begin, lay an eight to 10 weeks anesthetized C57 black six male mouse on the surgical bench. Using blunt forceps, pull out the tongue and hold it with fingers. Insert a laryngoscope into the mouth and visualize the vocal cord.Stabilize the mouse chin on the laryngoscope and hold the 20 gauge intravenous, or IV catheter with a guide wire inserted in it. Then slightly insert the guide wire into the vocal cord and slowly push the 20 gauge IV catheter into the trachea until its wing part is even with the nose tip. Once the mouse is connected to the ventilator, keep it in a lateral position with the right temporal area facing up.Maintain the rectal temperature at 37 degrees Celsius using a heating pad and a heat lamp. After locating the zygomatic arch and incising the temporal muscle, use two forceps to dissect the underlying zygomatic arch and expose the joint between the maxilla and zygomatic bones. Then using scissors, cut a three millimeter portion of the zygomatic arch and remove it.Finally, separate the masseter muscle from the skull base. Position four small retractors in different directions and expose the cranial skull base with one retractor pulling the trigeminal nerve branches laterally. Apply a saline drop to the skull above the middle cerebral artery, or MCA trunk and proximal to the rhinal cortex branch.Use an electrical grinder to thin the skull until a small fracture is visible. And with the tip of the forceps, remove the thin skull. Place a single-strand loop of black braided silk on top of the MCA.Then insert an 8-O microsurgical needle to lift the MCA trunk and tie the suture under the needle, leaving the needle's two ends on top of the silk thread loop knot. For transient middle cerebral artery occlusion, or MCAO, slightly tighten the silk thread knot under the needle to block arterial blood flow, representing MCAO onset. Use the forceps to hold the suture and slowly remove the needle at the end of the ischemia.The laser speckle contrast imaging confirmed the reduced blood supply in the right MCA. For transient MCAO after suture removal, the cerebral blood flow reperfusion was evident, and further improved after 24 hours. TTC staining of the brain after 24 hours of stroke demonstrated the generation of infarcted tissue in both the cortical and lateral striatum areas in young and aged mice.The infarct size was moderate compared to filament MCAO.

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Research

JoVE Journal

Neuroscience

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.

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. ...