A Rat Model of Middle Cerebral Artery Occlusion/Reperfusion Without Damaging the Anatomical Structure of Cerebral Vessels
Summary
The current protocol describes a rat model of middle cerebral artery occlusion/reperfusion that preserves the anatomical structure of cerebral vessels without causing damage.
Abstract
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.
Introduction
Stroke, the leading cause of death and long-term neurological dysfunction in adults worldwide, encompass various types, with ischemic stroke accounting for approximately 81.9% of cases, while cerebral hemorrhage and subarachnoid hemorrhage account for 14.9% and 3.1%, respectively1. Ischemic stroke often results from atherosclerosis or occlusion of the middle cerebral artery (MCA), leading to local cerebral blood flow reduction and subsequent brain damage. This reduction in nutrient supply to ischemic cells triggers energy depletion, causing membrane integrity loss, cell swelling, and eventual lysis. Additionally, factors like increased intracellular calcium, excitatory amino acid release, heightened free radical production, and inflammatory cell activation contribute to brain tissue injury post-stroke2.
The treatment goals for ischemic stroke are multifaceted: reduce ongoing neurological damage and lower mortality and long-term disability; prevent complications arising from immobility and neurological dysfunction; and minimize the risk of stroke recurrence. Given the complexity of ischemic stroke pathophysiology, a robust stroke model is essential for the research and development of treatments targeting ischemic stroke.
Currently, middle cerebral artery occlusion (MCAo) is a common animal model of ischemic stroke3. MCAo is typically performed by inserting a silicon-tipped or flame-blunted monofilament through the cervical vessels until it reaches the origin of the middle cerebral artery (MCA). In 1986, Koizumi and colleagues developed a technique involving the insertion of a silicon-tipped monofilament through an incision in the common carotid artery (CCA), allowing it to reach the entrance of the MCA and block its blood flow4. In 1989, Longa’s group reported another MCAo method that involved introducing a monofilament through an incision in the external carotid artery (ECA). The monofilament then traverses the bifurcation of the internal carotid artery (ICA) and ECA before reaching the starting point of the MCA5.
The limitations of the two existing methods for inducing MCAo are evident. Koizumi’s method necessitates the permanent blockage of the ipsilateral CCA, leading to reduced MCA perfusion since reperfusion solely relies on the circle of Willis4. Conversely, Longa’s method involves cutting the ECA, resulting in damage to the ECA system and subsequent impairment of facial muscle function5. This damage can impact food and water intake, as well as cause sensory dysfunction of the face, affecting rehabilitation and the assessment of neurological function post-ischemic stroke6.
The current protocol seeks to address these limitations by developing a new method to induce an ischemic stroke model without damaging the anatomical structure of cerebral vessels. This novel approach utilizes microsurgical techniques to repair the CCA, thus avoiding the drawbacks associated with previous methods.
Protocol
Representative Results
Discussion
MCAo is a widely used method to create animal models of stroke, effectively replicating the stroke process observed in patients by restoring blood flow after ischemia10,11,12. Our laboratory has developed an ischemic stroke model that enables reperfusion from the CCA without causing damage to the ECA. Following the induction of ischemia, microsurgical repair of the CCA is performed. Although direct measurement of CCA blood flow …
Disclosures
The authors have nothing to disclose.
Acknowledgements
This work was supported by the Hebei Province Introducing Foreign Intelligence Project in 2023. We thank Lisa Kreiner, PhD, from Liwen Bianji, (Edanz) (www.liwenbianji.cn), for editing the English text of a draft of this manuscript.
Materials
| 10% formalin | Beijing Labgic Technology Co.,Ltd. | BL913A | |
| 11-0 microsurgical sutures | Ningbo Medical Suture Needle Co., Ltd. | 3/8 1×5 | |
| 2,3,5-triphenyltetrazolium chloride (TTC) | SIGMA | T8877 | |
| 3-0 silk suture | Ningbo Medical Suture Needle Co., Ltd. | 3-0 | |
| 4% isoflurane | Tianjin Ringpu Bio-Technology Co., Ltd. | 20221102 | |
| 6-0 silk suture | Ningbo Medical Suture Needle Co., Ltd. | 6-0 | |
| 70% ethanol | Shandong Lierkang Medical Technology Co., Ltd. | 20230408 | |
| Filament | Xinong Technologies Co Ltd. | 2838-A5 | |
| Ketamine | Jiangsu Hengrui Pharmaceutical Co., Ltd. | 230613BL | |
| Male Sprague Dawley rats | Jinan Pengyue Experimental Animal Breeding Co., Ltd. | SCXK (Lu) 20190003 | |
| Microclips | Shanghai Jinzhong Medical Device Factory | W40150 | |
| Microscope | ZEISS | S100 OPMI pico | |
| Nursing box | Beijing Fuyi Electrical Appliance Co., Ltd. | FYL-YS-100L | |
| Xylazine | SIGMA | PHR3263 |
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