ProtocolAnesthetic methods in rats determine outcome after experimental focal cerebral ischemia: mechanical ventilation is required to obtain controlled experimental conditions
Section snippets
Type of research
The rat is one of the most widely used small animals in neuroscience research in general and in research in cerebral ischemia especially today [15], [34]. Most models of cerebral ischemia are surgically invasive and thus require anesthetics during the ischemic insult. Modern anesthesia in man combines volatile anesthetic gases, intravenous agents, and neuromuscular blocking agents to maintain unconsciousness while maintaining cardiovascular stability. In this context, a variety of commonly used
Time required
All described anesthetic procedures, intraperitoneal injection of chloral hydrate, or application of halothane via face-mask or an orotracheal tube, can be managed within a few minutes. After initial immobilization of the rat by restraining in a box or pharmacologically in a container with 4% halothane, all procedures lead to unconsciousness and immobility of the animals within 1–5 min. In experienced hands with adequate optical instruments, e.g. operational microscope, intubation can be made
Materials
A total of 35 male Sprague–Dawley rats (250–300 g body weight) were used for this study. Animals were purchased from Charles River Laboratory (Sulzfeld, Germany) and were cared for before and at all stages of the experiment in compliance with applicable institutional guidelines and regulations of the government of Bavaria.
Animal preparation and monitoring
Rats were fasted overnight before surgery with free access to water. After induction of anesthesia according to the experimental groups, all rats were subjected to 90 min of MCA occlusion by insertion of a silicone-coated 4-0 nylon monofilament via the external carotid artery as described by Koizumi et al. [25]. Reperfusion was achieved by withdrawing the filament into the external carotid artery after 90 min. Five animals with subarachnoid hemorrhage as confirmed by autopsy were excluded from
Physiological parameters
In group A, anesthesia with chloral hydrate within all subgroups resulted in hypercapnia with mean values between 52.1±6.9 and 57.3±6.3 mmHg without significant differences over time. All animals of group A showed respiratory acidosis during anesthesia with mean values between 7.32±0.02 and 7.37±0.05 without significant differences between groups or over time. Mean blood pressure ranged between 68.3±7.9 and 75.1±12.0 mmHg.
Anesthesia with halothane via face-mask (group B) also resulted in
Discussion
Anesthetic agents, blood pressure, arterial pH and blood gases have all been implicated in the pathophysiology of experimental stroke [7], [9], [47], [55], [56]. Despite their widespread usage, comparative studies on appropriate methods of rat anesthesia are scarce. We investigated the influence of various common anesthetic techniques on intra- and postischemic physiological variables, mortality and histological outcome in a model of transient focal cerebral ischemia.
In our study all anesthetic
Conclusion
Intra- and postischemic physiologic parameters such as blood pressure, pH and blood gases critically determine outcome after focal cerebral ischemia. All spontaneously breathing animals, whether anesthetized by chloral hydrate i.p. or by halothane via face-mask, exhibited hypercapnia, acidosis, and decreased blood pressure during anesthesia, resulting in high mortality and a significantly increased infarct volume compared to mechanically ventilated animals with halothane anesthesia. Although
Quick procedure
- 1.
Rats are anesthetized in a container with 4% halothane.
- 2.
Rats are then placed in supine position on an inclined plane and the vocal cords are visualized under the operating microscope by lifting the jaw and the tongue with a small surgical clamp, bent at the tip.
- 3.
The tube (A 16 G venous catheter, diameter 1.8 mm, with a 2.0-cm long conical silicone cuff), supplied with a blunted steel mandrin, is placed intratracheally and pushed forward until an increasing resistance due the silicone cuff is felt.
Essential literature references
Original papers: [7], [9], [17], [22], [29], [49], [54], [67]
Book chapters: [37]
Acknowledgments
This work was supported by the Deutsche Forschungsgemeinschaft (Schm1067/2-1) and Friedrich Baur Stiftung. Special thanks to Professor Lawrence M. Shuer, MD, Stanford University, CA for permission to reproduce the drawing of the face mask.
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