1. An Accessible and Humane Approach to Mouse Intubation E Tonsfeldt 1, DL Hickman 1, DM Van Winkle 2,3 1 Veterinary Medical Unit, 2 Anesthesiology Research, Portland VA Medical Center, Portland, OR; 3 Physiology and Pharmacology, Oregon Health and Sciences University, Portland, OR With several surgeons operating a complicated model of coronary occlusion/reperfusion requiring mechanical ventilation and using inhalant anesthesia, we required a novel approach to rapid, accurate mouse intubation that did not require specialized peripheral equipment. The method that we developed incorporates common laboratory implements and greatly reduces anesthesia time, user error, and tracheal damage. Mice anesthetized with Isoflurane are placed supine on the crook side of the vertical leg of an L-shaped plexiglass support (fabricated in-house, each leg 2” x 5”). The animal is restrained with a rubber band placed beneath the animal’s incisors and around the support. Trans-illumination is accomplished by aiming a horizontal microscope light at the mid-tracheal level. The animal’s tongue is held aside with a pair of forceps shielded with PE tubing and the tongue is flattened against the lower jaw with the bent end of a small weighing spatula. The backlit larynx is easily visualized and a 20-gauge, 1.25-in. Teflon i.v. catheter is inserted into the trachea. The animal is placed supine by rotating the plexiglass support 90 degrees about the vertex and endotracheal tube placement verified by presence of condensation on the tube and actuation of the lungs with a small disposable pipette. The animal is removed from the apparatus, and the endotracheal tube attached to a ventilator. With this technique, the animals move from induction chamber to supplemental ventilation and anesthesia very rapidly and with minimal equipment. Thus, the window for hypoxia and laryngeal trauma is reduced, addressing Refinement (3 R’s). As of May 2007, this technique has been employed by the surgical team for nearly 4 years, and has increased survivability by approximately 50%. The nature of this technique requires both mouse handling and anesthesia familiarity, and thus is ideally taught to an experienced technician. Additionally, this technique may be more easily taught with the use of injectable anesthesia, reducing the input required to maintain a surgical plane as the surgeon develops mechanical proficiency. Survival animal models requiring thoracotomy, by definition, depend on a reversible route of mechanical ventilation. Most animal subjects lend themselves easily to intubation: visualization of the larynx is easily accomplished in animals even as small as rats, and larger airways facilitate blind techniques. Mice, however, prove more difficult to intubate. Not only is visualization of the larynx difficult even when backlit, but a long airway, large, thick tongue, and light weight of the animal (confounding handling and airway management efforts) further complicate matters. In a model using strictly inhalant anesthesia (for scientific reasons, in our case), the difficulty of the task is compounded by the brief window of residual unconsciousness between induction and assisted ventilation. With several surgeons operating a complicated model of coronary occlusion/reperfusion, we required a novel approach to rapid, accurate mouse intubation that did not require specialized peripheral equipment. This streamlined technique has several benefits. The animals move from induction chamber to supplemental ventilation and anesthesia very rapidly. Thus, the window for hypoxia and laryngeal trauma is reduced, addressing Refinement (3 R’s). As of May 2007, this technique has been employed by the surgical team for nearly 4 years, and has increased survivability by approximately 50%. The procedure also requires minimal equipment, making it cost-effective and seamless to integrated into the surgical station. The biggest challenges with this technique revolve around the dexterity and experience required to perform it with predictable accuracy. The nature of this technique requires both mouse handling and anesthesia familiarity, and thus is ideally taught to an experienced technician. This technique is more easily taught with the use of injectable anesthesia, reducing the input required to maintain a surgical plane as the surgeon develops mechanical proficiency. Introduction Abstract Materials and Methods Discussion An Accessible And Humane Approach To Mouse Intubation E Tonsfeldt, DL Hickman, DM Van Winkle An Accessible And Humane Approach To Mouse Intubation E Tonsfeldt, DL Hickman, DM Van Winkle 12 All procedures were approved by the Institutional Animal Care and Use Committee of the Portland Veterans Affairs Medical Center prior to development. Male and female C57BL/6J and DBA/2J mice 4-52 weeks old are anesthetized with Isoflurane and placed supine on the crook side of the vertical leg of an L-shaped plexiglass support (fabricated in-house, each leg 2” x 5”) (fig.1). The animal is restrained with a rubber band placed beneath the animal’s incisors and around the support (fig. 2). Because the animals are ventilated immediately following intubation, this procedure is performed at the surgery station. The ventilator (Inspira, Harvard Apparatus Inc., South Natick, MA) is primed prior to induction. Our intubation tubes are slightly modified 20-gauge, 1.25-in. Teflon i.v. catheters: the catheters are slightly bent in a slow arc to maintain the view when inserted, and the stylets dulled with a mechanical grinder to prevent laryngeal trauma. Adequate lighting is essential for this procedure, and both surgical lights and microscope lights are employed to provide thorough illumination. Trans-illumination is accomplished by aiming a horizontal microscope light at the mid-tracheal level (fig. 2). The animal’s tongue is held aside with a pair of forceps shielded with PE tubing and the tongue is flattened against the lower jaw with the bent end of a small weighing spatula. The backlit larynx is easily visualized and intubation tube inserted into the trachea. When further magnification is needed for adequate visualization, a dissecting scope (already at the station) or ophthalmic loop is used to obtain an effective view (fig. 3). Once the tube has been placed, the animal is placed supine by rotating the plexiglass support 90 degrees about the vertex. Endotracheal tube placement is verified by presence of condensation on the tube and actuation of the lungs with a small disposable pipette. When placement has been verified, the endotracheal tube is attached to the ventilator. When appropriate ventilation and an adequate plane of anesthesia have been attained stable, the rubber band beneath the animal’s incisors is cut and the apparatus gently removed from beneath the animal. Fig. 3 Fig. 1 Fig. 2