Mucous membrane color: Should be pink, not blue or gray. 2009 Jun; 22(3):368-73. Recover the neonate by holding it in your gloved hand or placing it in a Petri dish on a warm water circulating heating pad or under a heat lamp for less than 5 minutes and not too close to the lamp watching carefully to avoid overheating. 5. Facility not large enough to hold your expanding transgenic colony? Due to the small respiratory capacity in mice, use a non-rebreathing system. Induction time: 2-4 minutes. The ice can be packed flat or a frozen block of ice covered with a thin lining can be used. Administration of local anesthetics prior to the painful stimulus (e.g., incision) would be considered an adjunct analgesic to opioid and NSAID analgesics. or greater, immersion in liquid nitrogen may be used only if preceded by anesthesia. Acetaminophen (0.1 ml PO of Children’s cherry flavored Tylenol Elixir) at 80 mg/2.5 ml once or twice a day may be acceptable and can be administered using a dropper or TB syringe.1, Boston University Research Support Cardio-pulmonary resuscitation and drugs to reverse anesthesia must be given or animal will die. Intramuscular (IM) injections are not recommended in mice as it can result in complications such as tissue irritation, lameness, and self-mutilation. Example: A mouse weighing 25.0 grams receives 6 microliters of the above anesthetic mixture per gram body weight for a total of 0.150 mL of anesthetic mixture injected intraperitoneally. The physiologic effects of isoflurane anesthesia in neonatal mice. E-Z Anesthesia http://www.ezanesthesia.com/ez_anesthesia/index.html. Mouse, Rat and Hamster Neonates over 10 days of age: Follow guidelines for adults. The effects of neonatal isoflurane exposure in mice on brain cell viability, adult behavior, and memory. A precision vaporizer provides the ability to precisely titrate the level of anesthesia during a procedure. Routes of administration include topical to mucous membranes (nose, eye, etc. Go to “Breathing Devices” EZ-103A Microflex Breather. University Animal Care Committee (UACC). If crushed ice is used, it must be carefully packed, to provide close contact between ice and pup. Returning neonates as a group (the entire litter) is best to minimize disturbing dam. Induction: 6.1.4.1. Lidocaine has a quick onset (1-2 minutes), but is short-acting (1-2 hours). Immerse pup in ice water or crushed ice for 3 to 4 minutes. Once the measuring unit has heated to the temperature, remove the mouse pup from the cage and wipe the thorax with 70% ethanol sprayed on a wipe. Isoflurane is the preferred inhaled anesthetic. Neonates must receive analgesia if the procedure can reasonably be expected to result in enough pain to interfere with the neonatal function of sleeping and suckling. Exceptions may be considered, if the disturbance of neonate and dam in order to administer the medication interferes more than warranted with these functions. Fur is now thicker; Females may show nipples (there are five pairs of mammories) Day 10. Combined ketamine/xylazine is the preferred injectable anesthetic in mice. McGill University. The choice of anesthetic agent will depend on the procedure to be performed, research aims, and other factors such as animal age; there are special considerations for neonatal mice. IACUC oversee... Approved January 7, 2011, Reviewed July 2019. Do not use incandescent light during the procedure, as it can warm the surgical field and cause animals to awaken from a surgical plane of anesthesia. compmed@umn.eduVeterinary Services Office: 612-624-9100Business Office: 612-624-6169, 420 Delaware St. SEMMC 351 (Campus Delivery Code)Minneapolis, MN 55455, Institutional Animal Care and Use Committee (IACUC)eProtocol (sign-in required)Institutional Biosafety Committee (IBC)Research Occupational Health Program (ROHP), Helping researchers external and internal to the University of Minnesota find services, facilities, and equipment, Research Ethics at the University of Minnesota. Gas anesthesia is recommended for longer, more invasive procedures and in neonates older than 5 days. For rodent use, dilute 1‐2% lidocaine to 0.5% and 0.5% bupivacaine to 0.25% to allow for feasible volumes to infuse at the incision site (1% solution is equal to 10 mg/mL). . Anesth.Analg. Parenteral agents are not advisable in very young animals. DJ Kohn, SK Wixon, WJ White, GJ Benson, Eds., Academic Press, 2008. Bupivacaine has a slow onset (10-15 minutes), but is long-acting (4-8 hours). Tattoos are permanent and will not alter the physiology of your animals. All procedures must have prior approval from the Institutional Animal Care and Use Committee (IACUC). Neonates are defined as mouse or rat pups up to 10 days of age. Gaertner et. A one-week acclimation period is recommended for all animals to prevent stress-induced disease. Consult your area veterinarian with questions about drug selection. RAT AND MOUSE ANESTHESIA AND ANALGESIA Formulary and General Drug Information Approved: March 2nd, 2016 Definitions General Anesthesia: Loss of consciousness in addition to loss of sensation Analgesia: Loss of sensitivity to pain. Allow the mouse to acclimate to the surface in the dark for approximately 2-5 minutes. Continue to monitor animals until they are fully recovered. Hoff is a training technician at the University of Michigan, 1150 W. Medical Center Dr., 018 ARF, Ann Arbor, MI 48109-0614. Guidelines: Consider providing warm subcutaneous (SQ) or intraperitoneal (IP) fluids, particularly for prolonged anesthetic events or animals that are ill, aged, or debilitated. Belly begins to show fur; Day 9. Analgesic Agents for Neonatal Rats and Mice, Administer a lower dose then would be administered to adult animals to start with. Guidelines: Contact your area veterinarian with questions or for additional information. Hypothermia allows less control of anesthetic depth compared to inhalant anesthesia. One Silber Way, 9th floor, Boston, MA 02215. Expect the birth of one pup every 15–30 ... (the latter is preferred if hypoxia alters the parameters to be tested in the mouse tissue) after delivery of her first pup. Find guidelines for mouse anesthesia, including anesthetic drugs and procedures and special considerations for neonatal mouse anesthesia. Find services, facilities, and equipment for researchers. Gentle handling for 5-minute intervals a few times a day. Duration of anesthesia. General guidelines and considerations for mouse anesthesia. PP.277-278. tribromoethanol Medicine & Life Sciences A 5-year-old male-castrated Cocker Spaniel presented to the Veterinary Teaching Hospital of the University of Georgia for a total ear canal ablation. Anesthesia: Pentobarbital Sodium The mice may be anesthetized with 65 mg/kg body weight This can lead to corneal drying and trauma. These opioids have been shown to be safe and effective analgesics in neonatal rodents.3. Inhalant anesthesia may be safe both for ultra short (<5 minutes=”” and=”” prolonged=”” greater=”” than=”” 6=”” hours=”” procedures=”” for=”” longer=”” periods=”” of=”” anesthesia=”” it=”” may=”” be=”” necessary=”” to=”” ventilate=”” the=”” animal=”” sup=””>2, 6, 7. Isoflurane: 3-4% for induction and 1-3% for maintenance. Make sure neonate is warm and pink and breathing and capable of spontaneous movement before returning it to dam. MOUSE ANESTHESIA SOP 110.05 - Mouse Anesthesia Page 1 of 4 1. Mouse, Rat and Hamster Neonates < 10 days of age. Parental cannibalism is a problem with neonatal rodent anesthesia. Experimental procedures on animals often require anesthesia and/or analgesia to obtain adequate immobilization and to reduce stress or pain. Animals should not be placed directly on ice (use latex glove or another substance between the animal and the underlying ice bath). The use of dry ice for hypothermia-induced anesthesia is prohibited. 2006, Jan; 102(1):75-80. The ribs and skin are closed using 7‐0 sutures, and the pup is rapidly rewarmed with a supply of oxygen. The anesthetic event lasts longer than five minutes. 2. Recover neonate on home cage/dam bedding so as to make it smell similar to what it did before removal from cage. We are committed to protecting research participants, upholding ethical standards, and improving our practice at every step of our work. Recover animals on paper towels (without bedding) in a clean cage. Animals have reached the proper plane of anesthesia when pedal reflex is lost. The described anesthetic device for stereotaxic operation on mouse pups consists of polydimethylsiloxane (PDMS). Valentim A.M., Guedes SR,Pereira AM, Antunes LM. The surgical procedure is expected to take 10 to 15 min per pup, with a further 5 min for recovery. Eds. 6.1.4. al. Immerse container holding pup in ice water or place on a barrier such as a glove or other water-proof material over crushed ice. BU IACUCInstitutional Animal Care and Use Committee Mice vary significantly in their sensitivity to various anesthetics. Per NIH guidelines, decapitation alone for this age group is an acceptable means of euthanasia, no confirmation required Cotton balls scented with various chemicals to be used (betadyne, isopropyl alcohol, chlorhexidine etc.) What protocols for P0 mouse pup fixation work best to resolve CNS immunohistochemistry? 2007. Eyes are open; Pups begin to nibble solid food; Day 13. Feb. 2008. When adding anesthetics to your protocol, it is advisable to provide a range to allow titration for the specific needs of each animal. Pre-anesthetic fasting is usually not necessary for mice due to their inability to vomit. MOUSE ANESTHESIA AND ANALGESIA RECOMMENDATIONS 2 Non-painful procedures (e.g., imaging, restraint) Recommendations include: 1. Hypothermia is appropriate for short, minor surgical procedures (5-15 minutes) in neonatal rats or mice up to 5 days of age. Pre-anesthetic fasting is usually not necessary for mice due to their inability to vomit. Dose: 4-5% for induction; 1-2% for maintenanceComments: 300μL in a 500ml container for drop jar anesthesia. Apply ophthalmic ointment (e.g., Paralube® or Lacrilube®) to eyes if: Regardless of the anesthetic administered, monitor mice under anesthesia to avoid excessive depression of cardiac and respiratory functions, or insufficient anesthesia. View the latest updates for researchers or visit our recovery toolkit. water-proof material covered ice cube or cold glass plate or a frozen ice block) to provide more constant level of hypothermia. Colored fuzz starting to cover pup; Day 8. xylazine 10-12.5 mg/kg IP. See how the Office of Research Compliance (ORC) provides support for institutional committees and Marquette faculty, staff and students for research involving human subjects, animal subjects, radioisotope use and research with recombinant DNA (rDNA). Advanced Research and Diagnostics Laboratory, Services for Researchers External to the University, Research & Support in the health sciences, special considerations for neonatal mouse anesthesia, IACUC guidelines on using hypothermia as anesthesia for neonatal rodents, Institutional Animal Care and Use Committee (IACUC), Research Occupational Health Program (ROHP). Immerse the container holding the pup into crushed ice or place the pup on a barrier, such as a glove or other water-proof material over crushed ice. Loepke, AW et.al. Protect pup in a glove or paper-lined tube to avoid damage to the skin. Please use the IACUC Anesthesia and Surgical Monitoring Form for Rodents or a substantially similar form. When the animal is ambulatory, return it to the home cage with immediate access to food and water. IP When used in combination (lidocaine plus bupivacaine in the same syringe), local anesthesia can have a fast onset with a relatively long duration of action. References 1. However, if fasting is employed, limit it to no more than two to three hours due to the high metabolic rate of small rodents. If additional anesthetic is needed, supplement with 1/3 dose of ketamine only. Am J Physiol Heart Circ Physiol 278: H652-H657. Or use the EZ-103A Microflex Breather and shave off a small amount to shorten the mask to fit the neonate. Do not re-dose xylazine. It is extremely important to accurately weigh the animal and calculate the correct dose. Teeth are beginning to erupt; Eyes start to open; Day 12. Parenteral agents are not advisable in very young animals. ** Anesthesia and Analgesia in Neonatal Mice and Rats. Animal Housing and Special Services Process: Submitting an ISR, Research Occupational Health Program (ROHP), Facilities and Administrative Costs and Analysis, Research Information Management System (RIMS), Find your Department and Research Administrators, Advisory Council for Research Administration (ACRA), Grant Management Training Videos and E-Learning, PAFO Onboarding Pack for Department Administrators, Information Systems and Web-Based Applications, Anesthesia and Analgesia: Neonatal Mice and Rats, Center for Antiracist Research Request For Proposals, Limited Submission Funding Opportunity: NEH American Rescue Plan: Humanities Organizations, Novartis Institutes for BioMedical Research Global Scholars Program 2021. 2. Ketamine/xylazine: intraperitoneal dose at 70-100 mg/kg ketamine + … Hypothermia Method: Mouse and rat pups up to 6 days of age may be anesthetized by hypothermia when inhalant anesthesia is not feasible. Rapid warming can cause tissue damage. Induction: 6.1.4.1. Guinea Pig Neonates: Follow guidelines foradults. Only for mice under 6 days of age Mouse pup must not be placed in direct contact with ice-water slurry Opioid analgesia Recommended: Buprenorphine 0.05 - 0.1 SC or IP For pain management after surgeries. If isoflurane anesthesia has been used, it may help to accustom dam to the smell for 1-2 days before surgery on neonate (see Preparation of Dam). Find guidelines for mouse anesthesia, including anesthetic drugs and procedures and special considerations for neonatal mouse anesthesia.Research Animal Resources (RAR) veterinary staff designed these guidelines to be general recommendations, not an inclusive list of all possible drug combinations that can be used in mice. Stage 4 — medullary paralysis with respiratory arrest, hypotension and imminent death. The blood collection method, interval between blood collection procedures, and volume of blood to remove must be listed in the approved protocol specific Anesth.Analg. Additional steps include exposing pups to soiled bedding from the mother’s cage, and placing the pup in the middle of the litter. Developmental changes of cardiac function and mass assessed with MRI in neonatal, juvenile, and adult mice. Acceptable Methods of Euthanasia Overdose of chemical anesthetics ( 2-3 times the anesthetic dose) Decapitation NOTE: you do not have to use CO 2 first. The vaporizer must be compatible with the specific inhaled anesthetic agent. Mice are particularly susceptible due to their high body surface area to body mass ratio. Before surgery start, the patient’s systoli… February 2000. Ketamine/xylazine* ketamine 80-100 mg/kg IP. The literature currently describe several anesthetic methods for use in neonatal rodents, including injectable anesthesia, inhalant anesthesia, and hypothermia. Oxygen saturation and heart rate: Can be monitored using a specialized rodent pulse oximeter, and should be greater than 95% and between 300-800 beats/minute, respectively. : for a 25 gram mouse, the maximum volume would be 0.25 ml Rat 23‐25g < 10 ml/kg i.e. Protect pup in a glove or paper-lined tube to avoid damageto the skin. Int. • Day 10 pups were anesthetized and stabilized in a non-invasive manner and successfully injected using the PDMS device. When using inhalant anesthesia, use a fume hood or an anesthetic system equipped with a gas scavenging system to minimize occupational exposure to exiting gases. The following doses are general guidelines that may vary significantly based on the aforementioned factors. Use as a primary analgesic is discouraged due to the short duration (hours). Dose: 1-2 mg/kg (0.4-0.8 ml/kg of a 0.25% solution)Route: Local infiltrationComments: Do not exceed 6 mg/kg total dose. Fur growth is complete; Pups are more active; Day 11. 26(1): 87-92. PURPOSE This Standard Operating Procedure (SOP) describes methods for anesthetizing mice. Dose: Ketamine: 80-120 mg/kg; Xylazine: 5-10 mg/kgRoute: Intraperitoneal (IP)Duration: 30-45 minutesComments: Thermal support is crucial. A thin material such as a latex glove, works better than a thicker material such as a neoprene glove. A funnel-shaped cone that makes an excellent anesthesia mask by cutting down on in-mask dead space without restricting access to the eyes or cranium. Ketamine/xylazine cocktail* KX mouse cocktail 0.1mL/20g mouse wt. PURPOSE This Standard Operating Procedure (SOP) describes methods for anesthetizing mice. Immerse pup in ice water or crushed ice for 3 to 4 minutes. Premedication was with carprofen 2.2 mg/kg SQ, hydromorphone 0.1 mg/kg IM, diazepam 0.2 mg/kg IM, and glycopyrrolate 0.01 mg/kg IM. Surface cooling is effective due to their small surface area and body size. 2 6. MOUSE ANESTHESIA SOP 110.05- Mouse Anesthesia Page 1 of 3 1. Fox, James G. et.al. Dose: 4 mg/kg (0.4 ml/kg of a 1% solution)Route: Local infiltrationComments: Do not exceed 7 mg/kg total dose. Observe pups closely during induction. During the recovery phase, turn off inhalant gas before oxygen. For more information contact veterinary staff for assistance. Continue to provide supplemental heat during recovery. Together they form a unique fingerprint. Consequently, guidelines do not reference specific 2 5. Anesthesia was induced by placing the pup in a syringe and adding a volume of isoflurane-saturated gas that produced an estimated level of 32% isoflurane. Parameters that can be monitored in an anesthetized mouse without specialized equipment include: All species are at risk for hypothermia while under anesthesia. Rapid onset, short duration. Research Animal Resources (RAR) veterinary staff designed these guidelines to be general recommendations, not an inclusive list of all possible drug combinations that can be used in mice. Repeated tail clips on a single mouse require the prior approval of the IACUC. Bertrand Mathon1, 2, Mérie Nassar2, Jean Simonnet2, Caroline Le Duigou2, Stéphane Clemenceau1, Richard Miles2, Desdemona Fricker2 1Department of Neurosurgery, Groupe Hospitalier Universitaire de La Pitié-Salpêtrière, APHP, F-75013, Paris, France Recover anesthetized animals alone in a cage. Never leave animal unattended during this time. Students can find additional information in the Undergraduate Student Guide and Graduate & Professional Student Guide. In Anesthesia and Analgesia 2nd Ed. FUp to P7 – No anesthesia/analgesia required FP7-12 – Anesthesia required FP13+ prohibited FFor ID purposes – justification required FAseptic technique – alcohol minimum FBleeding control – required FAs it has potential for pain/distress justification required P = Postnatal day Neonates are defined as mouse or rat pups up to 10 days of age. Protect pup by placing into finger of a glove or paper lined test tube. 2. * During the recovery phase, turn off inhalant gas before oxygen. Recovery period may be prolonged. Loepke, AW et al. mal, anesthesia is recommended when November 2000 Lab Animal Volume 29, No. Steps must be taken before, during, and after anesthesia to ensure animal safety and anesthesia efficacy. Gas anesthesia (isoflurane) or hypothermia may be used to anesthetize neonatal rats and mice. Dosage. Wiesmann, Frank et.al. Provide supplemental heat during all anesthetic events. Induction in a chamber generally requires 5% isoflurane, and maintenance (chamber or nose cone) requires 1-2% (flow rate = 0.5-1 L/min). Pups can be returned to dam once they are able to move without direct physical stimulation. Fingerprint Dive into the research topics of 'Refinements for embryo implantation surgery in the mouse: Comparison of injectable and inhalant anesthesias – tribromoethanol, ketamine and isoflurane – on pregnancy and pup survival'. Documentation is required for neonatal anesthesia, surgery and postoperative care. Local anesthetics are a type of injectable anesthetic. Only IP route of administration is allowed. 3. Wright-Williams SL, Courade JP, Richardson CA, Roughan JV, Flecknell PA. 2007. This minimizes the risk of tracheal obstruction or pneumonia. Induction of anesthesia with gas is typically achieved with < 2 min exposure to 3‐5% Isoflurane. Place neonate on warm pad or circulating warm water pad. SomnoSuite is our innovative low-flow anesthesia delivery system designed specifically for mice and rats. It may help with dam accepting the pups back after the procedure if all vestiges of blood and disinfectant are removed from the pups by gently wiping them with a wet gauze pad before returning them to the dam. Use an adult mouse facemask adjusted for neonate. Stage 3 — surgical stage of anesthesia, with loss of reflexes, muscle relaxation, deep and rhythmic breathing, planes 1-4 (light to deep). Exposure to isoflurane lasted 30 s. All the pups survived the anesthesia. Other commonly used injectable agents are listed below. Local anesthetics block nerve impulses by specifically binding the voltage‐gated Na+ channel in the nerve cell membrane. Hypothermia Anesthesia Age Altricial rodents up to 7 days old Induction 2-4 minutes. Very young hairless, altricial rodents function as poikilotherms and are resistant to brain damage associated with cephalic circulatory arrest. Return to top Pups must not come in direct contact with the cold source in order to prevent skin lesions secondary to frostbite. It has rapid and reliable onset and recovery. The patient was induced with lidocaine 2 mg/kg IV and etomidate 1 mg/kg IV and maintained with sevoflurane and a constant rate infusion consisting of lidocaine 0.05 mg/kg/min. When fitted in custom made flexible Bain-type circuits they are sometimes suitable for stereotaxic frames. Never place the animal directly on the heat source. References for these guidelines are available by request. Observe the pup closely. Hypothermia provides immobilization and mild analgesia (similar to local anesthesia resulting in nerve transmission blockage). This will avoid skin damage caused by direct contact with ice/cold water. Never restrict water. ), or an injection directly into tissues and around nerve bundles. Gas anesthesia (isoflurane) or hypothermia may be used to anesthetize neonatal rats and mice. Body temperature: Can be monitored with a rectal thermometer, and should be above 97℉. Neurosci. or gas anesthetic (isoflurane placed in cage for 1-2 minutes daily. Agent. Curr Opin Anesthesiolol. Unlike traditional vaporizers, the SomnoSuite is engineered with a precision syringe pump and integrated digital vaporizer which uses either room air or compressed gas to deliver anesthesia at low flow rates proportionate to the animal's size. Repeat administration is not advisable.Hypothermia must be avoided by keeping the neonate warmed as described above. Animals may be anesthetized for surgery, non-surgical procedures that may be painful, or non-painful procedures that require immobilization. Because the anesthetic concentration cannot be controlled in the chamber and can rapidly reach toxic concentrations, this method is reserved for very short procedures such as tail biopsies, subcutaneous tumor implantation, or induction prior to facemask anesthesia. J. Dev. Handling and olfactory conditioning of dam 7-10 days prior to birthing. Supplemental heat sources include circulating water blankets, electric heating pads, and commercial products that can be warmed in a microwave. Mouse eyes remain open under anesthesia. Jan; 108(1):90-104. Xylazine can be reversed with atipamizole (Antisedan®, 1-2 mg/kg SQ or IP), or yohimbine (Yobine®, 0.5-1 mg/kg IP). These procedures may help reduce dam cannibalizing or neglecting pups. Hypothermia allows less control of anesthetic depth compared to inhalant anesthesia. Neonatal rodents may have a longer induction time than adult rodents with inhalant anesthesia. Ketamine/xylazine is the recommended injectable anesthetic for mice >7 days of age. 10 TECHNIQUE Methods of Blood Collection in the Mouse Janet Hoff, LVT, RLATG The author outlines various meth-ods of blood collection from mice. Hypothermia for induction: Place the pup in a nitrile glove finger and immerse the glove finger in crushed ice and water (2-3 °C or 35-37°F) up to the level of the head so that the head of the pup is visible. Place the pup on the heated surface of the plastic. Sheldon, R. Ann et.al. Ketamine/xylazine cocktail may be considered for use in neonates but may not be as safe as hypothermia or isoflurane. 20-30 minutes. Neonatal Rodent Survival Surgery. Administer isoflurane at 1.5 – 2.5% in 100% oxygen. Following anesthesia, the animal should be re-warmed slowly. Consequently, guidelines do not reference specific research-associated concerns. old, general anesthesia and analgesia is required and only trained members should perform the procedure to ensure bone is not cut. Slower onset, longer duration. The mouse (Mus musculus) is currently the most popular laboratory animal in biomedical research. Pup can be kept on cooling platform (i.e. Advantages: Rapid induction and recovery. However, if fasting is employed, limit it to no more than two to three hours due to the high metabolic rate of small rodents. Anesthesia is being delivered by facemask. Once recovered, the pup is cleaned and rolled in a fecal bath to mask the smell of surgical interference before being returned to the mother. Starting with the lower dose range is recommended. Visit Back2BU for the latest updates and information on BU's response to COVID-19. Respiratory rate and pattern: The normal undisturbed rate is ~180/minute, a slow rate drop of 50% is acceptable during anesthesia.
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