Anesthesia and Analgesia Recommendations

The investigator should determine the best option for a particular study and include it in their proposal to the IACUC. Veterinary consultation is required when planning any potentially painful study.

It is not necessary or advisable to withhold water before anesthesia and surgery in any species. If water is withheld, it should be limited to one (1) hour.

Any deviations from these procedures must be stated in the protocol and approved by the IACUC..

Supplemental administration of warmed fluids (lactated Ringer’s solution or isotonic saline) and maintenance of body temperature may improve anesthetic recovery.

Rodent Anesthesia Monitoring: Anesthetic monitoring of small rodents includes testing of rear foot reflexes before any incision is made, and continual observation of respiratory pattern, mucous membrane color and responsiveness to manipulations throughout the procedure. It is recommended that rectal temperature and heart rate are monitored electronically if possible during long or involved procedures. Monitor the rodent continually, documenting findings every 15 minutes and note the following:

Toe pinch method: The toe pinch method to evaluate depth of anesthesia is useful but not enough in itself. One must use two fingers and give the toe/foot a good squeeze. If there is no withdrawal reaction, the animal is judged deep enough to commence surgery. Remember that after this has been done the fingers are not sterile anymore. A sterile gauze pad may be used to protect the sterile gloves. Alternatively, a hemostat may be used to squeeze toe/foot. In this case, one must be careful not to squeeze too hard. Remember that after the hemostat has been used to squeeze toe, it is not sterile anymore and must not be used for surgery.

Respiratory pattern: Anesthesia will cause a distinct slowing of respiratory rate (RR). The surgeon must evaluate if RR becomes too slow and the anesthesia needs to be lightened and if the depth of respiration becomes too shallow. Increasing RR indicates the need for supplemental anesthesia.

Mucous membranes (MM):MM are evaluated by the color of the pinnae (ears) and toes. If these become bluish this is an emergency, indicating that the animal does not have enough oxygen. Pink is good and red MM usually indicates that the animal is too warm. This is not likely to occur during surgery but may occur during recovery from anesthesia, especially if a heat lamp is used to keep the animal warm. In such a case, the animal recovering from anesthesia must be protected and the lamp moved.

Reaction to surgical manipulation:If the animal makes any kind of move in response to incision or manipulation of organs, surgery must be temporarily stopped and anesthesia supplemented
Injectable anesthetics are typically administered by the intraperitoneal (IP) route. Injectable analgesics and reversal agents are often administered by the subcutaneous or the IP route. Intramuscular (IM) injections must generally be avoided because of the small muscle mass in this species. Diluting drugs in sterile saline solution will make it easier to accurately measure volume for injection.
Safe doses of ketamine-xylazine and ketamine-dexmedetomidine combinations produce limited-duration surgical anesthesia that may be insufficient for major surgical procedures in some strains of mice. These combinations tend to produce more reliable anesthesia in rats. Redosing xylazine or dexmedetomidine is not recommended due to cumulative circulatory and cardiovascular effects. If an animal becomes too light during a surgical procedure, isoflurane may be used supplementally or ketamine may be re-dosed at half the original dose. Partial reversal of the xylazine or dexmedetomidine by using atipamezole is possible, and will restore cardiovascular status more quickly. See Dissociative Anesthetics, found in the Commonly Used Anesthetics and Analgesics section of this Policy.

Pain Control: Mice are nocturnal animals, and are frequently housed in groups of nearly identical animals. These two factors make diagnosis of mild to moderate pain challenging. Weight loss is frequently monitored in animals at risk for ongoing pain. Pre-emptive treatment of pain before signs of pain are obvious is strongly recommended.

Anticholinergics can also be administered during surgery in to correct bradycardia that occurs at the desired anesthetic plane. However, the first step in treating persistent bradycardia during anesthesia is normally to turn down the primary anesthetic if it can be titrated (inhalant or CRI) or reverse components that can be reversed, as this often means the animal is too deep.

Anticholinergics are not routinely used in rodents. Atropine, the older drug, is shorter acting and has more side effects, so glycopyrrolate is the recommended agent. In rabbits, glycopyrrolate is preferred, because many rabbits have atropinase, which metabolizes atropine making it inactive.

Isoflurane and Sevoflurane

Standard inhalant anesthetics for laboratory animal use are either isoflurane or sevoflurane, delivered to effect in concentrations of 1-3% in oxygen (up to 5% for initial induction), using a precision vaporizer.

Advantages: Advantages of inhalant agents include rapid induction and recovery, with the ability to precisely titrate the level of anesthesia.

Disadvantages: Disadvantages include the cost and logistics of using precision vaporizers, occupational exposure concerns, and the risk of fatal over-dosage if an open system is used instead of a precision vaporizer. In addition, once animals awaken from gas anesthesia, there is no residual analgesic activity.

Pre-emptive analgesia is mandatory with isoflurane and sevoflurane anesthesia. This is because the moment the animal recovers from anesthesia, which occurs rapidly with gas anesthesia, there is no pain control, unless it has been administered before surgery. Effective pre-emptive analgesia will also decrease the dose of anesthetic required to maintain a surgical plane.

Occupational safety is a serious concern. Inhalants must be directly vented out of the room, or (less reliable), adsorbed in f-air charcoal canister filter. Filters must be weighed and replaced before they reach target weight (usually an increase of 50 gm). Environmental Health and Safety (EHS) can assist in evaluating potential exposures.

Nitrous Oxide (N2O)

May be used 50:50 or 60:40 with oxygen as carrier gas for inhalant anesthetics such as isoflurane. Nitrous oxide is not acceptable as sole anesthetic agent for surgery, but it can be used to lower the required dose of inhalant. Occupational exposure is potentially dangerous so direct venting is required (charcoal filters do not absorb nitrous oxide).

Other inhaled agents

Other inhalants may only be used when specifically approved by the IACUC in the animal use protocol.

Ether is an irritant as well as a fire and explosion hazard, so it is not recommended as an anesthetic agent.

Ketamine & Tiletamine

Ketamine is a widely used anesthetic in a variety of species. In low doses, ketamine provides chemical restraint with some somatic analgesia (no visceral analgesia). In higher doses, it may provide short-term surgical anesthesia in some species. In most instances, ketamine is used in combination with other injectable agents. Tiletamine is similar to ketamine; it is primarily used in combination with zolazepam as the drug Telazol.

Advantages of ketamine: Advantages of ketamine are its wide margin of safety in most species and its analgesic action. In combination with other drugs, it can provide surgical plane of anesthesia for about one half hour.

Disadvantages of ketamine: Disadvantages of ketamine include some irritancy due to low pH, note that ketamine has an acidic pH of about 4 and stings when administered IM. If injected near the sciatic nerve in the gluteal muscles in rodents and rabbits it may cause neuronal damage evidenced by the animal losing sensation in the hind leg and self-mutilating.

It is not sufficient for anesthesia by itself, but is extensively used in NHP for non-painful procedures where the animal needs to be restrained, such as blood collection, Tb testing and physical exams. The sole use of ketamine in other species is discouraged, as the recovery may be very stressful and agitated. Ketamine is a Class III controlled substance and subject to regulatory requirements governed by the Drug Enforcement Agency (DEA).

Telazol is tiletamine-zolazepam, a dissociative-benzodiazepine combination.

Advantages of Telazol: A low volume of injection is required. Like ketamine combinations, it can occasionally produce short-term anesthesia, though rarely of sufficient depth for surgery. It is more useful as an induction agent prior to general inhalant anesthesia, or for chemical restraint for short non-surgical procedures.

Disadvantages of Telazol: Telazol should only be used for 7 days after reconstitution if stored at room temperature or 56 days after reconstitution if refrigerated. It must be stored in a lock box in either case since it is a Class III controlled substance. Never use telazol if the solution is not clear. It is not safe for use in rabbits (kidney damage).

Ketamine Combinations

Ketamine+ alpha2-agonists (Xylazine or dexmedetomidine): Αlpha-2 agonists are centrally acting anesthetics, with excellent analgesic properties. At lower doses they induce sedation. Their most common side effects are bradycardia and cardiac arrhythmia. When combined with ketamine, these properties are minimized and the combination of ketamine with the α2-agonists xylazine or dexmedetomidine in the same syringe produces a surgical level of anesthesia in many species, notably rodents and rabbits.
In large animals the combination of ketamine-alpha2-agonists may be appropriate to restrain the animal prior to inhalant anesthesia.

Advantages: Advantages of ketamine-a2-agonist combinations are that they may be combined in one syringe; that they may produce short-term surgical anesthesia with good analgesia, and that recovery can be hastened by reversing the a2-agonist with atipamezole. Administration of reversal agents may be useful but is not routinely recommended as the recovery may be agitated (see notes on ketamine as a sole agent) and analgesia is reversed along with sedation. .

Disadvantages: Disadvantages of ketamine-alpha2-agonist combinations are that they will not reliably reach surgical anesthesia in all cases, and that they can cause profound cardiac depression. They may cause vomiting, especially in cats.

Caution for use: if a ketamine-a2-agonist combination is used for surgery longer than 20 minutes, animals will likely require additional anesthetic. Redosing with ketamine alone rather than the combination may be safer, as the cardiovascular depression of a2-agonists is often longer-lasting than the sedation or analgesia produced.

Adding acepromazine to the ketamine-a2-agonist combination will result in deeper and/or longer plane of anesthesia in small rodents, especially rats, and possibly some strains of mice as well. Dose with caution because acepromazine also decreases in heart rate and blood pressure but is not reversible.

Ketamine+ benzodiazepine (midazolam or diazepam): ΑKetamine may be combined with the benzodiazepines—midazolam or diazepam —to produce a deep level of sedation. In most cases, this sedation will require an inhalant agent or other anesthetic to achieve surgical anesthesia. In most applications, midazolam is preferred, as it can be injected intramuscularly; intramuscular injection of propylene glycol (the carrier in injectable diazepam) can cause painful, sterile abscesses and is discouraged.

Advantages: Advantages of ketamine-benzodiazepine combinations are that they may be combined in one syringe and will produce deep sedation with moderate analgesia as well as amnesia. Recovery from ketamine-midazolam is often smoother than recovery from ketamine alone. Benzodiazepines, particularly diazepam, raise the seizure threshold and are used to treat seizures. .

Disadvantages: Disadvantages of ketamine- benzodiazepine combinations are that they will not reliably reach surgical anesthesia in most cases. Diazepam should be restricted to intravenous or intraperitoneal use when possible. Benzodiazepines are DEA Class IV controlled substances. Tolerance to benzodiazepines can occur quickly, so repeated doses may need to increase over time. Due to liver CP450 metabolism of benzodiazepines, use caution with other drugs that may inhibit similar hepatic microsomal enzymes and may lead to prolonged sedation.
Telazol is a dissociative-benzodiazepine combination.

Advantages: alpha2-agonists are that they produce profound analgesia of short duration, can be combined with ketamine (and in rodents, acepromazine) to produce deeper anesthesia, they are not controlled substances, and they are reversible with atipamezole (IP/SC in small rodents, IM in larger species, not IV). They are not irritant when injected via intramuscular or intraperitoneal routes.

Disadvantages: Disadvantages in most species include cardiovascular depression (decreased heart rate, decreased cardiac output, and hypotension), which is somewhat controlled by pre-treatment with atropine or glycopyrrolate. a2-agonists cause a transient hyperglycemia which may have research implications. Xylazine, in particular, often causes transient nausea and vomiting, but any alpha-2 agonist may do so in cats. Rapid IV administration of reversal agent has produced seizures in some species and is not recommended.

Caution for use: If a ketamine a2-agonist combination is used for surgery longer than 20 minutes, animals will likely require additional anesthetic. Redosing with ketamine rather than the combination may be safer. However, analgesia may not be adequate, as the cardiovascular depression of a2-agonists is often longer-lasting than the sedation or analgesia produced.

Disadvantages: If ketamine and alpha-2 agent have been used for anesthesia and the alpha2 agonist action is reversed, this may result in agitated recovery from anesthesia due to the ketamine (see disadvantages of ketamine as a sole agent). Reversal will quickly reverse analgesic effects of alpha-2 agonist, so additional analgesia should be administered prior to recovery if the animal has undergone a painful procedure. IV administration is usually not recommended because it may cause seizures or profound cardiovascular disturbances.

Sodium pentobarbital (Nembutal) is the most used barbiturate. Thiopental was historically used but is no longer available.

Advantages: Barbiturates do not depress cortical evoked responses to the extent that other anesthetics might. Animals do not feel pain when they are at a surgical plane of anesthesia. Once stable anesthesia has been achieved, it may be longer lasting than with most other injectable agents. Barbiturates are the most common of the injected euthanasia solutions, as they reliably produce unconsciousness before respiratory depression and death.

Disadvantages: Disadvantages of barbiturates include a narrow margin of safety, primarily associated with respiratory depression. Pain sensation is only decreased at surgical planes of unconsciousness, and may even be heightened (hyperalgesia) at subanesthetic doses. Larger animals may experience a distressful anesthetic recovery. Outside of the vein (perivascular, or intraperitoneal) injection of barbiturates can be irritating due to pH of ~11; barbiturates for IP injection should be diluted to a strength of 6 mg/kg. Barbiturates are Class II controlled substances, except for some Class III euthanasia solutions.

Advantages: Alfaxalone causes minimal cardiovascular and respiratory suppression, and it is short-lived. It has relatively smooth induction and recovery, even as a sole agent, good muscle relaxation, a wide margin of safety, and rapid elimination for safe repeated use.

Disadvantages: Action is short-lived, so it requires continuous or repeated administration. It may be inadequate as a sole agent for some procedures and species. It is a controlled substance (Schedule IV).

 

<strong>Advantages</strong>: Animals recover from propofol in minutes, even after prolonged administration.

<strong>Disadvantages</strong>: Propofol has minimal analgesia at sub-anesthetic doses. It can be a profound respiratory depressant, and may also cause hypotension. Because of its rapid elimination, it must be administered IV, and so is of limited use in small rodents. Once opened, propofol easily supports microbial growth, but some formulations are stabilized. Check packaging for how long it can be kept after opening.

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<strong>Advantages</strong>: Avertin may be easily administered by IP injection with good short- term surgical anesthesia, and it is not a controlled substance.

<strong>Disadvantages</strong>: Avertin is not commercially available in pharmaceutical grade, so it must be made from tribromoethanol and amylene hydrate. It can cause peritonitis in mice, and that risk increases with repeated use. Post-procedural analgesia has not been demonstrated, so using a preemptive or postoperative analgesic is required. Though surgical anesthesia is short (15-20 minutes), recovery is prolonged (~40 minutes), so the animal must be kept warm during that time.

<strong>Cautions for use</strong>: Avertin must be carefully prepared in the laboratory under aseptic conditions. Working with a dilution of 1.25% is recommended – this is best prepared fresh for use, or stored no more than one week. It degrades quickly with storage and exposure to light. Degradation products are toxic and potent GI irritants. Where possible, IACUC recommends inhalants, such as isoflurane, replace Avertin.

Buprenorphine is a longer-acting partial μ agonist (weak κ antagonist) and is good for most post-operative applications. There are also sustained or long-acting formulations that can provide several days of analgesia. However, buprenorphine has a ceiling effect and binds more strongly to μ receptors than pure agonists, so it can displace shorter-acting, more potent opioids leaving residual pain if a procedure or condition is very painful. This strong binding affinity is also why buprenorphine is sometimes used to reverse or dampen the activity of pure μ agonists (overdose) and why it cannot be reversed with opioid receptor antagonists, like naloxone. Butorphanol may be more efficacious than buprenorphine for birds and for cats. Butorphanol is a mixed κ/ μ receptor agonist/antagonist. Both buprenorphine and butorphanol produce a less profound respiratory depression than full agonists and exhibit a ceiling effect with increased doses.

Administered as a part of the anesthesia regimen as pre-emptive analgesia, opioids enable the anesthetist to lower the dose of other anesthetics and contribute to the comfort and safety of the patient. Opioids are most often administered by injection. Oral use is effective, but requires much higher doses because of “first-pass” liver metabolism when absorbed from the gut. Bioavailability varies by agent, route of administration and species.

<strong>Advantages</strong>: Opioids are potent analgesics. Concurrent use with inhalant or parenteral anesthetics for general anesthesia will lower the required dose of the anesthetic.

<strong>Disadvantages</strong>: Opioids can suppress respiration (more marked effect in fentanyl and morphine than in buprenorphine). Opioids may increase locomotor activity, and buprenorphine at high doses may cause pica (abnormal ingestion of non-food items such as bedding) in rats. Alternatively, they may sometimes cause sleepiness and slower recovery from general anesthesia. Fentanyl has a very short duration of action in most animal species and must be administered as an IV drip. <strong>Most opioids are controlled substances. Morphine, oxymorphone and fentanyl are Schedule II. Buprenorphine is Schedule III. Butorphanol are schedule IV.</strong>

<strong>Cautions for use</strong>: Buprenorphine has found favor as the longest-acting opioid analgesic. However, this duration of action is closer to 6 hours in most situations than it is to 12 hours. 12 hours is the absolute maximum dosing interval for use of buprenorphine for post-procedural pain. Simbadol™ (high concentration), Ethiqa XR™ (extended-release) and Buprenorphine SR-LAB (sustained-release) are buprenorphine formulations with extended durations of action, which have been developed and tested for analgesic efficacy in different species. Refer to veterinary staff or Huron Standard Procedures for up-to-date recommendations on these products to provide effective analgesia while reducing post-operative stress from repeated restraint and injection.

Pure mu receptor agonists (morphine, fentanyl, and oxymorphone) are not recommended in swine and should be used cautiously if at all in cats. This is because these drugs may cause excitement in these species. However, cats and swine do well with partial mu and kappa receptor agonists (buprenorphine and butorphanol).

<strong>Advantages</strong>: Carprofen, ketoprofen, ketorolac, and meloxicam may have duration of analgesic action up to 24 hours. They may be used concurrently with anesthetics, with opioid analgesics, and with

<strong>Disadvantages</strong>: NSAIDs may decrease clotting ability, of possible concern following surgery. Gastric upset and even ulceration may occur, especially with prolonged use. Prolonged use carries the risk of kidney or liver disease.

<strong>Cautions for use</strong>: Cats are particularly susceptible to toxic effects of NSAIDs. <strong>Never administer acetaminophen to cats.</strong> Other NSAIDs should be used only at the dose and frequency recommended.

Undesired side effects are more likely with increasing duration of usage – for most situations, limit use of NSAIDs to 3-4 days per animal, except under veterinary supervision. Do not use in dehydrated animals, or in animals with kidney or liver dysfunction.

Lidocaine cream (EMLA or ELAMax) is used topically on shaved or hairless skin, though it requires 30-60 minutes or more of contact with skin to reach full effect. It’s not generally required, but has been used historically to reduce discomfort from venipuncture.

Tricaine methanesulfonate (MS-222) is a related compound used as a general anesthetic for fish and frogs. Cetacaine is another similar agent used to reduce laryngeal spasm at the time of intubation in larger species.

<strong>Advantages:</strong> Intra-operative use can augment the pain relief of general anesthetics, and reduce the need for frequent redosing. Bupivacaine can augment the post-operative analgesic action of opioids and/or NSAIDs. They are not controlled substances. At appropriate doses, they have minimal cardiovascular effect.

<strong>Disadvantages</strong> Intramuscular and intravenous injection should both be avoided unless lidocaine is being used specifically for anti-arrhythmic effects. Systemic toxicity (including seizures and death) can result from over-dosage (more likely to occur with smaller subjects) and with accidental intravenous injection. Lidocaine may sting when first injected.

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Nonhuman Primate Formulary

Rabbit Formulary

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Swine Formulary