Commonly used anesthetics and analgesics
3. Commonly used anesthetics and analgesics
Anticholinergics are routinely used in human surgery and are strongly recommended for laboratory animals. The routine administration of atropine or glycopyrrolate before or at the time of induction for general anesthesia will protect cardiac function, and especially prevent bradycardia and cardiac arrhythmias. Anticholinergics are not routinely used in rodents, but there is no reason to omit them. Atropine, the older drug, is shorter acting and has more side effects, so glycopyrrolate is the recommended agent. In rabbits, glycopyrrolate is preferred, because a certain percentage of rabbits have atropinase, which metabolizes atropine making it inactive.
3.1.1 Isoflurane and Sevoflurane
The 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.
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). Office of Environmental Health and Safety (OEHS) can provide assistance in evaluating potential exposures.
3.2 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).
3.3 Other inhalant agents
Other agents and techniques may be used for inhalant anesthesia, only when specifically approved by the IACUC in the animal use protocol.
Ether is an irritant and a fire and explosion hazard, and its use is not allowed. The use of ether must be scientifically justified and then its use reviewed and approved by the IACUC.
Carbon dioxide is a potent anesthetic, but concentrations are difficult to control, making the margin of safety unacceptably low.
3.4.1 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).
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 must be stored under refrigeration once reconstituted. It is not safe for use in rabbits (kidney disease). Telazol is a Class III controlled substance
3.5 Ketamine combinations
3.5.1 Ketamine-alpha2-agonists (Xylazine or Medetomidine)
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 medetomidine in the same syringe produces a surgical level of anesthesia in many species, notably rodents and rabbits.
In swine the combination of Ketamine-alpha2-agonists is used to restrain the animal prior to induction with isoflurane.
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 or Yohimbine. Administration of reversal agents may be useful but is not routinely recommended as the recovery may be agitated and alpha2 analgesia is reversed.
Disadvantages: Disadvantages of ketamine-a2-agonist combinations are that they will not reliably reach surgical anesthesia in all cases, and that they can cause profound cardiac depression. Xylazine 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.
3.5.2 Ketamine-benzodiazepines (Midazolam or Diazepam)
Ketamine may be combined with the benzodiazepines Midazolam or Diazepam in the same syringe 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.
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. Ketamine is a Class III controlled substance while the benzodiazepines are in Class IV.
Pharmacologically, Telazol is a dissociate-benzodiazepine combination.
Though superseded in most applications by newer anesthetics, barbiturates still have their place in the animal laboratory. They are most frequently used in terminal or acute studies, as recovery can be prolonged and unpleasant, especially in larger animals. Barbiturates are often the anesthetic of choice when neuron-physiological recordings are being conducted, such as visual or auditory evoked responses. Concurrent use of an analgesic (opioid or non-steroidal anti-inflammatory drug) is encouraged as it may improve pain relief with barbiturate use, and lower the required dose of barbiturate.
Sodium pentobarbital (Nembutal) and sodium thiopental (Pentothal) are currently the two most commonly used barbiturates. The duration of action of pentobarbital is considerably longer than that of thiopental.
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.
3.7. a2-agonists (Xylazine or Medetomidine)
The a2-agonists (Xylazine or Medetomidine) are hypnotic analgesics with significant pain relief. Used as sole agents, they do not produce sufficient depth of anesthesia for even minor surgical procedures. Combined with Ketamine and possibly supplemented with inhalants or local or topical analgesics [link to local anesthetics later in document], they may be useful during surgery. In some species, medetomidine appears to lead to greater anesthetic depth than does xylazine, and it is more reliably antagonized by atipamezole.
Advantages: a2-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 IP or subcutaneous atipamezole (yohimbine is sometimes used for xylazine reversal). 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 pretreatment with atropine or glycopyrrolate. a2-agonists cause a transient hyperglycemia which may have research implications. Xylazine often causes transient nausea and vomiting, especially in cats. Rapid IV administration of reversal agent has produced seizures in some species.
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.
3.8 A2 Reversal agents:
Advantages: Administration of a reversal agent can rescue an animal which is in cardiac distress due to bradycardia or cardiac arrhythmia. It will also promote quicker recovery from a2 agent anesthesia.
Disadvantages: If ketamine and alpha2 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 remaining in the system not being softened by the a2 agonist.
Propofol can produce general anesthesia in animals, as a sole agent with continuous infusion for surgery, or as a pre-anesthetic for endotracheal intubation. It is valued for its fast recovery time, even after prolonged administration.
Advantages: Animals recover from propofol in minutes, even after prolonged administration.
Disadvantages: 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, therefore unused propofol from an opened ampule should be discarded after use and not stored for future use.
3.10 Tribromoethanol (Avertin)
Avertin has been the standard anesthetic in much mouse transgenic work. It produces short-term (15-20 minutes) surgical anesthesia with good muscle relaxation and moderate respiratory depression. It does not produce significant residual post-procedural analgesia. Unless strongly justified in the animal care and use protocol, use of Avertin is restricted to mice only, for a single survival anesthesia plus terminal/acute use.
Advantages: Advantages of Avertin are that it is easily administered via the intraperitoneal route, produces good short-term surgical anesthesia, and is not a controlled substance.
Disadvantages: Avertin is not commercially available as a pharmaceutical drug, and must be made in the laboratory from the reagents tribromoethanol and amylene hydrate. Avertin can cause peritonitis in mice, and the risk of peritonitis, including fatal peritonitis, increases with each time it is used. Post-procedural analgesia has not been demonstrated, so use of a preemptive or postoperative analgesic is generally required. Though surgical anesthesia is short (15-20 minutes), anesthetic recovery can take 40 minutes, during which time the animal must be continually attended and kept warm.
Cautions for use: Avertin must be carefully prepared in the laboratory under aseptic conditions. Working dilution of 1.25% is recommended — this is best prepared fresh for use, or stored for no more than one week. Avertin is used only for mice. It is not to be used twice in one animal on a survival basis (if used a second time that use should be terminal/acute). Where possible, IACUC recommends that inhalants replace Avertin.
Opioid drugs are important components of many surgical anesthesia regimens, and are the most potent available post-procedural analgesics. Drugs in this group vary in their potency as well as their duration of action. Fentanyl, oxymorphone, morphine, buprenorphine and butorphanol are the most commonly used opioids in laboratory animal medicine, though others may be used on occasion. Fentanyl is the most potent of the three μ agonists, but also the shortest acting. Buprenorphine is longer-acting and is good for most post-operative applications. Butorphanol may be more efficacious than buprenorphine for birds and for cats. Buprenorphine and butorphanol are mixed agonist/antagonists at different opioid receptors; they produce a less profound respiratory depression than full agonists, but also have a “ceiling effect” in the degree of analgesia produced with increasing doses. Administered as a part of the anesthesia regimen as pre-emptive analgesia, they 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.
Advantages: Opioids are potent analgesics. Concurrent use with inhalant or parenteral anesthetics for general anesthesia will lower the required dose of the anesthetic.
Disadvantages: 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. Opioids are controlled substances . Morphine, oxymorphone and fentanyl are Schedule II. Buprenorphine and butorphanol are schedule IV.
Cautions for use: 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. 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 kappa receptor agonists (buprenorphine and butorphanol).
3.12 Non-steroidal anti-inflammatory drugs (NSAIDs)
The advent of newer, more potent, more specific anti-inflammatory agents has increased their usefulness in laboratory animal use. Most reduce fever, reduce inflammation, and provide varying degrees of analgesia (acetaminophen does not significantly reduce inflammation).
Advantages: 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 local anesthetic/analgesics. Injectable NSAIDs are useful for accurate dosage and administration to small rodents. Oral flavored analgesics are useful for mild pain in nonhuman primates.
Disadvantages: 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.
Cautions for use: Cats are particularly susceptible to toxic effects of NSAIDs. Acetaminophen is never administered to cats; 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.
3.13 Local anesthetic/analgesic drugs (lidocaine and bupivacaine)
Local anesthetic/analgesic drugs (lidocaine and bupivacaine) may be useful both during surgery, and post-operatively. They block nerve conduction when applied locally at sufficient concentration. Lidocaine has a fast onset of action, and provides a couple of hours of analgesia. Bupivacaine has a slower onset of action (up to 30 minutes) but provides up to 12 hours of residual analgesia. Both are infiltrated subcutaneously at the surgical site, or (especially in larger animals) may be used regionally (epidural, intrathecal, intercostal).
Lidocaine cream (EMLA or ELAMax) is used topically on shaved, intact skin prior to venipuncture, though it requires 30-60 minutes or more of contact with skin to reach full effect. Tricaine methanesulfonate (MS-222) is a related compound used as a general anesthetic for fish and frogs.
Advantages: 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.
Disadvantages: Intramuscular and intravenous injection should both be avoided. 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.
3.14 Miscellaneous agents
Urethane, choral hydrate, equithesin, sodium thiamylal, a-chloralose have some specialized use in laboratory animal anesthesia. Their use should be discussed with a veterinarian.
Note that all of the doses listed in the formula tables are approximations and must be titrated to the animal’s strain, age, sex and individual responses. Significant departures from these doses should be discussed with a veterinarian. Doses will also vary depending on what other drugs are being administered concurrently.
All doses are listed as milligrams per kilogram (mg/kg) unless otherwise noted. Where a dose has not been determined, it is listed as TBD (To Be Determined).
Dilution of injected drugs allows more precise dosing, may make some drugs less irritating when injected, and may facilitate absorption, but also may shorten the shelf-life of the compound. Aseptic procedures must be observed as mixtures (cocktails) are prepared; this includes wiping the cover of each vial or bottle with 70% ethanol or isopropanol, diluting with sterile water or sterile saline and not reusing needles used for dilution or administration. Only new, sterile needles must be used for withdrawing aliquots from the cocktail. Diluted drugs must be labeled with the contents and concentration of the dilution, the preparer’s name or initials, and dated, then discarded after 1 month, at the expiration date of any of the components, or according to the manufacturer’s instructions.