The approach to rodent analgesia in laboratory animal medicine has undergone significant changes during the last decade,^1,2 and the need and benefits for analgesia in rodents has been clearly demonstrated. In spite of this, there are still personnel working with rodents who consider pain management to be unnecessary by making statement such as “rodents feel less pain than other mammals,” “pain is advantageous because it limits rodents' activity,” “analgesia conceals clinical deterioration,” or “alleviation of post-operative pain will result in the rodent injuring itself.” Obviously none of these statements are substantiated by scientific data. On the contrary, studies have shown that rodents are likely to experience pain at similar levels as other mammals. Pain may preclude rodents from being able to access food and water, and may lead to self-mutilation³— therefore, it must be addressed. Another stated justification for not using analgesia is that analgesics will adversely affect the results of an experiment.³ Analgesics do have an effect on a rodent’s physiology and behavior, however, this must be considered against the physiological effects of pain,which may include impairment of bowel motility and immune suppression.^4-9 Some may argue that the deleterious effects of pain far surpass any mild metabolic derangement caused by analgesia. The benefits of analgesia commonly surpass the rare and undesirable side effects which can arise from analgesic use. Because of the increased number of scientific studies regarding the detrimental side effects of pain, there is an increased use of appropriate analgesia in rodents.^1,2,10 Institutional Animal Care and Use Committees (IACUCs) must consider this argument and guard itself against “automatically” approving a surgical protocol without analgesics just because an investigator states that analgesics will interfere with data, even when references are provided.

Pain causes physiological and psychological changes in rodents such as ventilation, perfusion and cardiovascular abnormalities, increased metabolic demand, tissue catabolism, impaired immune function, increased risk of sepsis, delayed wound healing, a prolonged period of recovery, and abnormal behavior. ^4-9,11,12 Additionally, studies have demonstrated that a lack of control of acute pain increases the possibility of chronic pain.¹³ All of these have the potential to affect the results of an experiment, and increase morbidity and mortality.¹⁴ Beyond these reasons, we also have an ethical obligation to provide humane care and to comply with regulatory guidelines.^14,15

An understanding of the pharmacology, physiology, and specific applications of individual analgesics is vital in order to select the appropriate analgesic( s) for each case. The rodent’s health, age, strain, and the amount and duration of pain expected from a procedure are just a few of the considerations when choosing an analgesic protocol.^16-18 In some cases, like surgical procedures, the potential for pain is obvious. In other cases, the potential for pain is not as apparent. For example, more subtle sources of pain may occur from rodent models of arthritis, cancers (e.g. bone neoplasm), and infectious diseases (e.g. ulcerated lesions where pain can be chronic and subtle).^19-24 Every attempt should be made at treating pain preemptively, rather than waiting until a rodent is exhibiting signs of pain.²⁵ This prevents noxious stimuli from activating the central nervous system, which decreases the benefit and effectiveness effectiveness of the analgesic. Pre-emptive analgesia reduces peripheral inflammation, thereby decreasing the potential for central nervous system hypersensitivity. ²⁶ An analgesic protocol should include a multi-modal approach that incorporates analgesic agents from various classes such as opioids and nonsteroidal anti-inflammatory drugs (NSAIDS).^3,27 This approach allows analgesics to act at various points of the pain cascade, which leads to synergistic or additive activity and decreases overall drug toxicity. ^3,27 An optimal protocol of pain relief should include a preemptive and multi-modal approach with pre-, intra-, and post-procedural pain management.^3,25,27

Challenges of Pain Detection and Provision of Analgesia in Rodents
Detection of pain and the provision of analgesia in rodents is challenging because it is difficult to assess physiological and behavioral changes associated with pain in these animals. ^3,14,15,28 This is due to the fact that rodents are small, nocturnal animals, vocalize at ultrasonic frequencies and often mask signs of pain.^29-34 Therefore, pain detection in rodents utilizes subtle behavioral clues that alert personnel to their condition.^17,35-39 There is no universally accepted standard guideline or score sheet used to determine if a rodent is experiencing pain. ^15,35,38

Detecting painful behavior requires training and skilled observation. However, there can be large variations between individual observers. Fortunately, within the last decade scientific data has been introduced, which describes various behaviors associated with pain in rodents.^24,25,40-44 Observation of pain-associated behaviors and knowledge of normal behaviors allows for a better assessment of pain. In cases where specific data is not available, personnel can extrapolate the knowledge of the expected level of pain a procedure may cause. It is recommended that unless otherwise shown, if the procedure is expected to be painful to people it should be considered painful in animals.^14,45 The skill of the surgeon should be a consideration. There is an inverse relationship between the skill of the surgeon and the amount of postoperative pain. The expected level of pain a procedure may cause should be assessed against a surgeon’s skills when formulating an analgesic protocol.

Detection of Pain
Ideally, the animal’s behavior should be observed and, if an animal is determined to be in pain, then analgesia should be provided or the dose adjusted. An analgesic protocol for a group of rodents should be developed and then observation should be used to tailor the individual dosing.¹⁵ This is in contrast to providing an analgesic protocol for an entire group, which is a common practice with rodents due to the number of animals utilized per experiment. Since rodents are nocturnal, observation of behavior associated with pain only during the light phase of the light cycle can lead to poor pain assessment.⁴⁶ Initial rodent pain studies included red light monitoring of animals during the dark phase of the light cycle with video equipment. This might be ideal but may not be feasible or practical for most rodent facilities. Recent data indicates that it is possible to determine if a rat is in pain by observing it for 5-10 minutes post surgically during the light phase of the light cycle and by utilization of behavioral signs.³⁶ To further complicate pain recognition, variation in pain sensitivity and responsiveness to analgesics among different rodent genders and strains has been observed.⁴⁷ Additionally, clinical signs can be interpreted subjectively by different individuals. In order to simplify detection and make it more objective, pain scoring systems can be developed.¹⁵ Individual scoring systems must be developed for specific procedures, species, genders and strains. ^17,35-39 For example a scoring system for a laparotomy (incision into the abdomen)⁴⁸ will not be the same as that needed for a thoracotomy (incision into the thorax). Some analgesics are associated with specific rodent behavioral changes such as walking and grooming, which is seen under the influence of buprenorphine. Scratching and face washing is seen with ketoprofen and carprofen analgesia.³⁵ Therefore, it is recommended that observation of rodents should be performed for a minimum of 5-10 minutes at a time, and personnel observing the animals must be trained in the recognition of pain associated animal behavior. ³⁶

Recently, automated systems have been introduced and studied for detection of pain in rodents. ^39,49 These systems can provide a rapid analysis of rodent behavior. However, these methods require further validation. Automated systems would make the assessment of pain and analgesic effectiveness more feasible and efficient in rodent models than is currently possible with conventional analysis methods. ^39,49

Rodent-specific Behaviors Associated with Pain
Signs of pain in rodents that indicate the need for analgesics include back arching, staggering, pressing, writhing, and twitching of the skin and abdomen. ^35,36,38,50 Food intake, water intake, and body weight may all decrease postoperatively in a painful animal.⁵¹ Normal activities, like climbing and grooming, may also be decreased and the small rodents may appear hunched and show piloerection.⁵² Abnormal behaviors, such as paying excessive attention a particular body part, may be evidence of pain. ^53,54 For example, the rodent may stare at the body part, chew excessively at it, or even groom it excessively. It takes practice and careful observation in order to detect pain and monitor the effectiveness of pain control while using analgesics. Pain behaviors can be confused with normal behaviors. When using wound licking as one of the behaviors for signs of pain it must be remembered that licking is also performed by rats even with high doses of certain analgesics such as opioids.⁵⁵ In addition, one must distinguish between normal grooming behavior and excessive licking. In short, when assessing pain, as many behaviors as possible should be considered to determine if a rodent is indeed painful. ^27,52,56 Behavioral formulas could assist this process by adding up the frequency of behaviors associated with pain. Behavioral formulas can be designed to include not only signs of pain, but signs of normal behaviors as well.

Physiological Parameters
In addition to behaviors associated with pain, physiological signs associated with pain have been described, including fluctuations in respiratory rate, heart rate, body temperature, blood pressure, andweight.^15,57 Body weight change can be used to determine pain, since painful rodents lose more weight post-surgically than animals whose pain is alleviated.^58,59 If fluids are provided during or after the procedure, they may alter the weight of the animal and must be taken into consideration. Also, changes in mouse food consumption may not be as dramatic as in rats, therefore this parameter might not be as useful to assess of pain in mice.³¹

Appearance
A rodent’s appearance can provide additional insight into whether an animal might be painful. In order to perform this examination, the cage may need to be removed from the rack to provide optimal visualization. A poor hair coat could be a sign of pain and/or distress. In rats, porphyrin (reddish/brown) ocular or nasal staining could be a sign of pain and/or distress. This staining may be seen on the inside of the forepaws as rats tend to normally wipe the eyes and nose with their forepaws. ^29,31

Conclusion
Pain management in rodents is an integral aspect of laboratory animal medicine and surgery that offers many advantages to the animal and to the scientist. Not only is it unethical to ignore pain in rodent species, but ignoring it can lead to aberrant study results. Untreated pain may result in injury, illness, and induce abnormal physiological and behavioral changes, which can skew data. Pain management will generally result in improved data, compliance with laws and regulations, and most importantly improves animal welfare.

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Szczepan Baran, VMD, MS is the President for the Veterinary Bioscience Institute. Veterinary Bioscience Institute, 292 Main Street, #300, Harleysville, PA 19438; 206- 714-6208; szczepanb@vetbiotech.com; www.vetbiotech.com

Elizabeth Johnson, VMD is the Manager of Veterinary Services for Putney, Inc. Putney, Inc, 400 Congress Street, Portland, ME 04101.

Marcel Perret-Gentil, DVM, MS is the University Veterinarian and Director of the Laboratory Animal Resources Center at The University of Texas at San Antonio, One UTSA Circle, San Antonio, TX.