Module VIII Flashcards
- Give examples of behavioural changes and other signs of discomfort, pain, suffering, or distress in a rat or a mouse, as well as signs of positive well-being and principles of how pain, suffering and distress can be managed.
When animals are exposed to stress they will respond by modifying its behavior and physiology in attempt to cope with the stress.
- Acute stress (positive): a survival mechanism to aid the animal to adapts and cope to challenges.
- Chronic stress (negative): where the animal is unable to adapt completely to stressors, which will cause maladaptive behaviors.
Pain and stress assessment:
1. Physiological parameters:
- Heart rate, blood pressure, body temperature (will increase during stress).
- Biomarkers
- Adrenaline, noradrenaline, corticosteroids, oxidative stress markers. - Behavioral and clinical signs (most used to study animal in real time):
- We can look for signs such as: is the animal alert? Is it moving normally? Dies it drink or ear more or less? Is it protecting a certain part of the body? Is the animal’s posture abnormal? Does it sound when being handled? Does it bite or lick itself more than normal?
- Pain specific behaviour after laparotomy twitching, back-arching, falling, abdominal press, writhing.
- Ethogram and scoringsystem (a list of species-specific behaviors)
- Body weight loss, less food and water consumptions, more urination and defecation, fur quality.
The general signs that an animal is experiencing pain or distress depends on the animal so it is important to know the normal behavior in the specific species.
- Basic behaviors in mice are nesting, social behavior, exporation, grooming, and movement.
- Signs of pain and stress can be fear, anxiety, depression, anger/frustration, fatigue, tense muscles.
- Abnormal behaviour: stereotypic behaviour, when the animal try to cope or calm down.
Pain and suffering can be managed by:
- Anesthesia or analgesia.
- Prevent pain from occurring, by giving the analgesia beforehand.
- Good surgeon = less trauma and less pain.
- Good hygiene = avoid subclinical infection.
- Continue treatment after an appropriate time.
- Describe what a humane endpoint is. Identify criteria to be used to set humane endpoints. Define action to be taken when a humane endpoint is reached and consider possible options for refining methods to finish at an earlier endpoint. Discuss also factors to be considered and methods available for assessing and recording the welfare of animals e.g. score sheets.
The humane endpoint is the earliest point at which an experiment can or should be terminated to avoid unnecessary suffering for that animal. This will depend on the study, but could be cessation of treatment, administration of analgesics, euthanasia, etc. This is different from a scientific endpoint, which is the point when we have successfully collected the intended data from the study. You should reach the scientific endpoint before the humane endpoint because otherwise the study has no point.
Need to use early behavioral and clinical signs in combination with model-specific signs in order to introduce as early endpoints as possible but the endpoints may differ depending on the severity of the experiment.
To help set the human endpoint:
You can make a score sheet, which can be an important tool for determing humane endpoints. A score sheet is a document on which the clinical status of each individual experimental animal is recorded at regular predetermiend intervals. Score sheet should be specific for characteristics related to experiment/model, species and strain.
Quantitative data includes body weight, body temperature, respiration, heart rate, behavior.
Qualitative data includes quality of respiration, posture, appearance such as closed eyes, disturbed pelage as with ruffled fur, diarrhea, coughing.
E.g. humane endpoint in tumor models:
- Describe the severity classifications included in the Directive and give examples of each category; explain cumulative severity and the effect this may have on the severity classification.
Non-Recovery/Terminal: Procedures which are performed entirely under general anaesthesia from which the animal shall not recover consciousness shall be classified as ‘non-recovery’.
Mild Experiments (e.g. Administration of anaesthesia): Procedures on animals as a result of which the animals are likely to experience short-term mild pain, suffering or distress, as well as procedures with no significant impairment of the well-being or general condition of the animals shall be classified as ‘mild’.
Moderate Experiments (e.g. surgery under general anaesthesia): Procedures on animals as a result of which the animals are likely to experience short-term moderate pain, suffering or distress, or long -lasting mild pain, suffering or distress as well as procedures that are likely to cause moderate impairment of the well-being or general condition of the animals shall be classified as ‘moderate’.
Severe Experiments (e.g. severe restriction of movement over a long period or learned helplessness models): Procedures on animals as a result of which the animals are likely to experience severe pain, suffering or distress, or long-lasting moderate pain, suffering or distress as well as procedures, that are likely to cause severe impairment of the well - being or general condition of the animals shall be classified as ‘severe’.
● In this example, the period has to be defined by the competent authority and it is also mentioned in the directive that short term restriction is only categorized as mild.
Cumulative severity is a commonly used term but harms do not accumulate or add up, although animals become sensitised to certain procedures (e.g. repeated injections), so the distress associated with each one is increased.
Repeated intraperitoneal injections, for example, may result in a greater severity than the ‘mild’ classification that might be applied to a single injection of an appropriate volume of a non-irritant material.
- Describe the principles of humane killing (euthanasia) and give examples of different methods by which animals are allowed to be killed. In addition, explain why someone competent to kill animals should be available at all times.
Laboratory animals need to be killed at a defined time point (either experimental endpoint or humane endpoint). The animals need to be killed in a humane way with euthanasia which must be performed as quickly as possible without causing fear or stress in the animals. The method should be easily performed by the person doing it and it should also be safe for the person.
There are many methods (death should always be confirmed):
● Confirmation of permanent cessation of the circulation
● Destruction of the brain
● Dislocation of the neck
○ Mainly for mice and rats. The animal dies from rupture to the brainstem and the carotid arteries.
● Decapitation:
○ Mainly used for rats, the head is removed with a guillotine.
● Carbon dioxide:
○ Cheap, quick and easy. Gradually filling the chamber is important to avoid stress in the animals. Prior sedation or anesthesia may be beneficial but it is still debated.
● Exsanguination - draining of blood
● Confirmation of the onset of rigor mortis.
Immediate signs that the animal is dead are: heart beat stops, respiration stops, reflexes are absent, pale skin. Later signs are cold body surface and rigor mortis.
Someone competent to kill animals should be available at all times when handling animals or performing experiments as an accident may occur where the animal needs to be humanely euthanized immediately (e.g. animal injured after dropping the animal, injection of air bubble, etc.).
- Define sedation, local and general anaesthesia. Explain the triad of anaesthesia and the term balanced anaesthesia, and give examples of how to achieve this.
Sedation is the depression of awareness, whereby a patient response to external stimuli becomes limited. Sedation may be minimal, moderate or deep.
Local anesthesia involves numbing a specific area of the body to temporarily block sensation, typically for minor surgical procedures or medical interventions. It is just in the area that is going to be treated.
General anesthesia is a state of unconsciousness induced by medications to eliminate awareness, sensation, and memory during surgical procedures. It is done for people undergoing major treatment and the person is unconscious.
The anesthetic triad is anti-nociception/does not feel pain (not analgesia because this is the absence of pain and the subject is asleep), unconsciousness, and muscle relaxation. The basic elements are reversibility and controllability.
Balanced anaesthesia - combination of drugs to achieve the anaesthetic triad.
- Discuss the preanaesthetic considerations necessary for a successful anaesthetic procedure, including evaluation of the animal, as well as selection of agents.
● Evaluation of the animal (animal should be healthy, not carry infections, be acclimated to experimental facility).
● Practical considerations (assistance needed, fasting)
● Pre-anesthetic medication
● Choice of Anaesthesia
○ Pre-anaesthetic medication often consists of a cocktail of different drugs to reduce fear + stress, provides smoother induction of anesthesia, provides smoother recovery from anesthesia, reduces amount of other anesthetic agents required, reduced volume of salivary secretions, blocks vaso-vagal reflex.
■ Advantages:
● Lower dose of each drug can be given
● Minimization of the side effects from each drug while still fulfilling the anesthetic triad.
■ Disadvantages:
● The different side effects of each can interfere with the research protocol.
● The use of one single drug may be necessary in rare cases, where the interference from certain agents may affect the experiment negatively
○ Inhalation
■ Rapid induction and recovery
■ Easy to control anesthetic level
■ Can be used for prolonged surgery
■ Need for anesthetic machinery and technical competence
■ Could be harmful to the fetus in pregnant women
○ Injection
■ Simple, no requirements to equipment
■ Easy induction, allowing the animal to pass quietly into surgical anesthesia
■ Depth of anesthesia is less readily controlled
■ When a drug is injected, it has to be metabolized or excreted by the kidney - it is therefore easy to inject an overdose
■ Prolonged anesthesia calls for repeated injections or continuous infusion through intravenous access
- Discuss relevant maintenance of anaesthesia, and how to monitor the anaesthetic level. Describe what problems that might arise during anaesthesia and discuss what actions to be taken to prevent or counteract these.
Purpose of anaesthesia: unconsciousness, anti-nociception, muscle relaxation.
Before the anaesthesia: evaluation of the animal, clinical examination, fasting, pre-anaesthetic medication.
Monitoring the anaesthetic level: absence of reflexes (righting reflex, withdrawal reflex, corneal reflex, swallowing reflex), circulatory functions, respiratory functions, body temperature. Remember that the eyes are open during anaesthesia- use eye cream or ointment to prevent drying of the cornea.
Short and low risk procedures: Non-electronic monitoring of pulse rate, respiration, body temperature, color of skin or mucous membranes to assess arterial oxygenation, done with pulse oximetry, value should be above 95% oxygen saturation.
Long-term and highrisk procedures: In long-term & high-risk procedures, more sophisticated monitoring is needed. Circulatory monitoring (HR & rhythm using ECG; BP), respiratory monitoring (O2 & CO2 levels, blood gases). In larger animals, endotracheal intubation and ventilation is required, hypoxia can be avoided with an oxygen face mask, but CO2 levels cannot be controlled without ventilation leading to risk of acidosis.
Problem: animal is moving, or respond to noxious stimulus. Then provide more anesthesia.
Problem: poor respiration or circulation. Then reduce the anesthetic.
- Describe and discuss methods of optimising post anaesthetic recovery to ensure a smooth and rapid recovery from anaesthesia.
To ensure a smooth and rapid recovery:
● Pain
○ Provide analgesia
● Dehydration
○ Fluid therapy
○ Body temperature saline IV or SQ during anesthesia and/or post-operatively
○ Provide easy access to drinking water: extra long drinking nipples or jelly liquid for easy ingestion.
● Hypothermia
○ heating blankets (larger animals); infrared heating lamps or incubators (rodents)
○ body temperature must be maintained until animal is fully recovered
○ be aware not to overheat the animals!
● Infections
○ Asepsis
○ Observe wound healing
○ Provide antibiotics if necessary
● Post-anaesthetic restlessness and self injury can be caused by pain or severe distress, consult veterinarian, use sedatives or analgesic, if indicated.
Post-Operative Care: Large Animals vs. Rodents
● Large animals
○ Dedicated recovery room
○ Warm (20-25 C) and quiet
○ Supportive care
● Rodents
● Separate recovery area
● Incubator/warmed cage to maintain body temperature (25-30C for adults, 35- 37C for neonatals)
● Bedding should be soft and comfortable (avoid sawdust or similar – risk of sawdust in eyes and mouth)
● Avoid unnecessary human contact and handling
- Describe and discuss good working practices with regard to use, storage and disposal of anaesthetic and analgesic agents.
● Anaesthetic gases
○ Ensure adequate ventilation to scavenge waste gases
○ Handle equipment correctly
● Gas cylinders
○ Risk of explosion in case of fire
○ Heavy! Keep properly attached
● Needles
○ Careful handling
○ Disposal in designated waste containers
● Drugs
○ Store in locked cupboards
○ Keep record of purchase and usage
○ Destroy non-used drugs according to legislation
