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Summary LAS: Experimental Techniques and Behavioral observation in rat

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Laboratory Animal Science

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Universiteit Antwerpen (UA)

Laboratory Animal Science summary of lecture (see title) in English. The title is too long: '& mouse'.. Guaranteed to pass exam and get your certificate FELESA C. Written in ! There are always one or two questions from this part, usually big questions and in this summary I have explained everything...

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experimental techniques and behavioral observating in rat

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Laboratory Animal Science – Summary – partim Dr. D. van Dam

Lecture: Experimental Techniques

Marking lab animals
Lab animals can be marked based on:
- Short term marking & during behavioral observation where you color-code the
tail or the fur with a permanent marker.
- Permanently (long term) by:
o Ear punches
o Tattoo
o Cutting toes (not ideal when studying motor performance in lab animals)
o Metal numbered label in ear (currently forbidden!)
o Subcutaneous RF-chip with unique code that can be read by a chip reader

Marking animals is done to identify lab animals in a cage.

Tattoo tool to apply a tattoo to the feet of your animal or on the tail of your animal.

By cutting the toes of your animal, you can identify a large number of animals in your
cage. If you want to study the motor performance, this marking method is not preferred
since cutting the toes will influence the motor performance of the animals and this
method will interfere with the normal motor function and analysis of the motor
performance and affect your experimental results and outcomes.

There are techniques that evaluate motor performance and one of them studies the
footprint pattern of these mice e.g. the distance between the right and left foot. You look

Saif Haify Laboratory Animal Science – Summary 6-November-2015

,at the toespan, the distance between the toes. And if you have cut off the toes, you can
imagine you can’t study anything with these mice.

Metal ear tags. Because lab animals are housed close to each other, they might bite each
other and then the metal ear tags are ripped off and damage the ear or rip of the ear of
the mouse.

You lose your identification but also this damage to the ear will cause inflammation to
the ear.

The best two options in applying a permanent identification method to your lab animals:
- Ear punches

Simple, easy and cheap method. You punch holes in the ears of your animals. By
punching in different combinations in the ear, you can identify up to 8-10 animals
housed in the same cage.

- RF-chips

Saif Haify Laboratory Animal Science – Summary 6-November-2015

,Expensive method. RF response chips are subcutaneously implanted with a needle and
then you use a chip reader to read-out the unique code.

Immobilization of lab animals
Immobilization is required for some experimental procedures when you for example
want to add a compound to your animal. Immobilizing the animal is also done to avoid
stress in the animal but also to avoid injury to the researcher (such as biting or
scratching by the animal).

You can immobilize animals by:
- Restraining with your hands
- Restrainers (can be bought or self made)
- Anesthesia (through injection/inhalation) is used for long and painful
procedures. More further on in the course of prof. C. van Ginneken.

A very interesting website is that of AHWLA: Assessing the Welfare and Health of
Laboratory Animals with tutorials (photos and videos) on how to restrain the animals or
administer them a certain compound.

Immobilizing by hand

Rats
http://www.ahwla.org.uk/site/tutorials/BVA/BVA06-Rat/Rat.html

Rats are very docile, calm animals. They get used to being handled repeatedly and
actually when handling them more often, they become more calm and used to being
handled by the researcher.

You grab the animal around the shoulders and you fixate the head to avoid being
bitten.

Avoid grabbing the rats by their tails because the tail is very sensitive. But if you
have to grab the rat by the tail, grab it by the base of the tail and do this very quickly.
Never grab the rat by the point of the tail and especially never leave it hanging because
the skin can easily rip of.

Saif Haify Laboratory Animal Science – Summary 6-November-2015

,Mouse
http://www.ahwla.org.uk/site/tutorials/BVA/BVA05-Mouse/Mouse.html

Take the mouse out of the cage with one hand by grabbing the base of the tail (tip of the
tail is painful). Put it on the cage lid and let it grab the grid with the front paws; pull it a
little bit to the back and then by using your thumb and index finger (‘wijsvinger’) you
push the animal on the grid and grasp the skin around the neck. Finally you restrain the
tail between the palm of your hand and your pinky.

There are different strains available and the level of agitation depends on the strain. The
white albino mice are very docile and calm whereas the C57BL/6 mice are very agitated
and aggressive. Compared to a rat, a mouse will not habituate to repeated handling and
treating. The more you handle a mouse, the more agitated and nervous it gets.

Rabbits
http://www.ahwla.org.uk/site/tutorials/BVA/BVA08-Rabbit/Rabbit.html

NEVER PICK A RABBIT UP BY ITS EARS!

You grasp the back and scruff (lose skin) of the neck with one hand and with the other
hand you support the hindquarter of the animal. The rabbit becomes more calm when
you correctly restrain it and especially when you tuck away the head and cover the
eyes of the rabbit. Hold it firmly!

Saif Haify Laboratory Animal Science – Summary 6-November-2015

,Guinea pigs
http://www.ahwla.org.uk/site/tutorials/BVA/BVA07-GPig/GPig.html

Guinea pigs are usually docile and very calm. Grab the animal with one hand around
the shoulder and put your thumb under the chin to avoid being bitten and restrain the
animal. With your other hand you support the hindquarter of the animal. Place the
animal in an upright position because then they will feel most comfortable.

Hamsters
http://www.ahwla.org.uk/site/tutorials/BVA/BVA04-Hamster/Hamster.html

Restraining a hamster is similar to restraining a mouse. Place the animal beneath one
hand, take the scruff above the shoulder and back with the thumb and index finger
(‘wijsvinger’).

Immobilizing by restrainers

If you need your both hands free, for example when doing an intravenous (i.v.)
injection, you use species-specific restrainers like:

For rats and mice to do i.v. injections or administration of compounds in the tail.

Saif Haify Laboratory Animal Science – Summary 6-November-2015

,Again for rats and mice to perform injections/administrations via the tail.

Plastic restrainer for rabbits à the ears are used to administer a certain compound
or to draw blood in a rabbit à marginal vein can be used for this

The bunny snuggle à soft version where the ears are free and the eyes are covered to
calm down the animal and reduce stress.

Decapicones are triangle shaped plastic bags with an opening. These decapicones are
used when researchers want to decapitate their animal, by using a mini guillotine. An
advantage of these decapicones is that researcher also can do injections through the
plastic cone via an intraperitoneal (i.p.) injection.

Saif Haify Laboratory Animal Science – Summary 6-November-2015

,Simple and cheap solutions are also possible like using a tube or a towel to calm the
animals.

The type of administration of a chemical compound is determined by:
- Used species
- Duration of the planned experiment (acute vs. chronic administration)
- Chemical characteristics of the active compound such as water soluble or
maybe irritant for the animal
- Protocol or hypothesis; e.g. if you are studying a compound that has to be taken
orally by human, then during your experiment you also have to administer the
compound orally in your lab animal

Administration routes possible
I. Via the skin/mucous membranes
II. Enteral administration (via the GI system)
III. Parenteral administration
IV. Specific administration types
V. Chronic administration

I. Via skin/mucous membranes

Your compound must be in solution or in suspension (e.g. eye ointment) upon
administration to the skin or mucous membrane of the animal.

Advantages:
- Very easy access à you don’t need special skills to do this
- No stress for your animal
- No use of needles to puncture the skin or mucous membrane
- E.g. wound healing study

Disadvantages:
- Not so accurate à when you put some ointment on the skin of the animal, the
animal might lick it off or when many animals housed together, they rub against
each other and you lose some of your active compound. You don’t know exactly
how much of your active compound is taken up by the skin.
- Possible irritation to the skin or mucous membrane
- Limited research domain or application e.g. only in wound healing studies

II. Enteral administration (GI)

Two possibilities:
- Anal route (suppository = ‘zetpil’)
Can be done in all lab animals but usually only done in the larger lab animals e.g.
dogs. So a disadvantage is that it is not applicable in small animals such as rats, mice,
guinea pigs. But the advantage is the high accuracy in dose and administration time.

- Oral route
A better alternative to the anal route is administration via the oral route. This can be
done via supplementing the drinking water or feed of the animals.

Saif Haify Laboratory Animal Science – Summary 6-November-2015

, Advantages of supplementing the active compound in the drinking water or feed:
- It is very easy; no specific skill needed
- Less stressful for the animals

Disadvantages:
- Major disadvantage is the lack of accuracy (dose? Time of administration?). If
the animals are housed together, you don’t know the amount of active compound
taken up by the animal. You can overcome this problem by single housing the
animals, count the feed you gave the mouse and by the end of the day you see
how much feed is left. But this is expensive (single housing) and also not
accurate enough due to spillage of drinking water or feed. So you need to take
that into account when taking conclusions.
- Also time of administration is unknown. Rats and mice are nocturnal animals,
which means they are awake during our nights and they sleep during our day.
o Average mouse will drink 4-7 mL H2O/day and eat 3-6 g/day.
- Environment dependent (e.g. higher temperature will cause your mouse to
dehydrate and drink more and therefore consume more of the active compound).
- Hierarchy (e.g. group housed males: dominant male will always drink or eat first
and then the rest will follow. Again there is a variation in the uptake of active
compound).
- Potentially bad taste or irritation causing the animal to be reluctant (= ‘met
tegenzin’) when drinking or eating.
- Must be a chemically stable compound to be able to dissolve it in the feed.
- Must be water-soluble to be able to dissolve the active compound in the
drinking water.

If you are treating your animals for a long period of time, this method might be good to
use but it has many disadvantages and things to keep in mind when applying this
method.

When treating your animals for a short period of time, it is best to use a gavage
needle or catheter to be more accurate when dosing the animals. You administer the
active compound via the mouth, through the throat and esophagus directly in the
stomach of your animal.

Advantage:
- Very accurate: dose and administration time of active compound is known
Disadvantages:
- You need specific skills (skilled personnel) and a lot of training is required to do
this in the right way
- It is stressful for the animals

Saif Haify Laboratory Animal Science – Summary 6-November-2015

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