Transcript - ISpatula

Lab. No.1 :
Introduction to experimental
pharmacology , handling of
laboratory animals & drug
administration
Introduction:
 Animals are widely used experimentally in almost
every laboratory where a biological study is needed
 In pharmacological research, animals are being used
to test the effect of drugs before their introduction
to clinical trials where drugs are tested on humans
 In addition, animals are used in educational
institutions to demonstrate the effect of drugs for
teaching purposes.
• Experimental animals should always be:
A. Suitably bred.
B. Gently handled
C. Properly used
Ethical and legal responsibility
• Careful handling of animals in such a manner that
they do not suffer from unnecessary pain.
• Providing a good care towards the health and
well being of animals.
• It is important that animals are aware of the
handler’s presence before attempting to restrain
them, particularly if the animal is initially asleep.
This will reduce stress for the animal and help to
avoid bite injuries.
ANIMAL HOUSE
• Diurnal light cycles in animal holding rooms (12:12-h
light:dark cycle)
• Controlled temperature (22º C)
• Controlled humidity (range 30%-70%)
• Free access to water ‘ad libitum’ and chow diet
• Adaptation period
CAGES
Selection of an experimental animal
for a particular experiment depends
on:
1. Nature of the experiment.
2. Type of drug activity to be evaluated.
3. Specific character of the experimental
animals
Animals can be used in:
• “In Vivo” experiments when the whole intact
animal is used, or in
• “Ex Vivo” experiments when only a part of the
animal is used. This part could be an organ
(e.g. heart, uterus, trachea.etc.), a tissue (e.g.
skeletal muscle) or even some cells (e.g. blood
cells).
Types of animals:
• Large (monkey, pigs, dogs and cats)
OR
• Small (mice, rats, rabbits and frogs)
For teaching purposes the small animals are
commonly used.
Selection of species depends on many
factors:
• Technical issues, e.g. availability of the species in the
numbers needed, health and genetic status of the
species, animal size and volume of blood required
• Animal husbandry issues, e.g. availability of suitable
housing, ease of handling of the animals, training and
level of competence of personnel
• Public concern about use of certain species
• Financial considerations
Types of experimental animals:
• 1.Mouse
• 2. Rat
• 3. Guinea pig
• 4. Rabbit
MOUSE
Mice
• The most common mammal among laboratory animals.
• Characteristics:
1. Mice are small laboratory animals weighing about 18-30 g.
2. They are very sensitive and consume small doses of drugs.
3. They can be easily handled.
4. Highly exploring and mobile
5. Drugs are best injected intraperitoneally or intravenously into
one of their superficial tail veins.
Mice
• Uses in research:
1. In cancer and genetic research.
2. In acute toxicity studies.
3. In screening of various drug activities specially
central nervous system (CNS) activity.
Mice sexing (how to discriminate between male
and female mouse)
• The distance between the anal and genital orifices
is greater in the male (left) compared to the female
(right).
HANDLING & RESTRAINT OF MICE
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Instructions:
Gently but firmly.
Wear disposable gloves
Wash your hands prior to and after handling
Wear a clean laboratory coat
HANDLING & RESTRAINT OF MICE
• The animal should be grasped by the tail, and should then be
placed on a rough surface such as a cage top while holding the
tail firmly
To carry the mouse for injection , you can
use one-handed or two-handed methods:
• One-handed method :
• Place the mouse’s tail between the last two
fingers of the hand that is holding the nape.
• Two-handed method:
• The scruff can be grasped between the thumb
and forefinger whilst maintaining a grip on the
tail.
Injecting the mouse
1. Intraperitoneal injections:
• can be made into the lower quadrant of the
abdomen.
2. Subcutaneous injection
• Subcutaneous injection can be made into the
scruff of the neck. Care must be taken to
direct the needle into the scruff and not into
the handlers finger or thumb.
• 3. Intramuscular injections
• can be made into the quadriceps muscle
groups on the anterior of the thigh
GENETICALLY MODIFIED MICE
• Knockout mouse (KO):
• Is a genetically engineered mouse in which researchers have
inactivated, or "knocked out", an existing gene
• This causes changes in a mouse's phenotype (appearance,
behavior, physical and biochemical characteristics)
• Important animal models to study the role of genes that have
been sequenced but whose functions have not been
determined
• ….Causing a specific gene inactivation, and observing any
differences from normal behaviour or physiology, we can infer
its probable function
• May also be a useful experimental pathological model to test
the efficacy of new drugs
GENETICALLY MODIFIED MICE
• Knockin mouse
NUDE MOUSE
• Mouse from a strain with a genetic mutation that causes a
deteriorated or absent thymus, resulting in an inhibited
immune system due to a greatly reduced number of T cells.
• The phenotype (main outward appearance) of the mouse is a
lack of body hair…..nude" nickname.
• Valuable in research! it can receive different types of tissue
and tumor grafts (xenografts ), with no rejection response.
RAT
RAT
• Characteristics:
1. Rats have a larger weight than mice (about 300-500 g).
2. They can withstand long periods of experimentation under
anaesthesia.
3. They can be easily handled if treated kindly.
4. Drugs can be injected intravenously (I.V) into their tail veins,
subcutaneously (S.C), intramuscularly (I.M) or
intraperitonealy.
5. They can be also given drugs orally by means of stomach tube.
6. Isolated preparations can be taken from them easily.
(stomach, fundus, colon and uterus are the most commonly
used preparations)
Proper method of handling:
To initially restrain a rat, the handler should
gently grasp it around the shoulders.
Proper method of handling:
• The handler’s thumb can then be
placed under the rat’s mandible,
to prevent bites, and the rat’s hind
limb scan be supported with the
other hand. Restraint should be
firm but not too tight as this will
impede the animal’s respiration.
• Note: Bites from rats are
uncommon and will typically only
occur if the animal is stressed or in
pain.
Intravenous administration in rat
Intraperitonial injection
• Intraperitoneal injections may be made into
caudal half of the abdomen with the needle
directed along the line of the hind limb.
Gastric gavage
The guinea pig
The guinea pig
• Characteristics:
1. very docile and tame animals.
2. very susceptible to infections and highly sensitive to action of
many drugs, e.g. histamine and penicillin.
3. cannot synthesize endogenous vitamin C and this vitamin
should be supplied in their diet.
4. Some isolated muscle preparations can be taken from guinea
pigs, eg. Guinea pig Vas Deferens, tracheal chain or ileum.
5. Drugs can be injected intravenously (I.V) into their tail veins,
subcutaneously (S.C), intramuscularly (I.M) or
intraperitonealy.
6. They can be also given drugs orally by means of stomach tube.
The guinea pig
• Use in research:
1. Screening for histaminic, antihistaminic drugs,
respiratory or cardiovascular activity.
2. Testing the effect of drugs on their isolated
muscle preparations.
Handling and Restraint of Guinea Pig
• To initially restrain a guinea
pig, the handler should be
rapid and smooth, to avoid
frightening the animal.
• The handler’s thumb is placed
beneath the jaw of the guinea
pig. The hindquarters of the
guinea pig are supported by
the handler’s other hand
The rabbit
The rabbit
• Characteristics:
1. Rabbits are very docile animals.
2. They can be easily injected intravenously into
their marginal ear veins.
3. They can be also given drugs orally by means
of stomach tube.
4. Some isolated muscle preparations can be
taken from rabbits.
Intravenous injection in rabbit
• Rabbits may also be restrained for injection by wrapping
the animal in a drape or towel. If the rabbit is securely
wrapped, it will not struggle.
• Intravenous injections into the rabbit’s marginal ear vein
can be readily made if the rabbit is restrained with a
drape.
Handling and restraint of the rabbit:
• Rabbits are especially susceptible to the effects of
stress and should always be approached in a calm
and confident manner.
• The handler is restraining the rabbit firmly by the
scruff with the other hand ready to support the
animal’s hindquarters.
BLOOD SAMPLING IN MICE/RATS:
1.
2.
3.
4.
from tail vein
from orbital sinus
from cardiac puncture
from saphenous vein
1. Blood collection from Tail vein:
It may be used to collect a small volume of blood (Approximately
0.1 ml )
1. Carefully warm animal with heat lamp or disposable
hand warmers.
2. Place animal in an appropriate restraint device.
3. Clean withdrawal site with alcohol.
4. Using a sterile scalpel blade, nick the lateral tail vein as
shown in this picture.
5. Collect blood into an appropriate container.
6. Apply gentle pressure to stop bleeding.
2-Blood Collection From Orbital Sinus
• The Orbital Sinus method may be used to collect larger
volumes of blood(up to 0.5 ml)
• Appropriate anesthesia must be used for this procedure.
Animals may receive topical anesthetic or general anesthesia.
3-Blood Collection From Cardiac Puncture
• To collect up to 1 ml of blood within a short
period of time
• Must be performed under general anesthesia .
4-Blood Collection From Saphenous Vein
• The Saphenous Vein is located
along the lower portion of the
hind leg. This technique is
used for collecting small
volumes of blood.
• It can also be applied on rats
and guinea-pigs