INDEX
INDEX
| Sr.No. | Title of the Experiment | Page no. |
| 1. | General introduction to experimental. | 3 |
| 2. | To study different laboratory animals and their application in the experimental pharmacology. | 4 |
| 3. | Legal regulation for the use of experimental animals. | 10 |
| 4. | To study standard techniques for sacrifice and making animal unconscious. | 11 |
| 5. | Study of some basic instruments used for isolated tissue experiment. | 12 |
| 6. | Preparation and use of physiological salt solution. | 15 |
| 7. | Useful hints for ambitious students. | 17 |
| 8. | To study the relationship between dose of Acetycholine and contractile response by using chick ileum. | 18 |
| 9. | To record the effect of various spasmogenic and spasmolytic drugs on guinea pig ileum and scheme for identification of unknown drug. | 21 |
| 10. | To study the effect of various drugs on perfused | 25 |
| 11. | To study the effect of various drugs on isolated | 26 |
| 12. | Discussion of the effect of various drugs on frog | 27 |
| 13. | To study the scheme for identification of unknown drug using frogs heart. | 30 |
| 14. | To evaluate the local anaesthetic agent by various methods. | 31 |
| 15. | To study the convulsion action of strychnine on frog. | 34 |
| 16. | To study the hypnotic effect on mice. | 35 |
| 17. | To study the effects of Mydriatics and Miotics on rabbit eye. | 36 |
| 18. | To study the effect of anti-psychotic drugs using cooks pole climbing apparatus. | 39 |
| 19. | To evaluate antiepileptic activity of drug using maximum electro convulsive shock seizures ( M. E. S.) and chemical induced convulsions methods. | 40 |
| 20. | To study pyrogen testing on rabbit. | 41 |
| 21. | To study the taming effect of chlorpromazine in rat and mice. | 44 |
| 22. | To demonstrate the evaluation of the analgesic potency of drug. | 45 |
| 23. | Reference | 48 |
General introduction to experimental Pharmacology.[1]
Practical.No.: 1
Aim: General introduction to experimental Pharmacology.[1]
GENERAL CONSIDERATIONS :
Pharmacology is the study of the effects of drugs on living systems. It attempts to characterize the biological actions of chemical and to define the mechanisms by which these actions are evoked. A pharmacologist seeks to describe and quantify both the desirable and undesirable effects in laboratory animals and to establish the conditions under which such observations can be extrapolated to the human population. Experimental pharmacology thus helps in understanding the nature of drug action and the vulnerability of the living systems to the alternation by chemicals that serves as the basis on which a) New therapeutic agents are developed, and b) Toxic consequences of chemical exposure may be alleviated.
The experiments can be carried out in whole animal (In-vivo) or in isolated organs(In-vitro). In whole animal experiments (e.g. Dog-blood pressure) the effect of drugs on any organ is influenced by many factors such as metabolic state, blood supply, nervous control, hormonal influences, factors of bio-availability etc. It is difficult to estimate and control the influences of such a diversity of factors. Elucidation of action of drug may be hindered in such cases. The use of isolated ‘ in vitro’ preparations provides distinct advantage to facilitate the experimental investigations concerning mode of actions of drugs. The experimental conditions can be well controlled precisely in these experiments.
The effect of drug can be studied independent of other systems thus relating direct mechanism of action of drug. The isolated preparations are commonly used in experiments because of these advantages. The intact experiments (i.e. whole animal experiments) are also of importance to study the effect of drugs, the interaction between two drugs and between the body factors and drugs.
Before starting experiments in pharmacology it is essential to understand about laboratory and experimental animal . One should also know about their maintenance and handling.
To study common laboratory animals and their application in experimental pharmacology.[2]
PracticalNo.: 2
Aim: To study common laboratory animals and their application in experimental pharmacology.[2]
Common laboratory animals (rat, mice, guinea-pig, rabbit and hamster) are those animals, which can be bred and reared (maintained) in the laboratory under suitable conditions to provide a reliable supply of uniform and healthy animals for the purpose of experimentation. The other animals used for experimental purpose are cat, dog, frog, monkey, pigeon etc, which can be obtained from outside sources as and when required; but it is always better to keep them under quarantine for a few weeks before they are utilized.
Handling of laboratory animals involves two most important responsibilities on the part of the experimenter. First, the animal is handled with utmost care so that it does not suffer any pain, and second, a due regard is paid towards the health and well being of the animal colony. Even when they are killed at the end of the experiment, it should be done by a humane method, i.e. euthanasia, which means painless-killing.
RAT (Rattus norvegicus : Norway Rat)
Three strains are commonly available : Wistar strain, Sprague-Dawley and Wistar Kyoto. However two original strains of albino rats have been widely used through-out the world.
Wistar ratis a quite, moderately prolific strain rather resistant to the infection, and has a low incidence of spontaneous tumors. The head is wide, specially in the male, and the ears are long. The tail-length is always less than the body-length.
Sprague-Dwaley rat is more rapidly growing and more prolific than Wistar, has a longer and narrower head, and a longer tail, which may equal the body length. It is also less resistant to infection, specially to respiratory diseases.
Advantages and characteristics :
It is small in size compared to other animals so drugs are required in small quantity. Rodents do not vomit because the lack the vomiting center (so drug can be administered orally). Rats do not have tonsils or gall bladder. Because of the absence of gall bladder in rat there is continuous flow of bile into the intestine. This facilitates the study of drug acting on bile, cholesterol re-absorption etc. They have extremely diffuse pancreas and hence pancreactectomy is difficult to perform in order to produce a diabetic rat. The stomach shows an obvious division into two parts by a prominent white somewhat curved transverse ridge (limiting ridge). The upper two-fifth non-secretory portion (rumen) is translucent and thinner than the lower three-fifth glandular secretory portion. Histologically
also the two areas show distinct differences. The glandular portion is composed of two clearly defined areas that can be distinguished both by naked eyes as well as under a microscope. The antrum is roughly semicircular area straddling the lesser curvature below the limiting ridge. It is relatively thin and translucent, and has a slightly irregular inner lining. The remainder of the lower two third comprises the body of a small area above the pylorus. This is the thick portion of the stomach with the mucosa thrown into folds. This part of the stomach is analogous to the body of the stomach in the man rather than to the fundus both anatomically and functionally. The rat being omnivorous resembles man nutritionally. The gastric acid secretion is continuous.
Oestrus cycle : The cycle makes it’s appearance at puberty at the age of two to three months, and the whole cycle lasts for about four to five days and can be divided into four stages according to the cell types found in the vaginal smears as follows:
- Oestrus (9-15 hrs): It is characterized by sexual receptivity when the female will allow copulation.durin this period there is increased running activity, quivering of the ears and lordosis in the presence of another rat. The vaginal smear shows cornified epithelial cells only. It lasts for 9-15 hrs and ends with ovulation. Experimentally it can be induced by administration of 100 ug/kg i.p. Diethylstilbestrol.
- Metaoestrus (about 20 hrs): It occurs shortly after ovulation. Leucocytes starts appearing in this stage and predominate over cornified epithelial cells. It lasts for about 15-18 hrs.
- Diestrus (60-70 hrs.): It is the longest stage. Vaginal smear shows only leucocytes.
- Proestrus (about 20 hrs) : It is characterized by nucleated epithelial cells either singly or in groups (sheets).It is a preparatory phase preliminary to the next oestrus phase.
Vaginal smears may be taken in the following way to confirm the actual stage. Cotton swab made with a tooth picks is moistened with saline and gently inserted and slightly rotated within the vagina. The swab is then pressed in a drop of saline on a microscope slide and examined under low power. An alternative and better method is to take the vaginal wash with a few drops of saline with the help of a capillary pipette fitted with a rubber teat.
INDICATIONS IN THE ESTROUS CYCLE
| Stage | Superficial | Vaginal | Uterus | Ovary | Approxi | Remarks |
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| Genitalia | Smear |
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| Proest | Lips slightly | Epithelial | Increased | Ovarian | 12 | Heat may |
| rus | swollen | Cells only | distension | Growth |
| occur |
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| Follicle |
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| Vagina dry |
| fluid |
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| enlargement |
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| Estrus |