Published on June 8, 2019
1. Experimental Pharmacology
2. • Experimental pharmacology – great imp • Experimenter should take outmost care • Sacrificing - human • Euthanasia – painless killing • Laboratory animals – breaded and handled in laboratory •Mice •Rat •Guinea pig •Rabbits
3. RAT Species: Rattus norvegicus • Wister starin is commonly used • Other strains • Wistar rat • Sprague Dawley rat • Bio breeding rat • Long-Evans rat • Zucker rat • Hairless rats • RCS rats • Shaking rat Kawasaki
4. ADVANTAGES AND CHARACTERISTICS • Small in size • Drug to be tested required in small quantity • Vomiting center is absent – oral administration can be done • Gall bladder and tonsils are absent Continuous flow of bile into intestine This facilitates the study of the drugs acting on bile, cholesterol reabsorbing ect. • Pancreas are diffused therefore difficult to produce pancreatomy • Stomach, fundus and pyloric parts are clear lining between them • Gastric acid secretion is continuous
5. EXPERIMENTAL USE • psychopharmacological studies • Study of analgesics and anticonvulsants • Bioassay of various hormones, such as insulin, oxytocin, vasopressin • Study of oestrus cycle, mating behaviour and lactation • Isolated tissue Pre's • Chronic study • Gastric acid secretion • Hepatotoxicity studies • Study on mast cells
6. MICE ( Mus musculus) • ADVANTAGES • Smallest • Easy to handle • Cheap • Less drug required EXPERIMENTAL USES • Toxicological studies • Teratogenicity studies • Bioassay of insulin’ screening of analgesic and anticonvulsant • Screening of chemotherapeutic agents • Studies related to genetics and cancer research • Drugs action on CNS • Swiss albino mice are commonly used species • Other strains are – Balb/C and C-57
7. • Normal temperature:- 37.4 • Pulse rate:- 120 • Oestrous cycle:- 4-5 days • Gestation period:- 19-21 days • Weaning age:- 19-21 days • Mating age:- 6-8 weeks • Litters:- 8-12 yearly; average litter, 7-8 • Room temp.:- 20-21 • Humidity:- 50-60 % • Weight- weaning : 7g ,adult:- 25-28 g
8. Cage(Mouse) • Many different designs of mouse and no one pattern is the standard • Aluminium box approx. 6X12X6 in deep with tapering side to facilitate stacking . • The lids are made of steel sheet or of strong wire mesh and are designed so that hopper is built into them and accommodation provided to hold the drinking bottle. • The cages are light, durable and easily sterilized by dry or moist heat.
9. Feeding(Mouse) • Pelleted diets such as diet 86 of Howie(1952) or diet 41 of Bruce (1950) are satisfactory • Fresh water in drinking bottles must be provided
10. Handling(Mouse) • An assistant takes a grip on the middle of tail of the animal with the left hand and gently raises the hind limbs floor of the cage • A mouse held in the position cannot turn around the bite. • Then with the right finger and thumb a fold of skin is taken up as close as possible to the head. • The animal can now be lifted into convenient position for the operator to carry out simple inoculation procedures. • Place the animal on a rough surface and hold it by its tail with right hand, then pick up loose skin at the base of neck with the left forefinger and thumb, lift and turn the left hand palm uppermost at the same time catching the tail and pressing it against the palm with the left little finger. • The right hand is free to pick up the syringe
11. GUINEA PIGS ( CAVIA PORCELLUS ) • Docile animals • Highly susceptible to TB and anaphylaxis • Highly sensitive to histamine, penicillin • Required exogenous vit c in diet
12. • Rectal temp.:- 37.6-38.9 • Normal respiration rate:- 80 per minute • Pulse rate:- 150 per minute • Gestation period:- 59-72 days( avg. 63 days) • Weaning age:- 14-21 days • Mating age:- 12-30 weeks • Litters:- 3 yearly ;average litter,3 • Room temp:- 18.5- 21 • Humidity:- 45% • Weight –weaning :- 120g, adult:- 200-1000g
13. Cage (Guinea pig) • Stock runs should be abt. 4Å~6 ft. and 1 ft. 8 in high • One square foot of space should be allowed for each animal • not more than 25 animals should not be kept in any one pen. • For expt. Animals galvanized iron cages are recommended and sterilized. • A convenient size of 14X9X8 in fitting in a tray 1.5 in deep
14. Feeding(Guinea pig) • A diet in pelleted form is recommended in preference to mashes. • Diet of Bruce and Parks (1947) contains balanced proportions of protein, fats and carbohydrate with added vitamins salt and trace element. • Crushed oats 2 part+ Broken bran 1 part • Supplemented with cabbage and hay • Necessary to add fish or meat meal.
15. Handling • Place one hand across the back of the animal with thumb behind the shoulder and the other fingers well forward on the opposite side • Lift the animal gently and support its weight with other hand placed palm uppermost under the hind quarters.
16. EXPERIMENTAL USES • Evaluation of bronchodilators • Anaphylactic and immunological studies • Study of histamine and antihistamines • Bioassay of digitalis • Evaluation of local anaesthetics • Hearing experiments because of sensitive cochlea • Isolated tissues especially ileum, heart • Study on TB and ascorbic acid metabolism
17. RABBITS (Lupas cuniculus) • Docile animal • New Zealand white strains are widely used • It has huge caecum and long appendix • Enzyme atropine esterase is present in rabbit liver and plasma so it can tolerate large • Doses of belladonna (atropine)
18. • Rectal temperature:- 38.7 C-39.1 C • Normal respiratory rate:- 55 per minute • Pulse rate:- 135 per min • Gestation period:- 28-31 days • Weaning age:- 6-8 weeks • Mating age:- 6-9 months • Litters:- 4 yearly ; average 4 litter • Room temp:- 15.5-18.5 • Humidity:- 40 -45 percent • Weight – adult:- 0.9-6.75 kg
19. Cage • Cages are best made of galvanized iron. • The minimum size for a medium sized rabbit is 2X2X1(1/2)ft., • Young rabbits up to 3 months of age may be housed together but after that time sex should be separated. • From 8 -10 young rabbits may be kept together in pen similar to that used for guinea pigs.
20. Feeding(Rabbit) • Pelleted diet 18 of Bruce and Parkers(1947) or commercial breeders pellets are suitable • Daily supply of 2.5 0z(72 gm) of a mixture of one part oats and three parts bran may be fed as a slightly moist mash • Green stuffs or root vegetables • Clean drinking water
21. Handling(Rabbit) • Smooth ear of the rabbit back • pick up the ears and loose skin at the back of the neck with one hand in a firm grip • place the other hand under the hind quarter to support the weight and lift gently. • Never be lifted by ear alone • Should be placed on a non-slippery surface • If restraint is required during anaesthesia or inoculation , should be wrapped in a roller towel or placed in a special box.
22. EXPERIMENTAL USES • Pyrogen testing • Bioassay of anti-diabetic, curariform drugs and sex hormones • Screening of agents affecting capillary permeability • Drugs used in glaucoma • Pharmacokinetic studies • Studies related to antifertility agents • Isolated preparations like heart, duodenum, ileum, Pre's • Study on local anaesthetics • Study on meiotic and mydriatics
23. FROGS (RANA TIGRINA) • Most commonly used in physiology, pharmacology and toxicology • Used before 200 years • Amphibian animal safe to handle • Cannot breed in lab
24. EXPERIMENTAL USES • Isolated preparations, heart, rectus abdominis muscle • Drugs acting on CNS • Drugs acting on NMJ
26. • Reliability and irreversibility • Minimum pain, distress, anxiety or apprehension • Minimum delay until unconsciousness • Safety and emotional effect on personnel • Compatibility with requirement and purpose, including subsequent use of tissue and • • Compatibility with species, age and health status • Euthanasia methods are broadly classified as; • Chemical methods (Inhalants/Non-inhalants) • Physical methods.
27. Inhalant Anesthetics • Halothane • Enflurane • Sevoflurane • Methoxyflurane • Isoflurane • Desflurane are preferred for euthanasia in animals.
28. CO2 • Carbon dioxide is an effective and widely used agent to euthanize rodents due to rapid hypoxia which further leads to depression of vital centers. (Maintain 20% -70% of the chamber volume per minute). • This method is preferred in several animal but it is dangerous to use, so precaution should be taken while using it
29. Nitrous oxide • N2O is not preferred due to lack in fast onset of anesthesia, but may produce hypoxemia and cardiac or respiratory arrest. • However, it may be used in combination with other anesthetics to speed anesthesia onset.
30. Ether • Ether was formerly used extensively, but is now only acceptable conditionally. • The reason is being irritant to mucous membranes and risk of fire and explosion. • The use of ether is prohibited in many countries.
31. Non-inhalant Anesthetics • Barbiturates: • Sodium pentobarbital is the most rapid and reliable method of euthanasia for most experimental animals. • In non-rodent species, barbiturates are given intravenously to be most effective. • Intraperitoneal injection of barbiturates is acceptable for euthanasia in small mammals.
32. Potassium chloride (KCl) • KCl induces immediate cardiac arrest without any significant depression of the central nervous system. • Hence, it must only be used after the animal is deeply anesthetized
33. Magnesium sulphate (MgSO4) • MgSO4 produces its action through cardiac arrhythmia, neuromuscular blockade and deep anesthesia, hence ultimately animal gets euthanized due to cardiac arrest and neuromuscular blockade.
34. Neuromuscular Blocking Agents (Succinycholine, Curare, etc.) • These agents induce muscular paralysis and death because of suffocation. • Distress onset is more, hence less preferred for euthanasia.
35. Physical Methods • Physical methods are performed by skilled and experienced personnel with appropriate, well maintained equipment.
36. • Cervical dislocation: Humane technique for euthanasia which is frequently used for mice, rats, guinea pig, rabbits (weighing less than 1 kg) and other rodents. • Decapitation: Decapitation may be used to euthanize rodents and small rabbits. Except in neonatal animals, a guillotine is generally used.
37. Microwave irradiation • Special instruments designed (appropriate power and microwave distribution) for this purpose, which is used when an experiment requires fixation of mouse or rat brain metabolites in vivo without losing anatomic integrity of the brain.
38. Penetrating captive bolt • This method is conditionally acceptable and made for ruminants, horses, and swine when chemical agents are scientifically contraindicated. • This method is not employed in the laboratory animals.
39. Euthanasia of Poikilothermic (Cold-blooded) Animals • The euthanasia of poikilothermic animals is different due to differences in the pharmacokinetic, respiration and tolerance to cerebral hypoxia between these species and homeothermic animals. • Chemical agents: • Pentobarbital, • Tricainemethane sulfonate or • benzocaine HCl.
40. • Intraperitoneal administration of pentobarbital is an effective method of euthanasia in amphibians. • Tricaine methane sulfonate or benzocaine hydrochloride may be placed in the water of amphibians and fish to produce anesthesia and prolonged contact may produce death. • Inhalant anaesthetics may be used for amphibians and reptiles but due to the low oxygen requirements for amphibian, the onset of unconsciousness and death will be significantly lengthened.
41. Physical methods • Poikilotherms may be euthanized by stunning followed by decapitation or pithing to ensure death. • In frogs and toads, pithing the brain (single pithing) and spinal cord (double pithing) are effective and acceptable methods.