Principle
Rabbit intestine is a smooth muscle which shows regular pendular movement (i.e. continuous contraction and relaxation). Therefore, to study the effect of drugs on intestinal movement rabbit intestine is ideal preparation. Rabbit intestine is supplied by autonomic nervous system. Rabbit intestine contains muscarinic receptors and adrenergic receptors. Muscarinic receptor agonist like Ach produces contraction of rabbit intestine and physostigmine increases the spasm and pendular movements. The muscarinic actions and effects can be blocked by muscarinic blockers.
Adrenaline acts on α and β receptors and exhibit inhibitory effect on pendular movements. The actions of adrenaline are blocked by adrenoceptor blockers viz. α-receptor blockers and β-receptor blockers.
Equipment required
1) Kymograph
2) Student thermostatic organ bath
3) Aeration tube
4) Aerator
5) Aeration tube holder
6) Frontal writing lever
7) Lever holder
8) Screw clip
9) Haemostatic forceps
10) Mariotte bottle
11) Rubber tubes
12) Tuberculine syringe
13) 26 no. needle
14) Scissor
15) Forceps
16) Suturing needle
17) Carbogen gas cylinder with regulator.
Animal Required
Rabbit intestine.
Physiological solution required
Tyrode solution.
Drug solution required
1) Acetylcholine 20µg/ml
2) Physostigmine sulphate 10µg/ml
3) Atropine sulphate 4µg/ml
4) Adrenaline hydrogen tartrate 20µg/ml
5) Propranolol hydrochloride 100µg/ml
Procedure
1) Fast a rabbit for 12 hour period.
2) Set up assembly for rabbit intestine experiment.
3) Balance frontal writing lever using plasticine and apply a tension of 500 mg.
4) Fill the outer jacket of student organ bath with water.
5) Set the thermostat of the student organ bath at 370C and put the switch on.
6) Fill the mariotte bottle with tyrode solution and control the flow of tyrode solution to inner organ bath using haemostatic forceps.
7) When the temperature of the tyrode solution reaches 370C, sacrifice a rabbit by giving a blow on its head and cutting the carotid artery.
8) Open the abdominal region and identify the intestine.
9) Remove the intestine and place it in a petri dish containing tyrode solution at 370C and trim away the mesentry.
10) Cut a piece of intestine (approximately 3-4 cm) and using surgical suturing needle tie a thread at each end, taking care to see that the lumen of the intestine remained open.
11) Tie one end of the thread to the hook of the aeration tube and the other a frontal writing lever.
12) Stabilize the tissue for the 30 minutes period.
13) Aerate the rabbit intestine in the inner organ bath with a mixture of oxygen (95%) and CO2 (5%) (carbogen).
14) Required the normal pendular movement of the rabbit intestine on the drum for 30 seconds. At the end of 30 seconds add 0.1ml of Ach into the inner organ bath and record the spasmogenic effect of the drug for 30 seconds. After recording the response for 30 seconds switch off the kymograph. Immediately open the outlet of the inner organ bath and remove the tyrode solution present in the inner organ bath. Again fill the inner organ bath with fresh tyrode solution and keep it for 60 seconds. Repeat the washing procedure 3-4 times or till the writing point of frontal lever comes to the normal pendular movement base line.
15) Then record the response of 0.1ml physostigmine solution. Give 3-4 washing for recovery.
16) Record the response of 0.1 ml adrenaline solution. Give 3-4 washing for recovery.
17) Add 0.1 ml of atropine sulfate solution into the inner organ bath, allow it to act for 60 seconds then record the response of Ach in the presence of atropine sufate. Give 3-4 washing for recovery.
18) Add 0.1 ml of atropine sulfate solution into the inner organ bath, allow it to act for 60 seconds then record the response of physostigmine in the presence of atropine sufate. Give 3-4 washing for recovery.
19) Add 0.1 ml of propranolol solution into the inner organ bath, allow it to act for 60 seconds then record the response of adrenaline solution in the presence of atropine propranolol. Give 3-4 washing for recovery.
Observation and conclusion
Acetylcholine produces contraction in rabbit intestine. Atropine sulfate is a muscarinic receptor blocker; therefore it reduces the spasmogenic effect caused by Ach.
Physostigmine also produces mild spasmogenic effect and increases the peristaltic movement of the intestine. Atropine sulphate blocks the effect of the physostogmine.
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