INTERNATIONAL MEDICAL UNIVERSITYMALAYSIA

Bachelor of Pharmaceutical Chemistry (Hons)

Intake PC113Semester 2

General Pharmacology

Name: Mohd Yusri bin Mohd Yusoff

Student ID : PC 0913034179

Part I. Effect of Acetylcholine on the Guinea Pig Ileum

Part II. Effect of Atropine on responses to acetylcholine in the Guinea Pig Ileum

INTRODUCTION

This experiment is called the organ bath protocol. The main aim for this experiment is to observe the effect of an agonist, acetylcholine on the guinea-pig ileum preparation. Guinea-pig ileum contains a smooth muscle preparation exhibiting little spontaneous contractile activity. Besides that, the ileum also have numerous receptors on it. This experiment resembles an in-vitro tissue experiment.

Acetylcholine, the first neurotransmitter discovered, was originally described as "vagus stuff" by Otto Loewi because of its ability to mimic the electrical stimulation of the vagus nerve. It is an agonist where a substance that initiates a physiological response when combined with a receptor. It is now known to be a neurotransmitter at all autonomic ganglia, at many autonomically innervated organs, at the neuromuscular junction, and at many synapses in the CNS. The AcH acts on muscarinic receptors to cause contraction of the guinea pig ileum. Generally muscarinic receptors are found on myocardial muscle, certain smooth muscle, and in discrete CNS regions. Apart from that, the response is slow and prolonged. Only one factor affect the response which is the dose of AcH.

For the second part, it involves antagonist on muscarinic receptors which in this case we use atropine. A receptors antagonist functions by blocking agonist-mediated responses rather than provoking a  biological response itself upon binding to a receptor. Atropine is described as a competitive antagonist of the muscarinic cholinergic receptors. This makes it easily absorbed into the gastro intestinal tract and then excreted in the urine. Atropine is very useful when it comes to treating second degree heart block. By increasing the dose of acetylcholine, the antagonism by atropine can be overcome.

The structure of acetylcholine

The structure of atropine :

METHODOLOGY

Isolated tissue in physiological solution (Krebs-Hanseleit solution ) Guinea pig ileum Tissue-suspendor Continuous aeration (95% Oxygen, 5 % Carbon Dioxide) Temperature regulation (37 degree C ) Drugs to be injected into bath (max 5 ml) Stocks solution of drugs A/D converter –PowerLab

First we need to test whether the tissue is responsive or not : Add 0.3ml of 1E-4 M (FBC is 3E-6) leave drug in bath for 1 minute If there is no contraction, the tissue is dead If there is a contraction, allow it to peak for 1 minute. Wash away the drug When tissue’s response is about pre drug level, start the experiment.

Protocols

10ml Krebs aerated in organ bath Add to 200 ml of Krebs-Hanseleit solution into reservoir Drug dilution 10-2M to 10-9M prepared by serial dilution. Add 100microlitre , 300microlitre and 500microlitre of acetylcholine of

each concentration. Wash by overflow for 5 second

Final Bath Concentration= [stock]*volume addedBath volume

To start the second part, add 1E-8M of mepyramine to the reservoir Run the antagonist containing solution through the tubing into the organ

bath.

RESULTS Result for Part 1

Result for Part 2 ( Agonist and Antagonist )