THE EFFECTS OF DRUGS ON SMOOTH MUSCLE OF THE RESPIRATORY TRACT (THE TRACHEAL CHAIN PREPARATION)

1. INTERNATIONALISLAMIC UNIVERSITY MALAYSIA
KULLIYYAH OF PHARMACY
YEAR 2 SEMESTER 2
2013/2014 SESSION
PRACTICAL:
PHM 2252: BODY SYSTEM II: RESPIRATORY
TITLE: THE EFFECTS OF DRUGS ON SMOOTH MUSCLE OF
THE RESPIRATORY TRACT (THE TRACHEAL CHAIN
PREPARATION)
NAME : MUHAMMAD ‘IZZUDDIN BIN ZAMERY
MATRIC NO : 1217637
GROUP : A
DATE OF SUBMIT : 31ST MARCH 2014
LECTURER NAME : DR MARWAN AZ-ZUBAIDI

2. INTRODUCTION
Bronchodilators are used to treat bronchial asthma which characterized by the over
bronchoconstriction which complicates the inhalation and exhalation process. They act by
dilating the smooth muscle of the affected respiratory tract in bronchial asthma patient. Example
of drugs or bronchodilators available includes Adrenaline and Isoprenaline which are selective β2
adrenoceptor agonist, Ipratropium which is a Muscarinic receptor antagonists, and Theophylline,
Adenosine, A1 receptor antagonist. All of these drugs give effect on the relaxation of bronchial
smooth muscle and prevent or relieve bronchospasm in bronchial asthma patient.
The tracheal chain experiment using the organ bath technique demonstrates the effects of
various drugs and endogenous chemical mediators on the respiratory airways. In this experiment,
bronchodilators and bronchoconstrictors are used. The relative bronchodilator activity of drug
used in the treatment of bronchial asthma and the bronchoconstricting effects of known
(endogenous) mediator are demonstrated.
OBJECTIVES
- To exhibit the effect of bronchoconstriction of Acetylcholine, Histamine and 5-
hydroxytryptamine (Serotonin) on the smooth muscles of guinea pig trachea.
- To show the effect of bronchodilation of the Theophylline, Adrenaline/Isoprenaline on
the bronchial smooth muscle of guinea pig.
- To study the potency of the bronchodilators after the addition of bronchoconstrictor.
APPARATUS
- Syringe
- Organ bath
- Prepared Guinea-pig trachea smooth muscle
REAGENTS
- 10-6 M Acetylcholine
- 10-4 M Histamine
- 10-4 M 5-Hydroxytrptamine (serotonin creatinine sulphate monohydrate)
- 10-5 M Adrenaline
- 10-5 M Noradrenaline

3. - 5 x 10-6 M Isoprenaline (Isoproterenol)
- 5x 10-3 M Theophylline
PROCEDURE
1. A guinea-pig is euthanized by a blow on the head and the throat is cut.
2. Tracheal is dissected out, transferred to a dish containing Kreb’s solution and cut
transversely between the segments of the cartilage.
3. Several of the tracheal ring are tied together with a cotton thread so as to form a chain,
which is then mounted in Kreb’s solution
4. One end of the chain is attached to fixed pin in the bath and the other end is attached to a
pressure gauge.
5. Each drug (reagent) is left in contact with the tissue for 5 minutes.
6. The responses obtained with this preparation are slow in onset and last for a long period.
15 minutes cycle is enough for the tissue to completely recover from the effect of the
drug.
 Part 1
a) 0.1 ml dose of each drug is administered to the organ bath and the effects are measured
and recorded.
b) 15 minutes are need for effects of the drug to disappear before the administration of new
drug.
 Part 2
a) 0.1 ml dose of adrenaline / isoprenaline is administered and after it has acted for 3
minutes, without washing, acetylcholine is added and allowed to act for 5 minutes before
washing. The measurement is recorded.
b) 0.1 ml dose of adrenaline / isoprenaline is administered and after it has acted for 3
minutes, without washing, histamine is added and allowed to act for 5 minutes before
washing. The measurement is recorded.
c) 0.1 ml dose of adrenaline / isoprenaline is administered and after it has acted for 3
minutes, without washing, 5-hydroxytryptamine is added and allowed to act for 5 minutes
before washing. The measurement is recorded.
d) 0.1 ml dose of theophylline is administered and after it has acted for 3 minutes, without
washing, acetylcholine is added and allowed to act for 5 minutes before washing. The
measurement is recorded.

4. e) 0.1 ml dose of theophylline is administered and after it has acted for 3 minutes, without
washing, histamine is added and allowed to act for 5 minutes before washing. The
measurement is recorded.
f) 0.1 ml dose of theophylline is administered and after it has acted for 3 minutes, without
washing, 5-hydroxytryptamine is added and allowed to act for 5 minutes before washing.
The measurement is recorded.
RESULT
Table 1 : Direct effects of drugs on smooth muscle of respiratory tract
Drug Baseline (g) Effect (g) Baseline(g) Effect (g) Average(g)
Acetylcholine 1.02 1.08 1.03 1.20 1.14
Histamine 1.02 1.13 1.01 1.07 1.10
5-Hydroxytryptamine 1.02 1.20 1.02 1.09 1.15
Adrenaline 1.02 0.89 1.02 0.66 0.78
Noradrenaline 1.02 0.97 1.00 0.77 0.87
Isoprenaline 1.02 0.58 1.05 0.78 0.68
Theophyline 1.02 0.91 1.02 0.90 0.91
Table 2 :
Drug
Initial
effect (g)
Average
(g) [A]
Type of drug added Effect (g)
Average
(g) [B]
Difference
[A] – [B]
Acetylcholine 1.61 1.32 1.47 Adrenaline/Isoprenaline 0.70 1.15 0.93 0.54
Histamine 1.08 1.39 1.24 Adrenaline/Isoprenaline 0.86 0.99 0.93 0.31
5-Hydroxytryptamine 1.11 1.42 1.27 Adrenaline/Isoprenaline 0.84 1.19 1.02 0.25
Acetylcholine 1.37 1.24 1.31 Theophyline 1.32 1.20 1.26 0.05
histamine 1.28 1.24 1.26 Theophyline 1.22 1.21 1.22 0.04
5-Hydroxytryptamine 1.31 1.27 1.29 Theophyline 1.24 1.23 1.24 0.05
DISCUSSION

5. Different types of drug give different effect to the smooth muscle whether relaxation or
contraction depending on their classifications and affinities towards the smooth muscle. For
instance, they can be of constrictor or dilator and their extent of effect may be determined by the
distribution of specific receptors present on the smooth muscle. The class where the drugs
classified and their extent of effect towards any part of body can be tested by isolating any
smooth muscle then, observed the smooth muscle effect. In this practical, the smooth muscle of
guinea pig trachea is isolated and been tested with both classes of drug of different types to
exhibit their effect whether they will cause bronchoconstriction or bronchodilation on the smooth
muscles.
There are many types of receptor on the guinea pig trachea muscle in which the drugs can
bind to, such as Muscarinic receptor (M1 and M2 receptors), Histamine receptor (H1 receptor) and
Adrenergic receptor (β2 receptor). Drugs utilized in this experiment are Acetylcholine, Histamine
and 5-Hydroxytryptamine which are bronchoconstrictors as well Adrenaline and Theophyline
which are bronchodilators. Acetylcholine first have to bind to M1 and M3 Muscarinic receptors
on the smooth muscles and only then the muscle will exhibit contracting effect, thus causes
bronchoconstriction. This situation can be seen when one in resting and relax state. Similarly,
Histamine induces bronchoconstriction but the receptor it binds to is H1 receptor in which this
situation can be seen in the condition of hypersensitivity (allergy). 5-Hydroxytryptamine or also
known as Serotonin binds to 5-HT3 receptor which triggers the G-protein complex cascade to be
activated and leads to bronchoconstriction. The effects of these drugs are observed, recorded and
tabulated in the above table. It can be seen in table 1 and 2, after administrating those three drug,
they will cause the effect of increase in the contraction of muscle.
Conversely, Adrenaline/Isoprenaline and Theophyline have the opposite effects compare
with the previous three drugs. They give bronchodilation effect as they bind to β2 receptor and
this can be seen in the reduction of the reading from baseline and initial effect in table 1 and
table 2 respectively indicating the smooth muscle relaxation had occurred. Theoretically,
Noradrenalin should give little or no effect on the smooth muscles since it has low affinity to β2
receptor. Noradrenalin in table 1 shows only a little effect on muscle relaxation which is in line
with its theoretical effect.
Theophylline acts as a non-selective antagonist at Adenosine receptor (A1 receptor) and
give bronchodilation effect as it bind to the receptor. These compounds also act as
phosphodiesterase inhibitors, which produces additional anti-inflammatory effects, and makes
them medically useful for the treatment of conditions such as asthma. This effect was proven in
the tables above since the reading after the addition of Theophylline is decrease from the initial
effect and the baseline reading. Referring to the above table 2, the bronchodilation effect of
Adrenaline/Isoprenaline is more than of Theophylline which is indicated by the differences in the
reduction of readings (yellow highlighted) after the addition of both drugs.

6. There are some precautionary steps that have to be considered while conducting this
experiment. First of all, the organ bath should be set up accordingly in which the Kreb’s solution
should be added sufficiently until the smooth muscle fully immersed and then the aeration has to
be continuously maintained to make sure the smooth muscle is nourished and alive to give
accurate response. Then, the administration of different drug must be done by using different
syringe to avoid contamination of different drugs. Next, students have to ensure that the syringe
or hands do not touch the cotton thread since the thread of the pressure gauge is very sensitive to
vibration and it may affect the accuracy of the results.
Questions:
A. List the drugs that cause constriction and relaxation of the smooth muscle of the
respiratory tract in descending order of potency
Potency of the drugs is a measure of drug activity expressed in terms of the amount required to
produce an effect of given intensity. A highly potent drug evokes a larger response at low
concentrations, while a drug of lower potency evokes a small response at low concentrations. So,
according to this experiment:-
 Potency of the drugs that can cause relaxation of the smooth muscle in descending order
is as follows (according to table 1):
o Isoprenaline > Adrenaline > Theophylline
 Potency of the drugs that can cause constriction of the smooth muscle in descending
order is as follows (according to table 1):
o Serotonin > Acetylcholine > Histamine
B. Describe the effect of Adrenaline/Isoprenaline and Theophylline have on the actions
of acetylcholine, histamine and 5-hydroxytryptamine
According to table 2 above, the administration of Acetylcholine, Histamine and 5-
Hydroxytryptamine to the tissue causes a significant reading indicating the constriction effects.
Then, after 3 minutes, Adrenaline/Isoprenaline and Theophylline are added to the contracted
tissue which gives out the effect of the smooth muscle relaxation. However, from the highlighted

7. reading in table 2, it can be seen that, Adrenaline/Isoprenaline relaxes the smooth muscle more
than Theophylline.
C. What is the term used to describe this kind of drug interactions? Provide the basic
for such an answer.
This interaction is known as antagonism. Antagonism means that the effect of one drug say for
example bronchoconstrictor is opposed with the action of bronchodilator. The drugs which
opposed may act on same receptor as the already-binding drugs/ligands by displacing them or
may act on other receptor that can give out opposite effect. In this experiment, the
Adrenaline/Isoprenaline and Theophylline opposed the effect of Histamine, Acetylcholine and
Serotonin by binding to different receptor that give out opposite effect (constriction to
relaxation).
D. Discuss the use of the above drugs that relax the smooth muscle of the respiratory
tract in bronchial asthma.
Bronchial asthma is a complication characterized by the over constriction of bronchial smooth
muscle (over bronchoconstriction) which results to the difficulties in breathing.
Adrenaline/Isoprenaline acts as agonist of β2 receptor, the stimulation of β2 receptor causes the
relaxation of bronchial smooth muscles and lead to bronchodilation. The other drug which is
Theophylline acts as a non-selective antagonist at Adenosine receptor (A1 receptor) and give
bronchodilation effect as it bind to the receptor. As mentioned above, this drug also acts as
Phosphodiesterase inhibitors, which produces additional anti-inflammatory effects, and makes
them medically useful for the treatment of conditions such as asthma. Since,
Adrenaline/Isoprenaline and Theophylline can relax the bronchial smooth muscle, they can be
used as the treatment of bronchial asthma.
REFERENCES
Brenner, G. M., & Stevens, C. Bronchodilators. Pharmacology (4th ed). Elsevier Health Sciences
(2009). Retrieved from: https://www.inkling.com/read/pharmacology-brenner-stevens-
4th/chapter-27/bronchodilators

8. Townsend, E. A., Yim, P. D., Gallos, G., & Emala, C. W. Can We Find Better Bronchodilators
to Relieve Asthma Symptoms. New York: USA (2012). Retrieved from:
http://www.hindawi.com/journals/ja/2012/321949/
Caffeine and theophylline as adenosine receptor antagonists in humans. Retrieved from:
http://www.ncbi.nlm.nih.gov/pubmed/1865359