Basic Medical Sciences, Kulliyyah of Medicine, International Islamic University of Malaysia

THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

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Pharmacology : Autonomic nervous system agonists and antagonists Pharmacology : Routes of administration Elective Practicals

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NAME MATRIC NO

THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

AUTONOMIC NERVOUS SYSTEM : AGONISTS AND ANTAGONISTS

DATE GROUP MEMBERS NO 1. 2. 3. 4. NAME MATRIC NO

Objectives: To study the effects of Sympathetic and Parasympathetic agonists and antagonists on the anaesthetized cat. Procedure: The cat is anaesthetized with Pentobarbitone (40mg/kg i.p.) and the trachea is cannulated. In order to record the blood pressure changes, the carotid artery is cannulated and connected to a pressure transducer. The heart rate can also be recorded simultaneously. The vagus nerve is identified and is prepared for electrical stimulation. For recording the movements of the ileum, a balloon filled with water placed in the ileum of the cat and is then connected to a transducer. The nictitating membrane of the eye contains aipha-adrenergic receptors and its contraction on stimulation, can also be recorded via a transducer. The femoral vein is cannulated for administration of the drugs to be studied. Experiment 1. Sympathetic Cat la. When the conditions have stabilised in the l cat preparation, the basal BP, heart rate, and contraction of the nictitating membrane is recorded. lb. Noradrenalin 3 g/kg, Adrenaline 3 g/kg and Isoprenaline 3 g/kg are injected one after another and their effects on BP and HR are recorded. Ic. Next, Phenoxybenzamine 3mg/kg is injected.

1 d. Wait for 10 minute, stimulate the nerve to the nictitating membrane and repeat lb. 2a. 2b. In the 2nd cat, repeat procedures la and lb. Inject Propranolol 2mg/kg and wait for 10 minutes, and repeat lb.

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

Experiment 2. Parasympathetic Cat 3a. When conditions in the 3rd cat have stabilised, the BP, HR and the contractions of the ileum are recorded. 3b. 3c. 3d. 3e. 3f. 3g. 3f. Stimulate the vagus nerve and record the BP, HR and gut motility. Inject Acetylcholine 2g/kg and record the results. Inject Carbachol 2g/kg and record the results. Now inject Atropine 2mg/kg and wait for 10 minutes. Repeat procedures 3b, 3c, and 3d and record the results. Inject Acetylcholine 300g/kg and record the results. Inject Nicotine 200g/kg and record the results.

All students are required to enter their results with a short explanation of the results in the attached schedule list and submit them to the lecturer immediately for assessment.

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

PARASYMPATHETIC CAT

Blood Pressure Systolic/Diastolic Vagus Stimulation Control Test Acethycholine 2g/kg Control Test Carbachol 2g/kg Control Test Atropine3 mg/kg Control Test Acethycholine 2g/kg Control Test Carbachol 2g/kg Control Test Acethycholine 300g/kg Control Test Nicotine 200g/kg Control Test

Mean Arterial

Heart Rate ( BPM)

Gut Motility

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

Sympathetic Cat 1

Blood Pressure Systolic/Diastolic Noradrenaline 3g/kg Control Test Adrenaline 3g/kg Control Test Isoprenaline 3g/kg Control Test Phenoxybenzamine 3 mg/kg Control Test Noradrenaline 3g/kg Control Test Adrenaline 3g/kg Control Test Isoprenaline 3g/kg Control Test

Mean Arterial

Heart Rate ( BPM)

N. Membrane Contraction

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

Sympathetic Cat 2

Blood Pressure Systolic/Diastolic Noradrenaline 3g/kg Control Test Adrenaline 3g/kg Control Test Isoprenaline 3g/kg Control Test
Propranolol 2 mg/kg

Mean Arterial

Heart Rate ( BPM)

N. Membrane Contraction

Control Test Noradrenaline 3g/kg Control Test Adrenaline 3g/kg Control Test Isoprenaline 3g/kg Control Test

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

ROUTES OF ADMINISTRATION
DATE GROUP MEMBERS NO 1. 2. 3. 4. NAME MATRIC NO

General Instructions Students will be assigned to work in groups. Each group is given 10 mice. Handling of Mice Lift the mouse by grasping the tail at the caudal end with the right hand, and allow it to grip the wire mesh of the cage with its forelegs. With the left hand, grasp the nape of the animals neck between the thumb and the index finger. Place the tail from the right hand to the left small finger so that the mouse is held tightly in the left hand. The mouse is now ready for injection. Introduction The common routes of administration of drugs are oral, subcutaneous, intramuscular, intravenous, intrathecal, rectal, nasal, and topical application. Each of these routes has its particular advantages. Some substances may be effective when administered by one route but relatively or completely inert when given by another. The mode of entry of a drug into the body determines, to a large extent, the character, speed, dose and degree of action. A. ORAL ADMIINISRATION Administer by a stomach tube or by an oral feeding needle 35 mg/kg (0.1 ml/l0 gm body weight) of pentobarbital sodium (Nembutal) to a mouse. Note the time of administration of the drug and observe the animal for its effect. The mouse is held firmly and the feeding needle is held well towards the back of the mouth and near the upper palate. The needle is than passed-gently-not-forced down the esophagus. As it passes into the stomach (about 2 inches down) the animal will exhibit a definite and characteristic gagging. As much as 0.5m1/10 gm of animal may be administered. Note the several grades of response as follows: 1. Minimal spontaneous activity with normal responsiveness to stimulation; 2. Absence of spontaneous activity with uncoordinated locomotion upon stimulation; 3. No locomotion response to stimulation, but righting reflex retained. 4. Righting attempted without success; to

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

5. Immobility and righting not attempted In the table, record the nearest minute to the time of onset and the duration of loss of the righting reflex.

B. SUBCUTANEOUS INJECTION The drug is to be introduced directly underneath the skin. A 3/4 -l inch 26-gauge needle is employed. Insert the needle to its full length to avoid loss of liquid upon withdrawal. Inject 35 mg/kg (0.1 ml/l0 gm of body weight) of pentobarbital sodium under the skin of a male mouse followed by gentle massage of the site of injection. The common site employed is the dorsal neck area and that of the abdomen. Observe as under A. and record the data in the table provided. C. INTRAVENOUS INJECTION The mouse is to be held firmly by wrapping it in a cloth or by placing it in a suitable animal holder. With a little practice the tail veins can be recognized readily. It is helpful to dilate the veins by warming before attempting the injection. Employ a sharp, 26-gauge needle and slowly inject 35 mg/kg (0.1 ml/ 10 gm of body weight) of pentobarbital sodium into the mouse and record your observations. If the needle is not in the vein, resistance will be felt while injecting and blanching will be observed in the surrounding connective tissue. D. INTRAPERITONEAL INJECTION This is a common method to introduce drugs into animals. The drug is injected into the peritoneal cavity where absorption is rapid. By holding the animals head downward, inject into the lower half of the abdomen; intraintestinal or intravisceral injection may be avoided. Inject 35 mg/kg (0.1 ml/l0 gm of body weight) of pentobarbital sodium into the mouse, compare effects with those noted in previous sections, and record the data in the table. E. INTRAMUSCULAR INJECTION Inject 35 mg/kg (0.1 ml/l0 gm of body weight) of pentobarbital sodium into the gluteus maximus muscle of a mouse, using a 26-gauge needle. One must always pull back on the plunger to make certain the needle is not in a vein Make your observation and record the results in the table provided. At the end of the session, pool the data obtained from each student in your lab. Obtain the mean and standard deviations of the values that your lab group has obtained.

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

EFFECT OF DIFFERENT ROUTES OF ADMINISTRATION Route of Administration Group Onset Oral Subcutaneous Intravenous Intraperitoneal Intramuscular Data Duration Class Onset Average Duration

For group, calculate: 1. Mean time of onset standard deviation. 2. Mean duration of action standard deviation Comment and compare the time of onset and duration of action of each route of administered.

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

ELECTIVE PRACTICALS
In this section, you are invited to have cardiovascular parameters measured using the devices which are commonly used in patients management and in research. All you have to do is to take a few simple steps: 1. To call our Medical Lab Technologist Br Shahriman (ext 2382) to make an appointment for the measurement. 2. To answer a questionnaire on the day of measurement. 3. Following the measurement, you will receive a copy of your results.

4. To see any Pharmacology or Physiology lecturers to discuss on the results.

The devices are : 1. Stress test. 2. 24 hour Ambulatory blood pressure monitor. The stress test ECG leads are attached to your chest. Your blood pressure is measured, and the blood pressure cuff is left in place to allow regular blood pressure measurements during the test. Your heart rate, ECG pattern, and blood pressure will be monitored as you exercise (using a treadmill) at increasing levels of intensity.

The Ambulatory 24-hour blood pressure monitor. This will require you to wear a blood pressure monitoring for 24 hours (Yes! you are allowed to take the devices back to Mahallah). As the name implies, your BP is monitored throughout the 24 hour duration (even during solat and in sleep). There will be certain rules to follow while having the measurement taken.

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THE PRACTICAL BOOK FOR PHARMACOLOGY & PHYSIOLOGY LABORATORY EXPERIMENTS

SELF LEARNING Stress test 1. What is the information you can get from the stress test? ____________________________________________________ 2. What is the clinical indication for the stress test? ____________________________________________________ 3. What is the necessary precaution in performing this test, especially for patient? ______________________________________________________________ _

The Ambulatory 24-hour blood pressure monitor (ABPM). 1. Can you compare the information that we can get from using ABPM to that of using sphygmomanometer? ______________________________________________________________ ______________________________________________________________ ______________________________________________________________ _____ 2. Who will benefit from having their blood pressure measured by this way? ______________________________________________________________ ______________________________________________________________ ______________________________________________________________ _____ 3. In a drug clinical trial, what can ABPM tell us? ______________________________________________________________ ______________________________________________________________ ______________________________________________________________ _____

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