FAR 242/4ENDOCRINE SYSTEM AND METABOLISM

EXPERIMENT 1

THE EFFECTS OF HORMONES ON METABOLIC RATE IN RATS

NAME: KESHAMALINI A/P GOPALSAMYMATRIX NO: 119072LECTURER: ASSOC. PROF. MARIAM AHMADDATE : 17/03/2015

Introduction:The aim of this experiment is to determine the effects of the hormone thyroxine, which is released by the thyroid gland and also the antithyroid agent, thiouracil on the rate of O2 consumption in rats. Numerous studies have proven that thyroid hormones increases O2 consumption in test subjects as well as on isolated tissue preparations in vitro. Therefore, this observation corresponds to the theory that the function of the thyroid hormone is to increase the rates of activity of most of the chemical reactions that takes place in the cells of the body. In some cases, the thyroid gland produces excessive thyroid hormones for various reasons, e.g. Graves disease, toxic multinodular goiter, Plummers disease, drug induced and so forth. Therefore, an antithyroid agent is required to bring back the thyroxine levels to normal. Equipment: Jacketed oxygen consumption chamber U-tubeMaterials: Thyroxine Thiouracil ( 0.05%)Experimental procedure:Please refer to page 25-26 of FAR 242/4 Physiology Manual.

Results:Readings Control (cm)Thyroxine (cm)Thiouracil (cm)

Reading 12.504.001.00

Reading 22.303.00 2.50

Reading 32.40 3.80 1.50

Average2.403.60 1.67

Weight of rats (kg)0.18870.17130.2027

Table 1: Group 2 results showing the rise in water levels in manometer after 2 minutes and the weights of each rat.

GroupsAmount of O2 Consumption (ml/kg/hr)

ControlThyroxineThiouracil

11283.832099.13978.84

2763.121260.95494.33

31336.212197.051395.87

41389.081647.80908.43

51165.821703.351122.45

61352.871569.20785.48

71550.661552.121245.31

81224.002402.41116.90

Average1258.201804.001005.95

Table 2: Class results showing the average amount of O2 consumption.

Calculations:

When 20ml of air is injected into the jacket oxygen chamber, the rise in water level is 10cm.20 ml of air= 10.0 cm in water level2.0 ml of air= 1.0 cm of water levelAverage amount of O2 consumption:Volume of oxygen consumed (ml)= 2ml/1cm water level in the manometer after 2 minutes (cm)

Volume of O2 consumption (cm)

Control2/1 x 2.4 = 4.8

Thyroxine2/1 x 3.6 = 7.2

Thiouracil2/1 x 1.67 = 3.34

Table 3: Volume of O2 consumption by the rats.

Therefore, the amount of O2 consumption (ml/kg/hr) can be calculated from the formula:Volume of oxygen consumed (ml) x

Amount of O2 consumption (ml/kg/hr)

Control x = 763.12

Thyroxine x = 1260.95

Thiouracil x = 494.33

Table 4: Amount of O2 consumption (ml/kg/hr)

Discussion:The objective of this experiment is to investigate the effects of the thyroxine hormone and antithyroid drug (thiouracil) on the metabolic rate of rats in vivo. In this experiment, 3 rats were used, one as control and the other two as test subjects for thyroxine and thiouracil respectively. Thyroxine is a hormone produced by the thyroid gland as part of the endocrine system and it serves to regulate growth, energy levels, metabolism and mood stability. On the other hand, thiouracil and allied drugs are used in treating hyperthyroidism by lowering the amount of thyroid hormone produced by the thyroid gland (Bartels, 1948). By referring to Table 1, the amount of O2 consumption by the control rat was 763.12 ml/kg/hr, whereas it was 1260.95 ml/kg/hr and 494.33 ml/kg/hr respectively for the rats that were fed with thyroxine and thiouracil. Theoretically, the rat under the influence of thyroxine will have a higher consumption of O2 than the control rat, and the rat under the influence of thiouracil should have a lower consumption of O2 than the control rat. Based on the results obtained by my group (Group 2), it shows that our experimental results follow the theory proposed. However, when compared with the results from other groups, it seems there are vast deviations numerically. The amount of O2 consumption by my group (group 2) was the least when compared to other groups whom all obtained a value of more than 1000 ml/kg/hr. There are a few explanations for such differences in the results obtained by each group. This could be due to errors occurring during the experiment. For example, the state of health of the rats was not determined prior to the experiment, leakages at the junction of the U-tube are possible if the students did not fix them tightly. Inconsistent quantity of oxygen supplied into the jacketed oxygen consumption chamber may have resulted in such deviations of results between the groups. Nevertheless, 7 groups (excluding group3) managed to obtain results that corresponds to the theory taught in class; O2 consumption by the rat fed with thyroxine must be higher than the control rat and O2 consumption by the rat fed with thiouracil must be lower than the control rat. Therefore, the average amount of O2 consumption by the control rat was 1258.20 ml/kg/hr, whereas it was1804.00 ml/kg/hr and 494 ml/kg/hr respectively for the rats that were fed with thyroxine and thiouracil.Thyroxine (often abbreviated as tetraiodothyronine; T4 because it contains 4 iodines) is a lipid-soluble hormone that easily passes through the cell membrane. It is brought to target cells, either through passive diffusion or with the aid of protein carriers (1). Once T4 has entered the cytoplasm of the target cell, most of the T4 is converted into triiodothyronine; T3 (2). The remaining T4 and T3 enter the nucleus and bind to nuclear receptor proteins (3). The hormone-receptor complexes will then bind to specific hormone response elements (HRE) on DNA, eliciting hormonal responses (4). This may result in either stimulation, or in certain cases inhibition of gene transcription (5). The messenger RNA (mRNA) produced then codes for specific proteins (6). Thus, these proteins are responsible for mediating the thyroid hormone response (Tortora, 2011). Diseases of the thyroid gland occur when the levels of thyroid hormone secretion are abnormal. If the secretion of thyroid hormone is below the normal range, it is termed as hypothyroidism and if it is beyond the normal range, then it is termed hyperthyroidism. Therefore, an antithyroid agent has to be used in such cases to bring the levels of thyroxine back to normal. In the 1940s, hundreds of rats were tested with thiourea-related compounds and it was concluded that theory and theoretical were the most favorable compounds that possessed antithyroid activity (Astwood, 1943). The proposed mechanism of action of thiouracil is illustrated in Diagram 1. The thiourylene drugs are potent inhibitors of thyroperoxidase (TPO) enzyme. These drugs bind irreversibly to TPO, therefore inhibiting its ability to reduce iodine, I2 to I- and convalently attaching it to the tyrosine residue of thyroglobulin. Besides suppressing the coupling of iodothyronines to form T3 and T4, conversion of T4 to T3 are inhibited as well. MECHANISM OF ACTION OF THYROXINE

(1) (2)

(3) (4)

(5) (6)

McGrawHill Education. http://highered.mheducation.com/

Thyroxine and antithyroid drugs. Parakramawansha, Ruwan. 2013. http://www.pdn.ac.lk/.Diagram 1: The proposed mechanism of action of thiourylene/thioamide family.

Questions:1) Define basal metabolic rate (BMR). What is the effect of thyroxine on BMR?Basal metabolic rate is the rate of oxygen consumption under standard or basal conditions in order to maintain vital functions such as breathing and keeping warm (Tortora, 2011). BMR therefore refers to the minimum number of calories an individual needs to survive in the absence of any activity. Thyroxine controls the BMR and an increase in the thyroid hormone will increase the BMR by increasing the production of heat and by stimulating the use of cellular oxygen to produce ATP. Therefore, increase in basal metabolic rate results in an increase in metabolism of carbohydrate, lipids and proteins.

2) Discuss all the factors that affect BMR. Body size: Metabolic rate increases as weight, height and surface area increases. Body composition: Muscle tissue has a higher metabolic activity than fat tissue. Gender: Women generally have a lower BMR than men because women generally possess more body fat and less muscle mass. Hormone levels: Thyroxine, the key hormone released by the thyroid glands has a significant effect upon metabolic rate. Health: Fever, illness, or injury may increase resting metabolic rate twofold. Climate and body temperature: The BMR of people in tropical climates is generally 5 to 20 percent higher than their counterparts living in more temperate areas because it takes energy to keep the body cool. Exercise performed in hot weather also imposes an additional metabolic load. Fasting/Starvation - Fasting or starving will lower the BMR as calorie intake declines. Age: Metabolic rate declines with age. Caffeine and tobacco use: Increases metabolic activity. Pregnancy or breast feeding mothers: Increase metabolic rate

Conclusion:1) Thyroxine increases the O2 consumption and the basal metabolic rate.2) Thiouracil decreases the O2 consumption and the basal metabolic rate. Precautions:1) Make sure the system is airtight all the time (check all the junctions to ensure no air escapes).2) During the calibration process and during the experiment, make sure the rise in water level is measured accurately; measurement should be done by one person to minimize parallax error. 3) Rest the rat for 5 minutes before every repetition of the experiment and flush in oxygen for 30 seconds each time to obtain more accurate results. 4) Handle the manometer carefully as it is very fragile and breaks easily. 5) Label the jar correctly to indicate which jar contains the control rat and which jar contains the rat that has been fed with thyroxine and thiouracil. 6) Return the rats to the right cage after the experiment.

References:ASTWOOD, E. B. 1943. THE CHEMICAL NATURE OF COMPOUNDS WHICH INHIBIT THE FUNCTION OF THE THYROID GLAND. Journal of Pharmacology and Experimental Therapeutics, 78, 79-89.BARTELS, E. C. 1948. Thiouracil and Allied Drugs in Hyperthyroidism. New England Journal of Medicine, 238, 6-11.TORTORA, G. J.