a&pii-8-renal physiology

UMSD 1213ANATOMY & PHYSIOLOGY II

BSC. (HONS) BIOMEDICAL SCIENCE2010, YEAR 2 TRIMESTER 1

Name: Quah Chin Chew, Tony (0901751)

Date: 30th March 2010

Time: 9.00 am – 11.00 am

Experiment Number:

8

Lab Report Title: Renal Physiology

Lecturer/Tutor: Dr. Michelle Ng Yeen Tan

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 1Objective:Observe and analyze the alterations in urine composition and the rapidity with which kidney function can change following consumption of a large volume of hypotonic solution, isosmotic solution or a bicarbonate solution; investigate the responses of the kidneys to various solutions.

Introduction:Comprises of two kidneys, two ureters, a bladder and a urethra, the urinary system is the principal organ responsible for water and electrolyte homeostasis. The maintenance of homeostasis required that any input into a system is balanced by an equivalent output; the urinary system provides the mechanism by which excess water and electrolytes are eliminated from the body.

A second major function of the urinary system is the excretion of many toxic metabolic waste products, particularly the nitrogenous compounds urea and creatinine; this excretory function is intimately related to water and electrolyte elimination which provides an appropriate fluid vehicle. The end product of these processes is urine. Urine, in short, is produced in the kidneys and conducted by the ureters to the bladder, where it is stored until voided via the urethra.

Since all body fluids are maintained in dynamic equilibrium with one another via the circulatory system, any adjustment in the composition of the blood is reflected in complementary changes in the other fluid compartments of the body. Thus regulation if the osmotic concentration of blood plasma ensures the osmotic regulation of all other body fluids. The process, primarily performed by the urinary system, is called osmoregulation.

The functional units of the urinary system are the nephrons, of which there are approximately one million in each human kidney. Each nephron consists of two major components, the renal corpuscle and the renal tubule.

At the head of each nephron is a tuft of capillaries known as the glomerulus, the site of plasma filtration. This capillary bed is enclosed by the funnel-shaped Bowman's capsule. After leaving Bowman's capsule, the tubular fluid, or the filtrate, passes through the proximal convoluted tubule, descending limb and ascending limb of the loop of Henle, distal convoluted tubule and collecting duct. Cortical nephrons are located almost entirely within the cortex of the kidney, while juxtamedullary nephrons have long loops of Henle that descend into the renal medulla.

Nephrons perform the functions of osmoregulation and excretion by the following processes:1. Filtration of most relatively small molecules from blood plasma to form a filtrate;2. Selective reabsorption of most of the water and other molecules from the filtrate, leaving

behind excess and waste materials to be excreted;3. Secretion of some excretory products directly from blood into the filtrate.

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 2

The kidney is also involved in two other homeostatic mechanisms which are mediated via hormones. The rennin-angiotensin-aldosterone mechanism contributes to the maintenance of blood pressure, and the hormone erythropoietin stimulates erythrocyte production in bone marrow and hence contributes to the maintenance of the oxygen-carrying capacity of blood.

In brief, the functions of kidneys include the following:1. Regulation of blood ionic composition.

The kidneys help regulate the blood levels of several ions like sodium ion (Na+), potassium ion (K+), calcium ion (Ca2+), chloride ion (Cl-) and phosphate ion (HPO4

2-).

2. Regulation of blood pH.The kidneys excrete a variable amount of hydrogen ions (H+) into the urine and conserve bicarbonate ions (HCO3

-), which are an important buffer of H+ in the blood. Both of these activities help regulate blood pH at a level of pH7.4.

3. Regulation of blood volume. The kidneys adjust blood volume by conserving or eliminating water in the urine. An increase in blood volume increases blood pressure; a decrease in blood volume decreases blood pressure.

4. Regulation of blood pressure.The kidneys also help regulate blood pressure by secreting the enzume rennin, which activated the rennin-angiotensin-aldosterone pathway. Increases rennin causes an increase in blood pressure.

5. Maintenance of blood osmolarity.By separately regulating loss of water and loss of solutes in the urine, the kidneys maintain a relatively constant blood osmolarity close to 300 milliosmoles per liter (mOsm/liter).

6. Production of hormones.The kidneys produce Calcitriol, the active form of vitamin D which helps regulate calcium homeostasis, and erythropoietin stimulates the production of red blood cells.

7. Regulation of blood glucose level.Like the liver, the kidneys can use the amino acid glutamine in gluconeogenesis, the synthesis of new glucose molecules. They can then release glucose into the blood to help maintain a normal blood glucose level.

8. Excretion of waste and foreign substances.


By forming urine, the kidneys help excrete wastes – substances that have no useful function in the body. Some waste excreted in urine result from metabolic reactions in the body. These include ammonia and urea from the deamination of amino acids; bilirubin from the catabolism of hemoglobin; creatinine from the breakdown of creatine phosphate in muscle fibers; and uric acid from the catabolism of nucleic acids. Other wastes excreted in urine are foreign substances from the diet, such as drugs and environmental toxins.

Materials and Equipments: Water 300mOsm NaCl solution (isosmotic to ECF) 3%, 350mM NaHCO3

20% potassium chromate solution 2.9% silver nitrate solution Hydrometer (Urinometer) pH meter Thermometer

Methods:1. Four subjects with normal kidney function are subjected to the following treatments:

a. Subject 1: 700ml of waterb. Subject 2: 350ml of 300mOsm NaCl solutionc. Subject 3: 350ml of 3% NaHCO3

d. Subject 4: 350ml of water (control)2. The bladder of all four subjects is emptied prior to the experiment.3. 30 minutes later, bladder of the subjects is emptied again, with the full sample collected

and recorded as sample I (pre-treatment control).4. Immediately after step 3, the treatment solutions are consumed by respective subjects.5. The sample collected is measured and analyzed via the measurement steps as below.6. Step 3 is repeated 30 minutes after the consumption of the treatments, and the full sample

collected is then recorded as sample II (post-treatment 1).7. Similarly, after another 30 minutes, step 6 is repeated and the full sample collected is

recorded as sample III (post-treatment 2).

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 4Measurement steps:

1. Volume MeasurementThe volume of each urine sample collected is measured with a graduated cylinder.

2. Urine pH DeterminationThe pH of each urine sample collected is measured with a pH meter.

3. Specific Gravity MeasurementThe specific gravity of each urine sample collected is measured with a hydrometer.

a. The temperature of each urine sample is measured prior to the measurement.b. Sufficient sample is poured into the cylinder.c. The hydrometer is placed in the urine sample, ensuring it will float, and spin

slowly.d. The specific gravity indicated on the scale is recorded when the hydrometer stops

spinning.

4. Chloride Concentrationa. 10 drops of urine is placed in a test tube, and one drop of 20% potassium

chromate solution is added to the urine sample.b. The mixture is titrated with 2.9% silver nitrate solution drop-by-drop. The number

of drops required to obtain a cloudy reddish-brown solution is recorded.

Results:Table 1: Records of various measurements for four subjects’ sample I, II and III

Subject SampleUrine Produced

(ml)pH Temp (OC) Gravity*

Silver Nitrate

1I 7.0 6.02 26 1.064 22II 12.0 6.37 27 1.064 11III 180.0 6.57 34 1.016 2

2I 54.6 6.47 32 1.016 5II 60.0 6.64 34 1.008 5III 33.0 6.80 33 1.014 4

3I 7.0 5.88 28 -** 18II 14.0 6.97 32 1.030 16III 15.0 8.39 32 1.030 13

4I 51.0 5.80 26 1.011 7II 45.0 5.85 31 1.015 7III 100.0 5.50 34 1.009 3

* Specific Gravity values stated are after considering the temperature correction factor.** Specific Gravity for sample is undeterminable due to over low urine volume.

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 5Table 2: Calculations of various variables based on records on table 1

Subject SampleUrine Production

Rate (ml/min)pH

Urinary solids / Mass (g/L)

Sodium Chloride contents (g/L)

1I - 6.02

6.32170.24

127.6822.0

11.667II 0.43.2

6.37 170.24 11.0III 6.0 6.57 42.56 2.0

2I - 6.47

6.636742.56

33.6935.0

4.667II 2.01.55

6.64 21.28 5.0III 1.1 6.80 37.24 4.0

3I - 5.88

7.08-*

79.818.0

15.667II 0.4670.4835

6.97 79.8 16.0III 0.5 8.39 79.8 13.0

4I - 5.80

5.716729.26

31.0337.0

5.667II 1.52.4165

5.85 39.9 7.0III 3.333 5.50 23.94 3.0

* Specific Gravity for sample is undeterminable due to over low urine volume.

Calculations:1. Urine production rate (ml/min)

For subject 1, the urine volume for sample II is 12.0ml and for sample III is 180.0ml.Separately, 12.0ml of urine is produced 30 minutes after consumption of treatment with another 180.0ml of urine produced after another 30 minutes, giving a total of 192.0ml urine produced over the 60 minutes period.

The urine production rate in first 30 minutes after consumption of treatment is:Urine production rate = 12.0ml / 30min

= 0.4ml/min

The urine production rate in another 30 minutes is:Urine production rate = 180.0ml / 30min

= 6.0ml/min

Thus, the urine production rate in 60 minutes after consumption of treatment is:Urine production rate = 192.0ml / 60min

= 3.2ml/minWhich is also equals to the average value of the urine production rates calculated for first 30 minutes and another 30 minutes.




= 2.0ml/min


= 1.1ml/min





= 0.467ml/min


= 0.5ml/min



For subject 4, the urine volume for sample II is 45.0ml and for sample III is 100.0ml.


Separately, 45.0ml of urine is produced 30 minutes after consumption of treatment with another 100.0ml of urine produced after another 30 minutes, giving a total of 145.0ml urine produced over the 60 minutes period.


= 1.5ml/min


= 3.333ml/min



2. Urinary Solids / Mass (g/L)A temperature correction factor is necessary when determining specific gravity of a urine sample because hydrometers are calibrated for use at 15OC and the temperatures of the urine samples are higher. Measure the temperature of each sample at the time that its specific gravity is tested. For every 3OC about 15OC, 0.001 should be added to the specific gravity reading obtained from the scale of the hydrometer.

The formula to determine the urinary solids / mass based on the specific gravity values measured is as followed:

Urinary Solids (Mass of Solute in the Urine) in the unit of g/L= (Specific Gravity – 1) X 1000 X 2.66g

For subject 1, the specific gravities are measured and calculated to be 1.064 for both sample I and II, and 1.016 for sample III.For sample I and II, the urinary solids mass = (1.064 – 1) X 1000 X 2.66g

= 170.24g/LFor sample III, the urinary solids mass = (1.016 – 1) X 1000 X 2.66g

= 42.56g/LFor subject 2, the specific gravities are measured and calculated to be 1.016 for sample I, 1.008 for sample II, and 1.014 for sample III.For sample I, the urinary solids mass = (1.016 – 1) X 1000 X 2.66g

= 42.56g/L


For sample II, the urinary solids mass = (1.008 – 1) X 1000 X 2.66g= 21.28g/L

For sample III, the urinary solids mass = (1.014 – 1) X 1000 X 2.66g= 37.24g/L

For subject 3, the specific gravities are undeterminable for sample I, measured and calculated to be 1.030 for both sample II and III.For sample II and III, the urinary solids mass = (1.030 – 1) X 1000 X 2.66g

= 79.8g/L

For subject 4, the specific gravities are measured and calculated to be 1.011 for sample I, 1.015 for sample II, and 1.009 for sample III.For sample I, the urinary solids mass = (1.011 – 1) X 1000 X 2.66g



= 23.94g/L

3. Sodium Chloride contentEach drop of silver nitrate added during the titration represents 1.0g/L. So, the sodium chloride (NaCl) content per liter (g/L) in each sample simply equals to the number of drops of silver nitrate needed to change the whole solution to a cloudy reddish brown solution.

However, in order to calculate the NaCl content (g) in each sample, a formula has to be used, and the table below shown the calculated NaCl content (g):

NaCl content (g) = NaCl per liter of urine (g/L) X 1L/1000ml X urine produced (ml)

For Subject 1, sample I’s total NaCl content in urine= 22.0g/L X 1L/1000ml X 7.0ml= 0.154g

sample II’s total NaCl content in urine= 11.0g/L X 1L/1000ml X 12.0ml= 0.132gsample III’s total NaCl content in urine= 2.0g/L X 1L/1000ml X 180.0ml= 0.36g



sample II’s total NaCl content in urine= 5.0g/L X 1L/1000ml X 60.0ml= 0.3g

sample III’s total NaCl content in urine= 4.0g/L X 1L/1000ml X 33.0ml= 0.132g







Table 3: Calculated NaCl content (g) in each urine sample

Subject Sample Urine Produced (ml)NaCl per liter of

urine (g/L)Total NaCl content

in urine (g)1 I 7.0 22.0 0.154


II 12.0 11.0 0.132III 180.0 2.0 0.360

2I 54.6 5.0 0.273II 60.0 5.0 0.3III 33.0 4.0 0.132

3I 7.0 18.0 0.126II 14.0 16.0 0.224III 15.0 13.0 0.195

4I 51.0 7.0 0.357II 45.0 7.0 0.315III 100.0 3.0 0.3

Graphs:

Subject 1 Subject 2 Subject 3 Subject 4

Urine Produced in Sample I 7 54.6 7 51

Urine Produced in Sample II 12 60 14 45

Urine Produced in Sample III 180 33 15 100

Average Urine Produced 96 46.5 14.5 72.5

10

30

50

70

90

110

130

150

170

190

Urine Produced for Subject 1 - 4

Urin

e Pr

oduc

ed (m

l)



0.5

1

1.5

2

2.5

3

3.5

3.2

1.55

0.4835

2.4165

Urine Production Rate (after treatment) for Subject 1 - 4Urine Production Rate (ml/min)


pH in sample I 6.02 6.47 5.88 5.8

pH in sample II 6.37 6.64 6.97 5.85

pH in sample III 6.57 6.8 8.39 5.5

Overall Average pH 6.32 6.6367 7.08 5.7167

Average pH after treatment 6.46 6.72 7.68 5.675

5.25

5.75

6.25

6.75

7.25

7.75

8.25

8.75

pH values for Subject 1 - 4

pH



20

40

60

80

100

120

140

127.68

33.693

79.8

31.033

Average Urinary Solids for Subject 1 - 4Average Urinary Solids (g/L)


NaCl in sample I 22 5 18 7

NaCl in sample II 11 5 16 7

NaCl in sample III 2 4 13 3

Average NaCl 11.667 4.667 15.667 5.667

2.5

7.5

12.5

17.5

22.5

Urinary NaCl Content for Subject 1 - 4

NaC

l con

tent

(g/L

)



NaCl in sample I 0.154 0.273 0.126 0.357

NaCl in sample II 0.132 0.3 0.224 0.315

NaCl in Sample III 0.36 0.132 0.195 0.3

Average NaCl 0.2153 0.235 0.18167 0.324

0.025

0.075

0.125

0.175

0.225

0.275

0.325

0.375

Total NaCl Content in Urine for Subject 1 - 4

NaC

l con

tent

(g)

Discussions:Sodium chloride, NaCI which is also known as common salt, table salt or halite is an ionic compound. Sodium chloride is the salt plays an important role for the salinity of the ocean and the extracellular fluid of many multicellular organisms. It is commonly used as a condiment and food preservative. Therefore, salt is a dietary mineral composed primarily of sodium chloride that is essential for animal life but toxic to most land plants. Salt for human consumption is produced in different forms like unrefined salt such as sea salt, refined salt or table salt and iodized salt. It is a crystalline solid, white, pale pink or light gray in colour which normally obtained from sea water or rock deposits. Edible rock salts may be slightly grayish in colour because of this mineral content. Thus, Sodium chloride, NaCI plays a role as a isosmotic solution.

Salt helps in regulating the water content or fluid balance in the body. Salt cravings may caused by trace mineral deficiencies and deficiency of sodium chloride. Conversely, overconsumption of salt increases the risk of health problems such as high blood pressure. It also was demonstrated to ease nitric oxide production. Nitric oxide, NO contributes to vessel homeostasis by inhibiting vascular smooth muscle contraction and growth, platelet aggregation, and leukocyte adhesion to the endothelium.

Sodium bicarbonate, NaHCO3 or sodium hydrogen carbonate is a white solid that is crystalline but often appears as a fine powder. It has a slight alkaline taste. Besides that, it is also a component of the mineral natron and is found dissolved in many mineral springs. The natural

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 14mineral form is known as nahcolite. It is also produced artificially. It is used as baking soda, bread soda, cooking soda and bicarbonate of soda. Sodium bicarbonate, NaHCO3 acts as a bicarbonate solution.

Mostly, sodium bicarbonate is used in an aqueous solution as an antacid taken orally to treat acid indigestion, heartburn and governed intravenously for cases of acidosis or when there are insufficient sodium or bicarbonate ions in the blood. Moreover, it is also used to treat hyperkalemia, aspirin overdoses and tricyclic antidepressant overdose. Sometimes it also applied as a paste, with three parts baking soda to one part water, to relieve insect bites. Sodium bicarbonate is also used as an ingredient in some mouthwashes which cleans teeth and gums, neutralizes the production of acid in the mouth and also as an antiseptic to help prevent infections occurring.

However, the adverse reactions of sodium bicarbonate can include metabolic alkalosis, edema due to sodium overload, congestive heart failure, hyperosmolar syndrome, hypervolemic hypernatremia, and hypertension due to increased sodium. The use of sodium bicarbonate can cause milk-alkali syndrome which can result in metastatic calcification, kidney stones and kidney failure.

The regulations of salt and water balance by the kidneys often involve several negative feedback mechanisms, as shown below:

Atrial natriuretic hormone (ANH) which is also referred to as atriopeptin, secreted by the atria in response to increased atrial stretch, causes increased glomerular filtration rate and Na+ and water loss.

Antidiuretic hormone (ADH) which is also referred to as vasopressin, secreted by the posterior pituitary in response to a decrease in plasma volume or an increase in plasma osmolarity, causes increased water reabsorption across the walls of the collecting duct of the kidney, thereby decreasing urinary water loss and resulting in the production of hyperosmotic urine. Caffeine acts by inhibiting the secretion of antidiuretic hormone.

Aldosterone is secreted by the adrenal cortex in response to a regulatory cascade triggered by decreases in plasma Na+ concentration or plasma volume, causing increased recovery of Na+ in the distal tubule.

Under normal conditions, almost all of the bicarbonate ions (HCO3–) in the filtrate are

reabsorbed, 80-90% in proximal tubule, others in ascending loop of Henle and collecting duct. However, when a large amount of base is ingested, the plasma pH increases, which will further trigger coordinated responses in both the respiratory and urinary systems. The immediate response is a decrease in respiration, known as respiratory compensation, which leads to an increase in plasma PCO2. Renal compensation, in which part of the filtered load of HCO3

– in the filtrate is not absorbed and some HCO3

– spills into the urine, occurs more slowly.

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 15Based on results obtained, it may be observed that both treatment 1 and treatment 4 who drank water have higher urine production rate of 3.2ml/min and 2.4165ml/min respectively. The urine production rate for treatment 2 who drank 350ml of 300mOsm sodium chloride solution is 1.55ml/min while for treatment 3 who drank 350ml of 3%NaHCO3 is only 0.4835ml/min. Thus, it is observed that both isosmotic solution and basic solution do show some influences in decreasing the urine production rate, comparing to the control of 2.4165ml/min.

As for pH values, it is observed that most of the treatments – treatment 1, treatment 2 and treatment 4 – do not show much effects in the pH values. An average progressive increase of approximately 0.15-0.35 is observed for both treatment 1 and treatment 2, while the pH values of treatment 4 is considered quite stable and do not show any drastic fluctuation. As for pH values of treatment 3, a steady increase is observed. The pH value rose from 5.88 to 6.97 and 8.39 after 30 minutes and 60 minutes of treatment, changing its acidity from acidic to nearly neutral and to basic, respectively. It is thus observed that basic solution of NaHCO3 would cause an increase in the subject’s body fluid pH value.

Comparing to control treatment 4, all three treatments do show an extent of increase in the average urinary solids / mass values. The highest increase is observed in treatment 1, at the level of 127.68g/L, followed by 79.8g/L by treatment 3 and 33.693g/L of treatment 2; comparing to control treatment 4’s 31.033g/L. There might be some human errors while handling urine samples and measuring the samples – as theoretically, only treatment 2 and treatment 3 of different solutes will show increments in their average urinary solids, while treatment 1 should show decrement as higher urine volume is produced. Besides, due to insufficient amount of urine collected for treatment 3’s sample I, the results for this session may not be too accurate.

For the urinary sodium chloride content, treatment 2’s result of 4.667g/L is close to the control treatment 4’s result of 5.667g/L. Treatment 1 shows a doubling of the control, to the level of 11.667g/L while treatment 3 shows a tripling of the control, to the extent of 15.667g/L. For the total NaCl content in the urine samples, the control treatment 4 shows 0.324g in the average urine produced of 65.33ml. All other treatments show lesser total NaCl content in the urine samples. Treatment 1 shows only an average NaCl content of 0.2153g in the average urine amount of 65.33ml; treatment 2 shows an average NaCl content of 0.235g in the average urine amount of 49.2ml; while treatment 3 shows only an average NaCl content of 0.18167g in the average urine amount of 8.67ml.

“Link Theory to Practice~” ~Mr. Paul Davidson P a g e | 16Conclusion:Based on the results obtained, calculations made and graphs plotted, it can be concluded that:

Treatment 1 – 700ml Water – produce the highest urine amount of 12ml in 30 minutes and 180ml in another 30 minutes, giving a total of 192ml in the 60-minute duration, with the urine production rate of 3.2ml/min and averagely 96ml of urine produced every 30 minutes after treatment; while Treatment 3 – 350ml 3% NaHCO3 – produce the lowest urine amount of only 14ml in 30 minutes and 15ml in another 30 minutes, giving a total of only 29ml in the 60-minute duration, with the urine production rate of 0.4835ml/min and averagely 14.5ml of urine produced every 30 minutes after treatment.

The highest average pH value of 7.08 is shown by Treatment 3 as well. A progressive increase is noticed for the pH values increased from 5.88 to 6.97 and to 8.39 in sample I, sample II and sample III, respectively; while the lowest average pH value of 5.7167 is shown by the control Treatment 4 – 350ml Water. The pH values are considered stable 30 minutes after treatment (from 5.80 increased to 5.85), and shows a decrease to 5.50 after another 30 minutes.

Treatment 1 is showing the highest average urinary solids of 127.68g/L; while the lowest average urinary solids of 31.033 is shown by Treatment 4.

The highest average amount of NaCl excreted in urine is shown by Treatment 4, at 0.324g; while the lowest average amount is shown by Treatment 3, at 0.18167g.

As so, the urine conditions (volumes, pH, urinary solids and NaCl contents) are affected by the intake of various fluids and solutions.

References:1. Boron, W. F. & Boulpaep, E. L. 2007, Medical Physiology: A Cellular and Molecular

Approach (2nd Ed.), W. B. Saunders Company.2. Martin, T. R., Shier, D., Buttler, J. & Lewis, R. 2004, Hole’s Human Anatomy &

Physiology Laboratory Manual, McGraw-Hill.3. Tortora, G. J. & Derrickson, B. H. 2009, Principles of Anatomy and Physiology (12th

Ed.). Volume 2: Maintenance and Continuity of The Human Body, John Wiley & Sons.4. Waught, A. & Grant A. 2004, Ross and Wilson Anatomy & Physiology, Churchill

Livingstone.5. Wheather, P. R., Burkitt, H. G. & Daniels, V. G. 1988, Functional Histology: A Text and

Colour Atlas (2nd Ed.), Churchhill Livingstone.

a&pii-8-renal physiology

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