Human Kidney Functions And Osmotic Regulation Experimental Study

Human Kidney Functions and Anatomy

The experiment aimed at:

• Determining the osmotic regulation of the kidneys when it comes to maintaining a balance of fluids in the body
• Studying the physical characteristics of urine and establishing if a urine sample is normal

The human kidneys have a significant role to play in the human body as they aid in the regulation of the volume of plasma, the osmolality, pH, ionic composition and even elimination of waste products of metabolic processes. All these regulations work in harmony to ensure that homeostasis is maintained in the body¹. The main determinants of the composition of blood include cellular metabolism, urinary output as well as the diet. To the tune of 180 liters of blood plasma is filtered by the nephrons of the kidney in 24 hours through the glomeruli into the tubules in which it undergoes selective processing by the tubular reabsorption and secretion.

The human body has up to 5 liters of blood circulating at any given time out of which twenty percent which represents one liter of the blood circulates to the kidneys. The roles of the kidney are the billions of the nephrons that they are composed of which serve to filter blood. Each of the neurons of the kidney has renal corpuscle and renal tubules. In the renal corpuscles are the Bowman’s capsule and glomerulus while in the renal tubules contains the loop of Henle, proximal tubule, the distal convoluted tubules as well as the connecting tubules which connect the collecting duct².

There are two parts of the proximal tubule: proximal convoluted tubule and the proximal straight tubule. The loop of Henle is subdivided into three main parts: a thin ascending limb, the descending limb, and the thick ascending limb. The entry of blood is first through the afferent arteriole then into the glomerulus and finally, it’s filtered out to the Bowman’s capsule. The filtrate is referred to as the glomerular filtrate and is plasma that does not contain proteins.

Reabsorption and elimination of solutes, ions, and water take place in the kidneys. Whereas reabsorption defines the movement of water, ions, and nutrients back to the blood through the peritubular capillaries from the filtrate that is found in the tubules, a greater portion of it takes place in the proximal convoluted tubule and some fraction in the distal convoluted tubule.

Three main hormones aid in the proper functioning of the kidneys: Anti-Diuretic Hormone, Atrial Natriuretic Peptide, and Aldosterone³. Anti-Diuretic Hormone is termed as a vasoconstrictor which raises the blood sugar levels and increases the reabsorption of water, urine specific gravity, plasma volume as well as decreasing the flow rate of urine.

Regulation of Blood Composition

Stimulation of Anti-Diuretic Hormone targets the collecting duct and the proximal convoluted tubule resulting into reabsorption of water. Aldosterone is generated in the adrenal cortex where it stimulates Renin-Angiotensin-Aldosterone System upon a decrease in the volume, plasma osmolality and pressure occurrence thereby resulting into a enhancement n the reabsorption of sodium through the distal tubule⁴.

The role of this experimental study was to determine the kidneys’ osmotic regulation when it comes to maintaining a balance of fluids in the body as well as studying the various physical characteristics of urine. Two subjects were chosen for the first study in which one consumed fresh water and the other saline or salty water. The second part of the experiment involved comparing the properties of urine from three different subjects against the normal characteristics of urine. 

1. Every student was required to collect a morning sample of urine collected as soon as they got out of bed and labeled it t=morning. Students selected as subject 1 or 2 not to pass urine just before the experiment or take any caffeinated beverages.

Subject 1 would be undertaking the water test

Subject 2 would be undertaking the saline test.  

1. Immediately upon arrival to the practical, Subject 1and Subject 2 were required to completely empty their bladder into the sterile collection container (graduated cylinder) as soon as they arrive.  
2. Record these measurements in the table below.
3. Subject 1would drink 750 to 1000 ml of water quickly (within 5-10 minutes). Then wait 15 minutes and collect a urine sample. Collect a further 2 urine samples at 15 minute intervals over the next 45 minutes (i.e. t=15, t=30 and t=45). Ensure that the ‘recorder’ and subject are using the timer to keep an accurate measure of the timing⁵.
4. Subject 2would drink 750 to 1000 ml of 0.9% (isotonic) saline quickly (within 5-10 minutes). Similar to the Subject 1, collect a urine sample at three 15 minute intervals over the next 45 minutes (i.e. t=15, t=30 and t=45).
5. Wash down the area of the lab bench you are working on with a 70% ethanol solution.

One strip was removed from bottle and cap replaced. Completely immerse reagent areas of the strip in the sample and remove immediately.

1. In the process of elimination, the edge of the whole length of strip is run against the container rim to eliminate the excess sample. The strip is held in a horizontal position so as to avoid chances of mixing up of the chemicals from the nearby reagent areas and/or making the hands dirty with the sample.
2. Compare reagent areas to the corresponding color chart you were given at the times specified. Hold strip close to color blocks and match carefully. Avoid laying the stick directly on the color chart.
3. Taking proper readings of the time is very important for results of higher accuracy. The glucose and bilirubin test was read at 30 seconds upon dipping. The ketone test was read at 40 seconds; while that of the specific gravity done at 45 seconds; at 60 seconds the tests for Urobilinogen, pH, nitrite, protein blood were conducted; and finally at 2 minutes the test for leucocytes carried out⁶. The readings of the protein and pH areas may also be conducted immediately or any time within 2 minutes after dipping. The pH area of the sample is checked immediately upon dipping the strip. Should the colour on the pad be found to be non-uniform, the readings on the reagent area are taken immediately and a comparison is made of the darkest colour against the suitable colour chart. The readings of all the areas of the reagent may be taken excluding that of the leucocytes for the purposes of identification of the negative specimens and estimation of the specific gravity as well as the. A positive reaction on the leucocyte test even to the least degree of significance at less than 2 minutes may be treated as a likely indication of the presence of leucocytes in urine. Changes in colour take place after 2 minutes when there is no diagnostic value¹². 

                                                     

                                                                          Figure 1: Graph for water flow rate versus time

As can be observed on the graph, the rate of flow of water decreased with an increase in the time interval of collection of the water illustrating that the amount of the liquid that was leaving the body kept decreasing with an increase in time.

                                                       

                                                                    Figure 2: Graph for Water osmolality versus time

The graph indicates indirect proportionality such that as the time increased, the water osmolality decreases to a level that it becomes fairly constant.

                                                       

Hormones Involved in Kidney Functioning

                                                                     Figure 3: Graph for Saline flow rate versus time

As can be observed on the graph, the saline flow rate decreased with an increase in the time interval of collection of the water illustrating that the amount of saline that was leaving the body kept decreasing with an increase in time.

                                                   

                                                                  Figure 4: Graph Saline osmolality versus time

The graph indicates indirect proportionality such that as the time increased, the saline osmolality decreases to a level that it becomes fairly constant.

Physical Characteristics Colour	Pale yellow	Yellow: pale   medium   dark Other: ____________	Yellow: pale   medium   dark Other: ___very pale / cloudy _________	Yellow: pale medium dark Other ____________	Yellow: pale medium dark Other ____________
Transparency	Clear	Clear   slightly cloudy   cloudy	Clear   slightly cloudy   cloudy	Clear   slightly cloudy   cloudy	Clear   slightly cloudy   cloudy
Odour	Characteristic	Describe:	Describe:	Describe:	Describe:

Table 1: Physical characteristics of normal urine compared against the unknown samples.

The table illustrates the differences between the control sample urine and the unknown’s samples of urine with regard to the color, odour, and transparency. A lot of deviation is noticed in the transparency in which the normal urine is clear while the unknown samples are cloudy.

Glucose	negative	negative		negative		500-20	negative
Bilirubin	negative	negative		negative		negative	negative
Ketone	negative	negative		negative		negative	negative
Specific Gravity	1.001 – 1.035	1.015		1.02		1.015	1.02
Blood	negative	negative		negative		negative	80
pH	5 to 9 (abnormal and normal range)	6		6		6	6
Protein	negative	negative		100		negative	negative
Urobilinogen	0.2-1 U/dL 3.2–16 µmol/L	0.2/3.2		0.2/3.2		0.2/3.2	0.2/3.2
Nitrite	negative	negative		negative		negative	negative
Leucocytes	negative	negative		negative		negative	negative

Table 2: Various substances available in the control of urine samples and the unknown samples

The table shows the different substances found in the control urine sample and the unknown sample including protein, pH, nitrite, leucocytes, and blood among others. The table depicts a close similarity between the control sample and the unknown samples for most of the substances.

• The consumption of water had minor impacts on the ECF volume and ECF osmolality as any free water that was added to the ECF quickly somatically equilibrates with the available intercellular space which was a fluid compartment that is greater than the extracellular compartment⁷
• The resulting change in the ECF osmolarity illustrated more water available in the body. The hormone ADH was stimulated and more of it produced which targeted the collecting duct and distal tubule resulting into reabsorption of water.
• ADH enhanced the reabsorption back of water into the circulating by binding to the receptor cells in the collecting duct
• Treatment with water increased the flow of urine since the more excess supply of water in the body some of which had to be eliminated to maintain the osmotic balance of the body as well as the homeostasis
• A low ECF osmolality would mean less sodium concentration in the blood that would stimulate the production of aldosterone to facilitate the uptake of sodium ions⁸
• Aldosterone increased the uptake or absorption of sodium ions in the distal convoluted tubule

• The treatment with saline increased ECF volume and ECF osmolality as the saline treatment increased the isoosmotic of the solution.
• The amount of ANP produced increased with the resulting change in the ECF volume to counter or inhibit the reabsorption of sodium ions and enhance excretion of sodium.
• Treatment with saline reduced the flow of urine and urine osmolarity as it led to more reabsorption of water to achieve an osmotic equilibrium of body⁹.
• ANP served to inhibit the reabsorption of sodium ions in the collecting ducts and the distal tubule and foster excretion of sodium

• The osmolarity of the morning sample was higher than the other sample collected from subject 1 since before drinking water there was an osmotic balance of the solutes and solvent
• The morning urine sample had equal concentration of ADH and Aldosterone due to an isotonic concentration that was attained by the body throughout the night when there were no intakes of water of further reabsorption of osmolytes¹⁰

The urines of the unknowns were cloudy while those of the controls were clear. The cloudy nature could be attributed to conditions that result in too many crystalline substances or protein in the urine. Such conditions could be infections in any part of the urinary tract among them urethra or bladder¹¹.

Conclusion

The experiment achieved the aims which were to determine the osmotic regulation of the kidneys when it comes to maintaining a balance of fluids in the body as well as studying the various physical characteristics of urine. Various treatments lead to various changes in the ECF volume and ECF osmolality in which an increase in osmolytes results in an increase in ECF volume and ECF osmolarity. The cloudy color of urine is not normal and could be due to infections in the urinary tract

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