Urine Concentration and Dilution

1. Urine Concentration and Dilution

2. Learning objectives. 1. Identify the major routes of water loss from the body. 2. Identify the two most potent stimuli for ADH release and the negative feedback mechanism for each. 3. Describe the role of the ascending limb of the loop of Henle in producing high medullary interstitial fluid osmolality and dilute distal tubular fluid. 4. Describe the mechanisms of countercurrent multiplication and exchange. 5. Explain the importance of urea in formation of concentrated urine. 6. Explain the mechanisms by which ADH causes formation of concentrated urine. 7. Distinguish between central and nephrogenic Diabetes Insipidus based on the plasma ADH level and the response to injected ADH. 8. Define free water clearance. Given the urine and plasma osmolalities and urine flow rate, calculate osmolar and free water clearance. Identify expected free water clearance for an individual producing either a dilute or a concentrated urine.

3. Typical Daily Water Balance WATER BALANCE (loss = “intake”) Plasma osmolarity = 285 mOsm/L

4. Osmoreception Is Coupled To Anti-diuretic Hormone (ADH)(alias Arginine Vasopressin ; AVP)Secretion Thirst 12 ADH - V1 receptor (vasoconstriction) 8 Plasma ADH (pg/ml) 4 ADH - V2 receptor (water retention) 0 270 280 290 300 310 Plasma osmolality (mOsM)

5. Hypothalamus Via baroreceptors Via osmoreceptors ECF osmolarity ECF volume MOST IMPORTANT Posterior Pituitary ADH

6. Osmolarity gradient from the cortex down to the medulla Descending LH water permeable Ascending LH : impermeable to water + passive & active Na+ transport Collecting Duct : depends on [ADH]

7. Making Dilute Urine - “Free Water Excretion” Low ADH 1200 The medullary interstitium is hypertonic The ascending limb of Henle’s loop is the diluting segment Isosmotic reabsorption mOsmM 600 0 PT DL AL DCT CCD MCD

8. High ADH 1200 The cortico-medullary osmolality gradient is much bigger mOsmM 600 Osmotic equilibration is occurring in the collecting duct 0 MCD PT DL AL DCT CCD The loop of Henle deposits more salt than water in the intersitium, so: • Medullary interstitium becomes hypertonic – potential for urine concentration! • Tubular fluid becomes hypotonic – potential for urine dilution!

9. Summary of Ascending Limb of the Loop of Henle Activity as a result of solutes being removed from the filtrate and water being retained, there are two very important effects: The filtrate becomes increasingly dilute as it moves up the tubule. 2) The osmolarity of the interstitium is 200 milliosmoles greater than the osmolarity of the filtrate at any given level of the tubule. • Notice particularly that this has also formed an osmolarity gradient from the bottom of the interstitium to the top. In this medullary osmotic gradient, the osmolarity of the deeper region is greater than that of the region close to the cortex.

10. The Loop of Henle: A Countercurrent Multiplier • theory of how the loop of Henle forms the medullary osmotic gradient • called the countercurrent theory because of the opposing flow of filtrate within the two limbs of the loop. • • The complex interplay of the ascending and descending limbs forms and maintains an interstitial osmolarity with a gradient from approximately 1200 milliosmoles near the bottom of the loop to the normal 300 milliosmoles near the cortex. The gradient formed by this activity is essential for the concentration of urine. • The ascending limb of the loop actively transports sodium chloride into the interstitium, increasing its concentration, while restricting the diffusion of water. • The interstitium surrounding the tubule becomes more concentrated, while the fluid inside becomes more dilute. • As the fluid moves up the loop there is progressively less solute to pump out, so the highest concentrations of solutes are near the bottom of the loop. Ascending Limb.

11. Descending Limb. • The role of the descending limb in this process is to provide a continuous, concentrated supply of sodium chloride to the ascending limb. • The filtrate concentrates as it moves down the descending limb, because water diffuses into the higher osmolarityinterstitium created by the ascending limb. • The solutes remaining in the tube reach a concentration approximately four times greater than normal body fluid. (like salt lakes) • This salty filtrate provides a ready supply of sodium chloride for the ascending limb to maintain the osmotic gradient. Combined Activities. • The way the two limbs cooperate to multiply the solute concentration from the normal 300 milliosmoles of the cortex to a level four times higher in the medulla is called the countercurrentmultiplier mechanism. • This mechanism has also diluted the luminal fluid in the ascending limb to an osmolarity of approximately 100 milliosmoles; as a result, water is retained, while solutes are removed.

12. Urea is necessary for increasing osmolarity of Inner medulla

13. How Does The Counter-current Multiplier Work? mCD mTALH NaCl ADH H2O NaCl UREA UREA [ NaCl] 1200mOsm

14. The Vasa Recta: A Countercurrent Exchanger • How are the solutes in the medullaryinterstitium kept from being carried away by the blood? • capillaries of the vasa recta form ‘hair pin’ loops enabling them to function as countercurrent exchangers. • Blood in the vasa recta loops delivers nutrients such as glucose and oxygen to the cells of the region. • At the same time, the loops function as countercurrent exchangers of sodium chloride and water. Blood moving down the descending portion of the vasa recta loop passes through areas of increasing osmolarity. Water diffuses out in exchange for Na Cl . The viscous blood moves up the ascending portion of the vasa recta through areas of decreasing osmolarity. The blood regains the water by osmosis and loses most of the solutes back into the interstitium. • The net result is that the blood has brought nutrients to the cells of the medulla without carrying away extensive amounts of solute, which would weaken the osmotic gradient.

15. How Does ADH Control Water Reabsorption In The Collecting Duct? Lumen Blood Principal cell Aquaporin 2 - containing vesicles AQP3 Low ADH H2O High ADH AQP2 V2 H2O AQP3 AQP2

16. Water Conservation Fails In DIABETES INSIPIDUS (DI) • Nephrogenic DI: • V2- receptor mutations; AQP2 mutations • acquired e.g. Lithium therapy • Central DI: • congenital lack of ADH production • acquired e.g. Head trauma • Psychogenic polydipsia

17. FREE-WATER CLEARANCE Free water is defined as distilled water that is free of solutes (or solute-free water). In the nephron, free water is generated in the diluting segments, where solute is reabsorbed without water. The diluting segments of the nephron are the water-impermeable segments: the thick ascending limb and the early distal tubule. Measurement of free-water clearance (CH2O) provides a method for assessing the ability of the kidneys to dilute or concentrate the urine. The principles underlying this measurement are as follows: When ADH levels are low, all of the free water generated in the thick ascending limb and early distal tubule is excreted (since it cannot be reabsorbed by the collecting ducts). The urine is hyposmotic, and free-water clearance is positive. When ADH levels are high, all of the free water generated in the thick ascending limb and the early distal tubule is reabsorbed by the late distal tubule and collecting duct. The urine is hyperosmotic, and free-water clearance is negative.

18. Measurement of CH2O calculated by the following equation: CH2O can be zero, it can be positive, or it can be negative. CH2O is Zero– Isosthenuria where CH2O Free-water clearance (mL/min) V Urine flow rate (mL/min) Cosm Clearance of osmoles (mL/min) [U]osm Urine osmolarity (mOsm/L) [P]osm Plasma osmolarity (mOsm/L) CH2O is positive- Diabetes Insipidus. CH2O is negative- SIADH

19. Summary • 65% of the filtrate is reabsorbed in the proximal convoluted tubule through active and passive transport processes. • The filtrate is concentrated in the descending loop of Henle; water is lost while solutes are retained. • The filtrate is diluted in the ascending loop of Henle; solutes are lost while water is retained. • The asymmetrical pattern of water and sodium chloride reabsorption in the ascending and descending loop of Henle creates an osmotic gradient within the medullary region. • The vasa recta circulate blood through the medulla to provide nutrients without removing solutes and weakening the osmotic gradient.

21. Late Filtrate Processing- DCT & COLLECTING DUCTS.

22. • The final processing of filtrate in the late distal convoluted tubule and collecting ducts comes under direct physiological control. • precisely regulating the final balance of fluid and solutes returned to the blood. • • In this region, membrane permeabilities and cellular activities are altered in response to the body's need to retain or excrete specific substances.

23. Filtrate Processing in the CCD: Hydrogen Ion Secretion • • The epithelium of the collecting ducts consists of two cell types. Each of these cells plays a different role in the final processing of filtrate. • 1. Intercalated cells • The intercalated cells help to balance the blood pH by secreting hydrogen ions into the filtrate through ATPase pumps in the luminal membrane. • 2. Principal cells • The principal cells perform hormonally regulated water and sodium reabsorption and potassium secretion. • • The permeability of the principal cells to sodium ions and water is controlled by two hormones: aldosterone from the adrenal gland and antidiuretic hormone, or ADH, from the posterior pituitary gland.

24. Filtrate Processing in the CCD: Role of Aldosterone a decrease in the level of sodium ions or an increase in potassium ions in the blood will trigger the release of aldosterone. Role of Antidiuretic Hormone When stimulated by ADH, principal cells quickly insert luminal water channels, increasing their water permeability.

25. Response to Dehydration and Overhydration Dehydration: • In dehydration, which could be caused by hot weather, perspiration causes the body to lose both water and sodium. • In response ADH is released, it stimulates the kidney to conserve body fluid by increasing reabsorption of water from the filtrate. • Therefore, the volume of filtrate entering the medullary collecting duct is reduced, so urine volume decreases. Overhydration: Overhydration, which could be caused by drinking several cans of soda or other beverages, triggers a decrease in ADH. • As a result, membrane permeability for water and sodium ions decreases, reabsorption slows dramatically, and the volume of filtrate entering the medullary collecting duct increases above the normal level, causing urine volume to increase.

26. Progressive Change in Filtrate Osmolarity 300 milliosmole 3001000mlosmole 1000100mlosmole 100 milliosmole 300 milliosmole 3001200mlosmole 1200300mlosmole 300 milliosmole highly permeable Yes No Yes No Yes Yes highly permeable 65% reduction 15% reduction unchanged. 15% reduction

27. Urine Concentration: Medullary Collecting Duct • The last step in the formation of urine occurs as the filtrate passes down the medullary collecting duct. • Antidiuretic hormone regulates the final amount of water reabsorbed in the collecting duct, and thus determines the final concentration of urine.

28. • Normal Hydration and Normal ADH levels Conditions Affecting Final Urine Volume • water channels are present moderate water permeability. • • ADH also facilitates the diffusion of urea(40% of the medullary interstitial osmolarity) out of the medullary collecting duct into the interstitium. • • Normal urine has an osmolarity of about 600 milliosmoles or twice normal body osmolarity.( 1 & ½ liters/day) Dehydration Normal Hydration Overhydration • Dehydration • • Increase ADH leads to : • 1. additional luminal water channels  ↑permeability to water. • 2. increased permeability of the duct to urea  ↑ interstitial osmolaritydraws additional water from the filtrate. • • severe dehydration osmolarity1400 milliosmoles(4 times normal osmolarity / 400 ml/day ) Overhydration • ADH levels are very low or absent  impermeable to water and urea. • ↓ urea permeability decreases the medullary interstitial osmotic gradient, reducing the water-drawing power of the interstitium. • The final urine, which is dilute and high in volume, may have an osmolarity as low as 100 milliosmoles/ 22.5 liters /day Temporary/ Chronic conditions e.g.,Diabetesinsipidus.