human renal physiology

evidence that other factors are involved in potassium regulation

A steep relationship between plasma K+ and urinary K+ excretion is maintained even in adrenalectomised animals given constant aldosterone infusion. Direct effects of plasma [K+ ] appear to be responsible.

effect of calcitonin on kidney

Calcitonin (CT) does not appear to have any significant renal effects.

Renal blood supply

Blood (25% of cardiac output) reaches the kidneys in the renal arteries. Each renal artery divides to form interlobar arteries, these give rise to arcuate arteries which in turn supply cortical radial arteries (interlobular arteries). The cortical radial arteries give rise to afferent arterioles that supply the glomerular capillaries. Each cortical radial artery gives rise to a column of glomeruli. About 20% of the plasma is filtered from the glomerular capillaries. The remainder of the blood continues in the efferent arterioles that then divide into the peritubular capillaries. The efferent arterioles of cortical nephrons supply a profuse peritubular network of cortical capillaries. The efferent arterioles of the juxtamedullary nephrons supply capillary networks in the medulla. Most form the vascular bundle of the outer medulla, with counter-current flow in the afferent and efferent limbs of the capillaries. The inner medulla is supplied by long, hairpin capillary loops, the vasa recta, derived from the vascular bundles of the outer medulla. Blood flow is counter-current in the two limbs of each vas rectum. Blood from the peritubular capillaries drains into cortical radial veins, then arcuate veins, interlobar veins and finally the renal veins. Note that the efferent arteriole and its peritubular capillaries are usually dissociated from the tubule arising from the same glomerulus (i.e. supply a different tubule). In fact, any one tubule is supplied by peritubular capillaries from several efferent arterioles see image

measuring renal plasma flow using a clearance method

If a substance is completely removed from the plasma on one passage through the kidneys, its clearance will be equal to the renal plasma flow (RPF). Para-amino hippurate (PAH) is filtered and actively secreted into the tubules. If the concentration of PAH is kept low (to avoid saturating the secretory mechanism - see later), it is completely removed from the plasma in the peritubular capillaries. PAH clearance can be used as an estimate of RPF. In practice, PAH is only removed completely from blood flowing through the cortical peritubular capillary network. Blood flowing through non-secreting capillaries (such as renal fat and the medulla) is not cleared of PAH. PAH clearance therefore measures the effective renal plasma flow (ERPF). The ERPF is about 90% of the RPF. So RPF~ERPF/0.9. RPF in humans is about 700 ml min-1 . To more accurately determine RPF, the Fick principle can be used

measuring the GFR using clearance

If a substance is freely filtered, but is neither secreted nor reabsorbed, all of the substance filtered will appear in the urine and the volume of plasma cleared will be the volume filtered. Hence the clearance of a substance with these characteristics will be the GFR. Experimental estimation of GFR A substance used to estimate GFR using a clearance method must: Be freely filtered Not be secreted Not be reabsorbed Not be metabolised or synthesized by the body Not alter renal function (not alter the GFR) Be non-toxic Such a substance is inulin, a polymer of fructose (found in artichoke and dahlia tubers). Inulin has a molecular weight of 5,500 and a molecular radius of 1.48 x 10-9 m. Inulin is infused at a slow, steady rate until its concentration in arterial plasma becomes constant. At this time, the rate of infusion must equal the rate of excretion, so: GFR = Rate of infusion ( = Rate of excretion in urine) mg min-1 / Plasma concentration of inulin mg ml-1 The GFR in humans is about 125 ml min-1 .

detection and regulation in osmoregulation

Osmoreceptors in the hypothalamus detect very small changes in plasma osmolality and regulate the secretion of antidiuretic hormone (ADH). ADH regulates the water permeability of the connecting tubule and collecting duct, so determining water reabsorption driven by the high osmotic pressure created by countercurrent multiplication in the medulla. ADH also increases the urea permeability of the inner medullary collecting duct, which increases medullary osmotic pressure and so the concentrating ability of the kidney

Water handling by the kidney: DCT and connecting duct

More NaCl is reabsorbed in these segments, but little water. Na+ reabsorption here is regulated by the hormone, aldosterone. < 1% of the filtered NaCl remains by the end of the DCT

hormones influencing sodium excretion

The main hormones influencing Na+ excretion are angiotensin II, aldosterone and atrial natriuretic peptide.

vasa recta

The vasa recta allow solute and water absorbed into the medulla to be removed without destroying the osmotic gradient created by the loop of Henle. The vasa recta are long capillary loops. As blood flows down them and encounters medullary interstitial fluid with elevated osmotic pressure, water diffuses out and solute diffuses in. As blood flows back up, water diffuses in and solute diffuses out. This countercurrent exchange diffusion maintains the medullary gradient whilst the elevated colloid osmotic pressure of the blood in the vasa recta (after filtration) and the fact that equilibration across the endothelium is not complete, allow the vasa recta to carry away solute and water reabsorbed into the medulla.

examples of passive and active transport

Passive: Down an electrochemical gradient Diffusion (simple diffusion through or between cells) Facilitated diffusion (a carrier protein in the cell membrane allows a substance to cross) Solvent drag (movement of solute with water as it is reabsorbed) Active: Against an electrochemical gradient, requiring input of energy Primary Active Transport (carrier uses ATP directly) Secondary Active Transport (coupled transport systems where one substance moves up its gradient as another moves down its gradient. The carrier does not directly use ATP, but energy is put in maintaining the gradient for downhill movement of the coupled substance) Endocytosis (Invagination of the plasma membrane to form vesicles in the cytoplasm, requires ATP)

why regulate ecf vol?

increased ecf vol increases ABP inc ECF -> inc plasma vol -> inc MSFP -> inc Venous return -(starling mechanism)-> inc Stroke vol -> inc CO -> inc ABP

the van't Hoff equation

pi.V = nRT pi = osmotic pressure (Pascals = N m-2) V = volume (l) n = amount of solute (mol) R = gas constant (8.314 JK-1mol-1) T = absolute temperature (K)

effect of plasma concentration on glucose, PAH and inulin clearance

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gross structure of the kidney

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juxta-glomerular apparatus

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structure of the urinary system

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water handling by the kidney: diagram

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reabsorption of hydrogen carbonate in the proximal tubule

see image About 85% of filtered HCO3- is reabsorbed in the proximal tubule

glucose reabsorption in the proximal tubule

see image At normal plama glucose concentrations (about 3.0 - 7.0 mM) all glucose filtered is reabsorbed. If plasma glucose levels rise above normal, some glucose may appear in the urine (glucosuria). This occurs in those with poorly controlled diabetes mellitus, where there is failure to secrete sufficient insulin or to respond normally to it. Glucosuria occurs because the glucose carriers in the proximal tubule become saturated. There is a maximum rate at which glucose can be reabsorbed, the transport maximum or Tmax. The Tmax for glucose in man is about 375 mg min-1 see image The effects of varying plasma concentration of the rates of filtration, reabsorption and excretion of glucose are shown in fig. 22. Note that graphs for reabsorption and secretion are not the straight lines that might be anticipated (with no glucose excretion until the filtered load exceeds Tmax), but instead begin as curves. This so-called splay results from nephron heterogeneity: individual proximal tubules fail to completely reabsorb glucose at different plasma concentrations, because of variations in length or carrier density. Glucose begins to appear in the urine (the renal threshold is reached) at plasma concentrations around 2 mg ml -1 (10 - 11 mMolar).

colloid osmotic pressure effect on sodium excretion

see image Changes in colloid osmotic pressure (COP) affect both glomerular filtration and the uptake of reabsorbed fluid by the peritubular capillaries, since the balance of Starling forces across the capillary endothelium is affected. The expansion of the extracellular fluids by the addition of NaCl and water dilutes the plasma proteins and reduces the COP. As a result, GFR rises and the reabsorption of fluid from the proximal tubule falls (less fluid is drawn into the peritubular capillaries, reducing the net transepithelial uptake).

evidence for filtration

see image Fluid in Bowman's capsule can be sampled by micropuncture. Analysis shows almost identical composition to plasma (without protein). Also, filtration is affected by changes in pressure (e.g. it ceases if the ureters are clamped), but not by metabolic inhibitors (it is not directly active). Split oil-drop experiments (stopped-flow perfusion). Mineral oil injected into Bowman's capsule (fig. 19) so that some enters the PCT. A second micropipette is used to inject a test solution (isotonic NaCl), splitting the oil drops. With time, oil drops move towards each other indicating volume decrease. Test fluid can be resampled and shown to be isosmotic with that injected. [14C]inulin can be added to give measure of volume change. This type of experiment was originally performed on Necturus (amphibian) kidney. Varying the [Na+] by isosmotic replacement with mannitol (inert carbohydrate, cannot cross epithelium) showed dependence of water reabsorption on Na+ movement and its isosmotic nature. If luminal [Na+ ] (test solution) low, Na+ leaks into the tubule and volume of test fluid increases (remaining isosmotic).

Water handling by the kidney: cortical collecting duct

Water permeability here is regulated by ADH. Fluid entering the CCD is hypoosmotic to the plasma. When ADH levels are high, water is reabsorbed due to the higher osmotic pressure of the peritubular fluid and the tubule fluid becomes isosmotic with the plasma. Some NaCl is reabsorbed in the CCD. This drives further isosmotic fluid reabsorption (like in the proximal tubule), when water permeability is high. Hence when the kidney is concentrating, the CCD returns the tubular fluid to isosmotic with the plasma prior to further concentration in the medullary collecting duct. This reabsorption is important in limiting the volume of fluid which has to be processed by the medullary collecting duct, where substantial water reabsorption would tend to dissipate the accumulated solute. When ADH levels are low, further NaCl reabsorption makes the tubule fluid slightly more hyposmotic to the plasma than when it entered the CCD

general principles of osmoregulation

Extracellular osmolality is very finely controlled. The normal value is about 286 mOsmolal, range only 282 - 290 mOsmolal. (It is customary to use the round number of 300 mOsmoles - leaving undefined and ignoring the small difference between osmolality and osmolarity). Extracellular osmolality is controlled rapidly and normally at the expense of extracellular fluid volume. There are two major osmoregulatory mechanisms: the regulation of water loss by antidiuretic hormone and the regulation of water intake by thirst. Note control is by regulation of movements of water not solute. Under normal conditions, water intake far exceeds unregulated losses (via the lungs, skin and faeces), so that regulation is by the kidney. ADH secretion in osmoregulation is determined largely by osmoreceptors in the hypothalamus. These detect changes in osmotic pressure (but note that the osmotic pressure of the extracellular fluid is determined largely by the concentration of Na+ salts, nevertheless, the receptors are not [Na+ ] receptors).

evidence that LOH needed for urine concentration

Only species with a well developed loop of Henle can concentrate urine substantially. Amphibians, reptiles and birds lack loops of Henle and do not concentrate urine in their kidneys (reptiles and birds concentrate urine in a separate organ the cloaca). In mammals, which have loops of Henle, the length of the loop correlates with the extent to which urine can be concentrated (e.g. long loops in desert rats).

main functions of the kidney

(i) Regulation of electrolyte concentrations in extracellular fluid (ii) Regulation of extracellular fluid pH (in conjunction with breathing) (iii) Regulation of extracellular fluid volume and therefore: (iv) Long-term regulation of blood pressure (v) Regulation of extracellular fluid osmotic pressure Also: (vi) Excretion of metabolic wastes, ingested foreign substances (e.g. some drugs) and inactivated hormones (vii) Regulation of erythropoeisis (red cell production) (viii) Activation of vitamin D3 to 1,25-dihydroxycholecalciferol, which plays an important role in calcium homeostasis. (ix) Gluconeogenesis (in prolonged fasting, the kidneys synthesise glucose from amino acids and other precursors and release glucose into the blood).

Molar vs molal

1 mole of a substance in a total volume of 1 l of solution is at a concentration of 1 molar. 1 mole of a substance dissolved in 1 kg of solvent is at a concentration of 1 molal.

functions of calcium

1) Structural - bones and teeth. 2) Second messenger (e.g. exocytosis, excitation contraction coupling, regulation of many enzymes) 3) Stability of excitable cell membranes The latter is the most important reason for acute extracellular fluid calcium homeostasis. Hypocalcaemia (low ECF [Ca2+]) results in a reduced threshold for action potentials in excitable cells, which can result in spontaneous activity. Motor nerves are particularly susceptible, with spontaneous activity resulting in tetany in the skeletal muscles served. Death can result from asphyxiation due to tetanic contraction of muscles in the larynx. Hypercalcaemia results in sluggish CNS function and muscle weakness as excitable cell activity is suppressed. Calcium phosphates may precipitate, e.g. resulting in kidney stones. However, hypercalcaemia is not particularly dangerous in the short term

effect of 1,23-dihydroxycholecalciferol in the kidney

1,25-DHCC increases reabsorption of Ca2+ and phosphate, an effect compatible with enabling new bone mineralisation. 1,25-DHCC stimulated increased expression of TRPV5, TRPV6, Calbindin-D and the NCX in the distal convoluted tubule and collecting duct cells. It increases expression of the Type II Na+ /Pi transporter in the proximal straight tubule.

stimuli to aldosterone secretion

1. Increased plasma angiotensin II 2. Increased plasma [K+] 3. Decreased plasma [Na+] (unlikely to be an important physiological stimulus). ACTH (adrenocorticotrophic hormone) from the anterior pituitary gland regulates the secretion of glucocorticoids from the adrenal cortex. ACTH does not regulate the synthesis of aldosterone. However, aldosterone is synthesised from a glucocorticoid precursor, so ACTH is a permissive factor in aldosterone synthesis

effects of angiotensin II

1. Stimulates Na+ reabsorption in the proximal tubule (a direct effect). The main action appears to be via increased Na+/H+ exchange. Increased Na+ reabsorption results in increased water reabsorption. 2. Stimulates aldosterone synthesis via AT2 receptors (aldosterone is the only factor which stimulates Na+ reabsorption in the distal nephron). 3. Stimulates thirst. 4. Stimulates sodium appetite (desire to ingest NaCl). 5. Vasoconstriction. Angiotensin II contributes little to general vascular tone at normal circulating concentrations, but has a powerful, relatively selective effect on the efferent arteriole. Elevated levels after volume depletion contribute appreciably to the general increase in vascular tone. AII stimulates vascular smooth muscle via AT1 receptors. Efferent arteriolar constriction results in: i) a fall in hydrostatic pressure in the peritubular capillaries (so lowering renal interstitial hydrostatic pressure and promoting Na+ and fluid reabsorption mainly from the proximal tubule). ii) an increase in the filtration fraction (FF; so increasing peritubular capillary C.O.P., assisting fluid/Na+ reabsorption) and a decrease in renal blood flow (RBF). The decrease in RBF outweighs the increase FF so GFR falls. iii) a decreased blood flow rate through the vasa recta (so increasing the osmotic pressure of the medullary interstitial fluid and passive reabsorption of Na+ from the thin ascending limb of the loop of Henle). iv) stabilisation of GFR (may be important in maintaining sufficient filtration for excretion of waste products in severe hypovolaemia). The proximal tubules and efferent arterioles are more sensitive to angiotensin II than other tissues the hormone acts on. Since small amounts of angiotensin II appear to be generated within the kidney itself, the renin- angiotensin system forms a sensitive intrarenal mechanism for the control of Na+ excretion. The normal circulating level of angiotensin II is 500 - 600 pMolar. With severe Na+ (volume) depletion, its concentration may rise ten-fold. see image

stimuli to renin secretion

1. The afferent arteriole acts as an intrarenal baroreceptor. A fall in pressure in the afferent arteriole promotes renin secretion. 2. The renal sympathetic nerves stimulate renin secretion via ß1-adrenoreceptors (regulation arising from atrial/great vein volume receptors and arterial baroreceptors). 3. Change in composition/flow rate of fluid at macula densa (a fall in NaCl reabsorption at the macula densa occurs if GFR falls and NaCl reabsorption in the proximal tubule or ascending loop of Henle increase, this increases renin release).

distribution of renal blood flow

90% of the renal blood flow is through the cortical peritubular capillary network. 9% is through the vascular bundles of the outer medulla and only 1% through the vasa recta. The vascular structure described above means that the blood supplies to the cortex and medulla are largely independent, with the flows in the two capillary networks separate until reaching the arcuate veins. This is of considerable functional significance and an important part of the mechanism allowing the production of dilute or concentrated urine

calcium distribution in the body

About 1 kg of Ca2+ is present in bone as hydroxyapatite (Ca10(PO4)6(OH)2). Of this about 1 g lines the surface of canals filled with bone fluid, and so is available for exchange with the extracellular fluid. The extracellular fluid contains 1 g of Ca2+. The total concentration in the plasma is 2.5 mM. The free [Ca2+] is normally 1.075 mM. The remainder (1.2 mM) is bound to proteins or complexed with anions (0.225 mM). About 10 g of Ca2+ is present in the cells. Most is sequestered in organelles. The free [Ca2+]i is around 50 - 100 nM.

Water handling by the kidney: loop of henle

About 10% of the filtered water and 25% of the NaCl is reabsorbed in the loop of Henle. Fluid leaving the loop is hyposmotic to the plasma. The uncoupling of NaCl and water reabsorption in the loop of Henle is the key to the kidney's ability to vary the osmotic pressure of the urine, forming dilute or concentrated urine depending on whether water needs to be lost or conserved. The excess solute reabsorbed from the loop raises the osmotic pressure of the medullary interstitial fluid. This in turn allows a regulated reabsorption of water from the medullary collecting duct, determined by plasma levels of antidiuretic hormone (ADH). The loop of Henle raises the osmotic pressure in the surrounding interstitial fluid by counter-current multiplication (described in detail below). The thick ascending limb actively reabsorbs Na+ and Clfollows. This raises the osmotic pressure so that water leaves the thin descending limb by osmosis, increasing the concentration of solute in the tubular fluid left behind. When this more concentrated fluid reaches the thick ascending limb, the reduction in the solute gradient between the tubule lumen and the interstitium is reduced, allowing more NaCl to be actively reabsorbed. Hence more water leaves the thin descending limb and so a more concentrated fluid enters the thick ascending limb, and so on and so on.

Sodium reabsorption in the proximal tubule

About 65% of the filtered Na+ is reabsorbed in the proximal tubule (with water following osmotically). There is some regulation of the amount of Na+ reabsorbed by the hormone angiotensin II. Na+ reabsorption is also influenced by the Starling forces across the peritubular capillaries (if movement of Na+ and water into the peritubular capillaries from the peritubular fluid is reduced, this reduces reabsorption of Na+ and water from the proximal tubule since it has leaky tight junctions). In the distal nephron, the amount of Na+ reabsorbed is under hormonal control, in particular by aldosterone (see later). The plasma concentration of aldosterone determines the rate at which Na+ can be removed from the distal nephron. However, its effect can be overcome, particularly in the short term, by changes in the rate of Na+ delivery from the proximal nephron. Hence changes in GFR and in proximal Na+ reabsorption can both affect the rate of Na+ excretion

Water handling by the kidney: proximal tubule

About 65% of the filtered water is always reabsorbed in the proximal tubule, along with 65% of the Na+ (solute). This is isosmotic fluid reabsorption (the fluid remains isosmotic with the plasma all along the proximal tubule).

water reabsorption in the proximal tubule

About 65% of the filtered water is reabsorbed in the proximal tubule. The tubule here is water permeable so water follows the active reabsorption of Na+ (and other solutes). Hence the fluid in the proximal tubule remains isosmotic to the blood in the peritubular capillary network. In the distal nephron, water reabsorption depends on the physiological state of the animal. It is influenced mainly by the plasma concentration of the water conserving hormone, antidiuretic hormone (ADH), which increases the water permeability of the collecting ducts (see later)

aldosterone - deficiency and excess

Adrenal insufficiency is known as Addison's disease. Symptoms are complex as both aldosterone and glucocorticoids are deficient. A complete absence of aldosterone results in natriuresis, which leads to reduced extracellular fluid and so plasma volume, hypotension and circulatory collapse. Regulation of extracellular [K+ ] also fails. Survival is possible if salt intake is massively increased. Aldosterone excess (primary aldosteronism or Conn's disease) results in increased extracellular fluid volume, hypertension, potassium depletion and metabolic alkalosis.

aldosterone - effects

Aldosterone acts on the distal parts of the renal tubule, the main site of action being the cortical collecting duct. It may also act on the thick ascending loop of Henle. Aldosterone has several effects: 1. It promotes Na+ reabsorption. 2. It promotes K+ secretion and excretion. 3. It promotes H+ secretion. The relative importance of these actions is the subject of debate. Although aldosterone is required for normal Na+ reabsorption in the distal nephron, its major regulatory role is now considered to be over K+ excretion. (The older view that aldosterone is a major factor in the regulation Na+ excretion predates the discovery of direct effects of angiotensin II on Na+ reabsorption. At the end of the day, it is clear that both hormones influence Na+ excretion and so participate to a greater or lesser extent in extracellular fluid volume regulation)

aldosterone - secretion

Aldosterone is secreted by the zona glomerulosa, the outermost zone of the adrenal cortex. Other zones of the adrenal cortex secrete glucocorticoid hormones (e.g. cortisol) and some sex steroids. The adrenal medulla secretes adrenalin and noradrenalin.

Reabsorption of proteins

Although only a very small fraction of larger plasma proteins are filtered, this, together with the filtration of smaller proteins, could lead to substantial daily losses. Larger proteins (e.g. albumin) are partially degraded by enzymes on the surface of proximal tubule cells and then taken up by endocytosis. The vesicles formed fuse with lysosomes and the proteins and peptides are then hydrolysed to their constituent aas which leave across the basolateral membrane of the cells by facilitated diffusion. Smaller peptides (e.g. angiotensin II and ADH) are completely hydrolysed in the tubule lumen and the constituent aas reabsorbed together with those that have been filtered (see above). Note that the kidney is an important site of degradation of smaller protein and peptide hormones.

amino acid reabsorption

Amino acids (aa) are more or less fully reabsorbed. The mechanism is like that for glucose. The aa crosses the luminal (apical) membranes of the PCT cells by Na + -coupled, secondary active transport and leave across the basolateral membranes by facilitated diffusion. The are at least five different aa/Na+ symport carriers, one each for: Acidic aa (e.g. aspartic, glutamic) Basic aa (e.g. arginine, lysine) Neutral aa (alanine, leucine) Imino acids (proline, hydroxyproline) Glycine Molecular techniques have revealed a more complicated situation, with at least seven renal amino acid transporters having been identified to date. Overlapping substrate specificity is common.

filtration pressures

As for any capillary, the net filtration pressure across glomerular capillaries is the algebraic sum of the hydrostatic and colloid osmotic pressures acting across the capillary endothelium (the Starling forces). see images

natriuretic factors

Atrial myocytes contain granules of the precursor of atrial natriuretic peptide (ANP). ANP, a 28 amino acid peptide, is present in the plasma and its concentrations are increased when atrial stretch is increased (as in expansion of the extracellular fluid volume). ANP has a number of actions, including: • Inhibition of Na+ reabsorption in the medullary (and probably cortical) collecting duct (a direct action, related to increased intracellular levels of cGMP). • Inhibition of Na+ reabsorption in the proximal tubule (an indirect action, ANP stimulates the proximal tubule cells to secrete dopamine, which in turn inhibits Na+ reabsorption). • Inhibition of renin secretion (so reduced Angiotensin II and aldosterone levels, and reduced Na+ reabsorption). • Dilation of glomerular mesangial cells, which increases surface area for filtration, so Kf and GFR. • Inhibition of aldosterone secretion (a direct effect on the adrenal cortex, as well from reduced renin/angiotensin II) • Vasodilation of the afferent and efferent arterioles. The effect on the afferent is greatest. This raises glomerular capillary hydrostatic pressure, so GFR rises, increasing the amount of Na+ filtered. Peritubular capillary hydrostatic pressure also rises, so increasing renal interstitial hydrostatic pressure, reducing fluid and so Na+ reabsorption. • Inhibition of ADH secretion and the action of ADH on the collecting ducts, so increasing water loss. The combined effects of ANP result in natriuresis (excretion of sodium in urine) see image

PTH actions on the kidney - calcium

Calcium. Normally 99% of filtered Ca2+ is reabsorbed. 70% of the filtered load is reabsorbed by the proximal tubule, and another 20% by the thick ascending limb of the loop of Henle. These percentages are fixed. Regulation is exerted over the reabsorption of the remaining 10% of the filtered load, which is handled by the distal convoluted tubule and collecting duct. Two thirds of Ca2+ reabsorption in the proximal tubule and thick ascending limb occurs paracellularly (driven by the lumen positive transepithelial potential in both segments, and by solvent drag in the proximal tubule). The remaining third is reabsorbed transcellularly. Calcium enters the cells from the tubule lumen via two types of Ca2+ channel, TRPV5 (also called Epithelial Ca2+ Channel; ECaC1) and TRPV6 (also called Ca2+ Transport Protein 1; CaT1). Ca2+ is then ferried across the cell bound to proteins such as Calbindin-D. The Ca2+ is then transported out of the cell by a Ca2+ -ATPase and by a Na+ /Ca2+ exchanger (NCX). Ca2+ reabsorption in the distal convoluted tubule and collecting duct is transcellular (the lumen negative transepithelial potential in these segments precludes paracellular reabsorption). The mechanism is uncertain, but probably reflects the transcellular routes shown for the proximal tubule and thick ascending limb below (fig. 58). PTH decreases Ca2+ reabsorption in the proximal tubule and thick ascending limb, an effect which is secondary to reduced Na+ reabsorption. However, this effect is more than outweighed by stimulation of Ca2+ reabsorption in the distal convoluted tubule and collecting duct. The mechanism involves an increase in [cAMP] and PKA-dependent phosphorylation of the NCX, which increases the transport maximum for Ca2+ . see image

factors affecting GFR

Changes in any of the filtration pressures or the filtration coefficient will change GFR. (The changes mentioned here are those that would occur if a given factor could be changed in isolation). K - Increased surface area for filtration because of relaxation of mesangial cells. Increases GFR Pc - (a) Increased renal arterial pressure (b) Decreased afferent arteriole resistance Both increase GFR (c) Increased efferent arteriole resistance Decreases GFR (increased Pc is outweighed by decreased renal blood flow due to increased resistance Pb - Increased intratubular pressure caused by obstruction of tubule or extrarenal urinary system. Decreases GFR pic - (a) Increased systemic plasma colloid osmotic pressure (determines pic at start of glomerular capillaries) (b) Decreased renal plasma flow (determines rate of rise in pic along capillaries) Both decrease GFR

autoregulation of renal blood flow and GFR

Changes in arterial blood pressure might be expected to cause large changes in renal blood flow (RBF) and GFR. For any organ: Blood flow = Arteriovenous Pressure Difference / Resistance However, over a wide range of physiological blood pressures, relatively small changes in RBF and GFR occur. The kidneys respond to changes in pressure and flow rates to minimise the effect on RBF and GFR. This is autoregulation. Autoregulation is achieved by changing renal vascular resistance, mainly the resistance of the afferent arteriole. see image Two mechanisms are believed to contribute to autoregulation: a myogenic mechanism and tubulo-glomerular feedback.

two types of nephron

Cortical nephrons -renal corpuscles in the cortex (outer region) of the kidney. -no part in inner medulla -all of the loops of the cortical nephrons in outer medulla -The medullary rays contain the cortical parts of the collecting ducts, some of the proximal straight tubules of the cortical nephrons and some of the thick ascending limb of the loops of Henle of the cortical nephrons Juxtamedullary nephrons -renal corpuscles in the cortex, close to the junction with the medulla. -thin limbs of the juxtamedullary nephrons and large collecting ducts in inner medulla. -upper portions of the loops of Henle (including all the thick ascending limbs) in outer medulla The outer stripe of the outer medulla is where the proximal straight tubules penetrate the medulla. The medullary rays are finger-like extensions of the medulla up into the cortex. . Note that individual collecting ducts serve both types of nephron. see image

evidence that aldosterone is a major potassium (not sodium) regulating hormone

Dogs are adrenalectomised to stop endogenous aldosterone secretion and infused with aldosterone to maintain the normal plasma level of the hormone seen in Na+ replete animals. They are able to regulate sodium balance and blood pressure when severely deprived of dietary Na+ (so other mechanisms can cope alone when [aldosterone] cannot vary). In contrast, they are unable to regulate plasma [K+ ] when the amounts of K+ in the diet are varied. The ability to regulate Na+ without changes in aldosterone is not surprising given the many alternative mechanisms available. These experiments indicate that changes in aldosterone are not essential for Na+ homeostasis, rather than that they are not normally involved. see image

overview of Ca regulation

ECF free [Ca2+] is tightly regulated (around 1.075 mM in man). Hypocalcaemia is potentially fatal since it results in increased excitability in excitable cells, which can cause tetany in skeletal muscles and asphyxiation when respiratory muscles are involved. The major hypercalcaemic hormone is parathyroid hormone (PTH), which acts to mobilise Ca2+ from bone and increase Ca2+ reabsorption (and decrease phosphate reabsorption) in the kidney. PTH acts indirectly to increase Ca2+ absorption in the gut via the promotion of the synthesis of another hormone, 1,25-dihydroxycholecalciferol (1,25-DHCC), which is produced from vitamin D3. There is also a hypocalcaemic hormone, calcitonin (CT), whose main role may be to prevent excessive bone demineralisation, especially when Ca2+ demand is high in pregnancy and lactation. Non-renal aspects of calcium and phosphate handling were covered in Dr Mason's lectures on endocrinology.

effect of arterial blood pressure on sodium excretion

Expansion of the extracellular fluid volume raises plasma volume that tends to increase arterial blood pressure. The baroreceptor reflexes serve to minimise such changes in the short term, but they adapt over about 24 hours. A rise in arterial blood pressure increases the GFR that leads to increased Na+ loss. These changes occur despite autoregulation and glomerular-tubular balance, which are not perfect. The increased Na+ loss caused by an increase in arterial blood pressure is called a pressure natriuresis. The response has two components. Increased glomerular capillary hydrostatic pressure (Pc) increases net filtration pressure and so GFR. Increased peritubular capillary hydrostatic pressure reduces movement of fluid into these capillaries, raising renal interstitial hydrostatic pressure that in turn reduces fluid reabsorption from the tubule. The main effect is on the leaky proximal tubule, where increased renal interstitial hydrostatic pressure increases back leakage. see images Note that the effect of arterial pressure changes in the whole animal (in vivo) are greater than in perfused kidneys (in vitro), indicating that other factors play an important part in the integrated response.

structure of the renal corpuscle

Filtration occurs in the renal corpuscle as fluid is forced from the glomerular capillaries into the space of Bowman's capsule. see image

filtration barrier

Fluid from the lumen of the glomerular capillaries passes through fenestrations in the capillary endothelium, the capillary basement membrane and then filtration slits between the interdigitating processes of podocytes. see image

basic renal mechanisms: summary

Glomerular filtration About 20% of the plasma (water and crystalloids) are filtered into Bowman's capsule. Tubular reabsorption Most of the filtered material is reabsorbed. About two thirds is reabsorbed in the proximal convoluted tubule, most of the rest in later segments of the tubule. Reabsorbed material passes from the tubular fluid through and/or between the tubular cells to the peritubular fluid and thence enters the peritubular capillaries. Tubular secretion Some substances are secreted into the tubule and so are drawn out of the capillaries of the peritubular capillary network. Urinary excretion The fluid leaving the collecting ducts, carrying any filtered or secreted material not reabsorbed, passes down the ureters to the bladder and is excreted. see image

regulation of potassium excretion - aldosterone

High plasma [K+ ] directly stimulates aldosterone synthesis and so release. Aldosterone increases the amount of Na+ /K+ -ATPase in the principal cells. The resulting increased pumping of K+ into the cells increases the gradient for K+ efflux across the luminal membrane. The K+ efflux is promoted by the aldosterone-evoked increase in luminal K+ permeability (increased K+ channel density). Also, the increased luminal Na+ permeability (increased Na+ channel density) favours Na+ reabsorption, so K+ secretion.

dilution technique for volume determination

If a known amount of a substance A is mixed into a volume V and its final concentration is C, then: V (l)x C (mg l-1) = A (mg) To estimate the volume of a fluid compartment, a known amount of a marker substance is injected. While waiting for equilibration, any marker substance excreted from the body is collected and measured, and metabolic losses are estimated. The final concentration of the marker in the fluid of the compartment is measured and the volume of distribution calculated (correcting the amount injected for excretory and metabolic losses). To accurately estimate the volume of a fluid compartment, the marker substance should ideally: Be restricted to one compartment Distribute evenly throughout that compartment Not change the volume of the compartment Not be excreted or metabolised Be non-toxic Be easily measurable The simple dilution technique can be used to estimate total body water. Radioactive markers used are D2O (deuterium oxide) or HTO (tritiated water). Alternatively a drug, antipyrine, can be used. In practice, the criteria listed above can often only be partially met. Excretion and metabolism are usually a problem. Two modifications of the dilution method allow more accurate estimates to be made.

internal k balance

In humans, about 4 Moles of K+ are present intracellularly ([K+]i = 150 mMolal) and only 50 mMoles in the extracellular fluids. Shifts of K+ between the two compartments can present major challenges to plasma [K+] regulation. Hormones. The large intake of K+ after a meal would be fatal if all of it was present in the extracellular fluids at the same time. Temporary uptake by cells is essential since renal excretion of K+ is relatively slow. Insulin is the main factor promoting cellular uptake of K+ after a meal (larger [K+] rises in diabetes mellitus). Adrenalin and aldosterone also promote K+ uptake by cells. Acid-base balance Metabolic acidosis increases plasma [K+] as H+ enter cells and K+ leave. The reverse occurs in metabolic alkalosis. Plasma osmolality. A rise in extracellular osmolality causes cell shrinkage, so [K+]i rises and K+ leaves the cells. A fall in osmolality has the reverse effect. Cell lysis The lysis of cells (e.g. severe burns) releases large amounts of K+ into the extracellular fluid. Exercise. Plasma K+ rises during exercise as K+ leaves skeletal muscle cells during electrical activity. Adrenalin tends to oppose the effect (larger rises seen in people taking ß-blockers).

filtration equilibrium

In some species (e.g. rat), the net filtration pressure falls from about 10 mmHg (1.4 kPa) at the beginning of the glomerular capillaries to 0 mmHg at some point along the capillaries. When the net filtration pressure is zero, filtration ceases and filtration equilibrium is reached. In other species (e.g. dog and humans), filtration equilibrium is not achieved and filtration occurs along the whole length of the glomerular capillaries. It is believed that the situation in man is like that in the dog, but this is still not certain. A non-equilibrium condition means that the GFR is significantly influenced by small changes in the filtration coefficient of the barrier. see image

renal nerves in sodium excretion

Increased activity in the renal sympathetic nerves: Increases the secretion of renin (resulting in increased production of the Na+ retaining hormones, angiotensin II and aldosterone. Angiotensin II constricts the efferent arteriole more than the afferent, increasing the filtration fraction, raising peritubular capillary C.O.P. and lowering peritubular capillary hydrostatic pressure, so promoting Na+ (fluid) reabsorption). Constricts the renal arterioles (so reducing GFR and renal blood flow and increasing the filtration fraction, with consequences as above. So, less Na+ is filtered and a larger fraction is reabsorbed). Directly stimulates Na+ reabsorption (mainly in the proximal tubule, via a1-adrenoreceptors). (The effect appears to be via promotion of Na+ /H+ exchange). Renal sympathetic nerve activity is modulated in response to changed inputs to the CNS from volume receptors (atria and great veins) and baroreceptors (carotid sinus and aortic arch). Transplanted kidneys (which are denervated) function satisfactorily. This reflects the many systems influencing Na+ excretion, rather than a lack of importance of the renal nerves. In the absence of one system, changes in the activity of others can compensate. see image

Tubulo-glomerular feedback

Increased arterial pressure increases Pc and so GFR. This increases the rate of flow through the tubule. The increased flow rate increases the rate of delivery of Na+ and Clto the macula densa and so the rate at which the cells in this segment reabsorb Na+ and Cl- . The monitored variable is thought to be luminal fluid [Cl- ] (the Na+ sites on the Na+ /2Cl- /K+ co-transporter are always saturated, saturation of the Clsites varies over the physiological range of tubular [Cl- ]). The effect of increased flow at the macula densa is to cause increased release of ATP, which constricts the afferent arteriole. This constriction decreases Pc, so GFR, and reduces RBF. There may also be a role for adenosine in signalling from the macula densa to the afferent arteriole. The adenosine may be formed extracellularly from released ATP. The macula densa also signals to the glomerular mesangial cells. The signal may be chemical (ATP and/or adenosine) and may also involve intercellular signalling via extraglomerular mesangial cells. Contraction of the glomerular mesangial cells reduces glomerular surface area, so Kf and thus GFR. The role of the renin-angiotensin system (see later) in tubulo- glomerular feedback is uncertain. It may be important when ABP and GFR are low. The active end product of this system, angiotensin II, constricts the efferent arteriole more than the afferent. This raises Pc, helping to maintain GFR when ABP is low (e.g. after haemorrhage). It is essential that some filtration is maintained so that toxic waste products can be excreted. see image

regulation of potassium excretion - tubular fluid flow rate

Increased flow rate increases K+ secretion, a fall reduces it. As K+ enters the tubule lumen, [K+ ] rises, reducing the driving force for further K+ entry. A high flow rate reduces the [K+ ] rise, a slow flow rate increases it. The effect of tubular flow rate on K+ secretion helps explain why simultaneous Na+ (ECF volume) and K+ homeostasis can be achieved although the same cells (the principal cells) and hormone (aldosterone) are involved in both. For example, in hypovolaemia, Na+ reabsorption is promoted by high aldosterone. This would be expected to lead to K+ loss. However, it does not since the greater reabsorption of Na+ and water in earlier segments (promoted by multiple mechanisms) greatly reduces the rate of fluid delivery to the distal convoluted tubule and collecting duct. This reduced flow rate opposes K+ secretion. The reverse applies to hypervolaemia. Osmoregulation employing ADH might also be expected to disturb K+ homeostasis, but does not since ADH itself stimulates K+ secretion (by increasing luminal membrane K+ permeability). Hence in antidiuresis, when high ADH slows tubular flow rate, the effect of this in reducing K+ secretion is offset by the direct action of ADH on principal cell luminal K+ permeability. Equally, when ADH is low, the water diuresis does not increase K+ secretion since low ADH reduces the K+ permeability of the principal cell luminal membranes.

regulation of potassium excretion - plasma K+

Increased plasma K+ , as well as stimulating aldosterone secretion, promotes the activity of the Na+ /K+ -ATPase and so K+ loss across the luminal membrane. In addition, luminal membrane K+ permeability appears to increase (independently of increased aldosterone) when plasma [K+ ] rises. The mechanism is unknown.

sympathetic modulation of GFR

Increased renal sympathetic nerve activity and increased levels of circulating catecholamines evoke - adrenergic constriction of the afferent and to a greater extent the efferent arterioles. Although the filtration fraction rises due to an increase in Pc, except for small increases in sympathetic tone, renal blood flow is reduced to a greater extent, such that there is a fall in GFR.

external k balance :potassium reabsorption in the proximal tubules

K + reabsorption in the proximal tubule is by paracellular diffusion, the gradient for which is created by water reabsorption (hence by Na+ reabsorption)

external k balance : k reabsorption in the thick ascending loop of henle

K + reabsorption in the thick ascending loop of Henle occurs partially transcellularly by secondary active transport (K+ crosses the luminal membrane on the Na+ /2Cl- /K+ co-transporter and leaves across the basolateral membrane by co-transport with Cl- ). The remainder of the K+ reabsorption in this segment is paracellular, driven by the lumen positive transepithelial potential.

external k balance :renal potassium handling

K+ is freely filtered. 67% is reabsorbed in the proximal tubule and 20% in the loop of Henle. These fractions are constant. The distal tubule and collecting duct can show net reabsorption (in hypokalaemia) or net secretion (normal and in hyperkalaemia) of K+. Normally, 15% of filtered K+ is excreted. This amount can vary between 1% and 80%. Regulation is on K+ secretion in the distal tubule and collecting duct.

integrated response to expansion on ECF vol

Long term responses to correct volume are shown. In the short term, a reversal of the short term processes summarised under fig. 55 will minimise the effect of ECF volume expansion on blood volume, MCFP and ABP see image

Water handling by the kidney: urea movements

Many segments of the nephron are poorly permeable to urea, so as water is reabsorbed, urea is left behind and its concentration in the tubular fluid rises. ADH increases the urea permeability of the inner medullary collecting duct (IMCD). When ADH levels are high, urea passively diffuses out of the IMCD into the medullary fluids. Some of the urea diffuses into the thin parts of the loop of Henle (particularly the thin descending limb) and recycles (leaving the IMCD again). Urea recycling allows a high [urea] to be built up in the medulla. Urea can contribute 50% of the osmotic pressure of the medullary fluids in a maximally concentrating human kidney. A high medullary [urea] draws out water from the thin descending limb of the loop of Henle. More water leaves the descending limb in the outer medulla and, unlike in the diluting kidney, water also leaves the descending limb in the inner medulla. As a result the fluid reaching the apex of the loop of Henle in the concentrating kidney is more concentrated than in the diluting kidney. As fluid moves up the thin ascending limb, it is more concentrated than the fluid in the surrounding interstitium, so NaCl passively diffuses out of the thin ascending limb. Even after this passive NaCl movement, the fluid entering the thick ascending limb is more concentrated than in the diluting kidney, so more NaCl is actively reabsorbed into the medullary interstitium. The increased concentration of NaCl in the medullary interstitium draws out more water from the collecting ducts, resulting in the formation of a more concentrated urine when ADH levels are elevated. When ADH and so IMCD urea permeability is low, urea remains in the tubule and the medullary [urea] gradually dissipates as urea diffuses into the thin parts of the loop of Henle and is lost in the urine.

reabsorption in the proximal tubule

Micropuncture studies indicate that 70% of the glomerular filtrate is reabsorbed in the proximal tubule. see image Note that: (i) Tubular fluid remains isosmotic with respect to plasma (285 mOsmolal) all along the proximal tubule (water follows solute). (ii) [Na+] remains constant along the proximal tubule. (iii) Glucose, amino acids (), HCO3- , and phosphate are preferentially reabsorbed. The concentrations of glucose and aa normally become zero by the end of the proximal tubule. (iv) The preferential reabsorption of HCO3- results in a rise in [Cl-] along the proximal tubule, maintaining equality between anion and cation concentrations

PTH action on kidney - phosphate

Normally 90% of filtered phosphate is reabsorbed. 80% of the filtered load is reabsorbed by the proximal tubule and another 10% in the distal convoluted tubule. Reabsorption in the proximal tubule is transcellular. The mechanism in the distal tubule is uncertain. The properties of phosphate reabsorption resemble those of glucose, showing a transport maximum and splay, but differ in two ways. Firstly, the Tmax for phosphate is close to the normal filtered load, with the kidneys performing "overflow" control. Secondly, unlike glucose, the Tmax for phosphate is under hormonal control by PTH, which decreases it. Inhibition of phosphate reabsorption by PTH is important. Calcium phosphate is poorly soluble and the two ions exhibit a solubility product relationship ([Ca2+]3[PO4 3- ] 2 = constant), thus decreasing [PO4 3- ] favours the dissolution of calcium phosphate and a rise in ECF free [Ca2+]. Similarly, bone resorption releases Ca2+ and phosphate, which would precipitate if phosphate excretion was not increased. Normally 90% of filtered phosphate is reabsorbed. 80% of the filtered load is reabsorbed by the proximal tubule and another 10% in the distal convoluted tubule. Reabsorption in the proximal tubule is transcellular. The mechanism in the distal tubule is uncertain. see image The properties of phosphate reabsorption resemble those of glucose, showing a transport maximum and splay, but differ in two ways. Firstly, the Tmax for phosphate is close to the normal filtered load, with the kidneys performing "overflow" control. Secondly, unlike glucose, the Tmax for phosphate is under hormonal control by PTH, which decreases it. Inhibition of phosphate reabsorption by PTH is important. Calcium phosphate is poorly soluble and the two ions exhibit a solubility product relationship ([Ca2+]3[PO4 3- ] 2 = constant), thus decreasing [PO4 3- ] favours the dissolution of calcium phosphate and a rise in ECF free [Ca2+]. Similarly, bone resorption releases Ca2+ and phosphate, which would precipitate if phosphate excretion was not increased. The mechanism by which PTH regulates phosphate reabsorption is disputed and not fully understood. The main control over the Type IIa transporter is more likely via its removal from the luminal membrane. PKA and PKC phosphorylate the scaffolding protein NHERF-1 (sodium hydrogen exchanger regulatory factor 1), causing it to dissociate from the Type IIa transporter and making it available for removal from the luminal membrane by endocytosis. Luminal expression of the Type IIa transporter decreases in minutes after PTH stimulation. Luminal expression of the Type IIc transporter decreases over hours after PTH stimulation. The mechanism is uncertain but may be similar to that for the Type IIa transporter. It is unclear whether the Type III transporter (PiT-2) is regulated by PTH. 1,25-dihydroxycholecalciferol. PTH stimulates the synthesis of 1,25-DHCC in the kidney (see below).

units of osmotic concentration

Osmolality: Number of osmoles per kilogram of water (osmolal) Osmolarity: Number of osmoles per litre of solution (osmolar)

Potassium homeostasis - overview

Plasma [K+ ] is closely regulated at around 4.5 mMolal. Both hyperkalaemia ( > 5.5 mMolal) and hypokalaemia ( < 3.5 mMolal) are dangerous conditions because extracellular [K+ ] affects cell membrane potential and so the activity of excitable cells. Acid-base balance is also affected by failure of K+ homeostasis. The normal dietary intake of K+ is about 100 mMoles per day. About 10 mMoles is lost in the faeces and sweat (unregulated) and the remaining 90 mMoles is excreted in the urine (regulated). Aldosterone has an important role in regulating K+ secretion and so excretion. Changes in the flow rate of tubular fluid also play a part.

renin-angiotensin system

Renin is a proteolytic enzyme secreted by modified smooth muscle cells in the wall of the afferent arteriole. Renin catalyses the production of angiotensin I, a decapeptide, from a precursor plasma globulin, angiotensinogen. Angiotensin I is further cleaved to an octapeptide, angiotensin II, by angiotensin converting enzyme (ACE), found mainly in the capillaries of the lungs. see image

evidence for isosmotic fluid reabsorption in the proximal tubule

Simple Micropuncture. Samples from early and late PT reveal no change in osmotic pressure. If animal injected with inulin before sampling so that inulin is present in tubules, the [inulin] is found to rise. Since inulin is not secreted or reabsorbed, the increased [inulin] must be caused by fluid reabsorption. In other words, the inability of the tubules to transport inulin means that it can be used as an indicator of fluid (water) movement.

autoregulation: myogenic mechanism

Smooth muscle in the afferent arteriole contracts when stretched and relaxes when released from stretch.

osmotic comparisons

Solutions with the same osmotic pressure are said to be isosmotic. One solution is said to be hyperosmotic to another if it has a higher osmotic pressure or hyposomotic to another if it has a lower osmotic pressure

Clearance as an indicator of renal handling: clearance ratios

Substances with clearances less than inulin show net reabsorption (or are not freely filtered). Substances with clearances greater than inulin show net secretion. Such comparisons can be expressed as clearance ratios: clearance ratio of X = Clearance of X/Clearance of inulin = clearance of X/GFR

integrated response to volume depletion

Take the long term response to haemorrhage as an example, when isosmotic fluid loss occurs. Note that in the short term, autotransfusion, a shift of interstitial fluid into the blood due to reduced capillary hydrostatic pressure, occurs. This helps maintain blood volume, MCFP and ABP As do the rapid sympathetic cardiovascular reflexes and vasoconstriction evoked by ADH and angiotensin II. see image

estimating renal plasma flow using the fick principle

The Fick principle can be used to determine the blood flow (or plasma flow) through any organ which adds or removes a substance to or from the blood. flow rate (ml/min) = rate of removal (mmol min-1) / arteriovenous conc difference (mmol ml-1) see image Practically, PAH is infused at a slow, steady rate into a leg vein. Blood is sampled from an arm vein until the plasma concentration becomes constant. At this time, the rate of infusion must equal the rate of removal of PAH by the kidney (rate of excretion = R). A sample of arterial blood is taken to determine arterial plasma [PAH] ([PAH]a) and a sample of renal venous blood is drawn from a catheter in a renal vein to determine renal venous [PAH] ([PAH]v)

massive but selective reabsorption of filtered materials

The GFR is about 125 ml min-1 , but urine is only formed at a rate of about 1 ml min-1 . It follows that about 99% of the filtrate must be reabsorbed with only 1% being finally excreted. Urine composition is very different to that of the filtrate: Substances required by the body are selectively reabsorbed from the filtrate whilst some other substances are actively secreted into it see image

concept of clearance

The clearance of a substance is a measure of the rate at which it is being removed or cleared from the plasma by the kidneys. Clearance has units of ml min-1 and is calculated by dividing the rate of excretion of a substance in the urine by its plasma concentration. If a substance X is present in arterial plasma at a concentration [X]a mg ml-1 , the urine flow rate is v ml min-1 and the urinary concentration of X is [X]u mg ml-1, then: Rate of excretion of X is v[X]u mg min-1 Clearance of X is v[X]u/ [X]a ml min-1 In words: The clearance of a substance is the (hypothetical) volume of plasma which would have had to have been completely cleared of that substance each minute to account for the rate of excretion of that substance in the urine Clearance provides a means of comparing how the kidney handles different substances, by calculating back to find the volume of plasma which is theoretically cleared of that substance each minute. A substance with a large clearance is more avidly removed than a substance with a low clearance. substance in the urine

external k balance : potassium transport in the distal tubule and collecting duct

The distal convoluted tubule and cortical collecting ducts are the segments where K+ homeostasis is achieved, and may show net secretion or reabsorption. K+ is secreted by the principal cells and reabsorbed by the type-A intercalated cells. Regulation is achieved by varying the activity of the principal cells. Secretion of K+ by the principal cells depends on: The activity of the basolateral membrane Na+ /K+ -ATPase. The electrochemical gradient for K+ exit across the luminal membrane. The permeability of the luminal membrane to Na+ (Na+ entry stimulates the Na+ /K+ -ATPase). The permeability of the luminal membrane to K+ . Reabsorption of K+ by the type-A intercalated cells is driven by the luminal membrane H+ /K+ -ATPase. K+ leaves the cell down its concentration gradient via K+ channels.

aldosterone - mechanism of action

The effects of aldosterone on Na+ reabsorption and K+ excretion in the distal nephron are due to it stimulating new protein synthesis in the principal cells. As such, its actions are slow (taking about an hour to produce significant effects). The resulting aldosterone-induced proteins include channels for Na+ and K+, and, at least in the long term, more Na+/K+-ATPase (fig. 53). The Epithelial Na+ channel (ENaC), found in many epithelia including mammalian distal nephron, shows increased activity and is increased in density when aldosterone levels rise. Small conductance K+ channels (SK channels) believed to be responsible for K+secretion increase in density in rat nephron after aldosterone treatment. An increase in large conductance K+ channels (maxi-K or BK channels) has been demonstrated in amphibian kidney, but it is not clear that these channels are involved in K+ secretion. see image The additional Na+ channels (ENaC) increase the rate of entry of Na+ across the luminal membrane and the resulting rise in cytosolic [Na+] stimulates the active removal of Na+ across the basolateral membrane by the Na+/K+-ATPase. The new Na+/K+-ATPases increase the overall Na+ pumping capacity. The increased extrusion of Na+ from the cell is coupled to increased pumping of K+ into the cell. This favours increased diffusion of K+ into the tubule lumen, which is aided by the additional K+ channels and an enhanced lumen-negative transepithelial potential. Increased H+ secretion results from the action of aldosterone on the type-A intercalated (acid-secreting) cells of the distal nephron. K+/H+-ATPase activity is increased, an action which does not appear to be dependent on new protein synthesis. The greater lumen-negative transepithelial potential also favours K+/H+ -ATPase activity.

principles of volume regulation

The extracellular fluid represents about 38% of total body water. Na+ is the main cation and since it is excluded from the cells (low membrane permeability and activity of the Na+/K+-ATPase), the volume of the extracellular fluid is determined by its Na+ content. The osmoregulatory system controls water content to ensure appropriate dilution of the extracellular (and so other) fluids. The osmoregulatory system finely controls osmotic pressure. Hence extracellular fluid volume cannot be tightly controlled and shows short-term (acute) changes of about -10% to +20%, caused by absorption of water and salt, fluid loss in urine between drinking, production of secretions, etc. In other words, volume regulation is normally subordinate to osmoregulation and osmoregulation is at the expense of volume regulation. Most processes regulating extracellular fluid volume are slow, acting over hours or days, although sudden, large changes can be met with more rapid compensations. Regulation is mainly achieved by varying the loss of Na+ in the urine against a normal intake. In addition there appears to be some regulation of intake in response to severe volume depletion (sodium appetite). Note control is by regulation of movements of solute not water

composition of the glomerular filtrate

The glomerular filtrate is almost protein free (~10 mgl-1protein). However, given the large volume of filtrate, this would lead to considerable loss (~1.8 g day-1) without reabsorption. In essence, all the normal constituents of plasma except proteins pass through the filtration barrier. see table

glomerular filtration rate

The glomerular filtration rate (GFR) depends on the net filtration pressure, the hydraulic permeability (water permeability) of the filtration barrier and the surface area of the filter. Kf = filtration coefficient = hydraulic permeability x surface area GFR = Kf[(Pc - Pb) - (pic - pib)] = Kf[(Pc - Pb) - pic] (since pib = 0)

other organs and processes involved in regulation of excretion

The kidney is the major regulator of excretion, determining the rate of loss of substances in the urine, but works in conjunction with other processes:- (i) Ingestion - There are regulated drives for ingestion of water, food and salt (thirst, hunger and sodium appetite) (ii) Other Routes For Excretion - Breathing (CO2, water, other volatile chemicals); Faeces (bile components, other gastro-intestinal secretions); Secretions (sweat, milk, tears, etc) (iii) Changes In Metabolism - The liver alters the composition of various substances to allow their excretion in urine or bile (iv) Controls On Absorption - Iron and zinc uptake are controlled by the intestinal epithelium.

counter current multiplication - thin ascending limb

The model above ignores the thin ascending limb, which unlike the thick limb, does not actively reabsorb solute. However, the thin ascending limb is permeable to NaCl and is relatively impermeable to water. When urea is present in the medullary interstitium (see below), more water leaves The thick ascending limb of the loop of Henle actively reabsorbs NaCl, but since it is impermeable to water, water does not follow and a hypotonic fluid (about 100 mMolal) is produced. Na+ is actively pumped across the basolateral membrane and enters across the luminal membrane via a Na+ /2Cl- /K+ symporter. K+ and Clleave across the basolateral membrane via a symporter, but some K+ re-enters the lumen via a K+ channel. This generates a positive transepithelial potential (lumen +ve), which drives further Na+ (and other cation reabsorption) paracellularly (via the tight junctions). 32 the thin descending limb of the loop of Henle than in the diluting kidney. As a result, the tubular fluid in the thin ascending limb, flowing up from the apex of the loop, has a higher [NaCl] than the surrounding medullary interstitial fluid, so NaCl diffuses out. This passive reabsorption of NaCl explains why in the concentrating kidney the osmotic gradient in the medulla extends beyond the thick segments of the loop of Henle

Sodium homeostasis

The normal daily intake of Na+ in the diet exceeds the normal non-renal losses, so regulation of body Na+ content is by the kidney. see image

physical factors influencing sodium excretion

The physical factors which influence Na+ excretion can be seen in part as automatic compensations for extracellular fluid volume changes which assist direct nervous and hormonal mechanisms in restoring the ECF volume to normal. However, nerves and hormones also exert some of their effects via changes in physical factors ABP Colloid osmotic pressure

secretion of organic ions by the proximal tubule

The proximal tubule actively secretes a number of organic anions, including bile salts, oxalate, prostaglandins and urate. There appears to be only one or a small number of poorly specific carriers for these anions (hence foreign substances such as some anionic drugs - e.g. penicillin, aspirin, furosemide - are secreted). Para-amino hippurate (PAH - used to estimate RPF) is an anion secreted by this system. Since the process is carrier mediated, is shows a transport maximum. Similar mechanisms operate for organic cations. Endogenous substances transported include acetylcholine, adrenalin, noradrenalin and dopamine. Drugs secreted using the same carriers include atropine, isoprenaline, cimetidine and morphine.

the renal tubule

The renal tubule consists of the nephron (which includes Bowman's capsule) and the collecting duct system. The term nephron is commonly used to describe both parts. The nephron is the basic functional unit of the kidney. In recent years there have been attempts to standardise the nomenclature applied to the nephron. The preferred terms which will be used here are given in the figure below. see image

thick ascending limb - in counter current multiplication

The thick ascending limb of the loop of Henle actively reabsorbs NaCl, but since it is impermeable to water, water does not follow and a hypotonic fluid (about 100 mMolal) is produced. Na+ is actively pumped across the basolateral membrane and enters across the luminal membrane via a Na+/2Cl-/K+ symporter. K+ and Clleave across the basolateral membrane via a symporter, but some K+ re-enters the lumen via a K+ channel. This generates a positive transepithelial potential (lumen +ve), which drives further Na+ (and other cation reabsorption) paracellularly (via the tight junctions)

counter current multiplication - thin descending limb

The thin descending limb of the loop of Henle does not reabsorb solute. It is relatively impermeable to solute but it is permeable to water. As fluid flows down the thin descending limb, water is withdrawn osmotically due to the increasing medullary interstitial fluid solute concentration created by the thick ascending limb. This fluid then passes around the apex of the loop, through the thin ascending limb and is fed into the thick ascending limb. Imagine the loop starting completely full of fluid isosmotic with the plasma (Fig. 25). The thick ascending limb creates the maximum transepithelial gradient of 200 mOsm difference. Then fluid which has been concentrated as it moves down the thin descending limb flows into the thick ascending limb. So more solute is pumped out until the maximum gradient is re-established. Raising the basal solute concentration in the thick ascending limb has allowed the medullary interstitial fluid concentration to be raised. The counter-current system can create a much more concentrated fluid than the single effect. Notice however, that at any point along the loop of Henle, the concentration gradient across the epithelium never exceeds 200 mOsm. The counter-current flow allows a relatively modest transverse gradient to be multiplied into a steep longitudinal gradient.

role of urea in the concentrating mechanism

The thin parts of the loop of Henle are relatively permeable to urea as is the inner medullary collecting duct, which becomes very permeable to urea when ADH levels are elevated. The thick ascending limb of the loop of Henle, the distal tubule and early parts of the collecting system are relatively impermeable to urea. In the presence of ADH, urea becomes concentrated due to water reabsorption as fluid moves through the cortical and outer medullary collecting ducts. Urea then diffuses out of the inner medullary collecting ducts, which ADH has made highly permeable to urea. Some of the urea diffuses into the thin parts of the loop of Henle (particularly the thin descending limb) and recycles, gradually building up the concentration of urea in the medullary interstitial fluids. The increased medullary [urea] draws additional water out of the thin descending limb of the loop of Henle. This includes the thin descending limb in the inner medulla, where in the diluting kidney there is no osmotic gradient to cause water movement. As a result, the [NaCl] in the tubule fluid is elevated more than in the absence of urea, now reaching a maximum concentration at the apex of the loop. As a result, NaCl passively diffuses out of the thin ascending limb, extending the medullary solute gradient all the way to the innermost part of the medulla. Even after passive NaCl reabsorption from the thin ascending limb, the fluid entering the thick ascending limb is more concentrated than in a diluting kidney, so additional NaCl is pumped out into the inner medulla, improving counter-current multiplication. Therefore, with elevated ADH, the concentrations of both urea and NaCl in the medullary interstitial fluid rise. The increased [NaCl] results in additional water reabsorption from the collecting ducts. Hence in the presence of urea the concentrating ability of the kidney is increased. It is important to appreciate that the high urea concentration in the medullary interstitium is not directly responsible for the increased water reabsorption from the collecting ducts. Since the collecting duct epithelium is highly permeable to urea in the presence of ADH, urea cannot cause water to cross the collecting duct epithelium. It is an ineffective osmole in this situation. When ADH levels are low, urea enters the thin parts of the loop of Henle (particularly the thin descending limb) and is lost in the urine. NaCl no longer diffuses out of the thin ascending limb but NaCl continues to be lost gradually via the vasa recta. So the concentrations of urea and thus NaCl in the medullary interstitium fall and the concentrating ability of the kidney is reduced.

Distribution of water in the body

The total body water of a 70 kg person is about 42 l. Of this about 25 l is intracellular, 16 l is extracellular and 1 l is transcellular. The extracellular fluid consists of the plasma (3 l), rapidly exchangeable interstitial fluid including lymph (12 l) and relatively inaccessible / slowly exchangeable interstitial fluid in dense connective tissue (1 l). The main transcellular fluids (found in special cavities) are cerebrospinal fluid (brain and spinal cord) and synovial fluid (joints). 50-65% of the body weight is water. 63% in young adult males, 52% in young adult females (because females have more subcutaneous adipose tissue). In neonates, about 75% of body weight is water, which decreases towards 60% over a year. Measurement of body fluid volumes Total body water, the extracellular fluid volume and the plasma volume can be estimated directly. The intracellular fluid volume can be estimated from the difference between total body water and extracellular fluid volume. see image

regulation of ECF vol - overview

The volume of the extracellular fluid is determined by its Na+ content, with the osmoregulatory system ensuring that what is essentially a solution of sodium salts is appropriately diluted. Volume regulation is achieved by varying Na+ loss in the urine. Multiple, interrelated mechanisms are involved. Physical factors, such as glomerular and peritubular capillary hydrostatic pressure and colloid osmotic pressure, influence the GFR and tubular reabsorption. Hormones are also involved, particularly the renin-angiotensin-aldosterone system. Angiotensin II is an important regulator of Na+ excretion. Aldosterone is also involved in normal Na+ homeostasis, but K+ homeostasis is now considered its more important role. Natriuretic factors ( atrial natriuretic peptide) also contribute. Sympathetic activity in the renal nerves, varied by reflexes from the cardiovascular system, also influence Na+ excretion.

Counter current multiplication - why the need

There is a limit to the osmotic gradient that can be generated by active solute pumping across the thick ascending limb epithelium because of the leakage of water and back diffusion of solute. The maximum gradient that can be created by the basic mechanism (the single effect) is a difference of about 200 mOsm. Starting with fluid isosmotic with the plasma (300 mOsm), as received from the proximal tubule, the tubular fluid concentration in the lumen might be lowered to about 200 mOsm, so the maximum medullary interstitial fluid concentration would be about 400 mOsm. The fluid in the collecting ducts can never be more concentrated than the medullary fluid it equilibrates with (when ADH is present), yet urine can be concentrated to more much more than 400 mOsm. The reason for this lies in the countercurrent flow in the two arms of the loop of Henle, and the different solute and water permeabilities of these segments.

thirst

Thirst is stimulated by: High plasma osmotic pressure Osmoreceptors respond to high osmotic pressure as a result of cellular dehydration and shrinkage. They are thought to be different to the receptors controlling ADH secretion. They are less sensitive, requiring an increase in osmotic pressure of 2% Reduced extracellular fluid volume Large ( > 10%) falls in ECF volume cause thirst. This results from reduced inhibition arising from volume and baroreceptors in the circulation. Angiotensin II, which rises after volume depletion, is a stimulant to thirst (is a dipsogen). Dry throat Nervous input from mouth and throat. Not regulatory. The various inputs to the thirst mechanism are integrated in a "thirst centre" in the hypothalamus. Thirst initiates drinking, which appears to be terminated by uncertain mechanisms when sufficient water has been drunk.

constant perfusion method

This is suitable for measuring the extracellular fluid volume with substances that are rapidly excreted. A priming injection of the marker is given and then the marker is infused at a constant rate. The concentration Ce of the marker in the fluid becomes constant when the rate of infusion equals the rate of excretion. Once the concentration is steady (at time X), the perfusion is stopped and the urine is collected until all the marker has been excreted. The amount of marker excreted is measured and used to calculate the volume of distribution by dividing by the steady concentration reached. see image Markers used for estimating extracellular fluid volume using the constant perfusion method include inulin, mannitol, thiosulphate and radioisotopes of Na+ or Cl-. Different markers give different estimates (e.g. inulin underestimates because of slow penetration into dense connective tissue; radioisotopes of Na+ or Cloverestimate because some enters the cells).

single injection method

This is suitable for substances that are not rapidly excreted or metabolised. A single injection of the marker into the compartment to be measured is made and the concentration of the marker measured at intervals. The data are plotted and extrapolated to estimate the concentration C0 that would have resulted at the time of injection if distribution had been even and instantaneous. see image The single injection technique can be used to estimate plasma volume, by injecting Evans Blue (a dye which binds to plasma albumin) or radio-iodinated serum albumin (albumin labelled with 131I). Some albumin leaks from the capillaries and some is metabolised. This method corrects for these losses. The blood volume can be estimated from the plasma volume and haematocrit (fraction of blood occupied by red cells). Blood volume = Plasma volume ÷ (1- Haematocrit).

isosmotic fluid reabsorption in the proximal tubule

This segment of the tubule is water permeable (the tight junctions between the cells are leaky). Water reabsorption directly follows solute reabsorption. Na+ is pumped across the basolateral membranes of the cells by the Na+ /K+ -ATPase. Na+ enters the cells across the luminal (apical) membrane through leakage pathways and via Na+-coupled transport mechanisms (symport with glucose or ; antiport with H+). In the early proximal tubule, little Cl is reabsorbed as HCO3- and certain organic anions are selectively reabsorbed. This raises the luminal [Cl-] In later parts of the proximal tubule, Clis passively reabsorbed paracellularly as it moves down its concentration gradient. Some Clenters the cells in exchange for secreted anions (e.g. oxalate) and leaves across the basolateral membrane by symport with K+ The anions may combine with H+ antiported with Na+, forming their acids which diffuse back into the cell and recycle. This process is thus tertiary active transport of Cl- The effect of the solute reabsorption is to slightly elevate the osmotic pressure in the peritubular fluid so that water moves by osmosis through the leaky tight junctions (paracellularly) and to some extent through the cells (transcellularly). Transcellular movement of water occurs through the water channel, aquaporin 1 (AQP1), which is present in both the luminal and basolateral membranes.

ultrafiltration

Ultrafiltration is the bulk flow of water and substances in solution across a filter. Particles with molecular weights greater than about 70,000 Da are not filtered (plasma albumin = 69,000 Da). There is no hindrance to movement of particles of molecular weights < 7000 Da. Filtration becomes progressively smaller as particle size increases from molecular weights of 7000 Da. This is often called sieving.

Water handling by the kidney: medullary collecting duct

When ADH levels are high and the duct water permeable, water is drawn out by osmosis due to the high osmotic pressure of the medullary interstitial fluid (created by the action of the loop of Henle). Urine hyperosmotic to the plasma (hypertonic urine) results. When ADH levels are low and the medullary collecting duct poorly water permeable, the hyposmotic tubular fluid continues on to form urine hyposmotic to plasma (hypotonic urine).

factors affecting rate of filtration

Whether a particle is filtered or not does not just depend on its size. The particles charge is also important. All three components of the filtration barrier have a fixed negative charge, so that negatively charged macromolecules are filtered less than positively charged ones of the same size. Proteins are anionic at physiological pH. Hence even small proteins normally have a low filterability see image The charge on the filtration barrier has little direct effect on the movement of small inorganic ions, but the retention of negatively charged proteins "holds back" inorganic cations and favours filtration of inorganic anions. Hence the concentrations of these ions is slightly different in plasma and the glomerular filtrate (a Donnan equilibrium exists).

antidiuretic hormone

cyclic nonapeptide MW = 1084 half life of 6 - 10 min. The normal plasma level in human plasma is 3.5 pmolal. The major physiological action of ADH is to increase water reabsorption by increasing the water permeability of all parts of the collecting duct system (including the cortical collecting duct). This effect is reinforced by the action of ADH to increase the urea permeability of the inner medullary collecting duct. Another action of ADH is as a vasoconstrictor (hence its alternative name, vasopressin). Different receptors mediate the osmoregulatory (V2) and vasoconstrictor (V1) effects. These differ in affinity for the hormone. The vasoconstrictor effect only occurs significantly at plasma [ADH] well above the normal osmoregulatory range. The major physiological stimulus controlling ADH release is extracellular fluid osmolality, detected by hypothalamic osmoreceptors. Reflexes from the gut and liver inhibit ADH release during drinking and water absorption. ADH release is also influenced by signals from the circulatory system. The atrial/great vein volume receptors and the arterial baroreceptors inhibit ADH secretion. This inhibition is reduced if there are large falls in blood volume ( > 10%) or arterial blood pressure. There is thus a large rise in plasma [ADH], sufficient to cause vasoconstriction. ADH acts on the principal cells of the collecting duct, which contain vesicles which have the water channel aquaporin 2 (AQP2) present in their membranes. ADH binds to V2 receptors on the basolateral membranes of the principal cells. The V2 receptor is G protein coupled to adenylyl cyclase (AC), which is thus activated, resulting in the formation of the intracellular second messenger, cyclic AMP (cAMP). Cyclic AMP activates protein kinase A (PKA), which phosphorylates AQP2 on serine residue 256 (Ser 256). (Other proteins may also be phosphorylated). This triggers the fusion of the AQP2-containing vesicles with the luminal plasma membrane, thus inserting more AQP2 into the luminal membrane, increasing its water permeability. The water permeability of the basolateral membrane is always high due to the constitutive (permanent) presence of AQP3 and AQP4. The rate-limiting step in water reabsorption is at the luminal membrane. ADH increases urea permeability in the inner medullary collecting duct by stimulating the phosphorylation and so activation of urea transporters (UTs) in the luminal membrane. The main urea transporter is UT-A1. UT-A3 also plays a role. UT-A2 is present in the thin descending limb. ADH activates UT-A2 in the inner medullary thin descending limb, promoting urea recycling.

evidence for osmotic gradient in the medulla

microcryoscopy A kidney from an animal producing concentrated urine is rapidly frozen and sectioned. Solute concentrations can be estimated from the melting points of different regions (solute depresses melting point). The osmotic gradient can be mapped. Osmotic pressure is constant in the cortex and starts to rise from the cortico-medullary boundary, reaching its maximum at the tip of the renal papilla (inner medulla).

daily ca turnover between compartments

see image The figure above represents the situation in a normal adult in calcium balance. Positive calcium balance (intake > loss) occurs in growing children, in pregnancy and during bone healing. Negative calcium balance (loss > intake) occurs in old age, during prolonged bed rest and during prolonged weightlessness (Dr Mike Mason's Easter term Microgravity lecture).