Renal Transport Mechanisms: NaCl And Water Reabsorption Along The Nephron Notes

Renal Transport Mechanisms: Nacl And Water Reabsorption Along The Nephron

The formation of urine involves three basic processes:

(1) Ultrafiltration of plasma by the glomerulus,

(2) Reabsorption of water and solutes from the ultrafiltrate, and

(3) Secretion of select solutes into the tubular fluid. Although an average of 115 to 180 L of fluid for women and 130 to 200 L of fluid for men is filtered by the human glomeruli each day, less than 1% of the filtered water and sodium chloride (NaCl) and variable amounts of other solutes are excreted in the urine.

By the processes of reabsorption and secretion, the renal tubules regulate the volume and composition of urine.

As a result, the tubules precisely regulate the pH, composition, osmolality, and volume of the intracellular and extracellular fluid compartments.

The reabsorption and secretion of solutes and water in the kidneys are mediated by transport proteins in the cell membranes of the nephron.

Many renal diseases are caused by genetic and acquired defects in transport proteins, which are encoded by approximately 5% to 10% of all human genes.

Furthermore, numerous transport proteins are critical drug targets. This chapter addresses the reabsorption of NaCl and water, the transport of organic anion and cation, the transport proteins that are involved in the transport of solutes and water, and a few of the factors and hormones that regulate NaCl transport.

Details on acid-base transport and on \(\mathrm{K}^{+}\), \(\mathrm{Ca}^{++}\), and inorganic phosphate (P_i) transport and their regulation.

General Principles Of Membrane Transport

Passive mechanisms, active transport mechanisms, or endocytosis may be employed to transport solutes across cell membranes.

In mammals, solute movement is facilitated by both passive and active mechanisms, while all water movement is passive. If the movement of a solute across a membrane occurs spontaneously and does not necessitate the direct expenditure of metabolic energy, it is considered passive.

Uncharged solutes undergo passive transport (diffusion) from an area of elevated concentration to one of lower concentration, i.e., down its chemical concentration gradient.

The passive diffusion of ions (but not uncharged solutes, such as glucose and urea) is influenced by the electrical potential difference (i.e., electrical gradient) across cell membranes and the renal tubules, in addition to concentration gradients).

Cations (for example, \(\mathrm{Na}^{+}\) and \(\mathrm{K}^{+}\)) move to the negative side of the membrane, whereas anions (for example, \(\mathrm{Cl}^{-}\) and bicarbonate \(\left[\mathrm{HCO}_3^{-}\right]\)) move to the positive side of the membrane.

Diffusion of water (osmosis) occurs through aquaporin (AQP) water channels in the cell membrane and is driven by osmotic pressure gradients.

When water is reabsorbed across tubule segments, the solutes dissolved in the water also are carried along with the water. This process is called sol-vent drag and can account for a substantial amount of solute reabsorption across the proximal tubule.

Traditionally, it was thought that the biologically important gases O₂, carbon dioxide (CO₂), and ammonia (NH₃) diffused across the lipid bilayer of plasma membranes.

It is now known that these gases also move across the membrane via specific membrane transport proteins (for example, CO₂ and NH₃ have been shown to cross the membrane via the AQP1 water channel).

In facilitated diffusion, transport depends on the interaction of the solute with a specific protein in the membrane that facilitates its movement across the membrane.

If defined broadly, the term “facilitated diffusion” can be used to describe several different types of membrane transporters. For example, one form of facilitated diffusion is the diffusion of ions, such as \(\mathrm{Na}^{+}\) and \(\mathrm{K}^{+}\), through aqueous-filled channels created by proteins that span the plasma membrane.

Also, the movement of a single molecule across the membrane by a transport protein (uniport), such as occurs with urea and glucose, is a form of facilitated diffusion.

Another form of facilitated diffusion is coupled transport, in which the movement of two or more solutes across a membrane depends on their interaction with a specific transport protein.

Coupled transport of two or more solutes in the same direction is mediated by a symport mechanism. Examples of symport mechanisms in the kidneys include \(\mathrm{Na}^{+}\)-glucose, \(\mathrm{Na}^{+}\)-amino acid, and \(\mathrm{Na}^{+}\)–\(\mathrm{P}_{i}\) symporters in the proximal tubule and the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-2\(\mathrm{Cl}^{-}\)symporter in the thick ascending limb of Henle’s loop.

Coupled transport of two or more solutes in opposite directions is mediated by an antiport mechanism. An \(\mathrm{H}^{+}\)–\(\mathrm{Na}^{+}\) antiporter in the proximal tubule mediates Na+ reabsorption and H+ secretion. With coupled transporters, at least one of the solutes usually is transported against its electrochemical gradient.

• The energy for this uphill movement is derived from the passive downhill movement of at least one of the other solutes into the cell. For example, in the proximal tubule, operation of the \(\mathrm{Na}^{+}\)–\(\mathrm{H}^{+}\) antiporter in the apical membrane of the cell results in the movement of \(\mathrm{H}^{+}\) against its electrochemical gradient out of the cell into the tubular lumen.
• The movement of \(\mathrm{Na}^{+}\) from the tubular lumen into the cell, down its electrochemical gradient, drives the uphill movement of \(\mathrm{H}^{+}\).
• The uphill movement of \(\mathrm{H}^{+}\) is termed secondary active transport to reflect the fact that the movement of \(\mathrm{H}^{+}\) is not directly coupled to the hydrolysis of adenosine triphosphate (ATP).
• Instead, the energy is derived from the gradient of the other coupled ion (in this example, \(\mathrm{Na}^{+}\)).
• Transport is active if it is coupled directly to energy derived from metabolic processes (i.e., it consumes ATP). Active transport of solutes usually takes place from an area of lower concentration to an area of higher concentration.
• In the kidneys, the most prevalent active transport mechanism is sodium-potassium adenosine triphosphatase (\(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase) (or the sodium pump), which is located in the basolateral membrane.

The \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase mechanism is made up of several proteins that together actively move \(\mathrm{Na}^{+}\) out of the cell and \(\mathrm{K}^{+}\) into the cell.

Other active transport mechanisms in the kidneys include the \(\mathrm{H}^{+}\)-ATPase and \(\mathrm{H}^{+}\)–\(\mathrm{K}^{+}\)-ATPase mechanisms, which are responsible for \(\mathrm{H}^{+}\) secretion in the collecting duct, and the \(\mathrm{Ca}^{++}\)-ATPase mechanism, which is responsible for \(\mathrm{Ca}^{++}\) movement from the cell cytoplasm into the blood.

In addition to these transport ATPases, another large group of ATP-dependent transporters exists that is called ATP-binding cassette, or ABC transporters.

To date, seven groups and more than 40 specific ABC transporters have been identified in humans. They transport a diverse group of solutes, including \(\mathrm{Cl}^{-}\), cholesterol, bile acids, drugs, iron, and organic anions and cations.

Endocytosis is the process by which a substance is transported across the plasma membrane. This process involves the invagination of a portion of the membrane until it completely splits off and forms a vesicle in the cytoplasm. This mechanism is crucial for the reabsorption of macromolecules and small proteins by the proximal tubule. Endocytosis is classified as an active transport method due to its dependence on ATP.

General Principles Of Transepithelial Solute And Water Transport

The renal cells are held together by rigid junctions, as demonstrated. The cells are separated by lateral intercellular spaces below the rigid junctions.

The apical membranes are distinguished from the basolateral membranes by the rigid junctions. An epithelium can be likened to a six-pack of soda, in which the cans represent the cells and the plastic receptacle, the tight junctions.

In the nephron, a substance can be reabsorbed or secreted through cells, a process known as the transcellular pathway, or between cells, a process known as the paracellular pathway..

Na⁺ reabsorption by the proximal tubule is a good example of transport by the transcellular pathway. \(\mathrm{Na}^{+}\) reabsorption in this nephron segment depends on the operation of the \(\mathrm{K}^{+}\)–\(\mathrm{Na}^{+}\)– ATPase pump.

The \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump, which is located exclusively in the basolateral membrane, moves \(\mathrm{Na}^{+}\) out of the cell into the blood and \(\mathrm{K}^{+}\) into the cell.

Thus the operation of the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)– ATPase pump lowers intracellular [latex]\mathrm{Na}^{+}[/latex] and increases intracellular [latex]\mathrm{K}^{+}[/latex].

Because intracellular [latex]\mathrm{Na}^{+}[/latex] is low (12 mEq/L) and the [latex]\mathrm{Na}^{+}[/latex] in a tubular fluid is high (140 mEq/L), \(\mathrm{Na}^{+}\) moves across the apical cell membrane, down a chemical concentration gradient from the tubular lumen into the cell.

• Because the interior of the cell is electrically negative with respect to the tubular lumen, and depending on the \(\mathrm{Na}^{+}\) transporter, the energy in this electrical gradient also can drive \(\mathrm{Na}^{+}\) into the cell across the apical membrane.
• The \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)– ATPase pump senses the addition of \(\mathrm{Na}^{+}\) to the cell and is stimulated to increase its rate of \(\mathrm{Na}^{+}\) extrusion into the blood, thereby returning intracellular \(\mathrm{Na}^{+}\) to normal levels.
• Thus transcellular \(\mathrm{Na}^{+}\) reabsorption by the proximal tubule is a two-step process:
• Movement across the apical membrane into the cell, down a chemical concentration gradient and/or an electrochemical gradient established by the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump
• Movement across the basolateral membrane against an electrochemical gradient through the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump
• The reabsorption of \(\mathrm{Ca}^{++}\) and \(\mathrm{K}^{+}\) across the proximal tubule is a good example of paracellular transport.
• Some of the water reabsorbed across the proximal tubule traverses the paracellular pathway. Some solutes dissolved in this water, particularly \(\mathrm{Ca}^{++}\) and \(\mathrm{K}^{+}\), are entrained in the reabsorbed fluid and thereby reabsorbed by the process of solvent drag.

The tight junction in renal epithelial cells is a specialized membrane domain that creates a barrier that regulates the paracellular diffusion of solutes across the epithelia.

Tight junctions are composed of linear arrays of several integral membrane proteins, including occludins, claudins, and several members of the immunoglobulin superfamily.

The tight junction complex of proteins has biophysical properties of ion channels, including the ability to allow ions to diffuse selectively across the complex based on size and charge.

Nacl, Solute, And Water Reabsorption Along The Nephron

Quantitatively, the reabsorption of NaCl and water represents the major function of nephrons. Approximately 25,000 mEq/day of \(\mathrm{Na}^{+}\) and 179 L/day of water are reabsorbed by the renal tubules.

In addition, renal transport of many other important solutes is linked either directly or indirectly to \(\mathrm{Na}^{+}\) reabsorption.

In the following sections the NaCl and water transport processes of each nephron segment and its regulation by hormones, along with other factors, are presented.

Proximal Tubule

The proximal tubule reabsorbs approximately 67% of filtered water, \(\mathrm{Na}^{+}\), \(\mathrm{Cl}^{-}\), \(\mathrm{K}^{+}\), and other solutes.

In addition, the proximal tubule reabsorbs virtually all the glucose and amino acids filtered by the glomerulus. The key element in proximal tubule reabsorption is the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump in the basolateral membrane.

The reabsorption of every substance, including water, is linked in some manner to the operation of the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump.

Na⁺ Reabsorption

Na⁺ is reabsorbed by different mechanisms in the first and the second halves of the proximal tubule.

In the first half of the proximal tubule, \(\mathrm{Na}^{+}\) is reabsorbed primarily with \(\mathrm{HCO}_3^{-}\) and a number of other solutes (for example, glucose, amino acids, \(\mathrm{P}_{i}\), and lactate).

In contrast, in the second half, \(\mathrm{Na}^{+}\) is reabsorbed mainly with \(\mathrm{Cl}^{-}\). This disparity is mediated by differences in the Na+ transport systems in the first and second halves of the proximal tubule and by differences in the composition of tubular fluid at these sites.

In the first half of the proximal tubule, \(\mathrm{Na}^{+}\) uptake into the cell is coupled with either \(\mathrm{H}^{+}\) or organic solutes.

Specific transport proteins mediate Na+ entry into the cell across the apical membrane. For example, the \(\mathrm{Na}^{+}\)–\(\mathrm{H}^{+}\) antiporter couples \(\mathrm{Na}^{+}\) entry with \(\mathrm{H}^{+}\) extrusion from the cell.

Na⁺ secretion results in sodium bicarbonate (\(\mathrm{NaHCO}_3^{-}\)) reabsorption. \(\mathrm{Na}^{+}\) also enters proximal tubule cells by several symporter mechanisms, including \(\mathrm{Na}^{+}\)-glucose, \(\mathrm{Na}^{+}\)-amino acid, \(\mathrm{Na}^{+}\)–\(\mathrm{P}_{i}\), and \(\mathrm{Na}^{+}\)– lactate.

The glucose and other organic solutes that enter the cell with Na+ leave the cell across the basolateral membrane by passive transport mechanisms.

Any \(\mathrm{Na}^{+}\) that enters the cell across the apical membrane leaves the cell and enters the blood by the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase mechanism.

In brief, the reabsorption of \(\mathrm{Na}^{+}\) in the first half of the proximal tubule is coupled to that of \(\mathrm{HCO}_3^{-}, \mathrm{P}_{\mathrm{i}},\) and a number of organic molecules, and this process generates a negative transepithelial voltage across the proximal tubule that provides the driving force for the paracellular reabsorption of \(\mathrm{Cl}^{-}\).

The reabsorption of many organic molecules is so avid that they are almost completely removed from the tubular fluid in the first half of the proximal tubule.

The reabsorption of \(\mathrm{NaHCO}_3 \mathrm{Na}^{+}-\mathrm{P}_{\mathrm{i}} \text {, }\) and \(\mathrm{Na}^{+}\)-organic solutes across the proximal tubule establishes a transtubular osmotic gradient (i.e., the osmolality of the interstitial fluid bathing the basolateral side of the cells is slightly higher than the osmolality of tubule fluid) that provides the driving force for the passive reabsorption of water by osmosis.

Because more water than \(\mathrm{Cl}^{-}\) is reabsorbed in the first half of the proximal tubule, the \(\mathrm{Cl}^{-}\) concentration in tubular fluid rises along the length of the proximal tubule.

In the second half of the proximal tubule, \(\mathrm{Na}^{+}\) is mainly reabsorbed with \(\mathrm{Cl}^{-}\) across the transcellular pathway.

Na⁺ is primarily reabsorbed with \(\mathrm{Cl}^{-}\) rather than organic solutes or \(\mathrm{HCO}_3^{-}\) as the accompanying anion because the \(\mathrm{Na}^{+}\) transport mechanisms in the second half of the proximal tubule differ from those in the first half.

Furthermore, the tubular fluid that enters the second half contains very little glucose and amino acids, but the high concentration of \(\mathrm{Cl}^{-}\) (140 mEq/L) in tubule fluid exceeds that in the first half (105 mEq/L).

The high \(\mathrm{Cl}^{-}\) concentration is due to the preferential reabsorption of \(\mathrm{Na}^{+}\) with \(\mathrm{HCO}_3^{-}\) and organic solutes in the first half of the proximal tubule.

The mechanism of transcellular \(\mathrm{Na}^{+}\) reabsorption in the second half of the proximal tubule is shown. \(\mathrm{Na}^{+}\) enters the cell across the luminal membrane primarily through the parallel operation of a \(\mathrm{Na}^{+}\)–\(\mathrm{H}^{+}\) antiporter and one or more \(\mathrm{Cl}^{-}\)-base anti-porters.

Because the secreted \(\mathrm{H}^{+}\) and base combine in the tubular fluid and reenter the cell, the operation of the \(\mathrm{Na}^{+}\)–\(\mathrm{H}^{+}\) and \(\mathrm{Cl}^{-}\)-base antiporters is equivalent to NaCl uptake from tubular fluid into the cell.

Na⁺ leaves the cell through the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase mechanism, and \(\mathrm{Cl}^{-}\) leaves the cell and enters the blood through a \(\mathrm{K}^{+}\)–\(\mathrm{Cl}^{-}\) symporter and a \(\mathrm{Cl}^{-}\) channel in the basolateral membrane.

Some NaCl also is reabsorbed across the second half of the proximal tubule by a paracellular route. Paracellular NaCl reabsorption occurs because the rise in the \(\mathrm{Cl}^{-}\) concentration in the tubule fluid in the first half of the proximal tubule creates a \(\mathrm{Cl}^{-}\) concentration gradient (140 mEq/L in the tubule lumen and 105 mEq/L in the interstitium).

This concentration gradient favors the diffusion of \(\mathrm{Cl}^{-}\) from the tubular lumen across the tight junctions into the lateral intercellular space. Movement of the negatively charged \(\mathrm{Cl}^{-}\) causes the tubular fluid to become positively charged relative to the blood.

This positive transepithelial voltage causes the diffusion of positively charged \(\mathrm{Na}^{+}\) out of the tubular fluid across the tight junction into the blood.

Thus in the second half of the proximal tubule, some \(\mathrm{Na}^{+}\) and \(\mathrm{Cl}^{-}\) are reabsorbed across the tight junctions (the paracellular pathway) by passive diffusion.

The reabsorption of NaCl establishes a transit-tubular osmotic gradient that provides the driving force for the passive reabsorption of water by osmosis.

In summary, the reabsorption of \(\mathrm{Na}^{+}\) and \(\mathrm{Cl}^{-}\) in the proximal tubule occurs across paracellular and transcellular pathways.

Approximately 67% of the NaCl filtered each day is reabsorbed in the proximal tubule. Of this amount, two-thirds move across the transcellular pathway, and the remaining one-third move across the paracellular pathway.

Water Reabsorption

The proximal tubule reabsorbs 67% of the filtered water. The driving force for water reabsorption is a transtubular osmotic gradient established by solute reabsorption (for example, NaCl and \(\mathrm{Na}^{+}\)-glucose).

The reabsorption of \(\mathrm{Na}^{+}\) along with organic solutes, \(\mathrm{HCO}_3^{-}, \mathrm{P}_{\mathrm{i}} \text {, and } \mathrm{Cl}^{-}\) from the tubular fluid into the lateral intercellular spaces reduces the osmolality of the tubular fluid and increases the osmolality of the lateral intercellular space.

Because the proximal tubule is highly permeable to water, primarily because of the expression of aquaporin water channels (AQP1) in the apical and basolateral membranes, water is reabsorbed across cells by osmosis. However, because the tight junctions in the proximal tubule also are permeable to water, some water is reabsorbed across the paracellular pathway between proximal tubular cells.

The accumulation of fluid and solutes within the lateral intercellular space increases the hydrostatic pressure in this compartment. This increased hydrostatic pressure forces fluid and solutes into the capillaries.

Thus water reabsorption follows solute reabsorption in the proximal tubule. The reabsorbed fluid is slightly hyperosmotic to plasma. However, this difference in osmolality is so small that it is commonly said that proximal tubule reabsorption is isosmotic (i.e., ~67% of both the filtered solute and water are reabsorbed).

Indeed, little difference is seen in the osmolality of tubular fluid at the start and end of the proximal tubule. An important consequence of osmotic water flow across the proximal tubule is that some solutes, especially \(\mathrm{K}^{+}\) and \(\mathrm{Ca}^{++}\), are entrained in the reabsorbed fluid and thereby are reabsorbed by the process of solvent drag.

The reabsorption of virtually all organic solutes, \(\mathrm{HCO}_3^{-}, \mathrm{Cl}^{-}, \mathrm{P}_{\mathrm{i}}\), and other ions, and water is coupled to \(\mathrm{Na}^{+}\) reabsorption.

Therefore changes in Na⁺ reabsorption influence the reabsorption of water and other solutes by the proximal tubule. This point will be discussed later, notably, and is especially relevant during volume depletion when increased \(\mathrm{Na}^{+}\) reabsorption by the proximal tubule is accompanied by a parallel increase in \(\mathrm{HCO}_3^{-}\) reabsorption, which can lead to metabolic alkalosis (i.e., volume contraction alkalosis).

Protein Reabsorption

Proteins filtered across the glomerulus are reabsorbed in the proximal tubule. As mentioned previously, peptide hormones, small proteins, and small amounts of large proteins such as albumin are filtered by the glomerulus.

Overall, only a small percentage of proteins cross the glomerulus and enter Bowman’s space (i.e., the concentration of proteins in the glomerular ultrafiltrate is ~40 mg/L). However, the amount of protein filtered per day is significant because the glomerular filtration rate (GFR) is so high:

Filtered protein = GFR X [Protein] in the ultrafiltrate

Filtered protein = 180 L/day X 40 mg/L (eq 1)

= 7200 mg/day, or 7.2 g/day

Water channels called aquaporins (AQPs) mediate the transcellular reabsorption of water across many nephron segments. In 2003, Dr. Peter Agre received the Nobel Prize in Chemistry for his discovery that AQPs regulate and facilitate water transport across cell membranes, a process essential to all living organisms.

To date, 14 aquaporins have been identified. The AQP family is divided into two groups on the basis of their permeability characteristics. One group (AQPs) is permeable to water (AQP0, AQP1, AQP2, AQP4, AQP5, AQP6, AQP8, AQP11, and AQP12).

The other group (aquaglyceroporins) is permeable to water and small solutes, especially glycerol (AQP3, AQP5, AQP7, AQP9, and AQP10).

AQPs form tetramers in the plasma membrane of cells, with each subunit forming a water channel. In the kidneys, AQP1 is expressed in the apical and basolateral membranes of the proximal tubule and portions of the descending thin limb of the loop of Henle.

The importance of AQP1 in renal water reabsorption is underscored by studies in which the AQP1 gene was “knocked out” in mice. These mice have increased urine output (polyuria) and a reduced ability to concentrate the urine.

In addition, the osmotic water permeability of the proximal tubule is fivefold less in mice lacking APQ1 than in normal mice. AQP7 and AQP8 also are expressed in the proximal tubule.

AQP2 is expressed in the apical plasma membrane of principal cells in the collecting duct, and its expression in the membrane is regulated by AVP. AQP3 and AQP4 are expressed in the basolateral membrane of principal cells in the collecting duct.

• Mice deficient in AQP3 or AQP4 (i.e., knockout mice) have defects in the ability to concentrate urine. AQPs also are expressed in many other organs in the body, including the lung, eye, skin, secretory glands, and brain, where they play key physiological roles.
• For example, AQP4 is expressed in cells that form the blood-brain barrier. Knockout of AQP4 in mice affects the water permeability of the blood-brain barrier such that brain edema is reduced after acute water loading and hyponatremia.
• The endocytosis of protein by the proximal tubule is mediated by apical membrane receptors that specifically bind luminal proteins and peptides. These receptors, called multiligand endocytic receptors, can bind a wide range of peptides and proteins and thereby mediate their endocytosis.
• Megalin and cubilin mediate protein and peptide endocytosis in the proximal tubule. Both are glycoproteins, with megalin being a member of the low-density lipoprotein receptor gene family.
• Filtered proteins are either endocytosed intact or are endocytosed after being partially degraded by enzymes on the surface of the proximal tubule cells.
• Once the proteins and peptides are inside the cell, enzymes digest them into their constituent amino acids, which then leave the cell across the basolateral membrane by transport proteins and are returned to the blood.
• Normally this mechanism reabsorbs virtually all of the proteins filtered, and hence the urine is essentially protein-free. However, because the mechanism is easily saturated, an increase in filtered proteins causes proteinuria (i.e., the appearance of protein in the urine).

Disruption of the glomerular filtration barrier to proteins increases the filtration of proteins and results in proteinuria, which is frequently found in persons with kidney disease.

Urinalysis is an important and routine tool in disease detection. A thorough analysis of the urine includes macroscopic and microscopic assessments. These assessments are performed by visual assessment of the urine, microscopic examination, and chemical evaluation, which is conducted with the use of dipstick reagent strips.

The dipstick test is inexpensive and fast (i.e., it can be performed in less than 5 minutes). Dipstick reagent strips test the urine for the presence of many substances, including bilirubin, blood, glucose, ketones, pH, and protein.

It is normal to find trace amounts of protein in the urine. Trace amounts of protein in the urine can be derived from two sources: (1) filtration and incomplete reabsorption by the prox¬imal tubule and (2) synthesis by the thick ascending limb of the loop of Henle. Cells in the thick ascending limb produce Tamm-Horsfall glycoprotein and secrete it into the tubular fluid.

Because the mechanism for protein reabsorption is “upstream” of the thick ascending limb (i.e., proximal tubule), the secreted Tamm-Horsfall glycoprotein appears in the urine. However, more than trace amounts of protein in the urine are indicative of renal disease.

Organic Anion And Organic Cation Secretion

Cells of the proximal tubule also secrete organic anions and organic cations into the tubule fluid. Secretion of organic anions and cations by the proximal tubule plays a key role in regulating the plasma levels of xenobiotics (for example, a variety of antibiotics, diuretics, statins, antivirals, antineoplastics, immunosuppressants, neurotransmitters, and nonsteroidal antiinflammatory agents) and toxic compounds derived from endogenous and exogenous sources.

Many of the organic anions and cations secreted by the proximal tubule are end products of metabolism that circulate in the plasma. Many of these organic compounds are bound to plasma proteins and are not readily filtered.

Therefore only a small portion of these potentially toxic substances are eliminated from the body by excretion resulting from filtration alone. Such substances also are secreted from the peritubular capillary into the tubular fluid.

These secretory mechanisms are very powerful and can remove virtually all organic anions and cations from the plasma that enters the kidneys. Hence these substances are removed from the plasma by both filtration and secretion.

Thus it is important to note that when kidney function is reduced by disease, the urinary excretion of organic anions and cations is severely reduced, which can lead to increased plasma levels of xenobiotics and metabolites.

• Illustrates the mechanisms of organic anion (\(\mathrm{OA}^{-}\)) transport across the proximal tubule. These secretory pathways have a maximum transport rate and a low specificity (i.e., they transport many organic anions) and are responsible for the secretion of all organic anions listed. \(\mathrm{OA}^{-}\)-s are taken up into the cell, across the basolateral membrane, and against their chemical gradient in exchange for α-ketoglutarate (α-KG) by several \(\mathrm{OA}^{-}\)-a-KG antiport mechanisms, including OAT1, OAT2, and OAT3.
• α-KG accumulates inside the cells by the metabolism of glutamate and by an \(\mathrm{NA}^{+}\)-α-KG symporter (i.e., the \(\mathrm{NA}^{+}\)-dicarboxylate transporter [NaDC3]) also present in the basolateral membrane.
• Thus OA- uptake into the cell against an electrochemical gradient is coupled to the exit of α-KG out of the cell, down its chemical gradient generated by the \(\mathrm{OA}^{-}\)-α-KG antiport mechanism.
• The exit of OA- across the luminal membrane into the tubular fluid is mediated by the multidrug resistance proteins 2 and 4 (MRP2/4) and breast cancer resistance protein 1 (BCRP), which require ATP for their operation (i.e., they are ABC transporters).
• Recent studies reveal that OAT4 mediates the reabsorption of urate, the end product of purine catabolism, and the secretion of several drugs.
• Because many organic anions compete for the same secretory pathways, elevated plasma levels of one anion often inhibit the secretion of the others.
• For example, infusing para-amino hippurate (PAH) can reduce penicillin secretion by the proximal tubule. Because the kidneys are responsible for eliminating penicillin, the infusion of PAH into persons who receive penicillin reduces penicillin excretion and thereby extends the biological half-life of the drug.

In World War II, when penicillin was in short supply, hippurates were given with penicillin to extend the drug’s therapeutic effect. Similar competition for secretion by the proximal tubule occurs for organic cations.

For example, elevated plasma levels of one cation often inhibit the secretion of the others. The H₂ antagonist cimetidine is used to treat gastric ulcers, and organic cation transport mecha¬nisms in the proximal tubule secrete cimetidine.

When cimetidine is given to patients who also receive procainamide (a drug used to treat cardiac arrhythmias), cimetidine reduces the urinary excretion of procainamide (also an organic cation) by competing with this antiarrhythmic drug for the secretory pathway.

Thus the coadministration of organic cations can increase the plasma concentrations of both drugs to levels much higher than those seen when the drugs are given alone. This effect can lead to drug toxicity.

Illustrates the mechanism of organic cation (\(\mathrm{OC}^{+}\)) transport across the proximal tubule. \(\mathrm{OC}^{+}\)s, including xenobiotics such as the antidiabetic agent metformin, the antiviral agent lamivudine, and the anticancer drug oxaliplatin, along with many important monoamine neurotransmitters, including dopamine, epinephrine, histamine, and norepinephrine, are secreted by the proximal tubule.

OC⁺ s are taken up into the cell, across the basolateral membrane, primarily by the \(\mathrm{OC}^{+}\) transporter 2 (OCT2). The uptake of organic cations is driven by the magnitude of the cell-negative potential difference across the basolateral membrane.

Organic cation transport across the luminal membrane into the tubular fluid, which is the rate-limiting step in secretion, is mediated primarily by the electrically neutral multidrug and toxin extrusion transporters (MATEs) and the ABC transporter MDR1 (also known as P-glycoprotein).

These transport mechanisms mediating OC⁺ secretion are nonspecific; several OC⁺s usually compete for each transport pathway.

Henle’s Loop

Henle’s loop reabsorbs approximately 25% of the filtered NaCl and 15% of the filtered water. The reabsorption of NaCl in the loop of Henle occurs in both the thin ascending and thick ascending limbs.

The descending thin limb does not reabsorb NaCl. Water reabsorption occurs exclusively in some portions of the descending thin limb through AQP1 water channels. The ascending limb is impermeable to water. In addition, \(\mathrm{Ca}^{++}\) and \(\mathrm{HCO}_3^{-}\) are reabsorbed in the loop of Henle

The thin ascending limb reabsorbs NaCl by a passive mechanism. The reabsorption of water but not NaCl in the descending thin limb increases the [NaCl] in tubule fluid entering the ascending thin limb.

As the NaCl-rich fluid moves toward the cortex, NaCl diffuses out of tubule fluid across the ascending thin limb into the medullary interstitial fluid, down a concentration gradient directed from tubule fluid to interstitium.

The key element in solute reabsorption by the thick ascending limb is the \(\mathrm{NA}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump in the basolateral membrane.

As with reabsorption in the proximal tubule, the reabsorption of every solute by the thick ascending limb is linked to \(\mathrm{NA}^{+}\)–\(\mathrm{NA}^{+}\)– ATPase. This transporter maintains a low intracellular [latex]\mathrm{NA}^{+}[/latex], which provides a favorable chemical gradient for the movement of Na+ from the tubular fluid into the cell.

The movement of Na+ across the apical membrane into the cell is mediated by the \(\mathrm{NA}^{+}\)–\(\mathrm{K}^{+}\)-2\(\mathrm{Cl}^{-}\) symporter (NKCC2), which couples the movement of \(\mathrm{NA}^{+}\) with \(\mathrm{K}^{+}\) and 2\(\mathrm{Cl}^{-}\).

Using the potential energy released by the downhill movement of \(\mathrm{NA}^{+}\) and \(\mathrm{Cl}^{-}\), this symporter drives the uphill movement of \(\mathrm{K}^{+}\) into the cell.

The \(\mathrm{K}^{+}\) channel in the apical plasma mem¬brane plays an important role in NaCl reabsorption by the thick ascending limb.

This \(\mathrm{K}^{+}\) channel allows the \(\mathrm{K}^{+}\) transported into the cell by NKCC2 to recycle back into tubule fluid. Because the [latex]\mathrm{K}^{+}[/latex] in tubule fluid is relatively low, \(\mathrm{K}^{+}\) recycling is required for the continued operation of NKCC2.

An \(\mathrm{NA}^{+}\)–\(\mathrm{H}^{+}\) antiporter in the apical cell membrane also mediates \(\mathrm{NA}^{+}\) reabsorption as well as \(\mathrm{H}^{+}\) secretion (\(\mathrm{HCO}_3^{-}\) reabsorption) in the thick ascending limb.

Na leaves the cell across the basolateral membrane through the \(\mathrm{NA}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump, whereas \(\mathrm{K}^{+}\), \(\mathrm{Cl}^{-}\), and \(\mathrm{HCO}_3^{-}\) leave the cell across the basolateral membrane by sepa¬rate pathways.

The voltage across the thick ascending limb is important for the reabsorption of several cations. The tubular fluid is positively charged relative to blood because of the unique location of transport proteins in the apical and basolateral membranes. Two points are important:

Increased NaCl transport by the thick ascending limb increases the magnitude of the positive voltage in the lumen, and

This voltage is an important driving force for the reabsorption of several cations, including \(\mathrm{NA}^{+}\), \(\mathrm{K}^{+}\), \(\mathrm{Mg}^{++}\), and \(\mathrm{Ca}^{++}\) across the paracellular pathway.

The importance of the paracellular pathway to solute reabsorption is underscored by the observation that inactivating mutations of the tight junction protein claudin-16 reduces \(\mathrm{Mg}^{++}\) and \(\mathrm{Ca}^{++}\) reabsorption by the ascending thick limb even when the lumen-positive transepithelial voltage is positive.

In summary, salt reabsorption across the thick ascending limb occurs by the transcellular and paracellular pathways. A total of 50% of NaCl reabsorption is transcellular, and 50% is paracellular.

The thick ascending limb does not reabsorb water because it does not express water channels (i.e., AQPs), and thus the reabsorption of NaCl and other solutes reduces the osmolality of tubular fluid to less than 150 mOsm/kg H₂O.

Thus because the thick ascending limb produces a fluid that is dilute relative to plasma, the ascending limb of the Henle loop is called the diluting segment.

Epithelial cells are joined at their apical surfaces by tight junctions (also known as zonula occludens). A number of proteins now have been identified as components of the tight junction.

These proteins include those that span the membrane of one cell and link to the extracellular portion of the same molecule in the adjacent cell (for example, occludins and claudins), as well as cytoplasmic linker proteins (for example, ZO-1, ZO-2, and ZO-3) that link the mem¬branespanning proteins to the cytoskeleton of the cell.

Of these junctional proteins, claudins appear to be important in determining the permeability characteristics of the tight junction. Claudin-16 and claudin-1 9 are critical for determining the divalent cation permeability of the tight junctions in the thick ascending limb of the Henle loop.

Mutations in human claudin-16 and claudin-19 cause familial hypomagnesemia with hypercalciuria and nephrolithiasis. Claudin-2 is permeable to water and may allow paracellular water reabsorption across the proximal tubule.

In cultured kidney cells, claudin-4 has been shown to control the \(\mathrm{NA}^{+}\) permeability of the tight junction, and claudin-15 determines whether a tight junction is permeable to cations or anions.

Thus the permeability characteristics of the tight junctions in different nephron segments are determined, at least in part, by the specific claudins expressed by the cells in that segment.

Bartter syndrome is a set of autosomal recessive genetic diseases characterized by hypokalemia, metabolic alkalosis, and hyperaldosteronism.

Inactivating mutations in the gene coding for the \(\mathrm{NA}^{+}\)–\(\mathrm{K}^{+}\)-2\(\mathrm{Cl}^{-}\) symporter (SLC12A1), the apical K+ channel (KCNJ1), or the basolateral \(\mathrm{Cl}^{-}\) channel (CICNKB) decrease both sodium chloride (NaCl) and \(\mathrm{K}^{+}\) reabsorption by the ascending thick limb, which in turn causes hypokalemia (i.e., a low plasma [latex]\mathrm{K}^{+}[/latex]) and a decrease in the extracellular fluid (ECF) volume.

The decrease in ECF volume stimulates aldosterone secretion, which in turn stimulates NaCl reabsorption and \(\mathrm{H}^{+}\) secretion by the distal tubule and collecting duct.

Distal Tubule And Collecting Duct

The distal tubule and collecting duct reabsorb approximately 8% of the filtered NaCl, secrete variable amounts of \(\mathrm{K}^{+}\) and \(\mathrm{H}^{+}\), and reabsorb a variable amount of water (~8% to 17%).

The initial segment of the distal tubule (the early distal tubule) reabsorbs \(\mathrm{NA}^{+}\), \(\mathrm{Cl}^{-}\), and \(\mathrm{Ca}^{++}\) and is impermeable to water.

NaCl entry into the cell across the apical membrane is mediated by a \(\mathrm{NA}^{+}\)–\(\mathrm{Cl}^{-}\) symporter (NCC). \(\mathrm{NA}^{+}\) leaves the cell through the action of \(\mathrm{NA}^{+}\)–\(\mathrm{K}^{+}\)-ATPase, and

Gitelman syndrome is an autosomal recessive disorder characterized by metabolic alkalosis, hypokalemia, and hypocalciuria (hypomagnesemia also is seen often).

• It results from an inactivating mutation of the \(\mathrm{NA}^{+}\)–\(\mathrm{Cl}^{-}\) symporter gene (SIC12A3) that is expressed in the early portion of the distal tubule.
• The fluid and electrolyte disturbances seen in patients with Gitelman syndrome can be mimicked by the administration of thiazide diuretics, which act by inhibiting the sodium chloride transporter.
• Cl^– leaves the cell by diffusion through \(\mathrm{Cl}^{-}\) channels. NaCl reabsorption is reduced by thiazide diuretics, which inhibit NCC. Thus dilution of the tubular fluid begins in the thick ascending limb and continues in the early segment of the distal tubule.
• The last segment of the distal tubule (late distal tubule) and the collecting duct are composed of two cell types: principal cells and intercalated cells.
• As illustrated in A, principal cells reabsorb \(\mathrm{NA}^{+}\) and water and secrete \(\mathrm{K}^{+}\).
• The a-intercalated cell secretes \(\mathrm{H}^{+}\) and reabsorbs \(\mathrm{HCO}_3^{-}\) and \(\mathrm{K}^{+}\) and thus is important in regulating acid-base balance and \(\mathrm{K}^{+}\) balance.
• Intercalated cells secrete \(\mathrm{HCO}_3^{-}\) and reabsorb \(\mathrm{H}^{+}\) and \(\mathrm{Cl}^{-}\). β-Intercalated cells also reabsorb \(\mathrm{K}^{+}\) by the operation of an \(\mathrm{H}^{+}\)–\(\mathrm{K}^{+}\)-ATPase mechanism located in the apical plasma membrane.
• Both \(\mathrm{Na}^{+}\) reabsorption and \(\mathrm{K}^{+}\) secretion by principal cells depend on the activity of \(\mathrm{K}^{+}\)–\(\mathrm{H}^{+}\)-ATPase in the basolateral membrane. By maintaining a low intracellular [latex]\mathrm{Na}^{+}[/latex], this transporter provides a favorable chemical gradient for the movement of \(\mathrm{Na}^{+}\) from the tubular fluid into the cell.
• Because \(\mathrm{Na}^{+}\) enters the cell across the apical membrane by diffusion through \(\mathrm{Na}^{+}\)-selective channels in the apical membrane, the negative voltage inside the cell facilitates \(\mathrm{Na}^{+}\) entry.

Na⁺ leaves the cell across the basolateral membrane and enters the blood through the action of \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase.

Na⁺ reabsorption generates a lumen negative voltage across the late distal tubule and collecting duct, which provides the driving force for \(\mathrm{Cl}^{-}\) reabsorption across the paracellular pathway.

As previously noted, \(\mathrm{Cl}^{-}\) also is reabsorbed by p-intercalated cells. Chloride enters the p-intercalated cell across the apical membrane via a \(\mathrm{Cl}^{-}\)/\(\mathrm{HCO}_3^{-}\) antiporter (Pendrin) and leaves the cell across the basolateral membrane via a \(\mathrm{Cl}^{-}\) channel.

A variable amount of water is reabsorbed across principal cells in the late distal tubule and collecting duct. Water reabsorption is mediated by the AQP2 water channel located in the apical plasma membrane and AQP3 and AQP4 water channels located in the basolateral membrane of principal cells.

In the presence of arginine vasopressin (AVP), water is reabsorbed. By contrast, in the absence of AVP, the distal tubule and collecting duct reabsorb little water.

K⁺ is secreted from the blood into the tubular fluid by principal cells in two steps. First, \(\mathrm{K}^{+}\) uptake across the basolateral membrane is mediated by the action of the \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump.

Second, \(\mathrm{K}^{+}\) leaves the cell by passive diffusion. Because the \(\mathrm{K}^{+}\) concentration inside the cells is high (~ 150 mEq/L) and the \(\mathrm{K}^{+}\) concentration in tubular fluid is low (~10 mEq/L), \(\mathrm{K}^{+}\) diffuses down its concentration gradient through apical cell membrane \(\mathrm{K}^{+}\) channels into the tubular fluid.

Although the negative potential inside the cells tends to retain \(\mathrm{K}^{+}\) within the cell, the electrochemical gradient across the apical membrane favors \(\mathrm{K}^{+}\) secretion from the cell into the tubular fluid.

K⁺ reabsorption by a-intercalated cells is mediated by an \(\mathrm{H}^{+}\)–\(\mathrm{K}^{+}\)-ATPase pump located in the apical cell membrane.

Regulation Of Nacl And Water Reabsorption

Quantitatively, angiotensin II, aldosterone, catecholamines, natriuretic peptides, and uroguanylin are the most important hormones that regulate NaCl reabsorption and thereby urinary NaCl excretion.

However, other hormones (including dopamine and adrenomedullin), Starling forces, and the phenomenon of glomerulotubular balance influence NaCl reabsorption. AVP is the only major hormone that directly regulates the amount of water excreted by the kidneys.

Angiotensin II has a potent stimulatory effect on NaCl and water reabsorption in the proximal tubule. It also has been shown to stimulate \(\mathrm{Na}^{+}\) reabsorption in the thick ascending limb of the Henle loop, as well as the distal tubule and collecting duct.

Angiotensin II is one of the most potent hormones that stimulates NaCl and water reabsorption in the proximal tubule. A decrease in the extracellular fluid (ECF) volume activates the renin-angiotensin-aldosterone system, thereby increasing the plasma concentration of angiotensin II.

Aldosterone, which is synthesized by the glomerulosa cells of the adrenal cortex, stimulates NaCl reabsorption. It acts on the thick ascending limb of the loop of Henle, the late segment of the distal tubule, and the collecting duct.

The late distal tubule and collecting duct are collectively termed the aldosterone-sensitive distal nephron (ASDN). Aldosterone also stimulates \(\mathrm{K}^{+}\) secretion by the late segment of the distal tubule and collecting duct.

Aldosterone enhances NaCl reabsorption across principal cells in the ASDN by four mechanisms:

Increasing the amount of \(\mathrm{Na}^{+}\)– \(\mathrm{K}^{+}\)-ATPase in the basolateral membrane;

Increasing the expression of the epithelial sodium channel (ENaC) in the apical cell membrane;

Elevating serum glucocorticoid-stimulated kinase (Sgk) levels, which also increases the expression of ENaC in the apical cell membrane; and

Stimulating channel-activating protease (CAP1, also called prostatin), a serine protease, directly activates ENaC channels by proteolysis.

Taken together, these actions increase the uptake of \(\mathrm{Na}^{+}\) across the apical cell membrane and facilitate the exit of \(\mathrm{Na}^{+}\) from the cell interior into the blood.

The increase in the reabsorption of \(\mathrm{Na}^{+}\) generates a lumen-negative transepithelial voltage across the late segment of the distal tubule and collecting duct.

This lumen-negative voltage provides the electrochemical driving force for \(\mathrm{Cl}^{-}\) reabsorption across the tight junctions (i.e., paracellular pathway) in the ASDN.

Aldosterone secretion is enhanced by hyperkalemia and by volume contraction (i.e., reduced ECF volume) via increased angiotensin II (after activation of the renin-angiotensin system).

Aldosterone secretion is decreased by hypokalemia and natriuretic peptides (discussed later in this chapter). Through its stimulation of NaCl reabsorption in the collecting duct, aldosterone also indirectly increases water reabsorption by this nephron segment.

Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) inhibit NaCl and water reabsorption. Secretion of ANP by the cardiac atria and BNP by the cardiac ventricles is stimulated by a rise in blood pressure and an increase in the ECF volume.

ANP and BNP reduce blood pressure by decreasing the total peripheral resistance and by enhancing urinary NaCl and water excretion, primarily by increasing GFR and renal blood flow (RBF).

These natriuretic peptides vasodilate the afferent and efferent arterioles, which increases GFR and thus the filtration of NaCl, thereby increasing NaCl excretion.

In addition, the increase in RBF decreases the concentration of NaCl in the medullary interstitium, which in turn reduces passive NaCl reabsorption by the thin ascending limb of the loop of Henle.

ANP and BNP also inhibit NaCl reabsorption by the medullary portion of the collecting duct and inhibit AVP-stimulated water reabsorption across the collecting duct.

Moreover, ANP and BNP also reduce the secretion of AVP from the posterior pituitary. These actions of ANP and BNP are mediated by activation of membrane-bound guanylyl cyclase receptors, which increase intracellular levels of the second messenger cyclic guanosine monophosphate (cGMP). ANP induces a more profound natriuresis and diuresis than does BNP.

Urodilatin and ANP are encoded by the same gene and have similar amino acid sequences. Urodi-latin is a 32-amino-acid hormone that differs from ANP by the addition of four amino acids to the amino terminus.

Urodilatin is secreted by the distal tubule and collecting duct and is not present in the systemic circulation; thus urodilatin influences only the function of the kidneys.

Urodilatin secretion is stimulated by a rise in blood pressure and an increase in the ECF volume. It inhibits NaCl and water reabsorption across the medullary portion of the collecting duct.

Urodilatin is a more potent natriuretic and diuretic hormone than ANP because some of the ANP that enters the kidneys in the blood is degraded by a neutral endopeptidase that has no effect on urodilatin.

Uroguanylin and guanylin are produced by neuroendocrine cells in the intestine in response to the oral ingestion of NaCl. These hormones enter the circulation and inhibit NaCl and water reabsorption by the kidneys by activation of membrane-bound guanylyl cyclase receptors, which increases intracellular levels of cGMP.

The natriuretic response of the kidneys to a salt load is more pronounced when given orally than when delivered intravenously because oral administration of salt causes the intestinal secretion of uroguanylin and guanylin.

Catecholamines stimulate NaCl reabsorption. Catecholamines released from the sympathetic nerves (norepinephrine) and the adrenal medulla (epinephrine) stimulate NaCl and water reabsorption by the proximal tubule, the thick ascending limb of the loop of Henle, the distal tubule, and the collecting duct.

Although sympathetic nerves are not very active when the ECF volume is normal, when ECF volume declines (for example, after hemorrhage), sympathetic nerve activity rises dramatically and stimulates NaCl and water reabsorption by these four nephron segments.

Dopamine, a catecholamine, is synthesized by cells of the proximal tubule. The action of dopamine is opposite to that of norepinephrine and epinephrine.

Dopamine secretion is stimulated by an increase in ECF volume, and its secretion directly inhibits NaCl and water reabsorption in the proximal tubule.

Adrenomedullin is a 52-amino-acid peptide hormone that is produced by a variety of organs including the kidneys. Adrenomedullin induces marked diuresis and natriuresis, and its secretion is stimulated by congestive heart failure and hypertension.

The major effect of adrenomedullin on the kidneys is to increase GFR and RBF, thereby indirectly stimulating the excretion of NaCl and water (see the previous discussion regarding ANP/BNP).

AVP regulates water reabsorption and is the most important hormone that regulates water reabsorption in the kidneys. This hormone is secreted by the posterior pituitary gland in response to an increase in plasma osmolality (1% or more) or a decrease in the ECF volume (>5% to 10% of normal).

AVP increases the permeability of the collecting duct to water and thereby increases osmotic water reabsorption across the collecting duct because of the osmotic gradient that exists between the tubular fluid and the interstitium. AVP has little effect on urinary NaCl excretion.

Starling forces regulate NaCl and water reabsorption across the proximal tubule. As previously described, \(\mathrm{Na}^{+}, \mathrm{Cl}^{-}, \mathrm{HCO}_3^{-}\) amino acids, glucose, and water are transported into the intercellular space of the proximal tubule.

Starling forces between this space and the peritubular capillaries facilitate the movement of the reabsorbed fluid into the capillaries.

Starling forces across the wall of the peritubular capillaries are the hydrostatic pressures in the peritubular capillary (\(\mathrm{P}_{pc}\)) and lateral intercellular space (\(\mathrm{P}_{i}\)) and the oncotic pressures in the peritubular capillary (\(\pi_{pc}\)) and lateral intercellular space (\(\pi_{i}\)).

Thus the reabsorption of water resulting from \(\mathrm{Na}^{+}\) transport from tubular fluid into the lateral intercellular space is modified by the Star-ling forces.

Thus: \(\mathrm{Q}=\mathrm{K}_{\mathrm{f}}\left[\left(\mathrm{P}_{\mathrm{Pc}}-\mathrm{P}_{\mathrm{i}}\right)+\sigma\left(\pi_{\mathrm{Pc}}-\pi_{\mathrm{i}}\right)\right]\) (eq 2)

where Q is flow (positive numbers indicate flow from the intercellular space into the blood). Starling forces that favor movement from the interstitium into the peritubular capillaries are \(\pi_{\mathrm{pc}} \text { and } \mathrm{P}_{\mathrm{i}}\).

The opposing Starling forces are \(\pi_{\mathrm{i}} \text { and } \mathrm{P}_{\mathrm{Pc}}\). Normally the sum of the Starling forces favors the movement of solute and water from the interstitial space into the capillary.

• However, some of the solutes and fluid that enter the lateral intercellular space leak back into the proximal tubular fluid. Starling forces do not affect transport by the loop of Henle, distal tubule, and collecting duct because these segments are less permeable to water than the proximal tubule.
• A number of factors can alter the Starling forces across the peritubular capillaries surrounding the proximal tubule. For example, dilation of the efferent arteriole increases \(P_{pc}\), whereas constriction of the efferent arteriole decreases it. An increase in \(P_{pc}\) inhibits solute and water reabsorption by increasing the back-leak of NaCl and water across the tight junction, whereas a decrease stimulates reabsorption by decreasing back-leak across the tight junction.
• The \(\pi_{pc}\) is partially determined by the rate of formation of the glomerular ultrafiltrate. For example, if one assumes a constant plasma flow in the afferent arteriole, the plasma proteins become less concentrated in the plasma that enters the efferent arteriole and peritubular capillary because less ultrafiltrate is formed (i.e., as GFR decreases). Hence the npc decreases.
• The \(\pi_{pc}\) is directly related to the filtration fraction (FF) (FF = GFR/RPF). A decrease in the FF resulting from a decrease in GFR, at constant RPF, decreases the \(\pi_{pc}\).
• This phenomenon in turn increases the backflow of NaCl and water from the lateral intercellular space into the tubular fluid and thereby decreases net solute and water reabsorption across the proximal tubule. An increase in the FF has the opposite effect.
• The importance of Starling forces in regulating solute and water reabsorption by the proximal tubule is underscored by the phenomenon of glomerulotubu-lar (G-T) balance. Spontaneous changes in GFR markedly alter the filtered load of \(\mathrm{Na}^{+}\) (filtered load = GFR x \(\mathrm{Na}^{+}\)).
• Without rapid adjustments in \(\mathrm{Na}^{+}\) reabsorption to counter the changes in the filtration of \(\mathrm{Na}^{+}\), urine \(\mathrm{Na}^{+}\) excretion would fluctuate widely, disturb the N\(\mathrm{Na}^{+}\) balance of the body, and thus alter ECF volume and blood pressure.
• However, spontaneous changes in GFR do not alter \(\mathrm{Na}^{+}\) excretion in the urine or \(\mathrm{Na}^{+}\) balance when ECF volume is normal because of the phenomenon of G-T balance.

When body \(\mathrm{Na}^{+}\) balance is normal (i.e., ECF volume is normal), G-T balance refers to the fact that \(\mathrm{Na}^{+}\) and water reabsorption increases in proportion to the increase in GFR and filtered load of \(\mathrm{Na}^{+}\).

Thus a constant fraction of the filtered \(\mathrm{Na}^{+}\) and water is reabsorbed from the proximal tubule despite variations in GFR. The net result of G-T balance is to reduce the impact of GFR changes on the amount of \(\mathrm{Na}^{+}\) and water excreted in the urine.

Two mechanisms are responsible for G-T balance. One is related to the oncotic and hydrostatic pressure differences between the peritubular capillaries and the lateral intercellular space (i.e., Starling forces).

For example, an increase in the GFR (at constant renal blood flow) raises the protein concentration in the glomerular capillary plasma above normal. This protein-rich plasma leaves the glomerular capillaries, flows through the efferent arterioles, and enters the peritubular capillaries.

The increased NPC augments the movement of solute and fluid from the lateral intercellular space into the peritubular capillaries. This action increases net solute and water reabsorption by the proximal tubule.

The second mechanism responsible for G-T balance is initiated by an increase in the filtered load of glucose and amino acids. As discussed earlier, the reabsorption of Na⁺ in the first half of the proximal tubule is coupled to that of glucose and amino acids. The rate of

Na⁺ reabsorption therefore partially depends on the filtered load of glucose and amino acids. As the GFR and filtered load of glucose and amino acids increase, Na⁺ and water reabsorption also rise.

In addition to G-T balance, another mechanism minimizes changes in the filtered load of \(\mathrm{Na}^{+}\). An increase in the GFR (and thus in the amount of \(\mathrm{Na}^{+}\) filtered by the glomerulus) activates the tubuloglomerular feedback mechanism.

This action returns the GFR and filtration of Na⁺ to normal values. Thus spontaneous changes in the GFR (for example, caused by changes in posture and blood pressure) increase the amount of \(\mathrm{Na}^{+}\) filtered for only a few minutes.

The mechanisms that underlie G-T balance maintain urinary \(\mathrm{Na}^{+}\) excretion constant and thereby maintain \(\mathrm{Na}^{+}\) homeostasis (and ECF volume and blood pressure) until the GFR returns to normal.

As previously noted, aldosterone stimulates both sodium chloride (NaCl) reabsorption and K⁺ secretion by the collecting duct. Although both a reduction in the extracellular fluid (ECF) volume and hyperkalemia increase aldosterone levels, the physiologic response of the kidneys with regard to NaCl and \(\mathrm{K}^{+}\) excretion differs in these two conditions.

During reduction in the ECF volume, NaCl excretion by the kidneys is reduced, to restore ECF volume, without a change in \(\mathrm{K}^{+}\) excretion. By contrast, during hyperkalemia, \(\mathrm{K}^{+}\) excretion by the kidneys is increased, to return plasma [latex]\mathrm{K}^{+}[/latex] to normal, without a change in NaCl excretion.

This phenomenon, the apparent independent effects of aldosterone on net urinary \(\mathrm{Na}^{+}\) and \(\mathrm{K}^{+}\) excretion, is called the aldosterone paradox.

The paradox can be explained by the observation that although aldosterone increases in both conditions, angiotensin II levels increase only during ECF volume contraction and not during hyperkalemia, and the fact that aldosterone and angiotensin II differentially regulate a number of transport proteins in several nephron segments.

The integrated physiological response to a reduction in the ECF volume is depicted, A. During volume contraction, angiotensin II stimulates NaCl reabsorption by the proximal tubule and by the distal tubule (early segment) by activating with no lysine (K) kinase (WNK), which enhances NaCl reabsorption by activating NCC.

• Aldosterone stimulates Na⁺ reabsorption in principal cells of the collecting duct by activating SGK1, which increases the abundance of epithelial sodium channel (ENaC) channels in the apical plasma membrane.
• Both effects stimulate NaCl reabsorption. Moreover, angiotensin II activates WNK in principal cells, which inhibits \(\mathrm{K}^{+}\) secretion via the renal outer medullary \(\mathrm{K}^{+}\) (ROMK) channel, thereby preventing an increase in \(\mathrm{K}^{+}\) excretion even though aldosterone levels are elevated.
• The integrated physiological response to hyperkalemia is depicted, in B. During hyperkalemia, aldosterone stimulates ROMK-mediated \(\mathrm{K}^{+}\) secretion by principal cells in the collecting duct by activating WNK (a different WNK than the one that inhibits ROMK), thereby increasing \(\mathrm{K}^{+}\) excretion by the kidneys.
• Because the distal tubule (early segment) is not responsive to aldosterone, this hormone does not stimulate NaCl reabsorption in this nephron segment. In fact, because angiotensin II levels are not elevated during hyperkalemia, the basal activity of WNK is low, resulting in reduced NaCl reabsorption via NCC.
• The lack of effect of WNK on NaCl reabsorption in the distal tubule offsets the stimulatory effect of aldosterone on NaCl reabsorption in principal cells of the collecting duct, resulting in no net change in urinary NaCl excretion during hyperkalemia.
• Serum glucocorticoid-stimulated kinase (Sgk), a serine/threonine kinase, plays an important role in maintaining sodium chloride (NaCl) and \(\mathrm{K}^{+}\) homeostasis by regulating NaCl and K⁺ excretion by the kidneys.
• Studies in Sgk1 knockout mice reveal that this kinase is required for animals to survive severe NaCl restriction and \(\mathrm{K}^{+}\) loading. NaCl restriction and \(\mathrm{K}^{+}\) loading enhance plasma (aldosterone), which rapidly (in minutes) increases Sgk1 protein expression and phosphorylation.

Phosphorylated Sgk1 enhances epithelial sodium channel (ENaC)-mediated sodium reabsorption in the collecting duct, primarily by increasing the number of ENaC channels in the apical plasma membrane of principal cells and also by increasing the number of \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\) ATPase pumps in the basolateral membrane.

Phosphorylated Sgk1 inhibits Nedd4-2, a ubiquitin ligase, which monoubiquitinylates ENaC subunits, targeting them for endocytic removal from the plasma membrane and subsequent destruction in lysosomes. Inhibition of Nedd4-2 by Sgk1 reduces the monoubiquitinylation of ENaC, thereby reducing endocytosis and increasing the number of channels in the membrane.

Sgk1 induces a translocation of ROMK from an intracellular pool to the plasma membrane and thereby enhances ROMK-mediated K⁺ secretion by principal cells.

These effects of Sgk1 precede the aldosterone-stimulated increase in ENaC, ROMK, and \(\mathrm{Na}^{+}\)–\(\mathrm{K}^{+}\) ATPase abundance, which leads to a delayed (>4 hours), secondary increase in NaCl and \(\mathrm{K}^{+}\) transport by the collecting duct.

Activating polymorphisms in Sgk1 cause an increase in blood pressure, presumably by enhanced NaCl reabsorption by the collecting duct, which increases the extracellular fluid volume and thereby blood pressure.

Liddle syndrome is a rare genetic disorder characterized by an increase in the extracellular fluid (ECF) volume that causes an increase in blood pressure (i.e., hypertension).

Liddle syndrome is caused by activating mutations in either the β or γ subunit of the epithelial Na+ channel gene (ENaC), which is composed of three subunits, a, |3, and y. These mutations increase the number of Na+ channels in the apical cell membrane of principal cells and thereby the amount of Na⁺ reabsorbed by each channel.

In persons with Liddle syndrome, the rate of renal Na⁺ reabsorption is inappropriately high, which leads to an increase in the ECF volume and hypertension.

• Two different forms of pseudohypoaldosteronism type 1 (PHA1) exist (i.e., the kidneys waste sodium chloride [NaCl] as they do when aldosterone levels are reduced; however, in PHA1, aldosterone levels are elevated).
• The autosomal recessive form is caused by inactivating mutations in the α, β, or γ subunit of ENaC. The etiology of the autosomal dominant form is an inactivating mutation in the mineralocorticoid receptor. PHA is characterized by an increase in \(\mathrm{Na}^{+}\) excretion, a reduction in the ECF volume, hyperkalemia, and hypotension.
• Some persons with expanded ECF volume and elevated blood pressure are treated with drugs that inhibit angiotensin-converting enzyme (ACE) inhibitors (for example, captopril, enalapril, and lisinopril) and thereby lower fluid volume and blood pressure.
• The inhibition of ACE blocks the degradation of angiotensin I to angiotensin II and thereby lowers plasma angiotensin II levels. The decline in plasma angiotensin II concentration has three effects.
• First, NaCl and water reabsorption by the nephron (especially the proximal tubule) decreases. Second, aldosterone secretion decreases, thus reducing NaCl reabsorption in the thick ascending limb, distal tubule, and collecting duct.

Third, because angiotensin is a potent vasoconstrictor, a reduction in its concentration permits the systemic arterioles to dilate and thereby lower arterial blood pressure. ACE also degrades the vasodilator hormone bradykinin; thus ACE inhibitors increase the concentration of bradykinin, a vasodilatory hormone.

ACE inhibitors decrease the extracellular fluid volume and arterial blood pressure by promoting renal NaCl and water excretion and by reducing total peripheral vascular resistance.