Chapter 25: Regulation of Renal Function and Vascular Volume

What engineer, wishing to regulate the composition of the internal environment of the body on which the function of every bone, gland, muscle, and nerve depends, would devise a scheme that operated by throwing the whole thing out 16 times a day and rely on grabbing from it, as it fell to earth, only those precious elements which he wanted to keep?

   Homer Smith, From Fish to Philosopher

The basic urine-forming unit of the kidney is the nephron, which consists of a filtering apparatus, the glomerulus, connected to a long tubular portion that reabsorbs and conditions the glomerular ultrafiltrate. Each human kidney is composed of ~1 million nephrons. The nomenclature for segments of the tubular portion of the nephron has become increasingly complex as renal physiologists have subdivided the nephron into shorter and shorter named segments. These subdivisions were based initially on the axial location of the segments but increasingly have been based on the morphology of the epithelial cells lining the various nephron segments. Figure 25–1 illustrates subdivisions of the nephron.

Glomerular Filtration. In the glomerular capillaries, a portion of plasma water is forced through a filter that has three basic components: the fenestrated capillary endothelial cells, a basement membrane lying just beneath the endothelial cells, and the filtration slit diaphragms formed by epithelial cells that cover the basement membrane on its urinary space side. Solutes of small size flow with filtered water (solvent drag) into the urinary (Bowman's) space, whereas formed elements and macromolecules are retained by the filtration barrier. For each nephron unit, the rate of filtration (single-nephron glomerular filtration rate [SNGFR]) is a function of the hydrostatic pressure in the glomerular capillaries (P_GC), the hydrostatic pressure in Bowman's space (which can be equated with pressure in the proximal tubule, P_T), the mean colloid osmotic pressure in the glomerular capillaries (Π_GC), the colloid osmotic pressure in the proximal tubule (Π_T), and the ultra-filtration coefficient (K_f), according to the equation

(25-1)

If P_GC − P_T is defined as the transcapillary hydraulic pressure difference (ΔP), and if Π_T is negligible (as it usually is because little protein is filtered), then

(25-2)

This latter equation succinctly expresses the three major determinants of SNGFR. However, each of these three determinants can be influenced by a number of other variables. K_f is determined by the physicochemical properties of the filtering membrane and by the surface area available for filtration. ΔP is determined primarily by the arterial blood pressure and by the proportion of the arterial pressure that is transmitted to the glomerular capillaries. This is governed by the relative resistances of preglomerular and postglomerular vessels. Π_GC is determined by two variables, the concentration of protein in the arterial blood entering the glomerulus and the single-nephron blood flow (Q_A). Q_A influences Π_GC because as blood transverses the glomerular capillary bed, filtration concentrates proteins in the capillaries, causing Π_GC to rise with distance along the glomerular bed. When Q_A is high, this effect is reduced; however, when Q_A is low, Π_GC may increase to the point that Π_GC = ΔP, and filtration ceases (a condition known as filtration equilibrium).

The proximal tubule is contiguous with Bowman's capsule and takes a tortuous path until finally forming a straight portion that dives into the renal medulla. Based on the morphology of the epithelial cells lining the tubule, the proximal tubule has been subdivided into S1, S2, and S3 segments. Normally, ~65% of filtered Na⁺ is reabsorbed in the proximal tubule, and since this part of the tubule is highly permeable to water, reabsorption is essentially isotonic.

Between the outer and inner strips of the outer medulla, the tubule abruptly changes morphology to become the descending thin limb (DTL), which penetrates the inner medulla, makes a hairpin turn, and then forms the ascending thin limb (ATL). At the juncture between the inner and outer medulla, the tubule once again changes morphology and becomes the thick ascending limb, which is made up of three segments: a medullary portion (MTAL), a cortical portion (CTAL), and a postmacular segment. Together the proximal straight tubule, DTL, ATL, MTAL, CTAL, and postmacular segment are known as the loop of Henle. The DTL is highly permeable to water, yet its permeability to NaCl and urea is low. In contrast, the ATL is permeable to NaCl and urea but is impermeable to water. The thick ascending limb actively reabsorbs NaCl but is impermeable to water and urea. Approximately 25% of filtered Na⁺ is reabsorbed in the loop of Henle, mostly in the thick ascending limb, which has a large reabsorptive capacity.

The thick ascending limb passes between the afferent and efferent arterioles and makes contact with the afferent arteriole by means of a cluster of specialized columnar epithelial cells known as the macula densa. The macula densa is strategically located to sense concentrations of NaCl leaving the loop of Henle. If the concentration of NaCl is too high, the macula densa sends a chemical signal (perhaps adenosine or ATP) to the afferent arteriole of the same nephron, causing it to constrict. This, in turn, causes a reduction in P_GC and Q_A and decreases SNGFR. This homeostatic mechanism, known as tubuloglomerular feedback (TGF), serves to protect the organism from salt and volume wasting. Besides mediating the TGF response, the macula densa also regulates renin release from the adjacent juxtaglomerular cells in the wall of the afferent arteriole.

Approximately 0.2 mm past the macula densa, the tubule changes morphology once again to become the distal convoluted tubule (DCT). The postmacular segment of the thick ascending limb and the distal convoluted tubule often are referred to as the early distal tubule. Like the thick ascending limb, the DCT actively transports NaCl and is impermeable to water. Since these characteristics impart the ability to produce a dilute urine, the thick ascending limb and the DCT are collectively called the diluting segment of the nephron, and the tubular fluid in the DCT is hypotonic regardless of hydration status. However, unlike the thick ascending limb, the DCT does not contribute to the countercurrent-induced hypertonicity of the medullary interstitium (described later in this section). There are marked species differences in transporter expression in the DCT. The DCT has been subdivided into DCT1 and DCT2. DCT1 expresses the thiazide-sensitive NaCl co-transporter but does not express genes involved in transepithelial Ca²⁺ transport such as the Ca²⁺ entry channel, TRPV5, and the Na⁺/Ca²⁺ exchanger. DCT2 expresses Ca²⁺ transport proteins and amiloride-sensitive epithelial Na⁺ channels.

The collecting duct system (connecting tubule + initial collecting tubule + cortical collecting duct + outer and inner medullary collecting ducts—segments 10–14 in Figure 25–1) is an area of fine control of ultrafiltrate composition and volume. It is here that final adjustments in electrolyte composition are made, a process modulated by the adrenal steroid aldosterone. In addition, antidiuretic hormone modulates permeability of this part of the nephron to water.

The more distal portions of the collecting duct pass through the renal medulla, where the interstitial fluid is markedly hypertonic. In the absence of ADH, the collecting duct system is impermeable to water, and a dilute urine is excreted. In the presence of ADH, the collecting duct system is permeable to water, so water is reabsorbed. The movement of water out of the tubule is driven by the steep concentration gradient that exists between tubular fluid and medullary interstitium.

The hypertonicity of the medullary interstitium plays a vital role in the ability of mammals and birds to concentrate urine, and therefore is a key adaptation necessary for living in a terrestrial environment. This is accomplished by a combination of the unique topography of the loop of Henle and the specialized permeability features of the loop's subsegments. Although the precise mechanisms giving rise to the medullary hypertonicity have remained elusive, the "passive countercurrent multiplier hypothesis" is an intuitively attractive model that is qualitatively accurate. According to this hypothesis, the process begins with active transport in the thick ascending limb, which concentrates NaCl in the interstitium of the outer medulla. Since this segment of the nephron is impermeable to water, active transport in the ascending limb dilutes the tubular fluid. As the dilute fluid passes into the collecting-duct system, water is extracted if, and only if, ADH is present. Since the cortical and outer medullary collecting ducts have a low permeability to urea, urea is concentrated in the tubular fluid. The inner medullary collecting duct, however, is permeable to urea, so urea diffuses into the inner medulla, where it is trapped by countercurrent exchange in the vasa recta. Since the DTL is impermeable to salt and urea, the high urea concentration in the inner medulla extracts water from the DTL and concentrates NaCl in the tubular fluid of the DTL. As the tubular fluid enters the ATL, NaCl diffuses out of the salt-permeable ATL, thus contributing to the hypertonicity of the medullary interstitium.

General Mechanism of Renal Epithelial Transport. Figure 25–2 illustrates seven mechanisms by which solutes may cross renal epithelial cell membranes (see also Figure 5–4). If bulk water flow occurs across a membrane, solute molecules will be transferred by convection across the membrane, a process known as solvent drag. Solutes with sufficient lipid solubility also may dissolve in the membrane and diffuse across the membrane down their electrochemical gradients (simple diffusion). Many solutes, however, have limited lipid solubility, and transport must rely on integral proteins embedded in the cell membrane. In some cases the integral protein merely provides a conductive pathway (pore) through which the solute may diffuse passively (channel-mediated diffusion). In other cases the solute may bind to the integral protein and, owing to a conformational change in the protein, be transferred across the cell membrane down an electrochemical gradient (carrier-mediated or facilitated diffusion, also called uniport). However, this process will not result in net movement of solute against an electrochemical gradient. If solute must be moved "uphill" against an electrochemical gradient, then either primary active transport or secondary active transport is required. With primary active transport, ATP hydrolysis is coupled directly to conformational changes in the integral protein, thus providing the necessary free energy (ATP-mediated transport). Often, ATP-mediated transport is used to create an electrochemical gradient for a given solute, and the free energy of that solute gradient is then released to drive the "uphill" transport of other solutes. This process requires symport (co-transport of solute species in the same direction) or antiport (countertransport of solute species in opposite directions) and is known as secondary active transport.

The kinds of transport achieved in a particular nephron segment depend mainly on which transporters are present and whether they are embedded in the luminal or basolateral membrane. A general model of renal tubular transport is shown in Figure 25–3 and can be summarized as follows:

1. Na⁺, K⁺-ATPase (sodium pump) in the basolateral membrane hydrolyzes ATP, which results in the transport of Na⁺ into the intercellular and interstitial spaces, the movement of K⁺ into the cell, and the establishment and maintenance of an electrochemical gradient for Na⁺ across the cell membrane directed inward. Although other ATPases exist in selected renal epithelial cells and participate in the transport of specific solutes (e.g., Ca²⁺-ATPase and H⁺-ATPase), the bulk of all transport in the kidney is due to the abundant supply of Na⁺, K⁺-ATPase in the basolateral membranes of renal epithelial cells and the separation of Na⁺ and K⁺ across the cell membrane.

2. Na⁺ may diffuse across the luminal membrane by means of Na⁺ channels into the epithelial cell down the electrochemical gradient for Na⁺ that is established by the basolateral Na⁺, K⁺-ATPase. In addition, free energy available in the electrochemical gradient for Na⁺ is tapped by integral proteins in the luminal membrane, resulting in co-transport of various solutes against their electrochemical gradients by symporters (e.g., Na⁺-glucose, Na⁺-H₂PO₄⁻, and Na⁺-amino acid). This process results in movement of Na⁺ and co-transported solutes out of the tubular lumen into the cell. Also, antiporters (e.g., Na⁺-H⁺) move Na⁺ out of and some solutes into the tubular lumen.

3. Na⁺ exits the basolateral membrane into the intercellular and interstitial spaces by means of the Na⁺ pump or symporters or antiporters in the basolateral membrane.

4. The action of Na⁺-linked symporters in the luminal membrane causes the concentration of substrates for these symporters to rise in the epithelial cell. These electrochemical gradients then permit simple diffusion or mediated transport (e.g., symporters, antiporters, uniporters, and channels) of solutes into the intercellular and interstitial spaces.

5. Accumulation of Na⁺ and other solutes in the intercellular space creates a small osmotic pressure differential across the epithelial cell. In water-permeable epithelium, water moves into the intercellular spaces driven by the osmotic pressure differential. Water moves through aqueous pores in both the luminal and the basolateral cell membranes, as well as through tight junctions (paracellular pathway). Bulk water flow carries some solutes into the intercellular space by solvent drag.

6. Movement of water into the intercellular space concentrates other solutes in the tubular fluid, resulting in an electrochemical gradient for these substances across the epithelium. Membrane-permeable solutes then move down their electrochemical gradients into the intercellular space by both the transcellular (e.g., simple diffusion, symporters, antiporters, uniporters, and channels) and paracellular pathways. Membrane-impermeable solutes remain in the tubular lumen and are excreted in the urine with an obligatory amount of water.

7. As water and solutes accumulate in the intercellular space, hydrostatic pressure increases, thus providing a driving force for bulk water flow. Bulk water flow carries solute (solute convection) out of the intercellular space into the interstitial space and, finally, into the peritubular capillaries. The movement of fluid into peritubular capillaries is governed by the same Starling forces that determine transcapillary fluid movement for any capillary bed.

Mechanism of Organic Acid and Organic Base Secretion. The kidney is a major organ involved in the elimination of organic chemicals from the body. Organic molecules may enter the renal tubules by glomerular filtration of molecules not bound to plasma proteins or may be actively secreted directly into tubules. The proximal tubule has a highly efficient transport system for organic acids and an equally efficient but separate transport system for organic bases. Current models for these secretory systems are illustrated in Figure 25–4. Both systems are powered by the sodium pump in the basolateral membrane, involve secondary and tertiary active transport, and use a facilitated-diffusion step. There are at least nine different organic acid and five different organic base transporters; the precise roles that these transporters play in organic acid and base transport remain ill defined (Dresser et al., 2001). A family of organic anion transporters (OATs) countertransport organic anions with dicarboxylates (Figure 25–4A). OATs most likely exist as α-helical dodecaspans connected by short segments of ~10 or fewer amino acids, except for large interconnecting stretches of amino acids between helices 1 and 2 and helices 6 and 7 (Eraly et al., 2004); see Chapter 5.

Renal Handling of Specific Anions and Cations. Reabsorption of Cl⁻ generally follows reabsorption of Na⁺. In segments of the tubule with low-resistance tight junctions (i.e., "leaky" epithelium), such as the proximal tubule and thick ascending limb, Cl⁻ movement can occur paracellularly. With regard to transcellular Cl⁻ flux, Cl⁻crosses the luminal membrane by antiport with formate and oxalate (proximal tubule), symport with Na⁺/K⁺ (thick ascending limb), symport with Na⁺ (DCT), and antiport with HCO₃⁻ (collecting-duct system). Cl⁻ crosses the basolateral membrane by symport with K⁺ (proximal tubule and thick ascending limb), antiport with Na⁺/HCO₃⁻ (proximal tubule), and Cl⁻ channels (thick ascending limb, DCT, collecting-duct system).

Eighty to ninety percent of filtered K⁺ is reabsorbed in the proximal tubule (diffusion and solvent drag) and thick ascending limb (diffusion) largely through the paracellular pathway. In contrast, the DCT and collecting-duct system secrete variable amounts of K⁺ by a conductive (channel-mediated) pathway. Modulation of the rate of K⁺ secretion in the collecting-duct system, particularly by aldosterone, allows urinary K⁺ excretion to be matched with dietary intake. The transepithelial potential difference (V_T), lumen-positive in the thick ascending limb and lumen-negative in the collecting-duct system, provides an important driving force for K⁺ reabsorption and secretion, respectively.

Most of the filtered Ca²⁺ (~70%) is reabsorbed by the proximal tubule by passive diffusion through a paracellular route. Another 25% of filtered Ca²⁺ is reabsorbed by the thick ascending limb in part by a paracellular route driven by the lumen-positive V_T and in part by active transcellular Ca²⁺ reabsorption modulated by parathyroid hormone (PTH; see Chapter 44). Most of the remaining Ca²⁺ is reabsorbed in DCT by a transcellular pathway. The transcellular pathway in the thick ascending limb and DCT involves passive Ca²⁺ influx across the luminal membrane through Ca²⁺ channels (TRPV5), followed by Ca²⁺ extrusion across the basolateral membrane by a Ca²⁺-ATPase. Also, in DCT and CNT, Ca²⁺ crosses the basolateral membrane by Na⁺-Ca²⁺ exchanger (antiport).

Inorganic phosphate (P_i) is largely reabsorbed (80% of filtered load) by the proximal tubule. The Na⁺-P_i symporter uses the free energy of the Na⁺ electrochemical gradient to transport P_i into the cell. The Na⁺-P_i symporter is inhibited by PTH. P_i exits the basolateral membrane down its electrochemical gradient by a poorly understood transport system.

Only 20-25% of Mg²⁺ is reabsorbed in the proximal tubule, and only 5% is reabsorbed by the DCT and collecting-duct system. The bulk of Mg²⁺ is reabsorbed in the thick ascending limb by a paracellular pathway driven by the lumen-positive V_T. However, transcellular movement of Mg²⁺ also may occur with basolateral exit by Na⁺-Mg²⁺ antiport or an Mg²⁺-ATPase.

The renal tubules play an extremely important role in the reabsorption of HCO₃⁻ and secretion of protons (tubular acidification) and thus participate critically in the maintenance of acid-base balance. These processes are described in the section on carbonic anhydrase inhibitors.

Chemistry. When sulfanilamide was introduced as a chemotherapeutic agent, metabolic acidosis was recognized as a side effect. This observation led to the demonstration that sulfanilamide is an inhibitor of carbonic anhydrase. Subsequently, an enormous number of sulfonamides were synthesized and tested for their ability to inhibit carbonic anhydrase; of these compounds, acetazolamide has been studied most extensively. There are three orally administered carbonic anhydrase inhibitors—acetazolamide, dichlorphenamide (not marketed in the U.S.), and methazolamide. The common molecular motif of available carbonic anhydrase inhibitors is an unsubstituted sulfonamide moiety (Table 25–2).

In the proximal tubule, the free energy in the Na⁺ gradient established by the basolateral Na⁺ pump is used by an Na⁺-H⁺ antiporter (also referred to as an Na⁺-H⁺ exchanger [NHE]) in the luminal membrane to transport H⁺ into the tubular lumen in exchange for Na⁺ (Figure 25–6). In the lumen, H⁺ reacts with filtered HCO₃⁻ to form H₂CO₃, which decomposes rapidly to CO₂ and water in the presence of carbonic anhydrase in the brush border. Normally, the reaction occurs slowly, but carbonic anhydrase reversibly accelerates this reaction several thousand times. CO₂ is lipophilic and rapidly diffuses across the luminal membrane into the epithelial cell, where it reacts with water to form H₂CO₃, a reaction catalyzed by cytoplasmic carbonic anhydrase (Figure 25–6). Continued operation of the Na⁺-H⁺ antiporter maintains a low proton concentration in the cell, so H₂CO₃ ionizes spontaneously to form H⁺ and HCO₃⁻, creating an electrochemical gradient for HCO₃⁻ across the basolateral membrane. The electrochemical gradient for HCO₃⁻ is used by an Na⁺-HCO₃⁻ symporter (also referred to as the Na⁺-HCO₃⁻ co-transporter [NBC]) in the basolateral membrane to transport NaHCO₃ into the interstitial space. The net effect of this process is transport of NaHCO₃ from the tubular lumen to the interstitial space, followed by movement of water (isotonic reabsorption). Removal of water concentrates Cl⁻ in the tubular lumen, and consequently, Cl⁻ diffuses down its concentration gradient into the interstitium by the paracellular pathway.

Carbonic anhydrase inhibitors potently inhibit (IC₅₀ for acetazolamide is 10 nM) both the membrane-bound and cytoplasmic forms of carbonic anhydrase, resulting in nearly complete abolition of NaHCO₃ reabsorption in the proximal tubule. Studies with a high-molecular-weight carbonic anhydrase inhibitor that inhibits only luminal enzyme because of limited cellular permeability indicate that inhibition of both membrane-bound and cytoplasmic pools of carbonic anhydrase contributes to the diuretic activity of carbonic anhydrase inhibitors. Because of the large excess of carbonic anhydrase in proximal tubules, a high percentage of enzyme activity must be inhibited before an effect on electrolyte excretion is observed. Although the proximal tubule is the major site of action of carbonic anhydrase inhibitors, carbonic anhydrase also is involved in secretion of titratable acid in the collecting duct system (a process that involves a proton pump); therefore, the collecting duct system is a secondary site of action for this class of drugs.

Other mechanisms contributing to enhanced K⁺ excretion include flow-dependent enhancement of K⁺ secretion by the collecting duct, nonosmotic vasopressin release, and activation of the renin-angiotensin-aldosterone axis. Carbonic anhydrase inhibitors increase phosphate excretion (mechanism unknown) but have little or no effect on Ca²⁺ or Mg²⁺ excretion. The effects of carbonic anhydrase inhibitors on renal excretion are self-limiting, probably because the resulting metabolic acidosis decreases the filtered load of HCO₃⁻ to the point that the uncatalyzed reaction between CO₂ and water is sufficient to achieve HCO₃⁻ reabsorption. Studies in rabbits show that the K_i for luminal carbonic anhydrase inhibition in outer medulla is increased >100-fold over the concentration required in control tubules. These results suggest that acidosis induces a conformational change in the protein making it less sensitive to inhibition by acetazolamide (Tsuroka, 1998).

Other Actions. Carbonic anhydrase is present in a number of extrarenal tissues, including the eye, gastric mucosa, pancreas, central nervous system (CNS), and erythrocytes. Carbonic anhydrase in the ciliary processes of the eye mediates formation of large amounts of HCO₃⁻ in aqueous humor. Inhibition of carbonic anhydrase decreases the rate of formation of aqueous humor and consequently reduces intraocular pressure. Acetazolamide frequently causes paresthesias and somnolence, suggesting an action of carbonic anhydrase inhibitors in the CNS. The efficacy of acetazolamide in epilepsy is due in part to the production of metabolic acidosis; however, direct actions of acetazolamide in the CNS also contribute to its anticonvulsant action. Owing to interference with carbonic anhydrase activity in erythrocytes, carbonic anhydrase inhibitors increase CO₂ levels in peripheral tissues and decrease CO₂ levels in expired gas. Large doses of carbonic anhydrase inhibitors reduce gastric acid secretion, but this has no therapeutic application. Acetazolamide causes vasodilation by opening vascular Ca²⁺-activated K⁺ channels; however, the clinical significance of this effect is unclear.

Toxicity, Adverse Effects, Contraindications, Drug Interactions. Serious toxic reactions to carbonic anhydrase inhibitors are infrequent; however, these drugs are sulfonamide derivatives and, like other sulfonamides, may cause bone marrow depression, skin toxicity, sulfonamide-like renal lesions, and allergic reactions in patients hypersensitive to sulfonamides. With large doses, many patients exhibit drowsiness and paresthesias. Most adverse effects, contraindications, and drug interactions are secondary to urinary alkalinization or metabolic acidosis, including:

• diversion of ammonia of renal origin from urine into the systemic circulation, a process that may induce or worsen hepatic encephalopathy (the drugs are contraindicated in patients with hepatic cirrhosis)

• calculus formation and ureteral colic owing to precipitation of calcium phosphate salts in an alkaline urine

• worsening of metabolic or respiratory acidosis (the drugs are contraindicated in patients with hyperchloremic acidosis or severe chronic obstructive pulmonary disease)

• reduction of the urinary excretion rate of weak organic bases

The major indication for carbonic anhydrase inhibitors is open-angle glaucoma. Two products developed specifically for this use are dorzolamide (trusopt, others) and brinzolamide (azopt), which are available only as ophthalmic drops. Carbonic anhydrase inhibitors also may be employed for secondary glaucoma and preoperatively in acute angle-closure glaucoma to lower intraocular pressure before surgery (Chapter 64). Acetazolamide also is used for the treatment of epilepsy (Chapter 21). The rapid development of tolerance, however, may limit the usefulness of carbonic anhydrase inhibitors for epilepsy.

Acetazolamide can provide symptomatic relief in patients with high-altitude illness or mountain sickness; however, it is more appropriate to give acetazolamide as a prophylactic measure. Acetazolamide also is useful in patients with familial periodic paralysis. The mechanism for the beneficial effects of acetazolamide in altitude sickness and familial periodic paralysis is not clear, but it may be related to the induction of a metabolic acidosis. Other off-label clinical uses include the treatment of dural ectasia in individuals with Marfan syndrome, of sleep apnea, and of idiopathic intracranial hypertension. Finally, carbonic anhydrase inhibitors can be useful for correcting a metabolic alkalosis, especially one caused by diuretic-induced increases in H⁺ excretion.

By extracting water from intracellular compartments, osmotic diuretics expand extracellular fluid volume, decrease blood viscosity, and inhibit renin release. These effects increase RBF, and the increase in renal medullary blood flow removes NaCl and urea from the renal medulla, thus reducing medullary tonicity. Under some circumstances, prostaglandins may contribute to the renal vasodilation and medullary washout induced by osmotic diuretics. A reduction in medullary tonicity causes a decrease in the extraction of water from the DTL, which in turn limits the concentration of NaCl in the tubular fluid entering the ATL. This latter effect diminishes the passive reabsorption of NaCl in the ATL. In addition, the marked ability of osmotic diuretics to inhibit Mg²⁺ reabsorption, a cation that is reabsorbed mainly in the thick ascending limb, suggests that osmotic diuretics also interfere with transport processes in the thick ascending limb. The mechanism of this effect is unknown.

Toxicity, Adverse Effects, Contraindications, and Drug Interactions. Osmotic diuretics are distributed in the extracellular fluid and contribute to the extracellular osmolality. Thus, water is extracted from intracellular compartments, and the extracellular fluid volume becomes expanded. In patients with heart failure or pulmonary congestion, this may cause frank pulmonary edema. Extraction of water also causes hyponatremia, which may explain the common adverse effects, including headache, nausea, and vomiting. On the other hand, loss of water in excess of electrolytes can cause hypernatremia and dehydration. Osmotic diuretics are contraindicated in patients who are anuric owing to severe renal disease. Urea may cause thrombosis or pain if extravasation occurs, and it should not be administered to patients with impaired liver function because of the risk of elevation of blood ammonia levels. Both mannitol and urea are contraindicated in patients with active cranial bleeding. Glycerin is metabolized and can cause hyperglycemia.

By increasing the osmotic pressure of plasma, osmotic diuretics extract water from the eye and brain. All osmotic diuretics are used to control intraocular pressure during acute attacks of glaucoma and for short-term reductions in intraocular pressure both preoperatively and postoperatively in patients who require ocular surgery. Also, mannitol and urea are used to reduce cerebral edema and brain mass before and after neurosurgery.

Chemistry. Inhibitors of Na⁺-K⁺-2Cl⁻ symport are a chemically diverse group (Table 25–4). Only furosemide (lasix, others), bumetanide (bumex, others), ethacrynic acid (edecrin, others), and torsemide (demadex, others) are available currently in the U.S. Furosemide and bumetanide contain a sulfonamide moiety. Ethacrynic acid is a phenoxyacetic acid derivative and torsemide is a sulfonylurea.

It was thought initially that Cl⁻ was transported by a primary active electrogenic transporter in the luminal membrane independent of Na⁺. Discovery of furosemide-sensitive Na⁺-K⁺-2Cl⁻ symport in other tissues prompted a more careful investigation of the Na⁺ dependence of Cl⁻ transport in isolated perfused rabbit CTAL. Scrupulous removal of Na⁺ from the luminal perfusate demonstrated the dependence of Cl⁻ transport on Na⁺.

It is now well accepted that flux of Na⁺, K⁺, and Cl⁻ from the lumen into epithelial cells in thick ascending limb is mediated by an Na⁺-K⁺-2Cl⁻ symporter (Figure 25–7). This symporter captures free energy in the Na⁺ electrochemical gradient established by the basolateral Na⁺ pump and provides for "uphill" transport of K⁺ and Cl⁻ into the cell. K⁺ channels in the luminal membrane (called ROMK) provide a conductive pathway for the apical recycling of this cation, and basolateral Cl⁻ channels (called CLC-Kb) provide a basolateral exit mechanism for Cl⁻. Luminal membranes of epithelial cells in thick ascending limb have a large conductive pathway (channels) for K⁺; therefore, apical membrane voltage is determined by the equilibrium potential for K⁺ (E_K) and is hyperpolarized. In contrast, the basolateral membrane has a large conductive pathway (channels) for Cl⁻, so the basolateral membrane voltage is less negative than E_K; that is, conductance for Cl⁻ depolarizes the basolateral membrane. Hyperpolarization of the luminal membrane and depolarization of the basolateral membrane result in a transepithelial potential difference of ~10 mV, with the lumen positive with respect to the interstitial space. This lumen-positive potential difference repels cations (Na⁺, Ca²⁺, and Mg²⁺) and thereby provides an important driving force for the paracellular flux of these cations into the interstitial space.

Inhibitors of Na⁺-K⁺-2Cl⁻ symport bind to the Na⁺-K⁺-2Cl⁻ symporter in the thick ascending limb and block its function, bringing salt transport in this segment of the nephron to a virtual standstill. The molecular mechanism by which this class of drugs blocks the Na⁺-K⁺-2Cl⁻ symporter is unknown, but evidence suggests that these drugs attach to the Cl⁻ binding site located in the symporter's transmembrane domain (Isenring and Forbush, 1997). Inhibitors of Na⁺-K⁺-2Cl⁻ symport also inhibit Ca²⁺ and Mg²⁺ reabsorption in the thick ascending limb by abolishing the transepithelial potential difference that is the dominant driving force for reabsorption of these cations.

Na⁺-K⁺-2Cl⁻ symporters are an important family of transport molecules found in many secretory and absorbing epithelia. The rectal gland of the dogfish shark is a particularly rich source of the protein, and a cDNA encoding an Na⁺-K⁺-2Cl⁻ symporter was isolated from a cDNA library obtained from the dogfish shark rectal gland by screening with antibodies to the shark symporter (Xu et al., 1994). Molecular cloning revealed a deduced amino acid sequence of 1191 residues containing 12 putative membrane-spanning domains flanked by long N and C termini in the cytoplasm. Expression of this protein resulted in Na⁺-K⁺-2Cl⁻ symport that was sensitive to bumetanide. The shark rectal gland Na⁺-K⁺-2Cl⁻ symporter cDNA was used subsequently to screen a human colonic cDNA library, and this provided Na⁺-K⁺-2Cl⁻ symporter cDNA probes from this tissue. These latter probes were used to screen rabbit renal cortical and renal medullary libraries, which allowed cloning of the rabbit renal Na⁺-K⁺-2Cl⁻ symporter (Payne and Forbush, 1994). This symporter is 1099 amino acids in length, is 61% identical to the dogfish shark secretory Na⁺-K⁺-2Cl⁻ symporter, has 12 predicted transmembrane helices, and contains large N- and C-terminal cytoplasmic regions. Subsequent studies demonstrated that Na⁺-K⁺-2Cl⁻ symporters are of two varieties (Kaplan et al., 1996). The "absorptive" symporter (called ENCC2, NKCC2, or BSCl) is expressed only in the kidney, is localized to the apical membrane and subapical intracellular vesicles of the thick ascending limb, and is regulated by cyclic AMP/PKA (Obermüller et al., 1996; Plata et al., 1999). At least six different isoforms of the absorptive symporter are generated by alternative mRNA splicing (Mount et al., 1999), and alternative splicing of the absorptive symporter determines the dependency of transport on K⁺ (Plata et al., 2001). The "secretory" symporter (called ENCC3, NKCCl, or BSC2) is a "housekeeping" protein that is expressed widely and, in epithelial cells, is localized to the basolateral membrane. The affinity of loop diuretics for the secretory symporter is somewhat less than for the absorptive symporter (e.g., 4-fold difference for bumetanide). A model of Na⁺-K⁺-2Cl⁻ symport has been proposed based on ordered binding of ions to the symporter (Lytle et al., 1998). Mutations in genes coding for the absorptive Na⁺-K⁺-2Cl⁻ symporter, the apical K⁺ channel, the basolateral Cl⁻ channel, or the chloride channel subunit Barttin are causes of Bartter syndrome (inherited hypokalemic alkalosis with salt wasting and hypotension) (Simon and Lifton, 1998).

Effects on Renal Hemodynamics. If volume depletion is prevented by replacing fluid losses, inhibitors of Na⁺-K⁺-2Cl⁻ symport generally increase total RBF and redistribute RBF to the midcortex. However, the effects on RBF are variable. The mechanism of the increase in RBF is not known but may involve prostaglandins. Nonsteroidal anti-inflammatory drugs (NSAIDs) attenuate the diuretic response to loop diuretics in part by preventing prostaglandin-mediated increases in RBF. Loop diuretics block TGF by inhibiting salt transport into the macula densa so that the macula densa no longer can detect NaCl concentrations in the tubular fluid. Therefore, unlike carbonic anhydrase inhibitors, loop diuretics do not decrease GFR by activating TGF. Loop diuretics are powerful stimulators of renin release. This effect is due to interference with NaCl transport by the macula densa and, if volume depletion occurs, to reflex activation of the sympathetic nervous system and stimulation of the intrarenal baroreceptor mechanism. Prostaglandins, particularly prostacyclin, may play an important role in mediating the renin-release response to loop diuretics.

Other Actions. Loop diuretics may cause direct vascular effects. Loop diuretics, particularly furosemide, acutely increase systemic venous capacitance and thereby decrease left ventricular filling pressure. This effect, which may be mediated by prostaglandins and requires intact kidneys, benefits patients with pulmonary edema even before diuresis ensues. Furosemide and ethacrynic acid can inhibit Na⁺,K⁺-ATPase, glycolysis, mitochondrial respiration, the microsomal Ca²⁺ pump, adenylyl cyclase, phosphodiesterase, and prostaglandin dehydrogenase; however, these effects do not have therapeutic implications. In vitro, high doses of inhibitors of Na⁺-K⁺-2Cl⁻ symport can inhibit electrolyte transport in many tissues. Only in the inner ear, where alterations in the electrolyte composition of endolymph may contribute to drug-induced ototoxicity, is this effect important clinically. Irreversible ototoxicity is more common at high doses, with rapid IV administration, and during concommitant therapy with other drugs known to be ototoxic (e.g., aminoglycoside antibiotics, cisplatin, vancomycin).

Approximately 65% of furosemide is excreted unchanged in urine, and the remainder is conjugated to glucuronic acid in the kidney. Thus, in patients with renal but not liver disease, the elimination t_1/2 of furosemide is prolonged. In contrast, bumetanide and torsemide have significant hepatic metabolism, so the elimination t_1/2 of these loop diuretics are prolonged by liver but not renal disease.

Although average oral availability of furosemide is ~60%, bioavailability varies (10-100%). In contrast, oral availabilities of bumetanide and torsemide are reliably high. Heart failure patients have fewer hospitalizations and better quality of life with torsemide than with furosemide, perhaps because of its more reliable absorption.

As a class, loop diuretics have short elimination t_1/2, and prolonged-release preparations are not available. Consequently, often the dosing interval is too short to maintain adequate levels of loop diuretics in the tubular lumen. Note that torsemide has a longer t_1/2 than other agents available in the U.S. (Table 25–4). As the concentration of loop diuretic in the tubular lumen declines, nephrons begin to avidly reabsorb Na⁺, which often nullifies the overall effect of the loop diuretic on total-body Na⁺. This phenomenon of "postdiuretic Na⁺ retention" can be overcome by restricting dietary Na⁺ intake or by more frequent administration of the loop diuretic (Ellison, 1999). Furosemide, with its short t_1/2, often produces a poor response when administered only once daily, a common clinical error.

This may manifest as hyponatremia and/or extracellular fluid volume depletion associated with hypotension, reduced GFR, circulatory collapse, thromboembolic episodes, and in patients with liver disease, hepatic encephalopathy. Increased Na⁺ delivery to the distal tubule, particularly when combined with activation of the rennin–angiotensin system, leads to increased urinary K⁺ and H⁺ excretion, causing a hypochloremic alkalosis. If dietary K⁺ intake is not sufficient, hypokalemia may develop, and this may induce cardiac arrhythmias, particularly in patients taking cardiac glycosides. Increased Mg²⁺ and Ca²⁺ excretion may result in hypomagnesemia (a risk factor for cardiac arrhythmias) and hypocalcemia (rarely leading to tetany). Recent evidence suggests that loop diuretics should be avoided in postmenopausal osteopenic women, in whom increased Ca²⁺ excretion may have deleterious effects on bone metabolism (Rejnmark et al., 2003).

Loop diuretics can cause ototoxicity that manifests as tinnitus, hearing impairment, deafness, vertigo, and a sense of fullness in the ears. Hearing impairment and deafness are usually, but not always, reversible. Ototoxicity occurs most frequently with rapid intravenous administration and least frequently with oral administration. Ethacrynic acid appears to induce ototoxicity more often than do other loop diuretics and should be reserved for use only in patients who cannot tolerate other loop diuretics. Loop diuretics also can cause hyperuricemia (occasionally leading to gout) and hyperglycemia (infrequently precipitating diabetes mellitus) and can increase plasma levels of low-density lipoprotein (LDL) cholesterol and triglycerides while decreasing plasma levels of high-density lipoprotein (HDL) cholesterol. Other adverse effects include skin rashes, photosensitivity, paresthesias, bone marrow depression, and GI disturbances. Glucose intolerance frequently can be demonstrated but precipitation of diabetes mellitus has been reported only rarely.

Contraindications to the use of loop diuretics include severe Na⁺ and volume depletion, hypersensitivity to sulfonamides (for sulfonamide-based loop diuretics), and anuria unresponsive to a trial dose of loop diuretic.

Drug interactions may occur when loop diuretics are co-administered with:

• aminoglycosides, carboplatin, paclitaxel and several other agents (synergism of ototoxicity)

• anticoagulants (increased anticoagulant activity)

• digitalis glycosides (increased digitalis-induced arrhythmias)

• lithium (increased plasma levels of lithium)

• propranolol (increased plasma levels of propranolol)

• sulfonylureas (hyperglycemia)

• cisplatin (increased risk of diuretic-induced ototoxicity)

• NSAIDs (blunted diuretic response and salicylate toxicity when given with high doses of salicylates)

• probenecid (blunted diuretic response)

• thiazide diuretics (synergism of diuretic activity of both drugs leading to profound diuresis)

• amphotericin B (increased potential for nephrotoxicity and toxicity and intensification of electrolyte imbalance)

The edema of nephrotic syndrome often is refractory to less potent diuretics, and loop diuretics often are the only drugs capable of reducing the massive edema associated with this renal disease. Loop diuretics also are employed in the treatment of edema and ascites of liver cirrhosis; however, care must be taken not to induce volume contraction. In patients with a drug overdose, loop diuretics can be used to induce a forced diuresis to facilitate more rapid renal elimination of the offending drug. Loop diuretics, combined with isotonic saline administration to prevent volume depletion, are used to treat hypercalcemia. Loop diuretics interfere with the kidney's capacity to produce a concentrated urine. Consequently, loop diuretics combined with hypertonic saline are useful for the treatment of life-threatening hyponatremia.

Loop diuretics also are used to treat edema associated with chronic kidney disease. Higher doses of loop diuretics are often required in patients with chronic kidney disease. The dose-response curve is shifted to the right (Figure 25–8). Animal studies have demonstrated that loop diuretics increase P_GC by activating the renin-angiotensin system, an effect that could accelerate renal injury (Lane et al., 1998). Most patients with ARF receive a trial dose of a loop diuretic in an attempt to convert oliguric ARF to nonoliguric ARF. However, there is no evidence that loop diuretics prevent ATN or improve outcome in patients with ARF, and as mentioned earlier, the appearance of undesired effects increases with dose.

Chemistry. Inhibitors of Na⁺-Cl⁻ symport are sulfonamides (Table 25–5), and many are analogs of 1,2,4-benzothiadiazine-1,1-dioxide. Because the original inhibitors of Na⁺-Cl⁻ symport were benzothiadiazine derivatives, this class of diuretics became known as thiazide diuretics. Subsequently, drugs that are pharmacologically similar to thiazide diuretics but are not thiazides were developed and are called thiazide-like diuretics. The term thiazide diuretics is used here to refer to all members of the class of inhibitors of Na⁺-Cl⁻ symport.

Figure 25–9 illustrates the current model of electrolyte transport in DCT. As with other nephron segments, transport is powered by a Na⁺ pump in the basolateral membrane. Free energy in the electrochemical gradient for Na⁺ is harnessed by an Na⁺-Cl⁻ symporter in the luminal membrane that moves Cl⁻ into the epithelial cell against its electrochemical gradient. Cl⁻ then exits the basolateral membrane passively by a Cl⁻ channel. Thiazide diuretics inhibit the Na⁺-Cl⁻ symporter. In this regard, Na⁺ or Cl⁻ binding to the Na⁺-Cl⁻ symporter modifies thiazide-induced inhibition of the symporter, suggesting that the thiazide-binding site is shared or altered by both Na⁺ and Cl⁻.

Using a functional expression strategy (Cl⁻-dependent Na⁺ uptake in Xenopus oocytes), Gamba and colleagues (1993) isolated a cDNA clone from the urinary bladder of the winter flounder that codes for an Na⁺-Cl⁻ symporter. This Na⁺-Cl⁻ symporter is inhibited by a number of thiazide diuretics (but not by furosemide, acetazolamide, or an amiloride derivative) and has 12 putative membrane-spanning domains, and its sequence is 47% identical to the cloned dogfish shark rectal gland Na⁺-K⁺-2Cl⁻ symporter. Subsequently, the rat and human Na⁺-Cl⁻ symporters were cloned. The human Na⁺-Cl⁻ symporter has a predicted sequence of 1021 amino acids, 12 transmembrane domains, and 2 intracellular hydrophilic amino and carboxyl termini and maps to chromosome 16q13. The Na⁺-Cl⁻ symporter (called ENCCl or TSC) is expressed predominantly in kidney and is localized to the apical membrane of DCT epithelial cells (Bachmann et al., 1995). Expression of the Na⁺-Cl⁻ symporter is regulated by aldosterone (Velázquez et al., 1996). Mutations in the Na⁺-Cl⁻ symporter cause a form of inherited hypokalemic alkalosis called Gitelman syndrome (Simon and Lifton, 1998).

Other Actions. Thiazide diuretics may inhibit cyclic nucleotide phosphodiesterases, mitochondrial O₂ consumption, and renal uptake of fatty acids; however, these effects are not clinically significant.

Note the wide range of half-lives for this class of drugs. Sulfonamides are organic acids and therefore are secreted into the proximal tubule by the organic acid secretory pathway. Since thiazides must gain access to the tubular lumen to inhibit the Na⁺-Cl⁻ symporter, drugs such as probenecid can attenuate the diuretic response to thiazides by competing for transport into proximal tubule. However, plasma protein binding varies considerably among thiazide diuretics, and this parameter determines the contribution that filtration makes to tubular delivery of a specific thiazide.

Toxicity, Adverse Effects, Contraindications, Drug Interactions. Thiazide diuretics rarely cause CNS (e.g., vertigo, headache, paresthesias, xanthopsia, and weakness), GI (e.g., anorexia, nausea, vomiting, cramping, diarrhea, constipation, cholecystitis, and pancreatitis), hematological (e.g., blood dyscrasias), and dermatological (e.g., photosensitivity and skin rashes) disorders. The incidence of erectile dysfunction is greater with Na⁺-Cl⁻ symport inhibitors than with several other antihypertensive agents (e.g., β adrenergic receptor antagonists, Ca²⁺-channel blockers, or angiotensin converting enzyme inhibitors) (Grimm et al., 1997), but usually is tolerable. As with loop diuretics, most serious adverse effects of thiazides are related to abnormalities of fluid and electrolyte balance. These adverse effects include extracellular volume depletion, hypotension, hypokalemia, hyponatremia, hypochloremia, metabolic alkalosis, hypomagnesemia, hypercalcemia, and hyperuricemia. Thiazide diuretics have caused fatal or near-fatal hyponatremia, and some patients are at recurrent risk of hyponatremia when rechallenged with thiazides.

Thiazide diuretics also decrease glucose tolerance, and latent diabetes mellitus may be unmasked during therapy. Recent concerns have also been raised in randomized prospective blood-pressure lowering trials regarding an increased incidence of type II diabetes mellitus compared to other antihypertensive agents such as angiotensin-converting enzyme inhibitors and angiotensin receptor blockers. The mechanism of impaired glucose tolerance is not completely understood but appears to involve reduced insulin secretion and alterations in glucose metabolism. Hyperglycemia may be related in some way to K⁺ depletion, in that hyperglycemia is reduced when K⁺ is given along with the diuretic. In addition to contributing to hyperglycemia, thiazide-induced hypokalemia impairs its antihypertensive effect and cardiovascular protection (Franse et al., 2000) afforded by thiazides in patients with hypertension. Thiazide diuretics also may increase plasma levels of LDL cholesterol, total cholesterol, and total triglycerides. Thiazide diuretics are contraindicated in individuals who are hypersensitive to sulfonamides.

With regard to drug interactions, thiazide diuretics may diminish the effects of anticoagulants, uricosuric agents used to treat gout, sulfonylureas, and insulin and may increase the effects of anesthetics, diazoxide, digitalis glycosides, lithium, loop diuretics, and vitamin D. The effectiveness of thiazide diuretics may be reduced by NSAIDs, nonselective or selective COX-2 inhibitors, and bile acid sequestrants (reduced absorption of thiazides). Amphotericin B and corticosteroids increase the risk of hypokalemia induced by thiazide diuretics.

A potentially lethal drug interaction warranting special emphasis is that involving thiazide diuretics and quinidine. Prolongation of the QT interval by quinidine can lead to the development of polymorphic ventricular tachycardia (torsades de pointes) owing to triggered activity originating from early after-depolarizations (Chapter 29). Torsades de pointes may deteriorate into fatal ventricular fibrillation. Hypokalemia increases the risk of quinidine-induced torsades de pointes, and thiazide diuretics cause hypokalemia. Thiazide diuretic–induced K⁺ depletion may account for many cases of quinidine-induced torsades de pointes. In addition, alkalinization of the urine by thiazides increases the systemic exposure to quinidine by reducing its elimination.

Thiazide diuretics decrease blood pressure in hypertensive patients by increasing the slope of the renal pressure-natriuresis relationship (Figure 26–7), and thiazide diuretics are used widely for the treatment of hypertension either alone or in combination with other antihypertensive drugs (Chapter 28). In this regard, thiazide diuretics are inexpensive, as efficacious as other classes of antihypertensive agents, and well tolerated. Thiazides can be administered once daily, do not require dose titration, and have few contraindications. Moreover, thiazides have additive or synergistic effects when combined with other classes of antihypertensive agents. A common dose for hypertension is 25 mg/day of hydrochlorothiazide or the dose equivalent of another thiazide. The ALLHAT study (ALLHAT Officers and Coordinators for the ALLHAT Collaborative Research Group, 2002) provides strong evidence that thiazide diuretics are the best initial therapy for uncomplicated hypertension, a conclusion endorsed by the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (Chobanian et al., 2003) (Chapter 27). Studies also suggest that the antihypertensive response to thiazides is influenced by polymorphisms in the angiotensin-converting enzyme and α-adducin genes (Sciarrone et al., 2003).

Thiazide diuretics, which reduce urinary Ca²⁺ excretion, sometimes are employed to treat Ca²⁺ nephrolithiasis and may be useful for treatment of osteoporosis (Chapter 44). Thiazide diuretics also are the mainstay for treatment of nephrogenic diabetes insipidus, reducing urine volume by up to 50%. Although it may seem counterintuitive to treat a disorder of increased urine volume with a diuretic, thiazides reduce the kidney's ability to excrete free water. They do so by increasing proximal tubular water reabsorption (secondary to volume contraction) and by blocking the ability of the distal convoluted tubule to form dilute urine. This latter effect results in an increase in urine osmolality. Since other halides are excreted by renal processes similar to those for Cl⁻, thiazide diuretics may be useful for the management of Br⁻ intoxication.

Chemistry. Amiloride is a pyrazinoylguanidine derivative, and triamterene is a pteridine (Table 25–6). Both drugs are organic bases and are transported by the organic base secretory mechanism in proximal tubule.

Principal cells in the late distal tubule and collecting duct have, in their luminal membranes, epithelial Na⁺ channels that provide a conductive pathway for Na⁺ entry into the cell down the electrochemical gradient created by the basolateral Na⁺ pump. The higher permeability of the luminal membrane for Na⁺ depolarizes the luminal membrane but not the basolateral membrane, creating a lumen-negative transepithelial potential difference. This transepithelial voltage provides an important driving force for the secretion of K⁺ into the lumen by K⁺ channels (ROMK) in the luminal membrane. Carbonic anhydrase inhibitors, loop diuretics, and thiazide diuretics increase Na⁺ delivery to the late distal tubule and collecting duct, a situation that often is associated with increased K⁺ and H⁺ excretion. It is likely that the elevation in luminal Na⁺ concentration in distal nephron induced by such diuretics augments depolarization of the luminal membrane and thereby enhances the lumen-negative V_T, which facilitates K⁺ excretion. In addition to principal cells, the collecting duct also contains type A intercalated cells that mediate H⁺ secretion into the tubular lumen. Tubular acidification is driven by a luminal H⁺-ATPase (proton pump), and this pump is aided by partial depolarization of the luminal membrane. The luminal H⁺-ATPase is of the vacuolar-type and is distinct from the gastric H⁺-K⁺-ATPase that is inhibited by drugs such as omeprazole. However, increased distal Na⁺ delivery is not the only mechanism by which diuretics increase K⁺ and H⁺ excretion. Activation of the renin-angiotensin-aldosterone axis by diuretics also contributes to diuretic-induced K⁺ and H⁺ excretion, as discussed later in the section on mineralocorticoid antagonists.

Considerable evidence indicates that amiloride blocks epithelial Na⁺ channels in the luminal membrane of principal cells in late distal tubule and collecting duct. The amiloride-sensitive Na⁺ channel (called ENaC) consists of three subunits (α, β, and γ) (Kleyman et al., 1999). Although the α subunit is sufficient for channel activity, maximal Na⁺ permeability is induced when all three subunits are coexpressed in the same cell, probably forming a tetrameric structure consisting of two α subunits, one β subunit, and one γ subunit. Studies in Xenopus oocytes expressing ENaC suggest that triamterene and amiloride bind ENaC by similar mechanisms. The K_i of amiloride for ENaC is submicromolar, and molecular studies identified critical domains in ENaC that participate in amiloride binding (Kleyman et al., 1999). Liddle syndrome is an autosomal dominant form of low-renin, volume-expanded hypertension that is due to mutations in the β or γ subunits, leading to increased basal ENaC activity.

Other Actions. Amiloride, at concentrations higher than needed to elicit therapeutic effects, also blocks the Na⁺-H⁺ and Na⁺-Ca²⁺ antiporters and inhibits Na⁺, K⁺-ATPase.

Amiloride is eliminated predominantly by urinary excretion of intact drug. Triamterene is metabolized extensively to an active metabolite, 4-hydroxytriamterene sulfate, and this metabolite is excreted in urine. The pharmacological activity of 4-hydroxytriamterene sulfate is comparable with that of the parent drug. Therefore, triamterene toxicity may be enhanced in both hepatic disease (decreased metabolism of triamterene) and renal failure (decreased urinary excretion of active metabolite).

Pentamidine and high-dose trimethoprim are used often to treat Pneumocystis jirovecii pneumonia in patients with acquired immune deficiency syndrome (AIDS). Because these compounds are weak inhibitors of ENaC, they too may cause hyperkalemia. Risk with trimethoprim is related to both the dosage employed and the underlying level of renal function. It was first reported with high-dose therapy (200 mg/kg/day) in 1983. Subsequent reports of hyperkalemia occurred in HIV-infected patients with normal renal function on high doses of the drug. In one series of 30 such patients, 50% developed a serum K⁺ concentration >5.0 mEq/L and 10% >6.0 mEq/L. In a prospective study of otherwise healthy outpatients, the frequency was lower with a serum K⁺ concentration >5.5 mEq/L in only 6%. Within 3-10 days after onset of treatment, 10-21% of patients develop a K⁺ concentration >5.5 mEq/L (Perazella, 2000). Risk factors for severe hyperkalemia are older age, high-dose therapy, renal impairment, hypoaldosteronism, and treatment with other drugs that impair renal K⁺ excretion (e.g., NSAIDs and ACE inhibitors). Serum K⁺ concentration should be monitored after 3-4 days of trimethoprim treatment especially in those at increased risk. Likewise, routine monitoring of the serum K⁺ level is essential in patients receiving K⁺-sparing diuretics. Cirrhotic patients are prone to megaloblastosis because of folic acid deficiency, and triamterene, a weak folic acid antagonist, may increase the likelihood of this adverse event. Triamterene also can reduce glucose tolerance and induce photosensitization and has been associated with interstitial nephritis and renal stones. Both drugs can cause CNS, GI, musculoskeletal, dermatological, and hematological adverse effects. The most common adverse effects of amiloride are nausea, vomiting, diarrhea, and headache; those of triamterene are nausea, vomiting, leg cramps, and dizziness.

Co-administration of an Na⁺-channel inhibitor augments the diuretic and antihypertensive response to thiazide and loop diuretics. More important, the ability of Na⁺-channel inhibitors to reduce K⁺ excretion tends to offset the kaliuretic effects of thiazide and loop diuretics; consequently, the combination of an Na⁺-channel inhibitor with a thiazide or loop diuretic tends to result in normal plasma K⁺ values. Liddle syndrome can be treated effectively with Na⁺-channel inhibitors. Approximately 5% of people of African origin carry a T594M polymorphism in the β subunit of ENaC, and amiloride is particularly effective in lowering blood pressure in patients with hypertension who carry this polymorphism (Baker et al., 2002). Aerosolized amiloride has been shown to improve mucociliary clearance in patients with cystic fibrosis. By inhibiting Na⁺ absorption from the surfaces of airway epithelial cells, amiloride augments hydration of respiratory secretions and thereby improves mucociliary clearance. Amiloride also is useful for lithium-induced nephrogenic diabetes insipidus because it blocks Li⁺ transport into collecting tubule cells.

The discovery of gene mutation responsible for rare monogenic diseases that cause hypertension such as Liddle syndrome and apparent mineralocorticoid excess helped clarify how aldosterone regulates Na⁺ transport in the distal nephron (Figure 25–11). Mutations in the carboxy-terminal PY motif of either the β or γ subunits of ENaC are associated with Liddle syndrome. The PY motif is an area involved in protein-protein interaction. The PY motif of ENaC interacts with the ubiquitin ligase Nedd4-2, a protein that ubiquitinates ENaC. This then results in internalization of ENaC and proteasome-mediated degradation. Subsequent studies revealed that Nedd4-2 is phosphorylated and inactivated by SGK1 (serum and glucocorticoid-stimulated kinase) and that SGK1 is up-regulated after ~30 minutes by aldosterone. By tracing back the pathophysiologic mechanism of Liddle syndrome, one of the mechanisms whereby aldosterone acts in the collecting duct was identified. Aldosterone up-regulates SGK1, which phosphorylates and inactivates Nedd4-2. As a result, ENaC is not ubiquitinated and removed from the membrane, thereby increasing Na⁺ reabsorption. When the mineralocorticoid receptor was cloned and tested in vitro it was noted to have equal affinity for mineralocorticoid and glucocorticoids. It had been assumed that the mineralocorticoid receptor would have specificity for mineralocorticoids, but surprisingly this was not the case. Given that glucocorticoids circulate at 100- to 1000-fold higher concentration than mineralocorticoids, it was unclear how mineralocorticoids would ever bind to their receptor. The mineralocorticoid receptor would be predominantly occupied by glucocorticoids. This mystery was solved with the cloning of the enzyme type II 11-β-hydroxysteroid dehydrogenase (HSD). Mineralocorticoid target tissues expresses type II 11-β-HSD, which converts cortisol to the inactive cortisone. This allows mineralocorticoids to bind to the receptor. The type II 11-β-HSD enzyme is genetically absent in the inherited disorder of apparent mineralocorticoid excess.

Drugs such as spironolactone and eplerenone competitively inhibit the binding of aldosterone to the MR. Unlike the MR-aldosterone complex, the MR-spironolactone complex is not able to induce the synthesis of AIPs. Since spironolactone and eplerenone block biological effects of aldosterone, these agents also are referred to as aldosterone antagonists.

Other Actions. Spironolactone has some affinity toward progesterone and androgen receptors and thereby induces side effects such as gynecomastia, impotence, and menstrual irregularities. Owing to the 9,11-epoxide group, eplerenone has very low affinity for progesterone and androgen receptors (<1% and <0.1%, respectively) compared with spironolactone. High spironolactone concentrations were reported to interfere with steroid biosynthesis by inhibiting steroid hydroxylases (e.g., CYPs 11A1, 11B1, 11B2, and C21; see Chapters 40 and 41). These effects have limited clinical relevance.

Spironolactone undergoes metabolism by both deacylation and dethioacetylation to a variety of metabolites with prolonged t_1/2; of these, 7α thiomethyl spirononlactone is the primary active metabolite.

Although not available in the U.S., canrenone and the K⁺ salt of canrenoate also are in clinical use. Canrenoate is not active per se but is converted to canrenone. Eplerenone has good oral availability and is eliminated primarily by metabolism by CYP3A4 to inactive metabolites, with a t_1/2 of ~5 hours.

Salicylates may reduce the tubular secretion of canrenone and decrease diuretic efficacy of spironolactone, and spironolactone may alter the clearance of digitalis glycosides. Owing to its affinity for other steroid receptors, spironolactone may cause gynecomastia, impotence, decreased libido, hirsutism, deepening of the voice, and menstrual irregularities. Spironolactone also may induce diarrhea, gastritis, gastric bleeding, and peptic ulcers (the drug is contraindicated in patients with peptic ulcers). CNS adverse effects include drowsiness, lethargy, ataxia, confusion, and headache. Spironolactone may cause skin rashes and, rarely, blood dyscrasias. Breast cancer has occurred in patients taking spironolactone chronically (cause and effect not established), and high doses of spironolactone are associated with malignant tumors in rats. Whether or not therapeutic spironolactone doses can induce malignancies remains an open question. Strong inhibitors of CYP3A4 may increase plasma levels of eplerenone, and such drugs should not be administered to patients taking eplerenone, and vice versa. Other than hyperkalemia and GI disorders, the rate of adverse events for eplerenone is similar to that of placebo (Pitt et al., 2003).

Clinical experience with eplerenone is less than that with spironolactone. Eplerenone appears to be a safe and effective antihypertensive drug (Ouzan et al., 2002). It is somewhat more specific for the MR and therefore the incidience of progesterone-related adverse effects (e.g., gynecomastia) is lower than with spironolactone. In patients with acute myocardial infarction complicated by left ventricular systolic dysfunction, addition of eplerenone to optimal medical therapy significantly reduces morbidity and mortality (Pitt et al., 2003).

Mechanism and Site of Action. The IMCD is the final site along the nephron where Na⁺ is reabsorbed. Up to 5% of the filtered Na⁺ load can be reabsorbed here. Na⁺ enters the IMCD cell across the apical membrane down an electrochemical gradient through Na⁺ channels and exits via the Na⁺/K⁺-ATPase (Figure 25–12). Two types of Na⁺ channels are expressed in IMCD. The first is an amiloride-sensitive, 28pS, nonselective, cyclic nucleotide gated cation (CNG) channel. This channel is highly selective for cations over anions, has equal permeability for Na⁺ and K⁺, and is inhibited by cGMP, PKG, PKC, ATP, and atrial natriuretic peptides via their capacity to stimulate membrane-bound guanylyl cyclase activity and elevate cellular cGMP. Its open probability is increased by rises in intracellular Ca²⁺ concentration. CNG channels are expressed in all nephron segments with the possible exception of the thin limb of Henle's loop. The second type of Na⁺ channel expressed in the IMCD is the low-conductance 4 pS highly-selective Na⁺ channel ENaC. It appears that the majority of Na⁺ reabsorption in the IMCD is mediated via the CNG channel.

ANP has a 17 amino acid core ring and a cysteine bridge. It is produced in the cardiac atria in response to wall stretch. It binds to the natriuretic peptide receptor (NPR) A and increases intracellular cyclic GMP resulting in inhibition of the CNG channel and natriuresis. It also inhibits production of renin and aldosterone. BNP is produced in the ventricle, also binds to the NPR-A receptor and acts in a similar fashion as ANP. CNP binds to the NPR-B receptor and increases cGMP in vascular smooth muscle and mediates vasodilation. Urodilatin arises from the same precursor molecule as ANP but has four additional amino acids at the N terminus. It binds with lower affinity than ANP to the NPR-B receptor and has effects in glomeruli and IMCD. As a result of these effects, natriuretic peptides have been utilized to treat congestive heart failure. Human recombinant ANP (carperitide) is available in Japan but not yet in the U.S., where human recombinant BNP (nesiritide) is available. Urodilatin (ularitide) remains in preclinical trials in the U.S. and Europe. Only the use of nesiritide is discussed in the following sections. Nesiritide effects renal Na⁺ excretion by inhibiting the CNG nonspecific-cation channel, as well as through inhibiting both the renin-angiotensin-aldosterone system and endothelin production.

Other Actions. Administration of nesiritide decreases systemic and pulmonary resistances and left ventricular filling pressure, and induces a secondary increase in cardiac output. In one study, mean arterial pressure, pulmonary capillary wedge pressure, and right atrial pressure declined 17%, 48%, and 56%, respectively. Declines in blood pressure with administration are dose related.

Elimination. Natriuretic peptides are administered intravenously and have short t_1/2 values. Nesiritide has a distribution t_1/2 of 2 minutes and a mean terminal t_1/2 of 18 minutes. Its volume of distribution is small, 0.19 L/kg. It is cleared via three mechanisms: binding to the NPR-C receptor, degradation via a neutral endopeptidase, and renal excretion. There is no need to adjust the dose for renal insufficiency.

Toxicity, Adverse Effects, Contraindications, Drug Interactions. There are concerns about adverse renal effects and reports of increased short-term mortality in patients treated with nesiritide. Increases in serum creatinine concentration may be related to decreases in extracellular fluid volume, higher doses of diuretics used, decreases in blood pressure, and activation of the renin-angiotensin-aldosterone system. The Vasodilation in the Management of Acute CHF (VAMC) trial showed no increased risk with low or moderate doses of diuretics but an increased risk with high-dose diuretics (>160 mg furosemide), rising with increasing doses. The risk of hypotension was 4.4% and lasted an average of 2.2 hours. Oral ACE inhibitors may increase the risk of hypotension with nesiritide. Data on whether 30-day mortality is increased by nesiritide are conflicting. There are no data to suggest that nesiritide reduces mortality in the short term or long term in patients with acute decompensated CHF.

In many clinical situations, edema is not caused by abnormal Na⁺ intake or by an altered renal Na⁺ handling. Rather, edema is the result of altered Starling forces at the capillary beds—a "Starling trap." Use of diuretics in these clinical settings represents a judicious compromise between the edematous state and the hypovolemic state. In such conditions, reducing total ECFV with diuretics will decrease edema but also will cause depletion of "sensed" ECFV, possibly leading to hypotension, malaise, and asthenia.

NSAID co-administration is a common preventable cause of diuretic resistance. An estimated 10% of the elderly each year have periods when NSAID and diuretic prescriptions overlap. Prostaglandin production, especially PGE₂, is an important counterregulatory mechanism in states of reduced renal perfusion such as volume contraction, congestive heart failure, and cirrhosis characterized by activation of the renin-angiotensin-aldosterone (RAA) and sympathetic nervous systems. Prostaglandins reduce NaCl reabsorption in the distal nephron, antagonize effects of ADH, and redistribute blood flow from cortex to juxtamedullary regions in this setting. NSAID administration can block these prostaglandin-mediated effects that counterbalance the RAA and sympathetic nervous system, resulting in salt and water retention. Interaction between NSAIDs and diuretics can result in important clinical consequences such as development or exacerbation of congestive heart failure. NSAIDs can also increase blood pressure, especially in the elderly. In one study of patients over age 55, the relative risk of hospitalization for congestive heart failure (CHF) during periods of concomitant diuretic and NSAID use was increased 2.2-fold over that with diuretic use alone. The majority of these admissions occurred in the first month of simultaneous use, most in the first 20 days. After 40 days, risk was similar to that before simultaneous use. This appeared to be a class effect with no significant differences between individual NSAIDs. Diuretic resistance also occurs with COX-2-selective inhibitors.

All currently available diuretics perturb K⁺ homeostasis. However, studies in animals established that blockade of adenosine A₁ receptors induces a brisk natriuresis without significantly increasing urinary K⁺ excretion (Kuan et al., 1993). Two clinical studies with FK453, a highly selective A₁-receptor antagonist, confirm that blockade of A₁ receptors induces natriuresis in humans with minimal effects on K⁺ excretion (van Buren et al., 1993). The natriuretic mechanism of this novel class of diuretics has been partially elucidated (Takeda et al., 1993). Elevated intracellular cyclic AMP reduces basolateral Na⁺-HCO₃⁻ symport in proximal tubular cells. Endogenous adenosine normally acts on A₁ receptors in these cells to inhibit adenylyl cyclase and reduce cyclic AMP accumulation. Blockade of A₁ receptors removes this inhibition, permits cellular cyclic AMP to rise, and results in reduced activity of the Na⁺-HCO₃⁻ symporter. Because A₁ receptors are involved in TGF, A₁ receptor antagonists uncouple increased distal Na⁺ delivery from activation of TGF. Other mechanisms, including an effect in the collecting tubules, contribute to the natriuretic response to A₁-receptor antagonists; however, it is not known why this class of diuretics has little effect on K⁺ excretion. In some patients, loop diuretics may compromise renal hemodynamics and actually reduce GFR, a phenomenon known as diuretic intolerance. Importantly, A₁ receptor antagonists tend to improve GFR in the setting of diuretic intolerance. A₁ receptor antagonists such as rolofylline are in clinical trials. However, the future utility of this class of diuretics is doubtful: the large PROTECT trial showed in 2009 that rolofylline was not superior to placebo in patients with acute heart failure; rolofylline also increased the incidence of seizures, an expected side-effect of A₁ receptor antagonists.

Anatomy. The antidiuretic mechanism in mammals involves two anatomical components: a CNS component for synthesis, transport, storage, and release of vasopressin and a renal collecting-duct system composed of epithelial cells that respond to vasopressin by increasing their water permeability. The CNS component of the antidiuretic mechanism is called the hypothalamiconeurohypophyseal system and consists of neurosecretory neurons with perikarya located predominantly in two specific hypothalamic nuclei, the supraoptic nucleus (SON) and paraventricular nucleus (PVN). Long axons of magnocellular neurons in SON and PVN transverse the external zone of the median eminence to terminate in the neural lobe of the posterior pituitary (neurohypophysis), where they release vasopressin and oxytocin. In addition, axons of parvicellular neurons project to the external zone of the median eminence and release vasopressin directly into the pituitary portal circulation.

Synthesis. Vasopressin and oxytocin are synthesized mainly in the perikarya of magnocellular neurons in the SON and PVN; the two hormones are synthesized predominantly in separate neurons. Parvicellular neurons in the PVN also synthesize vasopressin. Vasopressin synthesis appears to be regulated solely at the transcriptional level. In humans, a 168-amino acid preprohormone (Figure 25–16) is synthesized, and a signal peptide (residues −23 to −1) ensures incorporation of the nascent polypeptide into ribosomes. During synthesis, the signal peptide is removed to form vasopressin prohormone, which then is processed and incorporated into the Golgi compartment and then into membrane-associated granules. The prohormone contains three domains: vasopressin (residues 1-9), vasopressin (VP)-neurophysin (residues 13-105), and VP-glycopeptide (residues 107-145). The vasopressin domain is linked to the VP-neurophysin domain through a glycine-lysine-arginine-processing signal, and the VP-neurophysin is linked to the VP-glycopeptide domain by an arginine-processing signal. In secretory granules, an endopeptidase, exopeptidase, monooxygenase, and lyase act sequentially on the prohormone to produce vasopressin, VP-neurophysin (sometimes referred to as neurophysin II or MSEL-neurophysin), and VP-glycopeptide (sometimes called copeptin). The synthesis and transport of vasopressin depend on the preprohormone conformation. In particular, VP-neurophysin binds vasopressin and is critical to correct processing, transport, and storage of vasopressin. Genetic mutations in either the signal peptide or VP-neurophysin give rise to central diabetes insipidus.

Transport and Storage. The process of axonal transport of vasopressin- and oxytocin-containing granules is rapid, and these hormone-laden granules arrive at their destinations within 30 minutes, ready for release by exocytosis when the magnocellular or parvicellular neurons are stimulated appropriately.

Maximal release of vasopressin occurs when impulse frequency is ~12 spikes per second for 20 seconds. Higher frequencies or longer periods of stimulation lead to diminished hormone release (fatigue). Appropriately, vasopressin-releasing cells demonstrate an atypical pattern of spike activity characterized by rapid phasic bursts (5-12 spikes per second for 15-60 seconds) separated by quiescent periods (15-60 seconds in duration). This pattern is orchestrated by activation and inactivation of ion channels in magnocellular neurons and provides for optimal vasopressin release.

Vasopressin Synthesis Outside the CNS. Vasopressin also is synthesized by heart and adrenal gland. In heart, elevated wall stress increases vasopressin synthesis severalfold. Cardiac synthesis of vasopressin is predominantly vascular and perivascular and may contribute to impaired ventricular relaxation and coronary vasoconstriction. Vasopressin synthesis in the adrenal medulla stimulates catecholamine secretion from chromaffin cells and may promote adrenal cortical growth and stimulate aldosterone synthesis.

Hyperosmolality. The relationship between plasma osmolality and plasma vasopressin concentration is shown in Figure 25–17A, and the relationship between plasma vasopressin levels and urine osmolality is illustrated in Figure 25–17B. The osmolality threshold for secretion is ~280 mOsm/kg. Below the threshold, vasopressin is barely detectable in plasma, and above the threshold, vasopressin levels are a steep and relatively linear function of plasma osmolality. A small increase in plasma osmolality leads to enhanced vasopressin secretion. Indeed, a 2% elevation in plasma osmolality causes a 2- to 3-fold increase in plasma vasopressin levels, which in turn causes increased solute-free water reabsorption, with an increase in urine osmolality. Increases in plasma osmolality above 290 mOsm/kg lead to an intense desire for water (thirst). Thus, the vasopressin system affords the organism longer thirst-free periods and, in the event that water is unavailable, allows the organism to survive longer periods of water deprivation. It is important to point out, however, that above a plasma osmolality of ~290 mOsm/kg, plasma vasopressin levels exceed 5 pM. Since urinary concentration is maximal (~1200 mOsm/kg) when vasopressin levels exceed 5 pM, further defense against hypertonicity depends entirely on water intake rather than on decreases in water loss.

Several CNS structures are involved in osmotic stimulation of vasopressin release by the posterior pituitary; these structures are collectively referred to as the osmoreceptive complex. Although magnocellular neurons in SON and PVN are osmosensitive, afferent inputs from other components of the osmoreceptive complex are required for a normal vasopressin response. The SON and PVN receive projections from the subfornical organ (SFO) and the organum vasculosum of the lamina terminalis (OVLT) either directly or indirectly by means of the median preoptic nucleus (MnPO). Subgroups of neurons in the SFO, OVLT, and MnPO are either osmoreceptors or osmoresponders (i.e., are stimulated by osmoreceptive neurons located at other sites). Thus, a web of interconnecting neurons contributes to osmotically induced vasopressin secretion.

Aquaporin 4, a water-selective channel, is associated with CNS structures involved in osmoregulation and may confer osmosensitivity. In the CNS, aquaporin 4 resides on glial and ependymal cells rather than on neurons, suggesting that osmotic status may be communicated to the neuronal cell by a glial-neuron interaction (Wells, 1998).

Hepatic Portal Osmoreceptors. An oral salt load activates hepatic portal osmoreceptors leading to increased vasopressin release. This mechanism augments plasma vasopressin levels even before the oral salt load increases plasma osmolality.

Hypovolemia and Hypotension. Vasopressin secretion also is regulated hemodynamically by changes in effective blood volume and/or arterial blood pressure (Robertson, 1992). Regardless of the cause (e.g., hemorrhage, Na⁺ depletion, diuretics, heart failure, hepatic cirrhosis with ascites, adrenal insufficiency, or hypotensive drugs), reductions in effective blood volume and/or arterial blood pressure may be associated with high circulating vasopressin concentrations. However, unlike osmoregulation, hemodynamic regulation of vasopressin secretion is exponential; i.e., small decreases (5%) in blood volume and/or pressure have little effect on vasopressin secretion, whereas larger decreases (20-30%) can increase vasopressin levels to 20-30 times normal (exceeding the vasopressin concentration required to induce maximal antidiuresis). Vasopressin is one of the most potent vasoconstrictors known, and the vasopressin response to hypovolemia or hypotension serves as a mechanism to stave off cardiovascular collapse during periods of severe blood loss and/or hypotension. Hemodynamic regulation of vasopressin secretion does not disrupt osmotic regulation; rather, hypovolemia/hypotension alters the set point and slope of the plasma osmolality-plasma vasopressin relationship (Figure 25–18).

Neuronal pathways that mediate hemodynamic regulation of vasopressin release are different from those involved in osmoregulation. Baroreceptors in left atrium, left ventricle, and pulmonary veins sense blood volume (filling pressures), and baroreceptors in carotid sinus and aorta monitor arterial blood pressure. Nerve impulses reach brainstem nuclei predominantly through the vagal trunk and glossopharyngeal nerve; these signals are relayed to the solitary tract nucleus, then to the A₁-noradrenergic cell group in the caudal ventrolateral medulla, and finally to the SON and PVN.

Hormones and Neurotransmitters. Vasopressin-synthesizing magnocellular neurons have a large array of receptors on both perikarya and nerve terminals; therefore, vasopressin release can be accentuated or attenuated by chemical agents acting at both ends of the magnocellular neuron. Also, hormones and neurotransmitters can modulate vasopressin secretion by stimulating or inhibiting neurons in nuclei that project, either directly or indirectly, to the SON and PVN. Because of these complexities, results of any given investigation may depend critically on route of administration of the agent and on the experimental paradigm. In many cases, the precise mechanism by which a given agent modulates vasopressin secretion is either unknown or controversial, and the physiological relevance of the modulation of vasopressin secretion by most hormones and neurotransmitters is unclear.

Nonetheless, several agents are known to stimulate vasopressin secretion, including acetylcholine (by nicotinic receptors), histamine (by H₁ receptors), dopamine (by both D₁ and D₂ receptors), glutamine, aspartate, cholecystokinin, neuropeptide Y, substance P, vasoactive intestinal polypeptide, prostaglandins, and angiotensin II (AngII). Inhibitors of vasopressin secretion include atrial natriuretic peptide, γ-aminobutyric acid, and opioids (particularly dynorphin via κ receptors). The affects of AngII have received the most attention. AngII, applied directly to magnocellular neurons in the SON and PVN, increases neuronal excitability; when applied to the MnPO, AngII indirectly stimulates magnocellular neurons in the SON and PVN. In addition, AngII stimulates angiotensin-sensitive neurons in the OVLT and SFO (circumventricular nuclei lacking a blood-brain barrier) that project to the SON/PVN. Thus, AngII synthesized in the brain and circulating AngII may stimulate vasopressin release. Inhibition of the conversion of AngII to AngIII blocks AngII-induced vasopressin release, suggesting that AngIII is the main effector peptide of the brain renin-angiotensin system controlling vasopressin release (Reaux et al., 2001).

Pharmacological Agents. A number of drugs alter urine osmolality by stimulating or inhibiting vasopressin secretion. In some cases the mechanism by which a drug alters vasopressin secretion involves direct effects on one or more CNS structures that regulate vasopressin secretion. In other cases vasopressin secretion is altered indirectly by the effects of a drug on blood volume, arterial blood pressure, pain, or nausea. In most cases the mechanism is not known. Stimulators of vasopressin secretion include vincristine, cyclophosphamide, tricyclic antidepressants, nicotine, epinephrine, and high doses of morphine. Lithium, which inhibits the renal effects of vasopressin, also enhances vasopressin secretion. Inhibitors of vasopressin secretion include ethanol, phenytoin, low doses of morphine, glucocorticoids, fluphenazine, haloperidol, promethazine, oxilorphan, and butorphanol. Carbamazepine has a renal action to produce antidiuresis in patients with central diabetes insipidus but actually inhibits vasopressin secretion by a central action.

For maximum concentration of urine, large amounts of urea must be deposited in the interstitium of the inner medulla. It is not surprising, therefore, that V₂-receptor activation also increases urea permeability by 400% in the terminal portions of the inner medullary collecting duct. V₂ receptors increase urea permeability by activating a vasopressin-regulated urea transporter (termed VRUT, UT1, or UT-A1), most likely by PKA-induced phosphorylation (Sands, 2003). Kinetics of vasopressin-induced water and urea permeability differ, and vasopressin-induced regulation of VRUT does not entail vesicular trafficking to the plasma membrane (Inoue et al., 1999).

In addition to increasing water permeability of the collecting duct and urea permeability of the inner medullary collecting duct, V₂-receptor activation also increases Na⁺ transport in thick ascending limb and collecting duct. Increased Na⁺ transport in thick ascending limb is mediated by three mechanisms that affect the Na⁺-K⁺-2C1⁻ symporter: rapid phosphorylation of the symporter, translocation of the symporter into the luminal membrane, and increased expression of symporter protein (Ecelbarger et al., 2001). Enhanced Na⁺ transport in collecting duct is mediated by increased expression of subunits of the epithelial Na⁺ channel (Ecelbarger et al., 2001). The multiple mechanisms by which vasopressin increases water reabsorption are summarized in Figure 25–22.

The collecting-duct system is critical for water conservation. By the time tubular fluid arrives at the cortical collecting duct, it has been rendered hypotonic by the upstream diluting segments of the nephron that reabsorb NaCl without reabsorbing water. In the well-hydrated subject, plasma osmolality is in the normal range, concentrations of vasopressin are low, the entire collecting duct is relatively impermeable to water, and the urine is dilute. Under conditions of dehydration, plasma osmolality is increased, concentrations of vasopressin are elevated, and the collecting duct becomes permeable to water. The osmotic gradient between dilute tubular urine and hypertonic renal interstitial fluid (which becomes progressively more hypertonic in deeper regions of the renal medulla) provides for osmotic flux of water out of the collecting duct. The final osmolality of urine may be as high as 1200 mOsm/kg in humans, and a significant saving of solute-free water thus is possible.

Other renal actions mediated by V₂ receptors include increased urea transport in inner medullary collecting duct and increased Na⁺ transport in thick ascending limb; both effects contribute to the urine-concentrating ability of the kidney (Figure 25–22). V₂ receptors also increase Na⁺ transport in cortical collecting duct (Ecelbarger et al., 2001), and this may synergize with aldosterone to enhance Na⁺ reabsorption during hypovolemia.

Nonrenal Actions of Vasopressin. Vasopressin and related peptides are ancient hormones in evolutionary terms, and they are found in species that do not concentrate urine. Thus, it is not surprising that vasopressin has nonrenal actions in mammals.

Cardiovascular System. The cardiovascular effects of vasopressin are complex, and vasopressin's role in physiological situations is ill-defined. Vasopressin is a potent vasoconstrictor (V₁ receptor-mediated), and resistance vessels throughout the circulation may be affected. Vascular smooth muscle in the skin, skeletal muscle, fat, pancreas, and thyroid gland appear most sensitive, with significant vasoconstriction also occurring in GI tract, coronary vessels, and brain. Despite the potency of vasopressin as a direct vasoconstrictor, vasopressin-induced pressor responses in vivo are minimal and occur only with vasopressin concentrations significantly higher than those required for maximal antidiuresis. To a large extent, this is due to circulating vasopressin actions on V₁ receptors to inhibit sympathetic efferents and potentiate baroreflexes. In addition, V₂ receptors cause vasodilation in some blood vessels.

A large body of data supports the conclusion that vasopressin helps to maintain arterial blood pressure during episodes of severe hypovolemia/hypotension. At present, there is no convincing evidence for a role of vasopressin in essential hypertension in humans (Kawano et al., 1997).

The effects of vasopressin on heart (reduced cardiac output and heart rate) are largely indirect and result from coronary vasoconstriction, decreased coronary blood flow, and alterations in vagal and sympathetic tone. In humans, vasopressin effects on coronary blood flow can be demonstrated easily, especially if large doses are employed. The cardiac actions of the hormone are of more than academic interest. Some patients with coronary insufficiency experience angina even in response to the relatively small amounts of vasopressin required to control diabetes insipidus, and vasopressin-induced myocardial ischemia has led to severe reactions and even death.

CNS. It is likely that vasopressin plays a role as a neurotransmitter and/or neuromodulator. Vasopressin may participate in the acquisition of certain learned behaviors, in the development of some complex social processes, and in the pathogenesis of specific psychiatric diseases such as depression. However, the physiological/ pathophysiological relevance of these findings is controversial, and some actions of vasopressin on memory and learned behavior may be due to visceral autonomic effects. Many studies support a physiological role for vasopressin as a naturally occurring antipyretic factor. Although vasopressin can modulate CNS autonomic systems controlling heart rate, arterial blood pressure, respiration rate, and sleep patterns, the physiological significance of these actions is unclear. Finally, ACTH secretion is enhanced by vasopressin released from parvicellular neurons in the PVN and secreted into pituitary portal capillaries from axon terminals in the median eminence. Although vasopressin is not the principal corticotropin-releasing factor, vasopressin may provide for sustained activation of the hypothalamic-pituitary-adrenal axis during chronic stress. CNS effects of vasopressin appear to be mediated predominantly by V₁ receptors.

Blood Coagulation. Activation of V₂ receptors by desmopressin or vasopressin increases circulating levels of procoagulant factor VIII and of von Willebrand factor. These effects are mediated by extrarenal V₂ receptors (Bernat et al., 1997). Presumably, vasopressin stimulates secretion of von Willebrand factor and of factor VIII from storage sites in vascular endothelium. However, since release of von Willebrand factor does not occur when desmopressin is applied directly to cultured endothelial cells or to isolated blood vessels, intermediate factors are likely to be involved.

Other Nonrenal Effects of Vasopressin. At high concentrations, vasopressin stimulates smooth muscle contraction in uterus (by oxytocin receptors) and GI tract (by V₁ receptors). Vasopressin is stored in platelets, and activation of V₁ receptors stimulates platelet aggregation. Also, activation of V₁ receptors on hepatocytes stimulates glycogenolysis. The physiological significance of these effects of vasopressin in not known.

Many vasopressin analogs were synthesized with the goal of increasing duration of action and selectivity for vasopressin receptor subtypes (V₁ versus V₂ vasopressin receptors, which mediate pressor responses and antidiuretic responses, respectively). Deamination at position 1 increases duration of action and increases antidiuretic activity without increasing vasopressor activity. Substitution of D-arginine for L-arginine greatly reduces vasopressor activity without reducing antidiuretic activity. Thus, the antidiuretic-to-vasopressor ratio for 1-deamino-8-D-arginine vasopressin (Table 25–8), also called desmopressin (ddavp, minirin, stimate, others), is ~3000 times greater than that for vasopressin, and desmopressin now is the preferred drug for the treatment of central diabetes insipidus. Substitution of valine for glutamine in position 4 further increases the antidiuretic selectivity, and the antidiuretic to vasopressor ratio for deamino [Val⁴, D-Arg⁸]AVP (Table 25–8) is ~11,000 times greater than that for vasopressin. Nakamura and colleagues (2000) synthesized a nonpeptide V₂-receptor agonist (Table 25–8).

Increasing V₁ selectivity has proved more difficult than increasing V₂ selectivity, but a limited number of agonists with modest selectivity for V₁ receptors have been developed (Table 25–8). Vasopressin receptors in the adenohypophysis that mediate vasopressin-induced ACTH release are neither classical V₁ nor V₂ receptors. Since vasopressin receptors in the adenohypophysis appear to share a common signal-transduction mechanism with classical V₁ receptors, and since many vasopressin analogs with vasoconstrictor activity release ACTH, V₁ receptors have been subclassified into V_1a (vascular/hepatic) and V_1b (pituitary) receptors. V_1b receptors are also called V₃ receptors. Vasopressin analogs that are agonists selective for V_1a or V_1b receptors have been described (Table 25–8).

Chemistry of Vasopressin Receptor Antagonists. The impetus for the development of specific vasopressin receptor antagonists is the belief that such drugs may be useful in a number of clinical settings. Based on receptor physiology, selective V_1a antagonists may be beneficial when total peripheral resistance is increased (e.g., congestive heart failure and hypertension), whereas selective V₂ antagonists could be useful whenever reabsorption of solute-free water is excessive (e.g., the syndrome of inappropriate secretion of antidiuretic hormone [SIADH] and hyponatremia associated with a reduced effective blood volume). Combined V_1a/V₂ receptor antagonists might be beneficial in diseases associated with a combination of increased peripheral resistance and dilutional hyponatremia (e.g., congestive heart failure).

Highly selective V₁ and V₂ peptide antagonists that are structural analogs of vasopressin have been synthesized (see Table 25–9 for examples), including both cyclic and linear peptides. [1-(β-Mercapto-β, β-cyclopentamethyleneproprionic acid), 2-O-methyltyrosine] arginine vasopressin, also known as d(CH₂)₅[Tyr(Me)²] AVP, has a greater affinity for V_1a receptors than for either V_1b or V₂ receptors; this antagonist has been employed widely in physiological and pharmacological studies. Although [1-deaminopenicillamine, 2-O-methyltyrosine] arginine vasopressin, also called dP[Tyr(Me)²] AVP, is a potent V_1b receptor antagonist with little affinity for the V₂ receptor, it also blocks V_1a receptors. No truly selective peptide V_1b-receptor antagonist is available. Peptide antagonists have limited oral activity, and the potency of peptide V₂ antagonists is species dependent.

The urinary taste test for DI has been supplanted by the approach of observing whether the patient is able to reduce urine volume and increase urine osmolality after a period of carefully observed fluid deprivation. Central DI can be distinguished from nephrogenic DI by administration of desmopressin, which will increase urine osmolality in patients with central DI but have little or no effect in patients with nephrogenic DI. DI can be differentiated from primary polydipsia by measuring plasma osmolality, which will be low to low-normal in patients with primary polydipsia and high to high-normal in patients with DI. For a more complete discussion of diagnostic procedures, see Robertson (2001).

Antidiuretic peptides are the primary treatment for central DI, with desmopressin being the peptide of choice. For patients with central DI who cannot tolerate antidiuretic peptides because of side effects or allergic reactions, other treatment options are available. Chlorpropamide, an oral sulfonylurea, potentiates the action of small or residual amounts of circulating vasopressin and will reduce urine volume in more than half of all patients with central DI. Doses of 125-500 mg daily appear effective in patients with partial central DI. If polyuria is not controlled satisfactorily with chlorpropamide alone, addition of a thiazide diuretic to the regimen usually results in an adequate reduction in urine volume. Carbamazepine (800-1000 mg daily in divided doses) also reduces urine volume in patients with central DI. Long-term use may induce serious adverse effects; therefore, carbamazepine is used rarely to treat central DI. The antidiuretic mechanisms of chlorpropamide, carbamazepine, and clofibrate (not available in the U.S.) are not clear. These agents are not effective in nephrogenic DI, which indicates that functional V₂ receptors are required for the antidiuretic effect. Since carbamazepine inhibits and chlorpropamide has little effect on vasopressin secretion, it is likely that carbamazepine and chlorpropamide act directly on the kidney to enhance V₂-receptor-mediated antidiuresis.

X-linked nephrogenic DI is caused by mutations in the gene encoding the V₂ receptor, which maps to Xq28. A number of missense, nonsense, and frame-shift mutations in the gene encoding the V₂ receptor were identified in patients with this disorder. Mutations in the V₂-receptor gene may cause impaired routing of the V₂ receptor to the cell surface, defective coupling of the receptor to G proteins, or decreased receptor affinity for vasopressin. The effects of these mutations range from a complete loss of responsiveness to vasopressin to a shift to the left of the concentration-response curve. Autosomal recessive and dominant nephrogenic DI result from inactivating mutations in aquaporin 2. These findings indicate that aquaporin 2 is essential for the antidiuretic effect of vasopressin in humans.

Although the mainstay of treatment of nephrogenic DI is assurance of an adequate water intake, drugs also can be used to reduce polyuria. Amiloride blocks Li⁺ uptake by the Na⁺ channel in the collecting-duct system and may be effective in patients with mild to moderate concentrating defects. Patients with severe disease often do not respond. Thiazide diuretics reduce the polyuria of patients with DI and often are used to treat nephrogenic DI. The use of thiazide diuretics in infants with nephrogenic DI may be crucially important because uncontrolled polyuria may exceed the child's capacity to imbibe and absorb fluids. The antidiuretic mechanism of thiazides in DI is incompletely understood. It is possible that the natriuretic action of thiazides and resulting extracellular fluid volume depletion play an important role in thiazide-induced antidiuresis. In this regard, whenever extracellular fluid volume is reduced, compensatory mechanisms increase NaCl reabsorption in proximal tubule, reducing the volume delivered to the distal tubule. Consequently, less free water can be formed, which should diminish polyuria. However, studies in rats with vasopressin-deficient DI challenge this hypothesis. Nonetheless, the antidiuretic effects appear to parallel the thiazide's ability to cause natriuresis, and the drugs are given in doses similar to those used to mobilize edema fluid. In patients with DI, a 50% reduction of urine volume is a good response to thiazides. Moderate restriction of Na⁺ intake can enhance the antidiuretic effectiveness of thiazides.

A number of case reports describe the effectiveness of indomethacin in the treatment of nephrogenic DI; however, other prostaglandin synthase inhibitors (e.g., ibuprofen) appear to be less effective. The mechanism of the effect may involve a decrease in glomerular filtration rate, an increase in medullary solute concentration, and/or enhanced proximal fluid reabsorption. Also, since prostaglandins attenuate vasopressin-induced antidiuresis in patients with at least a partially intact V₂-receptor system, some of the anti-diuretic response to indomethacin may be due to diminution of the prostaglandin effect and enhancement of vasopressin effects on the principal cells of collecting duct.

Although Li⁺ can inhibit the renal actions of vasopressin, it is effective in only a minority of patients, may induce irreversible renal damage when used chronically, and has a low therapeutic index. Therefore, Li⁺ should be considered for use only in patients with symptomatic SIADH who cannot be controlled by other means or in whom tetracyclines are contraindicated (e.g., patients with liver disease). It is important to stress that the majority of patients with SIADH do not require therapy because plasma Na⁺ stabilizes in the range of 125-132 mM; such patients usually are asymptomatic. Only when symptomatic hypotonicity ensues, generally when plasma Na⁺ levels drop below 120 mM, should therapy with demeclocycline be initiated. Since hypotonicity, which causes an influx of water into cells with resulting cerebral swelling, is the cause of symptoms, the goal of therapy is simply to increase plasma osmolality toward normal. For a more complete description of the diagnosis and treatment of SIADH, see Robertson (2001).

Other Water-Retaining States. In patients with congestive heart failure, cirrhosis, or nephrotic syndrome, effective blood volume often is reduced, and hypovolemia frequently is exacerbated by the liberal use of diuretics. Since hypovolemia stimulates vasopressin release, patients may become hyponatremic owing to vasopressin-mediated retention of water. The development of potent orally active V₂ receptor antagonists and specific inhibitors of water channels in the collecting duct has provided a new therapeutic strategy not only in patients with SIADH but also in the more common setting of hyponatremia in patients with heart failure, liver cirrhosis, and nephrotic syndrome, as discussed in the next section.

The applications of V₁ receptor agonists can be accomplished with vasopressin; however, the use of vasopressin for all these indications is no longer recommended because of significant adverse reactions. Terlipressin (lucassin) is preferred for bleeding esophageal varices because of increased safety compared with vasopressin (Vlavianos and Westaby, 2001) and is designated as an orphan drug for this use. Moreover, terlipressin is effective in patients with hepatorenal syndrome, particularly when combined with albumin (Ortega et al., 2002). Terlipressin has been granted priority review, orphan drug status, and fast-track designation by the FDA for type I hepatorenal syndrome.

Vasopressin levels in patients with vasodilatory shock are inappropriately low, and such patients are extraordinarily sensitive to the pressor actions of vasopressin. The combination of vasopressin and norepinephrine is superior to norepinephrine alone in the management of catecholamine-resistant vasodilatory shock. However, recent clinical trials show that, in comparison to catecholamines alone, addition of vasopressin does not improve outcomes in either cardiac arrest or septic shock.

V₂ receptor-mediated therapeutic applications are based on the rationale that V₂ receptors cause water conservation and release of blood coagulation factors. Central but not nephrogenic DI can be treated with V₂ receptor agonists, and polyuria and polydipsia usually are well controlled by these agents. Some patients experience transient DI (e.g., in head injury or surgery in the area of the pituitary); however, therapy for most patients with DI is lifelong. Desmopressin is the drug of choice for the vast majority of patients, and numerous clinical trials demonstrated the efficacy and tolerability of desmopressin in both adults and children.

The duration of effect from a single intranasal dose is from 6-20 hours; twice-daily administration is effective in most patients. There is considerable variability in the intranasal dose of desmopressin required to maintain normal urine volume, and the dosage must be tailored individually. The usual intranasal dosage in adults is 10-40 μg daily either as a single dose or divided into two or three doses. In view of the high cost of the drug and the importance of avoiding water intoxication, the schedule of administration should be adjusted to the minimal amount required. An initial dose of 2.5 μg can be used, with therapy first directed toward control of nocturia. An equivalent or higher morning dose controls daytime polyuria in most patients, although a third dose occasionally may be needed in the afternoon. In some patients, chronic allergic rhinitis or other nasal pathology may preclude reliable peptide absorption following nasal administration. Oral administration of desmopressin in doses 10-20 times the intranasal dose provides adequate desmopressin blood levels to control polyuria. Subcutaneous administration of 1-2 μg daily of desmopressin also is effective in central DI.

Vasopressin has little, if any, place in the long-term therapy of DI because of its short duration of action and V₁ receptor-mediated side effects. Vasopressin can be used as an alternative to desmopressin in the initial diagnostic evaluation of patients with suspected DI and to control polyuria in patients with DI who recently have undergone surgery or experienced head trauma. Under these circumstances, polyuria may be transient, and long-acting agents may produce water intoxication.

An additional V₂ receptor-mediated therapeutic application is the use of desmopressin in bleeding disorders (Mannucci, 1997). In most patients with type I von Willebrand's disease (vWD) and in some with type IIn vWD, desmopressin will elevate von Willebrand factor and shorten bleeding time. However, desmopressin generally is ineffective in patients with types IIa, IIb, and III vWD. Desmopressin may cause a marked transient thrombocytopenia in individuals with type IIb vWD and is contraindicated in such patients. Desmopressin also increases factor VIII levels in patients with mild to moderate hemophilia A. Desmopressin is not indicated in patients with severe hemophilia A, those with hemophilia B, or those with factor VIII antibodies. Using a test dose of nasal spray, the response of any given patient with type I vWD or hemophilia A to desmopressin should be determined at the time of diagnosis or 1-2 weeks before elective surgery to assess the extent of increase in factor VIII or von Willebrand factor. Desmopressin is employed widely to treat the hemostatic abnormalities induced by uremia. In patients with renal insufficiency, desmopressin shortens bleeding time and increases circulating levels of factor VIII coagulant activity, factor VIII-related antigen, and ristocetin cofactor. It also induces the appearance of larger von Willebrand factor multimers. Desmopressin is effective in some patients with liver cirrhosis-induced or drug-induced (e.g., heparin, hirudin, and antiplatelet agents) bleeding disorders. Desmopressin, given intravenously at a dose of 0.3 μg/kg, increases factor VIII and von Willebrand factor for > 6 hours. Desmopressin can be given at intervals of 12-24 hours depending on the clinical response and severity of bleeding. Tachyphylaxis to desmopressin usually occurs after several days (owing to depletion of factor VIII and von Willebrand factor storage sites) and limits its usefulness to preoperative preparation, postoperative bleeding, excessive menstrual bleeding, and emergency situations.

Inactivation by peptidases in various tissues (particularly liver and kidney) results in a plasma t_1/2 of vasopressin of 17-35 minutes. Following intramuscular or subcutaneous injection, antidiuretic effects of vasopressin last 2-8 hours. The plasma t_1/2 of desmopressin has two components, a fast component (~8 min) and a slow component (30-117 min). Only 3% and 0.15%, respectively, of intranasally and orally administered desmopressin is absorbed.

After injection of large doses of vasopressin, marked facial pallor owing to cutaneous vasoconstriction is observed commonly. Increased intestinal activity is likely to cause nausea, belching, cramps, and an urge to defecate. Most serious, however, is the effect on the coronary circulation. Vasopressin should be administered only at low doses and with extreme caution in individuals suffering from vascular disease, especially coronary artery disease. Other cardiac complications include arrhythmia and decreased cardiac output. Peripheral vasoconstriction and gangrene were encountered in patients receiving large doses of vasopressin.

Mild facial flushing and headache are the most common adverse effects associated with desmopressin. Allergic reactions ranging from urticaria to anaphylaxis may occur with desmopressin or vasopressin. Intranasal administration may cause local adverse effects in the nasal passages, such as edema, rhinorrhea, congestion, irritation, pruritus, and ulceration.

Mozavaptan. Mozavaptan was the first nonpeptide V₂ receptor antagonist in clinical use. It was approved for treatment of SIADH secondary to cancer in Japan in 1996. Mozavaptan causes a dose-dependent increase in free water excretion with only slight increases in urinary Na⁺ and K⁺ excretion.

Pharmacokinetics

Mozavaptan. Mozavaptan is available in 30-mg tablets. Few data are available regarding its pharmacokinetics. It binds to the V₂R with much higher affinity than V_1aR, K_i 9.42 ± 0.9 nM versus 150 ± 15 nM, respectively. Its peak effects occur 60-90 minutes after an oral dose.

Tolvaptan. Tolvaptan is available in 15 and 30 mg tablets and inhibits the V₂R with a K_i of 0.43 ± 0.06 nM and the V_1aR with a K_i of 12.3 ± 0.8 nM. Tolvaptan is 29 times more selective for the V₂R than the V_1aR. Most of its effects on renal free water excretion occur in the first 6-8 hours after administration. Tolvaptan has a t_1/2 of 6-8 hours and <1% is excreted in the urine. The starting dose is 15 mg once daily, and this dose can be titrated up to a maximum of 60 mg daily. Tolvaptan is a substrate and inhibitor of P-glycoprotein and is eliminated entirely by CYP3A metabolism.

Conivaptan. Conivaptan inhibits the V_1aR with a K_i of 6.3 ± 1.8 nM and the V₂R with a K_i of 1.1 ± 0.2 nM. Total body clearance is 9.5-10.1 L/hr. It is highly protein bound and has a terminal elimination t_1/2 of 5-12 hours. Conivaptan is metabolized via CYP3A4 and is partially excreted by the kidney. Caution should be exercised in those with hepatic and renal disease. At higher doses clearance may be reduced in the elderly.

Toxicity, Adverse Effects, Contraindications, Drug Interactions

Mozavaptan. Mozavaptan's major side effects are dry mouth and abnormal liver function tests.

Tolvaptan. Tolvaptan is eliminated by CYP3A4. Plasma concentrations of the drug are increased by ketoconazole. There is no known affect on amiodarone, digoxin, or warfarin metabolism. Among the adverse effects listed for tolvaptan are GI effects, hyperglycemia, and pyrexia. Less common are cerebrovascular accident, deep vein thrombosis, disseminated intravascular coagulation, intracardiac thrombus, ventricular fibrillation, urethral hemorrhage, vaginal hemorrhage, pulmonary embolism, respiratory failure, diabetic ketoacidosis, ischemic colitis, increaese in prothrombin time and rhabdomyolysis.

Conivaptan. Conivaptan is metabolized by CYP3A4 and as a result is associated with a variety of drug-drug interactions. It should not be administered in patients receiving ketoconazole, itraconazole, ritonavir, indinavir, clarithromycin, or other strong CYP3A4 inhibitors. Conivaptan increases levels of simvastatin, digoxin, amlodipine, and midazolam. It is not known to interact with warfarin or prolong the QT_c interval. Due to multiple drug-drug interactions conivaptan is no longer being developed for chronic long-term oral use. The most common adverse effect of conivaptan is an infusion site reaction. The manufacturer recommends that the drug only be infused into large veins and that infusion sites be changed daily. Other adverse effects have included headache, hypertnesion, hypotension, hypokalemia, and pyrexia.