Physiology_Levy

Physiology_Levy. To understand what determines the magnitude of the Vm, it is important to recognize that any transporter that transfers charge across the membrane has the potential to influence the Vm. Such transporters are said to be fIn muscle cells, in which contraction is regulated by the intracellular [Ca++], the maintenance of a low intracellular [Ca++] during the relaxed state involves not only the activity of the plasma membrane 3Na+-Ca++ antiporter and the Ca++-ATPase but also a Ca++-ATPase molecule located in the smooth endoplasmic reticulum (see


Physiology_Levy

Physiology_Levy. Chapters 12 to 14 electrogenic. As might be expected, the contribution of various electrogenic transporters to the Vm is highly variable from cell to cell. For example, the Na+,K+-ATPase channel transports three Na+ and two K+ ions and thus transfers one net positive charge across the membrane. However, the direct contribution of the Na+,K+-ATPase to the Vm of most cells is only a few millivolts at the most. Similarly, the contribution of other electrogenic transporters, such as the 3Na+-Ca++ antiporter and the Na+-glucose symporter is minimal. The major determinants of the Vm are ion channels. The type (e.g., selectivity), number, and activity (e.g., gating) of these channels determine the magnitude of the Vm. As described in , rapid changes in ion channel activity underlies the action potential in neurons and other excitable cells, such as those of skeletal and cardiac muscle (see


Physiology_Levy

Physiology_Levy. As ions move across the membrane through a channel, they generate a current. As described in , this current can be measured, even at the level of a single channel. By convention, the current generated by the movement of cations into the cell, or the movement of anions out of the cell, is defined as negative current. Conversely, the movement of cations out of the cell, or the movement of anions into the cell, is defined as positive current. Also by convention, the magnitude of the Vm is expressed in relation to the outside of the cell; thus for a cell with a Vm of −80 mV, the interior of the cell is electrically negative in relation to the outside of the cell. The current carried by ions moving through a channel depends on the driving force for that ion and on the conductance of the channel. As described in , the driving force is determined by the energy in the concentration gradient for the ion across the membrane (Ei), as calculated by the Nernst equation (Eq. 1.5a ) and the Vm:


Physiology_Levy

Physiology_Levy. Equation 2.3 Driving force = Vm − Ei. Thus as defined by Ohm’s law, the ion current through the channel (Ii) is determined as follows: Equation 2.4 where gi is the conductance of the channel. For a cell, the conductance of the membrane to a particular ion (Gi) is determined by the number of ion channels in the membrane and by the amount of time each channel is in the open state. Fig. 2.4 , the Vm is the voltage at which there is no net ion flow into or out of the cell. Thus for a cell that has ion channels selective for Na+ , K+ , and Cl− , Equation 2.5 Equation 2.6 + [(Vm − E − )× G − ]= 0.


Physiology_Levy

Physiology_Levy. Equation 2.5 Equation 2.6 + [(Vm − E − )× G − ]= 0. •Fig. 2.4 Current-Voltage Relationship of a Hypothetical Cell Containing Na+-, K+-, and Cl−-Selective Channels. Membranecurrentsareplottedoverarangeofmembranevoltages(i.e.,current-voltagerelationships).EachioncurrentiscalculatedwiththeuseofOhm’slaw,theNernstequilibriumpotentialfortheion(ECl,EK,andENa),andthemembraneconductancefortheion.Thecurrent-voltagerelationshipforthewholecellisalsoshown.Totalcellcurrent(Icell)wascalculatedwiththechordconductanceequation(see Eq.2.7).Because80%of cellconductanceisduetoK+ ,therestingmembranevoltage(Vm)of−64.4mVisneartothatoftheNernstequilibriumpotentialforK+ . Equation 2.7 where ˜G = G ++ G ++ G −. Inspection of Eq. 2.7 , which is often called the chord conductance equation, reveals that the Vm will be near to the Nernst equilibrium potential of the ion to which the membrane has the highest conductance. In


Physiology_Levy

Physiology_Levy. Eq. 2.7 , which is often called the chord conductance equation, reveals that the Vm will be near to the Nernst equilibrium potential of the ion to which the membrane has the highest conductance. In Fig. 2.4 , 80% of the membrane conductance is attributable to K+; as a result, Vm is near to the Nernst equilibrium potential for K+ (EK+). For most cells at rest, the membrane has a high conductance to K+ , and thus the Vm approximates EK+. Moreover, the Vm is greatly influenced by the magnitude of EK+ , which in turn is greatly influenced by changes in the [K+] of the ECF. For example, if the intracellular [K+] is 120 mEq/L and the extracellular [K+] is 4 mEq/L, EK+ has a value of −90.8 mV. If the extracellular [K+] is increased to 7 mEq/L, EK+ would be −79.9 mV. This change in EK+ depolarizes the Vm (i.e., Vm is less negative). Conversely, if the extracellular [K+] is decreased to 2 mEq/L, EK+ becomes −109.4 mV, and the Vm hyperpolarizes (i.e., Vm is more negative).


Physiology_Levy

Physiology_Levy. •Fig. 2.5 Nerve Action Potential Showing the Changes in Na+ and K+ Conductances ( G and GK+ , Respectively) and the Membrane Potential (Vm). Atrest,themembranehasahighK+ conductance,andVmisneartheNernstequilibriumpotentialforK+ (EK+ ).Withtheinitiationoftheactionpotential,thereisalargeincreaseintheNa+ conductanceofthemembrane,andtheVmapproachestheNernstequilibriumpotentialforNa+ (ENa+ ).TheincreaseinNa+ conductanceistransient,andtheK+ conductancethenincreasesaboveitsvaluebeforetheactionpotential.ThishyperpolarizesthecellasVmapproachesEK+.AstheK+ conductancereturnstoitsbaselinevalue,Vmreturnstoitsrestingvalueof−70mV.(ModifiedfromLevyMN,KoeppenBM,StantonBA.Berne & Levy’s Principles of Physiology. 4thed.St.Louis:Mosby;2006.)


Physiology_Levy

Physiology_Levy. Changesintheextracellular[K+]canhaveimportanteffectsonexcitablecells,especiallythoseoftheheart.Adecreaseinextracellular[K+](hypokalemia) hyperpolarizestheVmofcardiacmyocytesand,insodoing,makesinitiatinganactionpotentialmoredifficult,becausealargerdepolarizingcurrentisneededtoreachthreshold(see ).Ifsevere,hypokalemiacanleadtocardiacarrhythmias,andeventuallytheheartcanstopcontracting(asystole). Anincreaseintheextracellular[K+](hyperkalemia) canbeequallydeleterioustocardiacfunction.Withhyperkalemia,theVmisdepolarized,anditiseasiertoinitiateanactionpotential.However,oncetheactionpotentialfiresthechannelsbecomeinactivated,andareunabletoinitiateanotheractionpotential,untiltheyarereactivatedbynormalrepolarizationoftheVm.BecausetheVmisdepolarizedinhyperkalemia,thechannelsstayinaninactivatedstate.ThusdepolarizationoftheVmwithhyperkalemiacanleadtocardiacarrhythmiasandlossofcardiacmusclecontraction. Eq. 2.7 also defines the limits for the membrane potential. In the example depicted in


Physiology_Levy

Physiology_Levy. Eq. 2.7 also defines the limits for the membrane potential. In the example depicted in Fig. 2.4, it is apparent that the Vm cannot be more negative than EK+ (−90.8 mV), as would be the case if the membrane were only conductive to K+ . Conversely, the Vm could not be more positive than ENa+ (66.6 mV); such a condition would be met if the membrane were conductive only to Na+ . The dependence of the Vm on the conductance of the membrane to specific ions is the basis by which action potentials in excitable cells are generated ( Fig. 2.5). As noted previously, in all excitable cells, the CHAPTER 2 Homeostasis: Volume and Composition of Body Fluid Compartments membrane at rest is conductive predominantly to K+ , and thus Vm is near EK+. When an action potential is initiated, Na+-channels open and the membrane is now conductive predominantly to Na+ . As a result, Vm now approaches ENa+. The generation of action potentials is discussed in more detail in


Physiology_Levy

Physiology_Levy. TheestablishmentoftheVmrequirestheseparationofchargeacrosstheplasmamembrane.However,thenumberofionsthatmustmoveacrossthemembraneisatinyfractionofthetotalnumberofionsinthecell.Forexample,considerasphericalcellwithadiameterof20µmandaVmof−80mV.Furthermore,assumethatthisVmof−80mVistheresultofthediffusionofK+ outofthecellandthattheintracellular[K+]is120mmol/L.TheamountofK+ thatwouldhavetodiffuseoutofthecelltoestablishtheVmof−80mVisthencalculatedasfollows. Firstthechargeseparationacrossthemembraneneedstobecalculated.Thisisdonewiththeknowledgethattheplasmamembranebehaveselectricallylikeacapacitor,thecapacitance(C)ofwhichisapproximately1µF/cm2,and whereQ= chargeandisexpressedinunitsofcoulombs.Ifthesurfaceareaofthecellis4πr2or1.26× 10−5cm2,thecapacitanceofthecelliscalculatedasfollows: 1× 10 F/cm × . × 10 cm = . × 10 126 126 F. Thusthechargeseparationacrossthemembraneiscalculatedasfollows: 1 01× 10−12 coulombs.


Physiology_Levy

Physiology_Levy. Thusthechargeseparationacrossthemembraneiscalculatedasfollows: 1 01× 10−12 coulombs. Because1moleofK+ contains96,480coulombs,theamountofK+ thathadtodiffuseacrossthemembranetoestablishtheVmof−80mViscalculatedasfollows: . coul = 1 05. × 10 mole ofK , ombs/mol Withacellvolumeof4.19× 10−12L(volume= 4πr3/3)andanintracellular[K+]of120mmol/L,thetotalintracellularK+ contentis 4 19 × 10 × . mol/L = 5 03 . 012 . × 10 moles Therefore,thediffusionof1.05× 10−17molesofK+ outofthecellrepresentsonlya0.002%changeintheintracellularK+ content: . mol −13 ° 0 002. % 5 03 × 10 es . mol Thustheintracellular[K+]ofthecellisnotappreciablyalteredbythediffusionofK+ outofthecell. Regulation of Cell Volume As already noted, changes in cell volume can lead to cell damage and death. Cells have developed mechanisms to regulate their volume. Most cells are highly permeable by water because of the presence of aquaporins in their plasma membranes. As discussed in


Physiology_Levy

Physiology_Levy. Chapter 1, osmotic pressure gradients across the cell membrane that are generated by effective osmoles cause water to move either into or out of the cell, which result in changes in cell volume. Thus cells swell when placed in hypotonic solutions and shrink when placed in hypertonic solutions (see the section ”). However, even when a cell is placed in an isotonic solution, the maintenance of cell volume is an active process requiring the expenditure of ATP and specifically the activity of the Na+,K+-ATPase.


Physiology_Levy

Physiology_Levy. The importance of the Na+,K+-ATPase in isotonic cell volume regulation can be appreciated by the observation that red blood cells swell when chilled (i.e., reduced ATP synthesis) or when the Na+,K+-ATPase is inhibited by cardiac glycosides (e.g., ouabain, digoxin [Lanoxin]). The necessity for energy expenditure to maintain cell volume in an isotonic solution is the result of the effect of intracellular proteins on the distribution of ions across the plasma membrane: the so-called Gibbs-Donnan effect ( Fig. 2.6


Physiology_Levy

Physiology_Levy. Fig. 2.6 The Gibbs-Donnan effect occurs when a membrane separating two solutions can be permeated by some but not all of the molecules in solution. As noted previously, this effect accounts for the small differences in the ionic compositions of the plasma and the interstitial fluid. In this case, the capillary endothelium represents the membrane, and the plasma proteins are the molecules whose ability to permeate across the capillary is restricted. For cells, the membrane is the plasma membrane, and the impermeant molecules are the intracellular proteins and organic molecules.


Physiology_Levy

Physiology_Levy. Fig. 2.6 , the presence of impermeant molecules (e.g., protein) in one compartment results over time in the accumulation of permeant molecules/ions in the same compartment. This increases the number of osmotically active particles in the compartment containing the impermeant anions, which in turn increases the osmotic pressure, and water thereby enters that compartment. For cells, the Gibbs-Donnan effect would increase the number of osmotically active particles in the cell, and result in cell swelling. However, the activity of the Na+,K+-ATPase counteracts the Gibbs-Donnan effect by actively extruding cations (three Na+ ions are extruded, whereas two K+ ions are brought into the cell). In addition, the K+ gradient established by the Na+,K+-ATPase allows for the development of the Vm (in which the cell’s interior is electrically negative), that in turn drives Cl− and other anions out of the cell. Thus through the activity of the Na+,K+-ATPase, the number of intracellular osmotically


Physiology_Levy

Physiology_Levy. which the cell’s interior is electrically negative), that in turn drives Cl− and other anions out of the cell. Thus through the activity of the Na+,K+-ATPase, the number of intracellular osmotically active particles is


Physiology_Levy

Physiology_Levy. B the intracellular osmotic pressure and thereby restoring cell volume to normal. Conversely with cell shrinking a regulatory volume increase response transports osmolytes into the cell, raising the intracellular osmotic pressure and thereby restoring cell volume to normal. These osmolytes include ions and organic molecules such as polyols (sorbitol and myo-inositol), methylamines (glycerophosphorylcholine and betaine), and some amino acids (taurine, glutamate, and β-alanine). If the cell is exposed to the nonisotonic ECF for an extended period of time, the cell alters the intracellular levels of the organic osmolytes through metabolic processes.


Physiology_Levy

Physiology_Levy. •Fig. 2.6 The Gibbs-Donnan Effect. Top, TwosolutionsareseparatedbyamembranethatispermeablebyNa+ ,Cl− ,andH2Obutnotpermeablebyprotein(P−).TheosmolalityofsolutionAisidenticaltothatofsolutionB.Bottom, Cl− diffusesfromcompartmentBtocompartmentAdownitsconcentrationgradient.ThiscausescompartmentAtobecomeelectricallynegativewithregardtocompartmentB.ThemembranevoltagethendrivesthediffusionofNa+ fromcompartmentBtocompartmentA.TheaccumulationofadditionalNa+ andCl− incompartmentAincreasesitsosmolalityandcauseswatertoflowfromcompartmentBtocompartmentA(Note:theincreasevolumeofcompartmentAresultsinalower[P−]).IfthecontainercontainingthetwosolutionsweresealedatthetopsothatwatercouldnotmovefromcompartmentBtocompartmentA,thepressureincompartmentAwouldincreaseasthenumberofosmoticallyactiveparticlesincreasesinthatcompartment. reduced from what would be caused by the Gibbs-Donnan effect, and cell volume is maintained in isotonic solutions.


Physiology_Levy

Physiology_Levy. reduced from what would be caused by the Gibbs-Donnan effect, and cell volume is maintained in isotonic solutions. Most cells throughout the body are bathed with isotonic ECF, the composition of which is tightly regulated (see ). However, certain regions within the body are not isotonic (e.g., the medulla of the kidney), and with disorders of water balance, the ECF can become either hypotonic or hypertonic. When this occurs, cells either swell or shrink. Cell swelling or shrinkage can result in cell damage or death, but many cells have mechanisms that limit the degree to which the cell volume changes. These mechanisms are particularly important for neurons, in which swelling within the confined space of the skull can lead to serious neurological damage. In general, when a cell is exposed to nonisotonic ECF, volume-regulatory responses are activated within seconds to minutes to restore cell volume (


Physiology_Levy

Physiology_Levy. In general, when a cell is exposed to nonisotonic ECF, volume-regulatory responses are activated within seconds to minutes to restore cell volume ( Fig. 2.7 ). With cell swelling, a regulatory volume decrease response transports osmotically active particles (osmolytes) out of the cell, reducing


Physiology_Levy

Physiology_Levy. Fig. 2.7 ). With cell swelling, a regulatory volume decrease response transports osmotically active particles (osmolytes) out of the cell, reducing TheECFofindividualswithdisordersinwaterbalancemaybeeitherhypotonic(positivewaterbalance)orhypertonic(negativewaterbalance).WithadecreaseinECFosmolality,neuronsandglialcellsswellaswaterentersthecell.Tominimizethisswelling,theneuronsandglialcellsreduceintracellularosmolytes.IftheECFosmolalityiscorrected(i.e.,increased)tooquickly,theneuronsandglialcellsthenshrinkbecauseofthereducednumberofintracellularosmolytes.ThisresponsetoarapidcorrectionofECFosmolalitycanleadtocelldamage.Damagetotheglialcellsthatsynthesizemyelinwithinthebraincanresultindemyelinization.Thisdemyelinizationresponse,termedosmotic demyelinization syndrome, canaffectanyofthewhitematterofthebrain,butespeciallyregionsofthepons.Theseeffectsareoftenirreversible.Therefore,correctionofdisordersofwaterbalanceisusuallyaccomplishedslowlytoavoidthisseriousneurologicalcomplication.


Physiology_Levy

Physiology_Levy. The regulatory volume increase response results in the rapid uptake of NaCl and a number of organic osmolytes. To increase cell volume there is an activation of the Na+-H+ antiporter (NHE-1), the 1Na+,1K+,2Cl− symporter (NKCC1), and a number of cation-selective channels, which together bring NaCl into the cell. The Na+,K+-ATPase then extrudes the Na+ in exchange for K+ , so that ultimately the KCl content of the cell is increased. Several organic osmolyte transporters are also activated to increase cell volume. These include a 3Na+,1Cl−-taurine symporter, a 3Na+,2Cl−-betaine symporter, a 2Na+–myo-inositol symporter, and a Na+– amino acid symporter. These transporters use the energy in the Na+ and Cl− gradients to drive the secondary active uptake of these organic osmolytes into cells.


Physiology_Levy

Physiology_Levy. The regulatory volume decrease response results in the loss of KCl and organic osmolytes from the cell. The loss of KCl occurs through the activation of a wide range of K+selective, Cl−-selective, and anion-selective channels (the specific channels involved vary depending on the cell), as well as through activation of K+-Cl− symporters. Some of the organic osmolytes appear to leave the cell via anion channels (e.g., volume-sensitive organic osmolyte-anion channels). Several mechanisms are involved in activation of these various transporters during the volume regulatory


Physiology_Levy

Physiology_Levy. CHAPTER 2 Homeostasis: Volume and Composition of Body Fluid Compartments 2Cl Na+, Cl– Na+ + •Fig. 2.7 Volume Regulation of Cells in Hypotonic and Hypertonic Media. Top, Whencellsareexposedtoahypotonicmedium,theyswellandthenundergoavolume-regulatorydecrease(RVD).TheRVDinvolveslossofKClandorganicosmolytesfromthecell.ThedecreaseincellularKClandorganicosmolytescausesintracellularosmoticpressuretodecrease,waterleavesthecell,andthecellreturnstonearlyitsoriginalvolume.Bottom, Whencellsareexposedtoahypertonicmedium,theyshrinkandthenundergoavolume-regulatoryincrease(RVI).DuringtheRVI,NaClandorganicosmolytesenterthecell.TheincreaseintheactivityofNa+,K+-ATPase(notdepicted)enhancestheexchangeNa+ forK+ sothattheK+ (andCl−)contentofthecellisincreased.TheincreaseincellularKCl,alongwithariseinintracellularorganicosmolytes,increasesintracellularosmoticpressure,whichbringswaterbackintothecell,andthecellvolumereturnstonearlyitsoriginalvolume.π,theoncoticpressureinsidethecell.


Physiology_Levy

Physiology_Levy. responses. Changes in cell volume appear to monitored by the cytoskeleton, by changes in macromolecular crowding and ionic strength of the cytoplasm, and by channels whose gating is influenced, either directly or indirectly, by stretch of the plasma membrane (e.g., stretch-activated cation channels). A number of second messenger systems may also be involved in these responses (e.g., intracellular [Ca++], calmodulin, protein kinase A, and protein kinase C), but the precise mechanisms have not been defined completely. Principles of Epithelial Transport


Physiology_Levy

Physiology_Levy. Principles of Epithelial Transport Epithelial cells are arranged in sheets and provide the interface between the external world and the internal environment (i.e., ECF) of the body. Depending on their location, epithelial cells serve many important functions, such as establishing a barrier to microorganisms (lungs, gastrointestinal tract, and skin), prevention of the loss of water from the body (skin), and maintenance of a constant internal environment (lungs, gastrointestinal tract, and kidneys). This latter function is a result of the ability of epithelial cells to carry out regulated vectorial transport (i.e., transport from one side of the epithelial cell sheet to the opposite side). In this section, the principles of epithelial transport are reviewed. The transport functions of specific epithelial cells are discussed in the appropriate chapters throughout this book.


Physiology_Levy

Physiology_Levy. Fig. 2.8 shows a schematic representation of an epithelial cell. The free surface of the epithelial layer is referred to as the apical membrane. It is in contact with the external environment (e.g., air within the alveoli and larger airways of the lungs and the contents of the gastrointestinal tract) or with extracellular fluids (e.g., glomerular filtrate in the nephrons of the kidneys and the secretions of the ducts of the pancreas or sweat glands). The basal side of the epithelium rests on a basal lamina, which is secreted by the epithelial cells, and this in turn is attached to the underlying connective tissue. Epithelial cells are connected to one another and to the underlying connective tissue by a number of specialized junctions (see


Physiology_Levy

Physiology_Levy. Epithelial cells are connected to one another and to the underlying connective tissue by a number of specialized junctions (see Fig. 2.8 ). The adhering junction, desmosomes, and hemidesmosomes provide mechanical adhesion by linking together the cytoskeleton of adjacent cells (adhering junction and desmosome) or to the underlying connective tissue (hemidesmosome). The gap junction and tight junction play important physiological roles. Gap junctions provide low-resistance connections between cells. The functional unit of the gap junction is the connexon. The connexon is composed of six integral gGap junctions are not limited to epithelial cells. A number of other cells also have gap junctions (e.g., cardiac myocytes and smooth muscle cells). •Fig. 2.8 Schematic of an Epithelial Cell, Illustrating the Various Adhering Junctions. Thetightjunctionseparatestheapicalmembranefrombasolateralmembrane(seetextfordetails).


Physiology_Levy

Physiology_Levy. •Fig. 2.8 Schematic of an Epithelial Cell, Illustrating the Various Adhering Junctions. Thetightjunctionseparatestheapicalmembranefrombasolateralmembrane(seetextfordetails). membrane protein subunits called connexins. A connexon in one cell is aligned with the connexon in the adjacent cell, forming a channel. The channel may be gated, and when it is open, it allows the movement of ions and small molecules between cells. Because of their low electrical resistance, they effectively couple electrically one cell to the adjacent cell.


Physiology_Levy

Physiology_Levy. The tight junction serves two main functions. It divides the cell into two membrane domains (apical and basolateral) and, in so doing, restricts the movement of membrane lipids and proteins between these two domains. This so-called fence function allows epithelial cells to carry out vectorial transport from one surface of the cell to the opposite surface by segregating membrane transporters to one or other of the membrane domains. They also serve as a pathway for the movement of water, ions, and small molecules across the epithelium. This pathway between the cells is referred to as the paracellular pathway, as opposed to the transcellular pathway through the cells. The apical surface of epithelial cells may have specific structural features. One such feature is microvilli (Fig.


Physiology_Levy

Physiology_Levy. The apical surface of epithelial cells may have specific structural features. One such feature is microvilli (Fig. 2.9A ). Microvilli are small (typically 1 to 3 µm in length), nonmotile projections of the apical plasma membrane that serve to increase surface area. They are commonly located on cells that must transport large quantities of ions, water, and molecules (e.g., epithelial cells lining the small intestine and cells of the renal proximal tubule). The core of the microvilli is composed of actin filaments and a number of accessory proteins. This actin core is connected to the cytoskeleton of the cell via the terminal web (a network of actin fibers at


Physiology_Levy


Physiology_Levy

Physiology_Levy. CHAPTER 2 Homeostasis: Volume and Composition of Body Fluid Compartments 29 • Fig. 2.9 Illustration of Apical Membrane Specializations of Epithelial Cells (Not Drawn to Scale). A, Microvilli 1 to 3 µm in length serve to increase the surface area of the apical membrane (e.g., those of the epithelial cells of the small intestine). B, Stereocilia can be up to 120 µm in length (e.g., those of the epididymis of the male reproductive tract). Both microvilli and stereocilia have a core structure composed primarily of actin, with a number of associated proteins. Both are nonmotile. (Redrawn from Pawlina, W. Histology: A Text and Atlas, with Correlated Cell and Molecular Biology. 7th ed. Philadelphia: Wolters Kluwer Health, 2016.) the base of the microvilli) and provides structural support for the microvilli. Another surface feature is stereocilia (see


Physiology_Levy

Physiology_Levy. Fig. 2.9B ). Stereocilia are long (up to 120 µm), nonmotile membrane projections that, like microvilli, increase the surface area of the apical membrane. They are found in the epididymis of the testis and in the “hair cells” of the inner ear. Their core also contains actin filaments and accessory proteins. A third apical membrane feature is cilia ( Fig. 2.10 ). Cilia may be either motile (called secondary cilia) or non-motile (called primary cilia). The motile cilia contain a microtubule core arranged in a characteristic “9+2” pattern (nine pairs of microtubules around the circumference of the cilium, and one pair of microtubules in the center). Dynein is the molecular motor that drives the movement of the cilium. Motile cilia are characteristic features of the epithelial cells that line the respiratory tract. They pulsate in a synchronized manner and serve to transport mucus and inhaled particulates out of the lung, a process termed mucociliary transport (see Chapter 26).


Physiology_Levy

Physiology_Levy. Chapter 26). Nonmotile cilia serve as mechanoreceptors and are involved in determining left-right asymmetry of organs during embryological development, as well as sensing the flow rate of fluid in the nephron of the kidneys (see ). Only a single nonmotile cilium is found in the apical membrane of cells. Nonmotile cilia have a microtubule core (“9+0” arrangement) and lack a motor protein. As noted previously, the tight junction effectively divides the plasma membrane of an epithelial cell into two domains: an apical surface and a basolateral surface. The basolateral membrane of many epithelial cells is folded or invaginated. This is especially so for epithelial cells that have high transport rates. These invaginations serve to increase the membrane surface area to accommodate the large number of membrane transporters (e.g., Na+,K+-ATPase) needed in the membrane.


Physiology_Levy

Physiology_Levy. Because the tight junction divides the plasma membrane into two domains (i.e., apical and basolateral), epithelial cells are capable of vectorial transport, whereby an ion or molecule can be transported from one side of the epithelial sheet to the opposite side ( Fig. 2.11 ). The accomplishment of vectorial transport requires that specific membrane transport proteins be targeted to and remain in one or the other of the membrane domains. In the example shown in


Physiology_Levy

Physiology_Levy. Fig. 2.11 , the Na+ channel is present only in the apical •Fig. 2.10 Ciliaareapicalmembranespecializationsofsomeepithelialcells.Ciliaare5to10µminlengthandcontainarraysofmicrotubules,asdepictedinthesecross-sectiondiagrams.Left, Theprimaryciliumhasnineperipheralmicrotubulearrays.Itisnonmotileandservesasamechanoreceptor(e.g.,cellsoftherenalcollectingduct).Cellsthathaveaprimaryciliumhaveonlyasinglecilium.Right, Thesecondaryciliumhasacentralpairofmicrotubulesinadditiontothenineperipheralmicrotubulearrays.Alsointhesecondarycilium,themotorproteindyneinisassociatedwiththemicrotubulearraysandthereforeismotile.Asinglecellcanhavethousandsofsecondaryciliaonitsapicalsurface(e.g.,epithelialcellsoftherespiratorytract).(RedrawnfromRodat-DespoixL,DelmasP.Ciliaryfunctionsinthe nephron.Pflugers Archiv.2009;458:179.) membrane, whereas the Na+,K+-ATPase and the K+ channels are confined to the basolateral membrane. The operation of the Na+,K+-ATPase channel and the leakage of K+ out of the cell across the


Physiology_Levy

Physiology_Levy. membrane, whereas the Na+,K+-ATPase and the K+ channels are confined to the basolateral membrane. The operation of the Na+,K+-ATPase channel and the leakage of K+ out of the cell across the basolateral membrane sets up a large electrochemical gradient for Na+ to enter the cell across the apical membrane through the Na+ channel (intracellular [Na+] < extracellular [Na+], and Vm which is oriented with the cell’s interior electrically negative with respect to the cell’s exterior). The Na+ is then pumped out of the cell by the Na+,K+-ATPase, and vectorial transport from the apical side of the epithelium to the basolateral side of the epithelium occurs. Transport from the apical side to the basolateral side of an epithelium is termed either absorption or reabsorption: For example, the uptake of nutrients from the lumen of the gastrointestinal tract is termed absorption, whereas the transport of NaCl and water from the lumen of the renal nephrons is termed reabsorption. Transport from the


Physiology_Levy

Physiology_Levy. of nutrients from the lumen of the gastrointestinal tract is termed absorption, whereas the transport of NaCl and water from the lumen of the renal nephrons is termed reabsorption. Transport from the basolateral side of the epithelium to the apical side is termed secretion.


Physiology_Levy

Physiology_Levy. As noted previously, the Na+,K+-ATPase and K+-selective channels play an important role in establishing cellular ion gradients for Na+ and K+ and in generating the Vm. In all epithelial cells except the choroid plexus and retinal pigment epithelium, the Na+,K+-ATPase channel is located in the basolateral membrane of the cell. Numerous K+selective channels are in epithelial cells and may be located in either membrane domain. Through the establishment of these chemical and voltage gradients, the transport of other ions and solutes can be driven (e.g., Na+-glucose symporter, Na+-H+ antiporter, 1Na+,1K+,2Cl− symporter, 1Na+-3HCO3 − symporter). The direction of transepithelial transport (reabsorption or secretion) depends simply on which membrane domain the transporters are located. Because of the dependence on the Na+,K+-ATPase, epithelial transport requires the expenditure of energy. Other ATP-dependent transporters, such as the H+-ATPase, H+,K+-ATPase, and a host of ABC


Physiology_Levy

Physiology_Levy. Because of the dependence on the Na+,K+-ATPase, epithelial transport requires the expenditure of energy. Other ATP-dependent transporters, such as the H+-ATPase, H+,K+-ATPase, and a host of ABC transporters—such as P-glycoprotein (PGP) and multidrug resistance-associated protein 2 (MRP2), which transport xenobiotics (drugs), and cystic fibrosis transmembrane conductance regulator (CFTR), which transports Cl−—are involved in epithelial transport.


Physiology_Levy

Physiology_Levy. hThe choroid plexus is located in the ventricles of the brain and secretes the cerebrospinal fluid. The Na+,K+-ATPase channel is located in the apical membrane of these cells. CHAPTER 2 Homeostasis: Volume and Composition of Body Fluid Compartments •Fig. 2.11 Insymmetricalcells(A; e.g.,redbloodcells),membranetransportproteinsaredistributedovertheentiresurfaceofthecell.Epithelialcells(B), incontrast,areasymmetricalandtargetvariousmembranetransportproteinstoeithertheapicalorthebasolateralmembrane.Whenthetransportersareconfinedtoamembranedomain,vectorialtransportcanoccur.Inthecelldepicted,Na+ istransportedfromtheapicalsurfacetothebasolateralsurface.ATP,adenosinetriphosphate.


Physiology_Levy

Physiology_Levy. Solutes and water can be transported across an epithelium by traversing both the apical and basolateral membranes (transcellular transport) or by moving between the cells across the tight junction (paracellular transport). Solute transport via the transcellular route is a two-step process, in which the solute molecule is transported across both the apical and basolateral membrane. Uptake into the cell, or transport out of the cell, may be either a passive or an active process. Typically, one of the steps is passive, and the other is active. For the example shown in


Physiology_Levy

Physiology_Levy. Fig. 2.11B , the uptake of Na+ into the cell across the apical membrane through the Na+-selective channel is passive and driven by the electrochemical gradient for Na+ . The exit of Na+ from the cell across the basolateral membrane is primary active transport via the Na+,K+-ATPase channel. Because a transepithelial gradient for Na+ can be generated by this process (i.e., the [Na+] in the apical compartment can be reduced below that of the basolateral compartment, the overall process of transepithelial Na+ transport is said to be active). Any solute that is actively transported across an epithelium must be transported via the transcellular pathway.


Physiology_Levy

Physiology_Levy. Depending on the epithelium, the paracellular pathway is an important route for transepithelial transport of solute and water. As noted, the permeability characteristics of the paracellular pathway are determined, in large part, by the specific claudins that are expressed by the cell. Thus the tight junction can have low permeability for solutes, water, or both, or it can have a high permeability. For epithelia in which there are high rates of transepithelial transport, •Fig. 2.12 The Electrical Profile Across an Epithelial Cell. Themagnitudeofthemembranevoltages,andthetransepithelialvoltagearedeterminedbythevariousmembranetransportproteinsintheapicalandbasolateralmembranes.Thetransepithelialvoltageisequaltothesumoftheapicalandbasolateralmembranevoltages(seetextfordetails).


Physiology_Levy

Physiology_Levy. the tight junctions typically have a high permeability (i.e., are leaky). Examples of such epithelia include the proximal tubule of the renal nephron and the early segments of the small intestine (e.g., duodenum and jejunum). If the epithelium must establish large transepithelial gradients for solutes, water, or both, the tight junctions typically have low permeability (i.e., are tight). Examples of this type of epithelium include the collecting duct of the renal nephron, the urinary bladder, and the terminal portion of the colon. In addition, the tight junction may be selective for certain solutes (e.g., cation versus anion selective).


Physiology_Levy

Physiology_Levy. All solute transport that occurs through the paracellular pathway is passive in nature. The two driving forces for this transport are the transepithelial concentration gradient for the solute and, if the solute is charged, the transepithelial voltage (Fig. 2.12 ). The transepithelial voltage may be oriented with the apical surface electrically negative in relation to the basolateral surface as shown in Fig. 2.12 , or it may be oriented with the apical surface electrically positive in relation to the basolateral surface. The polarity and magnitude of the transepithelial voltage is determined by the specific membrane transporters in the apical and basolateral membranes, as well as by the permeability characteristics of the tight junction. It is important to recognize that transcellular transport processes set up the transepithelial chemical and voltage gradients, which in turn can drive paracellular transport. This is illustrated in


Physiology_Levy

Physiology_Levy. It is important to recognize that transcellular transport processes set up the transepithelial chemical and voltage gradients, which in turn can drive paracellular transport. This is illustrated in Fig. 2.13 for an epithelium that reabsorbs NaCl and for an epithelium that secretes NaCl. In both epithelia, the transepithelial voltage is oriented with the apical surface electrically negative in relation to the basolateral surface. For the NaCl-reabsorbing epithelium, the transepithelial voltage is generated by the active, transcellular reabsorption of Na+ . This voltage in turn drives Cl− reabsorption through the paracellular pathway. In contrast, for the NaCl-secreting epithelium, the transepithelial voltage is generated by the active transcellular secretion of Cl− . Na+ is then secreted passively via the paracellular pathway, driven by the negative transepithelial voltage.


Physiology_Levy

Physiology_Levy. Water movement across epithelia is passive and driven by transepithelial osmotic pressure gradients. Water movement can occur by a transcellular route involving aquaporins in both the apical and basolateral membranes. In addition, water may also move through the paracellular pathway. In the NaCl-reabsorbing epithelium depicted in Fig. 2.13A , the reabsorption of NaCl from the apical compartment lowers the osmotic pressure in that compartment, whereas the addition of NaCl to the basolateral compartment raises the osmotic pressure in that compartment. As a result, a transepithelial osmotic pressure gradient is established that drives the movement of water from the apical to the basolateral compartment (i.e., reabsorption). The opposite occurs with NaCl-secreting epithelia (see Fig. 2.13B ), in which the transepithelial secretion of NaCl establishes a transepithelial osmotic pressure gradient that drives water secretion.


Physiology_Levy

Physiology_Levy. Fig. 2.13B ), in which the transepithelial secretion of NaCl establishes a transepithelial osmotic pressure gradient that drives water secretion. In some epithelia (e.g., proximal tubule of the renal nephron), the movement of water across the epithelium via the paracellular pathway can drive the movement of additional solute. This process is termed solvent drag and reflects the fact that solutes dissolved in the water will traverse the tight junction with the water. As is the case with the establishment of transepithelial concentration and voltage gradients, the establishment of transepithelial osmotic pressure gradients requires transcellular transport of solutes by the epithelial cells. iDifferent aquaporin isoforms are often expressed in the apical and basolateral membrane. In addition, multiple isoforms may be expressed in one or more of the membrane domains. CHAPTER 2 Homeostasis: Volume and Composition of Body Fluid Compartments


Physiology_Levy

Physiology_Levy. CHAPTER 2 Homeostasis: Volume and Composition of Body Fluid Compartments NaCl reabsorption •Fig. 2.13 The Role of the Paracellular Pathway in Epithelial Transport. A, Na+ transportthroughthecellgeneratesatransepithelial voltagethatthendrivesthepassivemovementofCl− throughthetightjunction.NaClreabsorptionresults.B, Cl− transportthroughthecellgeneratesatransepithelialvoltagethatthendrivesthepassivetransportofNa+ throughthetightjunction.NaClsecretionresults. Regulation of Epithelial Transport


Physiology_Levy

Physiology_Levy. Epithelial transport must be regulated to meet the homeostatic needs of the individual. Depending on the epithelium, this regulation involves neural or hormonal mechanisms, or both. For example, the enteric nervous system of the gastrointestinal tract regulates solute and water transport by the epithelial cells that line the intestine and colon. Similarly, the sympathetic nervous system regulates transport by the epithelial cells of the renal nephron. Aldosterone, a steroid hormone produced by the adrenal cortex (see ), is an example of a hormone that stimulates NaCl transport by the epithelial cells of the colon, renal nephron, and sweat ducts. Epithelial cell transport can also be regulated by locally produced and locally acting substances, a process termed paracrine regulation. The stimulation of HCl secretion in the stomach by histamine is an example of this process. Cells that are located near the epithelial cells of the stomach release histamine, which acts on the HCl-secreting


Physiology_Levy

Physiology_Levy. of HCl secretion in the stomach by histamine is an example of this process. Cells that are located near the epithelial cells of the stomach release histamine, which acts on the HCl-secreting cells of the stomach (parietal cells) and stimulates them to secrete HCl.


Physiology_Levy

Physiology_Levy. When acted upon by a regulatory signal, the epithelial cell may respond in several different ways, including: • Retrieval of transporters from the membrane, by endocytosis, or insertion of transporters into the membrane from an intracellular vesicular pool, by a process called Change in activity of membrane transporters (e.g., channel gating) Synthesis of specific transporters, and their insertion into the membrane Na+ The first two mechanisms can occur quite rapidly (seconds 2Cl– to minutes), but the synthesis of transporters takes addi-K+ tional time (minutes to days).


Physiology_Levy

Physiology_Levy. Na+ The first two mechanisms can occur quite rapidly (seconds 2Cl– to minutes), but the synthesis of transporters takes addi-K+ tional time (minutes to days). The body maintains steady-state balance for water and a number of important solutes. This occurs when input into the body equals output from the body. For each solute and water, there is a normal set point. Deviations from this set point are monitored (i.e., when input ≠ output), and effector mechanisms are activated that restore balance. This balance is achieved by adjustment of either intake or excretion of water and solutes. Thereafter, input and output are again equal to maintain balance. The Na+,K+-ATPase and K+-selective channels are critically important in establishing and maintaining the intracellular composition, the membrane potential (Vm), and cell volume. Na+,K+-ATPase converts the energy in


Physiology_Levy

Physiology_Levy. Altenberg GA, Ruess L. Mechanisms of water transport across cell membranes and epithelia. In: Alpern R, Moe O, Kaplan M, eds. Seldin and Giebisch’s The Kidney—Physiology and Pathophysiology. 5th ed. New York: Academic Press; 2013. Günzel D, Yu ASL. Claudins and the modulation of tight junction permeability. Physiol Rev. 2013;93:525-569. Hoffman EK, et al. Physiology of cell volume regulation in vertebrates. Physiol Rev. 2009;89:193-277. Lang F. Cell volume control. In: Alpern R, Moe O, Kaplan M, eds. Seldin and Giebisch’s The Kidney—Physiology and Pathophysiology. 5th ed. New York: Academic Press; 2013. ATP into potential energy of ion gradients and the membrane potential. The ion and electrical gradients created by this process are then used to drive the transport of other ions and other molecules, especially by solute carriers (i.e., symporters and antiporters).


Physiology_Levy

Physiology_Levy. • Epithelial cells constitute the interface between the external world and the internal environment of the body. Vectorial transport of solutes and water across epithelia helps maintain steady-state balance for water and a number of important solutes. Because the external environment constantly changes, and because dietary intake of food and water is highly variable, transport by epithelia is regulated to meet the homeostatic needs of the individual. Pawlina W. Histology: A Text and Atlas, with Correlated Cell and Molecular Biology. 7th ed. Philadelphia: Wolters Kluwer Health; 2016. Pedersen SF, Kapus A, Hoffmann EK. Osmosensory mechanisms in cellular and systemic volume regulation. J Am Soc Nephrol. 2011;22:1587-1597. Sackin H, Palmer LG. Electrophysiological analysis of transepithelial transport. In: Alpern R, Moe O, Kaplan M, eds. Seldin and Giebisch’s The Kidney—Physiology and Pathophysiology. 5th ed. New York: Academic Press; 2013.


Physiology_Levy

Physiology_Levy. 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression Upon completion of this chapter, the student should be able to answer the following questions: 1. How do cells communicate with each other? 2. What are the four classes of receptors, and what signal transduction pathways are associated with each class of receptors? 3. How do steroid and thyroid hormones, cyclic adenosine monophosphate, and receptor tyrosine kinases regulate gene expression?


Physiology_Levy

Physiology_Levy. he human body is composed of billions of cells, each with a distinct function. However, the function of cells is tightly coordinated and integrated by external chemical signals, including hormones, neurotransmitters, growth factors, odorants, and products of cellular metabolism that serve as chemical messengers and provide cell-to-cell communication. Mechanical and thermal stimuli and light are physical external signals that also coordinate cellular function. Chemical and physical messengers interact with receptors located in the plasma membrane, cytoplasm, and nucleus. Interaction of these messengers with receptors initiates a cascade of signaling events that mediate the response to each stimulus. These signaling pathways ensure that the cellular response to external messengers is specific, amplified, tightly regulated, and coordinated. This chapter provides an overview of how cells communicate via external messengers and a discussion of the signaling pathways that process external


Physiology_Levy

Physiology_Levy. amplified, tightly regulated, and coordinated. This chapter provides an overview of how cells communicate via external messengers and a discussion of the signaling pathways that process external information into a highly coordinated cellular response. In subsequent chapters, details on signaling pathways in the nervous system, muscular system, cardiovascular system, respiratory system, gastrointestinal system, renal system, and endocrine system are discussed in greater detail.


Physiology_Levy

Physiology_Levy. The significance of signaling pathways in medicine is illustrated by the following short list of popular drugs that act by regulating signaling pathways. Details on these pathways are presented later in this and other chapters. Aspirin, the first pharmaceutical (1899), inhibits cyclooxygenase-1 (COX1) and cyclooxygenase-2 (COX2) and therefore is antithrombotic (i.e., reduces the formation of blood clots). β-Adrenergic receptor agonists and antagonists are used to treat a variety of medical conditions. β1-Agonists increase cardiac contractility and heart rate in patients with low blood pressure. β2-Agonists dilate bronchi and are used to treat asthma and chronic obstructive lung disease. In contrast, β-adrenergic antagonists are used to treat hypertension, angina, cardiac arrhythmias, and congestive heart failure (see Chapter 18).


Physiology_Levy

Physiology_Levy. Chapter 18). Fluoxetine (Prozac) is an antidepressant medication that inhibits reuptake of the neurotransmitter serotonin into the presynaptic cell, which results in enhanced activation of serotonin receptors (see Several monoclonal antibodies are used to treat cancer caused by the activation of growth factor receptors in cancer cells. For example, trastuzumab (Herceptin) is a monoclonal antibody used to treat metastatic breast cancer in women who overexpress HER2/neu, a member of the family of epidermal growth factor (EGF) receptors, which stimulate cell growth and differentiation. Cetuximab (Erbitux) and bevacizumab (Avastin) are monoclonal antibodies that are used to treat metastatic colorectal cancer and cancers of the head and neck. These antibodies bind to and inhibit the EGF receptor and thereby inhibit EGF-induced cell growth in cancer cells.


Physiology_Levy

Physiology_Levy. Drugs that inhibit cyclic guanosine monophosphate (cGMP)–specific phosphodiesterase type 5, such as sildenafil (Viagra), tadalafil (Cialis), and vardenafil (Levitra), prolong the vasodilatory effects of nitric oxide and are used to treat erectile dysfunction and pulmonary arterial hypertension (see Chapter 17). An overview of how cells communicate with each other is presented in


Physiology_Levy

Physiology_Levy. Fig. 3.1 . Cells communicate by releasing extracellular signaling molecules (e.g., hormones and neurotransmitters) that bind to receptor proteins located in the plasma membrane, cytoplasm, or nucleus. This signal is transduced into the activation, or inactivation, of one or more intracellular messengers by interacting with receptors. Receptors interact with a variety of intracellular signaling proteins, including kinases, phosphatases, and guanosine triphosphate (GTP)–binding proteins (G proteins). These signaling proteins interact with and regulate the activity of target proteins and thereby modulate cellular function. Target proteins include, but are not limited to, ion channels and other transport proteins, metabolic enzymes, cytoskeletal proteins, gene regulatory proteins, and cell cycle proteins that regulate cell growth and division. Signaling pathways are characterized by (1) multiple, hierarchical steps; (2) amplification of the signal-receptor binding event, which magnifies


Physiology_Levy

Physiology_Levy. proteins that regulate cell growth and division. Signaling pathways are characterized by (1) multiple, hierarchical steps; (2) amplification of the signal-receptor binding event, which magnifies the response; (3) activation of multiple pathways and regulation of multiple cellular functions; mechanisms, which minimize the response and provide tight regulatory control over these signaling pathways. A brief description of how cells communicate follows. Readers who desire a more in-depth presentation of this material are encouraged to consult one of the many cellular and molecular biology textbooks currently available.


Physiology_Levy

Physiology_Levy. Cells in higher animals release into the extracellular space hundreds of chemicals, including (1) peptides and proteins (e.g., insulin); (2) amines (e.g., epinephrine and norepinephrine); (3) steroid hormones (e.g., aldosterone, estrogen); and (4) small molecules, including amino acids, nucleotides, ions (e.g., Ca++), and gases, such as nitric oxide and carbon dioxide. Secretion of signaling molecules is • Fig. 3.1 An Overview of How Cells Communicate. A signaling molecule (i.e., hormone or neurotransmitter) binds to a receptor, which may be in the plasma membrane, cytosol, or nucleus. Binding of ligand to a receptor activates intracellular signaling proteins, which interact with and regulate the activity of one or more target proteins to change cellular function. Signaling molecules regulate cell growth, division, and differentiation and influence cellular metabolism. In addition, they modulate the intracellular ionic composition by regulating the activity of ion channels and


Physiology_Levy

Physiology_Levy. regulate cell growth, division, and differentiation and influence cellular metabolism. In addition, they modulate the intracellular ionic composition by regulating the activity of ion channels and transport proteins. Signaling molecules also control cytoskeleton-associated events, including cell shape, division, and migration and cell-to-cell and cell-to-matrix adhesion. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.)


Physiology_Levy

Physiology_Levy. CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression 37 cell-type specific. For example, beta cells in the pancreas release insulin, which stimulates glucose uptake into cells. The ability of a cell to respond to a specific signaling molecule depends on the expression of receptors that bind the signaling molecule with high affinity and specificity. Receptors are located in the plasma membrane, the cytosol, and the nucleus ( Fig. 3.2 Fig. 3.3). Contact-dependent signaling, in which a membrane-bound signaling molecule of one cell binds directly to a plasma membrane receptor of another cell, is important during development, in immune responses, and in cancer (see Fig. 3.3A ). Molecules that are released and act locally are called paracrine (see Fig. 3.3B


Physiology_Levy

Physiology_Levy. Fig. 3.3C ) hormones. Paracrine signals are released by one type of cell and act on another type; they are usually taken up by target cells or rapidly degraded (within • Fig. 3.2 Signaling molecules, especially ones that are hydrophilic and cannot cross the plasma membrane, bind directly to their cognate receptors in the plasma membrane (A). Other signaling molecules— including steroid hormones, triiodothyronines, retinoic acids, and vitamin D—bind to carrier proteins in blood and readily diffuse across the plasma membrane, where they bind to cognate nuclear receptors in the cytosol or nucleus (B). Still other signaling molecules, including nitric oxide, can diffuse without carrier proteins and cross the membrane to act on intracellular protein targets (B). Both classes of receptors, when ligand bound, regulate gene transcription. mRNA, messenger RNA. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.) minutes) by enzymes. For


Physiology_Levy

Physiology_Levy. when ligand bound, regulate gene transcription. mRNA, messenger RNA. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.) minutes) by enzymes. For example, enterochromaffin-like cells in the stomach secrete histamine, which stimulates the production of acid by neighboring parietal cells (see for details). Autocrine signaling involves the release of a molecule that affects the same cell or other cells of the same type (e.g., cancer cells). In synaptic signaling (see


Physiology_Levy

Physiology_Levy. Fig. 3.3D ), neurons transmit electrical signals along their axons and release neurotransmitters at synapses that affect the function of other neurons or cells that are distant from the neuron cell body. The close physical relationship between the nerve terminal and the target cell ensures that the neurotransmitter is delivered to a specific cell. Details on synaptic signaling are discussed in Chapter 6. Endocrine signals are hormones that are secreted into the blood and are widely dispersed in the body (see Fig. 3.3E ). Details on endocrine signaling are discussed in


Physiology_Levy

Physiology_Levy. Chapter 6. Endocrine signals are hormones that are secreted into the blood and are widely dispersed in the body (see Fig. 3.3E ). Details on endocrine signaling are discussed in In addition to paracrine, autocrine, endocrine, and synaptic signaling, cell-to-cell communication also occurs via gap junctions that form between adjacent cells (see ). Gap junctions are specialized junctions that allow intracellular signaling molecules, generally less than 1200 D in size, to diffuse from the cytoplasm of one cell to an adjacent cell. The permeability of gap junctions is regulated by cytosolic [Ca++], [H+], and cyclic adenosine monophosphate (cAMP) and by the membrane potential. Gap junctions also allow cells to be electrically coupled, which is vitally important for the coordinated activity of cardiac and smooth muscle cells (see


Physiology_Levy

Physiology_Levy. The speed of a response to an extracellular signal depends on the mechanism of delivery. Endocrine signals are relatively slow (seconds to minutes) because time is required for diffusion and blood flow to the target cell, whereas synaptic signaling is extremely fast (milliseconds). If the response involves changes in the activity of proteins in the cell, the response may occur in milliseconds to seconds. However, if the response involves changes in gene expression and the de novo synthesis of proteins, the response may take hours to occur, and a maximal response may take days. For example, the stimulatory effect of aldosterone on sodium transport by the kidneys requires days to develop fully (see


Physiology_Levy

Physiology_Levy. The response to a particular signaling molecule also depends on the ability of the molecule to reach a particular cell, on expression of the cognate receptor (i.e., receptors that recognize a particular signaling molecule or ligand with a high degree of specificity), and on the cytoplasmic signaling molecules that interact with the receptor. Thus signaling molecules frequently have many different effects that are dependent on the cell type. For example, the neurotransmitter acetylcholine stimulates contraction of skeletal muscle but decreases the force of contraction in heart muscle. This is because skeletal muscle and heart cells express different acetylcholine receptors. aThe acetylcholine receptor in skeletal muscle is termed nicotinic because nicotine can mimic this action of the neurotransmitter. In contrast, the acetylcholine receptor in cardiac muscle is termed muscarinic because this effect is mimicked by muscarine, an alkaloid derived from the mushroom Amanita muscaria.


Physiology_Levy

Physiology_Levy. Amanita muscaria. • Fig. 3.3 Cell-to-cell communication is mediated by five basic mechanisms: contact-dependent (A), paracrine (B), autocrine (C), synaptic (D), and endocrine signaling (E). These mechanisms are described in detail in the text. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.) CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression Nuclear receptors Extracellular ligand: GABA ACh (muscle) ATP Glutamate: NMDA Intracellular ligand: Neurotransmitters (ACh) Peptides (PTH, oxytocin) Odorants Cytokines, lipids ANP TGF-β Insulin, EGF Interleukin-6, erythropoietin Steroid hormones: Miscellaneous hormones: Membrane currents: Cl− Na+ , K+ , Ca++ Ca++ , Na+ , K+ Na+ , K+ , Ca++ K+ Na+ , K+ Ca++ βγ Subunits activate ion channels α Subunit activates enzymes:


Physiology_Levy

Physiology_Levy. Miscellaneous hormones: Membrane currents: Cl− Na+ , K+ , Ca++ Ca++ , Na+ , K+ Na+ , K+ , Ca++ K+ Na+ , K+ Ca++ βγ Subunits activate ion channels α Subunit activates enzymes: Cyclases that generate cAMP, cGMP, phospholipases that generate InsP3 and diacylglycerol, and phospholipases that generate arachidonic acid and its metabolites. Monomeric G proteins Receptor guanylyl cyclase Receptor serine/threonine kinase Receptor tyrosine kinase Tyrosine kinase–associated receptor Bind to regulatory sequences in DNA and increase or Bind to regulatory sequences in DNA and increase or decrease gene transcription ACh, acetylcholine; ANP, atrial natriuretic peptide; ATP, adenosine triphosphate; cAMP, cyclic adenosine monophosphate; cGMP, cyclic guanosine monophosphate; EGF, epidermal growth factor; GABA, gamma-aminobutyric acid; InsP3, inositol 1,4,5-triphosphate; NMDA, N-methyl-D-aspartate; PTH, parathyroid hormone; TGF, transforming growth factor.


Physiology_Levy

Physiology_Levy. All signaling molecules bind to specific receptors that act as signal transducers, thereby converting a ligand-receptor binding event into intracellular signals that affect cellular function. Receptors can be divided into four basic classes on the basis of their structure and mechanism of action: (1) ligand-gated ion channels, (2) G protein–coupled receptors (GPCRs), (3) enzyme-linked receptors, and (4) nuclear receptors ( Table 3.1 Figs. 3.4 3.5). Fig. 3.4A). Neurotransmitters bind to receptors and either open or close ion channels, thereby changing the ionic permeability of the plasma membrane and altering the membrane potential. For examples and more details, see GPCRs regulate the activity of other proteins, such as enzymes and ion channels (see Fig. 3.4B ). In the example in


Physiology_Levy

Physiology_Levy. GPCRs regulate the activity of other proteins, such as enzymes and ion channels (see Fig. 3.4B ). In the example in Fig. 3.4B , the interaction between the receptor and the target protein is mediated by heterotrimeric G proteins, which are composed of α, β, and γ subunits. Stimulation of G proteins by ligand-bound receptors activates or inhibits downstream target proteins that regulate signaling pathways if the target protein is an enzyme or changes membrane ion permeability if the target protein is an ion channel. Enzyme-linked receptors either function as enzymes or are associated with and regulate enzymes (see Fig. 3.4C ). Most enzyme-linked receptors are protein kinases or are associated with protein kinases, and ligand binding causes the kinases to phosphorylate a specific subset of proteins on specific amino acids, which in turn activates or inhibits protein activity.


Physiology_Levy

Physiology_Levy. Nuclear receptors are small hydrophobic molecules, including steroid hormones, thyroid hormones, retinoids, and vitamin D, that have a long biological half-life (hours to days), diffuse across the plasma membrane, and bind to nuclear receptors or to cytoplasmic receptors that, once bound to their ligand, translocate to the nucleus (see Fig. 3.5 ). Some nuclear receptors, such as those that bind cortisol


Physiology_Levy

Physiology_Levy. Signal molecule as a dimer Signal molecule • Fig. 3.4 Three of the Four Classes of Plasma Membrane Receptors. See text for details. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.) and aldosterone, are located in the cytosol and enter the nucleus after binding to hormone, whereas other receptors, including the thyroid hormone receptor, are located in the nucleus. In both cases, inactive receptors are bound to inhibitory proteins, and binding of hormone results in dissociation of the inhibitory complex. Hormone binding causes the receptor to bind coactivator proteins that activate gene transcription. Once activated, the hormone-receptor complex regulates the transcription of specific genes. Activation of specific genes usually occurs in two steps: an early primary response (≈30 minutes), which activates genes that stimulate other genes to produce a delayed (hours to days) secondary response (see Fig. 3.5).


Physiology_Levy

Physiology_Levy. Fig. 3.5). Each hormone elicits a specific response that is based on cellular expression of the cognate receptor, as well as on cell type–specific expression of gene regulatory proteins that interact with the activated receptor to regulate the transcription of a specific set of genes (see for more details). In addition to steroid receptors that regulate gene expression, evidence also suggests the existence of membrane and juxtamembrane steroid receptors that mediate the rapid, nongenomic effects of steroid hormones. Some membrane proteins do not fit the classic definition of receptors, but they subserve a receptor-like function in that they recognize extracellular signals and transduce the signals into an intracellular second messenger that has a biological effect. For example, on activation by a ligand, some membrane proteins undergo regulated intramembrane proteolysis (RIP), which elaborates a cytosolic peptide fragment that enters the nucleus and regulates gene expression (


Physiology_Levy

Physiology_Levy. Fig. 3.6 ). In this signaling pathway, binding of ligand to a plasma membrane receptor leads to ectodomain shedding, facilitated by members of the metalloproteinase-disintegrin family, and produces a carboxy-terminal fragment that is the substrate for γ-secretase. γ-Secretase induces RIP, thereby causing the release of an intracellular domain of CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression 41 • Fig. 3.5 Steroid Hormones Stimulate the Transcription of Early-Response Genes and Late-Response Genes. See text for details. (Redrawn from Alberts B, et al: Molecular Biology of Cell. 6th ed. New York: Garland Science; 2015.) Translocation to the nucleus • Fig. 3.6 Regulated Intramembrane Proteolysis. See text for details. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.) the protein that enters the nucleus and regulates transcription (see


Physiology_Levy

Physiology_Levy. Fig. 3.6 ). The best characterized example of RIP is the sterol regulatory element–binding protein (SREBP), a transmembrane protein expressed in the membrane of the endoplasmic reticulum. When cellular cholesterol levels are low, SREBP undergoes RIP, and the proteolytically cleaved fragment is translocated into the nucleus, where it transcriptionally activates genes that promote cholesterol biosynthesis. When hormones bind to plasma membrane receptors, signals are relayed to effector proteins via intracellular signaling pathways. When hormones bind to nuclear or cytosolic receptors, they relay signals primarily through regulation of gene expression. Signaling pathways can amplify and integrate signals but can also downregulate and desensitize signals, reducing or terminating the response, even in the continued presence of hormone.


Physiology_Levy

Physiology_Levy. Alzheimer’s disease, a progressive neurodegenerative brain disease characterized by the formation of amyloid plaques, affects approximately 44 million people worldwide. In Alzheimer’s disease, regulated intramembrane proteolysis of amyloid β-protein precursor (APP) causes the accumulation of amyloid β-protein (Aβ), which forms amyloid plaques that contribute to the pathogenesis of Alzheimer’s disease. APP is a type I transmembrane protein (i.e., its spans the membrane only once). After ectodomain shedding, its sequential proteolysis by β-secretase and γ-secretase produces the Aβ40 and Aβ42 peptides that are normally produced throughout life but accumulate in individuals with Alzheimer’s disease. Missense mutations in presenilins, proteins that regulate γ-secretase protease activity, enhance the production of Aβ42, which is more hydrophobic and prone to aggregation into amyloid fibrils than is the more abundant Aβ40 protein.


Physiology_Levy

Physiology_Levy. Intracellular signaling molecules—so-called second messengers (the first messenger of the signal is the ligand that binds to the receptor)—include small molecules such as cAMP, cGMP, Ca++, and diacylglycerol. Signaling pathways often include dozens of small molecules that form complicated networks within the cell (Fig. 3.7 ). Some proteins in the intracellular signaling pathways relay the signal by passing the message directly to another protein (e.g., by phosphorylating a target, or by binding and causing an allosteric change). Such intracellular signaling proteins act as reversible molecular switches: When a signal is received, they switch from an inactive to an active form or vice versa, until another signaling molecule reverses the process. This principle of reversibility is central to many signaling pathways. In many cases, activation is achieved by reversing inhibition: For example, the thyroid hormone receptor is bound to an inhibitory protein in the absence of signal.


Physiology_Levy

Physiology_Levy. Signaling complexes, composed of multiple proteins that interact physically, enhance the speed, efficiency, and specificity of signaling. Many proteins, usually enzymes or ion channels, transduce the signal into a different chemical form and simultaneously amplify the signal either by producing large amounts of additional signaling molecules or by activating a large number of downstream signaling proteins. For example, adenylyl cyclase, the enzyme that makes cAMP, transduces a signal (receptor activation of G proteins) and amplifies the signal by generating large amounts of cAMP. Other types of signaling proteins include those that integrate multiple signals. Other proteins carry the signal from one region of the cell to another: for example, by translocating from the cytosol to the nucleus.


Physiology_Levy

Physiology_Levy. Cells can adjust rapidly to changing signals. Cells can respond quickly and in a graded manner to increasing concentrations of hormone, and the effect of a signaling molecule can be either long-or short-lived. Cells can also adjust their sensitivity to a signal by desensitization, whereby prolonged exposure to a hormone decreases the cell’s response over time. Desensitization is a reversible process that can involve a reduction in the number of receptors expressed in the plasma membrane, inactivation of receptors, or changes in signaling proteins that mediate the downstream effect of the receptors. Homologous desensitization involves a reduction in the response only to the signaling molecule that caused the response (e.g., opioid dependence and tolerance), whereas heterologous desensitization is when one ligand desensitizes the response to another ligand.


Physiology_Levy

Physiology_Levy. Table 3.1 summarizes the four general classes of receptors and provides a few examples of the signal transduction pathways associated with each class of receptors. This class of receptors transduces a chemical signal into an electrical signal, which elicits a response. For example, the ryanodine receptor, located in the membrane of the sarcoplasmic reticulum of skeletal muscle, is activated by Ca++ , caffeine, adenosine triphosphate (ATP), or metabolites of arachidonic acid to release Ca++ into the cytosol, which facilitates muscle contraction (see for details). In glutamergic synapses in which high levels of prior synaptic activity have led to partial membrane depolarization, activation of the N-methyl-D-aspartate receptor by glutamate stimulates Ca++ influx important for synaptic plasticity.


Physiology_Levy

Physiology_Levy. There are two classes of GTP-binding proteins (i.e., GTPases, which are named for their ability to hydrolyze GTP to guanosine diphosphate [GDP] and an inorganic phosphate): low-molecular-weight, monomeric G proteins and heterotrimeric G proteins composed of α, β, and γ subunits. GTP binding activates, whereas hydrolysis of GTP to GDP inactivates, GTP-binding proteins (Fig. 3.8A ). All GTPases are controlled by regulatory proteins, including GTPase-activating proteins, which induce the hydrolysis of GTP to GDP and thus inactivate the GTPase, and guanine nucleotide exchange factors (GEFs) that causes the GTPase to release GDP, which is rapidly replaced by GTP, thereby activating the GTPase (see Fig. 3.8B


Physiology_Levy

Physiology_Levy. Fig. 3.8B Monomeric G proteins are composed of a single 20to 40-kDa protein and can be membrane bound because of the addition of lipids posttranslationally. Monomeric G proteins have been classified into five families (Ras, Rho, Rab, Ran, and Arf), play a central role in many enzyme-linked receptor pathways, and regulate gene expression and cell proliferation, differentiation, and survival. Rho GTPases regulate actin cytoskeletal organization, cell cycle progression, and gene expression. The Rab GTPases regulate intravesicular transport and trafficking of proteins between organelles in the secretory and endocytic pathways. Ran GTPases regulate nucleocytoplasmic transport of RNA and


Physiology_Levy

Physiology_Levy. CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression 43 • Fig. 3.7 Illustration of How Intracellular Signals Are Amplified and Integrated. Signaling pathways often include dozens of proteins and small molecules that form complicated networks within the cell. Some signaling proteins relay the signal by passing the message to another protein. Many proteins amplify the signal either by producing large amounts of additional signaling molecules or by activating a large number of downstream signaling proteins. Other proteins carry the signal from one region of the cell to another. See text for more details. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.).


Physiology_Levy

Physiology_Levy. • Fig. 3.8 GTP-Binding Proteins. GTP binding activates whereas hydrolysis of GTP to GDP inactivates GTP-binding proteins (A). All GTPase are controlled by regulatory proteins, including GTPase-activating proteins (GAP), which induce the hydrolysis of GTP to GDP, thus inactivating the GTPase, and guanine nucleotide exchange factors, (GEF) that which cause the GTPase to release GDP, which is rapidly replaced by GTP, thereby activating the GTPase (B). (Redrawn from Kantrowitx ER, Lipscomb WN. Escherichia coli aspartate transcarbamoylase: the molecular basis for a concerted allosteric transition. Trends Biochem Sci. 1990;15:53-59.) proteins. Ras GTPases are involved in many signaling pathways that control cell division, proliferation and death. Arf GTPases, like Rab GTPases, regulate vesicular transport.


Physiology_Levy

Physiology_Levy. Sci. 1990;15:53-59.) proteins. Ras GTPases are involved in many signaling pathways that control cell division, proliferation and death. Arf GTPases, like Rab GTPases, regulate vesicular transport. Heterotrimeric G proteins couple to more than 1000 different receptors and thereby mediate the cellular response to an incredibly diverse set of signaling molecules, including hormones, neurotransmitters, peptides, and odorants. Like monomeric G proteins, they can be membrane bound because of the addition of lipids posttranslationally. Heterotrimeric complexes are composed of three subunits: α, β, and γ. There exist 16 α subunits, 5 β subunits, and 11 γ subunits, which can assemble into hundreds of different combinations and thereby interact with a diverse number of receptors and effectors. The assembly of subunits and the association with receptors and effectors depend on the cell type. An overview of heterotrimeric G protein activation is illustrated in


Physiology_Levy

Physiology_Levy. An overview of heterotrimeric G protein activation is illustrated in Fig. 3.9 . In the absence of ligand, these G proteins are inactive and form a heterotrimeric complex in which GDP binds to the α subunit. Binding of a signal molecule to an inactive GPCR induces a conformational change in the G protein that promotes the release of GDP and the subsequent binding of GTP to the α subunit. Binding of GTP to the α subunit stimulates dissociation of the α subunit from the heterotrimeric complex and results in release of the α subunit from the βγ dimer, each of which can interact with and regulate downstream effectors such as adenylyl cyclase and phospholipases (see Fig. 3.9 ). Activation of downstream effectors by the α subunit and βγ dimer is terminated when the α subunit hydrolyzes the bound GTP to GDP and inorganic phosphate (Pi). The α subunit bound to GDP associates with the βγ dimer and terminates the activation of effectors.


Physiology_Levy

Physiology_Levy. Another way to attenuate or terminate signaling through a GPCR involves desensitization and endocytic removal of receptors from the plasma membrane. Binding of hormone to a GPCR increases the ability of GPCR kinases to phosphorylate the intracellular domain of GPCRs, which recruits proteins called β-arrestins to bind to the GPCR. β-Arrestins inactivate the receptor and promote endocytic


Physiology_Levy

Physiology_Levy. CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression 45 • Fig. 3.9 Activation of a G Protein–Coupled Receptor (GPCR) and Effector Activation. In the absence of ligand, heterotrimeric G proteins are in an inactive state because GDP binds to the α subunit. Binding of a signal molecule to an inactive G protein–coupled receptor (GPCR) induces a conformational change in the G protein that promotes the release of GDP and the subsequent binding of GTP to the α subunit. Binding of GTP to the α subunit stimulates dissociation of the α subunit from the heterotrimeric complex and results in release of the α subunit from the βγ dimer, each of which can interact with and regulate downstream effectors. (Redrawn from Alberts B, et al: Molecular Biology of the Cell. 6th ed. New York:


Physiology_Levy

Physiology_Levy. Garland Science; 2015.) removal of the GPCR from the plasma membrane. GPCR kinase/β-arrestin inactivation with endocytosis of GPCRs is an important mechanism whereby cells downregulate (desensitize) a response during prolonged exposure to elevated hormone levels. One of the major benefits of β blockers, given for congestive heart failure, is that they reverse chronic desensitization and the associated recovery of adrenergic responsiveness. Activated G protein α subunits couple to a variety of effector proteins, including adenylyl cyclase, phosphodiesterases, and phospholipases (A2, C, and D). A very common downstream effector of heterotrimeric G proteins is adenylyl cyclase, which facilitates the conversion of ATP to cAMP (


Physiology_Levy

Physiology_Levy. Fig. 3.10 ). When a signal molecule binds to a GPCR that interacts with a G protein composed of an α subunit of the αs class, adenylyl cyclase is activated, which causes an increase in cAMP levels and, as a result, activation of protein kinase A (PKA). By phosphorylating specific serine and threonine residues on downstream effector proteins, PKA regulates effector protein activity. In contrast, when a ligand binds to a receptor that interacts with a G protein composed of a α subunit of the αi class, adenylyl cyclase is inhibited, which causes reductions in cAMP levels and, consequently, in PKA activity.


Physiology_Levy

Physiology_Levy. Some effector proteins, such as ion-gated channels, are also regulated directly by cAMP. cAMP is degraded to AMP activated, phosphorylated CREB •Fig. 3.10 GPCR Stimulation of Adenylyl Cyclase, cAMP, and Protein Kinase A.Binding of a signal molecule to a GPCR mediates Gs stimulation of adenylyl cyclase, which increases cytosolic cAMP, which in turn activates protein kinase A (PKA). Activated PKA phosphorylates a number of target proteins to elicit many effects. PKA also enters the nucleus where it phosphorylates CREB (cyclic adenosine mono-phosphate [cyclic AMP] response element–binding protein). Phosphorylated CREB recruits coactivator CBP, which stimulates gene transcription. (Redrawn from Alberts B, etal: Molecular Biology of the Cell. 6th ed. New York: Garland Science; 2015.) by cAMP phosphodiesterases, which are inhibited by caf-and phosphorylate and activate the transcription factor feine and other methylxanthines. Thus by interfering with cAMP response element–binding (CREB)


Physiology_Levy

Physiology_Levy. phosphodiesterases, which are inhibited by caf-and phosphorylate and activate the transcription factor feine and other methylxanthines. Thus by interfering with cAMP response element–binding (CREB) protein (see a constitutive “off” signal, caffeine can prolong a cellular


Physiology_Levy

Physiology_Levy. Fig. 3.10 ). Phospho-CREB protein increases the transcripresponse mediated by cAMP and PKA. Because these effects tion of many genes, which can in turn produce a distinct set target existing proteins, they can be extremely rapid (e.g., of responses with much slower kinetics. Hence, cAMP has adrenaline response). In addition to cytoplasmic signaling, many cellular effects, including direct and indirect effects the catalytic subunit of PKA can enter the nucleus of cells mediated by PKA. CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression


Physiology_Levy

Physiology_Levy. CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression Cholera toxin, secreted by Vibrio cholera, catalyzes the ADP ribosylation of the G-protein αs subunit, which inhibits the GTPase activity of αs. Thus αs remains in its activated, GTP-bound state, which in turn causes activation of adenylyl cyclase and an increase in cAMP/PKA levels. In the intestines, elevated levels of PKA increase cystic fibrosis transmembrane conductance regulator (CFTR)–mediated chloride secretion, which leads to secretory diarrhea and extensive loss of fluids, characteristic of cholera. Bordetella pertussis, the bacterium that causes whooping cough, secretes pertussis toxin, which ADP ribosylates the αi subunit. In this case, the ribosylation inactivates αi, reducing the inhibition of adenylyl cyclase and thus also leading to increased levels of cAMP/PKA.


Physiology_Levy

Physiology_Levy. Heterotrimeric G proteins also regulate phototransduction. In rod cells in the eye, absorption of light by rhodopsin activates the G protein transducin, which via the αt subunit activates cGMP phosphodiesterase. Activation of this phosphodiesterase lowers the concentration of cGMP and thereby closes a cGMP-activated cation channel. The ensuing change in cation channel activity alters the membrane voltage. The exquisite sensitivity of rods to light—rods can detect a single photon of light—is due to the abundance of rhodopsin in rods and amplification of the signal (a photon of light) by the G protein–cGMP phosphodiesterase–cGMP channel signaling pathway (see for more details).


Physiology_Levy

Physiology_Levy. Heterotrimeric G proteins also regulate phospholipases, a family of enzymes that modulate a variety of signaling pathways. Ligands that activate receptors that are coupled to the αq subunit stimulate phospholipase C, an enzyme that converts phosphatidylinositol 4,5-biphosphate to inositol 1,4,5-triphosphate (InsP3) and diacylglycerol. InsP3 is a second messenger that diffuses to the endoplasmic reticulum, where it activates a ligand-activated Ca++ channel to release Ca++ into the cytosol, whereas diacylglycerol activates protein kinase C, which phosphorylates effector proteins. As noted earlier, both Ca++ and protein kinase C influence effector proteins, as well as other signaling pathways, to elicit responses.


Physiology_Levy

Physiology_Levy. Ligand binding to GPCRs can also activate phospholipase A2, an enzyme that releases arachidonic acid from membrane phospholipids Arachidonic acid, which can also be released from diacylglycerol via an indirect pathway, can be released from cells and thereby regulate neighboring cells or stimulate inflammation. It can also be retained within cells, where it is incorporated into the plasma membrane or is metabolized in the cytosol to form intracellular second messengers that affect the activity of enzymes and ion channels. In one pathway, cytosolic cyclooxygenases facilitate the metabolism of arachidonic acid to prostaglandins, thromboxanes, and prostacyclins. Prostaglandins mediate aggregation of platelets, cause constriction of the airways, and induce inflammation. Thromboxanes also induce platelet aggregation and constrict blood vessels, whereas prostacyclin inhibits platelet aggregation and causes dilation of blood vessels. In a second pathway of arachidonic acid metabolism, the


Physiology_Levy

Physiology_Levy. platelet aggregation and constrict blood vessels, whereas prostacyclin inhibits platelet aggregation and causes dilation of blood vessels. In a second pathway of arachidonic acid metabolism, the enzyme 5-lipoxygenase initiates the conversion of arachidonic acid to leukotrienes, which participate in allergic and inflammatory responses, including those causing asthma, rheumatoid arthritis, and inflammatory bowel disease. The third pathway of arachidonic acid metabolism is initiated by epoxygenase, an enzyme that facilitates the generation of hydroxyeicosatetraenoic acid (HETE) and cis-epoxyeicosatrienoic acid (cis-EET). HETE and cis-EET and their metabolites increase release of Ca++ from the endoplasmic reticulum, stimulate cell proliferation, and regulate inflammatory responses.


Physiology_Levy

Physiology_Levy. Ca++ is also an intracellular messenger that elicits cellular effects via Ca++-binding proteins, most notably calmodulin (CaM). When Ca++ binds to CaM, its conformation is altered, and the structural change in CaM allows it to bind to and regulate other signaling proteins, including cAMP phosphodiesterase, an enzyme that degrades cAMP to AMP, which is inactive and unable to activate PKA. By binding to CaM-dependent kinases, CaM also phosphorylates specific serine and threonine residues in many proteins, including myosin light-chain kinase, which facilitates smooth muscle contraction (see Protein Phosphatases and Phosphodiesterases Counteract the Activation of Cyclic Nucleotide Kinases


Physiology_Levy

Physiology_Levy. Protein Phosphatases and Phosphodiesterases Counteract the Activation of Cyclic Nucleotide Kinases There are two ways to terminate a signal initiated by cAMP and cGMP: enhancing degradation of these cyclic nucleotides by phosphodiesterases and dephosphorylation of effectors by protein phosphatases. Phosphodiesterases facilitate the breakdown of cAMP and cGMP to AMP and GMP, respectively, and are activated by ligand activation of GPCRs. Phosphatases dephosphorylate effector proteins that were phosphorylated by kinases such as PKA. The balance between kinase-mediated phosphorylation and phosphatase-mediated dephosphorylation allows rapid and exquisite regulation of the phosphorylated state and thus the activity of signaling proteins.


Physiology_Levy

Physiology_Levy. There are several classes of receptors that have enzymatic activity or are intimately associated with proteins that have enzymatic activity. Four of these classes are discussed next, including receptors that mediate the cellular responses to atrial natriuretic peptide (ANP) and nitric oxide (guanylyl cyclase receptors); transforming growth factor-β (TGF-β; threonine/serine kinase receptors); EGF, platelet-derived growth factor (PDGF), and insulin (tyrosine kinase receptors); and interleukins (tyrosine kinase–associated receptors).


Physiology_Levy

Physiology_Levy. There are two isoforms of cyclooxygenase: COX1 and COX2. When activated in endothelial cells, COX1 facilitates the production of prostacyclins, which inhibit blood clots (thrombin). In vascular smooth muscle cells and platelets, COX1 facilitates the production of thromboxane A2, which is prothrombotic. Thus cardiovascular health depends in part on the balance between prostacyclins and thromboxane A2, generated by distinct cell types. Low doses of aspirin, a nonsteroidal antiinflammatory drug (NSAID), reduce thromboxane A2 production by platelets with little effect on endothelial prostacyclin production. Thus low-dose aspirin is antithrombotic (i.e., reduces blood clots). COX2 is activated by inflammatory stimuli. Thus the ability of NSAIDs (e.g., aspirin, ibuprofen, naproxen, acetaminophen, indomethacin) to suppress the inflammatory response is due to inhibition of COX2. Both COX1 and COX2 facilitate the production of prostanoids that protect the stomach. Several lines of evidence


Physiology_Levy

Physiology_Levy. indomethacin) to suppress the inflammatory response is due to inhibition of COX2. Both COX1 and COX2 facilitate the production of prostanoids that protect the stomach. Several lines of evidence suggest that both COX1 and COX2 must be inhibited to elicit damage to the gastrointestinal tract. Consequently, the negative effects of NSAIDs on the gastric mucosa (e.g., increased incidence of gastrointestinal bleeding) are most likely due to inhibition of COX1 and COX2 by these nonselective COX inhibitors.


Physiology_Levy

Physiology_Levy. Selective COX2 inhibitors (e.g., celecoxib, rofecoxib) are very effective in selectively inhibiting COX2 and are used extensively to reduce the inflammatory response. Because COX2 inhibitors are thought to lack the negative effects elicited by NSAIDs on the gastrointestinal tract, their use has increased dramatically. However, in 2005, the U.S. Food and Drug Administration (FDA) announced that selective COX2 inhibitors, as well as nonselective NSAIDs, increase the risk for heart attacks and strokes and required that COX2selective and nonselective NSAIDs carry a warning label on product packaging that highlighted the potential for the increased risk for adverse cardiovascular events and stroke. In addition, although much evidence suggests that COX2selective inhibitors do not cause gastrointestinal bleeding, in 2005 the FDA also required the pharmaceutical industry to add to the warning label on COX2-selective drugs a caution about the potential for increased risk for gastrointestinal


Physiology_Levy

Physiology_Levy. bleeding, in 2005 the FDA also required the pharmaceutical industry to add to the warning label on COX2-selective drugs a caution about the potential for increased risk for gastrointestinal bleeding. In 2015, the FDA strengthened warnings that both COX2-selective and COX2-nonselective NSAIDs increase the risk of heart attacks and strokes.


Physiology_Levy

Physiology_Levy. bSee U.S. Food and Drug Administration. FDA Drug Safety Communication: FDA Strengthens Warning That Non-aspirin Nonsteroidal Anti-inflammatory Drugs (NSAIDs) Can Cause Heart Attacks or Strokes. < http://www.fda.gov/ Drugs/DrugSafety/ucm451800.htm >; 2015 Accessed 25.07.16. ANP binds to the extracellular domain of the plasma membrane receptor guanylyl cyclase and induces a conformational change in the receptor that causes receptor dimerization and activation of guanylyl cyclase, which metabolizes GTP to cGMP. cGMP activates cGMP-dependent protein kinase, which phosphorylates proteins on specific serine and threonine residues. In the kidney, ANP inhibits reabsorption of sodium and water by the collecting duct (see Nitric oxide activates a soluble receptor guanylyl cyclase that converts GTP to cGMP, which relaxes smooth muscle. Because nitroglycerin increases blood concentrations of


Physiology_Levy

Physiology_Levy. Nitric oxide activates a soluble receptor guanylyl cyclase that converts GTP to cGMP, which relaxes smooth muscle. Because nitroglycerin increases blood concentrations of Ras GTPases, monomeric G proteins, are involved in many signaling pathways that control cell division, proliferation, and death. Many mutations of proteins in the Ras signaling pathway are oncogenic (cancer causing) or inactivate tumor suppressors. Mutations in Ras genes that inhibit GTPase activity, as well as overexpression of Ras proteins as a result of transcriptional activation, lead to continuous cell proliferation, a major step in the development of cancer in many organs, including the pancreas, colon, and lungs. In addition, mutations in and overexpression of GEFs, which facilitate exchange of GTP for GDP, and GTPase-activating proteins, which accelerate GTP hydrolysis, may also be oncogenic (see


Physiology_Levy

Physiology_Levy. Fig. 3.8B nitric oxide, which increases cGMP and thereby relaxes smooth muscle in coronary arteries, it has long been used to treat angina pectoris (i.e., chest pain caused by inadequate blood flow to heart muscle; see The TGF-β receptor is a threonine/serine kinase that has two subunits. Binding of TGF-β to the type II subunit induces it to phosphorylate the type I subunit on specific serine and threonine residues, which in turn, phosphorylates other downstream effector proteins on serine and threonine residues and thereby elicits cellular responses, including cell growth, cell differentiation, and apoptosis.


Physiology_Levy

Physiology_Levy. There are two classes of tyrosine kinase receptors. Nerve growth factor (NGF) receptors are typical examples of one class. Ligand binding to two NGF receptors facilitates their dimerization and thus enables the cytoplasmic tyrosine kinase domain of each monomer to phosphorylate and activate the other monomer. Once the other monomer is phosphorylated, the cytoplasmic domains can recruit GEFs such as growth factor-receptor bound protein 2 to the plasma membrane, which in turn activates Ras and downstream kinases that regulate gene transcription programs important for cell survival and proliferation.


Physiology_Levy

Physiology_Levy. Activation of the insulin receptor (which is tetrameric and composed of two α and two β subunits) by insulin is an example of the other type of tyrosine kinase receptor. Binding of insulin to the α subunits produces a conformational change that facilitates interaction between the two α and β pairs. Binding of insulin to its receptor causes autophosphorylation of tyrosine residues in the catalytic domains of the β subunits, and the activated receptor then phosphorylates cytoplasmic proteins to initiate its cellular effects, including stimulating the absorption of glucose from the blood into skeletal muscle and fat tissue. The tyrosine kinase–associated receptors have no intrinsic kinase activity but associate with proteins that do have


Physiology_Levy

Physiology_Levy. The tyrosine kinase–associated receptors have no intrinsic kinase activity but associate with proteins that do have CHAPTER 3 Signal Transduction, Membrane Receptors, Second Messengers, and Regulation of Gene Expression tyrosine kinase activity, including tyrosine kinases of the Src family and Janus family. Receptors in this class bind several cytokines, including interleukin-6, a proinflammatory cytokine that is necessary for resistance to bacterial infections, and erythropoietin, which stimulates the production of red blood cells. Tyrosine kinase–associated receptor subunits assemble into homodimers (αα), heterodimers (αβ), or heterotrimers (αβγ) when ligands bind. Subunit assembly enhances the binding of tyrosine kinases, which induces kinase activity and thereby phosphorylates tyrosine residues on the kinases, as well as on the receptor. Most polypeptide growth factors bind to tyrosine-kinase-associated receptors. Regulation of Gene Expression by Signal Transduction Pathways


Physiology_Levy

Physiology_Levy. Regulation of Gene Expression by Signal Transduction Pathways Steroid and thyroid hormones, cAMP, and receptor tyrosine kinases are transcription factors that regulate gene expression and thereby participate in signal transduction pathways. This section discusses the regulation of gene expression by steroid and thyroid hormones, cAMP, and receptor tyrosine kinases. The family of nuclear receptors includes more than 30 genes and has been divided into two subfamilies on the basis of structure and mechanism of action: (1) steroid hormone receptors and (2) receptors that bind retinoic acid, thyroid hormones (iodothyronines), and vitamin D. When ligands bind to these receptors, the ligand-receptor complex activates transcription factors that bind to DNA and regulate the expression of genes (see Figs. 3.2B, 3.5, and 3.7


Physiology_Levy

Physiology_Levy. Figs. 3.2B, 3.5, and 3.7 The location of nuclear receptors varies. Glucocorticoid and mineralocorticoid receptors are located in the cytoplasm, where they interact with chaperones (i.e., heat shock proteins; see Fig. 3.2B ). Binding of hormone to these receptors results in a conformational change that causes chaperones to dissociate from the receptor, thereby revealing a nuclear localization motif that facilitates translocation of the hormone-bound receptor complex to the nucleus. Estrogen and progesterone receptors are located primarily in the nucleus, and thyroid hormone and retinoic acid receptors are located in the nucleus bound to DNA.


Physiology_Levy

Physiology_Levy. When activated by hormone binding, nuclear receptors bind to specific DNA sequences in the regulatory regions of responsive genes called hormone response elements. Ligand-receptor binding to DNA causes a conformational change in DNA that initiates transcription. Nuclear receptors also regulate gene expression by acting as transcriptional repressors. For example, glucocorticoids suppress the transcription activator protein-1 (AP-1) and nuclear factor κB, which stimulate the expression of genes that cause inflammation. By this mechanism glucocorticoids reduce inflammation. As noted previously, cAMP is an important second messenger. In addition to its importance in activating PKA, which phosphorylates specific serine and threonine residues on proteins, cAMP stimulates the transcription of many genes, including those that code for hormones, including somatostatin, glucagon, and vasoactive intestinal polypeptide (see


Physiology_Levy

Physiology_Levy. Fig. 3.10 ). Many genes activated by cAMP have a cAMP response element (CRE) in their DNA. Increases in cAMP stimulate PKA, which not only acts in the cytoplasm but also can translocate to the nucleus, where it phosphorylates CREB and thereby increases its affinity for CREB-binding protein (CBP). The CREB-CBP complex activates transcription. The response is terminated when PKA phosphorylates a phosphatase that dephosphorylates CREB (see Fig. 3.10). Many growth factors, including EGF, PDGF, NGF, and insulin, bind to and activate enzyme-linked receptors that have tyrosine kinase activity. Activation of tyrosine kinases initiates a cascade of events that enhance the activity of the small GTP-binding protein Ras, which in a series of steps and intermediary proteins phosphorylates the mitogenactivated protein kinase, which then translocates to the nucleus and stimulates transcription of genes that stimulate cell growth.


Physiology_Levy

Physiology_Levy. Tyrosine kinase–associated receptors, as noted earlier, are activated by a variety of hormones, including cytokines, growth hormone, and interferon. Although these receptors do not have tyrosine kinase activity, they are associated with Janus family proteins, which do have tyrosine kinase activity. Once activated, hormone tyrosine kinase– associated receptors activate Janus family protein, which phosphorylates latent transcription factors called signal transducers and activators of transcription (STATs). When phosphorylated on tyrosine residues, STATs dimerize and then enter the nucleus and regulate transcription.


Physiology_Levy

Physiology_Levy. 1. The function of cells is tightly coordinated and integrated by external chemical signals, including hormones, neurotransmitters, growth factors, odorants, and products of cellular metabolism that serve as chemical messengers and provide cell-to-cell communication. Chemical and physical signals interact with receptors located in the plasma membrane, cytoplasm, and nucleus. Interaction of these signals with receptors initiates a cascade of events that mediate the response to each stimulus. These pathways ensure that the cellular response to external signals is specific, amplified, tightly regulated, and coordinated.