Answers
Session 5 /
Transport, Fluids, and Pumps/Renal I
M.A.S.T.E.R. Learning Program, UC Davis School of Medicine
Date revised: Feb 14, 2002
Revised by: Hanieh Rad and Arvind Sonik1. Draw the direction of leak of Na+ and K+ in and out of the cell and show how the Na/K pump functions. (25/7-8)
a. What powers the pump?
Answer: ATP (hydrolysis)
b. What kind of transport does this represent?
Answer : Primary active transport
c. Does this require a great deal of the cell's energy?
Answer: Yes, this uses much of the cell's energy.
d. Describe the intra- and extracellular ion concentrations that can affect the rate of the Na/K ATPase pump.
Answer: The Na/K ATPase activity is proportional to [K]o between 0-12(or 14)mM. A decrease in [K]i/[Na]i also increases Na/K ATPase activity. As [K]o decreases or the ratio of [K]i/[Na]i increases, pump activity decreases.
2. If there was no ATP available in the cell: (Lect 25/10)
a. What happens to the Na/K pump?
Answer: The pump stops working without ATP. Na+, K+ and ATP are substrates of this enzyme and without any one, the pump cannot work.
b. What happens to the Na and K gradients?
Answer: The Na+ and K+ gradients collapse in the absence of pump activity due to ions leaking across the plasma membrane, at a rate proportional to PNa and PK.
c. What happens to the membrane potential and why?
Answer: The membrane potential will remain negative because of the presence of intracellular anionic proteins (The Donnan Potential) Note that the Donnan potential is more negative than normal resting Em for most cells.
d. To offset the membrane potential, what happens OR Describe the concept of Donnan Equilibrium?
Answer: If Na+ and Cl- are to be in equilibrium across the membrane of a cell with a negative interior, [Na]i must be greater than [Na]o and [Cl]i must be less than [Cl]o. This will satisfy the electrochemical equilibrium but will cause an osmotic disequilibrium, as more Na particles are present relative to Cl. We now have more solute particles inside the cell than outside. Consequently, water will enter the cell and cause it to swell (The Donnan Equilibrium).
e. What is the final result?
Answer: The cell will burst.
f. What extracellular condition could cause the same net result?
Answer: If the bathing cellular environment were hypoosmotic, water would enter the cell and cause cell swelling. If the cell swells enough, the membrane will lyse. Although the osmolality at which this occurs depends upon the cell type, in general tonicity of less than 150 mOsm is sufficient to cause lysis. Most cells lyse at a volume just over two times the normal volume.
3. Draw a cell with a Na+/K+ pump and a Na+/H+ exchanger. Include the appropriate arrows to show the direction of ion transport. (Lect. 25/13, 26/4, 27/4)
Answer: See picture below.
a. What would happen to [K+]o if the pH in the cell became lower? What happens to [Na]i?
Answer: The pH drop is defined by an increase in the [H+]i. This increases the activity of the Na+/H+ exchanger bringing more Na+ into the cell as H+ is expelled. This decreases the Na+ gradient and decreases the [K+]i/[Na]i (causing increased Na+/K+ ATPase activity). There is a resultant increased K+ transport into the cell which may lead to hypokalemia.
b. Now add the Na+/Ca++ pump and Ca++ slow leak to the diagram. Write the relevant driving force for both the Na+/H+ and the Na+/Ca++ transporters. Discuss the effect of hypoxia on the cell [Ca++]i. (26/7-9, 27/5)
· Na+/H+ transporter is electroneutral. Driving force is DmNa+ - DmH+.
· The Na+/Ca++ transporter is electrogenic. Driving force is 3D˜mNa+ - D˜mCa++. It responds to and contributes to the membrane voltage by transporting one net positive charge (1+).
· Hypoxia forces the cell to use glycolysis and increases H+ production. Decreased intracellular pH activates the Na+/H+ exchanger which decreases the Na+ ion gradient. This decreases the driving force for the Na+/Ca++ which normally pumps Ca++ (secondary active transport) out of the cell to balance the slow leak of Ca++ into the cell. Due to the decrease in Na+/Ca++ transporter activity, Ca++ builds up in the cell and subsequently triggers a number of biochemical pathways which are harmful to the cell (i.e. superoxide production and lipid oxidation).
c. Digitalis is a drug that is often administered to patients with congestive heart failure, a condition characterized by the diminished ability of the heart to generate contractile force. Digitalis is a Na+/K+ pump inhibitor. Explain the mechanism by which digitalis increases cardiac muscle contractile force. (Lect 26/10)
Answer: Treatment of a patient with digitalis will cause partial inhibition of Na+/K+ ATPase activity. This results in increased [Na+]i which inhibits the Na/Ca exchange. This leads to an increased [Ca++]i, which interacts with the cardiac myocyte contractile elements to increase the force of contraction.
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4. Explain how the Na+/H+ exchanger causes absorption of Na+, HCO3-, K+, Cl- and H2O in leaky epithelia (i.e. the proximal tubule of the nephron). Draw a diagram and indicate the lumenal and basolateral membrane surfaces. (Lect 28/4-5)
Answer: Na+/H+ exchange transports H+ to the lumenal fluid and this drives the production of H2O and CO2 in the lumen. The plasma membrane is highly permeable to both water and CO2 and these substances freely cross the membrane to enter the cell and subsequently leave the cell via the basolateral membrane. The result of Na+/H+ exchange activity is the absorption of Na+ and HC03- together with osmotically obliged H2O. (Because H+ is continually extruded from the cell, HCO3- is functionally absorbed). As water leaves the lumen, the concentrations of K+ and Cl- in the lumen increase. The leaky junctional complexes between cells cannot support transmembrane concentration differences and K+, Cl- and water are thus absorbed across the intercellular junction or paracellular pathway.
5. Explain the rationale for the presence of increasingly tight junctional complexes in cells found in the distal portions of the GI tract and nephron. (Lect. 28/6 and summary)
Answer: At the beginning of these systems, the ion and solute concentrations in the lumen are high (equal to those in plasma). Consequently, the body needs to perform crude, mass absorption. In the distal regions of the nephron/GI tract, the tight epithelium and conductive transporters fine tune the concentrations of key ions and expend energy to perfect the lumenal and ECF concentrations before eliminating the residual lumenal contents. Maintenance of extreme concentration gradients between compartments requires energy and high transepithelial potential differences.
6. Draw and describe a cell from the colonic epithelium. Include the type of junction (leaky vs. tight), examples of the transporters present and the relative magnitude of the transepithelial potential. (Lect. 28-pick an example)
Answer: The transepithelial potential is approximately 80 mV (this is high). The potential is generated due to the presence of tight junctions and the conductive ion channels allow fine tuning of the cell/lumenal contents. Ion flow through these channels is coupled to the membrane potential and the concentration gradient. The conductive ion channels are responsive to aldosterone.
7. What is primary active transport? (Lect 25/11)
Answer: ATP is hydrolyzed directly to provide energy to pump ions against a gradient (i.e. Na+/K+ ATPase).
8. What is secondary active transport? (Lect 25/11)
Answer: No ATP is directly hydrolyzed. However, the cell uses the stored potential energy in an ion or solute gradient (previously created by the primary active process) to drive other ions against their concentration gradients.
9. What is electrogenic (conductive) transport? (Lect 27/1-5)
Answer: This is a transport process (active or passive) that results in a NET current flow that will directly change the membrane potential.
a. Provide several examples of electrogenic transporters.
Answer: Examples of electrogenic transporters include single ion channels, Na+/K+ ATPase (because the 3:2 ratio results in a charge difference) and the Na+/Ca++ transporter (3:1 ratio).
b. What two forces drive conductive transport?
Answer: The forces that drive conductive transport are (1) the chemical potential energy difference (the energy stored in the ion concentration gradient) and (2) the electrical potential difference (the voltage that exists across the membrane).
10. What is electroneutral transport? (Lect 27/3-4)
Answer: This is a form of transport (active or passive) that results in no direct change in electrical potential. Examples include the Cl-/HCO3- exchanger, the Na+/K+/2Cl- cotransporter and the H+/K+ ATPase found in the kidney.
11. When no concentration gradient exists for a given ion across a membrane, then by definition the ion is in equilibrium. True or false? (Lect 25/2-3)
Answer: False. This state would be defined as equilibrium for the ion only if the membrane voltage was also zero so that the electrochemical potential difference is zero. Both electrical and chemical forces determine the equilibrium of an ion.
12. Which type of transporter (electroneutral or conductive) is better suited to high capacity and/or regulatory processes in the cell? Why? (Lect 26/3)
Answer: The electroneutral transporter is better suited to high capacity transport because it is dependent only on concentration differences. If a conductive transporter was used, the membrane would eventually charge and the electrical potential would stop ion transport thus limiting the capacity or regulatory ability of the transporter.
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13. Explain how the condition of chloridorrhea (congenital absence of the Cl-/HCO3- exchanger) results in diarrhea and metabolic alkalosis. Draw a normal cell and show where the defect occurs in chloridorrhea. (Lect 28/6)
Answer: In chloridorrhea, the Cl-/HCO3- exchanger is not present in the ileum while the Na+/H+ exchanger continues to function. H+ is transported from the cell into the lumen where it combines with HCO3- to produce H2O and CO2. Water and CO2 freely cross the plasma membrane and normally Chlorine would be absorbed and the bicarbonate ion would be recycled via the parallel Na+/H+ and Cl-/HCO3- transporters. In patients with chloridorrhea, Cl-/HCO3- transporters are absent and thus cannot participate in the net absorption of Na+ and Cl-. This functional uncoupling of the pumps results in a buildup of HCO3- in the plasma leading to metabolic alkalosis. The activity of the Na+/H+ exchange is decreased due to the increased lumenal [H+] and Cl- is unable to enter the cell due to the absence of the exchanger. The result is a lumenal fluid which is highly acidic with a high chloride ion concentration. The mass of Cl- solute in the lumen causes an osmotic diuresis and massive, watery diarrhea.
14. Many years ago, a child was disciplined by being forced to drink water from a garden hose until her mother told her to stop. She unfortunately lapsed into a coma before her mother stopped her. Although she was brought to the hospital, she died shortly thereafter. (Lect 26/1-2)
a. What ions control the volume of cells in a hypo/hyperosmotic environment?
Answer: Swollen cells regulate volume as a result of net K and Cl loss, while shrunken cells gain volume as a result of net uptake of Na and Cl or Na, K, and Cl.
b. What is the problem with this volume control mechanism, particularly in a small child (with a small total water volume)?
Answer: This system can control cell volume when given sufficient time to adapt or when changes in the extracellular environment are not extreme. In a child who has a small total water volume, the addition of a liter of pure water can cause huge drops in the osmolarity of the extracellular fluid. The cellular volume regulatory mechanisms can not compensate for extremely large changes in osmolarity which occur rapidly.
c. Why did this child go into a coma and die?
Answer: The primary problem was that all of her cells became swollen. In most areas of the body, this is not a problem. However, because the brain is extremely limited in its capacity to expand (low extracellular space in CNS)without irreversible damage, swelling of her brain caused herniation which resulted in coma and death.
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d. Which of these transporters would the cell use to try to compensate for cell swelling? Draw each transporter and describe how it is limited.
i. Na+/K+/2Cl- cotransporter: will increase cell volume.
ii. K+/Cl- cotransporter : will act to reduce cell swelling.
The limitation of the K+/Cl- cotransporter is the [Cl-]i.
K+/Cl- cotransporter Na+/K+/2Cl- cotransporter
e. Which transporter might help the cell overcome the limitation of the K+/Cl- cotransporter and how will this further affect the cell?
Answer: The cell can use the Cl-/HCO3- transporter to move Cl- into the cell, which would reestablish the Cl driving gradient for pushing K out. This would also result in HCO3- moving out of the cell, making the interior more acidic and the extracellular fluid more alkalotic. The key concept to understand is that the cell is able to utilize functional coupling of ion transporters to produce changes in the cell's environment.
15. If [K]i is 125 mM and [K]o is 4 mM, what is the maximum value (and sign) of the membrane potential that could be expected to result from K+ flux? (Lect 25/3)
Answer: -91 mV (EK=RT/ZF ln [K]o/[K]i)
If [Na]i is 15 mM and [Na]o is 150 mM, what is the maximum value and sign that the membrane potential might attain as a result of Na+ flux?
Answer: +61 mV
16. Why is creatinine used as a marker substance for GFR determinations in clinical settings? (Lect 29/12) OR What are the advantages of using inulin instead of creatinine? Disadvantages? (Lect 29/11-12) Why is PAH (Para-amino Hippourate) used to measure Renal Plasma Flow? (Lect 29/11)
Answer: Creatinine is an endogenous substance, derived form the metabolism of creatine in skeletal muscle, that fulfills almost all of the requirements for a marker substance. Creatinine is released from muscle at a constant rate. This will result in a stable plasma concentration if it is also excreted at an equivalent rate. It is freely filterable, it is not metabolized, and is not reabsorbed once filtered. There is a small amount of tubular secretion that makes creatinine clearance a slight overestimation of the GFR. (This overestimate is only quantitatively significant at low levels of GFR).