Cell Biology Exam #1

QUESTION 12-10 Which of the following statements are correct? Explain your answers. A. The plasma membrane is highly impermeable to all charged molecules. B. Channels have specific binding pockets for the solute molecules they allow to pass. C. Transporters allow solutes to cross a membrane at much faster rates than do channels. D. Certain H+ pumps are fueled by light energy. E. The plasma membrane of many animal cells contains open K+ channels, yet the K+ concentration in the cytosol is much higher than outside the cell. F. A symport would function as an antiport if its orientation in the membrane were reversed (i.e., if the portion of the molecule normally exposed to the cytosol faced the outside of the cell instead). G. The membrane potential of an axon temporarily becomes more negative when an action potential excites it.

ANSWER 12-10 A. False. The plasma membrane contains transport proteins that confer selective permeability to many but not all charged molecules. In contrast, a pure lipid bilayer lacking proteins is highly impermeable to all charged molecules. B. False. Channels do not have binding pockets for the solute that passes through them. Selectivity of a channel is achieved by the size of the internal pore and by charged regions at the entrance of the pore that attract or repel ions of the appropriate charge. C. False. Transporters are slower. They have enzymelike properties, i.e., they bind solutes and need to undergo conformational changes during their functional cycle. This limits the maximal rate of transport to about 1000 solute molecules per second, whereas channels can pass up to 1,000,000 solute molecules per second. D. True. The bacteriorhodopsin of some photosynthetic bacteria pumps H+ out of the cell, using energy captured from visible light. E. True. Most animal cells contain K+ leak channels in their plasma membrane that are predominantly open. The K+ concentration inside the cell still remains higher than outside, because the membrane potential is negative and therefore inhibits the positively charged K+ from leaking out. K+ is also continually pumped into the cell by the Na+ pump. F. False. A symport binds two different solutes on the same side of the membrane. Turning it around would not change it into an antiport, which must also bind to different solutes, but on opposing sides of the membrane. G. False. The peak of an action potential corresponds to a transient shift of the membrane potential from a negative to a positive value. The influx of Na+ causes the membrane potential first to move toward zero and then to reverse, rendering the cell positively charged on its inside. Eventually, the resting potential is restored by an efflux of K+ through voltage-gated K+ channels and K+ leak channels.

QUESTION 12-11 List the following compounds in order of increasing lipid bilayer permeability: RNA, Ca2+, glucose, ethanol, N2, water.

ANSWER 12-11 The permeabilities are N2 (small and nonpolar) > ethanol (small and slightly polar) > water (small and polar) > glucose (large and polar) > Ca2+ (small and charged) > RNA (very large and charged).

QUESTION 12-12 Name at least one similarity and at least one difference between the following (it may help to review the definitions of the terms using the Glossary): A. Symport and antiport B. Active transport and passive transport C. Membrane potential and electrochemical gradient D. Pump and transporter E. Axon and telephone wire F. Solute and ion

ANSWER 12-12 A. Both couple the movement of two different solutes across a cell membrane. Symports transport both solutes in the same direction, whereas antiports transport the solutes in opposite directions. B. Both are mediated by membrane transport proteins. Passive transport of a solute occurs downhill, in the direction of its concentration or electrochemical gradient, whereas active transport occurs uphill and therefore needs an energy source. Active transport can be mediated by transporters but not by channels, whereas passive transport can be mediated by either. C. Both terms describe gradients across a membrane. The membrane potential refers to the voltage gradient; the electrochemical gradient is a composite of the voltage gradient and the concentration gradient of a specific charged solute (ion). The membrane potential is defined independently of the solute of interest, whereas an electrochemical gradient refers to the particular solute. D. A pump is a specialized transporter that uses energy to transport a solute uphill—against an electrochemical gradient for a charged solute or a concentration for an uncharged solute. E. Both transmit electrical signals by means of electrons in wires and ion movements across the plasma membrane in axons. Wires are made of copper, axons are not. The signal passing down an axon does not diminish in strength, because it is self-amplifying, whereas the signal in a wire decreases over distance (by leakage of current across the insulating sheath). F. Both affect the osmotic pressure in a cell. An ion is a solute that bears a charge.

QUESTION 12-13 Discuss the following statement: "The differences between a channel and a transporter are like the differences between a bridge and a ferry."

ANSWER 12-13 A bridge allows vehicles to pass over water in a steady stream; the entrance can be designed to exclude, for example, oversized trucks, and it can be intermittently closed to traffic by a gate. By analogy, gated channels allow ions to pass across a cell membrane, imposing size and charge restrictions. A ferry, in contrast, loads vehicles on one side of the body of water, crosses, and unloads on the other side—a slower process. During loading, particular vehicles could be selected from the waiting line because they fit particularly well on the car deck. By analogy, transporters bind solutes on one side of the membrane and then, after a conformational movement, release them on the other side. Specific binding selects the molecules to be transported. As in the case of coupled transport, sometimes you have to wait until the ferry is full before you can go.

QUESTION 12-14 The neurotransmitter acetylcholine is made in the cytosol and then transported into synaptic vesicles, where its concentration is more than 100-fold higher than in the cytosol. When synaptic vesicles are isolated from neurons, they can take up additional acetylcholine added to the solution in which they are suspended, but only when ATP is present. Na+ ions are not required for the uptake, but, curiously, raising the pH of the solution in which the synaptic vesicles are suspended increases the rate of uptake. Furthermore, transport is inhibited when drugs are added that make the membrane permeable to H+ ions. Suggest a mechanism that is consistent with all of these observations.

ANSWER 12-14 Acetylcholine is being transported into the vesicles by an H+-acetylcholine antiport in the vesicle membrane. The H+ gradient that drives the uptake is generated by an ATP-driven H+ pump in the vesicle + membrane, which pumps H into the vesicle (hence the dependence of the reaction on ATP). Raising the pH of the solution surrounding the vesicles decreases the H+ concentration of the solution, thereby increasing the outward gradient across the vesicle membrane, explaining the enhanced rate of acetylcholine uptake.

QUESTION 12-15 The resting membrane potential of a typical animal cell is about -70 mV, and the thickness of a lipid bilayer is about 4.5 nm. What is the strength of the electric field across the membrane in V/cm? What do you suppose would happen if you applied this field strength to two metal electrodes separated by a 1-cm air gap?

ANSWER 12-15 The voltage gradient across the membrane is about 150,000 V/cm (70 × 10-3 V/4.5 × 10-7 cm). This extremely powerful electric field is close to the limit at which insulating materials—such as the lipid bilayer— break down and cease to act as insulators. The large field indicates what a large amount of energy can be stored in electrical gradients across the membrane, as well as the extreme electrical forces that proteins can experience in a membrane. A voltage of 150,000 V would instantly discharge in an arc across a 1-cm-wide gap (that is, air would be an insufficient insulator for this strength of field).

QUESTION 12-16 Phospholipid bilayers form sealed spherical vesicles in water (discussed in Chapter 11). Assume you have constructed lipid vesicles that contain Na+ pumps as the sole membrane protein, and assume for the sake of simplicity that each pump transports one Na+ one way and one K+ the other way in each pumping cycle. All the Na+ pumps have the portion of the molecule that normally faces the cytosol oriented toward the outside of the vesicles. With the help of Figure 12-11, determine what would happen if: A. Your vesicles were suspended in a solution containing both Na+ and K+ ions and had a solution with the same ionic composition inside them. B. You add ATP to the suspension described in (A). C. You add ATP, but the solution—outside as well as inside the vesicles—contains only Na+ ions and no K+ ions. D. The concentrations of Na+ and K+ were as in (A), but half of the pump molecules embedded in the membrane of each vesicle were oriented the other way around so that the normally cytosolic portions of these molecules faced the inside of the vesicles. You then add ATP to the suspension. E. You add ATP to the suspension described in (A), but in addition to Na+ pumps, the membrane of your vesicles also contains K+ leak channels.

ANSWER 12-16 A. Nothing. You require ATP to drive the Na+ pump. B. The ATP becomes hydrolyzed, and Na+ is pumped into the vesicles, generating a concentration gradient of Na+ across the membrane. At the same time, K+ is pumped out of the vesicles, generating a concentration gradient of K+ of opposite polarity. When all the K+ is pumped out of the vesicle or the ATP runs out, the pump would stop. C. The pump would initiate a transport cycle and then cease. Because all reaction steps must occur strictly sequentially, dephosphorylation and the accompanying conformational switch cannot occur in the absence of K+. The Na+ pump will therefore become stuck in the phosphorylated state, waiting indefinitely for a potassium ion. The number of sodium ions transported would be minuscule, because each pump molecule would have functioned only a single time. Similar experiments, leaving out individual ions and analyzing the consequences, were used to determine the sequence of steps by which the Na+ pump works. D. ATP would become hydrolyzed, and Na+ and K+ would be pumped across the membrane as described in (B). However, the pump molecules that sit in the membrane in the reverse orientation would be completely inactive E. (i.e., they would not—as one might have erroneously assumed—pump ions in the opposite direction), because ATP would not have access to the site on these molecules where phosphorylation occurs, which is normally exposed to the cytosol. ATP is highly charged and cannot cross membranes without the help of specific transporters. ATP becomes hydrolyzed, and Na+ and K+ are pumped across the membrane, as described in (B). K+, however, immediately flows back into the vesicles through the K+ leak channels. K+ moves down the K+ concentration gradient formed by the action of the Na+ pump. With each K+ that moves into the vesicle through a leak channel, a positive charge is moved across the membrane, generating a membrane potential that is positive on the inside of the vesicles. Eventually, K+ will stop flowing through the leak channels when the membrane potential balances the K+ concentration gradient. The scenario described here is a slight oversimplification: the Na+ pump in mammalian cells actually moves three sodium ions out of cells for each two potassium ions that it pumps, thereby driving an electric current across the membrane and making a small additional contribution to the resting membrane potential (which therefore corresponds only approximately to a state of equilibrium for K+ moving via K+ leak channels).

QUESTION 12-17 Name the three ways in which an ion channel can be gated.

ANSWER 12-17 Ion channels can be ligand-gated, voltage- gated, or mechanically (stress) gated.

QUESTION 12-18 One thousand Ca2+ channels open in the plasma membrane of a cell that is 1000 m3 in size and has a cytosolic Ca2+ concentration of 100 nM. For how long would the channels need to stay open in order for the cytosolic Ca2+ concentration to rise to 5 M? There is virtually unlimited Ca2+ available in the outside medium (the extracellular Ca2+ concentration in which most animal cells live is a few millimolar), and each channel passes 106 Ca2+ ions per second.

ANSWER 12-18 The cell has a volume of 10-12 liters (= 10-15 m3) and thus contains 6 × 104 calcium ions (= 6 × 1023 molecules/mole × 100 × 10-9 moles/liter × 10-12 liters). Therefore, to raise the intracellular Ca2+ concentration fiftyfold, another 2,940,000 calcium ions have to enter the cell (note that at 5 M concentration there are 3 × 106 ions in the cell, of which 60,000 are already present before the channels are opened). Because each of the 1000 channels allows 106 ions to pass per second, each channel has to stay open for only 3 milliseconds.

QUESTION 12-19 Amino acids are taken up by animal cells using a symport in the plasma membrane. What is the most likely ion whose electrochemical gradient drives the import? Is ATP consumed in the process? If so, how?

ANSWER 12-19 Animal cells drive most transport processes across the plasma membrane with the electrochemical gradient of Na+. ATP is needed to fuel the Na+ pump to maintain the Na+ gradient.

QUESTION 12-20 We will see in Chapter 15 that endosomes, which are membrane-enclosed intracellular organelles, need an acidic lumen in order to function. Acidification is achieved by an H+ pump in the endosomal membrane, which also contains Cl- channels. If the channels do not function properly (e.g., because of a mutation in the genes encoding the channel proteins), acidification is also impaired. A. Can you explain how Cl- channels might help acidification? B. According to your explanation, would the Cl- channels be absolutely required to lower the pH inside the endosome?

ANSWER 12-20 A. If H+ is pumped across the membrane into the endosomes, an electrochemical gradient of H+ results— composed of both an H+ concentration gradient and a membrane potential, with the interior of the vesicle positive. Both of these components add to the energy that is stored in the gradient and that must be supplied to generate it. The electrochemical gradient will limit the transfer of more H+. If, however, the membrane also contains Cl- channels, the negatively charged Cl- in the cytosol will flow into the endosomes and diminish their membrane potential. It therefore becomes energetically less expensive to pump more H+ across the membrane, and the interior of the endosomes can become more acidic. B. No. As explained in (A), some acidification would still occur in their absence.

QUESTION 12-22 Acetylcholine-gated cation channels do not discriminate between Na+, K+, and Ca2+ ions, allowing all to pass through them freely. So why is it that when acetylcholine binds to this protein in the plasma membrane of muscle cells, the channel opens and there is a large net influx of primarily Na+ ions?

ANSWER 12-22 The membrane potential and the steep extracellular Na+ concentration provide a large inward electrochemical driving force and a large reservoir of Na+ ions, so that mostly Na+ ions enter the cell as acetylcholine receptors open. Ca2+ ions will also enter the cell, but their influx is much more limited because of their lower extracellular concentration. (Most of the Ca2+ that enters the cytosol to stimulate muscle contraction is released from intracellular stores, as we discuss in Chapter 17). Because of the high intracellular K+ concentration and the opposing direction of the membrane potential, there will be little if any movement of K+ ions upon opening of a cation channel.

QUESTION 12-23 The ion channels that are regulated by binding of neurotransmitters, such as acetylcholine, glutamate, GABA, or glycine, have a similar overall structure. Yet, each class of these channels consists of a very diverse set of subtypes with different transmitter affinities, different channel conductances, and different rates of opening and closing. Do you suppose that such extreme diversity is a good or a bad thing from the standpoint of the pharmaceutical industry?

ANSWER 12-23 The diversity of neurotransmitter-gated ion channels is a good thing for the industry, as it raises the possibility of developing new drugs specific for each channel type. Each of the diverse subtypes of these channels is expressed in a narrow subset of neurons. This narrow range of expression should make it possible, in principle, to discover or design drugs that affect particular receptor subtypes present in a selected set of neurons, thus to target particular brain functions with greater specificity.