5. Ion Channels Flashcards
What does it mean to say that the ion channels of nerve cells are heterogeneous?
In different parts of the nervous system different types of channels carry out specific signalling tasks.
What often results from the malfunctioning of ion channels?
Because of their key roles in electrical signalling, malfunctioning of ion channels can cause a wide variety of neurological diseases.
Why do ion channels play a crucial role in both the normal physiology and pathophysiology of the nervous system?
Ion channels are often the site of action of drugs, poisons, or toxins. Thus ion channels have crucial roles in both the normal physiology and pathophysiology of the nervous system.
In addition to ion channels, nerve cells contain a second important class of proteins with a similar function. What are these?
In addition to ion channels, nerve cells contain a second important class of proteins specialised for moving ions across cell membranes, the ion trans- porters or pumps.
What role do these ion pumps play in neuronal signalling?
These proteins do not participate in rapid neuronal signalling but rather are important for establishing and maintaining the concentration gradients of physiologically important ions between the inside and outside of the cell.
What three important properties do ion channels have?
(1) They recognise and select specific ions
(2) They open and close in response to specific electrical, mechanical, or chemical signals
(3) They conduct ions across the membrane.
What causes the large flow if electric charge during an action potential?
The channels in nerve and muscle conduct ions across the cell membrane at extremely rapid rates, thereby providing a large flow of electric charge. Up to 100 million ions can pass through a single channel each second. This current causes the rapid changes in membrane potential required for signalling
What is a surprising feature of these ion channels given the fast turnover rate?
In light of such an extraordinary rate of ion flow, channels are surprisingly selective for the ions they allow to permeate. Each type of channel allows only one or a few types of ions to pass.
How do ion channels ‘know’ when to open?
Many channels open and close in response to a specific event: Voltage-gated channels are regulated by changes in membrane potential, ligand-gated channels by chemical transmitters, and mechanically gated channels by pressure or stretch. However, some channels are normally open in the cell at rest. The ion flux through these “resting” channels contributes significantly to the resting potential.
How are ion channels limited in a manner in which ion pumps are not?
Ion channels are limited to catalyzing the passive movement of ions down their thermodynamic concentration and electrical gradients. For example, Na+ ions enter a cell through voltage-gated Na+ channels during an action potential because the external Na+ concentration is much greater than the internal concentration; the open channels allow Na+ to diffuse into the cell down its concentration gradient. With such passive ion movement the Na+ concentration gradient would eventually dissipate were it not for
ion pumps.
Name four ions in which ion pumps manage
Different types of ion pumps maintain the concentration gradients for Na+ , K+ , Ca 2+ , Cl- , and other ions.
These pumps differ from ion channels in two important details. Describe these
First, whereas open ion channels have a continuous water-filled pathway through which ions flow unimpeded from one side of the membrane to the other, each time a pump moves an ion, or a group of a few ions, across the membrane, it must undergo a series of conformational changes. As a result, the rate of ion flow through pumps is 100 to 100,000 times slower than through channels.
Second, pumps that maintain ion gradients use energy, often in the form of adenosine triphosphate (ATP), to transport ions against their electrical and chemical gradients. Such ion movements are termed active transport.
Describe the plasma membrane of nerve cells
The plasma membrane of all cells, including nerve cells, is approximately 6 to 8 nm thick and consists of a mosaic of lipids and proteins. The core of the membrane is formed by a double layer of phospholipids. Embedded within this continuous lipid sheet are integral membrane proteins, including ion channels.
How does the lipid membrane differ to that of the ions within and outside in terms of what they attract?
The lipids of the membrane do not mix with water—they are hydrophobic. In contrast, the ions within the cell and those outside strongly attract water molecules—they are hydrophilic.
Why do the ions attract water?
Ions attract water because water molecules are dipolar: Although the net charge on a water molecule is zero, charge is separated within the molecule. The oxygen atom in a water molecule tends to attract electrons and so bears a small net negative charge, whereas the hydrogen atoms tend to lose electrons and therefore carry a small net positive charge. As a result of this unequal distribution of charge, positively charged ions (cations) are strongly attracted electrostatically to the oxygen atom of water, and negatively charged ions (anions) are attracted to the hydrogen atoms. Similarly, ions attract water; in fact they become surrounded by electrostatically bound waters of hydration
What is meant by ‘electrostatic’? **
relating to stationary electric charges or fields as opposed to electric currents.
Can an ion move away from water into the noncharged hydrocarbon tails of the lipid bilayer in the membrane? Why not or how?
An ion cannot move away from water into the noncharged hydrocarbon tails of the lipid bilayer in the membrane unless a large amount of energy is expended to overcome the attraction between the ion and the surrounding water molecules. For this reason it is extremely unlikely that an ion will move from solution into the lipid bilayer, and therefore the bilayer itself is almost completely impermeable to ions.
Why can ion channel selectivity not be based solely on the diameter of the ion?
Selectivity cannot be based solely on the diameter of the ion because K , with a crystal radius of 0.133 nm, is larger than Na+ (crystal radius of 0.095 nm).
How can a water-filled channel conduct ions at high rates and yet be selective? How, for instance, does a channel allow K+ ions to pass while excluding Na+ ions?
Because ions in solution are surrounded by waters of hydration, the ease with which an ion moves in solution (its mobility or diffusion constant) depends on the size of the ion together with the shell of water surrounding it. The smaller an ion, the more highly localised is its charge and the stronger its electric field. As a result, smaller ions attract water more strongly. Thus, as Na+ moves through solution its stronger electrostatic attraction for water causes it to have a larger water shell, which tends to slow it down relative to K .
Because of its larger water shell, Na+ behaves as if it is larger than K+ . In fact, there is a precise relationship between the size of an ion and its mobility in solution: the smaller the ion, the lower its mobility. We therefore can construct a model of a channel that selects K+ rather than Na+ simply on the basis of the interaction of the two ions with water in a water-filled channel
Although this model explains how a channel can select K+ and exclude Na+ , where does it fall short? How has this developed since?
It does not explain how a channel could select Na+ and exclude K+ . Moreover, the model cannot account quantitatively for the very high ionic selectivity exhibited by biological K+ channels.
Even though we now know that ions can cross membranes by means of a variety of transport macro-molecules, the Na -K pump being a well-characterised example, many properties of membrane ion conductances do not fit the carrier model.
What is the most important characteristic of ion channels which is not accounted for in this model? How do we know of this characteristic?
Most important is the rapid rate of ion transfer across membranes. This transfer rate was first examined in the early 1970s by measuring the transmembrane current initiated when the neurotransmitter acetylcholine (ACh) binds its receptor in the cell membrane of skeletal muscle fibers at the synapse between nerve and muscle. Using measurements of membrane current noise, small statistical fluctuations in the mean ionic current induced by ACh, Bernard Katz and Ricardo Miledi concluded that the current conducted by a single ACh receptor is 10 million ions per second. In contrast, the Na+ -K+ pump transports at most 100 ions per second.
What is this rate of transport evidence against?
If the ACh receptor acted as a carrier, it would
have to shuttle an ion across the membrane in 0.1 μs
(one ten-millionth of a second), an implausibly fast
rate. The 100,000-fold difference in rates strongly suggests that the ACh receptor (and other ligand-gates receptors) must conduct ions through a channel.
But we are still left with the problem of what makes a channel selective. How did Bertil Hille explain selectivity?
To explain selectivity, Bertil Hille extended the pore theory by proposing that channels have narrow regions that act as molecular sieves. At this selectivity filter an ion must shed most of its waters of hydration to traverse the channel; in their place weak chemical bonds (electrostatic interactions) form with polar (charged) amino acid residues that line the walls of the channel. Because shedding its waters of hydration is energetically unfavourable, the ion will traverse a channel only if its energy of interaction with the selectivity filter compensates for the loss of the energy of interaction with its waters of hydration. Ions traversing the channel are normally bound to the selectivity filter for only a short time (less than 1 μs), after which electrostatic and diffusional forces propel the ion through the channel. In channels where the pore diameter is large enough to accommodate several water molecules, an ion need not be stripped completely of its water shell.
How is this chemical recognition and specificity of ion channels established? Describe one theory
According to one theory was developed in the early 1960s by George Eisenman, a binding site with a high negative field strength—for example, one formed by negatively charged carboxylic acid groups of glutamate or aspartate — will bind Na+ more tightly than K+. This selectivity results because the electrostatic interaction between two charged groups, as governed by Coulomb’s law, depends inversely on the distance between the two groups.
Because Na has a smaller crystal radius than K , it will approach a negative site more closely than K+ and thus will derive a more favourable free-energy change on binding. This compensates for the requirement that Na+ lose some of its waters of hydration to traverse the narrow selectivity filter. In contrast, a binding site with a low negative strength—one that is composed, for example, of polar carbonyl or hydroxyl oxygen atoms—would select K over Na .
At such a site the binding of Na+ would not provide a sufficient free-energy change to compensate for the loss of the ion’s waters of hydration, which Na+ holds strongly. However, such a site would be able to compensate for the loss of a K+ ion’s associated water molecules as the larger K+ ions interact more weakly with water.
What is Coloumb’s law? **
According to Coulomb, the electric force for charges at rest has the following properties: Like charges repel each other; unlike charges attract.
He figured out that the force between two charges is equal to some constant times the first charge times the second charge divided by the distance between them squared.
What is charge? **
It’s a property of objects and particles like mass. We know that like repels like and opposites attract but eve the names positive and negative are arbitrary. o one fundamentally knows what it is.
Is the current thinking that ion channels are selective because of specific chemical interactions or because of molecular sieving based on pore diameter?
It is currently thought that ion channels are selective both because of specific chemical interactions and because of molecular sieving based on pore diameter.
X-ray crystallographic and other structural analy- ses have been informative in the study of enzymes and other soluble proteins. Why have they only recently bee applied to membrane proteins?
They have only recently been applied to integral membrane proteins, such as ion channels, because their transmembrane hydrophobic regions make them difficult to crystallise.
What alternative method has been informative regarding the structure of these channels?
For the past 35 years single-channel recording has provided important functional information that has yielded significant insights into channel structure.
Before it became possible to resolve the small amount of current that flows through a single ion channel in biological membranes, how was channel function studied?
Before it became possible to resolve the small amount of current that flows through a single ion channel in biological membranes, channel function was studied in artificial lipid bilayers.
Describe how Paul Mueller and Donald Rudin developed a technique for studying lipid bilayers in the early 60’s
In the early 1960s Paul Mueller and Donald Rudin developed a technique for forming functional lipid bilayers by painting a thin drop of phospholipid over a hole in a nonconducting barrier that separates two salt solutions.
How did Paul Mueller and Donald Rudin study the permeability of this artificial lipid bilayer? What did they find?
Although lipid membranes are highly impermeable to ions, ionic permeability of the membrane increases dramatically when certain peptide antibiotics are added to the salt solution. Early studies with gramicidin A, a 15-amino acid cyclic peptide, were especially informative. Application of low concentrations of gramicidin A brings about small step- like changes in current across the membrane. These brief pulses of current reflect the all-or-none opening and closing of the single ion channel formed by the peptide.
What is the relationship between the current through a single gramicidin channel and the membrane potential?
The current through a single gramicidin channel varies with membrane potential in a linear manner; that is, the channel behaves as a simple resistor.
How can the amplitude of the single-channel current be obtained?
The amplitude of the single-channel current can thus be obtained from Ohm’s law, i = V/R, where i is the current through the single channel, V is the voltage across the membrane, and R is the resistance of the open channel.
What is the approximate resistance of a single open channel?
The resistance of a single open channel is approximately 8 × 10^10 ohms
We don’t usually speak of resistance when it comes to channels, what do we speak of instead and why?
In dealing with ion channels it is more useful to speak of the reciprocal of resistance or conductance (γ = 1/R), as this provides an electrical measure related to ion permeability.
How can conductance be used in Ohm’s law?
Thus, Ohm’s law can be expressed as
i = γ × V
What is the approximate conductance of the gramicidin A channel?
The conductance of the gramicidin A channel is approximately 12 × 10^−12 siemens (S), or 12 picosiemens (pS), where 1 S equals 1/ohm.
Have basic channel properties obtained from artificial membranes been confirmed? What is preventing this, or how has it been confirmed?
The insights into basic channel properties obtained from artificial membranes were later confirmed in biological membranes by the patch-clamp technique, developed by Erwin Neher and Bert Sakmann in 1976
Describe how the patch clamp technique was first applied to obtain evidence for these channel properties
A glass micropipette containing ACh—the neurotransmitter that activates the ACh receptors in the membrane of skeletal muscle—was pressed tightly against a frog muscle membrane. Small unitary current pulses representing the opening and closing of a single ACh receptor channel were recorded from the area of the membrane under the pipette tip. As with gramicidin A channels, the relation between current and voltage in the ACh receptor channel is linear, with a single-channel conductance of approximately 25 pS. This generates a unitary current of 2 pA (picoamperes) at a membrane potential of −80 mV (millivolts), which corresponds to a flux of around 12.5 million ions per second.
Christopher Miller independently developed a method for incorporating channels from biological membranes into artificial lipid bilayers. Describe this method
Biological membranes are first homogenised in a laboratory blender; centrifugation (a technique used for the separation of particles from a solution according to their size, shape, density, viscosity of the medium and rotor speed) of the homogenate then separates out a portion composed only of membrane vesicles. Under appropriate ionic conditions these membrane vesicles will fuse with a planar lipid membrane, incorporating any ion channel in the vesicle into the planar membrane.
This technique of incorporating channels from biological membranes into artificial lipid bilayers has two experimental advantages. Describe them
First, it allows recording from ion channels in regions of cells that are inaccessible to patch clamp; for example, Miller has successfully studied a K+ channel isolated from the internal membrane of skeletal muscle (the sarcoplasmic reticulum).
Second, it allows researchers to study how the composition of the membrane lipids influences channel function.
To what extent are all cells capable of signalling?
Most cells are capable of local signalling, but only nerve and muscle cells are specialised for rapid signalling over long distances.
How is the direction of the flux of ions through a channel determined?
The direction and eventual equilibrium for this flux are determined not by the channel itself, but rather by the electrostatic and diffusional driving forces across the membrane.
Ion channels allow particular types of ions to cross the membrane, selecting either cations or anions to permeate.
What are cations and anions?
Cation: a positively charged ion, i.e. one that would be attracted to the cathode in electrolysis.
Anion: Anions are ions that are negatively charged. Ions are charged atoms or molecules. If a balanced atom loses one or more electrons, it will become a positively charged cation.
What ions can pass through cation-selective channels? (4)
Some types of cation-selective channels
allow the cations that are usually present in extracellular fluid—Na+ , K+ , Ca2+ , and Mg2+ —to pass almost indiscriminately. However, many other cation- selective channels are permeable primarily to a single type of ion, whether it is Na+, K+ ,or Ca 2+.
What are most anion selective channels selective for?
Most types of anion-selective channels are also highly discriminating; they conduct only one physiological ion, chloride (Cl−).
How are the kinetic properties of ion permeation best described?
The kinetic properties of ion permeation are best described by the channel’s conductance, which is deter- mined by measuring the current (ion flux) through the open channel in response to an electrochemical driving force.
How is the net electrochemical driving force determined?
The net electrochemical driving force is determined by two factors: the electrical potential difference across the membrane and the concentration gradients of the permeant ions across the membrane. Changing either one can change the net driving force
Describe the relationship between the current and the driving force in ion channels in regards to their function characteristics
As we have seen, in some open channels the cur- rent varies linearly with driving force—that is, the channels behave as simple resistors. In others the cur- rent is a nonlinear function of driving force. This type of channel behaves as a rectifier—it conducts ions more readily in one direction than in the other because of asymmetry in the channel’s structure or environment. Whereas the conductance of a resistor-like channel is constant—it is the same at all voltages—the conductance of a rectifying channel is variable and must be determined by plotting current versus voltage over the entire physiological range of membrane potential (In figures)
How does the net electrochemical driving force depend on concentration gradients of the permeant ions across the membrane?
The rate of net ion flux (current) through a channel depends on the concentration of the permeant ions in the surrounding solution. At low concentrations the current increases almost linearly with concentration. At higher concentrations the current tends to reach a point at which it no longer increases. At this point the current is said to saturate.
This saturation effect is consistent with what idea involving transmembrane current?
This saturation effect is consistent with the idea that ion flux across the cell membrane is not strictly a function of laws of electrochemical diffusion in free solution but also involves the binding of ions to specific polar sites within the pore of the channel. A simple electrodiffusion model would predict that the ionic current should continue to increase as long as the ionic concentration also increases; that is, the more charge carriers in solution, the greater the current.
The relation between current and ionic concentration for a wide range of ion channels is well described by what equation? What could this suggest?
The relation between current and ionic concentration for a wide range of ion channels is well described by a simple chemical binding equation, identical to the Michaelis-Menten equation for enzymes, suggesting that a single ion binds within the channel during permeation. The ionic concentration at which current reaches half its maximum defines the dissociation constant, the concentration at which half of the channels will be occupied by a bound ion.
How strong is the binding between the ion and the channel and how do we know?
The dissociation constant in plots of current versus concentration is typically quite high, approximately 100 mM, indicating weak binding. (In typical interactions between enzymes and substrates the dissociation constant is below 1 μM.)
What does this weak binding of the ion and ion channel suggest?
This weak interaction indicates that the bonds between the ion and the channel are rapidly formed and broken. In fact, an ion typically stays bound in the channel for less than 1 μs. The rapid rate at which an ion unbinds is necessary for the channel to achieve the very high conduction rates responsible for the rapid changes in membrane potential during signalling.
What is meant by the term occlusion and how is it relevant to ion channels?
Some ion channels are susceptible to occlusion (the blockage or closing of a blood vessel or hollow organ) by certain free ions or molecules in the cytoplasm or extracellular fluid. Passage through the channel can be blocked by particles that bind either to the mouth of the aqueous pore or somewhere within the pore. If the blocker is an ionised molecule that binds to a site within the pore, it will be influenced by the membrane electric field as it enters the channel
Give an example of how free ion blocking a channel can be influenced by the membrane electric field as it enters the channel
If a positively charged blocker enters the channel from outside the membrane, then making the cytoplasmic side of the membrane more negative—which, according to convention, corresponds to a more negative membrane potential—will drive the blocker into the channel through an electrostatic attraction, increasing the block.