Ion channels and Membrane potentials

Question 1

Voltage gated channels of the KV, NaV and CaV families have ___ membrane-spanning domains, each of which contains ____ alpha helices labeled______.

Answer 1

four; six; S1-S6

Question 2

What is the structural difference between the assembly of the four subunits in the KV channels vs. the CaV/NaV channels?

Answer 2

In KV channels, each domain is a separate polypeptide, and four of these assemble to form the channel. In NaV and CaV the four domains (I, II, III, and IV) are linked into a single polypeptide.

Question 3

Describe specific structures of NaV and KV channels that serve as the voltage sensors.

Answer 3

Voltage sensing is accomplished by the S4 helices. These helices contain positively-charged Lys or Arg residues at every third position and translocate in response to changes in voltage across the membrane.

It is not known precisely how the translocation of the S4 helices (4 per channel) cause activation, although the movement corresponding to the
opening of the activation gate likely corresponds to a hinge-like motion of the S6 segments around a conserved glycine.

Question 4

What determines the selectivity of KV and NaV ion channels?

Answer 4

Selectivity occurs within a central, ion conducting pathway formed by the four KV subunits or four repeats of NaV, where this central pathway is surrounded by S5 and S6 helices and connecting P loop contributed by the each of the four subunits or repeats (see above for the structures of NaV and KV).

Question 5

Describe what structural features of NaV and KV lead to “sidedness” of agents that act on these channels and to “state-dependence” of action. (location of selectivity filter, vestibule, activation/inactivation gates)

Answer 5

The location of the selectivity filter near the extracellular side, and of the vestibule nearer the intracellular side of the KV/NaV channels, together with the location of the activation/inactivation gates near the intracellular side, has the result that many channel modifying reagents have access to their sites of action only from one side of the membrane, and that this access may require that the channel be open.

The amino and carboxy terminals are intracellular.

[For example, tetrodoxin (TTX) is a charged molecule that cannot cross the membrane. When it is added to the extracellular side, it binds within the entrance of the pore, just above the selectivity filter of NaV. The binding of TTX is essentially independent of the position of the activation/inactivation gates; TTX has no effect when added intracellularly.]

Question 6

What structural subunit serves as the inactivation gate for NaV?

Answer 6

The inactivation gate of NaV channels is formed by the cytoplasmic loop (P loop) which connects repeats III and IV. When this cytoplasmic III-IV linker folds over the inner end of the central ion-conducting pathway, the channels is in a closed/inactivated state.

Question 7

For both KV and NaV, gating is controlled by _____.

Answer 7

membrane potential (Vm)

Question 8

What is an ionotropic receptor?

Answer 8

Neurotransmitter receptors that activate second messenger pathways which can affect physically separate ion channels.

Question 9

Within the ionotropic category are the pentameric ligand gated channels, also called the Cys-loop family of neurotransmitter receptor channels, including the GABAARs, GlyRs, nAChRs and 5-HT3Rs (all of which are named after the ligand most important for controlling their gating). These channels are ______ (structure). Each subunit has four transmembrane alpha helices (M1 through M4), with M2 assembling around the central, ion-conducting pathway. These channels are either selective for the permeation of ____, or allow permeation of both ____ and ______.

Answer 9

heteropentamers; chloride; sodium and potassium (with a slight preference for sodium)

Question 10

Ionotropic glutamate receptors are _____ (structure, what type of gate). In the case of the kind of glutamate receptors called NMDA receptors, two of the four subunits bind glutamate and the other two bind glycine.

Answer 10

tetrameric ligand gated channels

Question 11

Chloride channels of the CLC family are dimers in which each subunit has an ion permeation pathway. Each permeation pathway can gate open and closed independently of the other, although there is also another gate which controls both pathways simultaneously. Some members of the CLC family are H+-Cl- exchangers. The CLC chloride channels are important for stabilizing the resting membrane potential. When these are dysfunctional what may result?

Answer 11

Myotonia is a symptom of a small handful of certain neuromuscular disorders characterized by delayed relaxation (prolonged contraction) of the skeletal muscles after voluntary contraction or electrical stimulation.

The fainting goats.

Question 12

______ are tetramers in which each of the subunits contains a permeation pathway for ______. Strictly speaking, these are “anti-ion” channels since they exclude all ions including protons. Aquaporins are expressed in cells/tissues where rapid movement of water is important, such as ________. In addition to the four water channels, the assemblage of the four subunits also produces a central pore which may allow ion permeation and be gated between open and closed.

Answer 12

Aquaporin water channels; water molecules; the kidney

Question 13

About _____ (ratio) ions permeating a KV channel is not K+

Answer 13

1 : 10,000

Question 14

CaV channels select for Ca2+ over Na+ by ______.

Answer 14

3000 : 1

Question 15

_____ (ratio) permeating NaV channels is not sodium.

Answer 15

1 ion in 12

Question 16

nicotinic AChRs show only a slight preference for Na+ over K+ _____.

Answer 16

1.3 fold

Question 17

What four features help determine selectivity of a channel?

Answer 17

Size
Charge
Dehydration
Multiple binding sites

[Size: ions larger than the narrowest part of the pore will be rejected. However, size is by no means the only determinant: the crystal ionic radius of sodium is smaller than that of potassium, yet KV channels are highly selective]

[Dehydration: Ions are energetically stabilized in solution by waters of hydration, which make the ions effectively larger in size. Furthermore, the waters of hydration essentially mask small differences in the size of the ions. Thus, ions must be substantially de-hydrated before they pass through the channel pore, To compensate for this dehydration, which is energetically unfavorable, the ion is stabilized within the pore by energetic interactions with the amino acids forming the pore (but not too much, or the ion would stay “stuck” in the pore). The energetic interactions of the ion with the pore can occur with amino acid side chains (positive/basic residues: lysine or arginine; negative/acidic residues glutamate or aspartate), with backbone carbonyls (negative) or alpha helix dipoles (N-terminal: positive; C-teminal: negative). These and other interactions depend on the sign, valence and size of the ion.]

[Multiple binding sites can increase selectivity: If an ion interacts with multiple sites while traversing the channel pore, even relatively slight differences in the strength of interaction between preferred and non-preferred ions at each site can result in a significant enhancement of overall selectivity for the preferred ion.]

Question 18

What is a metabotropic receptor?

Answer 18

Neurotransmitter receptors that are directly coupled to ion channels (i.e. the receptor and channel are part of the same protein)

Question 19

Regarding the KV gate: When the inside of the cell has a____ potential with respect to the outside, the gate is held in its____ position and the current is ___.

When the inside is made____, the gate rotates to its ___ position and K+ ions flow ____. This process is called____.

When the inside of the cell is made ____ the gate rotates to the _____ position and the current decays away. This process is the reverse of activation and is called ____.

Answer 19

negative; closed; zero

positive; open; out of the cell; activation

negative; closed; deactivation

Question 20

At negative potentials the NaV gate is ____. Making the inside of the cell ____ causes the NaV activation gate to swing open (“activate”) and sodium ions to flow ____ the cell.

The second gate, the inactivation gate, is ____ at the resting potential because the activation gate occludes access to a site within the inner end of the pore at which the inactivation gate can bind. However, after the activation gate opens, the inactivation gate ____, a process called “inactivation,” causing the current to ____.

Answer 20

closed; positive; into

open; closes; decay to zero during a maintained depolarization

[Note that inactivation is not the same process as deactivation. Also, the reversal of inactivation is called “removal of inactivation.]

Question 21

How does lidocaine work?

Answer 21

Blocks NaV from the intracellular side only. Lidocaine equilibrates between de-protonated (neutral) and protonated (positively charged) forms. The protonated form is dominant at physiological pH and cannot cross the membrane, whereas the de-protonated form can. Protonated lidocaine has no effect on NaV from the extracellular side but can block the channel from the intracellular side (thereby producing local anesthesia).