Potassium channels

Question 1

members of inward rectifier K channel family

Answer 1

Kir (provides IK1) KACh KATP

Question 2

describe the structure of channels for Ik1

Answer 2

Whereas Kv channels form as an assembly of four alpha-subunit proteins each consisting of six membrane-spanning alpha-helical segments, S1-S6, ►Kir channels are built from four subunits each composed of just two membrane spanning alpha heliced (M1 and M2) with a type of P loop (called the H5 region) joining the two together. The channels are generally homo-tetrameric but some have hetero-tetrameric composition. ►The voltage sensor or S4 segment is not present and thus Kir channels are not sensitive to voltage. ►Their opening and closing is directed by polyvalent cation blocking and unblocking, which are, however, voltage-dependent processes. ►There are seven subfamilies of Kir channels, denoted Kir1- Kir7, and each subfamily has a variable number of isoforms and degrees of rectification. Channels through which cardiac Ik1 flow are thought to be formed from Kir2.1 and Kir2.2, coded by KCNJ2 and KCNJ12 respectively.

Question 3

structure of the inward rectifier channels (general)

Answer 3

two transmembrane segments (M1 and M2) no S4 regions, so no voltage sensitivity

Question 4

proteins that form channels for: Ik1 = IKACh =

Answer 4

Ik1 = Kir2.1 IKACh =Kir3.1 and 3.4

Question 5

what confers the K selectivity in the channel protein Kir,)

Answer 5

K selectivity of the central pore is conferred by a specific amino acid motif, glycine-tyrosine-glycine, in an intermembrane loop lining the pore (P or H5 loop)

Question 6

What is the typical Em and current flow relationship for Kir?

Answer 6

Inward current at Em more negative than Ek Outward current at Em more positive than Ek

Question 7

Location of Kir channels?

Answer 7

>80% of channels that are active at hyperpolarisation are in T-tubules Abundant in myocytes in ventricles, atria, purkinje but NOT present in the nodal cells Density is similar across the myocardial wall

Question 8

Function of Kir channels:

Answer 8

1) stabilise the resting membrane potential 2) determine excitation threshold 3) modulate the repolarisation process

Question 9

functional properties of the channels, how they work:

Answer 9

1) they open with steep voltage dependence on HYPERPOLARISATION AND prevent excessive loss of K from the cell during the plateau phase of AP 2) the voltage at which they open depends on the EXTRACELLULAR [K] but not the intracellular 2)part of steep rectification seems instantaneous, occuring in <1ms *** their usual function is to produce OUTWARD current because Em rarely falls below Ek. have no intrinsic voltage sensitivity but DEPOLARISATION REDUCES Kir conductance because INTRACELLULAR Mg and polyamines inpede the inner mouth of the channel when the curent is flowing outwards (voltage-dependent channel plugging by charged particles)

Question 10

what is inward rectification>

Answer 10

the conductance of Kir channels increases with membrane hyperpolarisation but decreases with depolarisation to potentials above the potassium equilibrium potential Ek, i.e. acts like a valve favouring entry of K during hyperpolarisation

Question 11

what is the current voltage relationship for Ik1 and what does this mean?

Answer 11

The current-voltage relationship is relatively steep indicating a low membrane resistance. The low membrane resistance means that membrane potential is well clamped and stable until a large depolarisation event.

Question 12

Describe the activation of Ik1 and further in relation to Em changes during the AP:

Answer 12

►Following phase 0, the inward rectifying channels close immediately and remain closed for much of the plateau. The almost zero Ik1 at early plateau potentials prevents rapid termination of the AP and loss of K from the cells. ►►As repolarisation proceeds, the inward rectifying channels tend to open again at potentials negative to -20mV and contribute to terminating the plateau and producing phase 3 of repolarisation. Ik1 stabilises the resting Em and determines the resistance of the membrane at rest.

Question 13

THE EFFECTS OF EXTRACELLULAR [K]

Answer 13

Extracellular [K] can modulate the Ik1. In addition, [K] can change locally in the heart. ►This is because K ions accumulate in T-tubules. The T-tubule network has a mean diameter of about 200nm which restricts ion diffusion. The diffusion rate for K in the T-tubules is about nine times slower than in free solution so, following increases in heart rate or a period of ischaemia, K ions accumulate in the confined spaces  producing cell depolarisation because Ek becomes more positive. ►The depolarisation causes inactivation of a portion of Na channels so fewer of them are available for activation and thus less current is generated to form the upstroke of the AP. ►The result is that phase 0 is smaller in size and the voltage change less rapid.

Question 14

what are the three major delayed rectifier currents?

Answer 14

IKr IKs IKur named after their variable raters of ACTIVATION

Question 15

describe the actions of Ikr what blocks it? why it appears to be inwardly rectifying?

Answer 15

blocked by methanesulfonanilides. ►IKr is halfactivated at -30 mV. Voltage-dependent inactivation reduces outward IKr during the plateau of the AP, but channels rapidly recover from inactivation and current magnitude rebounds during phase 3 repolarization. ►►Slow deactivation of IKr after repolarization of atrial nodal myocytes also contributes to the slow diastolic depolarization of these pacemaker cells. The amplitude of activating IKr decreases markedly as depolarization potential is more positive, i.e., the current displays apparent striking inward rectification. Increasing extracellular [K] increases the amplitude of outward current (contrary to what is expected by simple Nernstian considerations). Finally, close inspection of deactivating tail currents reveals a rapid hook of outward current preceding relatively slow deactivation. These observations are all consistent with a model in which transitions between closed and open states are slow and those between open and inactivated states are much more rapid. Thus, with maintained depolarization, the channel distributes into open and inactivated states, and at more positive potentials, distribution into the inactivated state is favored, thereby generating the appearance of inward rectification

Question 16

IKr: with maintained depolarization, the channel distributes into open and inactivated states, and at more positive potentials, distribution into the inactivated state is favored, thereby generating the appearance of inward rectification. WHEN IS THIS MOST EVIDENT?

Answer 16

with maintained depolarization, the channel distributes into open and inactivated states, and at more positive potentials, distribution into the inactivated state is favored, thereby generating the appearance of inward rectification This is most readily appreciated if, during maintained depolarization, a cell is briefly hyperpolarized. This allows inactivated channels to move to the open state (a rapid process), but if the hyperpolarization is sufficiently brief, open channels do not move to the closed state. Therefore, on return to depolarized potentials, channels are distributed primarily into open states, and rapid inactivation can be readily appreciated. When this experiment is conducted over a range of potentials, IKr current-voltage relations are linear, indicating that inward rectification is, indeed, attributable to voltage-dependent fast inactivation. The unusual [K]o dependence of activating IKr is also corrected when inactivation is transiently removed by a brief hyperpolarization, indicating that transitions to the inactivated state are [KC] dependent.

Question 17

what else is unusual about the IKr?

Answer 17

A second mechanism for the unusual [KC]o dependence of IKr involves [NaC]o-[KC]o interactions in the pore of the channel. When [KC]o is removed, [NaC]o can be shown to be a potent blocker of outward IKr, with a 50% inhibitory concentration of 3.1 mM, and [KC]o competes with [NaC]o to relieve this block. Inactivation is removed by mutations within the P-loops, supporting a pore-mediated (non–N-type) inactivation mechanism.

Question 18

Describe the structure and function of IKs

Answer 18

Iks is also a delayed rectifier current and gradually increases as cells depolarise. The channel is Kv7.1 (aka KVLQT1) and the gene that codes this protein is KCNQ1 ►voltage gated. ►It shows very slow activation on depolarisation but plays a role in determining the later plateau amplitude, the initial phase of faster repolarisation at the end of phase 2. ►The time constant for activation of Iks is 250-350ms, and the conductance is small. They vary in density across the ventricular wall – less in the midmyocardium. Also, they are more densely expressed in the base than the apex of the heart, so  the longest APD is measured in the apical M cells. The channel is composed of the standard K channel tetramer with the voltage sensor located in S4.

Question 19

where is the longest APD in the heart and why?

Answer 19

Iks channels are more densely expressed in the base than the apex of the heart, so  the longest APD is measured in the apical M cells.

Question 20

Ikr channels are activate at what voltages? what is their function and behaviour like?

Answer 20

channels that are activated at Em more positive than -40mV, ie during the plateau phase. The V1/2 (voltage required for half-activation) is -30mV. ►The behaviour is not typical of delayed rectifier because at voltages positive to about 0mV, the channels very rapidly inactivate reducing outward current at this and more positive potentials. ►In terms of current-voltage relationship, the channel behaves like an inward rectifier shutting off at more positive voltages. The reason for this apparent behaviour is that channel inactivation develops faster than channel activation at positive potentials and so limits the amount of time that these channels spend in an open state. ►As the AP moves into late phase 2 and early phase 3, the Em becomes more negative than 0mV, the channels recover from inactivation and open again. This leads to further outward current that helps with early phase 3 repolarisation. The current then decreases near the end of action potential because of channel deactivation. As a result, the current-voltage relationship is bell-shaped, much like that of L-type ca channel.

Question 21

structure of Ikr channels

Answer 21

Ikr flows through the Kv11.1 (hERG) potassium ion channel codded by the gene KCNH2. ►The channel has the typical Kv family structure of a tetrameric complex of four alpha-subunits. If the function of this channel is inhibited or compromised, either by drugs or genetic mutation, the AP duration prolongs. In the whole heart this manifests as increased QT interval. Over 450 mutation in the coding gene KCNH2 have been described and these translate into a variety of functional abnormalities of the channel protein. These all cause loss of function which leads to abnormal repolarisation of the cardiomyocytes and differences in their refractory periods. ►►Epicardial myocytes can express different amounts of Ikr compared with endocardial myocytes. Therefore, patients with these mutations will exhibit a large range of AP durations amongst their myocytes with heterogeneous repolarisation times and refractory periods. ►If there are regions of the heart with longer refractory periods compared to other closely neighbouring areas, this presents opportunities for re-excitation of these regions before the normal excitatory event occurs  re-entrant arrhythmias. The longer APDs can also predispose to after-depolarisations, which are pro-arrhythmic. ►►Kv11.1 is very susceptible to blockade from a variety of drugs.

Question 22

IKur channels: function, structure, therapeutic options

Answer 22

.► Eg, IKur seems to be confined to atrial cells. Ikur is an outwardly rectifying current that rapidly activates when Em is in the range from 0 to +60mV within about 10ms of the voltage change and thereafter partially inactivates over about 250ms. ►The channel of Ikur is Kv1.5, encoded by KCNA5. ►The Kv1.5 alpha subunit contains the typical voltage-sensitive K channel template of a tetramer of six trans-membrane spanning segments with a P loop between S5 and S6. ►Kvbeta subunits bind to control channel trafficking and appear to control the activation and inactivation by moving the V1/2 to more negative potentials. ♦♦Kv1.5 channels could be promising drug targets for the treatment of atrial fibrillation.

Question 23

what is the primary repolarising current?

what highlights this role?

Answer 23

Ikr

highlited by the severity of long QT syndrome mutations that affect it (LQT2) compared to those that affect Iks (LQT1)

Question 24

what is the structure of IKr

Answer 24

they are formed of four alpha subunits, encoded by HERG, each containing six transmembrane domains (S1-S6)

Question 25

structurally, how is the activation of Ikr achieved?

Answer 25

as a result of S$ movement in response to a change in Vm

Question 26

structurally, how is Ikr inactivation achieved?

Answer 26

voltage-dependent inactivation is caused by conformational changes in the outer mouth of the channel that mechanistically resemble C-type inactivation in Shaker channels.

Inactivaiton occures preferentially from the open state.

Question 27

during most of the AP, there is a balance between Ikr activation and inactivation. which one is favoured and what is the consequence?

Answer 27

Inactivation is favoured >>> this prevents the development of a large current.

as Vm repolarises and reaches levels where significant channel recovery from inactivation occurs, open-state occupancy rises to maximum and Ikr intensifies, reaching a peak late during the AP.

Question 28

comparing Ikr and Iks, which one is the more important in the rate-dependent adaptation of ADP? Why?

Answer 28

because voltage dependence recovery is much stronger than time dpependence during the AP, peak Ikr is similar at slow and fast rates. thus Ikr is secondary to Iks in underlying rate-dependent adaptation of APD (shortening of APD at fast rate)

There is no change in Ikr at fast rate compared to slow rate.

BUT: Iks increases significantly at fast rate due to accumulation of channels in closed states near the open states, from which they can open rapidly. The population of Iks channels in these states is likely to be increased in the presence of beta-adrenergic tone.

Question 29

how does increased Beta-AR stimulation effect the IKs participation in AP repolarisation?

what’s the evidence?

Answer 29

beta-AR stimulation increases Iks participation in AP repolarisation,

evidenced by the predisposition of patients with LQT1 and LQT5, which affect Iks, to suffer arrhythmia while exercsing or undergoing emmotional stress.

Question 30

what are the channels carrying Iks current formed of?

Answer 30

formed from two subunits, both can carry LQT mutations:

alpha subunit KCNQ1 (LQT1) and

beta subunit KCNE1 (LQT5)

the channels can be formed by a tetramer of KCNQ1 subunits, each with six transmembrane segments (S1-S6). the channel includes a variable number of subunits of KCNE1, which is a short protein that contains a sigle transmembrane spanning alpha-helix

Question 31

what confers the adrenergic control of the Iks channels?

Answer 31

beta-AR control of Iks is conferred by phosphorylation of the KCNQ1 N terminus and requires bindin of several protein components in addition to KCNE1 for transduction.