4 Structural and Molecular Bases of

­Cardiac Inward Rectifier Potassium

Channel Function
Anatoli N. Lopatin and Justus M.B. Anumonwo

CHAPTER OUTLINE resulting channels depends partially or solely on the interactions

with additional (accessory) proteins.2 From a functional per-
Background 38
spective, there are three distinct classes of cardiac Kir channels
A Family of Genes Encodes Inward Rectifier Potassium (IK1, IKACh, and IKATP). As will be discussed later in this chap-
Channels 38 ter, based on the degree of inward rectification, the underlying
channels can be considered as strong (IK1; KACh) or weak (KATP)
Classical Cardiac Inward Rectifier Potassium Channels 38 inward rectifiers.2
Acetylcholine-Activated Potassium Channels 43 There is ample molecular and electrophysiological evidence
for the expression of Kir2, Kir3, and Kir6 subfamily members in
ATP-Sensitive Potassium Channels 45 myocardial tissue, subunits representing the molecular correlates
Conclusion 47 of IK1, IKACh, and IKATP, respectively. Given that the other mem-
bers of Kir family (Kir1, Kir4, Kir5, and Kir7) are thought to be
primarily important for K+ transport in other tissues, they will
not be discussed further in this chapter. 
Background
Inwardly rectifying potassium (Kir) channels are important Classical Cardiac Inward Rectifier
for stabilizing the resting membrane potential, establishing Potassium Channels
the threshold of excitation, and modulating the repolarization
phase of the cardiac action potential.1 Inward rectification is a
Structure and Function
process in which the conductance of the Kir channel increases Kir2 Subfamily Underlies Cardiac IK1
with membrane hyperpolarization, but decreases with depolar- The Kir2 subfamily consists of six members (Kir2.1-Kir2.6),
ization to potentials positive to the K+ equilibrium potential. In of which only Kir2.1-Kir2.4 are expressed in the mammalian
essence, Kir channels behave as “biodiodes,” preferentially pass- heart. As shown in Table 4.1, the following genes encode the
ing current in one direction. The molecular basis of rectification mammalian Kir channels: KCNJ2, KCNJ12, KCNJ4, KCNJ14,
in Kir channels is a physical occlusion of the ion permeation KCNJ17(KCNJN1), and KCNJ18 for Kir2.1 through Kir2.6,
pathway by depolarization-induced movement of intracellu- respectively. There is evidence that in the mammalian heart
lar cations, such as magnesium and polyamines.1 Only few Kir Kir2.1-Kir2.3 isoforms are expressed in cardiac myocytes and
channels, however, display strong rectification.2 Strong inward that Kir2.4 is probably only expressed in neuronal cells. It is now
rectification not only enables the Kir channels to stabilize the well established that members of the Kir2 subfamily underlie IK1,
resting membrane potential but also allows for the protection although the subunit composition varies among species and cell
of the cell against an excessive loss of K+ ions during the plateau types and that channel complexes are likely formed as heterotet-
phase of an action potential.1 The molecular correlates of these rameric structures. 
channels are primary subunits encoded by the KCNJ family of
genes.3 Mutations in KCNJ genes have been associated with Crystal Structure of Kir2 Channels
various channelopathies,2 demonstrating the importance of Kir In recent years, x-ray crystallographic structures of both bacte-
channels in normal cardiac excitation. This chapter will focus rial and mammalian homologs of several Kir channels have been
on three well-studied Kir channels in the myocardial cells: the obtained.7 Fig. 4.2 highlights some important and common fea-
classical inward rectifier K+ channels (IK1), the acetylcholine- tures of Kir channel structure based on the results of work with
activated K+ channels (KACh), and the adenosine triphosphate Kir2.1 and Kir2.2 mammalian channels.8 It is now firmly estab-
(ATP)-sensitive K+ channels (KATP). Additional information on lished that Kir channels are tetramers of distinct subunits, each
the topics covered in this chapter can be obtained from recent having two transmembrane domains (M1 and M2), relatively
review articles.2,4,5  small N-terminal, and large C-terminal cytoplasmic domains,
and a pore-forming structure between M1 and M2 (see Fig. 4.2).
A Family of Genes Encodes Inward The pore structure contains a pore helix directed toward the con-
duction pathway and the characteristic GYG (or GFG) motif,
Rectifier Potassium Channels also known as the K+ channel signature sequence, which contrib-
Channels belonging to the Kir family are structurally and func- utes to the selectivity filter in all potassium channels. The M1 and
tionally different from voltage-gated K+ channels.2,4 The genes M2 transmembrane domains in each subunit are arranged as an
that encode Kir channels are ascribed the KCNJ nomenclature antiparallel coiled-coil and make contact with each other. Kir2
and are categorized into seven subfamilies based on the gene channels have a negatively charged amino acid (D172 in Kir2.1)
products (Kir1-7; Table 4.1; Fig. 4.1). In general, Kir channels located in approximately the middle of the pore, which plays a
consist of homomeric or heteromeric complexes of the respec- critical role in the phenomenon of inward rectification discussed
tive Kir subunits, but as will be discussed, functionality of the later.

38

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 STRUCTURAL AND MOLECULAR BASES OF CARDIAC INWARD RECTIFIER POTASSIUM CHANNEL FUNCTION 39

TABLE 4.1  Diversity of α-subunit proteins in the family of inward rectifier potassium channels

SUBFAMILY
Kir1
PROTEIN
Kir1.1
GENE
KCNJ1
HUMAN
11q24
MOUSE
9 A4
CURRENT
?
4
Kir2 Kir2.1 KCNJ2 17q24.3 11 E2 IK1
Kir2.2 KCNJ12 17p11.2 11 B2 IK1
Kir2.3 KCNJ4 22q13.1 15 E1 IK1
Kir2.4 KCNJ14 19q13.4 7 B4 ?
Kir2.5 KCNJ17* 17p11.1
Kir2.6 KCNJ18 17p11.2
Kir3 Kir3.1 KCNJ3 2q24.1 2 C1.1 IKACh
Kir3.2 KCNJ6 21q22.1 16 C4
Kir3.3 KCNJ9 1q23.2 1 H3
Kir3.4 KCNJ5 11q24 9 A4 IKACh
Kir4 Kir4.1 KCNJ10 1q23.2 1 H3
Kir4.2 KCNJ15 21q22.2 16 C4
Kir5 Kir5.1 KCNJ16 17q24.3 11 E2
Kir6 Kir6.1 KCNJ8 12p11.23 6 G2 IKATP
Kir6.2 KCNJ11 11p15.1 7 B4 IKATP
Kir7 Kir7.1 KCNJ13 2q37 1D ?

*Also known as KCNJN1.6

Modified and updated from Nerbonne JM, Nichols CG, Schwarz TL, et al. Genetic manipulation of cardiac K+ channel function in mice: what have we learned, and
where do we go from here? Circ Res. 2001;89:944-956.

Kir1 Kir2 Kir3 Kir4 Kir5 Kir6 Kir7

SU
4.1 5.1 R
R
SU

5.1 4.1
SU
R
R
SU

SUR1 SUR2 Kir6.1 Kir6.2 KATP

Kir3.1 Kir3.2 Kir3.3 Kir3.4 KACh

Kir2.1 Kir2.2 Kir2.3 Kir2.4 Kir2.5 Kir2.6 IK1

FIGURE 4.1  The family of inward rectifier potassium channels.  All members of the Kir family share
significant structural similarity, but only the Kir2 and Kir3 subfamilies represent channels carrying
classical strongly rectifying currents. Six members of the Kir2 and four members of the Kir3 subfami-
lies were cloned in mammals. Heteromeric assemblies of the Kir2.1, Kir2.2, and Kir2.3 subunits un-
derlie the IK1 current, and heteromeric assembly of the Kir3.1 and Kir3.4 subunits underlies the IKACh
current. Weakly rectifying KATP channels are composed of pore-forming Kir6.1 and Kir6.2 subunits
as well as auxiliary SUR1 and SUR2 subunits. (Modified from Anumonwo JM, Lopatin AN. Cardiac
strong inward rectifier potassium channels. J Mol Cell Cardiol. 2010;48:45-54.)

There are at least three distinct regions of the intramembrane nature. The important functional role of the slide helix is high-
part of the pore: the selectivity filter, a centrally located water lighted by the fact that many loss-of-function mutations associ-
filled cavity of approximately 10 Å in diameter, and the narrowing ated with Andersen-Tawil syndrome (LQT7) are located in the
of the pore at the cytoplasmic side as the pore-lining M2 helixes slide helix.
come closer to each other (known as bundle-crossing). M2 helixes A large C-terminal domain provided by four Kir2.x subunits
also possess a highly conserved glycine residue (a “hinge”; G168 consists primarily of β-strands and potentially strongly interacts
in Kir2.1) that likely contributes to the channel gating by allow- with a smaller N-terminal domain. As shown in Fig. 4.2, C-ter-
ing the helixes to bend and change the size of the pore at the minal domain forms a large intracellular vestibule, approximately
bundle crossing. 30 Å in length, for easy ion passage and likely provides binding
The so-called slide helix formed by the N-terminus region sites for various intracellular agents. The cytoplasmic domain
just preceding the M1 helix is another unique and important harbors a number of residues (e.g., E224 and E299 in Fig. 4.2)
regulatory feature of both bacterial and mammalian Kir chan- known to contribute to inward rectification.
nels.9 This helix intercalates between the inner leaflet of the A membrane phospholipid phosphatidylinositol 4,5-bisphos-
plasma membrane and cytosol likely because of its amphiphilic phate (PIP2) is an important structural and regulatory component

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40 CARDIAC ELECTROPHYSIOLOGY

Pore helix
M2 helix
M2
Selectivity Inner helix G168
filter
M1 M1 helix
Outer helix
RWR
Water cavity D172
PIP2
Bundle Slide helix
crossing G-loop

N N E299
E224

Spm
Cytoplasmic pore
C C
C C

FIGURE 4.2  Architecture of a typical Kir channel.  X-ray structure of chicken Kir2.2 crystallized in
the presence of PIP2 (accession code 3SPI, available in the Protein Data Bank; from Hansen and col-
leagues8). Only two opposing subunits are shown for clarity. Right, The structure is rotated approxi-
mately 90 degrees to better present the intracellular ion conduction pathway and bound molecules of
PIP2. Numbering of the amino acids corresponds to that in Kir2.1. Red circles represent the K+ ions in
the selectivity filter. Blue circles show the location of three residues most critical for inward rectifi-
cation. Spermine molecule (Spm) is shown in the middle at the same scale as the Kir2.2 structure.
Analysis and visual presentation of the structures were performed using DeepView Swiss-PdbViewer
and PyMol software.

of Kir channels.10 In the membrane, PIP2 likely interacts with domain (D172; the rectification controller; see Fig. 4.2) and in
hydrophobic amino acids on both M1 and M2 transmembrane the C-terminal tail [E224 and E299; Kir2.1 amino acid (aa) num-
helices as well as with a well-conserved (among Kir channels) RWR bering; see Fig. 4.2]. Positively charged polyamines enter the Kir
motif located just at the end of the M1 domain (see Fig. 4.2). The channel pore to physically occlude it, which is a process aided
interaction of PIP2 with a specific region in the cytoplasmic domain by negative charges provided by aspartate and glutamate resi-
leads to an approximately 6 Å translation of the entire cytoplasmic dues. Both electrophysiological and structural data are consistent
domain toward the membrane associated with the movement of the with the idea that channel block occurs in two sequential steps. A
M1 helix, which ultimately leads to the opening of the gate.8  weakly voltage-dependent (shallow) blocking step involves entry
of polyamine into the Kir2.1 pore at a site provided by a ring
Mechanism of Polyamine-Induced Rectification of negative charges (E224 and E299) in the C-terminus. A more
The key experiments conducted in mid-1990s with the first strongly voltage-dependent blocking step reflects the movement
cloned members of the Kir2 subfamily have clearly demonstrated of polyamine to its deep binding site near the D172 residue. It
that almost all essential properties of classical strong inward rec- is also believed that the strong voltage dependence of the poly-
tification could be explained by a voltage-dependent block of the amine block arises not only from the high valency (z) of poly-
channel by the ubiquitous intracellular organic cations, the poly- amines (z≈4 for spermine) but also from a displacement (push)
amines.11 The micromolar concentrations of free polyamines of K+ ions through the pore.12 Among the polyamines, spermine
(primarily spermine and spermidine) are sufficient to reproduce is the most voltage-dependent blocker and also has the highest
the degree of rectification observed in native cells. The strength potency for blocking the channel.
of rectification varies among the different members of the Kir A characteristic feature of Kir2.1 and Kir3 channels is a flex-
subfamily, and although every Kir channel shows some degree of ible cytoplasmic pore-facing G-loop (see Fig. 4.2) that forms a
inward rectification, they can be broadly grouped as either strongly girdle around the central axis of the Kir channel.13 It was esti-
rectifying (Kir2 and Kir3) or weakly rectifying (Kir1 and Kir6) mated that this girdle constricts the ion permeation pathway to
channels. It is well established that the time-dependent activation approximately 3 Å. Mutations in the G-loop were shown to dis-
of strong, inward rectifiers during membrane hyperpolarization rupt inward rectification. In addition to the previously described
reflects the exit of polyamines (primarily sperimine) from the pore.  E224 and E299 residues, it was shown that A255 and A259
located farther away from the pore axis are also involved in chan-
Rectification Properties Are Related to Electrostatic nel rectification. Electrophysiological experiments using cyste-
Interactions in the Cytoplasmic Pore of a Kir Channel ine modifications in the pore region of a mutant Kir6.2 channel
Early work with cloned Kir channels established that strong (N160D; equivalent to D172 in Kir2.1) showed that spermine
inward rectification by intracellular polyamines depends on three binds at a deep site beyond the rectification controller residue
negatively charged residues located in the second transmembrane D172, a site that is close to the extracellular mouth of the pore.

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 STRUCTURAL AND MOLECULAR BASES OF CARDIAC INWARD RECTIFIER POTASSIUM CHANNEL FUNCTION 41

In another study, refinement of the crystal structures of bacte- with specific antibodies. Intercalated discs were not studied
rial KirBac1.1 and KirBac3.1 allowed identification of the shallow electrophysiologically in regard to the presence of IK1 chan-
polyamine binding sites at the cytoplasmic interface between the
two subunits.14 These observations notwithstanding, the precise
nels, although labeling with various Kir2 antibodies can clearly
be observed at this location. Moreover, there is evidence that 4
mechanism involved in rectification is still being worked out, and in canine ventricular and atrial myocytes, Kir2.3 subunits are
the exact location of polyamine binding sites in Kir channels is expressed at higher levels in intercalated disc membranes rela-
still a controversial issue.  tive to T-tubules. 

Differential Properties of Kir2.x Subfamily Kir2.x Macromolecular Complexes and

Consistent with their overall significant sequence homology, all
Kir2 channels have the three mentioned residues (D172, E224,
Channel Biogenesis
and E299) important for rectification at the equivalent positions. Understanding the composition, as well as assembly mechanisms
Recent studies, however, unexpectedly showed that rectification of ion channel complexes in cellular microenvironments, such as
properties are rather different in the three Kir2 isoforms.15 Spe- the sarcolemma, intercalated discs, T-tubular membranes, and
cifically, the data show that Kir2.2 channels display significantly lipid rafts, has gained significant attention in recent years.4,18
stronger voltage dependence of rectification than that observed Compelling experimental evidence demonstrates that Kir2.x
in Kir2.1 and Kir2.3 channels. The voltage dependence of steady- protein-protein and protein-lipid interactions in these micro-
state rectification is different between the Kir2.x channels, as are environments have important implications for normal channel
the single-channel conductance and kinetics properties of the cur- functional properties.8,19–22 Standard biochemical techniques,
rents. In particular, a polyamine unblock (activation) at negative cell electrophysiology, and proteomic analyses have shown that
membrane potentials in the Kir2.3 channels is several-fold slower Kir2.x channels, in macromolecular complexes, interact with
than in Kir2.1 and Kir2.2 channels. Single-channel conductance the membrane-associated guanylate kinase (MAGUK) scaf-
is the smallest in Kir2.3 (approximately 10 pS), medium in Kir.2.1 folding protein family (i.e., SAP97, PSD95, and CASK, Mint
(approximately 25 pS), and the largest in Kir2.2 (approximately and Veli) and with members of the dystrophin-associated pro-
35 pS).  tein complex.18,23,24 Kir2.x proteins interact with the MAGUK
proteins through the type 1 PDZ binding motif at the channel
C-terminal residues. The PDZ binding motif (S/T-X-Φ; X: any
Cellular and Membrane Localization amino acid, Φ: a hydrophobic amino acid) is a set of the last three
IK1 currents, and thus the underlying Kir2.x subunits, display (with the exception of Kir2.4) residues on the channel peptide.
a distinct regional distribution in the heart. It has been shown In one heterologous expression study of Kir2.3 isoform, it was
that inward IK1 is generally more prominent in ventricular myo- shown that coexpression with SAP97 caused significant cellu-
cytes and Purkinje fibers and is significantly smaller in atrial lar redistribution of the Kir2.3 channel protein, with a modest
myocytes (with one known exception of the mouse heart). The increase in the cell surface expression of the channels.23 It was
density of inward IK1 currents (i.e., currents normalized to also shown that the coexpression led to a wide distribution of
membrane capacitance) is small in the pacemaker cells of the single channel conductances, with three distinct peaks centered
sinoatrial node in mice and rats and essentially undetectable in at 16, 29, and 42 pS. The coexpression, however, did not alter
the rabbit sinoatrial nodes. IK1 cannot be detected in atrioven- the channel open probability. In another study in rat ventricular
tricular node of the rabbit but relatively large IK1 currents can myocytes,24 silencing of SAP97 using adenoviral short hairpin
be recorded in the guinea-pig atrioventricular node. Moreover, RNA reduced IK1 whole-cell current density without affecting
the density of inward IK1 varies across the ventricular myocar- channel unitary conductance properties. In the study, silencing
dium. For example, in the mouse heart inward, IK1 is larger in of the MAGUK protein blunted the sensitivity of IK1 to iso-
the apical myocytes compared with the epicardial cells, and proterenol. Thus these results clearly demonstrate that Kir2.x
IK1 is larger in the right ventricular than in the left ventricular channel assembly in such macromolecular platforms is criti-
myocytes. cal for normal channel function. Therefore an understanding
Molecular biological studies are also consistent with the of the details of the architecture of such macromolecular scaf-
location-dependent expression of specific Kir2 isoforms. In par- folds/platforms, especially the essential molecular components,
ticular, real-time reverse-transcriptase polymerase chain reac- is important for a better insight into channel biophysics and
tion analysis of Kir2 transcripts in the human heart showed the regulatory properties. These studies are significant, given that
following relative expression levels: in Purkinje fibers, Kir2.1 abnormalities in proper assembly of the platforms may play a
> Kir2.3 > Kir2.2, and in the right ventricle, Kir2.1 > Kir2.2 > role in arrhythmogenesis.23–25 
Kir2.3; the sequence was reversed in the right atrium: Kir2.3
> Kir2.2 > Kir2.1.16 Information on the expression patterns of
the Kir2 subunits can also be gleaned from functional data using
Pharmacology and Regulation
the unique properties of corresponding channels. For example, Kir2.x and IK1 channels can be regulated in a number of ways.2
in cardiac atrial and ventricular myocytes, unitary conductance Most studies on adrenergic stimulation show that inward IK1 cur-
values display a wide spectrum ranging from 10 to 15 pS (as in rents are suppressed by the activation of both α- and β-receptors,
the Kir2.3 channels) to as high as 40 to 45 pS (as in the Kir2.2 although opposite effects have also been described. In addition,
channels). adrenergic regulation is clearly dependent on the type of recep-
Evidence shows that IK1 channels are located not only in the tors and subunit composition of the channel.
non–T-tubular component of the sarcolemma of ventricular Both isoproterenol and forskolin inhibit IK1 in human ven-
myocytes, but also in the intercalated discs and in the T-tubules. tricular myocytes, suggesting involvement of protein kinase A
For example, accumulation and depletion of K+ in T-tubules (PKA)-mediated phosphorylation of underlying Kir2.x subunits.
lead to changes in whole-cell IK1.17 IK1 accumulation/depletion Molecular details of the phenomenon, however, are contradictory.
phenomena are not observed in atrial cells, which essentially lack For example, it has been shown that the application of a catalytic
T-tubules, and in ventricular myocytes, in which T-tubules are subunit of cyclic adenosine monophosphate (cAMP)-dependent
removed by osmotic shock.17 PKA leads to activation of Kir2.1 channels expressed in Xenopus
Alternatively, Kir2.1, Kir2.2, and Kir2.3 subunits were oocytes, but to Kir2.1 inhibition when the channels are expressed
localized to the T-tubular membrane using immunolabeling in a mammalian cell line (COS-7). The data on PKA regulation of

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42 CARDIAC ELECTROPHYSIOLOGY

native IK1 channels is limited and somewhat controversial; how-

ever, most of the studies show that IK1 channels are inhibited by
Channelopathies
the exposure of the cytosolic side of the membrane to the puri- There are at least four known channelopathies associated with
fied catalytic subunit of PKA. Studies in exogenous-expressing IK1 channels, all originating from mutations in KCNJ2: ATS1,
systems also showed the involvement of protein kinase C in the short QT (SQT) syndrome, familial atrial fibrillation (FAF), and
negative regulation of Kir2.1 channels. Consistent with the latter, catecholaminergic polymorphic ventricular tachycardia (CPVT)
experiments using human atrial myocytes show that α1-adrenergic (Fig. 4.3).2
stimulation likely reduces IK1 via a protein kinase C–dependent ATS is characterized by a triad of pathological clinical pheno-
mechanism. Kir channels are also targets for phosphorylation by types including the morphogenesis and functioning of skeletal and
tyrosine kinases. In Kir2.1 channels, the site of downregulation cardiac muscle. One of the prominent features of ATS is cardiac
was targeted to a single Y242 residue in the C-terminus.26 electrical abnormalities, including brief episodes of ventricular
PIP2 is an important component in membrane-delimited sec- tachycardia, multifocal ventricular ectopy induced by adrenergic
ond messenger signaling system and a powerful activator of Kir stimulation, and prolongation of the QT interval. ATS1 is used
channels.27 There is significant evidence that PIP2 regulates the to differentiate ATS patients carrying mutations in KCNJ2. Many
channel gating primarily through specific electrostatic interac- ATS1 patients display clear QT prolongation and have been
tions with the cytosolic part of the channel (see Fig. 4.2).8 It is referred to, perhaps questionably in some cases, as LQT7.
also clear that various properties of Kir2 channels are modulated LQT7/ATS1 mutations are numerous (see Fig. 4.3)5 and result
by PIP2. For example, the pH sensitivity of Kir2.3 channels is in nonfunctional channels when exogenously expressed in a heter-
strongly dependent on the strength of the channel-PIP2 inter- ologous system.31,32 Because affected patients are heterozygous for
action. Cholesterol is another prominent lipid involved in the the mutant and wild type (WT) alleles, and the channel is com-
regulation of Kir2 channels. In particular, it has been shown that posed of four subunits, a dominant-negative effect of the mutated
increased levels of cholesterol in the membrane lead to the sup- subunit can lead to reduced IK1 current in cardiac myocytes (and
pression of Kir2 currents.28 other cells). The dominant-negative effect of LQT7/ATS1 muta-
Various common cations, such as Ca2+ and H+, contribute to tions has been demonstrated using cloned channels, although the
the regulation of Kir2 channels as well.2 Intracellular Ca2+ blocks magnitude of current inhibition is variable in different mutants.
IK1 channels in a voltage-dependent manner. There is evidence SQT syndrome is characterized by an abnormally short QT
that a transient increase in intracellular Ca2+ during an action interval (less than 300 ms) and an increased risk of developing
potential can lead to significant blockage of IK1.29 Regulation fibrillation and sudden death. Currently, three forms of SQT syn-
of IK1 by intracellular pH (pHi) is dependent on the species and drome have been described. SQT1 and SQT2 syndromes result
the type of tissue. For example, rat and guinea pig ventricular from mutations in genes underlying two voltage-gated K+ chan-
IK1 is not sensitive to physiologically relevant changes in pHi. In nels, hERG (KCNH2) and IKs (KCNQ1). A third variant of SQT
contrast, IK1 in sheep ventricular myocytes is inhibited by intra- syndrome (SQT3) originating from a mutation in KCNJ2 has
cellular H+ with a pKa of approximately 7.4. The difference in been first described by Priori and colleagues in 2005.33 Genetic
pH sensitivity is likely due to differences in the subunit composi- analysis revealed a charge-neutralizing substitution (D172N) in
tion of IK1. For example, channels composed of Kir2.3 subunits the critical place of the channel responsible for the strong inward
exhibit strong sensitivity to pHi within a physiologically relevant rectification (see Fig. 4.2). Accordingly, coexpression of WT
range, whereas Kir2.1 and Kir2.2 channels are relatively insensi- and D172N mutant subunits showed decreased rectification of
tive to H+. the heteromeric channel. Computer simulations showed that
Pharmacological tools for the modulation of Kir2.x and IK1 reduced rectification of IK1 can explain some of the characteristic
channels are limited. The most useful research tool for study- features of an electrocardiograph, such as tall and asymmetric T
ing Kir2 channels is Ba2+ [inhibitory concentration of 50% (IC50) waves, observed in affected patients. Recently, three other muta-
for inward currents, 0.5 to 10 μM]. Ba2+ action is subunit depen- tions in the KCNJ2 gene associated with SQT syndrome have
dent. For example, Ba2+ blocks Kir2.2 channels fivefold to seven- been identified: M301K34 and E299V35 (see Fig. 4.2), both of
fold more efficiently than Kir2.1 channels. In the early 1990s, a which are located in the region of the channel responsible for the
compound RP 58866 and its active enantiomer, terikalant, were “shallow” polyamine binding site, and K346T36 which is located
shown to be selective blockers of IK1 (IC50, 5 to 20 μM). However, on the outer side of C-terminal domain.
later studies revealed that terikalant also inhibits many other K+ One study described the association of a single V93I mutation
channels, some with even higher potency (IC50 in submicromolar in KCNJ2 with familial atrial fibrillation.37 Electrophysiologi-
range for IKr channels). Similarly, it was found that a LY97119 cal experiments with cloned channels showed, in particular, that
compound (LY, a tertiary homolog of clofilium) blocks IK1 in the V93I mutation leads to a relative increase in the magnitude of the
low micromolar range, but it also blocked Ito (transient outward outward current, or a decrease in the strength of inward rectifica-
current) at submicromolar concentrations. Perhaps chloroquine tion. Regarding the inward rectification, the effect of the V91I
(an antimalarial drug) is the most potent blocker of IK1 with an mutation on Kir2.1 current resembles the mutation found in the
IC50 of approximately 0.5 μM. However, chloroquine does not D172N mutant channels. However, patients carrying the V93I
discriminate among IK1, IKACh, and IKATP, and it has been shown mutation display a normal QT interval in contrast to patients
to affect other currents (e.g., INa) in the low micromolar range. affected by the D172N mutation.
Activators of Kir2 and IK1 channels were also described. In par- Mutations in KCNJ2 were also linked to another type of excit-
ticular, flecainide, a widely used antiarrhythmic drug, increases ability disorder—catecholaminergic polymorphic ventricular
Kir2.1 currents by approximately 50% at a concentration of 1 tachycardia (CPVT).38 CPVT is a heritable disorder character-
μM, but has no effect on current carried through the Kir2.2 and ized by frequent ventricular arrhythmias and sudden cardiac death
Kir2.3 channels.30 Arachidonic acid and the antiinflammatory associated with physical activity or adrenergic stimulation. Nine
agent tenidap were shown to specifically activate Kir2.3 channels, KCNJ2 mutations associated with CPVT have been identified so
with a greater than twofold maximum increase in inward cur- far (see Fig. 4.3). These mutations can be found in both N- and
rent with an IC50 of approximately 0.5 and 1.3 μM, respectively. C-termini as well as in the extracellular loop of the Kir2.1 chan-
Zacopride, a gastrointestinal prokinetic drug, was recently found nel. The electrical phenotype of these mutations included, in par-
to be a selective IK1 channel agonist. The activating effect, how- ticular, prominent U waves, ventricular ectopy, and polymorphic
ever, is modest, with a maximum increase in IK1 of approximately ventricular tachycardia. It can be seen from Fig. 4.3 that there is a
34% at a concentration of 1 μM.  significant phenotypic overlap between ATS and CPVT.

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 STRUCTURAL AND MOLECULAR BASES OF CARDIAC INWARD RECTIFIER POTASSIUM CHANNEL FUNCTION 43

LQT7/ATS1 E138K
S136F
4
SQT3 CPVT T142P

CPVT/ATS1 T130M G144D G144A/D/S

V123G T145C
FAF Other
G146A/D/R/S

C154F/Y

C101R
∆ 95–98 ∆ 163–164
L94P M1 M2
∆ 91–94 D172N
V931

P186L
T305A/I/P/S
R82W R82Q/W K187R E303K T305I/S
R80C R189I V302del

D78G/Y N190I V302M M307I

T75A/M/R T192A/I ∆ 299–301 T309I

T74A L193P M301K R312C/H

D71N/V/Y G215D/R G300A/D/V ∆ 314–315

Y68D N216H E299V N318S

R67W R67Q/W L217P L298R W322C

C54F R218P/Q/W R260P K346T

G52V S220I S220I R260P S369X

R40X V227F T400M

R228X/insKSHLVEAHVR P351S
N C
FIGURE 4.3  Channelopathies of the classical inward rectifier channel, IK1, associated with muta-
tions in the Kir2.1 subunit.  Mutant residues are color coded to represent the long QT (LQT7/ATS1;
black), catecholaminergic polymorphic ventricular tachycardia (CPVT; red), familial atrial fibrillation
(FAF; green), short QT3 (SQT3; blue), and Other are mutations with less defined cardiovascular as-
sociations. (Modified from Anumonwo JM, Lopatin AN. Cardiac strong inward rectifier potassium
channels. J Mol Cell Cardiol. 2010;48:45-54.)

Finally, some mutations in KCNJ2 could not be easily classi- be a muscarinic (M2) or a purinergic (P1) receptor, which are
fied into the four categories mentioned previously, and for this activated by acetylcholine or adenosine, respectively. The ulti-
reason they are referred to as other in Fig. 4.3.  mate result of channel activation is the opening of the Kir3.1/
Kir3.4 channels, which permits K+ efflux and consequently
hyperpolarizes the cell membrane.
Acetylcholine-Activated Potassium As in a typical K+ channel, the ion selectivity in Kir3.x chan-
Channels nels is conferred by the presence of the signature sequence
(T-X-G-Y/F-G),4,40 and the mechanism of rectification involves
Structure and Function an asparagine or an aspartate residue for interactions with the
In the heart, Kir3.1/Kir3.4 channels are responsible for the effects polyamines (details of rectification have been discussed previ-
of acetylcholine and adenosine, and they act through a coupling ously). Thus the mechanism of rectification is similar to that
mechanism involving a receptor, a G protein (Go/Gi family), and described for Kir2 channels, and the Kir3.x channels belong to
the K+ channel.2,39 Because channel gating requires a G protein, the class of strong inward rectifier channels.2 Membrane topol-
Kir3.1/Kir3.4 channels are considered a type of KG channel.4 For ogy of Kir3 channels is similar to that described for Kir2 chan-
channel activation, the G-protein–coupled receptor (Fig. 4.4) can nels (see Fig. 4.2).

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44 CARDIAC ELECTROPHYSIOLOGY

GPCR K+
Agonist
(M2 or P1)
(ACh/Ado)

D E E
J J

GiDEJ GDP
GTP
A
Cell-attached Inside-out
ACh
or
Ado
Cell
GTP 100µM
3 min
ACh 1.1µM
–60 mV

GTP 100µM
2.5 min
Ado 10µM
–80 mV

B
FIGURE 4.4  Activation of the Kir3.1/Kir3.4 channels by muscarinic (M2) and purinergic (P1)
G-protein coupled receptors (GPCRs).  (A) The membrane-delimited pathway for activation of
acetylcholine (Ach) and adenosine (Ado) gated Kir channels. Ligand-GPCR interaction enhances the
guanosine triphosphate (GTP) association with a G-protein α-subunit, and results in the release of the
β/γ-subunits for the activation of the Kir channel. Potassium efflux hyperpolarizes the cell membrane.
(B) Requirement of GTP for Kir3.1/Kir3.4 activations by ACh (top single channel traces) and Ado
(bottom single channel traces). Holding potentials for experiments are annotated. Channel activity
is present in the cell-attached patch mode, with ACh or Ado present in the pipette (top inset). Patch
excision (at arrow; inside-out patch configuration) loss of GTP results in the loss of channel activity.
Subsequent application of GTP (100 μM; intracellular side) restored channel activity. ([A] Modified
from Breitwieser GE. GIRK channels: hierarchy of control. Am J Physiol Cell Physiol. 2005;289:C509-
C511. [B] Modified from Hibino H, Inanobe A, Furutani K, et al. Inwardly rectifying potassium chan-
nels: their structure, function, and physiological roles. Physiol Rev. 2010;90:291-366.)

Exactly how do Kir3.1/Kir3.4 channels and G proteins N-terminus is positioned between the two C-terminal domains.41
interact to cause channel opening? First, a brief discussion of The study also showed that for Kir3.1 channels, the cytoplasmic
G proteins is necessary. G proteins are complexes consisting of residues L262, L333, and E336 are important in gating, and the
an α- [molecular weight (MW), ∼40,000], a β- (MW, ∼35,000), equivalent residues are H64 and L262 in Kir3.4 channels. 
and a γ- (MW, ∼8000) subunit, which all transduce signals from
membrane receptors [e.g., muscarinic (M2)] to an effector, such
as a KG channel.4,39 Four subfamily members of the guanosine
Cellular and Membrane Localization
diphosphate (GDP) Gα subunit dictate selectivity of signaling There are tissue-dependent differences in the expression of IKACh
[activation of adenylate cyclase (Gαs), inhibition of adenylate in the myocardium, with a very high density of expression in
cyclase (Gαi), and activation of phospholipase (Gq)].39 Normally, nodal tissues.2,4 Following the cloning of genes underlying Kir3
Gα is bound to GDP in the absence of an agonist, and the Gα/ channels in the early to middle 1990s, investigators have probed
GDP complex is coupled to the receptor and has low guanosine myocardial tissues to examine the localization of the gene prod-
triphosphatase (GTPase) activity. With vagal stimulation and ucts. Overall, these studies reported an abundance of Kir3.1 and
ligand-receptor interaction, GDP is exchanged for guanosine tri- Kir3.4 mRNA in the nodal and atrial, but not ventricular, tis-
phosphate (GTP) and results in the uncoupling of Gβγ from Gα sues, which would be consistent with tissue distribution of IKACh.
(see Fig. 4.4A). The released Gβγ-subunits in turn activate the In one such study,42 a comprehensive analysis was performed
Kir3.1/Kir3.4 channels. There is a critical requirement for GTP using Western blot and immunofluorescence to examine chan-
for channel activation (see Fig. 4.4B). A variety of experiments nel distribution in sinus-nodal, atrial, and ventricular tissues, and
have determined the precise molecular interactions involved in showed similar results for the expression patterns across species
the Gβγ-induced Kir3.1/Kir3.4 channel gating, and have shown (rat, ferret, and guinea pig hearts). It was reported that, whereas
that the cytoplasmic region of the channel is intimately involved there was minimal expression of Kir3.1 in the ventricles of all
with gating. For example, crystal structure analyses of the cyto- species tested, Kir3.1 and Kir 3.4 were highly expressed in the
plasmic region of Kir3.1 suggest that the C-terminus of two atrial tissue of all the species. It was noted that although there
neighboring Kir3 channels subunits bind to each other and that an were relatively high levels of expression in the atria, significant

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 STRUCTURAL AND MOLECULAR BASES OF CARDIAC INWARD RECTIFIER POTASSIUM CHANNEL FUNCTION 45

quantitative differences in Kir3.1 and Kir3.4 protein levels were on inherited cardiac channelopathies associated with the Kir3.1/
found in the different species. Furthermore, it was demonstrated Kir3.4 channels. However, alterations in Kir3.1/Kir3.4 channel
that Kir3.1 and M2 receptor colocalized in the sinoatrial node. In
nodal and atrial tissue, immunofluorescence showed localization
activity have been reported in certain cardiac rhythm abnormali-
ties.50 Electrical remodeling in atrial fibrillation patients has been 4
of Kir3.x/M2 receptors more on the outer (lateral) membranes shown to increase the constitutively active component of IKACh,
than in T-tubular membranes.42 Similar quantitative differences which could cause an abbreviation of the action potential dura-
in expression have been reported for atrial versus ventricular tis- tion. More recently, a study was performed in a large Chinese
sues in the human myocardium.42,43  family (49 individuals) with autosomal-dominant long QT syn-
drome (LQTS).51 The locus of the LQTS-associated gene was
Kir3.x Macromolecular Complexes and mapped to chromosome 11q23.3-24.3. A combination of bio-
chemistry and cell electrophysiology in heterologous expression
Channel Biogenesis systems was used to demonstrate that a heterozygous G387R
Similar to Kir2.x channels, multiple signaling molecules and mutation on the Kir3.4 (KCNJ5) gene that was identified in all
cytoskeletal proteins directly or indirectly interact with cardiac affected family members was responsible for the reduced channel
Kir3.x channels in multiprotein scaffolds.4 Thus scaffolds con- expression on the sarcolemma. 
taining Kir3.x channels include a variety of clustering proteins,
actin and protein kinases, and some phosphatases.20,44 Although
experimental data shows that assembly of Kir3.x channels in the ATP-Sensitive Potassium Channels
multiprotein complexes is essential for normal channel function,
the details of Kir3.x protein-protein/lipid interactions remain
Structure and Function
unknown.4,20,45 Caveolae, a group of specialized sarcolemmal Among all inward rectifiers, KATP channels are the most unique
membrane domains of ∼50–100 nm, are a subset of cholesterol in their molecular architecture.52 As in other Kir channels, the ion
and sphingolipids-enriched lipid rafts that play a role in Kir3.x conduction pathway is provided by a tetrameric arrangement of
channel function. Named after the clustering protein caveolin, pore-forming subunits, but additional auxiliary regulatory subunits
these lipid rafts sequester Kir3.x channel complexes, a process are necessary for the channel to be fully functional (Fig. 4.5B). The
thought to exemplify the importance of channel clustering into two known isoforms of the pore-forming subunits are encoded by
specific microdomains. G-protein coupled receptors (adrenergic Kir6.1 (KCNJ8) and Kir6.2 (KCNJ11) genes. The two isoforms
and muscarinic) release Gβγ subunits following ligand biding; of auxiliary subunits are encoded by SUR1 (ABCC8) and SUR2
however, it is only the Gβ-subunits of the Gi/o, and not the Gs, (ABCC9) genes (the SUR name originates from “sulfonylureas,”
protein that are capable of activating the Kir3.x (IK  ACh) channels. a class of drugs known to inhibit KATP channels by acting on the
Therefore it has been hypothesized that lipid rafts may differen- auxiliary subunit). SUR2 gene can be alternatively spliced at the
tially cluster Gs and Gi proteins into distinct microdomains.  very C-terminus (last 42 aa) leading to the SUR2A and SUR2B
isoforms. The genomic arrangements of SUR and Kir6 genes are
unique as well. Specifically, SUR1 is followed by Kir6.2 in close
Pharmacology and Regulation proximity on chromosome 11pp15.1, whereas while SUR2 is fol-
It is well established that Kir3.1/Kir3.4 are generally insensitive to lowed by Kir6.1 on chromosome 12p12.1. The consequences of
classical K+ channel blockers such as or 4-AP or tetraethylammo- this arrangement regarding the regulation at a genomic level are
nium.4 However, experiments in cardiac myocytes isolated from not clear at this time. Although the heteromeric assembly of Kir6.x
rabbit hearts show that the toxin, tertiapin, is a selective blocker subunits produces functional channels in vitro, it remains unclear
of IKACh.46 A channel block is highly potent, with an affinity in whether heteromeric Kir6.x complexes exist in native tissues. In
the nanomolar range. Cardiac Kir3.1/Kir3.4 channels are also contrast, both SUR1/Kir6.2 and SUR2/Kir6.2 channels likely exist
inhibited by quinine, quinidine, and verapamil; however, affini- in native tissues. Membrane topology and general organization of
ties of the these drugs are in the micromolar range.4 Based on Kir6.x subunits is highly similar to that in well-characterized Kir2.x
the results of experiments in heterologous (oocyte) expression channels (see Fig. 4.2).
systems, current through Kir3.x channels is enhanced by “intoxi- KATP channels display weak rectification; however, it has been
cating” concentrations of ethanol, and an approximately 43-aa shown that just a single amino acid substitution in the so-called
stretch on the C-terminus has been identified as important for rectification controller region of inward rectifiers (see Fig. 4.2,
the ethanol effect.47 Similarly, in a heterologous expression sys- Kir2.1; N160D in Kir6.2) converts Kir6.2 into a strongly rec-
tem, Kir3.x channels displayed sensitivity to intracellular acidifica- tifying channel.53 A defining property of KATP channels is their
tion, an inhibitory effect that was dependent on histidine residues characteristic sensitivity to intracellular ATP (hence the name
in the N- and C-terminal regions of the channels.4 A variety of ATP-dependent K+ channels). Under normal conditions, both
other agents have also been shown to modulate Kir3.x channels. exogenously expressed Kir6.2/SUR2 channels and native chan-
For example, similar to all Kir channels, Kir3.1/Kir3.4 require nels in cardiac myocytes are inhibited by ATP in the micromolar
PIP2 for channel activity.4 Curiously, however, the PIP2 effect is range (10 to 100 μM) by direct binding to the channel rather than
enhanced by other factors, such as the G-protein–Gβγ complex, as through phosphorylation mechanisms. In the absence of intracel-
well as by intracellular cations. In general, Kir3.x channel activity lular Mg2+ ions, adenosine diphosphate (ADP) and other nucleo-
is also sensitive to mechanical stretch, can be modified by phos- tides also inhibit the channel. Modeling studies suggests that the
phorylating agents, and is sensitive (negatively) to regulators of ATP binding pocket is located at the interface of the N and C
G-protein signaling.4  terminus of each Kir6.x subunit; therefore the KATP channel pos-
sesses four ATP binding sites.
The overall structure of auxiliary SUR subunit is presented in
Channelopathies Fig. 4.5B. It is believed that SUR interacts with Kir6.x subunits
Over four decades ago, a mouse with a striking locomotor defi- to modulate channel gating through the TMD0 domain and L0
ciency (weaving) was described, and the defect has subsequently linker region. Experimental data are consistent with the idea that
been traced to a naturally occurring gain-in-function mutation in intracellular ATP induces dimerization of nucleotide-binding
the Kir3.2 channel.48 Additional neurological defects attendant to domains NBD1 and NBD2, converting them into a catalytically
this mutation earned the weaver mouse the title of the “most can- active site for the Mg2+-dependent hydrolysis of ATP (leading
tankerous rodent.”49 There is relatively little information available to MgATP). Hydrolysis of ATP is followed by a conformational

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46 CARDIAC ELECTROPHYSIOLOGY

P-helix
TM1 TM2 TMD0 TMD1 TMD2

SURx

88 Å
Slide helix
Kir6
N-terminus L0
NBD1 NBD2

A C-terminus B C
FIGURE 4.5  Molecular structure of K ATP channel.  (A) A pore-forming subunit of KATP channel is
encoded by the Kir6.x genes and contains two transmembrane helical domains, TM1 and TM2; a pore-
forming region (P-helix); a large C-terminal domain; and characteristic N-terminal domain (slide helix)
interfacing the inner leaflet of the membrane and cytoplasmic C-terminus. (B) An auxiliary subunit to
the channel is encoded by SUR1 and SUR 2A/B genes and consists of the seventeen transmembrane
helices organized in several distinct domains and several important cytoplasmic regions. The TMD0 do-
main and L0 region are responsible for the interaction with Kir6.x subunits and regulation of gating of
the channel. TMD1 and TMD2 domains are followed by nucleotide binding domains, NBD1 and NBD2,
which form two nucleotide binding sites at their interface. (C) The arrangement of Kir6.x and SURx
subunits in an octameric KATP channel complex. Modeling studies predict one adenosine triphosphate
binding site at each of the Kir6 interfaces and Mg2+-nucleotide binding sites in the NBD domains of
SUR subunit. (Modified from Flagg TP, Enkvetchakul D, Koster JC, et al. Muscle KATP channels: recent
insights to energy sensing and myoprotection. Physiol Rev. 2010;90:799-829.)

change that is then transduced to the Kir6.x subunit; however, KATP channels. Studies with exogenously expressing systems
MgADP is an even more potent activator of the KATP channel. showed that cloned KATP channels originating from coexpres-
The latter puts the hydrolysis hypothesis into question. Accord- sion of Kir6.1 and SUR2B subunits resemble native KATP chan-
ingly, it has been suggested that NBDs can be locked in a posthy- nels in smooth muscle, for the most part. A strongest support for
drolytic state by MgADP and other nucleotides to sustain the the Kir6.1/SUR2B composition of smooth muscle KATP chan-
active state of the channel.  nels comes from experiments with genetically modified mice.
Specifically, KATP currents cannot be recorded in aortic smooth
muscle cells isolated from either Kir6.1 or SUR2 knockout mice,
Cellular and Membrane Localization whereas the activity of the KATP channels is preserved in these
KATP channels are found in virtually every kind of cardiac tis- cells in Kir6.2 knockout mice.
sues, but they are most prominent in cardiac myocytes and Mitochondrial KATP channels received a lot of attention since
smooth muscle cells. Detailed experimental analysis reveals they were first described at this location in early 1990s. In con-
significant differences in various properties of native KATP trast to sarcolemmal KATP channels, however, their molecular
channels, suggesting a different subunit organization in every identity remains highly controversial.52 Although exogenously
case.54 A significant amount of evidence suggests that ventricu- coexpressed Kir6.1 and SUR1 subunits produce KATP channels
lar sarcolemmal KATP channels are likely composed of Kir6.2 with many properties resembling those found in mitochon-
and SUR2A subunits. In particular, functional sarcolemmal dria, the activity of the mitochondrial KATP channels in Kir6.1
KATP channels are absent in Kir6.2 knockout mice, and various knockout mice was not affected. Recent promising developments
channel properties (including large single channel conductance, in this area include the identification of mitochondria-specific
high sensitivity to pinacidil and cromakalim, and low sensitiv- short-form of SUR2 subunits generated by a nonconventional
ity to diazoxide) are similar to those obtained from ventricular intraexonic splicing, which can underlie the mitochondrial KATP
myocytes. The latter is also supported by a relatively low level channels.55 There is also strong evidence that the pore-form-
of SUR1 (a pancreatic isoform) in the ventricles, and the find- ing subunit of the mitochondrial KATP channels is encoded by
ing that the activity of KATP channels is essentially unaffected in KCNJ1 (Kir1.1).56 
ventricular myocytes from SUR1 knockout mice. Recent experi-
ments with mice showed, however, that SUR1 subunit of the Kir6.x/SURx Macromolecular Complexes and
KATP channel may be a dominant isoform in atrial tissue. In par-
ticular, the activity of the KATP channels could not be detected
Channel Biogenesis
in atrial myocytes isolated from SUR1 knockout mice, and the There is evidence that Kir6.x/SURx channels are assembled in
pharmacological profile of atrial KATP channels is reminiscent of multiprotein scaffolds that are necessary for normal channel func-
that conferred by SUR1 (higher sensitivity to diazoxide, lower tion. The Kir6.x/SURx macromolecular complexes are thought
to pinacidil) rather than SUR2 subunits. It remains unresolved to contain metabolically active protein subunits, including Na/K
whether this subunit composition exists in atrial tissue of other ATPases, as well as kinases.57,58 In one study,59 a number of gly-
animals and humans. colytic enzymes (glyceraldehyde-3-phosphate dehydrogenase,
KATP channels in smooth muscle display a number of proper- triose-phosphate isomerase, and pyruvate kinase), were described
ties distinct from those in cardiac myocytes, suggesting unique as protein subunits of the Kir6.x/SURx protein complex, which
subunit composition.52 Significantly smaller channel densities (up are obligatory for the normal function of the channel. Conse-
to 100-fold per cell), generally low single channel conductance quently, it has been hypothesized that this clustering of channel
(∼30 pS), and a lack of channel activity upon excision of the mem- proteins/enzymes complexes may be needed to modulate the ATP
brane patch are some of the common features of smooth muscle levels in the microenvironment of the channels. Furthermore,

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 STRUCTURAL AND MOLECULAR BASES OF CARDIAC INWARD RECTIFIER POTASSIUM CHANNEL FUNCTION 47

experiments conducted in isolated cardiac myocytes as well as

in heterologous expression systems, demonstrate that currents
Channelopathies
mediated by Kir6.x/SURx channels are modulated by lipid raft
environment.21,59,60 
Mutations in genes underlying the KATP channels have been
associated with cardiovascular disorders. Specifically, the F1524S 4
and A1513T substitutions in the SUR2A gene (missense and
frameshift, respectively) were linked to dilated cardiomyopathy.
Pharmacology and Regulation Both mutations were mapped to the locus of SUR2A, which is
The pharmacology of KATP channels is extensive, and the regu- responsible for the catalytic activity of the NBD2 domains, and
lation is complex relative to other members of Kir family, which likely exert their actions through reduced activation of the KATP
is in part due to the structural complexity of the channel.4 Car- channel. The other reported mutation in the SUR2A gene was
diac KATP channels are regulated by a variety of mechanisms, associated with adrenergic atrial fibrillation originating in the
and important quantitative details of their modulation surely vein of Marshall, a well-known location for this type of fibrilla-
depend on the specific subunit composition. The most promi- tion. As in the previous case, the underlying missense mutation
nent and characteristic mechanism involves regulation by intra- in the SUR2A gene (T1547I substitution) likely affects the func-
cellular nucleotides such as ATP and ADP. Intracellular ATP tioning of the NBD2 leading to a compromised channel regula-
is a potent blocker of the channel while ADP (in the presence tion by adenine nucleotides.
of Mg2+ ions) has an activating effect. An overwhelming level Recent studies have identified multiple mutations in SUR2
of ATP under normal conditions keeps the channel essentially gene. These mutations are associated with Cantu syndrome,
shut (native channels in cardiac myocytes are half blocked by which is a rare multiorgan disease that is characterized, in partic-
50 to 100 μM ATP), whereas metabolic disturbances leading to ular, by various abnormalities of cardiovascular system, including
both a drop in ATP levels and a consequent rise in ADP levels cardiomegaly, concentric ventricular hypertrophy, and other car-
result in channel opening (in the presence of intracellular Mg2+ diovascular malfunctions. The mechanistic link between Cantu
ions). Experiments with isolated membrane patches show that syndrome mutations and various cardiovascular conditions, how-
phospholipids, especially PIP2, are potent activators of native ever, remains to be established.
(and cloned) KATP channels while their hydrolysis reduces chan- Recently, the gain-of-function mutation (S422L substitution)
nel activity. Regulation of KATP channels by PKA-dependent in the pore-forming Kir6.1 subunit (KCNJ8 gene) was associated
phosphorylation is well documented in smooth muscles, but the with ventricular fibrillation and linked to J-wave syndrome sus-
data are limited in cardiac myocytes. It has been shown that the ceptibility,61 and an inframe deletion (E332del) and a missense
activity of atrial KATP channels can be increased by stretch sug- mutation (V346I) were liked to sudden infant death syndrome.62
gesting the involvement of channel-cytoskeleton interactions. Mutations in the KCNJ11 gene encoding pore-forming Kir6.2
Intracellular pH is another potent regulator of native KATP subunit of KATP channels were linked to diabetes mellitus and
channels with acidification leading to an increase in channel congenital hyperinsulinism although no associated cardiac abnor-
activity. malities were reported. 
KATP channels can be inhibited or activated by a variety
of drugs, all acting on the SUR subunit. Sulfonylureas such
as acetohexamide, glipizide, glibenclamide, tolbutamide, and
Conclusion
HMR 1098 are prominent inhibitors. Clinically, sulfonyl- The biophysical and regulatory properties of Kir channels are
ureas are used exclusively for the treatment of type 2 diabetes, crucial for cardiac electrical activity. Significant experimental evi-
but they are also a useful research tool in work with cardiac dence clearly implicates several members of the Kir subfamily as
preparations. molecular determinants underlying the three major inward recti-
KATP channel openers (KCO) include pinacidil, cromakalim, fier potassium currents in native cardiac cells: IK1, IKACh, and IKATP.
rimakalim, nicorandil, diazoxide, and minoxidil sulfate. In con- The general architecture of Kir channels has been well established,
trast to sulfonylureas, KCOs are useful in treating cardiovascu- and fine details of their structure and function have been revealed
lar disorders such as myocardial ischemia and congestive heart with the aid of several available crystal structures of the cloned
failure. KCOs display strong selectivity to subunit composition channels. Nevertheless, many important questions remain unan-
of KATP channels. In particular, KATP channels in pancreatic swered. For example, how do the differences in the biophysical
β-cells (SUR1 based) are strongly activated by diazoxide, but and regulatory properties of Kir2 isoforms affect the heteromeric
not affected by cromakalim or nicorandil while ventricular channel complexes that underlie the native IK1 in different species
KATP channels (SUR2A based) are strongly activated by cro- and in different parts of the heart? What is the subunit composi-
makalim or nicorandil but not by diazoxide. Smooth muscle tion of the mitochondrial KATP channel? How are Kir channels
KATP channels (SUR2B based) are activated by all these drugs. sorted into microdomains in the sarcolemma, such as T-tubules
Mitochondrial KATP channels are known to be potently acti- or intercalated discs, and how do they interact with other proteins
vated by diazoxide and inhibited by 5-hydroxydecanoate within these microdomains? These questions undoubtedly will be
(5-HD).  the focus of much investigation in the near future.

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48 CARDIAC ELECTROPHYSIOLOGY

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