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Vol. 52, Issue 4, 557-594, December 2000
Neurological and Urological Diseases Research, Pharmaceutical Products Division, Abbott Laboratories, Abbott Park, Illinois
Abstract
I. Background
A. Channel Diversity and Classification
1. Six Transmembrane One-Pore Channels.
a. Pore and Selectivity Filter.
b. Voltage Sensor and Channel Activation.
c. Inactivation.
d. Subunit Interaction and Assembly Domains.
2. Two Transmembrane One-Pore Channels.
3. Four Transmembrane Two-Pore Channels.
B. Auxiliary Subunits
C. Crystal Structure of K+ Channels
II. Pathophysiologic Regulation of K+ Channels: Genetically Linked Diseases
A. Cardiac Diseases
1. Long-QT1 and Long-QT5 Syndromes: KCNQ1 (KvLQT1) and minK.
2. Long-QT2 Syndrome and Human ether-a-go-go-Related K+ Channel.
B. Neuronal Diseases
1. Episodic Ataxia/Myokymia and Kv1.1.
2. Benign Familial Neonatal Convulsions and KCNQ2/KCNQ3.
3. Neurodegeneration and Kir3.2.
4. Schizophrenia and SK3 (hKCa3).
C. Hearing and Vestibular Diseases: Nonsyndromic Dominant Deafness and KCNQ4
D. Renal Diseases: Bartter's Syndrome and Kir1.1
E. Metabolic Diseases: Familial Persistent Hyperinsulinemic Hypoglycemia of Infancy and Sulfonylurea Receptor 1
III. Disease- and Drug-Induced Regulation of K+ Channels
A. Cardiac Failure and Hypertrophy
B. Atrial Fibrillation
C. Drug-Induced Long-QT Syndromes
D. Apoptosis and Oncogenesis
1. Apoptosis.
2. Oncogenesis.
E. Alzheimer's Disease
1.-Amyloid.
2.
3. Presenilins.
F. Neuromuscular Disorders
IV. Pharmacological Considerations
A. Voltage-Gated K+ Channels
1. Kv1.3 Channels.
2. Cardiac Delayed Rectifier K+ Channels.
3. KCNQ2/KCNQ3 Channels.
B. Calcium-Activated K+ Channels
1. Large Conductance Channels.
2. Intermediate Conductance Channels.
3. Small Conductance Channels.
C. ATP-Sensitive K+ Channels
D. Two-Pore K+ Channels
V. Concluding Remarks
References
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Abstract |
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Potassium channels play important roles in vital cellular signaling processes in both excitable and nonexcitable cells. Over 50 human genes encoding various K+ channels have been cloned during the past decade, and precise biophysical properties, subunit stoichiometry, channel assembly, and modulation by second messenger and ligands have been elucidated to a large extent. Recent advances in genetic linkage analysis have greatly facilitated the identification of many disease-producing loci, and naturally occurring mutations in various K+ channels have been identified in diseases such as long-QT syndromes, episodic ataxia/myokymia, familial convulsions, hearing and vestibular diseases, Bartter's syndrome, and familial persistent hyperinsulinemic hypoglycemia of infancy. In addition, changes in K+ channel function have been associated with cardiac hypertrophy and failure, apoptosis and oncogenesis, and various neurodegenerative and neuromuscular disorders. This review aims to 1) provide an understanding of K+ channel function at the molecular level in the context of disease processes and 2) discuss the progress, hurdles, challenges, and opportunities in the exploitation of K+ channels as therapeutic targets by pharmacological and emerging genetic approaches.
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I. Background |
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TopPreviousNextReferences
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Potassium channels are a diverse and ubiquitous
family of membrane proteins present in both excitable and nonexcitable
cells. Members of this channel family play critical roles in cellular signaling processes regulating neurotransmitter release, heart rate,
insulin secretion, neuronal excitability, epithelial electrolyte transport, smooth muscle contraction, and cell volume regulation. Over
50 human genes encoding various K+ channels have
been cloned during the past decade (Fig.
1), and precise biophysical properties,
subunit stoichiometry, channel assembly and modulation by second
messenger and ligands have been addressed to a large extent. More
recently, the crystal structure of a K+ channel
from Streptomyces lividans has become available (Doyle et
al., 1998
).
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Concurrent with this remarkable progress in our understanding of molecular diversity, structure, and function, a growing number of discoveries have linked K+ channel gene mutations with various diseases. Such diseases of the heart, kidney, pancreas, and central nervous system involve either mutation(s) in K+ channel gene(s) and/or altered regulation of K+ channel function. The enhanced understanding of these diseases, facilitated by a combination of genomic and biophysical approaches, has helped our understanding of how various mutations affect channel function, contributes to disease etiology, and rationalizes novel treatment strategies. In this review, we provide a comprehensive overview of our recent understanding of molecular defects of K+ channels in various diseases and its implications for the development of novel prophylactic or therapeutic approaches targeting distinct types of K+ channels.
A brief overview of the structural and functional diversity of
K+ channels is initially provided to enable
familiarity with the nomenclature and biophysical and pharmacological
characteristics of diverse K+ channels. Several
extensive reviews are already available on this subject that may be
consulted for additional details (Doupnik et al., 1995; Coetzee et al.,
1999). Diseases involving other voltage-gated ion channels have been
reviewed elsewhere (Ackerman and Clapham, 1997; Lehmann-Horn and
Rüdel, 1997; Cooper and Jan, 1999).
A. Channel Diversity and Classification
K+ channels are membrane-spanning proteins
that selectively conduct K+ ions across the cell
membrane along its electrochemical gradient at a rate of
106 to 108 ions/s. To
accomplish this, K+ channels are endowed with a
set of salient features: 1) a water-filled permeation pathway (pore)
that allows K+ ions to flow across the cell
membrane; 2) a selectivity filter that specifies
K+ as permeant ion species; and 3) a gating
mechanism that serves to switch between open and closed channel
conformations (Hille, 1992). Since the first gene encoding a
K+ channel was cloned from Drosophila
Shaker mutant (Papazian et al., 1987), more than 200 genes
encoding a variety of K+ channels have been
identified (Fig. 1), all containing a homologous pore segment (S5-S6
linker) selective for K+ ions (Hartmann et al.,
1991; Yellen et al., 1991). Accordingly, a general classification of
K+ channels into families is based upon the
primary amino acid sequence of the pore-containing subunit. Three
groups with six, four, or two putative transmembrane segments are
recognized. These include 1) voltage-gated K+
channels (Shaker-like) containing six transmembrane regions
(S1-S6) with a single pore; 2) inward rectifier
K+ channels containing only two transmembrane
regions and a single pore; and 3) two-pore K+
channels containing four transmembranes with two pore regions (Fig.
2). Table
1
lists a generalized classification of various cloned
K+ channel subunits.
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1. Six Transmembrane One-Pore Channels. Voltage-gated K+ channels (Kv), whose members include Shaker-related channels, human ether-a-go-go-related K+ channels (hERG), Ca2+-activated K+ channels, and KCNQ channels, are activated by depolarization.
a. Pore and Selectivity Filter. The tripeptide sequence motif G(Y/F)G located in the S5-S6 linker is common to the pore or P-loop of these and other K+ channels and hence is considered as the K+-selectivity signature motif (Heginbotham et al., 1994). The residues immediately adjacent to either
side of this motif are also generally conserved within the
K+ channel superfamily. Four of the pore loop
domains contribute to the formation of a functional
K+-conducting pore (MacKinnon, 1991).
Accordingly, the heteromultimeric complex of voltage-gated
K+ channels is thought to be composed of four
pore loop-containing 
-subunits arranged in a tetrameric fashion
(MacKinnon, 1995; Jan and Jan, 1997). The external entry to the channel
pore consisting of portions of the P-loop and adjacent residues in both
S5 and S6 segments constitutes binding sites for toxins and
K+ channel blockers (MacKinnon and Miller, 1988;
MacKinnon et al., 1990; Yellen et al., 1991; Goldstein et al., 1993;
Pascual et al., 1995). On the other hand, the internal vestibule of
pore composed of residues from S5 and S6 segments facing the
intracellular side contributes to binding sites for compounds such as
4-aminopyridine, tetraethylammonium, and quinidine (Choi et al., 1993;
Lopez et al., 1994; Shieh and Kirsch, 1994; Yeola et al., 1996). The
S4-S5 linker lies close to the permeation pathway and forms part of the
receptor for the inactivation ball (Isacoff et al., 1991).
b. Voltage Sensor and Channel Activation.
In
voltage-dependent ion channels, membrane depolarization is required to
cause conformational changes leading to channel opening, which allows
permeant ions to flow. The movement of this voltage sensor sensing
changes in membrane potential has been monitored electrically as the
gating current (Armstrong and Bezanilla, 1974). Mutational analysis and
gating current measurements have suggested that the transmembrane S4
segment represents the major component of the voltage sensor (Papazian
et al., 1991; Perozo et al., 1994). The S4 segment that contains
positively charged residues (lysine or arginine) at approximately every
third position resulting in a regularly spaced array of 5 to 7 positive
charges is conserved within the voltage-gated K+
channel family. The rearrangement of S4 in response to membrane depolarization has also been confirmed by the means of fluorescence techniques (Mannuzzu et al., 1996; Cha and Bezanilla, 1997). While the
S4 segment comprises the major part of the voltage sensor required for
the K+ channel activation, the electrostatic
interaction of negative charges in S2 and S3 with the S4 segment also
contributes to the gating mechanism (Papazian et al., 1995; Seoh
et al., 1996). The nature of the gate that ultimately controls access
of permeant ions to the pore is not conclusively established. Studies
involving mutational analysis, gating current measurements, and the
substituted cysteine accessibility method point to several residues in
the S5 and S6 segments that might form the activation gate regulating access of ions to the pore (Liu et al., 1997; Shieh et al., 1997; Kanevsky and Aldrich, 1999).
c. Inactivation.
Many voltage-dependent
K+ channels activate and inactivate rapidly when
membrane potential becomes more positive. Inactivation is a
nonconducting state during maintained depolarization. Three types of
inactivation, i.e., N-, P-, and C-type, have been characterized and
associated with distinct molecular domains of the channel. For example,
the N-terminal residues (amino acids 6-46) of the Shaker
K+ channel involved in N-type inactivation moves
into the internal vestibule (in S4-S5 linker) to occlude the pore when
the channel opens (Hoshi et al., 1990; Isacoff et al., 1991). After
removal of this N terminus region, inactivation can be restored in the mutant K+ channel by the corresponding synthetic
peptide (Zagotta et al., 1990). In contrast to the fast process of
N-type inactivation, the C- and P-type inactivation involves a slower
rearrangement of outer mouth and specific residues in the pore,
respectively (Hoshi et al., 1991; De Biasi et al., 1993; Yellen et al.,
1994; Liu et al., 1996).
d. Subunit Interaction and Assembly Domains.
As noted
previously, K+ channels contain four
-subunits, which surround a water-filled,
K+-selective pore (Fig. 2). Among diverse
voltage-gated K+ channels, only closely related
subfamilies of -subunits are capable of coassembling to form
heteromultimers. For example, in the Kv1 subfamily, a highly conserved
cytoplasmic sequence immediately preceding the first transmembrane
segment (amino acid residues 83 to 196) was identified as important to
subfamily-specific channel assembly (Li et al., 1992). In
Shaker channels, a conserved region (T1, or tetramerization
domain 1) in the first transmembrane segment is involved in formation
of tetramers (Shen et al., 1993). However, in more distantly related
voltage-gated K+ channels,
ether-a-go-go (EAG), hERG, and KCNQ1
K+ channel subfamilies, channel assembly
primarily involves C-terminal domains (Ludwig et al., 1997; Kupershmidt
et al., 1998). As discussed in the following sections, patients with
Jervell and Lange-Nielsen long-QT (LQT) syndrome are characterized by
the absence of KCNQ1 heteromultimers caused by mutations in the C
terminus that impair subunit assembly (Schmitt et al., 2000).
2. Two Transmembrane One-Pore Channels.
The inward rectifier
K+ channels (Kirs) belong to a distant
superfamily of channels with four subunits each containing a
two-transmembrane segment (M1 and M2) and a pore loop in between (Ho et
al., 1993; Kubo et al., 1993). These channels conduct
K+ currents more in the inward direction than
outward, and they are important in setting the resting membrane
potential. This inward rectification is attributed to gating mechanisms
by internal Mg2+ and polyamines (spermine,
spermidine, etc.) that occlude access of K+ to
the internal vestibule of a conducting pore (Matsuda, 1991; Ficker et
al., 1994; Lu and MacKinnon, 1994; Wible et al., 1994). Like the
voltage-gated K+ channels, these channels are
organized as tetramers (Yang et al., 1995), although a more complex
octameric arrangement has been described, as in the case of the
ATP-sensitive K+ channels involving four inward
rectifiers contributing to ion conducting pore and four peripheral
sulfonylurea receptors as regulatory subunits (Clement et al., 1997;
Inagaki et al., 1997; Shyng and Nichols, 1997).
3. Four Transmembrane Two-Pore Channels.
The more recently
discovered tandem-pore domain family are weak inward rectifiers with
four putative transmembrane domains and two pore domains
(Ketchum et al., 1995; Lesage et al., 1996a). They represent perhaps
the most abundant class of K+ channels (at least
in C. elegans), with >50 distinct members (Wang
et al., 1999). The G(Y/F)G residues of
K+-selective motif is preserved in the first pore
loop of the two-pore K+ channel, but it is
replaced by GFG or GLG in the second pore loop. Although all the
two-pore channels have a conserved core region between transmembrane
segments M1 and M4, the amino- and carboxyl-terminal domains are quite
diverse. With two-pore domain subunits, two such subunits would
presumably form a channel to retain the tetrameric arrangement.
B. Auxiliary Subunits
Auxiliary subunits that associate with many of the
pore-forming subunits have also been described (reviewed in Isom et
al., 1994). For example, the Kv1 channels associate with cytoplasmic 
-subunits to alter channel kinetics (reviewed in Xu and Li, 1998). More recently, chaperone proteins, such as KChAP, regulating the function and expression of some of the Kv channels, such as Kv2.1, Kv1.3, and Kv4.3, have been reported (Kuryshev et al., 2000b). Certain
other Kv channels, such as Kv5, Kv6, Kv8, and Kv9, do not form
functional channels themselves but associate with Kv2.1 channels to
alter the biophysical properties (Salinas et al., 1997; Kramer et al.,
1998). Other examples include distinct -subunits that associate with
the calcium-activated K+ channels (Tseng-Crank et
al., 1996; Wallner et al., 1999a; Behrens et al., 2000; Brenner et al.,
2000; Meera et al., 2000), sulfonylurea receptors for the inward
rectifiers Kir6.1 or Kir6.2 (Aguilar-Bryan et al., 1995; Inagaki et
al., 1995a), and minK and minK-related peptides (MiRPs) for the
cardiac delayed rectifier channels (Barhanin et al., 1996; Sanguinetti
et al., 1996; Abbott et al., 1999). These subunits play roles as
diverse as modulation of gating properties such as inactivation, cell
surface expression, and/or trafficking of the ion channel complex, to
serving as binding sites for both endogenous and exogenous ligands.
Given the diversity of K+ channel subunits and
the potential to vary the constituents to form diverse - or
- heteromeric channel complexes to alter expression, cellular
targeting, and biophysical and pharmacological properties in native
cell types, understanding the precise composition of channel complexes
in vivo remains a challenge.
C. Crystal Structure of K+ Channels
Initial studies of the structure and function of
K+ channels by a combination of mutagenesis and
biophysical approaches have revealed domains that are responsible for
K+ selectivity, gating, channel assembly, subunit
interaction, and drug binding sites. However, the three dimensional
structural implications remained largely speculative. Recent discovery
of the crystal structure of the KCsA channel established a
blueprint of K+ channel structure with 3.2 Å resolution (Doyle et al., 1998). The KCsA channel is encoded by a
bacterial gene cloned from S. lividans on the
basis of sequence homology to K+-selective motif
GYG in the P-loop (Schrempf et al., 1995). The KCsA channel contains
only two transmembrane domains with an intervening pore loop, although
at the amino acid sequence level, this channel is more similar to the
voltage-gated K+ channels. Functionally, it lacks
any hint of voltage gating because of the lack of S4 region. X-ray
analysis revealed that four identical subunits form a tetramer creating
an inverted cone, cradling the selectivity filter of the pore in its
outer end. The overall length of the conducting pore is 45 Å, and its
diameter is variable along its distance. The internal vestibule of the
pore begins as a tunnel of 18 Å in length that widens into a cavity
(~10 Å across) near the middle of the membrane, with the narrow
selectivity filter only 12 Å long. The remainder of the pore is wider
and lined with hydrophobic amino acids. The selectivity filter is lined
by the carbonyl oxygen atoms of the GYG signature sequence, which is held open by structural constraints to coordinate
K+ ions (~3 Å) but not smaller
Na+ ions because the diameter is too wide to
substitute for the hydration energy of the Na+
ions (Doyle et al., 1998). The crystal structure of KCsA channel provides the first three-dimensional structure of the conduction pore
that fits consistently with current understanding of the core
functionality of K+ channels. However, structural
information of the remaining transmembrane segments (S1-S4),
particularly the voltage sensor and the gate coupling to channel
opening and closing, remains to be elucidated. Nevertheless, the
understanding of structural information can be applied to design
selective compounds targeting K+ channels. For
example, a structure-based design strategy allowed several
charybdotoxin analogs to be prepared with about 20-fold higher affinity
to block Ca2+-activated K+
channels versus voltage-gated Kv1.3 channels (Rauer et al., 2000). It
is to be anticipated that a detailed understanding of the structural aspects would revolutionize and refine approaches targeting
K+ channels for therapeutic purposes.
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II. Pathophysiologic Regulation of K+ Channels: Genetically Linked Diseases |
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Advances in genetic linkage analysis during the past decade have greatly facilitated the identification of many disease-producing loci. Both positional cloning and candidate gene approaches have been used. Using positional cloning techniques, it has become possible to identify the location of genetic locus responsible for a given hereditary syndrome without prior knowledge of the biochemical or physiological abnormalities underlying the disease. Alternatively, following identification of genes encoding proteins that may be logically altered in a particular disease, the candidate gene approach may be used to examine genetic linkage to the hereditary disease of interest and screened for mutations.
As K+ channels play fundamental roles in the regulation of membrane excitability, it is to be expected that both genetic and acquired diseases involving altered functioning of neurons, smooth muscle, and cardiac cells could arise subsequent to abnormalities in K+ channel proteins. Genetically linked diseases of the cardiac, neuronal, renal, and metabolic systems involving members of voltage-gated K+ channels, inward rectifiers, and channel-associated proteins are discussed in the following sections (Table 1).
A. Cardiac Diseases
K+ channels are critical to cardiac
excitability because they play a fundamental role in
repolarization of the action potential. Unlike the action potentials of
nerves that last only a few milliseconds, the action potentials of
ventricular myocytes can last several hundred milliseconds. This
prolonged depolarization phase is essential for normal
excitation-contraction coupling process and renders the myocytes
relatively refractory to premature excitation. Various classes of
K+ channels with different time and voltage
dependencies and pharmacological properties function in concert to
regulate the heart rate by setting the resting membrane potential,
amplitude, and duration of action potential and its refractoriness
(Barry and Nerbonne, 1996; Roden and Kupershmidt, 1999; Snyders, 1999).
The Kir2.1 current sets the resting membrane potential and contributes
to the terminal phase of repolarization. The transient outward
K+ current (Kv4.3 or Kv1.4), which is
Ca2+-independent and expressed in a species- and
cell type-specific fashion, is important for the early phase of
repolarization. The long ventricular action potentials that result from
the slow onset of repolarization are controlled mainly by two types of
delayed rectifier K+ currents, i.e., IKs (derived
from KCNQ1/minK) and IKr (derived from hERG/MiRP1). Both genetic
linkage analysis and the candidate gene approach revealed that
mutations in these delayed rectifier K+ channel
subunits form the molecular basis of LQT syndromes (Curran et al.,
1995; Sanguinetti et al., 1995; Schott et al., 1995; Wang et al., 1996;
Neyroud et al., 1997; Splawski et al., 1997b; Abbott et al.,
1999).
The LQT syndromes are inherited genetic disorders characterized by
prolonged or delayed ventricular repolarization, manifested on the
electrocardiogram (ECG) as a prolongation of the QT interval. Table
2 lists K+ and
other ion channel genes involved in various forms of inherited LQT
syndromes, LQT1 through LQT6. The inherited LQT causes syncopal attacks
and high risk of sudden death as result of torsade de pointes
polymorphic ventricular tachycardia, typically triggered by adrenergic
arousal (Ackerman and Clapham, 1997; Sanguinetti and Spector, 1997;
Vincent et al., 1999). Based on genetic origins, two allelic diseases
are recognized: 1) the Romano-Ward syndrome inherited as a
dominant trait and 2) the autosomal recessive Jervell and Lange-Nielsen
syndrome. In the case of the latter, the patient suffers from a severe
congenital bilateral deafness in addition to the cardiac disorder
(Vincent et al., 1999). Note that in addition to genetically linked LQT
syndromes, many drugs are also known to cause QT prolongation leading
to torsade de pointes (see Section III.).
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1. Long-QT1 and Long-QT5 Syndromes: KCNQ1 (KvLQT1) and minK.
KvLQT1, encoded by the KCNQ1 gene, in association with the minK
subunit, a short peptide of 130 residues, constitutes the IKs
responsible for phase 3 repolarization in the heart (Barhanin et al.,
1996; Sanguinetti et al., 1996b). Several mutations in the
KCNQ1 gene, including missense mutations, intragenic
deletion, and insertions, are involved in chromosome 11-linked LQT1
syndrome, the most common form of inherited LQT in families with
Jervell and Lange-Nielsen and Romano-Ward syndromes (Russell et al.,
1996; Wang et al., 1996; Donger et al., 1997; Tanaka et al., 1997; van den Berg et al., 1997; Saarinen et al., 1998; Li et al., 1998; Neyroud
et al., 1999;). Functional analysis of mutant channels in COS cells
cotransfected with the minK subunit revealed that these mutations
either alter gating properties or fail to produce functional homomeric
channels and reduced K+ current when coexpressed
with the wild-type subunit (Chouabe et al., 1997; Shalaby et al., 1997;
Wollnik et al., 1997; Franqueza et al., 1999).
; Duggal et al., 1998). Again, functionally, these mutations yield
diminished IKs current when coinjected with KCNQ1 either
by suppressing channel function in a dominant-negative fashion,
increasing rate of channel deactivation, or by shifting the voltage
dependence of channel activation in a positive direction. It is likely
that the mutations in KCNQ1 associated with LQT1 will
decrease the availability of IKs by altering gating properties or by a
dominant-negative loss of channel function leading to a prolonged
ventricular repolarization. Accordingly, activators that restore the
function of IKs may prove useful in the treatment of LQT1 and LQT5
syndromes. Recently, Abitbol et al. (1999) have shown that stilbenes
and fenamates, by binding the extracellular domain flanking the minK
transmembrane segment, restored inactive IKs mutant channels, including
the naturally occurring LQT5 mutant, D76N.
Neyroud et al. (1997) have also identified a homozygous
deletion-insertion event in the C-terminal domain of KCNQ1
in three affected children from two families with congenital bilateral deafness associated with QT prolongation. By in situ hybridization studies in mice, it was shown that the KCNQ1 gene was
expressed by the marginal cells of the stria vascularis. It has been
suggested that, in conjunction with the minK subunit, KCNQ1 forms a
functional channel in marginal cells that is responsible for secretion
of endolymph, in the inner ear, which bathes the stereocilia of sensory hair cells. Thus, KCNQ1 plays a key role not only in the ventricular repolarization but also in normal hearing, probably via control of
endolymph homeostasis (see Section II.C.).
2. Long-QT2 Syndrome and Human ether-a-go-go-Related K+
Channel.
The hERG gene encoding a rapidly activating IKr is a
major subunit responsible for repolarization during cardiac action
potential (Sanguinetti et al., 1995). Interaction with hERG channels
has been shown to be a primary mechanism involved in the therapeutic actions of the class III antiarrhythmic agents and the potential cardiotoxicity of second generation H1 receptor
antagonists, such as terfenadine and astemizole, as well as certain
antidepressants and neuroleptics (Vincent et al., 1999).
first demonstrated that missense mutations, intragenic
deletions, and splice donor mutations in the hERG gene resulted in
chromosome 7-linked LQT2 syndrome. This finding was further confirmed
by studying several mutations in different regions of the hERG subunit
in families associated with LQT syndromes (Benson et al., 1996; Dausse
et al., 1996; Satler et al., 1996, 1998; Tanaka et al., 1997). Similar
to KCNQ1, mutations of hERG decrease repolarizing current and thus
lengthen the duration of cardiac action potential. The mutant hERG
cRNA, when expressed alone or in combination with wild-type channel,
yields nonfunctional channels or evokes dominant negative suppression
of hERG function (Sanguinetti et al., 1996a; Li et al., 1997; Babij et
al., 1998; Nakajima et al., 1998). By green fluorescent protein tagging
and Western blot analyses, it was found that the hERG-G601S mutant was
deficient in the trafficking of functional protein to the plasma
membrane (Furutani et al., 1999), which could explain the reduction in
functional channels available for repolarization of the cardiac action
potential. Other LQT-associated mutations identified in the
amino-terminal region of hERG form functional channels, but with
altered gating properties such as accelerated channel deactivation, and
positively shifted voltage dependence of channel open probability.
Collectively, these alterations lead to reduced outward current during
the repolarization phase of the cardiac action potential and prolonged
QT interval (Chen et al., 1999a). The diversity of mutations in the
hERG gene impairing channel function in varying proportions likely
contributes to variable degrees of clinical severity in LQT2 patients.
Although channels formed of hERG subunits appear similar to IKr, and
although mutations in hERG gene are associated with LQT2 syndrome, the
recombinant channels differ in gating, single channel conductance, and
sensitivity to antiarrhythmic drugs compared with native currents.
Another small membrane subunit, MiRP1, cloned by searching the
expressed sequence tag (EST) database, was found to assemble with hERG
to alter its function (Abbott et al., 1999). Injection of MiRP1 cRNA
alone into oocytes revealed no currents by itself, whereas MiRP1 had
significant effects on the properties of channels formed with hERG
subunits but not with other K+ channels,
including KCNQ1, Shaker, and Kv members. Coexpression of
MiRP1 with hERG revealed functional current with gating and sensitivity
to E-4031 similar to native cardiac IKr. Three missense mutations
associated with the LQT6 syndrome and ventricular fibrillation have
been identified in the MiRP1 gene. The mutant channels open slowly and
close rapidly, thereby evoking diminished K+
currents. One variant, associated with clarithromycin-induced arrhythmia, increases sensitivity to channel blockade by the
antibiotic. The latter finding reveals an important mechanism for
acquired arrhythmia wherein a genetically based reduction in
K+ currents remains silent until combined with
additional factors.
-Adrenoceptor antagonists have been used in the treatment of LQT1
and LQT2 syndromes since episodes of syncope and sudden death occur
more frequently with exercise and at times of adrenergic surges
(Vincent et al., 1999). The mechanism of dysfunction of hERG and MiRP1
associated with LQT suggests that activators for these channels may be
therapeutically useful. Expression of hERG alone reveals little outward
K+ current upon depolarization, whereas large
inward K+ currents are seen when the membrane
voltage is hyperpolarized due to removal of C-type inactivation (Smith
et al., 1996; Spector et al., 1996). Elevation of external
K+ levels reduces this C-type inactivation,
thereby increasing outward K+ currents and
reducing the prolongation of cardiac action potential with LQT2.
Indeed, Compton et al. (1996) have shown that elevation of serum
[K+] using K+ supplements
and spironolactone in patients with LQT2 demonstrated a significant
reduction of the QT interval. Although it is difficult to maintain an
elevated level of serum K+, these findings
suggest that the patients could avoid administration of drugs that
cause hypokalemia.
B. Neuronal Diseases
K+ channels are critical to neurotransmission in the nervous system. Alterations in the function of these channels lead to remarkable perturbations in membrane excitability and neuronal function. Significant progress has been made in linking many neuronal disorders, including episodic ataxia and benign familial neonatal convulsions, to K+ channel mutations.
1. Episodic Ataxia/Myokymia and Kv1.1. Episodic ataxia (EA) is an autosomal dominant disorder in which the affected individuals have brief episodes of ataxia triggered by physical or emotional stress. On the basis of the duration and severity of the attacks, two types of episodic ataxia are recognized. In EA type 1 with onset in early childhood, the ataxia occurs several times during the day, lasts for seconds to minutes, and is associated with dysarthria and motor neuron activity, which causes muscle rippling (myokymia) between and during attacks. In contrast, in EA type 2, the attacks last for hours to several days and are precipitated by emotional stress and exercise, but they do not startle. This type of ataxia is associated with nystagmus and cerebellar atrophy, unlike the EA-1 type in which the affected children do not develop persistent ataxia or cerebellar atrophy.
Linkage analysis has mapped episodic ataxia to two different ion channel genes. EA-2 is associated with missense mutations in CACNA1A, encoding a brain-specific P/Q-type Ca2+ channel located on chromosome 19p13, the same region associated with familial hemiplegic migraine, suggesting the possibility that both EA-2 and familial hemiplegic migraine are allelic disorders (Ophoff et al., 1996; Jen et al., 1999). By linkage
studies, Litt et al. (1994) localized the EA-1 gene to chromosome 12p,
where the KCNA1 gene encoding the voltage-gated
K+ channel in brain and peripheral nervous
systems has been mapped. Mutational analysis of KCNA1 in
several families with EA-1 has identified at least ten missense
mutations (Browne et al., 1994; Scheffer et al., 1998). These mutations
alter Kv1.1 function by reducing channel expression (dominant-negative
effect), altering gating properties by shifting the midpoint of current
activation some 10 to 40 mV in the depolarization direction, or
enhancing deactivation or C-type inactivation rates (Adelman et al.,
1995; Zerr et al., 1998; Boland et al., 1999; Bretschneider et al., 1999). Accordingly, it could be inferred that altered Kv1.1 function could impair the capacity of the affected neurons to repolarize effectively following an action potential. Further support for the
notion that the diminished function of KCNA1 leads to ataxia is obtained from gene knockout studies in which the homozygous mutant
mice exhibit attacks of tremors and marked ataxia after cold-temperature stress (Smart et al., 1998).
Acetazolamide, a carbonic anhydrase inhibitor, has been effective in
reducing attack episodes in some patients suffering from EA-1. However,
this compound did not affect Kv1.1 wild-type or mutant channels
(Bretschneider et al., 1999). Pharmacological agents that either shift
the voltage dependence of Kv1.1 channel activation to more negative
potentials or enhance the magnitude of current could, in principle,
prevent both ataxia and myokymia (Sanguinetti and Spector, 1997).
2. Benign Familial Neonatal Convulsions and KCNQ2/KCNQ3.
Recent application of genetic analysis to hereditary epilepsy has
provided the impetus for the identification of mutations in genes
encoding various ion channels, including K+
channels (Biervert et al., 1998; Charlier et al., 1998; Singh et al.,
1998). BFNC is an idiopathic form of epilepsy beginning within the
first six months after birth. Seizures are generalized and mixed,
starting with tonic posture, ocular symptoms, and apnea, and often
progress to clonic movements and motor automatisms. Seizures last 1 to
2 s and occur three to six times per day. Two forms of benign
familial neonatal convulsions, BFNC1 and BFNC2, are typically observed
in families as an autosomal dominant inheritance and have been
previously mapped into chromosomes 20q and 8q, respectively (Leppert
and Singh, 1999). By positional cloning techniques, the voltage-gated
K+ channel KCNQ2, spanning the
deletion region of chromosome 20q13.3 that cosegregates with seizures
in a BFNC family, was identified (Biervert et al., 1998; Singh et al.,
1998). Missense mutation, frameshifts, and splice-site mutations in
KCNQ2 were also found in other BFNC families. By a homology
search of expressed sequence tag database and genotyping approaches, a
missense mutation in the pore region of another voltage-gated
K+ channel, KCNQ3, was also identified
from families with BFNC2 previously linked to chromosome 8q24 (Biervert
et al., 1998; Charlier et al., 1998; Schroeder et al., 1998).
).
First described in the peripheral sympathetic neurons and subsequently
in the CNS, this channel is one of the most important regulators of
neuronal excitability because it plays a critical role in determining
the excitability threshold, firing properties, and responsiveness of
neurons to synaptic inputs. In the absence of acetylcholine, the
M-channel activity hyperpolarizes the cell membrane potential, leading
to a dampening of neuronal responsiveness to synaptic inputs. However,
in the presence of released acetylcholine, the M-channels are
inhibited. This change in M-channel activity provides a mechanism for
neurons to respond to synaptic input and to favor firing a burst of
spikes, rather than a single spike, upon excitation (Hille, 1992). By
voltage-clamp recording of neurons from rat superior cervical ganglion,
Marrion et al. (1989) determined that ACh-evoked suppression of the
M-channel is mediated by the activation of muscarinic M1 receptors.
Seizures in mice induced by a muscarinic agonist, pilocarpine, were
sensitive to inhibition by a muscarinic M1 antagonist, pirezepine
(Maslanski et al., 1994). Furthermore, in transgenic mice lacking
muscarinic M1 receptors, the robust suppression of the M-current
activity evoked by muscarinic agonists in sympathetic ganglion neurons
was eliminated. Both homozygous and heterozygous mutant mice were also
resistant to seizures evoked by systemic administration of pilocarpine
(Hamilton et al., 1997). Taken together, these studies suggest that
M-channels play a key role in controlling seizure activity.
Both KCNQ2 and KCNQ3 belong to the
KCNQ family of K+ channels that
includes KCNQ1 (KvLQT1), whose aberrant function leads to the congenital bilateral deafness associated with QT prolongation. The
KCNQ2 protein exhibits 62% identity with KCNQ3 within the coding
region and is also highly conserved with KCNQ1 in transmembrane S1-S6
region with 60% identity and 70% similarity (Biervert et al., 1998;
Charlier et al., 1998; Tinel et al., 1998). Unlike KCNQ1,
which is expressed strongly in human heart and pancreas, KCNQ2 and KCNQ3 transcripts are detectable only
in brain (Biervert et al., 1998, Wang et al., 1998; Yang et al., 1998)
and in rat sympathetic ganglia (Wang et al., 1998). Expression of human
KCNQ2 was found to be high in the hippocampus, caudate
nucleus, and amygdala, moderate in the thalamus, and weak in the
subthalamic nucleus, substantia nigra, and corpus callosum. A similar
expression pattern for KCNQ3 was found in the human brain
(Biervert et al., 1998; Tinel et al., 1998; Yang et al., 1998).
In human brain, four splice variants of KCNQ2 were identified, among
which only two forms generated K+-selective
currents when heterologously expressed in oocytes or COS cells (Tinel
et al., 1998). These currents resemble those of KCNQ1 in their
permeability sequence of cations, voltage dependence, and kinetics
(Biervert et al., 1998; Tinel et al., 1998). When expressed in
Xenopus oocytes, KCNQ3 elicited currents that were only
slightly above background but resembled the larger
depolarization-activated K+ currents observed
with KCNQ2 (Schroeder et al., 1998; Wang et al., 1998). Unlike
KCNQ1 (KvLQT1), where coinjection with minK (KCNE1) dramatically alters the amplitude and gating
kinetics of the KCNQ1 channel and produces current resembling cardiac
IKs, neither KCNQ2 or KCNQ3 currents were altered when coinjected with the minK subunit (Yang et al., 1998). However, when KCNQ2
and KCNQ3 mRNAs were coinjected in the Xenopus
oocytes, the resultant current was more than 10-fold larger than that
observed in cells injected with either KCNQ2 or KCNQ3 alone
(Schroeder et al., 1998; Wang et al., 1998; Yang et al., 1998). The
expressed K+ current by coinjection with
KCNQ2 and KCNQ3 has gating kinetics and
sensitivities to blockade by classical M-channel inhibitors such as
linopirdine and XE991, indicating that the M-channel is a
heteromultimer composed of KCNQ2 and KCNQ3 subunits (Wang et al.,
1998).
No detectable currents were expressed when cRNA of the truncated
KCNQ2 identified from families with BFNC1 alone were
injected. When mutant and wild-type cRNA were coinjected at a 1:1 ratio to mimic the situation in a heterozygous patient, the currents were
reduced, compared with those recorded from oocytes injected with
similar amounts of wild-type cRNA. Thus, although there was no obvious
dominant negative effect, haploinsufficiency may be enough to explain
the dominant mode of inheritance of this disorder, which generally
occurs transiently during infancy (Biervert et al., 1998). Two single
mutations in KCNQ2 (Y284C and A306T), as well as insertion
mutant associated with BFNC1, were analyzed for current amplitude when
coexpressed with KCNQ3. The function of these mutant
heteromeric channels was significantly reduced, and no dominant
negative effect was observed. Likewise, when the KCNQ3
mutant G310V was coexpressed with wild-type KCNQ2, a loss function effect rather than a dominant-negative effect was seen (Schroeder et al., 1998). Together, Schroeder et al. (1998) suggested that a 25% loss of heteromeric KCNQ2/KCNQ3 function is sufficient to
cause the hyperexcitability in BFNC. Recently, another missense mutation that replaced tryptophan with arginine (W309R) in the P-loop
of KCNQ3 was also reported from patients with BFNC (Hirose et al.,
2000).
The cytoplasmic N terminus of KCNQ2 contains a consensus
site for cAMP-dependent phosphorylation, and increases in intracellular cAMP concentration have been shown to enhance KCNQ2/KCNQ3 current by
50% (Schroeder et al., 1998). Compounds that open or enhance the
activity of the M-currents, such as retigabine, or elevate associated
intracellular cAMP levels may serve as useful antiepileptic agents. It
should be pointed out that in addition to M-channel mutations linking
to BFNC disorders, mutations in other ion channels have been associated
with varying forms of epilepsy (Steinlein, 1999). These include
mutations of the neuronal nicotinic acetylcholine receptor 4-subunit
(CHRNA4), identified to be responsible for the autosomal
dominant nocturnal frontal lobe epilepsy (Steinlein et al., 1995,
1997), and those involving voltage-gated sodium channel 1-subunit
(SCN1B) identified in families associated with generalized
epilepsy with febrile seizures (Wallace et al., 1998). Collectively,
the discoveries of these ion channels as epilepsy disease genes
emphasize the potential roles of ion channels in epilepsy and suggest
that compounds that directly or indirectly modulate these channels may
prove helpful in suppressing seizures.
3. Neurodegeneration and Kir3.2.
The progressive loss of
dopaminergic neurons in the weaver mouse is similar to the
pathological symptom of Parkinson's disease where cell death of
dopaminergic neurons in the substantia nigra is observed, leading to
striatal dopaminergic deficit and a clinical syndrome dominated by
disorders of movement (Yamada et al., 1990; Gaspar et al., 1994). The
weaver phenotype in mice is an autosomal recessive
neurological and reproductive disorder characterized behaviorally by
severe ataxia, hyperactivity, and tremors that are manifested within 2 weeks after birth. These behavioral changes are attributable to the
degeneration of cerebellar granule cells and dopaminergic neurons in
the substantia nigra (Rakic and Sidman, 1973a,b). In addition,
wv/wv genotype causes death or impaired function of
dopaminergic neurons in the substantia nigra, male infertility, and
sporadic tonic-clonic seizures (Hess, 1996; Harrison and
Roffler-Tarlov, 1998). While heterozygous mice are not ataxic, they
have seizures and a significant reduction in the number of granule cells.
). By a combination of physical and transcript mapping of the
homologous segment on human chromosome 21, Patil et al. (1995)
identified two potential candidate genes in this region: 1)
mmb, encoding a serine/threonine-specific protein
kinase, and 2) Kir3.2, encoding a G protein-gated inwardly
rectifying K+ channel. Sequence analysis yielded
no mutations in mmb, whereas a single missense mutation
replacing a glycine with serine at residue 156 (G156S) was observed in
Kir3.2 associated with weaver mouse in a location within the
pore-forming region, critical for ion selectivity and conserved within
the K+ channel family (MacKinnon, 1995). The
mutation renders the channel nonselective, leading to conduction of
Na+ ions instead of the highly selective
K+ ions (Navarro et al., 1996; Slesinger et al.,
1996). Ribonuclease protection and reverse transcriptase-polymerase
chain reaction studies have shown that the overall expression pattern
of Kir3.2 gene parallels the developmental loss of the cells in
cerebellum, substantia nigra, and testes (Patil et al., 1995; Slesinger
et al., 1996).
It has been shown that Kir3.2 coassembles with Kir3.1 to form the G
protein-gated, K+-selective inward rectifier
channels in neurons (Duprat et al., 1995; Liao et al., 1996;
Velimirovic et al., 1996). Immunohistochemical localization studies
indicate that Kir3.2 and Kir3.1 proteins are expressed in the
cerebellar neurons of mice at postnatal day 4, at a time when neurons
normally undergo differentiation (Slesinger et al., 1996). Functional
analysis of expression of wvKir3.2 and Kir3.2 in
Xenopus oocytes or Chinese hamster ovary cells revealed that
the mutant channel showed reduced sensitivity to muscarinic M2 receptor
activation, failed to respond to G
subunit,
and evoked diminished K+ currents. Furthermore,
the loss in selectivity for K+ and increased
basal current resulting from increased Na+
permeability leads to alterations in membrane excitability, cell differentiation, and ultimately cell death (Kofuji et al., 1996; Navarro et al., 1996; Silverman et al., 1996; Slesinger et al., 1996;
Rossi et al., 1998). Results from transgenic studies confirmed that the
weaver phenotypes arise from a gain-of-function mutation of
Kir3.2. Although the transgenic mice lacking Kir3.2 (
/) are morphologically indistinguishable from the wild type, they have much
reduced Kir3.1 expression in the brain, develop spontaneous seizures,
and are more susceptible to pharmacologically induced seizures induced
by pentylenetetrazol (Signorini et al., 1997).
The nonselective cation current in cells expressing wvKir3.2
can be blocked by MK-801 and calcium channel blockers (Kofuji et al.,
1996). These compounds have been shown to enhance cell viability and
neurite outgrowth of cultured weaver granule cells, but not
of wild-type granule cells. In addition, neurite outgrowth and
migration of the weaver granule neurons has also been shown to be enhanced by Fab2 fragments of antibodies raised against a neurite
outgrowth domain of the laminin B2 chain (Liesi and Wright, 1996).
As mentioned previously, the degeneration of noncalbindin-positive
dopaminergic neurons in substantia nigra of weaver mice shares similarity to Parkinson's disease, in which the dopaminergic neurons that are progressively lost in the substantia nigra are also
noncalbindin-positive. These observations suggest the possibility of a
shared genetic defect in weaver mouse and Parkinson's
disease (Yamada et al., 1990; Gaspar et al., 1994). However, Bandmann et al. (1996) did not detect mutations by sequencing analysis of the
pore-forming region of Kir3.2 gene from patients with familial and
sporadic cases of Parkinson's disease, suggesting a differing etiology
of nigral cell loss in Parkinson's disease and weaver mice.
Nevertheless, the finding that weaver phenotype results from
a single amino acid mutation in Kir3.2 leading to alterations in
membrane excitability provides a reasonable avenue for understanding the molecular nature of this neuronal disorder.
4. Schizophrenia and SK3 (hKCa3).
Although initially
differentiated on the basis of biophysical and differential toxin
sensitivity, distinct genes are now known to encode various
calcium-activated K+ channels (Vergara et al.,
1998; Castle, 1999; Wallner et al., 1999b). Abnormal function of
calcium-activated K+ channels has been noted in
platelets of patients with Alzheimer's disease, although its relevance
to the pathology is not clear (de Silva et al., 1998). The CAG triplet
repeat in KCNN3 gene encoding a small conductance
calcium-activated K+ channel, hKCa3,
mapped to chromosome 1q21 has been reported to be associated with
schizophrenia (Chandy et al., 1998), although subsequent investigations
to confirm these findings have been met with mixed results (Austin et
al., 1999; Dror et al., 1999).
C. Hearing and Vestibular Diseases: Nonsyndromic Dominant Deafness and KCNQ4
Much progress has been made in the area of identifying genes
defective in hearing and balance disorders, with over 40 such genes
described (Holt and Corey, 1999). One of the genes reported to be the
locus for hereditary hearing impairment is another
K+ channel belonging to the KCNQ
channel superfamily, i.e., KCNQ4. The KCNQ4 gene,
isolated from a human retina library using KCNQ3 partial
cDNA as a probe, exhibits 38, 44, and 37% identity to KCNQ1, KCNQ2, and KCNQ3, respectively
(Kubisch et al., 1999). Reverse transcriptase-polymerase chain reaction
analysis revealed high expression of KCNQ4 in the vestibular
system and brain. In cochlea sections from mice at postnatal day P12,
sensory outer hair cells were strongly labeled with a KCNQ4
antisense probe but not in the inner hair cells and stria vascularis
where KCNQ1 expression was detected. Expression of
KCNQ4 in Xenopus oocytes generated a
voltage-dependent K+ current, similar to
KCNQ1, KCNQ2, and KCNQ3, except with slower activation. Unlike KCNQ1, KCNQ4 did not interact with minK.
However, coexpression of KCNQ3 with KCNQ4 yielded
currents resembling an M-channel, but with only weak inhibition (75%
inhibition at 200 µM) by linopirdine, unlike those observed with the
KCNQ2/KCNQ3 combination. The similarity of currents from
KCNQ3/KCNQ4 to M-channel indicated that
KCNQ3/KCNQ4 might potentially form another M-channel variant
in the nervous system (Kubisch et al., 1999).
Using fluorescence in situ hybridization to human chromosomes,
KCNQ4 was mapped to chromosome 1p34, a region also hosting DFNA2, a locus for autosomal dominant progressive hearing
loss (Kubisch et al., 1999). One 13-bp deletion mutation and four
missense mutations (G285S, G285C, W276S, and G321S) were identified
from families with autosomal dominant progressive hearing loss linked to the DFNA2 locus (Coucke et al., 1999; Kubisch et al.,
1999). The G285S and G285C mutations alter the first glycine residue in
the GYG signature sequence of K+ channel pore.
Mutations in these amino acids disrupt the selectivity filter and, in
most cases, abolish channel function. An identical change in amino
acids at the equivalent position has also been reported in the
KCNQ1 gene of a patient with the dominant LQT1 (Russell et
al., 1996). Functional analysis reveals that the mutant channel did not
produce current when the cRNA was injected into oocytes, whereas the
mutation exerted a dominant-negative effect when coexpressed with
wild-type KCNQ1. Whereas mutations in KCNQ1 affect endolymph secretion, the mechanism leading to
KCNQ4-related hearing loss appears to be in outer hair cells
(Kubisch et al., 1999), inner ear, and the central auditory pathway
(Kharkovets et al., 2000).
It must be pointed out that in addition to mutations in
KCNQ4, mutations in GJB3, which encodes the
connexin 31 component of gap junctions and was mapped to human
chromosome 1p33-p35, were identified from the DFNA2 family
with nonsyndromic autosomal dominant hearing loss (Xia et al., 1998).
Although at least two or three genes responsible for hearing impairment
are located close together on chromosome 1p34, KCNQ4
mutations may be a relatively frequent cause of autosomal dominant
hearing loss.
D. Renal Diseases: Bartter's Syndrome and Kir1.1
Several transporters and ion channels in the renal epithelium play
important roles in urine production, fluid balance, and electrolyte
metabolism. Genetic analysis reveals that dysfunction of an inward
rectifier K+ channel Kir1.1 is linked to
Bartter's syndrome, an autosomal recessive inherited renal tubular
disorder characterized by hypokalemia, metabolic alkalosis,
hyper-reninism and hyperaldosteronism. Patients have normal or low
blood pressure and renal salt loss despite increased plasma renin
activity and high serum aldosterone levels (Karolyi et al., 1998; Simon
and Lifton, 1998; Scheinman et al., 1999). At least three
phenotypically different renal tubulopathies have been identified:
antenatal Bartter's syndrome (hyperprostaglandin E syndrome), classic
Bartter's syndrome, and Gitelman's syndrome. Of these,
polyhydramnios, premature delivery, hypokalemic alkalosis, hypercalciuria, and dehydration at birth characterize the antenatal Bartter's syndrome (hypokalemic alkalosis with hypercalciuria). Children with the antenatal Bartter's syndrome present the typical pattern of impaired salt reabsorption in the thick ascending limb of
Henle's loop resulting in the marked ante- and postnatal salt wasting.
Genetic heterogeneity of antenatal Bartter's syndrome has been
demonstrated initially by identification of mutations in the SLC12A1 gene, encoding for the bumetanide-sensitive sodium
potassium 2 chloride cotransporter (NKCC2) leading to defective
reabsorption of sodium chloride in the thick ascending limb of Henle's
loop (Simon et al., 1996a; Vargas-Poussou et al., 1998). Subsequently, several mutations in KCNJ1, encoding the apical renal outer
medullary inward rectifying K+ channel (Kir1.1),
were identified in patients with antenatal Bartter's syndrome by
single strand conformation polymorphism analysis (Simon et al., 1996b;
Derst et al., 1997; Feldmann et al., 1998; Vollmer et al., 1998).
Functional studies revealed that mutant channels expressed none or
significantly reduced currents compared with the wild-type channel.
This impaired K+ flux and loss of tubular
K+ channel function probably prevents apical
membrane potassium recycling with secondary inhibition of Na-K-2Cl
cotransport in the thick ascending limb of Henle's loop (Derst et al.,
1997). The mechanisms underlying impaired Kir1.1 function involve
abnormalities in phosphorylation, proteolytic processing, and/or
protein trafficking (Schwalbe et al., 1998).
The signs and symptoms of Bartter's syndrome are usually a consequence
of hypokalemia. Maintaining normal serum K+
levels and limiting the degree of metabolic alkalosis are some of the
treatment approaches, and potassium supplements and potassium-sparing diuretics are frequently used (Gordon and Stokes, 1994).
E. Metabolic Diseases: Familial Persistent Hyperinsulinemic Hypoglycemia of Infancy and Sulfonylurea Receptor 1
Various types of ion channels are involved in the regulation of
electrical activity in the pancreatic -cell. Of these, the ATP-sensitive K+ (KATP)
channel plays a critical role in directly linking cellular metabolism
to the electrical activity. Opening the ATP-sensitive K+ channels leads to membrane hyperpolarization
and consequently suppression of insulin secretion. Recent genetic
analysis has revealed mutations in the ATP-sensitive
K+ channel subunits that may contribute to
inappropriate and excessive secretion of insulin.
PHHI is an autosomal recessive disorder characterized by increased
irregularity in insulin secretion leading to hypoglycemia, coma, and
severe brain damage in children. Both sporadic and familial variants of
PHHI are recognized; familial forms are common in communities with high
rates of consanguinity where the incidence may be as high as 1 in 2500 live births and is the most common cause of hypoglycemia in
newborns (Aynsley-Green and Hawdon, 1997). Recent genetic linkage
analysis has identified mutations in the KATP
channel complex that regulates insulin secretion from pancreatic -cells. The KATP channels predominantly
determine the resting potential of -cell and couple cellular
metabolism to electrical activity (Ashcroft and Rorsman, 1989; Dukes
and Philipson, 1996). When plasma glucose is elevated, increases in
intracellular ATP/ADP ratio lead to closure of
KATP channels and membrane depolarization that,
in turn, lead to the activation of voltage-dependent
Ca2+ channel, rise in intracellular
Ca2+, and insulin secretion.
The -cell KATP channel, like other
KATP channels described in neurons, cardiac,
smooth, and skeletal muscle, are inhibited by intracellular ATP, and
recent molecular cloning has shown that the channel is an octamer
composed of four subunits of the sulfonylurea receptor SUR1 coupled to
four subunits of the inward rectifier Kir6.2 (Inagaki et al., 1995a,
1997; Clement et al., 1997; Shyng and Nichols, 1997). Over 28 naturally
occurring mutations in SUR1 (Thomas et al., 1995b; Dunne et al., 1997;
Verkarre et al., 1998) and two different mutations in Kir6.2 subunits
have been identified in families with PHHI (Thomas et al., 1996a;
Nestorowicz et al., 1997; Meissner et al., 1999). No
KATP channel activity was observed in -cells
isolated from a homozygous patient or after coexpression of recombinant
Kir6.2 and mutant SUR1 (V187D) (Otonkoski et al., 1999). Detailed
functional analysis in COS cells by cotransfection of Kir6.2 with
various single mutations of SUR1 identified in the PHHI family
suggested this lack of KATP channel activity or reduction of KATP channel sensitivity to MgADP
(Shyng et al., 1998). In fact, patients with mutations in SUR1 either
failed to respond to diazoxide or showed diminished sensitivity to
treatment (Thornton et al., 1998).
The role of KATP channels in -cell function
has been evaluated in transgenic mice carrying a dominant-negative form
of Kir6.2 (G132S) generated by substituting the glycine lining the pore with serine (Miki et al., 1997). These mice develop hypoglycemia with
hyperinsulinemia in neonates and hyperglycemia with hypoinsulinemia and
decreased -cell population in adults. KATP
channel function was found to be impaired in the -cell of transgenic
mice with hyperglycemia. These results imply that the
KATP channel complex might play a significant
role in -cell survival and regulation in insulin secretion,
suggesting that modulation of Kir6.2 may offer additional opportunities
in treatment of diabetes and related conditions of abnormal glucose
regulation. More recently, it has been shown that the SUR1 knockout
mice, unlike the Kir6.2 counterpart, are not insulin-hypersensitive,
although their -cells lacks KATP channels and
show spontaneous Ca2+ transients similar to those
seen in PHHI patients. SUR1 knockout mice were normoglycemic until
stressed, unlike in PHHI patients whose glucose levels are persistently
low suggestive of a role for KATP-independent
pathways that regulate insulin secretion, at least in mice (Seghers et
al., 2000).
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III. Disease- and Drug-Induced Regulation of K+ Channels |
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A. Cardiac Failure and Hypertrophy
K+ channels are targets for the actions of
transmitters, hormones, or drugs that modulate cardiac functions.
Changes in the densities and/or properties of these
K+ channels that occur during the normal
development or as a result of damage or disease can have profound
physiological consequences (Matsubara et al., 1993; Xu et al., 1996;
Yao et al., 1999). Cardiac failure, a pathophysiologic condition with
numerous etiologies including myocardial infarction, hypertension, and
myocarditis (Wilson, 1997) is characterized by action potential
prolongation and, accordingly, altered expression of a variety of
depolarizing and hyperpolarizing membrane currents. In an attempt to
compensate for the reduction in cardiac function in cardiac failure,
the sympathetic nervous system, the renin-angiotensin-aldosterone systems, and other neurohumoral mechanisms are activated. Adaptive changes at the level of the cardiac myocyte include cellular
hypertrophy and altered gene expression. Electrical remodeling in
cardiac myocytes leading to action potential prolongation is a common finding in human heart failure and in animal models of cardiac hypertrophy. Changes in a wide range of plasma membrane receptors and
intracellular signals such as increased intracellular calcium, cAMP,
inositol phosphates, and diacylglycerol concentrations are associated
with cardiac hypertrophy and failure (Morgan and Baker, 1991;
Gopalakrishnan and Triggle, 1990; Wickenden et al., 1998).
A reduction in the current density of the transient outward current
(ITO) is the most consistent ionic current change
in cardiac hypertrophy and failure (Nabauer and Kaab, 1998; Wickenden
et al., 1998; Pinto and Boyden, 1999; Tomaselli and Marban, 1999). This
outward repolarizing K+ current activates and
inactivates rapidly with an inactivation constant of ~60 ms (Dixon et
al., 1996; Kong et al., 1998). The down-regulation of this current has
profound effects on phase 1 and the level of plateau of the action
potential, and it also alters currents that are subsequently active
along the cardiac action potential. The Kv4.3-containing channel is
thought to underlie the bulk of ITO found in the
mammalian heart, although Kv1.4 or Kv4.2 channels might represent
another fraction of ITO with distinct kinetics in
different regions of the heart (Dixon et al., 1996; Kong et al., 1998).
By ribonuclease protection assays and whole-cell electrophysiological
recording, Kaab et al. (1998) found that the level of Kv4.3 mRNA
decreased by 30% in human failing hearts compared with nonfailing
controls. This observation correlated with the reduction in peak
ITO density measured in ventricular myocytes
isolated from adjacent regions of the heart.
It has been known that action potential durations vary across the
myocardial wall and in different regions of the mammalian heart
(Litovsky and Antzelevitch, 1989; Fedida and Giles, 1991; Lukas and
Antzelevitch, 1993; Di Diego et al., 1996). The density of
ITO also varies regionally and transmurally in
the heart (Wettwer et al., 1994; Nabauer et al., 1996).
Electrophysiological recording from myocytes isolated from patients
with aortic stenosis and compensated left ventricular hypertrophy
indicates that macroscopic ITO was absent in
superficial subendocardial cells, whereas ITO current density was not significantly altered in the deeper layers (Bailly et al., 1997). A region-dependent alteration in the density of
ITO current was also observed in the
catecholamine-induced hypertrophy in animals (Bryant et al., 1999). It
is possible that this region-dependent suppression of
ITO current might, in part, underlie the regional
heterogeneity in action potential prolongation in cardiac hypertrophy
and may predispose to ventricular arrhythmias, a cause of sudden death
in patients with cardiac failure.
As discussed later, an approach to the treatment of heart failure would
be to normalize K+ channel gene expression by
gene transfer or pharmacologic modulation. Recent studies have shown
that thyroid hormone treatment can increase Kv4.2 or Kv4.3 expression
at the transcriptional level and enhance the recovery rate from the
inactivation of ITO in rat ventricular myocytes
(Shimoni et al., 1997; Wickenden et al., 1997). Accordingly, agents
with thyroid hormone-like properties might be useful in the treatment
of heart failure.
B. Atrial Fibrillation
Atrial fibrillation, the most common arrhythmia in man, is
characterized by a marked shortening of the action potential duration, effective refractory period of atria, and a decreased rate of atrial
repolarization resulting in increased dispersion of refractoriness as
well as changes in atrial conduction velocity (Zipes, 1997; Nattel,
1999). The development of atrial fibrillation can be triggered by
rapidly discharging atrial foci (mainly from pulmonary veins) or
degeneration of atrial flutter or atrial tachycardia into fibrillation (Chen et al., 1999b; Scheinman, 2000). Risk factors for atrial fibrillation include cardiac diseases such as congestive heart failure,
valvular heart disease, and myocardial infarction (Ryder and Benjamin,
1999).
It has been shown that sustained atrial tachycardia causes changes in
electrophysiological function to promote the occurrence and maintenance
of atrial fibrillation, a process referred to as atrial
electrophysiological remodeling (Morillo et al., 1995; Wijffels et al.,
1995). Recent studies have revealed that changes in ion channel
functions play important roles in atrial electrophysiological remodeling caused by atrial fibrillation. In the canine atrial fibrillation model induced by chronic atrial tachycardia (rapid pacing), isolated atrial myocytes showed significant reductions in
L-type Ca2+ current and ITO
densities, without changes in Kir2.1, hERG, KCNQ1-minK, Ca2+-dependent Cl
current, or T-type Ca2+ currents (Yue et al.,
1997). Consistent with this observation, reductions in mRNA levels for
Kv4.3, the 1-subunit of L-type Ca2+ channels, and the -subunit of cardiac
Na+ channels were noted with no changes in mRNA
levels for delayed rectifier K+ channel Kir2.1 or
the Na+/Ca2+ exchanger.
Western blot analysis further confirmed a reduction in protein
expression of Kv4.3 and Na+ channels, whereas
that of the Na+/Ca2+
exchanger was unchanged (Yue et al., 1999; Li et al., 2000). More
importantly and consistent with data from the canine atrial fibrillation model, significant reductions in ITO
(encoded by Kv4.3) and ultrarapid delayed rectifier
(IKur) (encoded by Kv1.5) as well as L-type
Ca2+ current densities were observed in atrial
myocytes isolated from patients in chronic atrial fibrillation.
Furthermore, quantitative Western blot analysis revealed that the
expression of Kv1.5 protein was reduced by >50% in both the left and
the right atrial appendages of atrial fibrillation (Van Wagoner et al.,
1997, 1999). Although abnormalities of K+
channels may be fundamentally implicated in atrial fibrillation, other
factors such as structural changes (Li et al., 1999) or heterogeneous
alterations in atrial sympathetic innervation (Jayachandran et al.,
2000) may also play critical roles in other forms of atrial fibrillation.
C. Drug-Induced Long-QT Syndromes
Drug-induced precipitation of polymorphic ventricular dysrhythmia,
the torsade de pointes, in susceptible individuals by certain H1 antagonists such as terfenadine has now been
linked to the prolongation of the QT interval consequent to inhibition
of the IKr channels encoded by the hERG gene (reviewed in Delpón
et al., 1999; Taglialatela et al., 2000). These drugs have been shown to block hERG channels in a concentration range similar to that found
in the plasma of subjects showing proarrhythmic effects. Similar
interactions have been reported for antipsychotics such as sertindole
(Rampe et al., 1998), tricyclic antidepressants, and certain
antibiotics and anti-emetic agents. Inhibition of another cardiac
delayed rectifier, Kv1.5, by H1 receptor
antagonists such as loratadine (Lacerda et al., 1997) and rupatadine
(Caballero et al., 1999) has also been suggested to contribute to
drug-induced cardiac arrhythmias.
D. Apoptosis and Oncogenesis
K+ channel activities play important roles
in signaling pathways leading to proliferation,
differentiation, and cell fusion. Increases in K+
channel activity and enhanced K+ efflux are
thought to sustain membrane hyperpolarization necessary to facilitate
Ca2+ entry (Santella, 1998), although additional
pathways, such as control of cellular volume by
K+ channels, might also be involved in cell
proliferation (Rouzaire-Dubois and Dubois, 1998; Vaur et al., 1998). A
number of studies have suggested membrane hyperpolarization as an
essential requirement for cell proliferation. For example, an increase
in expression levels of a Ca2+-dependent
K+ channel with strong inward rectification was
observed during the G1 phase of HeLa cells, which
progressively declined to a minimum in the S phase and then increased
in the M phase (Takahashi et al., 1993). Inhibition of
K+ channels by pharmacological agents has been
found to inhibit cell proliferation in normal human lymphocytes
(Amigorena et al., 1990; Lin et al., 1993; Rader et al., 1996; Jensen
et al., 1999), human melanoma cells (Nilius and Wohlrab, 1992;
Lepple-Wienhues et al., 1996), small lung cancer cells (Pancrazio et
al., 1993), breast cancer cells (Woodfork et al., 1995), and prostatic
cells (Skryma et al., 1997). Changes in expression of an inward
rectifying K+ channel and a noninactivating
delayed rectifier K+ channel are associated with
the time course of membrane fusion of myoblast to form multinucleated
skeletal muscle fibers (Shin et al., 1997; Occhiodoro et al., 1998).
Recently, a gene encoding the human EAG K+
channel was cloned from myoblasts, localized to chromosome 1q32-41 and
shown to be responsible, in part, for changes in membrane hyperpolarization during the myoblast fusion (Occhiodoro et al., 1998).
1. Apoptosis.
Apoptosis, or programmed cell death, is a
fundamental biological process involved in many physiological and
pathological phenomena. This process is predominantly catabolic in
nature where cellular macromolecules are broken down by distinct
enzymes to be later recycled in healthy cells. Activities of enzymes,
nucleases, and caspases that propagate and amplify death signals are
K+-dependent (Bortner et al., 1997; Hughes and
Cidlowski, 1999). Recent studies have shown that enhancement of
K+ current is directly involved in apoptosis (Yu
et al., 1997, 1999) and oncogenesis (Pardo et al., 1999). In mouse
neocortical neurons, a delayed rectifier and tetraethylammonium
(TEA)-sensitive K+ current responsible for
neuronal apoptosis was enhanced by serum deprivation or staurosporine.
Inhibition of outward K+ currents with TEA or
elevated extracellular K+, but not with blockers
of Ca2+, Cl, or other
K+ channels, reduced apoptosis. Exposure to the
K+ ionophore valinomycin or the
KATP channel opener cromakalim induced apoptosis
(Yu et al., 1997). Thus, enhanced K+ efflux
through increase in expression of a specific TEA-sensitive and delayed
rectifier K+ channel may mediate certain forms of
neuronal apoptosis in disease states. Thymocyte apoptosis induced by
dexamethasone, etoposide, 
-irradiation, or ceramide has also been
shown to be prevented by the K+ channel blocker
tetrapentylammonium (Dallaporta et al., 1999).
). In Drosophila,
reaper, grim, or hid gene expression
triggers apoptosis in a caspase-dependent manner. The peptides encoded
by these genes share a common feature in that their N termini are
similar to those of the Shaker
K+ channel that block channel and lead to fast
inactivation. Mutations that reduce the apoptotic activity of
reaper also reduced the peptide's ability to induce channel
inactivation. Thus, blocking a Shaker
K+ channel by peptides encoded by
reaper, grim, or hid gene was suggested to be involved in apoptosis (Avdonin et al., 1998).
2. Oncogenesis.
Modulation of K+
channels is involved in Ras/Raf signal transduction in
oncogenic transformation (Collin et al., 1990; Yatani et al., 1991;
Huang and Rane, 1994; Decker et al., 1998). Recent studies have shown a
high level of an intermediate conductance Ca2+-activated K+ current
(IKCa) in Ras-transformed fibroblasts but
not in the untransformed counterparts (Rane, 1991). High levels of
expression of IKCa have also been observed in rat
prostate cancer cell lines, AT2.1 and MatLyLyu, suggesting
hyperactivity of the Ras/MAPK pathway in prostatic cancer and that
IKCa plays important roles in regulating cell
growth (Rane, 2000). Similarly, the hERG was shown to be sequentially
expressed during neuronal development and to participate in the
regulation of membrane potential in mammalian neuroblastoma cells
(Arcangeli et al., 1995, 1997). The hERG, and the related ether-a-go-go K+ channels are
expressed in a variety of tumor cell lines (Bianchi et al., 1998; Pardo
et al., 1999), the inhibition of which causes a significant reduction
of cell proliferation. Moreover, the expression of rEAG favors tumor
progression when transfected cells are injected into immunosuppressed
mice, and overexpression of rEAG K+ channels in
Chinese hamster ovary or NIH 3T3 cells induces significant features
characteristic of malignant transformation (Pardo et al., 1999). Taken
together, these studies suggest that these K+
channels play crucial roles in oncogenesis.
E. Alzheimer's Disease
Alzheimer's disease is the most prevalent cause of progressive
declining cognitive function, loss of memory, and late stage decreasing
physical deterioration in the elderly. It is characterized pathologically by the presence of intracellular neurofibrillary tangles
and extracellular neuritic plaques consisting of deposits of the
-amyloid (A), a 39- to 43-amino acid peptide proteolytically derived from -amyloid protein precursor (-APP). In Alzheimer's disease, significant neuronal cell death is found in the temporal and
parietal cortex, hippocampus, amygdala, and basal forebrain cholinergic system. Several mechanisms have been linked to progressive neurodegenerative disorder, such as alterations in amyloid precursor protein metabolism, cholinergic transmission, calcium homeostasis, oxidative metabolism, and protein kinase C transduction systems (Mattson et al., 1993; Hensley et al., 1994; Ito et al., 1994; Yankner,
1996; Yu et al., 1998). As discussed below, dysfunction of
K+ channels in both central nervous systems and
peripheral tissues has been reported. It is plausible, however, that
any association of K+ channel defects with the
pathophysiology of Alzheimer's disease may be indirect or secondary in
nature consequent to generalized degeneration associated with the disease.
1. -Amyloid.
K+ channel
dysfunction in Alzheimer's disease was initially suggested by
radioligand binding studies using apamin, the bee venom octadecapeptide
that blocks small conductance Ca2+-activated
K+ channels responsible for
afterhyperpolarization of neurons (Ikeda et al., 1991). In hippocampus,
a reduction of 125I-apamin binding sites in the
subiculum and CA1 regions was found in patients with Alzheimer's
disease. The reduction of 125I-apamin binding
sites in the subiculum correlated with cell density but not neuritic
plaque density, indicating discrete loss of small conductance of
Ca2+-activated K+ channels
within the hippocampal formation. In hippocampal neurons from neonatal
rats, A was shown to inhibit voltage-dependent fast-inactivating
K+ currents (Good et al., 1996). This inhibition
results in abnormally large increases in intracellular
Ca2+ levels upon depolarization of the neuron
leading to neurotoxicity (Good and Murphy, 1996).
-induced abnormal K+
to the neuronal cell death was revealed by in vitro studies using a
cholinergic septal cell line, SN56 (Colom et al., 1998). These cells
exhibited a tetraethylammonium-sensitive outward
K+ current with delayed rectifier
characteristics. Addition of A increased K+
current density some 44 to 66% and decreased cell viability by 25 to
39%. TEA (10 to 20 mM) or K+ depolarization
inhibited outward currents, widened action potentials, elevated
[Ca2+]i, and inhibited
more than 68% of the A-induced toxicity. These data suggest that a
K+ channel with delayed rectifier characteristics
may play an important role in A-mediated toxicity in this septal
cholinergic cell line (Colom et al., 1998).
In peripheral tissues, K+ channel dysfunction was
initially identified in fibroblasts from patients with Alzheimer's
disease where a 113-pS TEA-sensitive K+ channel
was absent compared with normal human fibroblasts (Etcheberrigaray et
al., 1993). This defect was mimicked in normal fibroblasts by the
addition of -amyloid protein (Etcheberrigaray et al., 1994). TEA
depolarized and elevated intracellular Ca2+
levels in young and aged control fibroblasts but not in fibroblasts from Alzheimer's disease patients, supporting the dysfunction of
TEA-sensitive K+ channels in the disease.
Rb+ flux through apamin and
charybdotoxin-sensitive Ca2+-activated
K+ channels was selectively impaired in fresh,
noncultured platelets from patients with Alzheimer's-type dementia,
although the -dendrotoxin-sensitive voltage-dependent
K+ channel was not affected compared with
nondemented controls (de Silva et al., 1998). -Amyloid protein also
enhanced phytohemagglutinin-induced Ca2+ rise in
T-lymphocytes, consistent with the hypothesis that enhanced calcium
responses serve as a general feature of -amyloid neurotoxicity (Eckert et al., 1993). However, patch-clamp analysis indicated that
T-lymphocyte K+ channels are not functionally
deficient in Alzheimer's disease, and that -amyloid protein does
not mediate an alteration of their currents (Cohen et al., 1996),
suggesting A might induce toxicity through alternative pathways.
2. -Amyloid Protein Precursor.
-APP, the source of the
fibrillogenic A, is a membrane-spanning and multifunctional protein
that is widely expressed in the nervous system. -APP is axonally
transported and accumulates in presynaptic terminals and growth cones.
A secreted form of -APP (sAPP) is released from neurons in response
to electrical activity and plays important roles in learning, memory,
and cell survival (Roch et al., 1994; Mattson, 1997; Meziane et al.,
1998; Dodart et al., 2000). In addition to A-induced neurotoxicity via potential modulation of K+ channel function,
a study revealed that sAPP can suppress action potential and
hyperpolarize hippocampal neurons by activating large conductance
Ca2+-activated K+
channels leading to suppression of intracellular
Ca2+ concentration (Furukawa et al., 1996). These
results suggest that the effects of -APP on synaptogenesis and
synaptic plasticity might, in part, mediate through activation of
Ca2+-activated K+ channels
and that the abnormalities in -APP processing or sAPP might
contribute to the neurodegenerative process in Alzheimer's disease.
3. Presenilins.
The presenilins are proteins that contain
multiple transmembrane domains and localize primarily to the
endoplasmic reticulum and Golgi apparatus. Although the precise
functions of presenilins are not totally understood, presenilins are
involved in the proteolytic processing of -amyloid precursor
proteins and play important roles in the notch signaling during
embryonic development and/or cellular differentiation (Kim and Tanzi,
1997; Chan and Jan 1999; Haass and De Strooper, 1999; Czech et al.,
2000). Genetic linkage analysis showed that mutations in presenilin 1 (PS-1, mapped on chromosome 14) and presenilin 2 genes (PS-2 on
chromosome 1) yielding abnormal release of amyloidogenic peptide from
amyloid precursor protein have been linked to the autosomal dominant
early onset of familial Alzheimer's disease (Clark et al., 1995;
Rogaev et al., 1995; Schellenberg, 1995; Sherrington et al., 1995).
Based on the multiple membrane-spanning topology, it was proposed that presenilins might function as, or as part of, a channel, transporter, or pore (Li and Greenwald, 1996). Using in vitro expression in HEK-293
cells, a recent study has revealed that expression of wild-type PS-1 or
PS-2 increases outward K+ current densities. In
HEK-293 cells transiently transfected with PS-1 (S290C) or PS-1
(G209V), two missense mutations associated with early onset
Alzheimer's disease, mean outward K+ current
densities are also shown to be increased in HEK-293 cells expressing
the S290C mutant but not with the G209V mutant. Expression of wild-type
PS-1 in neonatal rat ventricular myocytes also results in increased
outward K+ currents, whereas no detectable
effects on membrane currents were seen in COS-7 cells transfected with
PS-1. These results suggest that the presenilins do not actually form
K+ channels, but rather that these proteins
up-regulate functional K+ channel expression
(Malin et al., 1998). Thus, presenilins could regulate neuronal
K+ channel expression, and mutations in PS-1 or
PS-2 can, in part, result in profound changes in neuronal excitability,
which may contribute to the cognitive decline commonly associated with
Alzheimer's disease to some extent.
F. Neuromuscular Disorders
Mutations in a variety of ion channels, including
Na+, Ca2+, and
Cl channels, have been found to underlie
various forms of human neuromuscular disorders. The defects of ion
channels lead to the aberrant excitability of muscle fibers that gives
rise to periodic paralysis or myotonia (for reviews see Cannon, 1996;
Engel et al., 1998). In addition to inherited genetic diseases, diverse neuromuscular disorders are attributed to antibody-mediated
autoimmunity where the extracellular domains of receptors or ion
channels are the primary targets of autoantibodies. For example,
myasthenia gravis is caused by autoantibodies to nicotinic
acetylcholine receptors at the neuromuscular junction, which cause
weakness of the skeletal muscle (Richman and Agius, 1994). The
autoantibodies that interfere with neurotransmitter release by binding
to presynaptic voltage-dependent Ca2+ channels
underlie the Lambert-Eaton myasthenic syndrome, which is often found in
patients with small cell lung cancer (Kim and Neher, 1988; Pelucchi et
al., 1993). In acquired neuromyotonia (Isaacs' syndrome), where
hyperexcitability of peripheral motor nerves leads to muscle twitching
during rest, cramps during muscle contraction, impaired muscle
relaxation, and muscle weakness, autobodies directed against
4-aminopyridine or -dendrotoxin-sensitive K+ channels in motor and sensory neurons were
detected (Shillito et al., 1995; Hart et al., 1997). These antibodies
mainly suppress voltage-gated K+ channels (Kv1.1
and Kv1.6) with no change in gating kinetics and lead to peripheral
nerve hyperexcitability (Nagado et al., 1999). In humans with
hypokalemic periodic paralysis caused by mutations of the
1,4-dihydropyridine receptor of the voltage-gated calcium channel,
diminished skeletal muscle KATP channel activity has also been reported (Tricarico et al., 1999).
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IV. Pharmacological Considerations |
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As discussed in the preceding sections, several genetically linked
and acquired diseases involve alterations in the function of
K+ channels. Genetic linkage studies have been
pivotal in elucidating the role of many K+
channels in pathophysiologic and physiologic conditions. More importantly, these findings provide a basis to develop
appropriate therapy for various diseases. Continuing pharmaceutical
interest revolves around the discovery and development of selective
organic modulators of various classes of K+
channels (Colatsky, 1998; Curran, 1998; Kaczorowski and Garcia, 1999). Enthusiasm in the K+ channel arena
is driven by the realization that class III antiarrhythmic agents and
antidiabetic sulfonylureas act as antagonists at specific K+ channel classes and that a variety of
K+ channel inhibitors and openers offer
significant therapeutic opportunities in areas ranging from cardiac,
vascular, and nonvascular muscle, neuronal, immune, and secretory
systems to modulation of hair follicle growth (Table
3). Gene delivery and selective targeting
of channel proteins by antisense oligonucleotides represent emerging
approaches. With advances in molecular biology and antisense technology, therapeutics based on gene delivery, with precise control
of the level and distribution of ion channel expression into mammalian
neuronal, cardiac, hair cells, and other cell types, are currently
being investigated (Holt et al., 1999; Johns et al., 1999; Hoppe et
al., 2000).
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However, key hurdles in targeting K+ channels remain to be resolved. Given the diversity of K+ channel subunits and the potential to vary the constituents to form heteromeric channel complexes to alter expression, cellular targeting, and biophysical and pharmacological properties in native cell types, it is difficult to know the precise composition of channel complexes in vivo. The latter, together with information on tissue-specific localization and the availability of high-throughput in vitro assays predictive of in vivo drug activity and selectivity, is seldom available. This is an important issue, which has not been addressed to the full extent, as efforts are launched to design openers and/or blockers of various classes of potassium channel modulators. Nevertheless, over the past decade or so, intense medicinal chemistry efforts have focused on the synthesis and development of modulators of various voltage-gated K+ channels, calcium-activated K+ channels and ATP-sensitive K+ channels (Figs. 3 and 4; Tables 3 and 4).
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A. Voltage-Gated K+ Channels
1. Kv1.3 Channels.
The Kv1.3 channels, members of the
voltage-gated K+ channel family expressed
predominantly in human lymphocytes, have been widely exploited as
pharmacological targets for immunosuppressive therapy. Selective
blockers of these channels depolarize membrane to attenuate calcium
influx and inhibition of T cell activation in vitro and
immunosuppression in vivo (Cahalan and Chandy, 1997 2. Cardiac Delayed Rectifier K+ Channels.
The
goal of developing a class III antiarrhythmic agent effective against
ventricular arrhythmias while reducing hemodynamic liabilities remains
to be realized, but should now be accelerated with the understanding of
the molecular components of cardiac delayed rectifiers, i.e., IKs
(KvLQT1-minK), IKr (hERG), and IKur (Kv1.5)
channels. The currently available class III drugs amiodarone (Kodama et
al., 1999). Many peptides
isolated from scorpion venoms and sea anemone potently block Kv1.3
channels and inhibit T-lymphocyte activation. Inhibition of these
channels by margatoxin was initially shown to prevent T cell activation
and attenuate immune responses in vivo (Koo et al., 1997). Several
nonpeptide analogs, such as dihydroquinolines, WIN 17317-3 (Hill et
al., 1995) and CP-339,818 (Nguyen et al., 1996), piperidines, UK
78,282, (Hanson et al., 1999), and certain alkoxypsoralenes (Wulff et
al., 1998) have been shown to block Kv1.3 channels and/or inhibit human
T cell activation in vitro. Despite this in vitro evidence, there has
been little in vivo demonstration until recently that blockade of Kv1.3
will attenuate immune responses, the latter possibly due to species
differences, since in many rodent peripheral T cells these channels do
not appear to set membrane potential. However, these channels appear to
be present on peripheral T cells of minipigs, and Koo et al. (1999)
have shown that the nortriterpene, correolide, and its analogs
extracted from the tree Spachea correae can block Kv1.3 channels and inhibit delayed-type hypersensitivity response to tuberculin in minipigs (Koo et al., 1999). The Kv1.3 modulators described thus far could serve as tools for the further design of
immunosuppressive agents because many of these compounds lack desirable
potencies, selectivity, and pharmacokinetic profile. For example, a
study with radiolabeled WIN 17317-3 has shown that this
compound is also a potent blocker of brain type IIa sodium channels
(Wanner et al., 1999).) and sotalol (Anderson and Prystowsky, 1999) possess
properties beyond the realm of a pure class III effect (Roden, 1993;
Nair and Grant, 1997; Sager, 1999).
) and antisense oligonucleotide studies in cultured adult human atrial myocytes (Feng et al., 1997). Association with Kv1.2 subunits can also alter functional properties of Kv1.5 channels (Majumder et al., 1995).
Selective blockers of Kv1.5 channels could be potentially beneficial
for the treatment of cardiac arrhythmias because such agents could
retard repolarization and prolong refractoriness selectively in cardiac
myocytes (Nattel et al., 1999).
Gene transfer of delayed rectifier K+
channels represents an emerging strategy for the control of arrhythmias
triggered by altered cardiac repolarization. Myocytes isolated from
adult rabbit ventricular myocytes in culture which demonstrate longer
action potentials and frequent early-after depolarizations when
maintained in culture, were reversed following adenoviral gene
transfer of the hERG gene (Nuss et al., 1999). Infection with a
recombinant adenovirus containing the hERG gene selectively enhanced
the E-4031-sensitive currents without affecting the density of
transient outward currents, suppressed early-after
depolarizations, and lengthened the refractory period. Action
potentials from failing dog hearts were also reversed after exposure to
an adenovirus that overexpresses Shaker
K+ channels (Nuss et al., 1996). Further
refinement of techniques to effectively control the level and to ensure
homogenous distribution of transgene expression at the target organ is
likely to be forthcoming (Hoppe et al., 2000).
As noted previously, it has also become increasingly important to avoid
interactions of many noncardiovascular medicinal products with cardiac
ion channels (reviewed in Pourrias et al., 1999). Certain
H1 antagonists, such as astemizole and
terfenadine, and the prokinetic agent cisapride are capable of
prolonging the QT interval and inducing torsade de pointes in
susceptible individuals through inhibition of IKr channels encoded by
hERG gene. Similar interactions have also been reported for certain
antipsychotics such as sertindole (Rampe et al., 1998), tricyclic
antidepressants, and some antibiotics. Inhibition of Kv1.5 channels by
H1 receptor antagonists such as loratadine
(Lacerda et al., 1997) and rupatadine, a dual antagonist of
H1, and platelet-activating factor receptors (Caballero et al., 1999) has also been suggested to contribute to
drug-induced cardiac arrhythmias. Prolongation of cardiac
repolarization reported with the 5HT3 receptor antagonist,
ondanestron, has been attributed to inhibition, albeit only 30%, of
hERG channels (Kuryshev et al., 2000a). Needless to note, these
pharmacologic misfortunes underscore the importance of evaluation of
potential inhibition of these cardiac channels during the early
developmental phase of novel compounds because drugs with minimal or no
potential to block hERG or Kv1.5 channels are likely to possess cardiac safety advantages.
3. KCNQ2/KCNQ3 Channels.
Unlike KCNQ1,
KCNQ2 and KCNQ3 are present exclusively in the nervous
system and coassemble to form heteromultimers that underlie the
M-current (Wang et al., 1998) critical to neuronal excitability in the
nervous system (Brown, 1988). The potential for targeting the
KCNQ2/KCNQ3 combination as a drug target is underscored by the findings
that compounds such as linopirdine [DuP 996, 3,3-bis(4-pyridinylmethyl)-1-phenylindolin-2-one] and XE991 developed
as cognition enhancers are blockers of cloned KCNQ channels (Lamas et
al., 1997; Eid and Rose, 1999). Linopirdine, a putative cognition
enhancing drug, increases acetylcholine release in rat brain tissue and
improves performance in animal models of learning and memory (Schnee
and Brown, 1998). Although clinical data with linopirdine were largely
inconclusive, analogs such as XE991 and DMP543 with superior
pharmacological and pharmacodynamic properties have entered development
as orally active acetylcholine-releasing agents with potential in
Alzheimer's disease (Zaczek et al., 1998). The KCNQ1/minK complex was
14- to 18-fold less sensitive to XE991 blockade compared with either
KCNQ1 alone or neuronal KCNQ2/KCNQ3 combination, revealing a much
desired degree of selectivity for this compound for neurotransmitter
release over cardiac function (Wang et al., 2000). More recently,
retigabine (D-23129), reportedly in phase II clinical studies for the
treatment of epilepsy, has been shown to activate KCNQ2/KCNQ3 channels
expressed in Chinese hamster ovary cells in a partially
linopirdine-sensitive manner, suggesting that M-channel activation may
be a novel mode of action for anticonvulsant drugs (Main et al., 2000;
Rundfeldt and Netzer, 2000).
B. Calcium-Activated K+ Channels
The recent molecular cloning of various calcium-activated
K+ channels has renewed enthusiasm for the
development of modulators for these channels. These channels,
critically dependent on intracellular calcium for channel opening, were
initially differentiated largely on the basis of biophysical
(conductance, voltage dependence) and differential toxin sensitivity
into large, intermediate, and small conductance
Ca2+-activated K+ channels.
Distinct genes are now known to encode the three subfamilies of
calcium-activated K+ channels, i.e., large
conductance (BKCa) (-subunit and its splice variants), small conductance (Sk1, Sk2, and Sk3), and intermediate conductance channels (reviewed in Vergara et al., 1998; Castle, 1999;
Wallner et al., 1999b). The search for organic modulators of various
Ca2+-activated K+ channels
with the potential to be developed as therapeutic agents has been
actively explored by functional screening using many of the recombinant
channels (Kaczorowski and Garcia, 1999).
1. Large Conductance Channels.
The
BKCa -subunit cloned from either
Drosophila (Slo) or mammalian (mSlo,
hSlo), in combination with different -subunits, 1, and more recently 2 to 4, now extends diversity of
BKCa channels. Initial modulators reported
include activators such as glycosylated triterpenes
(dehydrosoyasaponin-I) and several indole diterpene blockers, such as
paxilline, verruculogen, penitrem A, and aflatrem (Kaczorowski et al.,
1996). Activators of BKCa channels include the
benzimidazolones, such as NS-1619 and NS-004. However, these compounds
are, in general, not very potent or highly selective. More recently,
openers of BKCa channels have been developed as
neuroprotective agents. One such compound, BMS-204352, is
in advanced trials as a stroke neuroprotectant (Hewawasam et al., 2000). NS-8, a pyrrole derivative shown to activate
BKCa channels, is under investigation for the
treatment of urinary incontinence (Tanaka et al., 1998). The potential
for BKCa modulators in the treatment of erectile
dysfunction has been underscored by recent studies with the
BKCa channel -subunit (Christ et al., 1998). Intracavernous injection of hSlo DNA was capable of altering
nerve-stimulated penile erection and was associated with a significant
elevation in intracavernous pressure at least until two months
postinjection. Interestingly, the expression of the hSlo
message was highest in the corpus cavernosum tissue and minimal in
other tissues examined, raising the possibility that such localized
delivery of K+ channel genes may provide another
avenue for achieving end organ selectivity.
2. Intermediate Conductance Channels.
Blockers of the
IKCa channel have long been proposed for therapy
in sickle cell anemia, diarrhea, and rheumatoid arthritis; clotrimazole, an inhibitor of the IKCa channel in
red blood cells, has been used for this purpose (Brugnara et al., 1995;
de Franceschi et al., 1996). However, the inhibition of cytochrome P450
enzyme by clotrimazole limits its therapeutic applications. Recently, a
more selective and potent inhibitor of IKCa channel,
TRAM-34 (1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole),
with no effect on cytochrome P450 activity, has been reported (Fig. 3)
(Wulff et al., 2000). Although not highly specific,
1-ethyl-2-benzimidazolinone (1-EBIO) and the clinically used
benzoxazoles, chlorzoxazone and zoxazolamine, are described as
pharmacological activators of the IKCa channel
(Syme et al., 2000). Inhibitors of IKCa may also be useful as immunosuppressive agents because these channels are up-regulated following antigenic or mitogenic stimulation (Khanna et
al., 1999). IKCa channels may also serve as an
effector for mitogenic Ras/MAPK signaling in fibroblasts and other cell
types, including prostate cancer cells (Rane, 2000). Openers of
IKCa channels may be therapeutically beneficial
in cystic fibrosis and peripheral vascular disease, as well (Edwards,
1998).
3. Small Conductance Channels.
The
SKCa channel, first identified in cultured rat
skeletal muscle, was shown to be the receptor inhibited by the bee
venom peptide apamin (Blatz and Magleby, 1986). Activation of
apamin-sensitive SKCa channels underlies a
component of the after hyperpolarization current in neurons that
parallels the rise and fall of intracellular calcium levels (Sah and
Clements, 1999). Besides apamin, other blockers of
SKCa channels, albeit less selective, include
tubocurarine and dequalinium. Many dequalinium analogs with varying
potencies and selectivities for blocking IKCa and
SKCa channels have been described (Malik-Hall et
al., 2000). For example, the bisaminoquinolium cyclophane UCL 1684 is
about 5000-fold more selective in inhibiting SKCa
channels compared with IKCa-type channels.
Recently, conditional overexpression of a small conductance
K+ channel, Sk3, induced abnormal breathing patterns during
hypoxia and compromised parturition in mice by changes in uterine
smooth muscle function (Bond et al., 2000). The availability of
selective SKCa modulators will permit evaluation
of their potential role in epilepsy, sleep apnea, neurodegenerative,
and smooth muscle disorders.
C. ATP-Sensitive K+ Channels
KATP channels, a family of weak inward
rectifiers inhibited by intracellular ATP that couple cellular energy
metabolism to membrane electrical activity, have perhaps been the most
widely explored K+ channels in terms of
therapeutic potential (Noma, 1983; Ashcroft and Ashcroft, 1990;
Gopalakrishnan et al., 1993). First generation K+
channel openers (KCOs), including cromakalim and pinacidil, have been
known to activate glyburide-sensitive KATP
channels in a variety of vascular and nonvascular tissues (Edwards and
Weston, 1993). A variety of structurally diverse KCOs, including
benzopyran (cromakalim), cyanoguanidines (pinacidil), and nitroethylene
analogs, have been evaluated as potential antihypertensive agents
during the past 15 years, although only nicorandil, and to a lesser
extent diazoxide, have been used in cardiovascular medicine, in part due to the availability of other classes of agents for these indications.
The recent cloning and expression of KATP channel
components has provided insight into the observed heterogeneity in the
pharmacologic profile of KCOs (reviewed in Aguilar-Bryan et al., 1998).
As noted previously, the KATP channel expressed
in pancreatic -cells is a multimeric complex composed of Kir6.2 and
the sulfonylurea receptor SUR1 (Clement et al., 1997; Lorenz et al.,
1998). From expression studies using rat or mouse SUR subunits, it is
thought that the molecular composition of the cardiac/skeletal muscle
channel is SUR2A/Kir6.2, whereas SUR2B is thought to be one of the
subunits constituting the smooth muscle type KATP
channels. More recently, SUR2 splice variants that lack either exon 14 or exon 17 have been identified by RNA analysis (Chutkow et al., 1999;
Davis-Taber et al., 2000). With the emerging diversity of
KATP channel combinations, it could be
anticipated that tissues may contain a predominance of certain isoforms
involved in various functions ranging from transmitter release to
ischemic protection and may be selectively targeted for development of
tissue-selective compounds for the treatment of several cardiac and
smooth muscle disorders.
Recent efforts have focused on the development of second
generation openers of KATP channels for
nonvascular indications including bladder overactivity, irritative
bowel syndrome, airway hyper-reactivity, erectile dysfunction, and as
cardioprotective agents for the ischemic myocardium (Morley, 1994;
Garlid et al., 1997). Compounds investigated for the treatment of
bladder overactivity such as ZM-244085, ZD-6169, or WAY-133537
have been shown to activate KATP channels,
relax bladder smooth muscle, and exhibit modest in vivo selectivity (Howe et al., 1995; Wojdan et al., 1999; Gopalakrishnan et al., 1999). Analogs derived from the benzopyran nucleus, including BMS-180448 and BMS-191095, display selectivity for cardioprotective over vasorelaxant effects relative to the nonselective KCO, cromakalim. BMS-180448 has been shown to have cardioprotective effects at concentrations that do not affect action potential shortening, indicative of activation of a KATP channel other
than the plasma membrane KATP channel. The
cardioprotective effects of the antianginal drug nicorandil have been
shown to be via activation of mitochondrial KATP
channels (Sato et al., 2000). Mammalian cells transfected with
KATP channel subunits Kir6.2 and SUR1 showed
resistance to hypoxia reoxygenation, and a therapeutic approach based
on gene delivery of KATP subunits in tissues
vulnerable to hypoxia reoxygenation and damage has also been suggested
(Jovanovic et al., 1998a,b). KCOs examined for airway hyper-reactivity
include SDZ 217-744, with reported improved selectivity of
inhibition of airway hyperactivity relative to cromakalim (Williams et
al., 1990). KATP channel openers have also been
investigated for the potential treatment of male erectile dysfunction.
Pinacidil, cromakalim, and nicorandil or its analogs have shown
increases in intracavernosal pressure by relaxing corporal smooth
muscle, which leads to initiation and maintenance of erection (Moon et
al., 1999; Vick et al., 2000), providing proof of principle that such
compounds, if delivered directly into the corpus smooth muscle, could
be a viable treatment option. The basis for the reported modest in vivo
selectivity of second generation KCOs could, in principle, arise from
interactions with distinct KATP channel
combinations or, more plausibly, from physiologic or pharmacokinetic
factors. For instance, studies aimed at elucidating the basis for the
cardioprotective effect of KCOs reveal a role for the mitochondrial
KATP channel, the molecular composition of which
appears to be somewhat distinct from sarcolemmal
KATP channels (Garlid et al., 1997; Szewczyk and
Marban, 1999).
Sulfonylureas such as glibenclamide and glipizide that block
KATP channels in pancreatic -cells have been
used for the treatment of type II diabetes for over 30 years, and newer
agents with diminished propensity for sustained hypoglycemic potential
continue to be developed. More recently, it has been demonstrated that
transfection of SUR1 and Kir6.2 into an insulin-secreting cell line
(NES 2Y -cells) from PHHI patients can restore glucose-dependent
insulin release. This opens up the potential for gene therapy to
alleviate -cell dysfunction in PHHI and diabetes (Dunne et al.,
1997; Macfarlane et al., 2000). Blockers of
KATP channels such as PNU-37883A have also been
evaluated as diuretics or as antiarrhythmic agents (Humphrey and
Ludens, 1998). More recent focus continues in the identification of
cardioselective KATP channel blockers for the
prevention of ischemia-induced ventricular fibrillation. This has been
underscored by the notion that during acute myocardial infarction,
activation of ATP-sensitive K+ currents results
in action potential duration shortening and elevation of interstitial
[K+] accumulation that may contribute to
reentry arrhythmias and cardiac death (Gögelein et al., 1998).
HMR 1883, a relatively cardioselective KATP
channel blocker with modest selectivity for cardiac
KATP over the pancreatic
KATP, prevented ventricular fibrillation in dogs
at doses that did not affect plasma insulin or blood glucose. Such
compounds may prove useful in the treatment of ventricular arrhythmias
without pancreatic side effects or the liabilities of nonselective
blockers under ischemic conditions.
D. Two-Pore K+ Channels
The more recently identified two-pore K+
channels, including TWIK, TREK, TASK, and TRAAK genes (Table 1),
thought to function as background channels involved in the modulation
of resting membrane potential in various cell types could emerge as
attractive targets for discovering novel neuroprotective and anesthetic
agents (Lesage and Lazdunski, 1999). The neuroprotective agent
riluzole, currently in use for the treatment of amyotrophic lateral
sclerosis, has been shown to be an activator of TREK-1 and TRAAK
channels (Duprat et al., 2000). Volatile general anesthetics such as
chloroform and isoflurane have also been shown to target TREK-1
channels (Patel et al., 1999). Further knowledge of the localization
and regulation of these channels by cellular and extrinsic signals will
be important in targeting specific two-pore channels for therapeutic intervention.
| |
V. Concluding Remarks |
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TopPreviousNextReferences
|
|---|
K+ channels are increasingly being
elucidated as molecular targets in a number of pathophysiologic states,
and they continue to trigger considerable enthusiasm as drug targets.
The pivotal role of K+ channels in various
physiological processes including neuronal signaling, vascular and
nonvascular muscle contractility, cardiac pacing, auditory function,
hormone secretion, immune function, and cell proliferation has been
underscored by the recent flurry of discoveries linking
K+ channel mutations to various inherited
disorders. Insight into the structure and function of channel proteins
coupled with the knowledge of genetic and disease-induced regulation of
K+ channels could undoubtedly improve diagnosis
and offer specific candidate genes for the development of appropriate
therapies. On the assumption that defined K+
channel mutations are linked to specific diseases, it may be feasible
to conduct a molecular diagnosis to evaluate whether the patient will
respond to a drug aimed at specific K+ channels.
It has been shown that differences in K+ currents
may underlie gender-based drug-induced cardiac arrhythmias; for
example, women are at far greater risk of torsade de pointes following
a variety of drugs including antihistamines, antibiotics, and
antiarrhythmic agents (Makkar et al., 1993). Analysis of the differential contribution of K+ currents
contributing to cardiac repolarization could help improve screening methodologies for individuals at risk for drug-induced arrhythmias and direct development of drugs with reduced incidence of
inducing arrhythmias. Knowledge of specific mutations may also lead to
validation of more suitable animal models of disease to help
preclinical assessment of novel compounds. In the coming years,
modulating K+ channel gene expression in diseased
tissues via various gene delivery approaches or antisense
oligonucleotides could present an additional avenue to treat various
diseases and/or, in combination with pharmacotherapy, to enhance the
selectivity of K+ channel modulators.
Additionally, unraveling precise in situ channel combinations,
localization, and channel regulation in disease pathologies could shed
light on developing better therapeutic strategies. Targeting diverse
auxiliary subunits or modulating the interactions of auxiliary subunits
with the pore-forming subunit may also provide alternate avenues for
identifying selective regulators of K+ channel
function. It is to be anticipated that these efforts could collectively
enhance the development of selective compounds that modulate the
various classes of K+ channels with promising
therapeutic and prophylactic utility.
| |
Footnotes |
|---|
1 Address for correspondence: Char-Chang Shieh, Ph.D., Dept. 47C, Bldg. AP9A 3rd Floor, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064. E-mail: char-chang.shieh{at}abbott.com
| |
Abbreviations |
|---|
Kv, voltage-gated K+
channel;
A, -amyloid;
-APP, -amyloid protein precursor;
BFNC, benign familial neonatal convulsion;
BKCa, large
conductance Ca2+-activated K+ channel;
EA, episodic ataxia;
EAG, ether-a-go-go K+
channel;
1-EBIO, 1-ethyl-2-benzimidazolinone;
hERG, human
ether-a-go-go-related K+ channel;
IKCa, intermediate conductance Ca2+-activated
K+ channel;
IKr, cardiac rapid delayed rectifier;
IKs, cardiac slow delayed rectifier;
IKur, ultrarapid delayed
rectifier;
ITO, transient outward delayed rectifier;
KATP, ATP-sensitive K+;
KCsA, K+
channel from Streptomyces lividans;
Kir, inward
rectifier K+ channel;
KCO, K+ channel opener;
LQT, long-QT syndrome;
M-channel, muscarine-sensitive K+
channel;
MiRP, minK related peptide;
PHHI, persistent hyperinsulinemic
hypoglycemia of infancy;
P-loop, pore loop;
PS, presenilin;
sAPP, secreted form of -amyloid precursor protein;
SKCa, small
conductance Ca2+-activated K+ channel;
SUR, sulfonylurea receptor;
TEA, tetraethylammonium;
TM, transmembrane
segment;
TREK, two-pore weak inward rectifier-related K+
channel.
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References |
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|
TopPrevious |
|---|
basic science and clinical disease.
New Engl J Med
336:
1575-1586proteolysis holds the key.
Science (Wash DC)
286:
916-919
p11.1 Cytogenet Cell Genet 79:85-87. q14.1.
Genomics
15:
463-464[Medline].
0031-6997/00/5204-0557$03.00/0
PHARMACOLOGICAL REVIEWS
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