A cellular test system expressing hERG K+ channels for assaying drug cardiotoxicity: Elaboration and analysis

Many drugs are cardiotoxic because they inhibit hERG K⁺ channels, thus prolonging the repolarization phase of the cardiomyocyte action potential giving rise to cardiac arrhythmias. Early detection of inhibiting effects of candidate drugs on the activity of K⁺ channels in cardiomyocytes is one of the main challenges in preclinical drug screening. The aim of this study was to obtain a cell line expressing recombinant hERG channels at a stable and reproducible level as a prerequisite to its further application as a test system.     

Rab11-dependent Recycling of the Human Ether-a-go-go-related Gene (hERG) Channel

The human ether-a-go-go-related gene (hERG) encodes the pore-forming subunit of the rapidly activating delayed rectifier potassium channel (I_Kr). A reduction in the hERG current causes long QT syndrome, which predisposes affected individuals to ventricular arrhythmias and sudden death. We reported previously that hERG channels in the plasma membrane undergo vigorous internalization under low K⁺ conditions. In the present study, we addressed whether hERG internalization occurs under normal K⁺ conditions and whether/how internalized channels are recycled back to the plasma membrane. Using patch clamp, Western blot, and confocal imaging analyses, we demonstrated that internalized hERG channels can effectively recycle back to the plasma membrane. Low K⁺-enhanced hERG internalization is accompanied by an increased rate of hERG recovery in the plasma membrane upon reculture following proteinase K-mediated clearance of cell-surface proteins. The increased recovery rate is not due to enhanced protein synthesis, as hERG mRNA expression was not altered by low K⁺ exposure, and the increased recovery was observed in the presence of the protein biosynthesis inhibitor cycloheximide. GTPase Rab11, but not Rab4, is involved in the recycling of hERG channels. Interfering with Rab11 function not only delayed hERG recovery in cells after exposure to low K⁺ medium but also decreased hERG expression and function in cells under normal culture conditions. We concluded that the recycling pathway plays an important role in the homeostasis of plasma membrane-bound hERG channels.     

Involvement of Caveolin in Low K+-induced Endocytic Degradation of Cell-surface Human Ether-a-go-go-related Gene (hERG) Channels

Reduction in the rapidly activating delayed rectifier K⁺ channel current (I_Kr) due to either mutations in the human ether-a-go-go-related gene (hERG) or drug block causes inherited or drug-induced long QT syndrome. A reduction in extracellular K⁺ concentration ([K⁺]_o) exacerbates long QT syndrome. Recently, we demonstrated that lowering [K⁺]_o promotes degradation of I_Kr in rabbit ventricular myocytes and of the hERG channel stably expressed in HEK 293 cells. In this study, we investigated the degradation pathways of hERG channels under low K⁺ conditions. We demonstrate that under low K⁺ conditions, mature hERG channels and caveolin-1 (Cav1) displayed a parallel time-dependent reduction. Mature hERG channels coprecipitated with Cav1 in co-immunoprecipitation analysis, and internalized hERG channels colocalized with Cav1 in immunocytochemistry analysis. Overexpression of Cav1 accelerated internalization of mature hERG channels in 0 mm K⁺_o, whereas knockdown of Cav1 impeded this process. In addition, knockdown of dynamin 2 using siRNA transfection significantly impeded hERG internalization and degradation under low K⁺_o conditions. In cultured neonatal rat ventricular myocytes, knockdown of caveolin-3 significantly impeded low K⁺_o-induced reduction of I_Kr. Our data indicate that a caveolin-dependent endocytic route is involved in low K⁺_o-induced degradation of mature hERG channels.     

The role of hERG1 K channels and a functional link between hERG1 K channels and SDF-1 in acute leukemic cell migration

Stromal cell-derived factor-1 (SDF-1) and its unique receptor, CXCR4, regulate stem/progenitor cell migration and retention in the bone marrow and are required for hematopoiesis. Recent studies found that hERG1 K⁺ channels were important regulators of tumor cell migration. In this study, we investigated whether SDF-1 induced acute leukemic cell migration associated with hERG1 K⁺ channels. Our results showed that E-4031, a specific hERG1 K⁺ channels inhibitor, significantly blocked SDF-1-induced migration of leukemic cell lines, primary acute leukemic cells, leukemic stem cells and HEK293T cells transfected with herg-pEGFP. The migration of phenotypically recognizable subsets gave the indication that lymphoblastic leukemic cells were inhibited more than myeloid cells while in the presence of E-4031 which maybe associated with herg expression. SDF-1 increased hERG1 K⁺ current expressed in oocytes and HEK293T cells transfected with herg-pEGFP. There were no significant changes of CXCR4 expression on both HL-60 cells and primary leukemic cells regardless if untreated or treated with E-4031 for 24 h (P > 0.05). The hERG1 K⁺ current increased by SDF-1 might contribute to the mechanism of SDF-1-induced leukemic cell migration. The data suggested that hERG1 K⁺ channels functionally linked to cell migration induced by SDF-1.     

The Life and Death of Breast Cancer Cells: Proposing a Role for the Effects of Phytoestrogens on Potassium Channels

Changes in the regulation of potassium channels are increasingly implicated in the altered activity of breast cancer cells. Increased or reduced expression of a number of K⁺ channels have been identified in numerous breast cancer cell lines and cancerous tissue biopsy samples, compared to normal tissue, and are associated with tumor formation and spread, enhanced levels of proliferation, and resistance to apoptotic stimuli. Through knockout or silencing of K⁺ channel genes, and use of specific or more broad pharmacologic K⁺ channel blockers, the growth of numerous cell lines, including breast cancer cells, has been modified. In this manner it has been proposed that in MCF7 breast cancer cells proliferation appears to be regulated by the activity of a number of K⁺ channels, including the Ca²⁺ activated K⁺ channels, and the voltage-gated K⁺ channels hEAG and K_v1.1. The effect of phytoestrogens on K⁺ channels has not been extensively studied but yields some interesting results. In a number of cell lines the phytoestrogen genistein inhibits K⁺ current through several channels including K_v1.3 and hERG. Where it has been used, structurally similar daidzein has little or no effect on K⁺ channel activity. Since many K⁺ channels have roles in proliferation and apoptosis in breast cancer cells, the impact of K⁺ channel regulation by phytoestrogens is of potentially great relevance.     

Differential modulation of cardiac potassium channels by Grb adaptor proteins

Scaffolding growth factor receptor-bound (Grb) adaptor proteins are components of macromolecular signaling complexes at the plasma membrane and thus are putative regulators of ion channel activity. The present study aimed to define the impact of Grb adaptor proteins on the function of cardiac K⁺ channels. To this end channel proteins were coinjected with the adaptor proteins in Xenopus oocytes and channel activity analyzed with two-electrode voltage-clamp. It is shown that coexpression of Grb adaptor proteins can reduce current amplitudes of coexpressed channels. Grb7 and 10 significantly inhibited functional currents generated by hERG, Kv1.5 and Kv4.3 channels. Only Grb10 significantly inhibited KCNQ1/KCNE1 K⁺ channels, and only Grb7 reduced Kir2.3 activity, whereas neither Grb protein significantly affected the closely related Kir2.1 and Kir2.2 channels. The present observations for the first time provide evidence for a selective and modulatory role of Grb adaptor proteins in the functional expression of cardiac K⁺ channels.     

Regulation of the Human Ether-a-go-go-related Gene (hERG) Channel by Rab4 Protein through Neural Precursor Cell-expressed Developmentally Down-regulated Protein 4-2 (Nedd4-2)

The human ether-a-go-go-related gene (hERG) encodes the pore-forming α-subunit of the rapidly activating delayed rectifier K⁺ channel in the heart, which plays a critical role in cardiac action potential repolarization. Dysfunction of I_Kr causes long QT syndrome, a cardiac electrical disorder that predisposes affected individuals to fatal arrhythmias and sudden death. The homeostasis of hERG channels in the plasma membrane depends on a balance between protein synthesis and degradation. Our recent data indicate that hERG channels undergo enhanced endocytic degradation under low potassium (hypokalemia) conditions. The GTPase Rab4 is known to mediate rapid recycling of various internalized proteins to the plasma membrane. In the present study, we investigated the effect of Rab4 on the expression level of hERG channels. Our data revealed that overexpression of Rab4 decreases the expression level of hERG in the plasma membrane. Rab4 does not affect the expression level of the Kv1.5 or EAG K⁺ channels. Mechanistically, our data demonstrate that overexpression of Rab4 increases the expression level of endogenous Nedd4-2, a ubiquitin ligase that targets hERG but not Kv1.5 or EAG channels for ubiquitination and degradation. Nedd4-2 undergoes self- ubiquitination and degradation. Rab4 interferes with Nedd4-2 degradation, resulting in an increased expression level of Nedd4-2, which targets hERG. In summary, the present study demonstrates a novel pathway for hERG regulation; Rab4 decreases the hERG density at the plasma membrane by increasing the endogenous Nedd4-2 expression.     

Stoichiometry of altered hERG1 channel gating by small molecule activators

Voltage-gated K⁺ channels are tetramers formed by coassembly of four identical or highly related subunits. All four subunits contribute to formation of the selectivity filter, the narrowest region of the channel pore which determines K⁺ selective conductance. In some K⁺ channels, the selectivity filter can undergo a conformational change to reduce K⁺ flux by a mechanism called C-type inactivation. In human ether-a-go-go–related gene 1 (hERG1) K⁺ channels, C-type inactivation is allosterically inhibited by ICA-105574, a substituted benzamide. PD-118057, a 2-(phenylamino) benzoic acid, alters selectivity filter gating to enhance open probability of channels. Both compounds bind to a hydrophobic pocket located between adjacent hERG1 subunits. Accordingly, a homotetrameric channel contains four identical activator binding sites. Here we determine the number of binding sites required for maximal drug effect and determine the role of subunit interactions in the modulation of hERG1 gating by these compounds. Concatenated tetramers were constructed to contain a variable number (zero to four) of wild-type and mutant hERG1 subunits, either L646E to inhibit PD-118057 binding or F557L to inhibit ICA-105574 binding. Enhancement of hERG1 channel current magnitude by PD-118057 and attenuated inactivation by ICA-105574 were mediated by cooperative subunit interactions. Maximal effects of the both compounds required the presence of all four binding sites. Understanding how hERG1 agonists allosterically modify channel gating may facilitate mechanism-based drug design of novel agents for treatment of long QT syndrome.     

Isoform-Specific Dominant-Negative Effects Associated with hERG1 G628S Mutation in Long QT Syndrome

Background

Mutations in the human ether-a-go-go-related gene 1 (hERG1) cause type 2 long QT syndrome (LQT2). The hERG1 gene encodes a K⁺ channel with properties similar to the rapidly activating delayed rectifying K⁺ current in the heart. Several hERG1 isoforms with unique structural and functional properties have been identified. To date, the pathogenic mechanisms of LQT2 mutations have been predominantly described in the context of the hERG1a isoform. In the present study, we investigated the functional consequences of the LQT2 mutation G628S in the hERG1b and hERG1a_USO isoforms.

Methods

A double-stable, mammalian expression system was developed to characterize isoform-specific dominant-negative effects of G628S-containing channels when co-expressed at equivalent levels with wild-type hERG1a. Western blot and co-immunoprecipitation studies were performed to study the trafficking and co-assembly of wild-type and mutant hERG1 isoforms. Patch-clamp electrophysiology was performed to characterize hERG1 channel function and the isoform-specific dominant-negative effects associated with the G628S mutation.

Conclusions

The non-functional hERG1a-G628S and hERG1b-G628S channels co-assembled with wild-type hERG1a and dominantly suppressed hERG1 current. In contrast, G628S-induced dominant-negative effects were absent in the context of the hERG1a_USO isoform. hERG1a_USO-G628S channels did not appreciably associate with hERG1a and did not significantly suppress hERG1 current when co-expressed at equivalent ratios or at ratios that approximate those found in cardiac tissue. These results suggest that the dominant-negative effects of LQT2 mutations may primarily occur in the context of the hERG1a and hERG1b isoforms.     

Novel roles for hERG K+ channels in cell proliferation and apoptosis

The human ether-a-go-go-related gene potassium channel (hERG, Kv11.1, KCNH2) has an essential role in cardiac action potential repolarization. Electrical dysfunction of the voltage-sensitive ion channel is associated with potentially lethal ventricular arrhythmias in humans. hERG K⁺ channels are also expressed in a variety of cancer cells where they control cell proliferation and apoptosis. In this review, we discuss molecular mechanisms of hERG-associated cell cycle regulation and cell death. In addition, the significance of hERG K⁺ channels as future drug target in anticancer therapy is highlighted.     

Blockade of permeation by potassium but normal gating of the G628S nonconducting hERG channel mutant

G628S is a mutation in the signature sequence that forms the selectivity filter of the human ether-a-go-go-related gene (hERG) channel (GFG) and is associated with long-QT2 syndrome. G628S channels are known to have a dominant-negative effect on hERG currents, and the mutant is therefore thought to be nonfunctional. This study aims to assess the physiological mechanism that prevents the surface-expressing G628S channels from conducting ions. We used voltage-clamp fluorimetry along with two-microelectrode voltage clamping in Xenopus oocytes to confirm that the channels express well at the surface, and to show that they are actually functional, with activation kinetics comparable to that of wild-type, and that the mutation leads to a reduced selectivity to potassium. Although ionic currents are not detected in physiological solutions, removing extracellular K⁺ results in the appearance of an inward Na⁺-dependent current. Using whole-cell patch clamp in mammalian transfected cells, we demonstrate that the G628S channels conduct Na⁺, but that this can be blocked by both intracellular and higher-than-physiological extracellular K⁺. Using solutions devoid of K⁺ allows the appearance of nA-sized Na⁺ currents with activation and inactivation gating analogous to wild-type channels. The G628S channels are functionally conducting but are normally blocked by intracellular K⁺.     

Predicting the potency of hERG K<Superscript>+</Superscript> channel inhibition by combining 3D-QSAR pharmacophore and 2D-QSAR models

Blockade of the hERG K⁺ channel has been identified as the most important mechanism of QT interval prolongation and thus inducing cardiac risk. In this work, an ensemble of 3D-QSAR pharmacophore models was constructed to provide insight into the determinants of the interactions between the hERG K⁺ channel and channel inhibitors. To predict hERG inhibitory activities, the predicted values from the ensemble of models were averaged, and the results thus obtained showed that the predictive ability of the combined 3D-QSAR pharmacophore model was greater that those of the individual models. Also, using the same training and test sets, a 2D-QSAR model based on a heuristic machine-learning method was developed in order to analyze the physicochemical characters of hERG inhibitors. The models indicated that the inhibitors have certain key inhibitory features in common, including hydrophobicity, aromaticity, and flexibility. A final model was developed by combining the combined 3D-QSAR pharmacophore with the 2D-QSAR model, and this final model outperformed any other individual model, showing the highest predictive ability and the lowest deviation. This model can not only predict hERG inhibitory potency accurately, thus allowing fast cardiac safety evaluation, but it provides an effective tool for avoiding hERG inhibitory liability and thus enhanced cardiac risk in the design and optimization of new chemical entities.     

Potential role of the membrane in hERG channel functioning and drug-induced long QT syndrome

The human ether-à-go-go related gene (hERG) potassium channels are located in the myocardium cell membrane where they ensure normal cardiac activity. The binding of drugs to this channel, a side effect known as drug-induced (acquired) long QT syndrome (ALQTS), can lead to arrhythmia or sudden cardiac death. The hERG channel is a unique member of the family of voltage-gated K⁺ channels because of the long extracellular loop connecting its transmembrane S5 helix to the pore helix in the pore domain. Considering the proximal position of the S5-P linker to the membrane surface, we have investigated the interaction of its central segment I⁵⁸³-Y⁵⁹⁷ with bicelles. Liquid and solid-state NMR experiments as well as circular dichroism results show a strong affinity of the I⁵⁸³-Y⁵⁹⁷ segment for the membrane where it would sit on the surface with no defined secondary structure. A structural dependence of this segment on model membrane composition was observed. A helical conformation is favoured in detergent micelles and in the presence of negative charges. Our results suggest that the interaction of the S5-P linker with the membrane could participate in the stabilization of transient channel conformations, but helix formation would be triggered by interactions with other hERG domains. Because potential drug binding sites on the S5-P linker have been identified, we have explored the role of this segment in ALQTS. Four LQTS-liable drugs were studied which showed more affinity for the membrane than this hERG segment. Our results, therefore, identify two possible roles for the membrane in channel functioning and ALQTS.     

Enhanced inhibitory effect of acidosis on hERG potassium channels that incorporate the hERG1b isoform

Extracellular acidosis occurs in the heart during myocardial ischemia and can lead to dangerous arrhythmias. Potassium channels encoded by hERG (human ether-à-go-go-related gene) mediate the cardiac rapid delayed rectifier K⁺ current (I_Kr), and impaired hERG function can exacerbate arrhythmia risk. Nearly all electrophysiological investigations of hERG have centred on the hERG1a isoform, although native I_Kr channels may be comprised of hERG1a and hERG1b, which has a unique shorter N-terminus. This study has characterised for the first time the effects of extracellular acidosis (an extracellular pH decrease from 7.4 to 6.3) on hERG channels incorporating the hERG1b isoform. Acidosis inhibited hERG1b current amplitude to a significantly greater extent than that of hERG1a, with intermediate effects on coexpressed hERG1a/1b. I_hERG tail deactivation was accelerated by acidosis for both isoforms. hERG1a/1b activation was positively voltage-shifted by acidosis, and the fully-activated current–voltage relation was reduced in amplitude and right-shifted (by ∼10 mV). Peak I_hERG1a/1b during both ventricular and atrial action potentials was both suppressed and positively voltage-shifted by acidosis. Differential expression of hERG isoforms may contribute to regional differences in I_Kr in the heart. Therefore inhibitory effects of acidosis on I_Kr could also differ regionally, depending on the relative expression levels of hERG1a and 1b, thereby increasing dispersion of repolarization and arrhythmia risk.     

The Eag Domain Regulates the Voltage-Dependent Inactivation of Rat Eag1 K+ Channels

Eag (Kv10) and Erg (Kv11) belong to two distinct subfamilies of the ether-à-go-go K⁺ channel family (KCNH). While Erg channels are characterized by an inward-rectifying current-voltage relationship that results from a C-type inactivation, mammalian Eag channels display little or no voltage-dependent inactivation. Although the amino (N)-terminal region such as the eag domain is not required for the C-type inactivation of Erg channels, an N-terminal deletion in mouse Eag1 has been shown to produce a voltage-dependent inactivation. To further discern the role of the eag domain in the inactivation of Eag1 channels, we generated N-terminal chimeras between rat Eag (rEag1) and human Erg (hERG1) channels that involved swapping the eag domain alone or the complete cytoplasmic N-terminal region. Functional analyses indicated that introduction of the homologous hERG1 eag domain led to both a fast phase and a slow phase of channel inactivation in the rEag1 chimeras. By contrast, the inactivation features were retained in the reverse hERG1 chimeras. Furthermore, an eag domain-lacking rEag1 deletion mutant also showed the fast phase of inactivation that was notably attenuated upon co-expression with the rEag1 eag domain fragment, but not with the hERG1 eag domain fragment. Additionally, we have identified a point mutation in the S4–S5 linker region of rEag1 that resulted in a similar inactivation phenotype. Biophysical analyses of these mutant constructs suggested that the inactivation gating of rEag1 was distinctly different from that of hERG1. Overall, our findings are consistent with the notion that the eag domain plays a critical role in regulating the inactivation gating of rEag1. We propose that the eag domain may destabilize or mask an inherent voltage-dependent inactivation of rEag1 K⁺ channels.     

Control of K+ channels by G proteins

Heterotrimeic G proteins are thought to couple receptors to ionic channels via cytoplasmic mediators such as cGMP in the case of retinal rods, cAMP in the case of olfactory cells, and the cAMP cascade in the case of cardiac myocytes. G protein-mediated second messenger effects on K⁺ channels are dealt with elsewhere in this series. Recently, membrane-delimited pathways have been uncovered and an hypothesis proposed in which the subunits of G proteins directly couple receptors to ionic channels, particularly K⁺ channels. While direct coupling has not been proven, the membrane-delimited nature has been established for specific G proteins and their specific K⁺ channel effectors.     

The voltage-sensing domain of a hERG1 mutant is a cation-selective channel

A cationic leak current known as an “omega current” may arise from mutations of the first charged residue in the S4 of the voltage sensor domains of sodium and potassium voltage-gated channels. The voltage-sensing domains (VSDs) in these mutated channels act as pores allowing nonspecific passage of cations, such as Li⁺, K⁺, Cs⁺, and guanidinium. Interestingly, no omega currents have been previously detected in the nonswapped voltage-gated potassium channels such as the human-ether-a-go-go-related (hERG1), hyperpolarization-activated cyclic nucleotide-gated, and ether-a-go-go channels. In this work, we discovered a novel omega current by mutating the first charged residue of the S4 of the hERG1, K525 to serine. To characterize this omega current, we used various probes, including the hERG1 pore domain blocker, dofetilide, to show that the omega current does not require cation flux via the canonical pore domain. In addition, the omega flux does not cross the conventional selectivity filter. We also show that the mutated channel (K525S hERG1) conducts guanidinium. These data are indicative of the formation of an omega current channel within the VSD. Using molecular dynamics simulations with replica-exchange umbrella sampling simulations of the wild-type hERG1 and the K525S hERG1, we explored the molecular underpinnings governing the cation flow in the VSD of the mutant. We also show that the wild-type hERG1 may form water crevices supported by the biophysical surface accessibility data. Overall, our multidisciplinary study demonstrates that the VSD of hERG1 may act as a cation-selective channel wherein a mutation of the first charged residue in the S4 generates an omega current. Our simulation uncovers the atomistic underpinning of this mechanism.     

Concerted All-or-none Subunit Interactions Mediate Slow Deactivation of Human ether-à-go-go-related Gene K+ Channels

During the repolarization phase of a cardiac action potential, hERG1 K⁺ channels rapidly recover from an inactivated state then slowly deactivate to a closed state. The resulting resurgence of outward current terminates the plateau phase and is thus a key regulator of action potential duration of cardiomyocytes. The intracellular N-terminal domain of the hERG1 subunit is required for slow deactivation of the channel as its removal accelerates deactivation 10-fold. Here we investigate the stoichiometry of hERG1 channel deactivation by characterizing the kinetic properties of concatenated tetramers containing a variable number of wild-type and mutant subunits. Three mutations known to accelerate deactivation were investigated, including R56Q and R4A/R5A in the N terminus and F656I in the S6 transmembrane segment. In all cases, a single mutant subunit induced the same rapid deactivation of a concatenated channel as that observed for homotetrameric mutant channels. We conclude that slow deactivation gating of hERG1 channels involves a concerted, fully cooperative interaction between all four wild-type channel subunits.     

hERG1a/1b heteromeric currents exhibit amplified attenuation of inactivation in variant 1 short QT syndrome

Potassium channels encoded by hERG (human ether-à-go-go-related gene) underlie the cardiac rapid delayed rectifier K⁺ current (I_Kr) and hERG mutations underpin clinically important repolarization disorders. Virtually all electrophysiological investigations of hERG mutations have studied exclusively the hERG1a isoform; however, recent evidence indicates that native I_Kr channels may be comprised of hERG1a together with the hERG1b variant, which has a shorter N-terminus. Here, for the first time, electrophysiological effects were studied of a gain-of-function hERG mutation (N588K; responsible for the ‘SQT1’ variant of the short QT syndrome) on current (I_hERG1a/1b) carried by co-expressed hERG1a/1b channels. There were no significant effects of N588K on I_hERG1a/1b activation or deactivation, but N588K I_hERG1a/1b showed little inactivation up to highly positive voltages (?+80 mV), a more marked effect than seen for hERG1a expressed alone. I_hERG1a/1b under action potential voltage-clamp, and the effects on this of the N588K mutation, also showed differences from those previously reported for hERG1a. The amplified attenuation of I_hERG inactivation for the N588K mutation reported here indicates that the study of co-expressed hERG1a/1b channels should be considered when investigating clinically relevant hERG channel mutations, even if these reside outside of the N-terminus region.     

Acute hypoxia differentially regulates K<Superscript>+</Superscript> channels. Implications with respect to cardiac arrhythmia

The first ion channels demonstrated to be sensitive to changes in oxygen tension were K⁺ channels in glomus cells of the carotid body. Since then a number of hypoxia-sensitive ion channels have been identified. However, not all K⁺ channels respond to hypoxia alike. This has raised some debate about how cells detect changes in oxygen tension. Because ion channels respond rapidly to hypoxia it has been proposed that the channel is itself an oxygen sensor. However, channel function can also be modified by thiol reducing and oxidizing agents, implicating reactive oxygen species as signals in hypoxic events. Cardiac ion channels can also be modified by hypoxia and redox agents. The rapid and slow components of the delayed rectifier K⁺ channel are differentially regulated by hypoxia and -adrenergic receptor stimulation. Mutations in the genes that encode the subunits for the channel are associated with Long QT syndrome and sudden cardiac death. The implications with respect to effects of hypoxia on the channel and triggering of cardiac arrhythmia will be discussed.