Fluid flow facilitates inward rectifier K+ current by convectively restoring [K+] at the cell membrane surface.
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ABSTRACT:
SUBMITTER: Kim JG
PROVIDER: S-EPMC5177964 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Background Methadone is associated with a disproportionate risk of sudden death and ventricular tachyarrhythmia despite only modest inhibition of delayed rectifier K+ current (IKr), the principal mechanism of drug-associated arrhythmia. Congenital defects of inward rectifier K+ current (IK1) have been linked to increased U-wave amplitude on ECG and fatal arrhythmia. We hypothesized that methadone may also be a potent inhibitor of IK1, contributing to delayed repolarization and manifesting on surface ECGs as augmented U-wave integrals. Methods and Results Using a whole-cell voltage clamp, methadone inhibited both recombinant and native IK1 with a half-maximal inhibitory concentration IC50) of 1.5 μmol/L, similar to that observed for IKr block (half-maximal inhibitory concentration of 2.9 μmol/L). Methadone modestly increased the action potential duration at 90% repolarization and slowed terminal repolarization at low concentrations. At higher concentrations, action potential duration at 90% repolarization lengthening was abolished, but its effect on terminal repolarization rose steadily and correlated with increased fluctuations of diastolic membrane potential. In parallel, patient ECGs were analyzed before and after methadone initiation, with 68% of patients having a markedly increased U-wave integral compared with premethadone (lead V3; mean +38%±15%, P=0.016), along with increased QT and TPeak to TEnd intervals, likely reflective of IKr block. Conclusions Methadone is a potent IK1 inhibitor that causes augmentation of U waves on surface ECG. We propose that increased membrane instability resulting from IK1 block may better explain methadone's arrhythmia risk beyond IKr inhibition alone. Drug-induced augmentation of U waves may represent evidence of blockade of multiple repolarizing ion channels, and evaluation of the effect of that agent on IK1 may be warranted.
| S-EPMC9238736 | biostudies-literature
Project description:Inwardly rectifying potassium (Kir) channels play a key role in controlling membrane potentials in excitable and unexcitable cells, thereby regulating a plethora of physiological processes. G-protein-gated Kir channels control heart rate and neuronal excitability via small hyperpolarizing outward K+ currents near the resting membrane potential. Despite recent breakthroughs in x-ray crystallography and cryo-EM, the gating and conduction mechanisms of these channels are poorly understood. MD simulations have provided unprecedented details concerning the gating and conduction mechanisms of voltage-gated K+ and Na+ channels. Here, we use multi-microsecond-timescale MD simulations based on the crystal structures of GIRK2 (Kir3.2) bound to phosphatidylinositol-4,5-bisphosphate to provide detailed insights into the channel's gating dynamics, including insights into the behavior of the G-loop gate. The simulations also elucidate the elementary steps that underlie the movement of K+ ions through an inward-rectifier K+ channel under an applied electric field. Our simulations suggest that K+ permeation might occur via direct knock-on, similar to the mechanism recently shown for Kv channels.
| S-EPMC6785732 | biostudies-literature
Project description:The purpose of this study was to investigate the functional role of G-protein-coupled inward rectifier potassium (GIRK) channels in the cardiac ventricle.Immunofluorescence experiments demonstrated that GIRK4 was localized in outer sarcolemmas and t-tubules in GIRK1 knockout (KO) mice, whereas GIRK4 labelling was not detected in GIRK4 KO mice. GIRK4 was localized in intercalated discs in rat ventricle, whereas it was expressed in intercalated discs and outer sarcolemmas in rat atrium. GIRK4 was localized in t-tubules and intercalated discs in human ventricular endocardium and epicardium, but absent in mid-myocardium. Electrophysiological recordings in rat ventricular tissue ex vivo showed that the adenosine A1 receptor agonist N6-cyclopentyladenosine (CPA) and acetylcholine (ACh) shortened action potential duration (APD), and that the APD shortening was reversed by either the GIRK channel blocker tertiapin-Q, the adenosine A1 receptor antagonist DPCPX or by the muscarinic M2 receptor antagonist AF-DX 116. Tertiapin-Q prolonged APD in the absence of the exogenous receptor activation. Furthermore, CPA and ACh decreased the effective refractory period and the effect was reversed by either tertiapin-Q, DPCPX or AF-DX 116. Receptor activation also hyperpolarized the resting membrane potential, an effect that was reversed by tertiapin-Q. In contrast, tertiapin-Q depolarized the resting membrane potential in the absence of the exogenous receptor activation.Confocal microscopy shows that among species GIRK4 is differentially localized in the cardiac ventricle, and that it is heterogeneously expressed across human ventricular wall. Electrophysiological recordings reveal that GIRK current may contribute significantly to ventricular repolarization and thereby to cardiac electrical stability.
| S-EPMC3868351 | biostudies-literature
Project description:Atrial fibrillation (AF) is a highly prevalent arrhythmia with pronounced morbidity and mortality. Inward-rectifier K+ current (IK1) is believed to be an important regulator of reentrant-spiral dynamics and a major component of AF-related electrical remodeling. MicroRNA-26 (miR-26) is predicted to target the gene encoding KIR2.1, KCNJ2. We found that miR-26 was downregulated in atrial samples from AF animals and patients and this downregulation was accompanied by upregulation of IK1/KIR2.1 protein. miR-26 overexpression suppressed expression of KCNJ2/KIR2.1. In contrast, miR-26 knockdown, inhibition, or binding-site mutation enhanced KCNJ2/KIR2.1 expression, establishing KCNJ2 as a miR-26 target. Knockdown of endogenous miR-26 promoted AF in mice, whereas adenovirus-mediated expression of miR-26 reduced AF vulnerability. Kcnj2-specific miR-masks eliminated miR-26-mediated reductions in Kcnj2, abolishing miR-26's protective effects, while coinjection of a Kcnj2-specific miR-mimic prevented miR-26 knockdown-associated AF in mice. Nuclear factor of activated T cells (NFAT), a known actor in AF-associated remodeling, was found to negatively regulate miR-26 transcription. Our results demonstrate that miR-26 controls the expression of KCNJ2 and suggest that this downregulation may promote AF.
| S-EPMC3635715 | biostudies-literature
Project description:Precise trafficking, localization, and activity of inward rectifier potassium Kir2 channels are important for shaping the electrical response of skeletal muscle. However, how coordinated trafficking occurs to target sites remains unclear. Kir2 channels are tetrameric assemblies of Kir2.x subunits. By immunocytochemistry we show that endogenous Kir2.1 and Kir2.2 are localized at the plasma membrane and T-tubules in rodent skeletal muscle. Recently, a new subunit, Kir2.6, present in human skeletal muscle, was identified as a gene in which mutations confer susceptibility to thyrotoxic hypokalemic periodic paralysis. Here we characterize the trafficking and interaction of wild type Kir2.6 with other Kir2.x in COS-1 cells and skeletal muscle in vivo. Immunocytochemical and electrophysiological data demonstrate that Kir2.6 is largely retained in the endoplasmic reticulum, despite high sequence identity with Kir2.2 and conserved endoplasmic reticulum and Golgi trafficking motifs shared with Kir2.1 and Kir2.2. We identify amino acids responsible for the trafficking differences of Kir2.6. Significantly, we show that Kir2.6 subunits can coassemble with Kir2.1 and Kir2.2 in vitro and in vivo. Notably, this interaction limits the surface expression of both Kir2.1 and Kir2.2. We provide evidence that Kir2.6 functions as a dominant negative, in which incorporation of Kir2.6 as a subunit in a Kir2 channel heterotetramer reduces the abundance of Kir2 channels on the plasma membrane.
| S-EPMC3059001 | biostudies-literature
Project description:Cardiac conduction disorders are common diseases which cause slow heart rate and syncope. The best way to treat these diseases by now is to implant electronic pacemakers, which, yet, have many disadvantages, such as the limited battery life and infection. Biopacemaker has been expected to replace the electronic devices. Automatic ventricular myocytes (VMs) could show pacemaker activity, which was induced by depressing inward-rectifier K(+) current (I K1). In this study, a 2D model of human biopacemaker was created from the ventricular endocardial myocytes. We examined the stability of the created biopacemaker and investigated its driving capability by finding the suitable size and spatial distribution of the pacemaker for robust pacing and driving the surrounding quiescent cardiomyocytes. Our results suggest that the rhythm of the pacemaker is similar to that of the single cell at final stable state. The driving force of the biopacemaker is closely related to the pattern of spatial distribution of the pacemaker.
| S-EPMC4779510 | biostudies-literature
Project description:Synapse-associated protein-97 (SAP97) is a membrane-associated guanylate kinase scaffolding protein expressed in cardiomyocytes. SAP97 has been shown to associate and modulate voltage-gated potassium (Kv) channel function. In contrast to Kv channels, little information is available on interactions involving SAP97 and inward rectifier potassium (Kir2.x) channels that underlie the classical inward rectifier current, I(K1). To investigate the functional effects of silencing SAP97 on I(K1) in adult rat ventricular myocytes, SAP97 was silenced using an adenoviral short hairpin RNA vector. Western blot analysis showed that SAP97 was silenced by approximately 85% on day 3 post-infection. Immunostaining showed that Kir2.1 and Kir2.2 co-localize with SAP97. Co-immunoprecipitation (co-IP) results demonstrated that Kir2.x channels associate with SAP97. Voltage clamp experiments showed that silencing SAP97 reduced I(K1) whole cell density by approximately 55%. I(K1) density at -100 mV was -1.45 +/- 0.15 pA/picofarads (n = 6) in SAP97-silenced cells as compared with -3.03 +/- 0.37 pA/picofarads (n = 5) in control cells. Unitary conductance properties of I(K1) were unaffected by SAP97 silencing. The major mechanism for the reduction of I(K1) density appears to be a decrease in Kir2.x channel abundance. Furthermore, SAP97 silencing impaired I(K1) regulation by beta(1)-adrenergic receptor (beta1-AR) stimulation. In control, isoproterenol reduced I(K1) amplitude by approximately 75%, an effect that was blunted following SAP97 silencing. Our co-IP data show that beta1-AR associates with SAP97 and Kir2.1 and also that Kir2.1 co-IPs with protein kinase A and beta1-AR. SAP97 immunolocalizes with protein kinase A and beta1-AR in the cardiac myocytes. Our results suggest that in cardiac myocytes SAP97 regulates surface expression of channels underlying I(K1), as well as assembles a signaling complex involved in beta1-AR regulation of I(K1).
| S-EPMC2934665 | biostudies-literature
Project description:Inward rectifier potassium (Kir) channels are integral membrane proteins charged with a key role in establishing the resting membrane potential of excitable cells through selective control of the permeation of K(+) ions across cell membranes. In conjunction with secondary anionic phospholipids, members of this family are directly regulated by phosphoinositides (PIPs) in the absence of other proteins or downstream signaling pathways. Different Kir isoforms display distinct specificities for the activating PIPs but all eukaryotic Kir channels are activated by PI(4,5)P2. On the other hand, the bacterial KirBac1.1 channel is inhibited by PIPs. Recent crystal structures of eukaryotic Kir channels in apo and lipid bound forms reveal one specific binding site per subunit, formed at the interface of N- and C-terminal domains, just beyond the transmembrane segments and clearly involving some of the key residues previously identified as controlling PI(4,5)P2 sensitivity. Computational, biochemical, and biophysical approaches have attempted to address the energetic determinants of PIP binding and selectivity among Kir channel isoforms, as well as the conformational changes that trigger channel gating. Here we review our current understanding of the molecular determinants of PIP regulation of Kir channel activity, including in context with other lipid modulators, and provide further discussion on the key questions that remain to be answered.
| S-EPMC3884141 | biostudies-literature
Project description:BackgroundRecent evidence suggests that atrial fibrillation (AF) is maintained by high-frequency reentrant sources with a left-to-right-dominant frequency gradient, particularly in patients with paroxysmal AF (pAF). Unequal left-to-right distribution of inward rectifier K(+) currents has been suggested to underlie this dominant frequency gradient, but this hypothesis has never been tested in humans.Methods and resultsCurrents were measured with whole-cell voltage-clamp in cardiomyocytes from right atrial (RA) and left (LA) atrial appendages of patients in sinus rhythm (SR) and patients with AF undergoing cardiac surgery. Western blot was used to quantify protein expression of I(K1) (Kir2.1 and Kir2.3) and I(K,ACh) (Kir3.1 and Kir3.4) subunits. Basal current was ≈2-fold larger in chronic AF (cAF) versus SR patients, without RA-LA differences. In pAF, basal current was ≈2-fold larger in LA versus RA, indicating a left-to-right atrial gradient. In both atria, Kir2.1 expression was ≈2-fold greater in cAF but comparable in pAF versus SR. Kir2.3 levels were unchanged in cAF and RA-pAF but showed a 51% decrease in LA-pAF. In SR, carbachol-activated (2 μmol/L) I(K,ACh) was 70% larger in RA versus LA. This right-to-left atrial gradient was decreased in pAF and cAF caused by reduced I(K,ACh) in RA only. Similarly, in SR, Kir3.1 and Kir3.4 proteins were greater in RA versus LA and decreased in RA of pAF and cAF. Kir3.1 and Kir3.4 expression was unchanged in LA of pAF and cAF.ConclusionOur results support the hypothesis that a left-to-right gradient in inward rectifier background current contributes to high-frequency sources in LA that maintain pAF. These findings have potentially important implications for development of atrial-selective therapeutic approaches.
| S-EPMC5832445 | biostudies-literature
Project description:Sensory transduction in the cochlea depends on regulated ion secretion and absorption. Results of whole-organ experiments suggested that Reissner's membrane may play a role in the control of luminal Cl(-). We tested for the presence of Cl(-) transport pathways in isolated mouse Reissner's membrane using whole-cell patch clamp recording and gene transcript analyses using RT-PCR. The current-voltage (I-V) relationship in the presence of symmetrical NMDG-Cl was strongly inward-rectifying at negative voltages, with a small outward current at positive voltages. The inward-rectifying component of the I-V curve had several properties similar to those of the ClC-2 Cl(-) channel. It was stimulated by extracellular acidity and inhibited by extracellular Cd2+, Zn2+ and intracellular ClC-2 antibody. Channel transcripts expressed include ClC-2, Slc26a7 and ClC-Ka, but not Cftr, ClC-1, ClCa1, ClCa2, ClCa3, ClCa4, Slc26a9, ClC-Kb, Best1, Best2, Best3 or the beta-subunit of ClC-K, barttin. ClC-2 is the only molecularly-identified channel present that is a strong inward rectifier. This study is the first report of conductive Cl(-) transport in epithelial cells of Reissner's membrane and is consistent with an important role in endolymph anion homeostasis.
| S-EPMC3086792 | biostudies-literature