Exenatide reduces atrial fibrillation susceptibility by inhibiting hKv1.5 and hNav1.5 channels.

Exenatide, a promising cardioprotective agent, protects against cardiac structural remodeling and diastolic dysfunction. Combined blockade of sodium and potassium channels is valuable for managing atrial fibrillation (AF). Here, we explored whether exenatide displayed anti-AF effects by inhibiting human Kv1.5 and Nav1.5 channels. We used the whole-cell patch-clamp technique to investigate the effects of exenatide on hKv1.5 and hNav1.5 channels expressed in human embryonic kidney 293 cells and studied the effects of exenatide on action potential (AP) and other cardiac ionic currents in rat atrial myocytes. Additionally, an electrical mapping system was used to explore the effects of exenatide on electrical properties and AF activity in isolated rat hearts. Finally, a rat AF model, established using acetylcholine and calcium chloride, was employed to evaluate the anti-AF potential of exenatide in rats. Exenatide reversibly suppressed IKv1.5 with IC50 of 3.08 µM, preferentially blocked the hKv1.5 channel in its closed state, and positively shifted the voltage-dependent activation curve. Exenatide also reversibly inhibited INav1.5 with IC50 of 3.30 µM, negatively shifted the voltage-dependent inactivation curve, and slowed its recovery from inactivation with significant use-dependency at 5 and 10 Hz. Furthermore, exenatide prolonged AP duration and suppressed the sustained K+ current (Iss) and transient outward K+ current (Ito), but without inhibition of L-type Ca2+ current (ICa,L) in rat atrial myocytes. Exenatide prevented AF incidence and duration in rat hearts and rats. These findings demonstrate that exenatide inhibits IKv1.5 and INav1.5in vitro and reduces AF susceptibility in isolated rat hearts and rats.

Inhibition of the voltage-gated potassium channel Kv1.5 by hydrogen sulfide attenuates remodeling through S-nitrosylation-mediated signaling.

The voltage-gated K+ channel plays a key role in atrial excitability, conducting the ultra-rapid rectifier K+ current (IKur) and contributing to the repolarization of the atrial action potential. In this study, we examine its regulation by hydrogen sulfide (H2S) in HL-1 cardiomyocytes and in HEK293 cells expressing human Kv1.5. Pacing induced remodeling resulted in shorting action potential duration, enhanced both Kv1.5 channel and H2S producing enzymes protein expression in HL-1 cardiomyocytes. H2S supplementation reduced these remodeling changes and restored action potential duration through inhibition of Kv1.5 channel. H2S also inhibited recombinant hKv1.5, lead to nitric oxide (NO) mediated S-nitrosylation and activated endothelial nitric oxide synthase (eNOS) by increased phosphorylation of Ser1177, prevention of NO formation precluded these effects. Regulation of Ikur by H2S has important cardiovascular implications and represents a novel and potential therapeutic target.

Effect of Terahertz Electromagnetic Field on the Permeability of Potassium Channel Kv1.2.

In this paper, the influence of external terahertz electromagnetic fields with different frequencies of 4 THz, 10 THz, 15 THz, and 20 THz on the permeability of the Kv1.2 voltage-gated potassium ion channel on the nerve cell membrane was studied using the combined model of the "Constant Electric Field-Ion Imbalance" method by molecular dynamics. We found that although the applied terahertz electric field does not produce strong resonance with the -C=O groups of the conservative sequence T-V-G-Y-G amino acid residue of the selective filter (SF) of the channel, it would affect the stability of the electrostatic bond between potassium ions and the carbonyl group of T-V-G-Y-G of SF, and it would affect the stability of the hydrogen bond between water molecules and oxygen atoms of the hydroxyl group of the 374THR side chain at the SF entrance, changing the potential and occupied states of ions in the SF and the occurrence probability of the permeation mode of ions and resulting in the change in the permeability of the channel. Compared with no external electric field, when the external electric field with 15 THz frequency is applied, the lifetime of the hydrogen bond is reduced by 29%, the probability of the "soft knock on" mode is decreased by 46.9%, and the ion flux of the channel is activated by 67.7%. Our research results support the view that compared to "direct knock-on", "soft knock-on" is a slower permeation mode.

Inhibitory effects of N-methyl-D-aspartate (NMDA) and α1-adrenergic receptor antagonist ifenprodil on human Kv1.5 channel.

Ifenprodil has been known to reduce cardiac contractility and cerebral vasodilation by antagonizing α1-adrenergic and N-methyl D-aspartate receptor-mediated intracellular signals. This study aimed to investigate the direct effect of ifenprodil on the human voltage-gated Kv1.5 channel (hKv1.5) by using a Xenopus oocyte expression system and a two-microelectrode voltage clamp technique. The amplitudes of hKv1.5 currents, including peak and steady state, were suppressed in a concentration-dependent manner (IC50; 43.1 and 35.5 µM, respectively) after 6 min of ifenprodil treatment. However, these effects were ~ 80% reversed by washout, suggesting that ifenprodil directly inhibited the hKv1.5 independent of membrane receptors or intracellular signals. The inhibition rate of steady state showed voltage dependence, wherein the rates increased according to test voltage depolarization. Ifenprodil reduced the time constants of hKv1.5 inactivation but has higher effects on activation. hKv1.5 inhibition by ifenprodil showed use dependency because the drug more rapidly reduced the current at the higher activation frequencies, and subsequent reduction in frequency after high activation frequency caused a partial channel block relief. Therefore, ifenprodil directly blocked the hKv1.5 in an open state and accelerated the time course of the channel inactivation, which provided a biophysical mechanism for the hKv1.5 blocking effects of ifenprodil.

Structure Elucidation of Two Intriguing Neo-Debromoaplysiatoxin Derivatives from Marine Cyanobacterium Lyngbya sp. Showing Strong Inhibition of Kv1.5 Potassium Channel and Differential Cytotoxicity.

Two aplysiatoxin derivatives, neo-debromoaplysiatoxin I (1) and neo-debromoaplysiatoxin J (2), were isolated from marine cyanobacterium Lyngbya sp. collected from the South China Sea. Their structures including absolute configurations were assigned by spectroscopic analysis, in combination with GIAO NMR shift calculation and DP4+ analysis. Structures of neo-debromoaplysiatoxin I and neo-debromoaplysiatoxin J contained a decahydro-5H-pyrano [2,3,4-de] chromen-5-one 6/6/6 ring skeleton and an intriguing peroxide bridge group, respectively, which are unprecedented structure scaffold and motif in aplysiatoxins. Two compounds displayed comparable inhibitory activities against Kv1.5 K+ channel with IC50 values of 2.59 ± 0.37 µM (1) and 1.64 ± 0.15 µM (2); however, they presented differential cytotoxic effects. It is worth noting that neo-debromoaplysiatoxin J, containing a peroxide bridge, showed remarkable cytotoxicity against four cancer cell lines including SW480, SGC7901, LoVo and PC-9 compared to the human normal cell line.

Novel Loss-of-Function KCNA5 Variants in Pulmonary Arterial Hypertension.

Reduced expression and/or activity of Kv1.5 channels (encoded by KCNA5) is a common hallmark in human or experimental pulmonary arterial hypertension (PAH). Likewise, genetic variants in KCNA5 have been found in patients with PAH, but their functional consequences and potential impact on the disease are largely unknown. Herein, this study aimed to characterize the functional consequences of seven KCNA5 variants found in a cohort of patients with PAH. Potassium currents were recorded by patch-clamp technique in HEK293 cells transfected with wild-type or mutant Kv1.5 cDNA. Flow cytometry, Western blot, and confocal microscopy techniques were used for measuring protein expression and cell apoptosis in HEK293 and human pulmonary artery smooth muscle cells. KCNA5 variants (namely, Arg184Pro and Gly384Arg) found in patients with PAH resulted in a clear loss of potassium channel function as assessed by electrophysiological and molecular modeling analyses. The Arg184Pro variant also resulted in a pronounced reduction of Kv1.5 expression. Transfection with Arg184Pro or Gly384Arg variants decreased apoptosis of human pulmonary artery smooth muscle cells compared with the wild-type cells, demonstrating that KCNA5 dysfunction in both variants affects cell viability. Thus, in addition to affecting channel activity, both variants were associated with impaired apoptosis, a crucial process linked to the disease. The estimated prevalence of dysfunctional KCNA5 variants in the PAH population analyzed was around 1%. The data indicate that some KCNA5 variants found in patients with PAH have critical consequences for channel function, supporting the idea that KCNA5 pathogenic variants may be a causative or contributing factor for PAH.

Artificial pore blocker acts specifically on voltage-gated potassium channel isoform KV1.6.

Among voltage-gated potassium channel (KV) isoforms, KV1.6 is one of the most widespread in the nervous system. However, there are little data concerning its physiological significance, in part due to the scarcity of specific ligands. The known high-affinity ligands of KV1.6 lack selectivity, and conversely, its selective ligands show low affinity. Here, we present a designer peptide with both high affinity and selectivity to KV1.6. Previously, we have demonstrated that KV isoform-selective peptides can be constructed based on the simplistic α-hairpinin scaffold, and we obtained a number of artificial Tk-hefu peptides showing selective blockage of KV1.3 in the submicromolar range. We have now proposed amino acid substitutions to enhance their activity. As a result, we have been able to produce Tk-hefu-11 that shows an EC50 of ≈70 nM against KV1.3. Quite surprisingly, Tk-hefu-11 turns out to block KV1.6 with even higher potency, presenting an EC50 of ≈10 nM. Furthermore, we have solved the peptide structure and used molecular dynamics to investigate the determinants of selective interactions between artificial α-hairpinins and KV channels to explain the dramatic increase in KV1.6 affinity. Since KV1.3 is not highly expressed in the nervous system, we hope that Tk-hefu-11 will be useful in studies of KV1.6 and its functions.

Interaction of KCNA5, CX43, and CX40 proteins in the atrial muscle of patients with atrial fibrillation.

The objective of the study was to investigate the expression levels of potassium voltage-gated channel subfamily A member 5 (KCNA5), connexin 43 (Cx43), and connexin 40 (Cx40) in the left atrial appendage of patients with atrial fibrillation (AF) and the interactions between them. We gathered tissue samples from patients with persistent AF and sinus rhythm and used fluorescence quantitative polymerase chain reaction to evaluate messenger RNA (mRNA) changes of KCNA5, Cx43, and Cx40. Then, we studied the protein levels of KCNA5, Cx43, and Cx40 by immunofluorescence and western blot analysis and the interactions between these proteins were identified by immunoprecipitation and immunofluorescence colocation, respectively. Compared with the control group, the mRNA and protein levels of KCNA5, Cx43, and Cx40 in the AF group were decreased and the positive expression of KCNA5, Cx43, and Cx40 protein was also decreased by immunofluorescence staining in the AF group. In addition, immunoprecipitation and immunofluorescence colocation revealed that KCNA5 was coexpressed with Cx43 and Cx40 proteins. The expressions of KCNA5, Cx43, and Cx40 were substantially downregulated in the myocardium of patients with AF and KCNA5 interacted with Cx43 and Cx40 proteins, respectively.

Kv1.5 channel mediates monosodium urate-induced activation of NLRP3 inflammasome in macrophages and arrhythmogenic effects of urate on cardiomyocytes.

BACKGROUND: Gout is usually found in patients with atrial fibrillation (AF). K+ efflux is a common trigger of NLRP3 inflammasome activation which is involved in the pathogenesis of AF. We investigated the role of the K+ channel Kv1.5 in monosodium urate crystal (MSU)-induced activation of the NLRP3 inflammasome and electrical remodeling in mouse and human macrophages J774.1 and THP-1, and mouse atrial myocytes HL-1. METHODS AND RESULTS: Macrophages, primed with lipopolysaccharide (LPS), were stimulated by MSU. HL-1 cells were incubated with the conditioned medium (CM) from MSU-stimulated macrophages. Western blot, ELISA and patch clamp were used. MSU induced caspase-1 expression in LPS-primed J774.1 cells and IL-1ß secretion, suggesting NLRP3 inflammasome activation. A selective Kv1.5 inhibitor, diphenyl phosphine oxide-1 (DPO-1), and siRNAs against Kv1.5 suppressed the levels of caspase-1 and IL-1ß. MSU reduced intracellular K+ concentration which was prevented by DPO-1 and siRNAs against Kv1.5. MSU increased expression of Hsp70, and Kv1.5 on the plasma membrane. siRNAs against Hsp70 were suppressed but heat shock increased the expression of Hsp70, caspase-1, IL-1ß, and Kv1.5 in MSU-stimulated J774.1 cells. The CM from MSU-stimulated macrophages enhanced the expression of caspase-1, IL-1ß and Kv1.5 with increased Kv1.5-mediated currents that shortened action potential duration in HL-1 cells. These responses were abolished by DPO-1 and a siRNA against Kv1.5. CONCLUSIONS: Kv1.5 regulates MSU-induced activation of NLRP3 inflammasome in macrophages. MSUrelated activation of NLRP3 inflammasome and electrical remodeling in HL-1 cells are via macrophages. Kv1.5 may have therapeutic value for diseases related to gout-induced activation of the NLRP3 inflammsome, including AF.

Design, synthesis, and biological evaluation of arylmethylpiperidines as Kv1.5 potassium channel inhibitors.

Kv1.5 potassium channel, encoded by KCNA5, is a promising target for the treatment of atrial fibrillation, one of the common arrhythmia. A new series of arylmethylpiperidines derivatives based on DDO-02001 were synthesised and evaluated for their ability to inhibit Kv1.5 channel. Among them, compound DDO-02005 showed good inhibitory activity (IC50 = 0.72 µM), preferable anti-arrhythmic effects and favoured safety. These results indicate that DDO-02005 can be a promising Kv1.5 inhibitor for further studies.

The voltage-gated potassium channel KV1.3 regulates neutrophil recruitment during inflammation.

AIMS: Neutrophil trafficking within the vasculature strongly relies on intracellular calcium signalling. Sustained Ca2+ influx into the cell requires a compensatory efflux of potassium to maintain membrane potential. Here, we aimed to investigate whether the voltage-gated potassium channel KV1.3 regulates neutrophil function during the acute inflammatory process by affecting sustained Ca2+ signalling. METHODS AND RESULTS: Using in vitro assays and electrophysiological techniques, we show that KV1.3 is functionally expressed in human neutrophils regulating sustained store-operated Ca2+ entry through membrane potential stabilizing K+ efflux. Inhibition of KV1.3 on neutrophils by the specific inhibitor 5-(4-Phenoxybutoxy)psoralen (PAP-1) impaired intracellular Ca2+ signalling, thereby preventing cellular spreading, adhesion strengthening, and appropriate crawling under flow conditions in vitro. Using intravital microscopy, we show that pharmacological blockade or genetic deletion of KV1.3 in mice decreased neutrophil adhesion in a blood flow dependent fashion in inflamed cremaster muscle venules. Furthermore, we identified KV1.3 as a critical component for neutrophil extravasation into the inflamed peritoneal cavity. Finally, we also revealed impaired phagocytosis of Escherichia coli particles by neutrophils in the absence of KV1.3. CONCLUSION: We show that the voltage-gated potassium channel KV1.3 is critical for Ca2+ signalling and neutrophil trafficking during acute inflammatory processes. Our findings do not only provide evidence for a role of KV1.3 for sustained calcium signalling in neutrophils affecting key functions of these cells, they also open up new therapeutic approaches to treat inflammatory disorders characterized by overwhelming neutrophil infiltration.

Absolute Structure Determination and Kv1.5 Ion Channel Inhibition Activities of New Debromoaplysiatoxin Analogues.

Potassium channel Kv1.5 has been considered a key target for new treatments of atrial tachyarrhythmias, with few side effects. Four new debromoaplysiatoxin analogues with a 6/6/12 fused ring system were isolated from marine cyanobacterium Lyngbya sp. Their planar structures were elucidated by HRESIMS, 1D and 2D NMR. The absolute configuration of oscillatoxin J (1) was determined by single-crystal X-ray diffraction, and the absolute configurations of oscillatoxin K (2), oscillatoxin L (3) and oscillatoxin M (4) were confirmed on the basis of GIAO NMR shift calculation followed by DP4 analysis. The current study confirmed the absolute configuration of the pivotal chiral positions (7S, 9S, 10S, 11R, 12S, 15S, 29R and 30R) at traditional ATXs with 6/12/6 tricyclic ring system. Compound 1, 2 and 4 exhibited blocking activities against Kv1.5 with IC50 values of 2.61 ± 0.91 µM, 3.86 ± 1.03 µM and 3.79 ± 1.01 µM, respectively. However, compound 3 exhibited a minimum effect on Kv1.5 at 10 µM. Furthermore, all of these new debromoaplysiatoxin analogs displayed no apparent activity in a brine shrimp toxicity assay.

Remodeling of Ion Channel Trafficking and Cardiac Arrhythmias.

Both inherited and acquired cardiac arrhythmias are often associated with the abnormal functional expression of ion channels at the cellular level. The complex machinery that continuously traffics, anchors, organizes, and recycles ion channels at the plasma membrane of a cardiomyocyte appears to be a major source of channel dysfunction during cardiac arrhythmias. This has been well established with the discovery of mutations in the genes encoding several ion channels and ion channel partners during inherited cardiac arrhythmias. Fibrosis, altered myocyte contacts, and post-transcriptional protein changes are common factors that disorganize normal channel trafficking during acquired cardiac arrhythmias. Channel availability, described notably for hERG and KV1.5 channels, could be another potent arrhythmogenic mechanism. From this molecular knowledge on cardiac arrhythmias will emerge novel antiarrhythmic strategies.

Transcriptomic profile of cationic channels in human pulmonary arterial hypertension.

The dysregulation of K+ channels is a hallmark of pulmonary arterial hypertension (PAH). Herein, the channelome was analyzed in lungs of patients with PAH in a public transcriptomic database. Sixty six (46%) mRNA encoding cationic channels were dysregulated in PAH with most of them downregulated (83%). The principal component analysis indicated that dysregulated cationic channel expression is a signature of the disease. Changes were very similar in idiopathic, connective tissue disease and congenital heart disease associated PAH. This analysis 1) is in agreement with the widely recognized pathophysiological role of TASK1 and KV1.5, 2) supports previous preliminary reports pointing to the dysregulation of several K+ channels including the downregulation of KV1.1, KV1.4, KV1.6, KV7.1, KV7.4, KV9.3 and TWIK2 and the upregulation of KCa1.1 and 3) points to other cationic channels dysregulated such as Kv7.3, TALK2, CaV1 and TRPV4 which might play a pathophysiological role in PAH. The significance of other changes found in Na+ and TRP channels remains to be investigated.

Atypical antipsychotic olanzapine inhibits voltage-dependent K+ channels in coronary arterial smooth muscle cells.

BACKGROUND: Olanzapine, an FDA-approved atypical antipsychotic, is widely used to treat schizophrenia and bipolar disorder. In this study, the inhibitory effect of olanzapine on voltage-dependent K+ (Kv) channels in rabbit coronary arterial smooth muscle cells was investigated. METHODS: Electrophysiological recordings were performed in freshly isolated coronary arterial smooth muscle cells. RESULTS: Olanzapine inhibited the Kv channels in a concentration-dependent manner with an IC50 value of 7.76 ± 1.80 µM and a Hill coefficient of 0.82 ± 0.09. Although olanzapine did not change the steady-state activation curve, it shifted the inactivation curve to a more negative potential, suggesting that it inhibited Kv currents by affecting the voltage sensor of the Kv channel. Application of 1 or 2 Hz train pulses did not affect the olanzapine-induced inhibition of Kv channels, suggesting that its effect on Kv channels occurs in a use (state)-independent manner. Pretreatment with DPO-1 (Kv1.5 subtype inhibitor) reduced the olanzapine-induced inhibition of Kv currents. In addition, pretreatment with guangxitoxin (Kv2.1 subtype inhibitor) and linopirdine (Kv7 subtype inhibitor) partially decreased the degree of Kv current inhibition. Olanzapine induced membrane depolarization. CONCLUSION: From these results, we suggest that olanzapine inhibits the Kv channels in a concentration-dependent, but state-independent, manner by affecting the gating properties of Kv channels. The primary Kv channel target of olanzapine is the Kv1.5 subtype.

Loperamide Toxicity Revealing Apical Hypertrophic Cardiomyopathy.

Loperamide, a µ-opioid receptor agonist, can cause cardiotoxicity by inhibiting the potassium ion channel and slowing cardiomyocyte repolarization. This, in turn, can lead to frequent early afterdepolarizations, the most common mechanism of drug-induced long QT syndrome and torsades de pointes. Apical hypertrophic cardiomyopathy (AHCM) is a nonobstructive hypertrophic cardiomyopathy rarely associated with malignant arrhythmias. We present a case of loperamide-induced malignant ventricular arrhythmia revealing underlying AHCM in a 25-year-old woman with a history of sudden cardiac arrest (SCA) and opioid use. It is important to evaluate for structural heart disease in all patients presenting with SCA, regardless of presumed etiology such as drug-induced cardiotoxicity, to prevent missed opportunities for adequate treatment. Furthermore, the diagnosis of AHCM in SCA warrants further genetic evaluation for variances with a predilection for malignant arrhythmias.

Inhibition of voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells by the class Ic antiarrhythmic agent lorcainide.

Voltage-dependent K+ (Kv) channels play the role of returning the membrane potential to the resting state, thereby maintaining the vascular tone. Here, we used native smooth-muscle cells from rabbit coronary arteries to investigate the inhibitory effect of lorcainide, a class Ic antiarrhythmic agent, on Kv channels. Lorcainide inhibited Kv channels in a concentration-dependent manner with an IC50 of 4.46 ± 0.15 µM and a Hill coefficient of 0.95 ± 0.01. Although application of lorcainide did not change the activation curve, it shifted the inactivation curve toward a more negative potential, implying that lorcainide inhibits Kv channels by changing the channels' voltage sensors. The recovery time constant from channel inactivation increased in the presence of lorcainide. Furthermore, application of train steps (of 1 or 2 Hz) in the presence of lorcainide progressively augmented the inhibition of Kv currents, implying that lorcainide-induced inhibition of Kv channels is use (state)-dependent. Pretreatment with Kv1.5 or Kv2.1/2.2 inhibitors effectively reduced the amplitude of the Kv current but did not affect the inhibitory effect of lorcainide. Based on these results, we conclude that lorcainide inhibits vascular Kv channels in a concentration and use (state)-dependent manner by changing their inactivation gating properties. Considering the clinical efficacy of lorcainide, and the pathophysiological significance of vascular Kv channels, our findings should be considered when prescribing lorcainide to patients with arrhythmia and vascular disease.

8-hydroxypinoresinol-4-O-ß-D-glucoside from Valeriana officinalis L. Is a Novel Kv1.5 Channel Blocker.

ETHNOPHARMACOLOGY RELEVANCE: In folkloric medicine of many cultures, one of the medical uses of Valeriana officinalis Linn is to treat heart-related disease. Recently, it was shown that the ethanol extracts from V. officinalis could effectively prevent auricular fibrillation, and 8-hydroxypinoresinol-4-O-ß-D-glucoside (HPG) from the extracts is one of the two active compounds showing antiarrhythmia activities. AIM OF THE STUDY: The human Kv1.5 channel (hKv1.5) has potential antiarrhythmia activities, and this study arms at investigating the current blocking effects of HPG on hKv1.5 channel. MATERIAL AND METHODS: HPG was obtained from V. officinalis extracts, and hKv1.5 channels were expressed in HEK 293 cells. HPG was perfused while recording the current through hKv1.5 channels. Patch-clamp recording techniques were used to study the effects of HPG at various concentrations (10 µM, 30 µM, and 50 µM) on hKv1.5 channels. RESULTS: The present study demonstrated that HPG inhibited hKv1.5 channel current in a concentration-dependent manner; the higher the concentration, the greater is the inhibition at each depolarization potential. During washout, the channels did not full recover indicating that the un-coupling between HPG and hKv1.5 channels is a slow process. CONCLUSION: HPG may be an effective and safe active ingredient for AF having translational potential.

Kv1.5 channels are regulated by PKC-mediated endocytic degradation.

The voltage-gated potassium channel Kv1.5 plays important roles in the repolarization of atrial action potentials and regulation of the vascular tone. While the modulation of Kv1.5 function has been well studied, less is known about how the protein levels of Kv1.5 on the cell membrane are regulated. Here, through electrophysiological and biochemical analyses of Kv1.5 channels heterologously expressed in HEK293 cells and neonatal rat ventricular myocytes, as well as native Kv1.5 in human induced pluripotent stem cell (iPSC)-derived atrial cardiomyocytes, we found that activation of protein kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA, 10 nM) diminished Kv1.5 current (IKv1.5) and protein levels of Kv1.5 in the plasma membrane. Mechanistically, PKC activation led to monoubiquitination and degradation of the mature Kv1.5 proteins. Overexpression of Vps24, a protein that sorts transmembrane proteins into lysosomes via the multivesicular body (MVB) pathway, accelerated, whereas the lysosome inhibitor bafilomycin A1 completely prevented PKC-mediated Kv1.5 degradation. Kv1.5, but not Kv1.1, Kv1.2, Kv1.3, or Kv1.4, was uniquely sensitive to PMA treatment. Sequence alignments suggested that residues within the N terminus of Kv1.5 are essential for PKC-mediated Kv1.5 reduction. Using N-terminal truncation as well as site-directed mutagenesis, we identified that Thr15 is the target site for PKC that mediates endocytic degradation of Kv1.5 channels. These findings indicate that alteration of protein levels in the plasma membrane represents an important regulatory mechanism of Kv1.5 channel function under PKC activation conditions.