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Sample records for cell potassium channels

  1. KV7 potassium channels

    DEFF Research Database (Denmark)

    Stott, Jennifer B; Jepps, Thomas Andrew; Greenwood, Iain A

    2014-01-01

    Potassium channels are key regulators of smooth muscle tone, with increases in activity resulting in hyperpolarisation of the cell membrane, which acts to oppose vasoconstriction. Several potassium channels exist within smooth muscle, but the KV7 family of voltage-gated potassium channels have been...

  2. Molecular basis of potassium channels in pancreatic duct epithelial cells

    DEFF Research Database (Denmark)

    Hayashi, M.; Novak, Ivana

    2013-01-01

    Potassium channels regulate excitability, epithelial ion transport, proliferation, and apoptosis. In pancreatic ducts, K channels hyperpolarize the membrane potential and provide the driving force for anion secretion. This review focuses on the molecular candidates of functional K channels...... other cell types, preferably in epithelia, and, where known, their identification and functions in pancreatic ducts and in adenocarcinoma cells. We conclude by pointing out some outstanding questions and future directions in pancreatic K channel research with respect to the physiology of secretion...

  3. Cell-based potassium ion channel screening using the FluxOR assay.

    Science.gov (United States)

    Beacham, Daniel W; Blackmer, Trillium; O' Grady, Michael; Hanson, George T

    2010-04-01

    FluxOR technology is a cell-based assay used for high-throughput screening measurements of potassium channel activity. Using thallium influx as a surrogate indicator of potassium ion channel activity, the FluxOR Potassium Ion Channel Assay is based on the activation of a novel fluorescent dye. This indicator reports channel activity with a large fluorogenic response and is proportional to the number of open potassium channels on the cell, making it extremely useful for studying K(+) channel targets. In contrast to BTC-AM ester, FluxOR dye is roughly 10-fold more thallium sensitive, requiring much lower thallium for a larger signal window. This also means that the assay is carried out in a physiological, normal-chloride saline. In this article, the authors describe how they used BacMam gene delivery to express Kv7.2 and 7.3 (KCNQ), Kir2.1, or Kv11.1 (hERG) potassium ion channels in U2-OS cells. Using these cells, they ran the FluxOR assay to identify and characterize channel-specific inhibitory compounds discovered within the library (Tocriscreen Mini 1200 and Sigma Sodium/Potassium Modulators Ligand set). The FluxOR assay was able to identify several known specific inhibitors of Kv7.2/7.3 or hERG, highlighting its potential to identify novel and more efficacious small-molecule modulators.

  4. Cholesterol influences voltage-gated calcium channels and BK-type potassium channels in auditory hair cells.

    Directory of Open Access Journals (Sweden)

    Erin K Purcell

    Full Text Available The influence of membrane cholesterol content on a variety of ion channel conductances in numerous cell models has been shown, but studies exploring its role in auditory hair cell physiology are scarce. Recent evidence shows that cholesterol depletion affects outer hair cell electromotility and the voltage-gated potassium currents underlying tall hair cell development, but the effects of cholesterol on the major ionic currents governing auditory hair cell excitability are unknown. We investigated the effects of a cholesterol-depleting agent (methyl beta cyclodextrin, MβCD on ion channels necessary for the early stages of sound processing. Large-conductance BK-type potassium channels underlie temporal processing and open in a voltage- and calcium-dependent manner. Voltage-gated calcium channels (VGCCs are responsible for calcium-dependent exocytosis and synaptic transmission to the auditory nerve. Our results demonstrate that cholesterol depletion reduced peak steady-state calcium-sensitive (BK-type potassium current by 50% in chick cochlear hair cells. In contrast, MβCD treatment increased peak inward calcium current (~30%, ruling out loss of calcium channel expression or function as a cause of reduced calcium-sensitive outward current. Changes in maximal conductance indicated a direct impact of cholesterol on channel number or unitary conductance. Immunoblotting following sucrose-gradient ultracentrifugation revealed BK expression in cholesterol-enriched microdomains. Both direct impacts of cholesterol on channel biophysics, as well as channel localization in the membrane, may contribute to the influence of cholesterol on hair cell physiology. Our results reveal a new role for cholesterol in the regulation of auditory calcium and calcium-activated potassium channels and add to the growing evidence that cholesterol is a key determinant in auditory physiology.

  5. Differentially Expressed Potassium Channels Are Associated with Function of Human Effector Memory CD8+T Cells.

    Science.gov (United States)

    Sim, Ji Hyun; Kim, Kyung Soo; Park, Hyoungjun; Kim, Kyung-Jin; Lin, Haiyue; Kim, Tae-Joo; Shin, Hyun Mu; Kim, Gwanghun; Lee, Dong-Sup; Park, Chan-Wook; Lee, Dong Hun; Kang, Insoo; Kim, Sung Joon; Cho, Chung-Hyun; Doh, Junsang; Kim, Hang-Rae

    2017-01-01

    The voltage-gated potassium channel, Kv1.3, and the Ca 2+ -activated potassium channel, KCa3.1, regulate membrane potentials in T cells, thereby controlling T cell activation and cytokine production. However, little is known about the expression and function of potassium channels in human effector memory (EM) CD8 + T cells that can be further divided into functionally distinct subsets based on the expression of the interleukin (IL)-7 receptor alpha (IL-7Rα) chain. Herein, we investigated the functional expression and roles of Kv1.3 and KCa3.1 in EM CD8 + T cells that express high or low levels of the IL-7 receptor alpha chain (IL-7Rα high and IL-7Rα low , respectively). In contrast to the significant activity of Kv1.3 and KCa3.1 in IL-7Rα high EM CD8 + T cells, IL-7Rα low EM CD8 + T cells showed lower expression of Kv1.3 and insignificant expression of KCa3.1. Kv1.3 was involved in the modulation of cell proliferation and IL-2 production, whereas KCa3.1 affected the motility of EM CD8 + T cells. The lower motility of IL-7Rα low EM CD8 + T cells was demonstrated using transendothelial migration and motility assays with intercellular adhesion molecule 1- and/or chemokine stromal cell-derived factor-1α-coated surfaces. Consistent with the lower migration property, IL-7Rα low EM CD8 + T cells were found less frequently in human skin. Stimulating IL-7Rα low EM CD8 + T cells with IL-2 or IL-15 increased their motility and recovery of KCa3.1 activity. Our findings demonstrate that Kv1.3 and KCa3.1 are differentially involved in the functions of EM CD8 + T cells. The weak expression of potassium channels in IL-7Rα low EM CD8 + T cells can be revived by stimulation with IL-2 or IL-15, which restores the associated functions. This study suggests that IL-7Rα high EM CD8 + T cells with functional potassium channels may serve as a reservoir for effector CD8 + T cells during peripheral inflammation.

  6. Cardiac potassium channel subtypes

    DEFF Research Database (Denmark)

    Schmitt, Nicole; Grunnet, Morten; Olesen, Søren-Peter

    2014-01-01

    . The underlying posttranscriptional and posttranslational remodeling of the individual K(+) channels changes their activity and significance relative to each other, and they must be viewed together to understand their role in keeping a stable heart rhythm, also under menacing conditions like attacks of reentry......About 10 distinct potassium channels in the heart are involved in shaping the action potential. Some of the K(+) channels are primarily responsible for early repolarization, whereas others drive late repolarization and still others are open throughout the cardiac cycle. Three main K(+) channels...... that they could constitute targets for new pharmacological treatment of atrial fibrillation. The interplay between the different K(+) channel subtypes in both atria and ventricle is dynamic, and a significant up- and downregulation occurs in disease states such as atrial fibrillation or heart failure...

  7. Development of New Openers of ATP-Sensitive Potassium Channels of the Cell Membranes

    Directory of Open Access Journals (Sweden)

    Strutynskyi, R.B.

    2016-07-01

    Full Text Available Two innovative libraries (413 cyclosulfamides and 709 orthopyridine sulfamides of potential new openers of ATP-sensitive potassium channels of cell membranes were developed. It is shown experimentally that at least ten new original compounds have properties of pharmacological openers of the channels. Seven compounds, namely Z851154982, Z56762024, Z1269122570, Z31153162, Z45679561, Z756371174 and Z649723638, open channels of both types — sarcoplasmic as well as mitochondrial membranes: Simultaneously, Z734043408 compound is a potent activator of aforementioned channels of sarcolemmal membrane only. Z31197374 and Z666664306 compounds show the affinity onlyto КATP-channels of mitochondrial type. The results of the work can be used by pharmaceutical companies and scientific research institutes.

  8. The Arabidopsis guard cell outward potassium channel GORK is regulated by CPK33.

    Science.gov (United States)

    Corratgé-Faillie, Claire; Ronzier, Elsa; Sanchez, Frédéric; Prado, Karine; Kim, Jeong-Hyeon; Lanciano, Sophie; Leonhardt, Nathalie; Lacombe, Benoît; Xiong, Tou Cheu

    2017-07-01

    A complex signaling network involving voltage-gated potassium channels from the Shaker family contributes to the regulation of stomatal aperture. Several kinases and phosphatases have been shown to be crucial for ABA-dependent regulation of the ion transporters. To date, the Ca 2+ -dependent regulation of Shaker channels by Ca 2+ -dependent protein kinases (CPKs) is still elusive. A functional screen in Xenopus oocytes was launched to identify such CPKs able to regulate the three main guard cell Shaker channels KAT1, KAT2, and GORK. Seven guard cell CPKs were tested and multiple CPK/Shaker couples were identified. Further work on CPK33 indicates that GORK activity is enhanced by CPK33 and unaffected by a nonfunctional CPK33 (CPK33-K102M). Furthermore, Ca 2+ -induced stomatal closure is impaired in two cpk33 mutant plants. © 2017 Federation of European Biochemical Societies.

  9. Multi-walled carbon nanotubes suppress potassium channel activities in PC12 cells

    Science.gov (United States)

    Xu, Haifei; Bai, Juan; Meng, Jie; Hao, Wei; Xu, Haiyan; Cao, Ji-Min

    2009-07-01

    The advancement in nanotechnology has produced technological and conceptual breakthroughs but the effects nanomaterials have on organisms at the cellular level are poorly understood. Here we report that carboxyl-terminated multi-walled carbon nanotubes (MWCNTs) act as antagonists of three types of potassium channels as assessed by whole-cell patch clamp electrophysiology on undifferentiated pheochromocytoma (PC12) cells. Our results showed that carboxyl-terminated MWCNTs suppress the current densities of Ito, IK and IK1 in a time-dependent and irreversible manner. The suppressions were most distinct 24 h after incubation with MWCNTs. However, MWCNTs did not significantly change the expression levels of reactive oxygen species (ROS) or intracellular free calcium and also did not alter the mitochondrial membrane potential (ΔΨm) in PC12 cells. These results suggest that oxidative stress was not involved in the MWCNTs suppression of Ito, IK and IK1 current densities. Nonetheless, the suppression of potassium currents by MWCNTs will impact on electrical signaling of excitable cells such as neurons and muscles.

  10. Potassium Channels in Neurofbromatosis-1

    National Research Council Canada - National Science Library

    Chen, Mingkui

    2006-01-01

    .... We were the first to investigate potential mechanisms of cognitive impairment in NF-1 at the molecular level involving potassium channels, and demonstrated a possible mechanism for the learning deficits seen in NF1...

  11. A thallium-sensitive, fluorescence-based assay for detecting and characterizing potassium channel modulators in mammalian cells.

    Science.gov (United States)

    Weaver, C David; Harden, David; Dworetzky, Steven I; Robertson, Barbara; Knox, Ronald J

    2004-12-01

    Potassium channels have been identified as targets for a large number of therapeutic indications. The ability to use a high-throughput functional assay for the detection and characterization of small-molecule modulators of potassium channels is very desirable. However, present techniques capable of screening very large chemical libraries are limited in terms of data quality, temporal resolution, ease of use, and requirements for specialized instrumentation. To address these issues, the authors have developed a fluorescence-based thallium flux assay. This assay is capable of detecting modulators of both voltage and ligand-gated potassium channels expressed in mammalian cells. The thallium flux assay can use instruments standard to most high-throughput screening laboratories, and using such equipment has been successfully employed to screen large chemical libraries consisting of hundreds of thousands of compounds.

  12. Functional coupling between large-conductance potassium channels and Cav3.2 voltage-dependent calcium channels participates in prostate cancer cell growth

    Directory of Open Access Journals (Sweden)

    Florian Gackière

    2013-07-01

    It is strongly suspected that potassium (K+ channels are involved in various aspects of prostate cancer development, such as cell growth. However, the molecular nature of those K+ channels implicated in prostate cancer cell proliferation and the mechanisms through which they control proliferation are still unknown. This study uses pharmacological, biophysical and molecular approaches to show that the main voltage-dependent K+ current in prostate cancer LNCaP cells is carried by large-conductance BK channels. Indeed, most of the voltage-dependent current was inhibited by inhibitors of BK channels (paxillin and iberiotoxin and by siRNA targeting BK channels. In addition, we reveal that BK channels constitute the main K+ channel family involved in setting the resting membrane potential in LNCaP cells at around −40 mV. This consequently promotes a constitutive calcium entry through T-type Cav3.2 calcium channels. We demonstrate, using single-channel recording, confocal imaging and co-immunoprecipitation approaches, that both channels form macromolecular complexes. Finally, using flow cytometry cell cycle measurements, cell survival assays and Ki67 immunofluorescent staining, we show that both BK and Cav3.2 channels participate in the proliferation of prostate cancer cells.

  13. Basolateral localisation of KCNQ1 potassium channels in MDCK cells: molecular identification of an N-terminal targeting motif

    DEFF Research Database (Denmark)

    Jespersen, Thomas; Rasmussen, Hanne B; Grunnet, Morten

    2004-01-01

    of the tyrosine residue at position 51 resulted in a non-polarized steady-state distribution of the channel. The importance of tyrosine 51 in basolateral localisation was emphasized by the fact that a short peptide comprising this tyrosine was able to redirect the p75 neurotrophin receptor, an otherwise apically......KCNQ1 potassium channels are expressed in many epithelial tissues as well as in the heart. In epithelia KCNQ1 channels play an important role in salt and water transport and the channel has been reported to be located apically in some cell types and basolaterally in others. Here we show that KCNQ1...... channels are located basolaterally when expressed in polarised MDCK cells. The basolateral localisation of KCNQ1 is not affected by co-expression of any of the five KCNE beta-subunits. We characterise two independent basolateral sorting signals present in the N-terminal tail of KCNQ1. Mutation...

  14. Molecular and functional identification of cyclic AMP-sensitive BKCa potassium channels (ZERO variant) and L-type voltage-dependent calcium channels in single rat juxtaglomerular cells

    DEFF Research Database (Denmark)

    Friis, Ulla G; Jørgensen, Finn; Andreasen, Ditte

    2003-01-01

    This study aimed at identifying the type and functional significance of potassium channels and voltage-dependent calcium channels (Ca(v)) in single rat JG cells using whole-cell patch clamp. Single JG cells displayed outward rectification at positive membrane potentials and limited net currents......, respectively. Double immunofluorescence confirmed the presence of BKCa and renin in the same cell. cAMP increased the outward current by 1.6-fold, and this was inhibited by 74% with iberiotoxin. Expression of the cAMP-sensitive splice variant (ZERO) of BKCa was confirmed in single-sampled JG cells by RT...... no effect. We conclude that JG cells express functional cAMP-sensitive BKCa channels (the ZERO splice variant) and voltage-dependent L-type Ca2+ channels....

  15. Silencing of Kv4.1 potassium channels inhibits cell proliferation of tumorigenic human mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Soo Hwa [Laboratories of Veterinary Pharmacology, College of Veterinary Medicine, Seoul National University, San 56-1 Sillim-Dong Kwanak-Gu, Seoul 151-742 (Korea, Republic of); Choi, Changsun [Department of Food and Nutrition, College of Human Ecology, Chung-Ang University, Anseong, Gyeonggi (Korea, Republic of); Hong, Seong-Geun; Yarishkin, Oleg V. [Department of Physiology, College of Medicine, Gyeongsang National University, Jinju (Korea, Republic of); Bae, Young Min; Kim, Jae Gon [Department of Physiology, College of Medicine, Konkuk University, Seoul (Korea, Republic of); O' Grady, Scott M. [Department of Physiology, 495 Animal Science/Veterinary Medicine Bldg., St. Paul, University of Minnesota, MN (United States); Yoon, Kyong-Ah [Research Institute and Hospital, National Cancer Center, Goyang, Gyeonggi (Korea, Republic of); Kang, Kyung-Sun [Veterinary Public Health, College of Veterinary Medicine, Seoul National University, San 56-1 Sillim-Dong Kwanak-Gu, Seoul (Korea, Republic of); Ryu, Pan Dong [Laboratories of Veterinary Pharmacology, College of Veterinary Medicine, Seoul National University, San 56-1 Sillim-Dong Kwanak-Gu, Seoul 151-742 (Korea, Republic of); Lee, So Yeong, E-mail: leeso@snu.ac.kr [Laboratories of Veterinary Pharmacology, College of Veterinary Medicine, Seoul National University, San 56-1 Sillim-Dong Kwanak-Gu, Seoul 151-742 (Korea, Republic of)

    2009-06-26

    Potassium channel activity has been shown to facilitate cell proliferation in cancer cells. In the present study, the role of Kv4.1 channels in immortal and tumorigenic human mammary epithelial cells was investigated. Kv4.1 protein expression was positively correlated with tumorigenicity. Moreover, transfection with siRNAs targeting Kv4.1 mRNA suppressed proliferation of tumorigenic mammary epithelial cells. Experiments using mRNA isolated from human breast cancer tissues revealed that the level of Kv4.1 mRNA expression varied depending on the stage of the tumor. Kv4.1 protein expression increased during stages T2 and T3 compared to normal tissue. These results demonstrated that Kv4.1 plays a role in proliferation of tumorigenic human mammary epithelial cells. In addition, elevated Kv4.1 expression may be useful as a diagnostic marker for staging mammary tumors and selective blockers of Kv4.1 may serve to suppress tumor cell proliferation.

  16. Functional coupling between sodium-activated potassium channels and voltage-dependent persistent sodium currents in cricket Kenyon cells.

    Science.gov (United States)

    Takahashi, Izumi; Yoshino, Masami

    2015-10-01

    In this study, we examined the functional coupling between Na(+)-activated potassium (KNa) channels and Na(+) influx through voltage-dependent Na(+) channels in Kenyon cells isolated from the mushroom body of the cricket Gryllus bimaculatus. Single-channel activity of KNa channels was recorded with the cell-attached patch configuration. The open probability (Po) of KNa channels increased with increasing Na(+) concentration in a bath solution, whereas it decreased by the substitution of Na(+) with an equimolar concentration of Li(+). The Po of KNa channels was also found to be reduced by bath application of a high concentration of TTX (1 μM) and riluzole (100 μM), which inhibits both fast (INaf) and persistent (INaP) Na(+) currents, whereas it was unaffected by a low concentration of TTX (10 nM), which selectively blocks INaf. Bath application of Cd(2+) at a low concentration (50 μM), as an inhibitor of INaP, also decreased the Po of KNa channels. Conversely, bath application of the inorganic Ca(2+)-channel blockers Co(2+) and Ni(2+) at high concentrations (500 μM) had little effect on the Po of KNa channels, although Cd(2+) (500 μM) reduced the Po of KNa channels. Perforated whole cell clamp analysis further indicated the presence of sustained outward currents for which amplitude was dependent on the amount of Na(+) influx. Taken together, these results indicate that KNa channels could be activated by Na(+) influx passing through voltage-dependent persistent Na(+) channels. The functional significance of this coupling mechanism was discussed in relation to the membrane excitability of Kenyon cells and its possible role in the formation of long-term memory. Copyright © 2015 the American Physiological Society.

  17. A novel potassium channel in photosynthetic cyanobacteria.

    Directory of Open Access Journals (Sweden)

    Manuela Zanetti

    Full Text Available Elucidation of the structure-function relationship of a small number of prokaryotic ion channels characterized so far greatly contributed to our knowledge on basic mechanisms of ion conduction. We identified a new potassium channel (SynK in the genome of the cyanobacterium Synechocystis sp. PCC6803, a photosynthetic model organism. SynK, when expressed in a K(+-uptake-system deficient E. coli strain, was able to recover growth of these organisms. The protein functions as a potassium selective ion channel when expressed in Chinese hamster ovary cells. The location of SynK in cyanobacteria in both thylakoid and plasmamembranes was revealed by immunogold electron microscopy and Western blotting of isolated membrane fractions. SynK seems to be conserved during evolution, giving rise to a TPK (two-pore K(+ channel family member which is shown here to be located in the thylakoid membrane of Arabidopsis. Our work characterizes a novel cyanobacterial potassium channel and indicates the molecular nature of the first higher plant thylakoid cation channel, opening the way to functional studies.

  18. Osteopontin activates the diabetes-associated potassium channel TALK-1 in pancreatic β-cells.

    Directory of Open Access Journals (Sweden)

    Matthew T Dickerson

    Full Text Available Glucose-stimulated insulin secretion (GSIS relies on β-cell Ca2+ influx, which is modulated by the two-pore-domain K+ (K2P channel, TALK-1. A gain-of-function polymorphism in KCNK16, the gene encoding TALK-1, increases risk for developing type-2 diabetes. While TALK-1 serves an important role in modulating GSIS, the regulatory mechanism(s that control β-cell TALK-1 channels are unknown. Therefore, we employed a membrane-specific yeast two-hybrid (MYTH assay to identify TALK-1-interacting proteins in human islets, which will assist in determining signaling modalities that modulate TALK-1 function. Twenty-one proteins from a human islet cDNA library interacted with TALK-1. Some of these interactions increased TALK-1 activity, including intracellular osteopontin (iOPN. Intracellular OPN is highly expressed in β-cells and is upregulated under pre-diabetic conditions to help maintain normal β-cell function; however, the functional role of iOPN in β-cells is poorly understood. We found that iOPN colocalized with TALK-1 in pancreatic sections and coimmunoprecipitated with human islet TALK-1 channels. As human β-cells express two K+ channel-forming variants of TALK-1, regulation of these TALK-1 variants by iOPN was assessed. At physiological voltages iOPN activated TALK-1 transcript variant 3 channels but not TALK-1 transcript variant 2 channels. Activation of TALK-1 channels by iOPN also hyperpolarized resting membrane potential (Vm in HEK293 cells and in primary mouse β-cells. Intracellular OPN was also knocked down in β-cells to test its effect on β-cell TALK-1 channel activity. Reducing β-cell iOPN significantly decreased TALK-1 K+ currents and increased glucose-stimulated Ca2+ influx. Importantly, iOPN did not affect the function of other K2P channels or alter Ca2+ influx into TALK-1 deficient β-cells. These results reveal the first protein interactions with the TALK-1 channel and found that an interaction with iOPN increased β-cell

  19. ATP-sensitive potassium channel: a novel target for protection against UV-induced human skin cell damage.

    Science.gov (United States)

    Cao, Cong; Healey, Sarah; Amaral, Ashley; Lee-Couture, Avery; Wan, Shu; Kouttab, Nicola; Chu, Wenming; Wan, Yinsheng

    2007-07-01

    Ultraviolet radiation (UV) induces cell damages leading to skin photoaging and skin cancer. ATP-sensitive potassium (K(ATP)) channel openers (KCOs) have been shown to exert significant myocardial preservation and neuroprotection in vitro and in vivo, and yet the potential role of those KCOs in protection against UV-induced skin cell damage is unknown. We investigated the effects of pinacidil and diazoxide, two classical KCOs, on UV-induced cell death using cultured human keratinocytes (HaCat cells). Here, we demonstrated for the first time that Kir 6.1, Kir 6.2 and SUR2 subunits of K(ATP) channels are functionally expressed in HaCaT cells and both non-selective K(ATP) channel opener pinacidil and mitoK(ATP) (mitochondrial K(ATP)) channel opener diazoxide attenuated UV-induced keratinocytes cell death. The protective effects were abolished by both non-selective K(ATP) channel blocker glibenclamide and selective mitoK(ATP) channel blocker 5-hydroxydecanoate (5-HD). Also, activation of K(ATP) channel with pinacidil or diazoxide resulted in suppressive effects on UV-induced MAPK activation and reactive oxygen species (ROS) production. Unexpectedly, we found that the level of intracellular ROS was slightly elevated in HaCaT cells when treated with pinacidil or diazoxide alone. Furthermore, UV-induced mitochondrial membrane potential loss, cytochrome c release and ultimately apoptotic cell death were also inhibited by preconditioning with pinacidil and diazoxide, and their effects were reversed by glibenclamide and 5-HD. Taken together, we contend that mitoK(ATP) is likely to contribute the protection against UV-induced keratinocytes cell damage. Our findings suggest that K(ATP) openers such as pinacidil and diazoxide may be utilized to prevent from UV-induced skin aging.

  20. Calcitriol inhibits Ether-a go-go potassium channel expression and cell proliferation in human breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Garcia-Becerra, Rocio [Department of Reproductive Biology, Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Vasco de Quiroga No. 15, Tlalpan 14000 Mexico, D.F. (Mexico); Diaz, Lorenza, E-mail: lorenzadiaz@gmail.com [Department of Reproductive Biology, Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Vasco de Quiroga No. 15, Tlalpan 14000 Mexico, D.F. (Mexico); Camacho, Javier [Department of Pharmacology, Centro de Investigacion y de Estudios Avanzados, Instituto Politecnico Nacional, Av. Instituto Politecnico Nacional 2508, San Pedro Zacatenco 07360, Mexico, D.F. (Mexico); Barrera, David; Ordaz-Rosado, David; Morales, Angelica [Department of Reproductive Biology, Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Vasco de Quiroga No. 15, Tlalpan 14000 Mexico, D.F. (Mexico); Ortiz, Cindy Sharon [Department of Pathology, Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Vasco de Quiroga No. 15, Tlalpan 14000 Mexico, D.F. (Mexico); Avila, Euclides [Department of Reproductive Biology, Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Vasco de Quiroga No. 15, Tlalpan 14000 Mexico, D.F. (Mexico); Bargallo, Enrique [Department of Breast Tumors, Instituto Nacional de Cancerologia, Av. San Fernando No. 22, Tlalpan 14080, Mexico, D.F. (Mexico); Arrecillas, Myrna [Department of Pathology, Instituto Nacional de Cancerologia, Av. San Fernando No. 22, Tlalpan 14080, Mexico, D.F. (Mexico); Halhali, Ali; Larrea, Fernando [Department of Reproductive Biology, Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Vasco de Quiroga No. 15, Tlalpan 14000 Mexico, D.F. (Mexico)

    2010-02-01

    Antiproliferative actions of calcitriol have been shown to occur in many cell types; however, little is known regarding the molecular basis of this process in breast carcinoma. Ether-a-go-go (Eag1) potassium channels promote oncogenesis and are implicated in breast cancer cell proliferation. Since calcitriol displays antineoplastic effects while Eag1 promotes tumorigenesis, and both factors antagonically regulate cell cycle progression, we investigated a possible regulatory effect of calcitriol upon Eag1 as a mean to uncover new molecular events involved in the antiproliferative activity of this hormone in human breast tumor-derived cells. RT real-time PCR and immunocytochemistry showed that calcitriol suppressed Eag1 expression by a vitamin D receptor (VDR)-dependent mechanism. This effect was accompanied by inhibition of cell proliferation, which was potentiated by astemizole, a nonspecific Eag1 inhibitor. Immunohistochemistry and Western blot demonstrated that Eag1 and VDR abundance was higher in invasive-ductal carcinoma than in fibroadenoma, and immunoreactivity of both proteins was located in ductal epithelial cells. Our results provide evidence of a novel mechanism involved in the antiproliferative effects of calcitriol and highlight VDR as a cancer therapeutic target for breast cancer treatment and prevention.

  1. Calcitriol inhibits Ether-a go-go potassium channel expression and cell proliferation in human breast cancer cells

    International Nuclear Information System (INIS)

    Garcia-Becerra, Rocio; Diaz, Lorenza; Camacho, Javier; Barrera, David; Ordaz-Rosado, David; Morales, Angelica; Ortiz, Cindy Sharon; Avila, Euclides; Bargallo, Enrique; Arrecillas, Myrna; Halhali, Ali; Larrea, Fernando

    2010-01-01

    Antiproliferative actions of calcitriol have been shown to occur in many cell types; however, little is known regarding the molecular basis of this process in breast carcinoma. Ether-a-go-go (Eag1) potassium channels promote oncogenesis and are implicated in breast cancer cell proliferation. Since calcitriol displays antineoplastic effects while Eag1 promotes tumorigenesis, and both factors antagonically regulate cell cycle progression, we investigated a possible regulatory effect of calcitriol upon Eag1 as a mean to uncover new molecular events involved in the antiproliferative activity of this hormone in human breast tumor-derived cells. RT real-time PCR and immunocytochemistry showed that calcitriol suppressed Eag1 expression by a vitamin D receptor (VDR)-dependent mechanism. This effect was accompanied by inhibition of cell proliferation, which was potentiated by astemizole, a nonspecific Eag1 inhibitor. Immunohistochemistry and Western blot demonstrated that Eag1 and VDR abundance was higher in invasive-ductal carcinoma than in fibroadenoma, and immunoreactivity of both proteins was located in ductal epithelial cells. Our results provide evidence of a novel mechanism involved in the antiproliferative effects of calcitriol and highlight VDR as a cancer therapeutic target for breast cancer treatment and prevention.

  2. Cloning and expression analysis of potassium channel gene NKT3 ...

    African Journals Online (AJOL)

    Potassium (K+) is the predominant inorganic ion of plant cells. K+ channels in higher plant cells play an important role in regulating the influx and efflux of K+ from cells, and activity of these channels might be involved in plant stress resistance. A completely new K+ channel gene of Nicotiana tabacum was obtained through ...

  3. Intractable hyperkalemia due to nicorandil induced potassium channel syndrome

    Directory of Open Access Journals (Sweden)

    Vivek Chowdhry

    2015-01-01

    Full Text Available Nicorandil is a commonly used antianginal agent, which has both nitrate-like and ATP-sensitive potassium (K ATP channel activator properties. Activation of potassium channels by nicorandil causes expulsion of potassium ions into the extracellular space leading to membrane hyperpolarization, closure of voltage-gated calcium channels and finally vasodilatation. However, on the other hand, being an activator of K ATP channel, it can expel K + ions out of the cells and can cause hyperkalemia. Here, we report a case of nicorandil induced hyperkalemia unresponsive to medical treatment in a patient with diabetic nephropathy.

  4. Inhibition of UV-B induced apoptosis in corneal epithelial cells by potassium channel modulators.

    Science.gov (United States)

    Ubels, John L; Schotanus, Mark P; Bardolph, Susan L; Haarsma, Loren D; Koetje, Leah R; Louters, Julienne R

    2010-02-01

    The goal of this study was to determine whether prevention of K(+) loss can protect human corneal-limbal epithelial (HCLE) cells from UV-B induced apoptosis. Immunostaining for activated caspase-3 of HCLE cells exposed to 150-200 mJ/cm(2) UV-B demonstrated induction of apoptosis 6 h after exposure. The number of apoptotic cells was decreased by incubation in medium with 25 or 100 mM K(+). If this protection is due to a reduction of UV-induced K(+) loss then K(+) channel blockers should also protect HCLE cells from UV-B. Caspase-8 activity induced by exposure to UV-B at 150 mJ/cm(2) was significantly reduced when the cells were incubated in 0.3 microM BDS-I or 0.05-1 mM quinidine. Caspase-3 was also activated by UV-B and a reduction in activity was observed after incubation in 0.1-0.3 microM BDS-I and 0.1-1 mM quinidine. Induction of DNA fragmentation, as measured by the TUNEL assay, was decreased by treatment with 0.3 microM BDS-I and 0.01-0.05 mM quinidine. Patch-clamp recording showed activation of K(+) channels after exposure to UV-B and a decrease in outward K(+) current was observed following application of BDS-I. Quinidine did not block K(+) currents in HCLE cells, suggesting that the protective effect of quinidine occurs by a mechanism other than via K(+) channels. The effect of the K(+) channel blocker BDS-1 on HCLE cells exposed to UV-B confirms that preventing K(+) efflux protects corneal epithelial cells from apoptosis. This suggests the elevated [K(+)] in tears may protect the corneal epithelium from effects of ambient UV-B. Copyright 2009 Elsevier Ltd. All rights reserved.

  5. Inhibition of UV-B Induced Apoptosis in Corneal Epithelial Cells by Potassium Channel Modulators

    Science.gov (United States)

    Ubels, John L.; Schotanus, Mark P.; Bardolph, Susan L.; Haarsma, Loren D.; Koetje, Leah R.; Louters, Julienne R.

    2009-01-01

    The goal of this study was to determine whether prevention of K+ loss can protect human corneal-limbal epithelial (HCLE) cells from UV-B induced apoptosis. Immunostaining for activated caspase-3 of HCLE cells exposed to 150 – 200 mJ/cm2 UV-B demonstrated induction of apoptosis 6 hrs after exposure. The number of apoptotic cells was decreased by incubation in medium with 25 or 100 mM K+. If this protection is due to a reduction of UV induced K+ loss then K+ channel blockers should also protect HCLE cells from UV-B. Caspase-8 activity induced by exposure to UV-B at 150 mJ/cm2 was significantly reduced when the cells were incubated in 0.3 µM BDS-I or 0.05–1 mM quinidine. Caspase-3 was also activated by UV-B and a reduction in activity was observed after incubation in 0.1–0.3 µM BDS-I and 0.1–1mM quinidine. Induction of DNA fragmentation, as measured by the TUNEL assay, was decreased by treatment with 0.3 µM BDS-I and 0.01–0.05 mM quinidine. Patch-clamp recording showed activation of K+ channels after exposure to UV-B and a decrease in outward K+ current was observed following application of BDS-I. Quinidine did not block K+ currents in HCLE cells, suggesting that the protective effect of quinidine occurs by a mechanism other than via K+ channels. The effect of the K+ channel blocker BDS-1 on HCLE cells exposed to UV-B confirms that preventing K+ efflux protects corneal epithelial cells from apoptosis. This suggests the elevated [K+] in tears may protect the corneal epithelium from effects of ambient UV-B. PMID:19874821

  6. VKCDB: Voltage-gated potassium channel database

    Directory of Open Access Journals (Sweden)

    Gallin Warren J

    2004-01-01

    Full Text Available Abstract Background The family of voltage-gated potassium channels comprises a functionally diverse group of membrane proteins. They help maintain and regulate the potassium ion-based component of the membrane potential and are thus central to many critical physiological processes. VKCDB (Voltage-gated potassium [K] Channel DataBase is a database of structural and functional data on these channels. It is designed as a resource for research on the molecular basis of voltage-gated potassium channel function. Description Voltage-gated potassium channel sequences were identified by using BLASTP to search GENBANK and SWISSPROT. Annotations for all voltage-gated potassium channels were selectively parsed and integrated into VKCDB. Electrophysiological and pharmacological data for the channels were collected from published journal articles. Transmembrane domain predictions by TMHMM and PHD are included for each VKCDB entry. Multiple sequence alignments of conserved domains of channels of the four Kv families and the KCNQ family are also included. Currently VKCDB contains 346 channel entries. It can be browsed and searched using a set of functionally relevant categories. Protein sequences can also be searched using a local BLAST engine. Conclusions VKCDB is a resource for comparative studies of voltage-gated potassium channels. The methods used to construct VKCDB are general; they can be used to create specialized databases for other protein families. VKCDB is accessible at http://vkcdb.biology.ualberta.ca.

  7. Gap-junction coupling and ATP-sensitive potassium channels in human β -cell clusters: Effects on emergent dynamics

    Science.gov (United States)

    Loppini, A.; Pedersen, M. G.; Braun, M.; Filippi, S.

    2017-09-01

    The importance of gap-junction coupling between β cells in pancreatic islets is well established in mouse. Such ultrastructural connections synchronize cellular activity, confine biological heterogeneity, and enhance insulin pulsatility. Dysfunction of coupling has been associated with diabetes and altered β -cell function. However, the role of gap junctions between human β cells is still largely unexplored. By using patch-clamp recordings of β cells from human donors, we previously estimated electrical properties of these channels by mathematical modeling of pairs of human β cells. In this work we revise our estimate by modeling triplet configurations and larger heterogeneous clusters. We find that a coupling conductance in the range 0.005 -0.020 nS/pF can reproduce experiments in almost all the simulated arrangements. We finally explore the consequence of gap-junction coupling of this magnitude between β cells with mutant variants of the ATP-sensitive potassium channels involved in some metabolic disorders and diabetic conditions, translating studies performed on rodents to the human case. Our results are finally discussed from the perspective of therapeutic strategies. In summary, modeling of more realistic clusters with more than two β cells slightly lowers our previous estimate of gap-junction conductance and gives rise to patterns that more closely resemble experimental traces.

  8. Clinical relevance of ATP-dependent potassium channels

    NARCIS (Netherlands)

    Ligtenberg, JJM; vanHaeften, TW; Links, TP; Smit, AJ

    1995-01-01

    Many cells are equipped with so-called potassium (K+) channels which have an important role in maintaining transmembrane potential. Closure of these channels leads to membrane depolarization, which can be followed by cell-specific activity such as contraction of vascular smooth muscle, or secretion

  9. [Expression and properties of potassium channels in human mammary epithelial cell line MCF10A and its possible role in proliferation].

    Science.gov (United States)

    Liu, Jia; Feng, Shuang; Zhang, Lei; Wu, Zheng; Chen, Qian; Cheng, Wei; Wang, Shi-Qiang; Zou, Wei

    2010-06-25

    Voltage-dependent potassium channels (Kv) are involved in proliferation and transformation in mammary epithelial cells. In previous studies, several groups have detected various potassium channels in breast cancer cells, and they assumed that potassium channels are related to the development of breast carcinoma, although the precise mechanisms are still unknown. We have previously reported that 4-aminopyridine (4-AP), one kind of potassium channel (K(+) channel) blocker, could affect the proliferation of MCF10A cells. The aim of the present study is to explore the expression and properties of K(+) channels in human mammary epithelial cells (MCF10A) and whether Kv channels are required for the proliferation of MCF10A cell. Electrophysiological, MTT analysis, PCR and Western blot methods were used to identify a K(+) conductance which is involved in tumorigenesis and not yet be described in MCF10A cells. A voltage-dependent, outward rectification and 4-AP-sensitive K(+) current was observed in these cells. The perfusion of 5 mmol/L 4-AP significantly decreased the amplitude of Kv current from (912.5+/-0.6) pA to (275+/-0.8) pA (n=5, PMCF10A and MCF7 cells. Furthermore, the expression of Kv1.5 was much higher in MCF10A than that in MCF7. Inhibitory effect of 4-AP on cell proliferation was dosage-dependent. Incubation with 5 mmol/L 4-AP reduced MCF10A cell proliferation to 25.29% in 48 h. Western blot analysis showed the activation of ERK1/2 which related to cell proliferation was enhanced, while p38 activation was decreased by 4-AP treatment for 10 min. These data provided the first evidence of the Kv channels expression in MCF10A cell and 4-AP could inhibit the proliferation of MCF10A through blocking the potassium channels, and the mechanism may be related to regulating the activity of different members of cell proliferation signaling pathway of MEK/ERK.

  10. Tyrosine phosphatases epsilon and alpha perform specific and overlapping functions in regulation of voltage-gated potassium channels in Schwann cells

    DEFF Research Database (Denmark)

    Tiran, Zohar; Peretz, Asher; Sines, Tal

    2006-01-01

    Tyrosine phosphatases (PTPs) epsilon and alpha are closely related and share several molecular functions, such as regulation of Src family kinases and voltage-gated potassium (Kv) channels. Functional interrelationships between PTPepsilon and PTPalpha and the mechanisms by which they regulate K......+ channels and Src were analyzed in vivo in mice lacking either or both PTPs. Lack of either PTP increases Kv channel activity and phosphorylation in Schwann cells, indicating these PTPs inhibit Kv current amplitude in vivo. Open probability and unitary conductance of Kv channels are unchanged, suggesting...... an effect on channel number or organization. PTPalpha inhibits Kv channels more strongly than PTPepsilon; this correlates with constitutive association of PTPalpha with Kv2.1, driven by membranal localization of PTPalpha. PTPalpha, but not PTPepsilon, activates Src in sciatic nerve extracts, suggesting Src...

  11. Inhibition of SK4 Potassium Channels Suppresses Cell Proliferation, Migration and the Epithelial-Mesenchymal Transition in Triple-Negative Breast Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Panshi Zhang

    Full Text Available Treatments for triple-negative breast cancer (TNBC are limited; intermediate-conductance calcium-activated potassium (SK4 channels are closely involved in tumor progression, but little is known about these channels in TNBC. We aimed to investigate whether SK4 channels affect TNBC. First, by immunohistochemistry (IHC and western blotting (WB, increased SK4 protein expression in breast tumor tissues was detected relative to that in non-tumor breast tissues, but there was no apparent expression difference between various subtypes of breast cancer (p>0.05. Next, functional SK4 channels were detected in the TNBC cell line MDA-MB-231 using WB, real-time PCR, immunofluorescence and patch-clamp recording. By employing SK4 specific siRNAs and blockers, including TRAM-34 and clotrimazole, in combination with an MTT assay, a colony-formation assay, flow cytometry and a cell motility assay, we found that the suppression of SK4 channels significantly inhibited cell proliferation and migration and promoted apoptosis in MDA-MB-231 cells (p<0.05. Further investigation revealed that treatment with epidermal growth factor (EGF/basic fibroblast growth factor (bFGF caused MDA-MB-231 cells to undergo the epithelial-mesenchymal transition (EMT and to show increased SK4 mRNA expression. In addition, the down-regulation of SK4 expression inhibited the EMT markers Vimentin and Snail1. Collectively, our findings suggest that SK4 channels are expressed in TNBC and are involved in the proliferation, apoptosis, migration and EMT processes of TNBC cells.

  12. The Ketogenic Diet and Potassium Channel Function

    Science.gov (United States)

    2015-11-01

    observation is not reversed by the KD which to some extent supports our initial hypothesis. 15. SUBJECT TERMS Epilepsy , Ketogenic Diet , Seizure Disorder... ketogenic diet (KD), which is used to treat epilepsy (primarily in children) exerts a positive effect on seizure activity by regulating neuronal... Epilepsy , Ketogenic Diet , Seizure Disorder, Potassium Channels, Neurophysiology 3. Overall Project Summary: To determine the impact of KD on

  13. Effects of cisplatin on potassium currents in CT26 cells

    OpenAIRE

    Naveen Sharma; Janardhan Prasad Bhattarai; Sun Young Kim; Pyoung Han Hwang; Min Sun Kim; Seong Kyu Han

    2016-01-01

    Aims: Cisplatin, a platinum-based drug, is an important weapon against many types of cancer. It is well-known that cisplatin induces apoptosis. Potassium channel plays very important role in several signaling pathways. To investigate the possibility that potassium channels also have a role in the cellular response to cisplatin, we examined the effect of cisplatin on the activity of potassium channels on CT26 cell, the colon carcinoma cell line. Materials and Methods: The cells were culture...

  14. Potassium channels and human epileptic phenotypes: an updated overview

    Directory of Open Access Journals (Sweden)

    Chiara eVilla

    2016-03-01

    Full Text Available Potassium (K+ channels are expressed in almost every cells and are ubiquitous in neuronal and glial cell membranes. These channels have been implicated in different disorders, in particular in epilepsy. K+ channel diversity depends on the presence in the human genome of a large number of genes either encoding pore-forming or accessory subunits. More than 80 genes encoding the K+ channels were cloned and they represent the largest group of ion channels regulating the electrical activity of cells in different tissues, including the brain. It is therefore not surprising that mutations in these genes lead to K+ channels dysfunctions linked to inherited epilepsy in humans and non-human model animals.This article reviews genetic and molecular progresses in exploring the pathogenesis of different human epilepsies, with special emphasis on the role of K+ channels in monogenic forms.

  15. Specification of subunit assembly by the hydrophilic amino-terminal domain of the Shaker potassium channel.

    Science.gov (United States)

    Li, M; Jan, Y N; Jan, L Y

    1992-08-28

    The functional heterogeneity of potassium channels in eukaryotic cells arises not only from the multiple potassium channel genes and splice variants but also from the combinatorial mixing of different potassium channel polypeptides to form heteromultimeric channels with distinct properties. One structural element that determines the compatibility of different potassium channel polypeptides in subunit assembly has now been localized to the hydrophilic amino-terminal domain. A Drosophila Shaker B (ShB) potassium channel truncated polypeptide that contains only the hydrophilic amino-terminal domain can form a homomultimer; the minimal requirement for the homophilic interaction has been localized to a fragment of 114 amino acids. Substitution of the amino-terminal domain of a distantly related mammalian potassium channel polypeptide (DRK1) with that of ShB permits the chimeric DRK1 polypeptide to coassemble with ShB.

  16. Intermediate conductance calcium-activated potassium channels modulate summation of parallel fiber input in cerebellar Purkinje cells

    Science.gov (United States)

    Engbers, Jordan D. T.; Anderson, Dustin; Asmara, Hadhimulya; Rehak, Renata; Mehaffey, W. Hamish; Hameed, Shahid; McKay, Bruce E.; Kruskic, Mirna; Zamponi, Gerald W.; Turner, Ray W.

    2012-01-01

    Encoding sensory input requires the expression of postsynaptic ion channels to transform key features of afferent input to an appropriate pattern of spike output. Although Ca2+-activated K+ channels are known to control spike frequency in central neurons, Ca2+-activated K+ channels of intermediate conductance (KCa3.1) are believed to be restricted to peripheral neurons. We now report that cerebellar Purkinje cells express KCa3.1 channels, as evidenced through single-cell RT-PCR, immunocytochemistry, pharmacology, and single-channel recordings. Furthermore, KCa3.1 channels coimmunoprecipitate and interact with low voltage-activated Cav3.2 Ca2+ channels at the nanodomain level to support a previously undescribed transient voltage- and Ca2+-dependent current. As a result, subthreshold parallel fiber excitatory postsynaptic potentials (EPSPs) activate Cav3 Ca2+ influx to trigger a KCa3.1-mediated regulation of the EPSP and subsequent after-hyperpolarization. The Cav3-KCa3.1 complex provides powerful control over temporal summation of EPSPs, effectively suppressing low frequencies of parallel fiber input. KCa3.1 channels thus contribute to a high-pass filter that allows Purkinje cells to respond preferentially to high-frequency parallel fiber bursts characteristic of sensory input. PMID:22308379

  17. Optogenetics. Engineering of a light-gated potassium channel.

    Science.gov (United States)

    Cosentino, Cristian; Alberio, Laura; Gazzarrini, Sabrina; Aquila, Marco; Romano, Edoardo; Cermenati, Solei; Zuccolini, Paolo; Petersen, Jan; Beltrame, Monica; Van Etten, James L; Christie, John M; Thiel, Gerhard; Moroni, Anna

    2015-05-08

    The present palette of opsin-based optogenetic tools lacks a light-gated potassium (K(+)) channel desirable for silencing of excitable cells. Here, we describe the construction of a blue-light-induced K(+) channel 1 (BLINK1) engineered by fusing the plant LOV2-Jα photosensory module to the small viral K(+) channel Kcv. BLINK1 exhibits biophysical features of Kcv, including K(+) selectivity and high single-channel conductance but reversibly photoactivates in blue light. Opening of BLINK1 channels hyperpolarizes the cell to the K(+) equilibrium potential. Ectopic expression of BLINK1 reversibly inhibits the escape response in light-exposed zebrafish larvae. BLINK1 therefore provides a single-component optogenetic tool that can establish prolonged, physiological hyperpolarization of cells at low light intensities. Copyright © 2015, American Association for the Advancement of Science.

  18. Effects of cisplatin on potassium currents in CT26 cells

    Directory of Open Access Journals (Sweden)

    Naveen Sharma

    2016-01-01

    Conclusion: Potassium currents were detected in CT26 cells and the currents were reduced by the application of tetraethylammonium (TEA chloride, iberiotoxin, a big conductance calcium-activated potassium channel blocker and barium. The potassium currents were enhanced to 192< by the application of cisplatin (0.5 mM. Moreover, the increase of potassium currents by cisplatin was further inhibited by the application of TEA confirming the action of cisplatin on potassium channels. In addition, relative current induced by cisplatin in CT26 cells was bit larger than in normal IEC-6 cells.

  19. Proapoptotic Role of Potassium Ions in Liver Cells

    Directory of Open Access Journals (Sweden)

    Zhenglin Xia

    2016-01-01

    Full Text Available Potassium channels are transmembrane proteins that selectively promote the infiltration of potassium ions. The significance of these channels for tumor biology has become obvious. However, the effects of potassium ions on the tumor or normal cells have seldom been studied. To address this problem, we studied the biological effects of L02 and HepG2 cells with ectogenous potassium ions. Cell proliferation, cell cycle, and apoptosis rate were analyzed. Our results indicated that potassium ions inhibited proliferation of L02 and HepG2 cells and promoted their apoptosis. Potassium ions induced apoptosis through regulating Bcl-2 family members and depolarized the mitochondrial membrane, especially for HepG2 cell. These biological effects were associated with channel protein HERG. By facilitating expression of channel protein HERG, potassium ions may prevent it from being shunted to procancerous pathways by inducing apoptosis. These results demonstrated that potassium ions may be a key regulator of liver cell function. Thus, our findings suggest that potassium ions could inhibit tumorigenesis through inducing apoptosis of hepatoma cells by upregulating potassium ions transport channel proteins HERG and VDAC1.

  20. Effects of cisplatin on potassium currents in CT26 cells.

    Science.gov (United States)

    Sharma, Naveen; Bhattarai, Janardhan Prasad; Kim, Sun Young; Hwang, Pyoung Han; Kim, Min Sun; Han, Seong Kyu

    2016-01-01

    Cisplatin, a platinum-based drug, is an important weapon against many types of cancer. It is well-known that cisplatin induces apoptosis. Potassium channel plays very important role in several signaling pathways. To investigate the possibility that potassium channels also have a role in the cellular response to cisplatin, we examined the effect of cisplatin on the activity of potassium channels on CT26 cell, the colon carcinoma cell line. The cells were cultured in DMEM, supplemented with 10experiment with more than two groups. Potassium currents were detected in CT26 cells and the currents were reduced by the application of tetraethylammonium (TEA) chloride, iberiotoxin, a big conductance calcium-activated potassium channel blocker and barium. The potassium currents were enhanced to 192cisplatin (0.5 mM). Moreover, the increase of potassium currents by cisplatin was further inhibited by the application of TEA confirming the action of cisplatin on potassium channels. In addition, relative current induced by cisplatin in CT26 cells was bit larger than in normal IEC-6 cells. Potassium currents were detected in CT26 cells and the currents were reduced by the application of tetraethylammonium (TEA) chloride, iberiotoxin, a big conductance calcium-activated potassium channel blocker and barium. The potassium currents were enhanced to 192cisplatin (0.5 mM). Moreover, the increase of potassium currents by cisplatin was further inhibited by the application of TEA confirming the action of cisplatin on potassium channels. In addition, relative current induced by cisplatin in CT26 cells was bit larger than in normal IEC-6 cells.

  1. Pore size matters for potassium channel conductance

    Science.gov (United States)

    Moldenhauer, Hans; Pincuntureo, Matías

    2016-01-01

    Ion channels are membrane proteins that mediate efficient ion transport across the hydrophobic core of cell membranes, an unlikely process in their absence. K+ channels discriminate K+ over cations with similar radii with extraordinary selectivity and display a wide diversity of ion transport rates, covering differences of two orders of magnitude in unitary conductance. The pore domains of large- and small-conductance K+ channels share a general architectural design comprising a conserved narrow selectivity filter, which forms intimate interactions with permeant ions, flanked by two wider vestibules toward the internal and external openings. In large-conductance K+ channels, the inner vestibule is wide, whereas in small-conductance channels it is narrow. Here we raise the idea that the physical dimensions of the hydrophobic internal vestibule limit ion transport in K+ channels, accounting for their diversity in unitary conductance. PMID:27619418

  2. Extracellular potassium inhibits Kv7.1 potassium channels by stabilizing an inactivated state

    DEFF Research Database (Denmark)

    Larsen, Anders Peter; Steffensen, Annette Buur; Grunnet, Morten

    2011-01-01

    Kv7.1 (KCNQ1) channels are regulators of several physiological processes including vasodilatation, repolarization of cardiomyocytes, and control of secretory processes. A number of Kv7.1 pore mutants are sensitive to extracellular potassium. We hypothesized that extracellular potassium also...... modulates wild-type Kv7.1 channels. The Kv7.1 currents were measured in Xenopus laevis oocytes at different concentrations of extracellular potassium (1-50 mM). As extracellular potassium was elevated, Kv7.1 currents were reduced significantly more than expected from theoretical calculations based...... on the Goldman-Hodgkin-Katz flux equation. Potassium inhibited the steady-state current with an IC(50) of 6.0 ± 0.2 mM. Analysis of tail-currents showed that potassium increased the fraction of channels in the inactivated state. Similarly, the recovery from inactivation was slowed by potassium, suggesting...

  3. Relationship between Pore Occupancy and Gating in BK Potassium Channels

    OpenAIRE

    Piskorowski, Rebecca A.; Aldrich, Richard W.

    2006-01-01

    Permeant ions can have significant effects on ion channel conformational changes. To further understand the relationship between ion occupancy and gating conformational changes, we have studied macroscopic and single-channel gating of BK potassium channels with different permeant monovalent cations. While the slopes of the conductance?voltage curve were reduced with respect to potassium for all permeant ions, BK channels required stronger depolarization to open only when thallium was the perm...

  4. The small conductance calcium-activated potassium channel 3 (SK3) is a molecular target for Edelfosine to reduce the invasive potential of urothelial carcinoma cells.

    Science.gov (United States)

    Steinestel, Konrad; Eder, Stefan; Ehinger, Konstantin; Schneider, Juliane; Genze, Felicitas; Winkler, Eva; Wardelmann, Eva; Schrader, Andres J; Steinestel, Julie

    2016-05-01

    Metastasis is the survival-determining factor in urothelial carcinoma (UC) of the urinary bladder. The small conductance calcium-activated potassium channel 3 (SK3) enhances tumor cell invasion in breast cancer and malignant melanoma. Since Edelfosine, a glycerophospholipid with antitumoral properties, effectively inhibits SK3 channel activity, our goal was to evaluate SK3 as a potential molecular target to inhibit the gain of an invasive phenotype in UC. SK3 protein expression was analyzed in 208 tissue samples and UC cell lines. Effects of Edelfosine on SK3 expression and intracellular calcium levels as well as on cell morphology, cell survival and proliferation were assessed using immunoblotting, potentiometric fluorescence microscopy, and clonogenic/cell survival assay; furthermore, we analyzed the effect of Edelfosine and SK3 RNAi knockdown on tumor cell migration and invasion in vitro and in vivo. We found that SK3 is strongly expressed in muscle-invasive UC and in the RT112 cellular tumor model. Higher concentrations of Edelfosine have a strong antitumoral effect on UC cells, while 1 μM effectively inhibits migration/invasion of UC cells in vitro and in vivo comparable to the SK3 knockdown phenotype. Taken together, our results show strong expression of SK3 in muscle-invasive UC, consistent with the postulated role of the protein in tumor cell invasion. Edelfosine is able to effectively inhibit migration and invasion of UC cells in vitro and in vivo in an SK3-dependent way, pointing towards a possible role for Edelfosine as an antiinvasive drug to effectively inhibit UC cell invasion and metastasis.

  5. Confirming a Role for α9nAChRs and SK Potassium Channels in Type II Hair Cells of the Turtle Posterior Crista

    Directory of Open Access Journals (Sweden)

    Xiaorong Xu Parks

    2017-11-01

    Full Text Available In turtle posterior cristae, cholinergic vestibular efferent neurons (VENs synapse on type II hair cells, bouton afferents innervating type II hair cells, and afferent calyces innervating type I hair cells. Electrical stimulation of VENs releases acetylcholine (ACh at these synapses to exert diverse effects on afferent background discharge including rapid inhibition of bouton afferents and excitation of calyx-bearing afferents. Efferent-mediated inhibition is most pronounced in bouton afferents innervating type II hair cells near the torus, but becomes progressively smaller and briefer when moving longitudinally through the crista toward afferents innervating the planum. Sharp-electrode recordings have inferred that efferent-mediated inhibition of bouton afferents requires the sequential activation of alpha9-containing nicotinic ACh receptors (α9*nAChRs and small-conductance, calcium-dependent potassium channels (SK in type II hair cells. Gradations in the strength of efferent-mediated inhibition across the crista likely reflect variations in α9*nAChRs and/or SK activation in type II hair cells from those different regions. However, in turtle cristae, neither inference has been confirmed with direct recordings from type II hair cells. To address these gaps, we performed whole-cell, patch-clamp recordings from type II hair cells within a split-epithelial preparation of the turtle posterior crista. Here, we can easily visualize and record hair cells while maintaining their native location within the neuroepithelium. Consistent with α9*nAChR/SK activation, ACh-sensitive currents in type II hair cells were inward at hyperpolarizing potentials but reversed near −90 mV to produce outward currents that typically peaked around −20 mV. ACh-sensitive currents were largest in torus hair cells but absent from hair cells near the planum. In current clamp recordings under zero-current conditions, ACh robustly hyperpolarized type II hair cells. ACh

  6. TRESK potassium channel in human T lymphoblasts

    Energy Technology Data Exchange (ETDEWEB)

    Sánchez-Miguel, Dénison Selene, E-mail: amurusk@hotmail.com [Center for Biomedical Research, University of Colima, Av. 25 de Julio 965, Villa San Sebastian, C.P. 28045 Colima (Mexico); García-Dolores, Fernando, E-mail: garciaddf@yahoo.com [Department of Pathology, Institute of Forensic Sciences, Av. Niños Héroes 130, Col. Doctores, C.P. 06720 Mexico, DF (Mexico); Rosa Flores-Márquez, María, E-mail: mariafo31@yahoo.com.mx [National Medical Center of Occident (CMNO) IMSS, Belisario Dominguez 735, Col. Independencia Oriente, C.P. 44340 Guadalajara, Jalisco (Mexico); Delgado-Enciso, Iván [University of Colima, School of Medicine, Av. Universidad 333, Col. Las Viboras, C.P. 28040 Colima (Mexico); Pottosin, Igor, E-mail: pottosin@ucol.mx [Center for Biomedical Research, University of Colima, Av. 25 de Julio 965, Villa San Sebastian, C.P. 28045 Colima (Mexico); Dobrovinskaya, Oxana, E-mail: oxana@ucol.mx [Center for Biomedical Research, University of Colima, Av. 25 de Julio 965, Villa San Sebastian, C.P. 28045 Colima (Mexico)

    2013-05-03

    Highlights: • TRESK (KCNK18) mRNA is present in different T lymphoblastic cell lines. • KCNK18 mRNA was not found in resting peripheral blood lymphocytes. • Clinical samples of T lymphoblastic leukemias and lymphomas were positive for TRESK. • TRESK in T lymphoblasts has dual localization, in plasma membrane and intracellular. -- Abstract: TRESK (TWIK-related spinal cord K{sup +}) channel, encoded by KCNK18 gene, belongs to the double-pore domain K{sup +} channel family and in normal conditions is expressed predominantly in the central nervous system. In our previous patch-clamp study on Jurkat T lymphoblasts we have characterized highly selective K{sup +} channel with pharmacological profile identical to TRESK. In the present work, the presence of KCNK18 mRNA was confirmed in T lymphoblastic cell lines (Jurkat, JCaM, H9) but not in resting peripheral blood lymphocytes of healthy donors. Positive immunostaining for TRESK was demonstrated in lymphoblastic cell lines, in germinal centers of non-tumoral lymph nodes, and in clinical samples of T acute lymphoblastic leukemias/lymphomas. Besides detection in the plasma membrane, intracellular TRESK localization was also revealed. Possible involvement of TRESK channel in lymphocyte proliferation and tumorigenesis is discussed.

  7. Role of ATP sensitive potassium channel in extracellular potassium accumulation and cardiac arrhythmias during myocardial ischaemia.

    Science.gov (United States)

    Billman, G E

    1994-06-01

    Extracellular potassium rises rapidly during myocardial ischaemia, correlating with the onset of ventricular arrhythmias. The extracellular accumulation of potassium can induce abnormalities in both impulse conduction and impulse generation. Inhomogeneities of potassium conductance will elicit regional differences in action potential duration and repolarisation. The resulting spatial dispersion of refractory period will allow for fragmentation of impulse conduction on ensuing beats, the formation of irregular reentrant pathways and ventricular fibrillation. In a similar manner, the spread of injury current from the ischaemic tissue to surrounding normal tissue can trigger extrasystoles (depolarisation induced automaticity). It has been hypothesised that the activation of the ATP sensitive potassium channel contributes significantly to reductions in action potential duration and increases in extracellular potassium accumulation during myocardial ischaemia. ATP sensitive potassium channel antagonists prevent ischaemically induced reductions in action potential duration and the dispersion of refractory period but may induce oscillatory afterpotentials under some conditions (for example, calcium overload). In contrast, potassium channel agonists enhance the dispersion of refractory period ischaemia, which promotes the formation of re-entrant arrhythmias. The pharmacological modulation of the ATP sensitive potassium channels could therefore offer a novel approach for the management of cardiac arrhythmias in patients with ischaemic heart disease. In general, channel antagonists prevent ventricular fibrillation, while high (hypotensive) doses of channel agonists can induce malignant arrhythmias during ischaemia in animal models. However, recent evidence also suggests that potassium channel agonists may promote a better preservation of myocardial mechanical performance during reperfusion while ATP sensitive potassium channel antagonists exacerbate mechanical depression

  8. General Sensitization of melanoma cells for TRAIL-induced apoptosis by the potassium channel inhibitor TRAM-34 depends on release of SMAC.

    Directory of Open Access Journals (Sweden)

    Sandra-Annika Quast

    Full Text Available The death ligand TRAIL represents a promising therapeutic strategy for metastatic melanoma, however prevalent and inducible resistance limit its applicability. A new approach is presented here for sensitization to TRAIL. It is based on inhibition of the membrane potassium channel KCa3.1 (IK1, which serves fundamental cellular functions related to membrane potential. The selective inhibitor TRAM-34 did not induce apoptosis by itself but synergistically enhanced TRAIL sensitivity and overrode TRAIL resistance in a large panel of melanoma cell lines. Expression of IK1 was also found in mitochondria, and its inhibition resulted in mitochondrial membrane hyperpolarization and an early activation of Bax. The combination of TRAM-34 and TRAIL resulted in massive release of mitochondrial factors, cytochrome c, AIF and SMAC/DIABLO. Bax knockdown and Bcl-2 overexpression abolished apoptosis. Overexpression of XIAP diminished apoptosis by two-fold, and SMAC knockdown almost completely abolished apoptosis. These data uncover the existence of a rheostat in melanoma cells, consisting of inhibitor of apoptosis proteins and SMAC, which regulates TRAIL sensitivity. Thus, a new strategy is described based on mitochondrial membrane channels, which correspond to Bax activation. As both TRAIL and IK1 inhibitors had shown only minor side effects in clinical trials, a clinical application of this combination is conceivable.

  9. Optogenetic techniques for the study of native potassium channels

    Directory of Open Access Journals (Sweden)

    Guillaume Eric Sandoz

    2013-04-01

    Full Text Available Optogenetic tools were originally designed to target specific neurons for remote control of their activity by light and have largely been built around opsin-based channels and pumps. These naturally photosensitive opsins are microbial in origin and are unable to mimic the properties of native neuronal receptors and channels. Over the last 8 years, photoswitchable-tethered ligands (PTLs have enabled fast and reversible control of mammalian ion channels, allowing optical control of neuronal activity. One such PTL, MAQ, contains a maleimide (M to tether the molecule to a genetically engineered cysteine, a photoisomerizable azobenzene (A linker and a pore-blocking quaternary ammonium group (Q. MAQ was originally used to photo-control SPARK, an engineered light-gated potassium channel derived from Shaker. Potassium channel photo-block by MAQ has recently been extended to a diverse set of mammalian potassium channels including channels in the voltage-gated and K2P families. Photoswitchable potassium channels, which maintain native properties, pave the way for the optical control of specific aspects of neuronal function and for high precision probing of a specific channel’s physiological functions. To extend optical control to natively expressed channels, without overexpression, one possibility is to develop a knock-in mouse in which the wild type channel gene is replaced by its light-gated version. Alternatively, the recently developed photoswitchable-conditional-subunit technique (PCS provides photocontrol of the channel of interest by molecular replacement of wild type complexes. Finally, photochromic ligands (PCLs also allow photocontrol of potassium channels without genetic manipulation using soluble compounds. In this review we discuss different techniques for optical control of native potassium channels and their associated advantages and disadvantages.

  10. Expression of T-cell KV1.3 potassium channel correlates with pro-inflammatory cytokines and disease activity in ulcerative colitis

    DEFF Research Database (Denmark)

    Koch Hansen, Lars; Møller, Linda Maria Sevelsted; Rabjerg, Maj

    2014-01-01

    BACKGROUND AND AIMS: Potassium channels, KV1.3 and KCa3.1, have been suggested to control T-cell activation, proliferation, and cytokine production and may thus constitute targets for anti-inflammatory therapy. Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized...... by excessive T-cell infiltration and cytokine production. It is unknown if KV1.3 and KCa3.1 in the inflamed mucosa are markers of active UC. We hypothesized that KV1.3 and KCa3.1 correlate with disease activity and cytokine production in patients with UC. METHODS: Mucosal biopsies were collected from patients...... with active UC (n=33) and controls (n=15). Protein and mRNA expression of KV1.3 and KCa3.1, immune cell markers, and pro-inflammatory cytokines were determined by quantitative-real-time-polymerase-chain-reaction (qPCR) and immunofluorescence, and correlated with clinical parameters of inflammation. In...

  11. Effects of dalfampridine and its metabolites on cloned human potassium channels Kv 1.1, Kv 1.2, and Kv 1.4 expressed in human embryonic kidney cells.

    Science.gov (United States)

    Caggiano, Anthony; Blight, Andrew; Parry, Tom J

    2013-01-01

    Dalfampridine (4-aminopyridine; 4-AP) is a potassium channel blocker that has been available in the United States as a treatment to improve walking in patients with multiple sclerosis. 4-AP is well-characterized in vitro with regard to inhibition of neuronal potassium channels, but the potential contribution of its metabolites to clinical activity has not been determined. This study evaluated the concentration-response of 4-AP and its two primary metabolites, 3-hydroxy-4-aminopyridine and 3-hydroxy-4-aminopyridine sulfate, for inhibition of the potassium channels Kv 1.1, Kv 1.2, and Kv 1.4, which are considered candidates for mediating effects of 4-AP on action potential conduction because of their presence in axonal membranes. Stable transfection of cDNA for Kv 1.1, Kv 1.2, and Kv 1.4 was performed into HEK293 cells, and colonies of cells containing each channel were selected and maintained under appropriate cell culture conditions. Electrophysiological measurements were performed using a patch-clamp technique in at least three cells for each concentration (50, 500, 5000, and 50,000 μM) of 4-AP and the two metabolites, with each cell acting as its own control. Concentration-response curves were constructed for 4-AP and each metabolite. Data were analyzed using nonlinear least-squares fit, and concentrations inhibiting the channels by 50% (IC50) were estimated. 4-AP induced similar concentration-dependent inhibition profiles of all three potassium channels, resulting in a narrow range of IC50 values across channels (242 µM to 399 µM). Across the three channels, the IC50 values of 3-hydroxy-4-aminopyridine and 3-hydroxy-4-aminopyridine sulfate were 1-2 orders of magnitude higher (less potent) than those of 4-AP. 3-Hydroxy-4-aminopyridine and 3-hydroxy-4-aminopyridine sulfate demonstrated low in vitro potency for Kv 1.1, Kv 1.2, and Kv 1.4 inhibition, suggesting that these metabolites are unlikely to contribute to the positive pharmacodynamic effects of 4-AP

  12. Importance of large conductance calcium-activated potassium channels (BKCa) in interleukin-1b-induced adhesion of monocytes to endothelial cells.

    Science.gov (United States)

    Burgazli, K M; Venker, C J; Mericliler, M; Atmaca, N; Parahuleva, M; Erdogan, A

    2014-01-01

    The present study investigated the role of the large conductance calcium-activated potassium channels (BKCa) in interleukin-1b (IL-1b) induced inflammation. Human umbilical vein endothelial cells (HUVECs) were isolated and cultured. Endothelial cell membrane potential measurements were accomplished using the fluorescent dye DiBAC4(3). The role of BKCa was assessed using iberiotoxin, a highly selective BKCa inhibitor. Changes in the calcium intracellular calcium were investigated using Fura-2-AM imaging. Fluorescent dyes DCF-AM and DAF-AM were further used in order to measure the formation of reactive oxygen species (ROS) and nitric oxide (NO) synthesis, respectively. Endothelial cell adhesion tests were conducted with BCECF-AM adhesion assay and tritium thymidine uptake using human monocytic cells (U937). Expression of cellular adhesion molecules (ICAM-1, VCAM-1) was determined by flow cytometer. Interleukin-1b induced a BKCa dependent hyperpolarization of HUVECs. This was followed by an increase in the intracellular calcium concentration. Furthermore, IL-1b significantly increased the synthesis of NO and ROS. The increase of intracellular calcium, radicals and NO resulted in a BKCa dependent adhesion of monocytes to HUVECs. Endothelial cells treated with IL-1b expressed both ICAM-1 and VCAM-1 in significantly higher amounts as when compared to controls. It was further shown that the cellular adhesion molecules ICAM-1 and VCAM-1 were responsible for the BKCa-dependent increase in cellular adhesion. Additionally, inhibition of the NADPH oxidase with DPI led to a significant downregulation of IL-1b-induced expression of ICAM and VCAM, as well as inhibition of eNOS by L-NMMA, and intracellular calcium by BAPTA. Activation of the endothelial BKCa plays an important role in the IL-1b-induced monocyte adhesion to endothelial cells.

  13. The KCNQ1 potassium channel: from gene to physiological function

    DEFF Research Database (Denmark)

    Jespersen, Thomas; Grunnet, Morten; Olesen, Søren-Peter

    2005-01-01

    The voltage-gated KCNQ1 (KvLQT1, Kv7.1) potassium channel plays a crucial role in shaping the cardiac action potential as well as in controlling the water and salt homeostasis in several epithelial tissues. KCNQ1 channels in these tissues are tightly regulated by auxiliary proteins and accessory...

  14. Data on the construction of a recombinant HEK293 cell line overexpressing hERG potassium channel and examining the presence of hERG mRNA and protein expression

    Directory of Open Access Journals (Sweden)

    Yi Fan Teah

    2017-10-01

    Full Text Available The data presented in this article are related to the research article entitled “The effects of deoxyelephantopin on the cardiac delayed rectifier potassium channel current (IKr and human ether-a-go-go-related gene (hERG expression” (Y.F. Teah, M.A. Abduraman, A. Amanah, M.I. Adenan, S.F. Sulaiman, M.L. Tan [1], which the possible hERG blocking properties of deoxyelephantopin were investigated. This article describes the construction of human embryonic kidney 293 (HEK293 cells overexpressing HERG potassium channel and verification of the presence of hERG mRNA and protein expression in this recombinant cell line.

  15. 18β-Glycyrrhetinic acid potently inhibits Kv1.3 potassium channels and T cell activation in human Jurkat T cells.

    Science.gov (United States)

    Fu, Xiao-Xing; Du, Li-Li; Zhao, Ning; Dong, Qian; Liao, Yu-Hua; Du, Yi-Mei

    2013-07-09

    Licorice has been extensively used in traditional medicines for treatment of many diseases, including inflammations and immunological disorders. Recent studies have shown that the anti-inflammatory and immunomodulation activities of licorice have been attributed to its active component, glycyrretinic acid (GA). GA consists of two isoforms, 18α- and 18β-. However, its mechanism remains poorly understood. We compared the effects of two isoforms on Kv1.3 channels in Jurkat T cells and further characterized the inhibition of Kv1.3 channels by 18β-GA in CHO cells. In addition, we examined the effects of 18β-GA on Kv1.3 gene expression, Ca(2+) influx, proliferation, as well as IL-2 production in Jurkat T cells. Whole-cell patch-clamp technique was applied to record Kv1.3 currents in Jurkat T or CHO cells. Real-time PCR and Western blotting were used to detect gene expression. Fluo-4, CCK-8 kit and ELISA kit were used to measure Ca(2+) influx, proliferation, and IL-2 secretion in Jurkat T cells, respectively. Superfusion of 18β-GA (10-100 µM) blocked Kv1.3 currents in Jurkat T cells, while 18α-GA at the same concentration had no effect. The 18β-GA induced inhibition had a voltage- and concentration-dependent manner with an IC50 of 23.9±1.5 µM at +40 mV in CHO cells. Furthermore, 18β-GA significantly inhibited Kv1.3 gene expression. In addition, paralleling Kv1.3 inhibition, 18β-GA also inhibited Ca(2+) influx, proliferation as well as IL-2 production in Jurkat T cells. 18β-GA blocks Kv1.3 channels, which probably involves its anti-inflammatory and immunomodulation effects. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  16. PKC and AMPK regulation of Kv1.5 potassium channels

    DEFF Research Database (Denmark)

    Andersen, Martin Nybo; Skibsbye, Lasse; Tang, Chuyi

    2015-01-01

    The voltage-gated Kv1.5 potassium channel, conducting the ultra-rapid rectifier K(+) current (IKur), is regulated through several pathways. Here we investigate if Kv1.5 surface expression is controlled by the 2 kinases PKC and AMPK, using Xenopus oocytes, MDCK cells and atrial derived HL-1 cells...

  17. Flow activates an endothelial potassium channel to release an endogenous nitrovasodilator.

    Science.gov (United States)

    Cooke, J P; Rossitch, E; Andon, N A; Loscalzo, J; Dzau, V J

    1991-01-01

    Flow-mediated vasodilation is endothelium dependent. We hypothesized that flow activates a potassium channel on the endothelium, and that activation of this channel leads to the release of the endogenous nitrovasodilator, nitric oxide. To test this hypothesis, rabbit iliac arteries were perfused at varying flow rates, at a constant pressure of 60 mm Hg. Increments in flow induced proportional increases in vessel diameter, which were abolished by L,N-mono-methylarginine (the antagonist of nitric-oxide synthesis). Barium chloride, depolarizing solutions of potassium, verapamil, calcium-free medium, and antagonists of the KCa channel (charybdotoxin, iberiotoxin) also blocked flow-mediated vasodilation. Conversely, responses to other agonists of endothelium-dependent and independent vasodilation were unaffected by charybdotoxin or iberiotoxin. To confirm that flow activated a specific potassium channel to induce the release of nitric oxide, endothelial cells cultured on micro-carrier beads were added to a flow chamber containing a vascular ring without endothelium. Flow-stimulated endothelial cells released a diffusible vasodilator; the degree of vasorelaxation was dependent upon the flow rate. Relaxation was abrogated by barium, tetraethylammonium ion, or charybdotoxin, but was not affected by apamin, glybenclamide, tetrodotoxin, or ouabain. The data suggest that transmission of a hyperpolarizing current from endothelium to the vascular smooth muscle is not necessary for flow-mediated vasodilation. Flow activates a potassium channel (possibly the KCa channel) on the endothelial cell membrane that leads to the release of nitric oxide. Images PMID:1719029

  18. Inhibitory actions by ibandronate sodium, a nitrogen-containing bisphosphonate, on calcium-activated potassium channels in Madin–Darby canine kidney cells

    Directory of Open Access Journals (Sweden)

    Sheng-Nan Wu

    2015-01-01

    Full Text Available The nitrogen-containing bisphosphonates used for management of the patients with osteoporosis were reported to influence the function of renal tubular cells. However, how nitrogen-containing bisphosphates exert any effects on ion currents remains controversial. The effects of ibandronate (Iban, a nitrogen-containing bisphosphonate, on ionic channels, including two types of Ca2+-activated K+ (KCa channels, namely, large-conductance KCa (BKCa and intermediate-conductance KCa (IKCa channels, were investigated in Madin–Darby canine kidney (MDCK cells. In whole-cell current recordings, Iban suppressed the amplitude of voltage-gated K+ current elicited by long ramp pulse. Addition of Iban caused a reduction of BKCa channels accompanied by a right shift in the activation curve of BKCa channels, despite no change in single-channel conductance. Ca2+ sensitivity of these channels was modified in the presence of this compound; however, the magnitude of Iban-mediated decrease in BKCa-channel activity under membrane stretch with different negative pressure remained unchanged. Iban suppressed the probability of BKCa-channel openings linked primarily to a shortening in the slow component of mean open time in these channels. The dissociation constant needed for Iban-mediated suppression of mean open time in MDCK cells was 12.2 μM. Additionally, cell exposure to Iban suppressed the activity of IKCa channels, and DC-EBIO or 9-phenanthrol effectively reversed its suppression. Under current-clamp configuration, Iban depolarized the cells and DC-EBIO or PF573228 reversed its depolarizing effect. Taken together, the inhibitory action of Iban on KCa-channel activity may contribute to the underlying mechanism of pharmacological or toxicological actions of Iban and its structurally similar bisphosphonates on renal tubular cells occurring in vivo.

  19. Mechanism of Proarrhythmic Effects of Potassium Channel Blockers

    DEFF Research Database (Denmark)

    Skibsbye, Lasse; Ravens, Ursula

    2016-01-01

    Any disturbance of electrical impulse formation in the heart and of impulse conduction or action potential (AP) repolarization can lead to rhythm disorders. Potassium (K(+)) channels play a prominent role in the AP repolarization process. In this review we describe the causes and mechanisms...

  20. Modeling removal of accumulated potassium from T-tubules by inward rectifier potassium channels

    NARCIS (Netherlands)

    Wallinga, W.; Vliek, M.; Wienk, E.D.; Alberink, M.J.; Ypey, D.L.; Ypey, D.L.

    1996-01-01

    The membrane models of Cannon et al. (1993) and Alberink et al. (1995) for mammalian skeletal muscle fibers are based upon Hodgkin-Huxley descriptions of sodium, potassium delayed rectifier and leak conductances and the capacitive current taking into account fast inactivation of sodium channels. Now

  1. Regulation of Kv1.4 potassium channels by PKC and AMPK kinases

    DEFF Research Database (Denmark)

    Andersen, Martin Nybo; Skibsbye, Lasse; Saljic, Arnela

    2018-01-01

    Over the last years extensive kinase-mediated regulation of a number of voltage-gated potassium (Kv) channels important in cardiac electrophysiology has been reported. This includes regulation of Kv1.5, Kv7.1 and Kv11.1 cell surface expression, where the kinase-mediated regulation appears to center......, but is unaffected by PI3K-SGK1 signaling, as well as Nedd4-1/Nedd4-2 activity. In the light of previous reports, our results demonstrate an impressive heterogeneity in the molecular pathways controlling the surface expression of highly related potassium channel subunits....

  2. Effect of Potassium Channel Modulators on Morphine Withdrawal in Mice

    Directory of Open Access Journals (Sweden)

    Vikas Seth

    2010-01-01

    Full Text Available The present study was conducted to investigate the effect of potassium channel openers and blockers on morphine withdrawal syndrome. Mice were rendered dependent on morphine by subcutaneous injection of morphine; four hours later, withdrawal was induced by using an opioid antagonist, naloxone. Mice were observed for 30 minutes for the withdrawal signs ie, the characteristic jumping, hyperactivity, urination and diarrhea. ATP-dependent potassium (K + ATP channel modulators were injected intraperitoneally (i.p. 30 minutes before the naloxone. It was found that a K + ATP channel opener, minoxidil (12.5–50 mg/kg i.p., suppressed the morphine withdrawal significantly. On the other hand, the K + ATP channel blocker glibenclamide (12.5–50 mg/kg i.p. caused a significant facilitation of the withdrawal. Glibenclamide was also found to abolish the minoxidil's inhibitory effect on morphine withdrawal. The study concludes that K + ATP channels play an important role in the genesis of morphine withdrawal and K + ATP channel openers could be useful in the management of opioid withdrawal. As morphine opens K + ATP channels in neurons, the channel openers possibly act by mimicking the effects of morphine on neuronal K + currents.

  3. Potassium and ANO1/TMEM16A chloride channel profiles distinguish atypical and typical smooth muscle cells from interstitial cells in the mouse renal pelvis

    Science.gov (United States)

    Iqbal, Javed; Tonta, Mary A; Mitsui, Retsu; Li, Qun; Kett, Michelle; Li, Jinhua; Parkington, Helena C; Hashitani, Hikaru; Lang, Richard J

    2012-01-01

    BACKGROUND AND PURPOSE Although atypical smooth muscle cells (SMCs) in the proximal renal pelvis are thought to generate the pacemaker signals that drive pyeloureteric peristalsis, their location and electrical properties remain obscure. EXPERIMENTAL APPROACH Standard patch clamp, intracellular microelectrode and immunohistochemistry techniques were used. To unequivocally identify SMCs, transgenic mice with enhanced yellow fluorescent protein (eYFP) expressed in cells containing α-smooth muscle actin (α-SMA) were sometimes used. KEY RESULTS Atypical SMCs were distinguished from typical SMCs by the absence of both a transient 4-aminopyridine-sensitive K+ current (IKA) and spontaneous transient outward currents (STOCs) upon the opening of large-conductance Ca2+-activated K+ (BK) channels. Many typical SMCs displayed a slowly activating, slowly decaying Cl- current blocked by niflumic acid (NFA). Immunostaining for KV4.3 and ANO1/ TMEM16A Cl- channel subunits co-localized with α-SMA immunoreactive product predominately in the distal renal pelvis. Atypical SMCs fired spontaneous inward currents that were either selective for Cl- and blocked by NFA, or cation-selective and blocked by La3+. α-SMA- interstitial cells (ICs) were distinguished by the presence of a Xe991-sensitive KV7 current, BK channel STOCs and Cl- selective, NFA-sensitive spontaneous transient inward currents (STICs). Intense ANO1/ TMEM16A and KV7.5 immunostaining was present in Kit-α-SMA- ICs in the suburothelial and adventitial regions of the renal pelvis. CONCLUSIONS AND IMPLICATIONS We conclude that KV4.3+α-SMA+ SMCs are typical SMCs that facilitate muscle wall contraction, that ANO1/ TMEM16A and KV7.5 immunoreactivity may be selective markers of Kit- ICs and that atypical SMCs which discharge spontaneous inward currents are the pelviureteric pacemakers. PMID:22014103

  4. Neuronal trafficking of voltage-gated potassium channels

    DEFF Research Database (Denmark)

    Jensen, Camilla S; Rasmussen, Hanne Borger; Misonou, Hiroaki

    2011-01-01

    The computational ability of CNS neurons depends critically on the specific localization of ion channels in the somatodendritic and axonal membranes. Neuronal dendrites receive synaptic inputs at numerous spines and integrate them in time and space. The integration of synaptic potentials...... is regulated by voltage-gated potassium (Kv) channels, such as Kv4.2, which are specifically localized in the dendritic membrane. The synaptic potentials eventually depolarize the membrane of the axon initial segment, thereby activating voltage-gated sodium channels to generate action potentials. Specific Kv...

  5. Specific Sorting and Post-Golgi trafficking of Dendritic Potassium Channels in Living Neurons

    DEFF Research Database (Denmark)

    Jensen, Camilla Stampe; Watanabe, Shoji; Rasmussen, Hanne Borger

    2014-01-01

    localization in distinct dendritic sub-compartments are largely unknown. Here, we developed a quantitative live-cell imaging method to analyze protein sorting and post-Golgi vesicular trafficking. We focused on two dendritic voltage-gated potassium channels which exhibit distinct localizations; Kv2...

  6. Estradiol rapidly induces the translocation and activation of the intermediate conductance calcium activated potassium channel in human eccrine sweat gland cells.

    LENUS (Irish Health Repository)

    Muchekehu, Ruth W

    2009-02-01

    Steroid hormones target K+ channels as a means of regulating electrolyte and fluid transport. In this study, ion transporter targets of Estradiol (E2) were investigated in the human eccrine sweat gland cell line NCL-SG3.

  7. Glucose- and interleukin-1beta-induced beta-cell apoptosis requires Ca2+ influx and extracellular signal-regulated kinase (ERK) 1/2 activation and is prevented by a sulfonylurea receptor 1/inwardly rectifying K+ channel 6.2 (SUR/Kir6.2) selective potassium channel opener in human islets

    DEFF Research Database (Denmark)

    Maedler, Kathrin; Størling, Joachim; Sturis, Jeppe

    2004-01-01

    for the beta-cell potassium channel SUR1/Kir6.2, on glucose- and IL-1beta-induced apoptosis and impaired function in human beta-cells. Exposure of human islets for 4 days to 11.1 and 33.3 mmol/l glucose, 2 ng/ml IL-1beta, or 10 and 100 micromol/l of the sulfonylurea tolbutamide induced beta-cell apoptosis...

  8. Characterization of potassium channel modulators with QPatch automated patch-clamp technology: system characteristics and performance

    DEFF Research Database (Denmark)

    Kutchinsky, Jonatan; Friis, Søren; Asmild, Margit

    2003-01-01

    developed using this technology. A single-channel prototype of the QPatch system was used for validation of the patch-clamp chip technology. We present here data on the quality of patch-clamp recordings and from actual drug screening studies of human potassium channels expressed in cultured cell lines....... Using Chinese hamster ovary (CHO) and human embryonic kidney cells (HEK), gigaseals of 4.1 +/- 0.4 GOmega (n = 146) and high-quality whole-cell current recordings were obtained from hERG and KCNQ4 potassium channels. Success rates for gigaseal recordings varied from 40 to 95%, and 67% of the whole......-response relationship characterizations of verapamil and rBeKm-1 blockage of hERG currents provided IC(50) values similar to values reported in the literature....

  9. Differential effect of brief electrical stimulation on voltage-gated potassium channels.

    Science.gov (United States)

    Cameron, Morven A; Al Abed, Amr; Buskila, Yossi; Dokos, Socrates; Lovell, Nigel H; Morley, John W

    2017-05-01

    Electrical stimulation of neuronal tissue is a promising strategy to treat a variety of neurological disorders. The mechanism of neuronal activation by external electrical stimulation is governed by voltage-gated ion channels. This stimulus, typically brief in nature, leads to membrane potential depolarization, which increases ion flow across the membrane by increasing the open probability of these voltage-gated channels. In spiking neurons, it is activation of voltage-gated sodium channels (Na V channels) that leads to action potential generation. However, several other types of voltage-gated channels are expressed that also respond to electrical stimulation. In this study, we examine the response of voltage-gated potassium channels (K V channels) to brief electrical stimulation by whole cell patch-clamp electrophysiology and computational modeling. We show that nonspiking amacrine neurons of the retina exhibit a large variety of responses to stimulation, driven by different K V -channel subtypes. Computational modeling reveals substantial differences in the response of specific K V -channel subtypes that is dependent on channel kinetics. This suggests that the expression levels of different K V -channel subtypes in retinal neurons are a crucial predictor of the response that can be obtained. These data expand our knowledge of the mechanisms of neuronal activation and suggest that K V -channel expression is an important determinant of the sensitivity of neurons to electrical stimulation. NEW & NOTEWORTHY This paper describes the response of various voltage-gated potassium channels (K V channels) to brief electrical stimulation, such as is applied during prosthetic electrical stimulation. We show that the pattern of response greatly varies between K V channel subtypes depending on activation and inactivation kinetics of each channel. Our data suggest that problems encountered when artificially stimulating neurons such as cessation in firing at high frequencies, or

  10. Protective roles for potassium SK/KCa2 channels in microglia and neurons

    Directory of Open Access Journals (Sweden)

    Amalia M Dolga

    2012-11-01

    Full Text Available New concepts on potassium channel function in neuroinflammation suggest that they regulate mechanisms of microglial activation, including intracellular calcium homeostasis, morphological alterations, pro-inflammatory cytokine release, antigen presentation, and phagocytosis. Although little is known about voltage independent potassium channels in microglia, special attention emerges on small (SK/KCNN1-3/KCa2 and intermediate (IK/KCNN4/KCa3.1-conductance calcium-activated potassium channels as regulators of microglial activation in the field of research on neuroinflammation and neurodegeneration. In particular, recent findings suggested that SK/KCa2 channels, by regulating calcium homeostasis, may elicit a dual mechanism of action with protective properties in neurons and inhibition of inflammatory responses in microglia. Thus, modulating SK/KCa2 channels and calcium signaling may provide novel therapeutic strategies in neurological disorders, where neuronal cell death and inflammatory responses concomitantly contribute to disease progression. Here, we review the particular role of SK/KCa2 channels for [Ca2+]i regulation in microglia and neurons, and we discuss the potential impact for further experimental approaches addressing novel therapeutic strategies in neurological diseases, where neuronal cell death and neuroinflammatory processes are prominent.

  11. Study of the interaction of potassium ion channel protein with micelle by molecular dynamics simulation

    Science.gov (United States)

    Shantappa, Anil; Talukdar, Keka

    2018-04-01

    Ion channels are proteins forming pore inside the body of all living organisms. This potassium ion channel known as KcsA channel and it is found in the each cell and nervous system. Flow of various ions is regulated by the function of the ion channels. The nerve ion channel protein with protein data bank entry 1BL8, which is basically an ion channel protein in Streptomyces Lividans and which is taken up to form micelle-protein system and the system is analyzed by using molecular dynamics simulation. Firstly, ion channel pore is engineered by CHARMM potential and then Micelle-protein system is subjected to molecular dynamics simulation. For some specific micelle concentration, the protein unfolding is observed.

  12. The potassium channel Kir4.1 associates with the dystrophin-glycoprotein complex via alpha-syntrophin in glia.

    Science.gov (United States)

    Connors, Nathan C; Adams, Marvin E; Froehner, Stanley C; Kofuji, Paulo

    2004-07-02

    One of the major physiological roles of potassium channels in glial cells is to promote "potassium spatial buffering" in the central nervous system, a process necessary to maintain an optimal potassium concentration in the extracellular environment. This process requires the precise distribution of potassium channels accumulated at high density in discrete subdomains of glial cell membranes. To obtain a better understanding of how glial cells selectively target potassium channels to discrete membrane subdomains, we addressed the question of whether the glial inwardly rectifying potassium channel Kir4.1 associates with the dystrophin-glycoprotein complex (DGC). Immunoprecipitation experiments revealed that Kir4.1 is associated with the DGC in mouse brain and cultured cortical astrocytes. In vitro immunoprecipitation and pull-down assays demonstrated that Kir4.1 can bind directly to alpha-syntrophin, requiring the presence of the last three amino acids of the channel (SNV), a consensus PDZ domain-binding motif. Furthermore, Kir4.1 failed to associate with the DGC in brains from alpha-syntrophin knockout mice. These results suggest that Kir4.1 is localized in glial cells by its association with the DGC through a PDZ domain-mediated interaction with alpha-syntrophin and suggest an important role for the DGC in central nervous system physiology.

  13. Mechanism of electromechanical coupling in voltage-gated potassium channels

    Directory of Open Access Journals (Sweden)

    Rikard eBlunck

    2012-09-01

    Full Text Available Voltage-gated ion channels play a central role in the generation of action potentials in the nervous system. They are selective for one type of ion – sodium, calcium or potassium. Voltage-gated ion channels are composed of a central pore that allows ions to pass through the membrane and four peripheral voltage sensing domains that respond to changes in the membrane potential. Upon depolarization, voltage sensors in voltage-gated potassium channels (Kv undergo conformational changes driven by positive charges in the S4 segment and aided by pairwise electrostatic interactions with the surrounding voltage sensor. Structure-function relations of Kv channels have been investigated in detail, and the resulting models on the movement of the voltage sensors now converge to a consensus; the S4 segment undergoes a combined movement of rotation, tilt and vertical displacement in order to bring 3-4 e+ each through the electric field focused in this region. Nevertheless, the mechanism by which the voltage sensor movement leads to pore opening, the electromechanical coupling, is still not fully understood. Thus, recently, electromechanical coupling in different Kv channels has been investigated with a multitude of techniques including electrophysiology, 3D crystal structures, fluorescence spectroscopy and molecular dynamics simulations. Evidently, the S4-S5 linker, the covalent link between the voltage sensor and pore, plays a crucial role. The linker transfers the energy from the voltage sensor movement to the pore domain via an interaction with the S6 C-termini, which are pulled open during gating. In addition, other contact regions have been proposed. This review aims to provide (i an in-depth comparison of the molecular mechanisms of electromechanical coupling in different Kv channels; (ii insight as to how the voltage sensor and pore domain influence one another; and (iii theoretical predictions on the movement of the cytosolic face of the KV channels

  14. Voltage-gated potassium channels regulate calcium-dependent pathways involved in human T lymphocyte activation.

    Science.gov (United States)

    Lin, C S; Boltz, R C; Blake, J T; Nguyen, M; Talento, A; Fischer, P A; Springer, M S; Sigal, N H; Slaughter, R S; Garcia, M L

    1993-03-01

    The role that potassium channels play in human T lymphocyte activation has been investigated by using specific potassium channel probes. Charybdotoxin (ChTX), a blocker of small conductance Ca(2+)-activated potassium channels (PK,Ca) and voltage-gated potassium channels (PK,V) that are present in human T cells, inhibits the activation of these cells. ChTX blocks T cell activation induced by signals (e.g., anti-CD2, anti-CD3, ionomycin) that elicit a rise in intracellular calcium ([Ca2+]i) by preventing the elevation of [Ca2+]i in a dose-dependent manner. However, ChTX has no effect on the activation pathways (e.g., anti-CD28, interleukin 2 [IL-2]) that are independent of a rise in [Ca2+]i. In the former case, both proliferative response and lymphokine production (IL-2 and interferon gamma) are inhibited by ChTX. The inhibitory effect of ChTX can be demonstrated when added simultaneously, or up to 4 h after the addition of the stimulants. Since ChTX inhibits both PK,Ca and PK,V, we investigated which channel is responsible for these immunosuppressive effects with the use of two other peptides, noxiustoxin (NxTX) and margatoxin (MgTX), which are specific for PK,V. These studies demonstrate that, similar to ChTX, both NxTX and MgTX inhibit lymphokine production and the rise in [Ca2+]i. Taken together, these data provide evidence that blockade of PK,V affects the Ca(2+)-dependent pathways involved in T lymphocyte proliferation and lymphokine production by diminishing the rise in [Ca2+]i that occurs upon T cell activation.

  15. [Construction of acid-sensitive potassium channel-3 eukaryotic expression plasmid and its express in SH-SY5Y cells].

    Science.gov (United States)

    Wei, Lin-yu; Li, Xin-juan; Mei, Yi-wen; Wang, Guo-hong; Wang, Qi; Li, Dong-liang; Li, Chao-kun

    2015-05-01

    To construct the acid-sensitive potassium hannel-3(TASK3) eukaryotic expression plasmid and to establish a stable SH-SY5Y cell line expressing enhanced green fluorescent protein (EGFP)-tagged TASK3. TASK3 coding region was subcloned into pEGFP-N1 plasmid to construct a recombinant vector alled pEGFP-TASK3. The correct recombinant expressing plasmid was transfected with X-feet transfection reagent to SH-SY5Y cells. The cell line stably expressiing EGFP tagged-TASK3 gene was established by screening with antibiotic G418 and fluorescence microscope. The expression and localization of the EGFP tagged-TASK3 fusion protein was detected by Western blot and confocal microscope. Exposure of the SH-SY5Y cell line expressing stably TASK3-eGFP fusion proteins was exposed to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability was assessed with cell counting Kit-8 (CCK-8). All the results of identification by PCR, digestion with restriction endonuclease and sequencing indicated that the recombinant eukaryotic expression plasmid pEGFP-TASK3 was constructed correctly. The stable SH-SY5Y cell line expressing EGFP tagged-TASK3 fusion protein was successfully established. Exposure of the wild type SH-SY5Y cells and the stable SH-SY5Y-GFP tag-TASK3 cell line to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability of two group cells significantly reduced with pH declining, and the difference was statistically significant (P cells, the cell viability of stable SH-SYSY-GFP tag-TASK3 cell line increased significantly with the same pH media, and the difference was statistically significant (P eukaryotic expression vector pEGFP-TASK3 is successfully constructed and the cell line stably expressing TASK3-eGFP fusion is established which is important for their fundamental research and potential applications.

  16. A new pH-sensitive rectifying potassium channel in mitochondria from the embryonic rat hippocampus.

    Science.gov (United States)

    Kajma, Anna; Szewczyk, Adam

    2012-10-01

    Patch-clamp single-channel studies on mitochondria isolated from embryonic rat hippocampus revealed the presence of two different potassium ion channels: a large-conductance (288±4pS) calcium-activated potassium channel and second potassium channel with outwardly rectifying activity under symmetric conditions (150/150mM KCl). At positive voltages, this channel displayed a conductance of 67.84pS and a strong voltage dependence at holding potentials from -80mV to +80mV. The open probability was higher at positive than at negative voltages. Patch-clamp studies at the mitoplast-attached mode showed that the channel was not sensitive to activators and inhibitors of mitochondrial potassium channels but was regulated by pH. Moreover, we demonstrated that the channel activity was not affected by the application of lidocaine, an inhibitor of two-pore domain potassium channels, or by tertiapin, an inhibitor of inwardly rectifying potassium channels. In summary, based on the single-channel recordings, we characterised for the first time mitochondrial pH-sensitive ion channel that is selective for cations, permeable to potassium ions, displays voltage sensitivity and does not correspond to any previously described potassium ion channels in the inner mitochondrial membrane. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012). Copyright © 2012 Elsevier B.V. All rights reserved.

  17. Potassium Channels Blockers from the Venom of Androctonus mauretanicus mauretanicus

    Directory of Open Access Journals (Sweden)

    Marie-France Martin-Eauclaire

    2012-01-01

    Full Text Available K+ channels selectively transport K+ ions across cell membranes and play a key role in regulating the physiology of excitable and nonexcitable cells. Their activation allows the cell to repolarize after action potential firing and reduces excitability, whereas channel inhibition increases excitability. In eukaryotes, the pharmacology and pore topology of several structural classes of K+ channels have been well characterized in the past two decades. This information has come about through the extensive use of scorpion toxins. We have participated in the isolation and in the characterization of several structurally distinct families of scorpion toxin peptides exhibiting different K+ channel blocking functions. In particular, the venom from the Moroccan scorpion Androctonus mauretanicus mauretanicus provided several high-affinity blockers selective for diverse K+ channels  (SKCa,  Kv4.x, and  Kv1.x K+ channel families. In this paper, we summarize our work on these toxin/channel interactions.

  18. Structural and functional diversity of acidic scorpion potassium channel toxins.

    Directory of Open Access Journals (Sweden)

    Zong-Yun Chen

    Full Text Available BACKGROUND: Although the basic scorpion K(+ channel toxins (KTxs are well-known pharmacological tools and potential drug candidates, characterization the acidic KTxs still has the great significance for their potential selectivity towards different K(+ channel subtypes. Unfortunately, research on the acidic KTxs has been ignored for several years and progressed slowly. PRINCIPAL FINDINGS: Here, we describe the identification of nine new acidic KTxs by cDNA cloning and bioinformatic analyses. Seven of these toxins belong to three new α-KTx subfamilies (α-KTx28, α-KTx29, and α-KTx30, and two are new members of the known κ-KTx2 subfamily. ImKTx104 containing three disulfide bridges, the first member of the α-KTx28 subfamily, has a low sequence homology with other known KTxs, and its NMR structure suggests ImKTx104 adopts a modified cystine-stabilized α-helix-loop-β-sheet (CS-α/β fold motif that has no apparent α-helixs and β-sheets, but still stabilized by three disulfide bridges. These newly described acidic KTxs exhibit differential pharmacological effects on potassium channels. Acidic scorpion toxin ImKTx104 was the first peptide inhibitor found to affect KCNQ1 channel, which is insensitive to the basic KTxs and is strongly associated with human cardiac abnormalities. ImKTx104 selectively inhibited KCNQ1 channel with a K(d of 11.69 µM, but was less effective against the basic KTxs-sensitive potassium channels. In addition to the ImKTx104 toxin, HeTx204 peptide, containing a cystine-stabilized α-helix-loop-helix (CS-α/α fold scaffold motif, blocked both Kv1.3 and KCNQ1 channels. StKTx23 toxin, with a cystine-stabilized α-helix-loop-β-sheet (CS-α/β fold motif, could inhibit Kv1.3 channel, but not the KCNQ1 channel. CONCLUSIONS/SIGNIFICANCE: These findings characterize the structural and functional diversity of acidic KTxs, and could accelerate the development and clinical use of acidic KTxs as pharmacological tools and

  19. The inhibitor of volume-regulated anion channels DCPIB activates TREK potassium channels in cultured astrocytes

    Czech Academy of Sciences Publication Activity Database

    Minieri, L.; Pivoňková, Helena; Caprini, M.; Harantová, Lenka; Anděrová, Miroslava; Ferroni, S.

    2013-01-01

    Roč. 168, č. 5 (2013), s. 1240-1254 ISSN 0007-1188 R&D Projects: GA ČR GAP303/10/1338 Institutional support: RVO:68378041 Keywords : two-pore-domain potassium channels * patch clamp * neuroprotection Subject RIV: FH - Neurology Impact factor: 4.990, year: 2013

  20. POTASSIUM CHANNELS AS DRUGS TARGETS IN THERAPY OF CARDIOVASCULAR DESEASES: 25 YEARS LATER

    Directory of Open Access Journals (Sweden)

    Protić Dragana

    2013-01-01

    Full Text Available Potassium channels are the most variable ion channel group. They participate in numerous cardiovascular functions, for example regulation of vascular tone, maintenance of resting cardiac membrane potential and excitability of cardiac conduction tissue. Both drugs and endogenous ligands could modulate potassium channel function, belonging to the potassium channel blockers or openers. Modulation of potassium channels could be a therapeutic or adverse drug action. Class III antiarrhythmic agents block the potassium channels, thereby prolonging repolarization phase of action potential with resulting prolongation of effective refractory period. Their effectiveness against supraventricular and ventricular arrhythmias should be weighted against their proarrhythmogenic potential. In addition, numerous other antiarrhythmic agents could modulate potassium channels as well. Diazoxide, minoxidil and nicorandil (well known arterial vasodilators, as well as numerous newly synthesized substances with still unknown therapeutic potential, belong to the potassium channel activators/ openers. Therapeutic use of such vasodilators may involve treatment of hypertension (diazoxide, minoxidil and stable angina (nicorandil. Their use might be accompanied with side effects, such as vasodilation, edema, hypotension and reflex tachycardia. Potassium channel openers have also an important role in the treatment of peripheral vascular disease and pulmonary hypertension. In the future, drugs with selective effects on the vascular or cardiac potassium channels could be useful therapeutic agents.

  1. Potassium channels as drugs targets in therapy of cardiovascular diseases: 25 years later

    Directory of Open Access Journals (Sweden)

    Protić Dragana

    2013-03-01

    Full Text Available Potassium channels are the most variable ion channel group. They participate in numerous cardiovascular functions, for example regulation of vascular tone, maintenance of resting cardiac membrane potential and excitability of cardiac conduction tissue. Both drugs and endogenous ligands could modulate potassium channel function, belonging to the potassium channel blockers or openers. Modulation of potassium channels could be a therapeutic or adverse drug action. Class III antiarrhythmic agents block the potassium channels, thereby prolonging repolarization phase of action potential with resulting prolongation of effective refractory period. Their effectiveness against supraventricular and ventricular arrhythmias should be weighted against their proarrhythmogenic potential. In addition, numerous other antiarrhythmic agents could modulate potassium channels as well. Diazoxide, minoxidil and nicorandil (well known arterial vasodilators, as well as numerous newly synthesized substances with still unknown therapeutic potential, belong to the potassium channel activators/openers. Therapeutic use of such vasodilators may involve treatment of hypertension (diazoxide, minoxidil and stable angina (nicorandil. Their use might be accompanied with side effects, such as vasodilation, edema, hypotension and reflex tachycardia. Potassium channel openers have also an important role in the treatment of peripheral vascular disease and pulmonary hypertension. In the future, drugs with selective effects on the vascular or cardiac potassium channels could be useful therapeutic agents.

  2. Calcium-Activated Potassium Channels in Ischemia Reperfusion: A Brief Update

    Directory of Open Access Journals (Sweden)

    Jean-Yves eTano

    2014-10-01

    Full Text Available Ischemia and reperfusion (IR injury constitutes one of the major causes of cardiovascular morbidity and mortality. The discovery of new therapies to block/mediate the effects of IR is therefore an important goal in the biomedical sciences. Dysfunction associated with IR involves modification of calcium-activated potassium channels (KCa through different mechanisms, which are still under study. Respectively, the KCa family, major contributors to plasma membrane calcium influx in cells and essential players in the regulation of the vascular tone are interesting candidates. This family is divided into two groups including the large conductance (BKCa and the small/intermediate conductance (SKCa/IKCa K+ channels. In the heart and brain, these channels have been described to offer protection against IR injury. BKCa and SKCa channels deserve special attention since new data demonstrate that these channels are also expressed in mitochondria. More studies are however needed to fully determine their potential use as therapeutic targets.

  3. Immunolocalization and expression of small-conductance calcium-activated potassium channels in human myometrium

    DEFF Research Database (Denmark)

    Rosenbaum, Sofia T; Svalø, Julie; Nielsen, Karsten

    2012-01-01

    Small-conductance calcium-activated potassium (SK3) channels have been detected in human myometrium and we have previously shown a functional role of SK channels in human myometrium in vitro. The aims of this study were to identify the precise localization of SK3 channels and to quantify SK3 m...... muscle cells, and that the molecular expression of SK3 channels is higher in non-pregnant compared to pregnant myometrium. On the basis of our previous study and the present findings, we propose that SK3 activators reduce contractility in human myometrium by modulating telocyte function....... This is the first report to provide evidence for a possible role of SK3 channels in human uterine telocytes....

  4. The large-conductance calcium-activated potassium channel holds the key to the conundrum of familial hypokalemic periodic paralysis

    Science.gov (United States)

    Kim, Sung-Jo; Kang, Sun-Yang; Yi, Jin Woong; Kim, Seung-Min

    2014-01-01

    Purpose Familial hypokalemic periodic paralysis (HOKPP) is an autosomal dominant channelopathy characterized by episodic attacks of muscle weakness and hypokalemia. Mutations in the calcium channel gene, CACNA1S, or the sodium channel gene, SCN4A, have been found to be responsible for HOKPP; however, the mechanism that causes hypokalemia remains to be determined. The aim of this study was to improve the understanding of this mechanism by investigating the expression of calcium-activated potassium (KCa) channel genes in HOKPP patients. Methods We measured the intracellular calcium concentration with fura-2-acetoxymethyl ester in skeletal muscle cells of HOKPP patients and healthy individuals. We examined the mRNA and protein expression of KCa channel genes (KCNMA1, KCNN1, KCNN2, KCNN3, and KCNN4) in both cell types. Results Patient cells exhibited higher cytosolic calcium levels than normal cells. Quantitative reverse transcription polymerase chain reaction analysis showed that the mRNA levels of the KCa channel genes did not significantly differ between patient and normal cells. However, western blot analysis showed that protein levels of the KCNMA1 gene, which encodes KCa1.1 channels (also called big potassium channels), were significantly lower in the membrane fraction and higher in the cytosolic fraction of patient cells than normal cells. When patient cells were exposed to 50 mM potassium buffer, which was used to induce depolarization, the altered subcellular distribution of BK channels remained unchanged. Conclusion These findings suggest a novel mechanism for the development of hypokalemia and paralysis in HOKPP and demonstrate a connection between disease-associated mutations in calcium/sodium channels and pathogenic changes in nonmutant potassium channels. PMID:25379045

  5. Diclofenac distinguishes among homomeric and heteromeric potassium channels composed of KCNQ4 and KCNQ5 subunits.

    Science.gov (United States)

    Brueggemann, Lioubov I; Mackie, Alexander R; Martin, Jody L; Cribbs, Leanne L; Byron, Kenneth L

    2011-01-01

    KCNQ4 and KCNQ5 potassium channel subunits are expressed in vascular smooth muscle cells, although it remains uncertain how these subunits assemble to form functional channels. Using patch-clamp techniques, we compared the electrophysiological characteristics and effects of diclofenac, a known KCNQ channel activator, on human KCNQ4 and KCNQ5 channels expressed individually or together in A7r5 rat aortic smooth muscle cells. The conductance curves of the overexpressed channels were fitted by a single Boltzmann function in each case (V(0.5) values: -31, -44, and -38 mV for KCNQ4, KCNQ5, and KCNQ4/5, respectively). Diclofenac (100 μM) inhibited KCNQ5 channels, reducing maximum conductance by 53%, but increased maximum conductance of KCNQ4 channels by 38%. The opposite effects of diclofenac on KCNQ4 and KCNQ5 could not be attributed to the presence of a basic residue (lysine) in the voltage-sensing domain of KCNQ5, because mutation of this residue to neutral glycine (the residue present in KCNQ4) resulted in a more effective block of the channel. Differences in deactivation rates and distinct voltage-dependent effects of diclofenac on channel activation and deactivation observed with each of the subunit combinations (KCNQ4, KCNQ5, and KCNQ4/5) were used as diagnostic tools to evaluate native KCNQ currents in vascular smooth muscle cells. A7r5 cells express only KCNQ5 channels endogenously, and their responses to diclofenac closely resembled those of the overexpressed KCNQ5 currents. In contrast, mesenteric artery myocytes, which express both KCNQ4 and KCNQ5 channels, displayed whole-cell KCNQ currents with properties and diclofenac responses characteristic of overexpressed heteromeric KCNQ4/5 channels.

  6. Fear conditioning suppresses large-conductance calcium-activated potassium channels in lateral amygdala neurons.

    Science.gov (United States)

    Sun, P; Zhang, Q; Zhang, Y; Wang, F; Wang, L; Yamamoto, R; Sugai, T; Kato, N

    2015-01-01

    It was previously shown that depression-like behavior is accompanied with suppression of the large-conductance calcium activated potassium (BK) channel in cingulate cortex pyramidal cells. To test whether BK channels are also involved in fear conditioning, we studied neuronal properties of amygdala principal cells in fear conditioned mice. After behavior, we made brain slices containing the amygdala, the structure critically relevant to fear memory. The resting membrane potential in lateral amygdala (LA) neurons obtained from fear conditioned mice (FC group) was more depolarized than in neurons from naïve controls. The frequencies of spikes evoked by current injections were higher in neurons from FC mice, demonstrating that excitability of LA neurons was elevated by fear conditioning. The depolarization in neurons from FC mice was shown to depend on BK channels by using the BK channel blocker charybdotoxin. Suppression of BK channels in LA neurons from the FC group was further confirmed on the basis of the spike width, since BK channels affect the descending phase of spikes. Spikes were broader in the FC group than those in the naïve control in a manner dependent on BK channels. Consistently, quantitative real-time PCR revealed a decreased expression of BK channel mRNA. The present findings suggest that emotional disorder manifested in the forms of fear conditioning is accompanied with BK channel suppression in the amygdala, the brain structure critical to this emotional disorder. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Novel Leishmania and Malaria Potassium Channels: Candidate Therapeutic Targets

    National Research Council Canada - National Science Library

    McDonald, Thomas V

    2005-01-01

    .... major and T. cruzi). Using a combination of cultured mammalian cells and Xenopus oocytes for heterologous expression we have evidence that 2 channels from malaria [PFK1 & PFK22] and Leishmania [LMK1 & LMK2] generate K+...

  8. Long-pore Electrostatics in Inward-rectifier Potassium Channels

    Science.gov (United States)

    Robertson, Janice L.; Palmer, Lawrence G.; Roux, Benoît

    2008-01-01

    Inward-rectifier potassium (Kir) channels differ from the canonical K+ channel structure in that they possess a long extended pore (∼85 Å) for ion conduction that reaches deeply into the cytoplasm. This unique structural feature is presumably involved in regulating functional properties specific to Kir channels, such as conductance, rectification block, and ligand-dependent gating. To elucidate the underpinnings of these functional roles, we examine the electrostatics of an ion along this extended pore. Homology models are constructed based on the open-state model of KirBac1.1 for four mammalian Kir channels: Kir1.1/ROMK, Kir2.1/IRK, Kir3.1/GIRK, and Kir6.2/KATP. By solving the Poisson-Boltzmann equation, the electrostatic free energy of a K+ ion is determined along each pore, revealing that mammalian Kir channels provide a favorable environment for cations and suggesting the existence of high-density regions in the cytoplasmic domain and cavity. The contribution from the reaction field (the self-energy arising from the dielectric polarization induced by the ion's charge in the complex geometry of the pore) is unfavorable inside the long pore. However, this is well compensated by the electrostatic interaction with the static field arising from the protein charges and shielded by the dielectric surrounding. Decomposition of the static field provides a list of residues that display remarkable correspondence with existing mutagenesis data identifying amino acids that affect conduction and rectification. Many of these residues demonstrate interactions with the ion over long distances, up to 40 Å, suggesting that mutations potentially affect ion or blocker energetics over the entire pore. These results provide a foundation for understanding ion interactions in Kir channels and extend to the study of ion permeation, block, and gating in long, cation-specific pores. PMID:19001143

  9. Potassium channel gene mutations rarely cause atrial fibrillation

    Directory of Open Access Journals (Sweden)

    Nam Edwin G

    2006-08-01

    Full Text Available Abstract Background Mutations in several potassium channel subunits have been associated with rare forms of atrial fibrillation. In order to explore the role of potassium channels in inherited typical forms of the arrhythmia, we have screened a cohort of patients from a referral clinic for mutations in the channel subunit genes implicated in the arrhythmia. We sought to determine if mutations in KCNJ2 and KCNE1-5 are a common cause of atrial fibrillation. Methods Serial patients with lone atrial fibrillation or atrial fibrillation with hypertension were enrolled between June 1, 2001 and January 6, 2005. Each patient underwent a standardized interview and physical examination. An electrocardiogram, echocardiogram and blood sample for genetic analysis were also obtained. Patients with a family history of AF were screened for mutations in KCNJ2 and KCNE1-5 using automated sequencing. Results 96 patients with familial atrial fibrillation were enrolled. Eighty-three patients had lone atrial fibrillation and 13 had atrial fibrillation and hypertension. Patients had a mean age of 56 years at enrollment and 46 years at onset of atrial fibrillation. Eighty-one percent of patients had paroxysmal atrial fibrillation at enrollment. Unlike patients with an activating mutation in KCNQ1, the patients had a normal QTc interval with a mean of 412 ± 42 ms. Echocardiography revealed a normal mean ejection fraction of 62.0 ± 7.2 % and mean left atrial dimension of 39.9 ± 7.0 mm. A number of common polymorphisms in KCNJ2 and KCNE1-5 were identified, but no mutations were detected. Conclusion Mutations in KCNJ2 and KCNE1-5 rarely cause typical atrial fibrillation in a referral clinic population.

  10. Inwardly rectifying potassium channels influence Drosophila wing morphogenesis by regulating Dpp release.

    Science.gov (United States)

    Dahal, Giri Raj; Pradhan, Sarala Joshi; Bates, Emily Anne

    2017-08-01

    Loss of embryonic ion channel function leads to morphological defects, but the underlying reason for these defects remains elusive. Here, we show that inwardly rectifying potassium (Irk) channels regulate release of the Drosophila bone morphogenetic protein Dpp in the developing fly wing and that this is necessary for developmental signaling. Inhibition of Irk channels decreases the incidence of distinct Dpp-GFP release events above baseline fluorescence while leading to a broader distribution of Dpp-GFP. Work by others in different cell types has shown that Irk channels regulate peptide release by modulating membrane potential and calcium levels. We found calcium transients in the developing wing, and inhibition of Irk channels reduces the duration and amplitude of calcium transients. Depolarization with high extracellular potassium evokes Dpp release. Taken together, our data implicate Irk channels as a requirement for regulated release of Dpp, highlighting the importance of the temporal pattern of Dpp presentation for morphogenesis of the wing. © 2017. Published by The Company of Biologists Ltd.

  11. Inhibition of HERG potassium channels by celecoxib and its mechanism.

    Directory of Open Access Journals (Sweden)

    Roman V Frolov

    Full Text Available Celecoxib (Celebrex, a widely prescribed selective inhibitor of cyclooxygenase-2, can modulate ion channels independently of cyclooxygenase inhibition. Clinically relevant concentrations of celecoxib can affect ionic currents and alter functioning of neurons and myocytes. In particular, inhibition of Kv2.1 channels by celecoxib leads to arrhythmic beating of Drosophila heart and of rat heart cells in culture. However, the spectrum of ion channels involved in human cardiac excitability differs from that in animal models, including mammalian models, making it difficult to evaluate the relevance of these observations to humans. Our aim was to examine the effects of celecoxib on hERG and other human channels critically involved in regulating human cardiac rhythm, and to explore the mechanisms of any observed effect on the hERG channels.Celecoxib inhibited the hERG, SCN5A, KCNQ1 and KCNQ1/MinK channels expressed in HEK-293 cells with IC(50s of 6.0 µM, 7.5 µM, 3.5 µM and 3.7 µM respectively, and the KCND3/KChiP2 channels expressed in CHO cells with an IC(50 of 10.6 µM. Analysis of celecoxib's effects on hERG channels suggested gating modification as the mechanism of drug action.The above channels play a significant role in drug-induced long QT syndrome (LQTS and short QT syndrome (SQTS. Regulatory guidelines require that all new drugs under development be tested for effects on the hERG channel prior to first administration in humans. Our observations raise the question of celecoxib's potential to induce cardiac arrhythmias or other channel related adverse effects, and make a case for examining such possibilities.

  12. Differential distribution of the sodium-activated potassium channels slick and slack in mouse brain.

    Science.gov (United States)

    Rizzi, Sandra; Knaus, Hans-Günther; Schwarzer, Christoph

    2016-07-01

    The sodium-activated potassium channels Slick (Slo2.1, KCNT2) and Slack (Slo2.2, KCNT1) are high-conductance potassium channels of the Slo family. In neurons, Slick and Slack channels are involved in the generation of slow afterhyperpolarization, in the regulation of firing patterns, and in setting and stabilizing the resting membrane potential. The distribution and subcellular localization of Slick and Slack channels in the mouse brain have not yet been established in detail. The present study addresses this issue through in situ hybridization and immunohistochemistry. Both channels were widely distributed and exhibited distinct distribution patterns. However, in some brain regions, their expression overlapped. Intense Slick channel immunoreactivity was observed in processes, varicosities, and neuronal cell bodies of the olfactory bulb, granular zones of cortical regions, hippocampus, amygdala, lateral septal nuclei, certain hypothalamic and midbrain nuclei, and several regions of the brainstem. The Slack channel showed primarily a diffuse immunostaining pattern, and labeling of cell somata and processes was observed only occasionally. The highest Slack channel expression was detected in the olfactory bulb, lateral septal nuclei, basal ganglia, and distinct areas of the midbrain, brainstem, and cerebellar cortex. In addition, comparing our data obtained from mouse brain with a previously published study on rat brain revealed some differences in the expression and distribution of Slick and Slack channels in these species. J. Comp. Neurol. 524:2093-2116, 2016. © 2015 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc. © 2015 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc.

  13. Potassium channel activator attenuates salicylate-induced cochlear hearing loss potentially ameliorating tinnitus.

    Science.gov (United States)

    Sun, Wei; Liu, Jun; Zhang, Chao; Zhou, Na; Manohar, Senthilvelan; Winchester, Wendy; Miranda, Jason A; Salvi, Richard J

    2015-01-01

    High dose sodium salicylate causes moderate, reversible hearing loss and tinnitus. Salicylate-induced hearing loss is believed to arise from a reduction in the electromotile response of outer hair cells (OHCs) and/or reduction of KCNQ4 potassium currents in OHCs, which decreases the driving force for the transduction current. Therefore, enhancing OHC potassium currents could potentially prevent salicylate-induced temporary hearing loss. In this study, we tested whether opening voltage-gated potassium channels using ICA-105665, a novel small molecule that opens KCNQ2/3 and KCNQ3/5 channels, can reduce salicylate-induced hearing loss. We found that systemic application of ICA-105665 at 10 mg/kg prevented the salicylate-induced amplitude reduction and threshold shift in the compound action potentials recorded at the round window of the cochlea. ICA-105665 also prevented the salicylate-induced reduction of distortion-product otoacoustic emission. These results suggest that ICA-105665 partially compensates for salicylate-induced cochlear hearing loss by enhancing KCNQ2/3 and KCNQ3/5 potassium currents and the motility of OHCs.

  14. The neuroglial potassium cycle during neurotransmission: role of Kir4.1 channels.

    Directory of Open Access Journals (Sweden)

    Jérémie Sibille

    2015-03-01

    Full Text Available Neuronal excitability relies on inward sodium and outward potassium fluxes during action potentials. To prevent neuronal hyperexcitability, potassium ions have to be taken up quickly. However, the dynamics of the activity-dependent potassium fluxes and the molecular pathways underlying extracellular potassium homeostasis remain elusive. To decipher the specific and acute contribution of astroglial Kir4.1 channels in controlling potassium homeostasis and the moment to moment neurotransmission, we built a tri-compartment model accounting for potassium dynamics between neurons, astrocytes and the extracellular space. We here demonstrate that astroglial Kir4.1 channels are sufficient to account for the slow membrane depolarization of hippocampal astrocytes and crucially contribute to extracellular potassium clearance during basal and high activity. By quantifying the dynamics of potassium levels in neuron-glia-extracellular space compartments, we show that astrocytes buffer within 6 to 9 seconds more than 80% of the potassium released by neurons in response to basal, repetitive and tetanic stimulations. Astroglial Kir4.1 channels directly lead to recovery of basal extracellular potassium levels and neuronal excitability, especially during repetitive stimulation, thereby preventing the generation of epileptiform activity. Remarkably, we also show that Kir4.1 channels strongly regulate neuronal excitability for slow 3 to 10 Hz rhythmic activity resulting from probabilistic firing activity induced by sub-firing stimulation coupled to Brownian noise. Altogether, these data suggest that astroglial Kir4.1 channels are crucially involved in extracellular potassium homeostasis regulating theta rhythmic activity.

  15. Terbinafine is a novel and selective activator of the two-pore domain potassium channel TASK3

    OpenAIRE

    Wright, Paul D.; Veale, Emma L.; McCoull, David; Tickle, David C.; Large, Jonathan M.; Ococks, Emma; Gothard, Gemma; Kettleborough, Catherine; Mathie, Alistair; Jerman, Jeffrey

    2017-01-01

    Two-pore domain potassium channels (K2Ps) are characterized by their four transmembrane domain and two-pore topology. They carry background (or leak) potassium current in a variety of cell types. Despite a number of important roles there is currently a lack of pharmacological tools with which to further probe K2P function. We have developed a cell-based thallium flux assay, using baculovirus delivered TASK3 (TWIK-related acid-sensitive K+ channel 3, KCNK9, K2P9.1) with the aim of identifying ...

  16. The potassium channel KCa3.1 as new therapeutic target for the prevention of obliterative airway disease

    DEFF Research Database (Denmark)

    Hua, Xiaoqin; Deuse, Tobias; Chen, Yi-Je

    2013-01-01

    The calcium-activated potassium channel KCa3.1 is critically involved in T-cell activation as well as in the proliferation of smooth muscle cells and fibroblasts. We sought to investigate whether KCa3.1 contributes to the pathogenesis of obliterative airway disease (OAD) and whether knockout or p...

  17. The endosomal trafficking factors CORVET and ESCRT suppress plasma membrane residence of the renal outer medullary potassium channel (ROMK).

    Science.gov (United States)

    Mackie, Timothy D; Kim, Bo-Young; Subramanya, Arohan R; Bain, Daniel J; O'Donnell, Allyson F; Welling, Paul A; Brodsky, Jeffrey L

    2018-03-02

    Protein trafficking can act as the primary regulatory mechanism for ion channels with high open probabilities, such as the r enal o uter m edullary (ROMK) channel. ROMK, also known as Kir1.1 (KCNJ1), is the major route for potassium secretion into the pro-urine and plays an indispensable role in regulating serum potassium and urinary concentrations. However, the cellular machinery that regulates ROMK trafficking has not been fully defined. To identify regulators of the cell-surface population of ROMK, we expressed a pH-insensitive version of the channel in the budding yeast Saccharomyces cerevisiae We determined that ROMK primarily resides in the endoplasmic reticulum (ER), as it does in mammalian cells, and is subject to ER-associated degradation (ERAD). However, sufficient ROMK levels on the plasma membrane rescued growth on low-potassium medium of yeast cells lacking endogenous potassium channels. Next, we aimed to identify the biological pathways most important for ROMK regulation. Therefore, we used a synthetic genetic array to identify non-essential genes that reduce the plasma membrane pool of ROMK in potassium-sensitive yeast cells. Genes identified in this screen included several members of the endosomal complexes required for transport (ESCRT) and the class-C core vacuole/endosome tethering (CORVET) complexes. Mass spectroscopy analysis confirmed that yeast cells lacking an ESCRT component accumulate higher potassium concentrations. Moreover, silencing of ESCRT and CORVET components increased ROMK levels at the plasma membrane in HEK293 cells. Our results indicate that components of the post-endocytic pathway influence the cell-surface density of ROMK and establish that components in this pathway modulate channel activity.

  18. Mechanisms underlying KCNQ1channel cell volume sensitivity

    DEFF Research Database (Denmark)

    Hammami, Sofia

    Cells are constantly exposed to changes in cell volume during cell metabolism, nutrient uptake, cell proliferation, cell migration and salt and water transport. In order to cope with these perturbations, potassium channels in line with chloride channels have been shown to be likely contributors t...

  19. Slo1 is the principal potassium channel of human spermatozoa

    Science.gov (United States)

    Mannowetz, Nadja; Naidoo, Natasha M; Choo, Seung-A Sara; Smith, James F; Lishko, Polina V

    2013-01-01

    Mammalian spermatozoa gain competence to fertilize an oocyte as they travel through the female reproductive tract. This process is accompanied by an elevation of sperm intracellular calcium and a membrane hyperpolarization. The latter is evoked by K+ efflux; however, the molecular identity of the potassium channel of human spermatozoa (hKSper) is unknown. Here, we characterize hKSper, reporting that it is regulated by intracellular calcium but is insensitive to intracellular alkalinization. We also show that human KSper is inhibited by charybdotoxin, iberiotoxin, and paxilline, while mouse KSper is insensitive to these compounds. Such unique properties suggest that the Slo1 ion channel is the molecular determinant for hKSper. We show that Slo1 is localized to the sperm flagellum and is inhibited by progesterone. Inhibition of hKSper by progesterone may depolarize the spermatozoon to open the calcium channel CatSper, thus raising [Ca2+] to produce hyperactivation and allowing sperm to fertilize an oocyte. DOI: http://dx.doi.org/10.7554/eLife.01009.001 PMID:24137539

  20. Hydrogen bonds as molecular timers for slow inactivation in voltage-gated potassium channels

    DEFF Research Database (Denmark)

    Pless, Stephan Alexander; Galpin, Jason D; Niciforovic, Ana P

    2013-01-01

    Voltage-gated potassium (Kv) channels enable potassium efflux and membrane repolarization in excitable tissues. Many Kv channels undergo a progressive loss of ion conductance in the presence of a prolonged voltage stimulus, termed slow inactivation, but the atomic determinants that regulate the k...... subunit(s). DOI: http://dx.doi.org/10.7554/eLife.01289.001....

  1. KCNE3 is an inhibitory subunit of the Kv4.3 potassium channel

    DEFF Research Database (Denmark)

    Lundby, Alicia; Olesen, Søren-Peter

    2006-01-01

    The mammalian Kv4.3 potassium channel is a fast activating and inactivating K+ channel widely distributed in mammalian tissues. Kv4.3 is the major component of various physiologically important currents ranging from A-type currents in the CNS to the transient outward potassium conductance...

  2. The inwardly rectifying potassium channel Kir1.1: development of functional assays to identify and characterize channel inhibitors.

    Science.gov (United States)

    Felix, John P; Priest, Birgit T; Solly, Kelli; Bailey, Timothy; Brochu, Richard M; Liu, Chou J; Kohler, Martin G; Kiss, Laszlo; Alonso-Galicia, Magdalena; Tang, Haifeng; Pasternak, Alexander; Kaczorowski, Gregory J; Garcia, Maria L

    2012-10-01

    The renal outer medullary potassium (ROMK) channel is a member of the inwardly rectifying family of potassium (Kir) channels. ROMK (Kir1.1) is predominantly expressed in kidney where it plays a major role in the salt reabsorption process. Loss-of-function mutations in the human Kir1.1 channel are associated with antenatal Bartter's syndrome type II, a life-threatening salt and water balance disorder. Heterozygous carriers of Kir1.1 mutations associated with antenatal Bartter's syndrome have reduced blood pressure and a decreased risk of developing hypertension by age 60. These data suggest that Kir1.1 inhibitors could represent novel diuretics for the treatment of hypertension. Because little is known about the molecular pharmacology of Kir1.1 channels, assays that provide a robust, reliable readout of channel activity-while operating in high-capacity mode-are needed. In the present study, we describe high-capacity, 384- and 1,536-well plate, functional thallium flux, and IonWorks electrophysiology assays for the Kir1.1 channel that fulfill these criteria. In addition, 96-well (86)Rb(+) flux assays were established that can operate in the presence of 100% serum, and can provide an indication of the effect of a serum shift on compound potencies. The ability to grow Madin-Darby canine kidney cells expressing Kir1.1 in Transwell supports provides a polarized cell system that can be used to study the mechanism of Kir1.1 inhibition by different agents. All these functional Kir1.1 assays together can play an important role in supporting different aspects of drug development efforts during lead identification and/or optimization.

  3. Free RCK arrangement in Kch, a putative escherichia coli potassium channel, as suggested by electron crystallography.

    Science.gov (United States)

    Kuang, Qie; Purhonen, Pasi; Jegerschöld, Caroline; Koeck, Philip J B; Hebert, Hans

    2015-01-06

    The ligand-gated potassium channels are stimulated by various kinds of messengers. Previous studies showed that ligand-gated potassium channels containing RCK domains (the regulator of the conductance of potassium ion) form a dimer of tetramer structure through the RCK octameric gating ring in the presence of detergent. Here, we have analyzed the structure of Kch, a channel of this type from Escherichia coli, in a lipid environment using electron crystallography. By combining information from the 3D map of the transmembrane part of the protein and docking of an atomic model of a potassium channel, we conclude that the RCK domains face the solution and that an RCK octameric gating ring arrangement does not form under our crystallization condition. Our findings may be applied to other potassium channels that have an RCK gating ring arrangement. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Potassium channels in the central nervous system of the snail, Helix pomatia: localization and functional characterization.

    Science.gov (United States)

    Battonyai, I; Krajcs, N; Serfőző, Z; Kiss, T; Elekes, K

    2014-05-30

    The distribution and functional presence of three voltage-dependent potassium channels, Kv2.1, Kv3.4, Kv4.3, respectively, were studied in the central nervous system of the snail Helix pomatia by immunohistochemical and electrophysiological methods. Cell clusters displaying immunoreactivity for the different channels were observed in all parts of the CNS, although their localization and number partly varied. Differences were also found in their intracellular, perikaryonal and axonal localization, as well as in their presence in non-neuronal tissues nearby the CNS, such as the perineurium and the aorta wall. At ultrastructural level Kv4.3 channel immunolabeling was observed in axon profiles containing large 80-100nm granular vesicles. Blotting analyses revealed specific signals for the Kv2.1, Kv3.4 and Kv4.3 channels, confirming the presence of the channels in the Helix CNS. Voltage-clamp recordings proved that outward currents obtained from neurons displaying Kv3.4 or Kv4.3 immunoreactivity contained transient components while Kv2.1 immunoreactive cells were characterized by delayed currents. The distribution of the K(+)-channels containing neurons suggests specific roles in intercellular signaling processes in the Helix CNS, most probably related to well-defined, partly local events. The cellular localization of the K(+)-channels studied supports their involvement in both pre- and postsynaptic events at perikaryonal and axonal levels. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  5. [Mechanism of potassium channel in hypoxia-ischemic brain edema: experiment with neonatal rat astrocyte].

    Science.gov (United States)

    Fu, Xue-mei; Xiang, Long; Liao, Da-qing; Feng, Zhi-chun; Mu, De-zhi

    2008-11-04

    To investigate the mechanism of potassium channel in brain edema caused by hypoxia-ischemia (HI). Astrocytes were obtained from 3-day-old SD rats, cultured, and randomly divided into 2 groups: normoxia group, cultured under normoxic condition, and hypoxic-ischemic group, cultured under hypoxic-ischemic condition. The cell volume was measured by radiologic method. Patch-clamp technique was used to observe the electric physiological properties of the voltage-gated potassium channels (Kv) in a whole cell configuration, and the change of voltage-gated potassium channel current (IKv) was recorded in cultured neonatal rat astrocyte during HI. Aquaporin 4 (AQP4) expression vector was constructed from pSUPER vector and transfected into the astrocytes (AQP4 RNAi) to construct AQP4 knockdown (AQP4-/-) cells. cellular volume was determined using [3H]-3-O-methyl-D-glucose uptake in both AQP4-/- and AQP4+/+ cells under the condition of HI. Real time PCR and Western blotting were used to detect the mRNA and protein expression of AQP4. The percentages of the AQP4+/+ and AQP4-/- astrocyte volumes in the condition of HI for 0.5, 1, 2, and 4 h were 104+/-7, 109+/-6, 126+/-12, and 152+/-9 times, and 97+/-7, 105+/-9, 109+/-7, and 132+/-6 times as those of their corresponding control groups (all Pastrocytes significantly increased during HI and the degrees of edema mediated by AQP4 knockdown at different time points were all significantly milder (all Pastrocytes via aquaporin-4 and then cell swelling.

  6. Differential expression of the Kv1 voltage-gated potassium channel family in the rat nephron.

    Science.gov (United States)

    Carrisoza-Gaytán, Rolando; Salvador, Carolina; Diaz-Bello, Beatriz; Escobar, Laura I

    2014-10-01

    Several potassium (K(+)) channels contribute to maintaining the resting membrane potential of renal epithelial cells. Apart from buffering the cell membrane potential and cell volume, K(+) channels allow sodium reabsorption in the proximal tubule (PT), K(+) recycling and K(+) reabsorption in the thick ascending limb (TAL) and K(+) secretion and K(+) reabsorption in the distal convoluted tubule (DCT), connecting tubule (CNT) and collecting duct. Previously, we identified Kv.1.1, Kv1.3 and Kv1.6 channels in collecting ducts of the rat inner medulla. We also detected intracellular Kv1.3 channel in the acid secretory intercalated cells, which is trafficked to the apical membrane in response to dietary K(+) to function as a secretory K(+) channel. In this work we sought to characterize the expression of all members of the Kv1 family in the rat nephron. mRNA and protein expression were detected for all Kv1 channels. Immunoblots identified differential expression of each Kv1 in the cortex, outer and inner medulla. Immunofluorescence labeling detected Kv1.5 in Bowman´s capsule and endothelial cells and Kv1.7 in podocytes, endothelial cells and macula densa in glomeruli; Kv1.4, Kv1.5 and Kv1.7 in PT; Kv1.2, Kv1.4 and Kv1.6 in TAL; Kv1.1, Kv1.4 and Kv1.6 in DCT and CNT and Kv1.3 in DCT, and all the Kv1 family in the cortical and medullary collecting ducts. Recently, some hereditary renal syndromes have been attributed to mutations in K(+) channels. Our results expand the repertoire of K(+) channels that contribute to K(+) homeostasis to include the Kv1 family.

  7. Nitric oxide regulates neuronal activity via calcium-activated potassium channels.

    Directory of Open Access Journals (Sweden)

    Lei Ray Zhong

    Full Text Available Nitric oxide (NO is an unconventional membrane-permeable messenger molecule that has been shown to play various roles in the nervous system. How NO modulates ion channels to affect neuronal functions is not well understood. In gastropods, NO has been implicated in regulating the feeding motor program. The buccal motoneuron, B19, of the freshwater pond snail Helisoma trivolvis is active during the hyper-retraction phase of the feeding motor program and is located in the vicinity of NO-producing neurons in the buccal ganglion. Here, we asked whether B19 neurons might serve as direct targets of NO signaling. Previous work established NO as a key regulator of growth cone motility and neuronal excitability in another buccal neuron involved in feeding, the B5 neuron. This raised the question whether NO might modulate the electrical activity and neuronal excitability of B19 neurons as well, and if so whether NO acted on the same or a different set of ion channels in both neurons. To study specific responses of NO on B19 neurons and to eliminate indirect effects contributed by other cells, the majority of experiments were performed on single cultured B19 neurons. Addition of NO donors caused a prolonged depolarization of the membrane potential and an increase in neuronal excitability. The effects of NO could mainly be attributed to the inhibition of two types of calcium-activated potassium channels, apamin-sensitive and iberiotoxin-sensitive potassium channels. NO was found to also cause a depolarization in B19 neurons in situ, but only after NO synthase activity in buccal ganglia had been blocked. The results suggest that NO acts as a critical modulator of neuronal excitability in B19 neurons, and that calcium-activated potassium channels may serve as a common target of NO in neurons.

  8. Scorpion Toxins Specific for Potassium (K+ Channels: A Historical Overview of Peptide Bioengineering

    Directory of Open Access Journals (Sweden)

    Zachary L. Bergeron

    2012-11-01

    Full Text Available Scorpion toxins have been central to the investigation and understanding of the physiological role of potassium (K+ channels and their expansive function in membrane biophysics. As highly specific probes, toxins have revealed a great deal about channel structure and the correlation between mutations, altered regulation and a number of human pathologies. Radio- and fluorescently-labeled toxin isoforms have contributed to localization studies of channel subtypes in expressing cells, and have been further used in competitive displacement assays for the identification of additional novel ligands for use in research and medicine. Chimeric toxins have been designed from multiple peptide scaffolds to probe channel isoform specificity, while advanced epitope chimerization has aided in the development of novel molecular therapeutics. Peptide backbone cyclization has been utilized to enhance therapeutic efficiency by augmenting serum stability and toxin half-life in vivo as a number of K+-channel isoforms have been identified with essential roles in disease states ranging from HIV, T-cell mediated autoimmune disease and hypertension to various cardiac arrhythmias and Malaria. Bioengineered scorpion toxins have been monumental to the evolution of channel science, and are now serving as templates for the development of invaluable experimental molecular therapeutics.

  9. The KCNQ1 potassium channel is down-regulated by ubiquitylating enzymes of the Nedd4/Nedd4-like family

    DEFF Research Database (Denmark)

    Jespersen, Thomas; Membrez, Mathieu; Nicolas, Céline S

    2007-01-01

    OBJECTIVE: The voltage-gated KCNQ1 potassium channel regulates key physiological functions in a number of tissues. In the heart, KCNQ1 alpha-subunits assemble with KCNE1 beta-subunits forming a channel complex constituting the delayed rectifier current I(Ks). In epithelia, KCNQ1 channels...... that KCNQ1 internalization and stability is physiologically regulated by its Nedd4/Nedd4-like-dependent ubiquitylation. This mechanism may thereby be important in regulating the surface density of the KCNQ1 channels in cardiomyocytes and other cell types....

  10. Pulmonary vasoconstrictor action of KCNQ potassium channel blockers

    Directory of Open Access Journals (Sweden)

    Balan Prabhu

    2006-02-01

    Full Text Available Abstract Background KCNQ channels have been widely studied in the nervous system, heart and inner ear, where they have important physiological functions. Recent reports indicate that KCNQ channels may also be expressed in portal vein where they are suggested to influence spontaneous contractile activity. The biophysical properties of K+ currents mediated by KCNQ channels resemble a current underlying the resting K+ conductance and resting potential of pulmonary artery smooth muscle cells. We therefore investigated a possible role of KCNQ channels in regulating the function of pulmonary arteries by determining the ability of the selective KCNQ channel blockers, linopirdine and XE991, to promote pulmonary vasoconstriction. Methods The tension developed by rat and mouse intrapulmonary or mesenteric arteries was measured using small vessel myography. Contractile responses to linopirdine and XE991 were measured in intact and endothelium denuded vessels. Experiments were also carried out under conditions that prevent the contractile effects of nerve released noradrenaline or ATP, or block various Ca2+ influx pathways, in order to investigate the mechanisms underlying contraction. Results Linopirdine and XE991 both contracted rat and mouse pulmonary arteries but had little effect on mesenteric arteries. In each case the maximum contraction was almost as large as the response to 50 mM K+. Linopirdine had an EC50 of around 1 μM and XE991 was almost 10-fold more potent. Neither removal of the endothelium nor exposure to phentolamine or α,β-methylene ATP, to block α1-adrenoceptors or P2X receptors, respectively, affected the contraction. Contraction was abolished in Ca2+-free solution and in the presence of 1 μM nifedipine or 10 μM levcromakalim. Conclusion The KCNQ channel blockers are potent and powerful constrictors of pulmonary arteries. This action may be selective for the pulmonary circulation as mesenteric arteries showed little response. The

  11. Escitalopram block of hERG potassium channels.

    Science.gov (United States)

    Chae, Yun Ju; Jeon, Ji Hyun; Lee, Hong Joon; Kim, In-Beom; Choi, Jin-Sung; Sung, Ki-Wug; Hahn, Sang June

    2014-01-01

    Escitalopram, a selective serotonin reuptake inhibitor, is the pharmacologically active S-enantiomer of the racemic mixture of RS-citalopram and is widely used in the treatment of depression. The effects of escitalopram and citalopram on the human ether-a-go-go-related gene (hERG) channels expressed in human embryonic kidney cells were investigated using voltage-clamp and Western blot analyses. Both drugs blocked hERG currents in a concentration-dependent manner with an IC50 value of 2.6 μM for escitalopram and an IC50 value of 3.2 μM for citalopram. The blocking of hERG by escitalopram was voltage-dependent, with a steep increase across the voltage range of channel activation. However, voltage independence was observed over the full range of activation. The blocking by escitalopram was frequency dependent. A rapid application of escitalopram induced a rapid and reversible blocking of the tail current of hERG. The extent of the blocking by escitalopram during the depolarizing pulse was less than that during the repolarizing pulse, suggesting that escitalopram has a high affinity for the open state of the hERG channel, with a relatively lower affinity for the inactivated state. Both escitalopram and citalopram produced a reduction of hERG channel protein trafficking to the plasma membrane but did not affect the short-term internalization of the hERG channel. These results suggest that escitalopram blocked hERG currents at a supratherapeutic concentration and that it did so by preferentially binding to both the open and the inactivated states of the channels and by inhibiting the trafficking of hERG channel protein to the plasma membrane.

  12. Cloxyquin (5-chloroquinolin-8-ol) is an activator of the two-pore domain potassium channel TRESK.

    Science.gov (United States)

    Wright, Paul D; Weir, Gregory; Cartland, Jamie; Tickle, David; Kettleborough, Catherine; Cader, M Zameel; Jerman, Jeff

    2013-11-15

    TRESK is a two-pore domain potassium channel. Loss of function mutations have been linked to typical migraine with aura and due to TRESK’s expression pattern and role in neuronal excitability it represents a promising therapeutic target. We developed a cell based assay using baculovirus transduced U20S cells to screen for activators of TRESK. Using a thallium flux system to measure TRESK channel activity we identified Cloxyquin as a novel activator. Cloxyquin was shown to have an EC50 of 3.8 μM in the thallium assay and displayed good selectivity against other potassium channels tested. Activity was confirmed using whole cell patch electrophysiology, with Cloxyquin causing a near two fold increase in outward current. The strategy presented here will be used to screen larger compound libraries with the aim of identifying novel chemical series which may be developed into new migraine prophylactics.

  13. Inhibition by acrolein of light-induced stomatal opening through inhibition of inward-rectifying potassium channels in Arabidopsis thaliana.

    Science.gov (United States)

    Islam, Md Moshiul; Ye, Wenxiu; Matsushima, Daiki; Khokon, Md Atiqur Rahman; Munemasa, Shintaro; Nakamura, Yoshimasa; Murata, Yoshiyuki

    2015-01-01

    Acrolein is a reactive α,β-unsaturated aldehyde derived from lipid peroxides, which are produced in plants under a variety of stress. We investigated effects of acrolein on light-induced stomatal opening using Arabidopsis thaliana. Acrolein inhibited light-induced stomatal opening in a dose-dependent manner. Acrolein at 100 μM inhibited plasma membrane inward-rectifying potassium (Kin) channels in guard cells. Acrolein at 100 μM inhibited Kin channel KAT1 expressed in a heterologous system using Xenopus leaves oocytes. These results suggest that acrolein inhibits light-induced stomatal opening through inhibition of Kin channels in guard cells.

  14. The marine algal toxin azaspiracid is an open state blocker of hERG potassium channels

    Science.gov (United States)

    Twiner, Michael J.; Doucette, Gregory J.; Rasky, Andrew; Huang, Xi-Ping; Roth, Bryan L.; Sanguinetti, Michael C.

    2012-01-01

    Azaspiracids (AZA) are polyether marine dinoflagellate toxins that accumulate in shellfish and represent an emerging human health risk. Although human exposure is primarily manifested by severe and protracted diarrhea, this toxin class has been shown to be highly cytotoxic, a teratogen to developing fish, and a possible carcinogen in mice. Until now, AZA's molecular target(s) has not yet been determined. Using three independent methods (voltage clamp, channel binding assay, and thallium flux assay), we have for the first time demonstrated that AZA1, AZA2, and AZA3 each bind to and block the hERG (human ether-à-go-go related gene) potassium channel heterologously expressed in HEK-293 mammalian cells. Inhibition of K+ current for each AZA analogue was concentration-dependent (IC50 value range: 0.64 - 0.84 μM). The mechanism of hERG channel inhibition by AZA1 was investigated further in Xenopus oocytes where it was shown to be an open state-dependent blocker and, using mutant channels, to interact with F656 but not with Y652 within the S6 transmembrane domain that forms the channel's central pore. AZA1, AZA2, and AZA3 were each shown to inhibit [3H]dofetilide binding to the hERG channel and thallium ion flux through the channel (IC50 value range: 2.1 – 6.6 μM). AZA1 did not block K+ current of the closely related EAG1 channel. Collectively, these data suggest that the AZAs physically block the K+ conductance pathway of hERG1 channels by occluding the cytoplasmic mouth of the open pore. Although the concentrations necessary to block hERG channels are relatively high, AZA-induced blockage may prove to contribute to the toxicological properties of the AZAs. PMID:22856456

  15. Marine algal toxin azaspiracid is an open-state blocker of hERG potassium channels.

    Science.gov (United States)

    Twiner, Michael J; Doucette, Gregory J; Rasky, Andrew; Huang, Xi-Ping; Roth, Bryan L; Sanguinetti, Michael C

    2012-09-17

    Azaspiracids (AZA) are polyether marine dinoflagellate toxins that accumulate in shellfish and represent an emerging human health risk. Although human exposure is primarily manifested by severe and protracted diarrhea, this toxin class has been shown to be highly cytotoxic, a teratogen to developing fish, and a possible carcinogen in mice. Until now, AZA's molecular target has not yet been determined. Using three independent methods (voltage clamp, channel binding assay, and thallium flux assay), we have for the first time demonstrated that AZA1, AZA2, and AZA3 each bind to and block the hERG (human ether-à-go-go related gene) potassium channel heterologously expressed in HEK-293 mammalian cells. Inhibition of K(+) current for each AZA analogue was concentration-dependent (IC(50) value range: 0.64-0.84 μM). The mechanism of hERG channel inhibition by AZA1 was investigated further in Xenopus oocytes where it was shown to be an open-state-dependent blocker and, using mutant channels, to interact with F656 but not with Y652 within the S6 transmembrane domain that forms the channel's central pore. AZA1, AZA2, and AZA3 were each shown to inhibit [(3)H]dofetilide binding to the hERG channel and thallium ion flux through the channel (IC(50) value range: 2.1-6.6 μM). AZA1 did not block the K(+) current of the closely related EAG1 channel. Collectively, these data suggest that the AZAs physically block the K(+) conductance pathway of hERG1 channels by occluding the cytoplasmic mouth of the open pore. Although the concentrations necessary to block hERG channels are relatively high, AZA-induced blockage may prove to contribute to the toxicological properties of the AZAs.

  16. Voltage dependent potassium channel remodeling in murine intestinal smooth muscle hypertrophy induced by partial obstruction.

    Science.gov (United States)

    Liu, Dong-Hai; Huang, Xu; Guo, Xin; Meng, Xiang-Min; Wu, Yi-Song; Lu, Hong-Li; Zhang, Chun-Mei; Kim, Young-chul; Xu, Wen-Xie

    2014-01-01

    Partial obstruction of the small intestine causes obvious hypertrophy of smooth muscle cells and motility disorder in the bowel proximate to the obstruction. To identify electric remodeling of hypertrophic smooth muscles in partially obstructed murine small intestine, the patch-clamp and intracellular microelectrode recording methods were used to identify the possible electric remodeling and Western blot, immunofluorescence and immunoprecipitation were utilized to examine the channel protein expression and phosphorylation level changes in this research. After 14 days of obstruction, partial obstruction caused obvious smooth muscle hypertrophy in the proximally located intestine. The slow waves of intestinal smooth muscles in the dilated region were significantly suppressed, their amplitude and frequency were reduced, whilst the resting membrane potentials were depolarized compared with normal and sham animals. The current density of voltage dependent potassium channel (KV) was significantly decreased in the hypertrophic smooth muscle cells and the voltage sensitivity of KV activation was altered. The sensitivity of KV currents (IKV) to TEA, a nonselective potassium channel blocker, increased significantly, but the sensitivity of IKv to 4-AP, a KV blocker, stays the same. The protein levels of KV4.3 and KV2.2 were up-regulated in the hypertrophic smooth muscle cell membrane. The serine and threonine phosphorylation levels of KV4.3 and KV2.2 were significantly increased in the hypertrophic smooth muscle cells. Thus this study represents the first identification of KV channel remodeling in murine small intestinal smooth muscle hypertrophy induced by partial obstruction. The enhanced phosphorylations of KV4.3 and KV2.2 may be involved in this process.

  17. Voltage dependent potassium channel remodeling in murine intestinal smooth muscle hypertrophy induced by partial obstruction.

    Directory of Open Access Journals (Sweden)

    Dong-Hai Liu

    Full Text Available Partial obstruction of the small intestine causes obvious hypertrophy of smooth muscle cells and motility disorder in the bowel proximate to the obstruction. To identify electric remodeling of hypertrophic smooth muscles in partially obstructed murine small intestine, the patch-clamp and intracellular microelectrode recording methods were used to identify the possible electric remodeling and Western blot, immunofluorescence and immunoprecipitation were utilized to examine the channel protein expression and phosphorylation level changes in this research. After 14 days of obstruction, partial obstruction caused obvious smooth muscle hypertrophy in the proximally located intestine. The slow waves of intestinal smooth muscles in the dilated region were significantly suppressed, their amplitude and frequency were reduced, whilst the resting membrane potentials were depolarized compared with normal and sham animals. The current density of voltage dependent potassium channel (KV was significantly decreased in the hypertrophic smooth muscle cells and the voltage sensitivity of KV activation was altered. The sensitivity of KV currents (IKV to TEA, a nonselective potassium channel blocker, increased significantly, but the sensitivity of IKv to 4-AP, a KV blocker, stays the same. The protein levels of KV4.3 and KV2.2 were up-regulated in the hypertrophic smooth muscle cell membrane. The serine and threonine phosphorylation levels of KV4.3 and KV2.2 were significantly increased in the hypertrophic smooth muscle cells. Thus this study represents the first identification of KV channel remodeling in murine small intestinal smooth muscle hypertrophy induced by partial obstruction. The enhanced phosphorylations of KV4.3 and KV2.2 may be involved in this process.

  18. The role of voltage-gated potassium channels in the regulation of mouse uterine contractility

    Directory of Open Access Journals (Sweden)

    Abel Peter W

    2007-11-01

    Full Text Available Abstract Background Uterine smooth muscle cells exhibit ionic currents that appear to be important in the control of uterine contractility, but how these currents might produce the changes in contractile activity seen in pregnant myometrium has not been established. There are conflicting reports concerning the role of voltage-gated potassium (Kv channels and large-conductance, calcium-activated potassium (BK channels in the regulation of uterine contractility. In this study we provide molecular and functional evidence for a role for Kv channels in the regulation of spontaneous contractile activity in mouse myometrium, and also demonstrate a change in Kv channel regulation of contractility in pregnant mouse myometrium. Methods Functional assays which evaluated the effects of channel blockers and various contractile agonists were accomplished by quantifying contractility of isolated uterine smooth muscle obtained from nonpregnant mice as well as mice at various stages of pregnancy. Expression of Kv channel proteins in isolated uterine smooth muscle was evaluated by Western blots. Results The Kv channel blocker 4-aminopyridine (4-AP caused contractions in nonpregnant mouse myometrium (EC50 = 54 micromolar, maximal effect at 300 micromolar but this effect disappeared in pregnant mice; similarly, the Kv4.2/Kv4.3 blocker phrixotoxin-2 caused contractions in nonpregnant, but not pregnant, myometrium. Contractile responses to 4-AP were not dependent upon nerves, as neither tetrodotoxin nor storage of tissues at room temperature significantly altered these responses, nor were responses dependent upon the presence of the endometrium. Spontaneous contractions and contractions in response to 4-AP did not appear to be mediated by BK, as the BK channel-selective blockers iberiotoxin, verruculogen, or tetraethylammonium failed to affect either spontaneous contractions or 4-AP-elicited responses. A number of different Kv channel alpha subunit proteins were

  19. Effects of donepezil on hERG potassium channels.

    Science.gov (United States)

    Chae, Yun Ju; Lee, Hong Joon; Jeon, Ji Hyun; Kim, In-Beom; Choi, Jin-Sung; Sung, Ki-Wug; Hahn, Sang June

    2015-02-09

    Donepezil is a potent, selective inhibitor of acetylcholinesterase, which is used for the treatment of Alzheimer's disease. Whole-cell patch-clamp technique and Western blot analyses were used to study the effects of donepezil on the human ether-a-go-go-related gene (hERG) channel. Donepezil inhibited the tail current of the hERG in a concentration-dependent manner with an IC50 of 1.3 μM. The metabolites of donepezil, 6-ODD and 5-ODD, inhibited the hERG currents in a similar concentration-dependent manner; the IC50 values were 1.0 and 1.5 μM, respectively. A fast drug perfusion system demonstrated that donepezil interacted with both the open and inactivated states of the hERG. A fast application of donepezil during the tail currents inhibited the open state of the hERG in a concentration-dependent manner with an IC50 of 2.7 μM. Kinetic analysis of donepezil in an open state of the hERG yielded blocking and unblocking rate constants of 0.54 µM(-1)s(-1) and 1.82 s(-1), respectively. The block of the hERG by donepezil was voltage-dependent with a steep increase across the voltage range of channel activation. Donepezil caused a reduction in the hERG channel protein trafficking to the plasma membrane at low concentration, but decreased the channel protein expression at higher concentrations. These results suggest that donepezil inhibited the hERG at a supratherapeutic concentration, and that it did so by preferentially binding to the activated (open and/or inactivated) states of the channels and by inhibiting the trafficking and expression of the hERG channel protein in the plasma membrane. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Remote and reversible inhibition of neurons and circuits by small molecule induced potassium channel stabilization.

    Science.gov (United States)

    Auffenberg, Eva; Jurik, Angela; Mattusch, Corinna; Stoffel, Rainer; Genewsky, Andreas; Namendorf, Christian; Schmid, Roland M; Rammes, Gerhard; Biel, Martin; Uhr, Manfred; Moosmang, Sven; Michalakis, Stylianos; Wotjak, Carsten T; Thoeringer, Christoph K

    2016-01-13

    Manipulating the function of neurons and circuits that translate electrical and chemical signals into behavior represents a major challenges in neuroscience. In addition to optogenetic methods using light-activatable channels, pharmacogenetic methods with ligand induced modulation of cell signaling and excitability have been developed. However, they are largely based on ectopic expression of exogenous or chimera proteins. Now, we describe the remote and reversible expression of a Kir2.1 type potassium channel using the chemogenetic technique of small molecule induced protein stabilization. Based on shield1-mediated shedding of a destabilizing domain fused to a protein of interest and inhibition of protein degradation, this principle has been adopted for biomedicine, but not in neuroscience so far. Here, we apply this chemogenetic approach in brain research for the first time in order to control a potassium channel in a remote and reversible manner. We could show that shield1-mediated ectopic Kir2.1 stabilization induces neuronal silencing in vitro and in vivo in the mouse brain. We also validated this novel pharmacogenetic method in different neurobehavioral paradigms.The DD-Kir2.1 may complement the existing portfolio of pharmaco- and optogenetic techniques for specific neuron manipulation, but it may also provide an example for future applications of this principle in neuroscience research.

  1. The voltage-gated potassium channel subunit, Kv1.3, is expressed in epithelia

    DEFF Research Database (Denmark)

    Grunnet, Morten; Rasmussen, Hanne B; Hay-Schmidt, Anders

    2003-01-01

    The Shaker-type voltage-gated potassium channel, Kv1.3, is believed to be restricted in distribution to lymphocytes and neurons. In lymphocytes, this channel has gained intense attention since it has been proven that inhibition of Kv1.3 channels compromise T lymphocyte activation. To investigate...

  2. Deubiquitylating enzyme USP2 counteracts Nedd4-2-mediated downregulation of KCNQ1 potassium channels

    DEFF Research Database (Denmark)

    Krzystanek, Katarzyna; Rasmussen, Hanne Borger; Grunnet, Morten

    2012-01-01

    with the KCNQ1 potassium channel through a PY motif located in the C terminus of KCNQ1. This interaction induces ubiquitylation of KCNQ1, resulting in a reduced surface density of the channel. It was reported recently that the epithelial sodium channel is regulated by the reverse process...

  3. Coupling of activation and inactivation gate in a K+-channel: potassium and ligand sensitivity

    NARCIS (Netherlands)

    Ader, C.; Schneider, R.; Hornig, S.; Velisetty, P.; Vardanyan, V.; Giller, K.; Ohmert, I.; Becker, S.; Pongs, O.; Baldus, M.

    2009-01-01

    Potassium (K+)-channel gating is choreographed by a complex interplay between external stimuli, K+ concentration and lipidic environment. We combined solid-state NMR and electrophysiological experiments on a chimeric KcsA–Kv1.3 channel to delineate K+, pH and blocker effects on channel structure and

  4. The Sodium-Activated Potassium Channel Slack Is Required for Optimal Cognitive Flexibility in Mice

    Science.gov (United States)

    Bausch, Anne E.; Dieter, Rebekka; Nann, Yvette; Hausmann, Mario; Meyerdierks, Nora; Kaczmarek, Leonard K.; Ruth, Peter; Lukowski, Robert

    2015-01-01

    "Kcnt1" encoded sodium-activated potassium channels (Slack channels) are highly expressed throughout the brain where they modulate the firing patterns and general excitability of many types of neurons. Increasing evidence suggests that Slack channels may be important for higher brain functions such as cognition and normal intellectual…

  5. Increase of ATP-sensitive potassium (KATP channels in the heart of type-1 diabetic rats

    Directory of Open Access Journals (Sweden)

    Chen Zhih-Cherng

    2012-01-01

    Full Text Available Abstract Background An impairment of cardiovascular function in streptozotocin (STZ-diabetic rats has been mentioned within 5 days-to-3 months of induction. ATP-sensitive potassium (KATP channels are expressed on cardiac sarcolemmal membranes. It is highly responsive to metabolic fluctuations and can have effects on cardiac contractility. The present study attempted to clarify the changes of cardiac KATP channels in diabetic disorders. Methods Streptozotocin-induced diabetic rats and neonatal rat cardiomyocytes treated with a high concentration of glucose (a D-glucose concentration of 30 mM was used and cells were cultured for 24 hr were used to examine the effect of hyperglycemia on cardiac function and the expression of KATP channels. KATP channels expression was found to be linked to cardiac tonic dysfunction, and we evaluated the expression levels of KATP channels by Western blot and Northern blot analysis. Results The result shows diazoxide produced a marked reduction of heart rate in control group. Furthermore, the methods of Northern blotting and Western blotting were employed to identify the gene expression of KATP channel. Two subunits of cardiac KATP channel (SUR2A and kir 6.2 were purchased as indicators and showed significantly decreased in both diabetic rats and high glucose treated rat cardiac myocytes. Correction of hyperglycemia by insulin or phlorizin restored the gene expression of cardiac KATP in these diabetic rats. Conclusions Both mRNA and protein expression of cardiac KATP channels are decreased in diabetic rats induced by STZ for 8 weeks. This phenomenon leads to result in desensitization of some KATP channel drugs.

  6. Free energy dissipation of the spontaneous gating of a single voltage-gated potassium channel

    Science.gov (United States)

    Wang, Jia-Zeng; Wang, Rui-Zhen

    2018-02-01

    Potassium channels mainly contribute to the resting potential and re-polarizations, with the potassium electrochemical gradient being maintained by the pump Na+/K+-ATPase. In this paper, we construct a stochastic model mimicking the kinetics of a potassium channel, which integrates temporal evolving of the membrane voltage and the spontaneous gating of the channel. Its stationary probability density functions (PDFs) are found to be singular at the boundaries, which result from the fact that the evolving rates of voltage are greater than the gating rates of the channel. We apply PDFs to calculate the power dissipations of the potassium current, the leakage, and the gating currents. On a physical perspective, the essential role of the system is the K+-battery charging the leakage (L-)battery. A part of power will inevitably be dissipated among the process. So, the efficiency of energy transference is calculated.

  7. Structural analysis of the S4-S5 linker of the human KCNQ1 potassium channel.

    Science.gov (United States)

    Gayen, Shovanlal; Li, Qingxin; Kang, CongBao

    2015-01-02

    KCNQ1 plays important roles in the cardiac action potential and consists of an N-terminal domain, a voltage-sensor domain, a pore domain and a C-terminal domain. KCNQ1 is a voltage-gated potassium channel and its channel activity is regulated by membrane potentials. The linker between transmembrane helices 4 and 5 (S4-S5 linker) is important for transferring the conformational changes from the voltage-sensor domain to the pore domain. In this study, the structure of the S4-S5 linker of KCNQ1 was investigated by solution NMR, circular dichroism and fluorescence spectroscopic studies. The S4-S5 linker adopted a helical structure in detergent micelles. The W248 may interact with the cell membrane. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Psychiatric presentation of voltage-gated potassium channel antibody-associated encephalopathy

    OpenAIRE

    PARTHASARATHI, U. D.; HARROWER, T.; TEMPEST, M.; HODGES, J. R.; WALSH, C.; McKENNA, P. J.; FLETCHER, P.C.

    2006-01-01

    Voltage-gated potassium channel antibody encephalopathy, a rare cause of limbic encephalopathy, typically presents with memory impairment and seizures. Psychiatric symptoms have not been emphasised in the literature. Here we describe a 58-year-old man who presented with panic attacks and psychogenic non-epileptic seizures and, later on, developed delusions and hallucinations and then confusion.He was found to have antibodies to voltage-gated potassium channels.Treatment with immuno-modulatory...

  9. Developmental expression of Kv1 voltage-gated potassium channels in the avian hypothalamus.

    Science.gov (United States)

    Doczi, Megan A; Vitzthum, Carl M; Forehand, Cynthia J

    2016-03-11

    Specialized hypothalamic neurons integrate the homeostatic balance between food intake and energy expenditure, processes that may become dysregulated during the development of diabetes, obesity, and other metabolic disorders. Shaker family voltage-gated potassium channels (Kv1) contribute to the maintenance of resting membrane potential, action potential characteristics, and neurotransmitter release in many populations of neurons, although hypothalamic Kv1 channel expression has been largely unexplored. Whole-cell patch clamp recordings from avian hypothalamic brain slices demonstrate a developmental shift in the electrophysiological properties of avian arcuate nucleus neurons, identifying an increase in outward ionic current that corresponds with action potential maturation. Additionally, RT-PCR experiments identified the early expression of Kv1.2, Kv1.3, and Kv1.5 mRNA in the embryonic avian hypothalamus, suggesting that these channels may underlie the electrophysiological changes observed in these neurons. Real-time quantitative PCR analysis on intact microdissections of embryonic hypothalamic tissue revealed a concomitant increase in Kv1.2 and Kv1.5 gene expression at key electrophysiological time points during development. This study is the first to demonstrate hypothalamic mRNA expression of Kv1 channels in developing avian embryos and may suggest a role for voltage-gated ion channel regulation in the physiological patterning of embryonic hypothalamic circuits governing energy homeostasis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  10. Effect of tyrphostin AG879 on Kv 4.2 and Kv 4.3 potassium channels.

    Science.gov (United States)

    Yu, Haibo; Zou, Beiyan; Wang, Xiaoliang; Li, Min

    2015-07-01

    A-type potassium channels (IA) are important proteins for modulating neuronal membrane excitability. The expression and activity of Kv 4.2 channels are critical for neurological functions and pharmacological inhibitors of Kv 4.2 channels may have therapeutic potential for Fragile X syndrome. While screening various compounds, we identified tyrphostin AG879, a tyrosine kinase inhibitor, as a Kv 4.2 inhibitor from. In the present study we characterized the effect of AG879 on cloned Kv 4.2/Kv channel-interacting protein 2 (KChIP2) channels. To screen the library of pharmacologically active compounds, the thallium flux assay was performed on HEK-293 cells transiently-transfected with Kv 4.2 cDNA using the Maxcyte transfection system. The effects of AG879 were further examined on CHO-K1 cells expressing Kv 4.2/KChIP2 channels using a whole-cell patch-clamp technique. Tyrphostin AG879 selectively and dose-dependently inhibited Kv 4.2 and Kv 4.3 channels. In Kv 4.2/KChIP2 channels, AG879 induced prominent acceleration of the inactivation rate, use-dependent block and slowed the recovery from inactivation. AG879 induced a hyperpolarizing shift in the voltage-dependence of the steady-state inactivation of Kv 4.2 channels without apparent effect on the V1/2 of the voltage-dependent activation. The blocking effect of AG879 was enhanced as channel inactivation increased. Furthermore, AG879 significantly inhibited the A-type potassium currents in the cultured hippocampus neurons. AG879 was identified as a selective and potent inhibitor the Kv 4.2 channel. AG879 inhibited Kv 4.2 channels by preferentially interacting with the open state and further accelerating their inactivation. © 2015 The British Pharmacological Society.

  11. Inhibition of cloned hERG potassium channels by risperidone and paliperidone.

    Science.gov (United States)

    Lee, Hong Joon; Choi, Jin-Sung; Choi, Bok Hee; Hahn, Sang June

    2017-06-01

    Risperidone and one of its active metabolites, paliperidone, are widely used for the treatment of schizophrenia. We used a patch-clamp study to investigate the effects of paliperidone on hERG potassium channels expressed in HEK cells. Western blot analyses were used to study the effects of risperidone and paliperidone on hERG and hERG 3.1 isoform channel trafficking. Risperidone and paliperidone inhibited the hERG tail currents in a concentration-dependent manner with IC 50 values of 0.16 and 0.57 μM, respectively. The block of hERG currents by paliperidone was voltage-dependent, increasing over a range of voltages for channel activation. A fast application of paliperidone inhibited the hERG current elicited by a 5-s depolarizing pulse to +60 mV to fully inactivate the hERG currents, suggesting an inactivated state block. A fast application of paliperidone during repolarization reversibly inhibited the hERG tail currents in a concentration-dependent manner with a IC 50 value of 1.26 μM. Kinetic analysis of paliperidone interaction with the open state of the hERG channels showed that the rate constants of association (k +1 ) and dissociation (k -1 ) for paliperidone were 0.45 μM -1  s -1 and 1.07 s -1 , respectively. Paliperidone shifted the steady-state inactivation curve of the hERG currents in a hyperpolarizing direction and also produced a use-dependent block. Risperidone and paliperidone had no effect on hERG and hERG 3.1 channel trafficking to the cell membrane. Our results indicated that paliperidone inhibited the hERG current by preferentially interacting with the open and inactivated states of the channel, but not by disruption of hERG channel protein trafficking.

  12. Terbinafine is a novel and selective activator of the two-pore domain potassium channel TASK3.

    Science.gov (United States)

    Wright, Paul D; Veale, Emma L; McCoull, David; Tickle, David C; Large, Jonathan M; Ococks, Emma; Gothard, Gemma; Kettleborough, Catherine; Mathie, Alistair; Jerman, Jeffrey

    2017-11-04

    Two-pore domain potassium channels (K2Ps) are characterized by their four transmembrane domain and two-pore topology. They carry background (or leak) potassium current in a variety of cell types. Despite a number of important roles there is currently a lack of pharmacological tools with which to further probe K2P function. We have developed a cell-based thallium flux assay, using baculovirus delivered TASK3 (TWIK-related acid-sensitive K + channel 3, KCNK9, K2P9.1) with the aim of identifying novel, selective TASK3 activators. After screening a library of 1000 compounds, including drug-like and FDA approved molecules, we identified Terbinafine as an activator of TASK3. In a thallium flux assay a pEC50 of 6.2 ( ±0.12) was observed. When Terbinafine was screened against TASK2, TREK2, THIK1, TWIK1 and TRESK no activation was observed in thallium flux assays. Several analogues of Terbinafine were also purchased and structure activity relationships examined. To confirm Terbinafine's activation of TASK3 whole cell patch clamp electrophysiology was carried out and clear potentiation observed in both the wild type channel and the pathophysiological, Birk-Barel syndrome associated, G236R TASK3 mutant. No activity at TASK1 was observed in electrophysiology studies. In conclusion, we have identified the first selective activator of the two-pore domain potassium channel TASK3. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. In Vitro Contractile Response of Rabbit Myometrium to BKCa and KATP Potassium Channel Openers

    Directory of Open Access Journals (Sweden)

    Soňa Fraňová

    2009-01-01

    Full Text Available The aim of the study was to evaluate the participation of ligand-sensitive potassium large conductance calcium-activated channels (BKCa and ATP-sensitive potassium channels in uterine smooth muscle reactivity during different stages of the experimentally induced proliferatory and secretory phase in the sexual cycle in ovariectomised rabbits in vitro. The myometrial reactivity to oxytocin (10-6 mol l-1 was investigated by an in vitro method in female rabbits 14 days after ovariectomy treated with 17β-estradiol - 1 mg/kg/day i.m. for 7 days, or with a combination of progesterone 2 mg/kg/day s.c. for 7 days and 17β-estradiol - 0.2 mg/ kg/day (day 3–7. The strips of myometrial smooth muscle were incubated with a specific opener (NS 1619 and an antagonist (TEA of potassium large conductance calcium-activated channel, or with a specific opener (pinacidil and an antagonist (glybenclamide of ATP-sensitive potassium channels before the administration of oxytocin. NS1619 produced more potent inhibition of the oxytocin-induced contraction during the gestagen dominance (experimental secretory phase than the one observed during the oestrogen dominance (experimental proliferatory phase. TEA antagonized the NS1619 induced inhibition of the myometrial contraction. In the matter of KATP potassium channels, after the administration of pinacidil we observed a similar situation in the changes of myometrial contractility. Pinacidil produced more pronounced inhibition of oxytocin-induced contraction during the secretory phase, and its effect was abolished by the selective inhibitor glybenclamide. Our experimental results indicate that both potassium large conductance calcium-activated channels and ATP-sensitive potassium channels significantly participate in the regulation of myometrial oxytocin-induced contractions and the activity of these channels is probably influenced by the levels of oestrogens and gestagens.

  14. Cell-cell channels

    National Research Council Canada - National Science Library

    Baluška, F; Volkmann, Dieter; Barlow, P. W

    2006-01-01

    ...-Negative Bacteria Pheromone-Responsive Conjugative Plasmids in E. faecalis Nonpheromone-Responsive Plasmids in G+ Bacteria 21 22 27 28 Section II: Ciliate Cells 3. The Tetrahymena Conjugation Ju...

  15. Chronic Alcohol, Intrinsic Excitability, and Potassium Channels: Neuroadaptations and Drinking Behavior.

    Science.gov (United States)

    Cannady, Reginald; Rinker, Jennifer A; Nimitvilai, Sudarat; Woodward, John J; Mulholland, Patrick J

    2018-01-28

    Neural mechanisms underlying alcohol use disorder remain elusive, and this lack of understanding has slowed the development of efficacious treatment strategies for reducing relapse rates and prolonging abstinence. While synaptic adaptations produced by chronic alcohol exposure have been extensively characterized in a variety of brain regions, changes in intrinsic excitability of critical projection neurons are understudied. Accumulating evidence suggests that prolonged alcohol drinking and alcohol dependence produce plasticity of intrinsic excitability as measured by changes in evoked action potential firing and after-hyperpolarization amplitude. In this chapter, we describe functional changes in cell firing of projection neurons after long-term alcohol exposure that occur across species and in multiple brain regions. Adaptations in calcium-activated (K Ca 2), voltage-dependent (K V 7), and G protein-coupled inwardly rectifying (K ir 3 or GIRK) potassium channels that regulate the evoked firing and after-hyperpolarization parallel functional changes in intrinsic excitability induced by chronic alcohol. Moreover, there are strong genetic links between alcohol-related behaviors and genes encoding K Ca 2, K V 7, and GIRK channels, and pharmacologically targeting these channels reduces alcohol consumption and alcohol-related behaviors. Together, these studies demonstrate that chronic alcohol drinking produces adaptations in K Ca 2, K V 7, and GIRK channels leading to impaired regulation of the after-hyperpolarization and aberrant cell firing. Correcting the deficit in the after-hyperpolarization with positive modulators of K Ca 2 and K V 7 channels and altering the GIRK channel binding pocket to block the access of alcohol represent a potentially highly effective pharmacological approach that can restore changes in intrinsic excitability and reduce alcohol consumption in affected individuals.

  16. Activation of ERG2 potassium channels by the diphenylurea NS1643

    DEFF Research Database (Denmark)

    Elmedyb, Pernille; Olesen, Søren-Peter; Grunnet, Morten

    2007-01-01

    Three members of the ERG potassium channel family have been described (ERG1-3 or Kv 11.1-3). ERG1 is by far the best characterized subtype and it constitutes the molecular component of the cardiac I(Kr) current. All three channel subtypes are expressed in neurons but their function remains unclear...

  17. Ionic channels in plants: potassium transport Canais iônicos em plantas: o transporte de potássio

    Directory of Open Access Journals (Sweden)

    Antonio Costa de Oliveira

    1995-01-01

    Full Text Available The discovery of potassium channels on the plasma membrane has helped to elucidate important mechanisms in animal and plant physiology. Plant growth and development associated mechanisms, such as germination, leaf movements, stomatal action, ion uptake in roots, phloem transport and nutrient storage are linked to potassium transport. Studies describing potassium transport regulation by abscisic acid (ABA, Ca++, light and other factors are presented here. Also the types of channels that regulate potassium uptake and efflux in the cell, and the interaction of these channels with external signals, are discussed.A descoberta de canais iônicos presentes na membrana plasmática tem ajudado a elucidar importantes mecanismos fisiológicos em animais e plantas. Mecanismos associados ao crescimento e desenvolvimento das plantas, tais como germinação, movimento foliar, abertura e fechamento de estômatos, absorção de íons pelas raízes e armazenamento de nutrientes estão ligados ao transporte de potássio. Estudos descrevendo a regulação do transporte deste nutriente por ácido abscísico (ABA, Ca++, luz e outros fatores são apresentados. Os tipos de canais que regulam a saída e entrada de potássio na célula, e as interações destes com os sinais externos, são discutidos.

  18. Effects of the hypoglycaemic drugs repaglinide and glibenclamide on ATP-sensitive potassium-channels and cytosolic calcium levels in beta TC3 cells and rat pancreatic beta cells

    DEFF Research Database (Denmark)

    Gromada, J; Dissing, S; Kofod, Hans

    1995-01-01

    increase in intracellular free Ca2+ concentration ([Ca2+]i) with a half-maximal effect at 0.5 nmol/l for both drugs in long-term experiments (30 min). The rise in [Ca2+]i results from Ca2+ entry through voltage-dependent L-type Ca(2+)-channels since it is inhibited by verapamil (10 mumol/l). The effect......The present study demonstrates the action of the hypoglycaemic drugs repaglinide and glibenclamide in cultured newborn rat islet cells and mouse beta TC3 cells. In cell-attached membrane patches of newborn rat islet cells repaglinide (10 nmol/l) and glibenclamide (20 nmol/l) decrease the open...... probability of single ATP-sensitive K(+)-channels to approximately 10% of the activity prior to addition of the drugs in short-term experiments (

  19. Toxic effects of environmental rare earth elements on delayed outward potassium channels and their mechanisms from a microscopic perspective.

    Science.gov (United States)

    Wang, Lihong; He, Jingfang; Xia, Ao; Cheng, Mengzhu; Yang, Qing; Du, Chunlei; Wei, Haiyan; Huang, Xiaohua; Zhou, Qing

    2017-08-01

    The wide applications cause a large amount of rare earth elements (REEs) to be released into the environment, and ultimately into the human body through food chain. Toxic effects of REEs on humans have been extensively studied, but their toxic effects and binding targets in cells are not understood. Delayed outward potassium channels (K + channels) are good targets for exogenous substances or clinical drugs. To evaluate cellular toxicities of REEs and clarify toxic mechanisms, the toxicities of REEs on the K + channel and their structural basis were investigated. The results showed that delayed outward potassium channels on the plasma membrane are the targets of REEs acting on living organisms, and the changes in the thermodynamic and kinetic characteristics of the K + channel are the reasons of diseases induced by REEs. Two types of REEs, a light REE La 3+ and a heavy REE Tb 3+ , displayed different intensity of toxicities on the K + channel, in which the toxicity of Tb 3+ was stronger than that of La 3+ . More interestingly, in comparison with that of heavy metal Cd 2+ , the cytotoxicities of the light and heavy REEs showed discriminative differences, and the cytotoxicity of Tb 3+ was higher than that of Cd 2+ , while the cytotoxicity of La 3+ was lower than that of Cd 2+ . These different cytotoxicities of La 3+ , Tb 3+ and Cd 2+ on human resulted from the varying binding abilities of the metals to this channel protein. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Targeting the Small- and Intermediate-Conductance Ca2+-Activated Potassium Channels: The Drug-Binding Pocket at the Channel/Calmodulin Interface

    Directory of Open Access Journals (Sweden)

    Meng Cui

    2014-10-01

    Full Text Available The small- and intermediate-conductance Ca2+-activated potassium (SK/IK channels play important roles in the regulation of excitable cells in both the central nervous and cardiovascular systems. Evidence from animal models has implicated SK/IK channels in neurological conditions such as ataxia and alcohol use disorders. Further, genome-wide association studies have suggested that cardiovascular abnormalities such as arrhythmias and hypertension are associated with single nucleotide polymorphisms that occur within the genes encoding the SK/IK channels. The Ca2+ sensitivity of the SK/IK channels stems from a constitutively bound Ca2+-binding protein: calmodulin. Small-molecule positive modulators of SK/IK channels have been developed over the past decade, and recent structural studies have revealed that the binding pocket of these positive modulators is located at the interface between the channel and calmodulin. SK/IK channel positive modulators can potentiate channel activity by enhancing the coupling between Ca2+ sensing via calmodulin and mechanical opening of the channel. Here, we review binding pocket studies that have provided structural insight into the mechanism of action for SK/IK channel positive modulators. These studies lay the foundation for structure-based drug discovery efforts that can identify novel SK/IK channel positive modulators. © 2014 S. Karger AG, Basel

  1. Location of the β4 transmembrane helices in the BK potassium channel

    Science.gov (United States)

    Wu, Roland S.; Chudasama, Neelesh; Zakharov, Sergey I.; Doshi, Darshan; Motoike, Howard; Liu, Guoxia; Yao, Yongneng; Niu, Xiaowei; Deng, Shi-Xian; Landry, Donald W.; Karlin, Arthur; Marx, Steven O.

    2009-01-01

    Large-conductance, voltage- and Ca2+-gated potassium (BK) channels control excitability in a number of cell types. BK channels are composed of α subunits, which contain the voltage-sensor domains and the Ca2+- sensor domains, and form the pore, and often one of four types of β subunits, which modulate the channel in a cell-specific manner. β4 is expressed in neurons throughout the brain. Deletion of β4 in mice causes temporal lobe epilepsy. Compared to channels composed of α alone, channels composed of α and β4 activate and deactivate more slowly. We inferred the locations of the two β4 transmembrane (TM) helices, TM1 and TM2, relative to the seven αTM helices, S0-S6, from the extent of disulfide bond formation between cysteines substituted in the extracellular flanks of these TM helices. We found that β4 TM2 is close to α S0 and that β4 TM1 is close to both α S1 and S2. At least at their extracellular ends, TM1 and TM2 are not close to S3 through S6. In six of eight of the most highly crosslinked cysteine pairs, four crosslinks from TM2 to S0 and one each from TM1 to S1 and S2 had small effects on the V50 and on the rates of activation and deactivation. That disulfide crosslinking caused only small functional perturbations is consistent with the proximity of the extracellular ends of TM2 to S0 and of TM1 to S1 and to S2, in both the open and closed states. PMID:19571123

  2. Scorpion Potassium Channel-blocking Defensin Highlights a Functional Link with Neurotoxin.

    Science.gov (United States)

    Meng, Lanxia; Xie, Zili; Zhang, Qian; Li, Yang; Yang, Fan; Chen, Zongyun; Li, Wenxin; Cao, Zhijian; Wu, Yingliang

    2016-03-25

    The structural similarity between defensins and scorpion neurotoxins suggests that they might have evolved from a common ancestor. However, there is no direct experimental evidence demonstrating a functional link between scorpion neurotoxins and defensins. The scorpion defensin BmKDfsin4 from Mesobuthus martensiiKarsch contains 37 amino acid residues and a conserved cystine-stabilized α/β structural fold. The recombinant BmKDfsin4, a classical defensin, has been found to have inhibitory activity against Gram-positive bacteria such as Staphylococcus aureus, Bacillus subtilis, and Micrococcus luteusas well as methicillin-resistant Staphylococcus aureus Interestingly, electrophysiological experiments showed that BmKDfsin4,like scorpion potassium channel neurotoxins, could effectively inhibit Kv1.1, Kv1.2, and Kv1.3 channel currents, and its IC50value for the Kv1.3 channel was 510.2 nm Similar to the structure-function relationships of classical scorpion potassium channel-blocking toxins, basic residues (Lys-13 and Arg-19) of BmKDfsin4 play critical roles in peptide-Kv1.3 channel interactions. Furthermore, mutagenesis and electrophysiological experiments demonstrated that the channel extracellular pore region is the binding site of BmKDfsin4, indicating that BmKDfsin4 adopts the same mechanism for blocking potassium channel currents as classical scorpion toxins. Taken together, our work identifies scorpion BmKDfsin4 as the first invertebrate defensin to block potassium channels. These findings not only demonstrate that defensins from invertebrate animals are a novel type of potassium channel blockers but also provide evidence of a functional link between defensins and neurotoxins. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Diclofenac Distinguishes among Homomeric and Heteromeric Potassium Channels Composed of KCNQ4 and KCNQ5 SubunitsS⃞

    Science.gov (United States)

    Brueggemann, Lioubov I.; Mackie, Alexander R.; Martin, Jody L.; Cribbs, Leanne L.

    2011-01-01

    KCNQ4 and KCNQ5 potassium channel subunits are expressed in vascular smooth muscle cells, although it remains uncertain how these subunits assemble to form functional channels. Using patch-clamp techniques, we compared the electrophysiological characteristics and effects of diclofenac, a known KCNQ channel activator, on human KCNQ4 and KCNQ5 channels expressed individually or together in A7r5 rat aortic smooth muscle cells. The conductance curves of the overexpressed channels were fitted by a single Boltzmann function in each case (V0.5 values: −31, −44, and −38 mV for KCNQ4, KCNQ5, and KCNQ4/5, respectively). Diclofenac (100 μM) inhibited KCNQ5 channels, reducing maximum conductance by 53%, but increased maximum conductance of KCNQ4 channels by 38%. The opposite effects of diclofenac on KCNQ4 and KCNQ5 could not be attributed to the presence of a basic residue (lysine) in the voltage-sensing domain of KCNQ5, because mutation of this residue to neutral glycine (the residue present in KCNQ4) resulted in a more effective block of the channel. Differences in deactivation rates and distinct voltage-dependent effects of diclofenac on channel activation and deactivation observed with each of the subunit combinations (KCNQ4, KCNQ5, and KCNQ4/5) were used as diagnostic tools to evaluate native KCNQ currents in vascular smooth muscle cells. A7r5 cells express only KCNQ5 channels endogenously, and their responses to diclofenac closely resembled those of the overexpressed KCNQ5 currents. In contrast, mesenteric artery myocytes, which express both KCNQ4 and KCNQ5 channels, displayed whole-cell KCNQ currents with properties and diclofenac responses characteristic of overexpressed heteromeric KCNQ4/5 channels. PMID:20876743

  4. The role of ATP-sensitive potassium channels in cellular function and protection in the cardiovascular system.

    Science.gov (United States)

    Tinker, Andrew; Aziz, Qadeer; Thomas, Alison

    2014-01-01

    ATP-sensitive potassium channels (K(ATP)) are widely distributed and present in a number of tissues including muscle, pancreatic beta cells and the brain. Their activity is regulated by adenine nucleotides, characteristically being activated by falling ATP and rising ADP levels. Thus, they link cellular metabolism with membrane excitability. Recent studies using genetically modified mice and genomic studies in patients have implicated K(ATP) channels in a number of physiological and pathological processes. In this review, we focus on their role in cellular function and protection particularly in the cardiovascular system. © 2013 The British Pharmacological Society.

  5. Effect of methamphetamine on the microglial damage: role of potassium channel Kv1.3.

    Directory of Open Access Journals (Sweden)

    Jun Wang

    Full Text Available Methamphetamine (Meth abusing represents a major public health problem worldwide. Meth has long been known to induce neurotoxicity. However, the mechanism is still remained poorly understood. Growing evidences indicated that the voltage-gated potassium channels (Kv were participated in neuronal damage and microglia function. With the whole cell patch clamp, we found that Meth significantly increased the outward K⁺ currents, therefore, we explored whether Kv1.3, one of the major K⁺ channels expressed in microglia, was involved in Meth-induced microglia damage. Our study showed that Meth significantly increased the cell viability in a dose dependent manner, while the Kv blocker, tetraethylamine (TEA, 4-Aminopyridine (4-AP and Kv1.3 specific antagonist margatoxin (MgTx, prevented against the damage mediated by Meth. Interestingly, treatment of cells with Meth resulted in increasing expression of Kv1.3 rather than Kv1.5, at both mRNA and protein level, which is partially blocked by MgTx. Furthermore, Meth also stimulated a significant increased expression of IL-6 and TNF-α at protein level, which was significantly inhibited by MgTx. Taken together, these results demonstrated that Kv1.3 was involved in Meth-mediated microglial damage, providing the potential target for the development of therapeutic strategies for Meth abuse.

  6. Distribution, expression and functional effects of small conductance Ca-activated potassium (SK) channels in rat myometrium.

    Science.gov (United States)

    Noble, Karen; Floyd, Rachel; Shmygol, Andre; Shmygol, Anatoly; Mobasheri, A; Wray, Susan

    2010-01-01

    Calcium-activated potassium channels are important in a variety of smooth muscles, contributing to excitability and contractility. In the myometrium previous work has focussed on the large conductance channels (BK), and the role of small conductance channels (SK) has received scant attention, despite the finding that over-expression of an SK channel isoform (SK3) results in uterine dysfunction and delayed parturition. This study therefore characterises the expression of the three SK channel isoforms (SK1-3) in rat myometrium throughout pregnancy and investigates their effect on cytosolic [Ca] and force and compares this with that of BK channels. Consistent expression of all SK isoform transcripts and clear immunostaining of SK1-3 was found. Inhibition of SK1-3 channels (apamin, scyllatoxin) significantly inhibited outward current, caused membrane depolarisation and elicited action potentials in previously quiescent cells. Apamin or scyllatoxin increased the amplitude of [Ca] and force in spontaneously contracting myometrial strips throughout gestation. The functional effect of SK inhibition was larger than that of BK channel inhibition. Thus we show for the first time that SK1-3 channels are expressed and translated throughout pregnancy and contribute to outward current, regulate membrane potential and hence Ca signals in pregnant rat myometrium. They contribute more to quiescence that BK channels. 2009 Elsevier Ltd. All rights reserved.

  7. Potassium

    Science.gov (United States)

    ... and blackberries Root vegetables, such as carrots and potatoes Citrus fruits, such as oranges and grapefruit Your kidneys help to keep the right amount of potassium in your body. If you have chronic kidney disease, your kidneys may not remove extra potassium from ...

  8. Potassium Channel Subfamily K Member 3 (KCNK3) Contributes to the Development of Pulmonary Arterial Hypertension.

    Science.gov (United States)

    Antigny, Fabrice; Hautefort, Aurélie; Meloche, Jolyane; Belacel-Ouari, Milia; Manoury, Boris; Rucker-Martin, Catherine; Péchoux, Christine; Potus, François; Nadeau, Valérie; Tremblay, Eve; Ruffenach, Grégoire; Bourgeois, Alice; Dorfmüller, Peter; Breuils-Bonnet, Sandra; Fadel, Elie; Ranchoux, Benoît; Jourdon, Philippe; Girerd, Barbara; Montani, David; Provencher, Steeve; Bonnet, Sébastien; Simonneau, Gérald; Humbert, Marc; Perros, Frédéric

    2016-04-05

    Mutations in the KCNK3 gene have been identified in some patients suffering from heritable pulmonary arterial hypertension (PAH). KCNK3 encodes an outward rectifier K(+) channel, and each identified mutation leads to a loss of function. However, the pathophysiological role of potassium channel subfamily K member 3 (KCNK3) in PAH is unclear. We hypothesized that loss of function of KCNK3 is a hallmark of idiopathic and heritable PAH and contributes to dysfunction of pulmonary artery smooth muscle cells and pulmonary artery endothelial cells, leading to pulmonary artery remodeling: consequently, restoring KCNK3 function could alleviate experimental pulmonary hypertension (PH). We demonstrated that KCNK3 expression and function were reduced in human PAH and in monocrotaline-induced PH in rats. Using a patch-clamp technique in freshly isolated (not cultured) pulmonary artery smooth muscle cells and pulmonary artery endothelial cells, we found that KCNK3 current decreased progressively during the development of monocrotaline-induced PH and correlated with plasma-membrane depolarization. We demonstrated that KCNK3 modulated pulmonary arterial tone. Long-term inhibition of KCNK3 in rats induced distal neomuscularization and early hemodynamic signs of PH, which were related to exaggerated proliferation of pulmonary artery endothelial cells, pulmonary artery smooth muscle cell, adventitial fibroblasts, and pulmonary and systemic inflammation. Lastly, in vivo pharmacological activation of KCNK3 significantly reversed monocrotaline-induced PH in rats. In PAH and experimental PH, KCNK3 expression and activity are strongly reduced in pulmonary artery smooth muscle cells and endothelial cells. KCNK3 inhibition promoted increased proliferation, vasoconstriction, and inflammation. In vivo pharmacological activation of KCNK3 alleviated monocrotaline-induced PH, thus demonstrating that loss of KCNK3 is a key event in PAH pathogenesis and thus could be therapeutically targeted.

  9. Inwardly rectifying potassium channels (Kir) in central nervous system glia: a special role for Kir4.1 in glial functions

    OpenAIRE

    Butt, Arthur M; Kalsi, Amanpreet

    2007-01-01

    Glia in the central nervous system (CNS) express diverse inward rectifying potassium channels (Kir). The major function of Kir is in establishing the high potassium (K+) selectivity of the glial cell membrane and strongly negative resting membrane potential (RMP), which are characteristic physiological properties of glia. The classical property of Kir is that K+ flows inwards when the RMP is negative to the equilibrium potential for K+ (Ek), but at more positive potentials outward currents ar...

  10. Inward-rectifying potassium (Kir) channels regulate pacemaker activity in spinal nociceptive circuits during early life

    Science.gov (United States)

    Li, Jie; Blankenship, Meredith L.; Baccei, Mark L.

    2013-01-01

    Pacemaker neurons in neonatal spinal nociceptive circuits generate intrinsic burst-firing and are distinguished by a lower “leak” membrane conductance compared to adjacent, non-bursting neurons. However, little is known about which subtypes of leak channels regulate the level of pacemaker activity within the developing rat superficial dorsal horn (SDH). Here we demonstrate that a hallmark feature of lamina I pacemaker neurons is a reduced conductance through inward-rectifying potassium (Kir) channels at physiological membrane potentials. Differences in the strength of inward rectification between pacemakers and non-pacemakers indicate the presence of functionally distinct Kir currents in these two populations at room temperature. However, Kir currents in both groups showed high sensitivity to block by extracellular Ba2+ (IC50 ~ 10 µM), which suggests the presence of ‘classical’ Kir (Kir2.x) channels in the neonatal SDH. The reduced Kir conductance within pacemakers is unlikely to be explained by an absence of particular Kir2.x isoforms, as immunohistochemical analysis revealed the expression of Kir2.1, Kir2.2 and Kir2.3 within spontaneously bursting neurons. Importantly, Ba2+ application unmasked rhythmic burst-firing in ~42% of non-bursting lamina I neurons, suggesting that pacemaker activity is a latent property of a sizeable population of SDH cells during early life. In addition, the prevalence of spontaneous burst-firing within lamina I was enhanced in the presence of high internal concentrations of free Mg2+, consistent with its documented ability to block Kir channels from the intracellular side. Collectively, the results indicate that Kir channels are key modulators of pacemaker activity in newborn central pain networks. PMID:23426663

  11. Mechanism of inhibition of mouse Slo3 (KCa 5.1) potassium channels by quinine, quinidine and barium.

    Science.gov (United States)

    Wrighton, David C; Muench, Stephen P; Lippiat, Jonathan D

    2015-09-01

    The Slo3 (KCa 5.1) channel is a major component of mammalian KSper (sperm potassium conductance) channels and inhibition of these channels by quinine and barium alters sperm motility. The aim of this investigation was to determine the mechanism by which these drugs inhibit Slo3 channels. Mouse (m) Slo3 (KCa 5.1) channels or mutant forms were expressed in Xenopus oocytes and currents recorded with 2-electrode voltage-clamp. Gain-of-function mSlo3 mutations were used to explore the state-dependence of the inhibition. The interaction between quinidine and mSlo3 channels was modelled by in silico docking. Several drugs known to block KSper also affected mSlo3 channels with similar levels of inhibition. The inhibition induced by extracellular barium was prevented by increasing the extracellular potassium concentration. R196Q and F304Y mutations in the mSlo3 voltage sensor and pore, respectively, both increased channel activity. The F304Y mutation did not alter the effects of barium, but increased the potency of inhibition by both quinine and quinidine approximately 10-fold; this effect was not observed with the R196Q mutation. Block of mSlo3 channels by quinine, quinidine and barium is not state-dependent. Barium inhibits mSlo3 outside the cell by interacting with the selectivity filter, whereas quinine and quinidine act from the inside, by binding in a hydrophobic pocket formed by the S6 segment of each subunit. Furthermore, we propose that the Slo3 channel activation gate lies deep within the pore between F304 in the S6 segment and the selectivity filter. © 2015 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of The British Pharmacological Society.

  12. Cardamom extract induces cell proliferation by increasing potassium currents in NIH3T3 cell line.

    Science.gov (United States)

    Siddiqui, Sonia; Hassan, Sohail; Imran, Sumaira; Khan, Faisal; Ahmed, Faheem; Dar, Asim

    2017-11-01

    Amommum subulatum (Roxb.) or Cardamom extract is known to have anti-inflammatory and neuroprotective effects towards many gastrointestinal related problems. However, uptill now different fractions of cardamom extract on fibroblasts with respect to potassium channel activity have not been investigated. Therefore, present study investigated the effects of different fractions of cardamom extract on potassium channels in non-tumor NIH3T3 cell line. Phytochemical analysis of hydroalcoholic, n-hexane, butane and ethyl acetate fractions of cardamom extracts were purified and isolated by thin layer chromatography (TLC). 3T3 cells were cultured and incubated with hydroalcohol (1-2 μ/ml), n-hexane (1 μ/ml), butane (2 μ/ml) and ethyl acetate (1-2 μ/ml) for 5 hrs at 37°C. Modulation in potassium currents were recorded by whole-cell patch clamp method. The data showed two constituents Cineol (C 10 H 18 O) and Terpinyl acetate (C10H17OOCCH3) by TLC method. The present study shows that the constituents in n-hexane, hydro alcohol (1 μ/ml) and ethyl acetate (2 μ/ml) significantly increased (p<0.01) the potassium outward rectifying currents from NIH3T3 cells when compared to untreated controls cells. Whereas, butanol fraction (2 μ/ml) significantly decreased (p<0.01) the inward rectifying currents when compared to controls. Moreover hydroalcoholic and n-hexane fractions have increased the proliferation in 3T3 cell line. On the other hand butanol and ethyl acetate did not induce proliferation in 3T3 cells. Taken together, our data suggested that cardamom extract contains constituents that increased K+ currents, cell migration and proliferation and are involved in wound healing.

  13. Calcium-activated potassium channels mediated blood-brain tumor barrier opening in a rat metastatic brain tumor model

    Directory of Open Access Journals (Sweden)

    Ong John M

    2007-03-01

    Full Text Available Abstract Background The blood-brain tumor barrier (BTB impedes the delivery of therapeutic agents to brain tumors. While adequate delivery of drugs occurs in systemic tumors, the BTB limits delivery of anti-tumor agents into brain metastases. Results In this study, we examined the function and regulation of calcium-activated potassium (KCa channels in a rat metastatic brain tumor model. We showed that intravenous infusion of NS1619, a KCa channel agonist, and bradykinin selectively enhanced BTB permeability in brain tumors, but not in normal brain. Iberiotoxin, a KCa channel antagonist, significantly attenuated NS1619-induced BTB permeability increase. We found KCa channels and bradykinin type 2 receptors (B2R expressed in cultured human metastatic brain tumor cells (CRL-5904, non-small cell lung cancer, metastasized to brain, human brain microvessel endothelial cells (HBMEC and human lung cancer brain metastasis tissues. Potentiometric assays demonstrated the activity of KCa channels in metastatic brain tumor cells and HBMEC. Furthermore, we detected higher expression of KCa channels in the metastatic brain tumor tissue and tumor capillary endothelia as compared to normal brain tissue. Co-culture of metastatic brain tumor cells and brain microvessel endothelial cells showed an upregulation of KCa channels, which may contribute to the overexpression of KCa channels in tumor microvessels and selectivity of BTB opening. Conclusion These findings suggest that KCa channels in metastatic brain tumors may serve as an effective target for biochemical modulation of BTB permeability to enhance selective delivery of chemotherapeutic drugs to metastatic brain tumors.

  14. Role of calcium activated potassium channels in atrial fibrillation pathophysiology and therapy

    DEFF Research Database (Denmark)

    Diness, Jonas G.; Bentzen, Bo H.; S. Sørensen, Ulrik

    2015-01-01

    Small-conductance Ca2+-activated potassium (SK) channels are relative newcomers within the field of cardiac electrophysiology. In recent years, an increased focus has been given to these channels since they might constitute a relatively atrial selective target. The present review will give...... a general introduction to SK channels followed by their proposed function in the heart under normal and pathophysiological conditions. It is revealed how anti-arrhythmic effects can be obtained by SK channel inhibition in a number of species in situations of atrial fibrillation. On the contrary......, the beneficial effects of SK channel inhibition in situations of heart failure are questionable and still needs investigation. The understanding of cardiac SK channels is rapidly increasing these years, and hopefully this will clarify whether SK channel inhibition has potential as a new anti-atrial fibrillation...

  15. Role of renal vascular potassium channels in physiology and pathophysiology

    DEFF Research Database (Denmark)

    Salomonsson, Max; Brasen, Jens Christian; Sorensen, Charlotte Mehlin

    2017-01-01

    The control of renal vascular tone is important for the regulation of salt and water balance, blood pressure and the protection against damaging elevated glomerular pressure. The K+ conductance is a major factor in the regulation of the membrane potential (Vm ) in vascular smooth muscle (VSMC...... the ambiguous in vitro and in vivo results. We discuss the role of single types of K+ channels and the integrated function of several classes. We also deal with the possible role of renal vascular K+ channels in the pathophysiology of hypertension, diabetes mellitus and sepsis. This article is protected...

  16. The acrylamide (S)-1 differentially affects Kv7 (KCNQ) potassium channels

    DEFF Research Database (Denmark)

    Bentzen, Bo Hjorth; Schmitt, Nicole; Calloe, Kirstine

    2006-01-01

    .g., retigabine) for treatment of epilepsy and neuropathic pain. We investigated the effect of a Bristol-Myers Squibb compound (S)-N-[1-(3-morpholin-4-yl-phenyl)-ethyl]-3-phenyl-acrylamide [(S)-1] on cloned human Kv7.1-5 potassium channels expressed in Xenopus laevis oocytes. Using two-electrode voltage...

  17. Oestrogen promotes KCNQ1 potassium channel endocytosis and postendocytic trafficking in colonic epithelium

    Science.gov (United States)

    Rapetti-Mauss, Raphael; O'Mahony, Fiona; Sepulveda, Francisco V; Urbach, Valerie; Harvey, Brian J

    2013-01-01

    The cAMP-regulated potassium channel KCNQ1:KCNE3 plays an essential role in transepithelial Cl− secretion. Recycling of K+ across the basolateral membrane provides the driving force necessary to maintain apical Cl− secretion. The steroid hormone oestrogen (17β-oestradiol; E2), produces a female-specific antisecretory response in rat distal colon through the inhibition of the KCNQ1:KCNE3 channel. It has previously been shown that rapid inhibition of the channel conductance results from E2-induced uncoupling of the KCNE3 regulatory subunit from the KCNQ1 channel pore complex. The purpose of this study was to determine the mechanism required for sustained inhibition of the channel function. We found that E2 plays a role in regulation of KCNQ1 cell membrane abundance by endocytosis. Ussing chamber experiments have shown that E2 inhibits both Cl− secretion and KCNQ1 current in a colonic cell line, HT29cl.19A, when cultured as a confluent epithelium. Following E2 treatment, KCNQ1 was retrieved from the plasma membrane by a clathrin-mediated endocytosis, which involved the association between KCNQ1 and the clathrin adaptor, AP-2. Following endocytosis, KCNQ1 was accumulated in early endosomes. Following E2-induced endocytosis, rather than being degraded, KCNQ1 was recycled by a biphasic mechanism involving Rab4 and Rab11. Protein kinase Cδ and AMP-dependent kinase were rapidly phosphorylated in response to E2 on their activating phosphorylation sites, Ser643 and Thr172, respectively (as previously shown). Both kinases are necessary for the E2-induced endocytosis, because E2 failed to induce KCNQ1 internalization following pretreatment with specific inhibitors of both protein kinase Cδ and AMP-dependent kinase. The ubiquitin ligase Nedd4.2 binds KCNQ1 in response to E2 to induce channel internalization. This study has provided the first demonstration of hormonal regulation of KCNQ1 trafficking. In conclusion, we propose that internalization of KCNQ1 is a key event

  18. Resveratrol attenuates cortical neuron activity: roles of large conductance calcium-activated potassium channels and voltage-gated sodium channels.

    Science.gov (United States)

    Wang, Ya-Jean; Chan, Ming-Huan; Chen, Linyi; Wu, Sheng-Nan; Chen, Hwei-Hisen

    2016-05-21

    Resveratrol, a phytoalexin found in grapes and red wine, exhibits diverse pharmacological activities. However, relatively little is known about whether resveratrol modulates the ion channels in cortical neurons. The large-conductance calcium-activated potassium channels (BKCa) and voltage-gated sodium channels were expressed in cortical neurons and play important roles in regulation of neuronal excitability. The present study aimed to determine the effects of resveratrol on BKCa currents and voltage-gated sodium currents in cortical neurons. Resveratrol concentration-dependently increased the current amplitude and the opening activity of BKCa channels, but suppressed the amplitude of voltage-gated sodium currents. Similar to the BKCa channel opener NS1619, resveratrol decreased the firing rate of action potentials. In addition, the enhancing effects of BKCa channel blockers tetraethylammonium (TEA) and paxilline on action potential firing were sensitive to resveratrol. Our results indicated that the attenuation of action potential firing rate by resveratrol might be mediated through opening the BKCa channels and closing the voltage-gated sodium channels. As BKCa channels and sodium channels are critical molecular determinants for seizure generation, our findings suggest that regulation of these two channels in cortical neurons probably makes a considerable contribution to the antiseizure activity of resveratrol.

  19. Role of renal vascular potassium channels in physiology and pathophysiology

    DEFF Research Database (Denmark)

    Salomonsson, Max; Brasen, Jens Christian; Sorensen, Charlotte Mehlin

    2017-01-01

    of the renal parenchyma. The main classes of K+ channels (calcium activated (KCa ), inward rectifier (Kir ), voltage activated (Kv ) and ATP sensitive (KATP )) have been found in the renal vessels. In this review, we summarize results available in the literature and our own studies in the field. We compare...

  20. Active Sites of Spinoxin, a Potassium Channel Scorpion Toxin, Elucidated by Systematic Alanine Scanning.

    Science.gov (United States)

    Peigneur, Steve; Yamaguchi, Yoko; Kawano, Chihiro; Nose, Takeru; Nirthanan, Selvanayagam; Gopalakrishnakone, Ponnampalam; Tytgat, Jan; Sato, Kazuki

    2016-05-31

    Peptide toxins from scorpion venoms constitute the largest group of toxins that target the voltage-gated potassium channel (Kv). Spinoxin (SPX) isolated from the venom of scorpion Heterometrus spinifer is a 34-residue peptide neurotoxin cross-linked by four disulfide bridges. SPX is a potent inhibitor of Kv1.3 potassium channels (IC50 = 63 nM), which are considered to be valid molecular targets in the diagnostics and therapy of various autoimmune disorders and cancers. Here we synthesized 25 analogues of SPX and analyzed the role of each amino acid in SPX using alanine scanning to study its structure-function relationships. All synthetic analogues showed similar disulfide bond pairings and secondary structures as native SPX. Alanine replacements at Lys(23), Asn(26), and Lys(30) resulted in loss of activity against Kv1.3 potassium channels, whereas replacements at Arg(7), Met(14), Lys(27), and Tyr(32) also largely reduced inhibitory activity. These results suggest that the side chains of these amino acids in SPX play an important role in its interaction with Kv1.3 channels. In particular, Lys(23) appears to be a key residue that underpins Kv1.3 channel inhibition. Of these seven amino acid residues, four are basic amino acids, suggesting that the positive electrostatic potential on the surface of SPX is likely required for high affinity interaction with Kv1.3 channels. This study provides insight into the structure-function relationships of SPX with implications for the rational design of new lead compounds targeting potassium channels with high potency.

  1. Atomic basis for therapeutic activation of neuronal potassium channels

    DEFF Research Database (Denmark)

    Kim, Robin Y; Yau, Michael C; Galpin, Jason D

    2015-01-01

    with fluorinated Trp analogues, with increased H-bonding propensity, strengthens retigabine potency. In addition, potency of numerous retigabine analogues correlates with the negative electrostatic surface potential of a carbonyl/carbamate oxygen atom present in most KCNQ activators. These findings functionally...... pinpoint an atomic-scale interaction essential for effects of retigabine and provide stringent constraints that may guide rational improvement of the emerging drug class of KCNQ channel activators....

  2. Analogue pattern matching in a dendritic spine model based on phosphorylation of potassium channels.

    Science.gov (United States)

    Yang, K H; Blackwell, K T

    2000-11-01

    Modification of potassium channels by protein phosphorylation has been shown to play a role in learning and memory. If such memory storage machinery were part of dendritic spines, then a set of spines could act as an 'analogue pattern matching' device by learning a repeatedly presented pattern of synaptic activation. In this study, the plausibility of such analogue pattern matching is investigated in a detailed circuit model of a set of spines attached to a dendritic branch. Each spine head contains an AMPA synaptic channel in parallel with a calcium-dependent potassium channel whose sensitivity depends on its phosphorylation state. Repeated presentation of synaptic activity results in calcium activation of protein kinases and subsequent channel phosphorylation. Simulations demonstrate that signal strength is greatest when the synaptic input pattern is equal to the previously learned pattern, and smaller when components of the synaptic input pattern are either smaller or larger than corresponding components of the previously learned pattern. Therefore, our results indicate that dendritic spines may act as an analogue pattern matching device, and suggest that modulation of potassium channels by protein kinases may mediate neuronal pattern recognition.

  3. A novel crystallization method for visualizing the membrane localization of potassium channels.

    OpenAIRE

    Lopatin, A N; Makhina, E N; Nichols, C G

    1998-01-01

    The high permeability of K+ channels to monovalent thallium (Tl+) ions and the low solubility of thallium bromide salt were used to develop a simple yet very sensitive approach to the study of membrane localization of potassium channels. K+ channels (Kir1.1, Kir2.1, Kir2.3, Kv2.1), were expressed in Xenopus oocytes and loaded with Br ions by microinjection. Oocytes were then exposed to extracellular thallium. Under conditions favoring influx of Tl+ ions (negative membrane potential under volt...

  4. Microelectrode array measurement of potassium ion channel remodeling on the field action potential duration in rapid atrial pacing rabbits model.

    Science.gov (United States)

    Sun, Juan; Yan, Huang; Wugeti, Najina; Guo, Yujun; Zhang, Ling; Ma, Mei; Guo, Xingui; Jiao, Changan; Xu, Wenli; Li, Tianqi

    2015-01-01

    Atrial fibrillation (AF) arises from abnormalities in atrial structure and electrical activity. Microelectrode arrays (MEA) is a real-time, nondestructive measurement of the resting and action potential signal, from myocardial cells, to the peripheral circuit of electrophysiological activity. This study examined the field action potential duration (fAPD) of the right atrial appendage (RAA) by MEA in rapid atrial pacing (RAP) in the right atrium of rabbits. In addition, this study also investigated the effect of potassium ion channel blockers on fAPD. 40 New Zealand white rabbits of either sex were randomly divided into 3 groups: 1) the control, 2) potassium ion channel blocker (TEA, 4-Ap and BaCl2), and 3) amiodarone groups. The hearts were quickly removed and right atrial appendage sectioned (slice thickness 500 μm). Each slice was perfused with Tyrode's solution and continuously stimulated for 30 minutes. Sections from the control group were superfused with Tyrode's solution for 10 minutes, while the blocker groups and amiodarone were both treated with their respective compounds for 10 minutes each. The fAPD of RAA and action field action potential morphology were measured using MEA. In non-pace (control) groups, fAPD was 188.33 ± 18.29 ms after Tyrode's solution superfusion, and 173.91 ± 6.83 ms after RAP. In pace/potassium ion channel groups, TEA and BaCl2 superfusion prolonged atrial field action potential (fAPD) (control vs blocker: 176.67 ± 8.66 ms vs 196.11 ± 10.76 ms, 182.22 ± 12.87 ms vs 191.11 ± 13.09 ms with TEA and BaCl2 superfusion, respectively, P action potential in animal heart slices. After superfusing potassium ion channel blockers, fAPD was prolonged. These results suggest that Ito, IKur and IK1 remodel and mediate RAP-induced atrial electrical remodeling. Amiodarone alter potassium ion channel activity (Ito, IKur, IK1 and IKs), shortening fAPD.

  5. KCNE4 is an inhibitory subunit to Kv1.1 and Kv1.3 potassium channels

    DEFF Research Database (Denmark)

    Grunnet, Morten; Rasmussen, Hannne B; Hay-Schmidt, Anders

    2003-01-01

    Kv1 potassium channels are widely distributed in mammalian tissues and are involved in a variety of functions from controlling the firing rate of neurons to maturation of T-lymphocytes. Here we show that the newly described KCNE4 beta-subunit has a drastic inhibitory effect on currents generated...... is detected in the heart and in five different parts of the brain. Having the broad distribution of Kv1 channels in mind, the demonstrated inhibitory property of KCNE4-subunits could locally and/or transiently have a dramatic influence on cellular excitability and on setting resting membrane potentials....... by Kv1.1 and Kv1.3 potassium channels. The inhibition is found on channels expressed heterologously in both Xenopus oocytes and mammalian HEK293 cells. mKCNE4 does not inhibit Kv1.2, Kv1.4, Kv1.5, or Kv4.3 homomeric complexes, but it does significantly reduce current through Kv1.1/Kv1.2 and Kv1.2/Kv1...

  6. Slick (Kcnt2 Sodium-Activated Potassium Channels Limit Peptidergic Nociceptor Excitability and Hyperalgesia

    Directory of Open Access Journals (Sweden)

    Danielle L Tomasello

    2017-09-01

    Full Text Available The Slick (Kcnt2 sodium-activated potassium (K Na channel is a rapidly gating and weakly voltage-dependent and sodium-dependent potassium channel with no clearly defined physiological function. Within the dorsal root ganglia (DRGs, we show Slick channels are exclusively expressed in small-sized and medium-sized calcitonin gene–related peptide (CGRP-containing DRG neurons, and a pool of channels are localized to large dense-core vesicles (LDCV-containing CGRP. We stimulated DRG neurons for CGRP release and found Slick channels contained within CGRP-positive LDCV translocated to the neuronal membrane. Behavioral studies in Slick knockout (KO mice indicated increased basal heat detection and exacerbated thermal hyperalgesia compared with wild-type littermate controls during neuropathic and chronic inflammatory pain. Electrophysiologic recordings of DRG neurons from Slick KO mice revealed that Slick channels contribute to outward current, propensity to fire action potentials (APs, and to AP properties. Our data suggest that Slick channels restrain the excitability of CGRP-containing neurons, diminishing pain behavior after inflammation and injury.

  7. Activation of human ether-a-go-go-related gene potassium channels by the diphenylurea 1,3-bis-(2-hydroxy-5-trifluoromethyl-phenyl)-urea (NS1643)

    DEFF Research Database (Denmark)

    Hansen, Rie Schultz; Diness, Thomas Goldin; Christ, Torsten

    2005-01-01

    increased both steady-state and tail current at all voltages tested. The EC(50) value for HERG channel activation was 10.5 microM. These results were reproduced on HERG channels expressed in mammalian human embryonic kidney 293 cells. In guinea pig cardiomyocytes, studied by patch clamp, application of 10......-a-go-go-related gene (ERG) potassium channels. We have developed the diphenylurea compound 1,3-bis-(2-hydroxy-5-trifluoromethyl-phenyl)-urea (NS1643) and tested whether this small organic molecule could increase the activity of human ERG (HERG) channels expressed heterologously. In Xenopus laevis oocytes, NS1643...

  8. Ropivacaine-Induced Contraction Is Attenuated by Both Endothelial Nitric Oxide and Voltage-Dependent Potassium Channels in Isolated Rat Aortae

    Directory of Open Access Journals (Sweden)

    Seong-Ho Ok

    2013-01-01

    Full Text Available This study investigated endothelium-derived vasodilators and potassium channels involved in the modulation of ropivacaine-induced contraction. In endothelium-intact rat aortae, ropivacaine concentration-response curves were generated in the presence or absence of the following inhibitors: the nonspecific nitric oxide synthase (NOS inhibitor Nω-nitro-L-arginine methyl ester (L-NAME, the neuronal NOS inhibitor Nω-propyl-L-arginine hydrochloride, the inducible NOS inhibitor 1400W dihydrochloride, the nitric oxide-sensitive guanylyl cyclase (GC inhibitor ODQ, the NOS and GC inhibitor methylene blue, the phosphoinositide-3 kinase inhibitor wortmannin, the cytochrome p450 epoxygenase inhibitor fluconazole, the voltage-dependent potassium channel inhibitor 4-aminopyridine (4-AP, the calcium-activated potassium channel inhibitor tetraethylammonium (TEA, the inward-rectifying potassium channel inhibitor barium chloride, and the ATP-sensitive potassium channel inhibitor glibenclamide. The effect of ropivacaine on endothelial nitric oxide synthase (eNOS phosphorylation in human umbilical vein endothelial cells was examined by western blotting. Ropivacaine-induced contraction was weaker in endothelium-intact aortae than in endothelium-denuded aortae. L-NAME, ODQ, and methylene blue enhanced ropivacaine-induced contraction, whereas wortmannin, Nω-propyl-L-arginine hydrochloride, 1400W dihydrochloride, and fluconazole had no effect. 4-AP and TEA enhanced ropivacaine-induced contraction; however, barium chloride and glibenclamide had no effect. eNOS phosphorylation was induced by ropivacaine. These results suggest that ropivacaine-induced contraction is attenuated primarily by both endothelial nitric oxide and voltage-dependent potassium channels.

  9. A naturally occurring omega current in a Kv3 family potassium channel from a platyhelminth

    Directory of Open Access Journals (Sweden)

    Spencer Andrew N

    2008-06-01

    Full Text Available Abstract Background Voltage-gated ion channels are membrane proteins containing a selective pore that allows permeable ions to transit the membrane in response to a change in the transmembrane voltage. The typical selectivity filter in potassium channels is formed by a tetrameric arrangement of the carbonyl groups of the conserved amino-acid sequence Gly-Tyr-Gly. This canonical pore is opened or closed by conformational changes that originate in the voltage sensor (S4, a transmembrane helix with a series of positively charged amino acids. This sensor moves through a gating pore formed by elements of the S1, S2 and S3 helices, across the plane of the membrane, without allowing ions to pass through the membrane at that site. Recently, synthetic mutagenesis studies in the Drosophila melanogaster Shaker channel and analysis of human disease-causing mutations in sodium channels have identified amino acid residues that are integral parts of the gating-pore; when these residues are mutated the proteins allow a non-specific cation current, known as the omega current, to pass through the gating-pore with relatively low selectivity. Results The N.at-Kv3.2 potassium channel has an unusual weak inward rectifier phenotype. Several mutations of two amino acids in the voltage sensing (S4 transmembrane helix change the phenotype to a typical delayed rectifier. The inward rectifier channels (wild-type and mutant are sensitive to 4-aminopyridine (4-AP but not tetra-ethyl ammonium (TEA, whereas the delayed rectifier mutants are sensitive to TEA but not 4-AP. The inward rectifier channels also manifest low cation selectivity. The relative selectivity for different cations is sensitive to specific mutations in the S4 helix, Conclusion N.at-Kv3.2, a naturally occurring potassium channel of the Kv3 sequence family, mediates ion permeation through a modified gating pore, not the canonical, highly selective pore typical of potassium channels. This channel has evolved to

  10. Expression of inward rectifier potassium channel subunits in optic nerve glia

    OpenAIRE

    Brasko, Csilla

    2013-01-01

    In glia inward rectifying potassium channels (Kir) are predominantly responsible for the high selective membrane permeability to K+, for the maintenance of the RMP close to the EK and for the clearance of excess K+ released during action potentials by the process of K+ buffering. In this study I have investigated the expression, subcellular localisation and heteromer forming ability of Kir4.1, Kir5.1 and Kir2.1 subunits in optic nerve glia. Immunocytochemistry results demonstrated the e...

  11. Encephalitis due to antibodies to voltage gated potassium channel (VGKC with cerebellar involvement in a teenager

    Directory of Open Access Journals (Sweden)

    Megan M Langille

    2015-01-01

    Full Text Available Encephalitis due to antibodies to voltage gated potassium channel (VGKC typically presents with limbic encephalitis and medial temporal lobe involvement on neuroimaging. We describe a case of 13 year girl female with encephalitis due to antibodies to VGKC with signal changes in the cerebellar dentate nuclei bilaterally and clinical features that suggested predominant cerebellar involvement. These have never been reported previously in the literature. Our case expands the phenotypic spectrum of this rare condition.

  12. The potassium channel Ether à go-go is a novel prognostic factor with functional relevance in acute myeloid leukemia

    Directory of Open Access Journals (Sweden)

    Stühmer Walter

    2010-01-01

    Full Text Available Abstract Background The voltage-gated potassium channel hEag1 (KV10.1 has been related to cancer biology. The physiological expression of the human channel is restricted to the brain but it is frequently and abundantly expressed in many solid tumors, thereby making it a promising target for a specific diagnosis and therapy. Because chronic lymphatic leukemia has been described not to express hEag1, it has been assumed that the channel is not expressed in hematopoietic neoplasms in general. Results Here we show that this assumption is not correct, because the channel is up-regulated in myelodysplastic syndromes, chronic myeloid leukemia and almost half of the tested acute myeloid leukemias in a subtype-dependent fashion. Most interestingly, channel expression strongly correlated with increasing age, higher relapse rates and a significantly shorter overall survival. Multivariate Cox regression analysis revealed hEag1 expression levels in AML as an independent predictive factor for reduced disease-free and overall survival; such an association had not been reported before. As a functional correlate, specific hEag1 blockade inhibited the proliferation and migration of several AML cell lines and primary cultured AML cells in vitro. Conclusion Our observations implicate hEag1 as novel target for diagnostic, prognostic and/or therapeutic approaches in AML.

  13. Kir2.6 Regulates the Surface Expression of Kir2.x Inward Rectifier Potassium Channels*

    OpenAIRE

    Dassau, Lior; Conti, Lisa R.; Radeke, Carolyn M.; Ptáček, Louis J.; Vandenberg, Carol A.

    2011-01-01

    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 skele...

  14. Probing potassium channel function in vivo by intracellular delivery of antibodies in a rat model of retinal neurodegeneration

    Science.gov (United States)

    Raz-Prag, Dorit; Grimes, William N.; Fariss, Robert N.; Vijayasarathy, Camasamudram; Campos, Maria M.; Bush, Ronald A.; Diamond, Jeffrey S.; Sieving, Paul A.

    2010-01-01

    Inward rectifying potassium (Kir) channels participate in regulating potassium concentration (K+) in the central nervous system (CNS), including in the retina. We explored the contribution of Kir channels to retinal function by delivering Kir antibodies (Kir-Abs) into the rat eye in vivo to interrupt channel activity. Kir-Abs were coupled to a peptide carrier to reach intracellular epitopes. Functional effects were evaluated by recording the scotopic threshold response (STR) and photopic negative response (PhNR) of the electroretinogram (ERG) noninvasively with an electrode on the cornea to determine activity of the rod and cone pathways, respectively. Intravitreal delivery of Kir2.1-Ab coupled to the peptide carrier diminished these ERG responses equivalent to dimming the stimulus 10- to 100-fold. Immunohistochemistry (IHC) showed Kir2.1 immunostaining of retinal bipolar cells (BCs) matching the labeling pattern obtained with conventional IHC of applying Kir2.1-Ab to fixed retinal sections postmortem. Whole-cell voltage-clamp BC recordings in rat acute retinal slices showed suppression of barium-sensitive Kir2.1 currents upon inclusion of Kir2.1-Ab in the patch pipette. The in vivo functional and structural results implicate a contribution of Kir2.1 channel activity in these electronegative ERG potentials. Studies with Kir4.1-Ab administered in vivo also suppressed the ERG components and showed immunostaining of Müller cells. The strategy of administering Kir antibodies in vivo, coupled to a peptide carrier to facilitate intracellular delivery, identifies roles for Kir2.1 and Kir4.1 in ERG components arising in the proximal retina and suggests this approach could be of further value in research. PMID:20616020

  15. Mitochondria from rat uterine smooth muscle possess ATP-sensitive potassium channel

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    Olga B. Vadzyuk

    2018-03-01

    Full Text Available The objective of this study was to detect ATP-sensitive K+ uptake in rat uterine smooth muscle mitochondria and to determine possible effects of its activation on mitochondrial physiology. By means of fluorescent technique with usage of K+-sensitive fluorescent probe PBFI (potassium-binding benzofuran isophthalate we showed that accumulation of K ions in isolated mitochondria from rat myometrium is sensitive to effectors of KATP-channel (ATP-sensitive K+-channel – ATP, diazoxide, glibenclamide and 5HD (5-hydroxydecanoate. Our data demonstrates that K+ uptake in isolated myometrium mitochondria results in a slight decrease in membrane potential, enhancement of generation of ROS (reactive oxygen species and mitochondrial swelling. Particularly, the addition of ATP into incubation medium led to a decrease in mitochondrial swelling and ROS production, and an increase in membrane potential. These effects were eliminated by diazoxide. If blockers of KATP-channel were added along with diazoxide, the effects of diazoxide were removed. So, we postulate the existence of KATP-channels in rat uterus mitochondria and assume that their functioning may regulate physiological conditions of mitochondria, such as matrix volume, ROS generation and polarization of mitochondrial membrane. Keywords: ATP-sensitive potassium channel, Diazoxide, 5-hydroxydecanoate, Myometrium, Mitochondria, Mitochondrial swelling, Mitochondrial membrane potential, ROS

  16. Gain-of-function mutations in potassium channel subunit KCNE2 associated with early-onset lone atrial fibrillation

    DEFF Research Database (Denmark)

    Nielsen, Jonas Bille; Bentzen, Bo Hjorth; Olesen, Morten Salling

    2014-01-01

    Aims: Atrial fibrillation (AF) is the most common cardiac arrhythmia. Disturbances in cardiac potassium conductance are considered as one of the disease mechanisms in AF. We aimed to investigate if mutations in potassium-channel β-subunits KCNE2 and KCNE3 are associated with early-onset lone AF. ...

  17. Role of inward rectifier potassium channels in salivary gland function and sugar feeding of the fruit fly, Drosophila melanogaster

    Science.gov (United States)

    The arthropod salivary gland is of critical importance for horizontal transmission of pathogens, yet a detailed understanding of the ion conductance pathways responsible for saliva production and excretion is lacking. A superfamily of potassium ion channels, known as inward rectifying potassium (Ki...

  18. DPPX potassium channel antibody: frequency, clinical accompaniments, and outcomes in 20 patients.

    Science.gov (United States)

    Tobin, William Oliver; Lennon, Vanda A; Komorowski, Lars; Probst, Christian; Clardy, Stacey Lynn; Aksamit, Allen J; Appendino, Juan Pablo; Lucchinetti, Claudia F; Matsumoto, Joseph Y; Pittock, Sean J; Sandroni, Paola; Tippmann-Peikert, Maja; Wirrell, Elaine C; McKeon, Andrew

    2014-11-11

    To describe the detection frequency and clinical associations of immunoglobulin G (IgG) targeting dipeptidyl-peptidase-like protein-6 (DPPX), a regulatory subunit of neuronal Kv4.2 potassium channels. Specimens from 20 patients evaluated on a service basis by tissue-based immunofluorescence yielded a synaptic immunostaining pattern consistent with DPPX-IgG (serum, 20; CSF, all 7 available). Transfected HEK293 cell-based assay confirmed DPPX specificity in all specimens. Sixty-nine patients with stiff-person syndrome and related disorders were also evaluated by DPPX-IgG cell-based assay. Of 20 seropositive patients, 12 were men; median symptom onset age was 53 years (range, 13-75). Symptom onset was insidious in 15 and subacute in 5. Twelve patients reported prodromal weight loss. Neurologic disorders were multifocal. All had one or more brain or brainstem manifestations: amnesia (16), delirium (8), psychosis (4), depression (4), seizures (2), and brainstem disorders (15; eye movement disturbances [8], ataxia [7], dysphagia [6], dysarthria [4], respiratory failure [3]). Nine patients reported sleep disturbance. Manifestations of central hyperexcitability included myoclonus (8), exaggerated startle (6), diffuse rigidity (6), and hyperreflexia (6). Dysautonomia involved the gastrointestinal tract (9; diarrhea [6], gastroparesis, and constipation [3]), bladder (7), cardiac conduction system (3), and thermoregulation (1). Two patients had B-cell neoplasms: gastrointestinal lymphoma (1), and chronic lymphocytic leukemia (1). Substantial neurologic improvements followed immunotherapy in 7 of 11 patients with available treatment data. DPPX-IgG was not detected in any of the stiff-person syndrome patients. DPPX-IgG is a biomarker for an immunotherapy-responsive multifocal neurologic disorder of the central and autonomic nervous systems. © 2014 American Academy of Neurology.

  19. Cell volume and membrane stretch independently control K+ channel activity

    DEFF Research Database (Denmark)

    Bomholtz, Sofia Hammami; Willumsen, Niels J; Olsen, Hervør L

    2009-01-01

    A number of potassium channels including members of the KCNQ family and the Ca(2+) activated IK and SK, but not BK, are strongly and reversibly regulated by small changes in cell volume. It has been argued that this general regulation is mediated through sensitivity to changes in membrane stretch....... To test this hypothesis we have studied the regulation of KCNQ1 and BK channels after expression in Xenopus oocytes. Results from cell-attached patch clamp studies (approximately 50 microm(2) macropatches) in oocytes expressing BK channels demonstrate that the macroscopic volume-insensitive BK current...... that stretch and volume sensitivity can be considered two independent regulatory mechanisms....

  20. Quantitative Confocal Microscopy Analysis as a Basis for Search and Study of Potassium Kv1.x Channel Blockers

    Science.gov (United States)

    Feofanov, Alexey V.; Kudryashova, Kseniya S.; Nekrasova, Oksana V.; Vassilevski, Alexander A.; Kuzmenkov, Alexey I.; Korolkova, Yuliya V.; Grishin, Eugene V.; Kirpichnikov, Mikhail P.

    Artificial KcsA-Kv1.x (x = 1, 3) receptors were recently designed by transferring the ligand-binding site from human Kv1.x voltage-gated potassium channels into corresponding domain of the bacterial KscA channel. We found that KcsA-Kv1.x receptors expressed in E. coli cells are embedded into cell membrane and bind ligands when the cells are transformed to spheroplasts. We supposed that E. coli spheroplasts with membrane-embedded KcsA-Kv1.x and fluorescently labeled ligand agitoxin-2 (R-AgTx2) can be used as elements of an advanced analytical system for search and study of Kv1-channel blockers. To realize this idea, special procedures were developed for measurement and quantitative treatment of fluorescence signals obtained from spheroplast membrane using confocal laser scanning microscopy (CLSM). The worked out analytical "mix and read" systems supported by quantitative CLSM analysis were demonstrated to be reliable alternative to radioligand and electrophysiology techniques in the search and study of selective Kv1.x channel blockers of high scientific and medical importance.

  1. A single conserved basic residue in the potassium channel filter region controls KCNQ1 insensitivity toward scorpion toxins.

    Science.gov (United States)

    Chen, Zongyun; Hu, Youtian; Wang, Bin; Cao, Zhijian; Li, Wenxin; Wu, Yingliang

    2015-09-01

    Although many studies concerning the sensitivity mechanism of scorpion toxin-potassium channel interactions have been reported, few have explored the biochemical insensitivity mechanisms of potassium channel receptors toward natural scorpion toxin peptides, such as the KCNQ1 channel. Here, by sequence alignment analyses of the human KCNQ1 channel and scorpion potassium channel MmKv2, which is completely insensitive to scorpion toxins, we proposed that the insensitivity mechanism of KCNQ1 toward natural scorpion toxins might involve two functional regions, the turret and filter regions. Based on this observation, a series of KCNQ1 mutants were constructed to study molecular mechanisms of the KCNQ1 channel insensitivity toward natural scorpion toxins. Electrophysiological studies of chimera channels showed that the channel filter region controls KCNQ1 insensitivity toward the classical scorpion toxin ChTX. Interestingly, further residue mutant experiments showed that a single basic residue in the filter region determined the insensitivity of KCNQ1 channels toward scorpion toxins. Our present work showed that amino acid residue diversification at common sites controls the sensitivity and insensitivity of potassium channels toward scorpion toxins. The unique insensitivity mechanism of KCNQ1 toward natural scorpion toxins will accelerate the rational design of potent peptide inhibitors toward this channel.

  2. Thalamic microinfusion of antibody to a voltage-gated potassium channel restores consciousness during anesthesia.

    Science.gov (United States)

    Alkire, Michael T; Asher, Christopher D; Franciscus, Amanda M; Hahn, Emily L

    2009-04-01

    The Drosophila Shaker mutant fruit-fly, with its malfunctioning voltage-gated potassium channel, exhibits anesthetic requirements that are more than twice normal. Shaker mutants with an abnormal Kv1.2 channel also demonstrate significantly reduced sleep. Given the important role the thalamus plays in both sleep and arousal, the authors investigated whether localized central medial thalamic (CMT) microinfusion of an antibody designed to block the pore of the Kv1.2 channel might awaken anesthetized rats. Male Sprague-Dawley rats were implanted with a cannula aimed at the CMT or lateral thalamus. One week later, unconsciousness was induced with either desflurane (3.6 +/- 0.2%; n = 55) or sevoflurane (1.2 +/- 0.1%; n = 51). Arousal effects of a single 0.5-microl infusion of Kv1.2 potassium channel blocking antibody (0.1- 0.2 mg/ml) or a control infusion of Arc-protein antibody (0.2 mg/ml) were then determined. The Kv1.2 antibody, but not the control antibody, temporarily restored consciousness in 17% of all animals and in 75% of those animals where infusions occurred within the CMT (P Consciousness returned on average (+/- SD) 170 +/- 99 s after infusion and lasted a median time of 398 s (interquartile range: 279-510 s). Temporary seizures, without apparent consciousness, predominated in 33% of all animals. These findings support the idea that the CMT plays a role in modulating levels of arousal during anesthesia and further suggest that voltage-gated potassium channels in the CMT may contribute to regulating arousal or may even be relevant targets of anesthetic action.

  3. Modulation of Potassium Channel Activity in the Balance of ROS and ATP Production by Durum Wheat Mitochondria - An amazing defence tool against hyperosmotic stress

    Directory of Open Access Journals (Sweden)

    Daniela eTrono

    2015-12-01

    Full Text Available In plants, the existence of a mitochondrial potassium channel was firstly demonstrated about fifteen years ago in durum wheat as an ATP-dependent potassium channel (PmitoKATP. Since then, both properties of the original PmitoKATP and occurrence of different mitochondrial potassium channels in a number of plant species (monocotyledonous and dicotyledonous and tissues/organs (etiolated and green have been shown. Here, an overview of the current knowledge is reported; in particular, the issue of PmitoKATP physiological modulation is addressed. Similarities and differences with other potassium channels, as well as possible cross-regulation with other mitochondrial proteins (Plant Uncoupling Protein, Alternative Oxidase, Plant Inner Membrane Anion Channel are also described. PmitoKATP is inhibited by ATP and activated by superoxide anion, as well as by free fatty acids (FFAs and acyl-CoAs. Interestingly, channel activation increases electrophoretic potassium uptake across the inner membrane towards the matrix, so collapsing membrane potential (ΔΨ, the main component of the protonmotive force (Δp in plant mitochondria; moreover, cooperation between PmitoKATP and the K+/H+ antiporter allows a potassium cycle able to dissipate also ΔpH. Interestingly, ΔΨ collapse matches with an active control of mitochondrial reactive oxygen species (ROS production. Fully open channel is able to lower superoxide anion up to 35-fold compared to a condition of ATP-inhibited channel. On the other hand, ΔΨ collapse by PmitoKATP was unexpectedly found to not affect ATP synthesis via oxidative phosphorylation. This may probably occur by means of a controlled collapse due to ATP inhibition of PmitoKATP; this brake to the channel activity may allow a loss of the bulk phase Δp, but may preserve a non-classically detectable localized driving force for ATP synthesis. This ability may become crucial under environmental/oxidative stress. In particular, under moderate

  4. Ion channel profile of TRPM8 cold receptors reveals a novel role of TASK-3 potassium channels in thermosensation

    Science.gov (United States)

    Morenilla-Palao, Cruz; Luis, Enoch; Fernández-Peña, Carlos; Quintero, Eva; Weaver, Janelle L.; Bayliss, Douglas A.; Viana, Félix

    2017-01-01

    Summary Animals sense cold ambient temperatures through the activation of peripheral thermoreceptors that express TRPM8, a cold- and menthol-activated ion channel. These receptors can discriminate a very wide range of temperatures from innocuous to noxious. The molecular mechanism responsible for the variable sensitivity of individual cold receptors to temperature is unclear. To address this question, we performed a detailed ion channel expression analysis of cold sensitive neurons, combining BAC transgenesis with a molecular profiling approach in FACS purified TRPM8 neurons. We found that TASK-3 leak potassium channels are highly enriched in a subpopulation of these sensory neurons. The thermal threshold of TRPM8 cold neurons is decreased during TASK-3 blockade and in mice lacking TASK-3 and, most importantly, these mice display hypersensitivity to cold. Our results demonstrate a novel role of TASK-3 channels in thermosensation, showing that a channel-based combinatorial strategy in TRPM8 cold thermoreceptors leads to molecular specialization and functional diversity. PMID:25199828

  5. Ethanol affects network activity in cultured rat hippocampus: mediation by potassium channels.

    Directory of Open Access Journals (Sweden)

    Eduard Korkotian

    Full Text Available The effects of ethanol on neuronal network activity were studied in dissociated cultures of rat hippocampus. Exposure to low (0.25-0.5% ethanol concentrations caused an increase in synchronized network spikes, and a decrease in the duration of individual spikes. Ethanol also caused an increase in rate of miniature spontaneous excitatory postsynaptic currents. Higher concentrations of ethanol eliminated network spikes. These effects were reversible upon wash. The effects of the high, but not the low ethanol were blocked by the GABA antagonist bicuculline. The enhancing action of low ethanol was blocked by apamin, an SK potassium channel antagonist, and mimicked by 1-EBIO, an SK channel opener. It is proposed that in cultured hippocampal networks low concentration of ethanol is associated with SK channel activity, rather than the GABAergic receptor.

  6. [Reconstitution of large conductance calcium-activated potassium channels into artificial planar lipid bilayers].

    Science.gov (United States)

    Cheng, Jun; Zeng, Xiao-Rong; Tan, Xiao-Qiu; Li, Peng-Yun; Wen, Jing; Mao, Liang; Yang, Yan

    2017-06-25

    This study was aimed to establish a method to create a stable planar lipid bilayer membranes (PLBMs), in which large conductance calcium-activated potassium channels (BK Ca ) were reconstituted. Using spreading method, PLBMs were prepared by decane lipid fluid consisting of N 2 -weathered mixture of phosphatidylcholine and cholesterol at 3:1 ratio. After successful incorporation of BK Ca channel into PLBMs, single channel characteristics of BK Ca were studied by patch clamp method. The results showed that i) the single channel conductance of BK Ca was (206.8 ± 16.9) pS; ii) the activities of BK Ca channel were voltage dependent; iii) in the bath solution without Ca 2+ , there was almost no BK Ca channel activities regardless of under hyperpolarization or repolarization conditions; iv) under the condition of +40 mV membrane potential, BK Ca channels were activated in a Ca 2+ concentration dependent manner; v) when [Ca 2+ ] was increased from 1 μmol/L to 100 μmol/L, both the channel open probability and the average open time were increased, and the average close time was decreased from (32.2 ± 2.8) ms to (2.1 ± 1.8) ms; vi) the reverse potential of the reconstituted BK Ca was -30 mV when [K + ] was at 40/140 mmol/L (Cis/Trans). These results suggest that the spreading method could serve as a new method for preparing PLBMs and the reconstituted BK Ca into PLBMs showed similar electrophysiological characteristics to natural BK Ca channels, so the PLBMs with incorporated BK Ca can be used in the studies of pharmacology and dynamics of BK Ca channel.

  7. [Role of calcium activated-potassium channels in the injury to rat alveolar macrophages induced by quartz].

    Science.gov (United States)

    Li, Jun; Sun, Jingzhi; Yang, Li; Zhao, Jing; Wang, Zhenglun; Yang, Lei

    2014-01-01

    To investigate the role of calcium activated-potassium channels (KCa) in the injury to rat alveolar macrophages induced by quartz. The experiments were conducted on a rat alveolar macrophage cell line (NR8383) in vitro, where crystal silica (100 üg/ml) and amorphous silica (100 üg/ml) were used as the test substances and the cells without any treatment as negative controls. At first the effects of two kinds of quartz were compared. Then KCa special inhibitors (Paxilline for BK, Tram-34 for IK, Apamin for SK) were added in different doses to the in vitro test system with 100 üg/ml crystal quartz as matrix, to observe the function of such channels. Cell viability, lactate dehydrogenase (LDH), interleukin-1β (IL-1β) and tumor necrosis factor-a (TNF-α) were tested. Comparing to the negative control group, cell viability reduced, LDH leakage, IL-1β and TNF-α release increased significantly in the amorphous quartz group, furthermore, the effects by crystal quartz were much more serious than those by amorphous quartz, with a statistical significance (P quartz group, IK blockers (Tram-34) led to increase in cell viability significantly, with a statistical significance (P quartz in the rat alveolar macrophages cell line in vitro, which might serve as a signal in the early regulation of inflammatory responses by quartz.

  8. The Molecular Basis of Polyunsaturated Fatty Acid Interactions with the Shaker Voltage-Gated Potassium Channel.

    Directory of Open Access Journals (Sweden)

    Samira Yazdi

    2016-01-01

    Full Text Available Voltage-gated potassium (KV channels are membrane proteins that respond to changes in membrane potential by enabling K+ ion flux across the membrane. Polyunsaturated fatty acids (PUFAs induce channel opening by modulating the voltage-sensitivity, which can provide effective treatment against refractory epilepsy by means of a ketogenic diet. While PUFAs have been reported to influence the gating mechanism by electrostatic interactions to the voltage-sensor domain (VSD, the exact PUFA-protein interactions are still elusive. In this study, we report on the interactions between the Shaker KV channel in open and closed states and a PUFA-enriched lipid bilayer using microsecond molecular dynamics simulations. We determined a putative PUFA binding site in the open state of the channel located at the protein-lipid interface in the vicinity of the extracellular halves of the S3 and S4 helices of the VSD. In particular, the lipophilic PUFA tail covered a wide range of non-specific hydrophobic interactions in the hydrophobic central core of the protein-lipid interface, while the carboxylic head group displayed more specific interactions to polar/charged residues at the extracellular regions of the S3 and S4 helices, encompassing the S3-S4 linker. Moreover, by studying the interactions between saturated fatty acids (SFA and the Shaker KV channel, our study confirmed an increased conformational flexibility in the polyunsaturated carbon tails compared to saturated carbon chains, which may explain the specificity of PUFA action on channel proteins.

  9. Clinical spectrum and diagnostic value of antibodies against the potassium channel-related protein complex☆

    Science.gov (United States)

    Montojo, M.T.; Petit-Pedrol, M.; Graus, F.; Dalmau, J.

    2016-01-01

    Introduction Antibodies against a protein complex that includes voltage-gated potassium channels (VGKC) have been reported in patients with limbic encephalitis, peripheral nerve hyperexcitability, Morvan's syndrome, and a large variety of neurological syndromes. Review summary In this article, a review is presented of the syndromes associated with antibodies against VGKC-related proteins and the main antigens of this protein complex, the proteins LGI1 (leucine rich glioma inactivated protein 1) and Caspr2 (contactin-associated protein-like 2). The conceptual problems and clinical implications of the description of antibodies against VGKC-related proteins other than LGI1 and Caspr2 are also discussed. Although initial studies indicated the occurrence of antibodies against VGKC, recent investigations have shown that the main antigens are a neuronal secreted protein known as LGI1 which modulates synaptic excitability, and a protein called Caspr2 located on the cell surface and processes of neurons of different brain regions, and at the juxtaparanodal region of myelinated axons. While antibodies against LGI1 preferentially associate with classical limbic encephalitis, antibodies against Caspr2 associate with a wider spectrum of symptoms, including Morvan's syndrome, peripheral nerve hyperexcitability or neuromyotonia, and limbic or more extensive encephalitis. In addition there are reports of patients with antibodies against VGKC-related proteins that are different from LGI1 or Caspr2. In these cases, the identity and location of the antigens are unknown, the syndrome association is not specific, and the response to treatment uncertain. Conclusions The discovery of antigens such as LGI1 and Caspr2 has resulted in a clinical and molecular definition of the broad group of diseases previously attributed to antibodies against VGKC. Considering the literature that describes the presence of antibodies against VGKC other than LGI1 and Caspr2 proteins, we propose a practical

  10. Effects of potassium channel opener on the kinetics of thallium-201 in in-vitro and in-vivo

    Energy Technology Data Exchange (ETDEWEB)

    Lee, J.; Kim, E. J.; Ahn, B. C.; Chae, S. C.; Lee, K. B. [College of Medicine, Kyungpook National Univ., Taegu (Korea, Republic of); Kim, C. K. [Mt. Sinai Medical School, New York (United States)

    1997-07-01

    Potassium channel opener (K-opener) opens membrane ATP-sensitive K{sup +}-channel and induces and increase in potassium efflux from cells. K-openers are powerful smooth muscle relaxants and currently used as antihypertensive, antianginal drugs or bronchodilators in clinic. Pharmacologic potency of newly synthesized K-opener is being evaluated with efflux capacity of preincubated Rb-83 from the isolated aortic vascular tissue preparation. Thallium has similar characteristics to those of rubidium and potassium in vivo. To evaluate the effect of pinacidil (a potent K-opener) on Tl-201 biokinetics, we have performed uptake/washout studies in cultured myocytes, and mice biodistribution study. Primary culture of spontaneous contracting myocytes was undertake from hearts of newborn Sprague-Dawley rat. Different concentration of pinacidil (100nM or 10uM) was co-incubated with Tl-201 in HBSS buffer to evaluate its effect on cellular uptake, or challenged to myocyte preparations pre-incubated with Tl-201 for washout study. Pinacidil was injected into mice simultaneous or 10-min after Tl-201 injection, and organ uptake and whole body retention ratio was measured using gamma counter or dose calibrator. Co-incubation of pinacidil with Tl-201 resulted in a decrease in Tl uptake into myocytes by 1.6 - 2.5 times, and an increase in washout by 1.6 - 3.1 times. Pinacidil injection resulted in mild decrease in blood, heart and liver uptake in mice, bur renal uptake was markedly decreased in a dose dependent manner. These results suggest that the pinacidil Tl-201 kinetics and may potentially affect the interpretation of Tl-201 myocardial imaging.

  11. Disruption of ATP-sensitive potassium channel function in skeletal muscles promotes production and secretion of musclin

    Energy Technology Data Exchange (ETDEWEB)

    Sierra, Ana, E-mail: ana-sierra@uiowa.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Subbotina, Ekaterina, E-mail: ekaterina-subbotina@uiowa.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Zhu, Zhiyong, E-mail: zhiyong-zhu@uiowa.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Gao, Zhan, E-mail: zhan-gao@uiowa.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Koganti, Siva Rama Krishna, E-mail: sivaramakrishna.koganti@ttuhc.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Coetzee, William A., E-mail: william.coetzee@nyumc.org [Department of Pediatrics, NYU School of Medicine, New York, NY 10016 (United States); Goldhamer, David J., E-mail: david.goldhamer@uconn.edu [Center for Regenerative Biology, Department of Molecular and Cell Biology, Advanced Technology Laboratory, University of Connecticut, 1392 Storrs Road Unit 4243, Storrs, Connecticut 06269 (United States); Hodgson-Zingman, Denice M., E-mail: denice-zingman@uiowa.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Fraternal Order of Eagles Diabetes Research Center, Carver College of Medicine, Iowa City, IA 52242 (United States); Zingman, Leonid V., E-mail: leonid-zingman@uiowa.edu [Department of Internal Medicine, University of Iowa, Carver College of Medicine, Iowa City, IA 52242 (United States); Fraternal Order of Eagles Diabetes Research Center, Carver College of Medicine, Iowa City, IA 52242 (United States); Department of Veterans Affairs, Medical Center, Iowa City, IA 52242 (United States)

    2016-02-26

    Sarcolemmal ATP-sensitive potassium (K{sub ATP}) channels control skeletal muscle energy use through their ability to adjust membrane excitability and related cell functions in accordance with cellular metabolic status. Mice with disrupted skeletal muscle K{sub ATP} channels exhibit reduced adipocyte size and increased fatty acid release into the circulation. As yet, the molecular mechanisms underlying this link between skeletal muscle K{sub ATP} channel function and adipose mobilization have not been established. Here, we demonstrate that skeletal muscle-specific disruption of K{sub ATP} channel function in transgenic (TG) mice promotes production and secretion of musclin. Musclin is a myokine with high homology to atrial natriuretic peptide (ANP) that enhances ANP signaling by competing for elimination. Augmented musclin production in TG mice is driven by a molecular cascade resulting in enhanced acetylation and nuclear exclusion of the transcription factor forkhead box O1 (FOXO1) – an inhibitor of transcription of the musclin encoding gene. Musclin production/secretion in TG is paired with increased mobilization of fatty acids and a clear trend toward increased circulating ANP, an activator of lipolysis. These data establish K{sub ATP} channel-dependent musclin production as a potential mechanistic link coupling “local” skeletal muscle energy consumption with mobilization of bodily resources from fat. Understanding such mechanisms is an important step toward designing interventions to manage metabolic disorders including those related to excess body fat and associated co-morbidities. - Highlights: • ATP-sensitive K{sup +} channels regulate musclin production by skeletal muscles. • Lipolytic ANP signaling is promoted by augmented skeletal muscle musclin production. • Skeletal muscle musclin transcription is promoted by a CaMKII/HDAC/FOXO1 pathway. • Musclin links adipose mobilization to energy use in K{sub ATP} channel deficient skeletal muscle.

  12. Ankyrin-3 is a novel binding partner of the voltage-gated potassium channel Kv1.1 implicated in renal magnesium handling

    DEFF Research Database (Denmark)

    San-Cristobal, Pedro; Lainez, Sergio; Dimke, Henrik

    2013-01-01

    The voltage-gated potassium channel, Kv1.1, was recently identified as a causative gene in isolated dominant hypomagnesemia. The channel is situated in the distal convoluted tubule, where it participates in maintaining a favorable electrical gradient for driving magnesium ion into the cell through...... as compared to whole kidney. Electrophysiology studies performed in HEK293 cells expressing Kv1.1 showed that ANK3 significantly inhibited Kv1.1-mediated currents (267 compared to 125 pA/pF) for control and ANK3, respectively. Finally, to evaluate a potential role of ANK3 in magnesium handling, the intrarenal...

  13. Potassium transport in developing fleshy fruits: the grapevine inward K(+) channel VvK1.2 is activated by CIPK-CBL complexes and induced in ripening berry flesh cells.

    Science.gov (United States)

    Cuéllar, Teresa; Azeem, Farrukh; Andrianteranagna, Mamy; Pascaud, François; Verdeil, Jean-Luc; Sentenac, Hervé; Zimmermann, Sabine; Gaillard, Isabelle

    2013-03-01

    The grape berry provides a model for investigating the physiology of non-climacteric fruits. Increased K(+) accumulation in the berry has a strong negative impact on fruit acidity (and quality). In maturing berries, we identified a K(+) channel from the Shaker family, VvK1.2, and two CBL-interacting protein kinase (CIPK)/calcineurin B-like calcium sensor (CBL) pairs, VvCIPK04-VvCBL01 and VvCIPK03-VvCBL02, that may control the activity of this channel. VvCBL01 and VvCIPK04 are homologues of Arabidopsis AtCBL1 and AtCIPK23, respectively, which form a complex that controls the activity of the Shaker K(+) channel AKT1 in Arabidopsis roots. VvK1.2 remained electrically silent when expressed alone in Xenopus oocytes, but gave rise to K(+) currents when co-expressed with the pairs VvCIPK03-VvCBL02 or VvCIPK04-VvCBL01, the second pair inducing much larger currents than the first one. Other tested CIPK-CBL pairs expressed in maturing berries were found to be unable to activate VvK1.2. When activated by its CIPK-CBL partners, VvK1.2 acts as a voltage-gated inwardly rectifying K(+) channel that is activated at voltages more negative than -100 mV and is stimulated upon external acidification. This channel is specifically expressed in the berry, where it displays a very strong induction at veraison (the inception of ripening) in flesh cells, phloem tissues and perivascular cells surrounding vascular bundles. Its expression in these tissues is further greatly increased upon mild drought stress. VvK1.2 is thus likely to mediate rapid K(+) transport in the berry and to contribute to the extensive re-organization of the translocation pathways and transport mechanisms that occurs at veraison. © 2012 The Authors The Plant Journal © 2012 Blackwell Publishing Ltd.

  14. TWIK-1 two-pore domain potassium channels change ion selectivity and conduct inward leak sodium currents in hypokalemia.

    Science.gov (United States)

    Ma, Liqun; Zhang, Xuexin; Chen, Haijun

    2011-06-07

    Background potassium (K+) channels, which are normally selectively permeable to K+, maintain the cardiac resting membrane potential at around -80 mV. In subphysiological extracellular K+ concentrations ([K+]o), which occur in pathological hypokalemia, the resting membrane potential of human cardiomyocytes can depolarize to around -50 mV, whereas rat and mouse cardiomyocytes become hyperpolarized, consistent with the Nernst equation for K+. This paradoxical depolarization of cardiomyocytes in subphysiological [K+]o, which may contribute to cardiac arrhythmias, is thought to involve an inward leak sodium (Na+) current. Here, we show that human cardiac TWIK-1 (also known as K2P1) two-pore domain K+ channels change ion selectivity, becoming permeable to external Na+, and conduct inward leak Na+ currents in subphysiological [K+]o. A specific threonine residue (Thr118) within the pore selectivity sequence TxGYG was required for this altered ion selectivity. Mouse cardiomyocyte-derived HL-1 cells exhibited paradoxical depolarization with ectopic expression of TWIK-1 channels, whereas TWIK-1 knockdown in human spherical primary cardiac myocytes eliminated paradoxical depolarization. These findings indicate that ion selectivity of TWIK-1 K+ channels changes during pathological hypokalemia, elucidate a molecular basis for inward leak Na+ currents that could trigger or contribute to cardiac paradoxical depolarization in lowered [K+]o, and identify a mechanism for regulating cardiac excitability.

  15. ATP-sensitive potassium channels in isolated rat aorta during physiologic, hypoxic, and low-glucose conditions

    NARCIS (Netherlands)

    Hüsken, B. C.; Pfaffendorf, M.; van Zwieten, P. A.

    1997-01-01

    In arterial smooth muscle, adenosine triphosphate (ATP)-sensitive potassium channels are the targets of a variety of synthetic and endogenous vasodilators. In this study, we evaluated the influence of glibenclamide, an ATP-sensitive K(+)-channel blocker, on various vasodilator responses, including

  16. Potassium.

    Science.gov (United States)

    Halperin, M L; Kamel, K S

    1998-07-11

    In a logical, stepwise approach to patients presenting with hypokalaemia or hyperkalaemia the clinician must first recognise circumstances in which the dyskalaemia represents a clinical emergency because therapy then takes precedence over diagnosis. If a dyskalaemia has been present for a long time, there is an abnormal renal handling of K+. The next step to analyse is the rate of excretion of K+ and, if necessary, its two components (urine flow rate and K+ concentration in the cortical collecting duct [CCD]) analysed independently. If the K+ concentration in the CCD is not in the expected range, its basis should be defined at the ion-channel level in the CCD from clinical information that can be used to deduce the relative rates of reabsorption of Na+ and Cl- in the CCD. This analysis provides the basis for diagnosis and may indicate where non-emergency therapy should then be directed.

  17. Overexpression of GmAKT2 potassium channel enhances resistance to soybean mosaic virus.

    Science.gov (United States)

    Zhou, Lian; He, Hongli; Liu, Ruifang; Han, Qiang; Shou, Huixia; Liu, Bao

    2014-06-03

    Soybean mosaic virus (SMV) is the most prevalent viral disease in many soybean production areas. Due to a large number of SMV resistant loci and alleles, SMV strains and the rapid evolution in avirulence/effector genes, traditional breeding for SMV resistance is complex. Genetic engineering is an effective alternative method for improving SMV resistance in soybean. Potassium (K+) is the most abundant inorganic solute in plant cells, and is involved in plant responses to abiotic and biotic stresses. Studies have shown that altering the level of K+ status can reduce the spread of the viral diseases. Thus K+ transporters are putative candidates to target for soybean virus resistance. The addition of K+ fertilizer significantly reduced SMV incidence. Analysis of K+ channel gene expression indicated that GmAKT2, the ortholog of Arabidopsis K+ weak channel encoding gene AKT2, was significantly induced by SMV inoculation in the SMV highly-resistant genotype Rsmv1, but not in the susceptible genotype Ssmv1. Transgenic soybean plants overexpressing GmAKT2 were produced and verified by Southern blot and RT-PCR analysis. Analysis of K+ concentrations on different leaves of both the transgenic and the wildtype (Williams 82) plants revealed that overexpression of GmAKT2 significantly increased K+ concentrations in young leaves of plants. In contrast, K+ concentrations in the old leaves of the GmAKT2-Oe plants were significantly lower than those in WT plants. These results indicated that GmAKT2 acted as a K+ transporter and affected the distribution of K+ in soybean plants. Starting from 14 days after inoculation (DAI) of SMV G7, severe mosaic symptoms were observed on the WT leaves. In contrast, the GmAKT2-Oe plants showed no symptom of SMV infection. At 14 and 28 DAI, the amount of SMV RNA in WT plants increased 200- and 260- fold relative to GmAKT2-Oe plants at each time point. Thus, SMV development was significantly retarded in GmAKT2-overexpressing transgenic soybean

  18. An inhibitor of K+ channels modulates human endometrial tumor-initiating cells

    Directory of Open Access Journals (Sweden)

    Leslie Kimberly K

    2011-08-01

    Full Text Available Abstract Background Many potassium ion (K+ channels function as oncogenes to sustain growth of solid tumors, but their role in cancer progression is not well understood. Emerging evidence suggests that the early progenitor cancer cell subpopulation, termed tumor initiating cells (TIC, are critical to cancer progression. Results A non-selective antagonist of multiple types of K+ channels, tetraethylammonium (TEA, was found to suppress colony formation in endometrial cancer cells via inhibition of putative TIC. The data also indicated that withdrawal of TEA results in a significant enhancement of tumorigenesis. When the TIC-enriched subpopulation was isolated from the endometrial cancer cells, TEA was also found to inhibit growth in vitro. Conclusions These studies suggest that the activity of potassium channels significantly contributes to the progression of endometrial tumors, and the antagonists of potassium channels are candidate anti-cancer drugs to specifically target tumor initiating cells in endometrial cancer therapy.

  19. Compartmentalized beta subunit distribution determines characteristics and ethanol sensitivity of somatic, dendritic, and terminal large-conductance calcium-activated potassium channels in the rat central nervous system.

    Science.gov (United States)

    Wynne, P M; Puig, S I; Martin, G E; Treistman, S N

    2009-06-01

    Neurons are highly differentiated and polarized cells, whose various functions depend upon the compartmentalization of ion channels. The rat hypothalamic-neurohypophysial system (HNS), in which cell bodies and dendrites reside in the hypothalamus, physically separated from their nerve terminals in the neurohypophysis, provides a particularly powerful preparation in which to study the distribution and regional properties of ion channel proteins. Using electrophysiological and immunohistochemical techniques, we characterized the large-conductance calcium-activated potassium (BK) channel in each of the three primary compartments (soma, dendrite, and terminal) of HNS neurons. We found that dendritic BK channels, in common with somatic channels but in contrast to nerve terminal channels, are insensitive to iberiotoxin. Furthermore, analysis of dendritic BK channel gating kinetics indicates that they, like somatic channels, have fast activation kinetics, in contrast to the slow gating of terminal channels. Dendritic and somatic channels are also more sensitive to calcium and have a greater conductance than terminal channels. Finally, although terminal BK channels are highly potentiated by ethanol, somatic and dendritic channels are insensitive to the drug. The biophysical and pharmacological properties of somatic and dendritic versus nerve terminal channels are consistent with the characteristics of exogenously expressed alphabeta1 versus alphabeta4 channels, respectively. Therefore, one possible explanation for our findings is a selective distribution of auxiliary beta1 subunits to the somatic and dendritic compartments and beta4 to the terminal compartment. This hypothesis is supported immunohistochemically by the appearance of distinct punctate beta1 or beta4 channel clusters in the membrane of somatic and dendritic or nerve terminal compartments, respectively.

  20. The interference of uptake of thallium-201 in cultured rat myocardial cells with existence of potassium related pharmaceuticals--a preliminary report.

    Science.gov (United States)

    Chen, Y W; Yeh, J L; Huang, Y C; Chen, I J; Huang, Y F; Liu, G C

    2000-03-01

    Thallium-201 myocardial perfusion imaging is wildly used to detect and assess the extent of jeopardized myocardial ischemia in the coronary artery disease and the viability of myocardium post infarction. In recent years, there has been a great deal of pharmacological development of blockers and openers of potassium channel. In this study, we will discuss the interference of uptake of thallium-201 ion in cultured neonatal rat myocytes with existence of a variety of pharmacological agents. The cultures of neonatal rat myocardial cells were incubated with different agents such as potassium chloride, sodium-potassium ATPase pump inhibitor (ouabain), cesium compound, variable potassium channel blockers (4 AP, TEA and glibenclamide) and their openers (minoxidil, and cromakalim). The radioactivity of intracellular thallium-201 that could enter rat myocardial cells was detected by gamma counter sixty minutes after thallium-201 was added. In this study we found that thallium and potassium ions behave in an analogous manner in cultured rat myocardial cells. Both 2.5 mM and 5 mM concentration of extracellular potassium ion significantly result in reduction of thallium-201 ion influx in rat myocardial cells. 0.5 mM ouabain, an inhibitor of sodium-potassium ATPase pump, reduced about 40% of influx of thallium-201 ion in cultured rat myocardial cells via active transport. Combination of both potassium ion and ouabain inhibit most of thallium-201 ions influx in myocardial cells, but it is not completely inhibited. Cesium, a potassium antagonist, also interferes with the uptake of thallium-201 in cultured rat myocytes in our study. The most interesting finding in our investigation is that potassium channel blockers such as TEA and glibenclamide, inhibit the influx of thallium-201 in myocytes. However, potassium channel openers have no overt effect on influx of thallium-201 in cultured rat myocytes. We indirectly observe about 60% of influx of thallium-201 ion into cultured rat

  1. Inward-rectifying potassium channelopathies: new insights into disorders of sodium and potassium homeostasis.

    Science.gov (United States)

    Cheng, Chih-Jen; Sung, Chih-Chien; Huang, Chou-Long; Lin, Shih-Hua

    2015-03-01

    Inward-rectifying potassium (Kir) channels allow more inward than outward potassium flux when channels are open in mammalian cells. At physiological resting membrane potentials, however, they predominantly mediate outward potassium flux and play important roles in regulating the resting membrane potential in diverse cell types and potassium secretion in the kidneys. Mutations of Kir channels cause human hereditary diseases collectively called Kir channelopathies, many of which are characterized by disorders of sodium and potassium homeostasis. Studies on these genetic Kir channelopathies have shed light on novel pathophysiological mechanisms, including renal sodium and potassium handling, potassium shifting in skeletal muscles, and aldosterone production in the adrenal glands. Here, we review several recent advances in Kir channels and their clinical implications in sodium and potassium homeostasis.

  2. Comparison of the Effects of a Transient Outward Potassium Channel Activator on Currents Recorded from Atrial and Ventricular Cardiomyocytes

    DEFF Research Database (Denmark)

    Callø, Kirstine; Nof, Eyal; Jespersen, Thomas

    2011-01-01

    NS5806 activates the transient outward potassium current (I(to) ) in canine ventricular cells. We compared the effects of NS5806 on canine atrial versus ventricular tissues and myocytes.......NS5806 activates the transient outward potassium current (I(to) ) in canine ventricular cells. We compared the effects of NS5806 on canine atrial versus ventricular tissues and myocytes....

  3. Role of ATP-sensitive potassium channels in the piracetam induced blockade of opioid effects.

    Science.gov (United States)

    Rehni, Ashish K; Singh, Nirmal; Jindal, Seema

    2007-12-01

    The present study has been designed to investigate the effect of piracetam on morphine/ buprenorphine-induced antinociception in rats and effect of piracetam on morphine or minoxidil induced relaxation in KCl-precontracted isolated rat aortic ring preparation. Nociceptive threshold was measured by the tail flick test in rats. The cumulative dose responses of morphine or minoxidil were recorded in KCl-precontracted isolated rat aortic ring preparation. Piracetam attenuated buprenorphine-induced antinociception in rats. Piracetam significantly reduced the morphine and minoxidil induced relaxation in KCl precontracted isolated rat aortic ring preparation suggesting that piracetam interferes with opioid receptor and ATP-sensitive potassium channel (KATP) opener mediated responses in vitro. Thus, it may be suggested that piracetam attenuates opioid effects by an opioid receptor-KATP channel linked mechanism.

  4. Crystallization and preliminary X-ray diffraction analysis of human dipeptidyl peptidase 10 (DPPY), a component of voltage-gated potassium channels

    International Nuclear Information System (INIS)

    Bezerra, Gustavo Arruda; Dobrovetsky, Elena; Seitova, Alma; Dhe-Paganon, Sirano; Gruber, Karl

    2012-01-01

    The expression, purification and crystallization of human dipeptidyl peptidase 10, a component of voltage-gated potassium channels, is described. Dipeptidyl peptidase 10 (DPP10, DPPY) is an inactive peptidase associated with voltage-gated potassium channels, acting as a modulator of their electrophysiological properties, cell-surface expression and subcellular localization. Because potassium channels are important disease targets, biochemical and structural characterization of their interaction partners was sought. DPP10 was cloned and expressed using an insect-cell system and the protein was purified via His-tag affinity and size-exclusion chromatography. Crystals obtained by the sitting-drop method were orthorhombic, belonging to space group P2 1 2 1 2 1 with unit-cell parameters a = 80.91, b = 143.73, c = 176.25 Å. A single solution with two molecules in the asymmetric unit was found using the structure of DPP6 (also called DPPX) as the search model in a molecular replacement protocol

  5. Mechanism of HERG potassium channel inhibition by tetra-n-octylammonium bromide and benzethonium chloride

    Energy Technology Data Exchange (ETDEWEB)

    Long, Yan; Lin, Zuoxian [Key Laboratory of Regenerative Biology, Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510530 (China); Xia, Menghang; Zheng, Wei [National Center for Advancing Translational Sciences, National Institutes of Health, Bethesda, MD 20892 (United States); Li, Zhiyuan, E-mail: li_zhiyuan@gibh.ac.cn [Key Laboratory of Regenerative Biology, Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510530 (China)

    2013-03-01

    Tetra-n-octylammonium bromide and benzethonium chloride are synthetic quaternary ammonium salts that are widely used in hospitals and industries for the disinfection and surface treatment and as the preservative agent. Recently, the activities of HERG channel inhibition by these compounds have been found to have potential risks to induce the long QT syndrome and cardiac arrhythmia, although the mechanism of action is still elusive. This study was conducted to investigate the mechanism of HERG channel inhibition by these compounds by using whole-cell patch clamp experiments in a CHO cell line stably expressing HERG channels. Tetra-n-octylammonium bromide and benzethonium chloride exhibited concentration-dependent inhibitions of HERG channel currents with IC{sub 50} values of 4 nM and 17 nM, respectively, which were also voltage-dependent and use-dependent. Both compounds shifted the channel activation I–V curves in a hyperpolarized direction for 10–15 mV and accelerated channel activation and inactivation processes by 2-fold. In addition, tetra-n-octylammonium bromide shifted the inactivation I–V curve in a hyperpolarized direction for 24.4 mV and slowed the rate of channel deactivation by 2-fold, whereas benzethonium chloride did not. The results indicate that tetra-n-octylammonium bromide and benzethonium chloride are open-channel blockers that inhibit HERG channels in the voltage-dependent, use-dependent and state-dependent manners. - Highlights: ► Tetra-n-octylammonium and benzethonium are potent HERG channel inhibitors. ► Channel activation and inactivation processes are accelerated by the two compounds. ► Both compounds are the open-channel blockers to HERG channels. ► HERG channel inhibition by both compounds is use-, voltage- and state dependent. ► The in vivo risk of QT prolongation needs to be studied for the two compounds.

  6. Moderate hypoxia influences potassium outward currents in adipose-derived stem cells.

    Directory of Open Access Journals (Sweden)

    Mayuri Prasad

    Full Text Available Moderate hypoxic preconditioning of adipose-derived stem cells (ASCs enhances properties such as proliferation and secretion of growth factors, representing a valuable strategy to increase the efficiency of cell-based therapies. In a wide variety of cells potassium (K+ channels are key elements involved in the cellular responses to hypoxia, suggesting that ASCs cultured under low oxygen conditions may display altered electrophysiological properties. Here, the effects of moderate hypoxic culture on proliferation, whole-cell currents, and ion channel expression were investigated using human ASCs cultured at 5% and 20% oxygen. Although cell proliferation was greatly enhanced, the dose-dependent growth inhibition by the K+ channel blocker tetraethylammonium (TEA was not significantly affected by hypoxia. Under both normoxic and hypoxic conditions, ASCs displayed outward K+ currents composed by Ca2+-activated, delayed rectifier, and transient components. Hypoxic culture reduced the slope of the current-voltage curves and caused a negative shift in the voltage activation threshold of the whole-cell currents. However, the TEA-mediated shift of voltage activation threshold was not affected by hypoxia. Semiquantitative real-time RT-PCR revealed that expression of genes encoding for various ion channels subunits related to oxygen sensing and proliferation remained unchanged after hypoxic culture. In conclusion, outward currents are influenced by moderate hypoxia in ASCs through a mechanism that is not likely the result of modulation of TEA-sensitive K+ channels.

  7. Huntington disease skeletal muscle is hyperexcitable owing to chloride and potassium channel dysfunction.

    Science.gov (United States)

    Waters, Christopher W; Varuzhanyan, Grigor; Talmadge, Robert J; Voss, Andrew A

    2013-05-28

    Huntington disease is a progressive and fatal genetic disorder with debilitating motor and cognitive defects. Chorea, rigidity, dystonia, and muscle weakness are characteristic motor defects of the disease that are commonly attributed to central neurodegeneration. However, no previous study has examined the membrane properties that control contraction in Huntington disease muscle. We show primary defects in ex vivo adult skeletal muscle from the R6/2 transgenic mouse model of Huntington disease. Action potentials in diseased fibers are more easily triggered and prolonged than in fibers from WT littermates. Furthermore, some action potentials in the diseased fibers self-trigger. These defects occur because of decreases in the resting chloride and potassium conductances. Consistent with this, the expression of the muscle chloride channel, ClC-1, in Huntington disease muscle was compromised by improper splicing and a corresponding reduction in total Clcn1 (gene for ClC-1) mRNA. Additionally, the total Kcnj2 (gene for the Kir2.1 potassium channel) mRNA was reduced in disease muscle. The resulting muscle hyperexcitability causes involuntary and prolonged contractions that may contribute to the chorea, rigidity, and dystonia that characterize Huntington disease.

  8. Ion Concentration- and Voltage-Dependent Push and Pull Mechanisms of Potassium Channel Ion Conduction.

    Directory of Open Access Journals (Sweden)

    Kota Kasahara

    Full Text Available The mechanism of ion conduction by potassium channels is one of the central issues in physiology. In particular, it is still unclear how the ion concentration and the membrane voltage drive ion conduction. We have investigated the dynamics of the ion conduction processes in the Kv1.2 pore domain, by molecular dynamics (MD simulations with several different voltages and ion concentrations. By focusing on the detailed ion movements through the pore including selectivity filter (SF and cavity, we found two major conduction mechanisms, called the III-IV-III and III-II-III mechanisms, and the balance between the ion concentration and the voltage determines the mechanism preference. In the III-IV-III mechanism, the outermost ion in the pore is pushed out by a new ion coming from the intracellular fluid, and four-ion states were transiently observed. In the III-II-III mechanism, the outermost ion is pulled out first, without pushing by incoming ions. Increases in the ion concentration and voltage accelerated ion conductions, but their mechanisms were different. The increase in the ion concentrations facilitated the III-IV-III conductions, while the higher voltages increased the III-II-III conductions, indicating that the pore domain of potassium channels permeates ions by using two different driving forces: a push by intracellular ions and a pull by voltage.

  9. The calcium-activated potassium channel KCa3.1 is an important modulator of hepatic injury

    DEFF Research Database (Denmark)

    Møller, Linda Maria Sevelsted; Fialla, Annette Dam; Schierwagen, Robert

    2016-01-01

    The calcium-activated potassium channel KCa3.1 controls different cellular processes such as proliferation and volume homeostasis. We investigated the role of KCa3.1 in experimental and human liver fibrosis. KCa3.1 gene expression was investigated in healthy and injured human and rodent liver....... Effect of genetic depletion and pharmacological inhibition of KCa3.1 was evaluated in mice during carbon tetrachloride induced hepatic fibrogenesis. Transcription, protein expression and localisation of KCa3.1 was analysed by reverse transcription polymerase chain reaction, Western blot...... and immunohistochemistry. Hemodynamic effects of KCa3.1 inhibition were investigated in bile duct-ligated and carbon tetrachloride intoxicated rats. In vitro experiments were performed in rat hepatic stellate cells and hepatocytes. KCa3.1 expression was increased in rodent and human liver fibrosis and was predominantly...

  10. Diet-induced obesity impairs endothelium-derived hyperpolarization via altered potassium channel signaling mechanisms.

    Directory of Open Access Journals (Sweden)

    Rebecca E Haddock

    Full Text Available BACKGROUND: The vascular endothelium plays a critical role in the control of blood flow. Altered endothelium-mediated vasodilator and vasoconstrictor mechanisms underlie key aspects of cardiovascular disease, including those in obesity. Whilst the mechanism of nitric oxide (NO-mediated vasodilation has been extensively studied in obesity, little is known about the impact of obesity on vasodilation to the endothelium-derived hyperpolarization (EDH mechanism; which predominates in smaller resistance vessels and is characterized in this study. METHODOLOGY/PRINCIPAL FINDINGS: Membrane potential, vessel diameter and luminal pressure were recorded in 4(th order mesenteric arteries with pressure-induced myogenic tone, in control and diet-induced obese rats. Obesity, reflecting that of human dietary etiology, was induced with a cafeteria-style diet (∼30 kJ, fat over 16-20 weeks. Age and sexed matched controls received standard chow (∼12 kJ, fat. Channel protein distribution, expression and vessel morphology were determined using immunohistochemistry, Western blotting and ultrastructural techniques. In control and obese rat vessels, acetylcholine-mediated EDH was abolished by small and intermediate conductance calcium-activated potassium channel (SK(Ca/IK(Ca inhibition; with such activity being impaired in obesity. SK(Ca-IK(Ca activation with cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl-6-methyl-pyrimidin-4-yl]-amine (CyPPA and 1-ethyl-2-benzimidazolinone (1-EBIO, respectively, hyperpolarized and relaxed vessels from control and obese rats. IK(Ca-mediated EDH contribution was increased in obesity, and associated with altered IK(Ca distribution and elevated expression. In contrast, the SK(Ca-dependent-EDH component was reduced in obesity. Inward-rectifying potassium channel (K(ir and Na(+/K(+-ATPase inhibition by barium/ouabain, respectively, attenuated and abolished EDH in arteries from control and obese rats, respectively; reflecting differential K

  11. Clinical spectrum and diagnostic value of antibodies against the potassium channel related protein complex.

    Science.gov (United States)

    Montojo, M T; Petit-Pedrol, M; Graus, F; Dalmau, J

    2015-06-01

    Antibodies against a protein complex that includes voltage-gated potassium channels (VGKC) have been reported in patients with limbic encephalitis, peripheral nerve hyperexcitability, Morvan's syndrome, and a large variety of neurological syndromes. In this article, a review is presented of the syndromes associated with antibodies against VGKC-related proteins and the main antigens of this protein complex, the proteins LGI1 (leucine rich glioma inactivated protein 1) and Caspr2 (contactin-associated protein-like 2). The conceptual problems and clinical implications of the description of antibodies against VGKC-related proteins other than LGI1 and Caspr2 are also discussed. Although initial studies indicated the occurrence of antibodies against VGKC, recent investigations have shown that the main antigens are a neuronal secreted protein known as LGI1 which modulates synaptic excitability, and a protein called Caspr2 located on the cell surface and processes of neurons of different brain regions, and at the juxtaparanodal region of myelinated axons. While antibodies against LGI1 preferentially associate with classical limbic encephalitis, antibodies against Caspr2 associate with a wider spectrum of symptoms, including Morvan's syndrome, peripheral nerve hyperexcitability or neuromyotonia, and limbic or more extensive encephalitis. In addition there are reports of patients with antibodies against VGKC-related proteins that are different from LGI1 or Caspr2. In these cases, the identity and location of the antigens are unknown, the syndrome association is not specific, and the response to treatment uncertain. The discovery of antigens such as LGI1 and Caspr2 has resulted in a clinical and molecular definition of the broad group of diseases previously attributed to antibodies against VGKC. Considering the literature that describes the presence of antibodies against VGKC other than LGI1 and Caspr2 proteins, we propose a practical algorithm for the diagnosis and treatment

  12. Sodium/Potassium Homeostasis in the Cell

    DEFF Research Database (Denmark)

    Clausen, Michael Jakob; Poulsen, Hanne

    2013-01-01

    Metallomics and the Cell provides in an authoritative and timely manner in 16 stimulating chapters, written by 37 internationally recognized experts from 9 nations, and supported by more than 3000 references, several tables, and 110 illustrations, mostly in color, a most up-to-date view of the "m...

  13. Kv7 potassium channel activation with ICA-105665 reduces photoparoxysmal EEG responses in patients with epilepsy

    Science.gov (United States)

    Kasteleijn-Nolst Trenité, Dorotheé G A; Biton, Victor; French, Jacqueline A; Abou-Khalil, Bassel; Rosenfeld, William E; Diventura, Bree; Moore, Elizabeth L; Hetherington, Seth V; Rigdon, Greg C

    2013-01-01

    Purpose To assess the effects of ICA-105665, an agonist of neuronal Kv7 potassium channels, on epileptiform EEG discharges, evoked by intermittent photic stimulation (IPS), the so-called photoparoxysmal responses (PPRs) in patients with epilepsy. Methods Male and female patients aged 18–60 years with reproducible PPRs were eligible for enrollment. The study was conducted as a single-blind, single-dose, multiple-cohort study. Four patients were enrolled in each of the first three cohorts. Six patients were enrolled in the fourth cohort and one patient was enrolled in the fifth cohort. PPR responses to 14 IPS frequencies (steps) were used to determine the standard photosensitivity range (SPR) following placebo on day 1 and ICA-105665 on day 2. The SPR was quantified for three eye conditions (eyes closing, eyes closed, and eyes open), and the most sensitive condition was used for assessment of efficacy. A partial response was defined as a reduction in the SPR of at least three units at three separate time points following ICA-105665 compared to the same time points following placebo with no time points with more than three units of increase. Complete suppression was defined by no PPRs in any eye condition at one or more time points. Key Findings Six individual patients participated in the first three cohorts (100, 200, and 400 mg). Six patients participated in the fourth cohort (500 mg), and one patient participated in the fifth cohort (600 mg). Decreases in SPR occurred in one patient at 100 mg, two patients receiving 400 mg ICA-105665 (complete abolishment of SPR occurred in one patient at 400 mg), and in four of six patients receiving 500 mg. The most common adverse events (AEs) were those related to the nervous system, and dizziness appeared to be the first emerging AE. The single patient in the 600 mg cohort developed a brief generalized seizure within 1 h of dosing, leading to the discontinuation of additional patients at this dose, per the

  14. Urocortin2 prolongs action potential duration and modulates potassium currents in guinea pig myocytes and HEK293 cells.

    Science.gov (United States)

    Yang, Li-Zhen; Zhu, Yi-Chun

    2015-07-05

    We previously reported that activation of corticotropin releasing factor receptor type 2 by urocortin2 up-regulates both L-type Ca(2+) channels and intracellular Ca(2+) concentration in ventricular myocytes and plays an important role in cardiac contractility and arrhythmogenesis. This study goal was to further test the hypothesis that urocortin2 may modulate action potentials as well as rapidly and slowly activating delayed rectifier potassium currents. With whole cell patch-clamp techniques, action potentials and slowly activating delayed rectifier potassium currents were recorded in isolated guinea pig ventricular myocytes, respectively. And rapidly activating delayed rectifier potassium currents were tested in hERG-HEK293 cells. Urocortin2 produced a time- and concentration-dependent prolongation of action potential duration. The EC50 values of action potential duration and action potential duration at 90% of repolarization were 14.73 and 24.3nM respectively. The prolongation of action potential duration of urocortin2 was almost completely or partly abolished by H-89 (protein kinase A inhibitor) or KB-R7943 (Na(+)/Ca(2+) exchange inhibitor) pretreatment respectively. And urocortin2 caused reduction of rapidly activating delayed rectifier potassium currents in hERG-HEK293 cells. In addition, urocortin2 slowed the rate of slowly activating delayed rectifier potassium channel activation, and rightward shifted the threshold of slowly activating delayed rectifier potassium currents to more positive potentials. Urocortin2 prolonged action potential duration via activation of protein kinase A and Na(+)/ Ca(2+) exchange in isolated guinea pig ventricular myocytes in a time- and concentration- dependent manner. In hERG-HEK293 cells, urocortin2 reduced rapidly activating delayed rectifier potassium current density which may contribute to action potential duration prolongation. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Novel selective positive modulator of calcium-activated potassium channels exerts beneficial effects in a mouse model of spinocerebellar ataxia type 2

    Science.gov (United States)

    Kasumu, AW; Hougaard, C; Rode, F; Jacobsen, TA; Sabatier, JM; Eriksen, BL; Strøbæk, D; Liang, X; Egorova, P; Vorontsova, D; Christophersen, P; Rønn, LCB; Bezprozvanny, I

    2012-01-01

    Spinocerebellar ataxia type 2 (SCA2) is a neurodegenerative disorder caused by a polyglutamine expansion within the Ataxin-2 (Atxn2) protein. Purkinje cells (PC) of the cerebellum fire irregularly and eventually die in SCA2. We show here that the type 2 small conductance calcium-activated potassium channel (SK2) play a key role in control of normal PC activity. Using cerebellar slices from transgenic SCA2 mice we demonstrate that SK channel modulators restore regular pacemaker activity of SCA2 PCs. Furthermore, we also show that oral delivery of a novel selective positive modulator of SK2/3 channels (NS13001) alleviates behavioural and neuropathological phenotypes of aging SCA2 transgenic mice. We conclude that SK2 channels constitute a novel target for SCA2 treatment and that the developed selective SK2/3 modulator NS13001 holds promise as a potential therapeutic agent for treatment of SCA2 and possibly other cerebellar ataxias. PMID:23102227

  16. Hexachlorophene is a potent KCNQ1/KCNE1 potassium channel activator which rescues LQTs mutants.

    Science.gov (United States)

    Zheng, Yueming; Zhu, Xuejing; Zhou, Pingzheng; Lan, Xi; Xu, Haiyan; Li, Min; Gao, Zhaobing

    2012-01-01

    The voltage-gated KCNQ1 potassium channel is expressed in cardiac tissues, and coassembly of KCNQ1 with an auxiliary KCNE1 subunit mediates a slowly activating current that accelerates the repolarization of action potential in cardiomyocytes. Mutations of KCNQ1 genes that result in reduction or loss of channel activity cause prolongation of repolarization during action potential, thereby causing long QT syndrome (LQTs). Small molecule activators of KCNQ1/KCNE1 are useful both for understanding the mechanism of the complex activity and for developing therapeutics for LQTs. In this study we report that hexachlorophene (HCP), the active component of the topical anti-infective prescription drug pHisoHex, is a KCNQ1/KCNE1 activator. HCP potently increases the current amplitude of KCNQ1/KCNE1 expressed by stabilizing the channel in an open state with an EC(50) of 4.61 ± 1.29 μM. Further studies in cardiomyocytes showed that HCP significantly shortens the action potential duration at 1 μM. In addition, HCP is capable of rescuing the loss of function of the LQTs mutants caused by either impaired activation gating or phosphatidylinositol-4,5-bisphosphate (PIP2) binding affinity. Our results indicate HCP is a novel KCNQ1/KCNE1 activator and may be a useful tool compound for the development of LQTs therapeutics.

  17. Hexachlorophene is a potent KCNQ1/KCNE1 potassium channel activator which rescues LQTs mutants.

    Directory of Open Access Journals (Sweden)

    Yueming Zheng

    Full Text Available The voltage-gated KCNQ1 potassium channel is expressed in cardiac tissues, and coassembly of KCNQ1 with an auxiliary KCNE1 subunit mediates a slowly activating current that accelerates the repolarization of action potential in cardiomyocytes. Mutations of KCNQ1 genes that result in reduction or loss of channel activity cause prolongation of repolarization during action potential, thereby causing long QT syndrome (LQTs. Small molecule activators of KCNQ1/KCNE1 are useful both for understanding the mechanism of the complex activity and for developing therapeutics for LQTs. In this study we report that hexachlorophene (HCP, the active component of the topical anti-infective prescription drug pHisoHex, is a KCNQ1/KCNE1 activator. HCP potently increases the current amplitude of KCNQ1/KCNE1 expressed by stabilizing the channel in an open state with an EC(50 of 4.61 ± 1.29 μM. Further studies in cardiomyocytes showed that HCP significantly shortens the action potential duration at 1 μM. In addition, HCP is capable of rescuing the loss of function of the LQTs mutants caused by either impaired activation gating or phosphatidylinositol-4,5-bisphosphate (PIP2 binding affinity. Our results indicate HCP is a novel KCNQ1/KCNE1 activator and may be a useful tool compound for the development of LQTs therapeutics.

  18. Hexachlorophene Is a Potent KCNQ1/KCNE1 Potassium Channel Activator Which Rescues LQTs Mutants

    Science.gov (United States)

    Zheng, Yueming; Zhu, Xuejing; Zhou, Pingzheng; Lan, Xi; Xu, Haiyan; Li, Min; Gao, Zhaobing

    2012-01-01

    The voltage-gated KCNQ1 potassium channel is expressed in cardiac tissues, and coassembly of KCNQ1 with an auxiliary KCNE1 subunit mediates a slowly activating current that accelerates the repolarization of action potential in cardiomyocytes. Mutations of KCNQ1 genes that result in reduction or loss of channel activity cause prolongation of repolarization during action potential, thereby causing long QT syndrome (LQTs). Small molecule activators of KCNQ1/KCNE1 are useful both for understanding the mechanism of the complex activity and for developing therapeutics for LQTs. In this study we report that hexachlorophene (HCP), the active component of the topical anti-infective prescription drug pHisoHex, is a KCNQ1/KCNE1 activator. HCP potently increases the current amplitude of KCNQ1/KCNE1 expressed by stabilizing the channel in an open state with an EC50 of 4.61±1.29 μM. Further studies in cardiomyocytes showed that HCP significantly shortens the action potential duration at 1 μM. In addition, HCP is capable of rescuing the loss of function of the LQTs mutants caused by either impaired activation gating or phosphatidylinositol-4,5-bisphosphate (PIP2) binding affinity. Our results indicate HCP is a novel KCNQ1/KCNE1 activator and may be a useful tool compound for the development of LQTs therapeutics. PMID:23251633

  19. Effects of fluoxetine on protein expression of potassium ion channels in the brain of chronic mild stress rats

    Directory of Open Access Journals (Sweden)

    Chunlin Chen

    2015-01-01

    Full Text Available The purpose of this study is to investigate the expression of major potassium channel subtypes in the brain of chronical mild stress (CMS rats and reveal the effects of fluoxetine on the expression of these channels. Rats were exposed to a variety of unpredictable stress for three weeks and induced anhedonia, lower sucrose preference, locomotor activity and lower body weight. The protein expressions were determined by Western blot. CMS significantly increased the expression of Kv2.1 channel in frontal cortex but not in hippocampus, and the expression level was normalized after fluoxetine treatment. The expression of TREK-1 channel was also obviously increased in frontal cortex in CMS rats. Fluoxetine treatment might prevent this increase. However, the expression of Kv3.1 and Kv4.2 channels was considerably decreased in hippocampus after CMS, and was not affected by fluoxetine. These results suggest that different subtypes of potassium channels are associated with the pathophysiology of depression and that the therapeutical effects of fluoxetine may relate to Kv2.1 and TREK-1 potassium channels.

  20. Circadian rhythm in QT interval is preserved in mice deficient of potassium channel interacting protein 2

    DEFF Research Database (Denmark)

    Gottlieb, Lisa A; Lubberding, Anniek; Larsen, Anders Peter

    2017-01-01

    Potassium Channel Interacting Protein 2 (KChIP2) is suggested to be responsible for the circadian rhythm in repolarization duration, ventricular arrhythmias, and sudden cardiac death. We investigated the hypothesis that there is no circadian rhythm in QT interval in the absence of KChIP2. Implanted...... cardiac deaths were observed. We find similar diurnal (light:dark) and circadian (darkness) rhythms of RR intervals in WT and KChIP2(-/-) mice. Circadian rhythms in QT100 intervals are present in both groups, but at physiological small amplitudes: 1.6 ± 0.2 and 1.0 ± 0.3 ms in WT and KChIP2...

  1. K+ CHANNELEPSY: progress in the neurobiology of potassium channels and epilepsy

    Directory of Open Access Journals (Sweden)

    Maria Cristina D'Adamo

    2013-09-01

    Full Text Available K+ channels are important determinants of seizure susceptibility. These membrane proteins, encoded by more than 70 genes, make the largest group of ion channels that fine-tune the electrical activity of neuronal and non-neuronal cells in the brain. Their ubiquity and extremely high genetic and functional diversity, unmatched by any other ion channel type, place K+ channels as primary targets of genetic variations or perturbations in K+-dependent homeostasis, even in the absence of a primary channel defect. It is therefore not surprising that numerous inherited or acquired K+ channels dysfunctions have been associated with several neurologic syndromes, including epilepsy, which often generate confusion in the classification of the associated diseases. Therefore, we propose to name the K+ channels defects underlying distinct epilepsies as K+ channelepsies, and introduce a new nomenclature (e.g. Kx.y-channelepsy, following the widely used K+ channel classification, which could be also adopted to easily identify other channelopathies involving Na+ (e.g. Navx.y-phenotype, Ca2+ (e.g. Cavx.y-phenotype, and Cl- channels. Furthermore, we discuss novel genetic defects in K+ channels and associated proteins that underlie distinct epileptic phenotypes in humans, and analyze critically the recent progress in the neurobiology of this disease that has also been provided by investigations on valuable animal models of epilepsy. The abundant and varied lines of evidence discussed here strongly foster assessments for variations in genes encoding for K+ channels and associated proteins in patients with idiopathic epilepsy, provide new avenues for future investigations, and highlight these proteins as critical pharmacological targets.

  2. High-throughput screening for small-molecule modulators of inward rectifier potassium channels.

    Science.gov (United States)

    Raphemot, Rene; Weaver, C David; Denton, Jerod S

    2013-01-27

    Specific members of the inward rectifier potassium (Kir) channel family are postulated drug targets for a variety of disorders, including hypertension, atrial fibrillation, and pain. For the most part, however, progress toward understanding their therapeutic potential or even basic physiological functions has been slowed by the lack of good pharmacological tools. Indeed, the molecular pharmacology of the inward rectifier family has lagged far behind that of the S4 superfamily of voltage-gated potassium (Kv) channels, for which a number of nanomolar-affinity and highly selective peptide toxin modulators have been discovered. The bee venom toxin tertiapin and its derivatives are potent inhibitors of Kir1.1 and Kir3 channels, but peptides are of limited use therapeutically as well as experimentally due to their antigenic properties and poor bioavailability, metabolic stability and tissue penetrance. The development of potent and selective small-molecule probes with improved pharmacological properties will be a key to fully understanding the physiology and therapeutic potential of Kir channels. The Molecular Libraries Probes Production Center Network (MLPCN) supported by the National Institutes of Health (NIH) Common Fund has created opportunities for academic scientists to initiate probe discovery campaigns for molecular targets and signaling pathways in need of better pharmacology. The MLPCN provides researchers access to industry-scale screening centers and medicinal chemistry and informatics support to develop small-molecule probes to elucidate the function of genes and gene networks. The critical step in gaining entry to the MLPCN is the development of a robust target- or pathway-specific assay that is amenable for high-throughput screening (HTS). Here, we describe how to develop a fluorescence-based thallium (Tl(+)) flux assay of Kir channel function for high-throughput compound screening. The assay is based on the permeability of the K(+) channel pore to the K

  3. Potassium channel and NKCC cotransporter involvement in ocular refractive control mechanisms.

    Directory of Open Access Journals (Sweden)

    Sheila G Crewther

    Full Text Available Myopia affects well over 30% of adult humans globally. However, the underlying physiological mechanism is little understood. This study tested the hypothesis that ocular growth and refractive compensation to optical defocus can be controlled by manipulation of potassium and chloride ion-driven transretinal fluid movements to the choroid. Chicks were raised with +/-10D or zero power optical defocus rendering the focal plane of the eye in front of, behind, or at the level of the retinal photoreceptors respectively. Intravitreal injections of barium chloride, a non-specific inhibitor of potassium channels in the retina and RPE or bumetanide, a selective inhibitor of the sodium-potassium-chloride cotransporter were made, targeting fluid control mechanisms. Comparison of refractive compensation to 5 mM Ba(2+ and 10(-5 M bumetanide compared with control saline injected eyes shows significant change for both positive and negative lens defocus for Ba(2+ but significant change only for negative lens defocus with bumetanide (Rx(SAL(-10D = -8.6 +/- .9 D; Rx(Ba2+(-10D = -2.9 +/- .9 D; Rx(Bum(-10D = -2.9 +/- .9 D; Rx(SAL(+10D = +8.2 +/- .9 D; Rx(Ba2+(+10D = +2.8 +/- 1.3 D; Rx(Bum(+10D = +8.0 +/- .7 D. Vitreous chamber depths showed a main effect for drug conditions with less depth change in response to defocus shown for Ba(2+ relative to Saline, while bumetanide injected eyes showed a trend to increased depth without a significant interaction with applied defocus. The results indicate that both K channels and the NKCC cotransporter play a role in refractive compensation with NKCC blockade showing far more specificity for negative, compared with positive, lens defocus. Probable sites of action relevant to refractive control include the apical retinal pigment epithelium membrane and the photoreceptor/ON bipolar synapse. The similarities between the biometric effects of NKCC inhibition and biometric reports of the blockade of the retinal ON response, suggest a

  4. The KCNQ5 potassium channel from mouse: a broadly expressed M-current like potassium channel modulated by zinc, pH, and volume changes

    DEFF Research Database (Denmark)

    Jensen, Henrik Sindal; Callø, Kirstine; Jespersen, Thomas

    2005-01-01

    importance of the KCNQ channel family is emphasized by the fact that mutations in four of the five genes have been linked to human pathologies (KCNQ1 to 4). Here, we present the cloning and characterization of a novel KCNQ5 ortholog from mouse isolated by homology cloning from total mouse brain RNA (Gen......Bank accession number: AY679158). The predicted protein is 95% identical to human KCNQ5. Upon expression in Xenopus oocytes, these proteins form voltage-dependent slowly activating channels with half-maximal activation at -21 mV. Our functional characterization revealed three novel modes of modulation: p......H-dependent potentiation by Zn2+ (EC50 = 21.8 microM at pH 7.4), inhibition by acidification (IC50 = 0.75 microM; pKa = 6.1), and regulation by small changes in cell volume. Furthermore, the channels are activated by the anti-convulsant drug retigabine (EC50 = 2.0 microM) and inhibited by the M-current blockers...

  5. Oxidation of KCNB1 Potassium Channels Causes Neurotoxicity and Cognitive Impairment in a Mouse Model of Traumatic Brain Injury.

    Science.gov (United States)

    Yu, Wei; Parakramaweera, Randika; Teng, Shavonne; Gowda, Manasa; Sharad, Yashsavi; Thakker-Varia, Smita; Alder, Janet; Sesti, Federico

    2016-10-26

    The delayed rectifier potassium (K + ) channel KCNB1 (Kv2.1), which conducts a major somatodendritic current in cortex and hippocampus, is known to undergo oxidation in the brain, but whether this can cause neurodegeneration and cognitive impairment is not known. Here, we used transgenic mice harboring human KCNB1 wild-type (Tg-WT) or a nonoxidable C73A mutant (Tg-C73A) in cortex and hippocampus to determine whether oxidized KCNB1 channels affect brain function. Animals were subjected to moderate traumatic brain injury (TBI), a condition characterized by extensive oxidative stress. Dasatinib, a Food and Drug Administration-approved inhibitor of Src tyrosine kinases, was used to impinge on the proapoptotic signaling pathway activated by oxidized KCNB1 channels. Thus, typical lesions of brain injury, namely, inflammation (astrocytosis), neurodegeneration, and cell death, were markedly reduced in Tg-C73A and dasatinib-treated non-Tg animals. Accordingly, Tg-C73A mice and non-Tg mice treated with dasatinib exhibited improved behavioral outcomes in motor (rotarod) and cognitive (Morris water maze) assays compared to controls. Moreover, the activity of Src kinases, along with oxidative stress, were significantly diminished in Tg-C73A brains. Together, these data demonstrate that oxidation of KCNB1 channels is a contributing mechanism to cellular and behavioral deficits in vertebrates and suggest a new therapeutic approach to TBI. This study provides the first experimental evidence that oxidation of a K + channel constitutes a mechanism of neuronal and cognitive impairment in vertebrates. Specifically, the interaction of KCNB1 channels with reactive oxygen species plays a major role in the etiology of mouse model of traumatic brain injury (TBI), a condition associated with extensive oxidative stress. In addition, a Food and Drug Administration-approved drug ameliorates the outcome of TBI in mouse, by directly impinging on the toxic pathway activated in response to

  6. Calcium-activated potassium channel SK1 is widely expressed in the peripheral nervous system and sensory organs of adult zebrafish.

    Science.gov (United States)

    Cabo, R; Zichichi, R; Viña, E; Guerrera, M C; Vázquez, G; García-Suárez, O; Vega, J A; Germanà, A

    2013-10-25

    Sensory cells contain ion channels involved in the organ-specific transduction mechanisms that convert different types of stimuli into electric energy. Here we focus on small-conductance calcium-activated potassium channel 1 (SK1) which plays an important role in all excitable cells acting as feedback regulators in after-hyperpolarization. This study was undertaken to analyze the pattern of expression of SK1 in the zebrafish peripheral nervous system and sensory organs using RT-PRC, Westernblot and immunohistochemistry. Expression of SK1 mRNA was observed at all developmental stages analyzed (from 10 to 100 days post fertilization, dpf), and the antibody used identified a protein with a molecular weight of 70kDa, at 100dpf (regarded to be adult). Cell expressing SK1 in adult animals were neurons of dorsal root and cranial nerve sensory ganglia, sympathetic neurons, sensory cells in neuromasts of the lateral line system and taste buds, crypt olfactory neurons and photoreceptors. Present results report for the first time the expression and the distribution of SK1 in the peripheral nervous system and sensory organs of adult zebrafish, and may contribute to set zebrafish as an interesting experimental model for calcium-activated potassium channels research. Moreover these findings are of potential interest because the potential role of SK as targets for the treatment of neurological diseases and sensory disorders. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  7. Secondary structure, stability and tetramerisation of recombinant Kv1.1 potassium channel cytoplasmic N-terminal fragment.

    NARCIS (Netherlands)

    Abbott, G.W.; Bloemendal, M.; van Stokkum, I.H.M.; Mercer, E.A.J.; Miller, R.T.; Sewing, S.; Wolters, M.J.J.; Pongs, O.; Srai, S.K.S.

    1997-01-01

    The recombinant N-terminal fragment (amino acids 14-162) of a tetrameric voltage-gated potassium channel (K(V)1.1) has been studied using spectroscopic techniques. Evidence is presented that it forms a tetramer in aqueous solution, whereas when solubilised in 1% Triton X-100 it remains monomeric.

  8. Genetic variation in the two-pore domain potassium channel, TASK-1, may contribute to an atrial substrate for arrhythmogenesis

    DEFF Research Database (Denmark)

    Liang, Bo; Soka, Magdalena; Christensen, Alex Horby

    2013-01-01

    The two-pore domain potassium channel, K2P3.1 (TASK-1) modulates background conductance in isolated human atrial cardiomyocytes and has been proposed as a potential drug target for atrial fibrillation (AF). TASK-1 knockout mice have a predominantly ventricular phenotype however, and effects of TA...

  9. Protein self-assembly and lipid binding in the folding of the potassium channel KcsA

    NARCIS (Netherlands)

    Barrera, F.N.; Renard, M.L.; Poveda, J.A.; de Kruijff, B.; Killian, J.A.; González-Ros, J.M.

    2008-01-01

    Moderate concentrations of the alcohol 2,2,2-trifluoroethanol (TFE) cause the coupled unfolding and dissociation into subunits of the homotetrameric potassium channel KcsA, in a process that is partially irreversible when the protein is solubilized in plain dodecyl â-D-maltoside (DDM) micelles

  10. Role of calcium-activated potassium channels with small conductance in bradykinin-induced vasodilation of porcine retinal arterioles

    DEFF Research Database (Denmark)

    Dalsgaard, Thomas; Kroigaard, Christel; Bek, Toke

    2009-01-01

    PURPOSE: Endothelial dysfunction and impaired vasodilation may be involved in the pathogenesis of retinal vascular diseases. In the present study, the mechanisms underlying bradykinin vasodilation were examined and whether calcium-activated potassium channels of small (SK(Ca)) and intermediate (IK...

  11. Two inwardly rectifying potassium channels, Irk1 and Irk2, play redundant roles in Drosophila renal tubule function

    Science.gov (United States)

    Wu, Yipin; Baum, Michel; Huang, Chou-Long

    2015-01-01

    Inwardly rectifying potassium channels play essential roles in renal physiology across phyla. Barium-sensitive K+ conductances are found on the basolateral membrane of a variety of insect Malpighian (renal) tubules, including Drosophila melanogaster. We found that barium decreases the lumen-positive transepithelial potential difference in isolated perfused Drosophila tubules and decreases fluid secretion and transepithelial K+ flux. In those insect species in which it has been studied, transcripts from multiple genes encoding inwardly rectifying K+ channels are expressed in the renal (Malpighian) tubule. In Drosophila melanogaster, this includes transcripts of the Irk1, Irk2, and Irk3 genes. The role of each of these gene products in renal tubule function is unknown. We found that simultaneous knockdown of Irk1 and Irk2 in the principal cell of the fly tubule decreases transepithelial K+ flux, with no additive effect of Irk3 knockdown, and decreases barium sensitivity of transepithelial K+ flux by ∼50%. Knockdown of any of the three inwardly rectifying K+ channels individually has no effect, nor does knocking down Irk3 simultaneously with Irk1 or Irk2. Irk1/Irk2 principal cell double-knockdown tubules remain sensitive to the kaliuretic effect of cAMP. Inhibition of the Na+/K+-ATPase with ouabain and Irk1/Irk2 double knockdown have additive effects on K+ flux, and 75% of transepithelial K+ transport is due to Irk1/Irk2 or ouabain-sensitive pathways. In conclusion, Irk1 and Irk2 play redundant roles in transepithelial ion transport in the Drosophila melanogaster renal tubule and are additive to Na+/K+-ATPase-dependent pathways. PMID:26224687

  12. Social stress alters expression of large conductance calcium-activated potassium channel subunits in mouse adrenal medulla and pituitary glands.

    Science.gov (United States)

    Chatterjee, O; Taylor, L A; Ahmed, S; Nagaraj, S; Hall, J J; Finckbeiner, S M; Chan, P S; Suda, N; King, J T; Zeeman, M L; McCobb, D P

    2009-03-01

    Large conductance calcium-activated potassium (BK) channels are very prominently expressed in adrenal chromaffin and many anterior pituitary cells, where they shape intrinsic excitability complexly. Stress- and sex-steroids regulate alternative splicing of Slo-alpha, the pore-forming subunit of BK channels, and chronic behavioural stress has been shown to alter Slo splicing in tree shrew adrenals. In the present study, we focus on mice, measuring the effects of chronic behavioural stress on total mRNA expression of the Slo-alpha gene, two key BK channel beta subunit genes (beta2 and beta4), and the 'STREX' splice variant of Slo-alpha. As a chronic stressor, males of the relatively aggressive SJL strain were housed with a different unfamiliar SJL male every 24 h for 19 days. This 'social-instability' paradigm stressed all individuals, as demonstrated by reduced weight gain and elevated corticosterone levels. Five quantitative reverse transcriptase-polymerase chain assays were performed in parallel, including beta-actin, each calibrated against a dilution series of its corresponding cDNA template. Stress-related changes in BK expression were larger in mice tested at 6 weeks than 9 weeks. In younger animals, Slo-alpha mRNA levels were elevated 44% and 116% in the adrenal medulla and pituitary, respectively, compared to individually-housed controls. beta2 and beta4 mRNAs were elevated 162% and 194% in the pituitary, but slightly reduced in the adrenals of stressed animals. In the pituitary, dominance scores of stressed animals correlated negatively with alpha and beta subunit expression, with more subordinate individuals exhibiting levels that were three- to four-fold higher than controls or dominant individuals. STREX variant representation was lower in the subordinate subset. Thus, the combination of subunits responding to stress differs markedly between adrenal and pituitary glands. These data suggest that early stress will differentially affect neuroendocrine cell

  13. Free energy of a potassium ion in a model of the channel formed by an amphipathic leucine-serine peptide.

    Science.gov (United States)

    Smith, Graham R; Sansom, Mark S P

    2002-06-01

    We use molecular dynamics simulations to investigate the position-dependent free energy of a potassium ion in a model of an ion channel formed by the synthetic amphipathic leucine-serine peptide, LS3. The channel model is a parallel bundle of six LS3 helices around which are packed 146 methane-like spheres in order to mimic a membrane. At either end of and within the channel are 1051 water molecules, plus four ions (two potassium and two chloride). The free energy of a potassium ion in the channel was estimated using the weighted histogram analysis (WHAM) method. This is the first time to our knowledge that such a calculation has been carried out as a function of the position of an ion in three dimensions within a channel. The results indicate that for this channel, which is lined by hydrophilic serine sidechains, there is a relatively weak dependence of the free energy on the axial/off-axial position of the ion. There are some off-axis local minima, especially in the C-terminal half of the channel. Using the free energy results, a single channel current-voltage curve was estimated using a one-dimensional Nernst-Planck equation. Although reasonable agreement with experiment is achieved for K(+) ions flowing from the N-terminal to the C-terminal mouth, in the opposite direction the current is underestimated. This underestimation may be a consequence of under-sampling of the conformational dynamics of the channel. We suggest that our simulations may have captured, for example, a sub-conductance level (i.e. an incompletely open state) of the LS3 channel.

  14. Reversible antisense inhibition of Shaker-like Kv1.1 potassium channel expression impairs associative memory in mouse and rat

    Science.gov (United States)

    Meiri, Noam; Ghelardini, Carla; Tesco, Giuseppina; Galeotti, Nicoletta; Dahl, Dennis; Tomsic, Daniel; Cavallaro, Sebastiano; Quattrone, Alessandro; Capaccioli, Sergio; Bartolini, Alessandro; Alkon, Daniel L.

    1997-01-01

    Long-term memory is thought to be subserved by functional remodeling of neuronal circuits. Changes in the weights of existing synapses in networks might depend on voltage-gated potassium currents. We therefore studied the physiological role of potassium channels in memory, concentrating on the Shaker-like Kv1.1, a late rectifying potassium channel that is highly localized within dendrites of hippocampal CA3 pyramidal and dentate gyrus granular cells. Repeated intracerebroventricular injection of antisense oligodeoxyribonucleotide to Kv1.1 reduces expression of its particular intracellular mRNA target, decreases late rectifying K+ current(s) in dentate granule cells, and impairs memory but not other motor or sensory behaviors, in two different learning paradigms, mouse passive avoidance and rat spatial memory. The latter, hippocampal-dependent memory loss occurred in the absence of long-term potentiation changes recorded both from the dentate gyrus or CA1. The specificity of the reversible antisense targeting of mRNA in adult animal brains may avoid irreversible developmental and genetic background effects that accompany transgenic “knockouts”. PMID:9114006

  15. Reversible antisense inhibition of Shaker-like Kv1.1 potassium channel expression impairs associative memory in mouse and rat.

    Science.gov (United States)

    Meiri, N; Ghelardini, C; Tesco, G; Galeotti, N; Dahl, D; Tomsic, D; Cavallaro, S; Quattrone, A; Capaccioli, S; Bartolini, A; Alkon, D L

    1997-04-29

    Long-term memory is thought to be subserved by functional remodeling of neuronal circuits. Changes in the weights of existing synapses in networks might depend on voltage-gated potassium currents. We therefore studied the physiological role of potassium channels in memory, concentrating on the Shaker-like Kv1.1, a late rectifying potassium channel that is highly localized within dendrites of hippocampal CA3 pyramidal and dentate gyrus granular cells. Repeated intracerebroventricular injection of antisense oligodeoxyribonucleotide to Kv1.1 reduces expression of its particular intracellular mRNA target, decreases late rectifying K+ current(s) in dentate granule cells, and impairs memory but not other motor or sensory behaviors, in two different learning paradigms, mouse passive avoidance and rat spatial memory. The latter, hippocampal-dependent memory loss occurred in the absence of long-term potentiation changes recorded both from the dentate gyrus or CA1. The specificity of the reversible antisense targeting of mRNA in adult animal brains may avoid irreversible developmental and genetic background effects that accompany transgenic "knockouts".

  16. Calcium-activated potassium channels in insect pacemaker neurons as unexpected target site for the novel fumigant dimethyl disulfide.

    Science.gov (United States)

    Gautier, Hélène; Auger, Jacques; Legros, Christian; Lapied, Bruno

    2008-01-01

    Dimethyl disulfide (DMDS), a plant-derived insecticide, is a promising fumigant as a substitute for methyl bromide. To further understand the mode of action of DMDS, we examined its effect on cockroach octopaminergic neurosecretory cells, called dorsal unpaired median (DUM) neurons, using whole-cell patch-clamp technique, calcium imaging and antisense oligonucleotide strategy. At low concentration (1 microM), DMDS modified spontaneous regular spike discharge into clear bursting activity associated with a decrease of the amplitude of the afterhyperpolarization. This effect led us to suspect alterations of calcium-activated potassium currents (IKCa) and [Ca(2+)](i) changes. We showed that DMDS reduced amplitudes of both peak transient and sustained components of the total potassium current. IKCa was confirmed as a target of DMDS by using iberiotoxin, cadmium chloride, and pSlo antisense oligonucleotide. In addition, we showed that DMDS induced [Ca(2+)](i) rise in Fura-2-loaded DUM neurons. Using calcium-free solution, and (R,S)-(3,4-dihydro-6,7-dimethoxy-isoquinoline-1-yl)-2-phenyl-N,N-di-[2-(2,3,4-trimethoxy-phenyl)ethyl]-acetamide (LOE 908) [an inhibitor of transient receptor potential (TRP)gamma], we demonstrated that TRPgamma initiated calcium influx. By contrast, omega-conotoxin GVIA (an inhibitor of N-type high-voltage-activated calcium channels), did not affect the DMDS-induced [Ca(2+)](i) rise. Finally, the participation of the calcium-induced calcium release mechanism was investigated using thapsigargin, caffeine, and ryanodine. Our study revealed that DMDS-induced elevation in [Ca(2+)](i) modulated IKCa in an unexpected bell-shaped manner via intracellular calcium. In conclusion, DMDS affects multiple targets, which could be an effective way to improve pest control efficacy of fumigation.

  17. Overexpression of the potassium channel TPKb in small vacuoles confers osmotic and drought tolerance to rice.

    Science.gov (United States)

    Ahmad, Izhar; Devonshire, Jean; Mohamed, Radwa; Schultze, Michael; Maathuis, Frans J M

    2016-02-01

    Potassium (K(+) ) is the most important cationic nutrient for all living organisms. Vacuolar two-pore K(+) (TPK) channels are important players in the regulation of cellular levels of K(+) but have not been characterised in rice. In order to assess the role of OsTPKb, a K(+) selective ion channel predominantly expressed in the tonoplast of small vacuoles, we generated overexpressing (OX) lines using a constitutive promoter and compared their phenotypes with control plants. Relative to control plants, OX lines showed better growth when exposed to low-K(+) or water stress conditions. K(+) uptake was greater in OX lines which may be driven by increased AKT1 and HAK1 activity. The enhanced K(+) uptake led to tissue K(+) levels that were raised in roots and shoots. Furthermore, energy dispersive X-ray (EDX) analyses showed a higher cytoplasm: vacuole K(+) ratio which is likely to contribute to the increased stress tolerance. In all, the data suggest that TPKb can alter the K(+) status of small vacuoles, which is important for general cellular K(+) homeostasis which, in turn, affects stress tolerance. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

  18. Upregulation of basolateral small conductance potassium channels (KCNQ1/KCNE3) in ulcerative colitis.

    Science.gov (United States)

    Al-Hazza, Adel; Linley, John; Aziz, Qadeer; Hunter, Malcolm; Sandle, Geoffrey

    2016-02-05

    Basolateral K(+) channels hyperpolarize colonocytes to ensure Na(+) (and thus water) absorption. Small conductance basolateral (KCNQ1/KCNE3) K(+) channels have never been evaluated in human colon. We therefore evaluated KCNQ1/KCNE3 channels in distal colonic crypts obtained from normal and active ulcerative colitis (UC) patients. KCNQ1 and KCNE3 mRNA levels were determined by qPCR, and KCNQ1/KCNE3 channel activity in normal and UC crypts, and the effects of forskolin (activator of adenylate cyclase) and UC-related proinflammatory cytokines on normal crypts, studied by patch clamp recording. Whereas KCNQ1 and KCNE3 mRNA expression was similar in normal and UC crypts, single 6.8 pS channels were seen in 36% of basolateral patches in normal crypts, and to an even greater extent (74% of patches, P KCNQ1/KCNE3 channels make only a small contribution to basolateral conductance in normal colonic crypts, with increased channel activity in UC appearing insufficient to prevent colonic cell depolarization in this disease. This supports the proposal that defective Na(+) absorption rather than enhanced Cl(-) secretion, is the dominant pathophysiological mechanism of diarrhea in UC. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Sequence Alterations of I(Ks Potassium Channel Genes in Kazakhstani Patients with Atrial Fibrillation

    Directory of Open Access Journals (Sweden)

    Ainur Akilzhanova

    2014-12-01

    Full Text Available Introduction. Atrial fibrillation (AF is the most common sustained arrhythmia, and it results in significant morbidity and mortality. However, the pathogenesis of AF remains unclear to date. Recently, more pieces of evidence indicated that AF is a multifactorial disease resulting from the interaction between environmental factors and genetics. Recent studies suggest that genetic mutation of the slow delayed rectifier potassium channel (I(Ks may underlie AF.Objective. To investigate sequence alterations of I(Ks potassium channel genes KCNQ1, KCNE1 and KCNE2 in Kazakhstani patients with atrial fibrillation.Methods. Genomic DNA of 69 cases with atrial fibrillation and 27 relatives were analyzed for mutations in all protein-coding exons and their flanking splice site regions of the genes KCNQ1 (NM_000218.2 and NM_181798.1, KCNE1 (NM_000219.2, and KCNE2 (NM_172201.1 using bidirectional sequencing on the ABI 3730xL DNA Analyzer (Applied Biosystems, Foster City, CA, USA.Results. In total, a disease-causing mutation was identified in 39 of the 69 (56.5% index cases. Of these, altered sequence variants in the KCNQ1 gene accounted for 14.5% of the mutations, whereas a KCNE1 mutation accounted for 43.5% of the mutations and KCNE2 mutation accounted for 1.4% of the mutations. The majority of the distinct mutations were found in a single case (80%, whereas 20% of the mutations were observed more than once. We found two sequence variants in KCNQ1 exon 13 (S546S G1638A and exon 16 (Y662Y, C1986T in ten patients (14.5%. In KCNE1 gene in exon 3 mutation, S59G A280G was observed in 30 of 69 patients (43.5% and KCNE2 exon 2 T10K C29A in 1 patient (1.4%. Genetic cascade screening of 27 relatives to the 69 index cases with an identified mutation revealed 26.9% mutation carriers  who were at risk of cardiac events such as syncope or sudden unexpected death.Conclusion. In this cohort of Kazakhstani index cases with AF, a disease-causing mutation was identified in

  20. Nitroblue tetrazolium blocks BK channels in cerebrovascular smooth muscle cell membranes

    OpenAIRE

    Ye, D; Pospisilik, J A; Mathers, D A

    2000-01-01

    The effects of p-nitroblue tetrazolium (NBT) on large conductance, calcium-activated potassium channels (BK channels) in enzymatically dispersed rat cerebrovascular smooth muscle cells (CVSMCs) were examined.Patch clamp methods were employed to record single BK channel currents from inside-out patches of CVMC membrane maintained at 21–23°C.When applied to the cytoplasmic face of inside-out membrane patches (internally applied NBT), micromolar concentrations of NBT reversible reduced the mean ...

  1. Potassium accumulation in the globally ischemic mammalian heart. A role for the ATP-sensitive potassium channel

    NARCIS (Netherlands)

    Wilde, A. A.; Escande, D.; Schumacher, C. A.; Thuringer, D.; Mestre, M.; Fiolet, J. W.; Janse, M. J.

    1990-01-01

    We investigated the contribution of opening of the ATP-sensitive K+ channel to extracellular accumulation of K+ during ischemia with the use of glibenclamide, a specific blocker of this K+ channel. To characterize the electrophysiological effects of glibenclamide during metabolic inhibition (by

  2. PIST (GOPC modulates the oncogenic voltage-gated potassium channel KV10.1

    Directory of Open Access Journals (Sweden)

    Solveig eHerrmann

    2013-08-01

    Full Text Available Although crucial for their correct function, the mechanisms controlling surface expression of ion channels are poorly understood. In the case of the voltage-gated potassium channel KV10.1,this is determinant not only for its physiological function in brain, but also for its pathophysiology in tumors and possible use as a therapeutic target. The Golgi resident protein PIST binds several membrane proteins, thereby modulating their expression. Here we describe a PDZ domain-mediated interaction of KV10.1and PIST, which enhances surface levels ofKV10.1. The functional, but not the physical interaction of both proteins is dependent on the coiled-coil and PDZ domains of PIST; insertion of eight amino acids in the coiled-coil domain to render the neural form of PIST (nPIST and the corresponding short isoform in an as-of-yet unknown form abolishes the effect. In addition, two new isoforms of PIST (sPIST and nsPIST lacking nearly the complete PDZ domain were cloned and shown to be ubiquitously expressed. PIST and KV10.1 co-precipitate from native and expression systems. nPISTalso showed interaction, but did not alter the functional expression of the channel. We could not document physical interaction betweenKV10.1 and sPIST, but it reduced KV10.1 functional expression in a dominant-negative manner. nsPIST showed weak physical interaction and no functional effect on KV10.1. We propose these isoforms to work as modulators of PIST function via regulating the binding on interaction partners.

  3. Deafness and permanently reduced potassium channel gene expression and function in hypothyroid Pit1dw mutants

    Science.gov (United States)

    Mustapha, Mirna; Fang, Qing; Gong, Tzy-Wen; Dolan, David F.; Raphael, Yehoash; Camper, Sally A.; Duncan, R. Keith

    2012-01-01

    The absence of thyroid hormone (TH) during late gestation and early infancy can cause irreparable deafness in both humans and rodents. A variety of rodent models have been utilized in an effort to identify the underlying molecular mechanism. Here, we characterize a mouse model of secondary hypothyroidism, pituitary transcription factor 1 (Pit1dw), which has profound, congenital deafness that is rescued by oral TH replacement. These mutants have tectorial membrane abnormalities, including a prominent Hensen's stripe, elevated β-tectorin composition, and disrupted striated-sheet matrix. They lack distortion product otoacoustic emissions and cochlear microphonic responses, and exhibit reduced endocochlear potentials, suggesting defects in outer hair cell function and potassium recycling. Auditory system and hair cell physiology, histology and anatomy studies reveal novel defects of hormone deficiency related to deafness: (1) permanently impaired expression of KCNJ10 in the stria vascularis of Pit1dw mice, which likely contributes to the reduced endocochlear potential, (2) significant outer hair cell loss in the mutants, which may result from cellular stress induced by the lower KCNQ4 expression and current levels in Pit1dw mutant outer hair cells and (3) sensory and strial cell deterioration, which may have implications for thyroid hormone dysregulation in age related hearing impairment. In summary, we suggest that these defects in outer hair cell and strial cell function are important contributors to the hearing impairment in Pit1dw mice. PMID:19176829

  4. Biophysical characterization of KV3.1 potassium channel activating compounds

    DEFF Research Database (Denmark)

    Taskin, Bahar; von Schoubye, Nadia Lybøl; Sheykhzade, Majid

    2015-01-01

    The effect of two positive modulators, RE1 and EX15, on the voltage-gated K+ channel Kv3.1 was investigated using the whole-cell patch-clamp technique on HEK293 cells expressing Kv3.1a. RE1 and EX15 increased the Kv3.1 currents in a concentration-dependent manner with an EC50 value of 4.5 and 1.3...

  5. Interaction between the Cardiac Rapidly (IKr) and Slowly (IKs) Activating Delayed Rectifier Potassium Channels Revealed by Low K+-induced hERG Endocytic Degradation*

    Science.gov (United States)

    Guo, Jun; Wang, Tingzhong; Yang, Tonghua; Xu, Jianmin; Li, Wentao; Fridman, Michael D.; Fisher, John T.; Zhang, Shetuan

    2011-01-01

    Cardiac repolarization is controlled by the rapidly (IKr) and slowly (IKs) activating delayed rectifier potassium channels. The human ether-a-go-go-related gene (hERG) encodes IKr, whereas KCNQ1 and KCNE1 together encode IKs. Decreases in IKr or IKs cause long QT syndrome (LQTS), a cardiac disorder with a high risk of sudden death. A reduction in extracellular K+ concentration ([K+]o) induces LQTS and selectively causes endocytic degradation of mature hERG channels from the plasma membrane. In the present study, we investigated whether IKs compensates for the reduced IKr under low K+ conditions. Our data show that when hERG and KCNQ1 were expressed separately in human embryonic kidney (HEK) cells, exposure to 0 mm K+ for 6 h completely eliminated the mature hERG channel expression but had no effect on KCNQ1. When hERG and KCNQ1 were co-expressed, KCNQ1 significantly delayed 0 mm K+-induced hERG reduction. Also, hERG degradation led to a significant reduction in KCNQ1 in 0 mm K+ conditions. An interaction between hERG and KCNQ1 was identified in hERG+KCNQ1-expressing HEK cells. Furthermore, KCNQ1 preferentially co-immunoprecipitated with mature hERG channels that are localized in the plasma membrane. Biophysical and pharmacological analyses indicate that although hERG and KCNQ1 closely interact with each other, they form distinct hERG and KCNQ1 channels. These data extend our understanding of delayed rectifier potassium channel trafficking and regulation, as well as the pathology of LQTS. PMID:21844197

  6. THE OVEREXPRESSION OF GENE ENCODING RICE POTASSIUM CHANNEL - OsТРКa INCREASES THE SALT AND DROUGHT TOLERANCE OF PLANTS

    Directory of Open Access Journals (Sweden)

    S.V. Isayenkov

    2015-04-01

    Full Text Available The main aim of this study was to investigate the role of potassium TPK channel (OsTPKa from rice in regulation of salinity and drought stresses, potassium deficiency. In order to elevate the expression level of gene encoding two-pore potassium channel OsTPKa, the stable agrobacterium mediated transformation of plants was performed. The elevation of OsTPKa expression level in transformed plants leads to improvement of salt and drought tolerance of transformed plants was found during experiments. In conditions of potassium deficiency or salt stress the plants with elevated OsTPKa expression level exhibit better growth rates, decreasing of sodium accumulation in plant tissues.

  7. Potassium Channel Interacting Protein 2 (KChIP2) is not a transcriptional regulator of cardiac electrical remodeling

    DEFF Research Database (Denmark)

    Winther, Sine V; Tuomainen, Tomi; Borup, Rehannah

    2016-01-01

    The heart-failure relevant Potassium Channel Interacting Protein 2 (KChIP2) augments CaV1.2 and KV4.3. KChIP3 represses CaV1.2 transcription in cardiomyocytes via interaction with regulatory DNA elements. Hence, we tested nuclear presence of KChIP2 and if KChIP2 translocates into the nucleus...... in a Ca(2+) dependent manner. Cardiac biopsies from human heart-failure patients and healthy donor controls showed that nuclear KChIP2 abundance was significantly increased in heart failure; however, this was secondary to a large variation of total KChIP2 content. Administration of ouabain did...... not increase KChIP2 content in nuclear protein fractions in anesthetized mice. KChIP2 was expressed in cell lines, and Ca(2+) ionophores were applied in a concentration- and time-dependent manner. The cell lines had KChIP2-immunoreactive protein in the nucleus in the absence of treatments to modulate...

  8. Trafficking and intracellular regulation of Kv7.1 potassium channels in the heart

    DEFF Research Database (Denmark)

    Nielsen, Nathalie Hélix

    The electrical activity of the heart, measured by application of surface body electrodes and recorded as an electrocardiogram, is the result of a finely tuned balance of ion movement (K+, Na+, Ca2+). The ionic currents collectively constitute the cardiac action potential created in the cell....... The mutation is located in the C-terminus of the Kv7.1 channel protein in a calmodulin binding domain, where the methionine (M) at position 520 is replaced by an arginine (R). Our results show that although Kv7.1/calmodulin interaction is not impaired by the M520R mutation, the mutated channels are retained...

  9. Effects of calmodulin antagonists on calcium-activated potassium channels in pregnant rat myometrium.

    OpenAIRE

    Kihira, M.; Matsuzawa, K.; Tokuno, H.; Tomita, T.

    1990-01-01

    1. The effects of W-7, trifluoperazine, and W-5 on Ca2(+)-activated K(+)-channels were investigated with the inside-out patch-clamp method in smooth muscle cells freshly dispersed from pregnant rat myometrium. These drugs are known to have different potencies as calmodulin antagonists. 2. In the presence of 1 microM Ca2+ on the cytoplasmic side ([Ca2+]i), the fraction of time the channel was open (open probability, Po) was about 0.9 and the calmodulin antagonists (1-30 microM) applied to the ...

  10. Gain-of-function mutations in potassium channel subunit KCNE2 associated with early-onset lone atrial fibrillation

    DEFF Research Database (Denmark)

    Nielsen, Jonas Bille; Bentzen, Bo Hjorth; Olesen, Morten Salling

    2014-01-01

    Aims: Atrial fibrillation (AF) is the most common cardiac arrhythmia. Disturbances in cardiac potassium conductance are considered as one of the disease mechanisms in AF. We aimed to investigate if mutations in potassium-channel β-subunits KCNE2 and KCNE3 are associated with early-onset lone AF...... alleles). Electrophysiological investigations were performed for both mutations in combination with candidate pore-forming α-subunits KV7.1, KV11.1, KV4.3 and KV1.5. A significant gain-of-function effect was observed upon coexpression with KV7.1 and KV7.1 + KCNE1. Confocal imaging found no differences...... delayed rectifier potassium current might be involved in the pathogenesis of AF....

  11. Supratentorial white matter blurring associated with voltage-gated potassium channel-complex limbic encephalitis

    International Nuclear Information System (INIS)

    Urbach, H.; Mader, I.; Rauer, S.; Baumgartner, A.; Paus, S.; Wagner, J.; Malter, M.P.; Pruess, H.; Lewerenz, J.; Kassubek, J.; Hegen, H.; Auer, M.; Deisenhammer, F.; Ufer, F.; Bien, C.G.

    2015-01-01

    Limbic encephalitis (LE) associated with voltage-gated potassium channel-complex antibodies (VGKC-LE) is frequently non-paraneoplastic and associated with marked improvement following corticosteroid therapy. Mesial temporal lobe abnormalities are present in around 80 % of patients. If associated or preceded by faciobrachial dystonic seizures, basal ganglia signal changes may occur. In some patients, blurring of the supratentorial white matter on T2-weighted images (SWMB) may be seen. The purpose of this study was to evaluate the incidence of SWMB and whether it is specific for VGKC-LE. Two experienced neuroradiologists independently evaluated signal abnormalities on FLAIR MRI in 79 patients with LE while unaware on the antibody type. SWMB was independently assessed as present in 10 of 36 (28 %) compared to 2 (5 %) of 43 non-VGKC patients (p = 0.009). It was not related to the presence of LGI1 or CASPR2 proteins of VGKC antibodies. MRI showed increased temporomesial FLAIR signal in 22 (61 %) VGKC compared to 14 (33 %) non-VGKC patients (p = 0.013), and extratemporomesial structures were affected in one VGKC (3 %) compared to 11 (26 %) non-VGKC patients (p = 0.005). SWMB is a newly described MRI sign rather specific for VGKC-LE. (orig.)

  12. A novel 13 residue acyclic peptide from the marine snail, Conus monile, targets potassium channels.

    Science.gov (United States)

    Sudarslal, Sadasivannair; Singaravadivelan, Govindaswamy; Ramasamy, Palanisamy; Ananda, Kuppanna; Sarma, Siddhartha P; Sikdar, Sujit K; Krishnan, K S; Balaram, Padmanabhan

    2004-05-07

    A novel 13-residue peptide Mo1659 has been isolated from the venom of a vermivorous cone snail, Conus monile. HPLC fractions of the venom extract yielded an intense UV absorbing fraction with a mass of 1659Da. De novo sequencing using both matrix assisted laser desorption and ionization and electrospray MS/MS methods together with analysis of proteolytic fragments successfully yielded the amino acid sequence, FHGGSWYRFPWGY-NH(2). This was further confirmed by comparison with the chemically synthesized peptide and by conventional Edman sequencing. Mo1659 has an unusual sequence with a preponderance of aromatic residues and the absence of apolar, aliphatic residues like Ala, Val, Leu, and Ile. Mo1659 has no disulfide bridges distinguishing it from the conotoxins and bears no sequence similarity with any of the acyclic peptides isolated thus far from the venom of cone snails. Electrophysiological studies on the effect of Mo1659 on measured currents in dorsal root ganglion neurons suggest that the peptide targets non-inactivating voltage-dependent potassium channels.

  13. Supratentorial white matter blurring associated with voltage-gated potassium channel-complex limbic encephalitis

    Energy Technology Data Exchange (ETDEWEB)

    Urbach, H.; Mader, I. [University Medical Center Freiburg, Department of Neuroradiology, Freiburg (Germany); Rauer, S.; Baumgartner, A. [University Medical Center Freiburg, Department of Neurology, Freiburg (Germany); Paus, S. [University Medical Center, Department of Neurology, Bonn (Germany); Wagner, J. [University Medical Center, Department of Epileptology, Bonn (Germany); Malter, M.P. [University of Cologne, Department of Neurology, Cologne (Germany); Pruess, H. [Charite - Universitaetsmedizin Berlin, Department of Neurology, Berlin (Germany); Lewerenz, J.; Kassubek, J. [Ulm University, Department of Neurology, Ulm (Germany); Hegen, H.; Auer, M.; Deisenhammer, F. [University Innsbruck, Department of Neurology, Innsbruck (Austria); Ufer, F. [University Medical Center, Department of Neurology, Hamburg (Germany); Bien, C.G. [Epilepsy Centre Bethel, Bielefeld-Bethel (Germany)

    2015-12-15

    Limbic encephalitis (LE) associated with voltage-gated potassium channel-complex antibodies (VGKC-LE) is frequently non-paraneoplastic and associated with marked improvement following corticosteroid therapy. Mesial temporal lobe abnormalities are present in around 80 % of patients. If associated or preceded by faciobrachial dystonic seizures, basal ganglia signal changes may occur. In some patients, blurring of the supratentorial white matter on T2-weighted images (SWMB) may be seen. The purpose of this study was to evaluate the incidence of SWMB and whether it is specific for VGKC-LE. Two experienced neuroradiologists independently evaluated signal abnormalities on FLAIR MRI in 79 patients with LE while unaware on the antibody type. SWMB was independently assessed as present in 10 of 36 (28 %) compared to 2 (5 %) of 43 non-VGKC patients (p = 0.009). It was not related to the presence of LGI1 or CASPR2 proteins of VGKC antibodies. MRI showed increased temporomesial FLAIR signal in 22 (61 %) VGKC compared to 14 (33 %) non-VGKC patients (p = 0.013), and extratemporomesial structures were affected in one VGKC (3 %) compared to 11 (26 %) non-VGKC patients (p = 0.005). SWMB is a newly described MRI sign rather specific for VGKC-LE. (orig.)

  14. cAMP-dependent kinase does not modulate the Slack sodium-activated potassium channel.

    Science.gov (United States)

    Nuwer, Megan O; Picchione, Kelly E; Bhattacharjee, Arin

    2009-09-01

    The Slack gene encodes a Na(+)-activated K(+) channel and is expressed in many different types of neurons. Like the prokaryotic Ca(2+)-gated K(+) channel MthK, Slack contains two 'regulator of K(+) conductance' (RCK) domains within its carboxy terminal, domains likely involved in Na(+) binding and channel gating. It also contains multiple consensus protein kinase C (PKC) and protein kinase A (PKA) phosphorylation sites and although regulated by protein kinase C (PKC) phosphorylation, modulation by PKA has not been determined. To test if PKA directly regulates Slack, nystatin-perforated patch whole-cell currents were recorded from a human embryonic kidney (HEK-293) cell line stably expressing Slack. Bath application of forskolin, an adenylate cyclase activator, caused a rapid and complete inhibition of Slack currents however, the inactive homolog of forskolin, 1,9-dideoxyforskolin caused a similar effect. In contrast, bath application of 8-bromo-cAMP did not affect the amplitude nor the activation kinetics of Slack currents. In excised inside-out patch recordings, direct application of the PKA catalytic subunit to patches did not affect the open probability of Slack channels nor was open probability affected by direct application of protein phosphatase 2B. Preincubation of cells with the protein kinase A inhibitor KT5720 also did not change current density. Finally, mutating the consensus phosphorylation site located between RCK domain 1 and domain 2 from serine to glutamate did not affect current activation kinetics. We conclude that unlike PKC, phosphorylation by PKA does not acutely modulate the function and gating activation kinetics of Slack channels.

  15. Inwardly rectifying potassium channels (Kir) in central nervous system glia: a special role for Kir4.1 in glial functions.

    Science.gov (United States)

    Butt, Arthur M; Kalsi, Amanpreet

    2006-01-01

    Glia in the central nervous system (CNS) express diverse inward rectifying potassium channels (Kir). The major function of Kir is in establishing the high potassium (K+) selectivity of the glial cell membrane and strongly negative resting membrane potential (RMP), which are characteristic physiological properties of glia. The classical property of Kir is that K+ flows inwards when the RMP is negative to the equilibrium potential for K+ (E(K)), but at more positive potentials outward currents are inhibited. This provides the driving force for glial uptake of K+ released during neuronal activity, by the processes of "K+ spatial buffering" and "K+ siphoning", considered a key function of astrocytes, the main glial cell type in the CNS. Glia express multiple Kir channel subtypes, which are likely to have distinct functional roles related to their differences in conductance, and sensitivity to intracellular and extracellular factors, including pH, ATP, G-proteins, neurotransmitters and hormones. A feature of CNS glia is their specific expression of the Kir4.1 subtype, which is a major K+ conductance in glial cell membranes and has a key role in setting the glial RMP. It is proposed that Kir4.1 have a primary function in K+ regulation, both as homomeric channels and as heteromeric channels by co-assembly with Kir5.1 and probably Kir2.0 subtypes. Significantly, Kir4.1 are also expressed by oligodendrocytes, the myelin-forming cells of the CNS, and the genetic ablation of Kir4.1 results in severe hypomyelination. Hence, Kir, and in particular Kir4.1, are key regulators of glial functions, which in turn determine neuronal excitability and axonal conduction.

  16. Basolateral potassium channel in turtle colon. Evidence for single-file ion flow

    OpenAIRE

    1983-01-01

    Treatment of the apical surface of the isolated, ouabain-inhibited turtle colon with the polyene antibiotic amphotericin B permitted the properties of a barium-sensitive potassium conductance in the basolateral membrane to be discerned from the measurements of transepithelial fluxes and electrical currents. Simultaneous measurements of potassium currents and 42K fluxes showed that the movement of potassium was not in accord with simple diffusion. Two other cations, thallium and rubidium, were...

  17. Immunomodulatory effects of diclofenac in leukocytes through the targeting of Kv1.3 voltage-dependent potassium channels.

    Science.gov (United States)

    Villalonga, Núria; David, Miren; Bielańska, Joanna; González, Teresa; Parra, David; Soler, Concepció; Comes, Núria; Valenzuela, Carmen; Felipe, Antonio

    2010-09-15

    Kv1.3 plays a crucial role in the activation and proliferation of T-lymphocytes and macrophages. While Kv1.3 is responsible for the voltage-dependent potassium current in T-cells, in macrophages this K(+) current is generated by the association of Kv1.3 and Kv1.5. Patients with autoimmune diseases show a high number of effector memory T cells that are characterized by a high expression of Kv1.3 and Kv1.3 antagonists ameliorate autoimmune disorders in vivo. Diclofenac is a non-steroidal anti-inflammatory drug (NSAID) used in patients who suffer from painful autoimmune diseases such as rheumatoid arthritis. In this study, we show that diclofenac impairs immune response via a mechanism that involves Kv1.3. While diclofenac inhibited Kv1.3 expression in activated macrophages and T-lymphocytes, Kv1.5 remained unaffected. Diclofenac also decreased iNOS levels in Raw 264.7 cells, impairing their activation in response to lipopolysaccharide (LPS). LPS-induced macrophage migration and IL-2 production in stimulated Jurkat T-cells were also blocked by pharmacological doses of diclofenac. These effects were mimicked by Margatoxin, a specific Kv1.3 inhibitor, and Charybdotoxin, which blocks both Kv1.3 and Ca(2+)-activated K(+) channels (K(Ca)3.1). Because Kv1.3 is a very good target for autoimmune therapies, the effects of diclofenac on Kv1.3 are of high pharmacological relevance. Copyright 2010 Elsevier Inc. All rights reserved.

  18. Remote and reversible inhibition of neurons and circuits by small molecule induced potassium channel stabilization

    OpenAIRE

    Auffenberg, Eva; Jurik, Angela; Mattusch, Corinna; Stoffel, Rainer; Genewsky, Andreas; Namendorf, Christian; Schmid, Roland M.; Rammes, Gerhard; Biel, Martin; Uhr, Manfred; Moosmang, Sven; Michalakis, Stylianos; Wotjak, Carsten T.; Thoeringer, Christoph K.

    2016-01-01

    Manipulating the function of neurons and circuits that translate electrical and chemical signals into behavior represents a major challenges in neuroscience. In addition to optogenetic methods using light-activatable channels, pharmacogenetic methods with ligand induced modulation of cell signaling and excitability have been developed. However, they are largely based on ectopic expression of exogenous or chimera proteins. Now, we describe the remote and reversible expression of a Kir2.1 type ...

  19. Effects of Divalent Cations on Outward Potassium Currents in Leech Retzius Nerve Cells

    Directory of Open Access Journals (Sweden)

    Jovanovic Zorica

    2016-12-01

    Full Text Available The present study examines the effects of divalent metals, cadmium (Cd2+ and manganese (Mn2+, on the outward potassium currents of Retzius cells in the hirudinid leeches Haemopis sanguisuga using conventional two-microelectrode voltageclamp techniques. The outward potassium current is activated by depolarization and plays an important role in determining both the neuronal excitability and action potential duration. A strong inhibition of the fast current and a clear reduction in the late currents of the outward current with 1 mM Cd2+ were obtained, which indicated that both components are sensitive to this metal. Complete blockage of the fast and partial reduction of the slow outward currents was observed after adding 1 mM Mn2+ to the extracellular fluid. These data show that the outward K+ current in leech Retzius nerve cells comprises at least two components: a voltage-dependent K+ current and a Ca2+- activated K+ current. These observations also indicate that Cd2+ is more eff ective than Mn2+ in blocking ion fl ow through these channels and that suppressing Ca2+-activated K+ outward currents can prolong the action potential in nerve cells.

  20. Involvement of BKCa and KV potassium channels in cAMP-induced vasodilation: their insufficient function in genetic hypertension

    Czech Academy of Sciences Publication Activity Database

    Pintérová, Mária; Behuliak, Michal; Kuneš, Jaroslav; Zicha, Josef

    2014-01-01

    Roč. 63, č. 3 (2014), s. 275-285 ISSN 0862-8408 R&D Projects: GA ČR(CZ) GA305/09/0336; GA ČR(CZ) GAP304/12/0259; GA MŠk(CZ) 1M0510 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : isoprenaline * cAMP * potassium channels * calcium channels Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 1.293, year: 2014

  1. The role of K+ channels in uptake and redistribution of potassium in the model plant Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Tripti eSharma

    2013-06-01

    Full Text Available Potassium (K+ is inevitable for plant growth and development. It plays a crucial role in the regulation of enzyme activities, in adjusting the electrical membrane potential and the cellular turgor, in regulating cellular homeostasis and in the stabilization of protein synthesis. Uptake of K+ from the soil and its transport to growing organs is essential for a healthy plant development. Uptake and allocation of K+ are performed by K+ channels and transporters belonging to different protein families. In this review, we summarize the knowledge on the versatile physiological roles of plant K+ channels and their behaviour under stress conditions in the model plant Arabidopsis thaliana.

  2. The activation of mitochondrial BK potassium channels contributes to the protective effects of naringenin against myocardial ischemia/reperfusion injury.

    Science.gov (United States)

    Testai, L; Martelli, A; Marino, A; D'Antongiovanni, V; Ciregia, F; Giusti, L; Lucacchini, A; Chericoni, S; Breschi, M C; Calderone, V

    2013-06-01

    Naringenin (NAR), flavonoid abundant in the genus Citrus, has been reported to interact with the large-conductance calcium-activated potassium channels (BK). Since activators of BK channels expressed in cardiac mitochondria trigger protective effects in several models of myocardial ischemia/reperfusion (I/R), this work aimed to evaluate the potential cardioprotective effects of NAR and the involvement of mitochondrial BK channels. In an in vivo model of acute infarct in rats, NAR (100mg/kg i.p.) significantly reduced the heart injury induced by I/R. This effect was antagonized by the selective BK-blocker paxilline (PAX). The cardioprotective dose of NAR did not cause significant effects on the blood pressure. In Largendorff-perfused rat hearts submitted to ischemia/reperfusion, NAR improved the post-ischemic functional parameters (left ventricle developed pressure and dP/dt) with lower extension of myocardial injury. On isolated rat cardiac mitochondria, NAR caused a concentration-dependent depolarization of mitochondrial membrane and caused a trans-membrane flow of thallium (potassium-mimetic cation). Both these effects were antagonized by selective blockers of BK channels. Furthermore, NAR half-reduced the calcium accumulation into the matrix of cardiac mitochondria exposed to high calcium concentrations. In conclusion, NAR exerts anti-ischemic effects through a "pharmacological preconditioning" that it is likely to be mediated, at least in part, by the activation of mitochondrial BK channels. Copyright © 2013. Published by Elsevier Inc.

  3. [Co-location of ACh-sensitive BK channels and L-type calcium channels in type II vestibular hair cells of guinea pig].

    Science.gov (United States)

    Guo, Chang-Kai; Li, Guan-Qiao; Kong, Wei-Jia; Zhang, Song; Wu, Ting-Ting; Li, Jia-Li; Li, Qing-Tian

    2008-03-01

    To explore the mechanisms of the influx of calcium ions during the activation of ACh-sensitive BK channel (big conductance, calcium-dependent potassium channel) in type II vestibular hair cells of guinea pigs. Type II vestibular hair cells were isolated by collagenase type IA. Under the whole-cell patch mode, the sensitivity of ACh-sensitive BK current to the calcium channels blockers was investigated, the pharmacological property of L-type calcium channel activator-sensitive current and ACh-sensitive BK current was compared. Following application of ACh, type II vestibular hair cells displayed a sustained outward potassium current, with a reversal potential of (-70.5 +/- 10.6) mV (x +/- s, n = 10). At the holding potential of -50 mV, the current amplitude of ACh-sensitive potassium current activated by 100 micromol/L ACh was (267 +/- 106) pA (n = 11). ACh-sensitive potassium current was potently sensitive to the BK current blocker, IBTX (iberiotoxin, 200 nmol/L). Apamin, the well-known small conductance, calcium-dependent potassium current blocker, failed to inhibit the amplitude of ACh-sensitive potassium current at a dose of 1 micromol/L. ACh-sensitive BK current was sensitive to NiCl2 and potently inhibited by CdCl2. NiCl2 and CdCl2 showed a dose-dependent blocking effect with a half inhibition-maximal response of (135.5 +/- 18.5) micromol/L (n = 7) and (23.4 +/- 2.6) micromol/L (n = 7). The L-type calcium channel activator, (-) -Bay-K 8644 (10 micromol /L), mimicked the role of ACh and activated the IBTX-sensitive outward current. ACh-sensitive BK and L-type calcium channels are co-located in type II vestibular hair cells of guinea pigs.

  4. The Voltage-Gated Potassium Channel Subfamily KQT Member 4 (KCNQ4) Displays Parallel Evolution in Echolocating Bats

    Science.gov (United States)

    Liu, Yang; Han, Naijian; Franchini, Lucía F.; Xu, Huihui; Pisciottano, Francisco; Elgoyhen, Ana Belén; Rajan, Koilmani Emmanuvel; Zhang, Shuyi

    2012-01-01

    Bats are the only mammals that use highly developed laryngeal echolocation, a sensory mechanism based on the ability to emit laryngeal sounds and interpret the returning echoes to identify objects. Although this capability allows bats to orientate and hunt in complete darkness, endowing them with great survival advantages, the genetic bases underlying the evolution of bat echolocation are still largely unknown. Echolocation requires high-frequency hearing that in mammals is largely dependent on somatic electromotility of outer hair cells. Then, understanding the molecular evolution of outer hair cell genes might help to unravel the evolutionary history of echolocation. In this work, we analyzed the molecular evolution of two key outer hair cell genes: the voltage-gated potassium channel gene KCNQ4 and CHRNA10, the gene encoding the α10 nicotinic acetylcholine receptor subunit. We reconstructed the phylogeny of bats based on KCNQ4 and CHRNA10 protein and nucleotide sequences. A phylogenetic tree built using KCNQ4 amino acid sequences showed that two paraphyletic clades of laryngeal echolocating bats grouped together, with eight shared substitutions among particular lineages. In addition, our analyses indicated that two of these parallel substitutions, M388I and P406S, were probably fixed under positive selection and could have had a strong functional impact on KCNQ4. Moreover, our results indicated that KCNQ4 evolved under positive selection in the ancestral lineage leading to mammals, suggesting that this gene might have been important for the evolution of mammalian hearing. On the other hand, we found that CHRNA10, a gene that evolved adaptively in the mammalian lineage, was under strong purifying selection in bats. Thus, the CHRNA10 amino acid tree did not show echolocating bat monophyly and reproduced the bat species tree. These results suggest that only a subset of hearing genes could underlie the evolution of echolocation. The present work continues to

  5. The voltage-gated potassium channel subfamily KQT member 4 (KCNQ4) displays parallel evolution in echolocating bats.

    Science.gov (United States)

    Liu, Yang; Han, Naijian; Franchini, Lucía F; Xu, Huihui; Pisciottano, Francisco; Elgoyhen, Ana Belén; Rajan, Koilmani Emmanuvel; Zhang, Shuyi

    2012-05-01

    Bats are the only mammals that use highly developed laryngeal echolocation, a sensory mechanism based on the ability to emit laryngeal sounds and interpret the returning echoes to identify objects. Although this capability allows bats to orientate and hunt in complete darkness, endowing them with great survival advantages, the genetic bases underlying the evolution of bat echolocation are still largely unknown. Echolocation requires high-frequency hearing that in mammals is largely dependent on somatic electromotility of outer hair cells. Then, understanding the molecular evolution of outer hair cell genes might help to unravel the evolutionary history of echolocation. In this work, we analyzed the molecular evolution of two key outer hair cell genes: the voltage-gated potassium channel gene KCNQ4 and CHRNA10, the gene encoding the α10 nicotinic acetylcholine receptor subunit. We reconstructed the phylogeny of bats based on KCNQ4 and CHRNA10 protein and nucleotide sequences. A phylogenetic tree built using KCNQ4 amino acid sequences showed that two paraphyletic clades of laryngeal echolocating bats grouped together, with eight shared substitutions among particular lineages. In addition, our analyses indicated that two of these parallel substitutions, M388I and P406S, were probably fixed under positive selection and could have had a strong functional impact on KCNQ4. Moreover, our results indicated that KCNQ4 evolved under positive selection in the ancestral lineage leading to mammals, suggesting that this gene might have been important for the evolution of mammalian hearing. On the other hand, we found that CHRNA10, a gene that evolved adaptively in the mammalian lineage, was under strong purifying selection in bats. Thus, the CHRNA10 amino acid tree did not show echolocating bat monophyly and reproduced the bat species tree. These results suggest that only a subset of hearing genes could underlie the evolution of echolocation. The present work continues to

  6. Apigenin-induced nitric oxide production involves calcium-activated potassium channels and is responsible for antiangiogenic effects.

    Science.gov (United States)

    Erdogan, A; Most, A K; Wienecke, B; Fehsecke, A; Leckband, C; Voss, R; Grebe, M T; Tillmanns, H; Schaefer, C A; Kuhlmann, C R W

    2007-08-01

    The dietary flavonoid apigenin (Api) has been demonstrated to exert multiple beneficial effects upon the vascular endothelium. The aim of this study was to examine whether Ca(2+)-activated K(+) channels (K(Ca)) are involved in endothelial nitric oxide (NO) production and antiangiogenic effects. Endothelial NO generation was monitored using a cyclic guanosine monophosphate radioimmunoassay. K(Ca) activity and changes of the intracellular Ca(2+) concentration [Ca(2+)](i) were analyzed using the fluorescent dyes bis-barbituric acid oxonol, potassium-binding benzofuran isophthalate, and fluo-3. The endothelial angiogenic parameters measured were cell proliferation, [(3)H]-thymidine incorporation, and cell migration (scratch assay). Akt phosphorylation was examined using immunohistochemistry. Api caused a concentration-dependent increase in cyclic guanosine monophosphate levels, with a maximum effect at a concentration of 1 mum. Api-induced hyperpolarization was blocked by the small and large conductance K(Ca) inhibitors apamin and iberiotoxin, respectively. Furthermore, apamin and iberiotoxin blocked the late, long-lasting plateau phase of the Api-induced biphasic increase of [Ca(2+)](i). Inhibition of Ca(2+) signaling and the K(Ca) blockade both blocked NO production. Prevention of all three (NO, Ca(2+), and K(Ca) signaling) reversed the antiangiogenic effects of Api under both basal and basic fibroblast growth factor-induced culture conditions. Basic fibroblast growth factor-induced Akt phosphorylation was also reduced by Api. Based on our experimental results we propose the following signaling cascade for the effects of Api on endothelial cell signaling. Api activates small and large conductance K(Ca), leading to a hyperpolarization that is followed by a Ca(2+) influx. The increase of [Ca(2+)](i) is responsible for an increased NO production that mediates the antiangiogenic effects of Api via Akt dephosphorylation.

  7. Vascular ATP-sensitive potassium channels are over-expressed and partially regulated by nitric oxide in experimental septic shock.

    Science.gov (United States)

    Collin, Solène; Sennoun, Nacira; Dron, Anne-Gaëlle; de la Bourdonnaye, Mathilde; Montemont, Chantal; Asfar, Pierre; Lacolley, Patrick; Meziani, Ferhat; Levy, Bruno

    2011-05-01

    To study the activation and expression of vascular (aorta and small mesenteric arteries) potassium channels during septic shock with or without modulation of the NO pathway. Septic shock was induced in rats by peritonitis. Selective inhibitors of vascular K(ATP) (PNU-37883A) or BK(Ca) [iberiotoxin (IbTX)] channels were used to demonstrate their involvement in vascular hyporeactivity. Vascular response to phenylephrine was measured on aorta and small mesenteric arteries mounted on a wire myograph. Vascular expression of potassium channels was studied by PCR and Western blot, in the presence or absence of 1400W, an inducible NO synthase (iNOS) inhibitor. Aortic activation of the transcriptional factor nuclear factor-kappaB (NF-κB) was assessed by electrophoretic mobility shift assay. Arterial pressure as well as in vivo and ex vivo vascular reactivity were reduced by sepsis and improved by PNU-37883A but not by IbTX. Sepsis was associated with an up-regulation of mRNA and protein expression of vascular K(ATP) channels, while expression of vascular BK(Ca) channels remained unchanged. Selective iNOS inhibition blunted the sepsis-induced increase in aortic NO, decreased NF-κB activation, and down-regulated vascular K(ATP) channel expression. Vascular K(ATP) but not BK(Ca) channels are activated, over-expressed, and partially regulated by NO via NF-κB activation during septic shock. Their selective inhibition restores arterial pressure and vascular reactivity and decreases lactate concentration. The present data suggest that selective vascular K(ATP) channel inhibitors offer potential therapeutic perspectives for septic shock.

  8. MiRNA-135a regulates the expression of small conductance calcium-activated potassium (SK3) channels in epilepsy-like conditions

    NARCIS (Netherlands)

    Honrath, Birgit; Norwood, Braxton; Tanrioever, Gaye; Kuter, Katarzyna; Henshall, David C; Aksel-Aksoy, Ayla; Schratt, Gerhard; Pasterkamp, Jeroen; Dencher, Norbert A.; Nieweg, Katja; Culmsee, Carsten; Dolga, Amalia Mihalea

    2017-01-01

    Background Excessive and hypersynchronous neuronal discharges are key characteristics in the pathophysiology of neurological disorders such as epilepsy. Owing to their ability of regulating neuronal excitability, small conductance calcium-activated potassium (SK) channels have been implicated in

  9. SENSITIVE EFFECTS OF POTASSIUM AND CALCIUM CHANNEL BLOCKING AND ATP-SENSITIVE POTASSIUM CHANNEL ACTIVATORS ON SEMINAL VESICLE SMOOTH MUSCLE CONTRACTIONS

    Directory of Open Access Journals (Sweden)

    H SADRAEI

    2000-12-01

    Full Text Available Background. Seminal vesicle smooth muscle contraction is mediated through sympathetic and parasympathetic neurons activity. Although seminal vesicle plays an important role in male fertility, but little attention is given to mechanism involved in contraction of this organ.
    Methods. In this study effects of drugs which activate ATP - sensitive K channels and blockers of K and Ca channels were examined on contraction of guinea - pig isolated seminal vesicle due to electrical filled stimulation (EFS, noradrenaline, carbachol and KCI.
    Results. The K channel blocker tetraethyl ammonium potentate the EFS responses at all frequencies, while, the ATP - sensitive K channel inhibitor glibenclamide and the K channel opener levcromakalim, diazoxide, minoxidil and Ca channel blocker nifedipine all had relaxant effect on guinea - pig seminal vesicle.
    Discussion. This study indicate that activities of K and Ca channels is important in regulation of seminal vesicle contraction due to nerve stimulation, noradrenaline or carbachol.

  10. Effects of potassium on kesterite solar cells: Similarities, differences and synergies with sodium

    Directory of Open Access Journals (Sweden)

    S. G. Haass

    2018-01-01

    Full Text Available Addition of alkali dopants is essential for achieving high-efficiency conversion efficiency of thin film solar cells based on chalcogenide semiconductors like Cu(In,GaSe2 (CIGS and Cu2ZnSn(S,Se4 (CZTSSe also called kesterite. Whereas the treatment with potassium allows boosting the performance of CIGS solar cells as compared to the conventional sodium doping, it is debated if similar effects can be expected for kesterite solar cells. Here the influence of potassium is investigated by introducing the dopant during the solution processing of kesterite absorbers. It is confirmed that the presence of potassium leads to an enhanced grain growth and a ten-fold lower potassium concentration is sufficient for obtaining grain size similar to sodium-containing absorbers. Potassium is located predominantly at grain boundaries and it suppresses incorporation of sodium into the absorber layer. The potassium doping increases the apparent carrier concentration to ∼2×1016 cm-3 for a potassium concentration of 0.2 at%. The potassium-doped solar cells yield conversion efficiency close to 10%, on par with only sodium-doped samples. Co-doping with potassium and sodium has not revealed any beneficial synergetic effects and it is concluded that both dopants exhibit similar effects on the kesterite solar cell performance.

  11. Effect of antiarrhythmic drugs on small conductance calcium –activated potassium channels

    DEFF Research Database (Denmark)

    Simo Vicens, Rafel; Sauter, Daniel Rafael Peter; Grunnet, Morten

    2017-01-01

    Atrial fibrillation (AF) is the most common type of arrhythmia. Current pharmacological treatment for AF is moderately effective and/or increases the risk of serious ventricular adverse effects. To avoid ventricular adverse effects, a new target has been considered, the small conductance calcium....... Whether antiarrhythmic drugs (AADs) recommended for treating AF target KCa2.X channels is unknown. To this end, we tested a large number of AADs on the human KCa2.2 and KCa2.3 channels to assess their effect on this new target using automated whole-cell patch clamp. Of the AADs recommended for treatment...... for their antiarrhythmic effect is unlikely, as the calculated IC50 values are very high compared to the effective free therapeutic plasma concentration of the drugs when used for AF treatment, 40,000-fold for dofetilide and 140- fold higher for propafenone....

  12. G protein-coupled receptors form stable complexes with inwardly rectifying potassium channels and adenylyl cyclase.

    Science.gov (United States)

    Lavine, Natalie; Ethier, Nathalie; Oak, James N; Pei, Lin; Liu, Fang; Trieu, Phan; Rebois, R Victor; Bouvier, Michel; Hebert, Terence E; Van Tol, Hubert H M

    2002-11-29

    A large number of studies have demonstrated co-purification or co-immunoprecipitation of receptors with G proteins. We have begun to look for the presence of effector molecules in these receptor complexes. Co-expression of different channel and receptor permutations in COS-7 and HEK 293 cells in combination with co-immunoprecipitation experiments established that the dopamine D(2) and D(4), and beta(2)-adrenergic receptors (beta(2)-AR) form stable complexes with Kir3 channels. The D(4)/Kir3 and D(2) receptor/Kir3 interaction does not occur when the channel and receptor are expressed separately and mixed prior to immunoprecipitation, indicating that the interaction is not an artifact of the experimental protocol and reflects a biosynthetic event. The observed complexes are stable in that they are not disrupted by receptor activation or modulation of G protein alpha subunit function. However, using a peptide that binds Gbetagamma (betaARKct), we show that Gbetagamma is critical for dopamine receptor-Kir3 complex formation, but not for maintenance of the complex. We also provide evidence that Kir3 channels and another effector, adenylyl cyclase, are stably associated with the beta(2)-adrenergic receptor and can be co-immunoprecipitated by anti-receptor antibodies. Using bioluminescence resonance energy transfer, we have shown that in living cells under physiological conditions, beta(2)AR interacts directly with Kir3.1/3.4 and Kir3.1/3.2c heterotetramers as well as with adenylyl cyclase. All of these interactions are stable in the presence of receptor agonists, suggesting that these signaling complexes persist during signal transduction. In addition, we provide evidence that the receptor-effector complexes are also found in vivo. The observation that several G protein-coupled receptors form stable complexes with their effectors suggests that this arrangement might be a general feature of G protein-coupled signal transduction.

  13. Selection of Inhibitor-Resistant Viral Potassium Channels Identifies a Selectivity Filter Site that Affects Barium and Amantadine Block

    Science.gov (United States)

    Fujiwara, Yuichiro; Arrigoni, Cristina; Domigan, Courtney; Ferrara, Giuseppina; Pantoja, Carlos; Thiel, Gerhard; Moroni, Anna; Minor, Daniel L.

    2009-01-01

    Background Understanding the interactions between ion channels and blockers remains an important goal that has implications for delineating the basic mechanisms of ion channel function and for the discovery and development of ion channel directed drugs. Methodology/Principal Findings We used genetic selection methods to probe the interaction of two ion channel blockers, barium and amantadine, with the miniature viral potassium channel Kcv. Selection for Kcv mutants that were resistant to either blocker identified a mutant bearing multiple changes that was resistant to both. Implementation of a PCR shuffling and backcrossing procedure uncovered that the blocker resistance could be attributed to a single change, T63S, at a position that is likely to form the binding site for the inner ion in the selectivity filter (site 4). A combination of electrophysiological and biochemical assays revealed a distinct difference in the ability of the mutant channel to interact with the blockers. Studies of the analogous mutation in the mammalian inward rectifier Kir2.1 show that the T→S mutation affects barium block as well as the stability of the conductive state. Comparison of the effects of similar barium resistant mutations in Kcv and Kir2.1 shows that neighboring amino acids in the Kcv selectivity filter affect blocker binding. Conclusions/Significance The data support the idea that permeant ions have an integral role in stabilizing potassium channel structure, suggest that both barium and amantadine act at a similar site, and demonstrate how genetic selections can be used to map blocker binding sites and reveal mechanistic features. PMID:19834614

  14. Inhibition of Inwardly Rectifying Potassium (Kir 4.1 Channels Facilitates Brain-Derived Neurotrophic Factor (BDNF Expression in Astrocytes

    Directory of Open Access Journals (Sweden)

    Masato Kinboshi

    2017-12-01

    Full Text Available Inwardly rectifying potassium (Kir 4.1 channels in astrocytes regulate neuronal excitability by mediating spatial potassium buffering. Although dysfunction of astrocytic Kir4.1 channels is implicated in the development of epileptic seizures, the functional mechanisms of Kir4.1 channels in modulating epileptogenesis remain unknown. We herein evaluated the effects of Kir4.1 inhibition (blockade and knockdown on expression of brain-derived neurotrophic factor (BDNF, a key modulator of epileptogenesis, in the primary cultures of mouse astrocytes. For blockade of Kir4.1 channels, we tested several antidepressant agents which reportedly bound to and blocked Kir4.1 channels in a subunit-specific manner. Treatment of astrocytes with fluoxetine enhanced BDNF mRNA expression in a concentration-dependent manner and increased the BDNF protein level. Other antidepressants (e.g., sertraline and imipramine also increased the expression of BDNF mRNA with relative potencies similar to those for inhibition of Kir4.1 channels. In addition, suppression of Kir4.1 expression by the transfection of small interfering RNA (siRNA targeting Kir4.1 significantly increased the mRNA and protein levels of BDNF. The BDNF induction by Kir4.1 siRNA transfection was suppressed by the MEK1/2 inhibitor U0126, but not by the p38 MAPK inhibitor SB202190 or the JNK inhibitor SP600125. The present results demonstrated that inhibition of Kir4.1 channels facilitates BDNF expression in astrocytes primarily by activating the Ras/Raf/MEK/ERK pathway, which may be linked to the development of epilepsy and other neuropsychiatric disorders.

  15. Inhibitory effects of telmisartan on culture and proliferation of and Kv1.3 potassium channel expression in peripheral blood CD4+ T lymphocytes from Xinjiang Kazakh patients with hypertension

    Directory of Open Access Journals (Sweden)

    Sha-Sha Huang

    2016-10-01

    Full Text Available Introduction: Activation of T lymphocytes, for which potassium channels are essential, is involved in the development of hypertension. In this study, we explored the inhibitory effects of telmisartan on the culture and proliferation of and Kv1.3 potassium channel expression in peripheral blood CD4+ T lymphocytes derived from Xinjiang Kazakh patients with hypertension. Methods: CD4+ T-cell samples from hypertensive Kazakh patients and healthy Kazakh people were divided into healthy control, case control, telmisartan, and 4-aminopytidine groups. Changes in the expression levels of interleukin (IL-6 and IL-17 in the blood of the healthy control and case control subjects were detected by enzyme-linked immunosorbent assay. Peripheral blood CD4+ T lymphocytes were first activated and proliferated in vitro and then incubated for 0, 24, and 48 h under various treatment conditions. Thereafter, changes in CD4+ T-lymphocytic proliferation were determined using Cell Counting Kit-8 and microscope photography. Changes in messenger RNA (mRNA and protein expression of the Kv1.3 potassium channel in CD4+ T lymphocytes were detected using real-time quantitative polymerase chain reaction and Western blots, respectively. Results: The IL-6 and IL-17 expression levels were significantly higher in the blood of the hypertensive Kazakh patients than in the healthy Kazakh people. Telmisartan inhibited T-lymphocytic proliferation, as well as the mRNA and protein expression of the Kv1.3 potassium channel in CD4+ T lymphocytes, and the inhibitory effects were time-dependent, with the strongest inhibition observed after 48 h and significantly weaker inhibition observed after 24 h of treatment. Conclusions: Telmisartan may potentially regulate hypertensive inflammatory responses by inhibiting T-lymphocytic proliferation and Kv1.3 potassium channel expression in CD4+ T lymphocytes.

  16. POTASSIUM-DEPENDENT SODIUM EXTRUSION BY CELLS OF PORPHYRA PERFORATA, A RED MARINE ALGA

    Science.gov (United States)

    Eppley, Richard W.

    1958-01-01

    Potassium-free artificial sea water causes a loss of cell potassium and a gain of cell sodium in Porphyra perforata, which is not attributable to an inhibition of respiration. On adding KCl or RbCl to such low potassium, high sodium tissues, net accumulation of potassium or rubidium takes place, accompanied by net extrusion of sodium. Rates of potassium or rubidium accumulation and sodium extrusion are proportional to the amount of KCl or RbCl added only at low concentrations. Saturation of rates is evident at KCl or RbCl concentrations above 20–30 mM, suggesting the role of an ion carrier mechanism of transport. Evidence for and against mutually dependent sodium extrusion and potassium or rubidium accumulation is discussed. PMID:13587912

  17. Induction of stable ER-plasma-membrane junctions by Kv2.1 potassium channels.

    Science.gov (United States)

    Fox, Philip D; Haberkorn, Christopher J; Akin, Elizabeth J; Seel, Peter J; Krapf, Diego; Tamkun, Michael M

    2015-06-01

    Junctions between cortical endoplasmic reticulum (cER) and the plasma membrane are a subtle but ubiquitous feature in mammalian cells; however, very little is known about the functions and molecular interactions that are associated with neuronal ER-plasma-membrane junctions. Here, we report that Kv2.1 (also known as KCNB1), the primary delayed-rectifier K(+) channel in the mammalian brain, induces the formation of ER-plasma-membrane junctions. Kv2.1 localizes to dense, cell-surface clusters that contain non-conducting channels, indicating that they have a function that is unrelated to membrane-potential regulation. Accordingly, Kv2.1 clusters function as membrane-trafficking hubs, providing platforms for delivery and retrieval of multiple membrane proteins. Using both total internal reflection fluorescence and electron microscopy we demonstrate that the clustered Kv2.1 plays a direct structural role in the induction of stable ER-plasma-membrane junctions in both transfected HEK 293 cells and cultured hippocampal neurons. Glutamate exposure results in a loss of Kv2.1 clusters in neurons and subsequent retraction of the cER from the plasma membrane. We propose Kv2.1-induced ER-plasma-membrane junctions represent a new macromolecular plasma-membrane complex that is sensitive to excitotoxic insult and functions as a scaffolding site for both membrane trafficking and Ca(2+) signaling. © 2015. Published by The Company of Biologists Ltd.

  18. Renal fibrosis is attenuated by targeted disruption of KCa3.1 potassium channels

    DEFF Research Database (Denmark)

    Grgic, Ivica; Kiss, Eva; Kaistha, Brajesh P

    2009-01-01

    .1 blocker TRAM-34 similarly attenuated progression of UUO-induced renal fibrosis in wild-type mice and rats. In conclusion, our data demonstrate that K(Ca)3.1 is involved in renal fibroblast proliferation and fibrogenesis and suggest that K(Ca)3.1 may represent a therapeutic target for the treatment......Proliferation of interstitial fibroblasts is a hallmark of progressive renal fibrosis commonly resulting in chronic kidney failure. The intermediate-conductance Ca(2+)-activated K(+) channel (K(Ca)3.1) has been proposed to promote mitogenesis in several cell types and contribute to disease states...... of fibrotic kidney disease....

  19. Modification of sodium and potassium channel kinetics by diethyl ether and studies on sodium channel inactivation in the crayfish giant axon membrane

    Energy Technology Data Exchange (ETDEWEB)

    Bean, Bruce Palmer [Univ. of Rochester, NY (United States)

    1979-01-01

    The effects of ether and halothane on membrane currents in the voltage clamped crayfish giant axon membrane were investigated. Concentrations of ether up to 300 mM and of halothane up to 32 mM had no effect on resting potential or leakage conductance. Ether and halothane reduced the size of sodium currents without changing the voltage dependence of the peak currents or their reversal potential. Ether and halothane also produced a reversible, dose-dependent speeding of sodium current decay at all membrane potentials. Ether reduced the time constants for inactivation, and also shifted the midpoint of the steady-state inactivation curve in the hyperpolarizing direction. Potassium currents were smaller with ether present, with no change in the voltage dependence of steady-state currents. The activation of potassium channels was faster with ether present. There was no apparent change in the capacitance of the crayfish giant axon membrane with ether concentrations of up to 100 mM. Experiments on sodium channel inactivation kinetics were performed using 4-aminopyridine to block potassium currents. Sodium currents decayed with a time course generally fit well by a single exponential. The time constant of decay was a steep function of voltage, especially in the negative resistance region of the peak current vs voltage relation.The time course of inactivation was very similar to that of the decay of the current at the same potential. The measurement of steady-state inactivation curves with different test pulses showed no shifts along the voltage asix. The voltage-dependence of the integral of sodium conductance was measured to test models of sodium channel inactivation in which channels must open before inactivating; the results appear inconsistent with some of the simplest cases of such models.

  20. A novel mechanism for fine-tuning open-state stability in a voltage-gated potassium channel

    DEFF Research Database (Denmark)

    Pless, Stephan Alexander; Niciforovic, Ana P; Galpin, Jason D

    2013-01-01

    fluorinated derivatives of aromatic residues previously implicated in the gating of Shaker potassium channels. Here we show that stepwise dispersion of the negative electrostatic surface potential of only one site, Phe481, stabilizes the channel open state. Furthermore, these data suggest that this apparent...... stabilization is the consequence of the amelioration of an inherently repulsive open-state interaction between the partial negative charge on the face of Phe481 and a highly co-evolved acidic side chain, Glu395, and this interaction is potentially modulated through the Tyr485 hydroxyl. We propose...... that the intrinsic open-state destabilization via aromatic repulsion represents a new mechanism by which ion channels, and likely other proteins, fine-tune conformational equilibria....

  1. NO production and potassium channels activation induced by Crotalus durissus cascavella underlie mesenteric artery relaxation.

    Science.gov (United States)

    Santos, S S; Jesus, R L C; Simões, L O; Vasconcelos, W P; Medeiros, I A; Veras, R C; Casais-E-Silva, L L; Silva, D F

    2017-07-01

    Animal toxins are natural resources for pharmacological studies. The venom of Crotalus durissus cascavella (C.d. cascavella) may be a source in the bio-prospecting of new anti-hypertensive agents. The aim of this study was to investigate vascular effects of the venom of C.d. cascavella in normotensive rats. Studies were performed using isolated mesenteric artery segments and aortic endothelial cells. The cumulative administration of the venom of C.d. cascavella (0.001-30 μg/mL) on phenylephrine (Phe; 10 μM) pre-contracted rings induced a concentration-dependent vasorelaxation in the presence of vascular endothelium (E max  = 47.9 ± 5.0% n = 8), and its effect was almost abolished in the absence of endothelium (E max  = 5.8± 2.4% n = 5 ( ∗∗∗ p stable radical scavenger for nitric oxide (E max  = 10.77± 3.6%, n = 6). Moreover, the venom induced the release of NO by isolated aortic endothelial cells through amperometric studies. When assessing the participation of K + channels on the vasodilatory response of the venom, tyrode solution with 20 mM of KCl caused a significant reduction in the relaxation response (p channels (4-amynopiridine 1 mM; E max  = 9.5 ± 1.3, %, n = 5, ***p channels. These data illustrate that C.d. cascavella is a source of bioactive molecules and therefore has therapeutic potential in the treatment of cardiovascular diseases such as hypertension. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Selective Small Molecule Activators of TREK-2 Channels Stimulate Dorsal Root Ganglion c-Fiber Nociceptor Two-Pore-Domain Potassium Channel Currents and Limit Calcium Influx.

    Science.gov (United States)

    Dadi, Prasanna K; Vierra, Nicholas C; Days, Emily; Dickerson, Matthew T; Vinson, Paige N; Weaver, C David; Jacobson, David A

    2017-03-15

    The two-pore-domain potassium (K2P) channel TREK-2 serves to modulate plasma membrane potential in dorsal root ganglia c-fiber nociceptors, which tunes electrical excitability and nociception. Thus, TREK-2 channels are considered a potential therapeutic target for treating pain; however, there are currently no selective pharmacological tools for TREK-2 channels. Here we report the identification of the first TREK-2 selective activators using a high-throughput fluorescence-based thallium (Tl + ) flux screen (HTS). An initial pilot screen with a bioactive lipid library identified 11-deoxy prostaglandin F2α as a potent activator of TREK-2 channels (EC 50 ≈ 0.294 μM), which was utilized to optimize the TREK-2 Tl + flux assay (Z' = 0.752). A HTS was then performed with 76 575 structurally diverse small molecules. Many small molecules that selectively activate TREK-2 were discovered. As these molecules were able to activate single TREK-2 channels in excised membrane patches, they are likely direct TREK-2 activators. Furthermore, TREK-2 activators reduced primary dorsal root ganglion (DRG) c-fiber Ca 2+ influx. Interestingly, some of the selective TREK-2 activators such as 11-deoxy prostaglandin F2α were found to inhibit the K2P channel TREK-1. Utilizing chimeric channels containing portions of TREK-1 and TREK-2, the region of the TREK channels that allows for either small molecule activation or inhibition was identified. This region lies within the second pore domain containing extracellular loop and is predicted to play an important role in modulating TREK channel activity. Moreover, the selective TREK-2 activators identified in this HTS provide important tools for assessing human TREK-2 channel function and investigating their therapeutic potential for treating chronic pain.

  3. Effects of protein-protein interactions and ligand binding on the ion permeation in KCNQ1 potassium channel.

    Science.gov (United States)

    Jalily Hasani, Horia; Ganesan, Aravindhan; Ahmed, Marawan; Barakat, Khaled H

    2018-01-01

    The voltage-gated KCNQ1 potassium ion channel interacts with the type I transmembrane protein minK (KCNE1) to generate the slow delayed rectifier (IKs) current in the heart. Mutations in these transmembrane proteins have been linked with several heart-related issues, including long QT syndromes (LQTS), congenital atrial fibrillation, and short QT syndrome. Off-target interactions of several drugs with that of KCNQ1/KCNE1 ion channel complex have been known to cause fatal cardiac irregularities. Thus, KCNQ1/KCNE1 remains an important avenue for drug-design and discovery research. In this work, we present the structural and mechanistic details of potassium ion permeation through an open KCNQ1 structural model using the combined molecular dynamics and steered molecular dynamics simulations. We discuss the processes and key residues involved in the permeation of a potassium ion through the KCNQ1 ion channel, and how the ion permeation is affected by (i) the KCNQ1-KCNE1 interactions and (ii) the binding of chromanol 293B ligand and its derivatives into the complex. The results reveal that interactions between KCNQ1 with KCNE1 causes a pore constriction in the former, which in-turn forms small energetic barriers in the ion-permeation pathway. These findings correlate with the previous experimental reports that interactions of KCNE1 dramatically slows the activation of KCNQ1. Upon ligand-binding onto the complex, the energy-barriers along ion permeation path are more pronounced, as expected, therefore, requiring higher force in our steered-MD simulations. Nevertheless, pulling the ion when a weak blocker is bound to the channel does not necessitate high force in SMD. This indicates that our SMD simulations have been able to discern between strong and week blockers and reveal their influence on potassium ion permeation. The findings presented here will have some implications in understanding the potential off-target interactions of the drugs with the KCNQ1/KCNE1 channel

  4. Potassium channel antibody-associated encephalopathy: a potentially immunotherapy-responsive form of limbic encephalitis.

    Science.gov (United States)

    Vincent, Angela; Buckley, Camilla; Schott, Jonathan M; Baker, Ian; Dewar, Bonnie-Kate; Detert, Niels; Clover, Linda; Parkinson, Abigail; Bien, Christian G; Omer, Salah; Lang, Bethan; Rossor, Martin N; Palace, Jackie

    2004-03-01

    Patients presenting with subacute amnesia are frequently seen in acute neurological practice. Amongst the differential diagnoses, herpes simplex encephalitis, Korsakoff's syndrome and limbic encephalitis should be considered. Limbic encephalitis is typically a paraneoplastic syndrome with a poor prognosis; thus, identifying those patients with potentially reversible symptoms is important. Voltage-gated potassium channel antibodies (VGKC-Ab) have recently been reported in three cases of reversible limbic encephalitis. Here we review the clinical, immunological and neuropsychological features of 10 patients (nine male, one female; age range 44-79 years), eight of whom were identified in two centres over a period of 15 months. The patients presented with 1-52 week histories of memory loss, confusion and seizures. Low plasma sodium concentrations, initially resistant to treatment, were present in eight out of 10. Brain MRI at onset showed signal change in the medial temporal lobes in eight out of 10 cases. Paraneoplastic antibodies were negative, but VGKC-Ab ranged from 450 to 5128 pM (neurological and healthy controls memory, with sparing of general intellect in all but two patients, and of nominal functions in all but one. Variable regimes of steroids, plasma exchange and intravenous immunoglobulin were associated with variable falls in serum VGKC-Abs, to values between 2 and 88% of the initial values, together with marked improvement of neuropsychological functioning in six patients, slight improvement in three and none in one. The improvement in neuropsychological functioning in seven patients correlated broadly with the fall in antibodies. However, varying degrees of cerebral atrophy and residual cognitive impairment were common. Over the same period, only one paraneoplastic case of limbic encephalitis was identified between the two main centres. Thus, VGKC-Ab-associated encephalopathy is a relatively common form of autoimmune, non-paraneoplastic, potentially

  5. Discovery and characterization of a potent and selective inhibitor of Aedes aegypti inward rectifier potassium channels.

    Directory of Open Access Journals (Sweden)

    Rene Raphemot

    Full Text Available Vector-borne diseases such as dengue fever and malaria, which are transmitted by infected female mosquitoes, affect nearly half of the world's population. The emergence of insecticide-resistant mosquito populations is reducing the effectiveness of conventional insecticides and threatening current vector control strategies, which has created an urgent need to identify new molecular targets against which novel classes of insecticides can be developed. We previously demonstrated that small molecule inhibitors of mammalian Kir channels represent promising chemicals for new mosquitocide development. In this study, high-throughput screening of approximately 30,000 chemically diverse small-molecules was employed to discover potent and selective inhibitors of Aedes aegypti Kir1 (AeKir1 channels heterologously expressed in HEK293 cells. Of 283 confirmed screening 'hits', the small-molecule inhibitor VU625 was selected for lead optimization and in vivo studies based on its potency and selectivity toward AeKir1, and tractability for medicinal chemistry. In patch clamp electrophysiology experiments of HEK293 cells, VU625 inhibits AeKir1 with an IC50 value of 96.8 nM, making VU625 the most potent inhibitor of AeKir1 described to date. Furthermore, electrophysiology experiments in Xenopus oocytes revealed that VU625 is a weak inhibitor of AeKir2B. Surprisingly, injection of VU625 failed to elicit significant effects on mosquito behavior, urine excretion, or survival. However, when co-injected with probenecid, VU625 inhibited the excretory capacity of mosquitoes and was toxic, suggesting that the compound is a substrate of organic anion and/or ATP-binding cassette (ABC transporters. The dose-toxicity relationship of VU625 (when co-injected with probenecid is biphasic, which is consistent with the molecule inhibiting both AeKir1 and AeKir2B with different potencies. This study demonstrates proof-of-concept that potent and highly selective inhibitors of mosquito

  6. Side Fenestrations Provide an "Anchor" for a Stable Binding of A1899 to the Pore of TASK-1 Potassium Channels.

    Science.gov (United States)

    Ramírez, David; Arévalo, Bárbara; Martínez, Gonzalo; Rinné, Susanne; Sepúlveda, Francisco V; Decher, Niels; González, Wendy

    2017-07-03

    A1899 is a potent and selective inhibitor of the two-pore domain potassium (K 2P ) channel TASK-1. It was previously reported that A1899 acts as an open-channel blocker and binds to residues of the P1 and P2 regions, the M2 and M4 segments, and the halothane response element. The recently described crystal structures of K 2P channels together with the newly identified side fenestrations indicate that residues relevant for TASK-1 inhibition are not purely facing the central cavity as initially proposed. Accordingly, the TASK-1 binding site and the mechanism of inhibition might need a re-evaluation. We have used TASK-1 homology models based on recently crystallized K 2P channels and molecular dynamics simulation to demonstrate that the highly potent TASK-1 blocker A1899 requires binding to residues located in the side fenestrations. Unexpectedly, most of the previously described residues that interfere with TASK-1 blockade by A1899 project their side chains toward the fenestration lumina, underlining the relevance of these structures for drug binding in K 2P channels. Despite its hydrophobicity, A1899 does not seem to use the fenestrations to gain access to the central cavity from the lipid bilayer. In contrast, binding of A1899 to residues of the side fenestrations might provide a physical "anchor", reflecting an energetically favorable binding mode that after pore occlusion stabilizes the closed state of the channels.

  7. Opening of calcium-activated potassium channels improves long-term left-ventricular function after coronary artery occlusion in mice

    NARCIS (Netherlands)

    Behmenburg, Friederike; Hölscher, Nina; Flögel, Ulrich; Hollmann, Markus W.; Heinen, André; Huhn, Ragnar

    2017-01-01

    Background: Opening of mitochondrial calcium-activated potassium channels (BKCa) reduces infarct size after myocardial ischemia/reperfusion injury (I/R). It is unknown if targeting BKCa-channels improves cardiac performance in the long-term after I/R. Methods: Experiments were conducted in

  8. Localization of large conductance calcium-activated potassium channels and their effect on calcitonin gene-related peptide release in the rat trigemino-neuronal pathway

    DEFF Research Database (Denmark)

    Wulf-Johansson, H.; Amrutkar, D.V.; Hay-Schmidt, Anders

    2010-01-01

    Large conductance calcium-activated potassium (BK(Ca)) channels are membrane proteins contributing to electrical propagation through neurons. Calcitonin gene-related peptide (CGRP) is a neuropeptide found in the trigeminovascular system (TGVS). Both BK(Ca) channels and CGRP are involved in migrai...

  9. Estrogen modulates potassium currents and expression of the Kv4.2 subunit in GT1-7 cells.

    Science.gov (United States)

    Farkas, Imre; Varju, Patricia; Liposits, Zsolt

    2007-03-01

    The proper maintenance of reproduction requires the pulsatile secretion of gonadotropin-releasing hormone (GnRH), which is ensured by synchronized periodic firing of multiple GnRH neurons. Both hormone secretion and electrophysiological properties of GnRH cells are influenced by estrogen. The impact of 17beta-estradiol treatment on the function of voltage gated A- and K-type potassium channels, known modulators of firing rate, was therefore examined in our experiments using immortalized GnRH-producing GT1-7 neurons. Whole cell patch clamp recordings showed the absence of the A-type current in GT1-7 cells cultured in estrogen-free medium and after 8h 17beta-estradiol treatment. Exposure of the cells to 17beta-estradiol for 24 and 48 h, respectively, resulted in the appearance of the A-type current. The induction of the A-type current by 17beta-estradiol was dose-related (50 pM to 15 nM range). In contrast, the K-type potassium current was apparent in the estrogen-free environment and 17beta-estradiol administration significantly decreased its amplitude. Co-administration of 17beta-estradiol and estrogen receptor blocker, Faslodex (ICI 182,780; 1 microM) abolished the occurrence of the A-type current. Real-time PCR data demonstrated that expression of the Kv4.2 subunit of the A-type channel was low at 0, 0.5, 2 and 8h, peaked at 24h and diminished at 48 h 17beta-estradiol treatment (15 nM). These data indicate that potassium channels of GT1-7 neurons are regulated by estrogen a mechanism that might contribute to modulation of firing rate and hormone secretion in GnRH neurons.

  10. Synergistic activation of G protein-gated inwardly rectifying potassium channels by cholesterol and PI(4,5)P2.

    Science.gov (United States)

    Bukiya, Anna N; Rosenhouse-Dantsker, Avia

    2017-07-01

    G-protein gated inwardly rectifying potassium (GIRK or Kir3) channels play a major role in the control of the heart rate, and require the membrane phospholipid phosphatidylinositol-bis-phosphate (PI(4,5)P 2 ) for activation. Recently, we have shown that the activity of the heterotetrameric Kir3.1/Kir3.4 channel that underlies atrial K ACh currents was enhanced by cholesterol. Similarly, the activities of both the Kir3.4 homomer and its active pore mutant Kir3.4* (Kir3.4_S143T) were also enhanced by cholesterol. Here we employ planar lipid bilayers to investigate the crosstalk between PI(4,5)P 2 and cholesterol, and demonstrate that these two lipids act synergistically to activate Kir3.4* currents. Further studies using the Xenopus oocytes heterologous expression system suggest that PI(4,5)P 2 and cholesterol act via distinct binding sites. Whereas PI(4,5)P 2 binds to the cytosolic domain of the channel, the putative binding region of cholesterol is located at the center of the transmembrane domain overlapping the central glycine hinge region of the channel. Together, our data suggest that changes in the levels of two key membrane lipids - cholesterol and PI(4,5)P 2 - could act in concert to provide fine-tuning of Kir3 channel function. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Inwardly Rectifying Potassium (Kir) Channels Represent a Critical Ion Conductance Pathway in the Nervous Systems of Insects.

    Science.gov (United States)

    Chen, Rui; Swale, Daniel R

    2018-01-25

    A complete understanding of the physiological pathways critical for proper function of the insect nervous system is still lacking. The recent development of potent and selective small-molecule modulators of insect inward rectifier potassium (Kir) channels has enabled the interrogation of the physiological role and toxicological potential of Kir channels within various insect tissue systems. Therefore, we aimed to highlight the physiological and functional role of neural Kir channels the central nervous system, muscular system, and neuromuscular system through pharmacological and genetic manipulations. Our data provide significant evidence that Drosophila neural systems rely on the inward conductance of K + ions for proper function since pharmacological inhibition and genetic ablation of neural Kir channels yielded dramatic alterations of the CNS spike discharge frequency and broadening and reduced amplitude of the evoked EPSP at the neuromuscular junction. Based on these data, we conclude that neural Kir channels in insects (1) are critical for proper function of the insect nervous system, (2) represents an unexplored physiological pathway that is likely to shape the understanding of neuronal signaling, maintenance of membrane potentials, and maintenance of the ionic balance of insects, and (3) are capable of inducing acute toxicity to insects through neurological poisoning.

  12. Preclinical study of a Kv11.1 potassium channel activator as antineoplastic approach for breast cancer.

    Science.gov (United States)

    Fukushiro-Lopes, Daniela F; Hegel, Alexandra D; Rao, Vidhya; Wyatt, Debra; Baker, Andrew; Breuer, Eun-Kyoung; Osipo, Clodia; Zartman, Jeremiah J; Burnette, Miranda; Kaja, Simon; Kouzoukas, Dimitrios; Burris, Sarah; Jones, W Keith; Gentile, Saverio

    2018-01-09

    Potassium ion (K + ) channels have been recently found to play a critical role in cancer biology. Despite that pharmacologic manipulation of ion channels is recognized as an important therapeutic approach, very little is known about the effects of targeting of K + channels in cancer. In this study, we demonstrate that use of the Kv11.1 K + channel activator NS1643 inhibits tumor growth in an in vivo model of breast cancer. Tumors exposed to NS1643 had reduced levels of proliferation markers, high expression levels of senescence markers, increased production of ROS and DNA damage compared to tumors of untreated mice. Importantly, mice treated with NS1643 did not exhibit significant cardiac dysfunction. In conclusion, pharmacological stimulation of Kv11.1 activity produced arrested TNBC-derived tumor growth by generating DNA damage and senescence without significant side effects. We propose that use of Kv11.1 channels activators could be considered as a possible pharmacological strategy against breast tumors.

  13. Nicotine inhibits potassium currents in Aplysia bag cell neurons

    Science.gov (United States)

    White, Sean H.; Sturgeon, Raymond M.

    2016-01-01

    Acetylcholine and the archetypal cholinergic agonist, nicotine, are typically associated with the opening of ionotropic receptors. In the bag cell neurons, which govern the reproductive behavior of the marine snail, Aplysia californica, there are two cholinergic responses: a relatively large acetylcholine-induced current and a relatively small nicotine-induced current. Both currents are readily apparent at resting membrane potential and result from the opening of distinct ionotropic receptors. We now report a separate current response elicited by applying nicotine to cultured bag cell neurons under whole cell voltage-clamp. This current was ostensibly inward, best resolved at depolarized voltages, presented a noncooperative dose-response with a half-maximal concentration near 1.5 mM, and associated with a decrease in membrane conductance. The unique nicotine-evoked response was not altered by intracellular perfusion with the G protein blocker GDPβS or exposure to classical nicotinic antagonists but was occluded by replacing intracellular K+ with Cs+. Consistent with an underlying mechanism of direct inhibition of one or more K+ channels, nicotine was found to rapidly reduce the fast-inactivating A-type K+ current as well as both components of the delayed-rectifier K+ current. Finally, nicotine increased bag cell neuron excitability, which manifested as reduction in spike threshold, greater action potential height and width, and markedly more spiking to continuous depolarizing current injection. In contrast to conventional transient activation of nicotinic ionotropic receptors, block of K+ channels could represent a nonstandard means for nicotine to profoundly alter the electrical properties of neurons over prolonged periods of time. PMID:26864763

  14. Role of vascular potassium channels in the regulation of renal hemodynamics

    DEFF Research Database (Denmark)

    Sørensen, Charlotte Mehlin; Braunstein, Thomas Hartig; von Holstein-Rathlou, Niels-Henrik

    2012-01-01

    of one or more classes of K+ channels will lead to a change in hemodynamic resistance and therefore of renal blood flow and glomerular filtration pressure. Through these effects, the activity of renal vascular K+ channels influences renal salt and water excretion, fluid homeostasis, and ultimately blood...... function. Results from in vivo experiments are sparse. We discuss the role of the different classes of renal vascular K+ channels and their possible role in the integrated function of the renal microvasculature. Since several pathological conditions, among them hypertension, are associated with alterations...... in K+ channel function, the role of renal vascular K+ channels in the control of salt and water excretion deserves attention....

  15. Block of voltage-gated potassium channels by Pacific ciguatoxin-1 contributes to increased neuronal excitability in rat sensory neurons

    International Nuclear Information System (INIS)

    Birinyi-Strachan, Liesl C.; Gunning, Simon J.; Lewis, Richard J.; Nicholson, Graham M.

    2005-01-01

    The present study investigated the actions of the polyether marine toxin Pacific ciguatoxin-1 (P-CTX-1) on neuronal excitability in rat dorsal root ganglion (DRG) neurons using patch-clamp recording techniques. Under current-clamp conditions, bath application of 2-20 nM P-CTX-1 caused a rapid, concentration-dependent depolarization of the resting membrane potential in neurons expressing tetrodotoxin (TTX)-sensitive voltage-gated sodium (Na v ) channels. This action was completely suppressed by the addition of 200 nM TTX to the external solution, indicating that this effect was mediated through TTX-sensitive Na v channels. In addition, P-CTX-1 also prolonged action potential and afterhyperpolarization (AHP) duration. In a subpopulation of neurons, P-CTX-1 also produced tonic action potential firing, an effect that was not accompanied by significant oscillation of the resting membrane potential. Conversely, in neurons expressing TTX-resistant Na v currents, P-CTX-1 failed to alter any parameter of neuronal excitability examined in this study. Under voltage-clamp conditions in rat DRG neurons, P-CTX-1 inhibited both delayed-rectifier and 'A-type' potassium currents in a dose-dependent manner, actions that occurred in the absence of alterations to the voltage dependence of activation. These actions appear to underlie the prolongation of the action potential and AHP, and contribute to repetitive firing. These data indicate that a block of potassium channels contributes to the increase in neuronal excitability, associated with a modulation of Na v channel gating, observed clinically in response to ciguatera poisoning

  16. The lysosomal potassium channel TMEM175 adopts a novel tetrameric architecture

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Changkeun; Guo, Jiangtao; Zeng, Weizhong; Kim, Sunghoon; She, Ji; Cang, Chunlei; Ren, Dejian; Jiang , Youxing (UPENN); (UTSMC); (HHMI)

    2017-07-19

    TMEM175 is a lysosomal K+ channel that is important for maintaining the membrane potential and pH stability in lysosomes1. It contains two homologous copies of a six-transmembrane-helix (6-TM) domain, which has no sequence homology to the canonical tetrameric K+ channels and lacks the TVGYG selectivity filter motif found in these channels2, 3, 4. The prokaryotic TMEM175 channel, which is present in a subset of bacteria and archaea, contains only a single 6-TM domain and functions as a tetramer. Here, we present the crystal structure of a prokaryotic TMEM175 channel from Chamaesiphon minutus, CmTMEM175, the architecture of which represents a completely different fold from that of canonical K+ channels. All six transmembrane helices of CmTMEM175 are tightly packed within each subunit without undergoing domain swapping. The highly conserved TM1 helix acts as the pore-lining inner helix, creating an hourglass-shaped ion permeation pathway in the channel tetramer. Three layers of hydrophobic residues on the carboxy-terminal half of the TM1 helices form a bottleneck along the ion conduction pathway and serve as the selectivity filter of the channel. Mutagenesis analysis suggests that the first layer of the highly conserved isoleucine residues in the filter is primarily responsible for channel selectivity. Thus, the structure of CmTMEM175 represents a novel architecture of a tetrameric cation channel whose ion selectivity mechanism appears to be distinct from that of the classical K+ channel family.

  17. Excessive blinking and ataxia in a child with occult neuroblastoma and voltage-gated potassium channel antibodies.

    LENUS (Irish Health Repository)

    Allen, Nicholas M

    2012-05-01

    A previously healthy 9-year-old girl presented with a 10-day history of slowly progressive unsteadiness, slurred speech, and behavior change. On examination there was cerebellar ataxia and dysarthria, excessive blinking, subtle perioral myoclonus, and labile mood. The finding of oligoclonal bands in the cerebrospinal fluid prompted paraneoplastic serological evaluation and search for an occult neural crest tumor. Antineuronal nuclear autoantibody type 1 (anti-Hu) and voltage-gated potassium channel complex antibodies were detected in serum. Metaiodobenzylguanidine scan and computed tomography scan of the abdomen showed a localized abdominal mass in the region of the porta hepatis. A diagnosis of occult neuroblastoma was made. Resection of the stage 1 neuroblastoma and treatment with pulsed corticosteroids resulted in resolution of all symptoms and signs. Excessive blinking has rarely been described with neuroblastoma, and, when it is not an isolated finding, it may be a useful clue to this paraneoplastic syndrome. Although voltage-gated potassium channel complex autoimmunity has not been described previously in the setting of neuroblastoma, it is associated with a spectrum of paraneoplastic neurologic manifestations in adults, including peripheral nerve hyperexcitability disorders.

  18. Update on the implication of potassium channels in autism: K+ channelautism spectrum disorder

    Directory of Open Access Journals (Sweden)

    Luca eGuglielmi

    2015-03-01

    Full Text Available Autism spectrum disorders (ASDs are characterized by impaired ability to properly implement environmental stimuli that are essential to achieve a state of social and cultural exchange. Indeed, the main features of ASD are impairments of interpersonal relationships, verbal and non-verbal communication and restricted and repetitive behaviors. These aspects are often accompanied by several comorbidities such as motor delay, praxis impairment, gait abnormalities, insomnia and above all epilepsy. Genetic analyses of autistic individuals uncovered deleterious mutations in several K+ channel types strengthening the notion that their intrinsic dysfunction may play a central etiologic role in ASD. However, indirect implication of K+ channels in ASD has been also reported. For instance, loss of fragile X mental retardation protein (FMRP results in K+ channels deregulation, network dysfunction and ASD-like cognitive and behavioral symptoms. Therefore, this review provides an update on direct and indirect implications of K+ channels in ASDs. Owing to a mounting body of evidence associating a channelopathy pathogenesis to autism and that nearly 500 ion channel proteins are encoded by the human genome, we also propose to classify ASDs − whose susceptibility is significantly enhanced by ion channels defects, either in a monogenic or multigenic condition − in a new category named channelAutism Spectrum Disorder (channelASD; cASD and introduce a new taxonomy (e.g.: Kvx.y-channelASD and likewise Navx.y-channelASD, Cavx.y-channelASD; etc.. This review also highlights some degree of clinical and genetic overlap between K+ channelASDs and K+ channelepsies, whereby such correlation suggests that a subcategory characterized by a channelASD-channelepsy phenotype may be distinguished. Ultimately, this overview aims to further understand the different clinical subgroups and help parse out the distinct biological basis of autism that are essential to establish patient

  19. Effect of beta-adrenoceptor blockers on human ether-a-go-go-related gene (HERG) potassium channels

    DEFF Research Database (Denmark)

    Dupuis, Delphine S; Klaerke, Dan A; Olesen, Søren-Peter

    2005-01-01

    -1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol hydrochloride) blocked the HERG channel with similar affinity, whereas the beta1-receptor antagonists metoprolol and atenolol showed weak effects. Further, the four compounds blocked HERG channels expressed in a mammalian HEK293 cell line...

  20. Role of voltage-gated sodium, potassium and calcium channels in the development of cocaine-associated cardiac arrhythmias

    Science.gov (United States)

    O'Leary, Michael E; Hancox, Jules C

    2010-01-01

    Cocaine is a highly active stimulant that alters dopamine metabolism in the central nervous system resulting in a feeling of euphoria that with time can lead to addictive behaviours. Cocaine has numerous deleterious effects in humans including seizures, vasoconstriction, ischaemia, increased heart rate and blood pressure, cardiac arrhythmias and sudden death. The cardiotoxic effects of cocaine are indirectly mediated by an increase in sympathomimetic stimulation to the heart and coronary vasculature and by a direct effect on the ion channels responsible for maintaining the electrical excitability of the heart. The direct and indirect effects of cocaine work in tandem to disrupt the co-ordinated electrical activity of the heart and have been associated with life-threatening cardiac arrhythmias. This review focuses on the direct effects of cocaine on cardiac ion channels, with particular focus on sodium, potassium and calcium channels, and on the contributions of these channels to cocaine-induced arrhythmias. Companion articles in this edition of the journal examine the epidemiology of cocaine use (Wood & Dargan [1]) and the treatment of cocaine-associated arrhythmias (Hoffmann [2]). PMID:20573078

  1. Phycodnavirus potassium ion channel proteins question the virus molecular piracy hypothesis.

    Science.gov (United States)

    Hamacher, Kay; Greiner, Timo; Ogata, Hiroyuki; Van Etten, James L; Gebhardt, Manuela; Villarreal, Luis P; Cosentino, Cristian; Moroni, Anna; Thiel, Gerhard

    2012-01-01

    Phycodnaviruses are large dsDNA, algal-infecting viruses that encode many genes with homologs in prokaryotes and eukaryotes. Among the viral gene products are the smallest proteins known to form functional K(+) channels. To determine if these viral K(+) channels are the product of molecular piracy from their hosts, we compared the sequences of the K(+) channel pore modules from seven phycodnaviruses to the K(+) channels from Chlorella variabilis and Ectocarpus siliculosus, whose genomes have recently been sequenced. C. variabilis is the host for two of the viruses PBCV-1 and NY-2A and E. siliculosus is the host for the virus EsV-1. Systematic phylogenetic analyses consistently indicate that the viral K(+) channels are not related to any lineage of the host channel homologs and that they are more closely related to each other than to their host homologs. A consensus sequence of the viral channels resembles a protein of unknown function from a proteobacterium. However, the bacterial protein lacks the consensus motif of all K(+) channels and it does not form a functional channel in yeast, suggesting that the viral channels did not come from a proteobacterium. Collectively, our results indicate that the viruses did not acquire their K(+) channel-encoding genes from their current algal hosts by gene transfer; thus alternative explanations are required. One possibility is that the viral genes arose from ancient organisms, which served as their hosts before the viruses developed their current host specificity. Alternatively the viral proteins could be the origin of K(+) channels in algae and perhaps even all cellular organisms.

  2. Phycodnavirus potassium ion channel proteins question the virus molecular piracy hypothesis.

    Directory of Open Access Journals (Sweden)

    Kay Hamacher

    Full Text Available Phycodnaviruses are large dsDNA, algal-infecting viruses that encode many genes with homologs in prokaryotes and eukaryotes. Among the viral gene products are the smallest proteins known to form functional K(+ channels. To determine if these viral K(+ channels are the product of molecular piracy from their hosts, we compared the sequences of the K(+ channel pore modules from seven phycodnaviruses to the K(+ channels from Chlorella variabilis and Ectocarpus siliculosus, whose genomes have recently been sequenced. C. variabilis is the host for two of the viruses PBCV-1 and NY-2A and E. siliculosus is the host for the virus EsV-1. Systematic phylogenetic analyses consistently indicate that the viral K(+ channels are not related to any lineage of the host channel homologs and that they are more closely related to each other than to their host homologs. A consensus sequence of the viral channels resembles a protein of unknown function from a proteobacterium. However, the bacterial protein lacks the consensus motif of all K(+ channels and it does not form a functional channel in yeast, suggesting that the viral channels did not come from a proteobacterium. Collectively, our results indicate that the viruses did not acquire their K(+ channel-encoding genes from their current algal hosts by gene transfer; thus alternative explanations are required. One possibility is that the viral genes arose from ancient organisms, which served as their hosts before the viruses developed their current host specificity. Alternatively the viral proteins could be the origin of K(+ channels in algae and perhaps even all cellular organisms.

  3. Therapeutic targeting of two-pore-domain potassium (K(2P)) channels in the cardiovascular system.

    Science.gov (United States)

    Wiedmann, Felix; Schmidt, Constanze; Lugenbiel, Patrick; Staudacher, Ingo; Rahm, Ann-Kathrin; Seyler, Claudia; Schweizer, Patrick A; Katus, Hugo A; Thomas, Dierk

    2016-05-01

    The improvement of treatment strategies in cardiovascular medicine is an ongoing process that requires constant optimization. The ability of a therapeutic intervention to prevent cardiovascular pathology largely depends on its capacity to suppress the underlying mechanisms. Attenuation or reversal of disease-specific pathways has emerged as a promising paradigm, providing a mechanistic rationale for patient-tailored therapy. Two-pore-domain K(+) (K(2P)) channels conduct outward K(+) currents that stabilize the resting membrane potential and facilitate action potential repolarization. K(2P) expression in the cardiovascular system and polymodal K2P current regulation suggest functional significance and potential therapeutic roles of the channels. Recent work has focused primarily on K(2P)1.1 [tandem of pore domains in a weak inwardly rectifying K(+) channel (TWIK)-1], K(2P)2.1 [TWIK-related K(+) channel (TREK)-1], and K(2P)3.1 [TWIK-related acid-sensitive K(+) channel (TASK)-1] channels and their role in heart and vessels. K(2P) currents have been implicated in atrial and ventricular arrhythmogenesis and in setting the vascular tone. Furthermore, the association of genetic alterations in K(2P)3.1 channels with atrial fibrillation, cardiac conduction disorders and pulmonary arterial hypertension demonstrates the relevance of the channels in cardiovascular disease. The function, regulation and clinical significance of cardiovascular K(2P) channels are summarized in the present review, and therapeutic options are emphasized. © 2016 Authors; published by Portland Press Limited.

  4. The Role of Potassium Channels in Arabidopsis thaliana Long Distance Electrical Signalling: AKT2 Modulates Tissue Excitability While GORK Shapes Action Potentials

    Directory of Open Access Journals (Sweden)

    Tracey Ann Cuin

    2018-03-01

    Full Text Available Fast responses to an external threat depend on the rapid transmission of signals through a plant. Action potentials (APs are proposed as such signals. Plant APs share similarities with their animal counterparts; they are proposed to depend on the activity of voltage-gated ion channels. Nonetheless, despite their demonstrated role in (abiotic stress responses, the identities of the associated voltage-gated channels and transporters remain undefined in higher plants. By demonstrating the role of two potassium-selective channels in Arabidopsis thaliana in AP generation and shaping, we show that the plant AP does depend on similar Kv-like transport systems to those of the animal signal. We demonstrate that the outward-rectifying potassium-selective channel GORK limits the AP amplitude and duration, while the weakly-rectifying channel AKT2 affects membrane excitability. By computational modelling of plant APs, we reveal that the GORK activity not only determines the length of an AP but also the steepness of its rise and the maximal amplitude. Thus, outward-rectifying potassium channels contribute to both the repolarisation phase and the initial depolarisation phase of the signal. Additionally, from modelling considerations we provide indications that plant APs might be accompanied by potassium waves, which prime the excitability of the green cable.

  5. Structural Analysis and Deletion Mutagenesis Define Regions of QUIVER/SLEEPLESS that Are Responsible for Interactions with Shaker-Type Potassium Channels and Nicotinic Acetylcholine Receptors.

    Directory of Open Access Journals (Sweden)

    Meilin Wu

    Full Text Available Ly6 proteins are endogenous prototoxins found in most animals. They show striking structural and functional parallels to snake α-neurotoxins, including regulation of ion channels and cholinergic signaling. However, the structural contributions of Ly6 proteins to regulation of effector molecules is poorly understood. This question is particularly relevant to the Ly6 protein QUIVER/SLEEPLESS (QVR/SSS, which has previously been shown to suppress excitability and synaptic transmission by upregulating potassium (K channels and downregulating nicotinic acetylcholine receptors (nAChRs in wake-promoting neurons to facilitate sleep in Drosophila. Using deletion mutagenesis, co-immunoprecipitations, ion flux assays, surface labeling and confocal microscopy, we demonstrate that only loop 2 is required for many of the previously described properties of SSS in transfected cells, including interactions with K channels and nAChRs. Collectively our data suggest that QVR/SSS, and by extension perhaps other Ly6 proteins, target effector molecules using limited protein motifs. Mapping these motifs may be useful in rational design of drugs that mimic or suppress Ly6-effector interactions to modulate nervous system function.

  6. Structural Analysis and Deletion Mutagenesis Define Regions of QUIVER/SLEEPLESS that Are Responsible for Interactions with Shaker-Type Potassium Channels and Nicotinic Acetylcholine Receptors.

    Science.gov (United States)

    Wu, Meilin; Liu, Clifford Z; Joiner, William J

    2016-01-01

    Ly6 proteins are endogenous prototoxins found in most animals. They show striking structural and functional parallels to snake α-neurotoxins, including regulation of ion channels and cholinergic signaling. However, the structural contributions of Ly6 proteins to regulation of effector molecules is poorly understood. This question is particularly relevant to the Ly6 protein QUIVER/SLEEPLESS (QVR/SSS), which has previously been shown to suppress excitability and synaptic transmission by upregulating potassium (K) channels and downregulating nicotinic acetylcholine receptors (nAChRs) in wake-promoting neurons to facilitate sleep in Drosophila. Using deletion mutagenesis, co-immunoprecipitations, ion flux assays, surface labeling and confocal microscopy, we demonstrate that only loop 2 is required for many of the previously described properties of SSS in transfected cells, including interactions with K channels and nAChRs. Collectively our data suggest that QVR/SSS, and by extension perhaps other Ly6 proteins, target effector molecules using limited protein motifs. Mapping these motifs may be useful in rational design of drugs that mimic or suppress Ly6-effector interactions to modulate nervous system function.

  7. ATP-sensitive potassium (KATP channel openers diazoxide and nicorandil lower intraocular pressure by activating the Erk1/2 signaling pathway.

    Directory of Open Access Journals (Sweden)

    Uttio Roy Chowdhury

    Full Text Available Elevated intraocular pressure is the most prevalent and only treatable risk factor for glaucoma, a degenerative disease of the optic nerve. While treatment options to slow disease progression are available, all current therapeutic and surgical treatments have unwanted side effects or limited efficacy, resulting in the need to identify new options. Previous reports from our laboratory have established a novel ocular hypotensive effect of ATP-sensitive potassium channel (KATP openers including diazoxide (DZ and nicorandil (NCD. In the current study, we evaluated the role of Erk1/2 signaling pathway in KATP channel opener mediated reduction of intraocular pressure (IOP. Western blot analysis of DZ and NCD treated primary normal trabecular meshwork (NTM cells, human TM (isolated from perfusion cultures of human anterior segments and mouse eyes showed increased phosphorylation of Erk1/2 when compared to vehicle treated controls. DZ and NCD mediated pressure reduction (p0.1. Histologic evaluation of transmission electron micrographs from DZ + U0126 and NCD + U0126 treated eyes revealed no observable morphological changes in the ultrastructure of the conventional outflow pathway. Taken together, the results indicate that the Erk1/2 pathway is necessary for IOP reduction by KATP channel openers DZ and NCD.

  8. Stimulatory (insulin-mimetic) and inhibitory (ouabain-like) action of vanadate on potassium uptake and cellular sodium and potassium in heart cells in culture.

    Science.gov (United States)

    Werdan, K; Bauriedel, G; Fischer, B; Krawietz, W; Erdmann, E; Schmitz, W; Scholz, H

    1982-04-23

    (1) The influence of vanadate (Na3VO4) on sodium and potassium uptake as well as on cellular ion contents of sodium and potassium has been studied in heart muscle and non-muscle cells obtained from various species. An ouabain-like inhibition of potassium uptake (up to 50%), combined with a decrease of cellular potassium (up to 20%) has been observed by vanadate (10(-4)-10(-3) M) in heart non-muscle cells obtained from neonatal guinea pigs and chick embryos. In heart muscle and non-muscle cells prepared from neonatal rats, as well as in Girardi human heart cells, a vanadate-induced stimulation of potassium uptake (up to 100%), combined with a rise in cellular potassium (up to 20%) and without significant alteration of cellular sodium, has been found. A slight increase of 22Na+ influx can be measured in rat heart muscle cells and in Girardi human heart cells in the presence of vanadate (10(-4)--10(-3) M). (2) In beating rat heart muscle cells in culture, detrimental effects of serum deprivation--concerning beating properties, potassium uptake and cellular potassium--can at least in part be overcome by addition of vanadate. Furthermore, this compound prevents ouabain-induced signs of toxicity (contractures) in these cells. (3) The stimulatory effects of vanadate on potassium can be mimicked by insulin (1-10 mU/ml). Furthermore, vanadate produces an insulin-like stimulation of 2-deoxy-D-glucose uptake in rat heart muscle and non-muscle cells as well as in Girardi human heart cells. (4) The experimental data demonstrate an ouabain-like inhibition as well as an insulin-mimetic stimulation of potassium-uptake in various heart cells. The reason for this antagonistic mode of action may be due to the different capabilities of the heart cell types to reduce vanadium in the V-valence state of vanadium in the IV-valence state, thereby favouring either ouabain-like inhibition (vanadium V) or insulin-mimetic stimulation (vanadium IV) of potassium transport.

  9. Cholesterol up-regulates neuronal G protein-gated inwardly rectifying potassium (GIRK) channel activity in the hippocampus.

    Science.gov (United States)

    Bukiya, Anna N; Durdagi, Serdar; Noskov, Sergei; Rosenhouse-Dantsker, Avia

    2017-04-14

    Hypercholesterolemia is a well known risk factor for the development of neurodegenerative disease. However, the underlying mechanisms are mostly unknown. In recent years, it has become increasingly evident that cholesterol-driven effects on physiology and pathophysiology derive from its ability to alter the function of a variety of membrane proteins including ion channels. Yet, the effect of cholesterol on G protein-gated inwardly rectifying potassium (GIRK) channels expressed in the brain is unknown. GIRK channels mediate the actions of inhibitory brain neurotransmitters. As a result, loss of GIRK function can enhance neuron excitability, whereas gain of GIRK function can reduce neuronal activity. Here we show that in rats on a high-cholesterol diet, cholesterol levels in hippocampal neurons are increased. We also demonstrate that cholesterol plays a critical role in modulating neuronal GIRK currents. Specifically, cholesterol enrichment of rat hippocampal neurons resulted in enhanced channel activity. In accordance, elevated currents upon cholesterol enrichment were also observed in Xenopus oocytes expressing GIRK2 channels, the primary GIRK subunit expressed in the brain. Furthermore, using planar lipid bilayers, we show that although cholesterol did not affect the unitary conductance of GIRK2, it significantly enhanced the frequency of channel openings. Last, combining computational and functional approaches, we identified two putative cholesterol-binding sites in the transmembrane domain of GIRK2. These findings establish that cholesterol plays a critical role in modulating GIRK activity in the brain. Because up-regulation of GIRK function can reduce neuronal activity, our findings may lead to novel approaches for prevention and therapy of cholesterol-driven neurodegenerative disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Development and validation of fluorescence-based and automated patch clamp-based functional assays for the inward rectifier potassium channel Kir4.1.

    Science.gov (United States)

    Raphemot, Rene; Kadakia, Rishin J; Olsen, Michelle L; Banerjee, Sreedatta; Days, Emily; Smith, Stephen S; Weaver, C David; Denton, Jerod S

    2013-01-01

    The inward rectifier potassium (Kir) channel Kir4.1 plays essential roles in modulation of neurotransmission and renal sodium transport and may represent a novel drug target for temporal lobe epilepsy and hypertension. The molecular pharmacology of Kir4.1 is limited to neurological drugs, such as fluoxetine (Prozac(©)), exhibiting weak and nonspecific activity toward the channel. The development of potent and selective small-molecule probes would provide critically needed tools for exploring the integrative physiology and therapeutic potential of Kir4.1. A fluorescence-based thallium (Tl(+)) flux assay that utilizes a tetracycline-inducible T-Rex-HEK293-Kir4.1 cell line to enable high-throughput screening (HTS) of small-molecule libraries was developed. The assay is dimethyl sulfoxide tolerant and exhibits robust screening statistics (Z'=0.75±0.06). A pilot screen of 3,655 small molecules and lipids revealed 16 Kir4.1 inhibitors (0.4% hit rate). 3,3-Diphenyl-N-(1-phenylethyl)propan-1-amine, termed VU717, inhibits Kir4.1-mediated thallium flux with an IC50 of ∼6 μM. An automated patch clamp assay using the IonFlux HT workbench was developed to facilitate compound characterization. Leak-subtracted ensemble "loose patch" recordings revealed robust tetracycline-inducible and Kir4.1 currents that were inhibited by fluoxetine (IC50=10 μM), VU717 (IC50=6 μM), and structurally related calcium channel blocker prenylamine (IC50=6 μM). Finally, we demonstrate that VU717 inhibits Kir4.1 channel activity in cultured rat astrocytes, providing proof-of-concept that the Tl(+) flux and IonFlux HT assays can enable the discovery of antagonists that are active against native Kir4.1 channels.

  11. Small-conductance Ca2+-activated potassium type 2 channels regulate the formation of contextual fear memory.

    Directory of Open Access Journals (Sweden)

    Saravana R K Murthy

    Full Text Available Small-conductance, Ca2+ activated K+ channels (SK channels are expressed at high levels in brain regions responsible for learning and memory. In the current study we characterized the contribution of SK2 channels to synaptic plasticity and to different phases of hippocampal memory formation. Selective SK2 antisense-treatment facilitated basal synaptic transmission and theta-burst induced LTP in hippocampal brain slices. Using the selective SK2 antagonist Lei-Dab7 or SK2 antisense probes, we found that hippocampal SK2 channels are critical during two different time windows: 1 blockade of SK2 channels before the training impaired fear memory, whereas, 2 blockade of SK2 channels immediately after the training enhanced contextual fear memory. We provided the evidence that the post-training cleavage of the SK2 channels was responsible for the observed bidirectional effect of SK2 channel blockade on memory consolidation. Thus, Lei-Dab7-injection before training impaired the C-terminal cleavage of SK2 channels, while Lei-Dab7 given immediately after training facilitated the C-terminal cleavage. Application of the synthetic peptide comprising a leucine-zipper domain of the C-terminal fragment to Jurkat cells impaired SK2 channel-mediated currents, indicating that the endogenously cleaved fragment might exert its effects on memory formation by blocking SK2 channel-mediated currents. Our present findings suggest that SK2 channel proteins contribute to synaptic plasticity and memory not only as ion channels but also by additionally generating a SK2 C-terminal fragment, involved in both processes. The modulation of fear memory by down-regulating SK2 C-terminal cleavage might have applicability in the treatment of anxiety disorders in which fear conditioning is enhanced.

  12. Discovery and Characterization of 1H-Pyrazol-5-yl-2-phenylacetamides as Novel, Non-Urea-Containing GIRK1/2 Potassium Channel Activators.

    Science.gov (United States)

    Wieting, Joshua M; Vadukoot, Anish K; Sharma, Swagat; Abney, Kristopher K; Bridges, Thomas M; Daniels, J Scott; Morrison, Ryan D; Wickman, Kevin; Weaver, C David; Hopkins, Corey R

    2017-09-20

    The G protein-gated inwardly-rectifying potassium channels (GIRK, K ir 3) are a family of inward-rectifying potassium channels, and there is significant evidence supporting the roles of GIRKs in a number of physiological processes and as potential targets for numerous indications. Previously reported urea containing molecules as GIRK1/2 preferring activators have had significant pharmacokinetic (PK) liabilities. Here we report a novel series of 1H-pyrazolo-5-yl-2-phenylacetamides in an effort to improve upon the PK properties. This series of compounds display nanomolar potency as GIRK1/2 activators with improved brain distribution (rodent K p > 0.6).

  13. Functional regulation of BK potassium channels by γ1 auxiliary subunits.

    Science.gov (United States)

    Gonzalez-Perez, Vivian; Xia, Xiao-Ming; Lingle, Christopher J

    2014-04-01

    Many K(+) channels are oligomeric complexes with intrinsic structural symmetry arising from the homo-tetrameric core of their pore-forming subunits. Allosteric regulation of tetramerically symmetric proteins, whether by intrinsic sensing domains or associated auxiliary subunits, often mirrors the fourfold structural symmetry. Here, through patch-clamp recordings of channel population ensembles and also single channels, we examine regulation of the Ca(2+)- and voltage-activated large conductance Ca(2+)-activated K(+) (BK) channel by associated γ1-subunits. Through expression of differing ratios of γ1:α-subunits, the results reveal an all-or-none functional regulation of BK channels by γ-subunits: channels either exhibit a full gating shift or no shift at all. Furthermore, the γ1-induced shift exhibits a state-dependent labile behavior that recapitulates the fully shifted or unshifted behavior. The γ1-induced shift contrasts markedly to the incremental shifts in BK gating produced by 1-4 β-subunits and adds a new layer of complexity to the mechanisms by which BK channel functional diversity is generated.

  14. Engineering a peptide inhibitor towards the KCNQ1/KCNE1 potassium channel (IKs).

    Science.gov (United States)

    Hu, Youtian; Chen, Jing; Wang, Bin; Yang, Weishan; Zhang, Chuangeng; Hu, Jun; Xie, Zili; Cao, Zhijian; Li, Wenxin; Wu, Yingliang; Chen, Zongyun

    2015-09-01

    The KCNQ1/KCNE1 channel (IKs) plays important roles in the physiological and pathological process of heart, but no potent peptide acting on this channel has been reported. In this work, we found that the natural scorpion venom hardly inhibited KCNQ1/KCNE1 channel currents. Based on this observation, we attempted to use three natural scorpion toxins ChTX, BmKTX and OmTx2 with two different structural folds as templates to engineer potent peptide inhibitors towards the KCNQ1/KCNE1 channel. Pharmacological experiments showed that when we screen with 1μM MT2 peptide, an analog derived from BmKTX toxin, KCNQ1/KCNE1 channel currents could be effectively inhibited. Concentration-dependent experiments showed that MT2 inhibited the KCNQ1/KCNE1 channel with an IC50 value of 4.6±1.9μM. The mutagenesis experiments indicated that MT2 peptide likely used Lys26 residue to interact with the KCNQ1/KCNE1 channel. With MT2 as a new template, we further designed a more potent MT2-2 peptide, which selectively inhibited the KCNQ1/KCNE1 channel with an IC50 of 1.51±0.62μM. Together, this work provided a much potent KCNQ1/KCNE1 channel peptide inhibitor so far, and highlighted the role of molecular strategy in developing potent peptide inhibitors for the natural toxin-insensitive orphan receptors. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Purification and structural study of the voltage-sensor domain of the human KCNQ1 potassium ion channel.

    Science.gov (United States)

    Peng, Dungeng; Kim, Ji-Hun; Kroncke, Brett M; Law, Cheryl L; Xia, Yan; Droege, Kristin D; Van Horn, Wade D; Vanoye, Carlos G; Sanders, Charles R

    2014-04-01

    KCNQ1 (also known as KV7.1 or KVLQT1) is a voltage-gated potassium channel modulated by members of the KCNE protein family. Among multiple functions, KCNQ1 plays a critical role in the cardiac action potential. This channel is also subject to inherited mutations that cause certain cardiac arrhythmias and deafness. In this study, we report the overexpression, purification, and preliminary structural characterization of the voltage-sensor domain (VSD) of human KCNQ1 (Q1-VSD). Q1-VSD was expressed in Escherichia coli and purified into lyso-palmitoylphosphatidylglycerol micelles, conditions under which this tetraspan membrane protein yields excellent nuclear magnetic resonance (NMR) spectra. NMR studies reveal that Q1-VSD shares a common overall topology with other channel VSDs, with an S0 helix followed by transmembrane helices S1-S4. The exact sequential locations of the helical spans do, however, show significant variations from those of the homologous segments of previously characterized VSDs. The S4 segment of Q1-VSD was seen to be α-helical (with no 310 component) and underwent rapid backbone amide H-D exchange over most of its length. These results lay the foundation for more advanced structural studies and can be used to generate testable hypotheses for future structure-function experiments.

  16. Localization and function of ATP-sensitive potassium channels in human skeletal muscle

    DEFF Research Database (Denmark)

    Nielsen, Jens Jung; Kristensen, Michael; Hellsten, Ylva

    2003-01-01

    The present study investigated the localization of ATP-sensitive K+ (KATP) channels in human skeletal muscle and the functional importance of these channels for human muscle K+ distribution at rest and during muscle activity. Membrane fractionation based on the giant vesicle technique...... or the sucrose-gradient technique in combination with Western blotting demonstrated that the KATP channels are mainly located in the sarcolemma. This localization was confirmed by immunohistochemical measurements. With the microdialysis technique, it was demonstrated that local application of the KATP channel...... inhibitor glibenclamide reduced (P approximately 4.5 to 4.0 mM, whereas the concentration in the control leg remained constant. Glibenclamide had no effect on the interstitial K+ accumulation during knee-extensor exercise at a power output of 60 W. In contrast...

  17. Contributions of counter-charge in a potassium channel voltage-sensor domain

    DEFF Research Database (Denmark)

    Pless, Stephan Alexander; Galpin, Jason D; Niciforovic, Ana P

    2011-01-01

    Voltage-sensor domains couple membrane potential to conformational changes in voltage-gated ion channels and phosphatases. Highly coevolved acidic and aromatic side chains assist the transfer of cationic side chains across the transmembrane electric field during voltage sensing. We investigated...... the functional contribution of negative electrostatic potentials from these residues to channel gating and voltage sensing with unnatural amino acid mutagenesis, electrophysiology, voltage-clamp fluorometry and ab initio calculations. The data show that neutralization of two conserved acidic side chains...

  18. [Feature of ACh-sensitive potassium current in outer hair cells of guinea pig cochlea].

    Science.gov (United States)

    Guo, Chang-kai; Kong, Wei-jia; Yu, Qing-song; Li, Guan-qiao; Li, Jia-li; Li, Qing-tian

    2007-04-01

    To explore the feature of the ACh-sensitive potassium current in guinea pig cochlear outer hair cells. Cochlear outer hair cells of guinea pigs (n=38) were isolated by collagenase type IV. Under the whole-cell patch mode, the ions nature and the pharmacological properties of the ACh-sensitive potassium current were investigated by applying the inhibitors of calcium-dependent potassium currents and the inhibitors of nicotinic ACh receptor. Following application of ACh, cochlear outer hair cells displayed a rapidly activating outward potassium current with a fast desensitized kinetic and a reversal (x +/- s) potential of (-67.3 +/- 8.2) mV (n=10). At the holding potential of -50 mV, the current amplitude of ACh-sensitive potassium current activated by 100 micronmol/L ACh was (506.6 +/- 186.3) pA (n=9). ACh-sensitive potassium current was sensitive to TEA (tetraethylammonium chloride, 10 mmol/L) and potently inhibited by the small conductance calcium-dependent potassium current (SK) blocker, apamin (1 micromol/L). Iberiotoxin (IBTX), the well-known blocker of big conductance calcium-dependent potassium current (BK), failed to inhibit the amplitude of the ACh-sensitive potassium current at the dose of 200 nmol/L. The dose for half-maximal response (EC50) of the ACh-sensitive potassium current was (33.5 +/- 5.7) micromol/L (n=7). The ACh-sensitive potassium current was sensitive to the GABA (gamma-aminobutyric acid)-A receptor blocker, bicuculline, and strongly inhibited by the selective blocker of the alpha 9-nicotinic ACh receptor, strychnine. Strychnine and bicuculline showed the dose-dependent blocking effect with a half inhibition-maximal response (IC50) of (22.3 +/- 2.6) nmol/L (n=7) and (1.2 +/- 0.4) micromol/L (n=6), respectively. This work provides direct evidences that the ACh-sensitive SK current was present on guinea pig cochlear outer hair cells. The activation of the ACh-sensitive SK current was most possibly mediated by a alpha 9-nicotinic ACh receptor.

  19. Understanding cell passage through constricted microfluidic channels

    Science.gov (United States)

    Cartas-Ayala, Marco A.; Karnik, Rohit

    2012-11-01

    Recently, several microfluidic platforms have been proposed to characterize cells based on their behaviour during cell passage through constricted channels. Variables like transit time have been analyzed in disease states like sickle cell anemia, malaria and sepsis. Nevertheless, it is hard to make direct comparisons between different platforms and cell types. We present experimental results of the relationship between solid deformable particle properties, i.e. stiffness and relative particle size, and flow properties, i.e. particle's velocity. We measured the hydrodynamic variables during the flow of HL-60 cells, a white myeloid cell type, in narrow microfluidic square channels using a microfluidic differential manometer. We measured the flow force required to move cells of different sizes through microchannels and quantified friction forces opposing cell passage. We determined the non-dimensional parameters that influence the flow of cells and we used them to obtain a non dimensional expression that can be used to predict the forces needed to drive cells through microchannels. We found that the friction force needed to flow HL-60 through a microfluidic channel is the sum of two parts. The first part is a static friction force that is proportional to the force needed to keep the force compressed. The second part is a factor that is proportional to the cell velocity, hence a dynamic term, and slightly sensitive to the compressive force. We thank CONACYT (Mexican Science and Technology Council) for supporting this project, grant 205899.

  20. Allosteric gating mechanism underlies the flexible gating of KCNQ1 potassium channels

    Science.gov (United States)

    Osteen, Jeremiah D.; Barro-Soria, Rene; Robey, Seth; Sampson, Kevin J.; Kass, Robert S.; Larsson, H. Peter

    2012-01-01

    KCNQ1 (Kv7.1) is a unique member of the superfamily of voltage-gated K+ channels in that it displays a remarkable range of gating behaviors tuned by coassembly with different β subunits of the KCNE family of proteins. To better understand the basis for the biophysical diversity of KCNQ1 channels, we here investigate the basis of KCNQ1 gating in the absence of β subunits using voltage-clamp fluorometry (VCF). In our previous study, we found the kinetics and voltage dependence of voltage-sensor movements are very similar to those of the channel gate, as if multiple voltage-sensor movements are not required to precede gate opening. Here, we have tested two different hypotheses to explain KCNQ1 gating: (i) KCNQ1 voltage sensors undergo a single concerted movement that leads to channel opening, or (ii) individual voltage-sensor movements lead to channel opening before all voltage sensors have moved. Here, we find that KCNQ1 voltage sensors move relatively independently, but that the channel can conduct before all voltage sensors have activated. We explore a KCNQ1 point mutation that causes some channels to transition to the open state even in the absence of voltage-sensor movement. To interpret these results, we adopt an allosteric gating scheme wherein KCNQ1 is able to transition to the open state after zero to four voltage-sensor movements. This model allows for widely varying gating behavior, depending on the relative strength of the opening transition, and suggests how KCNQ1 could be controlled by coassembly with different KCNE family members. PMID:22509038

  1. Development and Validation of Fluorescence-Based and Automated Patch Clamp–Based Functional Assays for the Inward Rectifier Potassium Channel Kir4.1

    OpenAIRE

    Raphemot, Rene; Kadakia, Rishin J.; Olsen, Michelle L.; Banerjee, Sreedatta; Days, Emily; Smith, Stephen S.; Weaver, C. David; Denton, Jerod S.

    2013-01-01

    The inward rectifier potassium (Kir) channel Kir4.1 plays essential roles in modulation of neurotransmission and renal sodium transport and may represent a novel drug target for temporal lobe epilepsy and hypertension. The molecular pharmacology of Kir4.1 is limited to neurological drugs, such as fluoxetine (Prozac©), exhibiting weak and nonspecific activity toward the channel. The development of potent and selective small-molecule probes would provide critically needed tools for exploring th...

  2. Expression of Inwardly Rectifying Potassium Channel Subunits in Native Human Retinal Pigment Epithelium

    OpenAIRE

    Yang, Dongli; Zhang, Xiaoming; Hughes, Bret A.

    2008-01-01

    Previously, we demonstrated that the inwardly rectifying K+ (Kir) channel subunit Kir7.1 is highly expressed in bovine and human retinal pigment epithelium (RPE). The purpose of this study was to determine whether any of the 14 other members of the Kir gene family are expressed in native human RPE. Conventional reverse transcription-polymerase chain reaction (RT-PCR) analysis indicated that in addition to Kir7.1, 7 other Kir channel subunits (Kir1.1, Kir2.1, Kir2.2, Kir3.1, Kir3.4, Kir4.2 and...

  3. Association analysis of a highly polymorphic CAG Repeat in the human potassium channel gene KCNN3 and migraine susceptibility

    Directory of Open Access Journals (Sweden)

    Ovcaric Mick

    2005-09-01

    Full Text Available Abstract Background Migraine is a polygenic multifactorial disease, possessing environmental and genetic causative factors with multiple involved genes. Mutations in various ion channel genes are responsible for a number of neurological disorders. KCNN3 is a neuronal small conductance calcium-activated potassium channel gene that contains two polyglutamine tracts, encoded by polymorphic CAG repeats in the gene. This gene plays a critical role in determining the firing pattern of neurons and acts to regulate intracellular calcium channels. Methods The present association study tested whether length variations in the second (more 3' polymorphic CAG repeat in exon 1 of the KCNN3 gene, are involved in susceptibility to migraine with and without aura (MA and MO. In total 423 DNA samples from unrelated individuals, of which 202 consisted of migraine patients and 221 non-migraine controls, were genotyped and analysed using a fluorescence labelled primer set on an ABI310 Genetic Analyzer. Allele frequencies were calculated from observed genotype counts for the KCNN3 polymorphism. Analysis was performed using standard contingency table analysis, incorporating the chi-squared test of independence and CLUMP analysis. Results Overall, there was no convincing evidence that KCNN3 CAG lengths differ between Caucasian migraineurs and controls, with no significant difference in the allelic length distribution of CAG repeats between the population groups (P = 0.090. Also the MA and MO subtypes did not differ significantly between control allelic distributions (P > 0.05. The prevalence of the long CAG repeat (>19 repeats did not reach statistical significance in migraineurs (P = 0.15, nor was there a significant difference between the MA and MO subgroups observed compared to controls (P = 0.46 and P = 0.09, respectively, or between MA vs MO (P = 0.40. Conclusion This association study provides no evidence that length variations of the second polyglutamine array in

  4. Dynamic expression of genes encoding subunits of inward rectifier potassium (Kir) channels in the yellow fever mosquito Aedes aegypti.

    Science.gov (United States)

    Yang, Zhongxia; Statler, Bethanie-Michelle; Calkins, Travis L; Alfaro, Edna; Esquivel, Carlos J; Rouhier, Matthew F; Denton, Jerod S; Piermarini, Peter M

    2017-02-01

    Inward rectifier potassium (Kir) channels play fundamental roles in neuromuscular, epithelial, and endocrine function in mammals. Recent research in insects suggests that Kir channels play critical roles in the development, immune function, and excretory physiology of fruit flies and/or mosquitoes. Moreover, our group has demonstrated that mosquito Kir channels may serve as valuable targets for the development of novel insecticides. Here we characterize the molecular expression of 5 mRNAs encoding Kir channel subunits in the yellow fever mosquito, Aedes aegypti: Kir1, Kir2A-c, Kir2B, Kir2B', and Kir3. We demonstrate that 1) Kir mRNA expression is dynamic in whole mosquitoes, Malpighian tubules, and the midgut during development from 4th instar larvae to adult females, 2) Kir2B and Kir3 mRNA levels are reduced in 4th instar larvae when reared in water containing an elevated concentration (50mM) of KCl, but not NaCl, and 3) Kir mRNAs are differentially expressed in the Malpighian tubules, midgut, and ovaries within 24h after blood feeding. Furthermore, we provide the first characterization of Kir mRNA expression in the anal papillae of 4th instar larval mosquitoes, which indicates that Kir2A-c is the most abundant. Altogether, the data provide the first comprehensive characterization of Kir mRNA expression in Ae. aegypti and offer insights into the putative physiological roles of Kir subunits in this important disease vector. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. ML418: The First Selective, Sub-Micromolar Pore Blocker of Kir7.1 Potassium Channels.

    Science.gov (United States)

    Swale, Daniel R; Kurata, Haruto; Kharade, Sujay V; Sheehan, Jonathan; Raphemot, Rene; Voigtritter, Karl R; Figueroa, Eric E; Meiler, Jens; Blobaum, Anna L; Lindsley, Craig W; Hopkins, Corey R; Denton, Jerod S

    2016-07-20

    The inward rectifier potassium (Kir) channel Kir7.1 (KCNJ13) has recently emerged as a key regulator of melanocortin signaling in the brain, electrolyte homeostasis in the eye, and uterine muscle contractility during pregnancy. The pharmacological tools available for exploring the physiology and therapeutic potential of Kir7.1 have been limited to relatively weak and nonselective small-molecule inhibitors. Here, we report the discovery in a fluorescence-based high-throughput screen of a novel Kir7.1 channel inhibitor, VU714. Site-directed mutagenesis of pore-lining amino acid residues identified glutamate 149 and alanine 150 as essential determinants of VU714 activity. Lead optimization with medicinal chemistry generated ML418, which exhibits sub-micromolar activity (IC50 = 310 nM) and superior selectivity over other Kir channels (at least 17-fold selective over Kir1.1, Kir2.1, Kir2.2, Kir2.3, Kir3.1/3.2, and Kir4.1) except for Kir6.2/SUR1 (equally potent). Evaluation in the EuroFins Lead Profiling panel of 64 GPCRs, ion-channels, and transporters for off-target activity of ML418 revealed a relatively clean ancillary pharmacology. While ML418 exhibited low CLHEP in human microsomes which could be modulated with lipophilicity adjustments, it showed high CLHEP in rat microsomes regardless of lipophilicity. A subsequent in vivo PK study of ML418 by intraperitoneal (IP) administration (30 mg/kg dosage) revealed a suitable PK profile (Cmax = 0.20 μM and Tmax = 3 h) and favorable CNS distribution (mouse brain/plasma Kp of 10.9 to support in vivo studies. ML418, which represents the current state-of-the-art in Kir7.1 inhibitors, should be useful for exploring the physiology of Kir7.1 in vitro and in vivo.

  6. Opening of small and intermediate calcium-activated potassium channels induces relaxation mainly mediated by nitric-oxide release in large arteries and endothelium-derived hyperpolarizing factor in small arteries from rat

    DEFF Research Database (Denmark)

    Stankevicius, Edgaras; Dalsgaard, Thomas; Kroigaard, Christel

    2011-01-01

    This study was designed to investigate whether calcium-activated potassium channels of small (SK(Ca) or K(Ca)2) and intermediate (IK(Ca) or K(Ca)3.1) conductance activated by 6,7-dichloro-1H-indole-2,3-dione 3-oxime (NS309) are involved in both nitric oxide (NO) and endothelium-derived hyperpolar......This study was designed to investigate whether calcium-activated potassium channels of small (SK(Ca) or K(Ca)2) and intermediate (IK(Ca) or K(Ca)3.1) conductance activated by 6,7-dichloro-1H-indole-2,3-dione 3-oxime (NS309) are involved in both nitric oxide (NO) and endothelium...... in human umbilical vein endothelial cells (HUVECs), and calcium concentrations were investigated in both HUVECs and mesenteric arterial endothelial cells. In both superior (∼1093 μm) and small mesenteric (∼300 μm) arteries, NS309 evoked endothelium- and concentration-dependent relaxations. In superior....... In small mesenteric arteries, NS309 relaxations were reduced slightly by ADMA, whereas apamin plus an IK(Ca) channel blocker almost abolished relaxation. Iberiotoxin did not change NS309 relaxation. HUVECs expressed mRNA for SK(Ca) and IK(Ca) channels, and NS309 induced increases in calcium, outward...

  7. The inhibition of KCa3.1 channels activity reduces cell motility in glioblastoma derived cancer stem cells.

    Directory of Open Access Journals (Sweden)

    Paola Ruggieri

    Full Text Available In the present study we evaluated the expression of the intermediate conductance calcium-activated potassium (KCa3.1 channel in human glioblastoma stem-like cells (CSCs and investigated its role in cell motility. While the KCa3.1 channel is not expressed in neuronal- and glial-derived tissues of healthy individuals, both the KCa3.1 mRNA and protein are present in the glioblastoma tumor population, and are significantly enhanced in CSCs derived from both established cell line U87MG and a primary cell line, FCN9. Consistent with these data, voltage-independent and TRAM-34 sensitive potassium currents imputable to the KCa3.1 channel were recorded in the murine GL261 cell line and several primary human glioblastoma cells lines. Moreover, a significantly higher KCa3.1 current was recorded in U87MG-CD133 positive cells as compared to the U87MG-CD133 negative subpopulation. Further, we found that the tumor cell motility is strongly associated with KCa3.1 channel expression. Blockade of the KCa3.1 channel with the specific inhibitor TRAM-34 has in fact a greater impact on the motility of CSCs (reduction of 75%, which express a high level of KCa3.1 channel, than on the FCN9 parental population (reduction of 32%, where the KCa3.1 channel is expressed at lower level. Similar results were also observed with the CSCs derived from U87MG. Because invasion of surrounding tissues is one of the main causes of treatment failure in glioblastoma, these findings can be relevant for future development of novel cancer therapeutic drugs.

  8. The Mutation P.T613a in the Pore Helix of the Kv 11.1 Potassium Channel is Associated with Long Qt Syndrome

    DEFF Research Database (Denmark)

    Poulsen, Kristian L; Hotait, Mostafa; Calloe, Kirstine

    2015-01-01

    BACKGROUND: Loss-of-function mutations in the voltage gated potassium channel Kv 11.1 have been associated with the Long QT Syndrome (LQTS) type 2. We identified the p.T613A mutation in Kv 11.1 in a family with LQTS. T613A is located in the outer part of the pore helix, a structure that is involved...

  9. Expression of the Small Conductance Ca(2+)-Activated Potassium Channel Subtype 3 (SK3) in Rat Uterus after Stimulation with 17β-Estradiol

    DEFF Research Database (Denmark)

    Rahbek, Mette; Nazemi, Sasan; Odum, Lars

    2014-01-01

    of the myometrium. The small conductance Ca(2+)-activated potassium channel subtype 3 (SK3) has been identified in the myometrium of several species including humans, mice and rats, but with great inter species variation of the expression pattern and regulation. The aim of this study was to investigate...

  10. Activation of endothelial and epithelial K(Ca) 2.3 calcium-activated potassium channels by NS309 relaxes human small pulmonary arteries and bronchioles

    DEFF Research Database (Denmark)

    Kroigaard, Christel; Dalsgaard, Thomas; Nielsen, Gorm

    2012-01-01

    BACKGROUND AND PURPOSE: Small (K(Ca) 2) and intermediate (K(Ca) 3.1) conductance calcium-activated potassium channels (K(Ca) ) may contribute to both epithelium- and endothelium-dependent relaxations, but this has not been established in human pulmonary arteries and bronchioles. Therefore, we inv...... targets for treatment of pulmonary hypertension and chronic obstructive pulmonary disease....

  11. Influence of calcium-dependent potassium channel blockade and nitric oxide inhibition on norepinephrine-induced contractions in two forms of genetic hypertension

    Czech Academy of Sciences Publication Activity Database

    Líšková, Silvia; Petrová, M.; Karen, Petr; Kuneš, Jaroslav; Zicha, Josef

    2010-01-01

    Roč. 4, č. 3 (2010), s. 128-134 ISSN 1933-1711 R&D Projects: GA AV ČR(CZ) IAA500110902 Institutional research plan: CEZ:AV0Z50110509 Keywords : potassium channels * nitric oxide * norepinephrine Subject RIV: ED - Physiology Impact factor: 0.931, year: 2010

  12. Autoimmune encephalitis associated with voltage-gated potassium channels-complex and leucine-rich glioma-inactivated 1 antibodies - a national cohort study

    DEFF Research Database (Denmark)

    Celicanin, M; Blaabjerg, Morten; Maersk-Moller, C

    2017-01-01

    BACKGROUND AND PURPOSE: The aim of this study was to describe clinical and paraclinical characteristics of all Danish patients who tested positive for anti-voltage-gated potassium channels (VGKC)-complex, anti-leucine-rich glioma-inactivated 1 (LGI1) and anti-contactin-associated protein-2...

  13. K+ channels and cell cycle progression in tumor cells

    Directory of Open Access Journals (Sweden)

    HALIMA eOUADID-AHIDOUCH

    2013-08-01

    Full Text Available K+ ions play a major role in many cellular processes. The deregulation of K+ signaling is associated with a variety of diseases such as hypertension, atherosclerosis, or diabetes. K+ ions are important for setting the membrane potential, the driving force for Ca2+ influx, and regulate volume of growing cells. Moreover, it is increasingly recognized that K+ channels control cell proliferation through a novel signaling mechanisms triggered and modulated independently of ion fluxes. In cancer, aberrant expression, regulation and/or sublocalization of K+ channels can alter the downstream signals that converge on the cell cycle machinery. Various K+ channels are involved in cell cycle progression and are needed only at particular stages of the cell cycle. Consistent with this idea, the expression of Eag1 and HERG channels fluctuate along the cell cycle. Despite of acquired knowledge, our understanding of K+ channels functioning in cancer cells requires further studies. These include identifying the molecular mechanisms controling the cell cycle machinery. By understanding how K+ channels regulate cell cycle progression in cancer cells, we will gain insights into how cancer cells subvert the need for K+ signal and its downstream targets to proliferate.

  14. Frequency-dependent modulation of KCNQ1 and HERG1 potassium channels

    DEFF Research Database (Denmark)

    Diness, Thomas Goldin; Hansen, Rie Schultz; Olesen, Søren-Peter

    2006-01-01

    of the beta-subunits KCNE1 and KCNE2. In addition, the functional role of HERG1 in native guinea pig cardiac myocytes was demonstrated at different pacing frequencies by application of 10microM of the new HERG1 activator, NS1643. In conclusion, we have demonstrated that HERG1 and hKCNQ1 channels are inversely...

  15. Photobiomodulation on KATP Channels of Kir6.2-Transfected HEK-293 Cells

    Directory of Open Access Journals (Sweden)

    Fu-qing Zhong

    2014-01-01

    Full Text Available Background and Objective. ATP-sensitive potassium (KATP channel couples cell metabolism to excitability. To explore role of KATP channels in cellular photobiomodulation, we designed experiment to study effect of low intensity 808 nm laser irradiation on the activity of membrane KATP channel. Study Design/Materials and Methods. Plasmids encoding Kir6.2 was constructed and heterologously expressed in cultured mammalian HEK-293 cells. The patch-clamp and data acquisition systems were used to record KATP channel current before and after irradiation. A laser beam of Ga-As 808 nm at 5 mW/cm2 was used in experiments. A one-way ANOVA test followed by a post hoc Student-Newman-Keuls test was used to assess the statistical differences between data groups. Results. Obvious openings of KATP channels of Kir6.2-transfected HEK-293 cells and excised patches were recorded during and after low intensity 808 nm laser irradiation. Compared with the channels that did not undergo irradiation, open probability, current amplitude, and dwell time of KATP channels after irradiation improved. Conclusions. Low intensity 808 nm laser irradiation may activate membrane KATP channels of Kir6.2-transfected HEK-293 cells and in excised patches.

  16. Channels in cell membranes and synchrotron radiation

    International Nuclear Information System (INIS)

    Yan Xiaohui; Tian Liang; Zhang Xinyi

    2004-01-01

    For long time a lot of scientists have devoted to study how matter, such as water molecules and K + , Na + , Ca 2+ , Cl - ions, move through cell membranes and complete the matter exchange between the inside and outside of cells. Peter Agre discovered and characterized the first water channel protein in 1988 and Roderick MacKinnon elucidated the structural and mechanistic basis for ion channel function in 1998. These achievements have made it possible for us to 'see' these exquisitely designed molecular machines in action at the atomic level. The Nobel Prize in Chemistry for 2003 is shared between these two scientists. In determining the high resolution 3D structure of these channels, the synchrotron X-ray diffraction plays an important role

  17. Potassium Intake, Bioavailability, Hypertension, and Glucose Control

    Directory of Open Access Journals (Sweden)

    Michael S. Stone

    2016-07-01

    Full Text Available Potassium is an essential nutrient. It is the most abundant cation in intracellular fluid where it plays a key role in maintaining cell function. The gradient of potassium across the cell membrane determines cellular membrane potential, which is maintained in large part by the ubiquitous ion channel the sodium-potassium (Na+-K+ ATPase pump. Approximately 90% of potassium consumed (60–100 mEq is lost in the urine, with the other 10% excreted in the stool, and a very small amount lost in sweat. Little is known about the bioavailability of potassium, especially from dietary sources. Less is understood on how bioavailability may affect health outcomes. Hypertension (HTN is the leading cause of cardiovascular disease (CVD and a major financial burden ($50.6 billion to the US public health system, and has a significant impact on all-cause morbidity and mortality worldwide. The relationship between increased potassium supplementation and a decrease in HTN is relatively well understood, but the effect of increased potassium intake from dietary sources on blood pressure overall is less clear. In addition, treatment options for hypertensive individuals (e.g., thiazide diuretics may further compound chronic disease risk via impairments in potassium utilization and glucose control. Understanding potassium bioavailability from various sources may help to reveal how specific compounds and tissues influence potassium movement, and further the understanding of its role in health.

  18. Glucose- and interleukin-1beta-induced beta-cell apoptosis requires Ca2+ influx and extracellular signal-regulated kinase (ERK) 1/2 activation and is prevented by a sulfonylurea receptor 1/inwardly rectifying K+ channel 6.2 (SUR/Kir6.2) selective potassium channel opener in human islets

    DEFF Research Database (Denmark)

    Maedler, Kathrin; Størling, Joachim; Sturis, Jeppe

    2004-01-01

    Increasing evidence indicates that a progressive decrease in the functional beta-cell mass is the hallmark of both type 1 and type 2 diabetes. The underlying causes, beta-cell apoptosis and impaired secretory function, seem to be partly mediated by macrophage production of interleukin (IL)-1beta ...

  19. Rearrangement of potassium ions and Kv1.1/Kv1.2 potassium channels in regenerating axons following end-to-end neurorrhaphy: ionic images from TOF-SIMS.

    Science.gov (United States)

    Liu, Chiung-Hui; Chang, Hung-Ming; Wu, Tsung-Huan; Chen, Li-You; Yang, Yin-Shuo; Tseng, To-Jung; Liao, Wen-Chieh

    2017-10-01

    The voltage-gated potassium channels Kv1.1 and Kv1.2 that cluster at juxtaparanodal (JXP) regions are essential in the regulation of nerve excitability and play a critical role in axonal conduction. When demyelination occurs, Kv1.1/Kv1.2 activity increases, suppressing the membrane potential nearly to the equilibrium potential of K + , which results in an axonal conduction blockade. The recovery of K + -dependent communication signals and proper clustering of Kv1.1/Kv1.2 channels at JXP regions may directly reflect nerve regeneration following peripheral nerve injury. However, little is known about potassium channel expression and its relationship with the dynamic potassium ion distribution at the node of Ranvier during the regenerative process of peripheral nerve injury (PNI). In the present study, end-to-end neurorrhaphy (EEN) was performed using an in vivo model of PNI. The distribution of K + at regenerating axons following EEN was detected by time-of-flight secondary-ion mass spectrometry. The specific localization and expression of Kv1.1/Kv1.2 channels were examined by confocal microscopy and western blotting. Our data showed that the re-establishment of K + distribution and intensity was correlated with the functional recovery of compound muscle action potential morphology in EEN rats. Furthermore, the re-clustering of Kv1.1/1.2 channels 1 and 3 months after EEN at the nodal region of the regenerating nerve corresponded to changes in the K + distribution. This study provided direct evidence of K + distribution in regenerating axons for the first time. We proposed that the Kv1.1/Kv1.2 channels re-clustered at the JXP regions of regenerating axons are essential for modulating the proper patterns of K + distribution in axons for maintaining membrane potential stability after EEN.

  20. The marine algal toxin azaspiracid is an open state blocker of hERG potassium channels

    OpenAIRE

    Twiner, Michael J.; Doucette, Gregory J.; Rasky, Andrew; Huang, Xi-Ping; Roth, Bryan L.; Sanguinetti, Michael C.

    2012-01-01

    Azaspiracids (AZA) are polyether marine dinoflagellate toxins that accumulate in shellfish and represent an emerging human health risk. Although human exposure is primarily manifested by severe and protracted diarrhea, this toxin class has been shown to be highly cytotoxic, a teratogen to developing fish, and a possible carcinogen in mice. Until now, AZA's molecular target(s) has not yet been determined. Using three independent methods (voltage clamp, channel binding assay, and thallium flux ...

  1. Fluoxetine protection in decompression sickness in mice is enhanced by blocking TREK-1 potassium channel with the spadin antidepressant.

    Directory of Open Access Journals (Sweden)

    Nicolas eVallée

    2016-02-01

    Full Text Available In mice, disseminated coagulation, inflammation and ischemia induce neurological damages that can lead to the death. These symptoms result from circulating bubbles generated by a pathogenic decompression. An acute fluoxetine treatment or the presence of the TREK-1 potassium channel increased the survival rate when mice are subjected to an experimental dive/decompression protocol. This is a paradox because fluoxetine is a blocker of TREK-1 channels. First, we studied the effects of an acute dose of fluoxetine (50mg/kg in wild-type (WT and TREK-1 deficient mice (Knockout homozygous KO and heterozygous HET. Then, we combined the same fluoxetine treatment with a five-day treatment by spadin, in order to specifically block TREK-1 activity (KO-like mice. KO and KO-like mice could be regarded as antidepressed models.167 mice (45 WTcont 46 WTflux 30 HETflux and 46 KOflux constituting the flux-pool and 113 supplementary mice (27 KO-like 24 WTflux2 24 KO-likeflux 21 WTcont2 17 WTno dive constituting the spad-pool were included in this study. Only 7% of KO-TREK-1 treated with fluoxetine (KOflux and 4% of mice treated with both spadin and fluoxetine (KO-likeflux died from decompression sickness (DCS symptoms. These values are much lower than those of WT control (62% or KO-like mice (41%. After the decompression protocol, mice showed a significant consumption of their circulating platelets and leukocytes.Spadin antidepressed mice were more likely to declare DCS. Nevertheless, which had both blocked TREK-1 channel and were treated with fluoxetine were better protected against DCS. We conclude that the protective effect of such an acute dose of fluoxetine is enhanced when TREK-1 is inhibited. We confirmed that antidepressed models may have worse DCS outcomes, but a concomitant fluoxetine treatment not only decreases DCS severity but increases the survival rate.

  2. Altered Potassium Ion Channel Function as a Possible Mechanism of Increased Blood Pressure in Rats Fed Thermally Oxidized Palm Oil Diets.

    Science.gov (United States)

    Nkanu, Etah E; Owu, Daniel U; Osim, Eme E

    2017-12-27

    Intake of thermally oxidized palm oil leads to cytotoxicity and alteration of the potassium ion channel function. This study investigated the effects of fresh and thermally oxidized palm oil diets on blood pressure and potassium ion channel function in blood pressure regulation. Male Wistar rats were randomly divided into three groups of eight rats. Control group received normal feed; fresh palm oil (FPO) and thermally oxidized palm oil (TPO) groups were fed a diet mixed with 15% (weight/weight) fresh palm oil and five times heated palm oil, respectively, for 16 weeks. Blood pressure was measured; blood samples, hearts, and aortas were collected for biochemical and histological analyses. Thermally oxidized palm oil significantly elevated basal mean arterial pressure (MAP). Glibenclamide (10 -5 mmol/L) and tetraethylammonium (TEA; 10 -3 mmol/L) significantly raised blood pressure in TPO compared with FPO and control groups. Levcromakalim (10 -6 mmol/L) significantly (p palm oil increases MAP probably due to the attenuation of adenosine triphosphate-sensitive potassium (K ATP ) and large-conductance calcium-dependent potassium (BK Ca ) channels, tissue peroxidation, and altered histological structures of the heart and blood vessels.

  3. Relevance of lysine snorkeling in the outer transmembrane domain of small viral potassium ion channels.

    Science.gov (United States)

    Gebhardt, Manuela; Henkes, Leonhard M; Tayefeh, Sascha; Hertel, Brigitte; Greiner, Timo; Van Etten, James L; Baumeister, Dirk; Cosentino, Cristian; Moroni, Anna; Kast, Stefan M; Thiel, Gerhard

    2012-07-17

    Transmembrane domains (TMDs) are often flanked by Lys or Arg because they keep their aliphatic parts in the bilayer and their charged groups in the polar interface. Here we examine the relevance of this so-called "snorkeling" of a cationic amino acid, which is conserved in the outer TMD of small viral K(+) channels. Experimentally, snorkeling activity is not mandatory for Kcv(PBCV-1) because K29 can be replaced by most of the natural amino acids without any corruption of function. Two similar channels, Kcv(ATCV-1) and Kcv(MT325), lack a cytosolic N-terminus, and neutralization of their equivalent cationic amino acids inhibits their function. To understand the variable importance of the cationic amino acids, we reanalyzed molecular dynamics simulations of Kcv(PBCV-1) and N-terminally truncated mutants; the truncated mutants mimic Kcv(ATCV-1) and Kcv(MT325). Structures were analyzed with respect to membrane positioning in relation to the orientation of K29. The results indicate that the architecture of the protein (including the selectivity filter) is only weakly dependent on TMD length and protonation of K29. The penetration depth of Lys in a given protonation state is independent of the TMD architecture, which leads to a distortion of shorter proteins. The data imply that snorkeling can be important for K(+) channels; however, its significance depends on the architecture of the entire TMD. The observation that the most severe N-terminal truncation causes the outer TMD to move toward the cytosolic side suggests that snorkeling becomes more relevant if TMDs are not stabilized in the membrane by other domains.

  4. Potassium channels modulate the action but not the synthesis of hydrogen sulfide in rat corpus cavernosum.

    Science.gov (United States)

    Abd Elmoneim, H; Sharabi, F; Mohy El Din, M; Louedec, L; Norel, X; Senbel, A

    2017-11-15

    Hydrogen sulfide (H 2 S) is a newly-introduced gasotransmitter in penile tissues. However, its exact mechanism of action in mediating penile erection is not fully elucidated. The major aim of this study was to examine the role of different K + channels in mediating the responses to H 2 S in the corpus cavernosum. Tension studies using isolated rat corpus cavernosum strips were conducted. Endogenous H 2 S production was measured using polarographic technique. Results are expressed as mean±SEM. l-Cysteine (10 -2 M) stimulated rat corpus cavernosum to produce H 2 S. Blockade of CSE by BCA (10 -3 M) reduced the concentration of H 2 S produced from rat corpus cavernosum significantly. Addition of TEA (10 -2 M) or 4-AP (10 -3 M) didn't have a significant effect on the concentration of H 2 S produced. l-Cysteine (10 -6 -10 -2 M) elicited a concentration-dependent relaxation response which was significantly reduced by blockade of CSE using BCA (10 -3 M). TEA (10 -2 M), 4-AP (10 -3 M) and TEA (10 -4 M) attenuated l-cysteine-induced relaxation significantly. At 10 -4 M, l-cysteine resulted in percentage relaxation of 1.55±0.63, 10.94±1.93 and 1.93±0.80 in presence of TEA (10 -2 M), 4-AP (10 -3 M) and TEA (10 -4 M) respectively compared to 23.78±2.71 as control. Both glibenclamide (10 -5 M) and BaCl 2 (3×10 -5 M) failed to reduce these relaxations significantly. H 2 S-induced relaxation of rat corpus cavernosum may be mediated - at least in part - through BK ca and K V channels not by K ATP and K ir channels. It also seems that K + -channels do not contribute to the synthesis of H 2 S. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Genomic and structural characterization of Kunitz-type peptide LmKTT-1a highlights diversity and evolution of scorpion potassium channel toxins.

    Directory of Open Access Journals (Sweden)

    Zongyun Chen

    Full Text Available BACKGROUND: Recently, a new subfamily of long-chain toxins with a Kunitz-type fold was found in scorpion venom glands. Functionally, these toxins inhibit protease activity and block potassium channels. However, the genomic organization and three-dimensional (3-D structure of this kind of scorpion toxin has not been reported. PRINCIPAL FINDINGS: Here, we characterized the genomic organization and 3-D nuclear magnetic resonance structure of the scorpion Kunitz-type toxin, LmKTT-1a, which has a unique cysteine pattern. The LmKTT-1a gene contained three exons, which were interrupted by two introns located in the mature peptide region. Despite little similarity to other Kunitz-type toxins and a unique pattern of disulfide bridges, LmKTT-1a possessed a conserved Kunitz-type structural fold with one α-helix and two β-sheets. Comparison of the genomic organization, 3-D structure, and functional data of known toxins from the α-KTx, β-KTx, γ-KTx, and κ-KTx subfamily suggested that scorpion Kunitz-type potassium channel toxins might have evolved from a new ancestor that is completely different from the common ancestor of scorpion toxins with a CSα/β fold. Thus, these analyses provide evidence of a new scorpion potassium channel toxin subfamily, which we have named δ-KTx. CONCLUSIONS/SIGNIFICANCE: Our results highlight the genomic, structural, and evolutionary diversity of scorpion potassium channel toxins. These findings may accelerate the design and development of diagnostic and therapeutic peptide agents for human potassium channelopathies.

  6. Metamizol acts as an ATP sensitive potassium channel opener to inhibit the contracting response induced by angiotensin II but not to norepinephrine in rat thoracic aorta smooth muscle.

    Science.gov (United States)

    Valenzuela, Fermín; García-Saisó, Sebastián; Lemini, Cristina; Ramírez-Solares, Rafael; Vidrio, Horacio; Mendoza-Fernández, Víctor

    2005-08-01

    Clinically metamizol (MZ) has been related to alteration on haemodynamic parameters and modifications on blood pressure in humans when administered intravenously. These effects have been observed at MZ therapeutic doses. Experimentally, MZ is able to induce relaxation on several types of vascular smooth muscles and modulates the contraction induced by phenylephrine. However, the mechanism underlying the MZ effects on vascular reactivity is not clear. Potassium channels (K) present on vascular smooth muscle cells closely regulate the vascular reactivity and membrane potential. There are four described types of K in vascular tissue: K voltage sensitive (K(V)), K calcium sensitive (K(Ca)2+), K ATP sensitive (K(ATP) and K inward rectification (K(IR), voltage sensitive). The aim of this work was to investigate MZ effects on angiotensin II (AT II) and noradrenaline (NA) induced contraction and to evaluate the K participation on MZ modulating effect on vascular smooth muscle contraction, using isometric and patch clamp techniques. MZ induces relaxation in a concentration dependent manner. Furthermore, MZ strongly inhibits in a concentration dependent fashion the contraction induced by AT II. However, MZ inhibition on NA induced contraction was moderated compared with that observed on AT II. MZ effects on AT II induced contraction was blocked by glybenclamide (a specific K(ATP) blocker, 3 microM, *p < 0.01). In patch clamp experiments, MZ (3 mM) induces an increase on potassium current (K+) mediated by K(ATP) in similar way as diazoxide (a specific K(ATP) opener, 3 microM). Our results suggest that MZ induces relaxation and inhibits contraction induced by AT II acting as a K(ATP) opener.

  7. The Barium Site in a Potassium Channel by X-Ray Crystallography

    Science.gov (United States)

    Jiang, Youxing; MacKinnon, Roderick

    2000-01-01

    X-ray diffraction data were collected from frozen crystals (100°K) of the KcsA K+ channel equilibrated with solutions containing barium chloride. Difference electron density maps (Fbarium − Fnative, 5.0 Å resolution) show that Ba2+ resides at a single location within the selectivity filter. The Ba2+ blocking site corresponds to the internal aspect (adjacent to the central cavity) of the “inner ion” position where an alkali metal cation is found in the absence of the blocking Ba2+ ion. The location of Ba2+ with respect to Rb+ ions in the pore is in good agreement with the findings on the functional interaction of Ba2+ with K+ (and Rb+) in Ca2+-activated K+ channels (Neyton, J., and C. Miller. 1988. J. Gen. Physiol. 92:549–567). Taken together, these structural and functional data imply that at physiological ion concentrations a third ion may interact with two ions in the selectivity filter, perhaps by entering from one side and displacing an ion on the opposite side. PMID:10694255

  8. Effects of Potassium-Channel Opener on Thallium-201 Kinetics: In-vitro Study in Rat Myocyte Preparations and In-vivo Mice Biodistribution Study

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jae Tae; Kim, Eun Ji; Ahn, Byeong Cheol; Son, Kang Kyun; Lee, Kyu Bo [Kyungpook National University School of Medicine, Taegu (Korea, Republic of); Ha, Jeoung Hee [Youngnam University Medical School, Taegu (Korea, Republic of); Kim, Chun Ki [Mt. Sinai School of Medicine, New York (United States)

    1996-10-15

    Potassium channel opener (K-opener) opens ATP-sensitive K{sup +}-channel located at membrane and induces potassium efflux from cytosol, resulting in intracellular hyperpolarization. Newly synthesized K-opener is currently examined for pharmacologic potency by means of rubidium release test from smooth muscle strip preincubated with Rb-86. Since in-vive behavior of thallium is similar to that of rubidium, we hypothesized that K-opener can alter T1-201 kinetics in vivo. This study was prepared to investigate the effects of pinacidil (one of potent K-openers) on the T1-201 uptake and clearance in cultured myocyte, and in-vivo biodistribution in mice. Spontaneous contracting myocytes were prepared to imitate in-vivo condition from 20 hearts of 3-5 days old Sprague-Dawley rat and cultured for 3-5 days before use (5 X 105 cells/ml). Pinacidil was dissolved in 10% DMSO solution at a final concentration of 100nM or 10uM and was co-incubated with T1-201 in HBSS buffer for 20-min to evaluate its effect on cellular T1-uptake, or challenged to cell preparation pre-incubated with T1-201 for washout study. Two, 40 or 100 mg of pinacidil was injected intravenously into ICR mice at 10 min after 5 muCi T1-201 injection, and organ uptake and whole body retention rate were measured at different time points. Co-incubation of pinacidil with T1-201 resulted in a decrease in T1-201 uptake into cultured myocyte by 1.6 to 2.5 times, depending on pinacidil concentration and activity of T1-201 used. Pinacidil enhanced T1-201 washout by 1.6-3.1 times from myocyte preparations pre-incubated with T1-201. Pinacidil treatment appears to be resulted in mild decreases in blood and liver activity in normal mice, in contrast, renal and cardiac uptake were mildly decreased in a dose dependent manner. Whole body retention ratios of T1-201 were lower at 24 hour after injection with 100 mg of pinacidil than control. These results suggest that treatment with K-opener may affect the interpretation of T1

  9. Interaction of C-70 fullerene with the Kv1.2 potassium channel

    DEFF Research Database (Denmark)

    Monticelli, L.; Barnoud, J.; Orlowskid, A.

    2012-01-01

    Fullerene C-70 is known to partition into lipid membranes and change their physical properties. Together with gallic acid (GA), C-70 induces cell contraction and cell death. How C-70 and GA-induced perturbations of lipid membranes affect cellular function and membrane protein activity...

  10. Dysfunction of the heteromeric KV7.3/KV7.5 potassium channel is associated with autism spectrum disorders

    DEFF Research Database (Denmark)

    Nielsen, Mette Gilling; Rasmussen, Hanne Borger; Callø, Kirstine

    2013-01-01

    ) translocation truncating KCNQ3 in a boy with childhood autism. In addition, we identified a c.1720C¿>¿T [p.P574S] nucleotide change in three unrelated individuals with childhood autism and no history of convulsions. This nucleotide change was previously reported in patients with rolandic epilepsy or IGE and has.......3/2, KV7.3/4, or KV7.3/5 channels in HEK 293 cells or primary rat hippocampal neurons. Our results suggest that dysfunction of the heteromeric KV7.3/5 channel is implicated in the pathogenesis of some forms of autism spectrum disorders, epilepsy, and possibly other psychiatric disorders and therefore...

  11. Adrenaline reveals the torsadogenic effect of combined blockade of potassium channels in anaesthetized guinea pigs.

    Science.gov (United States)

    Michael, G; Kane, K A; Coker, S J

    2008-08-01

    Torsade de pointes (TdP) can be induced in several species by a reduction in cardiac repolarizing capacity. The aim of this study was to assess whether combined I(Kr) and I(Ks) blockade could induce TdP in anaesthetized guinea pigs and whether short-term variability (STV) or triangulation of action potentials could predict TdP. Experiments were performed in open-chest, pentobarbital-anaesthetized, adrenaline-stimulated male Dunkin Hartley guinea pigs, which received three consecutive i.v. infusions of either vehicle, the I(Kr) blocker E-4031 (3, 10 and 30 nmol kg(-1) min(-1)), the I(Ks) blocker HMR1556 (75, 250, 750 nmol kg(-1) min(-1)) or E-4031 and HMR1556 combined. Phenylephrine-stimulated guinea pigs were also treated with the K(+) channel blockers in combination. Arterial blood pressure, ECGs and epicardial monophasic action potential (MAP) were recorded. TdP was observed in 75% of adrenaline-stimulated guinea pigs given the K(+) channel blockers in combination, but was not observed in guinea pigs treated with either I(K) blocker alone, or in phenylephrine-stimulated guinea pigs. Salvos and ventricular tachycardia occurred with adrenaline but not with phenylephrine. No changes in STV or triangulation of the MAP signals were observed before TdP. Combined blockade of both I(Kr) and I(Ks) plus the addition of adrenaline were required to induce TdP in anaesthetized guinea pigs. This suggests that there must be sufficient depletion of repolarization reserve and an appropriate trigger for TdP to occur.

  12. Investigating the potassium interactions with the palytoxin induced channels in Na+/K+ pump.

    Science.gov (United States)

    Rodrigues, Antônio M; Almeida, Antônio-Carlos G; Infantosi, Antonio F C; Teixeira, Hewerson Z; Duarte, Mário A

    2009-02-01

    K(+) has been appointed as the main physiological inhibitor of the palytoxin (PTX) effect on the Na(+)/K(+) pump. This toxin acts opening monovalent cationic channels through the Na(+)/K(+) pump. We investigate, by means of computational modeling, the kinetic mechanisms related with K(+) interacting with the complex PTX-Na(+)/K(+) pump. First, a reaction model, with structure similar to Albers-Post model, describing the functional cycle of the pump, was proposed for describing K(+) interference on the complex PTX-Na(+)/K(+) pump in the presence of intracellular ATP. A mathematic model was derived from the reaction model and it was possible to solve numerically the associated differential equations and to simulate experimental maneuvers about the PTX induced currents in the presence of K(+) in the intra- and extracellular space as well as ATP in the intracellular. After the model adjusting to the experimental data, a Monte Carlo method for sensitivity analysis was used to analyze how each reaction parameter acts during each experimental maneuver involving PTX. For ATP and K(+) concentrations conditions, the simulations suggest that the enzyme substate with ATP bound to its high-affinity sites is the main substate for the PTX binding. The activation rate of the induced current is limited by the K(+) deocclusion from the PTX-Na(+)/K(+) pump complex. The K(+) occlusion in the PTX induced channels in the enzymes with ATP bound to its low-affinity sites is the main mechanism responsible for the reduction of the enzyme affinity to PTX.

  13. Blockade of the voltage-gated potassium channel Kv1.3 inhibits immune responses in vivo.

    Science.gov (United States)

    Koo, G C; Blake, J T; Talento, A; Nguyen, M; Lin, S; Sirotina, A; Shah, K; Mulvany, K; Hora, D; Cunningham, P; Wunderler, D L; McManus, O B; Slaughter, R; Bugianesi, R; Felix, J; Garcia, M; Williamson, J; Kaczorowski, G; Sigal, N H; Springer, M S; Feeney, W

    1997-06-01

    The voltage activated K+ channel (Kv1.3) has recently been identified as the molecule that sets the resting membrane potential of peripheral human T lymphoid cells. In vitro studies indicate that blockage of Kv1.3 inhibits T cell activation, suggesting that Kv1.3 may be a target for immunosuppression. However, despite the in vitro evidence, there has been no in vivo demonstration that blockade of Kv1.3 will attenuate an immune response. The difficulty is due to species differences, as the channel does not set the membrane potential in rodent peripheral T cells. In this study, we show that the channel is present on peripheral T cells of miniswine. Using the peptidyl Kv1.3 inhibitor, margatoxin, we demonstrate that Kv1.3 also regulates the resting membrane potential, and that blockade of Kv1.3 inhibits, in vivo, both a delayed-type hypersensitivity reaction and an Ab response to an allogeneic challenge. In addition, prolonged Kv1.3 blockade causes reduced thymic cellularity and inhibits the thymic development of T cell subsets. These results provide in vivo evidence that Kv1.3 is a novel target for immunomodulation.

  14. Cooperation between epithelial cells demonstrated by potassium transfer

    International Nuclear Information System (INIS)

    Ledbetter, M.L.; Young, G.J.; Wright, E.R.

    1986-01-01

    Junction-mediated communication can be measured in fibroblast cultures by determining the ability of mixed cultures of cells sensitive and resistant to ouabain to concentrate K+ in the presence of ouabain. We now report the extension of this assay procedure to cultured epithelial cells. Hamster kidney (HaK) cells maintain their ability to concentrate K+ in ouabain at levels inhibitory to dog kidney (MDCK) cells. When HaK and MDCK cells were cultured together in ouabain-containing medium, the K+ (measured as 86Rb+) in the mixed population was greater than expected if the cells were not interacting. The degree of enhancement, expressed as index of cooperation, depended on the numbers of cells in the cultures, their opportunity for cell-to-cell contact, and (above a certain permissive level) the concentration of ouabain. As with other cell types, protein synthesis in MDCK cells depends on maintenance of cell K+. Autoradiography of cells incubated with [3H]leucine demonstrated that MDCK cells in ouabain-treated mixed cultures were able to synthesize proteins only when physically adjacent to HaK cells. The transmission of labeled nucleosides among the cells provides independent evidence of the phenomenon of cooperation, probably mediated by gap junctions. This system offers promise for investigation of stimuli modulating junctional communication

  15. MicroRNA-Mediated Downregulation of the Potassium Channel Kv4.2 Contributes to Seizure Onset

    Directory of Open Access Journals (Sweden)

    Christina Gross

    2016-09-01

    Full Text Available Seizures are bursts of excessive synchronized neuronal activity, suggesting that mechanisms controlling brain excitability are compromised. The voltage-gated potassium channel Kv4.2, a major mediator of hyperpolarizing A-type currents in the brain, is a crucial regulator of neuronal excitability. Kv4.2 expression levels are reduced following seizures and in epilepsy, but the underlying mechanisms remain unclear. Here, we report that Kv4.2 mRNA is recruited to the RNA-induced silencing complex shortly after status epilepticus in mice and after kainic acid treatment of hippocampal neurons, coincident with reduction of Kv4.2 protein. We show that the microRNA miR-324-5p inhibits Kv4.2 protein expression and that antagonizing miR-324-5p is neuroprotective and seizure suppressive. MiR-324-5p inhibition also blocks kainic-acid-induced reduction of Kv4.2 protein in vitro and in vivo and delays kainic-acid-induced seizure onset in wild-type but not in Kcnd2 knockout mice. These results reveal an important role for miR-324-5p-mediated silencing of Kv4.2 in seizure onset.

  16. Subclinical Doses of ATP-Sensitive Potassium Channel Modulators Prevent Alterations in Memory and Synaptic Plasticity Induced by Amyloid-β.

    Science.gov (United States)

    Salgado-Puga, Karla; Rodríguez-Colorado, Javier; Prado-Alcalá, Roberto A; Peña-Ortega, Fernando

    2017-01-01

    In addition to coupling cell metabolism and excitability, ATP-sensitive potassium channels (KATP) are involved in neural function and plasticity. Moreover, alterations in KATP activity and expression have been observed in Alzheimer's disease (AD) and during amyloid-β (Aβ)-induced pathology. Thus, we tested whether KATP modulators can influence Aβ-induced deleterious effects on memory, hippocampal network function, and plasticity. We found that treating animals with subclinical doses (those that did not change glycemia) of a KATP blocker (Tolbutamide) or a KATP opener (Diazoxide) differentially restrained Aβ-induced memory deficit, hippocampal network activity inhibition, and long-term synaptic plasticity unbalance (i.e., inhibition of LTP and promotion of LTD). We found that the protective effect of Tolbutamide against Aβ-induced memory deficit was strong and correlated with the reestablishment of synaptic plasticity balance, whereas Diazoxide treatment produced a mild protection against Aβ-induced memory deficit, which was not related to a complete reestablishment of synaptic plasticity balance. Interestingly, treatment with both KATP modulators renders the hippocampus resistant to Aβ-induced inhibition of hippocampal network activity. These findings indicate that KATP are involved in Aβ-induced pathology and they heighten the potential role of KATP modulation as a plausible therapeutic strategy against AD.

  17. Orientations and proximities of the extracellular ends of transmembrane helices S0 and S4 in open and closed BK potassium channels.

    Directory of Open Access Journals (Sweden)

    Xiaowei Niu

    Full Text Available The large-conductance potassium channel (BK α subunit contains a transmembrane (TM helix S0 preceding the canonical TM helices S1 through S6. S0 lies between S4 and the TM2 helix of the regulatory β1 subunit. Pairs of Cys were substituted in the first helical turns in the membrane of BK α S0 and S4 and in β1 TM2. One such pair, W22C in S0 and W203C in S4, was 95% crosslinked endogenously. Under voltage-clamp conditions in outside-out patches, this crosslink was reduced by DTT and reoxidized by a membrane-impermeant bis-quaternary ammonium derivative of diamide. The rate constants for this reoxidation were not significantly different in the open and closed states of the channel. Thus, these two residues are approximately equally close in the two states. In addition, 90% crosslinking of a second pair, R20C in S0 and W203C in S4, had no effect on the V50 for opening. Taken together, these findings indicate that separation between residues at the extracellular ends of S0 and S4 is not required for voltage-sensor activation. On the contrary, even though W22C and W203C were equally likely to form a disulfide in the activated and deactivated states, relative immobilization by crosslinking of these two residues favored the activated state. Furthermore, the efficiency of recrosslinking of W22C and W203C on the cell surface was greater in the presence of the β1 subunit than in its absence, consistent with β1 acting through S0 to stabilize its immobilization relative to α S4.

  18. Voltage-gated potassium channel antibody limbic encephalitis: a case illustrating the neuropsychiatric and PET/CT features with clinical and imaging follow-up.

    Science.gov (United States)

    Celliers, Liesl; Hung, Te-Jui; Al-Ogaili, Zeyad; Moschilla, Girolamo; Knezevic, Wally

    2016-12-01

    To illustrate the neuropsychiatric and imaging findings in a confirmed case of voltage-gated potassium channel antibody limbic encephalitis. Case report and review of the literature. A 64-year-old man presented with several months' history of obsessive thoughts and compulsions associated with faciobrachial dystonic seizures. He had no significant past medical and psychiatric history. Physical examinations revealed only mildly increased tone in the left upper limb. Bedside cognitive testing was normal. Positron-emission tomography showed intense symmetrical uptake in the corpus striatum. No underlying malignancy was identified on whole body imaging. Magnetic resonance imaging, lumbar puncture and electroencephalogram were normal. Serum voltage-gated potassium channel antibodies were strongly positive. The patient had a favourable response to antiepileptic drugs, oral steroids and immunotherapy. Voltage-gated potassium channel limbic encephalitis characteristically presents with neuropsychiatric symptoms and temporal lobe seizures. Positron-emission tomography-computed tomography can be a useful adjunct to the clinical and biochemical work-up. © The Royal Australian and New Zealand College of Psychiatrists 2016.

  19. Potassium as a pluripotency-associated element identified through inorganic element profiling in human pluripotent stem cells.

    Science.gov (United States)

    Lin, Victor J T; Zolekar, Ashwini; Shi, Yi; Koneru, Bhuvaneswari; Dimitrijevich, Slobodan; Di Pasqua, Anthony J; Wang, Yu-Chieh

    2017-07-10

    Despite their well-known function in maintaining normal cell physiology, how inorganic elements are relevant to cellular pluripotency and differentiation in human pluripotent stem cells (hPSCs) has yet to be systematically explored. Using total reflection X-ray fluorescence (TXRF) spectrometry and inductively coupled plasma mass spectrometry (ICP-MS), we analyzed the inorganic components of human cells with isogenic backgrounds in distinct states of cellular pluripotency. The elemental profiles revealed that the potassium content of human cells significantly differs when their cellular pluripotency changes. Pharmacological treatment that alters cell membrane permeability to potassium affected the maintenance and establishment of cellular pluripotency via multiple mechanisms in bona fide hPSCs and reprogrammed cells. Collectively, we report that potassium is a pluripotency-associated inorganic element in human cells and provide novel insights into the manipulation of cellular pluripotency in hPSCs by regulating intracellular potassium.

  20. ATP-sensitive K(+-channels in muscle cells: features and physiological role

    Directory of Open Access Journals (Sweden)

    O. B. Vadzyuk

    2014-08-01

    Full Text Available ATP-sensitive K+-channels of plasma membranes belong to the inward rectifier potassium channels type. They are involved in coupling of electrical activity of muscle cell with its metabolic­ state. These channels are heterooctameric and consist of two types of subunits: four poreforming (Kir 6.х and four regulatory (SUR, sulfonylurea receptor. The Kir subunits contain highly selective K+ filter and provide for high-velocity K+ currents. The SUR subunits contain binding sites for activators and blockers and have metabolic sensor, which enables channel activation under conditions of metabolic stress. ATP blocks K+ currents through the ATP-sensitive K+-channels in the most types of muscle cells. However, functional activity of these channels does not depend on absolute concentration of ATP but on the АТР/ADP ratio and presence of Mg2+. Physiologically active substances, such as phosphatidylinositol bisphosphate and fatty acid esters can regulate the activity of these structures in muscle cells. Activation of these channels under ischemic conditions underlies their cytoprotective action, which results in prevention of Ca2+ overload in cytosol. In contrast to ATP-sensitive K+-channels of plasma membranes, the data regarding the structure and function of ATP-sensitive K+-channels of mitochondrial membrane are contradictory. Pore-forming subunits of this channel have not been firmly identified yet. ATP-sensitive K+ transport through the mitochondrial­ membrane is easily tested by different methods, which are briefly reviewed in this paper. Interaction of mitoKATP with physiological and pharmacological ligands is discussed as well.

  1. The Role of the Two-Pore Domain Potassium Channel TREK-1 in the Therapeutic Effects of Escitalopram in a Rat Model of Poststroke Depression.

    Science.gov (United States)

    Lin, Dai-Hua; Zhang, Xiang-Rong; Ye, Dong-Qing; Xi, Guang-Jun; Hui, Jiao-Jie; Liu, Shan-Shan; Li, Lin-Jiang; Zhang, Zhi-Jun

    2015-06-01

    Poststroke depression (PSD) is one of the most common neuropsychiatric complications after stroke. TREK-1, a two-pore-domain potassium channel, has been implicated in the pathogenesis of stroke and depression. The aim of this study was to investigate whether TREK-1 plays a role in the therapeutic effects of the selective serotonin reuptake inhibitor (SSRI) escitalopram in a rat PSD model. The whole-cell patch-clamp technique was performed to assess the effect of escitalopram on recombinant TREK-1 currents in HEK293 cells. The expression of TREK-1 mRNA and protein was measured in the hippocampus and prefrontal cortex (PFC), and neural stem cell (NSC) proliferation was detected in the hippocampal dentate gyrus (DG) in PSD rats after 3 weeks of escitalopram administration. Escitalopram reversibly inhibited TREK-1 currents in a concentration-dependent manner. Chronic treatment with escitalopram significantly reversed the reductions in weight gain, locomotor activity, and sucrose preference in PSD rats. The expressions of TREK-1 mRNA and protein were significantly increased in hippocampal CA1, CA3, DG, and PFC in PSD rats, with the exception of TREK-1 mRNA in hippocampal CA1. NSC proliferation was significantly decreased in hippocampal DG of PSD rats. Escitalopram significantly reversed the regional increases of TREK-1 expression and the reduction of hippocampal NSC proliferation in PSD rats. TREK-1 plays an important role in the therapeutic effects of the SSRI escitalopram in PSD model, making TREK-1 an attractive candidate molecule for further understanding the pathophysiology and treatment of PSD. © 2015 John Wiley & Sons Ltd.

  2. Tumor necrosis factor α modulates sodium-activated potassium channel SLICK in rat dorsal horn neurons via p38 MAPK activation pathway

    Directory of Open Access Journals (Sweden)

    Wang K

    2017-05-01

    Full Text Available Kun Wang,1 Feng Wang,1 Jun-Ping Bao,2 Zhi-Yang Xie,1 Lu Chen,1 Bao-Yi Zhou,1 Xin-Hui Xie,2 Xiao-Tao Wu1,2 1Medical School of Southeast University, 2Department of Orthopaedics, Zhongda Hospital, Southeast University, Nanjing, People’s Republic of China Abstract: The dorsal horn (DH of the spinal cord is the integrative center that processes and transmits pain sensation. Abnormal changes in ion channel expression can enhance the excitability of pain-related DH neurons. Sodium-activated potassium (KNa channels are highly expressed particularly in the central nervous system; however, information about whether rat DH neurons express the SLICK channel protein is lacking, and the direct effects on SLICK in response to inflammation and the potential signaling pathway mediating such effects are yet to be elucidated. Here, using cultured DH neurons, we have shown that tumor necrosis factor-α inhibits the total outward potassium current IK and the KNa current predominantly as well as induces a progressive loss of firing accommodation. However, we found that this change in channel activity is offset by the p38 inhibitor SB202190, thereby suggesting the modulation of SLICK channel activity via the p38 MAPK pathway. Furthermore, we have demonstrated that the tumor necrosis factor-α modulation of KNa channels does not occur at the level of SLICK channel gating but arises from possible posttranslational modification. Keywords: p38 MAPK, SLICK channel, neuropathic pain, dorsal horn, TNF-α

  3. Hyperkalemia caused by rapid red cell transfusion and the potassium absorption filter

    Directory of Open Access Journals (Sweden)

    Yasuhiko Imashuku

    2017-01-01

    Full Text Available We report a case of transient hyperkalemia during hysterectomy after cesarean section, due to preoperatively undiagnosed placenta accreta that caused unforeseen massive hemorrhage and required rapid red cell transfusion. Hyperkalemia-induced by rapid red cell transfusion is a well-known severe complication of transfusion; however, in patients with sudden massive hemorrhage, rapid red cell transfusion is necessary to save their life. In such cases, it is extremely important to monitor serum potassium levels. For an emergency situation, a system should be developed to ensure sufficient preparation for immediate transfusion and laboratory tests. Furthermore, sufficient stock of preparations to treat hyperkalemia, such as calcium preparations, diuretics, glucose, and insulin is required. Moreover, a transfusion filter that absorbs potassium has been developed and is now available for clinical use in Japan. The filter is easy to use and beneficial, and should be prepared when it is available.

  4. Negative inotropic effect of carbachol and interaction between acetylcholine receptor-operated potassium channel (K.ACh channel) and GTP binding protein in mouse isolated atrium--a novel methodological trial.

    Science.gov (United States)

    Okada, Muneyoshi; Noma, Chihiro; Yamawaki, Hideyuki; Hara, Yukio

    2013-01-01

    Interaction between acetylcholine receptor-operated potassium channel (K.ACh channel) and GTP binding protein was examined by an immunoprecipitation-Western blotting system in mouse isolated atrium. The carbachol-induced negative inotropic action in indomethacin-pretreated mouse atrium was significantly inhibited by a K.ACh channel blocker, tertiapin or atropine. Kir3.1 K.ACh channel (Kir3.1) was immunoprecipitated with a mouse anti-Kir3.1 antibody. Coprecipitating Gβ with Kir3.1, detected by Western blotting, was significantly augmented by carbachol. Atropine, but not tertiapin, significantly inhibited the carbachol-induced coprecipitating Gβ with Kir3.1. The data indicate that immunoprecipitation with Kir3.1 and Western blotting of Gβ system is a useful method for assessing interaction between K.ACh channel and GTP binding protein in mouse atrium.

  5. The Caenorhabditis elegans iodotyrosine deiodinase ortholog SUP-18 functions through a conserved channel SC-box to regulate the muscle two-pore domain potassium channel SUP-9.

    Directory of Open Access Journals (Sweden)

    Ignacio Perez de la Cruz

    2014-02-01

    Full Text Available Loss-of-function mutations in the Caenorhabditis elegans gene sup-18 suppress the defects in muscle contraction conferred by a gain-of-function mutation in SUP-10, a presumptive regulatory subunit of the SUP-9 two-pore domain K(+ channel associated with muscle membranes. We cloned sup-18 and found that it encodes the C. elegans ortholog of mammalian iodotyrosine deiodinase (IYD, an NADH oxidase/flavin reductase that functions in iodine recycling and is important for the biosynthesis of thyroid hormones that regulate metabolism. The FMN-binding site of mammalian IYD is conserved in SUP-18, which appears to require catalytic activity to function. Genetic analyses suggest that SUP-10 can function with SUP-18 to activate SUP-9 through a pathway that is independent of the presumptive SUP-9 regulatory subunit UNC-93. We identified a novel evolutionarily conserved serine-cysteine-rich region in the C-terminal cytoplasmic domain of SUP-9 required for its specific activation by SUP-10 and SUP-18 but not by UNC-93. Since two-pore domain K(+ channels regulate the resting membrane potentials of numerous cell types, we suggest that the SUP-18 IYD regulates the activity of the SUP-9 channel using NADH as a coenzyme and thus couples the metabolic state of muscle cells to muscle membrane excitability.

  6. Associations of two-pore domain potassium channels and triple negative breast cancer subtype in The Cancer Genome Atlas: systematic evaluation of gene expression and methylation.

    Science.gov (United States)

    Dookeran, Keith A; Zhang, Wei; Stayner, Leslie; Argos, Maria

    2017-09-12

    It is unclear whether 2-pore domain potassium channels are novel molecular markers with differential expression related to biologically aggressive triple-negative type breast tumors. Our objective was to systematically evaluate associations of 2-pore domain potassium channel gene expression and DNA methylation with triple-negative subtype in The Cancer Genome Atlas invasive breast cancer dataset. Methylation and expression data for all fifteen 2-pore domain potassium family genes were examined for 1040 women, and associations with triple-negative subtype (vs. luminal A) were evaluated using age/race adjusted generalized-linear models, with Bonferroni-corrected significance thresholds. Subtype associated CpG loci were evaluated for functionality related to expression using Spearman's correlation. Overexpression of KCNK5, KCNK9 and KCNK12, and underexpression of KCNK6 and KCNK15, were significantly associated with triple-negative subtype (Bonferroni-corrected p < 0.0033). A total of 195 (114 hypomethylated and 81 hypermethylated) CpG loci were found to be significantly associated with triple-negative subtype (Bonferroni-corrected p < 8.22 × 10 -8 ). Significantly negatively correlated expression patterns that were differentially observed in triple-negative vs. luminal A subtype were demonstrated for: KCNK2 (gene body: cg04923840, cg13916421), KCNK5 (gene body: cg05255811, cg18705155, cg09130674, cg21388745, cg00859574) and KCNK9 (TSS1500: cg21415530, cg12175729; KCNK9/TRAPPC9 intergenic region: cg17336929, cg25900813, cg03919980). CpG loci listed for KCNK5 and KCNK9 all showed relative hypomethylation for probability of triple-negative vs. luminal A subtype. Triple-negative subtype was associated with distinct 2-pore domain potassium channel expression patterns. Both KCNK5 and KCNK9 overexpression appeared to be functionally related to CpG loci hypomethylation.

  7. KCNQ1 channels sense small changes in cell volume

    DEFF Research Database (Denmark)

    Grunnet, Morten; Jespersen, Thomas; MacAulay, Nanna

    2003-01-01

    . Incubation of the oocytes in cytochalasin D and experiments with truncated KCNQ1 channels suggest that KCNQ1 channels sense cell volume changes through interactions between the cytoskeleton and the N-terminus of the channel protein. From our results we propose that KCNQ1 and KCNQ4 channels play an important......Many important physiological processes involve changes in cell volume, e.g. the transport of salt and water in epithelial cells and the contraction of cardiomyocytes. In this study, we show that voltage-gated KCNQ1 channels, which are strongly expressed in epithelial cells or cardiomyocytes......, and KCNQ4 channels, expressed in hair cells and the auditory tract, are tightly regulated by small cell volume changes when co-expressed with aquaporin 1 water-channels (AQP1) in Xenopus oocytes. The KCNQ1 and KCNQ4 current amplitudes precisely reflect the volume of the oocytes. By contrast, the related...

  8. The conserved potassium channel filter can have distinct ion binding profiles: structural analysis of rubidium, cesium, and barium binding in NaK2K.

    Science.gov (United States)

    Lam, Yee Ling; Zeng, Weizhong; Sauer, David Bryant; Jiang, Youxing

    2014-08-01

    Potassium channels are highly selective for K(+) over the smaller Na(+). Intriguingly, they are permeable to larger monovalent cations such as Rb(+) and Cs(+) but are specifically blocked by the similarly sized Ba(2+). In this study, we used structural analysis to determine the binding profiles for these permeant and blocking ions in the selectivity filter of the potassium-selective NaK channel mutant NaK2K and also performed permeation experiments using single-channel recordings. Our data revealed that some ion binding properties of NaK2K are distinct from those of the canonical K(+) channels KcsA and MthK. Rb(+) bound at sites 1, 3, and 4 in NaK2K, as it does in KcsA. Cs(+), however, bound predominantly at sites 1 and 3 in NaK2K, whereas it binds at sites 1, 3, and 4 in KcsA. Moreover, Ba(2+) binding in NaK2K was distinct from that which has been observed in KcsA and MthK, even though all of these channels show similar Ba(2+) block. In the presence of K(+), Ba(2+) bound to the NaK2K channel at site 3 in conjunction with a K(+) at site 1; this led to a prolonged block of the channel (the external K(+)-dependent Ba(2+) lock-in state). In the absence of K(+), however, Ba(2+) acts as a permeating blocker. We found that, under these conditions, Ba(2+) bound at sites 1 or 0 as well as site 3, allowing it to enter the filter from the intracellular side and exit from the extracellular side. The difference in the Ba(2+) binding profile in the presence and absence of K(+) thus provides a structural explanation for the short and prolonged Ba(2+) block observed in NaK2K. © 2014 Lam et al.

  9. Blockade of ATP-sensitive potassium channels prevents the attenuation of the exercise pressor reflex by tempol in rats with ligated femoral arteries.

    Science.gov (United States)

    Yamauchi, Katsuya; Stone, Audrey J; Stocker, Sean D; Kaufman, Marc P

    2012-08-01

    We reported previously that tempol attenuated the exercise pressor and muscle mechanoreceptor reflexes in rats whose femoral arteries were ligated, whereas tempol did not attenuate these reflexes in rats whose femoral arteries were freely perfused. Although the mechanism whereby tempol attenuated these reflexes in rats whose femoral artery was ligated was independent of its ability to scavenge reactive oxygen species, its nature remains unclear. An alternative explanation for the tempol-induced attenuation of these reflexes involves ATP-sensitive potassium channels (K(ATP)) and calcium-activated potassium channels (BK(Ca)), both of which are opened by tempol. We tested the likelihood of this explanation by measuring the effects of either glibenclamide (0.1 mg/kg), which blocks K(ATP) channels, or iberiotoxin (20 or 40 μg/kg), which blocks BK(Ca) channels, on the tempol-induced attenuation of the exercise pressor and muscle mechanoreceptor reflexes in decerebrated rats whose femoral arteries were ligated. We found that glibenclamide prevented the tempol-induced attenuation of both reflexes, whereas iberiotoxin did not. We also found that the amount of protein comprising the pore of the K(ATP) channel in the dorsal root ganglia innervating hindlimbs whose femoral artery was ligated was significantly greater than that in the dorsal root ganglia innervating hindlimbs whose femoral arteries were freely perfused. In contrast, the amounts of protein comprising the BK(Ca) channel in the dorsal root ganglia innervating the ligated and freely perfused hindlimbs were not different. We conclude that tempol attenuated both reflexes by opening K(ATP) channels, an effect that hyperpolarized muscle afferents stimulated by static contraction or tendon stretch.

  10. The mechanism of gentisic acid-induced relaxation of the guinea pig isolated trachea: the role of potassium channels and vasoactive intestinal peptide receptors.

    Science.gov (United States)

    Cunha, J F; Campestrini, F D; Calixto, J B; Scremin, A; Paulino, N

    2001-03-01

    We examined some of the mechanisms by which the aspirin metabolite and the naturally occurring metabolite gentisic acid induced relaxation of the guinea pig trachea in vitro. In preparations with or without epithelium and contracted by histamine, gentisic acid caused concentration-dependent and reproducible relaxation, with mean EC(50) values of 18 microM and E(max) of 100% (N = 10) or 20 microM and E(max) of 92% (N = 10), respectively. The relaxation caused by gentisic acid was of slow onset in comparison to that caused by norepinephrine, theophylline or vasoactive intestinal peptide (VIP). The relative rank order of potency was: salbutamol 7.9 > VIP 7.0 > gentisic acid 4.7 > theophylline 3.7. Gentisic acid-induced relaxation was markedly reduced (24 +/- 7.0, 43 +/- 3.9 and 78 +/- 5.6%) in preparations with elevated potassium concentration in the medium (20, 40 or 80 mM, respectively). Tetraethylammonium (100 microM), a nonselective blocker of the potassium channels, partially inhibited the relaxation response to gentisic acid, while 4-AP (10 microM), a blocker of the voltage potassium channel, inhibited gentisic acid-induced relaxation by 41 +/- 12%. Glibenclamide (1 or 3 microM), at a concentration which markedly inhibited the relaxation induced by the opener of ATP-sensitive K(+) channels, levcromakalim, had no effect on the relaxation induced by gentisic acid. Charybdotoxin (0.1 or 0.3 microM), a selective blocker of the large-conductance Ca(2+)-activated K(+) channels, caused rightward shifts (6- and 7-fold) of the gentisic acid concentration-relaxation curve. L-N(G)-nitroarginine (100 microM), a NO synthase inhibitor, had no effect on the relaxant effect of gentisic acid, and caused a slight displacement to the right in the relaxant effect of the gentisic acid curve at 300 microM, while methylene blue (10 or 30 microM) or ODQ (1 microM), the inhibitors of soluble guanylate cyclase, all failed to affect gentisic acid-induced relaxation. D-(P)-Cl-Phe(6),Leu(17

  11. Effects of calcium channel on ovarian cancer cells

    OpenAIRE

    Zhang, Chunyun; Li, Hailing

    2017-01-01

    The aim of the present study was to examine the effects of calcium channel protein on ovarian cancer cells. The expression of calcium channel protein in normal ovarian cells and ovarian cancer cells was detected by fluorescence quantitative PCR. Subsequently, the ovarian cancer cells were added to calcium channel protein activator media at various concentrations of 0, 1, 4, 8, 12, 16 and 20 mmol/l. The concentration of calcium ion in different samples was produced, and using an MTT assay, ova...

  12. Cortical effect of oxaliplatin associated with sustained neuropathic pain: exacerbation of cortical activity and down-regulation of potassium channel expression in somatosensory cortex.

    Science.gov (United States)

    Thibault, Karine; Calvino, Bernard; Dubacq, Sophie; Roualle-de-Rouville, Marie; Sordoillet, Vallier; Rivals, Isabelle; Pezet, Sophie

    2012-08-01

    Oxaliplatin is a third-generation platinum-based chemotherapy drug that has gained importance in the treatment of advanced metastatic colorectal cancer. Its dose-limiting side effect is the production of chronic peripheral neuropathy. Using a modified model of oxaliplatin-induced sensory neuropathy, we investigated plastic changes at the cortical level as possible mechanisms underlying the chronicity of pain sensation in this model. Changes in gene expression were studied using DNA microarray which revealed that when oxaliplatin-treated animals displayed clinical neuropathic pain symptoms, including mechanical and thermal hypersensitivity, approximately 900 were down-regulated in the somatosensory cortex. Because of the known role of potassium channels in neuronal excitability, the study further focussed on the down-regulation of these channels as the possible molecular origin of cortical hyperexcitability. Quantification of the magnitude of neuronal extracellular signal-regulated kinase (ERK) phosphorylation in cortical neurons as a marker of neuronal activity revealed a 10-fold increase induced by oxaliplatin treatment, suggesting that neurons of cortical areas involved in transmission of painful stimuli undergo a chronic cortical excitability. We further demonstrated, using cortical injection of lentiviral vector shRNA against Kv2.2, that down-regulation of this potassium channel in naive animals induced a sustained thermal and mechanical hypersensitivity. In conclusion, although the detailed mechanisms leading to this cortical excitability are still unknown, our study demonstrated that a cortical down regulation of potassium channels could underlie pain chronicity in this model of chemotherapy-induced neuropathic pain. Copyright © 2012 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  13. Potassium Channel Ether à go-go1 Is Aberrantly Expressed in Human Liposarcoma and Promotes Tumorigenesis

    Directory of Open Access Journals (Sweden)

    Jin Wu

    2014-01-01

    Full Text Available The ether à go-go1 (Eag1 channel is overexpressed in a variety of cancers. However, the expression and function of Eag1 in liposarcoma are poorly understood. In the present study, the mRNA expression of Eag1 in different adipose tissue samples was examined by real-time PCR. Then, the protein expression of Eag1 in 131 different adipose tissues from 109 patients was detected by immunohistochemistry. Next, the associations between Eag1 expression and clinicopathological features of liposarcoma were analyzed. In addition, the effects of Eag1 on liposarcoma cell proliferation and cycle were evaluated by CCK-8, colony formation, xenograft mouse model, and flow cytometry, respectively. Finally, the activation of p38 mitogen-activated protein kinase (MAPK was detected by Western blot analysis to explain the detailed mechanisms of oncogenic potential of Eag1 in liposarcoma. It was found that Eag1 was aberrantly expressed in over 67% liposarcomas, with a higher frequency than in lipoma, hyperplasia, inflammation, and normal adipose tissues. However, Eag1 expression was not correlated with clinicopathological features of liposarcoma. Eag1 inhibitor imipramine or Eag1-shRNA significantly suppressed the proliferation of liposarcoma cells in vitro and in vivo, accompanying with accumulation of cells in the G1 phase. These results suggest that Eag1 plays an important role in regulating the proliferation and cell cycle of liposarcoma cells and might be a potential therapeutic target for liposarcoma.

  14. Potassium ion influx measurements on cultured Chinese hamster cells exposed to 60-hertz electromagnetic fields

    International Nuclear Information System (INIS)

    Stevenson, A.P.; Tobey, R.A.

    1985-01-01

    Potassium ion influx was measured by monitoring 42 KCl uptake by Chinese hamster ovary (CHO) cells grown in suspension culture and exposed in the culture medium to 60-Hz electromagnetic fields up to 2.85 V/m. In the presence of the field CHO cells exhibited two components of uptake, the same as previously observed for those grown under normal conditions; both these components of influx were decreased when compared to sham-exposed cells. Although decreases were consistently observed in exposed cells when plotted as loge of uptake, the differences between the means of the calculated fluxes of exposed and sham-exposed cells were quite small (on the order of 4-7%). When standard deviations were calculated, there was no significant difference between these means; however, when time-paired uptake data were analyzed, the differences were found to be statistically significant. Cells exposed only to the magnetic field exhibited similar small decreases in influx rates when compared to sham-exposed cells, suggesting that the reduction in K+ uptake could be attributed to the magnetic field. Additionally, intracellular K+ levels were measured over a prolonged exposure period (96 h), and no apparent differences in intracellular K+ levels were observed between field-exposed and sham-exposed cultures. These results indicate that high-strength electric fields have a small effect on the rate of transport of potassium ions but no effect on long-term maintenance of intracellular K+

  15. Nitric oxide does not mediate promotion of cellular potassium release by phenolphthalein in COS-7 cells.

    Science.gov (United States)

    Mason, Robert W; Hopp, Laszlo; Lloyd, John B

    2004-04-01

    1. It has been proposed that phenolphthalein exerts its laxative effect via an intracellular cascade that begins with the activation of nitric oxide synthase (NOS) and ends with an inhibition of NaCl and water reabsorption from the colon. Phenolphthalein also promotes the release of potassium from cells, but it is not known how this is related to its effect on sodium and water uptake. 2. An established in vitro system was used to examine the role of nitric oxide (NO) in phenolphthalein-induced release of (86)Rb(+) from COS-7 cells. 3. Sodium nitroprusside, an NOS-independent NO source, was unable to mimic the effects of phenolphthalein and N(G)-nitro-L-arginine methyl ester, an NOS inhibitor, was unable to block the effect of phenolphthalein. 4. It is concluded that NO generation is not required for phenolphthalein-stimulated potassium release. It is proposed that the effect of phenolphthalein on cellular potassium release is mechanistically distinct from the effect on NaCl and water uptake by colonocytes.

  16. Ion Channels in Hematopoietic and Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Serena Pillozzi

    2012-01-01

    Full Text Available Hematopoietic stem cells (HSCs reside in bone marrow niches and give rise to hematopoietic precursor cells (HPCs. These have more restricted lineage potential and eventually differentiate into specific blood cell types. Bone marrow also contains mesenchymal stromal cells (MSCs, which present multilineage differentiation potential toward mesodermal cell types. In bone marrow niches, stem cell interaction with the extracellular matrix is mediated by integrin receptors. Ion channels regulate cell proliferation and differentiation by controlling intracellular Ca2+, cell volume, release of growth factors, and so forth. Although little evidence is available about the ion channel roles in true HSCs, increasing information is available about HPCs and MSCs, which present a complex pattern of K+ channel expression. K+ channels cooperate with Ca2+ and Cl− channels in regulating calcium entry and cell volume during mitosis. Other K+ channels modulate the integrin-dependent interaction between leukemic progenitor cells and the niche stroma. These channels can also regulate leukemia cell interaction with MSCs, which also involves integrin receptors and affects the MSC-mediated protection from chemotherapy. Ligand-gated channels are also implicated in these processes. Nicotinic acetylcholine receptors regulate cell proliferation and migration in HSCs and MSCs and may be implicated in the harmful effects of smoking.

  17. Clinical presentation of anti-N-methyl-d-aspartate receptor and anti-voltage-gated potassium channel complex antibodies in children: A series of 24 cases.

    Science.gov (United States)

    Konuskan, Bahadir; Yildirim, Mirac; Topaloglu, Haluk; Erol, Ilknur; Oztoprak, Ulkuhan; Tan, Huseyin; Gocmen, Rahsan; Anlar, Banu

    2018-01-01

    The symptomatology and paraclinical findings of antibody-mediated encephalitis, a relatively novel disorder, are still being characterized in adults and children. A high index of suspicion is needed in order to identify these cases among children presenting with various neurological symptoms. The aim of this study is to examine the clinical, demographic and laboratory findings and outcome of children with anti-NMDAR and anti-VGKC encephalitis for any typical or distinctive features. Cases diagnosed with anti-N-Methyl d-aspartate receptor (NMDAR) and anti-voltage gated potassium channel (VGKC) antibody-mediated encephalopathy in four major child neurology centers are described. In four years, 16 children with NMDAR and 8 children with VGKC antibody-associated disease were identified in the participating centers. The most frequent initial manifestation consisted of generalized seizures and cognitive symptoms in both groups. Movement abnormalities were frequent in anti-NMDAR patients and autonomic symptoms, in anti-VGKC patients. Cerebrospinal fluid (CSF) protein, cell count and IgG index were normal in 9/15 anti-NMDAR and 5/8 anti-VGKC patients tested. EEG and MRI findings were usually nonspecific and non-contributory. The rate and time of recovery was not related to age, sex, acute or subacute onset, antibody type, MRI, EEG or CSF results. Treatment within 3 months of onset was associated with normal neurological outcome. Our results suggest anti-NMDAR and VGKC encephalopathies mostly present with non-focal neurological symptoms longer than 3 weeks. In contrast with adult cases, routine CSF testing, MRI and EEG did not contribute to the diagnosis in this series. Copyright © 2017 European Paediatric Neurology Society. Published by Elsevier Ltd. All rights reserved.

  18. Cardiovascular action of insulin in health and disease: focus in endothelial L-arginine transport and cardiac voltage-dependent potassium channels.

    Directory of Open Access Journals (Sweden)

    Sebastián eDubó

    2016-03-01

    Full Text Available The impairment of insulin signaling on diabetes mellitus has been related to cardiovascular dysfunction, heart failure and sudden death. In human endothelium, cationic amino acid transporter 1 (hCAT-1 is related to the synthesis of nitric oxide (NO. Insulin has a vascular effect in endothelial cells through a signaling pathway that involved increases of hCAT-1 expression and L-arginine transport. This mechanism is disrupted in diabetes, a phenomenon potentiated by excessive accumulation of reactive oxygen species (ROS, which contributes to lower availability of NO and endothelial dysfunction. On the other hand, the electrical remodeling in cardiomyocytes is considered a key factor in heart failure progression associated to diabetes mellitus, generating a challenge to understand the specific role of insulin and the pathways involved in cardiac function. Studies on isolated mammalian cardiomyocytes have shown a prolongated action potential in ventricular repolarization phase that produces a long QT interval. The long QT generated is well explained by attenuation in the repolarizing potassium currents in cardiac ventricles. The impaired insulin signaling causes specific changes in these currents, such a decrease amplitude of the transient outward K+ (Ito and the ultra-rapid delayed rectifier (IKur currents where, together, a reduction of mRNA and protein expression levels of α-subunits (Ito, fast; Kv 4.2 and IKs; Kv 1.5 or β-subunits (KChIP2 and MiRP of K+ channels involved in these currents in a MAPK mediated pathway process have been described. These results support the hypothesis that the lack of insulin signaling can produce an abnormal repolarization in cardiomyocytes. Furthermore, the arrhythmogenic potential due to reduced Ito current can contribute to an increase in the incidence of sudden death in heart failure. This review aims to show, based on pathophysiological models, the regulatory function that would have insulin in vascular

  19. Potassium conductances mediate bidirectional state-dependent modulation of action potential evoked dendritic calcium signals in dentate gyrus granule cells

    Directory of Open Access Journals (Sweden)

    János Brunner

    2014-03-01

    Full Text Available Backpropagating action potentials (bAPs and local calcium signals that they trigger are fundamental for dendritic functions. Here we addressed the question what extent the changes of local dendritic membrane properties can contribute to the shaping of the coupling between dendritic action potentials and the local calcium responses. Using a combination of in vitro electrophysiological and confocal imaging techniques we found that activation of dendritic GIRK channels via mGlu2 or GABAB receptors enhanced the bAP¬-triggered calcium signals in the dendrites of dentate gyrus granule cells (GCs. The enhancement of calcium signals was significant only in those dendritic regions, where these receptors are predominantly expressed. Similarly to GIRK channel activation, somatic hyperpolarization by DC current injection (from -64 mV to -77 mV, significantly increased bAP-associated calcium signals in the proximal dendrites. The hyperpolarization was associated with a decrease in the input resistance due to the rectification of the membrane potential of GCs. The effect of hyperpolarization on the calcium signals was maintained when T-type calcium currents were blocked but it decreased when GIRK channels were inhibited. Simultaneous dual somato-dendritic recordings from GCs showed that somatic hyperpolarization accelerated the repolarization phase of dendritic bAP in the proximal region whereas the rising phase and peak amplitude was not affected. We hypothesize that the larger driving force for calcium ions during the faster repolarization can contribute to the increasing in calcium signals. Employment of previously recorded dendritic bAP waveforms from hyperpolarized membrane potential as voltage command evoked larger calcium currents in nucleated patches compared to bAP waveform from the same recording at depolarized membrane potential. Furthermore, addition of native, high-voltage activated, inactivating potassium conductance by somatic dynamic clamp

  20. Effect of ouabain, digoxin and digitoxigenin on potassium uptake and histamine release from rat peritoneal mast cells

    DEFF Research Database (Denmark)

    Knudsen, T; Ferjan, I; Johansen, Torben

    1993-01-01

    Rat peritoneal mast cells were used to study the effects of digitalis glycosides on potassium uptake and histamine release induced by compound 48/80, substance P and egg-albumin (immunological release). In the absence of calcium all glycosides inhibited potassium uptake. Ouabain and digoxin...... enhanced the histamine release while digitoxigenin either had no effect or was slightly inhibitory. In the presence of calcium, the glycosides only affected potassium uptake and histamine release slightly. In the presence of lithium or lanthanum the enhancement of the histamine release was counteracted....... Hydrophilic digitalis glycosides seem to enhance histamine release secondary to an increase in intracellular sodium. Lipophilic glycosides have no effect on the release....

  1. Oxygen-glucose deprivation enhancement of cell death/apoptosis in PC12 cells and hippocampal neurons correlates with changes in neuronal excitatory amino acid neurotransmitter signaling and potassium currents.

    Science.gov (United States)

    Wang, Yu-Xiang; Zhang, Feng; Ma, Xue-Ling; He, Cong-Cong; Tian, Kun; Wang, Hong-Gang; An, Di; Heng, Bin; Xie, Lai-Hua; Liu, Yan-Qiang

    2016-05-25

    Neuronal death is a pathophysiological process that is often caused by hypoxia/ischemia. However, the causes of hypoxia/ischemia-induced neuronal death are debated, and additional experimental data are needed to resolve this debate. In the present study, we applied oxygen-glucose deprivation (OGD) to PC12 cells and hippocampal neurons to establish a hypoxia/ischemia model. We evaluated the effects of OGD on cell death/apoptosis and on the levels of two excitatory amino acid neurotransmitters, aspartic acid and glutamic acid, in both hippocampal neurons and the medium used to culture the hippocampal neurons. We also evaluated GluR2 expression in hippocampal neurons as well as the effects of OGD on whole-cell potassium currents in PC12 cells and hippocampal neurons. Our experimental results showed that OGD significantly decreased cell viability and markedly enhanced apoptosis in PC12 cells and hippocampal neurons. OGD treatment for 3 h increased the levels of Asp and Glu in the medium used to culture hippocampal neurons, but decreased both the levels of Asp and Glu and GluR2 expression in hippocampal neurons. Furthermore, OGD altered the electrophysiological properties of voltage-dependent potassium channels in PC12 cells and hippocampal neurons in different ways; OGD decreased the voltage-dependent potassium current in PC12 cells, but increased this current in hippocampal neurons. On the basis of these results, we concluded that OGD enhanced neuronal cell death/apoptosis in addition to altering neuronal excitatory amino acid neurotransmitter signaling and whole-cell voltage-dependent potassium currents.

  2. Intracellular mediators of potassium-induced aldosterone secretion

    International Nuclear Information System (INIS)

    Ganguly, A.; Chiou, S.; Davis, J.S.

    1990-01-01

    We have investigated the intracellular messengers of potassium in eliciting aldosterone secretion in calf adrenal glomerulosa cells since there were unresolved issues relating to the role of phosphoinositides, cAMP and protein kinases. We observed no evidence of hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP 2 ) in 3 H-inositol labeled alf adrenal cells or increase of cAMP in response to potassium. Addition of calcium channel blocker, nitrendipine after stimulating adrenal glomerulosa cells with potassium, markedly inhibited aldosterone secretion. A calmodulin inhibitor (W-7) produced greater reduction of aldosterone secretion than an inhibitor of protein kinase C (H-7). These results suggest that a rise in cytosolic free calcium concentration through voltage-dependent calcium channel and calmodulin are the critical determinants of aldosterone secretion stimulated by potassium

  3. Ionic channel changes in glaucomatous retinal ganglion cells: multicompartment modeling.

    Science.gov (United States)

    Maturana, Matias I; Turpin, Andrew; McKendrick, Allison M; Kameneva, Tatiana

    2014-01-01

    This research takes a step towards discovering underlying ionic channel changes in the glaucomatous ganglion cells. Glaucoma is characterized by a gradual death of retinal ganglion cells. In this paper, we propose a hypothesis that the ionic channel concentrations change during the progression of glaucoma. We use computer simulation of a multi-compartment morphologically correct model of a mouse retinal ganglion cell to verify our hypothesis. Using published experimental data, we alter the morphology of healthy ganglion cells to replicate glaucomatous cells. Our results suggest that in glaucomatous cell, the sodium channel concentration decreases in the soma by 30% and by 60% in the dendrites, calcium channel concentration decreases by 10% in all compartments, and leak channel concentration increases by 40% in the soma and by 100% in the dendrites.

  4. Tungstate-Targeting of BKαβ1 Channels Tunes ERK Phosphorylation and Cell Proliferation in Human Vascular Smooth Muscle

    OpenAIRE

    López López, José Ramón; Fernández Mariño, Ana Isabel; Cidad, Pilar; Zafra, Delia; Nocito, Laura; Domínguez, Jorge; Oliván Viguera, Aida; Köhler, Ralf; Pérez García, María Teresa; Valverde, Miguel Ángel; Guinovart, Joan J.; Fernández Fernández, José Manuel

    2015-01-01

    Producción Científica Despite the substantial knowledge on the antidiabetic, antiobesity and antihypertensive actions of tungstate, information on its primary target/s is scarce. Tungstate activates both the ERK1/2 pathway and the vascular voltage- and Ca2+-dependent large-conductance BKαβ1 potassium channel, which modulates vascular smooth muscle cell (VSMC) proliferation and function, respectively. Here, we have assessed the possible involvement of BKαβ1 channels in the tungstate-induced...

  5. Potassium Channelopathies and Gastrointestinal Ulceration.

    Science.gov (United States)

    Han, Jaeyong; Lee, Seung Hun; Giebisch, Gerhard; Wang, Tong

    2016-11-15

    Potassium channels and transporters maintain potassium homeostasis and play significant roles in several different biological actions via potassium ion regulation. In previous decades, the key revelations that potassium channels and transporters are involved in the production of gastric acid and the regulation of secretion in the stomach have been recognized. Drugs used to treat peptic ulceration are often potassium transporter inhibitors. It has also been reported that potassium channels are involved in ulcerative colitis. Direct toxicity to the intestines from nonsteroidal anti-inflammatory drugs has been associated with altered potassium channel activities. Several reports have indicated that the long-term use of the antianginal drug Nicorandil, an adenosine triphosphate-sensitive potassium channel opener, increases the chances of ulceration and perforation from the oral to anal regions throughout the gastrointestinal (GI) tract. Several of these drug features provide further insights into the role of potassium channels in the occurrence of ulceration in the GI tract. The purpose of this review is to investigate whether potassium channelopathies are involved in the mechanisms responsible for ulceration that occurs throughout the GI tract.

  6. Modeling two-phase flow in PEM fuel cell channels

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yun; Basu, Suman; Wang, Chao-Yang [Electrochemical Engine Center (ECEC), and Department of Mechanical and Nuclear Engineering, The Pennsylvania State University, University Park, PA 16802 (United States)

    2008-05-01

    This paper is concerned with the simultaneous flow of liquid water and gaseous reactants in mini-channels of a proton exchange membrane (PEM) fuel cell. Envisaging the mini-channels as structured and ordered porous media, we develop a continuum model of two-phase channel flow based on two-phase Darcy's law and the M{sup 2} formalism, which allow estimate of the parameters key to fuel cell operation such as overall pressure drop and liquid saturation profiles along the axial flow direction. Analytical solutions of liquid water saturation and species concentrations along the channel are derived to explore the dependences of these physical variables vital to cell performance on operating parameters such as flow stoichiometric ratio and relative humility. The two-phase channel model is further implemented for three-dimensional numerical simulations of two-phase, multi-component transport in a single fuel-cell channel. Three issues critical to optimizing channel design and mitigating channel flooding in PEM fuel cells are fully discussed: liquid water buildup towards the fuel cell outlet, saturation spike in the vicinity of flow cross-sectional heterogeneity, and two-phase pressure drop. Both the two-phase model and analytical solutions presented in this paper may be applicable to more general two-phase flow phenomena through mini- and micro-channels. (author)

  7. The Peptide Toxin Amylosin of Bacillus amyloliquefaciens from Moisture-Damaged Buildings Is Immunotoxic, Induces Potassium Efflux from Mammalian Cells, and Has Antimicrobial Activity

    Science.gov (United States)

    Teplova, Vera V.; Andersson, Maria A.; Mikkola, Raimo; Kankkunen, Päivi; Matikainen, Sampsa; Gahmberg, Carl G.; Andersson, Leif C.; Salkinoja-Salonen, Mirja

    2015-01-01

    Amylosin, a heat-stable channel-forming non-ribosomally synthesized peptide toxin produced by strains of Bacillus amyloliquefaciens isolated from moisture-damaged buildings, is shown in this paper to have immunotoxic and cytotoxic effects on human cells as well as antagonistic effects on microbes. Human macrophages exposed to 50 ng of amylosin ml−1 secreted high levels of cytokines interleukin-1β (IL-1β) and IL-18 within 2 h, indicating activation of the NLRP3 inflammasome, an integral part of the innate immune system. At the same exposure level, expression of IL-1β and IL-18 mRNA increased. Amylosin caused dose-dependent potassium ion efflux from all tested mammalian cells (human monocytes and keratinocytes and porcine sperm cells) at 1 to 2 μM exposure. Amylosin also inhibited the motility of porcine sperm cells and depolarized the mitochondria of human keratinocytes. Amylosin may thus trigger the activation of the NLRP3 inflammasome and subsequently cytokine release by causing potassium efflux from exposed cells. The results of this study indicate that exposure to amylosin activates the innate immune system, which could offer an explanation for the inflammatory symptoms experienced by occupants of moisture-damaged buildings. In addition, the amylosin-producing B. amyloliquefaciens inhibited the growth of both prokaryotic and eukaryotic indoor microbes, and purified amylosin also had an antimicrobial effect. These antimicrobial effects could make amylosin producers dominant and therefore significant causal agents of health problems in some moisture-damaged sites. PMID:25681192

  8. Outward-Rectifying K^+ Channels in Stomatal Guard Cell Protoplasts

    OpenAIRE

    Shigeru, Hosoi; Moritoshi, Iino; Ken-ichiro, Shimazaki; Tsukuba Research Laboratory, Hamamatsu Photonics; Department of Biology, Tokyo Metropolitan University; Division of Environmental Biology, National Institute for Environmental Studies

    1988-01-01

    Ion channels in stomatal guard cell protoplasts from Vicia faba were examined using the patch-clamp technique. Most ion channels having unit conductance ranging between 10 and 30 pS showed clear outward-rectification in symmetrical 50 mM KCl. The large inside-out membranes contained these outward-rectifiers as the major and relatively stable channels. The channels were K^+ ion-selective. Kinetic analysis revealed that the channels have three conductance states: open, closed and inactivated. T...

  9. Loss of ATP-Sensitive Potassium Channel Surface Expression in Heart Failure Underlies Dysregulation of Action Potential Duration and Myocardial Vulnerability to Injury.

    Directory of Open Access Journals (Sweden)

    Zhan Gao

    Full Text Available The search for new approaches to treatment and prevention of heart failure is a major challenge in medicine. The adenosine triphosphate-sensitive potassium (KATP channel has been long associated with the ability to preserve myocardial function and viability under stress. High surface expression of membrane KATP channels ensures a rapid energy-sparing reduction in action potential duration (APD in response to metabolic challenges, while cellular signaling that reduces surface KATP channel expression blunts APD shortening, thus sacrificing energetic efficiency in exchange for greater cellular calcium entry and increased contractile force. In healthy hearts, calcium/calmodulin-dependent protein kinase II (CaMKII phosphorylates the Kir6.2 KATP channel subunit initiating a cascade responsible for KATP channel endocytosis. Here, activation of CaMKII in a transaortic banding (TAB model of heart failure is coupled with a 35-40% reduction in surface expression of KATP channels compared to hearts from sham-operated mice. Linkage between KATP channel expression and CaMKII is verified in isolated cardiomyocytes in which activation of CaMKII results in downregulation of KATP channel current. Accordingly, shortening of monophasic APD is slowed in response to hypoxia or heart rate acceleration in failing compared to non-failing hearts, a phenomenon previously shown to result in significant increases in oxygen consumption. Even in the absence of coronary artery disease, failing myocardium can be further injured by ischemia due to a mismatch between metabolic supply and demand. Ischemia-reperfusion injury, following ischemic preconditioning, is diminished in hearts with CaMKII inhibition compared to wild-type hearts and this advantage is largely eliminated when myocardial KATP channel expression is absent, supporting that the myocardial protective benefit of CaMKII inhibition in heart failure may be substantially mediated by KATP channels. Recognition of Ca

  10. The effect of diamide on potassium transport and cellular morphology in mouse L-cells

    International Nuclear Information System (INIS)

    Szekely, J.G.; Sargent, M.D.; Copps, T.P.; Lobreau, A.U.

    1982-06-01

    The effects of diamide (diazenedicarboxylic acid bis (N,N'-dimethylamide)) on the transport of potassium in mouse L-cells have been investigated using 86 Rb + as a tracer. Active, ouabain-sensitive uptake is reduced after 0.4 to 0.6 mol/L diamide treatment. The size of reduction depends on the temperature and the presence of glucose in the medium. These results suggest that the elimination of reduced glutathione by diamide is the major factor controlling the level of K + transport in treated L-cells. In addition to decreasing active transport, diamide produces dramatic changes in cellular ultrastructure, probably through altered Na + /K + balance and its action on tubulin. Clear organelle-free regions appear surrounded by vacuoles and swollen mitkchondria regions. The clear areas of cytoplasm eventually pinch off from the cell

  11. The mechanism of gentisic acid-induced relaxation of the guinea pig isolated trachea: the role of potassium channels and vasoactive intestinal peptide receptors

    Directory of Open Access Journals (Sweden)

    J.F. Cunha

    2001-03-01

    Full Text Available We examined some of the mechanisms by which the aspirin metabolite and the naturally occurring metabolite gentisic acid induced relaxation of the guinea pig trachea in vitro. In preparations with or without epithelium and contracted by histamine, gentisic acid caused concentration-dependent and reproducible relaxation, with mean EC50 values of 18 µM and Emax of 100% (N = 10 or 20 µM and Emax of 92% (N = 10, respectively. The relaxation caused by gentisic acid was of slow onset in comparison to that caused by norepinephrine, theophylline or vasoactive intestinal peptide (VIP. The relative rank order of potency was: salbutamol 7.9 > VIP 7.0 > gentisic acid 4.7 > theophylline 3.7. Gentisic acid-induced relaxation was markedly reduced (24 ± 7.0, 43 ± 3.9 and 78 ± 5.6% in preparations with elevated potassium concentration in the medium (20, 40 or 80 mM, respectively. Tetraethylammonium (100 µM, a nonselective blocker of the potassium channels, partially inhibited the relaxation response to gentisic acid, while 4-AP (10 µM, a blocker of the voltage potassium channel, inhibited gentisic acid-induced relaxation by 41 ± 12%. Glibenclamide (1 or 3 µM, at a concentration which markedly inhibited the relaxation induced by the opener of ATP-sensitive K+ channels, levcromakalim, had no effect on the relaxation induced by gentisic acid. Charybdotoxin (0.1 or 0.3 µM, a selective blocker of the large-conductance Ca2+-activated K+ channels, caused rightward shifts (6- and 7-fold of the gentisic acid concentration-relaxation curve. L-N G-nitroarginine (100 µM, a NO synthase inhibitor, had no effect on the relaxant effect of gentisic acid, and caused a slight displacement to the right in the relaxant effect of the gentisic acid curve at 300 µM, while methylene blue (10 or 30 µM or ODQ (1 µM, the inhibitors of soluble guanylate cyclase, all failed to affect gentisic acid-induced relaxation. D-P-Cl-Phe6,Leu17[VIP] (0.1 µM, a VIP receptor antagonist

  12. Structure, folding and stability of a minimal homologue from Anemonia sulcata of the sea anemone potassium channel blocker ShK.

    Science.gov (United States)

    Krishnarjuna, Bankala; MacRaild, Christopher A; Sunanda, Punnepalli; Morales, Rodrigo A V; Peigneur, Steve; Macrander, Jason; Yu, Heidi H; Daly, Marymegan; Raghothama, Srinivasarao; Dhawan, Vikas; Chauhan, Satendra; Tytgat, Jan; Pennington, Michael W; Norton, Raymond S

    2018-01-01

    Peptide toxins elaborated by sea anemones target various ion-channel sub-types. Recent transcriptomic studies of sea anemones have identified several novel candidate peptides, some of which have cysteine frameworks identical to those of previously reported sequences. One such peptide is AsK132958, which was identified in a transcriptomic study of Anemonia sulcata and has a cysteine framework similar to that of ShK from Stichodactyla helianthus, but is six amino acid residues shorter. We have determined the solution structure of this novel peptide using NMR spectroscopy. The disulfide connectivities and structural scaffold of AsK132958 are very similar to those of ShK but the structure is more constrained. Toxicity assays were performed using grass shrimp (Palaemonetes sp) and Artemia nauplii, and patch-clamp electrophysiology assays were performed to assess the activity of AsK132958 against a range of voltage-gated potassium (K V ) channels. AsK132958 showed no activity against grass shrimp, Artemia nauplii, or any of the K V channels tested, owing partly to the absence of a functional Lys-Tyr dyad. Three AsK132958 analogues, each containing a Tyr in the vicinity of Lys19, were therefore generated in an effort to restore binding, but none showed activity against any of K V channels tested. However, AsK132958 and its analogues are less susceptible to proteolysis than that of ShK. Our structure suggests that Lys19, which might be expected to occupy the pore of the channel, is not sufficiently accessible for binding, and therefore that AsK132958 must have a distinct functional role that does not involve K V channels. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Voltage-dependent antagonist/agonist actions of taurine on Ca(2+)-activated potassium channels of rat skeletal muscle fibers.

    Science.gov (United States)

    Tricarico, D; Barbieri, M; Conte Camerino, D

    2001-09-01

    Emerging evidence supports the idea that taurine exerts some of its actions through inhibition of inward rectifier K(+) channels, ATP-sensitive K(+) channels, and voltage-dependent K(+) channels. However, to date not much is known about the effects of this sulfonic amino acid on Ca(2+)-activated K(+) (K(Ca(2+))) channels, which are widely expressed in various tissues, including skeletal muscle. In the present work, the effects of taurine on K(Ca(2+)) channels of rat skeletal muscle fibers were investigated using the patch-clamp technique. The application of the amino acid to the internal side of the excised macropatches induced a dose-dependent decrease in the outward K(Ca(2+)) currents recorded at positive membrane potentials in the presence of 8 to 16 microM concentrations of free Ca(2+) ions in the bath with an IC(50) of 31.9. 10(-3) +/- 1 M (slope factor = 1.2) (n = 11 patches). In contrast, at negative membrane potentials taurine caused an enhancement of the muscular inward K(Ca(2+)) currents with a DE(50) (drug concentration needed to enhance the current by 50%) of 46.7. 10(-3) +/- 2 M (slope factor = 1.3) (n = 9 patches). Single channel analysis revealed that this effect was mediated by changes in the reversal potential of the K(Ca(2+)) channel for K(+) ions with no changes in the gating properties or in the sensitivity of the channel to Ca(2+) ions. Taurine also did not affect the single channel conductance. In conclusion, taurine shows a voltage-dependent dualistic action on K(Ca(2+)) channels, being an inhibitor of the channel at positive membrane potentials and an activator at negative membrane potentials.

  14. Beraprost sodium, a stable prostacyclin analogue, elicits dilation of isolated porcine retinal arterioles: roles of eNOS and potassium channels.

    Science.gov (United States)

    Ono, Shinji; Nagaoka, Taiji; Omae, Tsuneaki; Tanano, Ichiro; Kamiya, Takayuki; Otani, Shinichi; Ishibazawa, Akihiro; Yoshida, Akitoshi

    2014-07-31

    Prostacyclin (PGI2) is usually described as an endoEDRFsthelium-derived relaxing factor, but the vasoreactivity to PGI2 in the retinal arterioles and the underlying mechanisms are not fully understood. We examined the effects of PGI2 on the retinal microcirculation using beraprost sodium (BPS), a stable PGI2 analogue, and the signaling mechanisms involved in this vasomotor activity. Porcine retinal arterioles were isolated, cannulated, and pressurized without flow in vitro. Video microscopic techniques recorded the diametric responses to BPS. Beraprost sodium elicited dose-dependent (0.1 pM-0.1 μM) vasodilation of the retinal arterioles that was abolished by the PGI2 receptor (IP) antagonist CAY10441. Beraprost sodium-induced vasodilation decreased by 50% after the endothelium was removed and was inhibited by the nitric oxide (NO) synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) comparable with denudation. Inhibition of soluble guanylyl cyclase by 1H-1,2,4-oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and blockage of protein kinase A (PKA) by Rp-8-Br-cAMPS were comparable to L-NAME. Beraprost sodium-induced vasodilation was also inhibited by the nonselective potassium channel inhibitor, tetraethylammonium, and the adenosine triphosphate-sensitive potassium (KATP) channel blocker, glibenclamide. Residual vasodilation in the presence of glibenclamide decreased further with subsequent application of ODQ. Beraprost sodium, a stable PGI2 analogue, causes vasodilation of the retinal arterioles mediated via the IP receptor. The current findings suggest that BPS elicits endothelium-dependent and -independent dilation of the retinal arterioles mediated by NO induced by activation of PKA in the endothelium and the KATP channel activation in the vascular smooth muscle, respectively. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.

  15. A pharmacologically validated, high-capacity, functional thallium flux assay for the human Ether-à-go-go related gene potassium channel.

    Science.gov (United States)

    Schmalhofer, William A; Swensen, Andrew M; Thomas, Brande S; Felix, John P; Haedo, Rodolfo J; Solly, Kelli; Kiss, Laszlo; Kaczorowski, Gregory J; Garcia, Maria L

    2010-12-01

    The voltage-gated potassium channel, human Ether-à-go-go related gene (hERG), represents the molecular component of IKr, one of the potassium currents involved in cardiac action potential repolarization. Inhibition of IKr increases the duration of the ventricular action potential, reflected as a prolongation of the QT interval in the electrocardiogram, and increases the risk for potentially fatal ventricular arrhythmias. Because hERG is an appropriate surrogate for IKr, hERG assays that can identify potential safety liabilities of compounds during lead identification and optimization have been implemented. Although the gold standard for hERG evaluation is electrophysiology, this technique, even with the medium capacity, automated instruments that are currently available, does not meet the throughput demands for supporting typical medicinal chemistry efforts in the pharmaceutical environment. Assays that could provide reliable molecular pharmacology data, while operating in high capacity mode, are therefore desirable. In the present study, we describe a high-capacity, 384- and 1,536-well plate, functional thallium flux assay for the hERG channel that fulfills these criteria. This assay was optimized and validated using different structural classes of hERG inhibitors. An excellent correlation was found between the potency of these agents in the thallium flux assay and in electrophysiological recordings of channel activity using the QPatch automated patch platform. Extension of this study to include 991 medicinal chemistry compounds from different internal drug development programs indicated that the thallium flux assay was a good predictor of in vitro hERG activity. These data suggest that the hERG thallium flux assay can play an important role in supporting drug development efforts.

  16. A conserved residue cluster that governs kinetics of ATP-dependent gating of Kir6.2 potassium channels

    DEFF Research Database (Denmark)

    Zhang, Roger S; Wright, Jordan; Pless, Stephan Alexander

    2015-01-01

    elements that control the kinetics of ATP-dependent regulation of KATP (Kir6.2 + SUR1) channels using rapid concentration jumps. WT Kir6.2 channels re-open after rapid washout of ATP with a time constant of approximately 60 ms. Extending similar kinetic measurements to numerous mutants revealed fairly...... modest effects on gating kinetics despite significant changes in ATP sensitivity and open probability. However, we identified a pair of highly conserved neighboring amino acids (Trp68, Lys170) that control the rate of channel opening and inhibition in response to ATP. Paradoxically, mutations of Trp68...... or Lys170 markedly slow the kinetics of channel opening (500 ms and 700 ms for Trp68Leu and Lys170Asn, respectively), while increasing channel open probability. Examining the functional effects of these residues using phi-value analysis revealed a steep negative slope. This finding implies...

  17. Functional hair cell mechanotransducer channels are required for aminoglycoside ototoxicity.

    Directory of Open Access Journals (Sweden)

    Abdelrahman Alharazneh

    Full Text Available Aminoglycosides (AG are commonly prescribed antibiotics with potent bactericidal activities. One main side effect is permanent sensorineural hearing loss, induced by selective inner ear sensory hair cell death. Much work has focused on AG's initiating cell death processes, however, fewer studies exist defining mechanisms of AG uptake by hair cells. The current study investigated two proposed mechanisms of AG transport in mammalian hair cells: mechanotransducer (MET channels and endocytosis. To study these two mechanisms, rat cochlear explants were cultured as whole organs in gentamicin-containing media. Two-photon imaging of Texas Red conjugated gentamicin (GTTR uptake into live hair cells was rapid and selective. Hypocalcemia, which increases the open probability of MET channels, increased AG entry into hair cells. Three blockers of MET channels (curare, quinine, and amiloride significantly reduced GTTR uptake, whereas the endocytosis inhibitor concanavalin A did not. Dynosore quenched the fluorescence of GTTR and could not be tested. Pharmacologic blockade of MET channels with curare or quinine, but not concanavalin A or dynosore, prevented hair cell loss when challenged with gentamicin for up to 96 hours. Taken together, data indicate that the patency of MET channels mediated AG entry into hair cells and its toxicity. Results suggest that limiting permeation of AGs through MET channel or preventing their entry into endolymph are potential therapeutic targets for preventing hair cell death and hearing loss.

  18. Rotenone induces KATP channel opening in PC12 cells in association with the expression of tyrosine hydroxylase.

    Science.gov (United States)

    Bai, Qunhua; He, Junlin; Qiu, Jingfu; Wang, Yang; Wang, Shibo; Xiu, Yun; Yu, Chao

    2012-10-01

    The activation of ATP-sensitive potassium (KATP) channels in PC12 cells play a pivotal role in protection against the neurotoxic effect of rotenone. However, it remains unclear why rotenone seems to preferentially affect activation of KATP channels and if this could affect its physiological activity. In this study, we sought to determine how the different energy states caused by various doses of rotenone affect the KATP opening state and whether the KATP opening state influences the expression of tyrosine hydroxylase (TH) which is related with DA synthesis. With patch clamp technology, results showed that treatment of PC12 cells with rotenone (0.2-1 µg/ml) for 15 min can cause KATP channel opening with significantly increased intracellular ROS production. Treatment with rotenone (2-16 ng/ml) for 24 h also caused the channels to open with gently increased ROS. In order to study if the rather long-term action on KATP channel opening states could affect the specified function of PC12 cells, the KATP channel opener pinacidil and the inhibitor glibenclamide were used to treat cells for 24 h, and the expression of TH was detected. Our results showed that treatment of PC12 cells with glibenclamide for 24 h can notably promote TH expression and can also enhance the expression of TH which were reduced by rotenone. These data indicate that the energy states in PC12 induced by various doses of rotenone could significantly influence the opening states of KATP channels. However long-term energy stress may raise the opening rate and opening sensitivity of this channel. In addition, our results demonstrate for the first time that activation of plasma membrane KATP channels induced by rotenone inhibits TH expression which influences DA synthesis in PC12 cells.

  19. SK2 channel expression and function in cerebellar Purkinje cells

    Science.gov (United States)

    Hosy, Eric; Piochon, Claire; Teuling, Eva; Rinaldo, Lorenzo; Hansel, Christian

    2011-01-01

    Abstract Small-conductance calcium-activated K+ channels (SK channels) regulate the excitability of neurons and their responsiveness to synaptic input patterns. SK channels contribute to the afterhyperpolarization (AHP) following action potential bursts, and curtail excitatory postsynaptic potentials (EPSPs) in neuronal dendrites. Here we review evidence that SK2 channels are expressed in rat cerebellar Purkinje cells during development and throughout adulthood, and play a key role in diverse cellular processes such as the regulation of the spike firing frequency and the modulation of calcium transients in dendritic spines. In Purkinje cells as well as in other types of neurons, SK2 channel plasticity seems to provide an important mechanism allowing these cells to adjust their intrinsic excitability and to alter the probabilities for the induction of synaptic learning correlates, such as long-term potentiation (LTP). PMID:21521760

  20. A De Novo Mutation in the Sodium-Activated Potassium Channel KCNT2 Alters Ion Selectivity and Causes Epileptic Encephalopathy

    Directory of Open Access Journals (Sweden)

    Sushmitha Gururaj

    2017-10-01

    Full Text Available Early infantile epileptic encephalopathies (EOEE are a debilitating spectrum of disorders associated with cognitive impairments. We present a clinical report of a KCNT2 mutation in an EOEE patient. The de novo heterozygous variant Phe240Leu SLICK was identified by exome sequencing and confirmed by Sanger sequencing. Phe240Leu rSlick and hSLICK channels were electrophysiologically, heterologously characterized to reveal three significant alterations to channel function. First, [Cl−]i sensitivity was reversed in Phe240Leu channels. Second, predominantly K+-selective WT channels were made to favor Na+ over K+ by Phe240Leu. Third, and consequent to altered ion selectivity, Phe240Leu channels had larger inward conductance. Further, rSlick channels induced membrane hyperexcitability when expressed in primary neurons, resembling the cellular seizure phenotype. Taken together, our results confirm that Phe240Leu is a “change-of-function” KCNT2 mutation, demonstrating unusual altered selectivity in KNa channels. These findings establish pathogenicity of the Phe240Leu KCNT2 mutation in the reported EOEE patient.

  1. Taurine blocks ATP-sensitive potassium channels of rat skeletal muscle fibres interfering with the sulphonylurea receptor.

    Science.gov (United States)

    Tricarico, D; Barbieri, M; Camerino, D C

    2000-06-01

    Taurine is a sulphonic aminoacid present in high amounts in various tissues including cardiac and skeletal muscles showing different properties such as antioxidative, antimyotonic and anti-schaemic effects. The cellular mechanism of action of taurine is under investigation and appears to involve the interaction of the sulphonic aminoacid with several ion channels. Using the patch-clamp technique we studied the effects of taurine in rat skeletal muscle fibres on ATP-sensitive K(+) channel (K(ATP)) immediately after excision and on channels that underwent rundown. The cytoplasmic application of 20 mM of taurine reduced the K(ATP) current; this effect was reverted by washout of the drug solution. In this experimental condition the IC(50) was 20.1 mM. After rundown, taurine inhibited the K(ATP) current with similar efficacy. Competition experiments showed that taurine shifted the dose-response inhibition curve of glybenclamide to the left on the log-dose axis without significantly affecting those of ATP or Ca(2+) ion. Single channel recording revealed that taurine affects the close state of the channel prolonging it and reducing the bursts duration. Our data indicate that taurine inhibits the muscular K(ATP) channel interfering with the glybenclamide site on the sulphonylurea receptor of the channel or on the site allosterically coupled to it. During ischaemia and hypoxia, the skeletal and heart muscles undergo several changes; for example, the activation of K(ATP) channels and loss of the intracellular taurine content. The depletion of taurine during ischaemia would contribute to the early activation of K(ATP) channels and salvage the intracellular ATP content.

  2. Voltage-gated potassium channel (K(v) 1) autoantibodies in patients with chagasic gut dysmotility and distribution of K(v) 1 channels in human enteric neuromusculature (autoantibodies in GI dysmotility).

    Science.gov (United States)

    Hubball, A W; Lang, B; Souza, M A N; Curran, O D; Martin, J E; Knowles, C H

    2012-08-01

    Autoantibodies directed against specific neuronal antigens are found in a significant number of patients with gastrointestinal neuromuscular diseases (GINMDs) secondary to neoplasia. This study examined the presence of antineuronal antibodies in idiopathic GINMD and GINMD secondary to South American Trypanosomiasis. The GI distribution of voltage-gated potassium channels (VGKCs) was also investigated. Seventy-three patients were included in the study with diagnoses of primary achalasia, enteric dysmotility, chronic intestinal pseudo-obstruction, esophageal or colonic dysmotility secondary to Chagas' disease. Sera were screened for specific antibodies to glutamic acid decarboxylase, voltage-gated calcium channels (VGCCs; P/Q subtype), nicotinic acetylcholine receptors (nAChRs; α3 subtype), and voltage-gated potassium channels (VGKCs, K(V) 1 subtype) using validated immunoprecipitation assays. The distribution of six VGKC subunits (K(V) 1.1-1.6), including those known to be antigenic targets of anti-VGKC antibodies was immunohistochemically investigated in all main human GI tract regions. Three patients (14%) with chagasic GI dysmotility were found to have positive anti-VGKC antibody titers. No antibodies were detected in patients with idiopathic GINMD. The VGKCs were found in enteric neurons at every level of the gut in unique yet overlapping distributions. The VGKC expression in GI smooth muscle was found to be limited to the esophagus. A small proportion of patients with GI dysfunction secondary to Chagas' disease have antibodies against VGKCs. The presence of these channels in the human enteric nervous system may have pathological relevance to the growing number of GINMDs with which anti-VGKC antibodies have been associated. © 2012 Blackwell Publishing Ltd.

  3. Technology channel fuel cells; Reseau technologique piles a combustible

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2000-07-01

    This document presents the PACo channel, its research and development program and the calendar of the first year. The PACo channel aims at stimulate the technology innovation in the domain of the fuel cells and organize collaborations between enterprises and research laboratories. (A.L.B.)

  4. Cutaneous blood flow during intradermal NO administration in young and older adults: roles for calcium-activated potassium channels and cyclooxygenase?

    Science.gov (United States)

    Fujii, Naoto; Meade, Robert D; Minson, Christopher T; Brunt, Vienna E; Boulay, Pierre; Sigal, Ronald J; Kenny, Glen P

    2016-06-01

    Nitric oxide (NO) increases cutaneous blood flow; however, the underpinning mechanism(s) remains to be elucidated. We hypothesized that the cutaneous blood flow response during intradermal administration of sodium nitroprusside (SNP, a NO donor) is regulated by calcium-activated potassium (KCa) channels and cyclooxygenase (COX) in young adults. We also hypothesized that these contributions are diminished in older adults given that aging can downregulate KCa channels and reduce COX-derived vasodilator prostanoids. In 10 young (23 ± 5 yr) and 10 older (54 ± 4 yr) adults, cutaneous vascular conductance (CVC) was measured at four forearm skin sites infused with 1) Ringer (Control), 2) 50 mM tetraethylammonium (TEA), a nonspecific KCa channel blocker, 3) 10 mM ketorolac, a nonspecific COX inhibitor, or 4) 50 mM TEA + 10 mM ketorolac via intradermal microdialysis. All skin sites were coinfused with incremental doses of SNP (0.005, 0.05, 0.5, 5, and 50 mM each for 25 min). During SNP administration, CVC was similar at the ketorolac site (0.005-50 mM, all P > 0.05) relative to Control, but lower at the TEA and TEA + ketorolac sites (0.005-0.05 mM, all P 0.05). Furthermore, TEA alone did not modulate CVC during any concentration of SNP administration in older adults (all P > 0.05). We show that during low-dose NO administration (e.g., 0.005-0.05 mM), KCa channels contribute to cutaneous blood flow regulation in young adults; however, in older adults, COX inhibition increases cutaneous blood flow through a KCa channel-dependent mechanism. Copyright © 2016 the American Physiological Society.

  5. The polarization of the G-protein activated potassium channel GIRK5 to the vegetal pole of Xenopus laevis oocytes is driven by a di-leucine motif.

    Science.gov (United States)

    Díaz-Bello, Beatriz; Rangel-García, Claudia I; Salvador, Carolina; Carrisoza-Gaytán, Rolando; Escobar, Laura I

    2013-01-01

    The G protein-coupled inwardly-rectifying potassium channels (known as GIRK or Kir3) form functional heterotetramers gated by G-βγ subunits. GIRK channels participate in heart rate modulation and neuronal postsynaptic inhibition in mammals. In Xenopus laevis oocytes, GIRK5 is a functional homomultimer. Previously, we found that phosphorylation of a tyrosine (Y16) at its N-terminus downregulates the surface expression of GIRK5. In this work, we elucidated the subcellular localization and trafficking of GIRK5 in oocytes. Several EGFP-GIRK5 chimeras were produced and an ECFP construct was used to identify the endoplasmic reticulum (ER). Whereas GIRK5-WT was retained in the ER at the animal pole, the phospho-null GIRK5-Y16A was localized to the vegetal pole. Interestingly, a construct with an N-terminal Δ25 deletion produced an even distribution of the channel in the whole oocyte. Through an alanine-scan, we identified an acidic cluster/di-leucine sorting-signal recognition motif between E17 and I22. We quantified the effect of each amino acid residue within this di-leucine motif in determining the distribution of GIRK5 to the animal and vegetal poles. We found that Y16 and I22 contributed to functional expression and were dominant in the polarization of GIRK5. We thus conclude that the N-terminal acidic di-leucine motif of GIRK5 determines its retention and polarized trafficking within Xl oocytes.

  6. Taurine blocks ATP-sensitive potassium channels of rat skeletal muscle fibres interfering with the sulphonylurea receptor

    OpenAIRE

    Tricarico, Domenico; Barbieri, Mariagrazia; Camerino, Diana Conte

    2000-01-01

    Taurine is a sulphonic aminoacid present in high amounts in various tissues including cardiac and skeletal muscles showing different properties such as antioxidative, antimyotonic and anti-schaemic effects. The cellular mechanism of action of taurine is under investigation and appears to involve the interaction of the sulphonic aminoacid with several ion channels.Using the patch-clamp technique we studied the effects of taurine in rat skeletal muscle fibres on ATP-sensitive K+ channel (KATP) ...

  7. Patients with Long-QT Syndrome Caused by Impaired hERG -Encoded Kv11.1 Potassium Channel Have Exaggerated Endocrine Pancreatic and Incretin Function Associated with Reactive Hypoglycemia

    DEFF Research Database (Denmark)

    Hyltén-Cavallius, Louise; Iepsen, Eva W.; Wewer Albrechtsen, Nicolai J.

    2017-01-01

    Background: Loss-of-function mutations in hERG (encoding the Kv11.1 voltage-gated potassium channel) cause long-QT syndrome type 2 (LQT2) because of prolonged cardiac repolarization. However, Kv11.1 is also present in pancreatic and β cells and intestinal L and K cells, secreting glucagon, insulin...... to glucose ingestion in LQT2 patients with functional mutations in hERG and matched healthy participants, testing the hypothesis that LQT2 patients have increased incretin and β-cell function and decreased -cell function, and thus lower glucose levels. Methods: Eleven patients with LQT2 and 22 sex-, age...... in controls (4398 [95% confidence interval, 2259-8562] versus 2156 [1961-3201], P=0.03). Pharmacological Kv11.1 blockade (dofetilide) in rats had similar effect, and small interfering RNA inhibition of hERG in β and L cells increased insulin and GLP-1 secretion up to 50%. Glucose ingestion caused cardiac...

  8. Physiology and Regulation of Calcium Channels in Stomatal Guard Cells

    Energy Technology Data Exchange (ETDEWEB)

    Schroeder, Julian I.

    2007-05-02

    Stomatal pores in the epidermis of leaves regulate the diffusion of CO2 into leaves for photosynthetic carbon fixation and control water loss of plants during drought periods. Guard cells sense CO2, water status, light and other environmental conditions to regulate stomatal apertures for optimization of CO2 intake and plant growth under drought stress. The cytosolic second messenger calcium contributes to stomatal movements by transducing signals and regulating ion channels in guard cells. Studies suggest that both plasma membrane Ca2+ influx channels and vacuolar/organellar Ca2+ release channels contribute to ABA-induced Ca2+ elevations in guard cells. Recent research in the P.I.'s laboratory has led to identification of a novel major cation-selective Ca2+-permeable influx channel (Ica) in the plasma membrane of Arabidopsis guard cells. These advances will allow detailed characterization of Ica plasma membrane Ca2+ influx channels in guard cells. The long term goal of this research project is to gain a first detailed characterization of these novel plasma membrane Ca2+-permeable channel currents in Arabidopsis guard cells. The proposed research will investigate the hypothesis that Ica represents an important Ca2+ influx pathway for ABA and CO2 signal transduction in Arabidopsis guard cells. These studies will lead to elucidation of key signal transduction mechanisms by which plants balance CO2 influx into leaves and transpirational water loss and may contribute to future strategies for manipulating gas exchange for improved growth of crop plants and for biomass production.

  9. Preliminary Studies of Acute Cadmium Administration Effects on the Calcium-Activated Potassium (SKCa and BKCa) Channels and Na+/K+-ATPase Activity in Isolated Aortic Rings of Rats.

    Science.gov (United States)

    Vassallo, Dalton V; Almenara, Camila C P; Broseghini-Filho, Gilson Brás; Teixeira, Ariane Calazans; da Silva, David Chaves F; Angeli, Jhuli K; Padilha, Alessandra S

    2017-09-13

    Cadmium is an environmental pollutant closely linked with cardiovascular diseases that seems to involve endothelium dysfunction and reduced nitric oxide (NO) bioavailability. Knowing that NO causes dilatation through the activation of potassium channels and Na + /K + -ATPase, we aimed to determine whether acute cadmium administration (10 μM) alters the participation of K + channels, voltage-activated calcium channel, and Na + /K + -ATPase activity in vascular function of isolated aortic rings of rats. Cadmium did not modify the acetylcholine-induced relaxation. After L-NAME addition, the relaxation induced by acetylcholine was abolished in presence or absence of cadmium, suggesting that acutely, this metal did not change NO release. However, tetraethylammonium (a nonselective K + channels blocker) reduced acetylcholine-induced relaxation but this effect was lower in the preparations with cadmium, suggesting a decrease of K + channels function in acetylcholine response after cadmium incubation. Apamin (a selective blocker of small Ca 2+ -activated K + channels-SK Ca ), iberiotoxin (a selective blocker of large-conductance Ca 2+ -activated K + channels-BK Ca ), and verapamil (a blocker of calcium channel) reduced the endothelium-dependent relaxation only in the absence of cadmium. Finally, cadmium decreases Na + /K + -ATPase activity. Our results provide evidence that the cadmium acute incubation unaffected the calcium-activated potassium channels (SK Ca and BK Ca ) and voltage-calcium channels on the acetylcholine vasodilatation. In addition, acute cadmium incubation seems to reduce the Na + /K + -ATPase activity.

  10. Intermediate-Conductance-Ca2-Activated K Channel IKCa1 Is Upregulated and Promotes Cell Proliferation in Cervical Cancer

    Science.gov (United States)

    Liu, Ling; Zhan, Ping; Nie, Dan; Fan, Lingye; Lin, Hairui; Gao, Lanyang; Mao, Xiguang

    2017-03-10

    BACKGROUND Accumulating data point to intermediate-conductance calcium-activated potassium channel (IKCa1) as a key player in controlling cell cycle progression and proliferation of human cancer cells. However, the role that IKCa1 plays in the growth of human cervical cancer cells is largely unexplored. MATERIAL AND METHODS In this study, Western blot analysis, immunohistochemical staining, and RT-PCR were first used for IKCa1protein and gene expression assays in cervical cancer tissues and HeLa cells. Then, IKCa1 channel blocker and siRNA were employed to inhibit the functionality of IKCa1 and downregulate gene expression in HeLa cells, respectively. After these treatments, we examined the level of cell proliferation by MTT method and measured IKCa1 currents by conventional whole-cell patch clamp technique. Cell apoptosis was assessed using the Annexin V-FITC/Propidium Iodide (PI) double-staining apoptosis detection kit. RESULTS We demonstrated that IKCa1 mRNA and protein are preferentially expressed in cervical cancer tissues and HeLa cells. We also showed that the IKCa1 channel blocker, clotrimazole, and IKCa1 channel siRNA can be used to suppress cervical cancer cell proliferation and decrease IKCa1 channel current. IKCa1 downregulation by specific siRNAs induced a significant increase in the proportion of apoptotic cells in HeLa cells. CONCLUSIONS IKCa1 is overexpressed in cervical cancer tissues, and IKCa1 upregulation in cervical cancer cell linea enhances cell proliferation, partly by reducing the proportion of apoptotic cells.

  11. An intermediate-conductance Ca2+-activated K+ channel is important for secretion in pancreatic duct cells

    DEFF Research Database (Denmark)

    Hayashi, Mikio; Wang, Jing; Hede, Susanne Edeling

    2012-01-01

    the molecular basis of functional K(+) channels in rodent and human pancreatic ducts (Capan-1, PANC-1, and CFPAC-1) using molecular and electrophysiological techniques. RT-PCR analysis revealed mRNAs for KCNQ1, KCNH2, KCNH5, KCNT1, and KCNT2, as well as KCNN4 coding for the following channels: KVLQT1; HERG; EAG......2; Slack; Slick; and an intermediate-conductance Ca(2+)-activated K(+) (IK) channel (K(Ca)3.1). The following functional studies were focused on the IK channel. 5,6-Dichloro-1-ethyl-1,3-dihydro-2H-benzimidazole-2-one (DC-EBIO), an activator of IK channel, increased equivalent short-circuit current...... revealed IK channels with an average conductance of 80 pS in freshly isolated rodent duct cells. These results indicated that the IK channels may, at least in part, be involved in setting the resting membrane potential. Furthermore, the IK channels are involved in anion and potassium transport...

  12. [Influence of the Potassium Channels Activator Nicorandil to the Quality of Life in Patients With Ishemic Heart Disease and Stable Angina Pectoris].

    Science.gov (United States)

    Sizova, Zh M; Zakharova, V L; Kozlova, N V; Kuchkina, T S

    2016-06-01

    The aim of the study to assess the influence of medicamentous therapy to the quality of life of ischemic heart disease patients with stable angina pectoris by activator potassium channels nicorandil in comparison with traditional therapy by isosorbide dinitrate. The study included 84 ischemic heart disease patients. Authors consider quality of life as an estimated category of state of the subject in an illness situation. The dynamic of physical and psychological components of quality of life are compared in ischemic heart disease patients under the treatment by nicorandil and isosorbide dinitrate. Indicators of quality of life, defined on the basis of a questionnaires of SAQ and GHQ supplementing an illness picture, are an multiple-factor criterion of an assessment of a condition of this category of patients. The benefits of nicorandil in influence on quality of life indicators were revealed in the study.

  13. Increase of the cytotoxic effect of Bothrops jararacussu venom on mouse extensor digitorum longus and soleus by potassium channel blockers and by Na(+)/K(+)-ATPase inhibition.

    Science.gov (United States)

    Tomaz, Marcelo A; Fernandes, Fabrício F A; El-Kik, Camila Z; Moraes, Raphael A M; Calil-Elias, Sabrina; Saturnino-Oliveira, Jeison; Martinez, Ana Maria B; Ownby, Charlotte L; Melo, Paulo A

    2008-09-15

    We investigated the myotoxicity of Bothrops jararacussu crude venom and other cytolytic agents on mouse isolated extensor digitorum longus (EDL) and soleus (SOL) muscles, which present distinct properties: EDL is a fast-twitch, white muscle with predominantly glycolytic fibers, while SOL is slow-twitch, red muscle with predominantly oxidative fibers. Muscles were exposed to B. jararacussu crude venom (25 microg/ml) and other crotaline venoms (Agkistrodon contortrix laticinctus; Crotalus viridis viridis; Crotalus durissus terrificus) at the same concentration. Basal creatine kinase (CK) release to bathing solution was 0.43+/-0.06 for EDL and 0.29+/-0.06 for SOL (U g(-)(1) h(-)(1), n=36 for each muscle). Sixty minutes after exposure to B. jararacussu venom, EDL presented higher increase in the rate of CK release than SOL, respectively, 13.2+/-1.5 and 2.9+/-0.7 U g(-)(1)h(-)(1), n=10-12. Muscle denervation, despite decreasing CK content, did not affect sensitivities to B. jararacussu venom. Ouabain and potassium channel blockers (TEA; clotrimazole; glibenclamide) increased the rate of CK release by B. jararacussu in EDL and SOL muscles, decreasing and almost abolishing the different sensitivity. When we exposed EDL or SOL muscles to Naja naja, Apis mellifera venoms (25 microg/ml), or Triton X-100 (0.01%), they showed similar rate of CK release. Our present data suggest that a mechanism involving intracellular calcium regulation or potassium channels may participate in the different sensitivity of EDL and SOL to B. jararacussu venom.

  14. Evolutionary analyses of KCNQ1 and HERG voltage-gated potassium channel sequences reveal location-specific susceptibility and augmented chemical severities of arrhythmogenic mutations

    Directory of Open Access Journals (Sweden)

    Accili Eric A

    2008-06-01

    Full Text Available Abstract Background Mutations in HERG and KCNQ1 potassium channels have been associated with Long QT syndrome and atrial fibrillation, and more recently with sudden infant death syndrome and sudden unexplained death. In other proteins, disease-associated amino acid mutations have been analyzed according to the chemical severity of the changes and the locations of the altered amino acids according to their conservation over metazoan evolution. Here, we present the first such analysis of arrhythmia-associated mutations (AAMs in the HERG and KCNQ1 potassium channels. Results Using evolutionary analyses, AAMs in HERG and KCNQ1 were preferentially found at evolutionarily conserved sites and unevenly distributed among functionally conserved domains. Non-synonymous single nucleotide polymorphisms (nsSNPs are under-represented at evolutionarily conserved sites in HERG, but distribute randomly in KCNQ1. AAMs are chemically more severe, according to Grantham's Scale, than changes observed in evolution and their severity correlates with the expected chemical severity of the involved codon. Expected chemical severity of a given amino acid also correlates with its relative contribution to arrhythmias. At evolutionarily variable sites, the chemical severity of the changes is also correlated with the expected chemical severity of the involved codon. Conclusion Unlike nsSNPs, AAMs preferentially locate to evolutionarily conserved, and functionally important, sites and regions within HERG and KCNQ1, and are chemically more severe than changes which occur in evolution. Expected chemical severity may contribute to the overrepresentation of certain residues in AAMs, as well as to evolutionary change.

  15. Expression and function of Kv1.1 potassium channels in human atria from patients with atrial fibrillation.

    Science.gov (United States)

    Glasscock, Edward; Voigt, Niels; McCauley, Mark D; Sun, Qiang; Li, Na; Chiang, David Y; Zhou, Xiao-Bo; Molina, Cristina E; Thomas, Dierk; Schmidt, Constanze; Skapura, Darlene G; Noebels, Jeffrey L; Dobrev, Dobromir; Wehrens, Xander H T

    2015-09-01

    Voltage-gated Kv1.1 channels encoded by the Kcna1 gene are traditionally regarded as being neural-specific with no known expression or intrinsic functional role in the heart. However, recent studies in mice reveal low-level Kv1.1 expression in heart and cardiac abnormalities associated with Kv1.1-deficiency suggesting that the channel may have a previously unrecognized cardiac role. Therefore, this study tests the hypothesis that Kv1.1 channels are associated with arrhythmogenesis and contribute to intrinsic cardiac function. In intra-atrial burst pacing experiments, Kcna1-null mice exhibited increased susceptibility to atrial fibrillation (AF). The atria of Kcna1-null mice showed minimal Kv1 family ion channel remodeling and fibrosis as measured by qRT-PCR and Masson's trichrome histology, respectively. Using RT-PCR, immunocytochemistry, and immunoblotting, KCNA1 mRNA and protein were detected in isolated mouse cardiomyocytes and human atria for the first time. Patients with chronic AF (cAF) showed no changes in KCNA1 mRNA levels relative to controls; however, they exhibited increases in atrial Kv1.1 protein levels, not seen in paroxysmal AF patients. Patch-clamp recordings of isolated human atrial myocytes revealed significant dendrotoxin-K (DTX-K)-sensitive outward current components that were significantly increased in cAF patients, reflecting a contribution by Kv1.1 channels. The concomitant increases in Kv1.1 protein and DTX-K-sensitive currents in atria of cAF patients suggest that the channel contributes to the pathological mechanisms of persistent AF. These findings provide evidence of an intrinsic cardiac role of Kv1.1 channels and indicate that they may contribute to atrial repolarization and AF susceptibility.

  16. Voltage-gated sodium channels in taste bud cells.

    Science.gov (United States)

    Gao, Na; Lu, Min; Echeverri, Fernando; Laita, Bianca; Kalabat, Dalia; Williams, Mark E; Hevezi, Peter; Zlotnik, Albert; Moyer, Bryan D

    2009-03-12

    Taste bud cells transmit information regarding the contents of food from taste receptors embedded in apical microvilli to gustatory nerve fibers innervating basolateral membranes. In particular, taste cells depolarize, activate voltage-gated sodium channels, and fire action potentials in response to tastants. Initial cell depolarization is attributable to sodium influx through TRPM5 in sweet, bitter, and umami cells and an undetermined cation influx through an ion channel in sour cells expressing PKD2L1, a candidate sour taste receptor. The molecular identity of the voltage-gated sodium channels that sense depolarizing signals and subsequently initiate action potentials coding taste information to gustatory nerve fibers is unknown. We describe the molecular and histological expression profiles of cation channels involved in electrical signal transmission from apical to basolateral membrane domains. TRPM5 was positioned immediately beneath tight junctions to receive calcium signals originating from sweet, bitter, and umami receptor activation, while PKD2L1 was positioned at the taste pore. Using mouse taste bud and lingual epithelial cells collected by laser capture microdissection, SCN2A, SCN3A, and SCN9A voltage-gated sodium channel transcripts were expressed in taste tissue. SCN2A, SCN3A, and SCN9A were expressed beneath tight junctions in subsets of taste cells. SCN3A and SCN9A were expressed in TRPM5 cells, while SCN2A was expressed in TRPM5 and PKD2L1 cells. HCN4, a gene previously implicated in sour taste, was expressed in PKD2L1 cells and localized to cell processes beneath the taste pore. SCN2A, SCN3A and SCN9A voltage-gated sodium channels are positioned to sense initial depolarizing signals stemming from taste receptor activation and initiate taste cell action potentials. SCN2A, SCN3A and SCN9A gene products likely account for the tetrodotoxin-sensitive sodium currents in taste receptor cells.

  17. Tungstate-targeting of BKαβ1 channels tunes ERK phosphorylation and cell proliferation in human vascular smooth muscle.

    Directory of Open Access Journals (Sweden)

    Ana Isabel Fernández-Mariño

    Full Text Available Despite the substantial knowledge on the antidiabetic, antiobesity and antihypertensive actions of tungstate, information on its primary target/s is scarce. Tungstate activates both the ERK1/2 pathway and the vascular voltage- and Ca2+-dependent large-conductance BKαβ1 potassium channel, which modulates vascular smooth muscle cell (VSMC proliferation and function, respectively. Here, we have assessed the possible involvement of BKαβ1 channels in the tungstate-induced ERK phosphorylation and its relevance for VSMC proliferation. Western blot analysis in HEK cell lines showed that expression of vascular BKαβ1 channels potentiates the tungstate-induced ERK1/2 phosphorylation in a Gi/o protein-dependent manner. Tungstate activated BKαβ1 channels upstream of G proteins as channel activation was not altered by the inhibition of G proteins with GDPβS or pertussis toxin. Moreover, analysis of Gi/o protein activation measuring the FRET among heterologously expressed Gi protein subunits suggested that tungstate-targeting of BKαβ1 channels promotes G protein activation. Single channel recordings on VSMCs from wild-type and β1-knockout mice indicated that the presence of the regulatory β1 subunit was essential for the tungstate-mediated activation of BK channels in VSMCs. Moreover, the specific BK channel blocker iberiotoxin lowered tungstate-induced ERK phosphorylation by 55% and partially reverted (by 51% the tungstate-produced reduction of platelet-derived growth factor (PDGF-induced proliferation in human VSMCs. Our observations indicate that tungstate-targeting of BKαβ1 channels promotes activation of PTX-sensitive Gi proteins to enhance the tungstate-induced phosphorylation of ERK, and inhibits PDGF-stimulated cell proliferation in human vascular smooth muscle.

  18. Ion Channels Involved in Cell Volume Regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay

    2011-01-01

    This mini review outlines studies of cell volume regulation in two closely related mammalian cell lines: nonadherent Ehrlich ascites tumour cells (EATC) and adherent Ehrlich Lettre ascites (ELA) cells. Focus is on the regulatory volume decrease (RVD) that occurs after cell swelling, the volume...

  19. Transient receptor potential channels in mechanosensing and cell volume regulation

    DEFF Research Database (Denmark)

    Pedersen, Stine Falsig; Nilius, Bernd

    2007-01-01

    channels, and it is becoming increasingly apparent that Ca(2+) influx via TRP channels plays a crucial role in the response to mechanical and osmotic perturbations in a wide range of cell types. Although the events translating mechanical and osmotic stimuli into regulation of TRP channels are still...... incompletely understood, the specific mechanisms employed vary between different TRP isoforms, and probably include changes in the tension and/or curvature of the lipid bilayer, changes in the cortical cytoskeleton, and signaling events such as lipid metabolism and protein phosphorylation...... and involvement in cell volume regulation....

  20. Potassium and calcium channel gene expression in small arteries in porcine and rat models of diet-induced obesity (Poster)

    DEFF Research Database (Denmark)

    Jensen, Lars Jørn; Salomonsson, Max; Sørensen, Charlotte Mehlin

    2014-01-01

    Obesity is an increasing problem worldwide leading to cardiovascular morbidity. Only limited information exists on the transcriptional regulation of arterial K+ and Ca2+ channels in obesity. We quantified, by real-time PCR, mRNA expression of K+ channels and L-type Ca2+ channels (LTCC) in small...... mesenteric (MA), middle cerebral (MCA), and left coronary arteries (LCA) of lean vs. obese rats and minipigs. Male Sprague Dawley rats were fed a high-fat (FAT; N=5), high-fructose (FRUC; N=7), high-fat/high-fructose (FAT/FRUC; N=7) or standard diet (STD; N=7-11) for 28 Weeks. FAT and FAT/FRUC became obese......, whereas FRUC and STD were lean. Systolic blood pressure (SBP) averaged over 14 weeks was increased (P

  1. Effects of calcium channel on ovarian cancer cells.

    Science.gov (United States)

    Zhang, Chunyun; Li, Hailing

    2017-12-01

    The aim of the present study was to examine the effects of calcium channel protein on ovarian cancer cells. The expression of calcium channel protein in normal ovarian cells and ovarian cancer cells was detected by fluorescence quantitative PCR. Subsequently, the ovarian cancer cells were added to calcium channel protein activator media at various concentrations of 0, 1, 4, 8, 12, 16 and 20 mmol/l. The concentration of calcium ion in different samples was produced, and using an MTT assay, ovarian cancer cell activity in various samples was detected. Finally, a flow cytometer was used to explore the apoptosis rate. It was found that there was a significant difference between the expression of calcium channel protein in normal ovarian tissue and ovarian cancer cells (Pcancer cells to a certain extent, in a dose-dependent manner, especially when the concentration was at 12 mmol/l at which the intracellular calcium concentration was similar to that in normal ovarian cells. In conclusion, calcium ions play an important role in promoting cell proliferation of ovarian cancer cells, and they were involved in apoptosis of ovarian cancer cells to some extent, which regulates apoptosis by controlling the content of intracellular calcium.

  2. Expression and high glucose-mediated regulation of K+ channel interacting protein 3 (KChIP3) and KV4 channels in retinal Müller glial cells.

    Science.gov (United States)

    Chavira-Suárez, Erika; Sandoval, Alejandro; Felix, Ricardo; Lamas, Mónica

    2011-01-14

    Normal vision depends on the correct function of retinal neurons and glia and it is impaired in the course of diabetic retinopathy. Müller cells, the main glial cells of the retina, suffer morphological and functional alterations during diabetes participating in the pathological retinal dysfunction. Recently, we showed that Müller cells express the pleiotropic protein potassium channel interacting protein 3 (KChIP3), an integral component of the voltage-gated K(+) channels K(V)4. Here, we sought to analyze the role of KChIP3 in the molecular mechanisms underlying hyperglycemia-induced phenotypic changes in the glial elements of the retina. The expression and function of KChIp3 was analyzed in vitro in rat Müller primary cultures grown under control (5.6 mM) or high glucose (25 mM) (diabetic-like) conditions. We show the up-regulation of KChIP3 expression in Müller cell cultures under high glucose conditions and demonstrate a previously unknown interaction between the K(V)4 channel and KChIP3 in Müller cells. We show evidence for the expression of a 4-AP-sensitive transient outward voltage-gated K(+) current and an alteration in the inactivation of the macroscopic outward K(+) currents expressed in high glucose-cultured Müller cells. Our data support the notion that induction of KChIP3 and functional changes of K(V)4 channels in Müller cells could exert a physiological role in the onset of diabetic retinopathy. Copyright © 2010 Elsevier Inc. All rights reserved.

  3. Characterization of hERG1a and hERG1b potassium channels-a possible role for hERG1b in the I (Kr) current

    DEFF Research Database (Denmark)

    Larsen, Anders Peter; Olesen, Søren-Peter; Grunnet, Morten

    2008-01-01

    I (Kr) is the fast component of the delayed rectifier potassium currents responsible for the repolarization of the cardiac muscle. The molecular correlate underlying the I (Kr) current has been identified as the hERG1 channel. Recently, two splice variants of the hERG1 alpha-subunit, hERG1a and h...

  4. C5b-9-activated, Kv1.3 channels mediate oligodendrocyte cell cycle activation and dedifferentiation

    Science.gov (United States)

    Tegla, Cosmin A.; Cudrici, Cornelia; Rozycka, Monika; Soloviova, Katerina; Ito, Takahiro; Singh, Anil K.; Khan, Aamer; Azimzadeh, Philippe; Andrian-Albescu, Maria; Khan, Anver; Niculescu, Florin; Rus, Violeta; Judge, Susan I. V.; Rus, Horea

    2011-01-01

    Voltage-gated potassium (Kv) channels play an important role in the regulation of growth factor-induced cell proliferation. We have previously shown that cell cycle activation is induced in oligodendrocytes (OLGs) by complement C5b-9, but the role of Kv channels in these cells had not been investigated. Differentiated OLGs were found to express Kv1.4 channels, but little Kv1.3. Exposure of OLGs to C5b-9 modulated Kv1.3 functional channels and increased protein expression, whereas C5b6 had no effect. Pretreatment with the recombinant scorpion toxin rOsK-1, a specific Kv1.3 inhibitor, blocked the expression of Kv1.3 induced by C5b-9. rOsK-1 inhibited Akt phosphorylation and activation by C5b-9 but had no effect on ERK1 activation. These data strongly suggest a role for Kv1.3 in controlling the Akt activation induced by C5b-9. Since Akt plays a major role in C5b-9-induced cell cycle activation, we also investigated the effect of inhibiting Kv1.3 channels on DNA synthesis. rOsK-1 significantly inhibited the DNA synthesis induced by C5b-9 in OLG, indicating that Kv1.3 plays an important role in the C5b-9-induced cell cycle. In addition, C5b-9-mediated myelin basic protein and proteolipid protein mRNA decay was completely abrogated by inhibition of Kv1.3 expression. In the brains of multiple sclerosis patients, C5b-9 co-localized with NG2+ OLG progenitor cells that expressed Kv1.3 channels. Taken together, these data suggest that Kv1.3 channels play an important role in controlling C5b-9-induced cell cycle activation and OLG dedifferentiation, both in vitro and in vivo. PMID:21540025

  5. KCNE4 is an inhibitory subunit to Kv1.1 and Kv1.3 potassium channels

    DEFF Research Database (Denmark)

    Grunnet, Morten; Rasmussen, Hannne B; Hay-Schmidt, Anders

    2003-01-01

    is detected in the heart and in five different parts of the brain. Having the broad distribution of Kv1 channels in mind, the demonstrated inhibitory property of KCNE4-subunits could locally and/or transiently have a dramatic influence on cellular excitability and on setting resting membrane potentials....

  6. A New Negative Allosteric Modulator AP14145 for the Study of Small Conductance Calcium-Activated Potassium Channels

    DEFF Research Database (Denmark)

    Simo Vicens, Rafel; Kirchhoff, Jeppe Egedal; Dolce, Bernardo

    2017-01-01

    Background and purpose: Small conductance Ca2+-activated K+ (KCa2) channels represent a promising atrial-selective target for treatment of atrial fibrillation (AF). Here, we establish the mechanism of KCa2 inhibition by the new compound AP14145. Experimental approach: Using site directed mutagene......Background and purpose: Small conductance Ca2+-activated K+ (KCa2) channels represent a promising atrial-selective target for treatment of atrial fibrillation (AF). Here, we establish the mechanism of KCa2 inhibition by the new compound AP14145. Experimental approach: Using site directed...... mutagenesis binding determinants for AP14145 inhibition were explored. AP14145 selectivity and mechanism of action were investigated by patch clamp recordings of heterologously expressed KCa2 channels. The biological efficacy of AP14145 was assessed by measuring atrial effective refractory period (AERP......) prolongation in anaesthetised rats and a beam walk test was performed in mice to determine acute CNS related effects of the drug. Key results: AP14145 was found to be an equipotent negative allosteric modulator of KCa2.2 and KCa2.3 channels (IC50 = 1.1 ± 0.3 μM L-1). The presence of AP14145 (10 μM L-1...

  7. A New Negative Allosteric Modulator AP14145 for the Study of Small Conductance Calcium-Activated Potassium Channels

    DEFF Research Database (Denmark)

    Simo Vicens, Rafel; Kirchhoff, Jeppe Egedal; Dolce, Bernardo

    2017-01-01

    Background and purpose: Small conductance Ca2+-activated K+ (KCa2) channels represent a promising atrial-selective target for treatment of atrial fibrillation (AF). Here, we establish the mechanism of KCa2 inhibition by the new compound AP14145. Experimental approach: Using site directed mutagene......Background and purpose: Small conductance Ca2+-activated K+ (KCa2) channels represent a promising atrial-selective target for treatment of atrial fibrillation (AF). Here, we establish the mechanism of KCa2 inhibition by the new compound AP14145. Experimental approach: Using site directed......) prolongation in anaesthetised rats and a beam walk test was performed in mice to determine acute CNS related effects of the drug. Key results: AP14145 was found to be an equipotent negative allosteric modulator of KCa2.2 and KCa2.3 channels (IC50 = 1.1 ± 0.3 μM L-1). The presence of AP14145 (10 μM L-1...... inhibition is accomplished at the molecular level will help future development of drugs targeting KCa2 channels....

  8. Definitive Endoderm Formation from Plucked Human Hair-Derived Induced Pluripotent Stem Cells and SK Channel Regulation

    Directory of Open Access Journals (Sweden)

    Anett Illing

    2013-01-01

    Full Text Available Pluripotent stem cells present an extraordinary powerful tool to investigate embryonic development in humans. Essentially, they provide a unique platform for dissecting the distinct mechanisms underlying pluripotency and subsequent lineage commitment. Modest information currently exists about the expression and the role of ion channels during human embryogenesis, organ development, and cell fate determination. Of note, small and intermediate conductance, calcium-activated potassium channels have been reported to modify stem cell behaviour and differentiation. These channels are broadly expressed throughout human tissues and are involved in various cellular processes, such as the after-hyperpolarization in excitable cells, and also in differentiation processes. To this end, human induced pluripotent stem cells (hiPSCs generated from plucked human hair keratinocytes have been exploited in vitro to recapitulate endoderm formation and, concomitantly, used to map the expression of the SK channel (SKCa subtypes over time. Thus, we report the successful generation of definitive endoderm from hiPSCs of ectodermal origin using a highly reproducible and robust differentiation system. Furthermore, we provide the first evidence that SKCas subtypes are dynamically regulated in the transition from a pluripotent stem cell to a more lineage restricted, endodermal progeny.

  9. Potassium-chloride cotransporter 3 interacts with Vav2 to synchronize the cell volume decrease response with cell protrusion dynamics.

    Directory of Open Access Journals (Sweden)

    Adèle Salin-Cantegrel

    Full Text Available Loss-of-function of the potassium-chloride cotransporter 3 (KCC3 causes hereditary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC, a severe neurodegenerative disease associated with defective midline crossing of commissural axons in the brain. Conversely, KCC3 over-expression in breast, ovarian and cervical cancer is associated with enhanced tumor cell malignancy and invasiveness. We identified a highly conserved proline-rich sequence within the C-terminus of the cotransporter which when mutated leads to loss of the KCC3-dependent regulatory volume decrease (RVD response in Xenopus Laevis oocytes. Using SH3 domain arrays, we found that this poly-proline motif is a binding site for SH3-domain containing proteins in vitro. This approach identified the guanine nucleotide exchange factor (GEF Vav2 as a candidate partner for KCC3. KCC3/Vav2 physical interaction was confirmed using GST-pull down assays and immuno-based experiments. In cultured cervical cancer cells, KCC3 co-localized with the active form of Vav2 in swelling-induced actin-rich protruding sites and within lamellipodia of spreading and migrating cells. These data provide evidence of a molecular and functional link between the potassium-chloride co-transporters and the Rho GTPase-dependent actin remodeling machinery in RVD, cell spreading and cell protrusion dynamics, thus providing new insights into KCC3's involvement in cancer cell malignancy and in corpus callosum agenesis in HMSN/ACC.

  10. Surface engineering of titanium with potassium hydroxide and its effects on the growth behavior of mesenchymal stem cells.

    Science.gov (United States)

    Cai, Kaiyong; Lai, Min; Yang, Weihu; Hu, Ran; Xin, Renlong; Liu, Qing; Sung, K L Paul

    2010-06-01

    To improve the corrosion resistance and biological performance of commercially pure titanium (cp-Ti) substrates, potassium hydroxide was employed to modify the surfaces of titanium substrates, followed by biomimetic deposition of apatite on the substrates in a simulated body fluid. The morphologies of native and treated titanium substrates were characterized by field emission scanning electron microscopy (FE-SEM). Treatment with potassium hydroxide led to the formation of intermediate layers of potassium titanate on the surfaces of titanium substrates, while apatite was subsequently deposited onto the intermediate layer. The formation of potassium titanate and apatite was confirmed by thin-film X-ray diffraction and FE-SEM equipped with energy dispersive spectroscopy, respectively. Electrochemical impedance spectroscopy showed that the formed potassium titanate layer improved the corrosion-resistance properties of titanium substrates. The influence of modified titanium substrates on the biological behavior of mesenchymal stem cells (MSCs), including osteogenic differentiation, was investigated in vitro. Compared with cp-Ti substrates, MSCs cultured onto alkali- and heat-treated titanium substrates and apatite-deposited titanium substrates displayed significantly higher (P<0.05 or P<0.01) proliferation and differentiation levels of alkaline phosphatase and osteocalcin in 7 and 14day cultures, respectively. More importantly, our results suggest that the modified titanium substrates have great potential for inducing MSCs to differentiate into osteoblasts. The approach presented here may be exploited to fabricate titanium-based implants. Copyright 2009 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  11. Potassium accumulation between type I hair cells and calyx terminals in mouse crista.

    Science.gov (United States)

    Lim, Rebecca; Kindig, Angela E; Donne, Scott W; Callister, Robert J; Brichta, Alan M

    2011-05-01

    The mode of synaptic transmission in the vestibular periphery, between type I hair cells and their associated calyx terminal, has been the subject of much debate. The close and extensive apposition of pre- and post-synaptic elements has led some to suggest potassium (K(+)) accumulates in the intercellular space and even plays a role in synaptic transmission. During patch clamp recordings from isolated and embedded hair cells in a semi-intact preparation of the mouse cristae, we noted marked differences in whole-cell currents. Embedded type I hair cells show a prominent droop during steady-state activation as well as a dramatic collapse in tail currents. Responses to a depolarizing voltage step (-124 to +16 mV) in embedded, but not isolated, hair cells resulted in a >40 mV shift of the K(+) equilibrium potential and a rise in effective K(+) concentration (>50 mM) in the intercellular space. Together these data suggest K(+) accumulation in the intercellular space accounts for the different responses in isolated and embedded type I hair cells. To test this notion, we exposed the preparation to hyperosmotic solutions to enlarge the intercellular space. As predicted, the K(+) accumulation effects were reduced; however, a fit of our data with a classic diffusion model suggested K(+) permeability, rather than the intercellular space, had been altered by the hyperosmotic change. These results support the notion that under depolarizing conditions substantial K(+) accumulation occurs in the space between type I hair cells and calyx. The extent of K(+) accumulation during normal synaptic transmission, however, remains to be determined.

  12. Blueberry juice causes potent relaxation of rat aortic rings via the activation of potassium channels and the H₂S pathway.

    Science.gov (United States)

    Horrigan, Louise A; Holohan, Catherine A; Lawless, Gráinne A; Murtagh, Melissa A; Williams, Carmel T; Webster, Christina M

    2013-02-26

    The objective of this study was to investigate the in vitro effects of blueberry juice on healthy rat aortic rings, and to explore the roles of potassium channels and of the hydrogen sulphide (H(2)S) pathway in mediating the effects of blueberry juice. Firstly, the antioxidant capacity of blueberry juice was compared to other popular juice drinks using the Folin-Ciocalteu and the DPPH assays. Blueberry juice had significantly higher total polyphenol content than any of the other drinks studied (p blueberry juice on noradrenaline-contracted aortic rings was then observed, and the juice caused significant inhibition of noradrenaline-induced contractions (p blueberry juice (p blueberry juice (p blueberry juice has potent vasorelaxing properties, and thus may be a useful dietary agent for the prevention and treatment of hypertension. This study also provides strong evidence that Kv channels and the CSE/H(2)S pathway may be responsible, at least in part, for mediating the effects of blueberry juice.

  13. The role of potassium BK channels in anticonvulsant effect of cannabidiol in pentylenetetrazole and maximal electroshock models of seizure in mice.

    Science.gov (United States)

    Shirazi-zand, Zahra; Ahmad-Molaei, Leila; Motamedi, Fereshteh; Naderi, Nima

    2013-07-01

    Cannabidiol is a nonpsychoactive member of phytocannabinoids that produces various pharmacological effects that are not mediated through putative CB1/CB2 cannabinoid receptors and their related effectors. In this study, we examined the effect of the i.c.v. administration of potassium BK channel blocker paxilline alone and in combination with cannabidiol in protection against pentylenetetrazol (PTZ)- and maximal electroshock (MES)-induced seizure in mice. In the PTZ-induced seizure model, i.c.v. administration of cannabidiol caused a significant increase in seizure threshold compared with the control group. Moreover, while i.c.v. administration of various doses of paxilline did not produce significant change in the PTZ-induced seizure threshold in mice, coadministration of cannabidiol and paxilline attenuated the antiseizure effect of cannabidiol in PTZ-induced tonic seizures. In the MES model of seizure, both cannabidiol and paxilline per se produced significant increase in percent protection against electroshock-induced seizure. However, coadministration of cannabidiol and paxilline did not produce significant interaction in their antiseizure effect in the MES test. The results of the present study showed a protective effect of cannabidiol in both PTZ and MES models of seizure. These results suggested a BK channel-mediated antiseizure action of cannabidiol in PTZ model of seizure. However, such an interaction might not exist in MES-induced convulsion. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. Active Dendrites and Differential Distribution of Calcium Channels Enable Functional Compartmentalization of Golgi Cells.

    Science.gov (United States)

    Rudolph, Stephanie; Hull, Court; Regehr, Wade G

    2015-11-25

    Interneurons are essential to controlling excitability, timing, and synaptic integration in neuronal networks. Golgi cells (GoCs) serve these roles at the input layer of the cerebellar cortex by releasing GABA to inhibit granule cells (grcs). GoCs are excited by mossy fibers (MFs) and grcs and provide feedforward and feedback inhibition to grcs. Here we investigate two important aspects of GoC physiology: the properties of GoC dendrites and the role of calcium signaling in regulating GoC spontaneous activity. Although GoC dendrites are extensive, previous studies concluded they are devoid of voltage-gated ion channels. Hence, the current view holds that somatic voltage signals decay passively within GoC dendrites, and grc synapses onto distal dendrites are not amplified and are therefore ineffective at firing GoCs because of strong passive attenuation. Using whole-cell recording and calcium imaging in rat slices, we find that dendritic voltage-gated sodium channels allow somatic action potentials to activate voltage-gated calcium channels (VGCCs) along the entire dendritic length, with R-type and T-type VGCCs preferentially located distally. We show that R- and T-type VGCCs located in the dendrites can boost distal synaptic inputs and promote burst firing. Active dendrites are thus critical to the regulation of GoC activity, and consequently, to the processing of input to the cerebellar cortex. In contrast, we find that N-type channels are preferentially located near the soma, and control the frequency and pattern of spontaneous firing through their close association with calcium-activated potassium (KCa) channels. Thus, VGCC types are differentially distributed and serve specialized functions within GoCs. Interneurons are essential to neural processing because they modulate excitability, timing, and synaptic integration within circuits. At the input layer of the cerebellar cortex, a single type of interneuron, the Golgi cell (GoC), carries these functions. The

  15. Monitoring single-channel water permeability in polarized cells.

    Science.gov (United States)

    Erokhova, Liudmila; Horner, Andreas; Kügler, Philipp; Pohl, Peter

    2011-11-18

    So far the determination of unitary permeability (p(f)) of water channels that are expressed in polarized cells is subject to large errors because the opening of a single water channel does not noticeably increase the water permeability of a membrane patch above the background. That is, in contrast to the patch clamp technique, where the single ion channel conductance may be derived from a single experiment, two experiments separated in time and/or space are required to obtain the single-channel water permeability p(f) as a function of the incremental water permeability (P(f,c)) and the number (n) of water channels that contributed to P(f,c). Although the unitary conductance of ion channels is measured in the native environment of the channel, p(f) is so far derived from reconstituted channels or channels expressed in oocytes. To determine the p(f) of channels from live epithelial monolayers, we exploit the fact that osmotic volume flow alters the concentration of aqueous reporter dyes adjacent to the epithelia. We measure these changes by fluorescence correlation spectroscopy, which allows the calculation of both P(f,c) and osmolyte dilution within the unstirred layer. Shifting the focus of the laser from the aqueous solution to the apical and basolateral membranes allowed the FCS-based determination of n. Here we validate the new technique by determining the p(f) of aquaporin 5 in Madin-Darby canine kidney cell monolayers. Because inhibition and subsequent activity rescue are monitored on the same sample, drug effects on exocytosis or endocytosis can be dissected from those on p(f).

  16. Fenamates block gap junction coupling and potentiate BKCa channels in guinea pig arteriolar cells

    Science.gov (United States)

    Li, Xin-Zhi; Ma, Ke-Tao; Guan, Bing-Cai; Li, Li; Zhao, Lei; Zhang, Zhong-Shuang; Si, Jun-Qiang; Jiang, Zhi-Gen

    2013-01-01

    We determined the actions of the fenamates, flufenamic acid (FFA) and niflumic acid (NFA), on gap junction-mediated intercellular coupling between vascular smooth muscle cells (VSMC) in situ of acutely isolated arteriole segments from the three vascular beds: the spiral modiolar artery (SMA), anterior inferior cerebellar artery (AICA) and mesenteric artery (MA), and on non-junctional membrane channels in dispersed VSMCs. Conventional whole-cell recording methods were used. FFA reversibly suppressed the input conductance (Ginput) or increased the input resistance (Rinput) in a concentration dependent manner, with slightly different IC50s for the SMA, AICA and MA segments (26, 33 and 56 μM respectively, P>0.05). Complete electrical isolation of the recorded VSMC was normally reached at ≥300 μM. NFA had a similar effect on gap junction among VSMCs with an IC50 of 40, 48 and 62 μM in SMA, AICA and MA segments, respectively. In dispersed VSMCs, FFA and NFA increased outward rectifier K+-current mediated by the big conductance calcium-activated potassium channel (BKCa) in a concentration-dependent manner, with a similar EC50 of ~300 μM for both FFA and NFA in the three vessels. Iberiotoxin, a selective blocker of the BKCa, suppressed the enhancement of the BKCa by FFA and NFA. The KV blocker 4-AP had no effect on the fenamates-induced K+-current enhancement. We conclude that FFA and NFA blocked the vascular gap junction mediated electrical couplings uniformly in arterioles of the three vascular beds, and complete electrical isolation of the recorded VSMC is obtained at ≧300 μM; FFA and NFA also activate BKCa channels in the arteriolar smooth muscle cells in addition to their known inhibitory effects on chloride channels. PMID:23420003

  17. Potassium Sensing by Renal Distal Tubules Requires Kir4.1.

    Science.gov (United States)

    Cuevas, Catherina A; Su, Xiao-Tong; Wang, Ming-Xiao; Terker, Andrew S; Lin, Dao-Hong; McCormick, James A; Yang, Chao-Ling; Ellison, David H; Wang, Wen-Hui

    2017-06-01

    The mammalian distal convoluted tubule (DCT) makes an important contribution to potassium homeostasis by modulating NaCl transport. The thiazide-sensitive Na + /Cl - cotransporter (NCC) is activated by low potassium intake and by hypokalemia. Coupled with suppression of aldosterone secretion, activation of NCC helps to retain potassium by increasing electroneutral NaCl reabsorption, therefore reducing Na + /K + exchange. Yet the mechanisms by which DCT cells sense plasma potassium concentration and transmit the information to the apical membrane are not clear. Here, we tested the hypothesis that the potassium channel Kir4.1 is the potassium sensor of DCT cells. We generated mice in which Kir4.1 could be deleted in the kidney after the mice are fully developed. Deletion of Kir4.1 in these mice led to moderate salt wasting, low BP, and profound potassium wasting. Basolateral membranes of DCT cells were depolarized, nearly devoid of conductive potassium transport, and unresponsive to plasma potassium concentration. Although renal WNK4 abundance increased after Kir4.1 deletion, NCC abundance and function decreased, suggesting that membrane depolarization uncouples WNK kinases from NCC. Together, these results indicate that Kir4.1 mediates potassium sensing by DCT cells and couples this signal to apical transport processes. Copyright © 2017 by the American Society of Nephrology.

  18. Charybdotoxin Unbinding from the mKv1.3 Potassium Channel: A Combined Computational and Experimental Study

    Czech Academy of Sciences Publication Activity Database

    Khabiri, Morteza; Nikouee, A.; Cwiklik, Lukasz; Grissmer, S.; Ettrich, Rüdiger

    2011-01-01

    Roč. 115, č. 39 (2011), s. 11490-11500 ISSN 1520-6106 R&D Projects: GA ČR GA203/08/0114 Institutional research plan: CEZ:AV0Z60870520; CEZ:AV0Z40400503 Keywords : molecular-dynamics simulations * Gated K+ Channels * induced conformational-changes * binding free-energy * C-type inactivation Subject RIV: EH - Ecology, Behaviour Impact factor: 3.696, year: 2011

  19. Discovery, characterization and structure-activity relationships of an inhibitor of inward rectifier potassium (Kir channels with preference for Kir2.3, Kir3.X and Kir7.1

    Directory of Open Access Journals (Sweden)

    Jerod S Denton

    2011-11-01

    Full Text Available The inward rectifier family of potassium (Kir channels is comprised of at least 16 family members exhibiting broad and often overlapping cellular, tissue or organ distributions. The discovery of disease-causing mutations in humans and experiments on knockout mice has underscored the importance of Kir channels in physiology and in some cases raised questions about their potential as drug targets. However, the paucity of potent and selective small-molecule modulators targeting specific family members has with few exceptions mired efforts to understand their physiology and assess their therapeutic potential. A growing body of evidence suggests that GIRK (G protein-regulated inward rectifier K channels of the Kir3.X subfamily may represent novel targets for the treatment of atrial fibrillation. In an effort to expand the molecular pharmacology of GIRK, we performed a thallium (Tl+ flux-based high-throughput screen (HTS of a Kir1.1 inhibitor library for modulators of GIRK. One compound, termed VU573, exhibited 10-fold selectivity for GIRK over Kir1.1 (IC50 = 1.9 M and 19 M, respectively and was therefore selected for further study. In electrophysiological experiments performed on Xenopus laevis oocytes and mammalian cells, VU573 inhibited Kir3.1/3.2 (neuronal GIRK and Kir3.1/3.4 (cardiac GIRK channels with equal potency and preferentially inhibited GIRK, Kir2.3 and Kir7.1 over Kir1.1 and Kir2.1. Tl+ flux assays were established for Kir2.3 and the M125R pore mutant of Kir7.1 to support medicinal chemistry efforts to develop more potent and selective analogs for these channels. The structure-activity relationships of VU573 revealed few analogs with improved potency, however two compounds retained most of their activity toward GIRK and Kir2.3 and lost activity toward Kir7.1. We anticipate that the VU573 series will be useful for exploring the physiology and structure-function relationships of these Kir channels.

  20. Increased tolerance to stress in cardiac expressed gain-of-function of adenosine triphosphate-sensitive potassium channel subunit Kir6.1.

    Science.gov (United States)

    Henn, Matthew C; Janjua, M Burhan; Zhang, Haixia; Kanter, Evelyn M; Makepeace, Carol M; Schuessler, Richard B; Nichols, Colin G; Lawton, Jennifer S

    2016-12-01

    The adenosine triphosphate-sensitive potassium (K ATP ) channel opener diazoxide (DZX) prevents myocyte volume derangement and reduced contractility secondary to stress. K ATP channels are composed of pore-forming (Kir6.1 or Kir6.2) and regulatory (sulfonylurea receptor, SUR1 or SUR2) subunits. Gain of function (GOF) of Kir6.1 subunits has been implicated in cardiac pathology in Cantu syndrome in humans (cardiomegaly, lymphedema, and pericardial effusions). We hypothesized that GOF of Kir6.1 subunits would result in altered myocyte response to stress. Isolated cardiac myocytes from wild type (WT) and transgenic Kir6.1GOF mice were exposed to Tyrode's physiologic solution for 20 min, test solution (Tyrode's or stress [hyperkalemic cardioplegia {CPG, known myocyte stress}] +/- K ATP channel opener DZX), followed by Tyrode's for 20 min. Myocyte volume and contractility were measured and compared. WT myocytes demonstrated significant swelling in response to stress, but significantly less swelling was seen in Kir6.1GOF myocytes. DZX prevented swelling secondary to CPG in WT but resulted in a nonsignificant reduction in swelling in Kir6.1GOF myocytes. Both WT and Kir6.1GOF myocytes demonstrated a reduction in contractility during stress, although this was only significant in Kir6.1GOF myocytes. DZX was not associated with an improvement in contractility in Kir6.1GOF myocytes following stress. Similar to previous results in Kir6.1(-/-) myocytes, Kir6.1GOF myocytes demonstrate resistance (less volume derangement) to stress of cardioplegia. Understanding the role of Kir6.1 in myocyte response to stress may aid in the treatment of patients with Cantu syndrome and warrants further investigation. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. The polarization of the G-protein activated potassium channel GIRK5 to the vegetal pole of Xenopus laevis oocytes is driven by a di-leucine motif.

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    Beatriz Díaz-Bello

    Full Text Available The G protein-coupled inwardly-rectifying potassium channels (known as GIRK or Kir3 form functional heterotetramers gated by G-βγ subunits. GIRK channels participate in heart rate modulation and neuronal postsynaptic inhibition in mammals. In Xenopus laevis oocytes, GIRK5 is a functional homomultimer. Previously, we found that phosphorylation of a tyrosine (Y16 at its N-terminus downregulates the surface expression of GIRK5. In this work, we elucidated the subcellular localization and trafficking of GIRK5 in oocytes. Several EGFP-GIRK5 chimeras were produced and an ECFP construct was used to identify the endoplasmic reticulum (ER. Whereas GIRK5-WT was retained in the ER at the animal pole, the phospho-null GIRK5-Y16A was localized to the vegetal pole. Interestingly, a construct with an N-terminal Δ25 deletion produced an even distribution of the channel in the whole oocyte. Through an alanine-scan, we identified an acidic cluster/di-leucine sorting-signal recognition motif between E17 and I22. We quantified the effect of each amino acid residue within this di-leucine motif in determining the distribution of GIRK5 to the animal and vegetal poles. We found that Y16 and I22 contributed to functional expression and were dominant in the polarization of GIRK5. We thus conclude that the N-terminal acidic di-leucine motif of GIRK5 determines its retention and polarized trafficking within Xl oocytes.

  2. Role of transient receptor potential melastatin-like subtype 5 channel in insulin secretion from rat β-cells.

    Science.gov (United States)

    Krishnan, Kalaiselvan; Ma, Zuheng; Björklund, Anneli; Islam, Md Shahidul

    2014-05-01

    Several studies have reported that the transient receptor potential melastatin-like subtype 5 (TRPM5) channel, a Ca(2+)-activated monovalent cation channel, is involved in the stimulus-secretion coupling in the mouse pancreatic β-cells. We have studied the role of the TRPM5 channel in regulating insulin secretion and cytoplasmic free Ca(2+) concentration ([Ca(2+)](i)) in the rat β-cells by using triphenylphosphine oxide, a selective inhibitor of the channel. Insulin secretion from islets from Sprague-Dawley rats was measured in batch incubations. Cytoplasmic free Ca(2+) concentration was measured from single β-cells by fura-2-based microfluorometry. Triphenylphosphine oxide did not alter insulin secretion and [Ca(2+)](i) response triggered by KCl or fructose. It inhibited insulin secretion in response to glucose, L-arginine, and glucagon-like peptide 1. It also inhibited glucose-induced insulin secretion by mechanisms that are independent of the adenosine triphosphate-sensitive potassium channels and [Ca(2+)](i) increase. Our results suggest that in the rat islets, TRPM5 is involved in mediating insulin secretion by glucose and l-arginine and in potentiating the glucose-induced insulin secretion by glucagon-like peptide 1.

  3. Activation of BK(Ca channels in zoledronic acid-induced apoptosis of MDA-MB-231 breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Yu-Guang Ma

    Full Text Available BACKGROUND: Zoledronic acid, one of the most potent nitrogen-containing biphosphonates, has been demonstrated to have direct anti-tumor and anti-metastatic properties in breast cancer in vitro and in vivo. In particular, tumor-cell apoptosis has been recognized to play an important role in the treatment of metastatic breast cancer with zoledronic acid. However, the precise mechanisms remain less clear. In the present study, we investigated the specific role of large conductance Ca(2+-activated potassium (BK(Ca channel in zoledronic acid-induced apoptosis of estrogen receptor (ER-negative MDA-MB-231 breast cancer cells. METHODOLOGY/PRINCIPAL FINDINGS: The action of zoledronic acid on BK(Ca channel was investigated by whole-cell and cell-attached patch clamp techniques. Cell apoptosis was assessed with immunocytochemistry, analysis of fragmented DNA by agarose gel electrophoresis, and flow cytometry assays. Cell proliferation was investigated by MTT test and immunocytochemistry. In addition, such findings were further confirmed with human embryonic kidney 293 (HEK293 cells which were transfected with functional BK(Ca α-subunit (hSloα. Our results clearly indicated that zoledronic acid directly increased the activities of BK(Ca channels, and then activation of BK(Ca channel by zoledronic acid contributed to induce apoptosis in MDA-MB-231 cells. The possible mechanisms were associated with the elevated level of intracellular Ca(2+ and a concomitant depolarization of mitochondrial membrane potential (Δψm in MDA-MB-231 cells. CONCLUSIONS: Activation of BK(Ca channel was here shown to be a novel molecular pathway involved in zoledronic acid-induced apoptosis of MDA-MB-231 cells in vitro.

  4. β-adrenergic Receptor Blocker ICI 118,551 Selectively Increases Intermediate-Conductance Calcium-Activated Potassium Channel (IKCa)-Mediated Relaxations in Rat Main Mesenteric Artery.

    Science.gov (United States)

    Ozkan, Melike Hacer; Uma, Serdar

    2017-12-26

    Endothelial IK C a and/or SK C a channels play an important role in the control of vascular tone by participating in endothelium-dependent relaxation. Whether β-AR antagonists, mainly used in hypertension, affect endothelial K C a channel function is unknown. In this study, we examined the effect of the β2-AR antagonist and inverse agonist ICI 118,551 on the IK C a /SK C a channel activity by assessing functional relaxation responses to several agonists that stimulate these channels. Mesenteric arterial rings isolated from male Sprague Dawley mounted to organ baths. Acetylcholine elicited IK C a - and SK C a -mediated relaxations that were abolished by TRAM-34 and apamin, respectively. ICI 118,551, which did not dilate the arteries per se, increased the IK C a -mediated relaxations, whereas SK C a -mediated relaxations remained unaltered. Same potentiating effect was also detected on the IK C a -mediated relaxations to carbachol and A23187, but not to NS309. Neither acetylcholine-induced nitric oxide-mediated relaxations nor SNP relaxations changed with ICI 118,551. The PKA inhibitor KT-5720, the selective β2-AR agonist salbutamol, the selective β2-AR antagonist butoxamine, the non-selective β-AR antagonist propranolol, and the inverse agonists carvedilol or nadolol failed to affect the IK C a -mediated relaxations. ICI 118,551-induced increase was not reversed by salbutamol or propranolol as well. Besides, low potassium-induced relaxations in endothelium-removed arteries remained the same in the presence of ICI 118,551. These data demonstrate a previously unrecognized action of ICI 118,551, the ability to potentiate endothelial IK C a channel-mediated vasodilation, through a mechanism independent of β2-AR antagonistic or inverse agonistic action. Instead, the enhancement of acetylcholine relaxation seems likely to occur by a mechanism secondary to endothelial calcium increase. © 2017 Nordic Association for the Publication of BCPT (former Nordic

  5. Prevention of isoflurane-induced preconditioning by 5-hydroxydecanoate and gadolinium: possible involvement of mitochondrial adenosine triphosphate-sensitive potassium and stretch-activated channels.

    Science.gov (United States)

    Piriou, V; Chiari, P; Knezynski, S; Bastien, O; Loufoua, J; Lehot, J J; Foëx, P; Annat, G; Ovize, M

    2000-09-01

    Both mitochondrial adenosine triphosphate-sensitive potassium (MKATP) channels (selectively blocked by 5-hydroxydecanoate) and stretch-activated channels (blocked by gadolinium) have been involved in the mechanism of ischemic preconditioning. Isoflurane can reproduce the protection afforded by ischemic preconditioning. We sought to determine whether isoflurane-induced preconditioning may involve MKATP and stretch-activated channels. Anesthetized open-chest rabbits underwent 30 min of coronary occlusion followed by 3 h of reperfusion. Before this, rabbits were randomized into one of six groups and underwent a treatment period consisting of either no intervention for 40 min (control group; n = 9) or 15 min of isoflurane inhalation (1.1% end tidal) followed by a 15-min washout period (isoflurane group; n = 9). The two groups received an intravenous bolus dose of either 5-hydroxydecanoate (5 mg/kg) or gadolinium (40 micromol/kg) before coronary occlusion and reperfusion (5-hydroxydecanoate, n = 9; gadolinium, n = 7). Two additional groups received 5-hydroxydecanoate or gadolinium before isoflurane exposure (isoflurane-5-hydroxydecanoate, n = 10; isoflurane-gadolinium, n = 8). Area at risk and infarct size were assessed by blue dye injection and tetrazolium chloride staining. Area at risk was comparable among the six groups (29 +/- 7, 30 +/- 5, 27 +/- 6, 35 +/- 7, 31 +/- 7, and 27 +/- 4% of the left ventricle in the control, isoflurane, isoflurane-5-hydroxydecanoate, 5-hydroxydecanoate, isoflurane-gadolinium, and gadolinium groups, respectively). Infarct size averaged 60 +/- 20% (SD) in untreated controls versus 54 +/- 27 and 65 +/- 15% of the risk zone in 5-hydroxydecanoate- and gadolinium-treated controls (P = nonsignificant). In contrast, infarct size in the isoflurane group was significantly reduced to 26 +/- 11% of the risk zone (P < 0.05 vs.control). Both 5-hydroxydecanoate and gadolinium prevented this attenuation: infarct size averaged 68 +/- 23 and 56 +/- 21

  6. Relaxant effect of a novel calcium-activated potassium channel modulator on human myometrial spontaneous contractility in vitro

    DEFF Research Database (Denmark)

    Rosenbaum, S.T.; Larsen, T.; Joergensen, J.C.

    2012-01-01

    Aim: To investigate the effect of 4,5-dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one (NS4591), a novel SK/IK channels positive modulator, on human myometrial activity. Methods: Organ bath studies were performed on myometrial preparations obtained from women undergoing elective caesarean....... Simultaneous vehicle controls were performed for all experiments. The effects of drugs were studied on spontaneous contractions. Results: NS4591 exerted an inhibitory effect on myometrial contractions in muscle strips from non-pregnant and pregnant women. The contractility in non-pregnant and pregnant...

  7. Performance enhancement of PV cells through micro-channel cooling

    Directory of Open Access Journals (Sweden)

    Muzaffar Ali

    2015-11-01

    Full Text Available Efficiency of a PV cell is strongly dependent on its surface temperature. The current study is focused to achieve maximum efficiency of PV cells even in scorching temperatures in hot climates like Pakistan where the cell surface temperatures can even rise up to around 80 ℃. The study includes both the CFD and real time experimental investigations of a solar panel using micro channel cooling. Initially, CFD analysis is performed by developing a 3D model of a Mono-Crystalline cell with micro-channels to analyze cell surface temperature distribution at different irradiance and water flow rates. Afterwards, an experimental setup is developed for performance investigations under the real conditions of an open climate of a Pakistan's city, Taxila. Two 35W panels are manufactured for the experiments; one is based on the standard manufacturing procedure while other cell is developed with 4mm thick aluminum sheet having micro-channels of cross-section of 1mm by 1mm. The whole setup also includes different sensors for the measurement of solar irradiance, cell power, surface temperature and water flow rates. The experimental results show that PV cell surface temperature drop of around 15 ℃ is achieved with power increment of around 14% at maximum applied water flow rate of 3 LPM. Additionally, a good agreement is also found between CFD and experimental results. Therefore, that study clearly shows that a significant performance improvement of PV cells can be achieved through the proposed cell cooling technique.

  8. Reduced KCNQ4-encoded voltage-dependent potassium channel activity underlies impaired ß-adrenoceptor-mediated relaxation of renal arteries in hypertension

    DEFF Research Database (Denmark)

    Chadha, Preet S; Zunke, Friederike; Zhu, Hai-Lei

    2012-01-01

    expression of KCNQ1, KCNQ3, KCNQ4, KCNQ5, and Kv7.1, Kv7.4, and Kv7.5 in rat renal artery. Isoproterenol produced concentration-dependent relaxation of precontracted renal arteries and increased Kv7 channel currents in isolated smooth muscle cells. Application of the Kv7 blocker linopirdine attenuated...

  9. Antinociceptive action of oxytocin involves inhibition of potassium channel currents in lamina II neurons of the rat spinal cord

    Directory of Open Access Journals (Sweden)

    Darbon Pascal

    2009-11-01

    Full Text Available Abstract Background Growing evidence in the literature shows that oxytocin (OT has a strong spinal anti-nociceptive action. Oxytocinergic axons originating from a subpopulation of paraventricular hypothalamic neurons establish synaptic contacts with lamina II interneurons but little is known about the functional role of OT with respect to neuronal firing and excitability. Results Using the patch-clamp technique, we have recorded lamina II interneurons in acute transverse lumbar spinal cord slices of rats (15 to 30 days old and analyzed the OT effects on action potential firing ability. In the current clamp mode, we found that bath application of a selective OT-receptor agonist (TGOT reduced firing in the majority of lamina II interneurons exhibiting a bursting firing profile, but never in those exhibiting a single spike discharge upon depolarization. Interestingly, OT-induced reduction in spike frequency and increase of firing threshold were often observed, leading to a conversion of the firing profile from repetitive and delayed profiles into phasic ones and sometimes further into single spike profile. The observed effects following OT-receptor activation were completely abolished when the OT-receptor agonist was co-applied with a selective OT-receptor antagonist. In current and voltage clamp modes, we show that these changes in firing are strongly controlled by voltage-gated potassium currents. More precisely, transient IA currents and delayed-rectifier currents were reduced in amplitude and transient IA current was predominantly inactivated after OT bath application. Conclusion This effect of OT on the firing profile of lamina II neurons is in good agreement with the antinociceptive and analgesic properties of OT described in vivo.

  10. Administration of prostacyclin modulates cutaneous blood flow but not sweating in young and older males: roles for nitric oxide and calcium-activated potassium channels.

    Science.gov (United States)

    Fujii, Naoto; Notley, Sean R; Minson, Christopher T; Kenny, Glen P

    2016-11-01

    In young adults, cyclooxygenase (COX) contributes to the heat loss responses of cutaneous vasodilatation and sweating, and this may be mediated by prostacyclin-induced activation of nitric oxide synthase (NOS) and calcium-activated potassium (KCa) channels. This prostacyclin-induced response may be diminished in older relative to young adults because ageing is known to attenuate COX-dependent heat loss responses. We observed that, although prostacyclin does not mediate sweating in young and older males, it does modulate cutaneous vasodilatation, although the magnitude of increase is similar between groups. We also found that, although NOS and KCa channels contribute to prostacyclin-induced cutaneous vasodilatation in young males, these contributions are diminished in older males. Our findings provide new insight into the mechanisms governing heat loss responses and suggest that the age-related diminished COX-dependent heat loss responses reported in previous studies may be a result of the reduced COX-derived production of prostanoids (e.g., prostacyclin) rather than the decreased sensitivity of prostanoid receptors. Cyclooxygenase (COX) contributes to the regulation of cutaneous vasodilatation and sweating; however, the mechanism(s) underpinning this response remain unresolved. We hypothesized that prostacyclin (a COX-derived product) may directly mediate cutaneous vasodilatation and sweating through nitric oxide synthase (NOS) and calcium-activated potassium (KCa) channels in young adults. However, these responses would be diminished in older adults because ageing attenuates COX-dependent cutaneous vasodilatation and sweating. In young (25 ± 4 years) and older (60 ± 6 years) males (nine per group), cutaneous vascular conductance (CVC) and sweat rate were evaluated at four intradermal forearm skin sites: (i) control; (ii) 10 mm N G -nitro-l-arginine (l-NNA), a non-specific NOS inhibitor; (iii) 50 mm tetraethylammonium (TEA), a non-specific KCa channel

  11. Administration of prostacyclin modulates cutaneous blood flow but not sweating in young and older males: roles for nitric oxide and calcium‐activated potassium channels

    Science.gov (United States)

    Fujii, Naoto; Notley, Sean R.; Minson, Christopher T.

    2016-01-01

    Key points In young adults, cyclooxygenase (COX) contributes to the heat loss responses of cutaneous vasodilatation and sweating, and this may be mediated by prostacyclin‐induced activation of nitric oxide synthase (NOS) and calcium‐activated potassium (KCa) channels.This prostacyclin‐induced response may be diminished in older relative to young adults because ageing is known to attenuate COX‐dependent heat loss responses.We observed that, although prostacyclin does not mediate sweating in young and older males, it does modulate cutaneous vasodilatation, although the magnitude of increase is similar between groups.We also found that, although NOS and KCa channels contribute to prostacyclin‐induced cutaneous vasodilatation in young males, these contributions are diminished in older males.Our findings provide new insight into the mechanisms governing heat loss responses and suggest that the age‐related diminished COX‐dependent heat loss responses reported in previous studies may be a result of the reduced COX‐derived production of prostanoids (e.g., prostacyclin) rather than the decreased sensitivity of prostanoid receptors. Abstract Cyclooxygenase (COX) contributes to the regulation of cutaneous vasodilatation and sweating; however, the mechanism(s) underpinning this response remain unresolved. We hypothesized that prostacyclin (a COX‐derived product) may directly mediate cutaneous vasodilatation and sweating through nitric oxide synthase (NOS) and calcium‐activated potassium (KCa) channels in young adults. However, these responses would be diminished in older adults because ageing attenuates COX‐dependent cutaneous vasodilatation and sweating. In young (25 ± 4 years) and older (60 ± 6 years) males (nine per group), cutaneous vascular conductance (CVC) and sweat rate were evaluated at four intradermal forearm skin sites: (i) control; (ii) 10 mm N G‐nitro‐l‐arginine (l‐NNA), a non‐specific NOS inhibitor; (iii) 50

  12. Anomalous genotoxic responses induced in mouse lymphoma L5178Y cells by potassium bromate

    International Nuclear Information System (INIS)

    Priestley, Catherine C.; Green, Richard M.; Fellows, Michael D.; Doherty, Ann T.; Hodges, Nikolas J.; O'Donovan, Michael R.

    2010-01-01

    Potassium bromate (KBrO 3 ) is a well-established rodent kidney carcinogen and its oxidising activity is considered to be a significant factor in its mechanism of action. Although it has also been shown to be clearly genotoxic in a range of in vivo and in vitro test systems, surprisingly, it is not readily detected in several cell lines using the standard alkaline Comet assay. However, previous results from this laboratory demonstrated huge increases in tail intensity by modifying the method to include incubation with either human 8-oxodeoxyguanosine DNA glycosylase-1 (hOGG1) or bacterial formamidopyrimidine DNA glycosylase (FPG) indicating that, as expected, significant amounts of 8-oxodeoxyguanosine (8-OHdG) were induced. The purpose of this work, therefore, was to investigate why KBrO 3 , in contrast to other oxidising agents, gives a relatively poor response in the standard Comet assay. Results confirmed that it is a potent genotoxin in mouse lymphoma L5178Y cells inducing micronuclei and mutation at the tk and hprt loci at relatively non-cytotoxic concentrations. Subsequent time-course studies demonstrated that substantial amounts of 8-OHdG appear to remain in cells 24 h after treatment with KBrO 3 but result in no increase in frank stand breaks (FSB) even though phosphorylated histone H2AX (γ-H2AX) antibody labelling confirmed the presence of double-strand breaks. Using bromodeoxyuracil (BrdU) incorporation together with measured increases in cell numbers, L5178Y cells also appeared to go through the cell cycle with unrepaired hOGG1-recognisable damage. Since unrepaired 8-OHdG can give rise to point mutations through G:C → T:A transversions, it was also surprising that mutation could not be detected at the Na + /K + ATPase locus as determined by ouabain resistance. Some increases in strand breakage could be seen in the Comet assay by increasing the unwinding time, but only at highly toxic concentrations and to a much smaller extent than would be expected

  13. Apamin-Sensitive Small Conductance Calcium-Activated Potassium Channels were Negatively Regulated by Captopril in Volume-Overload Heart Failure Rats.

    Science.gov (United States)

    Hongyuan, Bai; Xin, Dong; Jingwen, Zhang; Li, Gao; Yajuan, Ni

    2016-08-01

    In heart failure (HF), the malignant arrhythmias occur frequently; a study demonstrated that upregulation of I KAS resulted in recurrent spontaneous ventricular fibrillation in HF. However, the regulation of SK channels was poorly understood. The activation of SK channels