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Sample records for voltage-dependent calcium currents

  1. Development of voltage-dependent calcium, sodium, and potassium currents in Xenopus spinal neurons.

    Science.gov (United States)

    O'Dowd, D K; Ribera, A B; Spitzer, N C

    1988-03-01

    Action potentials of embryonic nerve and muscle cells often have a different ionic dependence and longer duration than those of mature cells. The action potential of spinal cord neurons from Xenopus laevis exhibits a prominent calcium component at early stages of development that diminishes with age as the impulse becomes principally sodium dependent. Whole-cell voltage-clamp analysis has been undertaken to characterize the changes in membrane currents during development of these neurons in culture. Four voltage-dependent currents of cells were identified and examined during the first day in vitro, when most of the change in the action potential occurs. There are no changes in the peak density of the calcium current (ICa), its voltage dependence, or time to half-maximal activation; a small increase in inactivation is apparent. The major change in sodium current (INa) is a 2-fold increase in its density. In addition, more subtle changes in the kinetics of the macroscopic sodium current were noted. The peak density of voltage-dependent potassium current (IKv) increases 3-fold, and this current becomes activated almost twice as fast. No changes were noted in the extent of its inactivation. The calcium-dependent potassium current (IKc) consists of an inactivating and a sustained component. The former increases 2-fold in peak current density, and the latter increases similarly at less depolarized voltages. The changes in these currents contribute to the decrease in duration and the change in ionic dependence of the impulse.

  2. IgG anti-GalNAc-GD1a antibody inhibits the voltage-dependent calcium channel currents in PC12 pheochromocytoma cells.

    Science.gov (United States)

    Nakatani, Yoshihiko; Nagaoka, Takumi; Hotta, Sayako; Utsunomiya, Iku; Yoshino, Hiide; Miyatake, Tadashi; Hoshi, Keiko; Taguchi, Kyoji

    2007-03-01

    We investigated the effects of IgG anti-GalNAc-GD1a antibodies, produced by immunizing rabbits with GalNAc-GD1a, on the voltage-dependent calcium channel (VDCCs) currents in nerve growth factor (NGF)-differentiated PC12 pheochromocytoma cells. VDCCs currents in NGF-differentiated PC12 cells were recorded using the whole-cell patch-clamp technique. Immunized rabbit serum that had a high titer of anti-GalNAc-GD1a antibodies inhibited the VDCCs currents in the NGF-differentiated PC12 cells (36.0+/-9.6% reduction). The inhibitory effect of this serum was reversed to some degree within 3-4 min by washing with bath solution. Similarly, application of purified IgG from rabbit serum immunized with GalNAc-GD1a significantly inhibited the VDCCs currents in PC12 cells (30.6+/-2.5% reduction), and this inhibition was recovered by washing with bath solution. Furthermore, the inhibitory effect was also observed in the GalNAc-GD1a affinity column binding fraction (reduction of 31.1+/-9.85%), while the GalNAc-GD1a affinity column pass-through fraction attenuated the inhibitory effect on VDCCs currents. Normal rabbit serum and normal rabbit IgG did not affect the VDCCs currents in the PC12 cells. In an immunocytochemical study using fluorescence staining, the PC12 cells were stained using GalNAc-GD1a binding fraction. These results indicate that anti-GalNAc-GD1a antibodies inhibit the VDCCs currents in NGF-differentiated PC12 cells.

  3. Alterations of voltage-dependent calcium channel currents in basilar artery smooth muscle cells at early stage of subarachnoid hemorrhage in a rabbit model.

    Directory of Open Access Journals (Sweden)

    Xianqing Shi

    Full Text Available OBJECTIVE: To investigate the changes in the currents of voltage-dependent calcium channels (VDCCs in smooth muscle cells of basilar artery in a rabbit model of subarachnoid hemorrhage (SAH. METHODS: New Zealand white rabbits were randomly divided into five groups: sham (C, normal (N, 24 hours (S1, 48 hours (S2 and 72 hours (S3 after SAH. Non-heparinized autologous arterial blood (1 ml/kg was injected into the cisterna magna to create SAH after intravenous anesthesia, and 1 ml/kg of saline was injected into cisterna magna in the sham group. Rabbits in group N received no injections. Basilar artery in S1, S2, S3 group were isolated at 24, 48, 72 hours after SAH. Basilar artery in group C was isolated at 72 hours after physiological saline injection. Basilar artery smooth muscle cells were isolated for all groups. Whole-cell patch-clamp technique was utilized to record cell membrane capacitance and VDCCs currents. The VDCCs antagonist nifedipine was added to the bath solution to block the Ca(++ channels currents. RESULTS: There were no significant differences in the number of cells isolated, the cell size and membrane capacitance among all the five groups. VDCC currents in the S1-S3 groups had higher amplitudes than those in control and sham groups. The significant change of current amplitude was observed at 72 hours after SAH, which was higher than those of 24 and 48 hours. The VDCCs were shown to expression in human artery smooth muscle cells. CONCLUSIONS: The changes of activation characteristics and voltage-current relationship at 72 hours after SAH might be an important event which leads to a series of molecular events in the microenvironment of the basilar artery smooth muscle cells. This may be the key time point for potential therapeutic intervention against subarachnoid hemorrhage.

  4. Calmodulin and calcium differentially regulate the neuronal Nav1.1 voltage-dependent sodium channel

    Energy Technology Data Exchange (ETDEWEB)

    Gaudioso, Christelle; Carlier, Edmond; Youssouf, Fahamoe [INSERM U641, Institut Jean Roche, Marseille F-13344 (France); Universite de la Mediterranee, Faculte de Medecine Secteur Nord, IFR 11, Marseille F-13344 (France); Clare, Jeffrey J. [Eaton Pharma Consulting, Eaton Socon, Cambridgeshire PE19 8EF (United Kingdom); Debanne, Dominique [INSERM U641, Institut Jean Roche, Marseille F-13344 (France); Universite de la Mediterranee, Faculte de Medecine Secteur Nord, IFR 11, Marseille F-13344 (France); Alcaraz, Gisele, E-mail: gisele.alcaraz@univmed.fr [INSERM U641, Institut Jean Roche, Marseille F-13344 (France); Universite de la Mediterranee, Faculte de Medecine Secteur Nord, IFR 11, Marseille F-13344 (France)

    2011-07-29

    Highlights: {yields} Both Ca{sup ++}-Calmodulin (CaM) and Ca{sup ++}-free CaM bind to the C-terminal region of Nav1.1. {yields} Ca{sup ++} and CaM have both opposite and convergent effects on I{sub Nav1.1}. {yields} Ca{sup ++}-CaM modulates I{sub Nav1.1} amplitude. {yields} CaM hyperpolarizes the voltage-dependence of activation, and increases the inactivation rate. {yields} Ca{sup ++} alone antagonizes CaM for both effects, and depolarizes the voltage-dependence of inactivation. -- Abstract: Mutations in the neuronal Nav1.1 voltage-gated sodium channel are responsible for mild to severe epileptic syndromes. The ubiquitous calcium sensor calmodulin (CaM) bound to rat brain Nav1.1 and to the human Nav1.1 channel expressed by a stably transfected HEK-293 cell line. The C-terminal region of the channel, as a fusion protein or in the yeast two-hybrid system, interacted with CaM via a consensus C-terminal motif, the IQ domain. Patch clamp experiments on HEK1.1 cells showed that CaM overexpression increased peak current in a calcium-dependent way. CaM had no effect on the voltage-dependence of fast inactivation, and accelerated the inactivation kinetics. Elevating Ca{sup ++} depolarized the voltage-dependence of fast inactivation and slowed down the fast inactivation kinetics, and for high concentrations this effect competed with the acceleration induced by CaM alone. Similarly, the depolarizing action of calcium antagonized the hyperpolarizing shift of the voltage-dependence of activation due to CaM overexpression. Fluorescence spectroscopy measurements suggested that Ca{sup ++} could bind the Nav1.1 C-terminal region with micromolar affinity.

  5. Eugenol dilates rat cerebral arteries by inhibiting smooth muscle cell voltage-dependent calcium channels.

    Science.gov (United States)

    Peixoto-Neves, Dieniffer; Leal-Cardoso, Jose Henrique; Jaggar, Jonathan H

    2014-11-01

    Plants high in eugenol, a phenylpropanoid compound, are used as folk medicines to alleviate diseases including hypertension. Eugenol has been demonstrated to relax conduit and ear arteries and reduce systemic blood pressure, but mechanisms involved are unclear. Here, we studied eugenol regulation of resistance-size cerebral arteries that control regional brain blood pressure and flow and investigated mechanisms involved. We demonstrate that eugenol dilates arteries constricted by either pressure or membrane depolarization (60 mM K) in a concentration-dependent manner. Experiments performed using patch-clamp electrophysiology demonstrated that eugenol inhibited voltage-dependent calcium (Ca) currents, when using Ba as a charge carrier, in isolated cerebral artery smooth muscle cells. Eugenol inhibition of voltage-dependent Ca currents involved pore block, a hyperpolarizing shift (∼-10 mV) in voltage-dependent inactivation, an increase in the proportion of steady-state inactivating current, and acceleration of inactivation rate. In summary, our data indicate that eugenol dilates cerebral arteries by means of multimodal inhibition of voltage-dependent Ca channels.

  6. G Protein-induced Trafficking of Voltage-dependent Calcium Channels

    National Research Council Canada - National Science Library

    Eugene Tombler; Nory Jun Cabanilla; Paul Carman; Natasha Permaul; John J. Hall; Ryan W. Richman; Jessica Lee; Jennifer Rodriguez; Dan P. Felsenfeld; Robert F. Hennigan; María A. Diversé-Pierluissi

    2006-01-01

    .... Here we report a novel mechanism for G protein-mediated modulation of neuronal voltage-dependent calcium channels that involves the destabilization and subsequent removal of calcium channels from the plasma membrane...

  7. Voltage-dependent calcium channels from brain incorporated into planar lipid bilayers

    Science.gov (United States)

    Nelson, Mark T.; French, Robert J.; Krueger, Bruce K.

    1984-03-01

    Many important physiological processes, including neurotransmitter release and muscle contraction1-3, are regulated by the concentration of Ca2+ ions in the cell. Levels of cytoplasmic Ca2+ can be elevated by the entry of Ca2+ ions through voltage-dependent channels which are selective for Ca2+, Ba2+ and Sr2+ ions4-14. We have measured currents through single, voltage-dependent calcium channels from rat brain that have been incorporated into planar lipid bilayers. Channel gating was voltage-dependent: membrane depolarization increased the channel open times and decreased the closed times. The channels were selective for divalent cations over monovalent ions. The well-known calcium channel blockers, lanthanum and cadmium, produced a concentration-dependent reduction of the apparent single-channel conductance. Contrary to expectations14, the nature of the divalent cation carrying current through the channel affected not only the single-channel conductance, but also the channel open times, with mean open times being shortest for barium.

  8. The effects of S4 segments on the voltage-dependence of inactivation for Cav3.1 calcium channels

    Institute of Scientific and Technical Information of China (English)

    LI JunYing

    2007-01-01

    T-type calcium channels exhibit fast voltage-dependent inactivation,for which the underlying structure-function relationship still remains unclear.To investigate the roles of S4 segments in voltage-dependent inactivation of T-type calcium channels,we created S4 replacement chimeras between Cav3.1 calcium channels(fast voltage-dependent inactivation)and Cav1.2 calcium channels(little oltage-dependent inactivation)by replacing S4s in Cav3.1 with the corresponding regions in Cav1.2.Wild type and chimeric channels were expressed in Xenopus oocytes and channel currents were recorded with two-electrode voltage-clamp.We showed that replacing S4 region in domain I shifted voltage-dependence for inactivation of Cav3.1 to the left,and the V0.5 inact and kinact value were significantly changed.However replacing S4s in domains Ⅱ-Ⅳ had no effects on the voltage-dependent inactivation properties.These results suggest that the roles of S4 segments in domains Ⅰ-Ⅳ are different,and S4 in domain I is likely to be involved in voltage-dependent Inactivation process.Its movement during membrane depolarization may trigger a conformational change in the inactivation gate.

  9. Voltage-dependent currents in microvillar receptor cells of the frog vomeronasal organ.

    Science.gov (United States)

    Trotier, D; Døving, K B; Rosin, J F

    1993-08-01

    Vomeronasal receptor cells are differentiated bipolar neurons with a long dendrite bearing numerous microvilli. Isolated cells (with a mean dendritic length of 65 microns) and cells in mucosal slices were studied using whole-cell and Nystatin-perforated patch-clamp recordings. At rest, the membrane potential was -61 +/- 13 mV (mean +/- SD; n = 61). Sixty-four per cent of the cells had a resting potential in the range of -60 to -86 mV, with almost no spontaneous action potential. The input resistance was in the G omega range and overshooting repetitive action potentials were elicited by injecting depolarizing current pulses in the range of 2-10 pA. Voltage-dependent currents were characterized under voltage-clamp conditions. A transient fast inward current activating near -45 mV was blocked by tetrodotoxin. In isolated cells, it was half-deactivated at a membrane potential near -75 mV. An outward K+ current was blocked by internal Cs+ ions or by external tetraethylammonium or Ba2+ ions. A calcium-activated voltage-dependent potassium current was blocked by external Cd2+ ions. A voltage-dependent Ca2+ current was observed in an iso-osmotic BaCl2 solution. Finally, a hyperpolarization-activated inward current was recorded. Voltage-dependent currents in these microvillar olfactory receptor neurons appear qualitatively similar to those already described in ciliated olfactory receptor cells located in the principal olfactory epithelium.

  10. Localization and pharmacological characterization of voltage dependent calcium channels in cultured neocortical neurons

    DEFF Research Database (Denmark)

    Timmermann, D B; Lund, T M; Belhage, B

    2001-01-01

    using the fluorescent calcium chelator fura-2. The types of calcium channels present at the synaptic terminal were determined by the inhibitory action of calcium channel blockers on potassium-induced [3H]GABA release in the same cell preparation. L-, N-, P-, Q- and R-/T-type voltage dependent calcium...... channels were differentially distributed in somata, neurites and nerve terminals. omega-conotoxin MVIIC (omega-CgTx MVIIC) inhibited approximately 40% of the Ca(2+)-rise in both somata and neurites and 60% of the potassium induced [3H]GABA release, indicating that the Q-type channel is the quantitatively...... in cytosolic calcium concentration. The results of this investigation demonstrate that pharmacologically distinct types of voltage dependent calcium channels are differentially localized in cell bodies, neurites and nerve terminals of mouse cortical neurons but that the Q-type calcium channel appears...

  11. G protein-induced trafficking of voltage-dependent calcium channels.

    Science.gov (United States)

    Tombler, Eugene; Cabanilla, Nory Jun; Carman, Paul; Permaul, Natasha; Hall, John J; Richman, Ryan W; Lee, Jessica; Rodriguez, Jennifer; Felsenfeld, Dan P; Hennigan, Robert F; Diversé-Pierluissi, María A

    2006-01-20

    Calcium channels are well known targets for inhibition by G protein-coupled receptors, and multiple forms of inhibition have been described. Here we report a novel mechanism for G protein-mediated modulation of neuronal voltage-dependent calcium channels that involves the destabilization and subsequent removal of calcium channels from the plasma membrane. Imaging experiments in living sensory neurons show that, within seconds of receptor activation, calcium channels are cleared from the membrane and sequestered in clathrin-coated vesicles. Disruption of the L1-CAM-ankyrin B complex with the calcium channel mimics transmitter-induced trafficking of the channels, reduces calcium influx, and decreases exocytosis. Our results suggest that G protein-induced removal of plasma membrane calcium channels is a consequence of disrupting channel-cytoskeleton interactions and might represent a novel mechanism of presynaptic inhibition.

  12. Lavender Oil-Potent Anxiolytic Properties via Modulating Voltage Dependent Calcium Channels

    OpenAIRE

    2013-01-01

    Recent clinical data support the clinical use of oral lavender oil in patients suffering from subsyndromal anxiety. We identified the molecular mechanism of action that will alter the perception of lavender oil as a nonspecific ingredient of aromatherapy to a potent anxiolytic inhibiting voltage dependent calcium channels (VOCCs) as highly selective drug target. In contrast to previous publications where exorbitant high concentrations were used, the effects of lavender oil in behavioral, bioc...

  13. Analytical Model for Voltage-Dependent Photo and Dark Currents in Bulk Heterojunction Organic Solar Cells

    OpenAIRE

    2016-01-01

    A physics-based explicit mathematical model for the external voltage-dependent forward dark current in bulk heterojunction (BHJ) organic solar cells is developed by considering Shockley-Read-Hall (SRH) recombination and solving the continuity equations for both electrons and holes. An analytical model for the external voltage-dependent photocurrent in BHJ organic solar cells is also proposed by incorporating exponential photon absorption, dissociation efficiency of bound electron-hole pairs (...

  14. Effects of different kinds of stretch on voltage-dependent calcium current in antrial circular smooth muscle cells of the guinea-pig%不同牵张刺激对豚鼠胃窦环行肌细胞电压依赖性钙电流的影响

    Institute of Scientific and Technical Information of China (English)

    许文燮; 李英; 吴龙仁; 李在琉

    2000-01-01

    利用全细胞膜片钳技术,在胃窦环行肌细胞上观察了不同方式的牵张刺激对电压依赖性钙电流的影响,探讨牵张刺激对胃窦平滑肌细胞电压依赖性钙电流的作用.用低渗性溶液灌流细胞引起的牵张刺激首先增加电压依赖性钙电流,接着激活一种内向性钳制电流.钙电流的增加发生在灌流后1 min内,而内向性钳制电流在细胞明显膨胀之后缓慢激活.低渗和正压引起的细胞膨胀明显增加电压依赖性钙离子电流,而利用两个电极直接牵拉细胞则不出现钙电流增加效应.结果提示: 细胞膜牵张增强电压依赖性钙通道的活性,而这一作用可能与牵拉引起的细胞所受的膜张力或/和牵拉的方向有关.%In order to elucidate the effect of membrane stretch on ionic currents, we employed the whole-cell patch-clamp technique to investigate the effects of different kinds of stretch on voltage-dependent calcium currents in antrial circular smooth muscle cells of the guinea-pig. The membrane stretch induced by superfusing the smooth muscle cells with hyposmotic bath solution enhanced voltage-operated calcium current and activated inward holding current. The increase in calcium current occurred within 1 minute of superfusion and the sustained inward holding current was slowly activated after prominent cell swelling. Voltage-dependent calcium currents (Ica) were significantly increased by membrane stretch which was induced by cell swelling and cell inflation, but was not affected by direct longitudinal stretch (110~130%) using two electrodes.The results suggest that the cell membrane stretch can increase voltage-dependent calcium channel activity and the effect of stretch on calcium channels was related to the membrane tension and/or the direction of membrane stretch.

  15. Selective modulation of cellular voltage dependent calcium channels by hyperbaric pressure - a suggested HPNS partial mechanism

    Directory of Open Access Journals (Sweden)

    Ben eAviner

    2014-05-01

    Full Text Available Professional deep sea divers experience motor and cognitive impairment, known as High Pressure Neurological Syndrome (HPNS, when exposed to pressures of 100 msw (1.1MPa and above, considered to be the result of synaptic transmission alteration. Previous studies have indicated modulation of presynaptic Ca2+ currents at high pressure. We directly measured for the first time pressure effects on the currents of voltage dependent Ca2+ channels (VDCCs expressed in Xenopus oocytes. Pressure selectivity augmented the current in CaV1.2 and depressed it in CaV3.2 channels. Pressure application also affected the channels' kinetics, such as ƮRise, ƮDecay. Pressure modulation of VDCCs seems to play an important role in generation of HPNS signs and symptoms.

  16. Lavender oil-potent anxiolytic properties via modulating voltage dependent calcium channels.

    Science.gov (United States)

    Schuwald, Anita M; Nöldner, Michael; Wilmes, Thomas; Klugbauer, Norbert; Leuner, Kristina; Müller, Walter E

    2013-01-01

    Recent clinical data support the clinical use of oral lavender oil in patients suffering from subsyndromal anxiety. We identified the molecular mechanism of action that will alter the perception of lavender oil as a nonspecific ingredient of aromatherapy to a potent anxiolytic inhibiting voltage dependent calcium channels (VOCCs) as highly selective drug target. In contrast to previous publications where exorbitant high concentrations were used, the effects of lavender oil in behavioral, biochemical, and electrophysiological experiments were investigated in physiological concentrations in the nanomolar range, which correlate to a single dosage of 80 mg/d in humans that was used in clinical trials. We show for the first time that lavender oil bears some similarities with the established anxiolytic pregabalin. Lavender oil inhibits VOCCs in synaptosomes, primary hippocampal neurons and stably overexpressing cell lines in the same range such as pregabalin. Interestingly, Silexan does not primarily bind to P/Q type calcium channels such as pregabalin and does not interact with the binding site of pregabalin, the α2δ subunit of VOCCs. Lavender oil reduces non-selectively the calcium influx through several different types of VOCCs such as the N-type, P/Q-type and T-type VOCCs. In the hippocampus, one brain region important for anxiety disorders, we show that inhibition by lavender oil is mainly mediated via N-type and P/Q-type VOCCs. Taken together, we provide a pharmacological and molecular rationale for the clinical use of the oral application of lavender oil in patients suffering from anxiety.

  17. Lavender oil-potent anxiolytic properties via modulating voltage dependent calcium channels.

    Directory of Open Access Journals (Sweden)

    Anita M Schuwald

    Full Text Available Recent clinical data support the clinical use of oral lavender oil in patients suffering from subsyndromal anxiety. We identified the molecular mechanism of action that will alter the perception of lavender oil as a nonspecific ingredient of aromatherapy to a potent anxiolytic inhibiting voltage dependent calcium channels (VOCCs as highly selective drug target. In contrast to previous publications where exorbitant high concentrations were used, the effects of lavender oil in behavioral, biochemical, and electrophysiological experiments were investigated in physiological concentrations in the nanomolar range, which correlate to a single dosage of 80 mg/d in humans that was used in clinical trials. We show for the first time that lavender oil bears some similarities with the established anxiolytic pregabalin. Lavender oil inhibits VOCCs in synaptosomes, primary hippocampal neurons and stably overexpressing cell lines in the same range such as pregabalin. Interestingly, Silexan does not primarily bind to P/Q type calcium channels such as pregabalin and does not interact with the binding site of pregabalin, the α2δ subunit of VOCCs. Lavender oil reduces non-selectively the calcium influx through several different types of VOCCs such as the N-type, P/Q-type and T-type VOCCs. In the hippocampus, one brain region important for anxiety disorders, we show that inhibition by lavender oil is mainly mediated via N-type and P/Q-type VOCCs. Taken together, we provide a pharmacological and molecular rationale for the clinical use of the oral application of lavender oil in patients suffering from anxiety.

  18. Effects of in vitro and in vivo lead exposure on voltage-dependent calcium channels in central neurons of Lymnaea stagnalis.

    Science.gov (United States)

    Audesirk, G

    1987-01-01

    Currents through calcium channels of members of an identified cluster of neurons (B cells) in the pond snail Lymnaea stagnalis were studied under voltage clamp. The normal physiological saline was modified to maximize the visibility of voltage-dependent calcium currents and minimize contamination by other currents. Barium was used as the charge carrier for the calcium channels. Depolarizing voltage steps induce an inward current, the magnitude of which varies with the barium concentration. In brains taken from animals not exposed in vivo to lead, in vitro addition of lead acetate to the recording medium (0.25 to 14 microM) inhibits the barium current by 59 +/- 14% (mean +/- s.d.), in a manner that is independent of the lead concentration. The magnitude of the residual current still varies with the barium concentration. The voltage dependence of the current appears to be unaffected by lead. In contrast to some other calcium-channel blockers, such as cobalt, the inhibition of barium currents by in vitro lead exposure is irreversible, at least in short-term experiments. Contrary to expectations based on these in vitro results, barium currents in B cells of animals exposed to 5 microM lead for 6 to 12 weeks in vivo were approximately twice as large as barium currents in B cells from unexposed controls, when both were recorded in lead-free saline. It is possible that chronic in vivo lead exposure causes an increase in the number of calcium channels in these neurons.

  19. State-dependent FRET reports calcium- and voltage-dependent gating-ring motions in BK channels

    OpenAIRE

    Miranda, Pablo; Contreras, Jorge E.; Plested, Andrew J. R.; Sigworth, Fred J.; Holmgren, Miguel; Giraldez, Teresa

    2013-01-01

    Large-conductance voltage- and calcium-dependent potassium channels (BK, “Big K+”) are important controllers of cell excitability. In the BK channel, a large C-terminal intracellular region containing a “gating-ring” structure has been proposed to transduce Ca2+ binding into channel opening. Using patch-clamp fluorometry, we have investigated the calcium and voltage dependence of conformational changes of the gating-ring region of BK channels, while simultaneously monitoring channel conductan...

  20. Actin Dynamics Regulates Voltage-Dependent Calcium-Permeable Channels of the Vicia faba Guard Cell Plasma Membrane

    Institute of Scientific and Technical Information of China (English)

    Wei Zhang; Liu-Min Fan

    2009-01-01

    Free cytosolic Ca~(2+) ([Ca~(2+)]_(cyt)) is an ubiquitous second messenger in plant cell signaling, and [Ca~(2+)]_(cyt) elevation is associated with Ca~(2+)-permeable channels in the plasma membrane and endomembranes regulated by a wide range of stimuli. However, knowledge regarding Ca~(2+) channels and their regulation remains limited in planta. A type of voltage-dependent Ca~(2+)-permeable channel was identified and characterized for the Vicia faba L. guard cell plasma membrane by using patch-clamp techniques. These channels are permeable to both Ba~(2+) and Ca~(2+), and their activities can be inhibited by micromolar Gd~(3+). The unitary conductance and the reversal potential of the channels depend on the Ca~(2+) or Ba~(2+) gradients across the plasma membrane. The inward whole-cell Ca~(2+) (Ba~(2+)) current, as well as the unitary current amplitude and NP. of the single Ca~(2+) channel, increase along with the membrane hyperpolarization. Pharmacological experiments suggest that actin dynamics may serve as an upstream regulator of this type of calcium channel of the guard cell plasma membrane. Cytochalasin D, an actin polymerization blocker, activated the NP_o of these channels at the single channel level and increased the current amplitude at the whole-cell level. But these channel activations and current increments could be restrained by pretreatment with an F-actin stabilizer, phalloidin. The potential physiological significance of this regulatory mechanism is also discussed.

  1. Analytical Model for Voltage-Dependent Photo and Dark Currents in Bulk Heterojunction Organic Solar Cells

    Directory of Open Access Journals (Sweden)

    Mesbahus Saleheen

    2016-05-01

    Full Text Available A physics-based explicit mathematical model for the external voltage-dependent forward dark current in bulk heterojunction (BHJ organic solar cells is developed by considering Shockley-Read-Hall (SRH recombination and solving the continuity equations for both electrons and holes. An analytical model for the external voltage-dependent photocurrent in BHJ organic solar cells is also proposed by incorporating exponential photon absorption, dissociation efficiency of bound electron-hole pairs (EHPs, carrier trapping, and carrier drift and diffusion in the photon absorption layer. Modified Braun’s model is used to compute the electric field-dependent dissociation efficiency of the bound EHPs. The overall net current is calculated considering the actual solar spectrum. The mathematical models are verified by comparing the model calculations with various published experimental results. We analyze the effects of the contact properties, blend compositions, charge carrier transport properties (carrier mobility and lifetime, and cell design on the current-voltage characteristics. The power conversion efficiency of BHJ organic solar cells mostly depends on electron transport properties of the acceptor layer. The results of this paper indicate that improvement of charge carrier transport (both mobility and lifetime and dissociation of bound EHPs in organic blend are critically important to increase the power conversion efficiency of the BHJ solar cells.

  2. VOLTAGE-DEPENDENT SODIUM AND POTASSIUM, BUT NO CALCIUM CONDUCTANCES IN DDT1 MF-2 SMOOTH-MUSCLE CELLS

    NARCIS (Netherlands)

    MOLLEMAN, A; NELEMANS, A; VANDENAKKER, J; DUIN, M; DENHERTOG, A

    1991-01-01

    Voltage-dependent inward and outward membrane currents were investigated in the DDT1 MF-2 smooth muscle cell line using the whole-cell patch-clamp technique. Application of a pulse protocol with subsequent depolarizing voltage steps elicited an inactivating inward current and a non-inactivating outw

  3. Voltage-dependent K+ currents contribute to heterogeneity of olfactory ensheathing cells

    Science.gov (United States)

    Rela, Lorena; Piantanida, Ana Paula; Bordey, Angelique; Greer, Charles A.

    2015-01-01

    The olfactory nerve is permissive for axon growth throughout life. This has been attributed in part to the olfactory ensheathing glial cells that encompass the olfactory sensory neuron fascicles. Olfactory ensheathing cells also promote axon growth in vitro and when transplanted in vivo to sites of injury. The mechanisms involved remain largely unidentified owing in part to the limited knowledge of the physiological properties of ensheathing cells. Glial cells rely for many functions on the properties of the potassium channels expressed; however, those expressed in ensheathing cells are unknown. Here we show that olfactory ensheathing cells express voltage-dependent potassium currents compatible with inward rectifier (Kir) and delayed rectifier (KDR) channels. Together with gap junction coupling, these contribute to the heterogeneity of membrane properties observed in olfactory ensheathing cells. The relevance of K+ currents expressed by ensheathing cells is discussed in relation to plasticity of the olfactory nerve. PMID:25856239

  4. Differential expression of T- and L-type voltage-dependent calcium channels in renal resistance vessels

    DEFF Research Database (Denmark)

    Hansen, Pernille B. Lærkegaard; Jensen, Boye L.; Andreasen, D;

    2001-01-01

    .2 protein was demonstrated by immunochemical labeling of rat preglomerular vasculature and juxtamedullary efferent arterioles and vasa recta. Cortical efferent arterioles were not immunopositive. Recordings of intracellular calcium concentration with digital fluorescence imaging microscopy showed......The distribution of voltage-dependent calcium channels in kidney pre- and postglomerular resistance vessels was determined at the molecular and functional levels. Reverse transcription-polymerase chain reaction analysis of microdissected rat preglomerular vessels and cultured smooth muscle cells...... showed coexpression of mRNAs for T-type subunits (Ca(V)3.1, Ca(V)3.2) and for an L-type subunit (Ca(V)1.2). The same expression pattern was observed in juxtamedullary efferent arterioles and outer medullary vasa recta. No calcium channel messages were detected in cortical efferent arterioles. Ca(V)1...

  5. Differential expression of T- and L-type voltage-dependent calcium channels in renal resistance vessels

    DEFF Research Database (Denmark)

    Hansen, Pernille B. Lærkegaard; Jensen, Boye L.; Andreasen, D

    2001-01-01

    The distribution of voltage-dependent calcium channels in kidney pre- and postglomerular resistance vessels was determined at the molecular and functional levels. Reverse transcription-polymerase chain reaction analysis of microdissected rat preglomerular vessels and cultured smooth muscle cells...... showed coexpression of mRNAs for T-type subunits (Ca(V)3.1, Ca(V)3.2) and for an L-type subunit (Ca(V)1.2). The same expression pattern was observed in juxtamedullary efferent arterioles and outer medullary vasa recta. No calcium channel messages were detected in cortical efferent arterioles. Ca(V)1.......2 protein was demonstrated by immunochemical labeling of rat preglomerular vasculature and juxtamedullary efferent arterioles and vasa recta. Cortical efferent arterioles were not immunopositive. Recordings of intracellular calcium concentration with digital fluorescence imaging microscopy showed...

  6. Tetrahydroacridine inhibits voltage-dependent Na+ current in guinea-pig ventricular myocytes

    Institute of Scientific and Technical Information of China (English)

    Wei WANG; Yi-ping WANG; Guo-yuan HU

    2004-01-01

    AIM: To study the effects of tetrahydroacridine (tacrine) on voltage-gated Na+ channels in cardiac tissues.METHODS: Single ventricular myocytes were enzymatically dissociated from adult guinea-pig heart. Voltagedependent Na+ current was recorded using whole cell voltage-clamp technique. RESULTS: (1) Tacrine reversibly inhibited Na+ current with an IC50 value of 120 μmol/L (95 % confidence range: 108-133 μmol/L). (2) The inhibitory effects of tacrine on Na+ current exhibited both a tonic nature and use-dependence. (3) Tacrine at 100 μmol/L caused a negative shift (about 10 mV) in the voltage-dependence of steady-state inactivation of Na+ current, and retarded its recovery from inactivation, but did not affect its activation curve. (4) Intracellular application of tacrine significantly inhibited Na+ current. CONCLUSION: In addition to blocking other voltage-gated ion channels,tacrine blocked Na+ channels in guinea-pig ventricular myocytes. Tactine acted as inactivation stabilizer of Na+channels in cardiac tissues.

  7. Voltage-dependent Calcium Channel Plays a Role in the Formation of Large-amplitude Miniature Excitatory Postsynaptic Current%电压依赖性钙通道参与大振幅微小兴奋性突触后电流形成的实验研究

    Institute of Scientific and Technical Information of China (English)

    黄福森; 杨小娟; 王儒蓉; 吴超然

    2012-01-01

    目的 观察电压依赖性钙通道是否作用于大鼠脊髓背角胶状质层(SG)神经元大振幅微小兴奋性突触后电流的形成.方法 选用成年雄性Sprague-Dawley (SD)大鼠,2%~3%异氟烷麻醉后,分离其腰骶部的脊髓,然后切片.采用全细胞电压钳技术,玻璃微电极的电阻为4~6 MΩ,钳制电压为-70 mV,记录胶状质层神经元微小兴奋性突触后电流( mEPSC)电流.将电流信号用Axopatch 200来放大并储存于电脑.对照组和用药结束后,持续采样mEPSC电流30 s.mEPSC电流的频率和振幅用Clampfit 8.1进行分析.结果 钳制电压为-70 mV时,所有SG神经元均有自发性的EPSC.辣椒素增加mEPSC发生的频率和波幅.钴离子抑制辣椒素诱导的大振幅mEPSC.钴离子抑制辣椒素诱导的mEPSC的平均振幅,而不抑制其发生频率.结论 电压依赖性钙离子通道参与了辣椒素引起的痛觉形成.%Objective To observe whether the voltage-dependent calcium channel contributes to the formation of capsaicin-induced miniature excitatory postsynaptic current (mEPSC) in rats. Methods Experiments were performed in adult male Sprague-Dawley rats. The lumbosacral portion of the spinal cord were separated after anesthesia by 2%-3% isoflurane, and the spinal cord slices were prepared. Whole-cell voltage-clamp technique was applied to substantia gelatinosa (SG) neurons with a glass patch-pipette having a resistance of 4-6 MΩ, holding potential -70 mV. Signals were amplified with an Axopatch 200 amplifier and then stored in a personal computer. The mEPSC in controls and immediately after the end of drug applications were sampled for 30 seconds and the frequency and amplitude were analyzed using Clampfit 8.1. Results All SG neurons in this database had spontaneous mEPSC with the holding potential of-70 mV. Capsaicin increased the frequency and mean amplitude of mEPSC. Cobalt inhibited the capsaicin-induced large-amplitude mEPSC, as well as the mean amplitude but

  8. 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.

  9. Cloning, chromosomal localization, and functional expression of the alpha 1 subunit of the L-type voltage-dependent calcium channel from normal human heart

    NARCIS (Netherlands)

    Schultz, D; Mikala, G; Yatani, A; Engle, D B; Iles, D E; Segers, B; Sinke, R J; Weghuis, D O; Klöckner, U; Wakamori, M

    1993-01-01

    A unique structural variant of the cardiac L-type voltage-dependent calcium channel alpha 1 subunit cDNA was isolated from libraries derived from normal human heart mRNA. The deduced amino acid sequence shows significant homology to other calcium channel alpha 1 subunits. However, differences from t

  10. Altered calcium homeostasis in motor neurons following AMPA receptor but not voltage-dependent calcium channels' activation in a genetic model of amyotrophic lateral sclerosis.

    Science.gov (United States)

    Guatteo, Ezia; Carunchio, Irene; Pieri, Massimo; Albo, Federica; Canu, Nadia; Mercuri, Nicola B; Zona, Cristina

    2007-10-01

    Amyotrophic lateral sclerosis (ALS) is a late-onset progressive neurodegenerative disease characterized by a substantial loss of motor neurons in the spinal cord, brain stem and motor cortex. By combining electrophysiological recordings with imaging techniques, clearance/buffering capacity of cultured spinal cord motor neurons after a calcium accumulation has been analyzed in response to AMPA receptors' (AMPARs') activation and to depolarizing stimuli in a genetic mouse model of ALS (G93A). Our studies demonstrate that the amplitude of the calcium signal in response to AMPARs' or voltage-dependent calcium channels' activation is not significantly different in controls and G93A motor neurons. On the contrary, in G93A motor neurons, the [Ca(2+)](i) recovery to basal level is significantly slower compared to control neurons following AMPARs but not voltage-dependent calcium channels' activation. This difference was not observed in G93A cultured cortical neurons. This observation is the first to indicate a specific alteration of the calcium clearance linked to AMPA receptors' activation in G93A motor neurons and the involvement of AMPA receptor regulatory proteins controlling both AMPA receptor functionality and the sequence of events connected to them.

  11. Neuroprotective activity of stiripentol with a possible involvement of voltage-dependent calcium and sodium channels.

    Science.gov (United States)

    Verleye, Marc; Buttigieg, Dorothée; Steinschneider, Rémy

    2016-02-01

    A growing body of data has shown that recurrent epileptic seizures may be caused by an excessive release of the excitatory neurotransmitter glutamate in the brain. Glutamatergic overstimulation results in massive neuronal influxes of calcium and sodium through N-methyl-D-aspartate (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid, and kainic acid glutamate subtype receptors and also through voltage-gated calcium and sodium channels. These persistent and abnormal sodium and calcium entry points have deleterious consequences (neurotoxicity) for neuronal function. The therapeutic value of an antiepileptic drug would include not only control of seizure activity but also protection of neuronal tissue. The present study examines the in vitro neuroprotective effects of stiripentol, an antiepileptic compound with γ-aminobutyric acidergic properties, on neuronal-astroglial cultures from rat cerebral cortex exposed to oxygen-glucose deprivation (OGD) or to glutamate (40 µM for 20 min), two in vitro models of brain injury. In addition, the affinity of stiripentol for the different glutamate receptor subtypes and the interaction with the cell influx of Na(+) and of Ca(2+) enhanced by veratridine and NMDA, respectively, are assessed. Stiripentol (10-100 µM) included in the culture medium during OGD or with glutamate significantly increased the number of surviving neurons relative to controls. Stiripentol displayed no binding affinity for different subtypes of glutamate receptors (IC50  >100 µM) but significantly blocked the entry of Na(+) and Ca(2+) activated by veratridine and NMDA, respectively. These results suggest that Na(+) and Ca(2+) channels could contribute to the neuroprotective properties of sitiripentol.

  12. The action of a phorbol ester on voltage-dependent parameters of the sodium current in isolated hippocampal neurons.

    Science.gov (United States)

    Chizhmakov, I V; Klee, M R

    1994-03-01

    The action of a phorbol ester (phorbol-12,13-diacetate) on the voltage-activated sodium current has been investigated by the voltage-clamp method in acutely isolated pyramidal neurons from rat hippocampus. The intracellular perfusion of isolated pyramidal neurons for 30-40 min induced a gradual 10-15 mV shift in both the current-voltage relationship and voltage-dependent steady-state inactivation to more negative potentials. The application of phorbol ester (1-10 microM) to isolated neurons for the same time increased the amplitude of sodium current by 15-20%, shifted the above-mentioned voltage-dependent parameters for an additional 10-15 mV in the same direction and changed the slope of the steady-state inactivation curve. In contrast, after prolonged incubation of slices in the phorbol ester-containing solution (1-10 microM) for 0.5-3 h, subsequent application of phorbol ester at the same concentration caused neither the addition shift of the voltage-dependent characteristics of sodium channels nor the change of the slope of the steady-state inactivation curve. However, in this case an increase in the amplitude of sodium current by 15-20% during 30-40 min intracellular perfusion was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

  13. Correlation character of ionic current fluctuations: analysis of ion current through a voltage-dependent potassium single channel.

    Science.gov (United States)

    Tong-Han, Lan; Huang, Xi; Jia-Rui, Lin

    2005-10-03

    The gating of ion channels has widely been modeled by assuming the transition between open and closed states is a memoryless process. Nevertheless, the statistical analysis of an ionic current signal recorded from voltage dependence K(+) single channel is presented. Calculating the sample auto-correlation function of the ionic current based on the digitized signals, rather than the sequence of open and closed states duration time. The results provide evidence for the existence of memory. For different voltages, the ion channel current fluctuation has different correlation attributions. The correlations in data generated by simulation of two Markov models, on one hand, auto-correlation function of the ionic current shows a weaker memory, after a delayed period of time, the attribute of memory does not exist; on the other hand, the correlation depends on the number of states in the Markov model. For V(p)=-60 mV pipette potential, spectral analysis of ion channel current was conducted, the result indicates that the spectrum is not a flat spectrum, the data set from ionic current fluctuations shows considerable variability with a broad 1/f -like spectrum, alpha=1.261+/-0.24. Thus the ion current fluctuations give information about the kinetics of the channel protein, the results suggest the correlation character of ion channel protein nonlinear kinetics regardless of whether the channel is in open or closed state.

  14. Development of a voltage-dependent current noise algorithm for conductance-based stochastic modelling of auditory nerve fibres.

    Science.gov (United States)

    Badenhorst, Werner; Hanekom, Tania; Hanekom, Johan J

    2016-12-01

    This study presents the development of an alternative noise current term and novel voltage-dependent current noise algorithm for conductance-based stochastic auditory nerve fibre (ANF) models. ANFs are known to have significant variance in threshold stimulus which affects temporal characteristics such as latency. This variance is primarily caused by the stochastic behaviour or microscopic fluctuations of the node of Ranvier's voltage-dependent sodium channels of which the intensity is a function of membrane voltage. Though easy to implement and low in computational cost, existing current noise models have two deficiencies: it is independent of membrane voltage, and it is unable to inherently determine the noise intensity required to produce in vivo measured discharge probability functions. The proposed algorithm overcomes these deficiencies while maintaining its low computational cost and ease of implementation compared to other conductance and Markovian-based stochastic models. The algorithm is applied to a Hodgkin-Huxley-based compartmental cat ANF model and validated via comparison of the threshold probability and latency distributions to measured cat ANF data. Simulation results show the algorithm's adherence to in vivo stochastic fibre characteristics such as an exponential relationship between the membrane noise and transmembrane voltage, a negative linear relationship between the log of the relative spread of the discharge probability and the log of the fibre diameter and a decrease in latency with an increase in stimulus intensity.

  15. Blockade of the voltage-dependent sodium current in isolated rat hippocampal neurons by tetrodotoxin and lidocaine.

    Science.gov (United States)

    Kaneda, M; Oyama, Y; Ikemoto, Y; Akaike, N

    1989-04-10

    The effects of tetrodotoxin and lidocaine on the voltage-dependent sodium current (INa) were studied in the CA1 pyramidal neurons isolated acutely from rat hippocampus using a 'concentration-clamp' technique which combines the intracellular perfusion with a rapid external solution change within a few ms. Tetrodotoxin (TTX) exerted its inhibitory action in time- and dose-dependent manner on the peak amplitude of INa without any apparent effects on both the current activation and inactivation processes of the current. The time course for reaching a steady-state of the inhibitory action shortened with increasing TTX concentration, but the time course of recovery from the inhibition after washing out the toxin was quite the same at any concentrations used. Lidocaine also inhibited dose-dependently the INa, though with slightly accelerating both the activation and inactivation processes. The time courses for reaching the steady-state inhibition and the recovery from the inhibition were much shorter than those in the case of TTX. The results indicate that the voltage-dependent sodium channel of mammalian brain neuron is TTX-sensitive as well as that of peripheral neuron and that the mode of TTX inhibition on the INa is quite different from that of lidocaine.

  16. A voltage-dependent persistent sodium current in mammalian hippocampal neurons

    OpenAIRE

    1990-01-01

    Currents generated by depolarizing voltage pulses were recorded in neurons from the pyramidal cell layer of the CA1 region of rat or guinea pig hippocampus with single electrode voltage-clamp or tight- seal whole-cell voltage-clamp techniques. In neurons in situ in slices, and in dissociated neurons, subtraction of currents generated by identical depolarizing voltage pulses before and after exposure to tetrodotoxin revealed a small, persistent current after the transient current. These curren...

  17. [Role of calcineurin in down-regulation of left ventricular transmural voltage- dependent K(+) currents in mice with heart failure].

    Science.gov (United States)

    Shi, Chen-Xia; Dong, Fang; Chang, Yan-Chao; Wang, Xiao-Feng; Xu, Yan-Fang

    2015-08-25

    The aim of the present study was to investigate the role of calcineurin in the down-regulation of left ventricular transmural voltage-dependent K(+) currents in heart failure. Transverse aorta was banded by using microsurgical techniques to create mouse heart failure model. Sham-operated (Sham) or aorta banded (Band) mice were randomized to receive calcineurin inhibitor cyclosporine A (CsA) or vehicle. The densities and kinetic properties of voltage-dependent K(+) currents, as well as action potential (AP), of left ventricular subendocardial (Endo) and subepicardial (Epi) myocytes were determined by using whole-cell patch-clamp technique. The results showed that calcineurin activity was significant higher in Endo myocytes than that in Epi ones in all the groups. Compared with Sham group, Band mice showed significantly increased calcineurin activity both in Endo and Epi myocytes. CsA significantly reduced calcineurin activity in Band mice. CsA treatment in Band mice partially reversed the down-regulation of Ito density, completely reversed the down-regulation of IK,slow density both in Endo and Epi myocytes, and Iss density in Endo myocytes. In addition, CsA treatment in Band mice partially antagonized the prolongation of action potential duration (APD), and APD at 50% (APD50) and 90% repolarization (APD90) were significantly reduced. Because of non-parallel shortening of APD in Endo and Epi myocytes, the ratio of Endo/Epi APD90 was reduced from 4.8:1 in Band mice to 2.6:1 in CsA-treated mice, which was close to that in Sham mice. The results suggest that non-parallel activation of calcineurin in Endo and Epi myocytes contributes to the down-regulation of transmural voltage-dependent K(+) currents and the amplification of transmural dispersion of repolarization (TDR) in left ventricular failure hearts. Inhibition of calcineurin may be a potential new therapeutic strategy to prevent and cure arrhythmias and sudden death in heart failure.

  18. A voltage-dependent persistent sodium current in mammalian hippocampal neurons.

    Science.gov (United States)

    French, C R; Sah, P; Buckett, K J; Gage, P W

    1990-06-01

    Currents generated by depolarizing voltage pulses were recorded in neurons from the pyramidal cell layer of the CA1 region of rat or guinea pig hippocampus with single electrode voltage-clamp or tight-seal whole-cell voltage-clamp techniques. In neurons in situ in slices, and in dissociated neurons, subtraction of currents generated by identical depolarizing voltage pulses before and after exposure to tetrodotoxin revealed a small, persistent current after the transient current. These currents could also be recorded directly in dissociated neurons in which other ionic currents were effectively suppressed. It was concluded that the persistent current was carried by sodium ions because it was blocked by TTX, decreased in amplitude when extracellular sodium concentration was reduced, and was not blocked by cadmium. The amplitude of the persistent sodium current varied with clamp potential, being detectable at potentials as negative as -70 mV and reaching a maximum at approximately -40 mV. The maximum amplitude at -40 mV in 21 cells in slices was -0.34 +/- 0.05 nA (mean +/- 1 SEM) and -0.21 +/- 0.05 nA in 10 dissociated neurons. Persistent sodium conductance increased sigmoidally with a potential between -70 and -30 mV and could be fitted with the Boltzmann equation, g = gmax/(1 + exp[(V' - V)/k)]). The average gmax was 7.8 +/- 1.1 nS in the 21 neurons in slices and 4.4 +/- 1.6 nS in the 10 dissociated cells that had lost their processes indicating that the channels responsible are probably most densely aggregated on or close to the soma. The half-maximum conductance occurred close to -50 mV, both in neurons in slices and in dissociated neurons, and the slope factor (k) was 5-9 mV. The persistent sodium current was much more resistant to inactivation by depolarization than the transient current and could be recorded at greater than 50% of its normal amplitude when the transient current was completely inactivated. Because the persistent sodium current activates at

  19. Quaternary Organic Amines Inhibit Na,K Pump Current in a Voltage-dependent Manner

    OpenAIRE

    2004-01-01

    The effects of organic quaternary amines, tetraethylammonium (TEA) chloride and benzyltriethylammonium (BTEA) chloride, on Na,K pump current were examined in rat cardiac myocytes superfused in extracellular Na+-free solutions and whole-cell voltage-clamped with patch electrodes containing a high Na+-salt solution. Extracellular application of these quaternary amines competitively inhibited extracellular K+ (K+ o) activation of Na,K pump current; however, the concentration for half maximal inh...

  20. 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.

  1. Effect of etomidate on voltage-dependent potassium currents in rat isolated hippocampal pyramidal neurons

    Institute of Scientific and Technical Information of China (English)

    TAN Hong-yu; SUN Li-na; WANG Xiao-liang; YE Tie-hu

    2010-01-01

    Background Previous studies demonstrated general anesthetics affect potassium ion channels, which may be one of the mechanisms of general anesthesia. Because the effect of etomidate on potassium channels in rat hippocampus which is involved in memory function has not been studied, we investigated the effects of etomidate on both delayed rectifier potassium current (I_((K(DR))) and transient outward potassium current (I_((K(A))) in acutely dissociated rat hippocampal pyramidal neurons.Methods Single rat hippocampal pyramidal neurons from male Wistar rats of 7-10 days were acutely dissociated by enzymatic digestion and mechanical dispersion according to the methods of Kay and Wong with slight modification. Voltage-clamp recordings were performed in the whole-cell patch clamp configuration. Currents were recorded with a List EPC-10 amplifier and data were stored in a computer using Pulse 8.5. Student's paired two-tail t test was used for data analysis. Results At the concentration of 100 μmol/L, etomidate significantly inhibited I_(K(DR)) by 49.2% at +40 mV when depolarized from -110 mV (P 0.05). The IC_(50) value of etomidate for blocking I_(K(DR)) was calculated as 5.4 μmol/L, with a Hill slope of 2.45. At the presence of 10 μmol/L etomidate, the V_(1/2) of activation curve was shifted from (17.3±1.5) mV to (10.7±9.9) mV (n=8, P <0.05), the V_(1/2) of inactivation curve was shifted from (-18.3±2.2) mV to (-45.3±9.4) mV (n=8, P <0.05). Etomidate 10 μmol/L shifted both the activation curve and inactivation curve of I_(K(DR)) to negative potential, but mainly affected the inactivation kinetics.Conclusions Etomidate potently inhibited I_(K(DR)) but not I_(K(A)) in rat hippocampal pyramidal neurons. I_(K(DR)) was inhibited by etomidate in a concentration-dependent manner, while I_(K(A)) remained unaffected.

  2. Decreases of voltage-dependent K+ currents densities in ventricular myocytes of guinea pigs by chronic oxidant stress

    Institute of Scientific and Technical Information of China (English)

    De-li DONG; Yan LIU; Yu-hong ZHOU; Wei-hua SONG; He WANG; Bao-feng YANG

    2004-01-01

    AIM: To determine the changes of delayed rectifier K+ currents (Ik) and inward rectifier K+ currents (Ik1) in the ventricular myocytes of guinea pigs during the gradual apoptotic process by the chronic oxidant stress treatment.METHODS: H2O250 μmol/L (24 h) was used for inducing apoptosis in the cardiomyocytes culture of neonatal rats and to treat the isolated ventricular myocytes of adult guinea pigs in vitro for 24 h. Apoptosis was evaluated by TUNEL methods and voltage-dependent K+ currents were recorded by patch-clamp techniques. RESULTS: H2O250 μmol/L (24 h) induced cell apoptosis in the cardiomyocytes culture of neonatal rats. This concentration was used to treat the isolated ventricular myocytes of adult guinea pigs in vitro for 24 h and the voltage-dependent K+currents densities (Ik, Ik1) were down-regulated. The densities of the delayed rectifier K+ currents (Ik) in 50 μmol/L H2O2 group were 2.52±0.57 pA/pF vs 5.73±1.84 pA/pF in the control group at +50 mV (n=8, P<0.01). The densities of the inward rectifier K+ currents (Ik1) in 50 μmol/L H2O2 group were -13.9±2.70 pA/pF, 2.52±0.57 pA/pF vs -59.7± 11.9 pA/pF, 5.73± 1.84 pA/pF in the control group at -120 mV (n=8, P<0.01) and -40 mV (n=8, P<0.05), respectively. The extent of inward rectifier property of Ik1 was weakened by 50μmol/L H2O2 treatment. CONCLUSION: The densities of Ik, Ik1 in the cardiomyocytes of guinea pigs were downregulated and the inward rectifier property of Ik1 was weakened during the gradual apoptotic process after 50 μmol/L H2O2 treatment for 24 h.

  3. Voltage-dependent potassium currents during fast spikes of rat cerebellar Purkinje neurons: inhibition by BDS-I toxin.

    Science.gov (United States)

    Martina, Marco; Metz, Alexia E; Bean, Bruce P

    2007-01-01

    We characterized the kinetics and pharmacological properties of voltage-activated potassium currents in rat cerebellar Purkinje neurons using recordings from nucleated patches, which allowed high resolution of activation and deactivation kinetics. Activation was exceptionally rapid, with 10-90% activation in about 400 mus at +30 mV, near the peak of the spike. Deactivation was also extremely rapid, with a decay time constant of about 300 mus near -80 mV. These rapid activation and deactivation kinetics are consistent with mediation by Kv3-family channels but are even faster than reported for Kv3-family channels in other neurons. The peptide toxin BDS-I had very little blocking effect on potassium currents elicited by 100-ms depolarizing steps, but the potassium current evoked by action potential waveforms was inhibited nearly completely. The mechanism of inhibition by BDS-I involves slowing of activation rather than total channel block, consistent with the effects described in cloned Kv3-family channels and this explains the dramatically different effects on currents evoked by short spikes versus voltage steps. As predicted from this mechanism, the effects of toxin on spike width were relatively modest (broadening by roughly 25%). These results show that BDS-I-sensitive channels with ultrafast activation and deactivation kinetics carry virtually all of the voltage-dependent potassium current underlying repolarization during normal Purkinje cell spikes.

  4. Scorpion toxin prolongs an inactivation phase of the voltage-dependent sodium current in rat isolated single hippocampal neurons.

    Science.gov (United States)

    Kaneda, M; Oyama, Y; Ikemoto, Y; Akaike, N

    1989-05-15

    The effects of scorpion toxin on the voltage-dependent sodium current (INa) of CA1 pyramidal neurons isolated from rat hippocampus were studied under the single-electrode voltage-clamp condition using a 'concentration-clamp' technique. The toxin increased the peak amplitude of INa and prolonged its inactivation phase in a time- and dose-dependent manner. Inactivation phase of INa proceeded with two exponential components in the absence (control) and presence of the toxin. In the toxin-treated neurons, both the time constant of slow component and its fractional contribution to the total current increased dose-dependently while the fractional contribution of the fast one decreased in a dose-dependent fashion without changing its time constant. Actions of scorpion toxin on the sodium channels of hippocampal pyramidal neurons were essentially similar to those of peripheral preparations. Therefore, it can be concluded that the sodium channels of mammalian brain neurons have structures and functions similar to peripheral channels.

  5. Inhibitory effect of resveratrol on the proliferation of GH3 pituitary-adenoma cells and voltage-dependent potassium current

    Institute of Scientific and Technical Information of China (English)

    Ming Chu; Lanlan Wei; Chao Wang; Yu Cheng; Kongbin Yang; Baofeng Yang

    2006-01-01

    BACKGROUND:Recent researches indicate that activation of potassium channel is likely to cause many kinds of cells to proliferate and differentiate;using chemical to block the potassium channel can restrain the proliferation of small lung-cancer cells.breast cancer.prostate cancer and human lymphocyte,etc.Previous researches proved that resveratrol(RE),a selective estrogen receptor modulator(SERM).could inhibit growth of GH3 calls,induce apoptosis,and resist tumor through interfering K+ channel.OBJECTIVE:To investigate the effects of RE on Voltage-dependent K+ current [Ik(v)] and cell proliferation in GH3 pituitary-tumor cells.DESIGN:Observational contrast study.SETTING:Department of Neurosurgery.the First Clinical Hospital of Harbin Medical University;Department of Microbiology,Harbin Medical University;Department of Pharmacology,Harbin Medical University.MATERIALS: GH3 pituitary-tumor cell line of rats was purchased from the American Type Culture Collection (ATCC).RE and[3-(4,5-dimethylthiazo1-2-y1)-2.5-diphenyl-tetrazolium bromide](MTT)were obtained from Sigma Chemical CO,St Louis,USA;Ham's F-10 medium from Gibco BRL;Equine serum and fetal bovine serum from Hyclone Laboratories,Logan,UT;FACSCalibur flow cytometer from BD Company,USA.RE was dissolved in ethanol and stored at-20 ℃.It was diluted to different concentrations (10.50,100 μmol/L)with medium and extra cellular solution when needed.rhe final concentration of ethanol was Jess than 0.01%.METHODS:The experiment was carried out in the Department of Microbiology and Pharmacology of Harbin Medical University from March 2005 to January 2006.①Cell preparation:Proliferating indexes affected by 10.50 and 100 μmol/L RE were measured with MTT,respectively.0.0001 volume fraction of ethan ol was added into control group.Inhibitory rate of cellular growth was calculated as the following formula:Inhibitory rate (%)=(1-A value in experimental group/A value in control group)x100%.The experiments mentioned above were

  6. Inflammatory cytokine signaling in insulin producing beta-cells enhances the colocalization correlation coefficient between L-type voltage-dependent calcium channel and calcium-sensing receptor.

    Science.gov (United States)

    Parkash, Jai

    2008-08-01

    The immunological processes in type 1 diabetes and metabolic/inflammatory disorder in type 2 diabetes converge on common signaling pathway(s) leading to beta-cell death in these two diseases. The cytokine-mediated beta-cell death seems to be dependent on voltage-dependent calcium channel (VDCC)-mediated Ca2+ entry. The Ca2+ handling molecular networks control the homeostasis of [Ca2+]i in the beta-cell. The activity and membrane density of VDCC are regulated by several mechanisms including G protein-coupled receptors (GPCRs). CaR is a 123-kDa seven transmembrane extracellular Ca2+ sensing protein that belongs to GPCR family C. Tumor necrosis factor-alpha (TNF-alpha), is a cytokine widely known to activate nuclear factor-kappaB (NF-kappaB) transcription in beta-cells. To obtain a better understanding of TNF-alpha-induced molecular interactions between CaR and VDCC, confocal fluorescence measurements were performed on insulin-producing beta-cells exposed to varying concentrations of TNF-alpha and the results are discussed in the light of increased colocalization correlation coefficient. The insulin producing beta-cells were exposed to 5, 10, 20, 30, and 50 ng/ml TNF-alpha for 24 h at 37 degrees . The cells were then immunolabelled with antibodies directed against CaR, VDCC, and NF-kappaB. The confocal fluorescence imaging data showed enhancement in the colocalization correlation coefficient between CaR and VDCC in beta-cells exposed to TNF-alpha thereby indicating increased membrane delimited spatial interactions between these two membrane proteins. TNF-alpha-induced colocalization of VDCC with CaR was inhibited by nimodipine, an inhibitor of L-type VDCC thereby suggesting that VDCC activity is required for spatial interactions with CaR. The 3-D confocal fluorescence imaging data also demonstrated that addition of TNF-alpha to RIN cells led to the translocation of NF-kappaB from the cytoplasm to the nucleus. Such molecular interactions between CaR and VDCC in tissues

  7. Dopamine Induces LTP Differentially in Apical and Basal Dendrites through BDNF and Voltage-Dependent Calcium Channels

    Science.gov (United States)

    Navakkode, Sheeja; Sajikumar, Sreedharan; Korte, Martin; Soong, Tuck Wah

    2012-01-01

    The dopaminergic modulation of long-term potentiation (LTP) has been studied well, but the mechanism by which dopamine induces LTP (DA-LTP) in CA1 pyramidal neurons is unknown. Here, we report that DA-LTP in basal dendrites is dependent while in apical dendrites it is independent of activation of L-type voltage-gated calcium channels (VDCC).…

  8. Dopamine Induces LTP Differentially in Apical and Basal Dendrites through BDNF and Voltage-Dependent Calcium Channels

    Science.gov (United States)

    Navakkode, Sheeja; Sajikumar, Sreedharan; Korte, Martin; Soong, Tuck Wah

    2012-01-01

    The dopaminergic modulation of long-term potentiation (LTP) has been studied well, but the mechanism by which dopamine induces LTP (DA-LTP) in CA1 pyramidal neurons is unknown. Here, we report that DA-LTP in basal dendrites is dependent while in apical dendrites it is independent of activation of L-type voltage-gated calcium channels (VDCC).…

  9. Coexpression of voltage-dependent calcium channels Cav1.2, 2.1a, and 2.1b in vascular myocytes

    DEFF Research Database (Denmark)

    Andreasen, Ditte; Friis, Ulla Glenert; Uhrenholt, Torben Rene

    2006-01-01

    , and blocking P-type currents (omega-agatoxin IVA 10 nmol/L) led to 20.2+/-3.0% inhibition, whereas 300 nmol/L of omega agatoxin IVA (blocking P/Q-type) inhibited 45.0+/-7.3%. In rat aortic smooth muscle cells (A7r5), blockade of L-type channels resulted in 28.5+/-6.1% inhibition, simultaneous blockade of L...... microscopy revealed expression of both channels in all of the smooth muscle cells. Whole-cell patch clamp on single preglomerular VSMCs from mice showed L-, P-, and Q-type currents. Blockade of the L-type currents by calciseptine (20 nmol/L) inhibited 35.6+/-3.9% of the voltage-dependent Ca2+ current......-type and P-type channels inhibited 58.0+/-11.8%, and simultaneous inhibition of L-, P-, and Q-type channels led to blockade (88.7+/-5.6%) of the Ca2+ current. We conclude that aortic and renal preglomerular smooth muscle cells express L-, P-, and Q-type voltage-dependent Ca2+ channels in the rat and mouse....

  10. "Slow" Voltage-Dependent Inactivation of CaV2.2 Calcium Channels Is Modulated by the PKC Activator Phorbol 12-Myristate 13-Acetate (PMA.

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    Lei Zhu

    Full Text Available CaV2.2 (N-type voltage-gated calcium channels (Ca2+ channels play key roles in neurons and neuroendocrine cells including the control of cellular excitability, neurotransmitter / hormone secretion, and gene expression. Calcium entry is precisely controlled by channel gating properties including multiple forms of inactivation. "Fast" voltage-dependent inactivation is relatively well-characterized and occurs over the tens-to- hundreds of milliseconds timeframe. Superimposed on this is the molecularly distinct, but poorly understood process of "slow" voltage-dependent inactivation, which develops / recovers over seconds-to-minutes. Protein kinases can modulate "slow" inactivation of sodium channels, but little is known about if/how second messengers control "slow" inactivation of Ca2+ channels. We investigated this using recombinant CaV2.2 channels expressed in HEK293 cells and native CaV2 channels endogenously expressed in adrenal chromaffin cells. The PKC activator phorbol 12-myristate 13-acetate (PMA dramatically prolonged recovery from "slow" inactivation, but an inactive control (4α-PMA had no effect. This effect of PMA was prevented by calphostin C, which targets the C1-domain on PKC, but only partially reduced by inhibitors that target the catalytic domain of PKC. The subtype of the channel β-subunit altered the kinetics of inactivation but not the magnitude of slowing produced by PMA. Intracellular GDP-β-S reduced the effect of PMA suggesting a role for G proteins in modulating "slow" inactivation. We postulate that the kinetics of recovery from "slow" inactivation could provide a molecular memory of recent cellular activity and help control CaV2 channel availability, electrical excitability, and neurotransmission in the seconds-to-minutes timeframe.

  11. Poly(ethylene glycol-cholesterol inhibits L-type Ca2+ channel currents and augments voltage-dependent inactivation in A7r5 cells.

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    Rikuo Ochi

    Full Text Available Cholesterol distributes at a high density in the membrane lipid raft and modulates ion channel currents. Poly(ethylene glycol cholesteryl ether (PEG-cholesterol is a nonionic amphipathic lipid consisting of lipophilic cholesterol and covalently bound hydrophilic PEG. PEG-cholesterol is used to formulate lipoplexes to transfect cultured cells, and liposomes for encapsulated drug delivery. PEG-cholesterol is dissolved in the external leaflet of the lipid bilayer, and expands it to flatten the caveolae and widen the gap between the two leaflets. We studied the effect of PEG-cholesterol on whole cell L-type Ca(2+ channel currents (I(Ca,L recorded from cultured A7r5 arterial smooth muscle cells. The pretreatment of cells with PEG-cholesterol decreased the density of ICa,L and augmented the voltage-dependent inactivation with acceleration of time course of inactivation and negative shift of steady-state inactivation curve. Methyl-β-cyclodextrin (MβCD is a cholesterol-binding oligosaccharide. The enrichment of cholesterol by the MβCD:cholesterol complex (cholesterol (MβCD caused inhibition of I(Ca,L but did not augment voltage-dependent inactivation. Incubation with MβCD increased I(Ca,L, slowed the time course of inactivation and shifted the inactivation curve to a positive direction. Additional pretreatment by a high concentration of MβCD of the cells initially pretreated with PEG-cholesterol, increased I(Ca,L to a greater level than the control, and removed the augmented voltage-dependent inactivation. Due to the enhancement of the voltage-dependent inactivation, PEG-cholesterol inhibited window I(Ca,L more strongly as compared with cholesterol (MβCD. Poly(ethylene glycol conferred to cholesterol the efficacy to induce sustained augmentation of voltage-dependent inactivation of I(Ca,L.

  12. Differential rescue of spatial memory deficits in aged rats by L-type voltage-dependent calcium channel and ryanodine receptor antagonism.

    Science.gov (United States)

    Hopp, S C; D'Angelo, H M; Royer, S E; Kaercher, R M; Adzovic, L; Wenk, G L

    2014-11-01

    Age-associated memory impairments may result as a consequence of neuroinflammatory induction of intracellular calcium (Ca(+2)) dysregulation. Altered L-type voltage-dependent calcium channel (L-VDCC) and ryanodine receptor (RyR) activity may underlie age-associated learning and memory impairments. Various neuroinflammatory markers are associated with increased activity of both L-VDCCs and RyRs, and increased neuroinflammation is associated with normal aging. In vitro, pharmacological blockade of L-VDCCs and RyRs has been shown to be anti-inflammatory. Here, we examined whether pharmacological blockade of L-VDCCs or RyRs with the drugs nimodipine and dantrolene, respectively, could improve spatial memory and reduce age-associated increases in microglia activation. Dantrolene and nimodipine differentially attenuated age-associated spatial memory deficits but were not anti-inflammatory in vivo. Furthermore, RyR gene expression was inversely correlated with spatial memory, highlighting the central role of Ca(+2) dysregulation in age-associated memory deficits.

  13. Gαi2- and Gαi3-specific regulation of voltage-dependent L-type calcium channels in cardiomyocytes.

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    Sara Dizayee

    Full Text Available BACKGROUND: Two pertussis toxin sensitive G(i proteins, G(i2 and G(i3, are expressed in cardiomyocytes and upregulated in heart failure. It has been proposed that the highly homologous G(i isoforms are functionally distinct. To test for isoform-specific functions of G(i proteins, we examined their role in the regulation of cardiac L-type voltage-dependent calcium channels (L-VDCC. METHODS: Ventricular tissues and isolated myocytes were obtained from mice with targeted deletion of either Gα(i2 (Gα(i2 (-/- or Gα(i3 (Gα(i3 (-/-. mRNA levels of Gα(i/o isoforms and L-VDCC subunits were quantified by real-time PCR. Gα(i and Ca(vα(1 protein levels as well as protein kinase B/Akt and extracellular signal-regulated kinases 1/2 (ERK1/2 phosphorylation levels were assessed by immunoblot analysis. L-VDCC function was assessed by whole-cell and single-channel current recordings. RESULTS: In cardiac tissue from Gα(i2 (-/- mice, Gα(i3 mRNA and protein expression was upregulated to 187 ± 21% and 567 ± 59%, respectively. In Gα(i3 (-/- mouse hearts, Gα(i2 mRNA (127 ± 5% and protein (131 ± 10% levels were slightly enhanced. Interestingly, L-VDCC current density in cardiomyocytes from Gα(i2 (-/- mice was lowered (-7.9 ± 0.6 pA/pF, n = 11, p<0.05 compared to wild-type cells (-10.7 ± 0.5 pA/pF, n = 22, whereas it was increased in myocytes from Gα(i3 (-/- mice (-14.3 ± 0.8 pA/pF, n = 14, p<0.05. Steady-state inactivation was shifted to negative potentials, and recovery kinetics slowed in the absence of Gα(i2 (but not of Gα(i3 and following treatment with pertussis toxin in Gα(i3 (-/-. The pore forming Ca(vα(1 protein level was unchanged in all mouse models analyzed, similar to mRNA levels of Ca(vα(1 and Ca(vβ(2 subunits. Interestingly, at the cellular signalling level, phosphorylation assays revealed abolished carbachol-triggered activation of ERK1/2 in mice lacking Gα(i2. CONCLUSION: Our data provide novel evidence for an isoform

  14. Both barium and calcium activate neuronal potassium currents.

    OpenAIRE

    Ribera, A B; Spitzer, N C

    1987-01-01

    Amphibian spinal neurons in culture possess both rapidly inactivating and sustained calcium-dependent potassium current components, similar to those described for other cells. Divalent cation-dependent whole-cell outward currents were isolated by subtracting the voltage-dependent potassium currents recorded from Xenopus laevis neurons in the presence of impermeant cadmium (100-500 microM) from the currents produced without cadmium but in the presence of permeant divalent cations (50-100 micro...

  15. Transient voltage-dependent potassium currents are reduced in NTS neurons isolated from renal wrap hypertensive rats.

    Science.gov (United States)

    Belugin, Sergei; Mifflin, Steve

    2005-12-01

    Whole cell patch-clamp measurements were made in neurons enzymatically dispersed from the nucleus of the solitary tract (NTS) to determine if alterations occur in voltage-dependent potassium channels from rats made hypertensive (HT) by unilateral nephrectomy/renal wrap for 4 wk. Some rats had the fluorescent tracer DiA applied to the aortic nerve before the experiment to identify NTS neurons receiving monosynaptic baroreceptor afferent inputs. Mean arterial pressure (MAP) was greater in 4-wk HT (165 +/- 5 mmHg, n = 26, P NTS neurons from NT and HT rats. At activation voltages from -10 to +10 mV, TOCs were significantly less in HT neurons compared with those observed in NT neurons (P NTS neurons from NT and HT rats and was not different comparing neurons from NT and HT rats. However, examination of the subset of NTS neurons exhibiting somatic DiA fluorescence revealed that DiA-labeled neurons from HT rats had a significantly shorter duration delayed excitation (n = 8 cells, P = 0.022) than DiA-labeled neurons from NT rats (n = 7 cells). Neurons with delayed excitation from HT rats had a significantly broader first action potential (AP) and a slower maximal downstroke velocity of repolarization compared with NT neurons with delayed excitation (P = 0.016 and P = 0.014, respectively). The number of APs in the first 200 ms of a sustained depolarization was greater in HT than NT neurons (P = 0.012). These results suggest that HT of 4-wk duration reduces TOCs in NTS neurons, and this contributes to reduced delayed excitation and increased AP responses to depolarizing inputs. Such changes could alter baroreflex function in hypertension.

  16. The agonist-specific voltage dependence of M2 muscarinic receptors modulates the deactivation of the acetylcholine-gated K(+) current (I KACh).

    Science.gov (United States)

    Moreno-Galindo, Eloy G; Alamilla, Javier; Sanchez-Chapula, José A; Tristani-Firouzi, Martin; Navarro-Polanco, Ricardo A

    2016-07-01

    Recently, it has been shown that G protein-coupled receptors (GPCRs) display intrinsic voltage sensitivity. We reported that the voltage sensitivity of M2 muscarinic receptor (M2R) is also ligand specific. Here, we provide additional evidence to understand the mechanism underlying the ligand-specific voltage sensitivity of the M2R. Using ACh, pilocarpine (Pilo), and bethanechol (Beth), we evaluated the agonist-specific effects of voltage by measuring the ACh-activated K(+) current (I KACh) in feline and rabbit atrial myocytes and in HEK-293 cells expressing M2R-Kir3.1/Kir3.4. The activation of I KACh by the muscarinic agonist Beth was voltage insensitive, suggesting that the voltage-induced conformational changes in M2R do not modify its affinity for this agonist. Moreover, deactivation of the Beth-evoked I KACh was voltage insensitive. By contrast, deactivation of the ACh-induced I KACh was significantly slower at -100 mV than at +50 mV, while an opposite effect was observed when I KACh was activated by Pilo. These findings are consistent with the voltage affinity pattern observed for these three agonists. Our findings suggest that independent of how voltage disturbs the receptor binding site, the voltage dependence of the signaling pathway is ultimately determined by the agonist. These observations emphasize the pharmacological potential to regulate the M2R-parasympathetic associated cardiac function and also other cellular signaling pathways by exploiting the voltage-dependent properties of GPCRs.

  17. Voltage dependence of cardiac excitation-contraction coupling: unitary Ca2+ current amplitude and open channel probability.

    Science.gov (United States)

    Altamirano, Julio; Bers, Donald M

    2007-09-14

    Excitation-contraction coupling in cardiac myocytes occurs by Ca2+-induced Ca2+ release, where L-type Ca2+ current evokes a larger sarcoplasmic reticulum (SR) Ca2+ release. The Ca2+-induced Ca2+ release amplification factor or gain (SR Ca2+ release/I(Ca)) is usually assessed by the V(m) dependence of current and Ca2+ transients. Gain rises at negative V(m), as does single channel I(Ca) (i(Ca)), which has led to the suggestion that the increases of i(Ca) amplitude enhances gain at more negative V(m). However, I(Ca) = NP(o) x i(Ca) (where NP(o) is the number of open channels), and NP(o) and i(Ca) both depend on V(m). To assess how i(Ca) and NP(o) separately influence Ca2+-induced Ca2+ release, we measured I(Ca) and junctional SR Ca2+ release in voltage-clamped rat ventricular myocytes using "Ca2+ spikes" (confocal microscopy). To vary i(Ca) alone, we changed [Ca2+](o) rapidly at constant test V(m) (0 mV) or abruptly repolarized from +120 mV to different V(m) (at constant [Ca2+](o)). To vary NP(o) alone, we altered Ca2+ channel availability by varying holding V(m) (at constant test V(m)). Reducing either i(Ca) or NP(o) alone increased excitation-contraction coupling gain. Thus, increasing i(Ca) does not increase gain at progressively negative test V(m). Such enhanced gain depends on lower NP(o) and reduced redundant Ca2+ channel openings (per junction) and a consequently smaller denominator in the gain equation. Furthermore, modest i(Ca) (at V(m) = 0 mV) may still effectively trigger SR Ca2+ release, whereas at positive V(m) (and smaller i(Ca)), high and well-synchronized channel openings are required for efficient excitation-contraction coupling. At very positive V(m), reduced i(Ca) must explain reduced SR Ca2+ release.

  18. Both barium and calcium activate neuronal potassium currents

    Energy Technology Data Exchange (ETDEWEB)

    Ribera, A.B.; Spitzer, N.C.

    1987-09-01

    Amphibian spinal neurons in culture possess both rapidly inactivating and sustained calcium-dependent potassium current components, similar to those described for other cells. Divalent cation-dependent whole-cell outward currents were isolated by subtracting the voltage-dependent potassium currents recorded from Xenopus laevis neurons in the presence of impermeant cadmium from the currents produced without cadmium but in the presence of permeant divalent cations. These concentrations of permeant ions were low enough to avoid contamination by macroscopic inward currents through calcium channels. Calcium-dependent potassium currents were reduced by 1 ..mu..M tetraethylammonium. These currents can also be activated by barium or strontium. Barium as well as calcium activated outward currents in young neurons (6-8 hr) and in relatively mature neurons (19-26 hr in vitro). However, barium influx appeared to suppress the sustained voltage-dependent potassium current in most cells. Barium also activated at least one class of potassium channels observed in excised membrane patches, whole blocking others. The blocking action may have masked and hindered detection of the stimulatory action of barium in other systems.

  19. A CACNA1C variant associated with reduced voltage-dependent inactivation, increased CaV1.2 channel window current, and arrhythmogenesis.

    Directory of Open Access Journals (Sweden)

    Jessica A Hennessey

    Full Text Available Mutations in CACNA1C that increase current through the CaV1.2 L-type Ca2+ channel underlie rare forms of long QT syndrome (LQTS, and Timothy syndrome (TS. We identified a variant in CACNA1C in a male child of Filipino descent with arrhythmias and extracardiac features by candidate gene sequencing and performed functional expression studies to electrophysiologically characterize the effects of the variant on CaV1.2 channels. As a baby, the subject developed seizures and displayed developmental delays at 30 months of age. At age 5 years, he displayed a QTc of 520 ms and experienced recurrent VT. Physical exam at 17 years of age was notable for microcephaly, short stature, lower extremity weakness and atrophy with hyperreflexia, spastic diplegia, multiple dental caries and episodes of rhabdomyolysis. Candidate gene sequencing identified a G>C transversion at position 5731 of CACNA1C (rs374528680 predicting a glycine>arginine substitution at residue 1911 (p.G1911R of CaV1.2. The allele frequency of this variant is 0.01 in Malays, but absent in 984 Caucasian alleles and in the 1000 genomes project. In electrophysiological analyses, the variant decreased voltage-dependent inactivation, thus causing a gain of function of CaV1.2. We also observed a negative shift of V1/2 of activation and positive shift of V1/2 of channel inactivation, resulting in an increase of the window current. Together, these suggest a gain-of-function effect on CaV1.2 and suggest increased susceptibility for arrhythmias in certain clinical settings. The p.G1911R variant was also identified in a case of sudden unexplained infant death (SUID, for which an increasing number of clinical observations have demonstrated can be associated with arrhythmogenic mutations in cardiac ion channels. In summary, the combined effects of the CACNA1C variant to diminish voltage-dependent inactivation of CaV1.2 and increase window current expand our appreciation of mechanisms by which a gain of

  20. [Role of voltage-dependent ion channels in epileptogenesis].

    Science.gov (United States)

    Ricard-Mousnier, B; Couraud, F

    1993-10-01

    The aim of this review is to gather information in favour of the involvement of voltage-dependent ion channels in epileptogenesis. Although, up to now, no study has shown that epilepsy is accompanied by a modification in the activity to these channels, the recently acquired knowledge of their physiology allows to presume would favor their involvement in epileptogenesis. The results from electrophysiological studies are as follows: a persistent sodium current increases neuronal excitability whereas potassium currents have an inhibitory role. In particular, calcium-dependent potassium current are involved in the post-hyperpolarization phases which follows PDS. Calcium currents are also involved in the genesis of the "bursting pacemaker" activity displayed by the neurons presumed to be inducers of the epileptic activity. Biochemical data has shown that as a consequence of epileptic activity, sodium and calcium channels are down regulated. This down-regulation could be a way to reduces neuronal hyperexcitability. Pharmacological data demonstrate the drugs which activate calcium channels or which inhibit potassium channels have a convusilvant effect. On the contrary, agents which block calcium or sodium channels or which properties. Among the latter ones, some antiepileptic drugs can be found. In summary situations which lead to increase in calcium and sodium currents and/or to an inhibition in potassium currents are potentially epileptogenic.

  1. Inhibition of calcium currents in cultured rat dorsal root ganglion neurones by (-)-baclofen.

    OpenAIRE

    Dolphin, A C; Scott, R.H.

    1986-01-01

    Voltage-dependent inward calcium currents (ICa) activated in cultured rat dorsal root ganglion neurones were reversibly reduced in a dose-dependent manner by (-)-baclofen (10 microM to 100 microM). Baclofen (100 microM) reduced the calcium-dependent slow outward potassium current (IK(Ca)). This current was abolished in calcium-free medium and by 300 microM cadmium chloride. The action of baclofen on IK(Ca) was reduced when the calcium concentration in the medium was increased from 5 mM to 30 ...

  2. Voltage Dependence of Supercapacitor Capacitance

    Directory of Open Access Journals (Sweden)

    Szewczyk Arkadiusz

    2016-09-01

    Full Text Available Electronic Double-Layer Capacitors (EDLC, called Supercapacitors (SC, are electronic devices that are capable to store a relatively high amount of energy in a small volume comparing to other types of capacitors. They are composed of an activated carbon layer and electrolyte solution. The charge is stored on electrodes, forming the Helmholtz layer, and in electrolyte. The capacitance of supercapacitor is voltage- dependent. We propose an experimental method, based on monitoring of charging and discharging a supercapacitor, which enables to evaluate the charge in an SC structure as well as the Capacitance-Voltage (C-V dependence. The measurement setup, method and experimental results of charging/discharging commercially available supercapacitors in various voltage and current conditions are presented. The total charge stored in an SC structure is proportional to the square of voltage at SC electrodes while the charge on electrodes increases linearly with the voltage on SC electrodes. The Helmholtz capacitance increases linearly with the voltage bias while a sublinear increase of total capacitance was found. The voltage on SC increases after the discharge of electrodes due to diffusion of charges from the electrolyte to the electrodes. We have found that the recovery voltage value is linearly proportional to the initial bias voltage value.

  3. Potentiation of prolactin secretion following lactotrope escape from dopamine action. II. Phosphorylation of the alpha(1) subunit of L-type, voltage-dependent calcium channels.

    Science.gov (United States)

    Hernández, M E; del Mar Hernández, M; Díaz-Muñoz, M; Clapp, C; de la Escalera, G M

    1999-07-01

    Modulation of Ca(2+) channels has been shown to alter cellular functions. It can play an important role in the amplification of signals in the endocrine system, including the pleiotropically regulated pituitary lactotropes. Prolactin (PRL) secretion is tonically inhibited by dopamine (DA), the escape from which triggers acute episodes of hormone secretion. The magnitude of these episodes is explained by a potentiation of the PRL-releasing action of secretagogues such as thyrotropin-releasing hormone (TRH). While the mechanisms of this potentiation are not fully understood, it is known to be mimicked by the dihydropyridine, L-type Ca(2+) channel agonist Bay K 8644 and blocked by nifedipine and methoxyverapamil. The potentiation is also blocked by inhibitors of cyclic AMP-dependent protein kinase and protein kinase C. We recently described that the escape from tonic actions of DA results in increased macroscopic Ca(2+) currents in GH(4)C(1) lactotropic clonal cells transfected with a cDNA encoding the long form of the human D(2)-DA receptor. Here we show that the withdrawal from DA potentiates the PRL-releasing action of TRH in GH(4)C(1)/D(2)-DAR cells to the same extent as in enriched lactotropes in primary culture. In both experimental models a low density of dihydropyridine receptors was shown by (+)-[(3)H]PN200-110 binding. Photoaffinity labelling with the dihydropyridine [(3)H]azidopine revealed a protein consistent with the alpha(1) subunit of L-type Ca(2+) channels of 165-170 kDa. In both experimental models, the facilitation of TRH action triggered by the escape from DA was correlated with an enhanced rate of phosphorylation of this putative alpha(1) subunit, the nature of which was further supported by immunoprecipitation with selective antibodies directed against the alpha(1C) and alpha(1D) subunit of voltage-gated calcium channels. We propose that PKA- and PKC-dependent phosphorylation of the alpha(1) subunit of high voltage activated, L-type Ca(2

  4. Genotypic to expression profiling of bovine calcium channel, voltage-dependent, alpha-2/delta subunit 1 gene, and their association with bovine mastitis among Frieswal (HFX Sahiwal) crossbred cattle of Indian origin.

    Science.gov (United States)

    Deb, Rajib; Singh, Umesh; Kumar, Sushil; Kumar, Arun; Singh, Rani; Sengar, Gyanendra; Mann, Sandeep; Sharma, Arjava

    2014-04-01

    Calcium channel, voltage-dependent, alpha-2/delta subunit 1 (CACNA2D1) gene is considered to be an important noncytokine candidate gene influencing mastitis. Scanty of reports are available until today regarding the role play of CACNA2D1 gene on the susceptibility of bovine mastitis. We interrogated the CACNA2D1 G519663A [A>G] SNP by PCR-RFLP among two hundreds Frieswal (HF X Sahiwal) crossbred cattle of Indian origin. Genotypic frequency of AA (51.5, n=101) was comparatively higher than AG (35, n=70) and GG (14.5, n=29). Association of Somatic cell score (SCS) with genotypes revealed that, GG genotypes showing lesser count (less susceptible to mastitis) compare to AA and AG. Relative expression of CACNA2D1 transcript (in milk samples) was significantly higher among GG than AG and AA. Further we have also isolated blood sample from the all groups and PBMCs were cultured from each blood sample as per the standard protocol. They were treated with Calcium channel blocker and the expression level of the CACNA2D1 gene was evaluated by Real Time PCR. Results show that expression level decline in each genotypic group after treatment and expression level of GG are again significantly higher than AA and AG. Thus, it may be concluded that GG genotypic animals are favorable for selecting disease resistant breeds.

  5. Calcium-dependent potassium current in barnacle photoreceptor.

    Science.gov (United States)

    Bolsover, S R

    1981-12-01

    When barnacle lateral eye photoreceptors are depolarized to membrane potentials of 0 to +50 mV in the dark, the plot of outward current through the cell membrane against time has two distinct maxima. The first maximum occurs 5-10 ms after the depolarization began. The current then decays to a minimum at approximately 500 ms after the onset of depolarization, and then increases to a second maximum 4-6 s after the depolarization began. If depolarization is maintained, the current again decays to reach a steady value approximately 1 min after depolarization began. The increase in current to the maximum at 4-6s from the minimum at approximately 500 ms is termed the "late current." It is maximum for depolarizations to around +25 mV and is reduced in amplitude at more positive potentials. It is not observed when the membrane is depolarized to potentials more positive than +60 mV. The late current is inhibited by external cobaltous ion and external tetraethylammonium ion, and shows a requirement for external calcium ion. When the calcium-sequestering agent EGTA is injected, the late current is abolished. Illumination of a cell under voltage clamp reduces the amplitude of the late current recorded subsequently in the dark. On the basis of the voltage dependence and pharmacology of the late current, it is proposed that the current is a calcium-dependent potassium current.

  6. Evidence for a distinct light-induced calcium-dependent potassium current in Hermissenda crassicornis.

    Science.gov (United States)

    Blackwell, K T

    2000-01-01

    A model of phototransduction is developed as a first step toward a model for investigating the critical interaction of light and turbulence stimuli within the type B photoreceptor of Hermissenda crassicronis. The model includes equations describing phototransduction, release of calcium from intracellular stores, and other calcium regulatory mechanisms, as well as equations describing ligand-gating of a rhabdomeric sodium current. The model is used to determine the sources of calcium in the soma, whether calcium or IP3 is a plausible ligand of the light-induced sodium current, and whether the light-induced potassium current is equivalent to the calcium-dependent potassium current activated by light-induced calcium release. Simulations show that the early light-induced calcium elevation is due to influx through voltage-dependent channels, whereas the later calcium elevation is due to release from intracellular stores. Simulations suggest that the ligand of the fast, light-induced sodium current is IP3 but that there is a smaller, prolonged component of the light-induced sodium current that is activated by calcium. In the model, the calcium-dependent potassium current, located in the soma, is activated only slightly by light-induced calcium elevation, leading to the prediction that a calcium-dependent potassium current, active at resting potential, is located in the rhabdomere and is responsible for the light-induced potassium current.

  7. The Voltage-Dependent Anion Channel 1 (AtVDAC1 Negatively Regulates Plant Cold Responses during Germination and Seedling Development in Arabidopsis and Interacts with Calcium Sensor CBL1

    Directory of Open Access Journals (Sweden)

    Zhi-Yong Li

    2013-01-01

    Full Text Available The voltage-dependent anion channel (VDAC, a highly conserved major mitochondrial outer membrane protein, plays crucial roles in energy metabolism and metabolite transport. However, knowledge about the roles of the VDAC family in plants is limited. In this study, we investigated the expression pattern of VDAC1 in Arabidopsis and found that cold stress promoted the accumulation of VDAC1 transcripts in imbibed seeds and mature plants. Overexpression of VDAC1 reduced tolerance to cold stress in Arabidopsis. Phenotype analysis of VDAC1 T-DNA insertion mutant plants indicated that a vdac1 mutant line had faster germination kinetics under cold treatment and showed enhanced tolerance to freezing. The yeast two-hybrid system revealed that VDAC1 interacts with CBL1, a calcium sensor in plants. Like the vdac1, a cbl1 mutant also exhibited a higher seed germination rate. We conclude that both VDAC1 and CBL1 regulate cold stress responses during seed germination and plant development.

  8. The sea anemone Bunodosoma caissarum toxin BcIII modulates the sodium current kinetics of rat dorsal root ganglia neurons and is displaced in a voltage-dependent manner.

    Science.gov (United States)

    Salceda, Emilio; López, Omar; Zaharenko, André J; Garateix, Anoland; Soto, Enrique

    2010-03-01

    Sea anemone toxins bind to site 3 of the sodium channels, which is partially formed by the extracellular linker connecting S3 and S4 segments of domain IV, slowing down the inactivation process. In this work we have characterized the actions of BcIII, a sea anemone polypeptide toxin isolated from Bunodosoma caissarum, on neuronal sodium currents using the patch clamp technique. Neurons of the dorsal root ganglia of Wistar rats (P5-9) in primary culture were used for this study (n=65). The main effects of BcIII were a concentration-dependent increase in the sodium current inactivation time course (IC(50)=2.8 microM) as well as an increase in the current peak amplitude. BcIII did not modify the voltage at which 50% of the channels are activated or inactivated, nor the reversal potential of sodium current. BcIII shows a voltage-dependent action. A progressive acceleration of sodium current fast inactivation with longer conditioning pulses was observed, which was steeper as more depolarizing were the prepulses. The same was observed for other two anemone toxins (CgNa, from Condylactis gigantea and ATX-II, from Anemonia viridis). These results suggest that the binding affinity of sea anemone toxins may be reduced in a voltage-dependent manner, as has been described for alpha-scorpion toxins.

  9. Inactivation kinetics and pharmacology distinguish two calcium currents in mouse pancreatic B-cells

    Energy Technology Data Exchange (ETDEWEB)

    Hopkins, W.F.; Satin, L.S.; Cook, D.L. (Univ. of Washington School of Medicine, Seattle (USA))

    1991-02-01

    Voltage-dependent calcium currents were studied in cultured adult mouse pancreatic B-cells using the whole-cell voltage-clamp technique. When calcium currents were elicited with 10-sec depolarizing command pulses, the time course of inactivation was well fit by the sum of two exponentials. The more rapidly-inactivating component had a time constant of 75 +/- 5 msec at 0 mV and displayed both calcium influx- and voltage-dependent inactivation, while the more slowly-inactivating component had a time constant of 2750 +/- 280 msec at 0 mV and inactivated primarily via voltage. The fast component was subject to greater steady-state inactivation at holding potentials between -100 and -40 mV and activated at a lower voltage threshold. This component was also significantly reduced by nimodipine (0.5 microM) when a holding potential of -100 mV was used, whereas the slow component was unaffected. In contrast, the slow component was greatly increased by replacing external calcium with barium, while the fast component was unchanged. Cadmium (1-10 microM) displayed a voltage-dependent block of calcium currents consistent with a greater effect on the high-threshold, more-slowly inactivating component. Taken together, the data suggest that cultured mouse B-cells, as with other insulin-secreting cells we have studied, possess at least two distinct calcium currents. The physiological significance of two calcium currents having distinct kinetic and steady-state inactivation characteristics for B-cell burst firing and insulin secretion is discussed.

  10. Glucocorticoids specifically enhance L-type calcium current amplitude and affect calcium channel subunit expression in the mouse hippocampus.

    Science.gov (United States)

    Chameau, Pascal; Qin, Yongjun; Spijker, Sabine; Smit, August Benjamin; Smit, Guus; Joëls, Marian

    2007-01-01

    Previous studies have shown that corticosterone enhances whole cell calcium currents in CA1 pyramidal neurons, through a pathway involving binding of glucocorticoid receptor homodimers to the DNA. We examined whether glucocorticoids show selectivity for L- over N-type of calcium currents. Moreover, we addressed the putative gene targets that eventually lead to the enhanced calcium currents. Electrophysiological recordings were performed in nucleated patches that allow excellent voltage control. Calcium currents in these patches almost exclusively involve N- and L-type channels. We found that L- but not N-type calcium currents were largely enhanced after treatment with a high dose of corticosterone sufficient to activate glucocorticoid receptors. Voltage dependency and kinetic properties of the currents were unaffected by the hormone. Nonstationary noise analysis suggests that the increased current is not caused by a larger unitary conductance, but rather to a doubling of the number of functional channels. Quantitative real-time PCR revealed that transcripts of the Ca(v)1 subunits encoding for the N- or L-type calcium channels are not upregulated in the mouse CA1 area; instead, a strong, direct, and consistent upregulation of the beta4 subunit was observed. This indicates that the corticosteroid-induced increase in number of L-type calcium channels is not caused by a simple transcriptional regulation of the pore-forming subunit of the channels.

  11. Voltage dependence of the Na-K pump.

    Science.gov (United States)

    De Weer, P; Gadsby, D C; Rakowski, R F

    1988-01-01

    Present evidence demonstrates that the Na-K pump rate is voltage dependent, whereas early work was largely inconclusive. The I-V relationship has a positive slope over a wide voltage range, and the existence of a negative slope region is now doubtful. Monotonic voltage dependence is consistent with the reaction cycle containing a single voltage-dependent step. Recent measurements suggest that this voltage-dependent step occurs during Na translocation and may be deocclusion of Na+. In addition, two results suggest that K translocation is voltage insensitive: (a) large positive potentials appear to have no influence on Rb-Rb exchange or associated conformational transitions; and (b) transient currents associated with Na translocation appear to involve movement of a single charge, which is sufficient for a 3Na-2K cycle. The simplest interpretation is that the pump's cation binding sites supply two negative charges. Pre-steady-state measurements demonstrate that Na translocation precedes the pump cycle's rate-limiting step, presumably K translocation. But, because K translocation seems voltage insensitive, the voltage dependence of the steady-state pump rate probably reflects that of the concentration of the intermediate entering this slow step. Further pump current and flux data (both transient and steady-state), carefully determined over a range of conditions, should increase our understanding of the voltage-dependent step(s) in the Na-K pump cycle.

  12. Endothelin induces two types of contractions of rat uterus: phasic contractions by way of voltage-dependent calcium channels and developing contractions through a second type of calcium channels

    Energy Technology Data Exchange (ETDEWEB)

    Kozuka, M.; Ito, T.; Hirose, S.; Takahashi, K.; Hagiwara, H.

    1989-02-28

    Effects of endothelin on nonvascular smooth muscle have been examined using rat uterine horns and two modes of endothelin action have been revealed. Endothelin (0.3 nM) caused rhythmic contractions of isolated uterus in the presence of extracellular calcium. The rhythmic contractions were completely inhibited by calcium channel antagonists. These characteristics of endothelin-induced contractions were very similar to those induced by oxytocin. Binding assays using /sup 125/I-endothelin showed that endothelin and the calcium channel blockers did not compete for the binding sites. However, endothelin was unique in that it caused, in addition to rhythmic contractions, a slowly developing monophasic contraction that was insensitive to calcium channel blockers. This developing contraction became dominant at higher concentrations of endothelin and was also calcium dependent.

  13. Effects of arsenic trioxide on voltage-dependent potassium channels and on cell proliferation of human multiple myeloma cells

    Institute of Scientific and Technical Information of China (English)

    ZHOU Jin; WANG Wei; WEI Qing-fang; FENG Tie-ming; TAN Li-jun; YANG Bao-feng

    2007-01-01

    @@ Arsenic trioxide (ATO) can induce cellular apoptosis and inhibit the activities of multiple myeloma (MM)cells in vitro,1 but how it works is not very clear. Recent studies showed that ATO worked on the voltagedependent potassium channel and L-type calcium channel in myocardial cells,2-5 but the effect of ATO on ion channels of tumor cells was rarely reported. As the potassium channel plays an important role in controlling cell proliferation,6 we studied the effects of ATO on the voltage-dependent potassium current (Ikv) of the voltage-dependent potassium channel in an MM cell line,and probed into the relationship between changes of the Ikv caused by ATO and cell proliferation.

  14. Calcium ion currents mediating oocyte maturation events

    Directory of Open Access Journals (Sweden)

    Tosti Elisabetta

    2006-05-01

    Full Text Available Abstract During maturation, the last phase of oogenesis, the oocyte undergoes several changes which prepare it to be ovulated and fertilized. Immature oocytes are arrested in the first meiotic process prophase, that is morphologically identified by a germinal vesicle. The removal of the first meiotic block marks the initiation of maturation. Although a large number of molecules are involved in complex sequences of events, there is evidence that a calcium increase plays a pivotal role in meiosis re-initiation. It is well established that, during this process, calcium is released from the intracellular stores, whereas less is known on the role of external calcium entering the cell through the plasma membrane ion channels. This review is focused on the functional role of calcium currents during oocyte maturation in all the species, from invertebrates to mammals. The emerging role of specific L-type calcium channels will be discussed.

  15. Bone morphogenetic protein 4 inhibits insulin secretion from rodent beta cells through regulation of calbindin1 expression and reduced voltage-dependent calcium currents

    DEFF Research Database (Denmark)

    Christensen, Gitte L.; Jacobsen, Maria L. B.; Wendt, Anna

    2015-01-01

    cells reduced GSIS, and the effect of BMP4 on GSIS was lost in islets from calbindin1 (Calb1) knockout mice. CONCLUSIONS/INTERPRETATION: We found BMP4 treatment to markedly inhibit GSIS from rodent pancreatic islets in a calbindin1-dependent manner. Calbindin1 is suggested to mediate the effect of BMP4...

  16. Variations in onset of action potential broadening: effects on calcium current studied in chick ciliary ganglion neurones.

    Science.gov (United States)

    Pattillo, J M; Artim, D E; Simples, J E; Meriney, S D

    1999-02-01

    1. The voltage dependence and kinetic properties of stage 40 ciliary ganglion calcium currents were determined using short (10 ms) voltage steps. These properties aided the interpretation of the action potential-evoked calcium current described below, and the comparison of our data with those observed in other preparations. 2. Three different natural action potential waveforms were modelled by a series of ramps to generate voltage clamp commands. Calcium currents evoked by these model action potentials were compared before and after alterations in the repolarization phase of each action potential. 3. Abrupt step repolarizations from various time points were used to estimate the time course of calcium current activation during each action potential. Calcium current evoked by fast action potentials (duration at half-amplitude, 0.5 or 1.0 ms) did not reach maximal activation until the action potential had repolarized by 40-50 %. In contrast, calcium current evoked by a slow action potential (duration at half-amplitude, 2.2 ms) was maximally activated near the peak of the action potential. 4. Slowing the rate of repolarization of the action potential (broadening) from different times was used to examine effects on peak and total calcium influx. With all three waveforms tested, broadening consistently increased total calcium influx (integral). However, peak calcium current was either increased or decreased depending on the duration of the control action potential tested and the specific timing of the initiation of broadening the repolarization phase. 5. The opposite effects on peak calcium current observed with action potential broadening beginning at different time points in repolarization may provide a mechanism for the variable effects of potassium channel blockers on transmitter release magnitude.

  17. Gated currents in isolated olfactory receptor neurons of the larval tiger salamander.

    OpenAIRE

    Firestein, S; Werblin, F S

    1987-01-01

    The electrical properties of enzymatically isolated olfactory receptor cells were studied with whole-cell patch clamp. Voltage-dependent currents could be separated into three ionic components: a transient inward sodium current, a sustained inward calcium current, and an outward potassium current. Three components of the outward current could be identified by their gating and kinetics: a calcium-dependent potassium current [IK(Ca)], a voltage-dependent potassium current [IK(V)], and a transie...

  18. Voltage-dependent gating of hERG potassium channels

    Directory of Open Access Journals (Sweden)

    Yen May eCheng

    2012-05-01

    Full Text Available The mechanisms by which voltage-gated channels sense changes in membrane voltage and energetically couple this with opening of the ion conducting pore has been the source of significant interest. In voltage-gated potassium (Kv channels, much of our knowledge in this area comes from Shaker-type channels, for which voltage-dependent gating is quite rapid. In these channels, activation and deactivation are associated with rapid reconfiguration of the voltage-sensing domain unit that is electromechanically coupled, via the S4-S5 linker helix, to the rate-limiting opening of an intracellular pore gate. However, fast voltage-dependent gating kinetics are not typical of all Kv channels, such as Kv11.1 (human ether-a-go-go related gene, hERG, which activates and deactivates very slowly. Compared to Shaker channels, our understanding of the mechanisms underlying slow hERG gating is much poorer. Here, we present a comparative review of the structure-function relationships underlying voltage-dependent gating in Shaker and hERG channels, with a focus on the roles of the voltage sensing domain and the S4-S5 linker that couples voltage sensor movements to the pore. Measurements of gating current kinetics and fluorimetric analysis of voltage sensor movement are consistent with models suggesting that the hERG activation pathway contains a voltage independent step, which limits voltage sensor transitions. Constraints upon hERG voltage sensor movement may result from loose packing of the S4 helices and additional intra-voltage sensor counter charge interactions. More recent data suggest that key amino acid differences in the hERG voltage sensing unit and S4-S5 linker, relative to fast activating Shaker-type Kv channels, may also contribute to the increased stability of the resting state of the voltage sensor.

  19. Manipulating the voltage dependence of tunneling spin torques

    KAUST Repository

    Manchon, Aurelien

    2012-10-01

    Voltage-driven spin transfer torques in magnetic tunnel junctions provide an outstanding tool to design advanced spin-based devices for memory and reprogrammable logic applications. The non-linear voltage dependence of the torque has a direct impact on current-driven magnetization dynamics and on devices performances. After a brief overview of the progress made to date in the theoretical description of the spin torque in tunnel junctions, I present different ways to alter and control the bias dependence of both components of the spin torque. Engineering the junction (barrier and electrodes) structural asymmetries or controlling the spin accumulation profile in the free layer offer promising tools to design effcient spin devices.

  20. Vector spin modeling for magnetic tunnel junctions with voltage dependent effects

    Energy Technology Data Exchange (ETDEWEB)

    Manipatruni, Sasikanth, E-mail: sasikanth.manipatruni@intel.com; Nikonov, Dmitri E.; Young, Ian A. [Exploratory Integrated Circuits, Components Research, Intel Corp., Hillsboro, Oregon 97124 (United States)

    2014-05-07

    Integration and co-design of CMOS and spin transfer devices requires accurate vector spin conduction modeling of magnetic tunnel junction (MTJ) devices. A physically realistic model of the MTJ should comprehend the spin torque dynamics of nanomagnet interacting with an injected vector spin current and the voltage dependent spin torque. Vector spin modeling allows for calculation of 3 component spin currents and potentials along with the charge currents/potentials in non-collinear magnetic systems. Here, we show 4-component vector spin conduction modeling of magnetic tunnel junction devices coupled with spin transfer torque in the nanomagnet. Nanomagnet dynamics, voltage dependent spin transport, and thermal noise are comprehended in a self-consistent fashion. We show comparison of the model with experimental magnetoresistance (MR) of MTJs and voltage degradation of MR with voltage. Proposed model enables MTJ circuit design that comprehends voltage dependent spin torque effects, switching error rates, spin degradation, and back hopping effects.

  1. Voltage-dependent regulation of CaV2.2 channels by Gq-coupled receptor is facilitated by membrane-localized β subunit.

    Science.gov (United States)

    Keum, Dongil; Baek, Christina; Kim, Dong-Il; Kweon, Hae-Jin; Suh, Byung-Chang

    2014-10-01

    G protein-coupled receptors (GPCRs) signal through molecular messengers, such as Gβγ, Ca(2+), and phosphatidylinositol 4,5-bisphosphate (PIP2), to modulate N-type voltage-gated Ca(2+) (CaV2.2) channels, playing a crucial role in regulating synaptic transmission. However, the cellular pathways through which GqPCRs inhibit CaV2.2 channel current are not completely understood. Here, we report that the location of CaV β subunits is key to determining the voltage dependence of CaV2.2 channel modulation by GqPCRs. Application of the muscarinic agonist oxotremorine-M to tsA-201 cells expressing M1 receptors, together with CaV N-type α1B, α2δ1, and membrane-localized β2a subunits, shifted the current-voltage relationship for CaV2.2 activation 5 mV to the right and slowed current activation. Muscarinic suppression of CaV2.2 activity was relieved by strong depolarizing prepulses. Moreover, when the C terminus of β-adrenergic receptor kinase (which binds Gβγ) was coexpressed with N-type channels, inhibition of CaV2.2 current after M1 receptor activation was markedly reduced and delayed, whereas the delay between PIP2 hydrolysis and inhibition of CaV2.2 current was decreased. When the Gβγ-insensitive CaV2.2 α1C-1B chimera was expressed, voltage-dependent inhibition of calcium current was virtually abolished, suggesting that M1 receptors act through Gβγ to inhibit CaV2.2 channels bearing membrane-localized CaV β2a subunits. Expression of cytosolic β subunits such as β2b and β3, as well as the palmitoylation-negative mutant β2a(C3,4S), reduced the voltage dependence of M1 muscarinic inhibition of CaV2.2 channels, whereas it increased inhibition mediated by PIP2 depletion. Together, our results indicate that, with membrane-localized CaV β subunits, CaV2.2 channels are subject to Gβγ-mediated voltage-dependent inhibition, whereas cytosol-localized β subunits confer more effective PIP2-mediated voltage-independent regulation. Thus, the voltage dependence of

  2. Estimation of presynaptic calcium currents and endogenous calcium buffers at the frog neuromuscular junction with two different calcium fluorescent dyes

    Directory of Open Access Journals (Sweden)

    Dmitry eSamigullin

    2015-01-01

    Full Text Available At the frog neuromuscular junction, under physiological conditions, the direct measurement of calcium currents and of the concentration of intracellular calcium buffers—which determine the kinetics of calcium concentration and neurotransmitter release from the nerve terminal—has hitherto been technically impossible. With the aim of quantifying both Ca2+ currents and the intracellular calcium buffers, we measured fluorescence signals from nerve terminals loaded with the low-affinity calcium dye Magnesium Green or the high-affinity dye Oregon Green BAPTA-1, simultaneously with microelectrode recordings of nerve-action potentials and end-plate currents. The action-potential-induced fluorescence signals in the nerve terminals developed much more slowly than the postsynaptic response. To clarify the reasons for this observation and to define a spatiotemporal profile of intracellular calcium and of the concentration of mobile and fixed calcium buffers, mathematical modeling was employed. The best approximations of the experimental calcium transients for both calcium dyes were obtained when the calcium current had an amplitude of 1.6 ± 0.08 рА and a half-decay time of 1.2 ± 0.06 ms, and when the concentrations of mobile and fixed calcium buffers were 250 ± 13 µM and 8 ± 0.4 mM, respectively. High concentrations of endogenous buffers define the time course of calcium transients after an action potential in the axoplasm, and may modify synaptic plasticity.

  3. Coronary artery calcium score: current status

    Science.gov (United States)

    Neves, Priscilla Ornellas; Andrade, Joalbo; Monção, Henry

    2017-01-01

    The coronary artery calcium score plays an Important role In cardiovascular risk stratification, showing a significant association with the medium- or long-term occurrence of major cardiovascular events. Here, we discuss the following: protocols for the acquisition and quantification of the coronary artery calcium score by multidetector computed tomography; the role of the coronary artery calcium score in coronary risk stratification and its comparison with other clinical scores; its indications, interpretation, and prognosis in asymptomatic patients; and its use in patients who are symptomatic or have diabetes. PMID:28670030

  4. Proper Voltage-Dependent Ion Channel Function in Dysferlin-Deficient Cardiomyocytes.

    Science.gov (United States)

    Rubi, Lena; Gawali, Vaibhavkumar S; Kubista, Helmut; Todt, Hannes; Hilber, Karlheinz; Koenig, Xaver

    2015-01-01

    Dysferlin plays a decisive role in calcium-dependent membrane repair in myocytes. Mutations in the encoding DYSF gene cause a number of myopathies, e.g. limb-girdle muscular dystrophy type 2B (LGMD2B). Besides skeletal muscle degenerative processes, dysferlin deficiency is also associated with cardiac complications. Thus, both LGMD2B patients and dysferlin-deficient mice develop a dilated cardiomyopathy. We and others have recently reported that dystrophin-deficient ventricular cardiomyocytes from mouse models of Duchenne muscular dystrophy show significant abnormalities in voltage-dependent ion channels, which may contribute to the pathophysiology in dystrophic cardiomyopathy. The aim of the present study was to investigate if dysferlin, like dystrophin, is a regulator of cardiac ion channels. By using the whole cell patch-clamp technique, we compared the properties of voltage-dependent calcium and sodium channels, as well as action potentials in ventricular cardiomyocytes isolated from the hearts of normal and dysferlin-deficient (dysf) mice. In contrast to dystrophin deficiency, the lack of dysferlin did not impair the ion channel properties and left action potential parameters unaltered. In connection with normal ECGs in dysf mice these results suggest that dysferlin deficiency does not perturb cardiac electrophysiology. Our study demonstrates that dysferlin does not regulate cardiac voltage-dependent ion channels, and implies that abnormalities in cardiac ion channels are not a universal characteristic of all muscular dystrophy types. © 2015 S. Karger AG, Basel.

  5. Proper Voltage-Dependent Ion Channel Function in Dysferlin-Deficient Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Lena Rubi

    2015-06-01

    Full Text Available Background/Aims: Dysferlin plays a decisive role in calcium-dependent membrane repair in myocytes. Mutations in the encoding DYSF gene cause a number of myopathies, e.g. limb-girdle muscular dystrophy type 2B (LGMD2B. Besides skeletal muscle degenerative processes, dysferlin deficiency is also associated with cardiac complications. Thus, both LGMD2B patients and dysferlin-deficient mice develop a dilated cardiomyopathy. We and others have recently reported that dystrophin-deficient ventricular cardiomyocytes from mouse models of Duchenne muscular dystrophy show significant abnormalities in voltage-dependent ion channels, which may contribute to the pathophysiology in dystrophic cardiomyopathy. The aim of the present study was to investigate if dysferlin, like dystrophin, is a regulator of cardiac ion channels. Methods and Results: By using the whole cell patch-clamp technique, we compared the properties of voltage-dependent calcium and sodium channels, as well as action potentials in ventricular cardiomyocytes isolated from the hearts of normal and dysferlin-deficient (dysf mice. In contrast to dystrophin deficiency, the lack of dysferlin did not impair the ion channel properties and left action potential parameters unaltered. In connection with normal ECGs in dysf mice these results suggest that dysferlin deficiency does not perturb cardiac electrophysiology. Conclusion: Our study demonstrates that dysferlin does not regulate cardiac voltage-dependent ion channels, and implies that abnormalities in cardiac ion channels are not a universal characteristic of all muscular dystrophy types.

  6. Effects of Angiotensin Ⅱ and ACE Inhibitor, Captopril on L-type Calcium Current and Sodium Current of Single Guinea Myocytes

    Institute of Scientific and Technical Information of China (English)

    徐延敏; 黄体钢; 陈元禄

    2002-01-01

    Objectives To investigateeffect of AngⅡ, captopril on single guinea myocytes onL - type calcium current and sodium current. MethodsMembrane patch clamp whole cell recording tech-nique was used to investigate effect of angⅡ, captoprilon L- Ca maximum current density and sodium maxi-mum current density. Resutls AngⅡ increased themaximum current density compared with control afterpeffused 5 min, 357.7±219.7 Vs 279.5±240.5PA/PF, increase rate is 27.9 %, the shape of current- voltage relationship curve was unchanged, peaked at+ 10 my, indicated that angⅡ increased L- Ca cur-rent density in voltage -dependent. After perfusedwith captopril, captopril ± angⅡ 3, 5 rmin, L-Cacurrent was recorded, results suggest L - Ca maximumcurrent density decreased significantly compared withcontrol, in captopril group, 128.4 ± 92.6Vs286.2 ±89.7, 66.7 ±68.3Vs 286.2 ± 89.7, respectively, rateof inhibition is 55.1%, 76.6 %, respectively. L - Cacurrent further decreased in captopril perfused 5 mincompared with 3 rmin, 66.7 ± 68.3 Vs 128.4 ± 92.6,in captopril + angⅡ group, L- Ca current decreasedgreatly in 3, 5 min than control, 143.4 ± 117.6Vs267.7±141.4, 96.4±82.5 Vs 267.7+141.4, re-spectively, rate of inhibition is 46.4 %, 63.9 % re-spectively. We also investigated effect of captopril onNa current, which decreased significantly in 1 rmin and3 rmin compared with control, 939.1 ±319. 1 Vs1398.0 ± 144.6 PA/PF, 469.95±314.9 Vs 1398.0±144.6 PA/PF, respectively, rate of inhibition is32.8 %, 66. 3 %, respectively. Na current density de-creased significantly in 3 min compared with 1 min,469.9 ± 314.9 Vs 939. 1 ± 319. 1PA/PF, rate of in-hibition is 49.9 % . Conclusions Angiotensin Ⅱexerts increased maximum current density of L - Ca involtage dependent, captopril decreased maximum cur-rent density of L - Ca in voltage dependent, decreasedsodium maximum current density, which is the promi-nently antiarrhythmia mechanisms through inhibition ofangiotensin Ⅱ evoked

  7. 增龄引起犬心房L型电压依赖型钙通道离子重构的分子机制%Aging-related ionic remodeling of L-type voltage dependent calcium channel in left atria of canine

    Institute of Scientific and Technical Information of China (English)

    周贤惠; 张健; 甘天翊; 许国军; 汤宝鹏

    2012-01-01

    目的 探讨增龄导致犬心房L型电压依赖型钙通道离子重构的分子机制.方法 采用全细胞膜片钳技术记录犬左心房肌细胞L型电压依赖型钙通道动作电位时限(APD90)、动作电位平台期电压和L型钙离子电流(ICa-L)特性.应用实时定量逆转录聚合酶链反应(RT-PCR)法测定犬左心耳L型电压依赖型钙通道α1亚单位(CaV1.2)、钙离子释放通道兰尼碱受体(RYR2)、肌浆网钙调控-Ca2+ ATP酶基因(SERCA2)、钙激活蛋白酶-Ⅰ(Calpain-Ⅰ)、磷酸受钠蛋白(PLN1)等的mRNA表达,用Western blot检测蛋白表达.结果 老年犬与成年犬比较,心房肌细胞L型电压依赖型钙通道APD90较长[(340.5±10.1)ms 比(320.0±7.9) ms,P<0.05];动作电位平台期电压较低[(-9.5±1.7)mV比(-6.4±1.1)mV,P<0.05];ICa-L电流密度较低[(-14.04±0.82)pA/pF比(-8.11±0.54)pA/pF,P<0.05].老年犬与成年犬比较,CaV1.2基因表达明显下调(0.90±0.35比2.38±0.40,P<0.05),RYR2基因表达明显上调(4.39±4.68比1.49±1.69,P<0.05),两组犬SERCA2、Calpain-Ⅰ、PLN1基因表达差异无统计学意义;CaV1.2蛋白表达明显下调(0.13±0.10比0.29±0.12,P<0.05),RYR2蛋白明显上调(0.18±0.21比0.08±0.36,P<0.05),两组犬SERCA2、Calpain-Ⅰ、PLN1蛋白表达无明显改变.结论 增龄导致犬心房肌细胞钙通道CaV1.2和RYR2基因和蛋白表达的改变是L型电压依赖型钙通道离子重构的分子机制,可能是老年相关性心房颤动的潜在机制之一.%Objective To investigate aging-related ionic remodeling of L-type voltage dependent calcium channel (LVDCC) in left atria of canine.Methods Seven adult (2.0-2.5 years ) and 10 aged (>8 years) dogs were used.The current of LVDCC was recorded by patch clamp technique in the whole cell mode.The action potential duration (APD90),amplitude of action potential plateau (APA),ICa-L peak current density of LVDCC were recorded. The mRNA and protein expressions of αlc subunit (CaV1

  8. Bradykinin modulates potassium and calcium currents in neuroblastoma hybrid cells via different pertussis toxin-insensitive pathways.

    Science.gov (United States)

    Wilk-Blaszczak, M A; Gutowski, S; Sternweis, P C; Belardetti, F

    1994-01-01

    In NG108-15 cells, bradykinin (BK) activates a potassium current (IK,BK) and inhibits the voltage-dependent calcium current (ICa,V). BK also stimulates a phosphatidylinositol-specific phospholipase C (PI-PLC). The subsequent release of inositol 1,4,5-trisphosphate and increase in intracellular calcium contribute to IK,BK, through activation of a calcium-dependent potassium current. In membranes from these cells, stimulation of PI-PLC by BK is mediated by Gq and/or G11, two homologous, pertussis toxin-insensitive G proteins. Here, we have investigated the role of Gq/11 in the electrical responses to BK. GTP gamma S mimicked and occluded both actions of BK, and both effects were insensitive to pertussis toxin. Perfusion of an anti-Gq/11 alpha antibody into the pipette suppressed IK,BK, but not the inhibition of ICa,V by BK. Thus, BK couples to IK,BK via Gq/11, but coupling to ICa,V is most likely via a different, pertussis toxin-insensitive G protein.

  9. Imaging fast calcium currents beyond the limitations of electrode techniques.

    Science.gov (United States)

    Jaafari, Nadia; De Waard, Michel; Canepari, Marco

    2014-09-16

    The current understanding of Ca(2+) channel function is derived from the use of the patch-clamp technique. In particular, the measurement of fast cellular Ca(2+) currents is routinely achieved using whole-cell voltage-clamp recordings. However, this experimental approach is not applicable to the study of local native Ca(2+) channels during physiological changes of membrane potential in complex cells, since the voltage-clamp configuration constrains the membrane potential to a given value. Here, we report for the first time to our knowledge that Ca(2+) currents from individual cells can be quantitatively measured beyond the limitations of the voltage-clamp approach using fast Ca(2+) imaging with low-affinity indicators. The optical measurement of the Ca(2+) current was correlated with the membrane potential, simultaneously measured with a voltage-sensitive dye to investigate the activation of Ca(2+) channels along the apical dendrite of the CA1 hippocampal pyramidal neuron during the back-propagation of an action potential. To validate the method, we analyzed the voltage dependence of high- and low-voltage-gated Ca(2+) channels. In particular, we measured the Ca(2+) current component mediated by T-type channels, and we investigated the mechanisms of recovery from inactivation of these channels. This method is expected to become a reference approach to investigate Ca(2+) channels in their native physiological environment.

  10. Hypotonicity activates a voltage-dependent membrane conductance in N2a neuroblastoma cells.

    Science.gov (United States)

    Taruno, Akiyuki; Marunaka, Yoshinori

    2017-03-04

    To maintain cellular and bodily homeostasis, cells respond to extracellular stimuli including osmotic stress by activating various ion channels, which have been implicated in many physiological and pathophysiological conditions. However, cellular osmosensory mechanisms remain elusive. Here, we report a novel voltage-dependent current in N2a cells activated by exposure to hypotonic stress. After a hypotonic challenge, N2a cells sequentially develop two distinct currents. The volume-regulated anion channel (VRAC) current emerges first and, after a delay, activation of a previously uncharacterized strongly outwardly rectifying current follows. The latter, delayed current (Id) is insensitive to NPPB, a nonspecific blocker of Cl(-) channels, and intracellular Mg(2+), which inhibits VRAC and swelling-activated TRPM3 and TRPM7 channels. Replacement of extracellular Na(+) with NMDG(+) reduces inward tail currents, suggesting that Id is mediated by cations. Finally, Id shows voltage-dependent activation with slow activation kinetics and half-maximal activation at +76 mV. These pharmacological and biophysical characteristics of Id are distinct from those of known osmotic cell swelling-activated ion channels. In conclusion, our data identify and characterize a novel osmotically-activated, voltage-dependent ion channel in N2a cells.

  11. The voltage dependence of Ih in human myelinated axons

    Science.gov (United States)

    Howells, James; Trevillion, Louise; Bostock, Hugh; Burke, David

    2012-01-01

    HCN channels are responsible for Ih, a voltage-gated inwardly rectifying current activated by hyperpolarization. This current appears to be more active in human sensory axons than motor and may play a role in the determination of threshold. Differences in Ih are likely to be responsible for the high variability in accommodation to hyperpolarization seen in different subjects. The aim of this study was to characterise this current in human axons, both motor and sensory. Recordings of multiple axonal excitability properties were performed in 10 subjects, with a focus on the changes in threshold evoked by longer and stronger hyperpolarizing currents than normally studied. The findings confirm that accommodation to hyperpolarization is greater in sensory than motor axons in all subjects, but the variability between subjects was greater than the modality difference. An existing model of motor axons was modified to take into account the behaviour seen with longer and stronger hyperpolarization, and a mathematical model of human sensory axons was developed based on the data collected. The differences in behaviour of sensory and motor axons and the differences between different subjects are best explained by modulation of the voltage dependence, along with a modest increase of expression of the underlying conductance of Ih. Accommodation to hyperpolarization for the mean sensory data is fitted well with a value of −94.2 mV for the mid-point of activation (V0.5) of Ih as compared to −107.3 mV for the mean motor data. The variation in response to hyperpolarization between subjects is accounted for by varying this parameter for each modality (sensory: −89.2 to −104.2 mV; motor −87.3 to −127.3 mV). These voltage differences are within the range that has been described for physiological modulation of Ih function. The presence of slowly activated Ih isoforms on both motor and sensory axons was suggested by modelling a large internodal leak current and a masking of

  12. Voltage Dependence of Conformational Dynamics and Subconducting States of VDAC-1.

    Science.gov (United States)

    Briones, Rodolfo; Weichbrodt, Conrad; Paltrinieri, Licia; Mey, Ingo; Villinger, Saskia; Giller, Karin; Lange, Adam; Zweckstetter, Markus; Griesinger, Christian; Becker, Stefan; Steinem, Claudia; de Groot, Bert L

    2016-09-20

    The voltage-dependent anion channel 1 (VDAC-1) is an important protein of the outer mitochondrial membrane that transports energy metabolites and is involved in apoptosis. The available structures of VDAC proteins show a wide β-stranded barrel pore, with its N-terminal α-helix (N-α) bound to its interior. Electrophysiology experiments revealed that voltage, its polarity, and membrane composition modulate VDAC currents. Experiments with VDAC-1 mutants identified amino acids that regulate the gating process. However, the mechanisms for how these factors regulate VDAC-1, and which changes they trigger in the channel, are still unknown. In this study, molecular dynamics simulations and single-channel experiments of VDAC-1 show agreement for the current-voltage relationships of an "open" channel and they also show several subconducting transient states that are more cation selective in the simulations. We observed voltage-dependent asymmetric distortions of the VDAC-1 barrel and the displacement of particular charged amino acids. We constructed conformational models of the protein voltage response and the pore changes that consistently explain the protein conformations observed at opposite voltage polarities, either in phosphatidylethanolamine or phosphatidylcholine membranes. The submicrosecond VDAC-1 voltage response shows intrinsic structural changes that explain the role of key gating amino acids and support some of the current gating hypotheses. These voltage-dependent protein changes include asymmetric barrel distortion, its interaction with the membrane, and significant displacement of N-α amino acids.

  13. Voltage dependence of Na translocation by the Na/K pump.

    Science.gov (United States)

    Nakao, M; Gadsby, D C

    During each complete reaction cycle, the Na/K pump transports three Na ions out across the cell membrane and two K ions in. The resulting net extrusion of positive charge generates outward membrane current but, until now, it was unclear how that net charge movement occurs. Reasonable possibilities included a single positive charge moving outwards during Na translocation; or a single negative charge moving inwards during K translocation; or either positive or negative charges moving during both translocation steps, but in unequal quantities. Any step that involves net charge movement through the membrane must have voltage-dependent transition rates. Here we report measurements of transient, voltage-dependent, displacement currents generated by the pump when its normal Na/K transport cycle has been interrupted by removal of external K and it is thus constrained to carry out Na/Na exchange. The quantity and voltage sensitivity of the charge moved during these transient currents suggests that Na translocation includes a voltage-dependent transition involving movement of one positive charge across the membrane. This single step can thus fully account for the electrogenic nature of Na/K exchange. The result provides important new insight into the molecular mechanism of active cation transport.

  14. Facilitation of calcium-dependent potassium current.

    Science.gov (United States)

    Thompson, S H

    1994-12-01

    The activation of Ca-dependent K+ current, Ic, was studied in macropatches on the cell bodies of molluscan neurons. When a depolarizing voltage-clamp pulse was applied repeatedly, Ic facilitated in a manner that resembled the facilitation of synaptic transmitter release. Facilitation was characterized by an increase in Ic amplitude, a progressive increase in instantaneous outward current, and a decrease in utilization time. Experiments were done to investigate the mechanism responsible for Ic facilitation. Facilitation was reduced by microinjection of an exogenous Ca2+ buffer into the cytoplasm, indicating that facilitation is a Ca(2+)-dependent process. It was also reduced at elevated temperatures. Conversely, facilitation was greatly potentiated by blocking the Na/Ca exchange mechanism. It is concluded that the facilitation of Ca-dependent K+ current results from the accumulation of Ca2+ at the inner face of the membrane during the repeated activation of Ca2+ channels by depolarization. The Ca2+ indicator fluo-3 was used in fluorescence imaging experiments to measure changes in [Ca]i near the cell membrane during repeated depolarizing pulses and the interpretation of these results was aided by numerical simulations of Ca2+ accumulation, diffusion, and buffering in the peripheral cytoplasm. These experiments showed that the time course of Ic facilitation matches the time course of Ca2+ accumulation at the membrane. It was found that the strength of Ic facilitation varies among patches on the same neuron, suggesting that the accumulation of Ca2+ is not uniform along the inner surface of the membrane and that gradients in [Ca]i develop and are maintained during trains of depolarizing pulses. Potential mechanisms that may lead to local differences in Ca2+ accumulation and Ic facilitation are discussed.

  15. A comparative study of the action of tolperisone on seven different voltage dependent sodium channel isoforms.

    Science.gov (United States)

    Hofer, Doris; Lohberger, Birgit; Steinecker, Bibiane; Schmidt, Kurt; Quasthoff, Stefan; Schreibmayer, Wolfgang

    2006-05-24

    The specific, acute interaction of tolperisone, an agent used as a muscle relaxant and for the treatment of chronic pain conditions, with the Na(v1.2), Na(v1.3), Na(v1.4), Na(v1.5), Na(v1.6), Na(v1.7), and Na(v1.8) isoforms of voltage dependent sodium channels was investigated and compared to that of lidocaine. Voltage dependent sodium channels were expressed in the Xenopus laevis oocyte expression system and sodium currents were recorded with the two electrode voltage clamp technique. Cumulative dose response relations revealed marked differences in IC(50) values between the two drugs on identical isoforms, as well as between isoforms. A detailed kinetic analysis uncovered that tolperisone as well as lidocaine exhibited their blocking action not only via state dependent association/dissociation with voltage dependent sodium channels, but a considerable fraction of inhibition is tonic, i.e. permanent and basic in nature. Voltage dependent activation was affected to a minor extent only. A shift in steady-state inactivation to more negative potentials could be observed for most drug/isoform combinations. The contribution of this shift to overall block was, however, small at drug concentrations resulting in considerable overall block. Recovery from inactivation was affected notably by both drugs. Lidocaine application led to a pronounced prolongation of the time constant of the fast recovery process for the Na(v1.3), Na(v1.5), and Na(v1.7) isoforms, indicating common structural properties in the local anesthetic receptor site of these three proteins. Interestingly, this characteristic drug action was not observed for tolperisone.

  16. PIP2 regulation of KCNQ channels: biophysical and molecular mechanisms for lipid modulation of voltage-dependent gating

    Directory of Open Access Journals (Sweden)

    Mark Alan Zaydman

    2014-05-01

    Full Text Available Voltage-gated potassium (Kv channels contain voltage-sensing (VSD and pore-gate (PGD structural domains. During voltage-dependent gating, conformational changes in the two domains are coupled giving rise to voltage-dependent opening of the channel. In addition to membrane voltage, KCNQ (Kv7 channel opening requires the membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2. Recent studies suggest that PIP2 serves as a cofactor to mediate VSD-PGD coupling in KCNQ1 channels. In this review, we put these findings in the context of the current understanding of voltage-dependent gating, lipid modulation of Kv channel activation, and PIP2-regulation of KCNQ channels. We suggest that lipid-mediated coupling of functional domains is a common mechanism among KCNQ channels that may be applicable to other Kv channels and membrane proteins.

  17. PIP2 regulation of KCNQ channels: biophysical and molecular mechanisms for lipid modulation of voltage-dependent gating.

    Science.gov (United States)

    Zaydman, Mark A; Cui, Jianmin

    2014-01-01

    Voltage-gated potassium (Kv) channels contain voltage-sensing (VSD) and pore-gate (PGD) structural domains. During voltage-dependent gating, conformational changes in the two domains are coupled giving rise to voltage-dependent opening of the channel. In addition to membrane voltage, KCNQ (Kv7) channel opening requires the membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2). Recent studies suggest that PIP2 serves as a cofactor to mediate VSD-PGD coupling in KCNQ1 channels. In this review, we put these findings in the context of the current understanding of voltage-dependent gating, lipid modulation of Kv channel activation, and PIP2-regulation of KCNQ channels. We suggest that lipid-mediated coupling of functional domains is a common mechanism among KCNQ channels that may be applicable to other Kv channels and membrane proteins.

  18. Pharmacology of the human cell voltage-dependent cation channel. Part II: inactivation and blocking

    DEFF Research Database (Denmark)

    Bennekou, Poul; Barksmann, Trine L.; Kristensen, Berit I.

    2004-01-01

    Human red cells; Nonselective voltage-dependent cation channel; NSVDC channel; Thiol group reagents......Human red cells; Nonselective voltage-dependent cation channel; NSVDC channel; Thiol group reagents...

  19. Inhibitors of protein kinase C prevent enhancement of calcium current and action potentials in peptidergic neurons of Aplysia.

    Science.gov (United States)

    Conn, P J; Strong, J A; Kaczmarek, L K

    1989-02-01

    Following brief stimulation of an afferent pathway, the bag cell neurons of Aplysia undergo a dramatic change in excitability, resulting in a prolonged discharge of spontaneous action potentials. During the discharge, the action potentials of the bag cell neurons become enhanced in height and width. The afterdischarge triggers release of neuroactive peptides that initiate egg-laying behavior in this animal. Evidence suggests that changes in excitability of the bag cell neurons may be mediated by activation of protein kinase C (PKC) and cAMP-dependent protein kinase (cAMP-PK). PKC activators, such as the phorbol ester TPA (12-O-tetradecanoyl-13-phorbol acetate), enhance the amplitude of action potentials in isolated bag cell neurons in cell culture. These agents act by unmasking a previously covert species of voltage-dependent calcium channel resulting in an increase in calcium current. In the accompanying paper (Conn et al., 1989), we showed that H-7, a protein kinase inhibitor, inhibits the effect of TPA, and is a selective inhibitor of PKC relative to cAMP-PK in these cells. We now report that another PKC inhibitor, sphinganine, also inhibits the effect of TPA on action potential height and calcium current in cultured bag cell neurons, and that N-acetylsphinganine, an inactive sphinganine analog, fails to inhibit the effects of PKC activators. Although both H-7 and sphinganine prevent the effects of TPA when added prior to TPA addition, neither compound reverses the effects of TPA when added after the action potentials have already become enhanced by TPA.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Charged Residues at the First Transmembrane Region Contribute to the Voltage Dependence of the Slow Gate of Connexins.

    Science.gov (United States)

    Pinto, Bernardo I; García, Isaac E; Pupo, Amaury; Retamal, Mauricio A; Martínez, Agustín D; Latorre, Ramón; González, Carlos

    2016-07-22

    Connexins (Cxs) are a family of membrane-spanning proteins that form gap junction channels and hemichannels. Connexin-based channels exhibit two distinct voltage-dependent gating mechanisms termed slow and fast gating. Residues located at the C terminus of the first transmembrane segment (TM-1) are important structural components of the slow gate. Here, we determined the role of the charged residues at the end of TM-1 in voltage sensing in Cx26, Cx46, and Cx50. Conductance/voltage curves obtained from tail currents together with kinetics analysis reveal that the fast and slow gates of Cx26 involves the movement of two and four charges across the electric field, respectively. Primary sequence alignment of different Cxs shows the presence of well conserved glutamate residues in the C terminus of TM-1; only Cx26 contains a lysine in that position (lysine 41). Neutralization of lysine 41 in Cx26 increases the voltage dependence of the slow gate. Swapping of lysine 41 with glutamate 42 maintains the voltage dependence. In Cx46, neutralization of negative charges or addition of a positive charge in the Cx26 equivalent region reduced the slow gate voltage dependence. In Cx50, the addition of a glutamate in the same region decreased the voltage dependence, and the neutralization of a negative charge increased it. These results indicate that the charges at the end of TM-1 are part of the slow gate voltage sensor in Cxs. The fact that Cx42, which has no charge in this region, still presents voltage-dependent slow gating suggests that charges still unidentified also contribute to the slow gate voltage sensitivity.

  1. Conducting and voltage-dependent behaviors of potassium ion channels reconstituted from diaphragm sarcoplasmic reticulum: comparison with the cardiac isoform.

    Science.gov (United States)

    Picher, M; Decrouy, A; Rousseau, E

    1996-02-21

    Sarcoplasmic reticulum (SR) K+ channels from canine diaphragm were studied upon fusion of longitudinal and junctional membrane vesicles into planar lipid bilayers (PLB). The large-conductance cation selective channel (gamma(max) = 250 pS; Km = 33 mM) displays long-lasting open events which are much more frequent at positive than at negative voltages. A major subconducting state about 45% of the fully-open state current amplitude was occasionally observed at all voltages. The voltage-dependence of the open probability displays a sigmoid relationship that was fitted by the Boltzmann equation and expressed in terms of thermodynamic parameters, namely the free energy (delta Gi) and the effective gating charge (Zs): delta Gi = 0.27 kcal/mol and Zs = -1.19 in 250 mM potassium gluconate (K-gluconate). Kinetic analyses also confirmed the voltage-dependent gating behavior of this channel, and indicate the implication of at least two open and three closed states. The diaphragm SR K+ channel shares several biophysical properties with the cardiac isoform: g = 180 pS, delta Gi = 0.75 kcal/mol, Zs = -1.45 in 150 mM K-gluconate, and a similar sigmoid P(o)/voltage relationship. Little is known about the regulation of the diaphragm and cardiac SR K+ channels. The conductance and gating of these channels were not influenced by physiological concentrations of Ca2+ (0.1 microM-1 mM) or Mg2+ (0.25-1 mM), as well as by cGMP (25-100 microM), lemakalim (1-100 microM), glyburide (up to 10 microM) or charybdotoxin (45-200 nM), added either to the cis or to the trans chamber. The apparent lack of biochemical or pharmacological modulation of these channels implies that they are not related to any of the well characterized surface membrane K+ channels. On the other hand, their voltage sensitivity strongly suggests that their activity could be modulated by putative changes in SR membrane potential that might occur during calcium fluxes.

  2. Voltage-dependent metabolic regulation of Kv2.1 channels in pancreatic beta-cells.

    Science.gov (United States)

    Yoshida, Masashi; Nakata, Masanori; Yamato, Shiho; Dezaki, Katsuya; Sugawara, Hitoshi; Ishikawa, San-e; Kawakami, Masanobu; Yada, Toshihiko; Kakei, Masafumi

    2010-05-28

    Voltage-gated potassium channels (Kv channels) play a crucial role in formation of action potentials in response to glucose stimulation in pancreatic beta-ells. We previously reported that the Kv channel is regulated by glucose metabolism, particularly by MgATP. We examined whether the regulation of Kv channels is voltage-dependent and mechanistically related with phosphorylation of the channels. In rat pancreatic beta-cells, suppression of glucose metabolism with low glucose concentrations of 2.8mM or less or by metabolic inhibitors decreased the Kv2.1-channel activity at positive membrane potentials, while increased it at potentials negative to -10 mV, suggesting that modulation of Kv channels by glucose metabolism is voltage-dependent. Similarly, in HEK293 cells expressing the recombinant Kv2.1 channels, 0mM but not 10mM MgATP modulated the channel activity in a manner similar to that in beta-cells. Both steady-state activation and inactivation kinetics of the channel were shifted toward the negative potential in association with the voltage-dependent modulation of the channels by cytosolic dialysis of alkaline phosphatase in beta-cells. The modulation of Kv-channel current-voltage relations were also observed during and after glucose-stimulated electrical excitation. These results suggest that the cellular metabolism including MgATP production and/or channel phosphorylation/dephosphorylation underlie the physiological modulation of Kv2.1 channels during glucose-induced insulin secretion.

  3. Outward potassium current oscillations in macrophage polykaryons: extracellular calcium entry and calcium-induced calcium release

    Directory of Open Access Journals (Sweden)

    Saraiva R.M.

    1997-01-01

    Full Text Available Outward current oscillations associated with transient membrane hyperpolarizations were induced in murine macrophage polykaryons by membrane depolarization in the absence of external Na+. Oscillations corresponded to a cyclic activation of Ca2+-dependent K+ currents (IKCa probably correlated with variations in intracellular Ca2+ concentration. Addition of external Na+ (8 mM immediately abolished the outward current oscillations, suggesting that the absence of the cation is necessary not only for their induction but also for their maintenance. Oscillations were completely blocked by nisoldipine. Ruthenium red and ryanodine reduced the number of outward current cycles in each episode, whereas quercetin prolonged the hyperpolarization 2- to 15-fold. Neither low molecular weight heparin nor the absence of a Na+ gradient across the membrane had any influence on oscillations. The evidence suggests that Ca2+ entry through a pathway sensitive to Ca2+ channel blockers is elicited by membrane depolarization in Na+-free medium and is essential to initiate oscillations, which are also dependent on the cyclic release of Ca2+ from intracellular Ca2+-sensitive stores; Ca2+ ATPase acts by reducing intracellular Ca2+, thus allowing slow deactivation of IKCa. Evidence is presented that neither a Na+/Ca2+ antiporter nor Ca2+ release from IP3-sensitive Ca2+ stores participate directly in the mechanism of oscillation

  4. Glucocorticoids specifically enhance L-type calcium current amplitude and affect calcium channel subunit expression in the mouse hippocampus.

    NARCIS (Netherlands)

    Chameau, P.J.P.; Qin, Y.J.; Smit, G.; Joëls, M.

    2007-01-01

    Previous studies have shown that corticosterone enhances whole cell calcium currents in CA1 pyramidal neurons, through a pathway involving binding of glucocorticoid receptor homodimers to the DNA. We examined whether glucocorticoids show selectivity for L- over N-type of calcium currents. Moreover,

  5. Visinin-like neuronal calcium sensor proteins regulate the slow calcium-activated afterhyperpolarizing current in the rat cerebral cortex

    Science.gov (United States)

    Villalobos, Claudio; Andrade, Rodrigo

    2010-01-01

    Many neurons in the nervous systems express afterhyperpolarizations that are mediated by a slow calcium-activated potassium current. This current shapes neuronal firing and is inhibited by neuromodulators, suggesting an important role in the regulation of neuronal function. Surprisingly, very little is currently known about the molecular basis for this current or how it is gated by calcium. Recently, the neuronal calcium sensor protein hippocalcin was identified as a calcium sensor for the slow afterhyperpolarizing current in the hippocampus. However, while hippocalcin is very strongly expressed in the hippocampus, this protein shows a relatively restricted distribution in the brain. Furthermore, the genetic deletion of this protein only partly reduces the slow hyperpolarizing current in hippocampus. These considerations question whether hippocalcin can be the sole calcium sensor for the slow afterhyperpolarizing current. Here we use loss of function and overexpression strategies to show that hippocalcin functions as a calcium sensor for the slow afterhyperpolarizing current in the cerebral cortex, an area where hippocalcin is expressed at much lower levels than in hippocampus. In addition we show that neurocalcin δ, but not VILIP-2, can also act as a calcium sensor for the slow afterhyperpolarizing current. Finally we show that hippocalcin and neurocalcin δ both increase the calcium sensitivity of the afterhyperpolarizing current but do not alter its sensitivity to inhibition by carbachol acting through the Gαq-11-PLCβ signaling cascade. These results point to a general role for a subgroup of visinin-like neuronal calcium sensor proteins in the activation of the slow calcium-activated afterhyperpolarizing current. PMID:20980592

  6. Two distinct voltage-sensing domains control voltage sensitivity and kinetics of current activation in CaV1.1 calcium channels.

    Science.gov (United States)

    Tuluc, Petronel; Benedetti, Bruno; Coste de Bagneaux, Pierre; Grabner, Manfred; Flucher, Bernhard E

    2016-06-01

    Alternative splicing of the skeletal muscle CaV1.1 voltage-gated calcium channel gives rise to two channel variants with very different gating properties. The currents of both channels activate slowly; however, insertion of exon 29 in the adult splice variant CaV1.1a causes an ∼30-mV right shift in the voltage dependence of activation. Existing evidence suggests that the S3-S4 linker in repeat IV (containing exon 29) regulates voltage sensitivity in this voltage-sensing domain (VSD) by modulating interactions between the adjacent transmembrane segments IVS3 and IVS4. However, activation kinetics are thought to be determined by corresponding structures in repeat I. Here, we use patch-clamp analysis of dysgenic (CaV1.1 null) myotubes reconstituted with CaV1.1 mutants and chimeras to identify the specific roles of these regions in regulating channel gating properties. Using site-directed mutagenesis, we demonstrate that the structure and/or hydrophobicity of the IVS3-S4 linker is critical for regulating voltage sensitivity in the IV VSD, but by itself cannot modulate voltage sensitivity in the I VSD. Swapping sequence domains between the I and the IV VSDs reveals that IVS4 plus the IVS3-S4 linker is sufficient to confer CaV1.1a-like voltage dependence to the I VSD and that the IS3-S4 linker plus IS4 is sufficient to transfer CaV1.1e-like voltage dependence to the IV VSD. Any mismatch of transmembrane helices S3 and S4 from the I and IV VSDs causes a right shift of voltage sensitivity, indicating that regulation of voltage sensitivity by the IVS3-S4 linker requires specific interaction of IVS4 with its corresponding IVS3 segment. In contrast, slow current kinetics are perturbed by any heterologous sequences inserted into the I VSD and cannot be transferred by moving VSD I sequences to VSD IV. Thus, CaV1.1 calcium channels are organized in a modular manner, and control of voltage sensitivity and activation kinetics is accomplished by specific molecular mechanisms

  7. Conductance hysteresis in the voltage-dependent anion channel.

    Science.gov (United States)

    Rappaport, Shay M; Teijido, Oscar; Hoogerheide, David P; Rostovtseva, Tatiana K; Berezhkovskii, Alexander M; Bezrukov, Sergey M

    2015-09-01

    Hysteresis in the conductance of voltage-sensitive ion channels is observed when the transmembrane voltage is periodically varied with time. Although this phenomenon has been used in studies of gating of the voltage-dependent anion channel, VDAC, from the outer mitochondrial membrane for nearly four decades, full hysteresis curves have never been reported, because the focus was solely on the channel opening branches of the hysteresis loops. We studied the hysteretic response of a multichannel VDAC system to a triangular voltage ramp the frequency of which was varied over three orders of magnitude, from 0.5 mHz to 0.2 Hz. We found that in this wide frequency range the area encircled by the hysteresis curves changes by less than a factor of three, suggesting broad distribution of the characteristic times and strongly non-equilibrium behavior. At the same time, quasi-equilibrium two-state behavior is observed for hysteresis branches corresponding to VDAC opening. This enables calculation of the usual equilibrium gating parameters, gating charge and voltage of equipartitioning, which were found to be almost insensitive to the ramp frequency. To rationalize this peculiarity, we hypothesize that during voltage-induced closure and opening the system explores different regions of the complex free energy landscape, and, in the opening branch, follows quasi-equilibrium paths.

  8. Pharmacological Investigation of Voltage-dependent Ca2+ Channels in Human Ejaculatory Sperm in vitro

    Institute of Scientific and Technical Information of China (English)

    LI Lu; LIU Jihong; LI Jiagui; YE Zhangqun

    2006-01-01

    The types of the voltage-dependent calcium channels (VDCCs) in human ejaculatory sperm and the effects of calcium channel blocker (CCB) on human sperm motility parameters in vitro were investigated. The human sperm motility parameters in vitro in response to the pharmacological agents nifedipine (NIF, inhibitor of L-type VDCC) and ω-conotoxin (GVIA, inhibitor of N-type VDCC) were compared and analyzed statistically. The results showed that NIF (1, 5, 10 μmol/L)could not only significantly affect human sperm's shape but also spermatozoa motility after incubated at least 10 min in vitro (P<0.001). GVIA (0.1, 0.5 and 1 μmol/L) could just only significantly affect human sperm's progressive motility (a %+b %) after incubated for 20 min in vitro (P<0.01), but they both could not significantly affect spermic abnormality rate. It is suggested that L-type VDCC, non L-type VDCCs and isoform of L-type VDCC exist in the cell membrane of human sperm solely or together, and they participate in the spermic physiological processes especially the spermic motility.

  9. Cellular elements for seeing in the dark: voltage-dependent conductances in cockroach photoreceptors

    Directory of Open Access Journals (Sweden)

    Salmela Iikka

    2012-08-01

    Full Text Available Abstract Background The importance of voltage-dependent conductances in sensory information processing is well-established in insect photoreceptors. Here we present the characterization of electrical properties in photoreceptors of the cockroach (Periplaneta americana, a nocturnal insect with a visual system adapted for dim light. Results Whole-cell patch-clamped photoreceptors had high capacitances and input resistances, indicating large photosensitive rhabdomeres suitable for efficient photon capture and amplification of small photocurrents at low light levels. Two voltage-dependent potassium conductances were found in the photoreceptors: a delayed rectifier type (KDR and a fast transient inactivating type (KA. Activation of KDR occurred during physiological voltage responses induced by light stimulation, whereas KA was nearly fully inactivated already at the dark resting potential. In addition, hyperpolarization of photoreceptors activated a small-amplitude inward-rectifying (IR current mediated at least partially by chloride. Computer simulations showed that KDR shapes light responses by opposing the light-induced depolarization and speeding up the membrane time constant, whereas KA and IR have a negligible role in the majority of cells. However, larger KA conductances were found in smaller and rapidly adapting photoreceptors, where KA could have a functional role. Conclusions The relative expression of KA and KDR in cockroach photoreceptors was opposite to the previously hypothesized framework for dark-active insects, necessitating further comparative work on the conductances. In general, the varying deployment of stereotypical K+ conductances in insect photoreceptors highlights their functional flexibility in neural coding.

  10. MCU encodes the pore conducting mitochondrial calcium currents.

    Science.gov (United States)

    Chaudhuri, Dipayan; Sancak, Yasemin; Mootha, Vamsi K; Clapham, David E

    2013-06-04

    Mitochondrial calcium (Ca(2+)) import is a well-described phenomenon regulating cell survival and ATP production. Of multiple pathways allowing such entry, the mitochondrial Ca(2+) uniporter is a highly Ca(2+)-selective channel complex encoded by several recently-discovered genes. However, the identity of the pore-forming subunit remains to be established, since knockdown of all the candidate uniporter genes inhibit Ca(2+) uptake in imaging assays, and reconstitution experiments have been equivocal. To definitively identify the channel, we use whole-mitoplast voltage-clamping, the technique that originally established the uniporter as a Ca(2+) channel. We show that RNAi-mediated knockdown of the mitochondrial calcium uniporter (MCU) gene reduces mitochondrial Ca(2+) current (I MiCa ), whereas overexpression increases it. Additionally, a classic feature of I MiCa , its sensitivity to ruthenium red inhibition, can be abolished by a point mutation in the putative pore domain without altering current magnitude. These analyses establish that MCU encodes the pore-forming subunit of the uniporter channel. DOI:http://dx.doi.org/10.7554/eLife.00704.001.

  11. Effects of caffeine on intracellular calcium, calcium current and calcium-dependent potassium current in anterior pituitary GH3 cells.

    Science.gov (United States)

    Kramer, R H; Mokkapatti, R; Levitan, E S

    1994-01-01

    Caffeine elicits physiological responses in a variety of cell types by triggering the mobilization of Ca2+ from intracellular organelles. Here we investigate the effects of caffeine on intracellular Ca2+ concentration ([Ca2+]i) and ionic currents in anterior pituitary cells (GH3) cells. Caffeine has a biphasic effect on Ca(2+)-activated K+ current [IK(Ca)]: it induces a transient increase superimposed upon a sustained inhibition. While the transient increase coincides with a rise in [Ca2+]i, the sustained inhibition of IK(Ca) is correlated with a sustained inhibition of the L-type Ca2+ current. The L-type Ca2+ current is also inhibited by other agents that mobilize intracellular Ca2+, including thyrotropin releasing hormone (TRH) and ryanodine, but in a matter distinct from caffeine. Unlike the caffeine effect, the TRH-induced inhibition "washes-out" under whole-cell patch-clamp conditions and is eliminated by intracellular Ca2+ chelators. Likewise, the ryanodine-induced inhibition desensitizes while the caffeine-induced inhibition does not. Simultaneous [Ca2+]i and Ca2+ current measurements show that caffeine can inhibit Ca2+ current without changing [Ca2+]i. Single-channel recordings show that caffeine reduces mean open time without affecting single-channel conductance of L-type channels. Hence the effects of caffeine on ion channels in GH3 cells are attributable both to mobilization of intracellular Ca2+ and to a direct effect on the gating of L-type Ca2+ channels.

  12. Butanol isomers exert distinct effects on voltage-gated calcium channel currents and thus catecholamine secretion in adrenal chromaffin cells.

    Directory of Open Access Journals (Sweden)

    Sarah McDavid

    Full Text Available Butanol (C4H10OH has been used both to dissect the molecular targets of alcohols/general anesthetics and to implicate phospholipase D (PLD signaling in a variety of cellular functions including neurotransmitter and hormone exocytosis. Like other primary alcohols, 1-butanol is a substrate for PLD and thereby disrupts formation of the intracellular signaling lipid phosphatidic acid. Because secondary and tertiary butanols do not undergo this transphosphatidylation, they have been used as controls for 1-butanol to implicate PLD signaling. Recently, selective pharmacological inhibitors of PLD have been developed and, in some cases, fail to block cellular functions previously ascribed to PLD using primary alcohols. For example, exocytosis of insulin and degranulation of mast cells are blocked by primary alcohols, but not by the PLD inhibitor FIPI. In this study we show that 1-butanol reduces catecholamine secretion from adrenal chromaffin cells to a much greater extent than tert-butanol, and that the PLD inhibitor VU0155056 has no effect. Using fluorescent imaging we show the effect of these drugs on depolarization-evoked calcium entry parallel those on secretion. Patch-clamp electrophysiology confirmed the peak amplitude of voltage-gated calcium channel currents (I(Ca is inhibited by 1-butanol, with little or no block by secondary or tert-butanol. Detailed comparison shows for the first time that the different butanol isomers exert distinct, and sometimes opposing, effects on the voltage-dependence and gating kinetics of I(Ca. We discuss these data with regard to PLD signaling in cellular physiology and the molecular targets of general anesthetics.

  13. Butanol isomers exert distinct effects on voltage-gated calcium channel currents and thus catecholamine secretion in adrenal chromaffin cells.

    Science.gov (United States)

    McDavid, Sarah; Bauer, Mary Beth; Brindley, Rebecca L; Jewell, Mark L; Currie, Kevin P M

    2014-01-01

    Butanol (C4H10OH) has been used both to dissect the molecular targets of alcohols/general anesthetics and to implicate phospholipase D (PLD) signaling in a variety of cellular functions including neurotransmitter and hormone exocytosis. Like other primary alcohols, 1-butanol is a substrate for PLD and thereby disrupts formation of the intracellular signaling lipid phosphatidic acid. Because secondary and tertiary butanols do not undergo this transphosphatidylation, they have been used as controls for 1-butanol to implicate PLD signaling. Recently, selective pharmacological inhibitors of PLD have been developed and, in some cases, fail to block cellular functions previously ascribed to PLD using primary alcohols. For example, exocytosis of insulin and degranulation of mast cells are blocked by primary alcohols, but not by the PLD inhibitor FIPI. In this study we show that 1-butanol reduces catecholamine secretion from adrenal chromaffin cells to a much greater extent than tert-butanol, and that the PLD inhibitor VU0155056 has no effect. Using fluorescent imaging we show the effect of these drugs on depolarization-evoked calcium entry parallel those on secretion. Patch-clamp electrophysiology confirmed the peak amplitude of voltage-gated calcium channel currents (I(Ca)) is inhibited by 1-butanol, with little or no block by secondary or tert-butanol. Detailed comparison shows for the first time that the different butanol isomers exert distinct, and sometimes opposing, effects on the voltage-dependence and gating kinetics of I(Ca). We discuss these data with regard to PLD signaling in cellular physiology and the molecular targets of general anesthetics.

  14. Phosphorylation of purified mitochondrial Voltage-Dependent Anion Channel by c-Jun N-terminal Kinase-3 modifies channel voltage-dependence

    Directory of Open Access Journals (Sweden)

    Rajeev Gupta

    2017-06-01

    Full Text Available Voltage-Dependent Anion Channel (VDAC phosphorylated by c-Jun N-terminal Kinase-3 (JNK3 was incorporated into the bilayer lipid membrane. Single-channel electrophysiological properties of the native and the phosphorylated VDAC were compared. The open probability versus voltage curve of the native VDAC displayed symmetry around the voltage axis, whereas that of the phosphorylated VDAC showed asymmetry. This result indicates that phosphorylation by JNK3 modifies voltage-dependence of VDAC.

  15. Sex and regional differences in rabbit right ventricular L-type calcium current levels and mathematical modelling of arrhythmia vulnerability.

    Science.gov (United States)

    Kalik, Zane M; Mike, Joshua L; Slipski, Cassandra; Wright, Moriah; Jalics, Jozsi Z; Womble, Mark D

    2017-07-01

    What is the central question of this study? Regional variations of ventricular L-type calcium current (ICa-L ) amplitude may underlie the increased arrhythmia risk in adult females. Current amplitude variations have been described for the left ventricle but not for the right ventricle. What is the main finding and its importance? Adult female rabbit right ventricular base myocytes exhibit elevated ICa-L compared with female apex or male myocytes. Oestrogen upregulated ICa-L in cultured female myocytes. Mathematical simulations modelling long QT syndrome type 2 demonstrated that elevated ICa-L prolonged action potentials and induced early after-depolarizations. Thus, ventricular arrhythmias in adult females may be associated with an oestrogen-induced upregulation of ICa-L . Previous studies have shown that adult rabbit left ventricular myocytes exhibit sex and regional differences in L-type calcium current (ICa-L ) levels that contribute to increased female susceptibility to arrhythmogenic early after-depolarizations (EADs). We used patch-clamp recordings from isolated adult male and female rabbit right ventricular myocytes to determine apex-base differences in ICa-L density and used mathematical modelling to examine the contribution of ICa-L to EAD formation. Current density measured at 0 mV in female base myocytes was 67% higher than in male base myocytes and 55% higher than in female apex myocytes. No differences were observed between male and female apex myocytes, between male apex and base myocytes, or in the voltage dependences of ICa-L activation or inactivation. The role of oestrogen was investigated using cultured adult female right ventricular base myocytes. After 2 days, 17β-estradiol (1 nm) produced a 65% increase in ICa-L density compared with untreated control myocytes, suggesting an oestrogen-induced upregulation of ICa-L . Action potential simulations using a modified Luo-Rudy cardiomyocyte model showed that increased ICa-L density, at the level

  16. The Eag domain regulates the voltage-dependent inactivation of rat Eag1 K+ channels.

    Directory of Open Access Journals (Sweden)

    Ting-Feng Lin

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

  17. Control of spontaneous firing patterns by the selective coupling of calcium currents to calcium-activated potassium currents in striatal cholinergic interneurons.

    Science.gov (United States)

    Goldberg, Joshua A; Wilson, Charles J

    2005-11-02

    The spontaneous firing patterns of striatal cholinergic interneurons are sculpted by potassium currents that give rise to prominent afterhyperpolarizations (AHPs). Large-conductance calcium-activated potassium (BK) channel currents contribute to action potential (AP) repolarization; small-conductance calcium-activated potassium channel currents generate an apamin-sensitive medium AHP (mAHP) after each AP; and bursts of APs generate long-lasting slow AHPs (sAHPs) attributable to apamin-insensitive currents. Because all these currents are calcium dependent, we conducted voltage- and current-clamp whole-cell recordings while pharmacologically manipulating calcium channels of the plasma membrane and intracellular stores to determine what sources of calcium activate the currents underlying AP repolarization and the AHPs. The Cav2.2 (N-type) blocker omega-conotoxin GVIA (1 microM) was the only blocker that significantly reduced the mAHP, and it induced a transition to rhythmic bursting in one-third of the cells tested. Cav1 (L-type) blockers (10 microM dihydropyridines) were the only ones that significantly reduced the sAHP. When applied to cells induced to burst with apamin, dihydropyridines reduced the sAHPs and abolished bursting. Depletion of intracellular stores with 10 mM caffeine also significantly reduced the sAHP current and reversibly regularized firing. Application of 1 microM omega-conotoxin MVIIC (a Cav2.1/2.2 blocker) broadened APs but had a negligible effect on APs in cells in which BK channels were already blocked by submillimolar tetraethylammonium chloride, indicating that Cav2.1 (Q-type) channels provide the calcium to activate BK channels that repolarize the AP. Thus, calcium currents are selectively coupled to the calcium-dependent potassium currents underlying the AHPs, thereby creating mechanisms for control of the spontaneous firing patterns of these neurons.

  18. The Mechanism of Voltage Dependent Gating of the NaChBac Prokaryotic Sodium Channel

    Science.gov (United States)

    Decaen, Paul G.

    Electrical signaling in cells depends on selective conductance of ions through membrane proteins called 'voltage gated ion channels'. These channels are characterized by their ability turn on and off the flow of ionic current by opening and closing their conductive pore in response to changes in membrane potential. The opening and closing of the pore is a mechanically linked to conformational movement of the positively charged fourth transmembrane segment (S4) in 'the voltage sensor' region. How the S4 moves in response to membrane potential is a controversial subject. In this thesis, we used the prokaryotic sodium channel NaChBac as our model sodium channel to study voltage dependent movement of the S4 in the voltage sensor. We use a disulfide-locking method where we introduced pairs of cysteines in the voltage sensor that crosslink and trap the S4 in its path after depolarization. We screened over one hundred mutations of the NaChBac channel in the whole cell patch clamp assay and demonstrated discrete and sequential voltage dependent ion pair interactions that occur in at least three states between the positively charged residues of the S4 segment and the acidic residues in the S1, S2 and S3 segments. In conjunction with structural modeling of the voltage sensor and our disulfide locking data, we propose that the S4 moves in and out of the plane of the membrane 8-13 A, forming distinct gating charge interactions with counter charges of the voltage sensor and adopts a 310 helix over a portion of its structure during activation. These findings are compatible with the sliding helix model and refine our understanding of the structural determinates of voltage sensor function in voltage gated ion channels.

  19. Do calcium-dependent ionic currents mediate ischemic ventricular fibrillation?

    Science.gov (United States)

    Clusin, W T; Bristow, M R; Karagueuzian, H S; Katzung, B G; Schroeder, J S

    1982-02-18

    Calcium ions mediate the adverse effects of myocardial ischemia and have been implicated in the genesis of arrhythmias. Calcium influx blocking drugs protect against early ventricular arrhythmias during experimental coronary occlusion, and recent studies suggest that this effect is at least partly due to inhibition of myocardial cell calcium influx. Most of the pharmacologic maneuvers used to simulate acute ischemic arrhythmias in vivo also produce intracellular calcium overload. Production of calcium overload in small myocardial cell clusters causes fibrillatory electrical and mechanical activity similar to that recorded from fibrillating hearts. Fibrillation in these cell clusters is mediated not by reentrant conduction, but by the same subcellular processes that give rise to depolarizing afterpotentials and abnormal automaticity. Agents favoring calcium influx, such as beta adrenergic agonists, accentuate these processes, while agents that depress calcium influx inhibit them. Although the relation of these experimental models to clinical ischemic arrhythmias has not been fully delineated, calcium influx blocking drugs may prove useful in reducing the incidence of sudden cardiac death.

  20. Action of niflumic acid on evoked and spontaneous calcium-activated chloride and potassium currents in smooth muscle cells from rabbit portal vein.

    Science.gov (United States)

    Hogg, R. C.; Wang, Q.; Large, W. A.

    1994-01-01

    1. The action of niflumic acid was studied on spontaneous and evoked calcium-activated chloride (ICl(Ca)) and potassium (IK(Ca)) currents in rabbit isolated portal vein cells. 2. With the nystatin perforated patch technique in potassium-containing solutions at a holding potential of -77 mV (the potassium equilibrium potential), niflumic acid produced a concentration-dependent inhibition of spontaneous transient inward current (STIC, calcium-activated chloride current) amplitude. The concentration to reduce the STIC amplitude by 50% (IC50) was 3.6 x 10(-6) M. 3. At -77 mV holding potential, niflumic acid converted the STIC decay from a single exponential to 2 exponential components. In niflumic acid the fast component of decay was faster, and the slow component was slower than the control decay time constant. Increasing the concentration of niflumic acid enhanced the decay rate of the fast component and reduced the decay rate of the slow component. 4. The effect of niflumic acid on STIC amplitude was voltage-dependent and at -50 and +50 mV the IC50 values were 2.3 x 10(-6) M and 1.1 x 10(-6) M respectively (cf. 3.6 x 10(-6) M at -77 mV). 5. In K-free solutions at potentials of -50 mV and +50 mV, niflumic acid did not induce a dual exponential STIC decay but just increased the decay time constant at both potentials in a concentration-dependent manner. 6. Niflumic acid, in concentrations up to 5 x 10(-5) M, had no effect on spontaneous calcium-activated potassium currents.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7921628

  1. Control of Spontaneous Firing Patterns by the Selective Coupling of Calcium Currents to Calcium Activated Potassium Currents in Striatal Cholinergic Interneurons

    OpenAIRE

    Goldberg, Joshua A.; Wilson, Charles J.

    2005-01-01

    The spontaneous firing patterns of striatal cholinergic interneurons are sculpted by potassium currents that give rise to prominent afterhyperpolarizations (AHPs): BK currents contribute to action potential (AP) repolarization; SK currents generate an apamin-sensitive medium AHP (mAHP) following each AP; and bursts of APs generate long-lasting slow AHPs (sAHPs) due to apamin-insensitive currents. As all these currents are calcium-dependent, we conducted voltage- and current-clamp whole-cell r...

  2. Voltage-dependent Ca2+ channel and Na+ channel in frog taste cells.

    Science.gov (United States)

    Kashiwayanagi, M; Miyake, M; Kurihara, K

    1983-01-01

    Frog taste cells were hyperpolarized by injecting an inward current pulse, and regenerative anode-break potentials were observed at the termination of the current pulse. The results obtained are as follows. 1) The magnitude of the anode-break potentials increased with the extent of hyperpolarization of taste cells and reached a saturation level around -200 mV. 2) The magnitudes of the anode-break potentials observed in 80 different taste cells hyperpolarized to about -200 mV were distributed widely from cell to cell. The average magnitude was 39 mV. 3) The anode-break potentials were recorded after the lingual artery was perfused with artificial solutions containing various channel blockers. The results indicated that the anode-break potentials are composed of Na+ and Ca2+ components. 4) The slope of the current-voltage relation obtained with cells hyperpolarized to 100 mV was appreciably decreased above -50 mV by application of tetrodotoxin to the perfusing solution. Discussion was made on possible roles of the voltage-dependent Na+ and Ca2+ channels in the electrotonic spreading of the depolarization at the receptor membranes to the synaptic area and in releasing a chemical transmitter.

  3. Pharmacology of the human red cell voltage-dependent cation channel Part I. Activation by clotrimazole and analogues

    DEFF Research Database (Denmark)

    Barksmann, Trine Lyberth; Kristensen, Berit I.; Christophersen, Palle.

    2004-01-01

    Human red cells, Nonselective voltage dependent cation channel, NSVDC channel, Gárdos channel blockers, NSVDC channel activators......Human red cells, Nonselective voltage dependent cation channel, NSVDC channel, Gárdos channel blockers, NSVDC channel activators...

  4. Dissociation enzyme effects on the biophysical properties of calcium current in acutely isolated rat ventricular myocytes

    Directory of Open Access Journals (Sweden)

    Julio Álvarez

    2013-04-01

    Full Text Available Proteolytic enzymes such as collagenase, trypsin and pronase E are widely used to acutely dissociate adult cardiomyocytes. There is some evidence that enzyme treatment can alter ionic channels. The aim of the present investigation was to compare the characteristics of the L-type Ca2+ current (ICaL of rat ventricular cardiomyocytes dissociated with two enzyme combinations: collagenase + trypsin (C+T and collagenase + pronase E (C+P. ICaL density (pA/pF was significantly lower (~ 2 pA/pF in myocytes isolated with the C+P combination. However, its inactivation time course was barely affected. As well, the voltage dependency of ICaL kinetics was not affected by the C+P treatment. Our results suggest that, compared to the C+T, treatment with the C+P enzyme combination could decrease the number of functional (expressed channels in the sarcolemma.

  5. Voltage dependence of proton pumping by bacteriorhodopsin mutants with altered lifetime of the M intermediate.

    Directory of Open Access Journals (Sweden)

    Sven Geibel

    Full Text Available The light-driven proton pump bacteriorhodopsin (BR from Halobacterium salinarum is tightly regulated by the [H(+] gradient and transmembrane potential. BR exhibits optoelectric properties, since spectral changes during the photocycle are kinetically controlled by voltage, which predestines BR for optical storage or processing devices. BR mutants with prolonged lifetime of the blue-shifted M intermediate would be advantageous, but the optoelectric properties of such mutants are still elusive. Using expression in Xenopus oocytes and two-electrode voltage-clamping, we analyzed photocurrents of BR mutants with kinetically destabilized (F171C, F219L or stabilized (D96N, D96G M intermediate in response to green light (to probe H(+ pumping and blue laser flashes (to probe accumulation/decay of M. These mutants have divergent M lifetimes. As for BR-WT, this strictly correlates with the voltage dependence of H(+ pumping. BR-F171C and BR-F219L showed photocurrents similar to BR-WT. Yet, BR-F171C showed a weaker voltage dependence of proton pumping. For both mutants, blue laser flashes applied during and after green-light illumination showed reduced M accumulation and shorter M lifetime. In contrast, BR-D96G and BR-D96N exhibited small photocurrents, with nonlinear current-voltage curves, which increased strongly in the presence of azide. Blue laser flashes showed heavy M accumulation and prolonged M lifetime, which accounts for the strongly reduced H(+ pumping rate. Hyperpolarizing potentials augmented these effects. The combination of M-stabilizing and -destabilizing mutations in BR-D96G/F171C/F219L (BR-tri shows that disruption of the primary proton donor Asp-96 is fatal for BR as a proton pump. Mechanistically, M destabilizing mutations cannot compensate for the disruption of Asp-96. Accordingly, BR-tri and BR-D96G photocurrents were similar. However, BR-tri showed negative blue laser flash-induced currents even without actinic green light, indicating

  6. Over Expression of Voltage Dependent Anion Channel 2 (VDAC2 in Muscles of Electrically Stunned Chickens

    Directory of Open Access Journals (Sweden)

    Norshahida Abu Samah, Azura Amid, and Faridah Yusof

    2011-12-01

    Full Text Available Water bath stunning is a common practice in commercial slaughterhouses. Such treatment is economic and in line with animal welfare practice. However, the conditions applied for the stunning process may vary from a slaughterhouse to another slaughterhouse. Such a loose regulation on the stunning procedure has opened up doors for food adulteration such as over dose stunning. In this study, a simple and reliable approach using proteomics have been developed to study the effect of different currents and voltages in stunning on the protein expression of the chickens. Protein profiles of the chickens were constructed in order to detect any differences in protein expression and modifications. The different voltage studied were 10 V, 40 V and 70 V while the values for current studied were 0.25 A, 0.5 A, and 0.75 A. After the proteomics analyses using 2D Platinum ImageMaster 6.0 and Matrix-assisted laser desorption ionization- time of flight (MALDI TOF spectrometry identification, Voltage dependent anion channel 2 (VDAC2 was identified to be over expressed in the muscle sample of over stunned chicken. The over expression of VDAC2 was confirmed at the transcriptional level of RNA expression. Real Time PCR showed that all over stunned samples contained higher mRNA expression level for VDAC2 genes. The mRNA level of VDAC2 was up-regulated by 59.87 fold change when normalized with housekeeping gene. In conclusion, VDAC2 could serve as potential biomarkers for identification of electrically stimulated chickens. The existence of these biomarkers will help to monitor the slaughtering and stunning process in the future. It will revolutionize the food authentication field and give a new breathe to the meat industry.ABSTRAK: Kaedah "waterbath stunning" merupakan amalan biasa di pusat-pusat penyembelihan. Kaedah ini adalah ekonomik dan selari dengan amalan kebajikan haiwan. Walaubagaimanapun, syarat-syarat yang digunakan untuk proses kejutan tersebut mungkin

  7. Calcium currents and conductances in the msucle membrane of the crayfish.

    Science.gov (United States)

    Hencek, M; Zachar, J

    1977-06-01

    1. Membrane currents in calcium type muscle membrane of the cray-fish Astacus fluviatilis were analysed by a method in which a membrane microarea was isolated by circulating sucrose rings contacting the fibre perpendicular to the fibre surface.2. The early calcium inward currents were separated from the total membrane currents by subtraction of the early and delayed potassium currents from the total membrane current.3. The isolated calcium currents show a time course characteristic for a transient change of calcium conductance. The presence of inactivation was further checked by the time course of the tail currents at the end of voltage clamp pulses of variable duration.4. The reversal potential of the early calcium currents determined from the current-voltage relations was +85 +/- 4.2 mV. The calcium potentials were used to express the calcium currents in the form of chord conductances.5. Calcium conductances (g(Ca)) as functions of time and voltage were found to be described quantitatively on the assumption that g(Ca) is determined by two variables (m and h), according to the equation g(Ca) = m(6)hg(Ca), where g(Ca) is a constant and m and h obey first order differential equations of the Hodgkin-Huxley type.6. The activation parameters of the g(Ca) were determined by fitting the solutions of the above equations to the experimental values of the g(Ca). This method was also used to check the inactivation parameters.7. The inactivation parameters of the g(Ca) were obtained from the inactivation curves, which were determined for several membrane potentials by variation of the duration of the conditioning step.8. The average calcium conductance constants were tabulated and compared with sodium conductance constants in excitable membranes.

  8. Tweeters and Woofers: The Complex Orchestra of Calcium Currents in T Lymphocytes

    Directory of Open Access Journals (Sweden)

    Lilian L Nohara

    2015-05-01

    Full Text Available Elevation of intracellular calcium ion (Ca2+ levels is a vital event that regulates T lymphocyte homeostasis, activation, proliferation, differentiation, and apoptosis. The mechanisms that regulate intracellular Ca2+ signalling in lymphocytes involve tightly controlled orchestration of multiple ion channels, membrane receptors, and signalling molecules. T cell receptor (TCR engagement results in depletion of endoplasmic reticulum (ER Ca2+ stores and subsequent sustained influx of extracellular Ca2+ through Ca2+ release-activated Ca2+ (CRAC channels in the plasma membrane. This process termed store-operated Ca2+ entry (SOCE involves the ER Ca2+ sensing molecule, stromal interaction molecule 1 (STIM1, and a pore-forming plasma membrane protein, ORAI1. However, several other important Ca2+ channels that are instrumental in T cell function also exist. In this review, we discuss the role of additional Ca2+ channel families expressed on the plasma membrane of T cells that likely contribute to Ca2+ influx following TCR engagement, which include the IP3 receptors, the P2X receptors, the NMDA receptors, and the TRP channels, with a focus on the voltage-dependent Ca2+ (CaV channels.

  9. N-type calcium current, Cav2.2, is enhanced in small-diameter sensory neurons isolated from Nf1+/- mice.

    Science.gov (United States)

    Duan, J-H; Hodgdon, K E; Hingtgen, C M; Nicol, G D

    2014-06-13

    Major aspects of neuronal function are regulated by Ca(2+) including neurotransmitter release, excitability, developmental plasticity, and gene expression. We reported previously that sensory neurons isolated from a mouse model with a heterozygous mutation of the Nf1 gene (Nf1+/-) exhibited both greater excitability and evoked release of neuropeptides compared to wildtype mice. Furthermore, augmented voltage-dependent sodium currents but not potassium currents contribute to the enhanced excitability. To determine the mechanisms giving rise to the enhanced release of substance P and calcitonin gene-related peptide in the Nf1+/- sensory neurons, the potential differences in the total voltage-dependent calcium current (ICa) as well as the contributions of individual Ca(2+) channel subtypes were assessed. Whole-cell patch-clamp recordings from small-diameter capsaicin-sensitive sensory neurons demonstrated that the average peak ICa densities were not different between the two genotypes. However, by using selective blockers of channel subtypes, the current density of N-type (Cav2.2) ICa was significantly larger in Nf1+/- neurons compared to wildtype neurons. In contrast, there were no significant differences in L-, P/Q- and R-type currents between the two genotypes. Quantitative real-time polymerase chain reaction measurements made from the isolated but intact dorsal root ganglia indicated that N-type (Cav2.2) and P/Q-type (Cav2.1) Ca(2+) channels exhibited the highest mRNA expression levels although there were no significant differences in the levels of mRNA expression between the genotypes. These results suggest that the augmented N-type (Cav2.2) ICa observed in the Nf1+/- sensory neurons does not result from genomic differences but may reflect post-translational or some other non-genomic modifications. Thus, our results demonstrate that sensory neurons from Nf1+/- mice, exhibit increased N-type ICa and likely account for the increased release of substance P and

  10. IgG from Amyotrophic Lateral Sclerosis Patients Increases Current Through P-Type Calcium Channels in Mammalian Cerebellar Purkinje Cells and in Isolated Channel Protein in Lipid Bilayer

    Science.gov (United States)

    Llinas, R.; Sugimori, M.; Cherksey, B. D.; Smith, R. Glenn; Delbono, O.; Stefani, E.; Appel, S.

    1993-12-01

    The effect of the IgG from amyotrophic lateral sclerosis (ALS) patients was tested on the voltage-dependent barium currents (IBa) in mammalian dissociated Purkinje cells and in isolated P-type calcium channels in lipid bilayers. Whole cell clamp of Purkinje cells demonstrates that ALS IgG increases the amplitude of IBa without modifying their voltage kinetics. This increased IBa could be blocked by a purified nonpeptide toxin from Agelenopsis aperta venom (purified funnel-web spider toxin) or by a synthetic polyamine analog (synthetic funnel-web spider toxin) and by a peptide toxin from the same spider venom, ω-Aga-IVA. Similar results were obtained on single-channel recordings from purified P channel protein. The addition of ALS IgG increased single-channel IBa open time without affecting slope conductance. The results described above were not seen with normal human IgG nor with boiled ALS IgG. It is concluded that ALS IgG enhances inward current through P-type calcium channels. Since P-type Ca2+ channels are present in motoneuron axon terminals, we propose that the enhanced calcium current triggered by ALS IgG may contribute to neuronal damage in ALS.

  11. Cortisone dissociates voltage-dependent K+ channel from its beta subunit

    Science.gov (United States)

    Pan, Yaping; Weng, Jun; Kabaleeswaran, Venkataraman; Li, Huiguang; Cao, Yu; Bhosle, Rahul C.; Zhou, Ming

    2009-01-01

    The Shaker family voltage-dependent potassium channels (Kv1) are expressed in a wide variety of cells and essential for cellular excitability. In humans, loss-of-function mutations of Kv1 channels lead to hyperexcitability and are directly linked to episodic ataxia and atrial fibrillation. All Kv1 channels assemble with beta subunits (Kvβ) and certain Kvβs, for example Kvβ1, have an N-terminal segment that closes a channel by the N-type inactivation mechanism. In principle dissociation of Kvβ1, although never reported, should eliminate inactivation and thus potentiate Kv1 current. We found that cortisone increases mammalian (rat) Kv1 channel activity by binding to Kvβ1. A crystal structure of the Kvβ-cortisone complex was solved to 1.82 Å resolution and revealed novel cortisone binding sites. Further studies demonstrated that cortisone promotes dissociation of Kvβ. The new mode of channel modulation may be explored by native or synthetic ligands to fine tune cellular excitability. PMID:18806782

  12. A calcium-dependent potassium current is increased by a single-gene mutation in Paramecium.

    Science.gov (United States)

    Hennessey, T M; Kung, C

    1987-01-01

    The membrane currents of wild type Paramecium tetraurelia and the behavioral mutant teaA were analyzed under voltage clamp. The teaA mutant was shown to have a greatly increased outward current which was blocked completely by the combined use of internally delivered Cs+ and external TEA+. This, along with previous work (Satow, Y., Kung, C., 1976, J. Exp. Biol. 65:51-63) identified this as a K+ current. It was further found to be a calcium-activated K+ current since this increased outward K+ current cannot be elicited when the internal calcium is buffered with injected EGTA. The mutation pwB, which blocks the inward calcium current, also blocks this increased outward K+ current in teaA. This shows that this mutant current is activated by calcium through the normal depolarization-sensitive calcium channel. While tail current decay kinetic analysis showed that the apparent inactivation rates for this calcium-dependent K+ current are the same for mutant and wild type, the teaA current activates extremely rapidly. It is fully activated within 2 msec. This early activation of such a large outward current causes a characteristic reduction in the amplitude of the action potential of the teaA mutant. The teaA mutation had no effect on any of the other electrophysiological parameters examined. The phenotype of the teaA mutant is therefore a general decrease in responsiveness to depolarizing stimuli because of a rapidly activating calcium-dependent K+ current which prematurely repolarizes the action potential.

  13. Lack of negatively charged residues at the external mouth of Kir2.2 channels enable the voltage-dependent block by external Mg2+.

    Directory of Open Access Journals (Sweden)

    Junwei Li

    Full Text Available Kir channels display voltage-dependent block by cytosolic cations such as Mg2+ and polyamines that causes inward rectification. In fact, cations can regulate K channel activity from both the extracellular and intracellular sides. Previous studies have provided insight into the up-regulation of Kir channel activity by extracellular K+ concentration. In contrast, extracellular Mg2+ has been found to reduce the amplitude of the single-channel current at milimolar concentrations. However, little is known about the molecular mechanism of Kir channel blockade by external Mg2+ and the relationship between the Mg2+ blockade and activity potentiation by permeant K+ ions. In this study, we applied an interactive approach between theory and experiment. Electrophysiological recordings on Kir2.2 and its mutants were performed by heterologous expression in Xenopus laevis oocytes. Our results confirmed that extracellular Mg2+ could reduce heterologously expressed WT Kir2.2 currents in a voltage dependent manner. The kinetics of inhibition and recovery of Mg2+ exhibit a 3∼4s time constant. Molecular dynamics simulation results revealed a Mg2+ binding site located at the extracellular mouth of Kir2.2 that showed voltage-dependent Mg2+ binding. The mutants, G119D, Q126E and H128D, increased the number of permeant K+ ions and reduced the voltage-dependent blockade of Kir2.2 by extracellular Mg2+.

  14. Sodium and calcium currents shape action potentials in immature mouse inner hair cells.

    Science.gov (United States)

    Marcotti, Walter; Johnson, Stuart L; Rusch, Alfons; Kros, Corne J

    2003-11-01

    Before the onset of hearing at postnatal day 12, mouse inner hair cells (IHCs) produce spontaneous and evoked action potentials. These spikes are likely to induce neurotransmitter release onto auditory nerve fibres. Since immature IHCs express both alpha1D (Cav1.3) Ca2+ and Na+ currents that activate near the resting potential, we examined whether these two conductances are involved in shaping the action potentials. Both had extremely rapid activation kinetics, followed by fast and complete voltage-dependent inactivation for the Na+ current, and slower, partially Ca2+-dependent inactivation for the Ca2+ current. Only the Ca2+ current is necessary for spontaneous and induced action potentials, and 29 % of cells lacked a Na+ current. The Na+ current does, however, shorten the time to reach the action-potential threshold, whereas the Ca2+ current is mainly involved, together with the K+ currents, in determining the speed and size of the spikes. Both currents increased in size up to the end of the first postnatal week. After this, the Ca2+ current reduced to about 30 % of its maximum size and persisted in mature IHCs. The Na+ current was downregulated around the onset of hearing, when the spiking is also known to disappear. Although the Na+ current was observed as early as embryonic day 16.5, its role in action-potential generation was only evident from just after birth, when the resting membrane potential became sufficiently negative to remove a sizeable fraction of the inactivation (half inactivation was at -71 mV). The size of both currents was positively correlated with the developmental change in action-potential frequency.

  15. Calcium

    Science.gov (United States)

    ... in luck if you like sardines and canned salmon with bones. Almond milk. previous continue Working Calcium ... drinks, and cereals. Other Considerations for Building Bones Vitamin D is essential for calcium absorption, so it's ...

  16. Novel expression and regulation of voltage-dependent potassium channels in placentas from women with preeclampsia.

    Science.gov (United States)

    Mistry, Hiten D; McCallum, Laura A; Kurlak, Lesia O; Greenwood, Iain A; Broughton Pipkin, Fiona; Tribe, Rachel M

    2011-09-01

    Preeclampsia is associated with structural/functional alterations in placental and maternal vasculature. Voltage-dependant potassium channels encoded by KCNQ1-5 genes have been detected in several types of blood vessels where they promote vascular relaxation. Voltage-dependant potassium channel function can be modulated by KCNE1-5-encoded accessory proteins. The aim of this study was to determine whether KCNQ and KCNE genes are differentially expressed in placentas from women with preeclampsia compared with normotensive controls and to examine any differences in those who delivered preterm (voltage-dependant potassium channels are expressed and markedly modulated in placentas from preeclamptic women. Differential expression of isoforms may lead to altered cell proliferation. The correlation between KCNQ3 and KCNE5 expression is indicative of a novel channel complex and warrants further investigation.

  17. Selective serotonin reuptake inhibitor sertraline inhibits voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells

    Indian Academy of Sciences (India)

    HAN SOL KIM; HONGLIANG LI; HYE WON KIM; SUNG EUN SHIN; IL-WHAN CHOI; AMY L FIRTH; HYOWEON BANG; YOUNG MIN BAE; WON SUN PARK

    2016-12-01

    We examined the effects of the selective serotonin reuptake inhibitor (SSRI) sertraline on voltage-dependent K+ (Kv)channels in freshly isolated rabbit coronary arterial smooth muscle cells using the voltage-clamp technique. Sertralinedecreased the Kv channel current in a dose-dependent manner, with an IC50 value of 0.18 μM and a slope value (Hillcoefficient) of 0.61. Although the application of 1 μM sertraline did not affect the steady-state activation curves,sertraline caused a significant, negative shift in the inactivation curves. Pretreatment with another SSRI, paroxetine,had no significant effect on Kv currents and did not alter the inhibitory effects of sertraline on Kv currents. From theseresults, we concluded that sertraline dose-dependently inhibited Kv currents independently of serotonin reuptakeinhibition by shifting inactivation curves to a more negative potential.

  18. Selective serotonin reuptake inhibitor sertraline inhibits voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells.

    Science.gov (United States)

    Kim, Han Sol; Li, Hongliang; Kim, Hye Won; Shin, Sung Eun; Choi, Il-Whan; Firth, Amy L; Bang, Hyoweon; Bae, Young Min; Park, Won Sun

    2016-12-01

    We examined the effects of the selective serotonin reuptake inhibitor (SSRI) sertraline on voltage-dependent K+ (Kv) channels in freshly isolated rabbit coronary arterial smooth muscle cells using the voltage-clamp technique. Sertraline decreased the Kv channel current in a dose-dependent manner, with an IC50 value of 0.18 mu M and a slope value (Hill coefficient) of 0.61. Although the application of 1 mu M sertraline did not affect the steady-state activation curves, sertraline caused a significant, negative shift in the inactivation curves. Pretreatment with another SSRI, paroxetine, had no significant effect on Kv currents and did not alter the inhibitory effects of sertraline on Kv currents. From these results, we concluded that sertraline dose-dependently inhibited Kv currents independently of serotonin reuptake inhibition by shifting inactivation curves to a more negative potential.

  19. Regulation of calcium currents and secretion by magnesium in crustacean peptidergic neurons.

    Science.gov (United States)

    Richmond, J E; Sher, E; Keller, R; Haylett, B; Reichwein, B; Cooke, I M

    1995-12-01

    The effect of varying the external Mg2+ concentration on Ca2+ currents through voltage-operated Ca2+ channels has been examined with the patch-clamp technique in acutely isolated neuronal somata from the X-organ-sinus gland (XOSG) of the crab, Cardisoma carnifex. Neurons from this neurosecretory system were selected for morphology associated with crustacean hyperglycemic hormone (CHH) content. In parallel, the effects of Mg2+ concentration on K(+)-evoked secretion of CHH from isolated, intact XOSGs have been assayed by ELISA. At physiological Ca2+ levels the high-voltage-activated Ca2+ currents were attenuated with increasing Mg2+ concentration, with 50% inhibition at approximately 75 mM. Mg2+ block was voltage-dependent, relief from block occurring with increasing depolarization. Thus, in 24 mM Mg2+ inhibition of the Ca2+ current was approximately 55% at -10 mV. Secretion of CHH varied almost linearly with the log of Mg2+ concentration; in 2.4 mM Mg2+ it was double that in 24 mM Mg2+ and almost completely inhibited in 100 mM. Thus, Mg2+ produces a parallel inhibition of Ca2+ currents and CHH secretion and may play a role as a physiological modulator of neuronal activity and secretion in the XOSG of these crabs.

  20. 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.

  1. 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.

  2. Whole-cell recordings of calcium and potassium currents in acutely isolated smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Qing Cai; Zhong-Liang Zhu; Xiao-Li Fan

    2006-01-01

    AIM: To record calcium and potassium currents in acutely isolated smooth muscle cells of mesenteric arterial branches in rats.METHODS: Smooth muscle cells were freshly isolated by collagenase digest and mechanical trituration with polished pipettes. Patch clamp technique in whole-cell mode was employed to record calcium and potassium currents.RESULTS: The procedure dissociated smooth muscle cells without impairing the electrophysiological characteristics of the cells. The voltage-gated Ca2+ and potassium currents were successfully recorded using whole-cell patch clamp configuration.CONCLUSION: The method dissociates smooth muscle cells from rat mesenteric arterial branches. Voltage-gated channel currents can be recorded in this preparation.

  3. Effects of calcium on the steady outward currents at the equator of the rat lens.

    Science.gov (United States)

    Parmelee, J T; Robinson, K R; Patterson, J W

    1985-10-01

    The relationships between calcium and the steady outward currents at the equator of the rat lens were studied using the vibrating probe technique. In a calcium-free medium, the current was greatly increased and it returned to its original level when calcium was restored to the medium. The Ca-free effect was not observed in Na-free medium. Iodoacetate (IAA) inhibited the initial current, but a current then returned which is referred to as a secondary current. The secondary current was not observed in a Ca-free medium and, therefore, it is thought to be a calcium-dependent potassium current. These responses are consistent with effects on potassium efflux measured by others and lend support to the interpretation that the outward currents observed at the equator of the rat lens are potassium currents. The currents are partially inhibited but not abolished in Na-free bathing medium. This is consistent with the view that the inward currents at the optical poles may be related to the influx of sodium.

  4. KCNQ1 Channels Voltage Dependence through a Voltage-dependent Binding of the S4-S5 Linker to the Pore Domain*

    OpenAIRE

    2010-01-01

    Voltage-dependent potassium (Kv) channels are tetramers of six transmembrane domain (S1–S6) proteins. Crystallographic data demonstrate that the tetrameric pore (S5–S6) is surrounded by four voltage sensor domains (S1–S4). One key question remains: how do voltage sensors (S4) regulate pore gating? Previous mutagenesis data obtained on the Kv channel KCNQ1 highlighted the critical role of specific residues in both the S4-S5 linker (S4S5L) and S6 C terminus (S6T). From these data, we hypothesiz...

  5. Voltage-dependent modulation of cardiac ryanodine receptors (RyR2 by protamine.

    Directory of Open Access Journals (Sweden)

    Paula L Diaz-Sylvester

    Full Text Available It has been reported that protamine (>10 microg/ml blocks single skeletal RyR1 channels and inhibits RyR1-mediated Ca2+ release from sarcoplasmic reticulum microsomes. We extended these studies to cardiac RyR2 reconstituted into planar lipid bilayers. We found that protamine (0.02-20 microg/ml added to the cytosolic surface of fully activated RyR2 affected channel activity in a voltage-dependent manner. At membrane voltage (V(m; SR lumen-cytosol = 0 mV, protamine induced conductance transitions to several intermediate states (substates as well as full block of RyR2. At V(m>10 mV, the substate with the highest level of conductance was predominant. Increasing V(m from 0 to +80 mV, decreased the number of transitions and residence of the channel in this substate. The drop in current amplitude (full opening to substate had the same magnitude at 0 and +80 mV despite the approximately 3-fold increase in amplitude of the full opening. This is more similar to rectification of channel conductance induced by other polycations than to the action of selective conductance modifiers (ryanoids, imperatoxin. A distinctive effect of protamine (which might be shared with polylysines and histones but not with non-peptidic polycations is the activation of RyR2 in the presence of nanomolar cytosolic Ca2+ and millimolar Mg2+ levels. Our results suggest that RyRs would be subject to dual modulation (activation and block by polycationic domains of neighboring proteins via electrostatic interactions. Understanding these interactions could be important as such anomalies may be associated with the increased RyR2-mediated Ca2+ leak observed in cardiac diseases.

  6. Vestibular integrator neurons have quadratic functions due to voltage dependent conductances.

    Science.gov (United States)

    Magnani, Christophe; Eugène, Daniel; Idoux, Erwin; Moore, Lee E

    2013-12-01

    The nonlinear properties of the dendrites of the prepositus hypoglossi nucleus (PHN) neurons are essential for the operation of the vestibular neural integrator that converts a head velocity signal to one that controls eye position. A novel system of frequency probing, namely quadratic sinusoidal analysis (QSA), was used to decode the intrinsic nonlinear behavior of these neurons under voltage clamp conditions. Voltage clamp currents were measured at harmonic and interactive frequencies using specific nonoverlapping stimulation frequencies. Eigenanalysis of the QSA matrix reduces it to a remarkably compact processing unit, composed of just one or two dominant components (eigenvalues). The QSA matrix of rat PHN neurons provides signatures of the voltage dependent conductances for their particular dendritic and somatic distributions. An important part of the nonlinear response is due to the persistent sodium conductance (gNaP), which is likely to be essential for sustained effects needed for a neural integrator. It was found that responses in the range of 10 mV peak to peak could be well described by quadratic nonlinearities suggesting that effects of higher degree nonlinearities would add only marginal improvement. Therefore, the quadratic response is likely to sufficiently capture most of the nonlinear behavior of neuronal systems except for extremely large synaptic inputs. Thus, neurons have two distinct linear and quadratic functions, which shows that piecewise linear + quadratic analysis is much more complete than just piecewise linear analysis; in addition quadratic analysis can be done at a single holding potential. Furthermore, the nonlinear neuronal responses contain more frequencies over a wider frequency band than the input signal. As a consequence, they convert limited amplitude and bandwidth input signals to wider bandwidth and more complex output responses. Finally, simulations at subthreshold membrane potentials with realistic PHN neuron models

  7. Calcium-dependent potassium current in barnacle photoreceptor

    OpenAIRE

    1981-01-01

    When barnacle lateral eye photoreceptors are depolarized to membrane potentials of 0 to +50 mV in the dark, the plot of outward current through the cell membrane against time has two distinct maxima. The first maximum occurs 5-10 ms after the depolarization began. The current then decays to a minimum at approximately 500 ms after the onset of depolarization, and then increases to a second maximum 4-6 s after the depolarization began. If depolarization is maintained, the current again decays t...

  8. Aging Reduces L-type Calcium Channel Current and the Vasodilatory Response of Small Mesenteric Arteries to Calcium Channel Blockers

    Directory of Open Access Journals (Sweden)

    Sulayma A Albarwani

    2016-05-01

    Full Text Available Calcium channel blockers are widely used to treat cardiovascular disease (CVD including hypertension. As aging is an independent risk factor for CVD, the use of calcium channel blockers increases with increasing age. Hence, this study was designed to evaluate the effect of aging on the sensitivity of small mesenteric arteries to L-type voltage-gated calcium channel (LTCC blockers and also to investigate whether there was a concomitant change in calcium current density. Third order mesenteric arteries from male F344 rats, aged 2.5 - 3 months (young and 22 - 26 months (old were mounted on wire myograph to measure the tension during isometric contraction. Arteries were contracted with 100 mM KCl and were then relaxed in a cumulative concentration-response dependent manner with nifedipine (0.1nM - 10 µM, verapamil (0.1nM-10 µM or diltiazem (0.1nM - 10µM. Relaxation-concentration response curves produced by cumulative concentrations of three different calcium channel blockers (CCBs in arteries of old rats were shifted to the right with statistically significant IC50s. pEC50 ± s.e.m: (8.37 ± 0.06 vs 8.04 ± 0.05 , 7.40 ± 0.07 vs 6.81 ± 0.04 and 6.58 ± 0.07 vs 6.34 ± 0.06 in young vs old. It was observed that the maximal contractions induced by 100 mM KCl, phenylephrine and reversed by sodium nitroprusside were not different between young and old groups. However, Bay K 8644 increased resting tension by 23±4.8% in young arteries and 4.7±1.6% in old arteries. LTCC current density were also significantly lower in old arteries (-2.77 ± 0.45 pA/pF compared to young arteries (-4.5 ± 0.40 pA/pF; with similar steady-state activation and inactivation curves. Parallel to this reduction, the expression of Cav1.2 protein was reduced by 57 ± 5% in arteries from old rats compared to those from young rats. In conclusion, our results suggest that aging reduces the response of small mesenteric arteries to the vasodilatory effect of the CCBs and this may

  9. The impact of calcium current reversal on neurotransmitter release in the electrically stimulated retina

    Science.gov (United States)

    Werginz, Paul; Rattay, Frank

    2016-08-01

    Objective. In spite of intense theoretical and experimental investigations on electrical nerve stimulation, the influence of reversed ion currents on network activity during extracellular stimulation has not been investigated so far. Approach. Here, the impact of calcium current reversal on neurotransmitter release during subretinal stimulation was analyzed with a computational multi-compartment model of a retinal bipolar cell (BC) that was coupled with a four-pool model for the exocytosis from its ribbon synapses. Emphasis was laid on calcium channel dynamics and how these channels influence synaptic release. Main results. Stronger stimulation with anodic pulses caused transmembrane voltages above the Nernst potential of calcium in the terminals and, by this means, forced calcium ions to flow in the reversed direction from inside to the outside of the cell. Consequently, intracellular calcium concentration decreased resulting in a reduced vesicle release or preventing release at all. This mechanism is expected to lead to a pronounced ring-shaped pattern of exocytosis within a group of neighbored BCs when the stronger stimulated cells close to the electrode fail in releasing vesicles. Significance. Stronger subretinal stimulation causes failure of synaptic exocytosis due to reversal of calcium flow into the extracellular space in cells close to the electrode.

  10. Potassium currents in isolated CA1 neurons of the rat after kindling epileptogenesis.

    Science.gov (United States)

    Vreugdenhil, M; Wadman, W J

    1995-06-01

    Daily tetanic stimulation of the Schaffer collaterals generates an epileptogenic focus in area CA1 of the rat hippocampus, ultimately leading to generalized tonic-clonic convulsions (kindling). Potassium currents were measured under voltage-clamp conditions in pyramidal neurons, acutely dissociated from the focus of fully kindled rats, one day and six weeks after the last generalized seizure. Their amplitude, kinetics, voltage dependence and calcium dependence were compared with controls. With Ca2+ influx blocked by 0.5 mM Ni2+, the sustained current (delayed rectifier) and the transient current (A-current) were not different after kindling. Calcium influx evoked an additional fast transient current component. This transient calcium-dependent current component was increased by 154%, but only immediately after the seizure. A second, slow calcium-dependent potassium current component was dependent on the intracellular calcium level, set by the pipette as well as on calcium influx. The peak amplitude of this slow calcium-dependent current was under optimal calcium conditions not different after kindling, but we found indications that either calcium homeostasis or the calcium sensitivity of the potassium channels was affected by the kindling process. In contrast to the previously described enhancement of calcium current, kindling epileptogenesis did not change the total potassium current amplitude. The minor changes that were observed can be related either to changes in calcium current or to changes in intracellular calcium homeostasis.

  11. Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density.

    Science.gov (United States)

    Perissinotti, Paula P; Ethington, Elizabeth G; Cribbs, Leanne; Koob, Michael D; Martin, Jody; Piedras-Rentería, Erika S

    2014-05-01

    The actin-binding protein Kelch-like 1 (KLHL1) can modulate voltage-gated calcium channels in vitro. KLHL1 interacts with actin and with the pore-forming subunits of Cav2.1 and CaV3.2 calcium channels, resulting in up-regulation of P/Q and T-type current density. Here we tested whether endogenous KLHL1 modulates voltage gated calcium currents in cultured hippocampal neurons by down-regulating the expression of KLHL1 via adenoviral delivery of shRNA targeted against KLHL1 (shKLHL1). Control adenoviruses did not affect any of the neuronal properties measured, yet down-regulation of KLHL1 resulted in HVA current densities ~68% smaller and LVA current densities 44% smaller than uninfected controls, with a concomitant reduction in α(1A) and α(1H) protein levels. Biophysical analysis and western blot experiments suggest Ca(V)3.1 and 3.3 currents are also present in shKLHL1-infected neurons. Synapsin I levels, miniature postsynaptic current frequency, and excitatory and inhibitory synapse number were reduced in KLHL1 knockdown. This study corroborates the physiological role of KLHL1 as a calcium channel modulator and demonstrates a novel, presynaptic role.

  12. Calcium current homeostasis and synaptic deficits in hippocampal neurons from Kelch-like 1 knockout mice

    Directory of Open Access Journals (Sweden)

    Paula Patricia Perissinotti

    2015-01-01

    Full Text Available Kelch-like 1 (KLHL1 is a neuronal actin-binding protein that modulates voltage-gated CaV2.1 (P/Q-type and CaV3.2 (α1H T-type calcium channels; KLHL1 knockdown experiments (KD cause down-regulation of both channel types and altered synaptic properties in cultured rat hippocampal neurons (Perissinotti et al., 2014. Here, we studied the effect of ablation of KLHL1 on calcium channel function and synaptic properties in cultured hippocampal neurons from KLHL1 knockout (KO mice. Western blot data showed the P/Q-type channel α1A subunit was less abundant in KO hippocampus compared to wildtype (WT; and PQ-type calcium currents were smaller in KO neurons than WT during early days in vitro, although this decrease was compensated for at late stages by increases in L-type calcium current. In contrast, T-type currents did not change in culture. However, biophysical properties and western blot analysis revealed a differential contribution of T-type channel isoforms in the KO, with CaV3.2 α1H subunit being down-regulated and CaV3.1 α1G up-regulated. Synapsin I levels were reduced in the KO hippocampus; cultured neurons displayed a concomitant reduction in synapsin I puncta and decreased miniature excitatory postsynaptic current (mEPSC frequency. In summary, genetic ablation of the calcium channel modulator resulted in compensatory mechanisms to maintain calcium current homeostasis in hippocampal KO neurons; however, synaptic alterations resulted in a reduction of excitatory synapse number, causing an imbalance of the excitatory-inhibitory synaptic input ratio favoring inhibition.

  13. Calcium Transient and Sodium-Calcium Exchange Current in Human versus Rabbit Sinoatrial Node Pacemaker Cells

    OpenAIRE

    Verkerk, Arie O.; Marcel M. G. J. van Borren; Ronald Wilders

    2013-01-01

    There is an ongoing debate on the mechanism underlying the pacemaker activity of sinoatrial node (SAN) cells, focusing on the relative importance of the “membrane clock” and the “Ca2+ clock” in the generation of the small net membrane current that depolarizes the cell towards the action potential threshold. Specifically, the debate centers around the question whether the membrane clock-driven hyperpolarization-activated current, I f , which is also known as the “funny current” or “pacemaker c...

  14. Zn2+ regulates Kv2.1 voltage-dependent gating and localization following ischemia

    OpenAIRE

    Aras, Mandar A.; Saadi, Robert A.; Aizenman, Elias

    2009-01-01

    The delayed-rectifier K+ channel Kv2.1 exists in highly phosphorylated somatodendritic clusters. Ischemia induces rapid Kv2.1 dephosphorylation and a dispersal of these clusters, accompanied by a hyperpolarizing shift in their voltage-dependent activation kinetics. Transient modulation of Kv2.1 activity and localization following ischemia is dependent on a rise in intracellular Ca2+and the protein phosphatase calcineurin. Here, we show that neuronal free Zn2+also plays a critical role in the ...

  15. A highly calcium-selective cation current activated by intracellular calcium release in MDCK cells.

    Science.gov (United States)

    Delles, C; Haller, T; Dietl, P

    1995-08-01

    1. The whole-cell patch clamp technique and fluorescence microscopy with the Ca2+ indicators fura-2 and fluo-3 were used to measure the whole-cell current and the free intracellular Ca2+ concentration ([Ca2+]i) in Madin-Darby canine kidney (MDCK) cells. 2. In a Ca(2+)-free bath solution, thapsigargin (TG) caused a transient increase of [Ca2+]i. Subsequent addition of Ca2+ caused a long lasting elevation of [Ca2+]i. 3. In a Ca(2+)-free bath solution, extracellular application of TG, ATP or ionomycin, or intracellular application of inositol 1,4,5-trisphosphate (IP3), caused a small but significant inward current (Iin) and a transient outward Ca(2+)-dependent K+ current (IK(Ca)), consistent with intracellular Ca2+ release. Subsequent addition of Ca2+ induced a prominent Iin with a current density of -4.2 +/- 0.7 pA pF-1. This Iin was unaffected by inositol 1,3,4,5-tetrakisphosphate (IP4). 4. Na+ replacement by mannitol, N-methyl-D-glucamine+ (NMG+), aminomethylidin-trimethanol+ (Tris+) or choline+ reduced Iin by 54, 65, 52 and 56%, respectively. This indicates an apparent Ca2+ selectivity over Na+ of 26:1. Iin was, however, unaffected by replacing Cl- with gluconate- or by the K+ channel blocker charybdotoxin (CTX). 5. Iin was completely blocked by La3+ (IC50 = 0.77 microM). Consistently, La3+ completely reversed the TG-induced elevation of [Ca2+]i. SK&F 96365 (1-[3-(4-methoxyphenyl)-propoxyl]-1-(4-methoxy-phenyl)-ethyl-1H-im idazole) HCl did not inhibit the TG-induced Iin. It did, however, exhibit a biphasic effect on [Ca2+]i, consisting of an initial Ca2+ decay and a subsequent Ca2+ elevation. La3+ completely reversed the SK&F 96365-induced elevation of [Ca2+]i. 6. In the absence of Na+, Iin was dependent on the bath Ca2+ concentration (EC50 = 1.02 mM). Ca2+ replacement by Ba2+ or Mn2+ resulted in a reduction of Iin by 95 and 94%, respectively. 7. From these experiments we conclude that Ca2+ release from intracellular Ca2+ stores, induced by different independent

  16. Endocytic regulation of voltage-dependent potassium channels in the heart.

    Science.gov (United States)

    Ishii, Kuniaki; Norota, Ikuo; Obara, Yutaro

    2012-01-01

    Understanding the regulation of cardiac ion channels is critical for the prevention of arrhythmia caused by abnormal excitability. Ion channels can be regulated by a change in function (qualitative) and a change in number (quantitative). Functional changes have been extensively investigated for many ion channels including cardiac voltage-dependent potassium channels. By contrast, the regulation of ion channel numbers has not been widely examined, particularly with respect to acute modulation of ion channels. This article briefly summarizes stimulus-induced endocytic regulation of major voltage-dependent potassium channels in the heart. The stimuli known to cause their endocytosis include receptor activation, drugs, and low extracellular [K(+)], following which the potassium channels undergo either clathrin-mediated or caveolin-mediated endocytosis. Receptor-mediated endocytic regulation has been demonstrated for Kv1.2, Kv1.5, KCNQ1 (Kv7.1), and Kv4.3, while drug-induced endocytosis has been demonstrated for Kv1.5 and hERG. Low [K(+)](o)-induced endocytosis might be unique for hERG channels, whose electrophysiological characteristics are known to be under strong influence of [K(+)](o). Although the precise mechanisms have not been elucidated, it is obvious that major cardiac voltage-dependent potassium channels are modulated by endocytosis, which leads to changes in cardiac excitability.

  17. Aspects of calcium-activated chloride currents: a neuronal perspective.

    Science.gov (United States)

    Scott, R H; Sutton, K G; Griffin, A; Stapleton, S R; Currie, K P

    1995-06-01

    Ca(2+)-activated Cl- channels are expressed in a variety of cell types, including central and peripheral neurones. These channels are activated by a rise in intracellular Ca2+ close to the cell membrane. This can be evoked by cellular events such as Ca2+ entry through voltage- and ligandgated channels or release of Ca2+ from intracellular stores. Additionally, these Ca(2+)-activated Cl currents (ICl(Ca)) can be activated by raising intracellular Ca2+ through artificial experimental procedures such as intracellular photorelease of Ca2+ from "caged" photolabile compounds (e.g. DM-nitrophen) or by treating cells with Ca2+ ionophores. The potential changes that result from activation of Ca(2+)-activated Cl- channels are dependent on resting membrane potential and the equilibrium potential for Cl-. Ca2+ entry during a single action potential is sufficient to produce substantial after potentials, suggesting that the activity of these Cl- channels can have profound effects on cell excitability. The whole cell ICl(Ca) can be identified by sensitivity to increased Ca2+ buffering capacity of the cell, anion substitution studies and reversal potential measurements, as well as by the actions of Cl- channel blockers. In cultured sensory neurones, there is evidence that the ICl(Ca) deactivates as Ca2+ is buffered or removed from the intracellular environment. To date, there is no evidence in mammalian neurones to suggest these Ca(2+)-sensitive Cl- channels undergo a process of inactivation. Therefore, ICl(Ca) can be used as a physiological index of intracellular Ca2+ close to the cell membrane. The ICl(Ca) has been shown to be activated or prolonged as a result of metabolic stress, as well as by drugs that disturb intracellular Ca2+ homeostatic mechanisms or release Ca2+ from intracellular stores. In addition to sensitivity to classic Cl- channel blockers such as niflumic acid, derivatives of stilbene (4,4'diisothiocyanostilbene-2,2'-disulphonic acid, 4-acetamido-4

  18. Voltage dependence of Hodgkin-Huxley rate functions for a multistage K+ channel voltage sensor within a membrane

    Science.gov (United States)

    Vaccaro, S. R.

    2014-11-01

    The activation of a K+channel sensor in two sequential stages during a voltage clamp may be described as the translocation of a Brownian particle in an energy landscape with two large barriers between states. A solution of the Smoluchowski equation for a square-well approximation to the potential function of the S4 voltage sensor satisfies a master equation and has two frequencies that may be determined from the forward and backward rate functions. When the higher-frequency terms have small amplitude, the solution reduces to the relaxation of a rate equation, where the derived two-state rate functions are dependent on the relative magnitude of the forward rates (α and γ ) and the backward rates (β and δ ) for each stage. In particular, the voltage dependence of the Hodgkin-Huxley rate functions for a K+channel may be derived by assuming that the rate functions of the first stage are large relative to those of the second stage—α ≫γ and β ≫δ . For a Shaker IR K+ channel, the first forward and backward transitions are rate limiting (α <γ and δ ≪β ), and for an activation process with either two or three stages, the derived two-state rate functions also have a voltage dependence that is of a similar form to that determined for the squid axon. The potential variation generated by the interaction between a two-stage K+ ion channel and a noninactivating Na+ ion channel is determined by the master equation for K+channel activation and the ionic current equation when the Na+channel activation time is small, and if β ≪δ and α ≪γ , the system may exhibit a small amplitude oscillation between spikes, or mixed-mode oscillation, in which the slow closed state modulates the K+ ion channel conductance in the membrane.

  19. trans-Caryophyllene, a Natural Sesquiterpene, Causes Tracheal Smooth Muscle Relaxation through Blockade of Voltage-Dependent Ca2+ Channels

    Directory of Open Access Journals (Sweden)

    Jader Santos Cruz

    2012-10-01

    Full Text Available trans-Caryophyllene is a major component in the essential oils of various species of medicinal plants used in popular medicine in Brazil. It belongs to the chemical class of the sesquiterpenes and has been the subject of a number of studies. Here, we evaluated the effects of this compound in airway smooth muscle. The biological activities of trans-caryophyllene were examined in isolated bath organs to investigate the effect in basal tonus. Electromechanical and pharmacomechanical couplings were evaluated through the responses to K+ depolarization and exposure to acetylcholine (ACh, respectively. Isolated cells of rat tracheal smooth muscle were used to investigate trans-caryophyllene effects on voltage-dependent Ca2+ channels by using the whole-cell voltage-clamp configuration of the patch-clamp technique. trans-Caryophyllene showed more efficiency in the blockade of electromechanical excitation-contraction coupling while it has only minor inhibitory effect on pharmacomechanical coupling. Epithelium removal does not modify tracheal smooth muscle response elicited by trans-caryophyllene in the pharmacomechanical coupling. Under Ca2+-free conditions, pre-exposure to trans-caryophyllene did not reduce the contraction induced by ACh in isolated rat tracheal smooth muscle, regardless of the presence of intact epithelium. In the whole-cell configuration, trans-caryophyllene (3 mM, inhibited the inward Ba2+ current (IBa to approximately 50% of control levels. Altogether, our results demonstrate that trans-caryophyllene has anti-spasmodic activity on rat tracheal smooth muscle which could be explained, at least in part, by the voltage-dependent Ca2+ channels blockade.

  20. Aging Reduces L-Type Calcium Channel Current and the Vasodilatory Response of Small Mesenteric Arteries to Calcium Channel Blockers

    Science.gov (United States)

    Albarwani, Sulayma A.; Mansour, Fathi; Khan, Abdul Aleem; Al-Lawati, Intisar; Al-Kaabi, Abdulla; Al-Busaidi, Al-Manar; Al-Hadhrami, Safa; Al-Husseini, Isehaq; Al-Siyabi, Sultan; Tanira, Musbah O.

    2016-01-01

    Calcium channel blockers (CCBs) are widely used to treat cardiovascular disease (CVD) including hypertension. As aging is an independent risk factor for CVD, the use of CCBs increases with increasing age. Hence, this study was designed to evaluate the effect of aging on the sensitivity of small mesenteric arteries to L-type voltage-gated calcium channel (LTCC) blockers and also to investigate whether there was a concomitant change in calcium current density. Third order mesenteric arteries from male F344 rats, aged 2.5–3 months (young) and 22–26 months (old) were mounted on wire myograph to measure the tension during isometric contraction. Arteries were contracted with 100 mM KCl and were then relaxed in a cumulative concentration-response dependent manner with nifedipine (0.1 nM–1 μM), verapamil (0.1 nM–10 μM), or diltiazem (0.1 nM–10 μM). Relaxation-concentration response curves produced by cumulative concentrations of three different CCBs in arteries of old rats were shifted to the right with statistically significant IC50s. pIC50 ± s.e.m: (8.37 ± 0.06 vs. 8.04 ± 0.05, 7.40 ± 0.07 vs. 6.81 ± 0.04, and 6.58 ± 0.07 vs. 6.34 ± 0.06) in young vs. old. It was observed that the maximal contractions induced by phenylephrine and reversed by sodium nitroprusside were not different between young and old groups. However, Bay K 8644 (1 μM) increased resting tension by 23 ± 4.8% in young arteries and 4.7 ± 1.6% in old arteries. LTCC current density were also significantly lower in old arteries (−2.77 ± 0.45 pA/pF) compared to young arteries (−4.5 ± 0.40 pA/pF); with similar steady-state activation and inactivation curves. Parallel to this reduction, the expression of Cav1.2 protein was reduced by 57 ± 5% in arteries from old rats compared to those from young rats. In conclusion, our results suggest that aging reduces the response of small mesenteric arteries to the vasodilatory effect of the CCBs and this may be due to, at least in part, reduced

  1. Effect of Shenmai Injection on L-type Calcium Current of Diaphragmatic Muscle in Rats

    Institute of Scientific and Technical Information of China (English)

    赵丽敏; 熊盛道; 牛汝楫; 徐永健; 张珍祥

    2004-01-01

    In this study, whole cell patch clamp recording technique was employed to investigate the effect of Shenmai Injection (SMI) on L-type calcium current of diaphragmatic muscle in rats. The result showed that when the diaphragmatic muscle cell was held at -80 mV and depolarized to +60 mV, 10 μl/ml, 50 μl/ml and 100μl/ml SMI enhanced the inner peak L-type calcium current from -(6.8±0.7) pA/pF (n=7) to -(7.3±0.8) pA/pF (P>0.05, n=7), -(8.6±1.0) pA/pF (P<0.05, n=7) and -(9.4±1.2) pA/pF (P<0.05, n=7), respectively. The rates of L-type calcium current were increased by (7. 34±2.37) %, (25. 72±5.94)% , and (38. 16±7.33)% ,respectively. However, it had no significant effect on maximal activation potential and reversal potential. Our results suggested that SMI could activate the calcium channel of the diaphragmatic fibers of the rats, increase the influx of Ca2+ , and enhance the contractility of diaphragmatic muscles.

  2. KCNQ1 channels voltage dependence through a voltage-dependent binding of the S4-S5 linker to the pore domain.

    Science.gov (United States)

    Choveau, Frank S; Rodriguez, Nicolas; Abderemane Ali, Fayal; Labro, Alain J; Rose, Thierry; Dahimène, Shehrazade; Boudin, Hélène; Le Hénaff, Carole; Escande, Denis; Snyders, Dirk J; Charpentier, Flavien; Mérot, Jean; Baró, Isabelle; Loussouarn, Gildas

    2011-01-07

    Voltage-dependent potassium (Kv) channels are tetramers of six transmembrane domain (S1-S6) proteins. Crystallographic data demonstrate that the tetrameric pore (S5-S6) is surrounded by four voltage sensor domains (S1-S4). One key question remains: how do voltage sensors (S4) regulate pore gating? Previous mutagenesis data obtained on the Kv channel KCNQ1 highlighted the critical role of specific residues in both the S4-S5 linker (S4S5(L)) and S6 C terminus (S6(T)). From these data, we hypothesized that S4S5(L) behaves like a ligand specifically interacting with S6(T) and stabilizing the closed state. To test this hypothesis, we designed plasmid-encoded peptides corresponding to portions of S4S5(L) and S6(T) of the voltage-gated potassium channel KCNQ1 and evaluated their effects on the channel activity in the presence and absence of the ancillary subunit KCNE1. We showed that S4S5(L) peptides inhibit KCNQ1, in a reversible and state-dependent manner. S4S5(L) peptides also inhibited a voltage-independent KCNQ1 mutant. This inhibition was competitively prevented by a peptide mimicking S6(T), consistent with S4S5(L) binding to S6(T). Val(254) in S4S5(L) is known to contact Leu(353) in S6(T) when the channel is closed, and mutations of these residues alter the coupling between the two regions. The same mutations introduced in peptides altered their effects, further confirming S4S5(L) binding to S6(T). Our results suggest a mechanistic model in which S4S5(L) acts as a voltage-dependent ligand bound to its receptor on S6 at rest. This interaction locks the channel in a closed state. Upon plasma membrane depolarization, S4 pulls S4S5(L) away from S6(T), allowing channel opening.

  3. Purification and Characterization of Two Voltage-Dependent Anion Channel Isoforms from Plant Seeds1

    Science.gov (United States)

    Abrecht, Helge; Wattiez, Ruddy; Ruysschaert, Jean-Marie; Homblé, Fabrice

    2000-01-01

    Mitochondria were isolated from imbibed seeds of lentil (Lens culinaris) and Phaseolus vulgaris. We copurified two voltage-dependent anion channel from detergent solubilized mitochondria in a single purification step using hydroxyapatite. The two isoforms from P. vulgaris were separated by chromatofocusing chromatography in 4 m urea without any loss of channel activity. Channel activity of each isoform was characterized upon reconstitution into diphytanoyl phosphatidylcholine planar lipid bilayers. Both isoforms form large conductance channels that are slightly anion selective and display cation selective substates. PMID:11080295

  4. Admittance Spectroscopy in CZTSSe: Metastability Behavior and Voltage Dependent Defect Study

    Energy Technology Data Exchange (ETDEWEB)

    Koeper, Mark J.; Hages, Charles J.; Li, Jian V.; Levi, Dean; Agrawal, Rakesh

    2016-11-21

    Admittance spectroscopy has been performed on a CZTSSe device with a carrier injection pretreatment and under electronically relaxed conditions to demonstrate metastability behavior. We show that the measurements with the carrier injection pretreatment demonstrate two admittance signatures while the relaxed measurement demonstrates only one admittance signature with a different activation energy. Additionally, voltage dependent admittance spectroscopy was performed using the carrier injection pretreatment method at each of the applied voltage bias. The activation energies of the two admittance signatures were calculated and are shown to be independent of the voltage bias.

  5. Effects of Arecoline on Calcium Channel Currents and Caffeine-induced Calcium Release in Isolated Single Ventricular Myocyte of Guinea Pig

    Institute of Scientific and Technical Information of China (English)

    林先明; 李真; 胡本容; 夏国瑾; 姚伟星; 向继洲

    2002-01-01

    Summary: The effects of Arecoline (Are) on calcium mobilization were investigated. In isolatedsingle ventricular myocyte of guinea pig, patch clamp whole cell recording techniques were used torecord the current of L-type calcium channel and cytosolic Ca2+ level ([Ca2+]i) labeled with fluo-rescence probe Fluo-3/AM was measured under a laser scanning confocal microscope. Results re-vealed that Are (3-100 μmol/L) could inhibit L-type calcium current in a concentration-depen-dent manner and the value of IC50 was 33. 73μmol/L (n= 5). In the absence of extracellular calci-um, the resting levels of [Ca2+]i was not affected by Are (n=6, P>0. 05), but pretreatmentwith Are (30 μmol/L) could significantly inhibit the [Ca2+]i elevation induced by caffeine (10mmol/L, n = 6, P < 0. 01). It was concluded that Are could inhibit not only calcium influxthrough L-type calcium channel but also calcium release from sarcoplasmic reticulum.

  6. Role of calcium mobilization in the regulation of spontaneous transient outward currents in porcine coronary artery myocytes

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The purpose of the present study was to further study the characteristics and regulation of spontaneous transient outward currents (STOCs) in freshly isolated porcine coronary artery smooth muscle cells (ASMCs). STOCs were recorded using the perforated whole-cell patch-clamp configuration. STOCs were voltage-dependent and superimposed stochastically onto whole-cell Ca2+-activated-K+ (BKCa) currents. Charybdotoxin (ChTX, 200 nmol/L), a selective blocker of BKCa channels, completely inhibited STOCs within 10 min. STOCs activity was greatly suppressed when extracellular Ca2+ concentration decreased from 1.8 mmol/L to 200 nmol/L, further removal of Ca2+ abolished STOCs activity. Ca2+ ionophore A23187 (10 μmol/L) increased STOCs activity significantly. Verapamil (20 μmol/L) and CdCl2 (200 μmol/L), two kinds of organic L-type voltage-dependent Ca2+ channels (L-VDCCs) antagonists, had little effect on STOCs. In addition, the ryanodine receptors (RyRs) agonist caffeine (5 mmol/L) significantly activated STOCs. Application of ryanodine (50 μmol/L) to block RyRs abolished STOCs, subsequent washout of ryanodine or application of caffeine failed to reproduce STOCs activity. Inhibition of inositol 1,4,5-trisphosphate receptors (IP3Rs) by 2APB (40 μmol/L) greatly suppressed the activity of STOCs, application of caffeine (5 mmol/L) in the presence of 2APB caused a burst of outward currents followed by inhibition of STOCs. These results suggest that STOCs in porcine coronary ASMCs are mediated by BKCa channels. Extracellular Ca2+ is essential for STOCs activity, while Ca2+ entry through L-VDCCs has little effect on STOCs. Intracellular Ca2+ release induced by RyRs is responsible for the regulation of STOCs, whereas IP3Rs might also be involved.

  7. Inhibition of rat hippocampal excitability by the plant alkaloid 3-acetylaconitine mediated by interaction with voltage-dependent sodium channels.

    Science.gov (United States)

    Ameri, A

    1997-02-01

    The effects of the Aconitum alkaloid 3-acetylaconitine on neuronal activity were investigated in the slice preparation and on cultivated neurons of rat hippocampus by extracellular and patch-clamp recordings, respectively. 3-Acetylaconitine (0.01-1 microM) diminished the orthodromic and antidromic population spike in a concentration-dependent manner. The inhibitory action of the drug was preceded by a transiently enhanced excitability. The latency of onset of the inhibition was accelerated by increased stimulation frequency, whereas recovery during washout of the alkaloid was accelerated by decreased stimulation frequency. Moreover, the inhibitory effect of 3-acetylaconitine was evaluated in two different models of epileptiform activity induced either by blockade of GABA receptors by bicuculline (10 microM) or by a nominal Mg(2+)-free bathing medium. In accordance with the activity-dependent mode of action, this compound abolished the synaptically evoked population spikes in the presence of bicuculline or nominal Mg(2+)-free bathing medium, respectively. Whole-cell patch-clamp recordings revealed an interaction of 3-acetylaconitine with the voltage-dependent sodium channel. At a concentration of 1 microM, 3-acetylaconitine did not affect the peak amplitude of the sodium current, but shifted the current-voltage relationship in the hyperpolarized direction such that sodium currents were already activated at the resting potential.

  8. Voltage dependence of rate functions for Na+ channel inactivation within a membrane

    CERN Document Server

    Vaccaro, Samuel R

    2015-01-01

    The inactivation of a Na+ channel occurs when the activation of the charged S4 segment of domain IV, with rate functions $\\alpha_{i}$ and $\\beta_{i}$, is followed by the binding of an intracellular hydrophobic motif which blocks conduction through the ion pore, with rate functions $\\gamma_{i}$ and $\\delta_{i}$. During a voltage clamp of the Na+ channel, the solution of the master equation for inactivation reduces to the relaxation of a rate equation when the binding of the inactivation motif is rate limiting ($\\alpha_{i} \\gg \\gamma_{i}$ and $\\beta_{i} \\gg \\delta_{i}$). The voltage dependence of the derived forward rate function for Na+ channel inactivation has an exponential dependence on the membrane potential for small depolarizations and approaches a constant value for larger depolarizations, whereas the voltage dependence of the backward rate function is exponential, and each rate has a similar form to the Hodgkin-Huxley empirical rate functions for Na+ channel inactivation in the squid axon.

  9. Accumulation of cytoplasmic calcium, but not apamin-sensitive afterhyperpolarization current, during high frequency firing in rat subthalamic nucleus cells.

    Science.gov (United States)

    Teagarden, Mark; Atherton, Jeremy F; Bevan, Mark D; Wilson, Charles J

    2008-02-01

    The autonomous firing pattern of neurons in the rat subthalamic nucleus (STN) is shaped by action potential afterhyperpolarization currents. One of these is an apamin-sensitive calcium-dependent potassium current (SK). The duration of SK current is usually considered to be limited by the clearance of calcium from the vicinity of the channel. When the cell is driven to fire faster, calcium is expected to accumulate, and this is expected to result in accumulation of calcium-dependent AHP current. We measured the time course of calcium transients in the soma and proximal dendrites of STN neurons during spontaneous firing and their accumulation during driven firing. We compared these to the time course and accumulation of AHP currents using whole-cell and perforated patch recordings. During spontaneous firing, a rise in free cytoplasmic calcium was seen after each action potential, and decayed with a time constant of about 200 ms in the soma, and 80 ms in the dendrites. At rates higher than 10 Hz, calcium transients accumulated as predicted. In addition, there was a slow calcium transient not predicted by summation of action potentials that became more pronounced at high firing frequency. Spike AHP currents were measured in voltage clamp as tail currents after 2 ms voltage pulses that triggered action currents. Apamin-sensitive AHP (SK) current was measured by subtraction of tail currents obtained before and after treatment with apamin. SK current peaked between 10 and 15 ms after an action potential, had a decay time constant of about 30 ms, and showed no accumulation. At frequencies between 5 and 200 spikes s(-1), the maximal SK current remained the same as that evoked by a single action potential. AHP current did not have time to decay between action potentials, so at frequencies above 50 spikes s(-1) the apamin-sensitive current was effectively constant. These results are inconsistent with the view that the decay of SK current is governed by calcium dynamics. They

  10. Osteoclast spreading kinetics are correlated with an oscillatory activation of a calcium-dependent potassium current.

    Science.gov (United States)

    Espinosa, Leon; Paret, Laurent; Ojeda, Carlos; Tourneur, Yves; Delmas, Pierre D; Chenu, Chantal

    2002-10-01

    Cell movement and spreading involve calcium-dependent processes and ionic channel activation. During bone resorption, osteoclasts alternate between spread, motile and resorptive phases. We investigated whether the electrical membrane properties of osteoclasts were linked to their membrane morphological changes. Rabbit osteoclasts were recorded by time-lapse videomicroscopy performed simultaneously with patch-clamp whole cell and single channel recordings. Original image analysis methods were developed and used to demonstrate for the first time an oscillatory activation of a spontaneous membrane current in osteoclasts, which is directly correlated to the membrane movement rate. This current was identified as a calcium-dependent potassium current (IK(Ca)) that is sensitive to both charybdotoxin and apamin and was generated by a channel with unitary conductance of approximately 25+/-2 pS. Blockade of this current also decreased osteoclast spreading and inhibited bone resorption in vitro, demonstrating a physiological role for this current in osteoclast activity. These results establish for the first time a temporal correlation between lamellipodia formation kinetics and spontaneous peaks of IK(Ca), which are both involved in the control of osteoclast spreading and bone resorption.

  11. Properties of a calcium-activated K(+) current on interneurons in the developing rat hippocampus.

    Science.gov (United States)

    Aoki, T; Baraban, S C

    2000-06-01

    Calcium-activated potassium currents have an essential role in regulating excitability in a variety of neurons. Although it is well established that mature CA1 pyramidal neurons possess a Ca(2+)-activated K(+) conductance (I(K(Ca))) with early and late components, modulation by various endogenous neurotransmitters, and sensitivity to K(+) channel toxins, the properties of I(K(Ca)) on hippocampal interneurons (or immature CA1 pyramidal neurons) are relatively unknown. To address this problem, whole-cell voltage-clamp recordings were made from visually identified interneurons in stratum lacunosum-moleculare (L-M) and CA1 pyramidal cells in hippocampal slices from immature rats (P3-P25). A biphasic calcium-activated K(+) tail current was elicited following a brief depolarization from the holding potential (-50 mV). Analysis of the kinetic properties of I(K(Ca)) suggests that an early current component differs between these two cell types. An early I(K(Ca)) with a large peak current amplitude (200.8 +/- 13.2 pA, mean +/- SE), slow time constant of decay (70.9 +/- 3.3 ms), and relatively rapid time to peak (within 15 ms) was observed on L-M interneurons (n = 88), whereas an early I(K(Ca)) with a small peak current amplitude (112.5 +/- 7.3 pA), a fast time constant of decay (39.4 +/- 1.6 ms), and a slower time-to-peak (within 26 ms) was observed on CA1 pyramidal neurons (n = 85). Removal of extracellular calcium or addition of inorganic Ca(2+) channel blockers (cadmium, nickel, or cobalt) was used to demonstrate the calcium dependence of these currents. Addition of norepinephrine, carbachol, and a variety of channel toxins (apamin, iberiotoxin, verruculogen, paxilline, penitrem A, and charybdotoxin) were used to further distinguish between I(K(Ca)) on these two hippocampal cell types. Verruculogen (100 nM), carbachol (100 microM), apamin (100 nM), TEA (1 mM), and iberiotoxin (50 nM) significantly reduced early I(K(Ca)) on CA1 pyramidal neurons; early I(K(Ca)) on L

  12. A vesicle-trafficking protein commandeers Kv channel voltage sensors for voltage-dependent secretion.

    Science.gov (United States)

    Grefen, Christopher; Karnik, Rucha; Larson, Emily; Lefoulon, Cécile; Wang, Yizhou; Waghmare, Sakharam; Zhang, Ben; Hills, Adrian; Blatt, Michael R

    2015-01-01

    Growth in plants depends on ion transport for osmotic solute uptake and secretory membrane trafficking to deliver material for wall remodelling and cell expansion. The coordination of these processes lies at the heart of the question, unresolved for more than a century, of how plants regulate cell volume and turgor. Here we report that the SNARE protein SYP121 (SYR1/PEN1), which mediates vesicle fusion at the Arabidopsis plasma membrane, binds the voltage sensor domains (VSDs) of K(+) channels to confer a voltage dependence on secretory traffic in parallel with K(+) uptake. VSD binding enhances secretion in vivo subject to voltage, and mutations affecting VSD conformation alter binding and secretion in parallel with channel gating, net K(+) concentration, osmotic content and growth. These results demonstrate a new and unexpected mechanism for secretory control, in which a subset of plant SNAREs commandeer K(+) channel VSDs to coordinate membrane trafficking with K(+) uptake for growth.

  13. Effects of motilin on potassium and calcium currents of rat's proximal colon smooth muscle cells%胃动素对大鼠近端结肠平滑肌细胞钙钾电流的影响

    Institute of Scientific and Technical Information of China (English)

    吴增祐; 罗和生; 梁成柏; 刘颖; 夏虹

    2012-01-01

    Objective To investigate the effects of motilin on the voltage dependent potassium channel and L-type calcium channel currents in rat proximal colon smooth muscle cells (PCSM) and to explore its mechanism in increasing colonic motility.Methods PCSM were isolated by collagenase.The voltage dependent potassium channel transit outward current (IKA ) and delayed rectifier current (IKdr) and L-type calcium currents (ICa(L)) were measured by whole cell patch clamp technique.Groups were analyzed by paired t-test.Results There was no significant effect of motilin on IKA and IKdr.L-type calcium channel was dose-dependently activated by motilin from 0.5 × 105 mmol/L to 10.0 ×10-5 mmol/L.At 6 × 10-5 mmol/L motilin and under - 10,0 and 10 mV stimulating voltage,maximum current density increased by 154.61%,62.69% and 21.02% respectively and activation kinetics curve obviously left shifted.Half activation voltage decreased from (2.740±1.211) mV prior administration to ( - 25.290 ± 0.614) mV (t =8.534,P =0.007 ) and there was no significant difference in slope factor. Conclusions Motilin increases colonic smooth muscle contraction by promoting calcium influx. However the frequency of colonic smooth muscle contraction could not change with frequency of equilibrium potential and action potential of colonic smooth muscle.%目的 研究胃动素对大鼠近端结肠平滑肌细胞(PCSM)膜电压依赖性钾通道及L型钙通道电流的影响,以探讨其增强结肠运动的机制.方法 采用酶解法分离大鼠PCSM,采用全细胞模式膜片钳技术测定PCSM电压依赖性钾离子通道快速激活型钾电流及延迟整流型钾电流和L型钙电流,组间比较采用配对t检验.结果 胃动素对快速激活型钾电流及延迟整流型钾电流无明显作用.(0.5~10.0)×10-5 mmol/L胃动素浓度依赖性地激活L型钙电流,6×10-5 mmol/L胃动素在-10、0及10 mV刺激电压下,使最大电流密度分别增加154.61%、62.69%及21.02%,激

  14. Rab3 interacting molecule 3 mutations associated with autism alter regulation of voltage-dependent Ca²⁺ channels.

    Science.gov (United States)

    Takada, Yoshinori; Hirano, Mitsuru; Kiyonaka, Shigeki; Ueda, Yoshifumi; Yamaguchi, Kazuma; Nakahara, Keiko; Mori, Masayuki X; Mori, Yasuo

    2015-09-01

    Autism is a neurodevelopmental psychiatric disorder characterized by impaired reciprocal social interaction, disrupted communication, and restricted and stereotyped patterns of interests. Autism is known to have a strong genetic component. Although mutations in several genes account for only a small proportion of individuals with autism, they provide insight into potential biological mechanisms that underlie autism, such as dysfunction in Ca(2+) signaling, synaptic dysfunction, and abnormal brain connectivity. In autism patients, two mutations have been reported in the Rab3 interacting molecule 3 (RIM3) gene. We have previously demonstrated that RIM3 physically and functionally interacts with voltage-dependent Ca(2+) channels (VDCCs) expressed in neurons via the β subunits, and increases neurotransmitter release. Here, by introducing corresponding autism-associated mutations that replace glutamic acid residue 176 with alanine (E176A) and methionine residue 259 with valine (M259V) into the C2B domain of mouse RIM3, we demonstrate that both mutations partly cancel the suppressive RIM3 effect on voltage-dependent inactivation of Ba(2+) currents through P/Q-type CaV2.1 recombinantly expressed in HEK293 cells. In recombinant N-type CaV2.2 VDCCs, the attenuation of the suppressive RIM3 effect on voltage-dependent inactivation is conserved for M259V but not E176A. Slowing of activation speed of P/Q-type CaV2.1 currents by RIM3 is abolished in E176A, while the physical interaction between RIM3 and β subunits is significantly attenuated in M259V. Moreover, increases by RIM3 in depolarization-induced Ca(2+) influx and acetylcholine release are significantly attenuated by E176A in rat pheochromocytoma PC12 cells. Thus, our data raise the interesting possibility that autism phenotypes are elicited by synaptic dysfunction via altered regulation of presynaptic VDCC function and neurotransmitter release.

  15. Contribution of presynaptic calcium-activated potassium currents to transmitter release regulation in cultured Xenopus nerve-muscle synapses.

    Science.gov (United States)

    Pattillo, J M; Yazejian, B; DiGregorio, D A; Vergara, J L; Grinnell, A D; Meriney, S D

    2001-01-01

    Using Xenopus nerve-muscle co-cultures, we have examined the contribution of calcium-activated potassium (K(Ca)) channels to the regulation of transmitter release evoked by single action potentials. The presynaptic varicosities that form on muscle cells in these cultures were studied directly using patch-clamp recording techniques. In these developing synapses, blockade of K(Ca) channels with iberiotoxin or charybdotoxin decreased transmitter release by an average of 35%. This effect would be expected to be caused by changes in the late phases of action potential repolarization. We hypothesize that these changes are due to a reduction in the driving force for calcium that is normally enhanced by the local hyperpolarization at the active zone caused by potassium current through the K(Ca) channels that co-localize with calcium channels. In support of this hypothesis, we have shown that when action potential waveforms were used as voltage-clamp commands to elicit calcium current in varicosities, peak calcium current was reduced only when these waveforms were broadened beginning when action potential repolarization was 20% complete. In contrast to peak calcium current, total calcium influx was consistently increased following action potential broadening. A model, based on previously reported properties of ion channels, faithfully reproduced predicted effects on action potential repolarization and calcium currents. From these data, we suggest that the large-conductance K(Ca) channels expressed at presynaptic varicosities regulate transmitter release magnitude during single action potentials by altering the rate of action potential repolarization, and thus the magnitude of peak calcium current.

  16. Submicromolar concentrations of zinc irreversibly reduce a calcium-dependent potassium current in rat hippocampal neurons in vitro.

    Science.gov (United States)

    Sim, J A; Cherubini, E

    1990-01-01

    The action of the endogenous divalent cation zinc on Ca2+ and Ca2(+)-dependent currents was studied in rat hippocampal CA1 and CA3 neurons in vitro, by means of a single electrode voltage clamp technique. Bath application of zinc (0.5-1 microM) produced a small membrane depolarization associated with an increase in synaptic noise and cell excitability and a depression of the afterhyperpolarization following a train of action potentials. The effects on the afterhyperpolarization, could not be reversed on washout. In voltage-clamped neurons, zinc induced a steady inward current and reduced, at resting membrane potential, the peak amplitude of the outward current underlying the afterhyperpolarization, IAHP. In caesium loaded neurons (in the presence of tetrodotoxin and tetraethylammonium), zinc reduced the slow inactivating Ca2+ current activated from a holding potential of -40 mV. Similar results were observed with nickel and cobalt at comparable concentrations, with Zn2+ greater than Ni2+ greater than Co2+, in their order of potency. In contrast to nickel and cobalt the effects of zinc did not reverse on washout. These results suggest that low concentrations of zinc enhance cell excitability by reducing IAHP. In addition, zinc reduces the slow inactivating voltage-dependent Ca2+ current. The irreversible effect of this metal ion is compatible with a toxic, intracellular site of action.

  17. A critical GxxxA motif in the γ6 calcium channel subunit mediates its inhibitory effect on Cav3.1 calcium current

    Science.gov (United States)

    Lin, Zuojun; Witschas, Katja; Garcia, Thomas; Chen, Ren-Shiang; Hansen, Jared P; Sellers, Zachary M; Kuzmenkina, Elza; Herzig, Stefan; Best, Philip M

    2008-01-01

    The eight members of the calcium channel γ subunit family are integral membrane proteins that regulate the expression and behaviour of voltage and ligand gated ion channels. While a subgroup consisting of γ2, γ3, γ4 and γ8 (the TARPs) modulate AMPA receptor localization and function, the γ1 and γ6 subunits conform to the original description of these proteins as regulators of voltage gated calcium channels. We have previously shown that the γ6 subunit is highly expressed in atrial myocytes and that it is capable of acting as a negative modulator of low voltage activated calcium current. In this study we extend our understanding of γ6 subunit modulation of low voltage activated calcium current. Using engineered chimeric constructs, we demonstrate that the first transmembrane domain (TM1) of γ6 is necessary for its inhibitory effect on Cav3.1 current. Mutational analysis is then used to identify a unique GxxxA motif within TM1 that is required for the function of the subunit strongly suggesting the involvement of helix–helix interactions in its effects. Results from co-immunoprecipitation experiments confirm a physical association of γ6 with the Cav3.1 channel in both HEK cells and atrial myocytes. Single channel analysis reveals that binding of γ6 reduces channel availability for activation. Taken together, the results of this study provide both a molecular and a mechanistic framework for understanding the unique ability of the γ6 calcium channel subunit to modulate low voltage activated (Cav3.1) calcium current density. PMID:18818244

  18. A critical GxxxA motif in the gamma6 calcium channel subunit mediates its inhibitory effect on Cav3.1 calcium current.

    Science.gov (United States)

    Lin, Zuojun; Witschas, Katja; Garcia, Thomas; Chen, Ren-Shiang; Hansen, Jared P; Sellers, Zachary M; Kuzmenkina, Elza; Herzig, Stefan; Best, Philip M

    2008-11-15

    The eight members of the calcium channel gamma subunit family are integral membrane proteins that regulate the expression and behaviour of voltage and ligand gated ion channels. While a subgroup consisting of gamma(2), gamma(3), gamma(4) and gamma(8) (the TARPs) modulate AMPA receptor localization and function, the gamma(1) and gamma(6) subunits conform to the original description of these proteins as regulators of voltage gated calcium channels. We have previously shown that the gamma(6) subunit is highly expressed in atrial myocytes and that it is capable of acting as a negative modulator of low voltage activated calcium current. In this study we extend our understanding of gamma(6) subunit modulation of low voltage activated calcium current. Using engineered chimeric constructs, we demonstrate that the first transmembrane domain (TM1) of gamma(6) is necessary for its inhibitory effect on Cav3.1 current. Mutational analysis is then used to identify a unique GxxxA motif within TM1 that is required for the function of the subunit strongly suggesting the involvement of helix-helix interactions in its effects. Results from co-immunoprecipitation experiments confirm a physical association of gamma(6) with the Cav3.1 channel in both HEK cells and atrial myocytes. Single channel analysis reveals that binding of gamma(6) reduces channel availability for activation. Taken together, the results of this study provide both a molecular and a mechanistic framework for understanding the unique ability of the gamma(6) calcium channel subunit to modulate low voltage activated (Cav3.1) calcium current density.

  19. Role of tumor necrosis factor-αin the regulation of T-type calcium channel current in HL-1 cells

    Institute of Scientific and Technical Information of China (English)

    RAO Fang; SHAN Zhi-xin; ZHU Jie-ning; XIE Zhi; WU Shu-lin; DENG Chun-yu; XUE Yu-mei; YU Xi-yong; WEI Wei; LIU Fang-zhou; YANG Hui; KUANG Su-juan; CHEN Shao-xian; XIAO Ding-zhang

    2016-01-01

    AIM:Increasing evidence indicates that inflammation contributes to the initiation and perpetuation of atrial fibrillation ( AF) .Al-though tumor necrosis factor ( TNF)-αlevels are increased in patients with AF , the role of TNF-αin the pathogenesis of AF remains unclear.Recent research has revealed that T-type Ca2+currents ( ICa,T ) play an important role in the pathogenesis of AF .METH-ODS:In this study , we used the whole-cell voltage-clamp technique and biochemical assays to explore the role of TNF-αin the regula-tion of ICa,T in atrial myocytes.RESULTS:We found that compared with sinus rhythm (SR) controls, T-type calcium channel (TCC) subunit mRNA levels were decreased , while TNF-αexpression levels were increased , in human atrial tissue from patients with AF .In murine atrial myocyte HL-1 cells, after cultured for 24 h, 12.5, 25 and 50 μg/L TNF-αsignificantly reduced the protein expression levels of the TCC α1G subunit in a concentration-dependent manner .The peak current was reduced by the application of 12.5 or 25μg/L TNF-αin a concentration-dependent manner [from ( -15.08 ±1.11) pA/pF in controls to ( -11.89 ±0.83) pA/pF and (-8.54 ±1.55) pA/pF in 12.5 and 25 μg/L TNF-αgroups, respectively].TNF-αapplication also inhibited voltage-dependent inactivation of ICa,T shifted the inactivation curve to the left .CONCLUSION:These results suggest that TNF-αis involved in the path-ogenesis of AF, probably via decreasing ICa,T function in atrium-derived myocytes through impaired channel function and down -regula-tion of channel protein expression .This pathway thus represents a potential pathogenic mechanism in AF .

  20. Enhanced currents through L-type calcium channels in cardiomyocytes disturb the electrophysiology of the dystrophic heart.

    Science.gov (United States)

    Koenig, Xaver; Rubi, Lena; Obermair, Gerald J; Cervenka, Rene; Dang, Xuan B; Lukacs, Peter; Kummer, Stefan; Bittner, Reginald E; Kubista, Helmut; Todt, Hannes; Hilber, Karlheinz

    2014-02-15

    Duchenne muscular dystrophy (DMD), induced by mutations in the gene encoding for the cytoskeletal protein dystrophin, is an inherited disease characterized by progressive muscle weakness. Besides the relatively well characterized skeletal muscle degenerative processes, DMD is also associated with cardiac complications. These include cardiomyopathy development and cardiac arrhythmias. The current understanding of the pathomechanisms in the heart is very limited, but recent research indicates that dysfunctional ion channels in dystrophic cardiomyocytes play a role. The aim of the present study was to characterize abnormalities in L-type calcium channel function in adult dystrophic ventricular cardiomyocytes. By using the whole cell patch-clamp technique, the properties of currents through calcium channels in ventricular cardiomyocytes isolated from the hearts of normal and dystrophic adult mice were compared. Besides the commonly used dystrophin-deficient mdx mouse model for human DMD, we also used mdx-utr mice, which are both dystrophin- and utrophin-deficient. We found that calcium channel currents were significantly increased, and channel inactivation was reduced in dystrophic cardiomyocytes. Both effects enhance the calcium influx during an action potential (AP). Whereas the AP in dystrophic mouse cardiomyocytes was nearly normal, implementation of the enhanced dystrophic calcium conductance in a computer model of a human ventricular cardiomyocyte considerably prolonged the AP. Finally, the described dystrophic calcium channel abnormalities entailed alterations in the electrocardiograms of dystrophic mice. We conclude that gain of function in cardiac L-type calcium channels may disturb the electrophysiology of the dystrophic heart and thereby cause arrhythmias.

  1. The slow calcium-dependent potassium current in a myenteric neurone of the guinea-pig ileum.

    Science.gov (United States)

    Hirst, G D; Johnson, S M; van Helden, D F

    1985-04-01

    Experiments were performed in current-clamped and voltage-clamped after-hyperpolarizing (AH) neurones of the guinea-pig myenteric plexus to examine the properties of the potassium conductance (gK, Ca) underlying the slow calcium-activated after-hyperpolarization (VK, Ca). The action potential plateau lengthened by the addition of tetraethylammonium chloride (TEA) to the bathing medium was compared to VK, Ca. Results were consistent with enhanced calcium entry causing an increase of VK, Ca. 4-Aminopyridine (4-AP) directly reduced VK, Ca. Voltage-clamp data of gK, Ca were well fitted by a process with a delay (approximately equal to 60 ms) followed by exponential activation (time constant approximately equal to 300 ms) and inactivation (time constant approximately equal to 2 s). The presence of a small, much slower inactivating process was noted. Values for time constants were similar to those reported by Morita, North & Tokimasa (1982) and North & Tokimasa (1983) where gK, Ca was measured during VK, Ca subsequent to action potential stimulation. The relation between gK, Ca (or the calcium-activated potassium current IK, Ca) and estimated calcium influx resulting from short-duration calcium currents elicited at various voltages was compared. Both the integral of the calcium current and gK, Ca showed a similar dependence on the depolarizations used to elicit IK, Ca except there was a positive shift of about 4 mV for the gK, Ca curve. This shift was attributed to a requirement for calcium ions to prime the gK, Ca mechanism. An inward ion charge movement of about 8 pC was required before significant activation of gK, Ca occurred. After the 'priming' condition had been established, the sensitivity of gK, Ca to inward calcium current measured at the resting potential was about 500 pS/pC of inward charge. Large calcium entry obtained by prolonged calcium currents caused saturation of the peak amplitude of IK, Ca and initiated currents with slower time to peak amplitude and

  2. Voltage-dependent ionic channels in differentiating neural precursor cells collected from adult mouse brains six hours post-mortem.

    Science.gov (United States)

    Bellardita, Carmelo; Bolzoni, Francesco; Sorosina, Melissa; Marfia, Giovanni; Carelli, Stephana; Gorio, Alfredo; Formenti, Alessandro

    2012-04-01

    A novel type of adult neural precursor cells (NPCs) has been isolated from the subventricular zone of the mouse 6 hr after animal death (T6-NPCs). This condition is supposed to select hypoxia-resistant cells of scientific and clinical interest. Ionic channels are ultimately the expression of the functional maturation of neurons, so the aim of this research was to characterize the pattern of the main voltage-dependent ionic channels in T6-NPCs differentiating to a neuronal phenotype, comparing it with NPCs isolated soon after death (T0-NPCs). T6- and T0-NPCs grow in medium containing epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF). Differentiation was performed in small wells without the addition of growth factors, in the presence of adhesion molecules, fetal bovine serum, and leukemia inhibitory factor. Ionic currents, recorded by means of whole-cell patch-clamp, namely, I(Ca2+) HVA, both L- and non-L-type, I(K+) delayed rectifying, I(K+) inward rectifier, transient I(K+A) , and TTX-sensitive I(Na+) have been found, although Na(+) currents were found in only a small percentage of cells and after the fifth week of differentiation. No significant differences in current types, density, orcell capacitance were observed between T6-NPCs and T0-NPCs. The sequence in which the markers appear in new neural cells is not necessarily a fixed program, but the discrepancies in morphological, biochemical, and electrophysiological maturation of mouse NPCs to neurons, possibly different in vivo, suggest that the various steps of the differentiation are independently regulated. Therefore, in addition to morphological and biochemical data, functional tests should be considered for characterizing the maturation of neurons.

  3. Inhibitors of arachidonate-regulated calcium channel signaling suppress triggered activity induced by the late sodium current.

    Science.gov (United States)

    Wolkowicz, Paul; Umeda, Patrick K; Sharifov, Oleg F; White, C Roger; Huang, Jian; Mahtani, Harry; Urthaler, Ferdinand

    2014-02-05

    Disturbances in myocyte calcium homeostasis are hypothesized to be one cause for cardiac arrhythmia. The full development of this hypothesis requires (i) the identification of all sources of arrhythmogenic calcium and (ii) an understanding of the mechanism(s) through which calcium initiates arrhythmia. To these ends we superfused rat left atria with the late sodium current activator type II Anemonia sulcata toxin (ATXII). This toxin prolonged atrial action potentials, induced early afterdepolarization, and provoked triggered activity. The calmodulin-dependent protein kinase II (CaMKII) inhibitor KN-93 (N-[2-[[[3-(4-chlorophenyl)-2-propenyl]methylamino]methyl]phenyl]-N-(2-hydroxyethyl)-4-methoxybenzenesulphon-amide) suppressed ATXII triggered activity but its inactive congener KN-92 (2-[N-(4-methoxy benzenesulfonyl)]amino-N-(4-chlorocinnamyl)-N-methylbenzylamine) did not. Neither drug affected normal atrial contractility. Calcium entry via L-type channels or calcium leakage from sarcoplasmic reticulum stores are not critical for this type of ectopy as neither verapamil ((RS)-2-(3,4-dimethoxyphenyl)-5-{[2-(3,4-dimethoxyphenyl)ethyl]-(methyl)amino}-2-prop-2-ylpentanenitrile) nor ryanodine affected ATXII triggered activity. By contrast, inhibitors of the voltage independent arachidonate-regulated calcium (ARC) channel and the store-operated calcium channel specifically suppressed ATXII triggered activity without normalizing action potentials or affecting atrial contractility. Inhibitors of cytosolic calcium-dependent phospholipase A2 also suppressed triggered activity suggesting that this lipase, which generates free arachidonate, plays a key role in ATXII ectopy. Thus, increased left atrial late sodium current appears to activate atrial Orai-linked ARC and store operated calcium channels, and these voltage-independent channels may be unexpected sources for the arrhythmogenic calcium that underlies triggered activity.

  4. Hydrogen sulfide inhibits L-type calcium currents depending upon the protein sulfhydryl state in rat cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Rongyuan Zhang

    Full Text Available Hydrogen sulfide (H(2S is a novel gasotransmitter that inhibits L-type calcium currents (I (Ca, L. However, the underlying molecular mechanisms are unclear. In particular, the targeting site in the L-type calcium channel where H(2S functions remains unknown. The study was designed to investigate if the sulfhydryl group could be the possible targeting site in the L-type calcium channel in rat cardiomyocytes. Cardiac function was measured in isolated perfused rat hearts. The L-type calcium currents were recorded by using a whole cell voltage clamp technique on the isolated cardiomyocytes. The L-type calcium channel containing free sulfhydryl groups in H9C2 cells were measured by using Western blot. The results showed that sodium hydrosulfide (NaHS, an H(2S donor produced a negative inotropic effect on cardiac function, which could be partly inhibited by the oxidant sulfhydryl modifier diamide (DM. H(2S donor inhibited the peak amplitude of I( Ca, L in a concentration-dependent manner. However, dithiothreitol (DTT, a reducing sulfhydryl modifier markedly reversed the H(2S donor-induced inhibition of I (Ca, L in cardiomyocytes. In contrast, in the presence of DM, H(2S donor could not alter cardiac function and L type calcium currents. After the isolated rat heart or the cardiomyocytes were treated with DTT, NaHS could markedly alter cardiac function and L-type calcium currents in cardiomyocytes. Furthermore, NaHS could decrease the functional free sulfhydryl group in the L-type Ca(2+ channel, which could be reversed by thiol reductant, either DTT or reduced glutathione. Therefore, our results suggest that H(2S might inhibit L-type calcium currents depending on the sulfhydryl group in rat cardiomyocytes.

  5. 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.

  6. RAS-RAF-MEK-dependent oxidative cell death involving voltage-dependent anion channels.

    Science.gov (United States)

    Yagoda, Nicholas; von Rechenberg, Moritz; Zaganjor, Elma; Bauer, Andras J; Yang, Wan Seok; Fridman, Daniel J; Wolpaw, Adam J; Smukste, Inese; Peltier, John M; Boniface, J Jay; Smith, Richard; Lessnick, Stephen L; Sahasrabudhe, Sudhir; Stockwell, Brent R

    2007-06-14

    Therapeutics that discriminate between the genetic makeup of normal cells and tumour cells are valuable for treating and understanding cancer. Small molecules with oncogene-selective lethality may reveal novel functions of oncoproteins and enable the creation of more selective drugs. Here we describe the mechanism of action of the selective anti-tumour agent erastin, involving the RAS-RAF-MEK signalling pathway functioning in cell proliferation, differentiation and survival. Erastin exhibits greater lethality in human tumour cells harbouring mutations in the oncogenes HRAS, KRAS or BRAF. Using affinity purification and mass spectrometry, we discovered that erastin acts through mitochondrial voltage-dependent anion channels (VDACs)--a novel target for anti-cancer drugs. We show that erastin treatment of cells harbouring oncogenic RAS causes the appearance of oxidative species and subsequent death through an oxidative, non-apoptotic mechanism. RNA-interference-mediated knockdown of VDAC2 or VDAC3 caused resistance to erastin, implicating these two VDAC isoforms in the mechanism of action of erastin. Moreover, using purified mitochondria expressing a single VDAC isoform, we found that erastin alters the permeability of the outer mitochondrial membrane. Finally, using a radiolabelled analogue and a filter-binding assay, we show that erastin binds directly to VDAC2. These results demonstrate that ligands to VDAC proteins can induce non-apoptotic cell death selectively in some tumour cells harbouring activating mutations in the RAS-RAF-MEK pathway.

  7. Functional unit size of the neurotoxin receptors on the voltage-dependent sodium channel.

    Science.gov (United States)

    Angelides, K J; Nutter, T J; Elmer, L W; Kempner, E S

    1985-03-25

    Radiation inactivation was used in situ to determine the functional unit sizes of the neurotoxin receptors of the voltage-dependent sodium channel from rat brain. Frozen or lyophilized synaptosomes were irradiated with high energy electrons generated by a linear accelerator and assayed for [3H]saxitoxin, 125I-Leiurus quinquestriatus quinquestriatus (alpha-scorpion toxin), 125I-Centruroides suffusus suffusus (beta-scorpion toxin), and batrachotoxinin-A 20 alpha-[3H]benzoate binding activity. The functional unit size of the neurotoxin receptors determined in situ by target analysis are 220,000 for saxitoxin, 263,000 for alpha-scorpion toxin, and 45,000 for beta-scorpion toxin. Analysis of the inactivation curve for batrachotoxinin-A 20 alpha-benzoate binding to the channel yields two target sizes of Mr approximately 287,000 (50%) and approximately 51,000 (50%). The results are independent of the purity of the membrane preparation. Comparison of the radiation inactivation data with the protein composition of the rat brain sodium channel indicates that there are at least two functional components.

  8. Regulation of mitochondrial function by voltage dependent anion channels in ethanol metabolism and the Warburg effect.

    Science.gov (United States)

    Lemasters, John J; Holmuhamedov, Ekhson L; Czerny, Christoph; Zhong, Zhi; Maldonado, Eduardo N

    2012-06-01

    Voltage dependent anion channels (VDAC) are highly conserved proteins that are responsible for permeability of the mitochondrial outer membrane to hydrophilic metabolites like ATP, ADP and respiratory substrates. Although previously assumed to remain open, VDAC closure is emerging as an important mechanism for regulation of global mitochondrial metabolism in apoptotic cells and also in cells that are not dying. During hepatic ethanol oxidation to acetaldehyde, VDAC closure suppresses exchange of mitochondrial metabolites, resulting in inhibition of ureagenesis. In vivo, VDAC closure after ethanol occurs coordinately with mitochondrial uncoupling. Since acetaldehyde passes through membranes independently of channels and transporters, VDAC closure and uncoupling together foster selective and more rapid oxidative metabolism of toxic acetaldehyde to nontoxic acetate by mitochondrial aldehyde dehydrogenase. In single reconstituted VDAC, tubulin decreases VDAC conductance, and in HepG2 hepatoma cells, free tubulin negatively modulates mitochondrial membrane potential, an effect enhanced by protein kinase A. Tubulin-dependent closure of VDAC in cancer cells contributes to suppression of mitochondrial metabolism and may underlie the Warburg phenomenon of aerobic glycolysis. This article is part of a Special Issue entitled: VDAC structure, function, and regulation of mitochondrial metabolism.

  9. Regulation of KV channel voltage-dependent activation by transmembrane β subunits

    Directory of Open Access Journals (Sweden)

    Xiaohui eSun

    2012-04-01

    Full Text Available Voltage-activated K+ (KV channels are important for shaping action potentials and maintaining resting membrane potential in excitable cells. KV channels contain a central pore-gate domain (PGD surrounded by four voltage-sensing domains (VSD. The VSDs will change conformation in response to alterations of the membrane potential thereby inducing the opening of the PGD. Many KV channels are heteromeric protein complexes containing auxiliary β subunits. These β subunits modulate channel expression and activity to increase functional diversity and render tissue specific phenotypes. This review focuses on the KV β subunits that contain transmembrane (TM segments including the KCNE family and the β subunits of large conductance, Ca2+- and voltage-activated K+ (BK channels. These TM β subunits affect the voltage-dependent activation of KV α subunits. Experimental and computational studies have described the structural location of these β subunits in the channel complexes and the biophysical effects on VSD activation, PGD opening and VSD-PGD coupling. These results reveal some common characteristics and mechanistic insights into KV channel modulation by TM β subunits.

  10. Control of the light-regulated current in rod photoreceptors by cyclic GMP, calcium, and l-cis-diltiazem.

    Science.gov (United States)

    Stern, J H; Kaupp, U B; MacLeish, P R

    1986-02-01

    The effect of calcium ions on the cGMP-activated current of outer segment membrane was examined by the excised-patch technique. Changes in the extracellular calcium concentration had marked effects on the cGMP-activated current, while changes in intracellular calcium concentration were ineffective. Changes in calcium concentration in the absence of cGMP had little, if any, effect on membrane conductance. These results suggest that both intracellular cGMP and extracellular calcium can directly affect the conductance underlying the light response in rod cells. The pharmacological agent l-cis-diltiazem reversibly inhibited the cGMP-activated current when applied to the intracellular side of an excised patch. When superfused over intact rod cells, l-cis-diltiazem reversibly blocked much of the normal light response. The isomer, d-cis-diltiazem, did not significantly affect either patches or intact rod cells. Thus, the light-regulated conductance has binding sites for both calcium and cGMP that may interact during the normal light response in rod cells and a site specific for l-cis-diltiazem that can be used to identify and further study the conductance mechanism.

  11. A cyclic GMP-dependent calcium-activated chloride current in smooth-muscle cells from rat mesenteric resistance arteries

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Aalkjær, Christian; Nilsson, Holger

    2004-01-01

    -PET-cGMP or with a peptide inhibitor of PKG, or with the nonhydrolysable ATP analogue AMP-PNP. Under biionic conditions, the anion permeability sequence of the channel was SCN- > Br- > I- > Cl- > acetate > F- >> aspartate, but the conductance sequence was I- > Br- > Cl- > acetate > F- > aspartate = SCN-. The current had...... conditions of high calcium in the patch-pipette solution, a current similar to the latter could be identified also in the mesenteric artery smooth-muscle cells. We conclude that smooth-muscle cells from rat mesenteric resistance arteries have a novel cGMP-dependent calcium-activated chloride current, which...

  12. Tubocurarine blocks a calcium-dependent potassium current in rat tumoral pituitary cells.

    Science.gov (United States)

    Vacher, P; Vacher, A M; Mollard, P

    1998-04-30

    We investigated the effects of potassium channel inhibitors on electrical activity, membrane ionic currents, intracellular calcium concentration ([Ca2+]i) and hormone release in GH3/B6 cells (a line of pituitary origin). Patch-clamp recordings show a two-component after hyperpolarization (AHP) following each action potential (current clamp) or a two-component tail current (voltage-clamp). Both components can be blocked by inhibiting Ca2+ influx. Application of D-tubocurarine (dTc) (20-500 microM) reversibly suppressed the slowly decaying Ca2+-activated K+ tail current (I AHPs) in a concentration-dependent manner. On the other hand, low doses of tetraethylammonium ions (TEA+) only blocked the rapidly decaying voltage- and Ca2+-activated K+ tail current (I AHPf). Therefore, GH3/B6 cells exhibit at least two quite distinct Ca2+-dependent K+ currents, which differ in size, voltage- and Ca2+-sensitivity, kinetics and pharmacology. These two currents also play quite separate roles in shaping the action potential. d-tubocurarine increased spontaneous Ca2+ action potential firing, whereas TEA increased action potential duration. Thus, both agents stimulated Ca2+ entry. I AHPs is activated by a transient increase in [Ca2+]i such as a thyrotrophin releasing hormone-induced Ca2+ mobilization. All the K+ channel inhibitors we tested: TEA, apamin, dTC and charybdotoxin, stimulated prolactin and growth hormone release in GH3/B6 cells. Our results show that I AHPs is a good sensor for subplasmalemmal Ca2+ and that dTc is a good pharmacological tool for studying this current.

  13. Role of low voltage activated calcium channels in neuritogenesis and active migration of embryonic neural progenitor cells.

    Science.gov (United States)

    Louhivuori, Lauri M; Louhivuori, Verna; Wigren, Henna-Kaisa; Hakala, Elina; Jansson, Linda C; Nordström, Tommy; Castrén, Maija L; Akerman, Karl E

    2013-04-15

    The central role of calcium influx and electrical activity in embryonic development raises important questions about the role and regulation of voltage-dependent calcium influx. Using cultured neural progenitor cell (NPC) preparations, we recorded barium currents through voltage-activated channels using the whole-cell configuration of the patch-clamp technique and monitored intracellular free calcium concentrations with Fura-2 digital imaging. We found that NPCs as well as expressing high-voltage-activated (HVA) calcium channels express functional low-threshold voltage-dependent calcium channels in the very early stages of differentiation (5 h to 1 day). The size of the currents recorded at -50 versus -20 mV after 1 day in differentiation was dependent on the nature of the charge carrier. Peak currents measured at -20 mV in the presence 10 mM Ca2+ instead of 10 mM Ba2+ had a tendency to be smaller, whereas the nature of the divalent species did not influence the amplitude measured at -50 mV. The T-type channel blockers mibefradil and NNC 55-0396 significantly reduced the calcium responses elicited by depolarizing with extracellular potassium, while the overall effect of the HVA calcium channel blockers was small at differentiation day 1. At differentiation day 20, the calcium responses were effectively blocked by nifedipine. Time-lapse imaging of differentiating neurospheres cultured in the presence of low-voltage-activated (LVA) blockers showed a significant decrease in the number of active migrating neuron-like cells and neurite extensions. Together, these data provide evidence that LVA calcium channels are involved in the physiology of differentiating and migrating NPCs.

  14. The human red cell voltage-dependent cation channel. Part III: Distribution homogeneity and pH dependence

    DEFF Research Database (Denmark)

    Bennekou, P.; Barksmann, T. L.; Christophersen, P.

    2006-01-01

    The homogeneity of the distribution of the non-selective voltage-dependent cation channel (the NSVDC channel) in the human erythrocyte, and the pH dependence was investigated. Activation of this channel caused a uniform cellular dehydration, which was characterized by the changes in the erythrocyte...

  15. Voltage dependence of Hodgkin-Huxley rate functions for a multistage K^{+} channel voltage sensor within a membrane.

    Science.gov (United States)

    Vaccaro, S R

    2014-11-01

    The activation of a K^{+} channel sensor in two sequential stages during a voltage clamp may be described as the translocation of a Brownian particle in an energy landscape with two large barriers between states. A solution of the Smoluchowski equation for a square-well approximation to the potential function of the S4 voltage sensor satisfies a master equation and has two frequencies that may be determined from the forward and backward rate functions. When the higher-frequency terms have small amplitude, the solution reduces to the relaxation of a rate equation, where the derived two-state rate functions are dependent on the relative magnitude of the forward rates (α and γ) and the backward rates (β and δ) for each stage. In particular, the voltage dependence of the Hodgkin-Huxley rate functions for a K^{+} channel may be derived by assuming that the rate functions of the first stage are large relative to those of the second stage-α≫γ and β≫δ. For a Shaker IR K^{+} channel, the first forward and backward transitions are rate limiting (αchannel and a noninactivating Na^{+} ion channel is determined by the master equation for K^{+} channel activation and the ionic current equation when the Na^{+} channel activation time is small, and if β≪δ and α≪γ, the system may exhibit a small amplitude oscillation between spikes, or mixed-mode oscillation, in which the slow closed state modulates the K^{+} ion channel conductance in the membrane.

  16. Thiazolidinedione insulin sensitizers alter lipid bilayer properties and voltage-dependent sodium channel function: implications for drug discovery.

    Science.gov (United States)

    Rusinova, Radda; Herold, Karl F; Sanford, R Lea; Greathouse, Denise V; Hemmings, Hugh C; Andersen, Olaf S

    2011-08-01

    The thiazolidinediones (TZDs) are used in the treatment of diabetes mellitus type 2. Their canonical effects are mediated by activation of the peroxisome proliferator-activated receptor γ (PPARγ) transcription factor. In addition to effects mediated by gene activation, the TZDs cause acute, transcription-independent changes in various membrane transport processes, including glucose transport, and they alter the function of a diverse group of membrane proteins, including ion channels. The basis for these off-target effects is unknown, but the TZDs are hydrophobic/amphiphilic and adsorb to the bilayer-water interface, which will alter bilayer properties, meaning that the TZDs may alter membrane protein function by bilayer-mediated mechanisms. We therefore explored whether the TZDs alter lipid bilayer properties sufficiently to be sensed by bilayer-spanning proteins, using gramicidin A (gA) channels as probes. The TZDs altered bilayer elastic properties with potencies that did not correlate with their affinity for PPARγ. At concentrations where they altered gA channel function, they also altered the function of voltage-dependent sodium channels, producing a prepulse-dependent current inhibition and hyperpolarizing shift in the steady-state inactivation curve. The shifts in the inactivation curve produced by the TZDs and other amphiphiles can be superimposed by plotting them as a function of the changes in gA channel lifetimes. The TZDs' partition coefficients into lipid bilayers were measured using isothermal titration calorimetry. The most potent bilayer modifier, troglitazone, alters bilayer properties at clinically relevant free concentrations; the least potent bilayer modifiers, pioglitazone and rosiglitazone, do not. Unlike other TZDs tested, ciglitazone behaves like a hydrophobic anion and alters the gA monomer-dimer equilibrium by more than one mechanism. Our results provide a possible mechanism for some off-target effects of an important group of drugs, and

  17. Origin of the voltage dependence of G-protein regulation of P/Q-type Ca2+ channels.

    Science.gov (United States)

    Zhang, Yun; Chen, Yu-Hang; Bangaru, Saroja D; He, Linling; Abele, Kathryn; Tanabe, Shihori; Kozasa, Tohru; Yang, Jian

    2008-12-24

    G-protein (Gbetagamma)-mediated voltage-dependent inhibition of N- and P/Q-type Ca(2+) channels contributes to presynaptic inhibition and short-term synaptic plasticity. The voltage dependence derives from the dissociation of Gbetagamma from the inhibited channels, but the underlying molecular and biophysical mechanisms remain largely unclear. In this study we investigated the role in this process of Ca(2+) channel beta subunit (Ca(v)beta) and a rigid alpha-helical structure between the alpha-interacting domain (AID), the primary Ca(v)beta docking site on the channel alpha(1) subunit, and the pore-lining IS6 segment. Gbetagamma inhibition of P/Q-type channels was reconstituted in giant inside-out membrane patches from Xenopus oocytes. Large populations of channels devoid of Ca(v)beta were produced by washing out a mutant Ca(v)beta with a reduced affinity for the AID. These beta-less channels were still inhibited by Gbetagamma, but without any voltage dependence, indicating that Ca(v)beta is indispensable for voltage-dependent Gbetagamma inhibition. A truncated Ca(v)beta containing only the AID-binding guanylate kinase (GK) domain could fully confer voltage dependence to Gbetagamma inhibition. Gbetagamma did not alter inactivation properties, and channels recovered from Gbetagamma inhibition exhibited the same activation property as un-inhibited channels, indicating that Gbetagamma does not dislodge Ca(v)beta from the inhibited channel. Furthermore, voltage-dependent Gbetagamma inhibition was abolished when the rigid alpha-helix between the AID and IS6 was disrupted by insertion of multiple glycines, which also eliminated Ca(v)beta regulation of channel gating, revealing a pivotal role of this rigid alpha-helix in both processes. These results suggest that depolarization-triggered movement of IS6, coupled to the subsequent conformational change of the Gbetagamma-binding pocket through a rigid alpha-helix induced partly by the Ca(v)beta GK domain, causes the

  18. Effect of resveratrol on L-type calcium current in rat ventricular myocytes

    Institute of Scientific and Technical Information of China (English)

    Li-ping ZHANG; Jing-xiang YIN; Zheng LIU; Yi ZHANG; Qing-shan WANG; Juan ZHAO

    2006-01-01

    Aim: To study the effect of resveratrol on L-type calcium current (ICa-L) in isolated rat ventricular myocytes and the mechanisms underlying these effects. Methods:ICa-L was examined in isolated single rat ventricular myocytes by using the whole cell patch-clamp recording technique. Results: Resveratrol (10-40 μmol/L) reduced the peak amplitude of ICa-L and shifted the current-voltage (I-V) curve upwards in a concentration-dependent manner. Resveratrol (10, 20, 40 μmol/L)decreased the peak amplitude of ICa-L from -14.2± 1.5 pA/pF to -10.5± 1.5 pA/pF (P<0.05), -7.5±2.4 pA/pF (P<0.01), and -5.2±1.2 pA/pF (P<0.01), respectively.Resveratrol (40 μmol/L) shifted the steady-state activation curve of ICa-L to the right and changed the half-activation potential (V0.5) from -19.4±0.4 mV to -15.4±1.9 mV (P<0.05). Resveratrol at a concentration of 40 μmol/L did not affect the steady-state inactivation curve of ICa-L, but did markedly shift the timedependent recovery curve of ICa-L to the right, and slow down the recovery of ICa-L from inactivation. Sodium orthovanadate (Na3VO4; 1 mmol/L), a potent inhibitor of tyrosine phosphatase, significantly inhibited the effects of resveratrol (P<0.01). Conclusion: Resveratrol inhibited ICa- L mainly by inhibiting the activation of L-type calcium channels and slowing down the recovery of L-type calcium channels from inactivation. This inhibitory effect of resveratrol was mediated by the inhibition of protein tyrosine kinase in rat ventricular myocytes.

  19. T-type Ca2+ current activity during oocyte growth and maturation in the ascidian Styela plicata.

    Directory of Open Access Journals (Sweden)

    Alessandra Gallo

    Full Text Available Voltage-dependent calcium currents play a fundamental role during oocyte maturation, mostly L-type calcium currents, whereas T-type calcium currents are involved in sperm physiology and cell growth. In this paper, using an electrophysiological and pharmacological approach, we demonstrated, for the first time in oocytes, that T-type calcium currents are present with functional consequences on the plasma membrane of growing immature oocytes of the ascidian Styela plicata. We classified three subtypes of immature oocytes at the germinal vesicle stage on the basis of their size, morphology and accessory cellular structures. These stages were clearly associated with an increased activity of T-type calcium currents and hyperpolarization of the plasma membrane. We also observed that T-type calcium currents oscillate in the post-fertilization embryonic stages, with minimal amplitude of the currents in the zygote and maximal at 8-cell stage. In addition, chemical inhibition of T-type calcium currents, obtained by applying specific antagonists, induced a significant reduction in the rate of cleavage and absence of larval formation. We suggest that calcium entry via T-type calcium channels may act as a potential pacemaker in regulating cytosolic calcium involved in fertilization and early developmental events.

  20. Optical recording of calcium currents during impulse conduction in cardiac tissue.

    Science.gov (United States)

    Jousset, Florian; Rohr, Stephan

    2015-04-01

    We explore the feasibility of obtaining a spatially resolved picture of [Formula: see text] inward currents ([Formula: see text]) in multicellular cardiac tissue by differentiating optically recorded [Formula: see text] transients that accompany propagating action potentials. Patterned growth strands of neonatal rat ventricular cardiomyocytes were stained with the [Formula: see text] indicators Fluo-4 or Fluo-4FF. Preparations were stimulated at 1 Hz, and [Formula: see text] transients were recorded with high spatiotemporal resolution ([Formula: see text], 2 kHz analog bandwidth) with a photodiode array. Signals were differentiated after appropriate digital filtering. Differentiation of [Formula: see text] transients resulted in optically recorded calcium currents (ORCCs) that carried the temporal and pharmacological signatures of L-type [Formula: see text] inward currents: the time to peak amounted to [Formula: see text] (Fluo-4FF) and [Formula: see text] (Fluo-4), full-width at half-maximum was [Formula: see text], and ORCCs were completely suppressed by [Formula: see text][Formula: see text]. Also, and as reported before from patch-clamp studies, caffeine reversibly depressed the amplitude of ORCCs. The results demonstrate that the differentiation of [Formula: see text] transients can be used to obtain a spatially resolved picture of the initial phase of [Formula: see text] in cardiac tissue and to assess relative changes of activation/fast inactivation of [Formula: see text] following pharmacological interventions.

  1. Modulation by extracellular pH of low- and high-voltage-activated calcium currents of rat thalamic relay neurons.

    Science.gov (United States)

    Shah, M J; Meis, S; Munsch, T; Pape, H C

    2001-03-01

    The effects of changes in the extracellular pH (pH(o)) on low-voltage- (LVA) and high-voltage- (HVA) activated calcium currents of acutely isolated relay neurons of the ventrobasal thalamic complex (VB) were examined using the whole cell patch-clamp technique. Modest extracellular alkalinization (pH 7.3 to 7.7) reversibly enlarged LVA calcium currents by 18.6 +/- 3.2% (mean +/- SE, n = 6), whereas extracellular acidification (pH 7.3 to 6.9) decreased the current by 24.8 +/- 3.1% (n = 9). Normalized current amplitudes (I/I(7.3)) fitted as a function of pH(o) revealed an apparent pK(a) of 6.9. Both, half-maximal activation voltage and steady-state inactivation were significantly shifted to more negative voltages by 2-4 mV on extracellular alkalinization and to more positive voltages by 2-3 mV on extracellular acidification, respectively. Recovery from inactivation of LVA calcium currents was not significantly affected by changes in pH(o). In contrast, HVA calcium currents were less sensitive to changes in pH(o). Although extracellular alkalinization increased maximal HVA current by 6.0 +/- 2.0% (n = 7) and extracellular acidification decreased it by 11.9 +/- 0.02% (n = 11), both activation and steady-state inactivation were only marginally affected by the moderate changes in pH(o) used in the present study. The results show that calcium currents of thalamic relay neurons exhibit different pH(o) sensitivity. Therefore activity-related extracellular pH transients might selectively modulate certain aspects of the electrogenic behavior of thalamic relay neurons.

  2. Calcium ion channel and epilepsy

    Institute of Scientific and Technical Information of China (English)

    Yudan Lü; Weihong Lin; Dihui Ma

    2006-01-01

    OBJECTIVE: To review the relationship between calcium ion channel and epilepsy for well investigating the pathogenesis of epilepsy and probing into the new therapeutic pathway of epilepsy.DATA SOURCES: A computer-based online research Calcium ion channel and epilepsy related articles published between January 1994 and December 2006 in the CKNI and Wanfang database with the key words of "calcium influxion, epilepsy, calcium-channel blocker". The language was limited to Chinese. At the same time,related articles published between January 1993 and December 2006 in Pubmed were searched for on online with the key words of "calcium influxion, epilepsy" in English.STUDY SELECTION: The materials were selected firstly. Inclusive criteria: ① Studies related to calcium ion channel and the pat1hogenesis of epilepsy. ② Studies on the application of calcium ion channel blocker in the treatment of epilepsy. Exclusive criteria: repetitive or irrelated studies.DATA EXTRACTION: According to the criteria, 123 articles were retrieved and 93 were excluded due to repetitive or irrelated studies. Altogether 30 articles met the inclusive criteria, 11 of them were about the structure and characters of calcium ion channel, 10 about calcium ion channel and the pathogenesis of epilepsy and 9 about calcium blocker and the treatment of epilepsy.DATA SYNTHESIS: Calcium ion channels mainly consist of voltage dependent calcium channel and receptor operated calcium channel. Depolarization caused by voltage gating channel-induced influxion is the pathological basis of epileptic attack, and it is found in many studies that many anti-epileptic drugs have potential and direct effect to rivalizing voltage-dependent calcium ion channel.CONCLUSION: Calcium influxion plays an important role in the seizure of epilepsy. Some calcium antagonists seen commonly are being tried in the clinical therapy of epilepsy that is being explored, not applied in clinical practice. If there are enough evidences to

  3. Distribution of voltage-dependent and intracellular Ca2+ channels in submucosal neurons from rat distal colon.

    Science.gov (United States)

    Rehn, Matthias; Bader, Sandra; Bell, Anna; Diener, Martin

    2013-09-01

    We recently observed a bradykinin-induced increase in the cytosolic Ca2+ concentration in submucosal neurons of rat colon, an increase inhibited by blockers of voltage-dependent Ca2+ (Ca(v)) channels. As the types of Ca(v) channels used by this part of the enteric nervous system are unknown, the expression of various Ca(v) subunits has been investigated in whole-mount submucosal preparations by immunohistochemistry. Submucosal neurons, identified by a neuronal marker (microtubule-associated protein 2), are immunoreactive for Ca(v)1.2, Ca(v)1.3 and Ca(v)2.2, expression being confirmed by reverse transcription plus the polymerase chain reaction. These data agree with previous observations that the inhibition of L- and N-type Ca2+ currents strongly inhibits the response to bradykinin. However, whole-cell patch-clamp experiments have revealed that bradykinin does not enhance Ca2+ inward currents under voltage-clamp conditions. Consequently, bradykinin does not directly interact with Ca(v) channels. Instead, the kinin-induced Ca2+ influx is caused indirectly by the membrane depolarization evoked by this peptide. As intracellular Ca2+ channels on Ca(2+)-storing organelles can also contribute to Ca2+ signaling, their expression has been investigated by imaging experiments and immunohistochemistry. Inositol 1,4,5-trisphosphate (IP3) receptors (IP3R) have been functionally demonstrated in submucosal neurons loaded with the Ca(2+)-sensitive fluorescent dye, fura-2. Histamine, a typical agonist coupled to the phospholipase C pathway, induces an increase in the fura-2 signal ratio, which is suppressed by 2-aminophenylborate, a blocker of IP3 receptors. The expression of IP3R1 has been confirmed by immunohistochemistry. In contrast, ryanodine, tested over a wide concentration range, evokes no increase in the cytosolic Ca2+ concentration nor is there immunohistochemical evidence for the expression of ryanodine receptors in these neurons. Thus, rat submucosal neurons are equipped

  4. Brief RU 38486 Treatment Normalizes the Effects of Chronic Stress on Calcium Currents in Rat Hippocampal CA1 Neurons.

    NARCIS (Netherlands)

    Karst, H.; Joëls, M.

    2007-01-01

    Chronic stress alters many properties in rat brain, like serotonin responsiveness and dendritic morphology. In the present study, we examined (i) whether unpredictable stress during 21 days affects calcium (Ca) currents of CA1 pyramidal neurons recorded on day 22; and (ii) if so, whether this change

  5. VOLTAGE STABILITY ASSESSMENT FOR WIND FARMS INTEGRATION INTO ELECTRICITY GRIDS WITH AND WITHOUT CONSIDERATION OF VOLTAGE DEPENDENT LOADS

    Directory of Open Access Journals (Sweden)

    TOMA R.

    2016-09-01

    Full Text Available The paper presents a comparative study between the effects on voltage stability of the integration of a wind farm into the electricity grid with or without voltage dependent loads in the context of different locations of a synchronous compensator from the grid. The P-V curves are built by using the PowerFactory DigSilent 15.2.2 and a DPL script that implements a simplified form of the Continuation Power Flow method.

  6. Fibroblast growth factor 23 dysregulates late sodium current and calcium homeostasis with enhanced arrhythmogenesis in pulmonary vein cardiomyocytes.

    Science.gov (United States)

    Huang, Shih-Yu; Chen, Yao-Chang; Kao, Yu-Hsun; Hsieh, Ming-Hsiung; Lin, Yung-Kuo; Chung, Cheng-Chih; Lee, Ting-I; Tsai, Wen-Chin; Chen, Shih-Ann; Chen, Yi-Jen

    2016-10-25

    Fibroblast growth factor 23 (FGF23), elevated in chronic renal failure, increases atrial arrhythmogenesis and dysregulates calcium homeostasis. Late sodium currents (INa-Late) critically induces ectopic activity of pulmoanry vein (the most important atrial fibrillation trigger). This study was to investigate whether FGF23 activates the INa-Late leading to calcium dysregulation and increases PV arrhythmogenesis. Patch clamp, western blot, and confocal microscopy were used to evaluate the electrical activities, calcium homeostasis, and mitochondrial reactive oxygen species (ROS) in PV cardiomyocytes with or without FGF23 (0.1 or 1 ng/mL) incubation for 4~6 h. Compared to the control, FGF23 (1 ng/mL, but not 0.1 ng/mL)-treated PV cardiomyocytes had a faster beating rate. FGF23 (1 ng/mL)-treated PV cardiomyocytes had larger INa-Late, calcium transients, and mitochondrial ROS than controls. However, ranolazine (an inhibitor of INa-Late) attenuated FGF23 (1 ng/mL)-increased beating rates, calcium transients and mitochondrial ROS. FGF23 (1 ng/mL)-treated PV cardiomyocytes exhibited larger phosphorylation of calcium/calmodulin-dependent protein kinase II (CaMKII). Chelerythrine chloride (an inhibitor of protein kinase C) decreased INa-Late in FGF23 (1 ng/mL)-treated PV cardiomyocytes. However, KN93 (a selective CaMKII blocker) decreased INa-Late in control and FGF23 (1 ng/mL)-treated PV cardiomyocytes to a similar extent. In conclusion, FGF23 increased PV arrhythmogenesis through sodium and calcium dysregulation by acting protein kinase C signaling.

  7. Phosphoinositide 5- and 3-phosphatase activities of a voltage-sensing phosphatase in living cells show identical voltage dependence.

    Science.gov (United States)

    Keum, Dongil; Kruse, Martin; Kim, Dong-Il; Hille, Bertil; Suh, Byung-Chang

    2016-06-28

    Voltage-sensing phosphatases (VSPs) are homologs of phosphatase and tensin homolog (PTEN), a phosphatidylinositol 3,4-bisphosphate [PI(3,4)P2] and phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3] 3-phosphatase. However, VSPs have a wider range of substrates, cleaving 3-phosphate from PI(3,4)P2 and probably PI(3,4,5)P3 as well as 5-phosphate from phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and PI(3,4,5)P3 in response to membrane depolarization. Recent proposals say these reactions have differing voltage dependence. Using Förster resonance energy transfer probes specific for different PIs in living cells with zebrafish VSP, we quantitate both voltage-dependent 5- and 3-phosphatase subreactions against endogenous substrates. These activities become apparent with different voltage thresholds, voltage sensitivities, and catalytic rates. As an analytical tool, we refine a kinetic model that includes the endogenous pools of phosphoinositides, endogenous phosphatase and kinase reactions connecting them, and four exogenous voltage-dependent 5- and 3-phosphatase subreactions of VSP. We show that apparent voltage threshold differences for seeing effects of the 5- and 3-phosphatase activities in cells are not due to different intrinsic voltage dependence of these reactions. Rather, the reactions have a common voltage dependence, and apparent differences arise only because each VSP subreaction has a different absolute catalytic rate that begins to surpass the respective endogenous enzyme activities at different voltages. For zebrafish VSP, our modeling revealed that 3-phosphatase activity against PI(3,4,5)P3 is 55-fold slower than 5-phosphatase activity against PI(4,5)P2; thus, PI(4,5)P2 generated more slowly from dephosphorylating PI(3,4,5)P3 might never accumulate. When 5-phosphatase activity was counteracted by coexpression of a phosphatidylinositol 4-phosphate 5-kinase, there was accumulation of PI(4,5)P2 in parallel to PI(3,4,5)P3 dephosphorylation

  8. Calcium-activated potassium current modulates ventricular repolarization in chronic heart failure.

    Directory of Open Access Journals (Sweden)

    Ingrid M Bonilla

    Full Text Available The role of I(KCa in cardiac repolarization remains controversial and varies across species. The relevance of the current as a therapeutic target is therefore undefined. We examined the cellular electrophysiologic effects of I(KCa blockade in controls, chronic heart failure (HF and HF with sustained atrial fibrillation. We used perforated patch action potential recordings to maintain intrinsic calcium cycling. The I(KCa blocker (apamin 100 nM was used to examine the role of the current in atrial and ventricular myocytes. A canine tachypacing induced model of HF (1 and 4 months, n = 5 per group was used, and compared to a group of 4 month HF with 6 weeks of superimposed atrial fibrillation (n = 7. A group of age-matched canine controls were used (n = 8. Human atrial and ventricular myocytes were isolated from explanted end-stage failing hearts which were obtained from transplant recipients, and studied in parallel. Atrial myocyte action potentials were unchanged by I(KCa blockade in all of the groups studied. I(KCa blockade did not affect ventricular myocyte repolarization in controls. HF caused prolongation of ventricular myocyte action potential repolarization. I(KCa blockade caused further prolongation of ventricular repolarization in HF and also caused repolarization instability and early afterdepolarizations. SK2 and SK3 expression in the atria and SK3 in the ventricle were increased in canine heart failure. We conclude that during HF, I(KCa blockade in ventricular myocytes results in cellular arrhythmias. Furthermore, our data suggest an important role for I(KCa in the maintenance of ventricular repolarization stability during chronic heart failure. Our findings suggest that novel antiarrhythmic therapies should have safety and efficacy evaluated in both atria and ventricles.

  9. Current data with inulin-type fructans and calcium, targeting bone health in adults.

    Science.gov (United States)

    Coxam, Véronique

    2007-11-01

    In humans, there is increasing evidence that the colon can absorb nutritionally significant amounts of calcium, and this process may be susceptible to dietary manipulation by fermentable substrates, especially inulin-type fructans. Inulin-type fructans can modulate calcium absorption because they are resistant to hydrolysis by mammalian enzymes and are fermented in the large intestine to produce short-chain fatty acids, which in turn reduce luminal pH and modify calcium speciation, and hence solubility, or exert a direct effect on the mucosal transport pathway. Quite a few intervention studies showed an improvement of calcium absorption in adolescents or young adults by inulin-type fructans. In the same way, a positive effect has been reported in older women.

  10. Current and calcium responses to local activation of axonal NMDA receptors in developing cerebellar molecular layer interneurons.

    Directory of Open Access Journals (Sweden)

    Bénédicte Rossi

    Full Text Available In developing cerebellar molecular layer interneurons (MLIs, NMDA increases spontaneous GABA release. This effect had been attributed to either direct activation of presynaptic NMDA receptors (preNMDARs or an indirect pathway involving activation of somato-dendritic NMDARs followed by passive spread of somatic depolarization along the axon and activation of axonal voltage dependent Ca(2+ channels (VDCCs. Using Ca(2+ imaging and electrophysiology, we searched for preNMDARs by uncaging NMDAR agonists either broadly throughout the whole field or locally at specific axonal locations. Releasing either NMDA or glutamate in the presence of NBQX using short laser pulses elicited current transients that were highly sensitive to the location of the spot and restricted to a small number of varicosities. The signal was abolished in the presence of high Mg(2+ or by the addition of APV. Similar paradigms yielded restricted Ca(2+ transients in interneurons loaded with a Ca(2+ indicator. We found that the synaptic effects of NMDA were not inhibited by blocking VDCCs but were impaired in the presence of the ryanodine receptor antagonist dantrolene. Furthermore, in voltage clamped cells, bath applied NMDA triggers Ca(2+ elevations and induces neurotransmitter release in the axonal compartment. Our results suggest the existence of preNMDARs in developing MLIs and propose their involvement in the NMDA-evoked increase in GABA release by triggering a Ca(2+-induced Ca(2+ release process mediated by presynaptic Ca(2+ stores. Such a mechanism is likely to exert a crucial role in various forms of Ca(2+-mediated synaptic plasticity.

  11. Chemotransduction in the Carotid Body: K+ Current Modulated by Po2 in Type I Chemoreceptor Cells

    Science.gov (United States)

    Lopez-Barneo, Jose; Lopez-Lopez, Jose R.; Urena, Juan; Gonzalez, Constancio

    1988-07-01

    The ionic currents of carotid body type I cells and their possible involvement in the detection of oxygen tension (Po2) in arterial blood are unknown. The electrical properties of these cells were studied with the whole-cell patch clamp technique, and the hypothesis that ionic conductances can be altered by changes in Po2 was tested. The results show that type I cells have voltage-dependent sodium, calcium, and potassium channels. Sodium and calcium currents were unaffected by a decrease in Po2 from 150 to 10 millimeters of mercury, whereas, with the same experimental protocol, potassium currents were reversibly reduced by 25 to 50 percent. The effect of hypoxia was independent of internal adenosine triphosphate and calcium. Thus, ionic conductances, and particularly the O2-sensitive potassium current, play a key role in the transduction mechanism of arterial chemoreceptors.

  12. Role of arachidonic acid in hyposmotic membrane stretch-induced increase in calcium-activated potassium currents in gastric myocytes

    Institute of Scientific and Technical Information of China (English)

    Meng YANG; Wen-xie XU; Xing-lan LI; Hui-ying XU; Jia-bin SUN; Bin MEI; Hai-feng ZHENG; Lian-hua PIAO; De-gang XING; Zhai-liu LI

    2005-01-01

    Aim: To study effects of arachidonic acid (AA) and its metabolites on the hyposmotic membrane stretch-induced increase in calcium-activated potassium currents (IKCa) in gastric myocytes. Methods: Membrane currents were recorded by using a conventional whole cell patch-clamp technique in gastric myocytes isolated with collagenase. Results: Hyposmotic membrane stretch and AA increased both IK(Ca) and spontaneous transient outward currents significantly.Exogenous AA could potentiate the hyposmotic membrane stretch-induced increase in IK(Ca). The hyposmotic membrane stretch-induced increase in IK(Ca) was significantly suppressed by dimethyleicosadienoic acid (100 μmol/L in pipette solution), an inhibitor of phospholipase A2. Nordihydroguaiaretic acid, a lipoxygenase inhibitor, significantly suppressed AA and hyposmotic membrane stretch-induced increases in IK(Ca). External calcium-free or gadolinium chloride, a blocker of stretch-activated channels, blocked the AA-induced increase in IK(Ca) significantly, but it was not blocked by nicardipine, an L-type calcium channel blocker. Ryanodine, a calcium-induced calcium release agonist, completely blocked the AA-induced increase in IK(Ca); however, heparin, a potent inhibitor of inositol triphosphate receptor, did not block the AA-induced increase in IK(Ca). Conclusion:Hyposmotic membrane stretch may activate phospholipase A2, which hydrolyzes membrane phospholipids to ultimately produce AA; AA as a second messenger mediates Ca2+ influx, which triggers Ca2+-induced Ca2+ release and elicits activation of IK(Ca) in gastric antral circular myocytes of the guinea pig.

  13. Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.

    Science.gov (United States)

    Boone, Adrienne N; Senatore, Adriano; Chemin, Jean; Monteil, Arnaud; Spafford, J David

    2014-01-01

    The properties of leaky patch currents in whole cell recording of HEK-293T cells were examined as a means to separate these control currents from expressed sodium and calcium leak channel currents from snail NALCN leak channels possessing both sodium (EKEE) and calcium (EEEE) selectivity filters. Leak currents were generated by the weakening of gigaohm patch seals by artificial membrane rupture using the ZAP function on the patch clamp amplifier. Surprisingly, we found that leak currents generated from the weakened membrane/glass seal can be surprisingly stable and exhibit behavior that is consistent with a sodium leak current derived from an expressible channel. Leaky patch currents differing by 10 fold in size were similarly reduced in size when external sodium ions were replaced with the large monovalent ion NMDG+. Leaky patch currents increased when external Ca2+ (1.2 mM) was lowered to 0.1 mM and were inhibited (>40% to >90%) with 10 µM Gd3+, 100 µM La3+, 1 mM Co2+ or 1 mM Cd2+. Leaky patch currents were relatively insensitive (Sylgard rubber.

  14. Potassium current kinetics in bursting secretory neurons: effects of intracellular calcium.

    Science.gov (United States)

    Martínez, J J; Onetti, C G; García, E; Hernández, S

    1991-11-01

    1. The kinetics of delayed rectifier (IK) and transient potassium (IA) currents and their modification by intracellular calcium ions in bursting X-organ neurons of the crayfish were studied with whole-cell patch-clamp technique. Activation and inactivation kinetics were analyzed according to Hodgkin and Huxley-type equations. 2. IK activates with sigmoidal time course at membrane potentials more positive than -38.4 +/- 3.5 (SD) mV (n = 5), and does not inactivate. The conductance through delayed rectifier channels (gK) is described by the equation gK = GKn2. 3. IA activates at membrane potentials close to the resting potential (-52.2 +/- 4.3 mV, n = 5) and, after a peak, inactivates completely. The conductance through A-channels (gA) can be described by the product of independent activation and inactivation parameters: gA = GAa4b. Both activation and inactivation processes are voltage and time dependent. 4. Steady-state activation of IK and IA as well as inactivation of IA can be described by Boltzmann distributions for single particles with valencies of 2.55 +/- 0.01 (n = 5), 1.60 +/- 0.25 (n = 5), and 3.87 +/- 0.39 (n = 3), respectively. 5. Increasing [Ca2+]i, we observed the following: 1) a considerable inactivation of IK during test pulses, 2) an increase of maximal conductance for IA, 3) a reduction of the valency of IA inactivation gating particle (from 3.87 to 2.27), 4) a reduction of the inactivation time constants of IA, and 5) a shift of the inactivation steady-state curve to more positive membrane potentials.(ABSTRACT TRUNCATED AT 250 WORDS)

  15. Protective Effect of Carvedilol on Abnormality of L-type Calcium Current Induced by Oxygen Free Radical in Cardiomyocytes

    Institute of Scientific and Technical Information of China (English)

    刘念; 喻荣辉; 阮燕菲; 周强; 卜军; 李泱

    2004-01-01

    The protective effect of carvedilol on abnormality of L-type calcium current induced by oxygen free radical in single guinea pig ventricular myocytes was studied. Whole-cell patch clamp technique was used to study the effect of H2 O2 (0.5 mmol/L) on L-type calcium current in single guinea pig ventricular myocytes and the action of pretreatment with carvedilol (0.5 μmol/L). 0.5μmol/L carvedilol had no significant effect on ICa,L and its channel dynamics. In the presence of 0.5 mmol/L H2O2, peak current of ICa,L was reduced significantly (P<0.001), the I-V curve of Ica,L was shifted upward, steady-state activation curve and steady-state deactivation curve of ICa,L were shifted left and recovery time of ICa,L was delayed significantly (P<0. 001). 0. 5 μmol/L carvedilol significantly alleviated the inhibitory effect of H2O2 on ICa,L as compared with that in H2O2 group (P<0.01). In addition, carvedilol reversed the changes of dynamics of ICa,L induced by H2O2. It was concluded that carvedilol could alleviate the abnormality of L-type calcium current induced by oxygen free radical in cardiomyocytes. It shows partly the possible mechanism of the special availability of carvedilol in chronic heart failure.

  16. Zebrafish CaV2.1 Calcium Channels Are Tailored for Fast Synchronous Neuromuscular Transmission

    Science.gov (United States)

    Naranjo, David; Wen, Hua; Brehm, Paul

    2015-01-01

    The CaV2.2 (N-type) and CaV2.1 (P/Q-type) voltage-dependent calcium channels are prevalent throughout the nervous system where they mediate synaptic transmission, but the basis for the selective presence at individual synapses still remains an open question. The CaV2.1 channels have been proposed to respond more effectively to brief action potentials (APs), an idea supported by computational modeling. However, the side-by-side comparison of CaV2.1 and CaV2.2 kinetics in intact neurons failed to reveal differences. As an alternative means for direct functional comparison we expressed zebrafish CaV2.1 and CaV2.2 α-subunits, along with their accessory subunits, in HEK293 cells. HEK cells lack calcium currents, thereby circumventing the need for pharmacological inhibition of mixed calcium channel isoforms present in neurons. HEK cells also have a simplified morphology compared to neurons, which improves voltage control. Our measurements revealed faster kinetics and shallower voltage-dependence of activation and deactivation for CaV2.1. Additionally, recordings of calcium current in response to a command waveform based on the motorneuron AP show, directly, more effective activation of CaV2.1. Analysis of calcium currents associated with the AP waveform indicate an approximately fourfold greater open probability (PO) for CaV2.1. The efficient activation of CaV2.1 channels during APs may contribute to the highly reliable transmission at zebrafish neuromuscular junctions. PMID:25650925

  17. Effect of angiotensin II-induced arterial hypertension on the voltage-dependent contractions of mouse arteries.

    Science.gov (United States)

    Fransen, Paul; Van Hove, Cor E; Leloup, Arthur J A; Schrijvers, Dorien M; De Meyer, Guido R Y; De Keulenaer, Gilles W

    2016-02-01

    Arterial hypertension (AHT) affects the voltage dependency of L-type Ca(2+) channels in cardiomyocytes. We analyzed the effect of angiotensin II (AngII)-induced AHT on L-type Ca(2+) channel-mediated isometric contractions in conduit arteries. AHT was induced in C57Bl6 mice with AngII-filled osmotic mini-pumps (4 weeks). Normotensive mice treated with saline-filled osmotic mini-pumps were used for comparison. Voltage-dependent contractions mediated by L-type Ca(2+) channels were studied in vaso-reactive studies in vitro in isolated aortic and femoral arteries by using extracellular K(+) concentration-response (KDR) experiments. In aortic segments, AngII-induced AHT significantly sensitized isometric contractions induced by elevated extracellular K(+) and depolarization. This sensitization was partly prevented by normalizing blood pressure with hydralazine, suggesting that it was caused by AHT rather than by direct AngII effects on aortic smooth muscle cells. The EC50 for extracellular K(+) obtained in vitro correlated significantly with the rise in arterial blood pressure induced by AngII in vivo. The AHT-induced sensitization persisted when aortic segments were exposed to levcromakalim or to inhibitors of basal nitric oxide release. Consistent with these observations, AngII-treatment also sensitized the vaso-relaxing effects of the L-type Ca(2+) channel blocker diltiazem during K(+)-induced contractions. Unlike aorta, AngII-treatment desensitized the isometric contractions to depolarization in femoral arteries pointing to vascular bed specific responses of arteries to hypertension. AHT affects the voltage-dependent L-type Ca(2+) channel-mediated contraction of conduit arteries. This effect may contribute to the decreased vascular compliance in AHT and explain the efficacy of Ca(2+) channel blockers to reduce vascular stiffness and central blood pressure in AHT.

  18. Effect of chronic stress and mifepristone treatment on voltage-dependent Ca2+ currents in rat hippocampal dentate gyrus.

    NARCIS (Netherlands)

    van Gemert, N.G.; Joëls, M.

    2006-01-01

    Chronic unpredictable stress affects many properties in rat brain. In the dentate gyrus, among other things, increased mRNA expression of the Ca2+ channel alpha1C subunit has been found after 21 days of unpredictable stress in combination with acute corticosterone application (100 nM). In the presen

  19. Analysis and Comparison of Voltage Dependent Charging Strategies for Single-Phase Electric Vehicles in an Unbalanced Danish Distribution Grid

    DEFF Research Database (Denmark)

    Álvarez, Jorge Nájera; Knezovic, Katarina; Marinelli, Mattia

    2016-01-01

    This paper studies four voltage dependent solutions for modulating the charging of multiple Electric Vehicles (EVs) in a real Danish network. Uncontrolled EV charging, especially in grid with high EV penetration, can result in overloaded lines and transformers, low-voltages and other performance......-in on phases with lower voltages are constrained during the charging period. In order to solve instability issues which may occur due to lack of communication between the controllers, several improvements are applied to the aforementioned droop control. Simulation results demonstrate the performance...

  20. Cyclic-AMP regulation of calcium-dependent K channels in an insect central neurone.

    Science.gov (United States)

    David, J A; Pitman, R M

    1996-01-26

    In the cockroach fast coxal depressor motoneurone, either the muscarinic agonist McN-A-343 or dibutyryl cAMP (Db-cAMP) induced a reduction in voltage-dependent outward current. The response to McN is due to suppression of a calcium-dependent potassium current (IK,Ca) produced secondarily to a reduction in voltage-dependent calcium current (ICa). The response to Db-cAMP was investigated in order to establish whether cAMP might mediate the response to McN. ICa was suppressed by 3-isobutyl-1-methylxanthine (IBMX) but not by Db-cAMP. The effects of IBMX were therefore unlikely to be the result of phosphodiesterase inhibition. Since caffeine also suppressed ICa, the observed effect of IBMX is probably due to release of Ca2+ from intracellular stores. IK,Ca, evoked by injection of Ca2+, was reduced by Db-cAMP or forskolin but not by McN. These results indicate that the electrical response to McN in this neurone is not mediated by changes in cAMP.

  1. Gated currents in isolated olfactory receptor neurons of the larval tiger salamander.

    Science.gov (United States)

    Firestein, S; Werblin, F S

    1987-09-01

    The electrical properties of enzymatically isolated olfactory receptor cells were studied with whole-cell patch clamp. Voltage-dependent currents could be separated into three ionic components: a transient inward sodium current, a sustained inward calcium current, and an outward potassium current. Three components of the outward current could be identified by their gating and kinetics: a calcium-dependent potassium current [IK(Ca)], a voltage-dependent potassium current [IK(V)], and a transient potassium current (Ia). Typical resting potentials were near -54 mV, and typical input resistance was 3-6 G omega. Thus, only 3 pA of injected current was required to depolarize the cell to spike threshold near -45 mV. The response to a current step consisted of either a single spike regardless of stimulus strength, or a train of less than 8 spikes, decrementing in amplitude and frequency over approximately equal to 250 msec. Thus, the receptor response cannot be finely graded with stimulus intensity.

  2. Evaluation of calcium alginate gel as electrode material for alternating current iontophoresis of lidocaine using excised rat skin.

    Science.gov (United States)

    Ebisawa, Tomoko; Nakajima, Atsushi; Haida, Haruka; Wakita, Ryo; Ando, Shizuka; Yoshioka, Tomohiko; Ikoma, Toshiyuki; Tanaka, Junzo; Fukayama, Haruhisa

    2014-06-27

    Iontophoresis (IOP) is a noninvasive method of delivering medication transcutaneously through the skin. The electrodes used in this method should tightly fit to rough and irregular surfaces and be biologically safe, easy to handle and prepare, and cost-effective. To satisfy these requirements, calcium alginate gel can be a candidate electrode for IOP. Using calcium alginate gel electrodes, we examined whether lidocaine can be effectively transported across an excised rat skin by squarewave alternating current (AC) application. A squarewave AC with either a 70% or 80% duty cycle was continuously applied to 0.5% calcium alginate gel electrodes containing 10% lidocaine at 10 V and 1 kHz for 60 min. Lidocaine concentration was measured using a spectrophotometer and the temperature of the gel was determined. The lidocaine concentrations for AC-IOP at the 70% and 80% duty cycles were significantly higher than that without AC-IOP. Furthermore, the group with the 80% duty cycle showed higher lidocaine concentrations than the group with the 70% duty cycle. The temperatures of all the groups were lower than 28 °C throughout the procedure. In conclusion, the calcium alginate gel can be used as a possible matrix for IOP electrodes.

  3. Lidocaine stabilizes the open state of CNS voltage-dependent sodium channels.

    Science.gov (United States)

    Castañeda-Castellanos, David R; Nikonorov, Igor; Kallen, Roland G; Recio-Pinto, E

    2002-03-28

    We have previously reported that the lidocaine action is different between CNS and muscle batrachotoxin-modified Na+ channels [Salazar et al., J. Gen. Physiol. 107 (1996) 743-754; Brain Res. 699 (1995) 305-314]. In this study we examined lidocaine action on CNS Na+ currents, to investigate the mechanism of lidocaine action on this channel isoform and to compare it with that proposed for muscle Na+ currents. Na+ currents were measured with the whole cell voltage clamp configuration in stably transfected cells expressing the brain alpha-subunit (type IIA) by itself (alpha-brain) or together with the brain beta(1)-subunit (alphabeta(1)-brain), or the cardiac alpha-subunit (hH1) (alpha-cardiac). Lidocaine (100 microM) produced comparable levels of Na+ current block at positive potentials and of hyperpolarizing shift of the steady-state inactivation curve in alpha-brain and alphabeta(1)-brain Na+ currents. Lidocaine accelerated the rates of activation and inactivation, produced an hyperpolarizing shift in the steady-state activation curve and increased the current magnitude at negative potentials in alpha-brain but not in alphabeta(1)-brain Na+ currents. The lidocaine action in alphabeta(1)-brain resembled that observed in alpha-cardiac Na+ currents. The lidocaine-induced increase in current magnitude at negative potentials and the hyperpolarizing shift in the steady-state activation curve of alpha-brain, are novel effects and suggest that lidocaine treatment does not always lead to current reduction/block when it interacts with Na+ channels. The data are explained by using a modified version of the model proposed by Vedantham and Cannon [J. Gen. Physiol., 113 (1999) 7-16] in which we postulate that the difference in lidocaine action between alpha-brain and alphabeta(1)-brain Na+ currents could be explained by differences in the lidocaine action on the open channel state.

  4. Whole-cell recordings of voltage-gated Calcium, Potassium and Sodium currents in acutely isolated hippocampal pyramidal neurons

    Institute of Scientific and Technical Information of China (English)

    Shuyun Huang; Qing Cai; Weitian Liu; Xiaoling Wang; Tao Wang

    2009-01-01

    Objective:To record Calcium, Potassium and Sodium currents in acutely isolated hippocampal pyramidal neurons. Methods:Hip-pocampal CA3 neurons were freshly isolated by 1 mg protease/3 ml SES and mechanical trituration with polished pipettes of progressively smaller tip diameters. Patch clamp technique in whole-cell mode was employed to record voltage-gated channel currents. Results:The procedure dissociated hippocampal neurons, preserving apical dendrites and several basal dendrites, without impairing the electrical characteristics of the neurons. Whole-cell patch clamp configuration was successfully used to record voltage-gated Ca2+ currents, delayed rectifier K+ current and voltage-gated Na+ currents. Conclusion:Protease combined with mechanical trituration may be used for the dissociation of neurons from rat hippocampus. Voltage-gated channels currents could be recorded using a patch clamp technique.

  5. Effects of Losartan on L-type Calcium Current in Hypertrophied RatMyocytes

    Institute of Scientific and Technical Information of China (English)

    FuLiying; LiYang; ChengLan; WangFang; XiaGuojin; YaoWeixing

    2001-01-01

    Objective To investigate the alterations of L-type calcium current (IcaL) in abdominal aorticligation-induced hypertrophied rat hearts and the effect of losartan on these alterations. METHODS Cardiachypertrophy was induced by abdominal aortic ligation in rats. To record IcaL, whole-cell patch-clamp technique wasused. RESULTS Membrane capacitance was larger in hypertrophied cells (148±29 pF) than in sham-operated cells(102±14 pF, P<0.01) and losartan-treated cells (118±27, P<0.01). The maximal peak IcaL Was increased from-835±124 pA in sham-operated cells to -1404+_417 pA in hypertrophied cells (P<0.01), the corresponding IcaL density was increased from -7.5±1.8 pA.pF1 to -10.5±2.2 pA.pF1 (P<0.01), while they were reduced to -956-2:170pF (P<0.01) and -8.2±1.6 pA.pF1 (P<0.05) respectively in losartan-treated cells. The membrane potential of halfmaximal activation of the hypertrophied cells (-20.6±1.0 mV) shifted to more negative potentials than sham-operatedcells (-15.6±1.6 mV, P<0.01) and lorsartan-treated cells (-17.4±1.0 mV, P<0.01). The slope of the activation curveof hypertrophied cells (5.7±0.4) was decreased slightly than sham-operated cells (6.4±0.5, P<0.05). The membranepotential of half maximal inactivation of hypertrophied cells (-27.6±1.9 mV) shifted to more positive potentials thansham-operated cells (-31.4±2.2 mV, P<0.05). The slope of inactivation curves were not different in the three groups.

  6. Calcium-activated potassium conductance noise in snail neurons.

    Science.gov (United States)

    Westerfield, M; Lux, H D

    1982-11-01

    Current fluctuations were measured in small, 3-6 micrometers-diameter patches of soma membrane in bursting neurons of the snail, Helix pomatia. The fluctuations dramatically increased in magnitude with depolarization of the membrane potential under voltage clamp conditions. Two components of conductance noise were identified in the power spectra calculated from the membrane currents. One component had a corner frequency which increased with depolarization. This component was blocked by intracellular injection of TEA and was relatively insensitive to extracellular calcium levels (as long as the total number of effective divalent cations remained constant). It was identified as fluctuations of the voltage-dependent component of delayed outward current. The second component of conductance noise had a corner frequency which decreased with depolarization. It was relatively unaffected by TEA injection and was reversibly blocked by substitution of extracellular calcium with magnesium, cobalt, or nickel. This second component of noise was identified as fluctuations of the calcium-dependent potassium current. The results suggest that the two components of delayed outward current are conducted through physically distinct channels.

  7. Potassium currents in auditory hair cells of the frog basilar papilla.

    Science.gov (United States)

    Smotherman, M S; Narins, P M

    1999-06-01

    The whole-cell patch-clamp technique was used to identify and characterize ionic currents in isolated hair cells of the leopard frog basilar papilla (BP). This end organ is responsible for encoding the upper limits of a frog's spectral sensitivity (1.25-2.0 kHz in the leopard frog). Isolated BP hair cells are the smallest hair cells in the frog auditory system, with spherical cell bodies typically less than 20 microm in diameter and exhibiting whole-cell capacitances of 4-7 pF. Hair cell zero-current resting potentials (Vz) varied around a mean of -65 mV. All hair cells possessed a non-inactivating, voltage-dependent calcium current (I(Ca)) that activates above a threshold of -55 mV. Similarly all hair cells possessed a rapidly activating, outward, calcium-dependent potassium current (I(K)(Ca)). Most hair cells also possessed a slowly activating, outward, voltage-dependent potassium current (I(K)), which is approximately 80% inactive at the hair cell Vz, and a fast-activating, inward-rectifying potassium current (I(K1)) which actively contributes to setting Vz. In a small subset of cells I(K) was replaced by a fast-inactivating, voltage-dependent potassium current (I(A)), which strongly resembled the A-current observed in hair cells of the frog sacculus and amphibian papilla. Most cells have very similar ionic currents, suggesting that the BP consists largely of one homogeneous population of hair cells. The kinetic properties of the ionic currents present (in particular the very slow I(K)) argue against electrical tuning, a specialized spectral filtering mechanism reported in the hair cells of birds, reptiles, and amphibians, as a contributor to frequency selectivity of this organ. Instead BP hair cells reflect a generalized strategy for the encoding of high-frequency auditory information in a primitive, mechanically tuned, terrestrial vertebrate auditory organ.

  8. Ca2+ channel inhibitor NNC 55-0396 inhibits voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells.

    Science.gov (United States)

    Son, Youn Kyoung; Hong, Da Hye; Li, Hongliang; Kim, Dae-Joong; Na, Sung Hun; Park, Hongzoo; Jung, Won-Kyo; Choi, Il-Whan; Park, Won Sun

    2014-01-01

    We demonstrated the inhibitory effect of NNC 55-0396, a T-type Ca(2+) channel inhibitor, on voltage-dependent K(+) (K(V)) channels in freshly isolated rabbit coronary arterial smooth muscle cells. NNC 55-0396 decreased the amplitude of K(V) currents in a concentration-dependent manner, with an IC(50) of 0.080 μM and a Hill coefficient of 0.76.NNC 55-0396 did not affect steady-state activation and inactivation curves, indicating that the compound does not affect the voltage sensitivity of K(V) channel gating. Both the K(V) currents and the inhibitory effect of NNC 55-0396 on K(V) channels were not altered by depletion of extracellular Ca(2+) or intracellular ATP, suggesting that the inhibitory effect of NNC 55-0396 is independent of Ca(2+)-channel activity and phosphorylation-dependent signaling cascades. From these results, we concluded that NNC 55-0396 dosedependently inhibits K(V) currents, independently of Ca(2+)-channel activity and intracellular signaling cascades.

  9. Reversal of HCN channel voltage dependence via bridging of the S4-S5 linker and Post-S6.

    Science.gov (United States)

    Prole, David L; Yellen, Gary

    2006-09-01

    Voltage-gated ion channels possess charged domains that move in response to changes in transmembrane voltage. How this movement is transduced into gating of the channel pore is largely unknown. Here we show directly that two functionally important regions of the spHCN1 pacemaker channel, the S4-S5 linker and the C-linker, come into close proximity during gating. Cross-linking these regions with high-affinity metal bridges or disulfide bridges dramatically alters channel gating in the absence of cAMP; after modification the polarity of voltage dependence is reversed. Instead of being closed at positive voltage and activating with hyperpolarization, modified channels are closed at negative voltage and activate with depolarization. Mechanistically, this reversal of voltage dependence occurs as a result of selectively eliminating channel deactivation, while retaining an existing inactivation process. Bridging also alters channel activation by cAMP, showing that interaction of these two regions can also affect the efficacy of physiological ligands.

  10. Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.

    Directory of Open Access Journals (Sweden)

    Adrienne N Boone

    Full Text Available The properties of leaky patch currents in whole cell recording of HEK-293T cells were examined as a means to separate these control currents from expressed sodium and calcium leak channel currents from snail NALCN leak channels possessing both sodium (EKEE and calcium (EEEE selectivity filters. Leak currents were generated by the weakening of gigaohm patch seals by artificial membrane rupture using the ZAP function on the patch clamp amplifier. Surprisingly, we found that leak currents generated from the weakened membrane/glass seal can be surprisingly stable and exhibit behavior that is consistent with a sodium leak current derived from an expressible channel. Leaky patch currents differing by 10 fold in size were similarly reduced in size when external sodium ions were replaced with the large monovalent ion NMDG+. Leaky patch currents increased when external Ca2+ (1.2 mM was lowered to 0.1 mM and were inhibited (>40% to >90% with 10 µM Gd3+, 100 µM La3+, 1 mM Co2+ or 1 mM Cd2+. Leaky patch currents were relatively insensitive (<30% to 1 mM Ni2+ and exhibited a variable amount of block with 1 mM verapamil and were insensitive to 100 µM mibefradil or 100 µM nifedipine. We hypothesize that the rapid changes in leak current size in response to changing external cations or drugs relates to their influences on the membrane seal adherence and the electro-osmotic flow of mobile cations channeling in crevices of a particular pore size in the interface between the negatively charged patch electrode and the lipid membrane. Observed sodium leak conductance currents in weak patch seals are reproducible between the electrode glass interface with cell membranes, artificial lipid or Sylgard rubber.

  11. 双苯氟嗪对豚鼠心室肌细胞L-钙电流的影响%Effect of dipfluzine on L-type calcium current in guinea pig ventricular myocytes

    Institute of Scientific and Technical Information of China (English)

    张永健; 李德培; 薛保健; 王永利; 何瑞荣

    2001-01-01

    AIM: To study the effect of dipfluzine (Dip) on L-type calcium current in guinea pig ventricular myocytes.METHODS: Single myocytes were dissociated by enzymatic dissociation method.The current was recorded with the whole-cell configuration of the patch-clamp technique.RESULTS: Dip (0.3-30 μmo l/L)reduced the voltage-dependently activated peak value of ICa-L in a concentration-dependent manner. The characteristics of I-V relationship were not greatly altered by Dip, and the maximal activation voltage of Ica-L in the presence of Dip was not different from that of control.Steady-state activation of Ica-L was not affected markedly, and the half activation potential (V0.5) and the slope factor (κ) in the presence of Dip 3 μmol/L were not markedly different from those of the control. V0.5 value was (-12.8 ±1.7) mV in the control and (-13.2±2.4) mV in the presence of Dip 3 μ mol/L. The κ value was (7.1±0.4) mV in the control and (7.5±0.5) mV in the presence of Dip 3 μmol/L (n=7 cells from 3 hearts, P>0.05). Dip 3 μmol/L markedly shifted the steady-state inactivation curve of ICa-L to the left, and accelerated the voltage-dependent steady-state inactivation of calcium current.V0.5 value was (-19.7±2.4) mV in the control and (-31±6)mV in the presence of Dip 3 μmol/L.The κ value was (3.6±0.3) mV in the control and (1.8±0.2) mV in the presence of Dip 3 μmol/L (n=4cells from 2 hearts, P<0.05). Dip 3 μ mol/L markedly delayed half-recovery time of Ca2 + channel from inactivation from (40±11) to (288±63) ms (n=4, P<0.01).CONCLUSION: Dip mainly acts on the inac ti vated state of L-type calcium channel, accelerates the inactivation of calcium channel, and slows the recovery of calcium channel from inactivated state in guinea pig ventricular myocytes, through which the Ica-L is inhibited.%目的:观察双苯氟嗪(Dip)对豚鼠心室肌细胞L-型钙电流(ICa-L)的影响.方法:酶解法制备单个心室肌细胞.应用全细胞膜片箝技术记录

  12. Lead interferes with calcium entry through membrane pores

    Energy Technology Data Exchange (ETDEWEB)

    Buesselberg, D.; Schirrmacher, K.; Domann, R.; Wiemann, M. [Universitaet-GH Essen, Inst. fuer Physiologie, Essen (Germany)

    1998-06-01

    Calcium is an important intracellular messenger in all cells, represented here by nerve cells and osteoblast-like (OBL) cells. In neurons the intracellular calcium signal is related, e.g., to bioelectric phenomena. In OBL cells the intracellular calcium concentration ([Ca{sup 2+}]{sub i}) plays a role in the intercellular communication via gap junction channels. [Ca{sup 2+}]{sub i} might be affected by lead (Pb{sup 2+}). In the nervous system even low Pb{sup 2+} concentrations impair learning and memory functions. Considering long-term potentiation (LTP) as a model for learning and memory it has been proven that the generation and maintenance of LTP is reduced by Pb{sup 2+} (1-10 {mu}M). As the induction of LTP depends on a rise of [Ca{sup 2+}]{sub i}, we examined the effects of Pb{sup 2+} on [Ca{sup 2+}]{sub i} and on currents through calcium permeable membrane pores in dorsal-root ganglion (DRG) neurons, using calcium measurements (Fura-2/ AM) and whole cell patch clamp techniques. To study the effects of Pb{sup 2+} on intercellular communication via gap junctions we used rat OBL cells investigating interactions of Pb{sup 2+} with electric cell coupling. Furthermore, we examined calcium release activated channel currents (CRACCs) of these cells.Lead (1-10 {mu}M) reduced the stimulated increase of [Ca{sup 2+}]{sub i} in a concentration dependent manner, by reducing both voltage-activated calcium channels (VACCs) and N-methyl-D-aspartate activated calcium channels (NACCs) in neurons. Voltage-activated calcium channel currents (VACCCs) were reduced by Pb{sup 2+} with an IC{sub 50} of 0.46 {mu}M. The effect was quite specific as voltage activated sodium and potassium channel currents were not significantly altered in the same concentration and voltage range. Furthermore, this effect was not voltage dependent and only partly reversible. A 100-fold higher concentration of Pb{sup 2+} (IC{sub 50} of 46 {mu}M) was found for the reduction of NACC currents. A small portion

  13. 线粒体电压依赖性阴离子通道与心血管疾病%Voltage-dependent Anion Channel and Cardiovascular Diseases

    Institute of Scientific and Technical Information of China (English)

    夏晶

    2013-01-01

    电压依赖性阴离子通道(VDAC)是位于线粒体外膜的通道蛋白,是线粒体与细胞质之间转运ATP以及其他代谢产物的主要通道,在线粒体代谢和细胞生长中发挥重要调控作用.近期研究发现,在心肌缺血再灌、糖尿病、心衰、高血压和动脉粥样硬化时,VDAC表达明显增加,引起细胞内钙离子循环紊乱、氧化应激,进而导致细胞凋亡,已成为心血管疾病研究的新热点.本文就VDAC的分子功能,调控及其在心血管疾病中的作用和相关机制进行综述.%The voltage-dependent anion channel (VDAC),a mitochondrial membrane channel protein located in the outer of mitochondrial membrane,is the main pathway between mitochondria and cytoplasm exchanging ADP,ATP,and other metabolites,and plays an important role in mitochondrial metabolism and cell growth.A growing evidence showed that VDAC was increased in cardiovascular diseases including myocardial ischemia and reperfusion,diabetes,heart failure,hypertension and atherosclerosis.The abnormal state of VDAC will result in cell death by inducing calcium cycling dysfunction and oxidative stress.And VDAC has become a hot topic in the field of cardiovascular diseases research.In this article,we will introduce the molecular function and regulation of VDAC and its role in cardiovascular diseases.

  14. Calcium channel blockers in cardiovascular pharmacotherapy.

    Science.gov (United States)

    Godfraind, Theophile

    2014-11-01

    This paper summarizes the pharmacological properties of calcium channel blockers (CCBs), their established therapeutic uses for cardiovascular disorders and the current improvement of their clinical effects through drug combinations. Their identification resulted from study of small molecules including coronary dilators, which were named calcium antagonists. Further experiments showed that they reduced contraction of arteries by inhibiting calcium entry and by interacting with binding sites identified on voltage-dependent calcium channels. This led to the denomination calcium channel blockers. In short-term studies, by decreasing total peripheral resistance, CCBs lower arterial pressure. By unloading the heart and increasing coronary blood flow, CCBs improve myocardial oxygenation. In long-term treatment, the decrease in blood pressure is more pronounced in hypertensive than in normotensive patients. A controversy on the safety of CCBs ended after a large antihypertensive trial (ALLHAT) sponsored by the National Heart, Lung, and Blood Institute. There are two main types of CCBs: dihydopyridine and non-dihydropyridine; the first type is vascular selective. Dihydropyrines are indicated for hypertension, chronic, stable and vasospastic angina. Non-dihydropyridines have the same indications plus antiarrythmic effects in atrial fibrillation or flutter and paroxysmal supraventricular tachycardia. In addition, CCBs reduced newly formed coronary lesions in atherosclerosis. In order to reach recommended blood pressure goals, there is a recent therapeutic move by combination of CCBs with other antihypertensive agents particularly with inhibitors acting at the level of the renin-angiotensin system. They are also combined with statins. Prevention of dementia has been reported in hypertensive patients treated with nitrendipine, opening a way for further studies on CCBs' beneficial effect in cognitive deterioration associated with aging.

  15. The effect of verapamil and diltiazem on cardiac stimulant effect of adrenaline and calcium chloride on isolated frog heart

    Directory of Open Access Journals (Sweden)

    Lakhavat Sudhakar, Naveen Kumar T, Tadvi NA, Venkata Rao Y

    2013-04-01

    Full Text Available Background: Calcium channel blockers block voltage dependent L-type of calcium channel and thus reduce the frequency of opening of these channels in response to depolarization. The result is a marked decrease in transmembrane calcium current associated with long lasting relaxation of vascular smooth muscle, reduction in contractility in cardiac muscle, decrease in pacemaker activity in the SA node and decrease in conduction velocity in the AV node. Among Calcium channel blockers verapamil, is cardio selective, nifedipine is vascular smooth muscle selective, while diltiazem exhibits intermediate selectivity. Methods: In the present study, the effect of two Ca++ channel blocker, Verapamil and Diltiazem were compared on the isolated frog heart by using adrenaline & calcium chloride as standard on frog heart contractility. Results and conclusion: Adrenaline and calcium chloride increased the amplitude of contraction of isolated perfused frog heart. The L- type of Ca2+ channel blockers verapamil and diltiazem produced dose dependent (2mg, 4mg, 8mg, and 16mg reduction in the amplitude of contraction produced by calcium chloride in isolated perfused frog heart. There was no statistical significant difference (p > 0.05 between the inhibitory effect of diltiazem and verapamil on calcium chloride induced contraction of isolated frog heart.

  16. The voltage dependence of GABAA receptor gating depends on extracellular pH.

    Science.gov (United States)

    Pytel, Maria; Mercik, Katarzyna; Mozrzymas, Jerzy W

    2005-11-28

    Recent studies have indicated that changes in extracellular pH and in membrane voltage affect the gamma-amino-n-butyric acid type A receptor gating mainly by altering desensitization and binding. To test whether the effects of membrane potential and pH are additive, their combined actions were investigated. By analyzing the current responses to rapid gamma-amino-n-butyric acid applications, we found that the current to voltage relationship was close to linear at acid pH but the increasing pH induced an inward rectification. Desensitization was enhanced at depolarizing potentials, but this strongly depended on pH, being weak at acidic and strong at basic pH values. A similar trend was observed for the onset rate of responses to saturating gamma-amino-n-butyric acid concentration. These data provide evidence that the voltage sensitivity of GABAA receptors depends on extracellular pH.

  17. The voltage dependence of GABAA receptor gating depends on extracellular pH

    Science.gov (United States)

    Pytel, Maria; Mercik, Katarzyna; Mozrzymas, Jerzy W.

    2007-01-01

    Recent studies have indicated that changes in extracellular pH and in membrane voltage affect the γ-amino-n-butyric acid type A receptor gating mainly by altering desensitization and binding. To test whether the effects of membrane potential and pH are additive, their combined actions were investigated. By analyzing the current responses to rapid γ-amino-n-butyric acid applications, we found that the current to voltage relationship was close to linear at acid pH but the increasing pH induced an inward rectification. Desensitization was enhanced at depolarizing potentials, but this strongly depended on pH, being weak at acidic and strong at basic pH values. A similar trend was observed for the onset rate of responses to saturating γ-amino-n-butyric acid concentration. These data provide evidence that the voltage sensitivity of GABAA receptors depends on extracellular pH. PMID:16272885

  18. Extracellular Linkers Completely Transplant the Voltage Dependence from Kv1.2 Ion Channels to Kv2.1.

    Science.gov (United States)

    Elinder, Fredrik; Madeja, Michael; Zeberg, Hugo; Århem, Peter

    2016-10-18

    The transmembrane voltage needed to open different voltage-gated K (Kv) channels differs by up to 50 mV from each other. In this study we test the hypothesis that the channels' voltage dependences to a large extent are set by charged amino-acid residues of the extracellular linkers of the Kv channels, which electrostatically affect the charged amino-acid residues of the voltage sensor S4. Extracellular cations shift the conductance-versus-voltage curve, G(V), by interfering with these extracellular charges. We have explored these issues by analyzing the effects of the divalent strontium ion (Sr(2+)) on the voltage dependence of the G(V) curves of wild-type and chimeric Kv channels expressed in Xenopus oocytes, using the voltage-clamp technique. Out of seven Kv channels, Kv1.2 was found to be most sensitive to Sr(2+) (50 mM shifted G(V) by +21.7 mV), and Kv2.1 to be the least sensitive (+7.8 mV). Experiments on 25 chimeras, constructed from Kv1.2 and Kv2.1, showed that the large Sr(2+)-induced G(V) shift of Kv1.2 can be transferred to Kv2.1 by exchanging the extracellular linker between S3 and S4 (L3/4) in combination with either the extracellular linker between S5 and the pore (L5/P) or that between the pore and S6 (LP/6). The effects of the linker substitutions were nonadditive, suggesting specific structural interactions. The free energy of these interactions was ∼20 kJ/mol, suggesting involvement of hydrophobic interactions and/or hydrogen bonds. Using principles from double-layer theory we derived an approximate linear equation (relating the voltage shifts to altered ionic strength), which proved to well match experimental data, suggesting that Sr(2+) acts on these channels mainly by screening surface charges. Taken together, these results highlight the extracellular surface potential at the voltage sensor as an important determinant of the channels' voltage dependence, making the extracellular linkers essential targets for evolutionary selection.

  19. Analysis of the effects of calcium or magnesium on voltage-clamp currents in perfused squid axons bathed in solutions of high potassium.

    Science.gov (United States)

    Rojas, E; Taylor, R E; Atwater, I; Bezanilla, F

    1969-10-01

    Isolated axons from the squid, Dosidicus gigas, were internally perfused with potassium fluoride solutions. Membrane currents were measured following step changes of membrane potential in a voltage-clamp arrangement with external isosmotic solution changes in the order: potassium-free artificial seawater; potassium chloride; potassium chloride containing 10, 25, 40 or 50, mM calcium or magnesium; and potassium-free artificial seawater. The following results suggest that the currents measured under voltage clamp with potassium outside and inside can be separated into two components and that one of them, the predominant one, is carried through the potassium system. (a) Outward currents in isosmotic potassium were strongly and reversibly reduced by tetraethylammonium chloride. (b) Without calcium or magnesium a progressive increase in the nontime-dependent component of the currents (leakage) occurred. (c) The restoration of calcium or magnesium within 15-30 min decreases this leakage. (d) With 50 mM divalent ions the steady-state current-voltage curve was nonlinear with negative resistance as observed in intact axons in isosmotic potassium. (e) The time-dependent components of the membrane currents were not clearly affected by calcium or magnesium. These results show a strong dependence of the leakage currents on external calcium or magnesium concentration but provide no support for the involvement of calcium or magnesium in the kinetics of the potassium system.

  20. Voltage-dependent effects of barnidipine in rat vascular smooth muscle.

    Science.gov (United States)

    Wegener, J W; Korstanje, C; Nawrath, H

    2003-08-01

    The effects of the dihydropyridine nifedipine and its more lipophilic congener, barnidipine, were investigated in smooth muscle preparations from the rat in resting and depolarizing conditions. Both drugs relaxed precontracted aortic rings more potently in depolarizing conditions, barnidipine being more potent than nifedipine. Currents through Ca2+ channels in rat vascular smooth muscle cells (A7r5) and in isolated rat cardiomyocytes were reduced more potently by both drugs at a holding potential of -40 mV than at -80 mV. However, barnidipine and nifedipine were more effective in reducing the current in A7r5 cells than in cardiomyocytes. The IC(50) obtained in aortic rings and in A7r5 cells were similar for barnidipine but an order of magnitude different for nifedipine. The results show that, in depolarizing conditions, barnidipine was more effective than nifedipine. It is suggested that the higher potency of barnidipine acting in vascular smooth muscle is related to both a higher affinity to the inactivated state of vascular Ca2+ channels and to a more lipophilic property as compared with nifedipine.

  1. Involvement of inositol 1,4,5-trisphosphate formation in the voltage-dependent regulation of the Ca(2+) concentration in porcine coronary arterial smooth muscle cells.

    Science.gov (United States)

    Yamamura, Hisao; Ohya, Susumu; Muraki, Katsuhiko; Imaizumi, Yuji

    2012-08-01

    The involvement of inositol 1,4,5-trisphosphate (IP(3)) formation in the voltage-dependent regulation of intracellular Ca(2+) concentration ([Ca(2+)](i)) was examined in smooth muscle cells of the porcine coronary artery. Slow ramp depolarization from -90 to 0 mV induced progressive [Ca(2+)](i) increase. The slope was reduced or increased in the presence of Cd(2+) or (±)-1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-[trifluoromethyl]-phenyl)pyridine-3-carboxlic acid methyl ester (Bay K 8644), respectively. The decrease in [Ca(2+)](i) via the membrane hyperpolarization induced by K(+) channel openers (levcromakalim and Evans blue) under current clamp was identical to that under voltage clamp. The step hyperpolarization from -40 to -80 mV reduced [Ca(2+)](i) uniformly over the whole-cell area with a time constant of ∼10 s. The [Ca(2+)](i) at either potential was unaffected by heparin, an inhibitor of IP(3) receptors. Alternatively, [Ca(2+)](i) rapidly increased in the peripheral regions by depolarization from -80 to 0 mV and stayed at that level (∼400 nM) during a 60-s pulse. When the pipette solution contained IP(3) pathway blockers [heparin, 2-aminoethoxydiphenylborate, xestospongin C, or 1-[6-[((17β)-3-methoxyestra-1,3,5[10]-trien-17-yl)amino]hexyl]-1H-pyrrole-2,5-dione (U73122)], the peak [Ca(2+)](i) was unchanged, but the sustained [Ca(2+)](i) was gradually reduced by ∼250 nM within 30 s. In the presence of Cd(2+), a long depolarization period slightly increased the [Ca(2+)](i), which was lower than that in the presence of heparin alone. In coronary arterial myocytes, the sustained increase in the [Ca(2+)](i) during depolarization was partly caused by the Ca(2+) release mediated by the enhanced formation of IP(3). The initial [Ca(2+)](i) elevation triggered by the Ca(2+) influx though voltage-dependent Ca(2+) channels may be predominantly responsible for the activation of phospholipase C for IP(3) formation.

  2. Characterization and functional analysis of voltage-dependent anion channel 1 (VDAC1) from orange-spotted grouper (Epinephelus coioides).

    Science.gov (United States)

    Shi, Yan; Zhao, Zhe; Hong, Xiaoyou; Chen, Kunci; Zhu, Xinping

    2014-07-01

    The voltage-dependent anion channel (VDAC) is a highly conserved integral protein of mitochondria in different eukaryotic species. It forms a selective channel in the mitochondrial outer membrane that serves as the controlled pathway for small metabolites and ions. In this study, a VDAC gene, EcVDAC1, was isolated from orange-spotted grouper (Epinephelus coioides). The EcVDAC1 exhibits ubiquitous expression in various tissues of orange-spotted grouper and is upregulated in liver, gill, and spleen after stimulation with lipopolysaccharides (LPS). Subcellular localization analysis shows that the EcVDAC1 protein colocalized with the mitochondria. A caspase-3 assay demonstrates that overexpression of the EcVDAC1 induced apoptotic cell death in fathead minnow cells. The data presented in this study provide new information regarding the relationship between LPS and the EcVDAC1 gene, suggesting that the fish VDAC1 gene may play an important role in antibacterial immune response.

  3. Temperature and bias voltage dependence of Co/Pd multilayer-based magnetic tunnel junctions with perpendicular magnetic anisotropy

    Energy Technology Data Exchange (ETDEWEB)

    Kugler, Zoe, E-mail: zkugler@physik.uni-bielefeld.d [Bielefeld University, Department of Physics, Universitaetsstr. 25, 33615 Bielefeld (Germany); Drewello, Volker; Schaefers, Markus; Schmalhorst, Jan; Reiss, Guenter; Thomas, Andy [Bielefeld University, Department of Physics, Universitaetsstr. 25, 33615 Bielefeld (Germany)

    2011-01-15

    Temperature- and bias voltage-dependent transport measurements of magnetic tunnel junctions (MTJs) with perpendicularly magnetized Co/Pd electrodes are presented. Magnetization measurements of the Co/Pd multilayers are performed to characterize the electrodes. The effects of the Co layer thickness in the Co/Pd bilayers, the annealing temperature, the Co thickness at the MgO barrier interface, and the number of bilayers on the tunneling magneto resistance (TMR) effect are investigated. TMR-ratios of about 11% at room temperature and 18.5% at 13 K are measured and two well-defined switching fields are observed. The results are compared to measurements of MTJs with Co-Fe-B electrodes and in-plane anisotropy.

  4. Modulation of intracellular calcium mobilization and GABAergic currents through subtype-specific metabotropic glutamate receptors in neonatal rat hippocampus.

    Science.gov (United States)

    Taketo, M; Matsuda, H

    2010-01-15

    Group I metabotropic glutamate receptors (mGluRs) are coupled to phosphoinositide hydrolysis, and are thought to modulate neuronal excitability, by mobilizing intracellular Ca(2+). Difference in Ca(2+) mobilization among subclasses of the receptors has been reported, and regarded as a possible cause of variant neuronal modifications. In hippocampal interneurons, several subclasses of mGluRs including mGluR1 and mGluR5 have been immunohistochemically identified. The subclass-specific physiological effects of mGluRs on neuronal transmission in hippocampus, however, have not been fully elucidated. In the present study, effects of group I mGluR agonist, (S)-3,5-dihydroxyphenylglycine (DHPG) on intracellular calcium concentration were examined in hippocampal interneurons. Application of DHPG increased fluorescence ratio in neonatal CA3 stratum oriens/alveus interneurons. The DHPG-induced calcium mobilization was markedly inhibited by mGluR1-specific antagonist, cyclopropan[b]chromen-1a-carboxylate (CPCCOEt). Inhibition of the calcium elevation by mGluR5-specific antagonist, 6-methyl-2-(phenylazo)-3-pyrindol (MPEP), was weaker than that of CPCCOEt. The fluorescence ratio was not significantly changed by application of mGluR5-specific agonist, (RS)-2-chloro-5-hydroxyphenylglycine (CHPG). DHPG induced calcium responses in CA1 interneurons as in CA3, and the responses were partially inhibited by MPEP treatment. Effects of group I mGluR agonist and antagonist were also investigated, on GABA(A) receptor-mediated spontaneous inhibitory postsynaptic currents (sIPSCs) in CA3 pyramidal neurons. The GABAergic sIPSCs were facilitated by DHPG perfusion, and the potentiation was reduced by CPCCOEt, and less distinctly by MPEP. The sIPSCs were not significantly potentiated by CHPG application. These results indicate that mGluR1 is functional in hippocampal interneurons, and DHPG exerts its effect mainly through this receptor at early developmental period.

  5. Voltage-dependent K channels in protoplasts of trap-lobe cells of Dionaea muscipula.

    Science.gov (United States)

    Iijima, T; Hagiwara, S

    1987-01-01

    The outward rectification of the K+ current in mesophyll cell protoplasts from trap-lobes of Dionaea muscipula was studied with the patch-clamp technique. The rectification had instantaneous and time-dependent components. Changes in [K+]i strongly affected the conductance voltage relation of the plasma membrane while changes in [K+]o had little effect on the relation. Thus, the outward rectification depends on the membrane voltage and the concentration of intracellular K+. Corresponding single-channel activities were observed both in the intact membrane (cell-attached recording) and in excised patches. The single-channel conductance was about 3.3 pS with symmetrical solutions containing 30 mM K+.

  6. The calmodulin inhibitor CGS 9343B inhibits voltage-dependent K{sup +} channels in rabbit coronary arterial smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Hongliang; Hong, Da Hye; Kim, Han Sol; Kim, Hye Won [Institute of Medical Sciences, Department of Physiology, Kangwon National University School of Medicine, Chuncheon, 200-701 (Korea, Republic of); Jung, Won-Kyo [Department of Biomedical Engineering, Center for Marine-Integrated Biomedical Technology (BK21 Plus), Pukyong National University, Busan 608-737 (Korea, Republic of); Na, Sung Hun [Institute of Medical Sciences, Department of Obstetrics and Gynecology, Kangwon National University Hospital, School of Medicine, Kangwon National University, Chuncheon, 200-701 (Korea, Republic of); Jung, In Duk; Park, Yeong-Min [Department of Immunology, Lab of Dendritic Cell Differentiation and Regulation, College of Medicine, Konkuk University, Chungju 380-701 (Korea, Republic of); Choi, Il-Whan, E-mail: cihima@inje.ac.kr [Department of Microbiology, Inje University College of Medicine, Busan, 614-735 (Korea, Republic of); Park, Won Sun, E-mail: parkws@kangwon.ac.kr [Institute of Medical Sciences, Department of Physiology, Kangwon National University School of Medicine, Chuncheon, 200-701 (Korea, Republic of)

    2015-06-15

    We investigated the effects of the calmodulin inhibitor CGS 9343B on voltage-dependent K{sup +} (Kv) channels using whole-cell patch clamp technique in freshly isolated rabbit coronary arterial smooth muscle cells. CGS 9343B inhibited Kv currents in a concentration-dependent manner, with a half-maximal inhibitory concentration (IC{sub 50}) value of 0.81 μM. The decay rate of Kv channel inactivation was accelerated by CGS 9343B. The rate constants of association and dissociation for CGS 9343B were 2.77 ± 0.04 μM{sup −1} s{sup −1} and 2.55 ± 1.50 s{sup −1}, respectively. CGS 9343B did not affect the steady-state activation curve, but shifted the inactivation curve toward to a more negative potential. Train pulses (1 or 2 Hz) application progressively increased the CGS 9343B-induced Kv channel inhibition. In addition, the inactivation recovery time constant was increased in the presence of CGS 9343B, suggesting that CGS 9343B-induced inhibition of Kv channel was use-dependent. Another calmodulin inhibitor, W-13, did not affect Kv currents, and did not change the inhibitory effect of CGS 9343B on Kv current. Our results demonstrated that CGS 9343B inhibited Kv currents in a state-, time-, and use-dependent manner, independent of calmodulin inhibition. - Highlights: • We investigated the effects of CGS 9394B on Kv channels. • CGS 9394B inhibited Kv current in a state-, time-, and use-dependent manner. • Caution is required when using CGS 9394B in vascular function studies.

  7. Regulation of granule cell excitability by a low-threshold calcium spike in turtle olfactory bulb

    DEFF Research Database (Denmark)

    Pinato, Giulietta; Midtgaard, Jens

    2003-01-01

    Granule cells excitability in the turtle olfactory bulb was analyzed using whole cell recordings in current- and voltage-clamp mode. Low-threshold spikes (LTSs) were evoked at potentials that are subthreshold for Na spikes in normal medium. The LTSs were evoked from rest, but hyperpolarization...... of the cell usually increased their amplitude so that they more easily boosted Na spike initiation. The LTS persisted in the presence of TTX but was antagonized by blockers of T-type calcium channels. The voltage dependence, kinetics, and inactivation properties of the LTS were characteristic of a low......-threshold calcium spike. The threshold of the LTS was slightly above the resting potential but well below the Na spike threshold, and the LTS was often evoked in isolation in normal medium. Tetraethylammonium (TEA) and 4-aminopyridine (4-AP) had only minimal effects on the LTS but revealed the presence of a high...

  8. Flow- and voltage-dependent blocking effect of ethosuximide on the inward rectifier K⁺ (Kir2.1) channel.

    Science.gov (United States)

    Huang, Chiung-Wei; Kuo, Chung-Chin

    2015-08-01

    Absence seizures are manifestations of abnormal thalamocortical oscillations characterized by spike-and-wave complexes in EEG. Ethosuximide (ETX) is one of the principal medications against absence seizures. We investigate the effect of ETX on the Kir2.1 channel, a prototypical inward rectifier K(+) channel possibly playing an important role in the setting of neuronal membrane potential. We demonstrate that the outward currents of Kir2.1 channels are significantly inhibited by intracellular ETX. We further show that the movement of neutral molecule ETX in the Kir2.1 channel is accompanied by ∼1.2 K(+), giving rise to the vivid voltage dependence of ETX unbinding rate. Moreover, the apparent affinity (K d ) of ETX in the channels are decreased by single-point mutations involving M183, E224, and S165, and especially by double mutations involving T141/S165, which always also disrupt the flux-coupling feature of ETX block. Molecular dynamics simulation demonstrates narrowing of the pore at ∼D172 by binding of ETX to S165 or T141. ETX block of the Kir2.1 channels may cause a modest but critical depolarization of the relevant neurons, decreasing available T-type Ca(2+) channels and consequently lessening pathological thalamocortical burst discharges.

  9. Action of angiotensin II, 5-hydroxytryptamine and adenosine triphosphate on ionic currents in single ear artery cells of the rabbit.

    Science.gov (United States)

    Hughes, A D; Bolton, T B

    1995-10-01

    1. Angiotensin II, 5-hydroxytryptamine (5-HT) and adenosine triphosphate (ATP) evoked a transient inward current in isolated single car artery cells of rabbit held at -60 mV by whole cell voltage clamp in physiological saline using a KCL-containing pipette solution. Under these conditions agonist did not activate a calcium-dependent potassium current. 2. Responses to each agonist were transient and desensitized rapidly. Inward current at -60 mV holding potential was not abolished by blockade of voltage-dependent calcium channels or by buffering intracellular calcium with BAPTA, a calcium chelator, or following depletion of intracellular calcium stores with ryanodine. 3. The shape of the current-voltage relationships and the reversal potentials of the current induced by angiotensin II, 5-HT and ATP were similar under a variety of ionic conditions. Agonist-induced current was unaffected by replacing intracellular chloride with citrate ions or by replacing intracellular sodium with caesium or extracellular sodium with barium or calcium. Replacement of extracellular sodium with Tris shifted the reversal potential in all cases by around 30 mV negatively. 4. These data suggest that angiotensin II, 5-HT and ATP activate similar cationic conductances which are relatively non-selective allowing mono- and divalent cations to cross the smooth muscle cell membrane. These channels may allow the influx of calcium under physiological conditions.

  10. Kinetic study of N-type calcium current modulation by delta-opioid receptor activation in the mammalian cell line NG108-15.

    Science.gov (United States)

    Toselli, M; Tosetti, P; Taglietti, V

    1999-01-01

    The voltage-dependent inhibition of N-type Ca2+ channel current by the delta-opioid agonist [D-pen2, D-pen5]-enkephalin (DPDPE) was investigated in the mammalian cell line NG108-15 with 10 microM nifedipine to block L-type channels, with whole-cell voltage clamp methods. In in vitro differentiated NG108-15 cells DPDPE reversibly decreased omega-conotoxin GVIA-sensitive Ba2+ currents in a concentration-dependent way. Inhibition was maximal with 1 microM DPDPE (66% at 0 mV) and was characterized by a slowing of Ba2+ current activation at low test potentials. Both inhibition and kinetic slowing were attenuated at more positive potentials and could be relieved up to 90% by strong conditioning depolarizations. The kinetics of removal of inhibition (de-inhibition) and of its retrieval (re-inhibition) were also voltage dependent. Both de-inhibition and re-inhibition were single exponentials and, in the voltage range from -20 to +10 mV, had significantly different time constants at a given membrane potential, the time course of re-inhibition being faster than that of de-inhibition. The kinetics of de-inhibition at -20 mV and of reinhibition at -40 mV were also concentration dependent, both processes becoming slower at lower agonist concentrations. The rate of de-inhibition at +80/+120 mV was similar to that of Ca2+ channel activation at the same potentials measured during application of DPDPE (approximately 7 ms), both processes being much slower than channel activation in controls (<1 ms). Moreover, the amplitude but not the time course of tail currents changed as the depolarization to +80/+120 mV was made longer. The state-dependent properties of DPDPE Ca2+ channel inhibition could be simulated by a model that assumes that inhibition by DPDPE results from voltage- and concentration-dependent binding of an inhibitory molecule to the N-type channel. PMID:10233071

  11. Accelerated inactivation of the L-type calcium current due to a mutation in CACNB2b underlies Brugada syndrome

    DEFF Research Database (Denmark)

    Cordeiro, Jonathan M; Marieb, Mark; Pfeiffer, Ryan

    2009-01-01

    Recent studies have demonstrated an association between mutations in CACNA1c or CACNB2b and Brugada syndrome (BrS). Previously described mutations all caused a loss of function secondary to a reduction of peak calcium current (I(Ca)). We describe a novel CACNB2b mutation associated with Br...... revealed brief episodes of very rapid ventricular tachycardia. He was also diagnosed with vasovagal syncope. Genomic DNA was isolated from lymphocytes. All exons and intron borders of 15 ion channel genes were amplified and sequenced. The only mutation uncovered was a missense mutation (T11I) in CACNB2b...... that the faster current decay results in a loss-of-function responsible for the Brugada phenotype...

  12. Sensitization of voltage activated calcium channel currents for capsaicin in nociceptive neurons by tumor-necrosis-factor-alpha.

    Science.gov (United States)

    Hagenacker, T; Czeschik, J C; Schäfers, M; Büsselberg, D

    2010-01-15

    It is known that application of tumor-necrosis-factor-alpha (TNF-alpha) sensitizes neuronal calcium channels for heat stimuli in rat models of neuropathic pain. This study examines whether TNF-alpha modulates the capsaicin-induced effects after transient receptor potential vanilloid (TRPV)-1 receptor activation on voltage activated calcium channel currents (I(Ca(V))). TRPV-1 receptors are activated by heat and play an important role in the pathogenesis of thermal hyperalgesia in neuropathic pain syndromes, while voltage activated channels are essential for transmission of neuronal signals. Eliciting I(Ca(V)) in DRG neurons of rats by a depolarization from the resting potential to 0 mV, TNF-alpha (100 ng/ml) reduces I(Ca(V)) by 16.9+/-2.2%, while capsaicin (0.1 microM) decreases currents by 27+/-4.3%. Pre-application of TNF-alpha (100 ng/ml) for 24h results in a sensitization of I(Ca(V)) to capsaicin (0.1 microM) with a reduction of 42.8+/-4.4% mediated by TRPV-1. While L-type (36.6+/-5.2%) and P/Q-type currents (35.6+/-4.1%) are also sensitized by TRPV-1 activation, N-type channel currents are most sensitive (74.5+/-7.3%). The capsaicin-induced shift towards the hyperpolarizing voltage range does not occur when TNF-alpha is applied. Summarizing, TNF-alpha sensitizes nociceptive neurons for capsaicin.

  13. A Drosophila mutation that eliminates a calcium-dependent potassium current.

    Science.gov (United States)

    Elkins, T; Ganetzky, B; Wu, C F

    1986-11-01

    A mutation of Drosophila, slowpoke (slo), specifically abolishes a Ca2+-dependent K+ current, IC, from dorsal longitudinal flight muscles of adult flies. Other K+ currents remain normal, providing evidence that IC is mediated by a molecularly distinguishable set of channels. The pharmacological properties of IC are similar to those of Ca2+-dependent currents in some vertebrate cells. The muscle action potential was significantly lengthened in slo flies, indicating that IC plays the major role in its repolarization.

  14. A Drosophila mutation that eliminates a calcium-dependent potassium current.

    OpenAIRE

    Elkins, T; Ganetzky, B; Wu, C F

    1986-01-01

    A mutation of Drosophila, slowpoke (slo), specifically abolishes a Ca2+-dependent K+ current, IC, from dorsal longitudinal flight muscles of adult flies. Other K+ currents remain normal, providing evidence that IC is mediated by a molecularly distinguishable set of channels. The pharmacological properties of IC are similar to those of Ca2+-dependent currents in some vertebrate cells. The muscle action potential was significantly lengthened in slo flies, indicating that IC plays the major role...

  15. The beta-adrenergic blocker carvedilol restores L-type calcium current in a myocardial infarction model of rabbit

    Institute of Scientific and Technical Information of China (English)

    LI Xia; HUANG Cong-xin; JIANG Hong; CAO Feng; WANG Teng

    2005-01-01

    Background Carvedilol, an antagonist of α1- and β-adrenergic receptors, has shown efficacy in reducing all-cause death and arrhythmia death for ischemic heart disease and congestive heart failure in several large-scale trials. It has been found to prevent ventricular remodeling, and recently was reported to reverse down-regulation of Na+ channel in a chronic heart failure model. This study was conducted to investigate whether carvedilol could reverse the ion remodeling in a myocardial infarction model of rabbit.Methods After the procedure of coronary ligation, animals were randomized to placebo or carvedilol treatment (5 mg/kg). Action potentials, L-type calcium current (Ica L) and the effect of isoproterenol stimulation on Ica L were measured using whole-cell patch method. Evaluation of the expression of calcium channel subunits was carried out by RT-PCR and Western blot. Results The results indicate that mean peak Ica L densities (pA/pF) at +10 mV was reduced in postinfarction myocytes (5.33±0.45, n=25) compared to sham myocytes (6.52±0.21, n=20). Treatment of myocardial infarction rabbits with carvedilol could restore it partially (5.91±0.39, n=20, P<0.05). However, steady-state activation parameters were similar in three groups. With stimulation by isoproterenol (1 μmol/L) Ica L increased in all three groups, but the increase was smaller in postinfarction myocytes. mRNA levels of calcium channel subunit CaA1 gene was decreased but CaB2a, CaB2b and CaB3 mRNA levels did not change after MI. Corresponding change in CaA1 protein was also observed. Conclusions The results demonstrate that carvedilol restores Ica L density and reverse the downregulation of CaA1 postinfarction.

  16. Lysophosphatidic acid increases the electrophysiological instability of adult rabbit ventricular myocardium by augmenting L-type calcium current.

    Directory of Open Access Journals (Sweden)

    Yong Wei

    Full Text Available Lysophosphatidic acid (LPA has diverse actions on the cardiovascular system and is widely reported to modulate multiple ion currents in some cell types. However, little is known about its electrophysiological effects on cardiac myocytes. This study investigated whether LPA has electrophysiological effects on isolated rabbit myocardial preparations. The results indicate that LPA prolongs action potential duration at 90% repolarization (APD(90 in a concentration- and frequency-dependent manner in isolated rabbit ventricular myocytes. The application of extracellular LPA significantly increases the coefficient of APD(90 variability. LPA increased L-type calcium current (I(Ca,L density without altering its activation or deactivation properties. In contrast, LPA has no effect on two other ventricular repolarizing currents, the transient outward potassium current (I(to and the delayed rectifier potassium current (I(K. In arterially perfused rabbit left ventricular wedge preparations, the monophasic action potential duration, QT interval, and Tpeak-end are prolonged by LPA. LPA treatment also significantly increases the incidence of ventricular tachycardia induced by S(1S(2 stimulation. Notably, the effects of LPA on action potentials and I(Ca,L are PTX-sensitive, suggesting LPA action requires a G(i-type G protein. In conclusion, LPA prolongs APD and increases electrophysiological instability in isolated rabbit myocardial preparations by increasing I(Ca,L in a G(i protein-dependent manner.

  17. Effect of Calcium-Activated Chloride Current Blockade on the Delayed Afterdepolarizations. Simulation Study

    Science.gov (United States)

    2007-11-02

    dog and rabbit ventricular cells [4, 6]. The ionic nature of Iti is still subject to debate. Several authors propose two ionic currents to contribute to...activated Cl currents in the heart: a computer model. Computers in Cardiology 1999; 26: 109-112. Julio Gomis-Tena Dolz U.P.V. Departamento de Ingeniería

  18. Midbrain dopaminergic neurons generate calcium and sodium currents and release dopamine in the striatum of pups

    Directory of Open Access Journals (Sweden)

    Diana Carolina Ferrari

    2012-03-01

    Full Text Available Midbrain dopaminergic neurons (mDA neurons are essential for the control of diverse motor and cognitive behaviors. However, our understanding of the activity of immature mDA neurons is rudimentary. Rodent mDA neurons migrate and differentiate early in embryonic life and dopaminergic axons enter the striatum and contact striatal neurons a few days before birth, but when these are functional is not known. Here, we recorded Ca2+ transients and Na+ spikes from embryonic (E16-E18 and early postnatal (P0-P7 mDA neurons with dynamic two photon imaging and patch clamp techniques in slices from tyrosine hydroxylase-GFP mice, and measured evoked dopamine release in the striatum with amperometry. We show that half of identified E16-P0 mDA neurons spontaneously generate non-synaptic, intrinsically-driven Ca2+ spikes and Ca2+ plateaus mediated by N- and L-type voltage-gated Ca2+ channels. Starting from E18-P0, half of the mDA neurons also reliably generate overshooting Na+ spikes with an abrupt maturation at birth (P0 = E19. At that stage (E18-P0, dopaminergic terminals release dopamine in a calcium-dependent manner in the striatum in response to local stimulation. We propose that the intrinsic spontaneous activity of mouse mDA neurons may impact the development/activity of the striatal network from birth.

  19. Reduced KCNQ4-encoded voltage-dependent potassium channel activity underlies impaired β-adrenoceptor-mediated relaxation of renal arteries in hypertension.

    Science.gov (United States)

    Chadha, Preet S; Zunke, Friederike; Zhu, Hai-Lei; Davis, Alison J; Jepps, Thomas A; Olesen, Søren P; Cole, William C; Moffatt, James D; Greenwood, Iain A

    2012-04-01

    KCNQ4-encoded voltage-dependent potassium (Kv7.4) channels are important regulators of vascular tone that are severely compromised in models of hypertension. However, there is no information as to the role of these channels in responses to endogenous vasodilators. We used a molecular knockdown strategy, as well as pharmacological tools, to examine the hypothesis that Kv7.4 channels contribute to β-adrenoceptor-mediated vasodilation in the renal vasculature and underlie the vascular deficit in spontaneously hypertensive rats. Quantitative PCR and immunohistochemistry confirmed gene and protein 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 isoproterenol-induced relaxation and current. Isoproterenol-induced relaxations were also reduced in arteries incubated with small interference RNAs targeted to KCNQ4 that produced a ≈60% decrease in Kv7.4 protein level. Relaxation to isoproterenol and the Kv7 activator S-1 were abolished in arteries from spontaneously hypertensive rats, which was associated with ≈60% decrease in Kv7.4 abundance. This study provides the first evidence that Kv7 channels contribute to β-adrenoceptor-mediated vasodilation in the renal vasculature and that abrogation of Kv7.4 channels is strongly implicated in the impaired β-adrenoceptor pathway in spontaneously hypertensive rats. These findings may provide a novel pathogenic link between arterial dysfunction and hypertension.

  20. Voltage-Dependent Anion Channel 1(VDAC1) Participates the Apoptosis of the Mitochondrial Dysfunction in Desminopathy

    Science.gov (United States)

    Mo, Yanqing; Gong, Qi; Jiang, Aihua; Zhao, Jing

    2016-01-01

    Desminopathies caused by the mutation in the gene coding for desmin are genetically protein aggregation myopathies. Mitochondrial dysfunction is one of pathological changes in the desminopathies at the earliest stage. The molecular mechanisms of mitochondria dysfunction in desminopathies remain exclusive. VDAC1 regulates mitochondrial uptake across the outer membrane and mitochondrial outer membrane permeabilization (MOMP). Relationships between desminopathies and Voltage-dependent anion channel 1 (VDAC1) remain unclear. Here we successfully constructed the desminopathy rat model, evaluated with conventional stains, containing hematoxylin and eosin (HE), Gomori Trichrome (MGT), (PAS), red oil (ORO), NADH-TR, SDH staining and immunohistochemistry. Immunofluorescence results showed that VDAC1 was accumulated in the desmin highly stained area of muscle fibers of desminopathy patients or desminopathy rat model compared to the normal ones. Meanwhile apoptosis related proteins bax and ATF2 were involved in desminopathy patients and desminopathy rat model, but not bcl-2, bcl-xl or HK2.VDAC1 and desmin are closely relevant in the tissue splices of deminopathies patients and rats with desminopathy at protein lever. Moreover, apoptotic proteins are also involved in the desminopathies, like bax, ATF2, but not bcl-2, bcl-xl or HK2. This pathological analysis presents the correlation between VDAC1 and desmin, and apoptosis related proteins are correlated in the desminopathy. Furthermore, we provide a rat model of desminopathy for the investigation of desmin related myopathy. PMID:27941998

  1. Multiphasic profiles for voltage-dependent K+ channels: Reanalysis of data of MacKinnon and coworkers

    CERN Document Server

    Nissen, Per

    2016-01-01

    In a study of the role that voltage-dependent K+ channels may have in the mechanosensation of living cells (Schmidt et al. Proc Soc Natl Acad Sci USA 109: 10352-10357. 2012), the data were as conventionally done fitted by a Boltzmann function. However, as also found for other data for ion channels, this interpretation must be rejected in favor of a multiphasic profile, a series of straight lines separated by discontinuous transitions, quite often in the form of noncontiguities (jumps). The data points in the present study are often very unevenly distributed around the curvilinear profiles. Thus, for 43 of the 75 profiles, the probability is less than 5% that the uneven distribution is due to chance, for 26 the probability is less than 1%, and for 12 the probability is less than 0.1%, giving a vanishingly low overall probability for all profiles. Especially at low voltages, the differences between the fits to curvilinear and multiphasic profiles may be huge. In the multiphasic profiles, adjacent lines are quit...

  2. Voltage-dependent motion of the catalytic region of voltage-sensing phosphatase monitored by a fluorescent amino acid.

    Science.gov (United States)

    Sakata, Souhei; Jinno, Yuka; Kawanabe, Akira; Okamura, Yasushi

    2016-07-05

    The cytoplasmic region of voltage-sensing phosphatase (VSP) derives the voltage dependence of its catalytic activity from coupling to a voltage sensor homologous to that of voltage-gated ion channels. To assess the conformational changes in the cytoplasmic region upon activation of the voltage sensor, we genetically incorporated a fluorescent unnatural amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (Anap), into the catalytic region of Ciona intestinalis VSP (Ci-VSP). Measurements of Anap fluorescence under voltage clamp in Xenopus oocytes revealed that the catalytic region assumes distinct conformations dependent on the degree of voltage-sensor activation. FRET analysis showed that the catalytic region remains situated beneath the plasma membrane, irrespective of the voltage level. Moreover, Anap fluorescence from a membrane-facing loop in the C2 domain showed a pattern reflecting substrate turnover. These results indicate that the voltage sensor regulates Ci-VSP catalytic activity by causing conformational changes in the entire catalytic region, without changing their distance from the plasma membrane.

  3. Voltage-dependent anion channels (VDACs, porin) expressed in the plasma membrane regulate the differentiation and function of human osteoclasts.

    Science.gov (United States)

    Kotake, Shigeru; Yago, Toru; Kawamoto, Manabu; Nanke, Yuki

    2013-01-01

    Fewer molecules have been identified on human than murine osteoclasts, the former differing from murine osteoclasts in many ways. We show that voltage-dependent anion channels (VDACs, porin) are expressed in the plasma membrane of human osteoclasts. A search for novel proteins expressed in the plasma membrane of human osteoclasts identified VDAC. Anti-VDAC antibodies inhibited human osteoclastogenesis in vitro. VDAC expression was detected in membranes by immunoelectron microscopy and immunocytochemical double staining. The VDAC protein functions as a Cl(-) channel. VDACs regulate bone resorption, which show using Osteologic™ plates. The epitope of the antibody lay within a 10-amino acid sequence in the VDAC. The findings suggest that the VDAC is, at least partly, a novel Cl(-) channel regulating the differentiation and function of human osteoclasts. VDACs may play a crucial role in acidifying the resorption lacunae between osteoclasts and bone. Inhibitors of VDACs could be used to treat diseases involving increased resorption, such as osteoporosis, rheumatoid arthritis, and Paget's disease. © 2012 International Federation for Cell Biology.

  4. The voltage-dependent K+ channels Kv1.3 and Kv1.5 in human cancer

    Science.gov (United States)

    Comes, Núria; Bielanska, Joanna; Vallejo-Gracia, Albert; Serrano-Albarrás, Antonio; Marruecos, Laura; Gómez, Diana; Soler, Concepció; Condom, Enric; Ramón y Cajal, Santiago; Hernández-Losa, Javier; Ferreres, Joan C.; Felipe, Antonio

    2013-01-01

    Voltage-dependent K+ channels (Kv) are involved in a number of physiological processes, including immunomodulation, cell volume regulation, apoptosis as well as differentiation. Some Kv channels participate in the proliferation and migration of normal and tumor cells, contributing to metastasis. Altered expression of Kv1.3 and Kv1.5 channels has been found in several types of tumors and cancer cells. In general, while the expression of Kv1.3 apparently exhibits no clear pattern, Kv1.5 is induced in many of the analyzed metastatic tissues. Interestingly, evidence indicates that Kv1.5 channel shows inversed correlation with malignancy in some gliomas and non-Hodgkin's lymphomas. However, Kv1.3 and Kv1.5 are similarly remodeled in some cancers. For instance, expression of Kv1.3 and Kv1.5 correlates with a certain grade of tumorigenicity in muscle sarcomas. Differential remodeling of Kv1.3 and Kv1.5 expression in human cancers may indicate their role in tumor growth and their importance as potential tumor markers. However, despite of this increasing body of information, which considers Kv1.3 and Kv1.5 as emerging tumoral markers, further research must be performed to reach any conclusion. In this review, we summarize what it has been lately documented about Kv1.3 and Kv1.5 channels in human cancer. PMID:24133455

  5. The voltage-dependent K(+) channels Kv1.3 and Kv1.5 in human cancer.

    Science.gov (United States)

    Comes, Núria; Bielanska, Joanna; Vallejo-Gracia, Albert; Serrano-Albarrás, Antonio; Marruecos, Laura; Gómez, Diana; Soler, Concepció; Condom, Enric; Ramón Y Cajal, Santiago; Hernández-Losa, Javier; Ferreres, Joan C; Felipe, Antonio

    2013-10-10

    Voltage-dependent K(+) channels (Kv) are involved in a number of physiological processes, including immunomodulation, cell volume regulation, apoptosis as well as differentiation. Some Kv channels participate in the proliferation and migration of normal and tumor cells, contributing to metastasis. Altered expression of Kv1.3 and Kv1.5 channels has been found in several types of tumors and cancer cells. In general, while the expression of Kv1.3 apparently exhibits no clear pattern, Kv1.5 is induced in many of the analyzed metastatic tissues. Interestingly, evidence indicates that Kv1.5 channel shows inversed correlation with malignancy in some gliomas and non-Hodgkin's lymphomas. However, Kv1.3 and Kv1.5 are similarly remodeled in some cancers. For instance, expression of Kv1.3 and Kv1.5 correlates with a certain grade of tumorigenicity in muscle sarcomas. Differential remodeling of Kv1.3 and Kv1.5 expression in human cancers may indicate their role in tumor growth and their importance as potential tumor markers. However, despite of this increasing body of information, which considers Kv1.3 and Kv1.5 as emerging tumoral markers, further research must be performed to reach any conclusion. In this review, we summarize what it has been lately documented about Kv1.3 and Kv1.5 channels in human cancer.

  6. Probing the gate--voltage-dependent surface potential of individual InAs nanowires using random telegraph signals.

    Science.gov (United States)

    Salfi, Joe; Paradiso, Nicola; Roddaro, Stefano; Heun, Stefan; Nair, Selvakumar V; Savelyev, Igor G; Blumin, Marina; Beltram, Fabio; Ruda, Harry E

    2011-03-22

    We report a novel method for probing the gate-voltage dependence of the surface potential of individual semiconductor nanowires. The statistics of electronic occupation of a single defect on the surface of the nanowire, determined from a random telegraph signal, is used as a measure for the local potential. The method is demonstrated for the case of one or two switching defects in indium arsenide (InAs) nanowire field effect transistors at temperatures T=25-77 K. Comparison with a self-consistent model shows that surface potential variation is retarded in the conducting regime due to screening by surface states with density Dss≈10(12) cm(-2) eV(-1). Temperature-dependent dynamics of electron capture and emission producing the random telegraph signals are also analyzed, and multiphonon emission is identified as the process responsible for capture and emission of electrons from the surface traps. Two defects studied in detail had capture activation energies of EB≈50 meV and EB≈110 meV and cross sections of σ∞≈3×10(-19) cm2 and σ∞≈2×10(-17) cm2, respectively. A lattice relaxation energy of Sℏω=187±15 meV was found for the first defect.

  7. Voltage-dependent anion channels modulate mitochondrial metabolism in cancer cells: regulation by free tubulin and erastin.

    Science.gov (United States)

    Maldonado, Eduardo N; Sheldon, Kely L; DeHart, David N; Patnaik, Jyoti; Manevich, Yefim; Townsend, Danyelle M; Bezrukov, Sergey M; Rostovtseva, Tatiana K; Lemasters, John J

    2013-04-26

    Respiratory substrates and adenine nucleotides cross the mitochondrial outer membrane through the voltage-dependent anion channel (VDAC), comprising three isoforms--VDAC1, 2, and 3. We characterized the role of individual isoforms in mitochondrial metabolism by HepG2 human hepatoma cells using siRNA. With VDAC3 to the greatest extent, all VDAC isoforms contributed to the maintenance of mitochondrial membrane potential, but only VDAC3 knockdown decreased ATP, ADP, NAD(P)H, and mitochondrial redox state. Cells expressing predominantly VDAC3 were least sensitive to depolarization induced by increased free tubulin. In planar lipid bilayers, free tubulin inhibited VDAC1 and VDAC2 but not VDAC3. Erastin, a compound that interacts with VDAC, blocked and reversed mitochondrial depolarization after microtubule destabilizers in intact cells and antagonized tubulin-induced VDAC blockage in planar bilayers. In conclusion, free tubulin inhibits VDAC1/2 and limits mitochondrial metabolism in HepG2 cells, contributing to the Warburg phenomenon. Reversal of tubulin-VDAC interaction by erastin antagonizes Warburg metabolism and restores oxidative mitochondrial metabolism.

  8. Transcriptional upregulation of α2δ-1 elevates arterial smooth muscle cell voltage-dependent Ca2+ channel surface expression and cerebrovascular constriction in genetic hypertension.

    Science.gov (United States)

    Bannister, John P; Bulley, Simon; Narayanan, Damodaran; Thomas-Gatewood, Candice; Luzny, Patrik; Pachuau, Judith; Jaggar, Jonathan H

    2012-10-01

    A hallmark of hypertension is an increase in arterial myocyte voltage-dependent Ca2+ (CaV1.2) currents that induces pathological vasoconstriction. CaV1.2 channels are heteromeric complexes composed of a pore-forming CaV1.2α1 with auxiliary α2δ and β subunits. Molecular mechanisms that elevate CaV1.2 currents during hypertension and the potential contribution of CaV1.2 auxiliary subunits are unclear. Here, we investigated the pathological significance of α2δ subunits in vasoconstriction associated with hypertension. Age-dependent development of hypertension in spontaneously hypertensive rats was associated with an unequal elevation in α2δ-1 and CaV1.2α1 mRNA and protein in cerebral artery myocytes, with α2δ-1 increasing more than CaV1.2α1. Other α2δ isoforms did not emerge in hypertension. Myocytes and arteries of hypertensive spontaneously hypertensive rats displayed higher surface-localized α2δ-1 and CaV1.2α1 proteins, surface α2δ-1:CaV1.2α1 ratio, CaV1.2 current density and noninactivating current, and pressure- and depolarization-induced vasoconstriction than those of Wistar-Kyoto controls. Pregabalin, an α2δ-1 ligand, did not alter α2δ-1 or CaV1.2α1 total protein but normalized α2δ-1 and CaV1.2α1 surface expression, surface α2δ-1:CaV1.2α1, CaV1.2 current density and inactivation, and vasoconstriction in myocytes and arteries of hypertensive rats to control levels. Genetic hypertension is associated with an elevation in α2δ-1 expression that promotes surface trafficking of CaV1.2 channels in cerebral artery myocytes. This leads to an increase in CaV1.2 current-density and a reduction in current inactivation that induces vasoconstriction. Data also suggest that α2δ-1 targeting is a novel strategy that may be used to reverse pathological CaV1.2 channel trafficking to induce cerebrovascular dilation in hypertension.

  9. The Calcium-dependent Potassium Current in Olfactory Interneurons of the Cockroach Periplaneta americana

    OpenAIRE

    Schleicher, Sabine

    2014-01-01

    The olfactory system of insects has already served as a suitable model to investigate mechanisms of general information processing. Thus, insect olfactory neurons were used to study physiology, transmitter content, from that evoked currents as well as sensory processing on a network level. As an important step towards understanding how distinct electrophysiological properties of neurons are generated by intrinsic currents, I used whole-cell patch-clamp recordings to analyze biophysical and ph...

  10. Somatostatinergic modulation of firing pattern and calcium-activated potassium currents in medium spiny neostriatal neurons.

    Science.gov (United States)

    Galarraga, E; Vilchis, C; Tkatch, T; Salgado, H; Tecuapetla, F; Perez-Rosello, T; Perez-Garci, E; Hernandez-Echeagaray, E; Surmeier, D J; Bargas, J

    2007-05-11

    Somatostatin is synthesized and released by aspiny GABAergic interneurons of the neostriatum, some of them identified as low threshold spike generating neurons (LTS-interneurons). These neurons make synaptic contacts with spiny neostriatal projection neurons. However, very few somatostatin actions on projection neurons have been described. The present work reports that somatostatin modulates the Ca(2+) activated K(+) currents (K(Ca) currents) expressed by projection cells. These actions contribute in designing the firing pattern of the spiny projection neuron; which is the output of the neostriatum. Small conductance (SK) and large conductance (BK) K(Ca) currents represent between 30% and 50% of the sustained outward current in spiny cells. Somatostatin reduces SK-type K(+) currents and at the same time enhances BK-type K(+) currents. This dual effect enhances the fast component of the after hyperpolarizing potential while reducing the slow component. Somatostatin then modifies the firing pattern of spiny neurons which changed from a tonic regular pattern to an interrupted "stuttering"-like pattern. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) tissue expression analysis of dorsal striatal somatostatinergic receptors (SSTR) mRNA revealed that all five SSTR mRNAs are present. However, single cell RT-PCR profiling suggests that the most probable receptor in charge of this modulation is the SSTR2 receptor. Interestingly, aspiny interneurons may exhibit a "stuttering"-like firing pattern. Therefore, somatostatin actions appear to be the entrainment of projection neurons to the rhythms generated by some interneurons. Somatostatin is then capable of modifying the processing and output of the neostriatum.

  11. Effect of Pinellia ternata Lectin on Membrane Currents of Mouse Motor Nerve Terminals

    Institute of Scientific and Technical Information of China (English)

    施玉樑; 徐幼芬; 张辉

    1994-01-01

    Pinellia ternata lectin (PTL) extracted from the fresh juice of rhizome of pinellia ternata used as a traditional Chinese medicine facilitated the quantal release of acetylcholine (ACh) in the mouse motor nerve terminals and formed cation channels in artificial lipid bilayer. Here we report the action of PTL on presynaptic membrane currents of motor nerve terminals.The experiments were performed on the intercostal nerve triangularis sterni muscle preparations. By means of the perineurial recording, the effects of PTL on the sodium current in the preterminal part , three potassium currents and two calcium currents generated from the nerve terminals were investigated. The results show that PTL increases voltage-dependent fast Ca2+ current (ICa,f), Na+ current (INa) and Ca2+-acti-vated K+ current (IK,Ca) without action on either the voltage-dependent fast K+ current (IK,f) or the slow K+ current (IK,S). These effects are irreversible, but can be reversed by mannan, the specific binding sugar for PTL.The to

  12. Simultaneous mapping of membrane voltage and calcium in zebrafish heart in vivo reveals chamber-specific developmental transitions in ionic currents

    Directory of Open Access Journals (Sweden)

    Jennifer H Hou

    2014-09-01

    Full Text Available The cardiac action potential (AP and the consequent cytosolic Ca2+ transient are key indicators of cardiac function. Natural developmental processes, as well as many drugs and pathologies change the waveform, propagation, or variability (between cells or over time of these parameters. Here we apply a genetically encoded dual-function calcium and voltage reporter (CaViar to study the development of the zebrafish heart in vivo between 1.5 and 4 days post fertilization (dpf. We developed a high-sensitivity spinning disk confocal microscope and associated software for simultaneous three-dimensional optical mapping of voltage and calcium. We produced a transgenic zebrafish line expressing CaViar under control of the heart-specific cmlc2 promoter, and applied ion channel blockers at a series of developmental stages to map the maturation of the action potential in vivo. Early in development, the AP initiated via a calcium current through L-type calcium channels. Between 90 – 102 hours post fertilization (hpf, the ventricular AP switched to a sodium-driven upswing, while the atrial AP remained calcium driven. In the adult zebrafish heart, a sodium current drives the AP in both the atrium and ventricle. Simultaneous voltage and calcium imaging with genetically encoded reporters provides a new approach for monitoring cardiac development, and the effects of drugs on cardiac function.

  13. A conserved threonine in the S1-S2 loop of KV7.2 and K V7.3 channels regulates voltage-dependent activation.

    Science.gov (United States)

    Füll, Yvonne; Seebohm, Guiscard; Lerche, Holger; Maljevic, Snezana

    2013-06-01

    The voltage-gated potassium channels KV7.2 and KV7.3 (KCNQ2/3 genes) play an important role in regulating neuronal excitability. More than 50 KCNQ2/3 mutations have been identified to cause an inherited form of epilepsy in newborns. For two of those (E119G and S122L) found in the S1-S2 region of KV7.2, we previously showed a decreased channel availability mainly at action potential subthreshold voltages caused by a slight depolarizing shift of the activation curve. Interestingly, recent studies revealed that a threonine residue within the S1-S2 loop, highly conserved among different classes of KV channels, is crucial for both their function and surface expression. To investigate the functional role of the homologous threonine residues in KV7.2 (T114) and KV7.3 (T144) channels, we replaced them with alanine and examined the electrophysiological properties using heterologous expression in CHO cells and whole cell patch clamping. Channels comprising mutant subunits yielded decreased potassium currents with slowed activation and accelerated deactivation kinetics. However, the most striking effect was a depolarizing shift in the voltage dependence of activation reaching +30 mV upon co-expression of both mutant subunits. Potential interactions of T114 within the channel were analyzed by creating a 3D homology model of KV7.2 in an open state suggesting that this residue plays a central role in the formation of a stable interface between the S1-S2 and the S5 segment helices. This could be the explanation why substitution of the conserved threonine in KV7.2 and KV7.3 channels destabilizes the open and favors the closed state of these channels.

  14. The episodic ataxia type 1 mutation I262T alters voltage-dependent gating and disrupts protein biosynthesis of human Kv1.1 potassium channels.

    Science.gov (United States)

    Chen, Szu-Han; Fu, Ssu-Ju; Huang, Jing-Jia; Tang, Chih-Yung

    2016-01-18

    Voltage-gated potassium (Kv) channels are essential for setting neuronal membrane excitability. Mutations in human Kv1.1 channels are linked to episodic ataxia type 1 (EA1). The EA1-associated mutation I262T was identified from a patient with atypical phenotypes. Although a previous report has characterized its suppression effect, several key questions regarding the impact of the I262T mutation on Kv1.1 as well as other members of the Kv1 subfamily remain unanswered. Herein we show that the dominant-negative effect of I262T on Kv1.1 current expression is not reversed by co-expression with Kvβ1.1 or Kvβ2 subunits. Biochemical examinations indicate that I262T displays enhanced protein degradation and impedes membrane trafficking of Kv1.1 wild-type subunits. I262T appears to be the first EA1 mutation directly associated with impaired protein stability. Further functional analyses demonstrate that I262T changes the voltage-dependent activation and Kvβ1.1-mediated inactivation, uncouples inactivation from activation gating, and decelerates the kinetics of cumulative inactivation of Kv1.1 channels. I262T also exerts similar dominant effects on the gating of Kv1.2 and Kv1.4 channels. Together our data suggest that I262T confers altered channel gating and reduced functional expression of Kv1 channels, which may account for some of the phenotypes of the EA1 patient.

  15. Accessory subunit KChIP2 modulates the cardiac L-type calcium current

    DEFF Research Database (Denmark)

    Thomsen, Morten B; Wang, Chaojian; Ozgen, Nazira

    2009-01-01

    Complex modulation of voltage-gated Ca2+ currents through the interplay among Ca2+ channels and various Ca(2+)-binding proteins is increasingly being recognized. The K+ channel interacting protein 2 (KChIP2), originally identified as an auxiliary subunit for K(V)4.2 and a component of the transient...... outward K+ channel (I(to)), is a Ca(2+)-binding protein whose regulatory functions do not appear restricted to K(V)4.2. Consequently, we hypothesized that KChIP2 is a direct regulator of the cardiac L-type Ca2+ current (I(Ca,L)). We found that I(Ca,L) density from KChIP2(-/-) myocytes is reduced by 28......% compared to I(Ca,L) recorded from wild-type myocytes (Pchannel current, as shown in a transfected cell line devoid of confounding cardiac ion currents. I(Ca,L) regulation by KChIP2 was independent of Ca2+ binding...

  16. Transient calcium-dependent potassium current in magnocellular neurosecretory cells of the rat supraoptic nucleus.

    Science.gov (United States)

    Bourque, C W

    1988-03-01

    1. Magnocellular neurosecretory neurones were impaled in the supraoptic nucleus of perfused explants of rat hypothalamus. Membrane currents were studied at 35 degrees C using the single-microelectrode voltage-clamp technique. 2. Depolarizing voltage steps applied from -100 mV evoked a transient outward current (TOC) from a threshold of -75 mV. From this potential, the amplitude of the current increased non-linearly with voltage. 3. Following its activation TOC reached a peak within 7 ms and subsequently decayed monotonically with a time constant of 30 ms. This time constant did not vary significantly with voltage between -75 and -55 mV. 4. The TOC showed complete steady-state inactivation at potentials positive to -55 mV. Inactivation was removed by hyperpolarization, with a mid-point near -80 mV. The removal of inactivation followed a complex time course with distinct fast (tens of milliseconds) and slow (hundreds of milliseconds) components. 5. Tail current measurements revealed that the TOC equilibrium potential (ETOC) lies near -97 mV in the presence of 3 mM [K+]o. Increasing [K+]o caused a decrease of TOC amplitude and a shift in ETOC of 57 mV/log [K+]o. The TOC is therefore predominantly a K+ current. 6. The TOC was unaffected by tetraethylammonium (up to 12 mM) but was reversibly reduced by 4-aminopyridine (ca. 50% block at 1.0 mM) and dendrotoxin (ca. 50% block at 4 nM). 7. The TOC was strongly inhibited (greater than 70%) by adding Co2+ or Mn2+ (1-3 mM) or Cd2+ (50-400 microM) to Ca-containing solutions, or by removal of Ca2+ from the perfusate. These effects were not accompanied by detectable changes in threshold voltage. The amplitude of TOC was also depressed by the organic Ca2+ channel blocker methoxyverapamil (D600). Finally replacement of Ca2+ by Ba2+ in the perfusate completely and reversibly abolished the TOC. 8. These findings suggest that neurosecretory neurones of the rat supraoptic nucleus display a transient K+ current which is strongly

  17. Pronase acutely modifies high voltage-activated calcium currents and cell properties of Lymnaea neurons.

    Science.gov (United States)

    Hermann, P M; Lukowiak, K; Wildering, W C; Bulloch, A G

    1997-12-01

    Pronase E ('pronase') is one of the proteolytic enzymes that are used in preparative procedures such as cell isolation and to soften the sheath of invertebrate ganglia. Although several effects of proteolytic enzymes on the physiology of non-neuronal tissues have been described, the effects of these enzymes on central neurons have received little attention. We examined the effects of bath-applied pronase on neurons in the Lymnaea central nervous system and in vitro. Pronase caused action potential broadening in neurons that exhibit a shoulder on the repolarization phase of their action potentials. This effect of pronase was accompanied by, although unrelated to, a depolarization and decrease in action potential interval. Some, but not all, effects of pronase in the central nervous system were reversible. For example, the decreases in membrane potential and action potential interval were both reversed after approximately 1 h of washing with saline. However, the effect of pronase on the action potential duration was not reversed after a period of 90 min. The modulation of action potential width prompted us to examine Ca2+ currents. Exposure to pronase resulted in an increase in both peak and late high voltage-activated Ca2+ currents in isolated neurons. Pronase neither changed the inactivation rate nor caused a shift in the current-voltage relationship of the current. The changes in action potential duration could be prevented by application of 0.1 mM Cd2+, indicating that the action potential broadening caused by pronase depends on Ca2+ influx. This is the first systematic study of the acute and direct actions of pronase on Ca2+ currents and cell properties both in the CNS and in vitro.

  18. Calcium sensing in exocytosis

    DEFF Research Database (Denmark)

    Gustavsson, Natalia; Wu, Bingbing; Han, Weiping

    2012-01-01

    an increase in intracellular calcium levels. Besides the triggering role, calcium signaling modulates the precise amount and kinetics of vesicle release. Thus, it is a central question to understand the molecular machineries responsible for calcium sensing in exocytosis. Here we provide an overview of our...... current understanding of calcium sensing in neurotransmitter release and hormone secretion....

  19. Altered ischemic cerebral injury in mice lacking αIE subunit of the voltage-dependent Ca2+ channel

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Objective ①To set up a stable and reproducible focal cerebral infarct modelin mice; (②To examine theinvolvement of αIE subunit of voltage-dependent Ca2 + channel in cerebral ischemic injury. Methods Male C57BL/6J Jclmice 8 ~ 12w and F4 ~ F6αIE subunit of Ca2+ channel mutant mice were both used in this study. All animals were allowedto freely access to food and water before and after operation. Animals were anesthetized with pentobarbital sodium 60mg/kg,ip. Rectal temperature was continuously monitored before, during and after operation, and maintained at (36.6 +0.1 )°C by a autoregulating pad. To produce pilot models, the middle cerebral artery (MCA) was occluded either by sur-gical ligation or electrical coagulation and in some models the common carotid artery (CCA) was surgically ligated in tan-dem. In our latter work the MCA was cut off soon after it was ligated or coagulated in order to make sure that the bloodflow was occluded completely. The MCA was coagulated or ligated with a bipolar coagulator or microsurgery suture at thesite just superior to the rhinal fissure. Twenty~four hours after the operation, the mice were anesthetized and decapitated,then their brains were dissected from the skull and put into cold artificial brain spinal fluid as soon as possible. Lmm thickcoronal sections were cut by vibratome and stained with 2% 2,3,5-triphenyltetrazolium chloride (TTC) at 37°C for30min. Every section was photographed positively and the whole infarction volume was calculated by summing up the in-farction volumes of all sections by NIH Image System. Infarction ratio ( % ) was also calculated by the following fommula:(contralateral volume-ipsilateral undamaged volume)/contralateral volume × 100% to eliminate the influence of edema.In brief, the mutant mice were produced with gene targeting technique. F4 ~ F6 mice were used in this experiment. Alloffsprings were genotyped by the polymerase chain reaction (PCR) and the genotypes remained umknown

  20. Extracellular calcium modulates persistent sodium current-dependent burst-firing in hippocampal pyramidal neurons.

    Science.gov (United States)

    Su, H; Alroy, G; Kirson, E D; Yaari, Y

    2001-06-15

    The generation of high-frequency spike bursts ("complex spikes"), either spontaneously or in response to depolarizing stimuli applied to the soma, is a notable feature in intracellular recordings from hippocampal CA1 pyramidal cells (PCs) in vivo. There is compelling evidence that the bursts are intrinsically generated by summation of large spike afterdepolarizations (ADPs). Using intracellular recordings in adult rat hippocampal slices, we show that intrinsic burst-firing in CA1 PCs is strongly dependent on the extracellular concentration of Ca(2+) ([Ca(2+)](o)). Thus, lowering [Ca(2+)](o) (by equimolar substitution with Mn(2+) or Mg(2+)) induced intrinsic bursting in nonbursters, whereas raising [Ca(2+)](o) suppressed intrinsic bursting in native bursters. The induction of intrinsic bursting by low [Ca(2+)](o) was associated with enlargement of the spike ADP. Low [Ca(2+)](o)-induced intrinsic bursts and their underlying ADPs were suppressed by drugs that reduce the persistent Na(+) current (I(NaP)), indicating that this current mediates the slow burst depolarization. Blocking Ca(2+)-activated K(+) currents with extracellular Ni(2+) or intracellular chelation of Ca(2+) did not induce intrinsic bursting. This and other evidence suggest that lowering [Ca(2+)](o) may induce intrinsic bursting by augmenting I(NaP). Because repetitive neuronal activity in the hippocampus is associated with marked decreases in [Ca(2+)](o), the regulation of intrinsic bursting by extracellular Ca(2+) may provide a mechanism for preferential recruitment of this firing mode during certain forms of hippocampal activation.

  1. Inhibition of potassium and calcium currents in neurones by molecularly-defined P2Y receptors.

    Science.gov (United States)

    Brown, D A; Filippov, A K; Barnard, E A

    2000-07-01

    Messenger RNAs and cDNAs for individual cloned P2Y(1), P2Y2 and P2Y(6) nucleotide receptors have been expressed by micro-injection into dissociated rat superior cervical sympathetic neurones and the effects of stimulating the expressed receptors on voltage-activated N-type Ca(2+) currents and M-type K(+) currents recorded. Both currents were reduced by stimulating all three receptors, with the following mean IC(50) values: P2Y(1) (agonist: ADP) - I(K(M)) 6.9 nM, I(Ca) 8.2 nM; P2Y(2) (agonist: UTP) - I(K(M)) 1.5 microM, I(Ca) 0.5 microM; P2Y(6) (agonist: UDP) - I(K(M)) 30 nM, I(Ca) 5.9 nM. Inhibition of I(K(M)) was voltage-independent and insensitive to Pertussis toxin; inhibition of I(Ca) showed both voltage-sensitive and insensitive, and Pertussis toxin-sensitive and insensitive components. It is concluded that these P2Y receptors can couple to more than one G protein and thereby modulate more than one ion channel. It is suggested that these effects on K(M) and Ca(N) channels may induce both postsynaptic excitory and presynaptic inhibitory responses.

  2. The Voltage-dependent Anion Channel 1 Mediates Amyloid β Toxicity and Represents a Potential Target for Alzheimer Disease Therapy.

    Science.gov (United States)

    Smilansky, Angela; Dangoor, Liron; Nakdimon, Itay; Ben-Hail, Danya; Mizrachi, Dario; Shoshan-Barmatz, Varda

    2015-12-25

    The voltage-dependent anion channel 1 (VDAC1), found in the mitochondrial outer membrane, forms the main interface between mitochondrial and cellular metabolisms, mediates the passage of a variety of molecules across the mitochondrial outer membrane, and is central to mitochondria-mediated apoptosis. VDAC1 is overexpressed in post-mortem brains of Alzheimer disease (AD) patients. The development and progress of AD are associated with mitochondrial dysfunction resulting from the cytotoxic effects of accumulated amyloid β (Aβ). In this study we demonstrate the involvement of VDAC1 and a VDAC1 N-terminal peptide (VDAC1-N-Ter) in Aβ cell penetration and cell death induction. Aβ directly interacted with VDAC1 and VDAC1-N-Ter, as monitored by VDAC1 channel conductance, surface plasmon resonance, and microscale thermophoresis. Preincubated Aβ interacted with bilayer-reconstituted VDAC1 and increased its conductance ∼ 2-fold. Incubation of cells with Aβ resulted in mitochondria-mediated apoptotic cell death. However, the presence of non-cell-penetrating VDAC1-N-Ter peptide prevented Aβ cellular entry and Aβ-induced mitochondria-mediated apoptosis. Likewise, silencing VDAC1 expression by specific siRNA prevented Aβ entry into the cytosol as well as Aβ-induced toxicity. Finally, the mode of Aβ-mediated action involves detachment of mitochondria-bound hexokinase, induction of VDAC1 oligomerization, and cytochrome c release, a sequence of events leading to apoptosis. As such, we suggest that Aβ-mediated toxicity involves mitochondrial and plasma membrane VDAC1, leading to mitochondrial dysfunction and apoptosis induction. The VDAC1-N-Ter peptide targeting Aβ cytotoxicity is thus a potential new therapeutic strategy for AD treatment.

  3. Photoaffinity labeling with cholesterol analogues precisely maps a cholesterol-binding site in voltage-dependent anion channel-1.

    Science.gov (United States)

    Budelier, Melissa M; Cheng, Wayland W L; Bergdoll, Lucie; Chen, Zi-Wei; Janetka, James W; Abramson, Jeff; Krishnan, Kathiresan; Mydock-McGrane, Laurel; Covey, Douglas F; Whitelegge, Julian P; Evers, Alex S

    2017-06-02

    Voltage-dependent anion channel-1 (VDAC1) is a highly regulated β-barrel membrane protein that mediates transport of ions and metabolites between the mitochondria and cytosol of the cell. VDAC1 co-purifies with cholesterol and is functionally regulated by cholesterol, among other endogenous lipids. Molecular modeling studies based on NMR observations have suggested five cholesterol-binding sites in VDAC1, but direct experimental evidence for these sites is lacking. Here, to determine the sites of cholesterol binding, we photolabeled purified mouse VDAC1 (mVDAC1) with photoactivatable cholesterol analogues and analyzed the photolabeled sites with both top-down mass spectrometry (MS), and bottom-up MS paired with a clickable, stable isotope-labeled tag, FLI-tag. Using cholesterol analogues with a diazirine in either the 7 position of the steroid ring (LKM38) or the aliphatic tail (KK174), we mapped a binding pocket in mVDAC1 localized to Thr(83) and Glu(73), respectively. When Glu(73) was mutated to a glutamine, KK174 no longer photolabeled this residue, but instead labeled the nearby Tyr(62) within this same binding pocket. The combination of analytical strategies employed in this work permits detailed molecular mapping of a cholesterol-binding site in a protein, including an orientation of the sterol within the site. Our work raises the interesting possibility that cholesterol-mediated regulation of VDAC1 may be facilitated through a specific binding site at the functionally important Glu(73) residue. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. Voltage-dependent changes in specific membrane capacitance caused by prestin, the outer hair cell lateral membrane motor.

    Science.gov (United States)

    Santos-Sacchi, Joseph; Navarrete, Enrique

    2002-05-01

    In the outer hair cell (OHC), membrane capacitance principally derives from two components - that associated with lateral membrane sensor/motor charge movement, and that proportional to the membrane surface area (C(sa)). We used measures of membrane capacitance to test a model hypothesis that OHC lateral membrane molecular motors, recently identified as the protein prestin, fluctuate between two area states. By measuring membrane capacitance in native OHCs or prestin-transfected HEK cells at extreme voltages (+/-200 mV) where motor-derived charge movement is small or absent, we observed that C(sa) depends on the state of the motors, or correspondingly on membrane voltage. Deiters cells or control HEK cells, which lack motors, do not show this dependence. We modeled the voltage-dependent change in C(sa) as a Boltzmann process with the same parameters that describe the charge movement of the motors' voltage sensors. C(sa) is 3.28+/-0.75 pF (mean +/-SD; n=23) larger during extreme hyperpolarization, and the number of motors in OHCs and prestin-transfected HEK cells correlates with the magnitude of Delta C(sa)( r=0.78). Although these data are consistent with the area motor model, the corresponding area change, assuming 0.5 microF/cm(2) under constant membrane thickness, is unphysiologically large, and indicates that the capacitance change must result from changes not only in lateral membrane area but also specific capacitance. Thus, we conclude that a conformational change in the lateral membrane motor, prestin, additionally alters the dielectric constant and/or thickness of the lateral plasma membrane.

  5. Charged residues distribution modulates selectivity of the open state of human isoforms of the voltage dependent anion-selective channel.

    Science.gov (United States)

    Amodeo, Giuseppe Federico; Scorciapino, Mariano Andrea; Messina, Angela; De Pinto, Vito; Ceccarelli, Matteo

    2014-01-01

    Voltage Dependent Anion-selective Channels (VDACs) are pore-forming proteins located in the outer mitochondrial membrane. They are responsible for the access of ions and energetic metabolites into the inner membrane transport systems. Three VDAC isoforms exist in mammalian, but their specific role is unknown. In this work we have performed extensive (overall ∼5 µs) Molecular Dynamics (MD) simulations of the human VDAC isoforms to detect structural and conformational variations among them, possibly related to specific functional roles of these proteins. Secondary structure analysis of the N-terminal domain shows a high similarity among the three human isoforms of VDAC but with a different plasticity. In particular, the N-terminal domain of the hVDAC1 is characterized by a higher plasticity, with a ∼20% occurrence for the 'unstructured' conformation throughout the folded segment, while hVDAC2, containing a peculiar extension of 11 amino acids at the N-terminal end, presents an additional 310-helical folded portion comprising residues 10' to 3, adhering to the barrel wall. The N-terminal sequences of hVDAC isoforms are predicted to have a low flexibility, with possible consequences in the dynamics of the human VDACs. Clear differences were found between hVDAC1 and hVDAC3 against hVDAC2: a significantly modified dynamics with possible important consequence on the voltage-gating mechanism. Charge distribution inside and at the mouth of the pore is responsible for a different preferential localization of ions with opposite charge and provide a valuable rationale for hVDAC1 and hVDAC3 having a Cl-/K+ selectivity ratio of 1.8, whereas hVDAC2 of 1.4. Our conclusion is that hVDAC isoforms, despite sharing a similar scaffold, have modified working features and a biological work is now requested to give evidence to the described dissimilarities.

  6. Calcium-dependent potassium current following penicillin-induced epileptiform discharges in the hippocampal slice.

    Science.gov (United States)

    Domann, R; Dorn, T; Witte, O W

    1989-01-01

    Penicillin-induced paroxysmal depolarization shifts (PDS) are followed by prolonged afterhyperpolarizations of about 2 seconds duration. Intracellular injection of EGTA blocked a late component of the afterhyperpolarizations; an early one lasting up to one second was only slightly reduced by EGTA. It is concluded that afterhyperpolarizations following penicillin-induced PDS comprise different components: an initial one lasting up to one second which is not Ca2+-dependent and a slow one lasting up to two seconds which is caused by a Ca2+-dependent K+ current.

  7. The Hyperpolarization-Activated Current Determines Synaptic Excitability, Calcium Activity and Specific Viability of Substantia Nigra Dopaminergic Neurons

    Directory of Open Access Journals (Sweden)

    Carmen Carbone

    2017-06-01

    Full Text Available Differential vulnerability between Substantia Nigra pars compacta (SNpc and Ventral Tegmental Area (VTA dopaminergic (DAergic neurons is a hallmark of Parkinson’s disease (PD. Understanding the molecular bases of this key histopathological aspect would foster the development of much-needed disease-modifying therapies. Non-heterogeneous DAergic degeneration is present in both toxin-based and genetic animal models, suggesting that cellular specificity, rather than causing factors, constitutes the background for differential vulnerability. In this regard, we previously demonstrated that MPP+, a neurotoxin able to cause selective nigrostriatal degeneration in animal rodents and primates, inhibits the Hyperpolarization-activated current (Ih in SNpc DAergic neurons and that pharmacological Ih antagonism causes potentiation of evoked Excitatory post-synaptic potentials (EPSPs. Of note, the magnitude of such potentiation is greater in the SNpc subfield, consistent with higher Ih density. In the present work, we show that Ih block-induced synaptic potentiation leads to the amplification of somatic calcium responses (SCRs in vitro. This effect is specific for the SNpc subfield and largely mediated by L-Type calcium channels, as indicated by sensitivity to the CaV 1 blocker isradipine. Furthermore, Ih is downregulated by low intracellular ATP and determines the efficacy of GABAergic inhibition in SNpc DAergic neurons. Finally, we show that stereotaxic administration of Ih blockers causes SNpc-specific neurodegeneration and hemiparkinsonian motor phenotype in rats. During PD progression, Ih downregulation may result from mitochondrial dysfunction and, in concert with PD-related disinhibition of excitatory inputs, determine a SNpc-specific disease pathway.

  8. Decrease of high voltage Ca(2+) currents in the dentate gyrus granule cells by entorhinal amyloidopathy is reversed by calcium channel blockade.

    Science.gov (United States)

    Pourbadie, Hamid Gholami; Naderi, Nima; Delavar, Hadi Mirzapour; Hosseinzadeh, Mahshid; Mehranfard, Nasrin; Khodagholi, Fariba; Janahmadi, Mahyar; Motamedi, Fereshteh

    2017-01-05

    In the Alzheimer's disease (AD), entorhinal-hippocampal circuit is one of the earliest affected networks. There are some evidences indicating abnormal neuronal excitability and impaired synaptic plasticity in the dentate gyrus (DG) of AD animal model. However, the underlying mechanism leading to DG dysfunction particularly in the early phase of AD is not known. Since calcium dyshomeostasis has a critical role in the etiology of AD, it is possible that this phenomenon precedes electrophysiological alteration in the DG. Here, the effect of the amyloid pathogenesis in the entorhinal cortex (EC) on high activated Ca(2+) currents in the DG granule cells was investigated. One week after bilaterally injection of amyloid beta (Aβ) 1-42 into the EC, Ca(2+) currents in the DG granule cells were assessed by whole cell patch clamp. Voltage clamp recording showed the amplitude of high voltage calcium currents in the DG granule cells was decreased following EC amyloidopathy. However, the Ca(2+) current decay was slower than control. Double-pulse recording revealed that Ca(2+)-dependent inactivation of calcium current (CDI) was more pronounced in the EC-Aβ group compared to the control group. However, chronic treatment by calcium channel blocker (CCBs), isradipine or nimodipine, reverse the Ca(2+) currents toward the control level. On the other hand, there was no significant difference in the calbindin level in the DG of different groups. In conclusion, our results suggest that Aβ in the EC independent of calbindin level triggers a decreased Ca(2+) currents along with increased CDI in the DG granule cells which may lead to further electrophysiological alterations in these cells, and treatment by CCBs could preserve normal calcium current and may ultimately normal function against the Aβ toxicity. Copyright © 2016. Published by Elsevier B.V.

  9. Arachidonic acid modulates hippocampal calcium current via protein kinase C and oxygen radicals.

    Science.gov (United States)

    Keyser, D O; Alger, B E

    1990-10-01

    Arachidonic acid (AA) is a second messenger liberated via receptor activation of phospholipase A2 or diacylglycerol-lipase. We used whole-cell voltage clamp of acutely isolated hippocampal CA1 pyramidal cells to investigate the hypothesis that AA modulates Ca2+ channel current (ICa) via activation of protein kinase C (PKC) and generation of free radicals. AA depressed ICa in a dose- and time-dependent manner similar to that previously reported for the action of phorbol esters on ICa. A similar depression was seen with a xanthine-based free radical generating system. The specific PKC inhibitor PKCI (19-36), the protein kinase inhibitor H-7, and the superoxide free radical scavenger SOD each blocked ICa depression by 70%-80%. Complete block of the AA response occurred when SOD was used simultaneously with a PKC inhibitor. These data suggest that PKC and free radicals play a role in AA-induced suppression of ICa.

  10. Cyclic AMP enhances calcium-dependent potassium current in Aplysia neurons.

    Science.gov (United States)

    Ewald, D; Eckert, R

    1983-12-01

    The effect on the Ca-dependent potassium current, IK(Ca), of procedures that increase intracellular cAMP levels was studied in Aplysia neurons using three different pharmacological approaches. Exposure to cAMP analogues which were either resistant to or protected from phosphodiesterase hydrolysis caused an increase in IK(Ca) from 30 to 50% in 10 min. The degree of reversibility of this effect varied from complete with db cAMP to very little with pcpt cAMP. Exposure to cholera toxin, which stimulates the synthesis of endogenous cAMP, increased IK(Ca) 25% in 10 min and the effect was not reversible. Both approaches were effective in all seven neuron types studied. Application of serotonin plus phosphodiesterase inhibitor caused an increase in IK(Ca) in neuron R15 but not in the other neuron types. Application of pentylene tetrazole (PTZ) led to a decrease in IK(Ca). It is proposed that elevation of cyclic AMP mediates an increased sensitivity of the IK(Ca) channel to Ca ions.

  11. Disruption of learned timing in P/Q calcium channel mutants.

    Directory of Open Access Journals (Sweden)

    Akira Katoh

    Full Text Available To optimize motor performance, both the amplitude and temporal properties of movements should be modifiable by motor learning. Here we report that the modification of movement timing is highly dependent on signaling through P/Q-type voltage-dependent calcium channels. Two lines of mutant mice heterozygous for P/Q-type voltage-dependent calcium channels exhibited impaired plasticity of eye movement timing, but relatively intact plasticity of movement amplitude during motor learning in the vestibulo-ocular reflex. The results thus demonstrate a distinction between the molecular signaling pathways regulating the timing versus amplitude of movements.

  12. Effect of calcium on nicotine-induced current expressed by an atypical alpha-bungarotoxin-insensitive nAChR2.

    Science.gov (United States)

    Thany, Steeve H; Courjaret, Raphael; Lapied, Bruno

    2008-06-27

    Two distinct native alpha-bungarotoxin (alpha-Bgt)-insensitive nicotinic acetylcholine receptors (nAChRs), named nAChR1 and nAChR2, were identified in the cockroach Periplaneta americana dorsal unpaired median (DUM) neurons. They differed in their electrophysiological, pharmacological properties and intracellular regulation pathways. nAChR2 being an atypical nicotinic receptor closed upon agonist application and its current-voltage relationship resulted from a reduction in potassium conductance. In this study, using whole-cell patch-clamp technique, we demonstrated that calcium modulated nAChR2-mediated nicotine response. Under 0.5 microM alpha-Bgt and 20 mM d-tubocurarine, the nicotine-induced inward current amplitude was strongly reduced in the presence of intracellularly applied BAPTA or bath application of calcium-free solution. In addition, using cadmium chloride, we showed that nicotine response was modulated by extracellular calcium through plasma membrane calcium channels. Moreover, extracellular application of caffeine and thapsigargin reduced nAChR2-mediated response. Together these experiments revealed a complex calcium-dependent regulation of nAChR2.

  13. Coronary artery calcium screening : current status and recommendations from the European Society of Cardiac Radiology and North American Society for Cardiovascular Imaging

    NARCIS (Netherlands)

    Oudkerk, Matthijs; Stillman, Arthur E.; Halliburton, Sandra S.; Kalender, Willi A.; Moehlenkamp, Stefan; McCollough, Cynthia H.; Vliegenthart, Rozemarijn; Shaw, Leslee J.; Stanford, William; Taylor, Allen J.; van Ooijen, Peter M. A.; Wexler, Lewis; Raggi, Paolo

    2008-01-01

    Current guidelines and literature on screening for coronary artery calcium for cardiac risk assessment are reviewed for both general and special populations. It is shown that for both general and special populations a zero score excludes most clinically relevant coronary artery disease. The importan

  14. Electrolytic deposition of calcium phosphate/chitosan coating on titanium alloy: Growth kinetics and influence of current density, acetic acid, and chitosan

    NARCIS (Netherlands)

    Wang, Jiawei; Apeldoorn, van Aart; Groot, de Klaas

    2006-01-01

    Electrolytically deposited calcium phosphate/chitosan coating demonstrated good bone marrow stromal cell attachment. The aim of this study was to understand the coating's growth kinetics as well as the effects of current density, acetic acid, and chitosan on the coating's formation. The scanning ele

  15. Orexin-A potentiates L-type calcium/barium currents in rat retinal ganglion cells.

    Science.gov (United States)

    Liu, F; Weng, S-J; Yang, X-L; Zhong, Y-M

    2015-10-01

    Two neuropeptides, orexin-A and orexin-B (also called hypocretin-1 and -2), have been implicated in sleep/wake regulation, feeding behaviors via the activation of two subtypes of G-protein-coupled receptors: orexin 1 and orexin 2 receptors (OX1R and OX2R). While the expression of orexins and orexin receptors is immunohistochemically revealed in retinal neurons, the function of these peptides in the retina is largely unknown. Using whole-cell patch-clamp recordings in rat retinal slices, we demonstrated that orexin-A increased L-type-like barium currents (IBa,L) in ganglion cells (GCs), and the effect was blocked by the selective OX1R antagonist SB334867, but not by the OX2R antagonist TCS OX2 29. The orexin-A effect was abolished by intracellular dialysis of GDP-β-S/GPAnt-2A, a Gq protein inhibitor, suggesting the mediation of Gq. Additionally, during internal dialysis of the phosphatidylinositol (PI)-phospholipase C (PLC) inhibitor U73122, orexin-A did not change the IBa,L of GCs, whereas the orexin-A effect persisted in the presence of the phosphatidylcholine (PC)-PLC inhibitor D609. The orexin-A-induced potentiation was not seen with internal infusion of Ca(2+)-free solution or when inositol 1,4,5-trisphosphate (IP3)-sensitive Ca(2+) release from intracellular stores was blocked by heparin/xestospongins-C. Moreover, the orexin-A effect was mimicked by the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate, but was eliminated when PKC was inhibited by bisindolylmaleimide IV (Bis-IV)/Gö6976. Neither adenosine 3',5'-cyclic monophosphate (cAMP)-protein kinase A (PKA) nor guanosine 3',5'-cyclic monophosphate (cGMP)-protein kinase G (PKG) signaling pathway was likely involved, as orexin-A persisted to potentiate the IBa,L of GCs no matter these two pathways were activated or inhibited. These results suggest that, by activating OX1R, orexin-A potentiates the IBa,L of rat GCs through a distinct Gq/PI-PLC/IP3/Ca(2+)/PKC signaling pathway.

  16. Effects of Ginkgolide B on action potential and calcium,potassium current in guinea pig ventricular myocytes

    Institute of Scientific and Technical Information of China (English)

    Xiao-yan QI; Zhi-xiong ZHANG; You-qiu XU

    2004-01-01

    AIM: To investigate the effect of Ginkgolide B (GB) on action potential (AP), delayed rectifier potassium current (IK), and L-type calcium current (ICa-L) in guinea pig ventricular myocytes. METHODS: Single ventricular myocytes were isolated by an enzymatic dissociation method. AP, IK, ICa-L were recorded by whole-cell patch-clamp technique in either current or voltage clamp mode. RESULTS: GB shortened APD in a concentration-dependent manner. GB 0.1, 1, and 10 μmol/L shortened APD50 by 7.9 % (n=5, P>0.05), 18.4 % (n=5, P<0.01), and 28.9 % (n=6, P<0.01), respectively; APD90 by 12.4 % (n=5, P>0.05), 17.6 % (n=5, P<0.01), 26.4 % (n=5, P<0.01),respectively. GB increased IK in a concentration-dependent manner. GB 0.1, 1, and l0 μmol/L increased IK by 20.1% (n=6, P<0.05), 43.1% (n=6, P<0.01), 55.6 % (n=6, P<0.05); increased IKtail by 10.7 % (n=6, P<0.05),25.1% (n=6, P<0.05), and 37.7 % (n=6, P<0.05), respectively at testing potential of +50 mV and shift the I-V curve of Ik upward. But GB had no significant effect on ICa-L at above concentrations. CONCLUSION: GB significantly shortened APD in a concentration-dependent manner which mainly due to increase of IK.

  17. The calcium current activated by T cell receptor and store depletion in human lymphocytes is absent in a primary immunodeficiency.

    Science.gov (United States)

    Partiseti, M; Le Deist, F; Hivroz, C; Fischer, A; Korn, H; Choquet, D

    1994-12-23

    Stimulation of antigen receptors of lymphocytes triggers a transitory release of Ca2+ from internal stores and the opening of a transmembrane Ca2+ conductive pathway. The latter underlies the sustained increase of intracellular free calcium concentration, and it seems to be a key event in the Ca(2+)-dependent biochemical cascade leading to T cell proliferation. Alternatively, pharmacological depletion of internal stores by itself activates Ca2+ influx. This has led to the hypothesis that antigen-triggered Ca2+ influx is secondary to Ca2+ release from internal stores. However, the precise relationship between antigen and Ca2+ release-activated Ca2+ currents remains unclear, particularly since neither of them has been electrophysiologically recorded in normal lymphocytes. Using the whole-cell and the perforated configurations of the patch clamp technique on peripheral blood lymphocytes, we found that a low amplitude Ca(2+)-selective current was triggered when intracellular stores were depleted by stimuli such as the intracellular perfusion of inositol triphosphate or thapsigargin and the extracellular perfusion of ionomycin. A similar current was elicited by the cross-linking of the T cell receptor-CD3 complex. This current displayed an inward rectification below 0 mV and was completely blocked by the divalent cation Cd2+. It was very selective for Ca2+ over Na+ and insensitive to changes in chloride concentration. The physiological relevance of this conductance was investigated with the analysis of abnormal Ca2+ signaling in lymphocytes from a patient suffering from a primary immunodeficiency associated with a defective T cell proliferation. Using fura-2 video imaging, an absence of Ca2+ influx was established in the patient's lymphocytes, whereas the Ca2+ release from internal stores was normal. This was the case whether cells were stimulated physiologically through their antigen receptors or with store depleting pharmacological agents. Most importantly, no Ca(2

  18. Temperature and voltage dependence of barrier height and ideality factor in Au/0.07 graphene-doped PVA/n-Si structures

    Science.gov (United States)

    Altındal Yerişkin, S.; Balbaşı, M.; Demirezen, S.

    2017-01-01

    In this study, Au/0.07 graphene-doped PVA/n-Si structures were fabricated and current conduction mechanism in these structures were investigated in the temperature range of 80-380 K through forward bias current-voltage (I-V) measurements. Main electrical parameters were extracted from I-V data. Zero-bias barrier height (overline{Φ}_{B0} ) and ideality factor (n) were found strong functions of temperature and their values ranged from 0.234 eV and 4.98 (at 80 K) to 0.882 eV and 1.15 (at 380 K), respectively. Φ ap versus q/2kT plot was drawn to obtain an evidence of a Gaussian distribution of the barrier heights (BHs) and it revealed two distinct linear regions with different slopes and intercepts. The mean values of BH (Φ Bo) and zero-bias standard deviation (σ s ) were obtained from the intercept and slope of the linear regions of this plot as 1.30 eV and 0.16 V for the first region (280-380 K) and 0.74 eV and 0.085 V for the second region (80-240 K), respectively. Thus, the values of overline{Φ}_{B0} and effective Richardson constant (A*) were also found from the intercept and slope of the modified Richardson plot [ln(I s /T 2) - q 2 σ {/o 2} /2k 2 T 2 vs q/kT] as 1.31 eV and 130 A/cm2 K2 for the first region and 0.76 eV and 922 A/cm2 K2 for the second region, respectively. The value of A* for the first region was very close to the theoretical value for n-Si (112 A/cm2 K2). The energy density distribution profile of surface states (Nss) was also extracted from the forward bias I-V data by taking into account voltage dependent effective BH (Φe) and n.

  19. Time-resolved photoluminescence measurements for determining voltage-dependent charge-separation efficiencies of subcells in triple-junction solar cells

    Science.gov (United States)

    Tex, David M.; Ihara, Toshiyuki; Akiyama, Hidefumi; Imaizumi, Mitsuru; Kanemitsu, Yoshihiko

    2015-01-01

    Conventional external quantum-efficiency measurement of solar cells provides charge-collection efficiency for approximate short-circuit conditions. Because this differs from actual operating voltages, the optimization of high-quality tandem solar cells is especially complicated. Here, we propose a contactless method, which allows for the determination of the voltage dependence of charge-collection efficiency for each subcell independently. By investigating the power dependence of photoluminescence decays, charge-separation and recombination-loss time constants are obtained. The upper limit of the charge-collection efficiencies at the operating points is then obtained by applying the uniform field model. This technique may complement electrical characterization of the voltage dependence of charge collection, since subcells are directly accessible.

  20. Venom of the Chilean Latrodectus mactans alters bovine spermatozoa calcium and function by blocking the TEA-sensitive K(+) current.

    Science.gov (United States)

    Navarrete, Patricia; Martínez-Torres, Ataúlfo; Gutiérrez, Raúl Sánchez; Mejía, Fernando Romero; Parodi, Jorge

    2010-08-01

    The morphology and size of spermatozoa make it difficult to study the functional properties of the plasma membrane, however, some studies have revealed the presence of a number of ion channels in this cell. We measured the calcium (Ca(++)) influx induced by depolarization of the plasma membrane and by venom isolated from the Chilean black widow spider (Latrodectus mactans), and functional changes in the presence of either high potassium or total venom. Our results indicate that the venom increased the Ca(++) influx, with an EC50 of 6.1 microg/mL and triggering the acrosome reaction in 43.26% of the cells. The application of potassium (10 mM K(+)) or total venom (10 microg/mL) did not affect the morphology or DNA stability of the sperm. The effects induced by high K(+) and venom suggest that direct blocking of K(+) currents alters the passive properties of the plasma membrane, leading to the entry of Ca(++). These results show the importance of functional changes induced by depolarizing the spermatozoa and by venom. This venom possesses one or more molecules that may be used as pharmacological tools for studies on spermatozoa and have potential applications in reproductive biotechnology.

  1. Enhanced effect of VEGF165 on L-type calcium currents in guinea-pig cardiac ventricular myocytes.

    Science.gov (United States)

    Xing, Wenlu; Gao, Chuanyu; Qi, Datun; Zhang, You; Hao, Peiyuan; Dai, Guoyou; Yan, Ganxin

    2017-01-01

    The mechanisms of vascular endothelial growth factor 165 (VEGF165) on electrical properties of cardiomyocytes have not been fully elucidated. The aim of this study is to test the hypothesis that VEGF165, an angiogenesis-initiating factor, affects L-type calcium currents (ICa,L) and cell membrane potential in cardiac myocytes by acting on VEGF type-2 receptors (VEGFR2). ICa,L and action potentials (AP) were recorded by the whole-cell patch clamp method in isolated guinea-pig ventricular myocytes treated with different concentrations of VEGF165 proteins. Using a VEGFR2 inhibitor, we also tested the receptor of VEGF165 in cardiomyocytes. We found that VEGF165 increased ICa,L in a concentration-dependent manner. SU5416, a VEGFR2 inhibitor, almost completely eliminated VEGF165-induced ICa,L increase. VEGF165 had no significant influence on action potential 90 (APD90) and other properties of AP. We conclude that in guinea-pig ventricular myocytes, ICa,L can be increased by VEGF165 in a concentration-dependent manner through binding to VEGFR2 without causing any significant alteration to action potential duration. Results of this study may further expound the safety of VEGF165 when used in the intervention of heart diseases.

  2. Investigation into in vitro anti-leishmanial combinations of calcium channel blockers and current anti-leishmanial drugs

    Directory of Open Access Journals (Sweden)

    Juliana Quero Reimão

    2011-12-01

    Full Text Available The need for drug combinations to treat visceral leishmaniasis (VL arose because of resistance to antimonials, the toxicity of current treatments and the length of the course of therapy. Calcium channel blockers (CCBs have shown anti-leishmanial activity; therefore their use in combination with standard drugs could provide new alternatives for the treatment of VL. In this work, in vitro isobolograms of Leishmania (Leishmania chagasi using promastigotes or intracellular amastigotes were utilised to identify the interactions between five CCBs and the standard drugs pentamidine, amphotericin B and glucantime. The drug interactions were assessed with a fixed ratio isobologram method and the fractional inhibitory concentrations (FICs, sum of FICs (ΣFICs and the overall mean ΣFIC were calculated for each combination. Graphical isobologram analysis showed that the combination of nimodipine and glucantime was the most promising in amastigotes with an overall mean ΣFIC value of 0.79. Interactions between CCBs and the anti-leishmanial drugs were classified as indifferent according to the overall mean ΣFIC and the isobologram graphic analysis.

  3. Calcium Electroporation

    DEFF Research Database (Denmark)

    Frandsen, Stine Krog; Gibot, Laure; Madi, Moinecha;

    2015-01-01

    BACKGROUND: Calcium electroporation describes the use of high voltage electric pulses to introduce supraphysiological calcium concentrations into cells. This promising method is currently in clinical trial as an anti-cancer treatment. One very important issue is the relation between tumor cell kill...... efficacy-and normal cell sensitivity. METHODS: Using a 3D spheroid cell culture model we have tested the effect of calcium electroporation and electrochemotherapy using bleomycin on three different human cancer cell lines: a colorectal adenocarcinoma (HT29), a bladder transitional cell carcinoma (SW780......), and a breast adenocarcinoma (MDA-MB231), as well as on primary normal human dermal fibroblasts (HDF-n). RESULTS: The results showed a clear reduction in spheroid size in all three cancer cell spheroids three days after treatment with respectively calcium electroporation (p

  4. β-Adrenoceptor activation enhances L-type calcium channel currents in anterior piriform cortex pyramidal cells of neonatal mice: implication for odor learning.

    Science.gov (United States)

    Ghosh, Abhinaba; Mukherjee, Bandhan; Chen, Xihua; Yuan, Qi

    2017-03-01

    Early odor preference learning occurs in one-week-old rodents when a novel odor is paired with a tactile stimulation mimicking maternal care. β-Adrenoceptors and L-type calcium channels (LTCCs) in the anterior piriform cortex (aPC) are critically involved in this learning. However, whether β-adrenoceptors interact directly with LTCCs in aPC pyramidal cells is unknown. Here we show that pyramidal cells expressed significant LTCC currents that declined with age. β-Adrenoceptor activation via isoproterenol age-dependently enhanced LTCC currents. Nifedipine-sensitive, isoproterenol enhancement of calcium currents was only observed in post-natal day 7-10 mice. APC β-adrenoceptor activation induced early odor preference learning was blocked by nifedipine coinfusion.

  5. Attenuation of G protein-mediated inhibition of N-type calcium currents by expression of caveolins in mammalian NG108–15 cells

    Science.gov (United States)

    Toselli, M; Taglietti, V; Parente, V; Flati, S; Pavan, A; Guzzi, F; Parenti, M

    2001-01-01

    Caveolins are integral proteins of glycolipid/cholesterol-rich plasmalemmal caveolae domains, where, they may function as a plasma membrane scaffold onto which many classes of signalling molecules, including receptors and heterotrimeric G proteins, can assemble. To ascertain whether caveolins influence G protein-mediated signal transduction, we stably expressed caveolin-1 and −3 isoforms in the neuroblastoma × glioma NG108–15 hybrid cell line, lacking endogenous caveolins. Subsequently, using whole-cell voltage clamp methods, we examined whether the modulation of N-type voltage-gated Ca2+ channels by Go protein-coupled, δ-type opioid receptors might be affected by recombinant caveolin expression. In transfected NG108–15 cells, caveolins localized at the plasma membrane and, upon subcellular fractionation on sucrose density gradients, they co-localized in Triton-resistant, low buoyancy fractions, with endogenous Go protein α-subunits. The voltage-dependent inhibition of ω-conotoxin GVIA-sensitive Ba2+ currents following either activation of δ-opioid receptors by the agonist [o-pen2,o-pen5]-enkephalin (DPDPE), or direct stimulation of G proteins with guanosine 5′-O-(thiotriphosphate) (GTPγS) was significantly attenuated in caveolin-expressing cells. The kinetics of Ca2+ channel inhibition were also modified by caveolins. Overall, these results suggest that caveolins may negatively affect G protein-dependent regulation of voltage-gated N-type Ca2+ channels, presumably by causing a reduction of the available pool of activated G proteins. PMID:11600672

  6. Conopressin affects excitability, firing, and action potential shape through stimulation of transient and persistent inward currents in mulluscan neurons.

    Science.gov (United States)

    van Soest, P F; Kits, K S

    1998-04-01

    The molluscan vasopressin/oxytocin-related neuropeptide conopressin activates two persistent inward currents in neurons from the anterior lobe of the right cerebral ganglion of Lymnaea stagnalis that are involved in the control of male copulatory behavior. The low-voltage-activated (LVA) current is activated at a wide range of membrane potentials, its amplitude being only weakly voltage dependent. The high-voltage-activated (HVA) current is activated at potentials positive to -40 mV only and shows a steep voltage dependence. Occurrence of both currents varies from cell to cell, some expressing both and others only the HVA current. In most neurons that have the LVA current, a conopressin-independent persistent inward current (INSR) is found that resembles the HVA current in its voltage dependence. The functional importance of the LVA and HVA currents was studied under current-clamp conditions in isolated anterior lobe neurons. In cells exhibiting both current types, the effect of activation of the LVA current alone was investigated as follows: previously recorded LVA current profiles were injected into the neurons, and the effects were compared with responses induced by conopressin. Both treatments resulted in a strong depolarization and firing activity. No differences in firing frequency and burst duration were observed, indicating that activation of the LVA current is sufficient to evoke bursts. In cells exhibiting only the HVA current, the effect of conopressin on the response to a depolarizing stimulus was tested. Conopressin reversibly increased the number of action potentials generated by the stimulus, suggesting that the HVA current enhances excitability and counteracts accommodation. Conopressin enhanced action potential broadening during depolarizing stimuli in many neurons. Voltage-clamp experiments performed under ion-selective conditions revealed the presence of transient sodium and calcium currents. Using the action potential clamp technique, it was

  7. Voltage-dependent capacitance behavior and underlying mechanisms in metal-insulator-metal capacitors with Al2O3-ZrO2-SiO2 nano-laminates

    Science.gov (United States)

    Zhu, Bao; Liu, Wen-Jun; Wei, Lei; Ding, Shi-Jin

    2016-04-01

    Nano-laminates consisting of high-permittivity dielectrics and SiO2 have been extensively studied for radio frequency metal-insulator-metal (MIM) capacitors because of their superior voltage linearity and low leakage current. However, there are no reports on the capacitance-voltage (C-V) characteristics at a high sweep voltage range. In this work, an interesting variation in the voltage-dependent capacitance that forms a ‘ω’-like shape is demonstrated for the MIM capacitors with Al2O3/ZrO2/SiO2 nano-laminates. As the thickness ratio of the SiO2 film to the total insulator increases to around 0.15, the C-V curve changes from an upward parabolic shape to a ‘ω’ shape. This can be explained based on the competition between the orientation polarization from SiO2 and the electrode polarization from Al2O3 and ZrO2. When the SiO2 film is very thin, the electrode polarization dominates in the MIM capacitor, generating a positive curvature C-V curve. When the thickness of SiO2 is increased, the orientation polarization is enhanced and thus both polarizations are operating in the MIM capacitors. This leads to the appearance of a multiple domain C-V curve containing positive and negative curvatures. Therefore, good consistency between the experimental results and the theoretical simulations is demonstrated. Such voltage-dependent capacitance behavior is not determined by the stack structure of the insulator, measurement frequency and oscillator voltage, but by the thickness ratio of the SiO2 film to the whole insulator. These findings are helpful to engineer MIM capacitors with good voltage linearity.

  8. Effects of unsaturated fatty acids on calcium-activated potassium current in gastric myocytes of guinea pigs

    Institute of Scientific and Technical Information of China (English)

    Hai-Feng Zheng; Xiang-Lan Li; Zheng-Yuan Jin; Jia-Bin Sun; Zai-Liu Li; Wen-Xie Xu

    2005-01-01

    AIM: To investigate the effects of exogenous unsaturated fatty acids on calcium-activated potassium current [Ik(Ca)]in gastric antral circular myocytes of guinea pigs.METHODS: Gastric myocytes were isolated by collagenase from the antral circular layer of guinea pig stomach. The whole-cell patch clamp technique was used to record Ik(Ca)in the isolated single smooth muscle cells with or without different concentrations of arachidonic acid (AA), linoleic acid (LA), and oleic acid (OA).RESULTS: AA at concentrations of 2,5 and 10 μmol/L markedly increased IK(ca)in a dose-dependent manner. LA at concentrations of 5, 10 and 20 μmol/L also enhanced IK(Ca)in a dose-dependent manner. The increasing potency of AA, LA, and oleic acid (OA) on Ik(Ca) at the same concentration(10 μmol/L) was in the order of AA>LA>OA. AA (10 μmol/L)-induced increase of Ik(Ca) was not blocked by H-7 (10 μmol/L), an inhibitor of protein kinase C (PKC), or indomethacin (10 μmol/L),an inhibitor of the cyclooxygenase pathway, and 17-octadecynoic acid (10 μmol/L), an inhibitor of the cytochrome P450 pathway, but weakened by nordihydroguaiaretic acid(10 μmol/L), an inhibitor of the lipoxygenase pathway.CONCLUSION: Unsaturated fatty acids markedly increase Ik(Ca), and the enhancing potencies are related to the number of double bonds in the fatty acid chain. The lipoxygenase pathway of unsaturated fatty acid metabolism is involved in the unsaturated fatty acid-induced increase of IK(Ca) in gastric antral circular myocytes of guinea pigs.

  9. Contribution of downregulation of L-type calcium currents to delayed neuronal death in rat hippocampus after global cerebral ischemia and reperfusion.

    Science.gov (United States)

    Li, Xiao-Ming; Yang, Jian-Ming; Hu, De-Hui; Hou, Feng-Qing; Zhao, Miao; Zhu, Xin-Hong; Wang, Ying; Li, Jian-Guo; Hu, Ping; Chen, Liang; Qin, Lu-Ning; Gao, Tian-Ming

    2007-05-09

    Transient forebrain ischemia induces delayed, selective neuronal death in the CA1 region of the hippocampus. The underlying molecular mechanisms are as yet unclear, but it is known that activation of L-type Ca2+ channels specifically increases the expression of a group of genes required for neuronal survival. Accordingly, we examined temporal changes in L-type calcium-channel activity in CA1 and CA3 pyramidal neurons of rat hippocampus after transient forebrain ischemia by patch-clamp techniques. In vulnerable CA1 neurons, L-type Ca2+-channel activity was persistently downregulated after ischemic insult, whereas in invulnerable CA3 neurons, no change occurred. Downregulation of L-type calcium channels was partially caused by oxidation modulation in postischemic channels. Furthermore, L-type but neither N-type nor P/Q-type Ca2+-channel antagonists alone significantly inhibited the survival of cultured hippocampal neurons. In contrast, specific L-type calcium-channel agonist remarkably reduced neuronal cell death and restored the inhibited channels induced by nitric oxide donor. More importantly, L-type calcium-channel agonist applied after reoxygenation or reperfusion significantly decreased neuronal injury in in vitro oxygen-glucose deprivation ischemic model and in animals subjected to forebrain ischemia-reperfusion. Together, the present results suggest that ischemia-induced inhibition of L-type calcium currents may give rise to delayed death of neurons in the CA1 region, possibly via oxidation mechanisms. Our findings may lead to a new perspective on neuronal death after ischemic insult and suggest that a novel therapeutic approach, activation of L-type calcium channels, could be tested at late stages of reperfusion for stroke treatment.

  10. Involvement of reactive oxygen species and high-voltage-activated calcium currents in nanoparticle zinc oxide-induced cytotoxicity in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Zhao Jingxia [Nankai University, College of Medicine (China); Yao Yang [Tianjin First Central Hospital (China); Liu Shichang [Nankai University, College of Medicine (China); Zhang Tao [Nankai University, College of Life Science (China); Ren Guogang [University of Hertfordshire, Science and Technology Research Institute (United Kingdom); Yang Zhuo, E-mail: zhuoyang@nankai.edu.cn [Nankai University, College of Medicine (China)

    2012-11-15

    This study was to determine the possible neurotoxicity and mechanisms underlying the effects of nano-ZnO with sizes of 20-80 nm on central nervous system (CNS). The cytotoxicity of nano-ZnO was investigated in PC12 cells. The viability of cells was observed by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and the generation of reactive oxygen species (ROS) for cells was evaluated by a fluorometry assay. The apoptosis of cells was detected and analyzed by flow cytometry. In addition, effects of nano-ZnO on the properties of high-voltage-activated (HVA) calcium currents were studied in acutely isolated rat hippocampal pyramidal neurons using the whole-cell patch clamp technique. The results of MTT assay showed that nano-ZnO (10{sup -4} g/mL) caused a significant decrease in cell viability (P < 0.05). Nano-ZnO induced intracellular accumulation of ROS and the apoptosis of PC12 cells with the increasing concentration of nano-ZnO in flow cytometric assay (P < 0.05). Further results of electrophysiological recording indicated that 10{sup -4} g/mL nano-ZnO first altered the current-voltage curve and the peak amplitudes of HVA calcium currents at 10 min of the recording, and the peak current amplitudes were increased significantly at the end of 30 min (P < 0.05). All these results suggested that the increase of intracellular ROS was one of potential mechanisms of cellular apoptosis induced by nano-ZnO. Nano-ZnO could cause the elevation of cytosolic calcium levels by enhancement of HVA calcium currents, which would increase the generation of intracellular ROS, and consequently promote the neuronal apoptosis.

  11. Long-term effects of imidapril on calcium and potassium currents in rabbit left ventricular hypertrophic myocytes

    Institute of Scientific and Technical Information of China (English)

    李泱; 陆再英; 肖建民; 张存泰; 马杰; 刘念

    2003-01-01

    Objective To investigate the long-term effects of imidapril (IMI) on action potential and calcium and potassium currents in rabbit left ventricular hypertrophic myocytes.Methods Rabbits were randomly divided into three groups: IMI-treated, hypertrophic and sham-operated control groups. Cardiac hypertrophy was induced in hypertrophy group by partial ligation of the abdominal aorta. In the IMI-treated group, the rabbits were administered IMI (1.5 mg·kg-1·d-1) for 8 weeks after surgery. In the sham-operated control group, the animals underwent an abdominal laparotomy without further procedure. Whole-cell patch clamp technique was used to record ionic currents.Results Membrane capacitance was larger in hypertrophic cells than in sham-operated cells or IMI-treated cells. Action potential duration was lengthened in hypertrophic cells and was remarkably shortened by IMI. The density of ICa,L was reduced from 12.8±0.7 pA/pF in the sham-operated cells, to 7.7±0.8 pA/pF in hypertrophic cells, while it resembled the control cells after IMI treatment (11.9±1.0 pA/pF). After IMI treatment, the density of IKs,tail was enhanced from 2.5±0.1 pA/pF in hypertrophic cells to 4.7±0.6 pA/pF (n=7, P<0.01), which was similar to the sham-operated cells. The densities of Ito and IK1 were significantly increased in IMI-treated cells, from 3.8±0.4 pA/pF and 3.7±0.5 pA/pF in the hypertrophic cells to 6.4±0.8 pA/pF and 6.5±0.3 pA/pF, respectively, but the IKr densities were not different in the three groups.Conclusion IMI could reverse the increase in membrane capacitance in hypertrophic cells, shorten action potential duration, and increase the densities of ICa, L, IKs, Ito and IK1 in hypertrophic cells.

  12. Calcium channels, neuromuscular synaptic transmission and neurological diseases.

    Science.gov (United States)

    Urbano, Francisco J; Pagani, Mario R; Uchitel, Osvaldo D

    2008-09-15

    Voltage-dependent calcium channels are essential in neuronal signaling and synaptic transmission, and their functional alterations underlie numerous human disorders whether monogenic (e.g., ataxia, migraine, etc.) or autoimmune. We review recent work on Ca(V)2.1 or P/Q channelopathies, mostly using neuromuscular junction preparations, and focus specially on the functional hierarchy among the calcium channels recruited to mediate neurotransmitter release when Ca(V)2.1 channels are mutated or depleted. In either case, synaptic transmission is greatly compromised; evidently, none of the reported functional replacements with other calcium channels compensates fully.

  13. Effects of in vitro lead exposure on voltage-sensitive calcium channels differ among cell types in central neurons of Lymnaea stagnalis.

    Science.gov (United States)

    Audesirk, G; Audesirk, T

    1989-01-01

    The effects of acute in vitro lead exposure on slowly inactivating voltage-sensitive calcium channels in central neurons of the freshwater pond snail Lymnaea stagnalis were studied under voltage clamp. Three physiologically distinct cell types were used: two subsets of the B cell cluster (Bpos and Bneg) and the pedal giant neuron (RPeD1). In Bpos neurons, 5 nM free Pb2+ irreversibly inhibited current flow through calcium channels by 38 +/- 10%. In Bneg neurons, 5 nM free Pb2+ slightly inhibited inward currents (12 +/- 6%) and may have shifted their voltage dependence to more depolarized voltages. The inhibition and voltage shift were irreversible. In RPeD1 neurons, Pb2+ caused a small, statistically insignificant inhibition of inward current (5 nM free Pb2+; 18 +/- 19%; 30 nM free Pb2+: 31 +/- 23%). The effects of Pb2+ were fully reversible. These data indicate that (1) voltage-sensitive calcium channels in Lymnaea neurons are inhibited by nanomolar concentrations of free Pb2+; (2) there are multiple types of calcium channels in Lymnaea neurons; and (3) the effects of in vitro lead exposure differ qualitatively among channel types.

  14. Role of a T-type calcium current in supporting a depolarizing potential, damped oscillations, and phasic firing in vasopressinergic guinea pig supraoptic neurons.

    Science.gov (United States)

    Erickson, K R; Ronnekleiv, O K; Kelly, M J

    1993-05-01

    Guinea pig magnocellular neurosecretory cells (MNCs) of the supraoptic nucleus (SON) were studied using the in vitro slice preparation. Intracellular recordings were made with biocytin-filled electrodes, permitting immunocytochemical identification of the recorded cells as arginine vasopressin- (AVP) versus oxytocin- (OT) containing. Only AVP cells displaying a depolarizing potential (DP) fired phasically. The DP was associated with a transient inward current measured in voltage clamp, which exhibited a number of properties of the T-type calcium current: activation threshold of -64 mV, time course of up to 250 ms, blockade by nickel and augmentation by barium chloride. This current has not been reported previously in SON neurons. The T-type current (IT) was always associated with a damped oscillation of the membrane following the offset from hyperpolarizing steps. In all cells tested, an apamin-sensitive afterhyperpolarization (AHP) was observed, similar to the calcium-dependent potassium current (IK, Ca) described in rat SON and other CNS regions. Therefore, as with other CNS regions displaying damped oscillations, guinea pig SON cells possess both an IT and an IK, Ca. We have previously described an Ih activating at hyperpolarized potentials in these cells, which depolarizes the membrane to a range in which the IT and IK, Ca can interactively support oscillations. In summary, the IT and associated depolarizing potential appears to be a requisite feature for phasic firing in AVP cells of guinea pig SON.

  15. Rare mutations of CACNB2 found in autism spectrum disease-affected families alter calcium channel function.

    Directory of Open Access Journals (Sweden)

    Alexandra F S Breitenkamp

    Full Text Available Autism Spectrum Disorders (ASD are complex neurodevelopmental diseases clinically defined by dysfunction of social interaction. Dysregulation of cellular calcium homeostasis might be involved in ASD pathogenesis, and genes coding for the L-type calcium channel subunits CaV1.2 (CACNA1C and CaVβ2 (CACNB2 were recently identified as risk loci for psychiatric diseases. Here, we present three rare missense mutations of CACNB2 (G167S, S197F, and F240L found in ASD-affected families, two of them described here for the first time (G167S and F240L. All these mutations affect highly conserved regions while being absent in a sample of ethnically matched controls. We suggest the mutations to be of physiological relevance since they modulate whole-cell Ba2+ currents through calcium channels when expressed in a recombinant system (HEK-293 cells. Two mutations displayed significantly decelerated time-dependent inactivation as well as increased sensitivity of voltage-dependent inactivation. In contrast, the third mutation (F240L showed significantly accelerated time-dependent inactivation. By altering the kinetic parameters, the mutations are reminiscent of the CACNA1C mutation causing Timothy Syndrome, a Mendelian disease presenting with ASD. In conclusion, the results of our first-time biophysical characterization of these three rare CACNB2 missense mutations identified in ASD patients support the hypothesis that calcium channel dysfunction may contribute to autism.

  16. Structural mapping of the voltage-dependent sodium channel. Distance between the tetrodotoxin and Centruroides suffusus suffusus II beta-scorpion toxin receptors.

    Science.gov (United States)

    Darbon, H; Angelides, K J

    1984-05-25

    A 7- dimethylaminocoumarin -4-acetate fluorescent derivative of toxin II from the venom of the scorpion Centruroides suffusus suffusus (Css II) has been prepared to study the structural, conformational, and cellular properties of the beta-neurotoxin receptor site on the voltage-dependent sodium channel. The derivative retains high affinity for its receptor site on the synaptosomal sodium channel with a KD of 7 nM and site capacity of 1.5 pmol/mg of synaptosomal protein. The fluorescent toxin is very environmentally sensitive and the fluorescence emission upon binding indicates that the Css II receptor is largely hydrophobic. Binding of tetrodotoxin or batrachotoxin does not alter the spectroscopic properties of bound Css II, whereas toxin V from Leiurus quinquestriatus effects a 10-nm blue shift to a more hydrophobic environment. This is the first direct indication of conformational coupling between these separate neurotoxin receptor sites. The distance between the tetrodotoxin and Css II scorpion toxin receptors on the sodium channel was measured by fluorescence resonance energy transfer. Efficiencies were measured by both donor quenching and acceptor-sensitized emission. The distance between these two neurotoxin sites is about 34 A. The implications of these receptor locations together with other known molecular distances are discussed in terms of a molecular structure of the voltage-dependent sodium channel.

  17. Current perspectives of bio-ceramic technology in endodontics: calcium enriched mixture cement - review of its composition, properties and applications

    OpenAIRE

    Utneja, Shivani; Nawal, Ruchika Roongta; Talwar, Sangeeta; Verma, Mahesh

    2014-01-01

    Advancements in bio-ceramic technology has revolutionised endodontic material science by enhancing the treatment outcome for patients. This class of dental materials conciliates excellent biocompatibility with high osseoconductivity that render them ideal for endodontic care. Few recently introduced bio-ceramic materials have shown considerable clinical success over their early generations in terms of good handling characteristics. Calcium enriched mixture (CEM) cement, Endosequence sealer, a...

  18. Comparison of the effects of nitric oxide and hydrogen sulfide on calcium and potassium currents of rat colonic smooth muscle%一氧化氮和硫化氢对大鼠结肠平滑肌钙钾电流影响的比较

    Institute of Scientific and Technical Information of China (English)

    全晓静; 罗和生; 陈炜; 崔凝; 夏虹; 余光

    2015-01-01

    [(-3.76 ± 0.66) pA/pF] and [(-4.13±0.29) pA/pF] to[(-2.67±0.42) pA/pF]and[(-2.73±0.76) pA/pF], respectively (P < 0.05).SNP had no effect on the voltage-dependent property of ICa, L, While the Ⅰ-Ⅴ relationship of L-type calcium channels was modified by NaHS, the peak of Ⅰ-Ⅴ curve was shifted to right.SNP did not affect the curves of the steady-state activation, but evoked a leftward shift of the steady-state inactivation curve.However, NaHS evoked a significant rightward shift of the steady-state activation curve and inhibited the inactivation of L-type calcium channels (P < 0.05).BKCa channel currents were significantly increased by SNP, the current density was increased from [(12.7 ± 1.9) pA/pF] to[(14.7 ± 2.1) pA/pF] (P < 0.05);while inhibited by NaHS (control [(15.5 ± 2.4) pA/pF] vs.NaHS [(12.4 ± 2.9) pA/pF] (P < 0.05).Conclusion Both NO and H2S inhibit the contraction of colonic SMCs, while the mechanisms are different.The relaxant effect of NO on rat colonic muscle is due to direct inhibition of L type calcium channels and activation of BKCa channels.The relaxant effect of H2S on colonic muscle may be associated with the direct inhibition of H2S on L-type calcium channels.H2S may involve in the regulation of calcium homeostasis in colonic SMCs of rat colon.

  19. Local calcium elevation and cell elongation initiate guided motility in electrically stimulated osteoblast-like cells.

    Directory of Open Access Journals (Sweden)

    Nurdan Ozkucur

    Full Text Available BACKGROUND: Investigation of the mechanisms of guided cell migration can contribute to our understanding of many crucial biological processes, such as development and regeneration. Endogenous and exogenous direct current electric fields (dcEF are known to induce directional cell migration, however the initial cellular responses to electrical stimulation are poorly understood. Ion fluxes, besides regulating intracellular homeostasis, have been implicated in many biological events, including regeneration. Therefore understanding intracellular ion kinetics during EF-directed cell migration can provide useful information for development and regeneration. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed the initial events during migration of two osteogenic cell types, rat calvarial and human SaOS-2 cells, exposed to strong (10-15 V/cm and weak (< or = 5 V/cm dcEFs. Cell elongation and perpendicular orientation to the EF vector occurred in a time- and voltage-dependent manner. Calvarial osteoblasts migrated to the cathode as they formed new filopodia or lamellipodia and reorganized their cytoskeleton on the cathodal side. SaOS-2 cells showed similar responses except towards the anode. Strong dcEFs triggered a rapid increase in intracellular calcium levels, whereas a steady state level of intracellular calcium was observed in weaker fields. Interestingly, we found that dcEF-induced intracellular calcium elevation was initiated with a local rise on opposite sides in calvarial and SaOS-2 cells, which may explain their preferred directionality. In calcium-free conditions, dcEFs induced neither intracellular calcium elevation nor directed migration, indicating an important role for calcium ions. Blocking studies using cadmium chloride revealed that voltage-gated calcium channels (VGCCs are involved in dcEF-induced intracellular calcium elevation. CONCLUSION/SIGNIFICANCE: Taken together, these data form a time scale of the morphological and physiological

  20. Current concepts in combination therapy for the treatment of hypertension: combined calcium channel blockers and RAAS inhibitors

    Directory of Open Access Journals (Sweden)

    Alberto F Rubio-Guerra

    2009-11-01

    Full Text Available Alberto F Rubio-Guerra1, David Castro-Serna2, Cesar I Elizalde Barrera2, Luz M Ramos-Brizuela21Metabolic and Research Clinic, 2Internal Medicine Department, Hospital General de Ticomán SS DF, MéxicoAbstract: Recent guidelines for the management of hypertension recommend target blood pressures <140/90 mmHg in hypertensive patients, or <130/80 mmHg in subjects with diabetes, chronic kidney disease, or coronary artery disease. Despite the availability and efficacy of antihypertensive drugs, most hypertensive patients do not reach the recommended treatment targets with monotherapy, making combination therapy necessary to achieve the therapeutic goal. Combination therapy with 2 or more agents is the most effective method for achieving strict blood pressure goals. Fixed-dose combination simplifies treatment, reduces costs, and improves adherence. There are many drug choices for combination therapy, but few data are available about the efficacy and safety of some specific combinations. Combination therapy of calcium antagonists and inhibitors of the renin-angiotensin-aldosterone system (RAAS are efficacious and safe, and have been considered rational by both the JNC 7 and the 2007 European Society of Hypertension – European Society of Cardiology guidelines for the management of arterial hypertension. The aim of this review is to discuss some relevant issues about the use of combinations with calcium channel blockers and RAAS inhibitors in the treatment of hypertension.Keywords: hypertension, calcium channel blockers, renin-angiotensin-aldosterone system inhibitors, fixed-dose combination, adherence

  1. Blockade of L-type calcium channel in myocardium and calcium-induced contractions of vascular smooth muscle by CPU 86017.

    Science.gov (United States)

    Dai, De-zai; Hu, Hui-juan; Zhao, Jing; Hao, Xue-mei; Yang, Dong-mei; Zhou, Pei-ai; Wu, Cai-hong

    2004-04-01

    To assess the blockade by CPU 86017 on the L-type calcium channels in the myocardium and on the Ca(2+)-related contractions of vascular smooth muscle. The whole-cell patch-clamp was applied to investigate the blocking effect of CPU 86017 on the L-type calcium current in isolated guinea pig myocytes and contractions by KCl or phenylephrine (Phe) of the isolated rat tail arteries were measured. Suppression of the L-type current of the isolated myocytes by CPU 86017 was moderate, in time- and concentration-dependent manner and with no influence on the activation and inactivation curves. The IC(50) was 11.5 micromol/L. Suppressive effect of CPU 86017 on vaso-contractions induced by KCl 100 mmol/L, phenylephrine 1 micromol/L in KH solution (phase 1), Ca(2+) free KH solution ( phase 2), and by addition of CaCl(2) into Ca(2+)-free KH solution (phase 3) were observed. The IC(50) to suppress vaso-contractions by calcium entry via the receptor operated channel (ROC) and voltage-dependent channel (VDC) was 0.324 micromol/L and 16.3 micromol/L, respectively. The relative potency of CPU 86017 to suppress vascular tone by Ca(2+) entry through ROC and VDC is 1/187 of prazosin and 1/37 of verapamil, respectively. The blocking effects of CPU 86017 on the L-type calcium channel of myocardium and vessel are moderate and non-selective. CPU 86017 is approximately 50 times more potent in inhibiting ROC than VDC.

  2. Blockade of L-type calcium channel in myocardium and calcium-induced contractions of vascular smooth muscle by by CPU 86017

    Institute of Scientific and Technical Information of China (English)

    De-zai DAI; Hui-juan HU; Jing ZHAO; Xue-mei HAO; Dong-mei YANG; Pei-ai ZHOU; Cai-hong WU

    2004-01-01

    AIM: To assess the blockade by CPU 86017 on the L-type calcium channels in the myocardium and on the Ca2+related contractions of vascular smooth muscle. METHODS: The whole-cell patch-clamp was applied to investigate the blocking effect of CPU 86017 on the L-type calcium current in isolated guinea pig myocytes and contractions by KC1 or phenylephrine (Phe) of the isolated rat tail arteries were measured. RESULTS: Suppression of the L-type current of the isolated myocytes by CPU 86017 was moderate, in time- and concentration-dependent manner and with no influence on the activation and inactivation curves. The IC50 was 11.5 μmol/L. Suppressive effect of CPU 86017 on vaso-contractions induced by KC1 100 mmol/L, phenylephrine I μmol/Lin KH solution (phase 1),Ca2+ free KH solution ( phase 2), and by addition of CaCI2 into Ca2+-free KH solution (phase 3) were observed. The IC50 to suppress vaso-contractions by calcium entry via the receptor operated channel (ROC) and Voltage-dependent channel (VDC) was 0.324 μmol/L and 16.3 μmol/L, respectively. The relative potency of CPU 86017 to suppress vascular tone by Ca2+ entry through ROC and VDC is 1/187 of prazosin and 1/37 of verapamil, respectively.CONCLUSION: The blocking effects of CPU 86017 on the L-type calcium channel of myocardium and vessel are moderate and non-selective. CPU 86017 is approximately 50 times more potent in inhibiting ROC than VDC.

  3. Physics-Based Compact Model for CIGS and CdTe Solar Cells: From Voltage-Dependent Carrier Collection to Light-Enhanced Reverse Breakdown: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Xingshu; Alam, Muhammad Ashraful; Raguse, John; Garris, Rebekah; Deline, Chris; Silverman, Timothy

    2015-10-15

    In this paper, we develop a physics-based compact model for copper indium gallium diselenide (CIGS) and cadmium telluride (CdTe) heterojunction solar cells that attributes the failure of superposition to voltage-dependent carrier collection in the absorber layer, and interprets light-enhanced reverse breakdown as a consequence of tunneling-assisted Poole-Frenkel conduction. The temperature dependence of the model is validated against both simulation and experimental data for the entire range of bias conditions. The model can be used to characterize device parameters, optimize new designs, and most importantly, predict performance and reliability of solar panels including the effects of self-heating and reverse breakdown due to partial-shading degradation.

  4. Voltage-Dependent Anion Channel 2 of Arabidopsis thaliana (AtVDAC2 Is Involved in ABA-Mediated Early Seedling Development

    Directory of Open Access Journals (Sweden)

    Xufeng Li

    2009-05-01

    Full Text Available The voltage-dependent anion channel (VDAC is the major transport protein in the outer membrane of mitochondria and plays crucial roles in energy metabolism, apoptosis, and metabolites transport. In plants, the expression of VDACs can be affected by different stresses, including drought, salinity and pathogen defense. In this study, we investigated the expression pattern of AtVDAC2 in A. thaliana and found ABA suppressed the accumulation of AtVDAC2 transcripts. Further, phenotype analysis of this VDAC deregulated-expression transgenic Arabidopsis plants indicated that AtVDAC2 anti-sense line showed an ABA-insensitivity phenotype during the early seedling development under ABA treatment. The results suggested that AtVDAC2 might be involved in ABA signaling in A. thaliana.

  5. 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

    KCNQ4-encoded voltage-dependent potassium (Kv7.4) channels are important regulators of vascular tone that are severely compromised in models of hypertension. However, there is no information as to the role of these channels in responses to endogenous vasodilators. We used a molecular knockdown...... strategy, as well as pharmacological tools, to examine the hypothesis that Kv7.4 channels contribute to ß-adrenoceptor-mediated vasodilation in the renal vasculature and underlie the vascular deficit in spontaneously hypertensive rats. Quantitative PCR and immunohistochemistry confirmed gene and protein...... spontaneously hypertensive rats, which was associated with ˜60% decrease in Kv7.4 abundance. This study provides the first evidence that Kv7 channels contribute to ß-adrenoceptor-mediated vasodilation in the renal vasculature and that abrogation of Kv7.4 channels is strongly implicated in the impaired ß...

  6. Potassium currents in type II vestibular hair cells isolated from the guinea-pig's crista ampullaris.

    Science.gov (United States)

    Griguer, C; Kros, C J; Sans, A; Lehouelleur, J

    1993-11-01

    Type II vestibular hair cells were isolated from cristae ampullares of guinea-pig and maintained in vitro for 2-3 h. Outward membrane currents were studied under whole-cell voltage-clamp conditions. Type II hair cells had resting potentials of about -45 mV. Depolarizing voltage steps from a holding potential of -80 or -90 mV induced time- and voltage-dependent outward currents which slowly decayed to a sustained level. Tail currents reversed at about -70 mV, indicating that the outward currents were mainly carried by potassium ions. The currents had an activation threshold around -50 mV. The transient component was completely removed by a depolarizing pre-pulse positive to -10 mV. While bath application of 4-aminopyridine (5 mM) reduced both components, extracellular tetraethylammonium (10 mM) or zero calcium preferentially diminished the sustained current. We conclude that at least two potassium conductances are present, a delayed rectifier with a relatively fast inactivation and a calcium-dependent potassium current. Depolarizing current injections induced an electrical resonance in the voltage responses, with a frequency of 25-100 Hz, larger currents causing higher frequencies.

  7. Quantum entanglement in the voltage dependent sodium channel can reproduce the salient features of neuronal action potential initiation

    CERN Document Server

    Summhammer, Johann

    2007-01-01

    We investigate the effects of a quantum entanglement regime within an ion conducting molecule (ion channel) of the neuronal plasma membrane on the onset dynamics of propagating nerve pulses (action potentials). In particular, we model the onset parameters of the sodium current in the Hodgkin Huxley equation as three similar but independent probabilistic mechanisms which become quantum entangled. The underlying physics is general and can involve entanglement between various degrees of freedom underlaying ion transition states or 'gating states' during conduction, e.g. Na$^+$ ions in different channel locations, or different 'affinity' states of ions with atoms lining the sub-regions of the channel protein ('filter-states'). We find that the 'quantum corrected' Hodgkin Huxley equation incorporating entangled systems states can reproduce action potential pulses with the critical onset dynamics observed recently in neocortical neurons in vivo by Naundorf et al. [Nature {\\bf 440}, 1060 (20 April 2006)]. Interestin...

  8. Adenine nucleotides and intracellular Ca2+ regulate a voltage-dependent and glucose-sensitive potassium channel in neurosecretory cells.

    Science.gov (United States)

    Onetti, C G; Lara, J; García, E

    1996-05-01

    Effects of membrane potential, intracellular Ca2+ and adenine nucleotides on glucose-sensitive channels from X organ (XO) neurons of the crayfish were studied in excised inside-out patches. Glucose- sensitive channels were selective to K+ ions; the unitary conductance was 112 pS in symmetrical K+, and the K+ permeability (PK) was 1.3 x 10(-13) cm x s(-1). An inward rectification was observed when intracellular K+ was reduced. Using a quasi-physiological K+ gradient, a non-linear K+ current/voltage relationship was found showing an outward rectification and a slope conductance of 51 pS. The open-state probability (Po) increased with membrane depolarization as a result of an enhancement of the mean open time and a shortening of the longer period of closures. In quasi-physio- logical K+ concentrations, the channel was activated from a threshold of about -60 mV, and the activation midpoint was -2 mV. Po decreased noticeably at 50 microM internal adenosine 5'-triphosphate (ATP), and single-channel activity was totally abolished at 1 mM ATP. Hill analysis shows that this inhibition was the result of simultaneous binding of two ATP molecules to the channel, and the half-blocking concentration of ATP was 174 microM. Internal application of 5'-adenylylimidodiphosphate (AMP-PNP) as well as glibenclamide also decreased Po. By contrast, the application of internal ADP (0.1 to 2 mM) activated this channel. An optimal range of internal free Ca2+ ions (0.1 to 10 microM) was required for the activation of this channel. The glucose--sensitive K+ channel of XO neurons could be considered as a subtype of ATP-sensitive K+ channel, contributing substantially to macroscopic outward current.

  9. Involvement of presynaptic voltage-dependent Kv3 channel in endothelin-1-induced inhibition of noradrenaline release from rat gastric sympathetic nerves.

    Science.gov (United States)

    Nakamura, Kumiko; Shimizu, Takahiro; Tanaka, Kenjiro; Taniuchi, Keisuke; Yokotani, Kunihiko

    2012-11-05

    We previously reported that two types of K(+) channels, the BK type Ca(2+)-activated K(+) channel coupled with phospholipase C (PLC) and the voltage-dependent K(+) channel (Kv channel), are, respectively, involved in the prostanoid TP receptor- and muscarinic M(2) receptor-mediated inhibition of noradrenaline (NA) release from rat gastric sympathetic nerves. In the present study, therefore, we examined whether these K(+) channels are involved in endothelin-1-induced inhibition of NA release, using an isolated, vascularly perfused rat stomach. The gastric sympathetic postganglionic nerves around the left gastric artery were electrically stimulated twice at 2.5 Hz for 1 min, and endothelin-1 was added during the second stimulation. Endothelin-1 (1, 2 and 10 nM) dose-dependently inhibited gastric NA release. Endothelin-1 (2 nM)-induced inhibition of NA release was neither attenuated by PLC inhibitors [U-73122 (3 μM) and ET-18-OCH(3) (3 μM)] nor by Ca(2+)-activated K(+) channel blockers [charybdotoxin (0.1 μM) (a blocker of BK type K(+) channel) and apamin (0.3 μM) (a blocker of SK type K(+) channel)]. The endothelin-1-induced inhibitory response was also not attenuated by α-dendrotoxin (0.1 μM) (a selective inhibitor of Kv1 channel), but abolished by 4-aminopyridine (20 μM) (a selectively inhibitory dose for Kv3 channel). These results suggest the involvement of a voltage-dependent Kv3 channel in the endothelin-1-induced inhibition of NA release from the gastric sympathetic nerves in rats.

  10. Heparin/heparan sulfates bind to and modulate neuronal L-type (Cav1.2) voltage-dependent Ca(2+) channels.

    Science.gov (United States)

    Garau, Gianpiero; Magotti, Paola; Heine, Martin; Korotchenko, Svetlana; Lievens, Patricia Marie-Jeanne; Berezin, Vladimir; Dityatev, Alexander

    2015-12-01

    Our previous studies revealed that L-type voltage-dependent Ca(2+) channels (Cav1.2 L-VDCCs) are modulated by the neural extracellular matrix backbone, polyanionic glycan hyaluronic acid. Here we used isothermal titration calorimetry and screened a set of peptides derived from the extracellular domains of Cav1.2α1 to identify putative binding sites between the channel and hyaluronic acid or another class of polyanionic glycans, such as heparin/heparan sulfates. None of the tested peptides showed detectable interaction with hyaluronic acid, but two peptides derived from the first pore-forming domain of Cav1.2α1 subunit bound to heparin. At 25 °C the binding of the peptide P7 (MGKMHKTCYN) was at ~50 μM, and that of the peptide P8 (GHGRQCQNGTVCKPGWDGPKHG) was at ~21 μM. The Cav1.2α1 first pore forming segment that contained both peptides maintained a high affinity for heparin (~23 μM), integrating their enthalpic and entropic binding contributions. Interaction between heparin and recombinant as well as native full-length neuronal Cav1.2α1 channels was confirmed using the heparin-agarose pull down assay. Whole cell patch clamp recordings in HEK293 cells transfected with neuronal Cav1.2 channels revealed that enzymatic digestion of highly sulfated heparan sulfates with heparinase 1 affects neither voltage-dependence of channel activation nor the level of steady state inactivation, but did speed up channel inactivation. Treatment of hippocampal cultures with heparinase 1 reduced the firing rate and led to appearance of long-lasting bursts in the same manner as treatment with the inhibitor of L-VDCC diltiazem. Thus, heparan sulfate proteoglycans may bind to and regulate L-VDCC inactivation and network activity.

  11. Cyanocobalamin, vitamin B12, depresses glutamate release through inhibition of voltage-dependent Ca2+ influx in rat cerebrocortical nerve terminals (synaptosomes).

    Science.gov (United States)

    Hung, Kun-Long; Wang, Chia-Chuan; Huang, Chia-Yu; Wang, Su-Jane

    2009-01-14

    The effect of cyanocobalamin, vitamin B12, on glutamate release in isolated nerve terminals (synaptosomes) prepared from rat prefrontal cortex was examined. Cyanocobalamin inhibited the release of glutamate evoked by 4-aminopyridine in a concentration-dependent manner. The inhibitory action of cyanocobalamin was blocked by the vesicular transporter inhibitor bafilomycin A1, not by the glutamate transporter inhibitor L-transpyrrolidine-2,4-dicarboxylic acid or the nontransportable glutamate inhibitor DL-threo-beta-benzyloxyaspartate, indicating that this release inhibition results from a reduction of vesicular exocytosis and not from an inhibition of Ca(2+)-independent efflux via glutamate transporter. Examination of the effect of cyanocobalamin on cytosolic free Ca(2+) concentration revealed that the inhibition of glutamate release could be attributed to a reduction in voltage-dependent Ca(2+) influx. Consistent with this, the N- and P/Q-type Ca(2+) channel blocker omega-conotoxin MVIIC, largely attenuated the inhibitory effect of cyanocobalamin on 4-aminopyridine-evoked glutamate release, but the Ca(2+) release inhibitor dantrolene had no effect. Cyanocobalamin did not alter the resting synaptosomal membrane potential or 4-aminopyridine-mediated depolarization; thus, the inhibition of 4-aminopyridine-evoked Ca(2+) influx and glutamate release produced by cyanocobalamin was not due to its decreasing synaptosomal excitability. In addition, cyanocobalamin-mediated inhibition of 4-aminopyridine-evoked Ca(2+) influx and glutamate release was significantly attenuated by protein kinase C inhibitors GF109203X and Ro318220. Furthermore, 4-aminopyridine-induced phosphorylation of protein kinase C was significantly reduced by cyanocobalamin. These results suggest that cyanocobalamin effects a decrease in protein kinase C activation, which subsequently reduces the Ca(2+) entry through voltage-dependent N- and P/Q-type Ca(2+) channels to cause a decrease in evoked glutamate

  12. Roscovitine, a cyclin-dependent kinase inhibitor, affects several gating mechanisms to inhibit cardiac L-type (Ca(V)1.2) calcium channels.

    Science.gov (United States)

    Yarotskyy, V; Elmslie, K S

    2007-10-01

    L-type calcium channels (Ca((V))1.2) play an important role in cardiac contraction. Roscovitine, a cyclin-dependent kinase inhibitor and promising anticancer drug, has been shown to affect Ca((V))1.2 by inhibiting current amplitude and slowing activation. This research investigates the mechanism by which roscovitine inhibits Ca((V))1.2 channels. Ca((V))1.2 channels were transfected into HEK 293 cells, using the calcium phosphate precipitation method, and currents were measured using the whole-cell patch clamp technique. Roscovitine slows activation at all voltages, which precludes one previously proposed mechanism. In addition, roscovitine enhances voltage-dependent, but not calcium-dependent inactivation. This enhancement resulted from both an acceleration of inactivation and a slowing of the recovery from inactivation. Internally applied roscovitine failed to affect Ca((V))1.2 currents, which supports a kinase-independent mechanism and extracellular binding site. Unlike the dihydropyridines, closed state inactivation was not affected by roscovitine. Inactivation was enhanced in a dose-dependent manner with an IC(50)=29.5+/-12 microM, which is close to that for slow activation and inhibition. We conclude that roscovitine binds to an extracellular site on Ca((V))1.2 channels to inhibit current by both slowing activation and enhancing inactivation. Purine-based drugs could become a new option for treatment of diseases that benefit from L-channel inhibition such as cardiac arrhythmias and hypertension.

  13. Series resistance mapping of Cu(In,Ga)Se{sub 2} solar cells by voltage dependent electroluminescence

    Energy Technology Data Exchange (ETDEWEB)

    Daume, Felix; Puttnins, Stefan [Solarion AG, Ostende 5, 04288 Leipzig (Germany); Institut fuer Experimentelle Physik II, Universitaet Leipzig, Linnestr. 5, 04103 Leipzig (Germany); Scheit, Christian; Rahm, Andreas [Solarion AG, Ostende 5, 04288 Leipzig (Germany); Grundmann, Marius [Institut fuer Experimentelle Physik II, Universitaet Leipzig, Linnestr. 5, 04103 Leipzig (Germany)

    2011-07-01

    Cu(In,Ga)Se{sub 2} (CIGSe) thin film solar cells deposited on flexible polyimide foil promising innovative applications and a fabrication in continuous roll-to-roll processes currently reach efficiencies up to 17.6 %. The optimization of the solar cell efficiency requires the reduction of inherent losses in the cell. In order to achieve this goal preferably spatially resolved access to parameters characterizing ohmic losses like series and shunt resistances are indispensable. We apply an interpretation method for electroluminescence (EL) images taken at different voltages which is known for solar cells made of crystalline silicon from literature to solar cells made of polycrystalline CIGSe. The theory of this method to obtain a mapping of the series resistance and the EL imaging process as well as the data interpretation ils reviewed and demonstrated on an example. Furthermore, the benefit of this method for the characterization of solar cells under accelerated aging conditions (damp heat) which is important for the estimation of the long-term stability is shown.

  14. Intramitochondrial accumulation of cationic Atto520-biotin proceeds via voltage-dependent slow permeation through lipid membrane.

    Science.gov (United States)

    Antonenko, Yuri N; Nechaeva, Natalya L; Baksheeva, Victoria E; Rokitskaya, Tatyana I; Plotnikov, Egor Y; Kotova, Elena A; Zorov, Dmitry B

    2015-06-01

    Conjugation to penetrating cations is a general approach for intramitochondrial delivery of physiologically active compounds, supported by a high membrane potential of mitochondria having negative sign on the matrix side. By using fluorescence correlation spectroscopy, we found here that Atto520-biotin, a conjugate of a fluorescent cationic rhodamine-based dye with the membrane-impermeable vitamin biotin, accumulated in energized mitochondria in contrast to biotin-rhodamine 110. The energy-dependent uptake of Atto520-biotin by mitochondria, being slower than that of the conventional mitochondrial dye tetramethyl-rhodamine ethyl ester, was enhanced by the hydrophobic anion tetraphenylborate (TPB). Atto520-biotin also exhibited accumulation in liposomes driven by membrane potential resulting from potassium ion gradient in the presence valinomycin. The induction of electrical current across planar bilayer lipid membrane by Atto520-biotin proved the ability of the compound to permeate through lipid membrane in a cationic form. Atto520-biotin stained mitochondria in a culture of L929 cells, and the staining was enhanced in the presence of TPB. Therefore, the fluorescent Atto520 moiety can serve as a vehicle for intramitochondrial delivery of hydrophilic drugs. Of importance for biotin-streptavidin technology, binding of Atto520-biotin to streptavidin was found to cause quenching of its fluorescence similar to the case of fluorescein-4-biotin.

  15. Actions of a hydrogen sulfide donor (NaHS) on transient sodium, persistent sodium, and voltage-gated calcium currents in neurons of the subfornical organ.

    Science.gov (United States)

    Kuksis, Markus; Ferguson, Alastair V

    2015-09-01

    Hydrogen sulfide (H2S) is an endogenously found gasotransmitter that has been implicated in a variety of beneficial physiological functions. This study was performed to investigate the cellular mechanisms underlying actions of H2S previously observed in subfornical organ (SFO), where H2S acts to regulate blood pressure through a depolarization of the membrane and an overall increase in the excitability of SFO neurons. We used whole cell patch-clamp electrophysiology in the voltage-clamp configuration to analyze the effect of 1 mM NaHS, an H2S donor, on voltage-gated potassium, sodium, and calcium currents. We observed no effect of NaHS on potassium currents; however, both voltage-gated sodium currents (persistent and transient) and the N-type calcium current had a depolarized activation curve and an enhanced peak-induced current in response to a series of voltage-step and ramp protocols run in the control and NaHS conditions. These effects were not responsible for the previously observed depolarization of the membrane potential, as depolarizing effects of H2S were still observed following block of these conductances with tetrodotoxin (5 μM) and ω-conotoxin-GVIA (100 nM). Our studies are the first to investigate the effect of H2S on a variety of voltage-gated conductances in a single brain area, and although they do not explain mechanisms underlying the depolarizing actions of H2S on SFO neurons, they provide evidence of potential mechanisms through which this gasotransmitter influences the excitability of neurons in this important brain area as a consequence of the modulation of multiple ion channels.

  16. A major role for calcium-dependent potassium current in action potential repolarization in adrenal chromaffin cells.

    Science.gov (United States)

    Pancrazio, J J; Johnson, P A; Lynch, C

    1994-12-30

    To determine the extent which Ca dependent K current (IKCa) contributes during an action potential (AP), bovine chromaffin cells were voltage-clamped using a pre-recorded AP as the command voltage waveform. Based on (1) differential sensitivity of IKCa and Ca-independent K current (IK) to tetraethylammonium; (2) measurements of AP currents under conditions where Ca activation of IKCa had been abolished; and (3) blockade by charybdotoxin, IKCa comprised 70-90% of the outward K current during AP repolarization. In addition, observations are made concerning the form of AP-evoked Ca current.

  17. Gentamicin Blocks the ACh-Induced BK Current in Guinea Pig Type II Vestibular Hair Cells by Competing with Ca2+ at the l-Type Calcium Channel

    Directory of Open Access Journals (Sweden)

    Hong Yu

    2014-04-01

    Full Text Available Type II vestibular hair cells (VHCs II contain big-conductance Ca2+-dependent K+ channels (BK and L-type calcium channels. Our previous studies in guinea pig VHCs II indicated that acetylcholine (ACh evoked the BK current by triggering the influx of Ca2+ ions through l-type Ca2+ channels, which was mediated by M2 muscarinic ACh receptor (mAChRs. Aminoglycoside antibiotics, such as gentamicin (GM, are known to have vestibulotoxicity, including damaging effects on the efferent nerve endings on VHCs II. This study used the whole-cell patch clamp technique to determine whether GM affects the vestibular efferent system at postsynaptic M2-mAChRs or the membrane ion channels. We found that GM could block the ACh-induced BK current and that inhibition was reversible, voltage-independent, and dose-dependent with an IC50 value of 36.3 ± 7.8 µM. Increasing the ACh concentration had little influence on GM blocking effect, but increasing the extracellular Ca2+ concentration ([Ca2+]o could antagonize it. Moreover, 50 µM GM potently blocked Ca2+ currents activated by (--Bay-K8644, but did not block BK currents induced by NS1619. These observations indicate that GM most likely blocks the M2 mAChR-mediated response by competing with Ca2+ at the l-type calcium channel. These results provide insights into the vestibulotoxicity of aminoglycoside antibiotics on mammalian VHCs II.

  18. Role of calcium-activated potassium currents in CNP-induced relaxation of gastric antral circular smooth muscle in guinea pigs

    Institute of Scientific and Technical Information of China (English)

    Hui-Shu Guo; Zheng-Xu Cai; Hai-Feng Zheng; Xiang-Lan Li; Yi-Feng Cui; Zuo-Yu Wang; Wen-Xie Xu; Sang-Jin Lee; Young-Chul Kim

    2003-01-01

    AIM: To investigate ion channel mechanism in CNP-induced relaxation of gastric circular smooth muscle in guinea pigs.METHODS: Spontaneous contraction of gastric smooth muscle was recorded by a four -channel physiograph. The whole cell patch-clamp technique was used to record calciumactivated potassium currents and membrane potential in the gastric myocytes isolated by collagenase.RESULTS: C-type natriuretic peptide (CNP) markedly inhibited the spontaneous contraction in a dose-dependent manner in gastric circular smooth muscle in guinea pigs.Ly83583, an inhibitor of guanylate cyclase, weakened CNPinduced inhibition on spontaneous contraction but Zaparinast, an inhibitor of cGMP sensitive phosphoesterase,potentiated CNP-induced inhibition in gastric circular smooth muscles. The inhibitory effects of CNP on spontaneous contraction were blocked by tetrathylammonium (TEA), a nonselective potassium channel blocker. CNP hyperpolarized membrane potential from -60.0 mV±2.0 mV to -68.3 meV±3.0 mV in a single gastric myocyte. CNP increased calcium-activated potassium currents (Ik(ca)) in a dose-dependent manner in gastric circular myocytes. CNP also increased the spontaneously transient outward currents (STOCs). Ly83583 partly blocked CNP-induced increase of calcium-activated potassium currents, but Zaparinast potented the effect.CONCLUSION: CNP inhibits spontaneous contraction, and potassium channel may be involved in the process in gastric circular smooth musde of guinea pigs. CNP-induced increase of Ik(ca) is mediated by a cGMP dependent pathway.

  19. CONSUMO DE CALCIO: EVOLUCIÓN Y SITUACIÓN ACTUAL The evolution and current situation of calcium consumption

    Directory of Open Access Journals (Sweden)

    María del Pilar Barrera P

    2012-03-01

    Full Text Available El calcio es un mineral esencial en la nutrición humana, contribuye a la formación de masa ósea, interviene en funciones metabólicas y en la contracción muscular. Debido a la importancia de este nutriente en la salud pública, se han realizado estudios en varios países, entre ellos Colombia, tendientes a determinar su consumo en diferentes grupos poblacionales, encontrando en la mayoría de ellos una ingesta insuficiente, que cubre menos del 50% de la recomendación, independientemente del desarrollo económico. Las Encuestas de la Situación Nutricional en Colombia (2005, 2010 reportaron déficit en el consumo de lácteos, especialmente en población de bajo estrato socioeconómico y en las regiones Pacífica, Amazonía y Orinoquía. Recientemente el Institute of Medicine (IOM estableció la ingesta diaria recomendada de calcio para cubrir las necesidades fisiológicas de la población. Esta medida es necesaria teniendo en cuenta que la evidencia científica señala una fuerte asociación entre baja ingesta y disminución de la densidad mineral ósea con incremento de la osteoporosis y de las fracturas asociadas. El IOM, también estableció los niveles máximos tolerables (UL para evitar problemas asociados al exceso con el uso de suplementos. En todos los países, las guías alimentarias tienen como objetivo promover una alimentación saludable, por lo tanto la divulgación de este instrumento es responsabilidad de todos los profesionales de la salud, especialmente de los nutricionistas. Se requiere fortalecer la investigación y desarrollar estrategias que involucren diferentes sectores para mejorar la ingesta de calcio y la salud ósea de la población.Calcium is an essential mineral in human nutrition; it contributes towards bone formation, intervenes in metabolic functions and muscular contraction. Studies have been carried out in several countries, including Colombia, due to this nutrient's importance in public health; they have

  20. Calcium handling by vascular myocytes in hypertension

    Directory of Open Access Journals (Sweden)

    R.C.A. Tostes

    1997-03-01

    Full Text Available Calcium ions (Ca2+ trigger the contraction of vascular myocytes and the level of free intracellular Ca2+ within the myocyte is precisely regulated by sequestration and extrusion mechanisms. Extensive evidence indicates that a defect in the regulation of intracellular Ca2+ plays a role in the augmented vascular reactivity characteristic of clinical and experimental hypertension. For example, arteries from spontaneously hypertensive rats (SHR have an increased contractile sensitivity to extracellular Ca2+ and intracellular Ca2+ levels are elevated in aortic smooth muscle cells of SHR. We hypothesize that these changes are due to an increase in membrane Ca2+ channel density and possibly function in vascular myocytes from hypertensive animals. Several observations using various experimental approaches support this hypothesis: 1 the contractile activity in response to depolarizing stimuli is increased in arteries from hypertensive animals demonstrating increased voltage-dependent Ca2+ channel activity in hypertension; 2 Ca2+ channel agonists such as Bay K 8644 produce contractions in isolated arterial segments from hypertensive rats and minimal contraction in those from normotensive rats; 3 intracellular Ca2+ concentration is abnormally increased in vascular myocytes from hypertensive animals following treatment with Ca2+ channel agonists and depolarizing interventions, and 4 using the voltage-clamp technique, the inward Ca2+ current in arterial myocytes from hypertensive rats is nearly twice as large as that from myocytes of normotensive rats. We suggest that an alteration in Ca2+ channel function and/or an increase in Ca2+ channel density, resulting from increased channel synthesis or reduced turnover, underlies the increased vascular reactivity characteristic of hypertension

  1. Current perspectives of bio-ceramic technology in endodontics: calcium enriched mixture cement - review of its composition, properties and applications

    Science.gov (United States)

    Nawal, Ruchika Roongta; Talwar, Sangeeta; Verma, Mahesh

    2015-01-01

    Advancements in bio-ceramic technology has revolutionised endodontic material science by enhancing the treatment outcome for patients. This class of dental materials conciliates excellent biocompatibility with high osseoconductivity that render them ideal for endodontic care. Few recently introduced bio-ceramic materials have shown considerable clinical success over their early generations in terms of good handling characteristics. Calcium enriched mixture (CEM) cement, Endosequence sealer, and root repair materials, Biodentine and BioAggregate are the new classes of bio-ceramic materials. The aim of this literature review is to present investigations regarding properties and applications of CEM cement in endodontics. A review of the existing literature was performed by using electronic and hand searching methods for CEM cement from January 2006 to December 2013. CEM cement has a different chemical composition from that of mineral trioxide aggregate (MTA) but has similar clinical applications. It combines the biocompatibility of MTA with more efficient characteristics, such as significantly shorter setting time, good handling characteristics, no staining of tooth and effective seal against bacterial leakage. PMID:25671207

  2. Current perspectives of bio-ceramic technology in endodontics: calcium enriched mixture cement - review of its composition, properties and applications

    Directory of Open Access Journals (Sweden)

    Shivani Utneja

    2015-02-01

    Full Text Available Advancements in bio-ceramic technology has revolutionised endodontic material science by enhancing the treatment outcome for patients. This class of dental materials conciliates excellent biocompatibility with high osseoconductivity that render them ideal for endodontic care. Few recently introduced bio-ceramic materials have shown considerable clinical success over their early generations in terms of good handling characteristics. Calcium enriched mixture (CEM cement, Endosequence sealer, and root repair materials, Biodentine and BioAggregate are the new classes of bio-ceramic materials. The aim of this literature review is to present investigations regarding properties and applications of CEM cement in endodontics. A review of the existing literature was performed by using electronic and hand searching methods for CEM cement from January 2006 to December 2013. CEM cement has a different chemical composition from that of mineral trioxide aggregate (MTA but has similar clinical applications. It combines the biocompatibility of MTA with more efficient characteristics, such as significantly shorter setting time, good handling characteristics, no staining of tooth and effective seal against bacterial leakage.

  3. Current perspectives of bio-ceramic technology in endodontics: calcium enriched mixture cement - review of its composition, properties and applications.

    Science.gov (United States)

    Utneja, Shivani; Nawal, Ruchika Roongta; Talwar, Sangeeta; Verma, Mahesh

    2015-02-01

    Advancements in bio-ceramic technology has revolutionised endodontic material science by enhancing the treatment outcome for patients. This class of dental materials conciliates excellent biocompatibility with high osseoconductivity that render them ideal for endodontic care. Few recently introduced bio-ceramic materials have shown considerable clinical success over their early generations in terms of good handling characteristics. Calcium enriched mixture (CEM) cement, Endosequence sealer, and root repair materials, Biodentine and BioAggregate are the new classes of bio-ceramic materials. The aim of this literature review is to present investigations regarding properties and applications of CEM cement in endodontics. A review of the existing literature was performed by using electronic and hand searching methods for CEM cement from January 2006 to December 2013. CEM cement has a different chemical composition from that of mineral trioxide aggregate (MTA) but has similar clinical applications. It combines the biocompatibility of MTA with more efficient characteristics, such as significantly shorter setting time, good handling characteristics, no staining of tooth and effective seal against bacterial leakage.

  4. Dissociation of the store-operated calcium current ICRAC and the Mg-nucleotide-regulated metal ion current MagNuM

    Science.gov (United States)

    Hermosura, Meredith C; Monteilh-Zoller, Mahealani K; Scharenberg, Andrew M; Penner, Reinhold; Fleig, Andrea

    2002-01-01

    Rat basophilic leukaemia cells (RBL-2H3-M1) were used to study the characteristics of the store-operated Ca2+ release-activated Ca2+ current (ICRAC) and the magnesium-nucleotide-regulated metal cation current (MagNuM) (which is conducted by the LTRPC7 channel). Pipette solutions containing 10 mm BAPTA and no added ATP induced both currents in the same cell, but the time to half-maximal activation for MagNuM was about two to three times slower than that of ICRAC. Differential suppression of ICRAC was achieved by buffering free [Ca2+]i to 90 nm and selective inhibition of MagNuM was accomplished by intracellular solutions containing 6 mm Mg.ATP, 1.2 mm free [Mg2+]i or 100 μm GTP-γ-S, allowing investigations on these currents in relative isolation. Removal of extracellular Ca2+ and Mg2+ caused both currents to be carried significantly by monovalent ions. In the absence or presence of free [Mg2+]i, ICRAC carried by monovalent ions inactivated more rapidly and more completely than MagNuM carried by monovalent ions. Since several studies have used divalent-free solutions on either side of the membrane to study selectivity and single-channel behaviour of ICRAC, these experimental conditions would have favoured the contribution of MagNuM to monovalent conductance and call for caution in interpreting results where both ICRAC and MagNuM are activated. PMID:11882677

  5. Calcium binding protein-mediated regulation of voltage-gated calcium channels linked to human diseases

    Institute of Scientific and Technical Information of China (English)

    Nasrin NFJATBAKHSH; Zhong-ping FENG

    2011-01-01

    Calcium ion entry through voltage-gated calcium channels is essential for cellular signalling in a wide variety of cells and multiple physiological processes. Perturbations of voltage-gated calcium channel function can lead to pathophysiological consequences. Calcium binding proteins serve as calcium sensors and regulate the calcium channel properties via feedback mechanisms. This review highlights the current evidences of calcium binding protein-mediated channel regulation in human diseases.

  6. Forgetting of long-term memory requires activation of NMDA receptors, L-type voltage-dependent Ca2+ channels, and calcineurin

    Science.gov (United States)

    Sachser, Ricardo Marcelo; Santana, Fabiana; Crestani, Ana Paula; Lunardi, Paula; Pedraza, Lizeth Katherine; Quillfeldt, Jorge Alberto; Hardt, Oliver; de Oliveira Alvares, Lucas

    2016-01-01

    In the past decades, the cellular and molecular mechanisms underlying memory consolidation, reconsolidation, and extinction have been well characterized. However, the neurobiological underpinnings of forgetting processes remain to be elucidated. Here we used behavioral, pharmacological and electrophysiological approaches to explore mechanisms controlling forgetting. We found that post-acquisition chronic inhibition of the N-methyl-D-aspartate receptor (NMDAR), L-type voltage-dependent Ca2+ channel (LVDCC), and protein phosphatase calcineurin (CaN), maintains long-term object location memory that otherwise would have been forgotten. We further show that NMDAR activation is necessary to induce forgetting of object recognition memory. Studying the role of NMDAR activation in the decay of the early phase of long-term potentiation (E-LTP) in the hippocampus, we found that ifenprodil infused 30 min after LTP induction in vivo blocks the decay of CA1-evoked postsynaptic plasticity, suggesting that GluN2B-containing NMDARs activation are critical to promote LTP decay. Taken together, these findings indicate that a well-regulated forgetting process, initiated by Ca2+ influx through LVDCCs and GluN2B-NMDARs followed by CaN activation, controls the maintenance of hippocampal LTP and long-term memories over time. PMID:26947131

  7. Evidence for functional interaction of plasma membrane electron transport, voltage-dependent anion channel and volume-regulated anion channel in frog aorta

    Indian Academy of Sciences (India)

    Rashmi P Rao; J Prakasa Rao

    2010-12-01

    Frog aortic tissue exhibits plasma membrane electron transport (PMET) owing to its ability to reduce ferricyanide even in the presence of mitochondrial poisons, such as cyanide and azide. Exposure to hypotonic solution (108 mOsmol/kg H2O) enhanced the reduction of ferricyanide in excised aortic tissue of frog. Increment in ferricyanide reductase activity was also brought about by the presence of homocysteine (100 M dissolved in isotonic frog Ringer solution), a redox active compound and a potent modulator of PMET. Two plasma-membrane-bound channels, the volume regulated anion channel (VRAC) and the voltage-dependent anion channel (VDAC), are involved in the response to hypotonic stress. The presence of VRAC and VDAC antagonists–tamoxifen, glibenclamide, fluoxetine and verapamil, and 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS), respectively–inhibited this enhanced activity brought about by either hypotonic stress or homocysteine. The blockers do not affect the ferricyanide reductase activity under isotonic conditions. Taken together, these findings indicate a functional interaction of the three plasma membrane proteins, namely, ferricyanide reductase (PMET), VDAC and VRAC.

  8. Correlation between Barrier Width, Barrier Height, and DC Bias Voltage Dependences on the Magnetoresistance Ratio in Ir-Mn Exchange Biased Single and Double Tunnel Junctions

    Science.gov (United States)

    Saito, Yoshiaki; Amano, Minoru; Nakajima, Kentaro; Takahashi, Shigeki; Sagoi, Masayuki; Inomata, Koichiro

    2000-10-01

    Dual spin-valve-type double tunnel junctions (DTJs) of Ir-Mn/CoFe/AlOx/Co90Fe10/AlOx/CoFe/Ir-Mn and spin-valve-type single tunnel junctions (STJs) of Ir-Mn/CoFe/AlOx/CoFe/Ni-Fe were fabricated using an ultrahigh vacuum sputtering system, conventional photolithography and ion-beam milling. The STJs could be fabricated with various barrier heights by changing the oxidization conditions during deposition and changing the annealing temperature after deposition, while the AlOx layer thickness remained unchanged. There was a correlation between barrier width, height estimated using Simmons’ expressions, and dc bias voltage dependence on the MR ratio. The VB dependence on the tunneling magnetoresistance (TMR) ratio was mainly related to the barrier width, and the decrease in the TMR ratio with increasing bias voltage is well explained, taking into account the spin-independent two-step tunneling via defect states in the barrier, as a main mechanism, at room temperature. Under optimized oxidization and annealing conditions, the maximum TMR ratio at a low bias voltage, and the dc bias voltage value at which the TMR ratio decreases in value by half (V1/2) were 42.4% and 952 mV in DTJs, and 49.0% and 425 mV in STJs, respectively.

  9. hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state.

    Science.gov (United States)

    Malak, Olfat A; Es-Salah-Lamoureux, Zeineb; Loussouarn, Gildas

    2017-12-01

    Delayed-rectifier potassium channels (hERG and KCNQ1) play a major role in cardiac repolarization. These channels are formed by a tetrameric pore (S5-S6) surrounded by four voltage sensor domains (S1-S4). Coupling between voltage sensor domains and the pore activation gate is critical for channel voltage-dependence. However, molecular mechanisms remain elusive. Herein, we demonstrate that covalently binding, through a disulfide bridge, a peptide mimicking the S4-S5 linker (S4-S5L) to the channel S6 C-terminus (S6T) completely inhibits hERG. This shows that channel S4-S5L is sufficient to stabilize the pore activation gate in its closed state. Conversely, covalently binding a peptide mimicking S6T to the channel S4-S5L prevents its inhibiting effect and renders the channel almost completely voltage-independent. This shows that the channel S4-S5L is necessary to stabilize the activation gate in its closed state. Altogether, our results provide chemical evidence that S4-S5L acts as a voltage-controlled ligand that binds S6T to lock the channel in a closed state, elucidating the coupling between voltage sensors and the gate in delayed rectifier potassium channels and potentially other voltage-gated channels.

  10. Forgetting of long-term memory requires activation of NMDA receptors, L-type voltage-dependent Ca2+ channels, and calcineurin.

    Science.gov (United States)

    Sachser, Ricardo Marcelo; Santana, Fabiana; Crestani, Ana Paula; Lunardi, Paula; Pedraza, Lizeth Katherine; Quillfeldt, Jorge Alberto; Hardt, Oliver; Alvares, Lucas de Oliveira

    2016-03-07

    In the past decades, the cellular and molecular mechanisms underlying memory consolidation, reconsolidation, and extinction have been well characterized. However, the neurobiological underpinnings of forgetting processes remain to be elucidated. Here we used behavioral, pharmacological and electrophysiological approaches to explore mechanisms controlling forgetting. We found that post-acquisition chronic inhibition of the N-methyl-D-aspartate receptor (NMDAR), L-type voltage-dependent Ca(2+) channel (LVDCC), and protein phosphatase calcineurin (CaN), maintains long-term object location memory that otherwise would have been forgotten. We further show that NMDAR activation is necessary to induce forgetting of object recognition memory. Studying the role of NMDAR activation in the decay of the early phase of long-term potentiation (E-LTP) in the hippocampus, we found that ifenprodil infused 30 min after LTP induction in vivo blocks the decay of CA1-evoked postsynaptic plasticity, suggesting that GluN2B-containing NMDARs activation are critical to promote LTP decay. Taken together, these findings indicate that a well-regulated forgetting process, initiated by Ca(2+) influx through LVDCCs and GluN2B-NMDARs followed by CaN activation, controls the maintenance of hippocampal LTP and long-term memories over time.

  11. α-Synuclein Shows High Affinity Interaction with Voltage-dependent Anion Channel, Suggesting Mechanisms of Mitochondrial Regulation and Toxicity in Parkinson Disease.

    Science.gov (United States)

    Rostovtseva, Tatiana K; Gurnev, Philip A; Protchenko, Olga; Hoogerheide, David P; Yap, Thai Leong; Philpott, Caroline C; Lee, Jennifer C; Bezrukov, Sergey M

    2015-07-24

    Participation of the small, intrinsically disordered protein α-synuclein (α-syn) in Parkinson disease (PD) pathogenesis has been well documented. Although recent research demonstrates the involvement of α-syn in mitochondrial dysfunction in neurodegeneration and suggests direct interaction of α-syn with mitochondria, the molecular mechanism(s) of α-syn toxicity and its effect on neuronal mitochondria remain vague. Here we report that at nanomolar concentrations, α-syn reversibly blocks the voltage-dependent anion channel (VDAC), the major channel of the mitochondrial outer membrane that controls most of the metabolite fluxes in and out of the mitochondria. Detailed analysis of the blockage kinetics of VDAC reconstituted into planar lipid membranes suggests that α-syn is able to translocate through the channel and thus target complexes of the mitochondrial respiratory chain in the inner mitochondrial membrane. Supporting our in vitro experiments, a yeast model of PD shows that α-syn toxicity in yeast depends on VDAC. The functional interactions between VDAC and α-syn, revealed by the present study, point toward the long sought after physiological and pathophysiological roles for monomeric α-syn in PD and in other α-synucleinopathies.

  12. Progesterone treatment inhibits and dihydrotestosterone (DHT) treatment potentiates voltage-gated calcium currents in gonadotropin-releasing hormone (GnRH) neurons.

    Science.gov (United States)

    Sun, Jianli; Moenter, Suzanne M

    2010-11-01

    GnRH neurons are central regulators of fertility, and their activity is modulated by steroid feedback. In normal females, GnRH secretion is regulated by estradiol and progesterone (P). Excess androgens present in hyperandrogenemic fertility disorders may disrupt communication of negative feedback signals from P and/or independently stimulate GnRH release. Voltage-gated calcium channels (VGCCs) are important in regulating excitability and hormone release. Estradiol alters VGCCs in a time-of-day-dependent manner. To further elucidate ovarian steroid modulation of GnRH neuron VGCCs, we studied the effects of dihydrotestosterone (DHT) and P. Adult mice were ovariectomized (OVX) or OVX and treated with implants containing DHT (OVXD), estradiol (OVXE), estradiol and DHT (OVXED), estradiol and P (OVXEP), or estradiol, DHT, and P (OVXEDP). Macroscopic calcium current (I(Ca)) was recorded in the morning or afternoon 8-12 d after surgery using whole-cell voltage-clamp. I(Ca) was increased in afternoon vs. morning in GnRH neurons from OVXE mice but this increase was abolished in cells from OVXEP mice. I(Ca) in cells from OVXD mice was increased regardless of time of day; there was no additional effect in OVXED mice. P reduced N-type and DHT potentiated N- and R-type VGCCs; P blocked the DHT potentiation of N-type-mediated current. These data suggest P and DHT have opposing actions on VGCCs in GnRH neurons, but in the presence of both steroids, P dominates. VGCCs are targets of ovarian steroid feedback modulation of GnRH neuron activity and, more specifically, a potential mechanism whereby androgens could activate GnRH neuronal function.

  13. Calcium electroporation in three cell lines; a comparison of bleomycin and calcium, calcium compounds, and pulsing conditions

    DEFF Research Database (Denmark)

    Frandsen, Stine Krog; Gissel, Hanne; Hojman, Pernille

    2013-01-01

    BACKGROUND: Electroporation with calcium (calcium electroporation) can induce ATP depletion-associated cellular death. In the clinical setting, the cytotoxic drug bleomycin is currently used with electroporation (electrochemotherapy) for palliative treatment of tumors. Calcium electroporation...... offers several advantages over standard treatment options: calcium is inexpensive and may readily be applied without special precautions, as is the case with cytostatic drugs. Therefore, details on the use of calcium electroporation are essential for carrying out clinical trials comparing calcium...

  14. Pharmacological profile of the ATP-mediated increase in L-type calcium current amplitude and activation of a non-specific cationic current in rat ventricular cells.

    OpenAIRE

    Scamps, F.; Vassort, G.

    1994-01-01

    1. The pharmacological profile of the ATP-induced increase in ICa amplitude and of ATP activation of a non-specific cationic current, IATP, was investigated in rat ventricular cells. 2. The EC50 values for ICa increase and IATP activation were 0.36 microM and 0.76 microM respectively. Suramin (10 microM) and cibacron blue (1 microM) competitively antagonized both effects of ATP. 3. The rank order of efficacy and potency of ATP analogues in increasing ICa amplitude was 2-methylthio-ATP approxi...

  15. 4-氨基吡啶对豚鼠心室肌钙和钠电流的影响%EFFECTS OF 4-AMINOPYRIDINE ON CALCIUM CURRENTS AND SODIUM CURRENTS IN GUINEA PIG VENTRICULAR MYOCYTES

    Institute of Scientific and Technical Information of China (English)

    傅丽英; 李泱; 夏国瑾; 姚伟星; 江明性

    2001-01-01

    目的研究4-氨基吡啶(4-AP)对心肌细胞L型钙通道和钠通道的影响。方法用全细胞膜片钳技术考察4-AP对豚鼠心室肌细胞L型钙电流和钠电流的作用。结果 4-AP 0.1, 0.5, 1.0 mmol*L-1浓度依赖性地抑制L型钙电流(ICa,L)和钠电流(INa),抑制率分别为(11.6±1.7)%,(37.5±8.3)%和(54.5±6.9)%以及(22.1±14.3)%,(39.4±8.8)%和(62.3±6.8)%。0.5 mmol*L-1 4-AP使ICa,L和INa I-V曲线均上移。结论 4-AP可浓度依赖性地阻滞豚鼠心室肌细胞L型钙通道和钠通道。%AIM To investigate the effect of 4-aminopyridine (4-AP) on ion channels of myocytes. METHODS L-type calcium channel and sodium channel currents were recorded in guinea pig single ventricular myocyte using whole-cell patch-clamp techniques. RESULTS 4-AP, 0.1, 0.5 and 1.0 mmol*L-1 were shown to inhibit L-type calcium channel currents (ICa,L) and sodium channel currents (INa) concentration-dependently. The percentage of inhibition were (11.6±1.7)%, (37.5±8.3)% and (54.5±6.9)% (P<0.01) respectively for ICa,L, and (22.1±14.3)% (P<0.05), (39.4±8.8)% and (62.3±6.8)% (P<0.01) respectively for INa. 4-AP 0.5 mmol*L-1 shifted the I-V curves of ICa,L and INa upwardly. CONCLUSION 4-AP blocked L-type calcium channel and sodium channels in guinea-pig ventricular myocytes concentration-dependently.

  16. RGS12 interacts with the SNARE-binding region of the Cav2.2 calcium channel.

    Science.gov (United States)

    Richman, Ryan W; Strock, Jesse; Hains, Melinda D; Cabanilla, Nory Jun; Lau, King-Kei; Siderovski, David P; Diversé-Pierluissi, María

    2005-01-14

    Activation of GABAB receptors in chick dorsal root ganglion (DRG) neurons inhibits the Cav2.2 calcium channel in both a voltage-dependent and voltage-independent manner. The voltage-independent inhibition requires activation of a tyrosine kinase that phosphorylates the alpha1 subunit of the channel and thereby recruits RGS12, a member of the "regulator of G protein signaling" (RGS) proteins. Here we report that RGS12 binds to the SNARE-binding or "synprint" region (amino acids 726-985) in loop II-III of the calcium channel alpha1 subunit. A recombinant protein encompassing the N-terminal PTB domain of RGS12 binds to the synprint region in protein overlay and surface plasmon resonance binding assays; this interaction is dependent on tyrosine phosphorylation and yet is within a sequence that differs from the canonical NPXY motif targeted by other PTB domains. In electrophysiological experiments, microinjection of DRG neurons with synprint-derived peptides containing the tyrosine residue Tyr-804 altered the rate of desensitization of neurotransmitter-mediated inhibition of the Cav2.2 calcium channel, whereas peptides centered about a second tyrosine residue, Tyr-815, were without effect. RGS12 from a DRG neuron lysate was precipitated using synprint peptides containing phosphorylated Tyr-804. The high degree of conservation of Tyr-804 in the SNARE-binding region of Cav2.1 and Cav2.2 calcium channels suggests that this region, in addition to the binding of SNARE proteins, is also important for determining the time course of the modulation of calcium current via tyrosine phosphorylation.

  17. Calcium window currents, periodic forcing, and chaos: Understanding single neuron response with a discontinuous one-dimensional map

    Science.gov (United States)

    Laudanski, J.; Sumner, C.; Coombes, S.

    2010-07-01

    Thalamocortical (TC) neurones are known to express the low-voltage activated, inactivating Ca2+ current IT . The triggering of this current underlies the generation of low threshold Ca2+ potentials that may evoke single or bursts of action potentials. Moreover, this current can contribute to an intrinsic slow (dynamics for the gating variables in the model of IT . This model can be analyzed in closed form and is shown to support an unstable set of periodic orbits. Trajectories are repelled from these organizing centers until they reach the threshold for firing. By determining the condition for a grazing bifurcation (at the border between a spiking and nonspiking event) we show how knowledge of the unstable periodic orbits (existence and stability) can be combined with the grazing condition to determine an effective one-dimensional map that captures the essentials of the chaotic behavior. This map is discontinuous and has strong similarities with the universal limit mapping in grazing bifurcations derived in the context of impacting mechanical systems [A. B. Nordmark, Phys. Rev. E 55, 266 (1997)10.1103/PhysRevE.55.266].

  18. Characterization of the Ca2+-gated and voltage-dependent K+-channel Slo-1 of nematodes and its interaction with emodepside.

    Science.gov (United States)

    Kulke, Daniel; von Samson-Himmelstjerna, Georg; Miltsch, Sandra M; Wolstenholme, Adrian J; Jex, Aaron R; Gasser, Robin B; Ballesteros, Cristina; Geary, Timothy G; Keiser, Jennifer; Townson, Simon; Harder, Achim; Krücken, Jürgen

    2014-12-01

    The cyclooctadepsipeptide emodepside and its parent compound PF1022A are broad-spectrum nematicidal drugs which are able to eliminate nematodes resistant to other anthelmintics. The mode of action of cyclooctadepsipeptides is only partially understood, but involves the latrophilin Lat-1 receptor and the voltage- and calcium-activated potassium channel Slo-1. Genetic evidence suggests that emodepside exerts its anthelmintic activity predominantly through Slo-1. Indeed, slo-1 deficient Caenorhabditis elegans strains are completely emodepside resistant. However, direct effects of emodepside on Slo-1 have not been reported and these channels have only been characterized for C. elegans and related Strongylida. Molecular and bioinformatic analyses identified full-length Slo-1 cDNAs of Ascaris suum, Parascaris equorum, Toxocara canis, Dirofilaria immitis, Brugia malayi, Onchocerca gutturosa and Strongyloides ratti. Two paralogs were identified in the trichocephalids Trichuris muris, Trichuris suis and Trichinella spiralis. Several splice variants encoding truncated channels were identified in Trichuris spp. Slo-1 channels of trichocephalids form a monophyletic group, showing that duplication occurred after the divergence of Enoplea and Chromadorea. To explore the function of a representative protein, C. elegans Slo-1a was expressed in Xenopus laevis oocytes and studied in electrophysiological (voltage-clamp) experiments. Incubation of oocytes with 1-10 µM emodepside caused significantly increased currents over a wide range of step potentials in the absence of experimentally increased intracellular Ca2+, suggesting that emodepside directly opens C. elegans Slo-1a. Emodepside wash-out did not reverse the effect and the Slo-1 inhibitor verruculogen was only effective when applied before, but not after, emodepside. The identification of several splice variants and paralogs in some parasitic nematodes suggests that there are substantial differences in channel properties among

  19. Characterization of the Ca2+-gated and voltage-dependent K+-channel Slo-1 of nematodes and its interaction with emodepside.

    Directory of Open Access Journals (Sweden)

    Daniel Kulke

    2014-12-01

    Full Text Available The cyclooctadepsipeptide emodepside and its parent compound PF1022A are broad-spectrum nematicidal drugs which are able to eliminate nematodes resistant to other anthelmintics. The mode of action of cyclooctadepsipeptides is only partially understood, but involves the latrophilin Lat-1 receptor and the voltage- and calcium-activated potassium channel Slo-1. Genetic evidence suggests that emodepside exerts its anthelmintic activity predominantly through Slo-1. Indeed, slo-1 deficient Caenorhabditis elegans strains are completely emodepside resistant. However, direct effects of emodepside on Slo-1 have not been reported and these channels have only been characterized for C. elegans and related Strongylida. Molecular and bioinformatic analyses identified full-length Slo-1 cDNAs of Ascaris suum, Parascaris equorum, Toxocara canis, Dirofilaria immitis, Brugia malayi, Onchocerca gutturosa and Strongyloides ratti. Two paralogs were identified in the trichocephalids Trichuris muris, Trichuris suis and Trichinella spiralis. Several splice variants encoding truncated channels were identified in Trichuris spp. Slo-1 channels of trichocephalids form a monophyletic group, showing that duplication occurred after the divergence of Enoplea and Chromadorea. To explore the function of a representative protein, C. elegans Slo-1a was expressed in Xenopus laevis oocytes and studied in electrophysiological (voltage-clamp experiments. Incubation of oocytes with 1-10 µM emodepside caused significantly increased currents over a wide range of step potentials in the absence of experimentally increased intracellular Ca2+, suggesting that emodepside directly opens C. elegans Slo-1a. Emodepside wash-out did not reverse the effect and the Slo-1 inhibitor verruculogen was only effective when applied before, but not after, emodepside. The identification of several splice variants and paralogs in some parasitic nematodes suggests that there are substantial differences in

  20. Auxiliary Subunit GSG1L Acts to Suppress Calcium-Permeable AMPA Receptor Function

    Science.gov (United States)

    McGee, Thomas P.; Bats, Cécile

    2015-01-01

    AMPA-type glutamate receptors are ligand-gated cation channels responsible for a majority of the fast excitatory synaptic transmission in the brain. Their behavior and calcium permeability depends critically on their subunit composition and the identity of associated auxiliary proteins. Calcium-permeable AMPA receptors (CP-AMPARs) contribute to various forms of synaptic plasticity, and their dysfunction underlies a number of serious neurological conditions. For CP-AMPARs, the prototypical transmembrane AMPAR regulatory protein stargazin, which acts as an auxiliary subunit, enhances receptor function by increasing single-channel conductance, slowing channel gating, increasing calcium permeability, and relieving the voltage-dependent block by endogenous intracellular polyamines. We find that, in contrast, GSG1L, a transmembrane auxiliary protein identified recently as being part of the AMPAR proteome, acts to reduce the weighted mean single-channel conductance and calcium permeability of recombinant CP-AMPARs, while increasing polyamine-dependent rectification. To examine the effects of GSG1L on native AMPARs, we manipulated its expression in cerebellar and hippocampal neurons. Transfection of GSG1L into mouse cultured cerebellar stellate cells that lack this protein increased the inward rectification of mEPSCs. Conversely, shRNA-mediated knockdown of endogenous GSG1L in rat cultured hippocampal pyramidal neurons led to an increase in mEPSC amplitude and in the underlying weighted mean single-channel conductance, revealing that GSG1L acts to suppress current flow through native CP-AMPARs. Thus, our data suggest that GSG1L extends the functional repertoire of AMPAR auxiliary subunits, which can act not only to enhance but also diminish current flow through their associated AMPARs. SIGNIFICANCE STATEMENT Calcium-permeable AMPA receptors (CP-AMPARs) are an important group of receptors for the neurotransmitter glutamate. These receptors contribute to various forms of

  1. Frequency and voltage dependence dielectric properties, ac electrical conductivity and electric modulus profiles in Al/Co{sub 3}O{sub 4}-PVA/p-Si structures

    Energy Technology Data Exchange (ETDEWEB)

    Bilkan, Çiğdem, E-mail: cigdembilkan@gmail.com [Department of Physics, Faculty of Sciences, The University of Çankırı Karatekin, 18100 Çankırı (Turkey); Azizian-Kalandaragh, Yashar [Department of Physics, Faculty of Science, The University of Mohaghegh Ardabili, Ardabil (Iran, Islamic Republic of); Altındal, Şemsettin [Department of Physics, Faculty of Sciences, The University of Gazi, 06500 Ankara (Turkey); Shokrani-Havigh, Roya [Department of Physics, Faculty of Science, The University of Mohaghegh Ardabili, Ardabil (Iran, Islamic Republic of)

    2016-11-01

    In this research a simple microwave-assisted method have been used for preparation of cobalt oxide nanostructures. The as-prepared sample has been investigated by UV–vis spectroscopy, X-ray diffraction (XRD), scanning electron microscopy (SEM). On the other hand, frequency and voltage dependence of both the real and imaginary parts of dielectric constants (ε′, ε″) and electric modulus (M′ and M″), loss tangent (tanδ), and ac electrical conductivity (σ{sub ac}) values of Al/Co{sub 3}O{sub 4}-PVA/p-Si structures were obtained in the wide range of frequency and voltage using capacitance (C) and conductance (G/ω) data at room temperature. The values of ε′, ε″ and tanδ were found to decrease with increasing frequency almost for each applied bias voltage, but the changes in these parameters become more effective in the depletion region at low frequencies due to the charges at surface states and their relaxation time and polarization effect. While the value of σ is almost constant at low frequency, increases almost as exponentially at high frequency which are corresponding to σ{sub dc} and σ{sub ac}, respectively. The M′ and M″ have low values at low frequencies region and then an increase with frequency due to short-range mobility of charge carriers. While the value of M′ increase with increasing frequency, the value of M″ shows two peak and the peaks positions shifts to higher frequency with increasing applied voltage due to the decrease of the polarization and N{sub ss} effects with increasing frequency.

  2. Delta receptors are required for full inhibitory coupling of mu-receptors to voltage-dependent Ca(2+) channels in dorsal root ganglion neurons.

    Science.gov (United States)

    Walwyn, Wendy; John, Scott; Maga, Matthew; Evans, Christopher J; Hales, Tim G

    2009-07-01

    Recombinant micro and delta opioid receptors expressed in cell lines can form heterodimers with distinctive properties and trafficking. However, a role for opioid receptor heterodimerization in neurons has yet to be identified. The inhibitory coupling of opioid receptors to voltage-dependent Ca(2+) channels (VDCCs) is a relatively inefficient process and therefore provides a sensitive assay of altered opioid receptor function and expression. We examined micro-receptor coupling to VDCCs in dorsal root ganglion neurons of delta(+/+), delta(+/-), and delta(-/-) mice. Neurons deficient in delta receptors exhibited reduced inhibition of VDCCs by morphine and [D-Ala(2),Phe(4),Gly(5)-ol]-enkephalin (DAMGO). An absence of delta receptors caused reduced efficacy of DAMGO without affecting potency. An absence of delta receptors reduced neither the density of VDCCs nor their inhibition by either the GABA(B) receptor agonist baclofen or intracellular guanosine 5'-O-(3-thio)triphosphate. Flow cytometry revealed a reduction in micro-receptor surface expression in delta(-/-) neurons without altered DAMGO-induced internalization. There was no change in micro-receptor mRNA levels. D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH(2)-sensitive mu-receptor-coupling efficacy was fully restored to delta(+/+) levels in delta(-/-) neurons by expression of recombinant delta receptors. However, the dimerization-deficient delta-15 construct expressed in delta(-/-) neurons failed to fully restore the inhibitory coupling of micro-receptors compared with that seen in delta(+/+) neurons, suggesting that, although not essential for micro-receptor function, micro-delta receptor dimerization contributes to full micro-agonist efficacy. Because DAMGO exhibited a similar potency in delta(+/+) and delta(-/-) neurons and caused similar levels of internalization, the role for heterodimerization is probably at the level of receptor biosynthesis.

  3. Voltage-dependent anion channels (VDACs) promote mitophagy to protect neuron from death in an early brain injury following a subarachnoid hemorrhage in rats.

    Science.gov (United States)

    Li, Jian; Lu, Jianfei; Mi, Yongjie; Shi, Zhao; Chen, Chunhua; Riley, John; Zhou, Changman

    2014-07-21

    The term mitophagy is coined to describe the selective removal of mitochondria by autophagy but the process itself is still contentious, especially in the early period following subarachnoid hemorrhage (SAH). In the present study, we investigated the role of mitophagy following 48h after SAH injury in rats. Specifically evaluating whether mitophagy, through voltage dependant anion channels (VDACs) interacting with microtubule-associated protein 1 light chain 3, could orchestrate the induction of apoptotic and necrotic cell death in neurons, a VDAC1siRNA and an activitor Rapamycian (RAPA), were engaged. One hundred and twelve male Sprague-Dawley rats were randomly divided into 4 groups: Sham, SAH, SAH+VDAC1siRNA, and SAH+RAPA. Outcomes measured included mortality rate, brain edema, BBB disruption, and neurobehavioral testing. We also used western blotting techniques to analyze the expressions of key mitophagic/autophagic proteins and pro-apoptotic protein such as ROS, VDAC1, LC-3II and Caspase-3. Rapamycin treatment significantly improved the mortality rate, cerebral edema, and neurobehavioral deficits; apoptotic and necrotic cell death in neurons were reduced by Rapamycin following SAH injury. However, VDAC1siRNA worsened the brain injury following SAH. Immunohistochemical staining and western blot analysis demonstrated a decreased expression of VDAC1, LC3II, and an increase of ROS and Caspase-3 followed by VDAC1siRNA administration. In conclusion, mitophagy induced by VDAC1 following SAH injury may in fact play a significant role in neuroprotection, the mechanism which may be through the attenuation of the apoptosic and necrosic molecular pathways. This translates a preservation of functional integrity and an improvement in mortality.

  4. Voltage-dependent gating of hyperpolarization-activated, cyclic nucleotide-gated pacemaker channels: molecular coupling between the S4-S5 and C-linkers.

    Science.gov (United States)

    Decher, Niels; Chen, Jun; Sanguinetti, Michael C

    2004-04-02

    Hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels have a transmembrane topology that is highly similar to voltage-gated K(+) channels, yet HCN channels open in response to membrane hyperpolarization instead of depolarization. The structural basis for the "inverted" voltage dependence of HCN gating and how voltage sensing by the S1-S4 domains is coupled to the opening of the intracellular gate formed by the S6 domain are unknown. Coupling could arise from interaction between specific residues or entire transmembrane domains. We previously reported that the mutation of specific residues in the S4-S5 linker of HCN2 (i.e. Tyr-331 and Arg-339) prevented normal channel closure presumably by disruption of a crucial interaction with the activation gate. Here we hypothesized that the C-linker, a carboxyl terminus segment that connects S6 to the cyclic nucleotide binding domain, interacts with specific residues of the S4-S5 linker to mediate coupling. The recently solved structure of the C-linker of HCN2 indicates that an alpha-helix (the A'-helix) is located near the end of each S6 domain, the presumed location of the activation gate. Ala-scanning mutagenesis of the end of S6 and the A'-helix identified five residues that were important for normal gating as mutations disrupted channel closure. However, partial deletion of the C-linker indicated that the presence of only two of these residues was required for normal coupling. Further mutation analyses suggested that a specific electrostatic interaction between Arg-339 of the S4-S5 linker and Asp-443 of the C-linker stabilizes the closed state and thus participates in the coupling of voltage sensing and activation gating in HCN channels.

  5. The N-Terminal Peptides of the Three Human Isoforms of the Mitochondrial Voltage-Dependent Anion Channel Have Different Helical Propensities.

    Science.gov (United States)

    Guardiani, Carlo; Scorciapino, Mariano Andrea; Amodeo, Giuseppe Federico; Grdadolnik, Joze; Pappalardo, Giuseppe; De Pinto, Vito; Ceccarelli, Matteo; Casu, Mariano

    2015-09-15

    The voltage-dependent anion channel (VDAC) is the main mitochondrial porin allowing the exchange of ions and metabolites between the cytosol and the mitochondrion. In addition, VDAC was found to actively interact with proteins playing a fundamental role in the regulation of apoptosis and being of central interest in cancer research. VDAC is a large transmembrane β-barrel channel, whose N-terminal helical fragment adheres to the channel interior, partially closing the pore. This fragment is considered to play a key role in protein stability and function as well as in the interaction with apoptosis-related proteins. Three VDAC isoforms are differently expressed in higher eukaryotes, for which distinct and complementary roles are proposed. In this work, the folding propensity of their N-terminal fragments has been compared. By using multiple spectroscopic techniques, and complementing the experimental results with theoretical computer-assisted approaches, we have characterized their conformational equilibrium. Significant differences were found in the intrinsic helical propensity of the three peptides, decreasing in the following order: hVDAC2 > hVDAC3 > hVDAC1. In light of the models proposed in the literature to explain voltage gating, selectivity, and permeability, as well as interactions with functionally related proteins, our results suggest that the different chemicophysical properties of the N-terminal domain are possibly correlated to different functions for the three isoforms. The overall emerging picture is that a similar transmembrane water accessible conduit has been equipped with not identical domains, whose differences can modulate the functional roles of the three VDAC isoforms.

  6. Frequency and voltage dependence dielectric properties, ac electrical conductivity and electric modulus profiles in Al/Co3O4-PVA/p-Si structures

    Science.gov (United States)

    Bilkan, Çiğdem; Azizian-Kalandaragh, Yashar; Altındal, Şemsettin; Shokrani-Havigh, Roya

    2016-11-01

    In this research a simple microwave-assisted method have been used for preparation of cobalt oxide nanostructures. The as-prepared sample has been investigated by UV-vis spectroscopy, X-ray diffraction (XRD), scanning electron microscopy (SEM). On the other hand, frequency and voltage dependence of both the real and imaginary parts of dielectric constants (ε‧, ε″) and electric modulus (M‧ and M″), loss tangent (tanδ), and ac electrical conductivity (σac) values of Al/Co3O4-PVA/p-Si structures were obtained in the wide range of frequency and voltage using capacitance (C) and conductance (G/ω) data at room temperature. The values of ε‧, ε″ and tanδ were found to decrease with increasing frequency almost for each applied bias voltage, but the changes in these parameters become more effective in the depletion region at low frequencies due to the charges at surface states and their relaxation time and polarization effect. While the value of σ is almost constant at low frequency, increases almost as exponentially at high frequency which are corresponding to σdc and σac, respectively. The M‧ and M″ have low values at low frequencies region and then an increase with frequency due to short-range mobility of charge carriers. While the value of M‧ increase with increasing frequency, the value of M″ shows two peak and the peaks positions shifts to higher frequency with increasing applied voltage due to the decrease of the polarization and Nss effects with increasing frequency.

  7. Effects of Wenxin Keli on the Action Potential and L-Type Calcium Current in Rats with Transverse Aortic Constriction-Induced Heart Failure

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    Yu Chen

    2013-01-01

    Full Text Available Objective. We investigated the effects of WXKL on the action potential (AP and the L-type calcium current (ICa-L in normal and hypertrophied myocytes. Methods. Forty male rats were randomly divided into two groups: the control group and the transverse aortic constriction- (TAC- induced heart failure group. Cardiac hypertrophy was induced by TAC surgery, whereas the control group underwent a sham operation. Eight weeks after surgery, single cardiac ventricular myocytes were isolated from the hearts of the rats. The APs and ICa-L were recorded using the whole-cell patch clamp technique. Results. The action potential duration (APD of the TAC group was prolonged compared with the control group and was markedly shortened by WXKL treatment in a dose-dependent manner. The current densities of the ICa-L in the TAC group treated with 5 g/L WXKL were significantly decreased compared with the TAC group. We also determined the effect of WXKL on the gating mechanism of the ICa-L in the TAC group. We found that WXKL decreased the ICa-L by accelerating the inactivation of the channels and delaying the recovery time from inactivation. Conclusions. The results suggest that WXKL affects the AP and blocked the ICa-L, which ultimately resulted in the treatment of arrhythmias.

  8. Antimigraine drug, zolmitriptan, inhibits high-voltage activated calcium currents in a population of acutely dissociated rat trigeminal sensory neurons

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    Matsuzawa Yoshiyasu

    2006-03-01

    Full Text Available Abstract Background Triptans, 5-HT1B/ID agonists, act on peripheral and/or central terminals of trigeminal ganglion neurons (TGNs and inhibit the release of neurotransmitters to second-order neurons, which is considered as one of key mechanisms for pain relief by triptans as antimigraine drugs. Although high-voltage activated (HVA Ca2+ channels contribute to the release of neurotransmitters from TGNs, electrical actions of triptans on the HVA Ca2+ channels are not yet documented. Results In the present study, actions of zolmitriptan, one of triptans, were examined on the HVA Ca2+ channels in acutely dissociated rat TGNs, by using whole-cell patch recording of Ba2+ currents (IBa passing through Ca2+ channels. Zolmitriptan (0.1–100 μM reduced the size of IBa in a concentration-dependent manner. This zolmitriptan-induced inhibitory action was blocked by GR127935, a 5-HT1B/1D antagonist, and by overnight pretreatment with pertussis toxin (PTX. P/Q-type Ca2+ channel blockers inhibited the inhibitory action of zolmitriptan on IBa, compared to N- and L-type blockers, and R-type blocker did, compared to L-type blocker, respectively (p 1B/1D receptor linked to Gi/o pathway. Conclusion It is concluded that this zolmitriptan inhibition of HVA Ca2+ channels may explain the reduction in the release of neurotransmitters including CGRP, possibly leading to antimigraine effects of zolmitriptan.

  9. Potassium Current Is Not Affected by Long-Term Exposure to Ghrelin or GHRP-6 in Somatotropes GC Cells

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    Belisario Domínguez Mancera

    2013-01-01

    Full Text Available Ghrelin is a growth hormone (GH secretagogue (GHS and GHRP-6 is a synthetic peptide analogue; both act through the GHS receptor. GH secretion depends directly on the intracellular concentration of Ca2+; this is determined from the intracellular reserves and by the entrance of Ca2+ through the voltage-dependent calcium channels, which are activated by the membrane depolarization. Membrane potential is mainly determined by K+ channels. In the present work, we investigated the effect of ghrelin (10 nM or GHRP-6 (100 nM for 96 h on functional expression of voltage-dependent K+ channels in rat somatotropes: GC cell line. Physiological patch-clamp whole-cell recording was used to register the K+ currents. With Cd2+ (1 mM and tetrodotoxin (1 μm in the bath solution recording, three types of currents were characterized on the basis of their biophysical and pharmacological properties. GC cells showed a K+ current with a transitory component sensitive to 4-aminopyridine, which represents ~40% of the total outgoing current; a sustained component named delayed rectifier , sensitive to tetraethylammonium; and a third type of K+ current was recorded at potentials more negative than −80 mV, permitting the entrance of K+ named inward rectifier (KIR. Chronic treatment with ghrelin or GHRP-6 did not modify the functional expression of K+ channels, without significant changes ( in the amplitudes of the three currents observed; in addition, there were no modifications in their biophysical properties and kinetic activation or inactivation.

  10. Impaired beta-adrenergic response and decreased L-type calcium current of hypertrophied left ventricular myocytes in postinfarction heart failure

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    R.M. Saraiva

    2003-05-01

    Full Text Available Infarct-induced heart failure is usually associated with cardiac hypertrophy and decreased ß-adrenergic responsiveness. However, conflicting results have been reported concerning the density of L-type calcium current (I Ca(L, and the mechanisms underlying the decreased ß-adrenergic inotropic response. We determined I Ca(L density, cytoplasmic calcium ([Ca2+]i transients, and the effects of ß-adrenergic stimulation (isoproterenol in a model of postinfarction heart failure in rats. Left ventricular myocytes were obtained by enzymatic digestion 8-10 weeks after infarction. Electrophysiological recordings were obtained using the patch-clamp technique. [Ca2+]i transients were investigated via fura-2 fluorescence. ß-Adrenergic receptor density was determined by [³H]-dihydroalprenolol binding to left ventricle homogenates. Postinfarction myocytes showed a significant 25% reduction in mean I Ca(L density (5.7 ± 0.28 vs 7.6 ± 0.32 pA/pF and a 19% reduction in mean peak [Ca2+]i transients (0.13 ± 0.007 vs 0.16 ± 0.009 compared to sham myocytes. The isoproterenol-stimulated increase in I Ca(L was significantly smaller in postinfarction myocytes (Emax: 63.6 ± 4.3 vs 123.3 ± 0.9% in sham myocytes, but EC50 was not altered. The isoproterenol-stimulated peak amplitude of [Ca2+]i transients was also blunted in postinfarction myocytes. Adenylate cyclase activation through forskolin produced similar I Ca(L increases in both groups. ß-Adrenergic receptor density was significantly reduced in homogenates from infarcted hearts (Bmax: 93.89 ± 20.22 vs 271.5 ± 31.43 fmol/mg protein in sham myocytes, while Kd values were similar. We conclude that postinfarction myocytes from large infarcts display reduced I Ca(L density and peak [Ca2+]i transients. The response to ß-adrenergic stimulation was also reduced and was probably related to ß-adrenergic receptor down-regulation and not to changes in adenylate cyclase activity.

  11. Lipoxin A4 stimulates calcium-activated chloride currents and increases airway surface liquid height in normal and cystic fibrosis airway epithelia.

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    Valia Verrière

    Full Text Available Cystic Fibrosis (CF is a genetic disease characterised by a deficit in epithelial Cl(- secretion which in the lung leads to airway dehydration and a reduced Airway Surface Liquid (ASL height. The endogenous lipoxin LXA(4 is a member of the newly identified eicosanoids playing a key role in ending the inflammatory process. Levels of LXA(4 are reported to be decreased in the airways of patients with CF. We have previously shown that in normal human bronchial epithelial cells, LXA(4 produced a rapid and transient increase in intracellular Ca(2+. We have investigated, the effect of LXA(4 on Cl(- secretion and the functional consequences on ASL generation in bronchial epithelial cells obtained from CF and non-CF patient biopsies and in bronchial epithelial cell lines. We found that LXA(4 stimulated a rapid intracellular Ca(2+ increase in all of the different CF bronchial epithelial cells tested. In non-CF and CF bronchial epithelia, LXA(4 stimulated whole-cell Cl(- currents which were inhibited by NPPB (calcium-activated Cl(- channel inhibitor, BAPTA-AM (chelator of intracellular Ca(2+ but not by CFTRinh-172 (CFTR inhibitor. We found, using confocal imaging, that LXA(4 increased the ASL height in non-CF and in CF airway bronchial epithelia. The LXA(4 effect on ASL height was sensitive to bumetanide, an inhibitor of transepithelial Cl(- secretion. The LXA(4 stimulation of intracellular Ca(2+, whole-cell Cl(- currents, conductances and ASL height were inhibited by Boc-2, a specific antagonist of the ALX/FPR2 receptor. Our results provide, for the first time, evidence for a novel role of LXA(4 in the stimulation of intracellular Ca(2+ signalling leading to Ca(2+-activated Cl(- secretion and enhanced ASL height in non-CF and CF bronchial epithelia.

  12. Lipoxin A4 stimulates calcium-activated chloride currents and increases airway surface liquid height in normal and cystic fibrosis airway epithelia.

    LENUS (Irish Health Repository)

    2012-01-01

    Cystic Fibrosis (CF) is a genetic disease characterised by a deficit in epithelial Cl(-) secretion which in the lung leads to airway dehydration and a reduced Airway Surface Liquid (ASL) height. The endogenous lipoxin LXA(4) is a member of the newly identified eicosanoids playing a key role in ending the inflammatory process. Levels of LXA(4) are reported to be decreased in the airways of patients with CF. We have previously shown that in normal human bronchial epithelial cells, LXA(4) produced a rapid and transient increase in intracellular Ca(2+). We have investigated, the effect of LXA(4) on Cl(-) secretion and the functional consequences on ASL generation in bronchial epithelial cells obtained from CF and non-CF patient biopsies and in bronchial epithelial cell lines. We found that LXA(4) stimulated a rapid intracellular Ca(2+) increase in all of the different CF bronchial epithelial cells tested. In non-CF and CF bronchial epithelia, LXA(4) stimulated whole-cell Cl(-) currents which were inhibited by NPPB (calcium-activated Cl(-) channel inhibitor), BAPTA-AM (chelator of intracellular Ca(2+)) but not by CFTRinh-172 (CFTR inhibitor). We found, using confocal imaging, that LXA(4) increased the ASL height in non-CF and in CF airway bronchial epithelia. The LXA(4) effect on ASL height was sensitive to bumetanide, an inhibitor of transepithelial Cl(-) secretion. The LXA(4) stimulation of intracellular Ca(2+), whole-cell Cl(-) currents, conductances and ASL height were inhibited by Boc-2, a specific antagonist of the ALX\\/FPR2 receptor. Our results provide, for the first time, evidence for a novel role of LXA(4) in the stimulation of intracellular Ca(2+) signalling leading to Ca(2+)-activated Cl(-) secretion and enhanced ASL height in non-CF and CF bronchial epithelia.

  13. Effects of acute and chronic nicotine on elevated plus maze in mice: involvement of calcium channels.

    Science.gov (United States)

    Biala, Grazyna; Budzynska, Barbara

    2006-05-30

    The current experiments examined the anxiety-related effects of acute and repeated nicotine administration using the elevated plus maze test in mice. Nicotine (0.1 mg/kg s.c., 5 and 30 min after injection; 0.5 mg/kg, s.c., 5 min after injection) had an anxiogenic effect, shown by specific decreases in the percentage of time spent on the open arms and in the percentage of open arm entries. Tolerance developed to this anxiogenic action after 6 days of daily nicotine administration (0.1 mg/kg, s.c.). Five minutes after the seventh injection, an anxiolytic effect was observed, i.e., specific increases in the percentage of time spent on the open arms and in the percentage of open arm entries. L-type voltage-dependent calcium channel antagonists nimodipine (5 and 10 mg/kg, i.p.), flunarizine (5 and 10 mg/kg, i.p.), verapamil (5, 10, 20 mg/kg) and diltiazem (5, 10, 20 mg/kg, i.p.) were also injected prior to an acute low dose of nicotine or to each injection of chronic nicotine. Our results revealed that calcium channel blockers dose-dependently attenuated both an anxiogenic effect of nicotine as well as the development of tolerance to this effect. Our results suggest that neural calcium-dependent mechanisms are involved in the anxiety-related responses to acute and chronic nicotine injection that may ultimately lead to addiction and smoking relapse in human smokers.

  14. Difference of Sodium Currents between Pediatric and Adult Human Atrial Myocytes: Evidence for Developmental Changes of Sodium Channels

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    Benzhi Cai, Xiaoqin Mu, Dongmei Gong, Shulin Jiang, Jianping Li, Qingxin Meng, Yunlong Bai, Yanju Liu, Xinyue Wang, Xueying Tan, Baofeng Yang, Yanjie Lu

    2011-01-01

    Full Text Available Voltage-gated calcium currents and potassium currents were shown to undergo developmental changes in postnatal human and animal cardiomocytes. However, so far, there is no evidence whether sodium currents also presented the developmental changes in postnatal human atrial cells. The aim of this study was to observe age-related changes of sodium currents between pediatric and adult atrial myocytes. Human atrial myocytes were acutely isolated and the whole-cell patch clamp technique was used to record sodium currents isolated from pediatric and adult atrial cardiomocytes. The peak amplitude of sodium currents recorded in adult atrial cells was significantly larger than that in pediatric atrial myocytes. However, there was no significant difference of the activation voltage for peak sodium currents between two kinds of atrial myocytes. The time constants for the activation and inactivation of sodium currents were smaller in adult atria than pediatric atria. The further study revealed that the voltage-dependent inactivation of sodium currents were more slow in adult atrial cardiomyocytes than pediatric atrial cells. A significant difference was also observed in the recovery process of sodium channel from inactivation. In summary, a few significant differences were demonstrated in sodium currents characteristics between pediatric and adult atrial myocytes, which indicates that sodium currents in human atria also undergo developmental changes.

  15. A genetic screen for dihydropyridine (DHP-resistant worms reveals new residues required for DHP-blockage of mammalian calcium channels.

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    Trevor C Y Kwok

    2008-05-01

    Full Text Available Dihydropyridines (DHPs are L-type calcium channel (Ca(v1 blockers prescribed to treat several diseases including hypertension. Ca(v1 channels normally exist in three states: a resting closed state, an open state that is triggered by membrane depolarization, followed by a non-conducting inactivated state that is triggered by the influx of calcium ions, and a rapid change in voltage. DHP binding is thought to alter the conformation of the channel, possibly by engaging a mechanism similar to voltage dependent inactivation, and locking a calcium ion in the pore, thereby blocking channel conductance. As a Ca(v1 channel crystal structure is lacking, the current model of DHP action has largely been achieved by investigating the role of candidate Ca(v1 residues in mediating DHP-sensitivity. To better understand DHP-block and identify additional Ca(v1 residues important for DHP-sensitivity, we screened 440,000 randomly mutated Caenorhabditis elegans genomes for worms resistant to DHP-induced growth defects. We identified 30 missense mutations in the worm Ca(v1 pore-forming (alpha(1 subunit, including eleven in conserved residues known to be necessary for DHP-binding. The remaining polymorphisms are in eight conserved residues not previously associated with DHP-sensitivity. Intriguingly, all of the worm mutants that we analyzed phenotypically exhibited increased channel activity. We also created orthologous mutations in the rat alpha(1C subunit and examined the DHP-block of current through the mutant channels in culture. Six of the seven mutant channels examined either decreased the DHP-sensitivity of the channel and/or exhibited significant residual current at DHP concentrations sufficient to block wild-type channels. Our results further support the idea that DHP-block is intimately associated with voltage dependent inactivation and underscores the utility of C. elegans as a screening tool to identify residues important for DHP interaction with mammalian

  16. —Part I. Interaction of Calcium and Copper-Calcium Alloy with Electrolyte

    Science.gov (United States)

    Zaikov, Yurii P.; Batukhtin, Victor P.; Shurov, Nikolay I.; Ivanovskii, Leonid E.; Suzdaltsev, Andrey V.

    2014-06-01

    This paper describes the interaction between calcium and molten CaCl2 and the solubility of calcium in this melt, depending on the calcium content in the copper-calcium alloy that comes in contact with the molten CaCl2. The negative influence of the dissolved calcium on the current efficiency was verified. The negative effects of moisture and CaO impurities on the calcium current efficiency were demonstrated. The dependence of the current efficiency and the purity of the metal obtained by the electrolysis conditions were studied in a laboratory electrolyzer (20 to 80 A).

  17. Rectification of Acetylcholine-Elicited Currents in PC12 Pheochromocytoma Cells

    Science.gov (United States)

    Ifune, C. K.; Steinbach, J. H.

    1990-06-01

    The current-voltage (I-V) relationship for acetylcholine-elicited currents in the rat pheochromocytoma cell line PC12 is nonlinear. Two voltage-dependent processes that could account for the whole-cell current rectification were examined, receptor channel gating and single receptor channel permeation. We found that both factors are involved in the rectification of the whole-cell currents. The voltage dependence of channel gating determines the shape of the I-V curve at negative potentials. The single-channel I-V relationship is inwardly rectifying and largely responsible for the characteristic shape of the whole-cell I-V curve at positive potentials. The rectification of the single-channel currents is produced by the voltage-dependent block of outward currents by intracellular Mg2+ ions.

  18. Fruit Calcium: Transport and Physiology

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    Bradleigh eHocking

    2016-04-01

    Full Text Available Calcium has well-documented roles in plant signaling, water relations and cell wall interactions. Significant research into how calcium impacts these individual processes in various tissues has been carried out; however, the influence of calcium on fruit ripening has not been thoroughly explored. Here, we review the current state of knowledge on how calcium may impact fruit development, physical traits and disease susceptibility through facilitating developmental and stress response signaling, stabilizing membranes, influencing water relations and modifying cell wall properties through cross-linking of de-esterified pectins. We explore the involvement of calcium in hormone signaling integral to ripening and the physiological mechanisms behind common disorders that have been associated with fruit calcium deficiency (e.g. blossom end rot in tomatoes or bitter pit in apples. This review works towards an improved understanding of how the many roles of calcium interact to influence fruit ripening, and proposes future research directions to fill knowledge gaps. Specifically, we focus mostly on grapes and present a model that integrates existing knowledge around these various functions of calcium in fruit, which provides a basis for understanding the physiological impacts of sub-optimal calcium nutrition in grapes. Calcium accumulation and distribution in fruit is shown to be highly dependent on water delivery and cell wall interactions in the apoplasm. Localized calcium deficiencies observed in particular species or varieties can result from differences in xylem morphology, fruit water relations and pectin composition, and can cause leaky membranes, irregular cell wall softening, impaired hormonal signaling and aberrant fruit development. We propose that the role of apoplasmic calcium-pectin crosslinking, particularly in the xylem, is an understudied area that may have a key influence on fruit water relations. Furthermore, we believe that improved

  19. 心房肌细胞钙通道和钾通道免疫电镜定位研究%The study of distribution of calcium channel and potassium channel in atrial myocytes by immunoelectromicroscopy

    Institute of Scientific and Technical Information of China (English)

    袁平; 张建成; 何爱华; 颜永碧

    2002-01-01

    @@ 离子通道是一种跨膜蛋白,迄今为止,尚未见关于离子通道的细胞内分布情况的报道.心脏L-型电压依赖钙通道(L-type voltage dependent calcium channel, LVDCC)和电压依赖KV4.3钾通道(voltage dependent potassium channel,VDKV4.3)对心肌细胞动作电位有重要作用.

  20. 穿孔膜片钳方法记录L型钙通道及脱氢紫堇碱对其影响的研究%Recording L-type calcium channel current by perforated patch clamp and effect of dehydrocorydaline on L-type calcium channel

    Institute of Scientific and Technical Information of China (English)

    孟红旭; 王宝; 刘建勋

    2011-01-01

    Aim To observe the difference of recording L-type calcium channel currents over time through by whole cell patch clamp and perforated patch clamp method and the effect of dehydrocorydaline on L-type calcium channel by perforated patch clamp method.Methods Whole cell patch clamp and perforated patch clamp were used to record L-type calcium channel current in acutely isolated rat ventricular myocytes.Results The L-type calcium channel current peak recorded by whole cell patch clamp decayed of the ( 34 ±23 )% ( n =10 ) within 15 minutes, while using perforated patch clamp, L-type calcium channel current peak attenuated ( 2. 7 ±3. 4 )% ( n =9 ) within 15 minutes; The inhibitory effect of ginsenoside Re( 100μmol · L-1 ) could be clearlv recorded by perforated patch clamp method. while the current decaying produced by whole cell patch clamp almost completely overshadowed the effects of ginsenoside Re. Dehydrocorydaline ( 10, 100 μmol · L-1 ) could inhibit L-type calcium channel current peak and the inhibitory rates were ( 9 ±7. 5 )%( n =5 )and ( 28. 6 ±8. 5 )%( n =5 )individually. Conclusions Perforated patch clamp method has more stability and accuracy than ordinary whole-cell patch clamp technique in recording L-type calcium channel current; dehydrocorydaline can suppress L-type calcium channel in a concentration-dependent manner.%目的 比较全细胞膜片钳和穿孔膜片钳方法记录大鼠心室肌细胞L型钙通道电流随时间经过的变化差异,并观察脱氢紫堇碱对L型钙通道的影响.方法 采用全细胞膜片钳和穿孔膜片钳方法记录急性分离的大鼠心室肌细胞L型钙通道电流.结果 采用全细胞膜片钳法记录到的L型钙通道电流峰值在15 min内衰减了(34±23)%(n=10),采用穿孔膜片钳方法记录到的L型钙通道电流峰值15 min内仅衰减了(2.7±3.4)%(n=9);采用穿孔膜片钳方法能够记录到人参皂苷Re(100 μmol·L-1)的抑制效应,而采用全细胞膜片钳方法产

  1. 心房颤动24 h和48 h对犬心房肌有效不应期及L型钙电流的影响%Effects of 24 h and 48 h atrial fibrillation on L-type calcium currents in canine atrial myocytes

    Institute of Scientific and Technical Information of China (English)

    张玲; 孙娟; 张广伟; 王欢; 侯月梅

    2013-01-01

    Objective To evaluate the effects of 24 h and 48 h atrial fibrillation(AF) on L-type calcium current (ICa-L)in atrial myocytes,and to investigate its possible mechanism and concordance.Methods Eighteen healthy adult mongrel canines were randomly divided into three groups:control group,24 h AF group and 48 h AF group,6 in each group.AF model was established by right atrial pacing with 600 bpm.The effective refractory period(ERP) of right atrial was measured by program stimulation.The atrial myocytes isolated from 24 h and 48 h AF canine,L-type calcium channel currents in atrial myocytes were recorded by patch clamp technique in the whole cell mode.The expressions of L-type voltage-dependent Ca2+ channel(LVDCC) α1c protein in atrium were measured by immunohistochemistry in three groups,semi-quantitative analysis of α1c protein expression was performed with pathological image analytic system.Results In 24 h AF group and 48 h AF group,AF can be induced in all the experiment animal.The pattern of ERP with an rapid drop in 6 h followed by a more gradual decline during hours 6 through 48 as the result of rapid pacing.The mean ERP was decreased by (12.13 ± 2.24) ms after 6 hours of pacing (from 141.00 ± 15.23 to 129.50 ± 8.64) ms (P < 0.01).Similarly,after 24 h pacing the mean ERP was decreased by (19.23 ± 2.14) ms (P< 0.01),and by (28.15 ±4.26) ms after 48 h pacing(P<0.01).Compared with control group(-6.69±0.08)pA/pF,the peak density of ICa-L was significantly reduced in 24 h AF group(-4.83±0.30)pA/pF,and it was reduced more in 48 h AF group(-3.70±0.50)pA/pF.Compared with control group,the protein expression of LVDCC α1c decreased significantly in group of 48 h and 24 h AF(P<0.05),but in the group of 48 h AF it decreased more (P<0.01).Conclusion The decrease of ICa-L may contribute to ionic basis of acute electrical remodeling of atrial cardiomyocytes caused by 24 h and 48 h AF in canine,which suggested that calcium channels blocker might prevent AF

  2. Aqueous leaf extract of Averrhoa carambola L. (Oxalidaceae reduces both the inotropic effect of BAY K 8644 on the guinea pig atrium and the calcium current on GH3cells

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    Carla M. L. Vasconcelos

    2008-12-01

    Full Text Available It was previously showed that aqueous leaf extract (AqEx of Averrhoa carambola depresses the guinea pig atrial inotropism. Therefore, experiments were carried out on guinea pig left atrium and on pituitary GH3 cells in order to evaluate the effect of AqEx on the cellular calcium influx. The atrium was mounted in an organ chamber (5 mL, Tyrode, 27 ± 0.1 ºC, 95 % O2, 5 % CO2, stretched to 10 mN, and paced at 2 Hz (0.5 ms, 400 V and GH3 cells were submitted to a whole cell voltage clamp configuration. In the atrium, the AqEx (1500 µg/mL shifted to the right the concentration-effect curve of the positive inotropic effect produced by (± BAY K 8644, an L-type calcium channel agonist. The AqEx increased EC50 (concentration required to promote 50% of the maximum effect of the inotropic effect of BAY K 8644 from 7.8 ± 0.38 to 115.1 ± 0.44 nM (N = 3; p < 0.05. In GH3 cells assayed with 500 µg/mL of AqEx, the L-type calcium inward current declined 30 % (from 282 to 190 pA. Nevertheless, the extract did not change the voltage correspondent to the peak current. These data suggest that, at least in part, the negative inotropic effect of AqEx on the guinea pig atrium is due to a reduction of the L-type calcium current.

  3. ß-Adrenoceptor Activation Enhances L-Type Calcium Channel Currents in Anterior Piriform Cortex Pyramidal Cells of Neonatal Mice: Implication for Odor Learning

    Science.gov (United States)

    Ghosh, Abhinaba; Mukherjee, Bandhan; Chen, Xihua; Yuan, Qi

    2017-01-01

    Early odor preference learning occurs in one-week-old rodents when a novel odor is paired with a tactile stimulation mimicking maternal care. ß-Adrenoceptors and L-type calcium channels (LTCCs) in the anterior piriform cortex (aPC) are critically involved in this learning. However, whether ß-adrenoceptors interact directly with LTCCs in aPC…

  4. Calcium signaling in neocortical development.

    Science.gov (United States)

    Uhlén, Per; Fritz, Nicolas; Smedler, Erik; Malmersjö, Seth; Kanatani, Shigeaki

    2015-04-01

    The calcium ion (Ca(2+) ) is an essential second messenger that plays a pivotal role in neurogenesis. In the ventricular zone (VZ) of the neocortex, neural stem cells linger to produce progenitor cells and subsequently neurons and glial cells, which together build up the entire adult brain. The radial glial cells, with their characteristic radial fibers that stretch from the inner ventricular wall to the outer cortex, are known to be the neural stem cells of the neocortex. Migrating neurons use these radial fibers to climb from the proliferative VZ in the inner part of the brain to the outer layers of the cortex, where differentiation processes continue. To establish the complex structures that constitute the adult cerebral cortex, proliferation, migration, and differentiation must be tightly controlled by various signaling events, including cytosolic Ca(2+) signaling. During development, cells regularly exhibit spontaneous Ca(2+) activity that stimulates downstream effectors, which can elicit these fundamental cell processes. Spontaneous Ca(2+) activity during early neocortical development depends heavily on gap junctions and voltage dependent Ca(2+) channels, whereas later in development neurotransmitters and synapses exert an influence. Here, we provide an overview of the literature on Ca(2+) signaling and its impact on cell proliferation, migration, and differentiation in the neocortex. We point out important historical studies and review recent progress in determining the role of Ca(2+) signaling in neocortical development.

  5. Calcium in diet

    Science.gov (United States)

    ... D is needed to help your body use calcium. Milk is fortified with vitamin D for this reason. ... of calcium dietary supplements include calcium citrate and calcium carbonate. Calcium citrate is the more expensive form of ...

  6. Calcium Phosphate Biomaterials: An Update

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Current calcium phosphate (CaP) biomaterials for bone repair, substitution, augmentation and regeneration include hydroxyapatite ( HA ) from synthetic or biologic origin, beta-tricalcium phosphate ( β-TCP ) , biphasic calcium phosphate (BCP), and are available as granules, porous blocks, components of composites (CaP/polymer) cements, and as coatings on orthopedic and dental implants. Experimental calcium phosphate biomaterials include CO3- and F-substituted apatites, Mg-and Zn-substituted β-TCP, calcium phosphate glasses. This paper is a brief review of the different types of CaP biomaterials and their properties such as bioactivity, osteoconductivity, osteoinductivity.

  7. Membrane Currents in Airway Smooth Muscle: Mechanisms and Therapeutic Implications

    Directory of Open Access Journals (Sweden)

    Luke J Janssen

    1997-01-01

    Full Text Available Electrophysiological and pharmacological techniques were used to characterize the membrane conductance changes underlying spasmogen-evoked depolarization in airway smooth muscle (ASM. Changes included a transient activation of chloride ion channels and prolonged suppression of potassium ion channels; both changes are triggered by release of internally sequestered calcium ion and in turn cause opening of voltage-dependent calcium channels. The resultant influx of calcium ions contributes to contraction as well as to refilling of the internal calcium ion pool. Bronchodilators, on the other hand, act in part through activation of potassium channels, with consequent closure of calcium channels. The tools used to study ion channels in ASM are described, and the investigations of the roles of ion channels in ASM physiology (autacoid-evoked depolarization and hyperpolarization and pathophysiology (airway hyperresponsiveness are summarized. Finally, how the relationship between ion channels and ASM function/dysfunction may relate to the treatment of asthma and related breathing disorders is discussed.

  8. Calcium sparks in the intact gerbil spiral modiolar artery

    Directory of Open Access Journals (Sweden)

    Berge Samantha

    2011-08-01

    Full Text Available Abstract Background Calcium sparks are ryanodine receptor mediated transient calcium signals that have been shown to hyperpolarize the membrane potential by activating large conductance calcium activated potassium (BK channels in vascular smooth muscle cells. Along with voltage-dependent calcium channels, they form a signaling unit that has a vasodilatory influence on vascular diameter and regulation of myogenic tone. The existence and role of calcium sparks has hitherto been unexplored in the spiral modiolar artery, the end artery that controls blood flow to the cochlea. The goal of the present study was to determine the presence and properties of calcium sparks in the intact gerbil spiral modiolar artery. Results Calcium sparks were recorded from smooth muscle cells of intact arteries loaded with fluo-4 AM. Calcium sparks occurred with a frequency of 2.6 Hz, a rise time of 17 ms and a time to half-decay of 20 ms. Ryanodine reduced spark frequency within 3 min from 2.6 to 0.6 Hz. Caffeine (1 mM increased spark frequency from 2.3 to 3.3 Hz and prolonged rise and half-decay times from 17 to 19 ms and from 20 to 23 ms, respectively. Elevation of potassium (3.6 to 37.5 mM, presumably via depolarization, increased spark frequency from 2.4 to 3.2 Hz. Neither ryanodine nor depolarization changed rise or decay times. Conclusions This is the first characterization of calcium sparks in smooth muscle cells of the spiral modiolar artery. The results suggest that calcium sparks may regulate the diameter of the spiral modiolar artery and cochlear blood flow.

  9. Beta-scorpion toxin effects suggest electrostatic interactions in domain II of voltage-dependent sodium channels. : Electrostatic interactions between segments IIS2, IIS3 and IIS4 of Na+ channel.

    OpenAIRE

    Mantegazza, Massimo; Cestèle, Sandrine

    2005-01-01

    International audience; Beta-scorpion toxins specifically modulate the voltage dependence of sodium channel activation by acting through a voltage-sensor trapping model. We used mutagenesis, functional analysis and the action of beta-toxin as tools to investigate the existence and role in channel activation of molecular interactions between the charged residues of the S2, S3 and S4 segments in domain II of sodium channels. Mutating to arginine the acidic residues of the S2 and S3 transmembran...

  10. Calcium supplements

    Science.gov (United States)

    ... Related Bone Diseases National Resource Center. Calcium and vitamin D: Important at every age. NIAMS.NIH.gov website. www.niams.nih.gov/Health_Info/Bone/Bone_Health/Nutrition . Updated May 2015. Accessed March ...

  11. Mechanism of store-operated calcium entry

    Indian Academy of Sciences (India)

    Devkanya Dutta

    2000-12-01

    Activation of receptors coupled to the phospholipase C/IP3 signalling pathway results in a rapid release of calcium from its intracellular stores, eventually leading to depletion of these stores. Calcium store depletion triggers an influx of extracellular calcium across the plasma membrane, a mechanism known as the store-operated calcium entry or capacitative calcium entry. Capacitative calcium current plays a key role in replenishing calcium stores and activating various physiological processes. Despite considerable efforts, very little is known about the molecular nature of the capacitative channel and the signalling pathway that activates it. This review summarizes our current knowledge about store operated calcium entry and suggests possible hypotheses for its mode of activation.

  12. Chronic electroconvulsive stimulation but not chronic restraint stress modulates mRNA expression of voltage-dependent potassium channels Kv7.2 and Kv11.1 in the rat piriform cortex

    DEFF Research Database (Denmark)

    Hjæresen, Marie-Louise; Hageman, Ida; Plenge, Per

    2008-01-01

    The mechanisms by which stress and electroconvulsive therapy exert opposite effects on the course of major depression are not known. Potential candidates might include the voltage-dependent potassium channels. Potassium channels play an important role in maintaining the resting membrane potential...... and controlling neuronal excitability. To explore this hypothesis, we examined the effects of one or several electroconvulsive stimulations and chronic restraint stress (6 h/day for 21 days) on the expression of voltage-dependent potassium channel Kv7.2, Kv11.1, and Kv11.3 mRNA in the rat brain using in situ...... hybridization. Repeated, but not acute, electroconvulsive stimulation increased Kv7.2 and Kv11.1 mRNA levels in the piriform cortex. In contrast, restraint stress had no significant effect on mRNA expression of Kv7.2, Kv11.1, or Kv11.3 in any of the brain regions examined. Thus, it appears that the investigated...

  13. Calcium channel blockers and Alzheimer's disease

    Institute of Scientific and Technical Information of China (English)

    Yi Tan; Yulin Deng; Hong Qing

    2012-01-01

    Alzheimer's disease is characterized by two pathological hallmarks: amyloid plaques and neurofi-brillary tangles. In addition, calcium homeostasis is disrupted in the course of human aging. Recent research shows that dense plaques can cause functional alteration of calcium signals in mice with Alzheimer's disease. Calcium channel blockers are effective therapeutics for treating Alzheimer's disease. This review provides an overview of the current research of calcium channel blockers in-volved in Alzheimer's disease therapy.

  14. Calcium regulates caveolin-1 expression at the transcriptional level

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Xiao-Yan; Huang, Cheng-Cheng; Kan, Qi-Ming [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Li, Yan [Experimental Animal Center, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Liu, Dan; Zhang, Xue-Cheng [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Sato, Toshinori [Department of Biosciences and Informatics, Keio University, Hiyoshi, Yokohama 223-8522 (Japan); Yamagata, Sadako [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China); Yamagata, Tatsuya, E-mail: tcyamagata@gmail.com [Laboratory of Tumor Biology and Glycobiology, Department of Life Sciences, Shenyang Pharmaceutical University, Shenyang 110016, People' s Republic of China (China)

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer Caveolin-1 expression is regulated by calcium signaling at the transcriptional level. Black-Right-Pointing-Pointer An inhibitor of or siRNA to L-type calcium channel suppressed caveolin-1 expression. Black-Right-Pointing-Pointer Cyclosporine A or an NFAT inhibitor markedly reduced caveolin-1 expression. Black-Right-Pointing-Pointer Caveolin-1 regulation by calcium signaling is observed in several mouse cell lines. -- Abstract: Caveolin-1, an indispensable component of caveolae serving as a transformation suppressor protein, is highly expressed in poorly metastatic mouse osteosarcoma FBJ-S1 cells while highly metastatic FBJ-LL cells express low levels of caveolin-1. Calcium concentration is higher in FBJ-S1 cells than in FBJ-LL cells; therefore, we investigated the possibility that calcium signaling positively regulates caveolin-1 in mouse FBJ-S1 cells. When cells were treated with the calcium channel blocker nifedipine, cyclosporin A (a calcineurin inhibitor), or INCA-6 (a nuclear factor of activated T-cells [NFAT] inhibitor), caveolin-1 expression at the mRNA and protein levels decreased. RNA silencing of voltage-dependent L-type calcium channel subunit alpha-1C resulted in suppression of caveolin-1 expression. This novel caveolin-1 regulation pathway was also identified in mouse NIH 3T3 cells and Lewis lung carcinoma cells. These results indicate that caveolin-1 is positively regulated at the transcriptional level through a novel calcium signaling pathway mediated by L-type calcium channel/Ca{sup 2+}/calcineurin/NFAT.

  15. Reconstruction of Cell Surface Densities of Ion Pumps, Exchangers, and Channels from mRNA Expression, Conductance Kinetics, Whole-Cell Calcium, and Current-Clamp Voltage Recordings, with an Application to Human Uterine Smooth Muscle Cells.

    Directory of Open Access Journals (Sweden)

    Jolene Atia

    2016-04-01

    Full Text Available Uterine smooth muscle cells remain quiescent throughout most of gestation, only generating spontaneous action potentials immediately prior to, and during, labor. This study presents a method that combines transcriptomics with biophysical recordings to characterise the conductance repertoire of these cells, the 'conductance repertoire' being the total complement of ion channels and transporters expressed by an electrically active cell. Transcriptomic analysis provides a set of potential electrogenic entities, of which the conductance repertoire is a subset. Each entity within the conductance repertoire was modeled independently and its gating parameter values were fixed using the available biophysical data. The only remaining free parameters were the surface densities for each entity. We characterise the space of combinations of surface densities (density vectors consistent with experimentally observed membrane potential and calcium waveforms. This yields insights on the functional redundancy of the system as well as its behavioral versatility. Our approach couples high-throughput transcriptomic data with physiological behaviors in health and disease, and provides a formal method to link genotype to phenotype in excitable systems. We accurately predict current densities and chart functional redundancy. For example, we find that to evoke the observed voltage waveform, the BK channel is functionally redundant whereas hERG is essential. Furthermore, our analysis suggests that activation of calcium-activated chloride conductances by intracellular calcium release is the key factor underlying spontaneous depolarisations.

  16. Reconstruction of Cell Surface Densities of Ion Pumps, Exchangers, and Channels from mRNA Expression, Conductance Kinetics, Whole-Cell Calcium, and Current-Clamp Voltage Recordings, with an Application to Human Uterine Smooth Muscle Cells.

    Science.gov (United States)

    Atia, Jolene; McCloskey, Conor; Shmygol, Anatoly S; Rand, David A; van den Berg, Hugo A; Blanks, Andrew M

    2016-04-01

    Uterine smooth muscle cells remain quiescent throughout most of gestation, only generating spontaneous action potentials immediately prior to, and during, labor. This study presents a method that combines transcriptomics with biophysical recordings to characterise the conductance repertoire of these cells, the 'conductance repertoire' being the total complement of ion channels and transporters expressed by an electrically active cell. Transcriptomic analysis provides a set of potential electrogenic entities, of which the conductance repertoire is a subset. Each entity within the conductance repertoire was modeled independently and its gating parameter values were fixed using the available biophysical data. The only remaining free parameters were the surface densities for each entity. We characterise the space of combinations of surface densities (density vectors) consistent with experimentally observed membrane potential and calcium waveforms. This yields insights on the functional redundancy of the system as well as its behavioral versatility. Our approach couples high-throughput transcriptomic data with physiological behaviors in health and disease, and provides a formal method to link genotype to phenotype in excitable systems. We accurately predict current densities and chart functional redundancy. For example, we find that to evoke the observed voltage waveform, the BK channel is functionally redundant whereas hERG is essential. Furthermore, our analysis suggests that activation of calcium-activated chloride conductances by intracellular calcium release is the key factor underlying spontaneous depolarisations.

  17. 钙离子通道与老龄心房颤动心房电重构的研究进展%Advances in Research of Calcium Channel Current and Atrial Electrical Remodeling of Atrial Fibrillation

    Institute of Scientific and Technical Information of China (English)

    李耀东; 汤宝鹏

    2011-01-01

    随着心房颤动电生理机制研究的广泛深人,已经认识到离子通道重构在心房颤动的发生和维持过程中起重要作用.L-型钙通道及其基因表达的改变可能是老年人容易发生心房颤动电生理重建的离子和分子基础.现就心脏钙离子通道及年龄与心房颤动的研究进展予以综述.%It is important to recognize that atrial fibrillation (AF) modifies the calcium channel of the atrium promotes its occurrence and maintenance. Changes of L-type calcium channels and their gene expression in the elderly may explain ion electrophysiology and molecular reconstruction in AF. This review analyzes the relationship between the heart calcium channel current and AF in the aged.

  18. The mechanism of hetero-synaptic interaction based on spatiotemporal intracellular calcium dynamics.

    Directory of Open Access Journals (Sweden)

    Daiki Futagi

    2014-03-01

    Full Text Available In recent physiological experiments focusing on synaptic plasticity, it is shown that synaptic modifications induced at one synapse are accompanied by hetero-synaptic changes at neighbor sites (Bi, 2002. These evidences imply that the hetero-synaptic interaction plays an important role in reconfiguration of synaptic connections to form and maintain functional neural circuits (Takahashi et al., 2012. Although the mechanism of the interaction is still unclear, some physiological studies suggest that the hetero-synaptic interaction could be caused by propagation of intracellular calcium signals (Nishiyama et al., 2000. Concretely, a spike-triggered calcium increase initiates calcium ion propagation along a dendrite through activation of molecular processes at neighboring sites. Here we hypothesized that the mechanism of the hetero-synaptic interaction was based on the intracellular calcium signaling, which is regulated by interactions between NMDA receptors (NMDARs, voltage-dependent calcium channels (VDCCs and Ryanodine receptors (RyRs on endoplasmic reticulum (ER. To assess realizability of the hypothesized interaction mechanism, we simulated intracellular calcium dynamics at a cellular level, using the computational model that integrated the model of intracellular calcium dynamics (Keizer and Levine, 1996 and the multi-compartment neuron model (Poirazi et al., 2003. Using the proposed computational model, we induced calcium influxes at a local site in postsynaptic dendrite by controlling the spike timings of pre- and postsynaptic neurons. As a result, synchronized calcium influxes through NMDARs and VDCCs caused calcium release from ER. According to the phase plane analysis, RyR-mediated calcium release occurred when the calcium concentration in cytoplasm sufficiently increased under the condition of a high calcium concentration in ER. An NMDAR-mediated calcium influx was slow and persistent, consequently responsible for maintaining a high

  19. Characterisation of marrubenol, a diterpene extracted from Marrubium vulgare, as an L-type calcium channel blocker.

    Science.gov (United States)

    El-Bardai, Sanae; Wibo, Maurice; Hamaide, Marie-Christine; Lyoussi, Badiaa; Quetin-Leclercq, Joelle; Morel, Nicole

    2003-12-01

    1. The objective of the present study was to investigate the mechanism of the relaxant activity of marrubenol, a diterpenoid extracted from Marrubium vulgare. In rat aorta, marrubenol was a more potent inhibitor of the contraction evoked by 100 mM KCl (IC50: 11.8+/-0.3 microM, maximum relaxation: 93+/-0.6%) than of the contraction evoked by noradrenaline (maximum relaxation: 30+/-1.5%). 2. In fura-2-loaded aorta, marrubenol simultaneously inhibited the Ca2+ signal and the contraction evoked by 100 mM KCl, and decreased the quenching rate of fura-2 fluorescence by Mn2+. 3. Patch-clamp data obtained in aortic smooth muscle cells (A7r5) indicated that marrubenol inhibited Ba2+ inward current in a voltage-dependent manner (KD: 8+/-2 and 40+/-6 microM at holding potentials of -50 and -100 mV, respectively). 4. These results showed that marrubenol inhibits smooth muscle contraction by blocking L-type calcium channels.

  20. A persistent sodium current contributes to oscillatory activity in heart interneurons of the medicinal leech.

    Science.gov (United States)

    Opdyke, C A; Calabrese, R L

    1994-12-01

    1. Normal activity in bilateral pairs of heart interneurons, from ganglia 3 or 4, in the medicinal leech (Hirudo medicinalis) is antiphasic due to their reciprocally inhibitory connections. However, Ca(++)-free Co(++)-containing salines lead to synchronous oscillations in these neurons. 2. Internal TEA+ allows expression of full plateaus during Co++ induced oscillations in heart interneurons; these plateaus are not blocked by Cs+. Similar plateaus are also observed with internal TEA+ alone, but under these conditions activity in heart interneurons from ganglia 3 or 4 is antiphasic. 3. Plateaus in heart interneurons induced by Co++ and internal TEA+ involve a conductance increase. 4. A voltage-dependent inward current, IP, showing little inactivation, was isolated using single-electrode voltage-clamp in heart interneurons. This current is carried at least in part by Na+; the current is reduced when external Na+ is reduced and is carried by Li++ when substituted for Na+. 5. Calcium channel blockers such as La3+ and Co++ block neither the TEA+ induced plateaus nor IP, suggesting that Na+ is not using Ca++ channels. Moreover, IP is enhanced by Ca(++)-free CO(++)-containing salines. Thus, IP is correlated with the TEA(+)- and Co(++)-induced plateau behavior.

  1. Calcium and bones

    Science.gov (United States)

    Bone strength and calcium ... calcium (as well as phosphorus) to make healthy bones. Bones are the main storage site of calcium in ... your body does not absorb enough calcium, your bones can get weak or will not grow properly. ...

  2. Differential regulation of voltage- and calcium-activated potassium channels in human B lymphocytes.

    Science.gov (United States)

    Partiseti, M; Choquet, D; Diu, A; Korn, H

    1992-06-01

    The expression and characteristics of K+ channels of human B lymphocytes were studied by using single and whole-cell patch-clamp recordings. They were gated by depolarization (voltage-gated potassium current, IKv, 11-20 pS) and by an increase in intracellular Ca2+ concentration (calcium-activated potassium current, IKCa, 26 pS), respectively. The level of expression of these channels was correlated with the activational status of the cell. Both conductances are blocked by tetraethylammonium, verapamil, and charybdotoxin, and are insensitive to apamin; 4-aminopyridine blocks IK, preferentially. We used a protein kinase C activator (PMA) or antibodies to membrane Ig (anti-mu) to activate resting splenocytes in culture. Although IKv was recorded in the majority of the resting lymphocytic population, less than 20% of the activated cells expressed this conductance. However, in this subset the magnitude of IKv was 20-fold larger than in resting cells. On the other hand, IKCa was detected in nearly one half of the resting cells, whereas all activated cells expressed this current. The magnitude of IKCa was, on average, 30 times larger in activated than in nonactivated cells. These results probably reflect that during the course of activation 1) the number of voltage-dependent K+ channels per cell decreases and increases in a small subset and 2) the number of Ca(2+)-dependent K+ channels per cell increases in all cells. We suggest that the expression of functional Ca(2+)- and voltage-activated K+ channels are under the control of different regulatory signals.

  3. The omega-atracotoxins: selective blockers of insect M-LVA and HVA calcium channels.

    Science.gov (United States)

    Chong, Youmie; Hayes, Jessica L; Sollod, Brianna; Wen, Suping; Wilson, David T; Hains, Peter G; Hodgson, Wayne C; Broady, Kevin W; King, Glenn F; Nicholson, Graham M

    2007-08-15

    The omega-atracotoxins (omega-ACTX) are a family of arthropod-selective peptide neurotoxins from Australian funnel-web spider venoms (Hexathelidae: Atracinae) that are candidates for development as biopesticides. We isolated a 37-residue insect-selective neurotoxin, omega-ACTX-Ar1a, from the venom of the Sydney funnel-web spider Atrax robustus, with high homology to several previously characterized members of the omega-ACTX-1 family. The peptide induced potent excitatory symptoms, followed by flaccid paralysis leading to death, in acute toxicity tests in house crickets. Using isolated smooth and skeletal nerve-muscle preparations, the toxin was shown to lack overt vertebrate toxicity at concentrations up to 1 microM. To further characterize the target of the omega-ACTXs, voltage-clamp analysis using the whole-cell patch-clamp technique was undertaken using cockroach dorsal unpaired median neurons. It is shown here for the first time that omega-ACTX-Ar1a, and its homolog omega-ACTX-Hv1a from Hadronyche versuta, reversibly block both mid-low- (M-LVA) and high-voltage-activated (HVA) insect calcium channel (Ca(v)) currents. This block occurred in the absence of alterations in the voltage-dependence of Ca(v) channel activation, and was voltage-independent, suggesting that omega-ACTX-1 family toxins are pore blockers rather than gating modifiers. At a concentration of 1 microM omega-ACTX-Ar1a failed to significantly affect global K(v) channel currents. However, 1 microM omega-ACTX-Ar1a caused a modest 18% block of insect Na(v) channel currents, similar to the minor block of Na(v) channels reported for other insect Ca(v) channel blockers such as omega-agatoxin IVA. These findings validate both M-LVA and HVA Ca(v) channels as potential targets for insecticides.

  4. Accelerated activation of SOCE current in myotubes from two mouse models of anesthetic- and heat-induced sudden death.

    Directory of Open Access Journals (Sweden)

    Viktor Yarotskyy

    Full Text Available Store-operated calcium entry (SOCE channels play an important role in Ca(2+ signaling. Recently, excessive SOCE was proposed to play a central role in the pathogenesis of malignant hyperthermia (MH, a pharmacogenic disorder of skeletal muscle. We tested this hypothesis by characterizing SOCE current (ISkCRAC magnitude, voltage dependence, and rate of activation in myotubes derived from two mouse models of anesthetic- and heat-induced sudden death: 1 type 1 ryanodine receptor (RyR1 knock-in mice (Y524S/+ and 2 calsequestrin 1 and 2 double knock-out (dCasq-null mice. ISkCRAC voltage dependence and magnitude at -80 mV were not significantly different in myotubes derived from wild type (WT, Y524S/+ and dCasq-null mice. However, the rate of ISkCRAC activation upon repetitive depolarization was significantly faster at room temperature in myotubes from Y524S/+ and dCasq-null mice. In addition, the maximum rate of ISkCRAC activation in dCasq-null myotubes was also faster than WT at more physiological temperatures (35-37°C. Azumolene (50 µM, a more water-soluble analog of dantrolene that is used to reverse MH crises, failed to alter ISkCRAC density or rate of activation. Together, these results indicate that while an increased rate of ISkCRAC activation is a common characteristic of myotubes derived from Y524S/+ and dCasq-null mice and that the protective effects of azumolene are not due to a direct inhibition of SOCE channels.

  5. Calcium Carbonate

    Science.gov (United States)

    ... doctor if you have or have ever had kidney disease or stomach conditions.tell your doctor if you are pregnant, plan to become pregnant, or are breast-feeding. If you become pregnant while taking calcium carbonate, call your doctor.

  6. Calcium Test

    Science.gov (United States)

    ... if a person has symptoms of a parathyroid disorder , malabsorption , or an overactive thyroid. A total calcium level is often measured as part of a routine health screening. It is included in the comprehensive metabolic panel (CMP) and the basic metabolic panel (BMP) , ...

  7. A study on separation of canine atrial myocytes and detection of their L-type calcium channel current%犬心房肌细胞分离及其L型钙通道电流检测的研究

    Institute of Scientific and Technical Information of China (English)

    邹帅; 龙毅; 范晋奇; 周亚南; 高大中; 殷跃辉

    2011-01-01

    目的 探讨一种稳定可靠的犬心房肌细胞的分离方法,提高膜片钳的实验成功率并检测L型钙通道电流.方法 采用改良Langderoff灌流系统行主动脉逆行灌注,采用选择性冠状动脉插管的方法,获得单个心房肌细胞.在显微镜下观察细胞形态,并应用膜片钳全细胞模式记录L型钙通道电流.结果 分离细胞存活率为70%~80%,复钙后细胞存活率为30%~40%,耐钙细胞形态呈杆状,折光性强,横纹清晰,并能稳定记录L型钙通道电流.结论 改良Langderoff技术有助于稳定分离出存活率高、具有正常电生理特性的犬心房肌细胞,能用于心房肌细胞离子通道的研究.%Objective To explore a stable and reliable method of canine atrial myocytes separation for improving the success rate of patch-clamp techniques and detecting the L-type calcium channel current. Methods Modified Langderoff perfusion system was adopted to perform retrograde aortic perfusion and selective coronary artery catheterization was used to obtain single atrial myot-cytes. Microscope was employed to observe the cellular morphology and the L-type calcium channel current was measured by whole-cell patch-clamp recording. Results The survival rate of separated cells was 70% to 80% ,and it was 30% to 40% after recalcifica-tion. A stable L-type calcium channel current could be recorded in calcium-tolerant cells which was rod-shaped with good refraction and clear horizontal stripe. Conclusion Modified Langderoff technique contribute to stable separation of canine atrial myocytes with high survival rate and normal electrophysiological properties,and can be used in the research of the iron channel of atrial myocytes.

  8. Autogenous vein graft thrombosis following exposure to calcium-free solutions (calcium paradox).

    Science.gov (United States)

    Nozick, J H; Farnsworth, P; Montefusco, C M; Parsonnet, V; Ruigrok, T J; Zimmerman, A N

    1981-01-01

    The morphological and functional effects of calcium-free and calcium-containing solutions on canine jugular vein intima were examined under conditions which closely resemble those techniques currently employed in peripheral vascular and aortocoronary bypass surgery. Veins that had been exposed only to calcium-containing solutions remained patent for the duration of the experimental period. Vein perfusion with a calcium-free solution, however, resulted in disruption of the jugular vein intima once calcium ions were reintroduced. Autogenous as a femoral arterial graft became thrombosed within 60 minutes. It is therefore suggested that vein grafts of autogenous origin be irrigated with calcium-containing solutions to prevent intimal damage and thrombosis.

  9. Current blocking and current collection in CIGSe solar cells depending on sodium content

    Energy Technology Data Exchange (ETDEWEB)

    Puttnins, Stefan; Daume, Felix [Solarion AG, Leipzig (Germany); Institut fuer Experimentelle Physik II, Universitaet Leipzig (Germany); Zachmann, Hendrik; Rahm, Andreas [Solarion AG, Leipzig (Germany); Grundmann, Marius [Institut fuer Experimentelle Physik II, Universitaet Leipzig (Germany)

    2010-07-01

    IV-curves of thin film solar cells often show non-idealites like voltage dependent carrier collection and current blocking behaviour. Sodium is long known to improve the efficiency of Cu(In,Ga)Se{sub 2} solar cells by increasing V{sub OC} and FF. However, the way in which sodium influences the electrical properties is still under discussion. We investigated the influence of sodium on voltage dependent carrier collection and current blocking behaviour. Losses caused by incomplete photocurrent collection can be reduced by increased sodium content in the CIGSe layer. Current blocking behaviour like the rollover effect is less pronounced with increased sodium content. The influences were analyzed both in detailed illumination intensity and temperature dependent IV-measurements as well as by extensive statistical analysis over thousands of produced flexible CIGSe solar cells. Theoretical models for this dependency were simulated with SCAPS-1D and show good agreement with respective measurements.

  10. Nicotinic acetylcholine receptor-mediated calcium signaling in the nervous system

    Institute of Scientific and Technical Information of China (English)

    Jian-xin SHEN; Jerrel L YAKEL

    2009-01-01

    Based on the composition of the five subunits forming functional neuronal nicotinic acetylcholine receptors (nAChRs), they are grouped into either heteromeric (comprising both α and β subunits) or homomeric (comprising only α subunits) recep-tors. The nAChRs are known to be differentially permeable to calcium ions, with the α7 nAChR subtype having one of the highest permeabilities to calcium. Calcium influx through nAChRs, particularly through the α-bungarotoxin-sensitive α7-containing nAChRs, is a very efficient way to raise cytoplasmic calcium levels. The activation of nAChRs can mediate three types of cytoplasmic calcium signals: (1) direct calcium influx through the nAChRs, (2) indirect calcium influx through voltage-dependent calcium channels (VDCCs) which are activated by the nAChR-mediated depolarization, and (3) calcium-induced calcium release (CICR) (triggered by the first two sources) from the endoplasmic reticulum (ER) through the ryanodine receptors and inositol (1,4,5)-triphosphate receptors (IP3Rs). Downstream signaling events mediated by nAChR-mediated calcium responses can be grouped into instantaneous effects (such as neurotransmitter release, which can occur in milliseconds after nAChR activation), short-term effects (such as the recovery of nAChR desensitization through cellular signaling cascades), and long-term effects (such as neuroprotection via gene expression). In addition, nAChR activity can be regulated by cytoplasmic calcium levels, suggesting a complex reciprocal relationship. Further advances in imaging techniques, animal models, and more potent and subtype-selective ligands for neuronal nAChRs would help in understand-ing the neuronal nAChR-mediated calcium signaling, and lead to the development of improved therapeutic treatments.

  11. [The characteristics and oxidative modulation of large-conductance calcium-activated potassium channels in guinea-pig colon smooth muscle cells.].

    Science.gov (United States)

    Huang, Wei-Feng; Ouyang, Shou; Zhang, Hui

    2009-06-25

    To investigate the characteristics of large-conductance calcium-activated potassium channels (BK(Ca)) and the effect of hydrogen peroxide (H2O2) on BK(Ca) in guinea-pig proximal colon smooth muscle cells, single smooth muscle cells of guinea-pig colon were enzymatically isolated in low calcium solution containing papain (3 mg/mL), DTT (2 mg/mL), and bovine serum albumin (BSA, 2 mg/mL). Tissues were incubated at 36 degrees C in enzyme solution for 15 min and were then suspended in enzyme-free low calcium solution. Inside-out single channel recording technique was used to record BK(Ca) current. The intracellular (bath) and microelectrode solution both contained symmetrical high potassium. The BK(Ca) in guinea-pig colon smooth muscle cell possesses: 1) voltage-dependence, 2) high selectivity for potassium ion, 3) large conductance (223.7 pS+/-9.2 pS), 4) dependence of [Ca(2+)](i). Intracellular application of H2O2 decreased the open probability (P(o)) of BK(Ca) at low concentration (Ca) at high concentration (5 mmol/L), without affecting the unitary conductance. The effects of H2O2 were reversed by reducing agent dithiothreitol (DTT). Similarly, cysteine specific oxidizing agent, DTNB, also increased or decreased P(o) of BK(Ca) and DTT partially reversed the effect of DTNB. It is thus suggested that H2O2 and DTNB may modulate P(o) of BK(Ca) via the oxidation of cysteine residue.

  12. Calcium channel antagonists suppress cross-tolerance to the anxiogenic effects of D-amphetamine and nicotine in the mouse elevated plus maze test.

    Science.gov (United States)

    Biala, Grazyna; Kruk, Marta

    2008-01-01

    The purpose of the current experiments was to examine the anxiety-related effects of repeated amphetamine and nicotine administration using the mouse elevated plus maze (EPM). d-amphetamine was administered daily for 8 days (2 mg/kg, i.p.). On the 9th day, mice were challenged with amphetamine (2 mg/kg, i.p.) or nicotine (0.1 mg/kg, s.c.), and were tested 30 min after this last injection. Additionally, a distinct group of mice was pretreated with nicotine (0.1 mg/kg, s.c., 6 days). These mice were subjected to nicotine (0.1 mg/kg, s.c.) or amphetamine (2 mg/kg, i.p.) challenge on the seventh day to see if full crossover effects developed after the pretreatment of both psychostimulant drugs. Moreover, the L-type voltage-dependent calcium channel antagonists nimodipine (5 and 10 mg/kg, i.p.), flunarizine (5 and 10 mg/kg, i.p.), verapamil (5 and 10 mg/kg, i.p.) and diltiazem (5 and 10 mg/kg, i.p.) were injected prior to each injection of chronic d-amphetamine or nicotine. We observed cross-tolerance to the anxiogenic effects of d-amphetamine and nicotine that was blunted by a pretreatment with calcium channel blockers. Overall our findings imply that similar neural calcium-dependent mechanisms are involved in the anxiety-related responses to chronic amphetamine and nicotine injections. As anxiety seems to be an important factor for the development of psychostimulant dependence, the L-type VDCC antagonists can offer an interesting approach for the pharmacotherapy of addiction, including amphetamine and/or nicotine dependence.

  13. Fast kinetics of calcium signaling and sensor design.

    Science.gov (United States)

    Tang, Shen; Reddish, Florence; Zhuo, You; Yang, Jenny J

    2015-08-01

    Fast calcium signaling is regulated by numerous calcium channels exhibiting high spatiotemporal profiles which are currently measured by fluorescent calcium sensors. There is still a strong need to improve the kinetics of genetically encoded calcium indicators (sensors) to capture calcium dynamics in the millisecond time frame. In this review, we summarize several major fast calcium signaling pathways and discuss the recent developments and application of genetically encoded calcium indicators to detect these pathways. A new class of genetically encoded calcium indicators designed with site-directed mutagenesis on the surface of beta-barrel fluorescent proteins to form a pentagonal bipyramidal-like calcium binding domain dramatically accelerates calcium binding kinetics. Furthermore, novel genetically encoded calcium indicators with significantly increased fluorescent lifetime change are advantageous in deep-field imaging with high light-scattering and notable morphology change.

  14. Calcium paradox and calcium entry blockers

    NARCIS (Netherlands)

    Ruigrok, T.J.C.; Slade, A.M.; Nayler, W.G.; Meijler, F.L.

    1984-01-01

    Reperfusion of isolated hearts with calcium-containing solution after a short period of calcium-free perfusion results in irreversible cell damage (calcium paradox). This phenomenon is characterized by an excessive influx of calcium into the cells, the rapid onset of myocardial contracture, exhausti

  15. Calcium Flux between the Endoplasmic Reticulum and Mitochondrion Contributes to Poliovirus-Induced Apoptosis▿

    Science.gov (United States)

    Brisac, Cynthia; Téoulé, François; Autret, Arnaud; Pelletier, Isabelle; Colbère-Garapin, Florence; Brenner, Catherine; Lemaire, Christophe; Blondel, Bruno

    2010-01-01

    We show that poliovirus (PV) infection induces an increase in cytosolic calcium (Ca2+) concentration in neuroblastoma IMR5 cells, at least partly through Ca2+ release from the endoplasmic reticulum lumen via the inositol 1,4,5-triphosphate receptor (IP3R) and ryanodine receptor (RyR) channels. This leads to Ca2+ accumulation in mitochondria through the mitochondrial Ca2+ uniporter and the voltage-dependent anion channel (VDAC). This increase in mitochondrial Ca2+ concentration in PV-infected cells leads to mitochondrial dysfunction and apoptosis. PMID:20861253

  16. Comparison of current docking tools for the simulation of inhibitor binding by the transmembrane domain of the sarco/endoplasmic reticulum calcium ATPase

    Science.gov (United States)

    Lape, Michael; Elam, Christopher; Paula, Stefan

    2010-01-01

    Inhibitors of the transmembrane protein sarco/endoplasmic reticulum calcium ATPase (SERCA) are invaluable tools for the study of the enzyme’s physiological functions and they have been recognized as a promising new class of anticancer agents. For the discovery of novel enzyme inhibitors, small molecule docking for virtual screens of large compound libraries has become increasingly important. Since the performance of various docking routines varies considerably, depending on the target and the chemical nature of the ligand, we critically evaluated the performance of four frequently used programs – GOLD, AutoDock, Surflex-Dock, and FRED – for the docking of SERCA inhibitors based on the structures of thapsigargin, di-tert-butylhydroquinone, and cyclopiazonic acid. Evaluation criteria were docking accuracy using crystal structures as references, docking reproducibility, and correlation between docking scores and known bioactivities. The best overall results were obtained by GOLD and FRED. Docking runs with conformationally flexible binding sites produced no significant improvement of the results. PMID:20167416

  17. Store-operated calcium signaling in neutrophils.

    Science.gov (United States)

    Clemens, Regina A; Lowell, Clifford A

    2015-10-01

    Calcium signals in neutrophils are initiated by a variety of cell-surface receptors, including formyl peptide and other GPCRs, FcRs, and integrins. The predominant pathway by which calcium enters immune cells is termed SOCE, whereby plasma membrane CRAC channels allow influx of extracellular calcium into the cytoplasm when intracellular ER stores are depleted. The identification of 2 key families of SOCE regulators, STIM calcium "sensors" and ORAI calcium channels, has allowed for genetic manipulation of SOCE pathways and provided valuable insight into the molecular mechanism of calcium signaling in immune cells, including neutrophils. This review focuses on our current knowledge of the molecules involved in neutrophil SOCE and how study of these molecules has further informed our understanding of the role of calcium signaling in neutrophil activation.

  18. Bioceramics of calcium orthophosphates.

    Science.gov (United States)

    Dorozhkin, Sergey V

    2010-03-01

    A strong interest in use of ceramics for biomedical applications appeared in the late 1960's. Used initially as alternatives to metals in order to increase a biocompatibility of implants, bioceramics have become a diverse class of biomaterials, presently including three basic types: relatively bioinert ceramics, bioactive (or surface reactive) and bioresorbable ones. Furthermore, any type of bioceramics could be porous to provide tissue ingrowth. This review is devoted to bioceramics prepared from calcium orthophosphates, which belong to the categories of bioresorbable and bioactive compounds. During the past 30-40 years, there have been a number of major advances in this field. Namely, after the initial work on development of bioceramics that was tolerated in the physiological environment, emphasis was shifted towards the use of bioceramics that interacted with bones by forming a direct chemical bond. By the structural and compositional control, it became possible to choose whether the bioceramics of calcium orthophosphates was biologically stable once incorporated within the skeletal structure or whether it was resorbed over time. At the turn of the millennium, a new concept of calcium orthophosphate bioceramics, which is able to regenerate bone tissues, has been developed. Current biomedical applications of calcium orthophosphate bioceramics include replacements for hips, knees, teeth, tendons and ligaments, as well as repair for periodontal disease, maxillofacial reconstruction, augmentation and stabilization of the jawbone, spinal fusion and bone fillers after tumor surgery. Potential future applications of calcium orthophosphate bioceramics will include drug-delivery systems, as well as they will become effective carriers of growth factors, bioactive peptides and/or various types of cells for tissue engineering purposes.

  19. Tetraethylammonium-sensitive K(+) current in the bovine spermatozoa and its blocking by the venom of the Chilean Latrodectus mactans.

    Science.gov (United States)

    Parodi, Jorge; Navarrete, Patricia; Marconi, Marcelo; Gutiérrez, Raúl Sánchez; Martínez-Torres, Ataúlfo; Mejías, Fernando Romero

    2010-02-01

    The morphology and size of spermatozoa hinder the study of the functional properties of the spermatozoa plasma membrane. However, some studies have revealed the presence of a number of ion channels in this cell. We set out to measure the endogenous currents and to study the effect of the venom of the Chilean black widow spider (Latrodectus mactans). By patch-clamping bovine spermatozoa our results indicate the presence of an outwardly rectifying current, sensitive to changes in K(+) concentration (30-140 mM) and to tetraethylammonium (TEA, 10-100 mM). The application of the venom (7.5 microg/ml) blocks these K+ currents and then alters the passive properties of the plasma membrane. This leads to the entry of Ca(++), reflected by a change in basal fluorescent units (5+/-2 at 35+/-10 FAU). The Ca(++) influx follows a reduction in the membrane conductance (control 22+/-2; venom 10+/-1 pS), as calcium channels open in accord with voltage dependence.

  20. Differential regulation of potassium currents by FGF-1 and FGF-2 in embryonic Xenopus laevis myocytes.

    Science.gov (United States)

    Chauhan-Patel, R; Spruce, A E

    1998-10-01

    1. Fibroblast growth factors (FGFs) are involved in the regulation of many aspects of muscle development. This study investigated their role in regulating voltage-dependent K+ currents in differentiating Xenopus laevis myocytes. Both FGF-1 and FGF-2 are expressed by developing muscle cells, so their actions were compared. Experiments were performed on cultured myocytes isolated from stage 15 embryos. 2. Long-term exposure of the embryonic myocytes to FGF-1 downregulated inward rectifier K+ current (IK(IR)) density as well as both sustained and inactivating voltage-dependent outward K+ currents (IK,S and IK,I, respectively) and their densities. In contrast, FGF-2 upregulated these currents, although, because of an increase in capacitance caused by FGF-2, current density did not change with this factor. 3. The regulation of IK(IR) by FGF-1 was prevented by the cytoplasmic tyrosine kinase inhibitor herbimycin A, but that of IK,S and IK,I was unaffected, indicating that FGF-1 achieves its regulatory effects on electrical development via separate signalling pathways. The receptor tyrosine kinase inhibitor genistein in isolation suppressed K+ currents, but this may have occurred through a channel-blocking mechanism. 4. In many cells, IK, S was found to be composed of two components with differing voltage dependencies of activation. The FGFs brought about an alteration in the amount of total IK,S by equal effects on each component. Conversely, herbimycin A increased the proportion of low voltage-activated current without affecting total current amplitude. Therefore, we suggest that a single species of channel whose voltage dependence is shifted by tyrosine phosphorylation generates IK,S. 5. In summary, FGF-1 and FGF-2 exert opposite effects on voltage-dependent K+ currents in embryonic myocytes and, furthermore, FGF-1 achieves its effects on different K+ currents via separate second messenger pathways.

  1. Ca2+ signals induced from calcium stores in pancreatic islet β cells

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    In single rat pancreatic β cells,using fura-2 microfluorometry to measure [Ca2+]i response upon different stimuli,the ways of calcium regulation have been studied.When the extracellular calcium concentration was 2.5 mmol/L,either 60 mmol/L KCl,20 mmol/L D-glucose or 0.1 mmol/L tolbutamide induced increase in [Ca2+]i.Such increase in [Ca2+]i was absent when the same stimuli were applied under zero extracellular calcium.These results indicate that the increase of [Ca2+]i is induced by the activation of voltage-dependent calcium channels in β cells.The manifold forms of [Ca2+]i change induced by glucose imply that the effects of glucose are complex.5 mmol/L caffeine or 5 mmol/L MCh increase the [Ca2+]i ,which is independent of the external calcium,suggesting that [Ca2+]i can be regulated by Ca2+ release from not only the IP3-sensitive but also the ryanodine sensitive calcium stores in β cells.The latency of Ca responses for IP3 pathway (5 s) is faster than that for ryanodine pathway (30 s).It is concluded that there are multiple calcium stores in rat pancreatic β cells.

  2. Endogenous and exogenous hydrogen sulfide facilitates T-type calcium channel currents in Cav3.2-expressing HEK293 cells.

    Science.gov (United States)

    Sekiguchi, Fumiko; Miyamoto, Yosuke; Kanaoka, Daiki; Ide, Hiroki; Yoshida, Shigeru; Ohkubo, Tsuyako; Kawabata, Atsufumi

    2014-02-28

    Hydrogen sulfide (H2S), a gasotransmitter, is formed from l-cysteine by multiple enzymes including cystathionine-γ-lyase (CSE). We have shown that an H2S donor, NaHS, causes hyperalgesia in rodents, an effect inhibited by knockdown of Cav3.2 T-type Ca(2+) channels (T-channels), and that NaHS facilitates T-channel-dependent currents (T-currents) in NG108-15 cells that naturally express Cav3.2. In the present study, we asked if endogenous and exogenous H2S participates in regulation of the channel functions in Cav3.2-transfected HEK293 (Cav3.2-HEK293) cells. dl-Propargylglycine (PPG), a CSE inhibitor, significantly decreased T-currents in Cav3.2-HEK293 cells, but not in NG108-15 cells. NaHS at 1.5mM did not affect T-currents in Cav3.2-HEK293 cells, but enhanced T-currents in NG108-15 cells. In the presence of PPG, NaHS at 1.5mM, but not 0.1-0.3mM, increased T-currents in Cav3.2-HEK293 cells. Similarly, Na2S, another H2S donor, at 0.1-0.3mM significantly increased T-currents in the presence, but not absence, of PPG in Cav3.2-HEK293 cells. Expression of CSE was detected at protein and mRNA levels in HEK293 cells. Intraplantar administration of Na2S, like NaHS, caused mechanical hyperalgesia, an effect blocked by NNC 55-0396, a T-channel inhibitor. The in vivo potency of Na2S was higher than NaHS. These results suggest that the function of Cav3.2 T-channels is tonically enhanced by endogenous H2S synthesized by CSE in Cav3.2-HEK293 cells, and that exogenous H2S is capable of enhancing Cav3.2 function when endogenous H2S production by CSE is inhibited. In addition, Na2S is considered a more potent H2S donor than NaHS in vitro as well as in vivo.

  3. Calcium Signals from the Vacuole

    Directory of Open Access Journals (Sweden)

    Gerald Schönknecht

    2013-10-01

    Full Text Available The vacuole is by far the largest intracellular Ca2+ store in most plant cells. Here, the current knowledge about the molecular mechanisms of vacuolar Ca2+ release and Ca2+ uptake is summarized, and how different vacuolar Ca2+ channels and Ca2+ pumps may contribute to Ca2+ signaling in plant cells is discussed. To provide a phylogenetic perspective, the distribution of potential vacuolar Ca2+ transporters is compared for different clades of photosynthetic eukaryotes. There are several candidates for vacuolar Ca2+ channels that could elicit cytosolic [Ca2+] transients. Typical second messengers, such as InsP3 and cADPR, seem to trigger vacuolar Ca2+ release, but the molecular mechanism of this Ca2+ release still awaits elucidation. Some vacuolar Ca2+ channels have been identified on a molecular level, the voltage-dependent SV/TPC1 channel, and recently two cyclic-nucleotide-gated cation channels. However, their function in Ca2+ signaling still has to be demonstrated. Ca2+ pumps in addition to establishing long-term Ca2+ homeostasis can shape cytosolic [Ca2+] transients by limiting their amplitude and duration, and may thus affect Ca2+ signaling.

  4. Dehydroepiandrosterone (DHEA) inhibits voltage-gated T-type calcium channels.

    Science.gov (United States)

    Chevalier, M; Gilbert, G; Lory, P; Marthan, R; Quignard, J F; Savineau, J P

    2012-06-01

    Dehydroepiandrosterone (DHEA) and its sulfated form, DHEAS, are the most abundant steroid hormones in the mammalian blood flow. DHEA may have beneficial effects in various pathophysiological conditions such as cardiovascular diseases or deterioration of the sense of well-being. However to date, the cellular mechanism underlying DHEA action remains elusive and may involve ion channel modulation. In this study, we have characterized the effect of DHEA on T-type voltage-activated calcium channels (T-channels), which are involved in several cardiovascular and neuronal diseases. Using the whole-cell patch-clamp technique, we demonstrate that DHEA inhibits the three recombinant T-channels (Ca(V)3.1, Ca(V)3.2 and Ca(V)3.3) expressed in NG108-15 cell line, as well as native T-channels in pulmonary artery smooth muscle cells. This effect of DHEA is both concentration (IC(50) between 2 and 7μM) and voltage-dependent and results in a significant shift of the steady-state inactivation curves toward hyperpolarized potentials. Consequently, DHEA reduces window T-current and inhibits membrane potential oscillations induced by Ca(V)3 channels. DHEA inhibition is not dependent on the activation of nuclear androgen or estrogen receptors and implicates a PTX-sensitive Gi protein pathway. Functionally, DHEA and the T-type inhibitor NNC 55-0396 inhibited KCl-induced contraction of pulmonary artery rings and their effect was not cumulative. Altogether, the present data demonstrate that DHEA inhibits T-channels by a Gi protein dependent pathway. DHEA-induced alteration in T-channel activity could thus account for its therapeutic action and/or physiological effects. Copyright © 2012 Elsevier Inc. All rights reserved.

  5. Impact of phosphomimetic and non-phosphorylatable mutations of phospholemman on L-type calcium channels gating in HEK 293T cells.

    Science.gov (United States)

    Guo, Kai; Wang, Yue-Peng; Zhou, Zhi-Wen; Jiang, Yi-Bo; Li, Wei; Chen, Xiao-Meng; Li, Yi-Gang

    2015-03-01

    Phospholemman (PLM) is an important phosphorylation substrate for protein kinases A and C in the heart. Until now, the association between PLM phosphorylation status and L-type calcium channels (LTCCs) gating has not been fully understood. We investigated the kinetics of LTCCs in HEK 293T cells expressing phosphomimetic or nonphosphorylatable PLM mutants. The LTCCs gating was measured in HEK 293T cells transfected with LTCC and wild-type (WT) PLM, phosphomimetic or nonphosphorylatable PLM mutants: 6263AA, 6869AA, AAAA, 6263DD, 6869DD or DDDD. WT PLM significantly slowed LTCCs activation and deactivation while enhanced voltage-dependent inactivation (VDI). PLM mutants 6869DD and DDDD significantly increased the peak of the currents. 6263DD accelerated channel activation, while 6263AA slowed it more than WT PLM. 6869DD significantly enhanced PLM-induced increase of VDI. AAAA slowed the channel activation more than 6263AA, and DDDD accelerated the channel VDI more than 6869DD. Our results demonstrate that phosphomimetic PLM could stimulate LTCCs and alter their dynamics, while PLM nonphosphorylatable mutant produced the opposite effects. © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  6. Frequency and voltage dependence of electric and dielectric properties of Au/TiO2/n-4H-SiC (metal-insulator-semiconductor) type Schottky barrier diodes

    Science.gov (United States)

    Tanrıkulu, E. E.; Yıldız, D. E.; Günen, A.; Altındal, Ş.

    2015-09-01

    The main electrical and dielectric properties of Au/TiO2/n-4H-SiC (MIS) type Schottky barrier diodes (SBDs) have been investigated as functions of frequency and applied bias voltage. We believe that the use of high dielectric interfacial layer between metal and semiconductor can improve the performance of Schottky diodes. From the experimental data, both electrical and dielectric parameters were found as strong function of frequency and applied bias voltage. The Fermi energy level (EF), the concentration of doping donor atoms (P), barrier height (ΦB) and series resistance (Rs) values were obtained from reverse and forward bias C-V characteristics. The changes in EF and ND with frequency are considerably low. Therefore, their values were taken at about constant. The real and imaginary parts of dielectric constant (\\varepsilon \\prime , \\varepsilon \\prime\\prime ), tangent loss (tanδ), ac electrical conductivity (σac), and real and imaginary parts of electric modulus (M‧ and M″) values were also obtained from reverse and forward bias C-V and G/ω-V characteristics. In addition, the voltage dependent profiles of all these electrical and dielectric parameters were drawn for each frequency. These results confirmed that both electrical and dielectric properties of Au/TiO2/n-4H-SiC (MIS) type SBD are quite sensitive to both the frequency and applied bias voltage due to surface polarization, density distribution of interface traps (Dit), and interfacial layer.

  7. Types of voltage—dependent calcium channels involved in high potassium depolarization—induced amylase secretion in the exocrine pancreatic tumour cell line AR4—2J

    Institute of Scientific and Technical Information of China (English)

    CUIZONGJIE

    1998-01-01

    In the perifused fura-2 loaded exocrine pancreatic acinar cell line AR4-2J pulses of high potassium induced repetitive increases in intracellular calcium,Attached cells when stimulated with high potassium secreted large amount of amylase.High potassium-induced secretion was dependent both on the concentration of potassium and duration of stimulation.High potassium induced increases in intracellular calcium were inhibited by voltage-dependent calcium channel anatagonists with an order of potency as follows:nifedipine>ω-agatoxin IVA>ω-conotoxin GVIA.In contrast,the L-type calcium channel anatagonist nifedipine almost completely inhibited potassium-induced amylase secretion,whereas the N-type channel antagonist ω-conotoxin GVIA was without effect.The P-type channel antagonist ω-agatoxin IVA had a small inhibitory effect,but this inhibition was not significant at the level of amylase secretion.In conclusion,the AR4-2J cell line posesses different voltage-dependent calcium channels(L,P,N)with the L-type predominantly involved in depolarization induced amylase secretion.

  8. Calcium D-saccharate

    DEFF Research Database (Denmark)

    Garcia, André Castilho; Hedegaard, Martina Vavrusova; Skibsted, Leif Horsfelt

    2016-01-01

    K-1. Equilibria in supersaturated solutions of calcium d-saccharate seem only to adjust slowly, as seen from calcium activity measurements in calcium d-saccharate solutions made supersaturated by cooling. Solutions formed by isothermal dissolution of calcium d-gluconate in aqueous potassium d......-saccharate becomes spontaneously supersaturated with both d-gluconate and d-saccharate calcium salts, from which only calcium d-saccharate slowly precipitates. Calcium d-saccharate is suggested to act as a stabilizer of supersaturated solutions of other calcium hydroxycarboxylates with endothermic complex formation...

  9. Calcium antagonists and vasospasm.

    Science.gov (United States)

    Meyer, F B

    1990-04-01

    A critical review of the clinical data supports the conclusion that nimodipine decreases the severity of neurologic deficits and improves outcome after subarachnoid hemorrhage. The mechanisms by which mortality and morbidity are reduced are still controversial. First, the frequency of vasospasm is not altered (Figs. 5 and 6). Second, the consistent reversal of vasospasm once present has not been demonstrated either angiographically or by noninvasive cerebral blood flow studies. These observations suggest that there is either modification of microcirculatory flow (i.e., dilation of pial conducting vessels or decreased platelet aggregation) or a direct neuronal protective effect. As suggested previously, support for either mechanism is not resolute, and further investigation is necessary. Currently, nimodipine has been the most thoroughly investigated calcium antagonist both from an experimental and clinical perspective. Oral administration has had few reported complications. Therefore, the benefit/risk ratio clearly supports the prophylactic use of this calcium antagonist in patients of all clinical grades after subarachnoid hemorrhage. Evidence also indicates that starting nimodipine after the onset of delayed ischemic deficits is of benefit. Finally, it can be predicted that in the future additional calcium antagonists with more selective vascular or neuronal effects will be developed for use in neurologic disorders.

  10. Calcium affects on vascular endpoints

    Directory of Open Access Journals (Sweden)

    Patel Vaishali B

    2012-03-01

    Full Text Available Abstract Calcium is one of the most abundant minerals in the body and its metabolism is one of the basic biologic processes in humans. Although historically linked primarily to bone structural development and maintenance, calcium is now recognized as a key component of many physiologic pathways necessary for optimum health including cardiovascular, neurological, endocrine, renal, and gastrointestinal systems. A recent meta-analysis published in August 2011 showed a potential increase in cardiovascular events related to calcium supplementation. The possible mechanism of action of this correlation has not been well elucidated. This topic has generated intense interest due to the widespread use of calcium supplements, particularly among the middle aged and elderly who are at the most risk from cardiac events. Prior studies did not control for potential confounding factors such as the use of statins, aspirin or other medications. These controversial results warrant additional well-designed studies to investigate the relationship between calcium supplementation and cardiovascular outcomes. The purpose of this review is to highlight the current literature in regards to calcium supplementation and cardiovascular health; and to identify areas of future research.

  11. Adding calcium improves lithium ferrite core

    Science.gov (United States)

    Lessoff, H.

    1969-01-01

    Adding calcium increases uniformity of grain growth over a wide range of sintering temperatures and reduces porosity within the grain. Ferrite cores containing calcium have square hysteresis loops and high curie temperatures, making them useful in coincident current memories of digital electronic computers.

  12. Blocking mitochondrial calcium release in Schwann cells prevents demyelinating neuropathies.

    Science.gov (United States)

    Gonzalez, Sergio; Berthelot, Jade; Jiner, Jennifer; Perrin-Tricaud, Claire; Fernando, Ruani; Chrast, Roman; Lenaers, Guy; Tricaud, Nicolas

    2016-03-01

    Schwann cells produce myelin sheath around peripheral nerve axons. Myelination is critical for rapid propagation of action potentials, as illustrated by the large number of acquired and hereditary peripheral neuropathies, such as diabetic neuropathy or Charcot-Marie-Tooth diseases, that are commonly associated with a process of demyelination. However, the early molecular events that trigger the demyelination program in these diseases remain unknown. Here, we used virally delivered fluorescent probes and in vivo time-lapse imaging in a mouse model of demyelination to investigate the underlying mechanisms of the demyelination process. We demonstrated that mitochondrial calcium released by voltage-dependent anion channel 1 (VDAC1) after sciatic nerve injury triggers Schwann cell demyelination via ERK1/2, p38, JNK, and c-JUN activation. In diabetic mice, VDAC1 activity was altered, resulting in a mitochondrial calcium leak in Schwann cell cytoplasm, thereby priming the cell for demyelination. Moreover, reduction of mitochondrial calcium release, either by shRNA-mediated VDAC1 silencing or pharmacological inhibition, prevented demyelination, leading to nerve conduction and neuromuscular performance recovery in rodent models of diabetic neuropathy and Charcot-Marie-Tooth diseases. Therefore, this study identifies mitochondria as the early key factor in the molecular mechanism of peripheral demyelination and opens a potential opportunity for the treatment of demyelinating peripheral neuropathies.

  13. Osteoblasts detect pericellular calcium concentration increase via neomycin-sensitive voltage gated calcium channels.

    Science.gov (United States)

    Sun, Xuanhao; Kishore, Vipuil; Fites, Kateri; Akkus, Ozan

    2012-11-01

    The mechanisms underlying the detection of critically loaded or micro-damaged regions of bone by bone cells are still a matter of debate. Our previous studies showed that calcium efflux originates from pre-failure regions of bone matrix and MC3T3-E1 osteoblasts respond to such efflux by an increase in the intracellular calcium concentration. The mechanisms by which the intracellular calcium concentration increases in response to an increase in the pericellular calcium concentration are unknown. Elevation of the intracellular calcium may occur via release from the internal calcium stores of the cell and/or via the membrane bound channels. The current study applied a wide range of pharmaceutical inhibitors to identify the calcium entry pathways involved in the process: internal calcium release from endoplasmic reticulum (ER, inhibited by thapsigargin and TMB-8), calcium receptor (CaSR, inhibited by calhex), stretch-activated calcium channel (SACC, inhibited by gadolinium), voltage-gated calcium channels (VGCC, inhibited by nifedipine, verapamil, neomycin, and ω-conotoxin), and calcium-induced-calcium-release channel (CICRC, inhibited by ryanodine and dantrolene). These inhibitors were screened for their effectiveness to block intracellular calcium increase by using a concentration gradient induced calcium efflux model which mimics calcium diffusion from the basal aspect of cells. The inhibitor(s) which reduced the intracellular calcium response was further tested on osteoblasts seeded on mechanically loaded notched cortical bone wafers undergoing damage. The results showed that only neomycin reduced the intracellular calcium response in osteoblasts, by 27%, upon extracellular calcium stimulus induced by concentration gradient. The inhibitory effect of neomycin was more pronounced (75% reduction in maximum fluorescence) for osteoblasts seeded on notched cortical bone wafers loaded mechanically to damaging load levels. These results imply that the increase in

  14. Persistent current oscillations produced by activation of metabotropic glutamate receptors in immature rat CA3 hippocampal neurons.

    Science.gov (United States)

    Aniksztejn, L; Sciancalepore, M; Ben Ari, Y; Cherubini, E

    1995-04-01

    1. The single-electrode voltage-clamp technique was used to study the effects of the metabotropic glutamate receptors (mGluRs) agonist 1S,3R-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD, ACPD, 3-10 microM) on CA3 hippocampal neurons during the 1st 10 days of postnatal (P) life and in adulthood. 2. Repeated applications of 1S,3R-ACPD, in the presence of tetrodotoxin (TTX, 1 microM), tetraethylammonium chloride (TEACl 10 mM), and CsCl (2 mM), induced in immature but not in adult neurons periodic inward currents (PICs) that persisted for several hours after the last application of the agonist. 3. PICs, which were generated by nonspecific cationic currents, reversed polarity at 2.8 +/- 3 (SD) mV. They were reversibly blocked by kynurenic acid (1 mM), suggesting that they were mediated by glutamate acting on ionotropic receptors. They were also abolished in a nominally Ca(2+)-free medium. 4. PICs were irreversibly abolished by thapsigargin (10 microM) but were unaffected by ryanodine (10-40 microM). Caffeine (2 mM) also reversibly blocked PICs; this effect was independent from adenosine 3',5'-cyclic monophosphate (cAMP) accumulation, inhibition of voltage-dependent Ca2+ current, or blockade of adenosine receptors. 5. We suggest that, in neonatal slices, mGluRs-induced PICs are triggered by elevation of [Ca2+]i, after mobilization of Ca2+ from inositol 1,4,5-trisphosphate (InsP3)-sensitive stores. This will lead to a persistent, pulsatile release of glutamate from presynaptic nerve terminals, a phenomenon that is probably maintained via a calcium-induced-calcium release process.

  15. Adaptation of mammalian auditory hair cell mechanotransduction is independent of calcium entry.

    Science.gov (United States)

    Peng, Anthony W; Effertz, Thomas; Ricci, Anthony J

    2013-11-20

    Adaptation is a hallmark of hair cell mechanotransduction, extending the sensory hair bundle dynamic range while providing mechanical filtering of incoming sound. In hair cells responsive to low frequencies, two distinct adaptation mechanisms exist, a fast component of debatable origin and a slow myosin-based component. It is generally believed that Ca(2+) entry through mechano-electric transducer channels is required for both forms of adaptation. This study investigates the calcium dependence of adaptation in the mammalian auditory system. Recordings from rat cochlear hair cells demonstrate that altering Ca(2+) entry or internal Ca(2+) buffering has little effect on either adaptation kinetics or steady-state adaptation responses. Two additional findings include a voltage-dependent process and an extracellular Ca(2+) binding site, both modulating the resting open probability independent of adaptation. These data suggest that slow motor adaptation is negligible in mammalian auditory cells and that the remaining adaptation process is independent of calcium entry.

  16. Imaging calcium in neurons.

    Science.gov (United States)

    Grienberger, Christine; Konnerth, Arthur

    2012-03-08

    Calcium ions generate versatile intracellular signals that control key functions in all types of neurons. Imaging calcium in neurons is particularly important because calcium signals exert their highly specific functions in well-defined cellular subcompartments. In this Primer, we briefly review the general mechanisms of neuronal calcium signaling. We then introduce the calcium imaging devices, including confocal and two-photon microscopy as well as miniaturized devices that are used in freely moving animals. We provide an overview of the classical chemical fluorescent calcium indicators and of the protein-based genetically encoded calcium indicators. Using application examples, we introduce new developments in the field, such as calcium imaging in awake, behaving animals and the use of calcium imaging for mapping single spine sensory inputs in cortical neurons in vivo. We conclude by providing an outlook on the prospects of calcium imaging for the analysis of neuronal signaling and plasticity in various animal models.

  17. Effect of naloxone on L-type calcium current in isolated rat ventricular myocytes%纳洛酮对大鼠心室肌细胞L型钙电流的影响

    Institute of Scientific and Technical Information of China (English)

    王群超; 闻庆平

    2011-01-01

    Objective To evaluate the effect of naloxone on L-type calcium current (ICa-L) in isolated rat ventricular myocytes.Methods Adult SD rats of both sexes aged 8 weeks weighing 200-250 g were used in this study.Single cardiac ventricular myocytes were enzymatically isolated from SD rats.ICa-L was measured in ventricular myocytes and recorded using whole cell patch-clamp technique.Different concentrations (20 and 100 μg/ml) of naloxone were added to the cardiomyocytes.The effect of naloxone on ICa-L was evaluated.Results The peak current of ICa-L Was inhibited by naloxone in a concentration-dependent manner.Naloxone had no significant effect on steady-state activation curve.Conclusion Naloxone inhibits the L-type calcium channel of ventricular myocytes and exerts negative effect on ventricular muscle function.%目的 评价纳洛酮对大鼠心室肌细胞L型钙电流(ICa-L)的影响.方法 采用急性酶解法分离SD大鼠单个心室肌细胞,以全细胞膜片钳技术测定ICa-L.纳洛酮采用累积给药法,记录不同浓度(20和100μg/ml)纳洛酮作用下心室肌细胞ICa-L和给药前、后半激活电压.结果 20和100 μg/ml的纳洛酮分别使L型钙通道峰电流强度从给药前(341±30) pA降至(270±23) pA和(173±21) pA,与20μg/ml纳洛酮相比,100μg/ml纳洛酮给药后L型钙通道峰电流强度降低(P<0.05).20和100μg/ml纳洛酮给药前、后半激活电压比较差异无统计学意义(P>0.05).结论 纳洛酮可抑制大鼠心室肌细胞L型钙通道,具有心室肌负性肌力作用.

  18. Structure and function of CrACA1, the major PM-type Ca2+-ATPase, expressed at the peak of the gravity-directed trans-cell calcium current in spores of the fern Ceratopteris richardii.

    Science.gov (United States)

    Bushart, T J; Cannon, A; Clark, G; Roux, S J

    2014-01-01

    Spores of the fern Ceratopteris richardii have proven to be a valuable single-cell system for studying gravity responses. The earliest cellular change directed by gravity in these cells is a trans-cell calcium current, which peaks near 10 h after the spores are induced to germinate. This current is needed for gravity-directed axis alignment, and its peak is coincident with the time period when gravity polarises the direction of subsequent nuclear migration and rhizoid growth. Transcriptomic analysis of genes expressed at the 10-h time point revealed several that encode proteins likely to be key components that either drive the current or regulate it. Notable among these is a plasma membrane (PM)-type Ca(2+) ATPase, CrACA1, whose activity pumping Ca(2+) out of cells is regulated by gravity. This report provides an initial characterisation of the structure and expression of this protein, and demonstrates its heterologous function complementing the K616 mutant of yeast, which is deficient in PM-type Ca(2+) pump activity. Gravity-induced changes in the trans-cell Ca(2+) current occur within seconds, a result consistent with the hypothesis that the force of gravity can rapidly alter the post-translational state of the channels and pumps that drive this current across spore cells. This report identifies a transporter likely to be a key driver of the current, CrACA1, and characterises the role of this protein in early germination and gravity-driven polarity fixation through analysis of expression levels, functional complementation and pharmacological treatments. These data, along with newly available transcriptomic data obtained at the 10-h time point, indicate that CrACA1 is present, functional and likely a major contributing component of the trans-cell Ca(2+) efflux. CrACA1 is not necessary for polar axis alignment, but pharmacological perturbations of it disrupt rhizoid development. These data support and help refine the post-translational modification model for

  19. Is Excess Calcium Harmful to Health?

    OpenAIRE

    Daly, Robin M.; Ebeling, Peter R.

    2010-01-01

    Most current guidelines recommend that older adults and the elderly strive for a total calcium intake (diet and supplements) of 1,000 to 1,300 mg/day to prevent osteoporosis and fractures. Traditionally, calcium supplements have been considered safe, effective and well tolerated, but their safety has recently been questioned due to potential adverse effects on vascular disease which may increase mortality. For example, the findings from a meta-analysis of randomized controlled trials (current...

  20. Altered calcium signaling following traumatic brain injury

    Directory of Open Access Journals (Sweden)

    John Thomas Weber

    2012-04-01

    Full Text Available Cell death and dysfunction after traumatic brain injury (TBI is caused by a primary phase, related to direct mechanical disruption of the brain, and a secondary phase which consists of delayed events initiated at the time of the physical insult. Arguably, the calcium ion contributes greatly to the delayed cell damage and death after TBI. A large, sustained influx of calcium into cells can initiate cell death signaling cascades, through activation of several degradative enzymes, such as proteases and endonucleases. However, a sustained level of intracellular free calcium is not necessarily lethal, but the specific route of calcium entry may couple calcium directly to cell death pathways. Other sources of calcium, such as intracellular calcium stores, can also contribute to cell damage. In addition, calcium-mediated signal transduction pathways in neurons may be perturbed following injury. These latter types of alterations may contribute to abnormal physiology in neurons that do not necessarily die after a traumatic episode. This review provides an overview of experimental evidence that has led to our current understanding of the role of calcium signaling in death and dysfunction following TBI.

  1. Myoscape controls cardiac calcium cycling and contractility via regulation of L-type calcium channel surface expression

    OpenAIRE

    Eden, Matthias; Meder, Benjamin; V?lkers, Mirko; Poomvanicha, Montatip; Domes, Katrin; Branchereau, M.; Marck, P.; Will, Rainer; Bernt, Alexander; Rangrez, Ashraf; Busch, Matthias; ,; Adler, Thure; Busch, Dirk H.; Antonio Aguilar-Pimentel, Juan

    2016-01-01

    Calcium signalling plays a critical role in the pathogenesis of heart failure. Here we describe a cardiac protein named Myoscape/FAM40B/STRIP2, which directly interacts with the L-type calcium channel. Knockdown of Myoscape in cardiomyocytes decreases calcium transients associated with smaller Ca2+ amplitudes and a lower diastolic Ca2+ content. Likewise, L-type calcium channel currents are significantly diminished on Myoscape ablation, and downregulation of Myoscape significantly reduces cont...

  2. Rapid responses to reverse T₃ hormone in immature rat Sertoli cells: calcium uptake and exocytosis mediated by integrin.

    Directory of Open Access Journals (Sweden)

    Ana Paula Zanatta

    Full Text Available There is increasing experimental evidence of the nongenomic action of thyroid hormones mediated by receptors located in the plasma membrane or inside cells. The aim of this work was to characterize the reverse T₃ (rT₃ action on calcium uptake and its involvement in immature rat Sertoli cell secretion. The results presented herein show that very low concentrations of rT₃ are able to increase calcium uptake after 1 min of exposure. The implication of T-type voltage-dependent calcium channels and chloride channels in the effect of rT₃ was evidenced using flunarizine and 9-anthracene, respectively. Also, the rT₃-induced calcium uptake was blocked in the presence of the RGD peptide (an inhibitor of integrin-ligand interactions. Therefore, our findings suggest that calcium uptake stimulated by rT₃ may be mediated by integrin αvβ₃. In addition, it was demonstrated that calcium uptake stimulated by rT₃ is PKC and ERK-dependent. Furthermore, the outcomes indicate that rT₃ also stimulates cellular secretion since the cells manifested a loss of fluorescence after 4 min incubation, indicating an exocytic quinacrine release that seems to be mediated by the integrin receptor. These findings indicate that rT₃ modulates the calcium entry and cellular secretion, which might play a role in the regulation of a plethora of intracellular processes involved in male reproductive physiology.

  3. Calcium in ciliated protozoa: sources, regulation, and calcium-regulated cell functions.

    Science.gov (United States)

    Plattner, H; Klauke, N

    2001-01-01

    In ciliates, a variety of processes are regulated by Ca2+, e.g., exocytosis, endocytosis, ciliary beat, cell contraction, and nuclear migration. Differential microdomain regulation may occur by activation of specific channels in different cell regions (e.g., voltage-dependent Ca2+ channels in cilia), by local, nonpropagated activation of subplasmalemmal Ca stores (alveolar sacs), by different sensitivity thresholds, and eventually by interplay with additional second messengers (cilia). During stimulus-secretion coupling, Ca2+ as the only known second messenger operates at approximately 5 microM, whereby mobilization from alveolar sacs is superimposed by "store-operated Ca2+ influx" (SOC), to drive exocytotic and endocytotic membrane fusion. (Content discharge requires binding of extracellular Ca2+ to some secretory proteins.) Ca2+ homeostasis is reestablished by binding to cytosolic Ca2+-binding proteins (e.g., calmodulin), by sequestration into mitochondria (perhaps by Ca2+ uniporter) and into endoplasmic reticulum and alveolar sacs (with a SERCA-type pump), and by extrusion via a plasmalemmal Ca2+ pump and a Na+/Ca2+ exchanger. Comparison of free vs total concentration, [Ca2+] vs [Ca], during activation, using time-resolved fluorochrome analysis and X-ray microanalysis, respectively, reveals that altogether activation requires a calcium flux that is orders of magnitude larger than that expected from the [Ca2+] actually required for local activation.

  4. Calcium channels and migraine.

    Science.gov (United States)

    Pietrobon, Daniela

    2013-07-01

    Missense mutations in CACNA1A, the gene that encodes the pore-forming α1 subunit of human voltage-gated Ca(V)2.1 (P/Q-type) calcium channels, cause a rare form of migraine with aura (familial hemiplegic migraine type 1: FHM1). Migraine is a common disabling brain disorder whose key manifestations are recurrent attacks of unilateral headache that may be preceded by transient neurological aura symptoms. This review, first, briefly summarizes current understanding of the pathophysiological mechanisms that are believed to underlie migraine headache, migraine aura and the onset of a migraine attack, and briefly describes the localization and function of neuronal Ca(V)2.1 channels in the brain regions that have been implicated in migraine pathogenesis. Then, the review describes and discusses i) the functional consequences of FHM1 mutations on the biophysical properties of recombinant human Ca(V)2.1 channels and native Ca(V)2.1 channels in neurons of knockin mouse models carrying the mild R192Q or severe S218L mutations in the orthologous gene, and ii) the functional consequences of these mutations on neurophysiological processes in the cerebral cortex and trigeminovascular system thought to be involved in the pathophysiology of migraine, and the insights into migraine mechanisms obtained from the functional analysis of these processes in FHM1 knockin mice. This article is part of a Special Issue entitled: Calcium channels. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. Integumentary loss of calcium.

    Science.gov (United States)

    Chu, J Y; Margen, S; Calloway, D H; Costa, F M

    1979-08-01

    Integumentary calcium loss was studied in 16 healthy young men. The daily loss by the 16 ambulatory but relatively sedentary young men in 52 determinations of 6-day periods each was 8.7 +/- 1.9 mg/m2 per day (average 15.8 mg/man per day). The amount lost was not influenced by calcium intake (0.1 to 2.3 g/day). In contrast to urinary calcium excretion, which is directly related to protein intake, there was no significant change in integumentary calcium loss with varying protein intakes (1 to 96 g nitrogen per day). No compensatory relationship between urinary and integumentary calcium excretion was noted. During strenuous exercise calcium loss increased to an average of 25 mg in 40 min. There was no compensatory decrease in urinary excretion on the day of strenuous exercise. It was also noted that integumentary calcium loss was not affected by general calcium balance.

  6. Study on intracellular calcium concentration and calcium current of osteoblast in Stat3 knockout mice%Stat3基因敲除小鼠成骨细胞内钙和钙通道电流变化特性的研究

    Institute of Scientific and Technical Information of China (English)

    李磊; 王俊成; 刘颖新; 任晖; 李春龙

    2015-01-01

    目的:研究Stat3基因敲除小鼠成骨细胞内钙和钙通道电流的变化特性。方法应用PCR技术对Stat3基因敲除小鼠进行鉴定;采用二次酶消化法分离、培养小鼠的原代成骨细胞;通过激光扫描共聚焦技术测定细胞内游离钙离子浓度([Ca2+]i);应用全细胞膜片钳技术记录成骨细胞膜钙通道电流( ICa)的变化。结果共聚焦结果显示Stat3基因敲除小鼠与正常小鼠相比,细胞内[ Ca2+] i 明显升高(P<0.01)。膜片钳结果显示,刺激电压为+10mV时,Stat3基因敲除组ICa为(-443.03±49.03)pA,与正常组[(-325.19±38.40)pA]相比,明显增加(P<0.01)。结论 Stat3基因敲除小鼠成骨细胞伴有明显的[ Ca2+] i的异常,其机制可能与细胞膜上钙通道活性改变有关。%Objective To investigate the character of intracellar calcium concentration ( [ Ca2+] ) i and calcium current ( ICa) of osteoblast in Stat3 knockout mice.Methods PCR was used to identify Stat3 knockout mice.The first generation of osteoblast was isolated and purified from the mice by sequential enzyme digestion. [ Ca2+] i was detected by laser scanning confocal microscopy, and the changes of ICa were recorded by whole-cell patch-clamp technique.Results Confocal experiments showed that [ Ca2+] i was markedly increased in Stat3 knockout mice, compared with that in control mice (P<0.01).Patch-clamp studies displayed that at +10mV, the ICa was (-443.03 ±49.03) pA and ( -325.19 ±38.40) pA in Stat3 knockout mice group and control group, respectively.ICa was obvious difference between two groups (P <0.01).Conclusions [Ca2+]i of osteoblast is abnormal in Stat3 knockout mice, and the mechanisms are associated with the changes of electrophysiological activity of calcium channel.

  7. Process for the preparation of calcium superoxide

    Science.gov (United States)

    Ballou, E. V.; Wood, P. C.; Wydeven, T. J.; Spitze, L. A. (Inventor)

    1978-01-01

    Calcium superoxide is prepared in high yields by spreading a quantity of calcium peroxide diperoxyhydrate on the surface of a container, positioning said container in a vacuum chamber on a support structure through which a coolant fluid can be circulated, partially evacuating said vacuum chamber, allowing the temperature of the diperoxyhydrate to reach the range of about 0 to about 40 C; maintaining the temperature selected for a period of time sufficient to complete the disproproriation of the diperoxyhydrate to calcium superoxide, calcium hydroxide, oxygen, and water; constantly and systematically removing the water as it is formed by sweeping the reacting material with a current of dry inert gas and/or by condensation of said water on a cold surface; backfilling the chamber with a dry inert gas; and finally, recovering the calcium superoxide produced.

  8. Application of Calcium Phosphate Materials in Dentistry

    Directory of Open Access Journals (Sweden)

    Jabr S. Al-Sanabani

    2013-01-01

    Full Text Available Calcium phosphate materials are similar to bone in composition and in having bioactive and osteoconductive properties. Calcium phosphate materials in different forms, as cements, composites, and coatings, are used in many medical and dental applications. This paper reviews the applications of these materials in dentistry. It presents a brief history, dental applications, and methods for improving their mechanical properties. Notable research is highlighted regarding (1 application of calcium phosphate into various fields in dentistry; (2 improving mechanical properties of calcium phosphate; (3 biomimetic process and functionally graded materials. This paper deals with most common types of the calcium phosphate materials such as hydroxyapatite and tricalcium phosphate which are currently used in dental and medical fields.

  9. Role of glycine residues highly conserved in the S2-S3 linkers of domains I and II of voltage-gated calcium channel alpha(1) subunits.

    Science.gov (United States)

    Teng, Jinfeng; Iida, Kazuko; Ito, Masanori; Izumi-Nakaseko, Hiroko; Kojima, Itaru; Adachi-Akahane, Satomi; Iida, Hidetoshi

    2010-05-01

    The pore-forming component of voltage-gated calcium channels, alpha(1) subunit, contains four structurally conserved domains (I-IV), each of which contains six transmembrane segments (S1-S6). We have shown previously that a Gly residue in the S2-S3 linker of domain III is completely conserved from yeasts to humans and important for channel activity. The Gly residues in the S2-S3 linkers of domains I and II, which correspond positionally to the Gly in the S2-S3 linker of domain III, are also highly conserved. Here, we investigated the role of the Gly residues in the S2-S3 linkers of domains I and II of Ca(v)1.2. Each of the Gly residues was replaced with Glu or Gln to produce mutant Ca(v)1.2s; G182E, G182Q, G579E, G579Q, and the resulting mutants were transfected into BHK6 cells. Whole-cell patch-clamp recordings showed that current-voltage relationships of the four mutants were the same as those of wild-type Ca(v)1.2. However, G182E and G182Q showed significantly smaller current densities because of mislocalization of the mutant proteins, suggesting that Gly(182) in domain I is involved in the membrane trafficking or surface expression of alpha(1) subunit. On the other hand, G579E showed a slower voltage-dependent current inactivation (VDI) compared to Ca(v)1.2, although G579Q showed a normal VDI, implying that Gly(579) in domain II is involved in the regulation of VDI and that the incorporation of a negative charge alters the VDI kinetics. Our findings indicate that the two conserved Gly residues are important for alpha(1) subunit to become functional.

  10. Changes in intracellular calcium in brain cells of aged rats

    Institute of Scientific and Technical Information of China (English)

    Yu Li; Yunpeng Cao

    2008-01-01

    BACKGROUND: Studies have shown that voltage-dependent calcium influx, and enhancement of certain calcium-dependent processes in neurons, is related to aging. OBJECTIVE: To observe changes in intracellular calcium ([Ca2+]i) in neurons of aged rats, and to compare with young rats. DESIGN, TIME AND SETTING: A randomized control experiment of neurophysiology was performed at the Central Laboratory of School of Pharmaceutical Science, China Medical University from June to August 2004. MATERIALS: Ten male, healthy, Wistar rats, 19 months old, were selected for the aged group. Ten male, 3-month-old, Wistar rats were selected for the young control group. Fura-2/AM was provided by the Institute of Pharmaceutical Research of Chinese Academy of Medical Sciences, and the F-2000 fluorospectrophotometer was a product of Hitachi, Japan. METHODS: Fluorescence Fura-2 spectrophotometer was used to measure [Ca2+]i in acutely dissociated brain cells of aged and young rats. The concentration of extracellular potassium was controlled by adding different volumes of chloridated potassium solution of high concentration. MAIN OUTCOME MEASURES: [Ca2+]i in neurons of young and aged rats in the presence of 1 mmol/L extracellular calcium concentration and 0 mmol/L (resting state), 5, 10, 20, and 40 mmol/L extracellular potassium. Absolute increase of [Ca2+]i in neurons of young and aged rats when extraceUular potassium was 5,10,20, 40 mmol/L. RESULTS: In the presence of 1 mmol/L extracellular Ca2+ and 0 mmol/L (resting state), 5, 10, 20, and 40 mmol/L extracellular potassium, [Ca2+]i in the neurons of aged rats was significantly less than that in young rats (P 0.05). CONCLUSION: The overload of [Ca2+]i in neurons of aged rats is greater than that of young rats under the same circumstances.

  11. Calcium and Mitosis

    Science.gov (United States)

    Hepler, P.

    1983-01-01

    Although the mechanism of calcium regulation is not understood, there is evidence that calcium plays a role in mitosis. Experiments conducted show that: (1) the spindle apparatus contains a highly developed membrane system that has many characteristics of sarcoplasmic reticulum of muscle; (2) this membrane system contains calcium; and (3) there are ionic fluxes occurring during mitosis which can be seen by a variety of fluorescence probes. Whether the process of mitosis can be modulated by experimentally modulating calcium is discussed.

  12. Calcium and Mitosis

    Science.gov (United States)

    Hepler, P.

    1983-01-01

    Although the mechanism of calcium regulation is not understood, there is evidence that calcium plays a role in mitosis. Experiments conducted show that: (1) the spindle apparatus contains a highly developed membrane system that has many characteristics of sarcoplasmic reticulum of muscle; (2) this membrane system contains calcium; and (3) there are ionic fluxes occurring during mitosis which can be seen by a variety of fluorescence probes. Whether the process of mitosis can be modulated by experimentally modulating calcium is discussed.

  13. Connexin 40 and ATP-dependent intercellular calcium wave in renal glomerular endothelial cells.

    Science.gov (United States)

    Toma, Ildikó; Bansal, Eric; Meer, Elliott J; Kang, Jung Julie; Vargas, Sarah L; Peti-Peterdi, János

    2008-06-01

    Endothelial intracellular calcium ([Ca(2+)](i)) plays an important role in the function of the juxtaglomerular vasculature. The present studies aimed to identify the existence and molecular elements of an endothelial calcium wave in cultured glomerular endothelial cells (GENC). GENCs on glass coverslips were loaded with Fluo-4/Fura red, and ratiometric [Ca(2+)](i) imaging was performed using fluorescence confocal microscopy. Mechanical stimulation of a single GENC caused a nine-fold increase in [Ca(2+)](i), which propagated from cell to cell throughout the monolayer (7.9 +/- 0.3 microm/s) in a regenerative manner (without decrement of amplitude, kinetics, and speed) over distances >400 microm. Inhibition of voltage-dependent calcium channels with nifedipine had no effect on the above parameters, but the removal of extracellular calcium reduced Delta[Ca(2+)](i) by 50%. Importantly, the gap junction uncoupler alpha-glycyrrhetinic acid or knockdown of connexin 40 (Cx40) by transfecting GENCs with Cx40 short interfering RNA (siRNA) almost completely eliminated Delta[Ca(2+)](i) and the calcium wave. Breakdown of extracellular ATP using a scavenger cocktail (apyrase and hexokinase) or nonselective inhibition of purinergic P2 receptors with suramin, had similar blocking effects. Scraping cells off along a line eliminated physical contact between cells but did not effect calcium wave propagation. Using an ATP biosensor technique, we detected a significant elevation in extracellular ATP (Delta = 76 +/- 2 microM) during calcium wave propagation, which was abolished by Cx40 siRNA treatment (Delta = 6 +/- 1 microM). These studies suggest that connexin 40 hemichannels and extracellular ATP are key molecular elements of the glomerular endothelial calcium wave, which may serve important juxtaglomerular functions.

  14. Calcium - Function and effects

    NARCIS (Netherlands)

    Liang, Jianfen; He, Yifan; Gao, Qian; Wang, Xuan; Nout, M.J.R.

    2016-01-01

    Rice is the primary food source for more than half of the world population. Levels of calcium contents and inhibitor - phytic acid are summarized in this chapter. Phytic acid has a very strong chelating ability and it is the main inhibit factor for calcium in rice products. Calcium contents in br

  15. Calcium en cardioplegie

    NARCIS (Netherlands)

    Ruigrok, T.J.C.; Meijler, F.L.

    1985-01-01

    Coronary perfusion with a calcium-free solution, followed by reperfusion with a calcium containing solution, may result in acute myocardial cell death and in irreversible loss of the e1ectrical and mechanical activity of the heart. This phenomenon is known as the calcium paradox. A number of cardiop

  16. Calcium signaling in pluripotent stem cells.

    Science.gov (United States)

    Apáti, Ágota; Pászty, Katalin; Erdei, Zsuzsa; Szebényi, Kornélia; Homolya, László; Sarkadi, Balázs

    2012-04-28

    Pluripotent stem cells represent a new source of biological material allowing the exploration of signaling phenomena during normal cell development and differentiation. Still, the calcium signaling pathways and intracellular calcium responses to various ligands or stress conditions have not been sufficiently explored as yet in embryonic or induced pluripotent stem cells and in their differentiated offspring. This is partly due to the special culturing conditions of these cell types, the rapid morphological and functional changes in heterogeneous cell populations during early differentiation, and methodological problems in cellular calcium measurements. In this paper, we review the currently available data in the literature on calcium signaling in pluripotent stem cells and discuss the potential shortcomings of these studies. Various assay methods are surveyed for obtaining reliable data both in undifferentiated embryonic stem cells and in specific, stem cell-derived human tissues. In this paper, we present the modulation of calcium signaling in human embryonic stem cells (hESC) and in their derivates; mesenchymal stem cell like (MSCl) cells and cardiac tissues using the fluorescent calcium indicator Fluo-4 and confocal microscopy. LPA, trypsin and angiotensin II were effective in inducing calcium signals both in HUES9 and MSCl cells. Histamine and thrombin induced calcium signal exclusively in the MSCl cells, while ATP was effective only in HUES9 cells. There was no calcium signal evoked by GABA, even at relatively high concentrations. In stem cell-derived cardiomyocytes a rapid increase in the beating rate and an increase of the calcium signal peaks could be observed after the addition of adrenaline, while verapamil led to a strong decrease in cellular calcium and stopped spontaneous contractions in a relaxed state.

  17. Calcium intake and calcium deficiency in toddlers in a slum population of Bhubaneswar

    Directory of Open Access Journals (Sweden)

    Sonali Kar

    2014-12-01

    Full Text Available Introduction: When considering their children's nutrition, parents often think more about fat grams, carbs, and calories, and forget about calcium, a mineral that is important to help build strong and healthy bones and collagen structures like teeth. The RDI recommendation for 1-3 years is minimum 400mg/day. Calcium is selectively present in milk and milk products besides vegetables like spinach and fruits like orange. The current study was undertaken in the urban field practice area of KIMS that caters to a slum population of nearly 20,000. Aims & Objectives: To assess the knowledge of mothers regarding calcium rich foods and its deficiency and their sociodemographic conditions; to assess the average intake of Calcium using the 7 day recall method; to find the prevalence of possible calcium deficiency in the study population ie 1-3 years of age. Methods: All the mothers with children in the age group 1-3 years were recruited in the study after due informed consent, the final sample being nearly 284. The male female child ratio was 56:44. Mostly women i.e. 83% had some formal education and out of the total nearly 65% had heard never heard of calcium and of those who had heard only 30% could say that milk was the best sour