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Sample records for calcium channel inactivation

  1. Comparison of the calcium release channel of cardiac and skeletal muscle sarcoplasmic reticulum by target inactivation analysis

    International Nuclear Information System (INIS)

    McGrew, S.G.; Inui, Makoto; Chadwick, C.C.; Boucek, R.J. Jr.; Jung, C.Y.; Fleischer, S.

    1989-01-01

    The calcium release channel of sarcoplasmic reticulum which triggers muscle contraction in excitation-contraction coupling has recently been isolated. The channel has been found to be morphologically identical with the feet structures of the junctional face membrane of terminal cisternae and consists of an oligomer of a unique high molecular weight polypeptide. In this study, the authors compare the target size of the calcium release channel from heart and skeletal muscle using target inactivation analysis. The target molecular weights of the calcium release channel estimated by measuring ryanodine binding after irradiation are similar for heart (139,000) and skeletal muscle (143,000) and are smaller than the monomeric unit (estimated to be about 360,000). The target size, estimated by measuring polypeptide remaining after irradiation, was essentially the same for heart and skeletal muscle, 1,061,000 and 1,070,000, respectively, indicating an oligomeric association of protomers. Thus, the calcium release channel of both cardiac and skeletal muscle reacts uniquely with regard to target inactivation analysis in that (1) the size by ryanodine binding is smaller than the monomeric unit and (2) a single hit leads to destruction of more than one polypeptide, by measuring polypeptide remaining. The target inactivation analysis studies indicate that heart and skeletal muscle receptors are structurally very similar

  2. Disruption of the IS6-AID linker affects voltage-gated calcium channel inactivation and facilitation.

    Science.gov (United States)

    Findeisen, Felix; Minor, Daniel L

    2009-03-01

    Two processes dominate voltage-gated calcium channel (Ca(V)) inactivation: voltage-dependent inactivation (VDI) and calcium-dependent inactivation (CDI). The Ca(V)beta/Ca(V)alpha(1)-I-II loop and Ca(2+)/calmodulin (CaM)/Ca(V)alpha(1)-C-terminal tail complexes have been shown to modulate each, respectively. Nevertheless, how each complex couples to the pore and whether each affects inactivation independently have remained unresolved. Here, we demonstrate that the IS6-alpha-interaction domain (AID) linker provides a rigid connection between the pore and Ca(V)beta/I-II loop complex by showing that IS6-AID linker polyglycine mutations accelerate Ca(V)1.2 (L-type) and Ca(V)2.1 (P/Q-type) VDI. Remarkably, mutations that either break the rigid IS6-AID linker connection or disrupt Ca(V)beta/I-II association sharply decelerate CDI and reduce a second Ca(2+)/CaM/Ca(V)alpha(1)-C-terminal-mediated process known as calcium-dependent facilitation. Collectively, the data strongly suggest that components traditionally associated solely with VDI, Ca(V)beta and the IS6-AID linker, are essential for calcium-dependent modulation, and that both Ca(V)beta-dependent and CaM-dependent components couple to the pore by a common mechanism requiring Ca(V)beta and an intact IS6-AID linker.

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

    Directory of Open Access Journals (Sweden)

    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.

  4. Calcium release-dependent inactivation precedes formation of the tubular system in developing rat cardiac myocytes.

    Science.gov (United States)

    Macková, Katarina; Zahradníková, Alexandra; Hoťka, Matej; Hoffmannová, Barbora; Zahradník, Ivan; Zahradníková, Alexandra

    2017-12-01

    Developing cardiac myocytes undergo substantial structural and functional changes transforming the mechanism of excitation-contraction coupling from the embryonic form, based on calcium influx through sarcolemmal DHPR calcium channels, to the adult form, relying on local calcium release through RYR calcium channels of sarcoplasmic reticulum stimulated by calcium influx. We characterized day-by-day the postnatal development of the structure of sarcolemma, using techniques of confocal fluorescence microscopy, and the development of the calcium current, measured by the whole-cell patch-clamp in isolated rat ventricular myocytes. We characterized the appearance and expansion of the t-tubule system and compared it with the appearance and progress of the calcium current inactivation induced by the release of calcium ions from sarcoplasmic reticulum as structural and functional measures of direct DHPR-RYR interaction. The release-dependent inactivation of calcium current preceded the development of the t-tubular system by several days, indicating formation of the first DHPR-RYR couplons at the surface sarcolemma and their later spreading close to contractile myofibrils with the growing t-tubules. Large variability of both of the measured parameters among individual myocytes indicates uneven maturation of myocytes within the growing myocardium.

  5. Inhibition of parathyroid hormone release by maitotoxin, a calcium channel activator

    International Nuclear Information System (INIS)

    Fitzpatrick, L.A.; Yasumoto, T.; Aurbach, G.D.

    1989-01-01

    Maitotoxin, a toxin derived from a marine dinoflagellate, is a potent activator of voltage-sensitive calcium channels. To further test the hypothesis that inhibition of PTH secretion by calcium is mediated via a calcium channel we studied the effect of maitotoxin on dispersed bovine parathyroid cells. Maitotoxin inhibited PTH release in a dose-dependent fashion, and inhibition was maximal at 1 ng/ml. Chelation of extracellular calcium by EGTA blocked the inhibition of PTH by maitotoxin. Maitotoxin enhanced the effects of the dihydropyridine calcium channel agonist (+)202-791 and increased the rate of radiocalcium uptake in parathyroid cells. Pertussis toxin, which ADP-ribosylates and inactivates a guanine nucleotide regulatory protein that interacts with calcium channels in the parathyroid cell, did not affect the inhibition of PTH secretion by maitotoxin. Maitotoxin, by its action on calcium channels allows entry of extracellular calcium and inhibits PTH release. Our results suggest that calcium channels are involved in the release of PTH. Inhibition of PTH release by maitotoxin is not sensitive to pertussis toxin, suggesting that maitotoxin may act distal to the site interacting with a guanine nucleotide regulatory protein, or maitotoxin could interact with other ions or second messengers to inhibit PTH release

  6. 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.

  7. Progress in the structural understanding of voltage-gated calcium channel (CaV) function and modulation.

    Science.gov (United States)

    Minor, Daniel L; Findeisen, Felix

    2010-01-01

    Voltage-gated calcium channels (CaVs) are large, transmembrane multiprotein complexes that couple membrane depolarization to cellular calcium entry. These channels are central to cardiac action potential propagation, neurotransmitter and hormone release, muscle contraction, and calcium-dependent gene transcription. Over the past six years, the advent of high-resolution structural studies of CaV components from different isoforms and CaV modulators has begun to reveal the architecture that underlies the exceptionally rich feedback modulation that controls CaV action. These descriptions of CaV molecular anatomy have provided new, structure-based insights into the mechanisms by which particular channel elements affect voltage-dependent inactivation (VDI), calcium‑dependent inactivation (CDI), and calcium‑dependent facilitation (CDF). The initial successes have been achieved through structural studies of soluble channel domains and modulator proteins and have proven most powerful when paired with biochemical and functional studies that validate ideas inspired by the structures. Here, we review the progress in this growing area and highlight some key open challenges for future efforts.

  8. Voltage-Gated Calcium Channels

    Science.gov (United States)

    Zamponi, Gerald Werner

    Voltage Gated Calcium Channels is the first comprehensive book in the calcium channel field, encompassing over thirty years of progress towards our understanding of calcium channel structure, function, regulation, physiology, pharmacology, and genetics. This book balances contributions from many of the leading authorities in the calcium channel field with fresh perspectives from risings stars in the area, taking into account the most recent literature and concepts. This is the only all-encompassing calcium channel book currently available, and is an essential resource for academic researchers at all levels in the areas neuroscience, biophysics, and cardiovascular sciences, as well as to researchers in the drug discovery area.

  9. Calcium channel blocker poisoning

    Directory of Open Access Journals (Sweden)

    Miran Brvar

    2005-04-01

    Full Text Available Background: Calcium channel blockers act at L-type calcium channels in cardiac and vascular smooth muscles by preventing calcium influx into cells with resultant decrease in vascular tone and cardiac inotropy, chronotropy and dromotropy. Poisoning with calcium channel blockers results in reduced cardiac output, bradycardia, atrioventricular block, hypotension and shock. The findings of hypotension and bradycardia should suggest poisoning with calcium channel blockers.Conclusions: Treatment includes immediate gastric lavage and whole-bowel irrigation in case of ingestion of sustainedrelease products. All patients should receive an activated charcoal orally. Specific treatment includes calcium, glucagone and insulin, which proved especially useful in shocked patients. Supportive care including the use of catecholamines is not always effective. In the setting of failure of pharmacological therapy transvenous pacing, balloon pump and cardiopulmonary by-pass may be necessary.

  10. 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

    S in which loss of function is caused by accelerated inactivation of I(Ca). The proband, a 32 year old male, displayed a Type I ST segment elevation in two right precordial ECG leads following a procainamide challenge. EP study was positive with induction of polymorphic VT/VF. Interrogation of implanted ICD...... significantly faster in mutant channels between 0 and + 20 mV. Action potential voltage clamp experiments showed that total charge was reduced by almost half compared to WT. We report the first BrS mutation in CaCNB2b resulting in accelerated inactivation of L-type calcium channel current. Our results suggest...

  11. Calcium channel blocker overdose

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/002580.htm Calcium-channel blocker overdose To use the sharing features on this page, please enable JavaScript. Calcium-channel blockers are a type of medicine used ...

  12. A Crash Course in Calcium Channels.

    Science.gov (United States)

    Zamponi, Gerald W

    2017-12-20

    Much progress has been made in understanding the molecular physiology and pharmacology of calcium channels. Recently, there have been tremendous advances in learning about calcium channel structure and function through crystallography and cryo-electron microscopy studies. Here, I will give an overview of our knowledge about calcium channels, and highlight two recent studies that give important insights into calcium channel structure.

  13. Apo states of calmodulin and CaBP1 control CaV1 voltage-gated calcium channel function through direct competition for the IQ domain.

    Science.gov (United States)

    Findeisen, Felix; Rumpf, Christine H; Minor, Daniel L

    2013-09-09

    In neurons, binding of calmodulin (CaM) or calcium-binding protein 1 (CaBP1) to the CaV1 (L-type) voltage-gated calcium channel IQ domain endows the channel with diametrically opposed properties. CaM causes calcium-dependent inactivation and limits calcium entry, whereas CaBP1 blocks calcium-dependent inactivation (CDI) and allows sustained calcium influx. Here, we combine isothermal titration calorimetry with cell-based functional measurements and mathematical modeling to show that these calcium sensors behave in a competitive manner that is explained quantitatively by their apo-state binding affinities for the IQ domain. This competition can be completely blocked by covalent tethering of CaM to the channel. Further, we show that Ca(2+)/CaM has a sub-picomolar affinity for the IQ domain that is achieved without drastic alteration of calcium-binding properties. The observation that the apo forms of CaM and CaBP1 compete with each other demonstrates a simple mechanism for direct modulation of CaV1 function and suggests a means by which excitable cells may dynamically tune CaV activity. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. Inactivation as a new regulatory mechanism for neuronal Kv7 channels

    DEFF Research Database (Denmark)

    Jensen, Henrik Sindal; Grunnet, Morten; Olesen, Søren-Peter

    2007-01-01

    neuronal channels and are important for controlling excitability. Kv7.1 channels have been considered the only Kv7 channels to undergo inactivation upon depolarization. However, here we demonstrate that inactivation is also an intrinsic property of Kv7.4 and Kv7.5 channels, which inactivate to a larger...

  15. 6-OHDA induced calcium influx through N-type calcium channel alters membrane properties via PKA pathway in substantia nigra pars compacta dopaminergic neurons.

    Science.gov (United States)

    Qu, Liang; Wang, Yuan; Zhang, Hai-Tao; Li, Nan; Wang, Qiang; Yang, Qian; Gao, Guo-Dong; Wang, Xue-Lian

    2014-07-11

    Voltage gated calcium channels (VGCC) are sensitive to oxidative stress, and their activation or inactivation can impact cell death. Although these channels have been extensively studied in expression systems, their role in the brain, particularly in the substantia nigra pars compacta (SNc), remain controversial. In this study, we assessed 6-hydroxydopamine (6-OHDA) induced transformation of firing pattern and functional changes of calcium channels in SNc dopaminergic neurons. Application of 6-OHDA (0.5-2mM) evoked a dose-dependent, desensitizing inward current and intracellular free calcium concentration ([Ca(2+)]i) rise. In voltage clamp, ω-conotoxin-sensitive Ca(2+) current modulation mediated by 6-OHDA reflected an altered sensitivity. Furthermore, we found that 6-OHDA modulated Ca(2+) currents through PKA pathway. These results provided evidence for the potential role of VGCCs and PKA involved in oxidative stress in degeneration of SNc neurons in Parkinson's disease (PD). Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  16. Apo calmodulin binding to the L-type voltage-gated calcium channel Cav1.2 IQ peptide

    International Nuclear Information System (INIS)

    Lian Luyun; Myatt, Daniel; Kitmitto, Ashraf

    2007-01-01

    The influx of calcium through the L-type voltage-gated calcium channels (LTCCs) is the trigger for the process of calcium-induced calcium release (CICR) from the sarcoplasmic recticulum, an essential step for cardiac contraction. There are two feedback mechanisms that regulate LTCC activity: calcium-dependent inactivation (CDI) and calcium-dependent facilitation (CDF), both of which are mediated by calmodulin (CaM) binding. The IQ domain (aa 1645-1668) housed within the cytoplasmic domain of the LTCC Ca v 1.2 subunit has been shown to bind both calcium-loaded (Ca 2+ CaM ) and calcium-free CaM (apoCaM). Here, we provide new data for the structural basis for the interaction of apoCaM with the IQ peptide using NMR, revealing that the apoCaM C-lobe residues are most significantly perturbed upon complex formation. In addition, we have employed transmission electron microscopy of purified LTCC complexes which shows that both apoCaM and Ca 2+ CaM can bind to the intact channel

  17. Control of Excitation/Inhibition Balance in a Hippocampal Circuit by Calcium Sensor Protein Regulation of Presynaptic Calcium Channels.

    Science.gov (United States)

    Nanou, Evanthia; Lee, Amy; Catterall, William A

    2018-05-02

    Activity-dependent regulation controls the balance of synaptic excitation to inhibition in neural circuits, and disruption of this regulation impairs learning and memory and causes many neurological disorders. The molecular mechanisms underlying short-term synaptic plasticity are incompletely understood, and their role in inhibitory synapses remains uncertain. Here we show that regulation of voltage-gated calcium (Ca 2+ ) channel type 2.1 (Ca V 2.1) by neuronal Ca 2+ sensor (CaS) proteins controls synaptic plasticity and excitation/inhibition balance in a hippocampal circuit. Prevention of CaS protein regulation by introducing the IM-AA mutation in Ca V 2.1 channels in male and female mice impairs short-term synaptic facilitation at excitatory synapses of CA3 pyramidal neurons onto parvalbumin (PV)-expressing basket cells. In sharp contrast, the IM-AA mutation abolishes rapid synaptic depression in the inhibitory synapses of PV basket cells onto CA1 pyramidal neurons. These results show that CaS protein regulation of facilitation and inactivation of Ca V 2.1 channels controls the direction of short-term plasticity at these two synapses. Deletion of the CaS protein CaBP1/caldendrin also blocks rapid depression at PV-CA1 synapses, implicating its upregulation of inactivation of Ca V 2.1 channels in control of short-term synaptic plasticity at this inhibitory synapse. Studies of local-circuit function revealed reduced inhibition of CA1 pyramidal neurons by the disynaptic pathway from CA3 pyramidal cells via PV basket cells and greatly increased excitation/inhibition ratio of the direct excitatory input versus indirect inhibitory input from CA3 pyramidal neurons to CA1 pyramidal neurons. This striking defect in local-circuit function may contribute to the dramatic impairment of spatial learning and memory in IM-AA mice. SIGNIFICANCE STATEMENT Many forms of short-term synaptic plasticity in neuronal circuits rely on regulation of presynaptic voltage-gated Ca 2+ (Ca V

  18. L-Type Calcium Channels Modulation by Estradiol.

    Science.gov (United States)

    Vega-Vela, Nelson E; Osorio, Daniel; Avila-Rodriguez, Marco; Gonzalez, Janneth; García-Segura, Luis Miguel; Echeverria, Valentina; Barreto, George E

    2017-09-01

    Voltage-gated calcium channels are key regulators of brain function, and their dysfunction has been associated with multiple conditions and neurodegenerative diseases because they couple membrane depolarization to the influx of calcium-and other processes such as gene expression-in excitable cells. L-type calcium channels, one of the three major classes and probably the best characterized of the voltage-gated calcium channels, act as an essential calcium binding proteins with a significant biological relevance. It is well known that estradiol can activate rapidly brain signaling pathways and modulatory/regulatory proteins through non-genomic (or non-transcriptional) mechanisms, which lead to an increase of intracellular calcium that activate multiple kinases and signaling cascades, in the same way as L-type calcium channels responses. In this context, estrogens-L-type calcium channels signaling raises intracellular calcium levels and activates the same signaling cascades in the brain probably through estrogen receptor-independent modulatory mechanisms. In this review, we discuss the available literature on this area, which seems to suggest that estradiol exerts dual effects/modulation on these channels in a concentration-dependent manner (as a potentiator of these channels in pM concentrations and as an inhibitor in nM concentrations). Indeed, estradiol may orchestrate multiple neurotrophic responses, which open a new avenue for the development of novel estrogen-based therapies to alleviate different neuropathologies. We also highlight that it is essential to determine through computational and/or experimental approaches the interaction between estradiol and L-type calcium channels to assist these developments, which is an interesting area of research that deserves a closer look in future biomedical research.

  19. Altered Elementary Calcium Release Events and Enhanced Calcium Release by Thymol in Rat Skeletal Muscle

    OpenAIRE

    Szentesi, Péter; Szappanos, Henrietta; Szegedi, Csaba; Gönczi, Monika; Jona, István; Cseri, Julianna; Kovács, László; Csernoch, László

    2004-01-01

    The effects of thymol on steps of excitation-contraction coupling were studied on fast-twitch muscles of rodents. Thymol was found to increase the depolarization-induced release of calcium from the sarcoplasmic reticulum, which could not be attributed to a decreased calcium-dependent inactivation of calcium release channels/ryanodine receptors or altered intramembrane charge movement, but rather to a more efficient coupling of depolarization to channel opening. Thymol increased ryanodine bind...

  20. Calcium channel blockers and Alzheimer's disease★

    Science.gov (United States)

    Tan, Yi; Deng, Yulin; Qing, Hong

    2012-01-01

    Alzheimer's disease is characterized by two pathological hallmarks: amyloid plaques and neurofibrillary 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 involved in Alzheimer's disease therapy. PMID:25767489

  1. Modeling-independent elucidation of inactivation pathways in recombinant and native A-type Kv channels

    Science.gov (United States)

    Fineberg, Jeffrey D.; Ritter, David M.

    2012-01-01

    A-type voltage-gated K+ (Kv) channels self-regulate their activity by inactivating directly from the open state (open-state inactivation [OSI]) or by inactivating before they open (closed-state inactivation [CSI]). To determine the inactivation pathways, it is often necessary to apply several pulse protocols, pore blockers, single-channel recording, and kinetic modeling. However, intrinsic hurdles may preclude the standardized application of these methods. Here, we implemented a simple method inspired by earlier studies of Na+ channels to analyze macroscopic inactivation and conclusively deduce the pathways of inactivation of recombinant and native A-type Kv channels. We investigated two distinct A-type Kv channels expressed heterologously (Kv3.4 and Kv4.2 with accessory subunits) and their native counterparts in dorsal root ganglion and cerebellar granule neurons. This approach applies two conventional pulse protocols to examine inactivation induced by (a) a simple step (single-pulse inactivation) and (b) a conditioning step (double-pulse inactivation). Consistent with OSI, the rate of Kv3.4 inactivation (i.e., the negative first derivative of double-pulse inactivation) precisely superimposes on the profile of the Kv3.4 current evoked by a single pulse because the channels must open to inactivate. In contrast, the rate of Kv4.2 inactivation is asynchronous, already changing at earlier times relative to the profile of the Kv4.2 current evoked by a single pulse. Thus, Kv4.2 inactivation occurs uncoupled from channel opening, indicating CSI. Furthermore, the inactivation time constant versus voltage relation of Kv3.4 decreases monotonically with depolarization and levels off, whereas that of Kv4.2 exhibits a J-shape profile. We also manipulated the inactivation phenotype by changing the subunit composition and show how CSI and CSI combined with OSI might affect spiking properties in a full computational model of the hippocampal CA1 neuron. This work unambiguously

  2. Calcium Channel Blockers

    Science.gov (United States)

    ... Certain calcium channel blockers interact with grapefruit products. Kaplan NM, et al. Treatment of hypertension: Drug therapy. In: Kaplan's Clinical Hypertension. 11th ed. Philadelphia, Pa.: Wolters Kluwer ...

  3. Structural basis for the differential effects of CaBP1 and calmodulin on CaV1.2 calcium-dependent inactivation

    Science.gov (United States)

    Findeisen, Felix; Minor, Daniel L.

    2010-01-01

    Calcium-binding protein 1 (CaBP1), a calmodulin (CaM) homolog, endows certain voltage-gated calcium channels (CaVs) with unusual properties. CaBP1 inhibits CaV1.2 calcium-dependent inactivation (CDI) and introduces calcium-dependent facilitation (CDF). Here, we show that the ability of CaBP1 to inhibit CaV1.2 CDI and induce CDF arises from interaction between the CaBP1 N-lobe and interlobe linker residue Glu94. Unlike CaM, where functional EF hands are essential for channel modulation, CDI inhibition does not require functional CaBP1 EF-hands. Furthermore, CaBP1-mediated CDF has different molecular requirements than CaM-mediated CDF. Overall, the data show that CaBP1 comprises two structural modules having separate functions: similar to CaM, the CaBP1 C-lobe serves as a high-affinity anchor that binds the CaV1.2 IQ domain at a site that overlaps with the Ca2+/CaM C-lobe site, whereas the N-lobe/linker module houses the elements required for channel modulation. Discovery of this division provides the framework for understanding how CaBP1 regulates CaVs. PMID:21134641

  4. Structural basis for the differential effects of CaBP1 and calmodulin on Ca(V)1.2 calcium-dependent inactivation.

    Science.gov (United States)

    Findeisen, Felix; Minor, Daniel L

    2010-12-08

    Calcium-binding protein 1 (CaBP1), a calmodulin (CaM) homolog, endows certain voltage-gated calcium channels (Ca(V)s) with unusual properties. CaBP1 inhibits Ca(V)1.2 calcium-dependent inactivation (CDI) and introduces calcium-dependent facilitation (CDF). Here, we show that the ability of CaBP1 to inhibit Ca(V)1.2 CDI and induce CDF arises from interaction between the CaBP1 N-lobe and interlobe linker residue Glu94. Unlike CaM, where functional EF hands are essential for channel modulation, CDI inhibition does not require functional CaBP1 EF hands. Furthermore, CaBP1-mediated CDF has different molecular requirements than CaM-mediated CDF. Overall, the data show that CaBP1 comprises two structural modules having separate functions: similar to CaM, the CaBP1 C-lobe serves as a high-affinity anchor that binds the Ca(V)1.2 IQ domain at a site that overlaps with the Ca²+/CaM C-lobe site, whereas the N-lobe/linker module houses the elements required for channel modulation. Discovery of this division provides the framework for understanding how CaBP1 regulates Ca(V)s. Copyright © 2010 Elsevier Ltd. All rights reserved.

  5. Mechanism of Cd2+-coordination during Slow Inactivation in Potassium Channels

    Science.gov (United States)

    Raghuraman, H.; Cordero-Morales, Julio F.; Jogini, Vishwanath; Pan, Albert C.; Kollewe, Astrid; Roux, Benoît; Perozo, Eduardo

    2013-01-01

    Summary In K+ channels, rearrangements of the pore outer-vestibule have been associated with C-type inactivation gating. Paradoxically, the crystal structure of Open/C-type inactivated KcsA suggest these movements to be modest in magnitude. Here, we show that under physiological conditions, the KcsA outer-vestibule undergoes relatively large dynamic rearrangements upon inactivation. External Cd2+ enhances the rate of C-type inactivation in an outer-vestibule cysteine mutant (Y82C) via metal-bridge formation. This effect is not present in a non-inactivating mutant (E71A/Y82C). Tandem dimer and tandem tetramer constructs of equivalent cysteine mutants in KcsA and Shaker K+ channels demonstrate that these Cd2+ metal bridges are formed only between adjacent subunits. This is well supported by molecular dynamics simulations. Based on the crystal structure of Cd2+-bound Y82C-KcsA in the closed state, together with EPR distance measurements in the KcsA outer-vestibule, we suggest that subunits must dynamically come in close proximity as the channels undergo inactivation. PMID:22771214

  6. A molecular switch driving inactivation in the cardiac K+ channel HERG.

    Directory of Open Access Journals (Sweden)

    David A Köpfer

    Full Text Available K(+ channels control transmembrane action potentials by gating open or closed in response to external stimuli. Inactivation gating, involving a conformational change at the K(+ selectivity filter, has recently been recognized as a major K(+ channel regulatory mechanism. In the K(+ channel hERG, inactivation controls the length of the human cardiac action potential. Mutations impairing hERG inactivation cause life-threatening cardiac arrhythmia, which also occur as undesired side effects of drugs. In this paper, we report atomistic molecular dynamics simulations, complemented by mutational and electrophysiological studies, which suggest that the selectivity filter adopts a collapsed conformation in the inactivated state of hERG. The selectivity filter is gated by an intricate hydrogen bond network around residues S620 and N629. Mutations of this hydrogen bond network are shown to cause inactivation deficiency in electrophysiological measurements. In addition, drug-related conformational changes around the central cavity and pore helix provide a functional mechanism for newly discovered hERG activators.

  7. 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.

  8. 43. Calmodulin regulating calcium sensitivity of Na channels

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    R. Vegiraju

    2016-07-01

    Full Text Available By extrapolating information from existing research and observing previous assumptions regarding the structure of the Na Channel, this experiment was conducted under the hypothesis that the Na Channel is in part regulated by the calmodulin protein, as a result proving calcium sensitivity of the Na Channel. Furthermore, we assume that there is a one to one stoichiometry between the Na Channel and the Calmodulin. There has been extensive research into the functionality and structure of sodium ion channels (Na channels, as several diseases are associated with the lack of regulation of sodium ions, that is caused by the disfunction of these Na channels. However, one highly controversial matter in the field is the importance of the protein calmodulin (CaM and calcium in Na channel function. Calmodulin is a protein that is well known for its role as a calcium binding messenger protein, and that association is believed to play an indirect role in regulating the Na channel through the Na channel’s supposed calcium sensitivity. While there are proponents for both sides, there has been relatively little research that provides strong evidence for either case. In this experiment, the effect of calmodulin on NaV 1.5 is tested by preparing a set of cardiac cells (of the human specie with the NaV 1.5 C-Termini and CaM protein, which were then to be placed in solutions with varying concentrations of calcium. We took special care to test multiple concentrations of calcium, as previous studies have tested very low concentrations, with Manu Ben-Johny’s team from the John Hopkins laboratory in particular testing up to a meager 50 micromolar, despite producing a well-respected paper (By comparison, the average Na channel can naturally sustain a concentration of almost 1-2 millimolar and on some occasions, reaching even higher concentrations. After using light scattering and observing the signals given off by the calcium interacting with these Nav1.5/Ca

  9. The functions of store-operated calcium channels.

    Science.gov (United States)

    Putney, James W; Steinckwich-Besançon, Natacha; Numaga-Tomita, Takuro; Davis, Felicity M; Desai, Pooja N; D'Agostin, Diane M; Wu, Shilan; Bird, Gary S

    2017-06-01

    Store-operated calcium channels provide calcium signals to the cytoplasm of a wide variety of cell types. The basic components of this signaling mechanism include a mechanism for discharging Ca 2+ stores (commonly but not exclusively phospholipase C and inositol 1,4,5-trisphosphate), a sensor in the endoplasmic reticulum that also serves as an activator of the plasma membrane channel (STIM1 and STIM2), and the store-operated channel (Orai1, 2 or 3). The advent of mice genetically altered to reduce store-operated calcium entry globally or in specific cell types has provided important tools to understand the functions of these widely encountered channels in specific and clinically important physiological systems. This review briefly discusses the history and cellular properties of store-operated calcium channels, and summarizes selected studies of their physiological functions in specific physiological or pathological contexts. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech. Published by Elsevier B.V.

  10. New Role of P/Q-type Voltage-gated Calcium Channels

    DEFF Research Database (Denmark)

    Hansen, Pernille B L

    2015-01-01

    Voltage-gated calcium channels are important for the depolarization-evoked contraction of vascular smooth muscle cells (SMCs), with L-type channels being the classical channel involved in this mechanism. However, it has been demonstrated that the CaV2.1 subunit, which encodes a neuronal isoform...... of the voltage-gated calcium channels (P/Q-type), is also expressed and contributes functionally to contraction of renal blood vessels in both mice and humans. Furthermore, preglomerular vascular SMCs and aortic SMCs coexpress L-, P-, and Q-type calcium channels within the same cell. Calcium channel blockers...... are widely used as pharmacological treatments. However, calcium channel antagonists vary in their selectivity for the various calcium channel subtypes, and the functional contribution from P/Q-type channels as compared with L-type should be considered. Confirming the presence of P/Q-type voltage...

  11. Activation of purified calcium channels by stoichiometric protein phosphorylation

    Energy Technology Data Exchange (ETDEWEB)

    Nunoki, K.; Florio, V.; Catterall, W.A. (Univ. of Washington, Seattle (USA))

    1989-09-01

    Purified dihydropyridine-sensitive calcium channels from rabbit skeletal muscle were reconstituted into phosphatidylcholine vesicles to evaluate the effect of phosphorylation by cyclic AMP-dependent protein kinase (PK-A) on their function. Both the rate and extent of {sup 45}Ca{sup 2+} uptake into vesicles containing reconstituted calcium channels were increased severalfold after incubation with ATP and PK-A. The degree of stimulation of {sup 45}Ca{sup 2+} uptake was linearly proportional to the extent of phosphorylation of the alpha 1 and beta subunits of the calcium channel up to a stoichiometry of approximately 1 mol of phosphate incorporated into each subunit. The calcium channels activated by phosphorylation were determined to be incorporated into the reconstituted vesicles in the inside-out orientation and were completely inhibited by low concentrations of dihydropyridines, phenylalkylamines, Cd{sup 2+}, Ni{sup 2+}, and Mg{sup 2+}. The results demonstrate a direct relationship between PK-A-catalyzed phosphorylation of the alpha 1 and beta subunits of the purified calcium channel and activation of the ion conductance activity of the dihydropyridine-sensitive calcium channels.

  12. Activation of purified calcium channels by stoichiometric protein phosphorylation

    International Nuclear Information System (INIS)

    Nunoki, K.; Florio, V.; Catterall, W.A.

    1989-01-01

    Purified dihydropyridine-sensitive calcium channels from rabbit skeletal muscle were reconstituted into phosphatidylcholine vesicles to evaluate the effect of phosphorylation by cyclic AMP-dependent protein kinase (PK-A) on their function. Both the rate and extent of 45 Ca 2+ uptake into vesicles containing reconstituted calcium channels were increased severalfold after incubation with ATP and PK-A. The degree of stimulation of 45 Ca 2+ uptake was linearly proportional to the extent of phosphorylation of the alpha 1 and beta subunits of the calcium channel up to a stoichiometry of approximately 1 mol of phosphate incorporated into each subunit. The calcium channels activated by phosphorylation were determined to be incorporated into the reconstituted vesicles in the inside-out orientation and were completely inhibited by low concentrations of dihydropyridines, phenylalkylamines, Cd 2+ , Ni 2+ , and Mg 2+ . The results demonstrate a direct relationship between PK-A-catalyzed phosphorylation of the alpha 1 and beta subunits of the purified calcium channel and activation of the ion conductance activity of the dihydropyridine-sensitive calcium channels

  13. Calcium microdomains near R-type calcium channels control the induction of presynaptic LTP at parallel fiber to Purkinje cell synapses

    Science.gov (United States)

    Myoga, Michael H.; Regehr, Wade G.

    2011-01-01

    R-type calcium channels in postsynaptic spines signal through functional calcium microdomains to regulate a calcium-calmodulin sensitive potassium channel that in turn regulates postsynaptic hippocampal LTP. Here we ask whether R-type calcium channels in presynaptic terminals also signal through calcium microdomains to control presynaptic LTP. We focus on presynaptic LTP at parallel fiber to Purkinje cell synapses in the cerebellum (PF-LTP), which is mediated by calcium/calmodulin-stimulated adenylyl cyclases. Although most presynaptic calcium influx is through N-type and P/Q-type calcium channels, blocking these channels does not disrupt PF-LTP, but blocking R-type calcium channels does. Moreover, global calcium signaling cannot account for the calcium dependence of PF-LTP because R-type channels contribute modestly to overall calcium entry. These findings indicate that within presynaptic terminals, R-type calcium channels produce calcium microdomains that evoke presynaptic LTP at moderate frequencies that do not greatly increase global calcium levels,. PMID:21471358

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

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    Erin K Purcell

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

  15. Apo-states of calmodulin and CaBP1 control CaV1 voltage-gated calcium channel function through direct competition for the IQ domain

    Science.gov (United States)

    Findeisen, Felix; Rumpf, Christine; Minor, Daniel L.

    2013-01-01

    In neurons, binding of calmodulin (CaM) or calcium-binding protein 1 (CaBP1) to the CaV1 (L-type) voltage-gated calcium channel IQ domain endows the channel with diametrically opposed properties. CaM causes calcium-dependent inactivation (CDI) and limits calcium entry, whereas CaBP1 blocks CDI and allows sustained calcium influx. Here, we combine isothermal titration calorimetry (ITC) with cell-based functional measurements and mathematical modeling to show that these calcium sensors behave in a competitive manner that is explained quantitatively by their apo-state binding affinities for the IQ domain. This competition can be completely blocked by covalent tethering of CaM to the channel. Further, we show that Ca2+/CaM has a sub-picomolar affinity for the IQ domain that is achieved without drastic alteration of calcium binding properties. The observation that the apo-forms of CaM and CaBP1 compete with each other demonstrates a simple mechanism for direct modulation of CaV1 function and suggests a means by which excitable cells may dynamically tune CaV activity. PMID:23811053

  16. SH Oxidation Stimulates Calcium Release Channels (Ryanodine Receptors From Excitable Cells

    Directory of Open Access Journals (Sweden)

    CECILIA HIDALGO

    2000-01-01

    Full Text Available The effects of redox reagents on the activity of the intracellular calcium release channels (ryanodine receptors of skeletal and cardiac muscle, or brain cortex neurons, was examined. In lipid bilayer experiments, oxidizing agents (2,2'-dithiodipyridine or thimerosal modified the calcium dependence of all single channels studied. After controlled oxidation channels became active at sub µM calcium concentrations and were not inhibited by increasing the calcium concentration to 0.5 mM. Subsequent reduction reversed these effects. Channels purified from amphibian skeletal muscle exhibited the same behavior, indicating that the SH groups responsible for modifying the calcium dependence belong to the channel protein. Parallel experiments that measured calcium release through these channels in sarcoplasmic reticulum vesicles showed that following oxidation, the channels were no longer inhibited by sub mM concentrations of Mg2+. It is proposed that channel redox state controls the high affinity sites responsible for calcium activation as well as the low affinity sites involved in Mg2+ inhibition of channel activity. The possible physiological and pathological implications of these results are discussed

  17. Low voltage-activated calcium channels gate transmitter release at the dorsal root ganglion sandwich synapse.

    Science.gov (United States)

    Rozanski, Gabriela M; Nath, Arup R; Adams, Michael E; Stanley, Elise F

    2013-11-15

    A subpopulation of dorsal root ganglion (DRG) neurons are intimately attached in pairs and separated solely by thin satellite glial cell membrane septa. Stimulation of one neuron leads to transglial activation of its pair by a bi-, purinergic/glutamatergic synaptic pathway, a transmission mechanism that we term sandwich synapse (SS) transmission. Release of ATP from the stimulated neuron can be attributed to a classical mechanism involving Ca(2+) entry via voltage-gated calcium channels (CaV) but via an unknown channel type. Specific blockers and toxins ruled out CaV1, 2.1 and 2.2. Transmission was, however, blocked by a moderate depolarization (-50 mV) or low-concentration Ni(2+) (0.1 mM). Transmission persisted using a voltage pulse to -40 mV from a holding potential of -80 mV, confirming the involvement of a low voltage-activated channel type and limiting the candidate channel type to either CaV3.2 or a subpopulation of inactivation- and Ni(2+)-sensitive CaV2.3 channels. Resistance of the neuron calcium current and SS transmission to SNX482 argue against the latter. Hence, we conclude that inter-somatic transmission at the DRG SS is gated by CaV3.2 type calcium channels. The use of CaV3 family channels to gate transmission has important implications for the biological function of the DRG SS as information transfer would be predicted to occur not only in response to action potentials but also to sub-threshold membrane voltage oscillations. Thus, the SS synapse may serve as a homeostatic signalling mechanism between select neurons in the DRG and could play a role in abnormal sensation such as neuropathic pain.

  18. The calmodulin-binding, short linear motif, NSCaTE is conserved in L-type channel ancestors of vertebrate Cav1.2 and Cav1.3 channels.

    Directory of Open Access Journals (Sweden)

    Valentina Taiakina

    Full Text Available NSCaTE is a short linear motif of (xWxxx(I or Lxxxx, composed of residues with a high helix-forming propensity within a mostly disordered N-terminus that is conserved in L-type calcium channels from protostome invertebrates to humans. NSCaTE is an optional, lower affinity and calcium-sensitive binding site for calmodulin (CaM which competes for CaM binding with a more ancient, C-terminal IQ domain on L-type channels. CaM bound to N- and C- terminal tails serve as dual detectors to changing intracellular Ca(2+ concentrations, promoting calcium-dependent inactivation of L-type calcium channels. NSCaTE is absent in some arthropod species, and is also lacking in vertebrate L-type isoforms, Cav1.1 and Cav1.4 channels. The pervasiveness of a methionine just downstream from NSCaTE suggests that L-type channels could generate alternative N-termini lacking NSCaTE through the choice of translational start sites. Long N-terminus with an NSCaTE motif in L-type calcium channel homolog LCav1 from pond snail Lymnaea stagnalis has a faster calcium-dependent inactivation than a shortened N-termini lacking NSCaTE. NSCaTE effects are present in low concentrations of internal buffer (0.5 mM EGTA, but disappears in high buffer conditions (10 mM EGTA. Snail and mammalian NSCaTE have an alpha-helical propensity upon binding Ca(2+-CaM and can saturate both CaM N-terminal and C-terminal domains in the absence of a competing IQ motif. NSCaTE evolved in ancestors of the first animals with internal organs for promoting a more rapid, calcium-sensitive inactivation of L-type channels.

  19. ALTERNATIVE EQUATIONS FOR DYNAMIC BEHAVIOR OF IONIC CHANNEL ACTIVATION AND INACTIVATION GATES

    Directory of Open Access Journals (Sweden)

    Mahmut ÖZER

    2003-03-01

    Full Text Available In this paper, alternative equations for dynamics of ionic channel activation and inactivation gates are proposed based on the path probability method. Dynamic behavior of a voltage-gated ionic channel is modeled by the conventional Hodgkin-Huxley (H-H mathematical formalism. In that model, conductance of the channel is defined in terms of activation and inactivation gates. Dynamics of the activation and inactivation gates is modeled by first-order differential equations dependent on the gate variable and the membrane potential. In the new approach proposed in this study, dynamic behavior of activation and inactivation gates is modeled by a firstorder differential equation dependent on internal energy and membrane potential by using the path probability method which is widely used in statistical physics. The new model doesn't require the time constant and steadystate values which are used explicitly in the H-H model. The numerical results show validity of the proposed method.

  20. Stac gets the skeletal L-type calcium channel unstuck

    Czech Academy of Sciences Publication Activity Database

    Weiss, Norbert

    2015-01-01

    Roč. 34, č. 2 (2015), s. 101-103 ISSN 0231-5882 Institutional support: RVO:61388963 Keywords : calcium channel * L-type calcium channel * Ca(v)1.1 channel * Stac adaptor protein * excitation- contraction coupling * trafficking Subject RIV: CE - Biochemistry Impact factor: 0.892, year: 2015

  1. Single channel recording of a mitochondrial calcium uniporter.

    Science.gov (United States)

    Wu, Guangyan; Li, Shunjin; Zong, Guangning; Liu, Xiaofen; Fei, Shuang; Shen, Linda; Guan, Xiangchen; Yang, Xue; Shen, Yuequan

    2018-01-29

    Mitochondrial calcium uniporter (MCU) is the pore-forming subunit of the entire uniporter complex and plays an important role in mitochondrial calcium uptake. However, the single channel recording of MCU remains controversial. Here, we expressed and purified different MCU proteins and then reconstituted them into planar lipid bilayers for single channel recording. We showed that MCU alone from Pyronema omphalodes (pMCU) is active with prominent single channel Ca 2+ currents. In sharp contrast, MCU alone from Homo sapiens (hMCU) is inactive. The essential MCU regulator (EMRE) activates hMCU, and therefore, the complex (hMCU-hEMRE) shows prominent single channel Ca 2+ currents. These single channel currents are sensitive to the specific MCU inhibitor Ruthenium Red. Our results clearly demonstrate that active MCU can conduct large amounts of calcium into the mitochondria. Copyright © 2018 Elsevier Inc. All rights reserved.

  2. Interaction of H2S with Calcium Permeable Channels and Transporters

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    Weihua Zhang

    2015-01-01

    Full Text Available A growing amount of evidence has suggested that hydrogen sulfide (H2S, as a gasotransmitter, is involved in intensive physiological and pathological processes. More and more research groups have found that H2S mediates diverse cellular biological functions related to regulating intracellular calcium concentration. These groups have demonstrated the reciprocal interaction between H2S and calcium ion channels and transporters, such as L-type calcium channels (LTCC, T-type calcium channels (TTCC, sodium/calcium exchangers (NCX, transient receptor potential (TRP channels, β-adrenergic receptors, and N-methyl-D-aspartate receptors (NMDAR in different cells. However, the understanding of the molecular targets and mechanisms is incomplete. Recently, some research groups demonstrated that H2S modulates the activity of calcium ion channels through protein S-sulfhydration and polysulfide reactions. In this review, we elucidate that H2S controls intracellular calcium homeostasis and the underlying mechanisms.

  3. Regulation of Na+ channel inactivation by the DIII and DIV voltage-sensing domains.

    Science.gov (United States)

    Hsu, Eric J; Zhu, Wandi; Schubert, Angela R; Voelker, Taylor; Varga, Zoltan; Silva, Jonathan R

    2017-03-06

    Functional eukaryotic voltage-gated Na + (Na V ) channels comprise four domains (DI-DIV), each containing six membrane-spanning segments (S1-S6). Voltage sensing is accomplished by the first four membrane-spanning segments (S1-S4), which together form a voltage-sensing domain (VSD). A critical Na V channel gating process, inactivation, has previously been linked to activation of the VSDs in DIII and DIV. Here, we probe this interaction by using voltage-clamp fluorometry to observe VSD kinetics in the presence of mutations at locations that have been shown to impair Na V channel inactivation. These locations include the DIII-DIV linker, the DIII S4-S5 linker, and the DIV S4-S5 linker. Our results show that, within the 10-ms timeframe of fast inactivation, the DIV-VSD is the primary regulator of inactivation. However, after longer 100-ms pulses, the DIII-DIV linker slows DIII-VSD deactivation, and the rate of DIII deactivation correlates strongly with the rate of recovery from inactivation. Our results imply that, over the course of an action potential, DIV-VSDs regulate the onset of fast inactivation while DIII-VSDs determine its recovery. © 2017 Hsu et al.

  4. Calcium channels in the brain as targets for the calcium-channel modulators used in the treatment of neurological disorders

    NARCIS (Netherlands)

    Peters, Thies; WILFFERT, B; VANHOUTTE, PM; VANZWIETEN, PA

    1991-01-01

    Recent investigations of calcium channels in brain cells by voltage-clamp techniques have revealed that, in spite of electrophysiological similarities, the pharmacological properties of these channels differ considerably from channels in peripheral tissues, e.g., heart and smooth muscle. Therefore,

  5. Discovery and Development of Calcium Channel Blockers

    Directory of Open Access Journals (Sweden)

    Théophile Godfraind

    2017-05-01

    Full Text Available In the mid 1960s, experimental work on molecules under screening as coronary dilators allowed the discovery of the mechanism of calcium entry blockade by drugs later named calcium channel blockers. This paper summarizes scientific research on these small molecules interacting directly with L-type voltage-operated calcium channels. It also reports on experimental approaches translated into understanding of their therapeutic actions. The importance of calcium in muscle contraction was discovered by Sidney Ringer who reported this fact in 1883. Interest in the intracellular role of calcium arose 60 years later out of Kamada (Japan and Heibrunn (USA experiments in the early 1940s. Studies on pharmacology of calcium function were initiated in the mid 1960s and their therapeutic applications globally occurred in the the 1980s. The first part of this report deals with basic pharmacology in the cardiovascular system particularly in isolated arteries. In the section entitled from calcium antagonists to calcium channel blockers, it is recalled that drugs of a series of diphenylpiperazines screened in vivo on coronary bed precontracted by angiotensin were initially named calcium antagonists on the basis of their effect in depolarized arteries contracted by calcium. Studies on arteries contracted by catecholamines showed that the vasorelaxation resulted from blockade of calcium entry. Radiochemical and electrophysiological studies performed with dihydropyridines allowed their cellular targets to be identified with L-type voltage-operated calcium channels. The modulated receptor theory helped the understanding of their variation in affinity dependent on arterial cell membrane potential and promoted the terminology calcium channel blocker (CCB of which the various chemical families are introduced in the paper. In the section entitled tissue selectivity of CCBs, it is shown that characteristics of the drug, properties of the tissue, and of the stimuli are

  6. Bell-shaped calcium-response curves of lns(l,4,5)P3- and calcium-gated channels from endoplasmic reticulum of cerebellum

    Science.gov (United States)

    Bezprozvanny, Llya; Watras, James; Ehrlich, Barbara E.

    1991-06-01

    RELEASE of calcium from intracellular stores occurs by two pathways, an inositol 1,4,5-trisphosphate (InsP3)-gated channel1-3 and a calcium-gated channel (ryanodine receptor)4-6. Using specific antibodies, both receptors were found in Purkinje cells of cerebellum7,8. We have now compared the functional properties of the channels corresponding to the two receptors by incorporating endoplasmic reticulum vesicles from canine cerebellum into planar bilayers. InsP3-gated channels were observed most frequently. Another channel type was activated by adenine nucleotides or caffeine, inhibited by ruthenium red, and modified by ryanodine, characteristics of the ryanodine receptor/channel6. The open probability of both channel types displayed a bell-shaped curve for dependence on calcium. For the InsP3-gated channel, the maximum probability of opening occurred at 0.2 µM free calcium, with sharp decreases on either side of the maximum. Maximum activity for the ryanodine receptor/channel was maintained between 1 and 100 µM calcium. Thus, within the physiological range of cytoplasmic calcium, the InsP3-gated channel itself allows positive feed-back and then negative feedback for calcium release, whereas the ryanodine receptor/channel behaves solely as a calcium-activated channel. The existence in the same cell of two channels with different responses to calcium and different ligand sensitivities provides a basis for complex patterns of intracellular calcium regulation.

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

    DEFF Research Database (Denmark)

    Timmermann, D B; Lund, Trine Meldgaard; Belhage, B

    2001-01-01

    The physiological significance and subcellular distribution of voltage dependent calcium channels was defined using calcium channel blockers to inhibit potassium induced rises in cytosolic calcium concentration in cultured mouse neocortical neurons. The cytosolic calcium concentration was measured...... 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...... most important voltage dependent calcium channel in all parts of the neuron. After treatment with thapsigargin the increase in cytosolic calcium was halved, indicating that calcium release from thapsigargin sensitive intracellular calcium stores is an important component of the potassium induced rise...

  8. A comprehensive search for calcium binding sites critical for TMEM16A calcium-activated chloride channel activity

    Science.gov (United States)

    Tien, Jason; Peters, Christian J; Wong, Xiu Ming; Cheng, Tong; Jan, Yuh Nung; Jan, Lily Yeh; Yang, Huanghe

    2014-01-01

    TMEM16A forms calcium-activated chloride channels (CaCCs) that regulate physiological processes such as the secretions of airway epithelia and exocrine glands, the contraction of smooth muscles, and the excitability of neurons. Notwithstanding intense interest in the mechanism behind TMEM16A-CaCC calcium-dependent gating, comprehensive surveys to identify and characterize potential calcium sensors of this channel are still lacking. By aligning distantly related calcium-activated ion channels in the TMEM16 family and conducting systematic mutagenesis of all conserved acidic residues thought to be exposed to the cytoplasm, we identify four acidic amino acids as putative calcium-binding residues. Alterations of the charge, polarity, and size of amino acid side chains at these sites alter the ability of different divalent cations to activate the channel. Furthermore, TMEM16A mutant channels containing double cysteine substitutions at these residues are sensitive to the redox potential of the internal solution, providing evidence for their physical proximity and solvent accessibility. DOI: http://dx.doi.org/10.7554/eLife.02772.001 PMID:24980701

  9. Domain IV voltage-sensor movement is both sufficient and rate limiting for fast inactivation in sodium channels.

    Science.gov (United States)

    Capes, Deborah L; Goldschen-Ohm, Marcel P; Arcisio-Miranda, Manoel; Bezanilla, Francisco; Chanda, Baron

    2013-08-01

    Voltage-gated sodium channels are critical for the generation and propagation of electrical signals in most excitable cells. Activation of Na(+) channels initiates an action potential, and fast inactivation facilitates repolarization of the membrane by the outward K(+) current. Fast inactivation is also the main determinant of the refractory period between successive electrical impulses. Although the voltage sensor of domain IV (DIV) has been implicated in fast inactivation, it remains unclear whether the activation of DIV alone is sufficient for fast inactivation to occur. Here, we functionally neutralize each specific voltage sensor by mutating several critical arginines in the S4 segment to glutamines. We assess the individual role of each voltage-sensing domain in the voltage dependence and kinetics of fast inactivation upon its specific inhibition. We show that movement of the DIV voltage sensor is the rate-limiting step for both development and recovery from fast inactivation. Our data suggest that activation of the DIV voltage sensor alone is sufficient for fast inactivation to occur, and that activation of DIV before channel opening is the molecular mechanism for closed-state inactivation. We propose a kinetic model of sodium channel gating that can account for our major findings over a wide voltage range by postulating that DIV movement is both necessary and sufficient for fast inactivation.

  10. Large-conductance calcium-dependent potassium channels prevent dendritic excitability in neocortical pyramidal neurons.

    Science.gov (United States)

    Benhassine, Narimane; Berger, Thomas

    2009-03-01

    Large-conductance calcium-dependent potassium channels (BK channels) are homogeneously distributed along the somatodendritic axis of layer 5 pyramidal neurons of the rat somatosensory cortex. The relevance of this conductance for dendritic calcium electrogenesis was studied in acute brain slices using somatodendritic patch clamp recordings and calcium imaging. BK channel activation reduces the occurrence of dendritic calcium spikes. This is reflected in an increased critical frequency of somatic spikes necessary to activate the distal initiation zone. Whilst BK channels repolarise the somatic spike, they dampen it only in the distal dendrite. Their activation reduces dendritic calcium influx via glutamate receptors. Furthermore, they prevent dendritic calcium electrogenesis and subsequent somatic burst discharges. However, the time window for coincident somatic action potential and dendritic input to elicit dendritic calcium events is not influenced by BK channels. Thus, BK channel activation in layer 5 pyramidal neurons affects cellular excitability primarily by establishing a high threshold at the distal action potential initiation zone.

  11. CaV3.1 isoform of T-type calcium channels supports excitability of rat and mouse ventral tegmental area neurons.

    Science.gov (United States)

    Tracy, Matthew E; Tesic, Vesna; Stamenic, Tamara Timic; Joksimovic, Srdjan M; Busquet, Nicolas; Jevtovic-Todorovic, Vesna; Todorovic, Slobodan M

    2018-03-23

    Recent data have implicated voltage-gated calcium channels in the regulation of the excitability of neurons within the mesolimbic reward system. While the attention of most research has centered on high voltage L-type calcium channel activity, the presence and role of the low voltage-gated T-type calcium channel (T-channels) has not been well explored. Hence, we investigated T-channel properties in the neurons of the ventral tegmental area (VTA) utilizing wild-type (WT) rats and mice, Ca V 3.1 knock-out (KO) mice, and TH-eGFP knock-in (KI) rats in acute horizontal brain slices of adolescent animals. In voltage-clamp experiments, we first assessed T-channel activity in WT rats with characteristic properties of voltage-dependent activation and inactivation, as well as characteristic crisscrossing patterns of macroscopic current kinetics. T-current kinetics were similar in WT mice and WT rats but T-currents were abolished in Ca V 3.1 KO mice. In ensuing current-clamp experiments, we observed the presence of hyperpolarization-induced rebound burst firing in a subset of neurons in WT rats, as well as dopaminergic and non-dopaminergic neurons in TH-eGFP KI rats. Following the application of a pan-selective T-channel blocker TTA-P2, rebound bursting was significantly inhibited in all tested cells. In a behavioral assessment, the acute locomotor increase induced by a MK-801 (Dizocilpine) injection in WT mice was abolished in Ca V 3.1 KO mice, suggesting a tangible role for 3.1 T-type channels in drug response. We conclude that pharmacological targeting of Ca V 3.1 isoform of T-channels may be a novel approach for the treatment of disorders of mesolimbic reward system. Copyright © 2018. Published by Elsevier Ltd.

  12. L-Type Calcium Channel Inhibition Contributes to the Proarrhythmic Effects of Aconitine in Human Cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Jianjun Wu

    Full Text Available Aconitine (ACO is well-known for causing lethal ventricular tachyarrhythmias. While cardiac Na+ channel opening during repolarization has long been documented in animal cardiac myocytes, the cellular effects and mechanism of ACO in human remain unexplored. This study aimed to assess the proarrhythmic effects of ACO in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs. ACO concentration-dependently (0.3 ~ 3.0 μM shortened the action potentials (AP durations (APD in ventricular-like hiPSC-CMs by > 40% and induced delayed after-depolarization. Laser-scanning confocal calcium imaging analysis showed that ACO decreased the duration and amplitude of [Ca2+]i transients and increased in the beating frequencies by over 60%. Moreover, ACO was found to markedly reduce the L-type calcium channel (LTCC currents (ICa,L in hiPSC-CMs associated with a positive-shift of activation and a negative shift of inactivation. ACO failed to alter the peak and late Na+ currents (INa in hiPSC-CMs while it drastically increased the late INa in Guinea-pig ventricular myocytes associated with enhanced activation/delayed inactivation of INa at -55 mV~ -85 mV. Further, the effects of ACO on ICa,L, INa and the rapid delayed rectifier potassium current (Ikr were validated in heterologous expression systems by automated voltage-clamping assays and a moderate suppression of Ikr was observed in addition to concentration-dependent ICa,L inhibition. Lastly, increased beating frequency, decreased Ca2+ wave and shortened field potential duration were recorded from hiPSC-CMs by microelectrode arrays assay. In summary, our data demonstrated that LTCC inhibition could play a main role in the proarrhythmic action of ACO in human cardiomyocytes.

  13. [Mechanism of action of neurotoxins acting on the inactivation of voltage-gated sodium channels].

    Science.gov (United States)

    Benoit, E

    1998-01-01

    This review focuses on the mechanism(s) of action of neurotoxins acting on the inactivation of voltage-gated Na channels. Na channels are transmembrane proteins which are fundamental for cellular communication. These proteins form pores in the plasma membrane allowing passive ionic movements to occur. Their opening and closing are controlled by gating systems which depend on both membrane potential and time. Na channels have three functional properties, mainly studied using electrophysiological and biochemical techniques, to ensure their role in the generation and propagation of action potentials: 1) a highly selectivity for Na ions, 2) a rapid opening ("activation"), responsible for the depolarizing phase of the action potential, and 3) a late closing ("inactivation") involved in the repolarizing phase of the action potential. As an essential protein for membrane excitability, the Na channel is the specific target of a number of vegetal and animal toxins which, by binding to the channel, alter its activity by affecting one or more of its properties. At least six toxin receptor sites have been identified on the neuronal Na channel on the basis of binding studies. However, only toxins interacting with four of these sites (sites 2, 3, 5 et 6) produce alterations of channel inactivation. The maximal percentage of Na channels modified by the binding of neurotoxins to sites 2 (batrachotoxin and some alkaloids), 3 (alpha-scorpion and sea anemone toxins), 5 (brevetoxins and ciguatoxins) et 6 (delta-conotoxins) is different according to the site considered. However, in all cases, these channels do not inactivate. Moreover, Na channels modified by toxins which bind to sites 2, 5 and 6 activate at membrane potentials more negative than do unmodified channels. The physiological consequences of Na channel modifications, induced by the binding of neurotoxins to sites 2, 3, 5 and 6, are (i) an inhibition of cellular excitability due to an important membrane depolarization (site

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1979-01-01

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

  15. Aberrant Splicing Induced by Dysregulated Rbfox2 Produces Enhanced Function of CaV1.2 Calcium Channel and Vascular Myogenic Tone in Hypertension.

    Science.gov (United States)

    Zhou, Yingying; Fan, Jia; Zhu, Huayuan; Ji, Li; Fan, Wenyong; Kapoor, Isha; Wang, Yue; Wang, Yuan; Zhu, Guoqing; Wang, Juejin

    2017-12-01

    Calcium influx from activated voltage-gated calcium channel Ca V 1.2 in vascular smooth muscle cells is indispensable for maintaining myogenic tone and blood pressure. The function of Ca V 1.2 channel can be optimized by alternative splicing, one of post-transcriptional modification mechanisms. The splicing factor Rbfox2 is known to regulate the Ca V 1.2 pre-mRNA alternative splicing events during neuronal development. However, Rbfox2's roles in modulating the key function of vascular Ca V 1.2 channel and in the pathogenesis of hypertension remain elusive. Here, we report that the proportion of Ca V 1.2 channels with alternative exon 9* is increased by 10.3%, whereas that with alternative exon 33 is decreased by 10.5% in hypertensive arteries. Surprisingly, the expression level of Rbfox2 is increased ≈3-folds, presumably because of the upregulation of a dominant-negative isoform of Rbfox2. In vascular smooth muscle cells, we find that knockdown of Rbfox2 dynamically increases alternative exon 9*, whereas decreases exon 33 inclusion of Ca V 1.2 channels. By patch-clamp studies, we show that diminished Rbfox2-induced alternative splicing shifts the steady-state activation and inactivation curves of vascular Ca V 1.2 calcium channel to hyperpolarization, which makes the window current potential to more negative. Moreover, siRNA-mediated knockdown of Rbfox2 increases the pressure-induced vascular myogenic tone of rat mesenteric artery. Taken together, our data indicate that Rbfox2 modulates the functions of vascular Ca V 1.2 calcium channel by dynamically regulating the expressions of alternative exons 9* and 33, which in turn affects the vascular myogenic tone. Therefore, our work suggests a key role for Rbfox2 in hypertension, which provides a rational basis for designing antihypertensive therapies. © 2017 American Heart Association, Inc.

  16. 5,6-EET potently inhibits T-type calcium channels

    DEFF Research Database (Denmark)

    Cazade, M.; Bidaud, I.; Hansen, Pernille B. Lærkegaard

    2014-01-01

    T-type calcium channels (T-channels) are important actors in neuronal pacemaking, in heart rhythm, and in the control of the vascular tone. T-channels are regulated by several endogenous lipids including the primary eicosanoid arachidonic acid (AA), which display an important role in vasodilation...

  17. Glycosylation of voltage-gated calcium channels in health and disease

    Czech Academy of Sciences Publication Activity Database

    Lazniewska, Joanna; Weiss, Norbert

    2017-01-01

    Roč. 1859, č. 5 (2017), s. 662-668 ISSN 0005-2736 R&D Projects: GA ČR GA15-13556S; GA MŠk 7AMB15FR015 Institutional support: RVO:61388963 Keywords : calcium channels * voltage-gated calcium channels * N-glycosylation * ancillary subunit * trafficking * stability Subject RIV: CE - Biochemistry OBOR OECD: Biochemistry and molecular biology Impact factor: 3.498, year: 2016

  18. Functional Importance of L- and P/Q-Type Voltage-Gated Calcium Channels in Human Renal Vasculature

    DEFF Research Database (Denmark)

    Hansen, Pernille B; Poulsen, Christian B; Walter, Steen

    2011-01-01

    Calcium channel blockers are widely used for treatment of hypertension, because they decrease peripheral vascular resistance through inhibition of voltage-gated calcium channels. Animal studies of renal vasculature have shown expression of several types of calcium channels that are involved......-type subtype (Ca(v) 3.1 and Ca(v) 3.2) voltage-gated calcium channels (Ca(v)s), and quantitative PCR showed highest expression of L-type channels in renal arteries and variable expression between patients of subtypes of calcium channels in intrarenal vessels. Immunohistochemical labeling of kidney sections...

  19. Binding of [125I]iodipine to parathyroid cell membranes: Evidence of a dihydropyridine-sensitive calcium channel

    International Nuclear Information System (INIS)

    Jones, J.I.; Fitzpatrick, L.A.

    1990-01-01

    The parathyroid cell is unusual, in that an increase in extracellular calcium concentrations inhibits PTH release. Calcium channels are glycoproteins that span cell membranes and allow entry of extracellular calcium into cells. We have demonstrated that the calcium channel agonist (+)202-791, which opens calcium channels, inhibits PTH release and that the antagonist (-)202-791, which closes calcium channels, stimulates PTH release. To identify the calcium channels responsible for these effects, we used a radioligand that specifically binds to calcium channels. Bovine parathyroid cell membranes were prepared and incubated under reduced lighting with [125I] iodipine (SA, 2000 Ci/mmol), which recognizes 1,4-dihydropyridine-sensitive calcium channels. Bound ligand was separated from free ligand by rapid filtration through Whatman GF/B filters. Nonspecific binding was measured by the inclusion of nifedipine at 10 microM. Specific binding represented approximately 40% of the total binding. The optimal temperature for [125I] iodipine binding was 4 C, and binding reached equilibrium by 30 min. The equilibrium dissociation constant (Kd) was approximately 550 pM, and the maximum number of binding sites was 780 fmol/mg protein. Both the calcium channel agonist (+)202-791 and antagonist (-)202-791 competitively inhibited [125I] iodipine binding, with 50% inhibition concentrations of 20 and 300 nM, respectively. These data indicate the presence of dihydropyridine-sensitive calcium channels on parathyroid cell membranes

  20. Calcium channel-dependent molecular maturation of photoreceptor synapses.

    Directory of Open Access Journals (Sweden)

    Nawal Zabouri

    Full Text Available Several studies have shown the importance of calcium channels in the development and/or maturation of synapses. The Ca(V1.4(α(1F knockout mouse is a unique model to study the role of calcium channels in photoreceptor synapse formation. It features abnormal ribbon synapses and aberrant cone morphology. We investigated the expression and targeting of several key elements of ribbon synapses and analyzed the cone morphology in the Ca(V1.4(α(1F knockout retina. Our data demonstrate that most abnormalities occur after eye opening. Indeed, scaffolding proteins such as Bassoon and RIM2 are properly targeted at first, but their expression and localization are not maintained in adulthood. This indicates that either calcium or the Ca(V1.4 channel, or both are necessary for the maintenance of their normal expression and distribution in photoreceptors. Other proteins, such as Veli3 and PSD-95, also display abnormal expression in rods prior to eye opening. Conversely, vesicle related proteins appear normal. Our data demonstrate that the Ca(V1.4 channel is important for maintaining scaffolding proteins in the ribbon synapse but less vital for proteins related to vesicular release. This study also confirms that in adult retinae, cones show developmental features such as sprouting and synaptogenesis. Overall we present evidence that in the absence of the Ca(V1.4 channel, photoreceptor synapses remain immature and are unable to stabilize.

  1. Calcium channel-dependent molecular maturation of photoreceptor synapses.

    Science.gov (United States)

    Zabouri, Nawal; Haverkamp, Silke

    2013-01-01

    Several studies have shown the importance of calcium channels in the development and/or maturation of synapses. The Ca(V)1.4(α(1F)) knockout mouse is a unique model to study the role of calcium channels in photoreceptor synapse formation. It features abnormal ribbon synapses and aberrant cone morphology. We investigated the expression and targeting of several key elements of ribbon synapses and analyzed the cone morphology in the Ca(V)1.4(α(1F)) knockout retina. Our data demonstrate that most abnormalities occur after eye opening. Indeed, scaffolding proteins such as Bassoon and RIM2 are properly targeted at first, but their expression and localization are not maintained in adulthood. This indicates that either calcium or the Ca(V)1.4 channel, or both are necessary for the maintenance of their normal expression and distribution in photoreceptors. Other proteins, such as Veli3 and PSD-95, also display abnormal expression in rods prior to eye opening. Conversely, vesicle related proteins appear normal. Our data demonstrate that the Ca(V)1.4 channel is important for maintaining scaffolding proteins in the ribbon synapse but less vital for proteins related to vesicular release. This study also confirms that in adult retinae, cones show developmental features such as sprouting and synaptogenesis. Overall we present evidence that in the absence of the Ca(V)1.4 channel, photoreceptor synapses remain immature and are unable to stabilize.

  2. Trafficking of neuronal calcium channels

    Czech Academy of Sciences Publication Activity Database

    Weiss, Norbert; Zamponi, G. W.

    2017-01-01

    Roč. 1, č. 1 (2017), č. článku NS20160003. ISSN 2059-6553 R&D Projects: GA ČR GA15-13556S; GA MŠk 7AMB15FR015 Institutional support: RVO:61388963 Keywords : calcium channel * neuron * trafficing Subject RIV: ED - Physiology OBOR OECD: Physiology (including cytology) http://www.neuronalsignaling.org/content/1/1/NS20160003

  3. Eslicarbazepine and the enhancement of slow inactivation of voltage-gated sodium channels: a comparison with carbamazepine, oxcarbazepine and lacosamide.

    Science.gov (United States)

    Hebeisen, Simon; Pires, Nuno; Loureiro, Ana I; Bonifácio, Maria João; Palma, Nuno; Whyment, Andrew; Spanswick, David; Soares-da-Silva, Patrício

    2015-02-01

    This study aimed at evaluating the effects of eslicarbazepine, carbamazepine (CBZ), oxcarbazepine (OXC) and lacosamide (LCM) on the fast and slow inactivated states of voltage-gated sodium channels (VGSC). The anti-epileptiform activity was evaluated in mouse isolated hippocampal slices. The anticonvulsant effects were evaluated in MES and the 6-Hz psychomotor tests. The whole-cell patch-clamp technique was used to investigate the effects of eslicarbazepine, CBZ, OXC and LCM on sodium channels endogenously expressed in N1E-115 mouse neuroblastoma cells. CBZ and eslicarbazepine exhibit similar concentration dependent suppression of epileptiform activity in hippocampal slices. In N1E-115 mouse neuroblastoma cells, at a concentration of 250 μM, the voltage dependence of the fast inactivation was not influenced by eslicarbazepine, whereas LCM, CBZ and OXC shifted the V0.5 value (mV) by -4.8, -12.0 and -16.6, respectively. Eslicarbazepine- and LCM-treated fast-inactivated channels recovered similarly to control conditions, whereas CBZ- and OXC-treated channels required longer pulses to recover. CBZ, eslicarbazepine and LCM shifted the voltage dependence of the slow inactivation (V0.5, mV) by -4.6, -31.2 and -53.3, respectively. For eslicarbazepine, LCM, CBZ and OXC, the affinity to the slow inactivated state was 5.9, 10.4, 1.7 and 1.8 times higher than to the channels in the resting state, respectively. In conclusion, eslicarbazepine did not share with CBZ and OXC the ability to alter fast inactivation of VGSC. Both eslicarbazepine and LCM reduce VGSC availability through enhancement of slow inactivation, but LCM demonstrated higher interaction with VGSC in the resting state and with fast inactivation gating. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. Spatial distribution of calcium-gated chloride channels in olfactory cilia.

    Science.gov (United States)

    French, Donald A; Badamdorj, Dorjsuren; Kleene, Steven J

    2010-12-30

    In vertebrate olfactory receptor neurons, sensory cilia transduce odor stimuli into changes in neuronal membrane potential. The voltage changes are primarily caused by the sequential openings of two types of channel: a cyclic-nucleotide-gated (CNG) cationic channel and a calcium-gated chloride channel. In frog, the cilia are 25 to 200 µm in length, so the spatial distributions of the channels may be an important determinant of odor sensitivity. To determine the spatial distribution of the chloride channels, we recorded from single cilia as calcium was allowed to diffuse down the length of the cilium and activate the channels. A computational model of this experiment allowed an estimate of the spatial distribution of the chloride channels. On average, the channels were concentrated in a narrow band centered at a distance of 29% of the ciliary length, measured from the base of the cilium. This matches the location of the CNG channels determined previously. This non-uniform distribution of transduction proteins is consistent with similar findings in other cilia. On average, the two types of olfactory transduction channel are concentrated in the same region of the cilium. This may contribute to the efficient detection of weak stimuli.

  5. Epithelial calcium channels: from identification to function and regulation.

    NARCIS (Netherlands)

    Hoenderop, J.G.J.; Nilius, B.; Bindels, R.J.M.

    2003-01-01

    The epithelial calcium channels TRPV5 and TRPV6 have been studied extensively in the epithelial tissues controlling Ca(2+) homeostasis and exhibit a range of distinctive properties that distinguish them from other transient receptor potential (TRP) channels. These two novel members of the

  6. Distinct molecular sites of anaesthetic action: pentobarbital block of human brain sodium channels is alleviated by removal of fast inactivation

    NARCIS (Netherlands)

    Wartenberg, H. C.; Urban, B. W.; Duch, D. S.

    1999-01-01

    Fast inactivation of sodium channel function is modified by anaesthetics. Its quantitative contribution to the overall anaesthetic effect is assessed by removing the fast inactivation mechanism enzymatically. Sodium channels from human brain cortex were incorporated into planar lipid bilayers. After

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

    DEFF Research Database (Denmark)

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

    2013-01-01

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

  8. Activation of a cGMP-sensitive calcium-dependent chloride channel may cause transition from calcium waves to whole-cell oscillations in smooth muscle cells

    DEFF Research Database (Denmark)

    Jacobsen, Jens Christian; Aalkjær, Christian; Nilsson, Holger

    2007-01-01

    approximately doubles. In this transition, the simulated results point to a key role for a recently discovered cGMP-sensitive calcium-dependent chloride channel. This channel depolarizes the membrane in response to calcium released from the SR. In turn, depolarization causes uniform opening of L-type calcium...... channels on the cell surface stimulating synchronized release of SR-calcium and inducing the shift from waves to whole-cell oscillations. The effect of the channel is therefore to couple the processes of the SR with those of the membrane. We hypothesize that the shift in oscillatory mode and the associated...

  9. Functional and pharmacological consequences of the distribution of voltage-gated calcium channels in the renal blood vessels

    DEFF Research Database (Denmark)

    Hansen, P B L

    2013-01-01

    Calcium channel blockers are widely used to treat hypertension because they inhibit voltage-gated calcium channels that mediate transmembrane calcium influx in, for example, vascular smooth muscle and cardiomyocytes. The calcium channel family consists of several subfamilies, of which the L......-type is usually associated with vascular contractility. However, the L-, T- and P-/Q-types of calcium channels are present in the renal vasculature and are differentially involved in controlling vascular contractility, thereby contributing to regulation of kidney function and blood pressure. In the preglomerular...... vascular bed, all the three channel families are present. However, the T-type channel is the only channel in cortical efferent arterioles which is in contrast to the juxtamedullary efferent arteriole, and that leads to diverse functional effects of L- and T-type channel inhibition. Furthermore...

  10. Drosophila QVR/SSS modulates the activation and C-type inactivation kinetics of Shaker K+ channels

    Science.gov (United States)

    Dean, Terry; Xu, Rong; Joiner, William; Sehgal, Amita; Hoshi, Toshinori

    2011-01-01

    The quiver/sleepless (qvr/sss) gene encodes a small, glycosylphosphatidylinositol-anchored protein that plays a critical role in the regulation of sleep in Drosophila. Loss-of-function mutations in qvr/sss severely suppress sleep and effect multiple changes in in situ Shaker K+ currents, including decreased magnitude, slower time-to-peak, and cumulative inactivation. Recently, we demonstrated that SLEEPLESS (SSS) protein modulates Shaker channel activity, possibly through a direct interaction at the plasma membrane. We show here that SSS accelerates the activation of heterologously expressed Shaker channels with no effect on deactivation or fast N-type inactivation. Furthermore, this SSS-induced acceleration is sensitive to the pharmacological disruption of lipid rafts and sufficiently accounts for the slower time-to-peak of in situ Shaker currents seen in qvr/sss mutants. We also find that SSS decreases the rate of C-type inactivation of heterologously expressed Shaker channels, providing a potential mechanism for the cumulative inactivation phenotype induced by qvr/sss loss of function mutations. Kinetic modeling based on the in vitro results suggests that the SSS-dependent regulation of channel kinetics accounts for nearly 40% of the decrease in Shaker current magnitude in flies lacking SSS. Sleep duration in qvr/sss null mutants is restored to normal by a qvr/sss transgene that fully rescues the Shaker kinetic phenotypes but only partially rescues the decrease in current magnitude. Together, these results suggest that the role of SSS in the regulation of sleep in Drosophila correlates more strongly with the effects of SSS on Shaker kinetics than current magnitude. PMID:21813698

  11. Drosophila QVR/SSS modulates the activation and C-type inactivation kinetics of Shaker K(+) channels.

    Science.gov (United States)

    Dean, Terry; Xu, Rong; Joiner, William; Sehgal, Amita; Hoshi, Toshinori

    2011-08-03

    The quiver/sleepless (qvr/sss) gene encodes a small, glycosylphosphatidylinositol-anchored protein that plays a critical role in the regulation of sleep in Drosophila. Loss-of-function mutations in qvr/sss severely suppress sleep and effect multiple changes in in situ Shaker K(+) currents, including decreased magnitude, slower time-to-peak, and cumulative inactivation. Recently, we demonstrated that SLEEPLESS (SSS) protein modulates Shaker channel activity, possibly through a direct interaction at the plasma membrane. We show here that SSS accelerates the activation of heterologously expressed Shaker channels with no effect on deactivation or fast N-type inactivation. Furthermore, this SSS-induced acceleration is sensitive to the pharmacological disruption of lipid rafts and sufficiently accounts for the slower time-to-peak of in situ Shaker currents seen in qvr/sss mutants. We also find that SSS decreases the rate of C-type inactivation of heterologously expressed Shaker channels, providing a potential mechanism for the cumulative inactivation phenotype induced by qvr/sss loss-of-function mutations. Kinetic modeling based on the in vitro results suggests that the SSS-dependent regulation of channel kinetics accounts for nearly 40% of the decrease in Shaker current magnitude in flies lacking SSS. Sleep duration in qvr/sss-null mutants is restored to normal by a qvr/sss transgene that fully rescues the Shaker kinetic phenotypes but only partially rescues the decrease in current magnitude. Together, these results suggest that the role of SSS in the regulation of sleep in Drosophila correlates more strongly with the effects of SSS on Shaker kinetics than current magnitude.

  12. Calcium Occupancy of N-terminal Sites within Calmodulin Induces Inhibition of the Ryanodine Receptor Calcium Release Channel

    Energy Technology Data Exchange (ETDEWEB)

    Boschek, Curt B; Jones, Terry E; Squier, Thomas C; Bigelow, Diana J

    2007-08-01

    Calmodulin (CaM) regulates calcium release from intracellular stores in skeletal muscle through its association with the ryanodine receptor (RyR1) calcium release channel, where CaM association enhances channel opening at resting calcium levels and its closing at micromolar calcium levels associated with muscle contraction. A high-affinity CaM-binding sequence (RyRp) has been identified in RyR1, which corresponds to a 30-residue sequence (i.e., K3614 – N3643) located within the central portion of the primary sequence. However, it is currently unclear whether the identified CaM-binding sequence a) senses calcium over the physiological range of calcium-concentrations associated with RyR1 regulation or b) plays a structural role unrelated to the calcium-dependent modulation of RyR1 function. Therefore, we have measured the calcium-dependent activation of the individual domains of CaM in association with RyRp and their relationship to the CaM-dependent regulation of RyR1. These measurements utilize an engineered CaM, permitting the site-specific incorporation of N-(1-pyrene) maleimide at either T34C (PyN-CaM) or T110C (PyC-CaM) in the N- and C-domains, respectively. Consistent with prior measurements, we observe a high-affinity association between both apo- and calcium-activated CaM and RyRp. Upon association with RyRp, fluorescence changes in PyN-CaM or PyC-CaM permit the measurement of the calcium-activation of these individual domains. Fluorescence changes upon calcium-activation of PyC-CaM in association with RyRp are indicative of high-affinity calcium-dependent activation of the C-terminal domain of CaM bound to RyRp at resting calcium levels and the activation of the N-terminal domain at levels of calcium associated cellular activation. In comparison, occupancy of calcium-binding sites in the N-domain of CaM mirrors the calcium-dependence of RyR1 inhibition observed at activating calcium levels, where [Ca]1/2 = 4.3 0.4 μM, suggesting a direct regulation of Ry

  13. Local calcium signalling is mediated by mechanosensitive ion channels in mesenchymal stem cells

    International Nuclear Information System (INIS)

    Chubinskiy-Nadezhdin, Vladislav I.; Vasileva, Valeria Y.; Pugovkina, Natalia A.; Vassilieva, Irina O.; Morachevskaya, Elena A.; Nikolsky, Nikolay N.; Negulyaev, Yuri A.

    2017-01-01

    Mechanical forces are implicated in key physiological processes in stem cells, including proliferation, differentiation and lineage switching. To date, there is an evident lack of understanding of how external mechanical cues are coupled with calcium signalling in stem cells. Mechanical reactions are of particular interest in adult mesenchymal stem cells because of their promising potential for use in tissue remodelling and clinical therapy. Here, single channel patch-clamp technique was employed to search for cation channels involved in mechanosensitivity in mesenchymal endometrial-derived stem cells (hMESCs). Functional expression of native mechanosensitive stretch-activated channels (SACs) and calcium-sensitive potassium channels of different conductances in hMESCs was shown. Single current analysis of stretch-induced channel activity revealed functional coupling of SACs and BK channels in plasma membrane. The combination of cell-attached and inside-out experiments have indicated that highly localized Ca 2+ entry via SACs triggers BK channel activity. At the same time, SK channels are not coupled with SACs despite of high calcium sensitivity as compared to BK. Our data demonstrate novel mechanism controlling BK channel activity in native cells. We conclude that SACs and BK channels are clusterized in functional mechanosensitive domains in the plasma membrane of hMESCs. Co-clustering of ion channels may significantly contribute to mechano-dependent calcium signalling in stem cells. - Highlights: • Stretch-induced channel activity in human mesenchymal stem cells was analyzed. • Functional expression of SACs and Ca 2+ -sensitive BK and SK channels was shown. • Local Ca 2+ influx via stretch-activated channels triggers BK channel activity. • SK channels are not coupled with SACs despite higher sensitivity to [Ca 2+ ] i . • Functional clustering of SACs and BK channels in stem cell membrane is proposed.

  14. Inotropic effect, binding properties, and calcium flux effects of the calcium channel agonist CGP 28392 in intact cultured embryonic chick ventricular cells

    International Nuclear Information System (INIS)

    Laurent, S.; Kim, D.; Smith, T.W.; Marsh, J.D.

    1985-01-01

    CGP 28392 is a recently described dihydropyridine derivative with positive inotropic properties. To study the mechanism of action of this putative calcium channel agonist, we have related the effects of CGP 28392 on contraction (measured with an optical video system) and radioactive calcium uptake to ligand-binding studies in cultured, spontaneously beating chick embryo ventricular cells. CGP 28392 produced a concentration-dependent increase in amplitude and velocity of contraction (EC 50 = 2 x 10(-7) M; maximum contractile effect = 85% of the calcium 3.6 mM response). Nifedipine produced a shift to the right of the concentration-effect curve for CGP 28392 without decreasing the maximum contractile response, suggesting competitive antagonism (pA2 = 8.3). Computer analysis of displacement of [ 3 H]nitrendipine binding to intact heart cells by unlabeled CGP 28392 indicated a K /sub D/ = 2.2 +/- 0.95 x 10(-7) M, in good agreement with the EC 50 for the inotropic effect. CGP 28392 increased the rate of radioactive calcium influx (+39% at 10 seconds) without altering beating rate, while nifedipine decreased radioactive calcium influx and antagonized the CGP 28392-induced increase in calcium influx. Our results indicate that, in intact cultured myocytes, CGP 28392 acts as a calcium channel agonist and competes for the dihydropyridine-binding site of the slow calcium channel. In contrast to calcium channel blockers, CGP 28392 increases calcium influx and enhances the contractile state

  15. Functional and pharmacological consequences of the distribution of voltage-gated calcium channels in the renal blood vessels.

    Science.gov (United States)

    Hansen, P B L

    2013-04-01

    Calcium channel blockers are widely used to treat hypertension because they inhibit voltage-gated calcium channels that mediate transmembrane calcium influx in, for example, vascular smooth muscle and cardiomyocytes. The calcium channel family consists of several subfamilies, of which the L-type is usually associated with vascular contractility. However, the L-, T- and P-/Q-types of calcium channels are present in the renal vasculature and are differentially involved in controlling vascular contractility, thereby contributing to regulation of kidney function and blood pressure. In the preglomerular vascular bed, all the three channel families are present. However, the T-type channel is the only channel in cortical efferent arterioles which is in contrast to the juxtamedullary efferent arteriole, and that leads to diverse functional effects of L- and T-type channel inhibition. Furthermore, by different mechanisms, T-type channels may contribute to both constriction and dilation of the arterioles. Finally, P-/Q-type channels are involved in the regulation of human intrarenal arterial contractility. The calcium blockers used in the clinic affect not only L-type but also P-/Q- and T-type channels. Therefore, the distinct effect obtained by inhibiting a given subtype or set of channels under experimental settings should be considered when choosing a calcium blocker for treatment. T-type channels seem to be crucial for regulating the GFR and the filtration fraction. Use of blockers is expected to lead to preferential efferent vasodilation, reduction of glomerular pressure and proteinuria. Therefore, renovascular T-type channels might provide novel therapeutic targets, and may have superior renoprotective effects compared to conventional calcium blockers. Acta Physiologica © 2013 Scandinavian Physiological Society.

  16. Expression of voltage-activated calcium channels in the early zebrafish embryo.

    Science.gov (United States)

    Sanhueza, Dayán; Montoya, Andro; Sierralta, Jimena; Kukuljan, Manuel

    2009-05-01

    Increases in cytosolic calcium concentrations regulate many cellular processes, including aspects of early development. Calcium release from intracellular stores and calcium entry through non-voltage-gated channels account for signalling in non-excitable cells, whereas voltage-gated calcium channels (CaV) are important in excitable cells. We report the expression of multiple transcripts of CaV, identified by its homology to other species, in the early embryo of the zebrafish, Danio rerio, at stages prior to the differentiation of excitable cells. CaV mRNAs and proteins were detected as early as the 2-cell stages, which indicate that they arise from both maternal and zygotic transcription. Exposure of embryos to pharmacological blockers of CaV does not perturb early development significantly, although late effects are appreciable. These results suggest that CaV may have a role in calcium homeostasis and control of cellular process during early embryonic development.

  17. Intracellular calcium modulation of voltage-gated sodium channels in ventricular myocytes

    NARCIS (Netherlands)

    Casini, Simona; Verkerk, Arie O.; van Borren, Marcel M. G. J.; van Ginneken, Antoni C. G.; Veldkamp, Marieke W.; de Bakker, Jacques M. T.; Tan, Hanno L.

    2009-01-01

    AIMS: Cardiac voltage-gated sodium channels control action potential (AP) upstroke and cell excitability. Intracellular calcium (Ca(i)(2+)) regulates AP properties by modulating various ion channels. Whether Ca(i)(2+) modulates sodium channels in ventricular myocytes, is unresolved. We studied

  18. Single molecule microscopy on Store-Operated Calcium channels

    International Nuclear Information System (INIS)

    Madl, J.

    2011-01-01

    Store-Operated Calcium Entry is essential for many signaling processes in non-excitable cells. The best studied Store-Operated Calcium current is the Calcium-Release-Activated-Calcium (CRAC) current in T-cells and mast cells, with Orai1 representing the essential pore forming subunit. Functional CRAC channels in store-depleted cells are composed of four Orai1 subunits. However, the stoichiometric composition in resting cells is still discussed controversially: both a tetrameric and a dimeric stoichiometry of resting-state Orai1 have been reported for immobilized or immobile Orai1 proteins. The aim of this thesis was to design a more versatile approach that allows reliable determination of the subunit stoichiometry of mobile Orai1 channels. The motive for this approach is that mobile sub-fractions of the entire Orai1 population provide the cleanest pool of data, devoid of contributions e.g. from immobile Orai1 clusters or Orai1-loaded vesicles attached to the plasma membrane. Moreover, resting-state Orai1 is predominantly mobile, and mobility appears critical for the lateral redistribution which occurs upon store depletion. The method per se is based on single molecule fluorescence microscopy and brightness analysis. Orai1 proteins were fused to a monomeric variant of Green Fluorescent Protein (mGFP) and over-expressed in a human cell line (T24). The 1:1 labeling stoichiometry allows using the brightness of individual Orai1-mGFP channels as a direct measure of the pore stoichiometry. Due to over-expression a potential mixing with endogenous Orai1 can be neglected. However, over-expression of Orai1-mGFP results in channel densities that are too high to allow for resolving single channels using diffraction limited optical microscopy. In order to overcome this challenge, I developed an experimental strategy that allows reduction of the density of actively fluorescent Orai1-mGFP channels without altering the labeling stoichiometry. In order to reduce the surface density

  19. Activation of a cGMP-sensitive calcium-dependent chloride channel may cause transition from calcium waves to whole-cell oscillations in smooth muscle cells

    DEFF Research Database (Denmark)

    Jacobsen, Jens Christian; Aalkjær, Christian; Nilsson, Holger

    2007-01-01

    waves sweeping through the cytoplasm when the SR is stimulated to release calcium. A rise in cyclic guanosine monophosphate (cGMP) leads to the experimentally observed transition from waves to whole-cell calcium oscillations. At the same time membrane potential starts to oscillate and the frequency...... approximately doubles. In this transition, the simulated results point to a key role for a recently discovered cGMP-sensitive calcium-dependent chloride channel. This channel depolarizes the membrane in response to calcium released from the SR. In turn, depolarization causes uniform opening of L-type calcium...... onset of oscillations in membrane potential within the individual cell may underlie sudden intercellular synchronization and the appearance of vasomotion. Key words: Vasomotion, Chloride channel, cGMP, Mathematical model, Calcium waves....

  20. Crystal structure of the epithelial calcium channel TRPV6.

    Science.gov (United States)

    Saotome, Kei; Singh, Appu K; Yelshanskaya, Maria V; Sobolevsky, Alexander I

    2016-06-23

    Precise regulation of calcium homeostasis is essential for many physiological functions. The Ca(2+)-selective transient receptor potential (TRP) channels TRPV5 and TRPV6 play vital roles in calcium homeostasis as Ca(2+) uptake channels in epithelial tissues. Detailed structural bases for their assembly and Ca(2+) permeation remain obscure. Here we report the crystal structure of rat TRPV6 at 3.25 Å resolution. The overall architecture of TRPV6 reveals shared and unique features compared with other TRP channels. Intracellular domains engage in extensive interactions to form an intracellular 'skirt' involved in allosteric modulation. In the K(+) channel-like transmembrane domain, Ca(2+) selectivity is determined by direct coordination of Ca(2+) by a ring of aspartate side chains in the selectivity filter. On the basis of crystallographically identified cation-binding sites at the pore axis and extracellular vestibule, we propose a Ca(2+) permeation mechanism. Our results provide a structural foundation for understanding the regulation of epithelial Ca(2+) uptake and its role in pathophysiology.

  1. Altered elementary calcium release events and enhanced calcium release by thymol in rat skeletal muscle.

    Science.gov (United States)

    Szentesi, Péter; Szappanos, Henrietta; Szegedi, Csaba; Gönczi, Monika; Jona, István; Cseri, Julianna; Kovács, László; Csernoch, László

    2004-03-01

    The effects of thymol on steps of excitation-contraction coupling were studied on fast-twitch muscles of rodents. Thymol was found to increase the depolarization-induced release of calcium from the sarcoplasmic reticulum, which could not be attributed to a decreased calcium-dependent inactivation of calcium release channels/ryanodine receptors or altered intramembrane charge movement, but rather to a more efficient coupling of depolarization to channel opening. Thymol increased ryanodine binding to heavy sarcoplasmic reticulum vesicles, with a half-activating concentration of 144 micro M and a Hill coefficient of 1.89, and the open probability of the isolated and reconstituted ryanodine receptors, from 0.09 +/- 0.03 to 0.22 +/- 0.04 at 30 micro M. At higher concentrations the drug induced long-lasting open events on a full conducting state. Elementary calcium release events imaged using laser scanning confocal microscopy in the line-scan mode were reduced in size, 0.92 +/- 0.01 vs. 0.70 +/- 0.01, but increased in duration, 56 +/- 1 vs. 79 +/- 1 ms, by 30 micro M thymol, with an increase in the relative proportion of lone embers. Higher concentrations favored long events, resembling embers in control, with duration often exceeding 500 ms. These findings provide direct experimental evidence that the opening of a single release channel will generate an ember, rather than a spark, in mammalian skeletal muscle.

  2. A Gate Hinge Controls the Epithelial Calcium Channel TRPV5

    OpenAIRE

    van der Wijst, Jenny; Leunissen, Elizabeth H.; Blanchard, Maxime G.; Venselaar, Hanka; Verkaart, Sjoerd; Paulsen, Candice E.; Bindels, Ren? J.; Hoenderop, Joost G.

    2017-01-01

    TRPV5 is unique within the large TRP channel family for displaying a high Ca2+ selectivity together with Ca2+-dependent inactivation. Our study aims to uncover novel insights into channel gating through in-depth structure-function analysis. We identify an exceptional tryptophan (W583) at the terminus of the intracellular pore that is unique for TRPV5 (and TRPV6). A combination of site-directed mutagenesis, biochemical and electrophysiological analysis, together with homology modeling, demonst...

  3. Membrane Incorporation, Channel Formation, and Disruption of Calcium Homeostasis by Alzheimer's β-Amyloid Protein

    Directory of Open Access Journals (Sweden)

    Masahiro Kawahara

    2011-01-01

    Full Text Available Oligomerization, conformational changes, and the consequent neurodegeneration of Alzheimer's β-amyloid protein (AβP play crucial roles in the pathogenesis of Alzheimer's disease (AD. Mounting evidence suggests that oligomeric AβPs cause the disruption of calcium homeostasis, eventually leading to neuronal death. We have demonstrated that oligomeric AβPs directly incorporate into neuronal membranes, form cation-sensitive ion channels (“amyloid channels”, and cause the disruption of calcium homeostasis via the amyloid channels. Other disease-related amyloidogenic proteins, such as prion protein in prion diseases or α-synuclein in dementia with Lewy bodies, exhibit similarities in the incorporation into membranes and the formation of calcium-permeable channels. Here, based on our experimental results and those of numerous other studies, we review the current understanding of the direct binding of AβP into membrane surfaces and the formation of calcium-permeable channels. The implication of composition of membrane lipids and the possible development of new drugs by influencing membrane properties and attenuating amyloid channels for the treatment and prevention of AD is also discussed.

  4. L-Carnitine for the treatment of a calcium channel blocker and metformin poisoning.

    Science.gov (United States)

    St-Onge, Maude; Ajmo, Ian; Poirier, Diane; Laliberté, Martin

    2013-09-01

    The object of the current communication is to discuss the theory and the evidence for the use of L-carnitine in calcium channel blocker and metformin poisonings. A 68-year-old male known for hypertension and type II diabetes was admitted to the critical care unit of a community hospital following an overdose of amlodipine and metformin. The patient was intubated, ventilated, and hemodynamically supported with vasopressors. Despite calcium, glucagon, high-dose insulin (HDI), and lipid emulsion for calcium channel blocker and bicarbonate for metabolic acidosis, the patient remained hemodynamically unstable. The patient was considered too unstable to initiate continuous renal replacement therapy; and without access to extracorporeal life support, the administration of L-carnitine was administered as a last resort. One hour after L-carnitine, the norepinephrine requirements started to decrease, the patient began to improve and was subsequently extubated successfully without apparent sequelae in less than 4 days. L-Carnitine combined with HDI may have helped with the calcium channel blocker (CCB) poisoning by decreasing insulin resistance, promoting intracellular glucose transport, facilitating the metabolism of free fatty acids, and increasing calcium channel sensitivity. It may have also stimulated oxidative utilization of glucose instead of converting pyruvate into lactate and contributed to decrease lactate production with metformin poisoning.

  5. Energetics of discrete selectivity bands and mutation-induced transitions in the calcium-sodium ion channels family.

    Science.gov (United States)

    Kaufman, I; Luchinsky, D G; Tindjong, R; McClintock, P V E; Eisenberg, R S

    2013-11-01

    We use Brownian dynamics (BD) simulations to study the ionic conduction and valence selectivity of a generic electrostatic model of a biological ion channel as functions of the fixed charge Q(f) at its selectivity filter. We are thus able to reconcile the discrete calcium conduction bands recently revealed in our BD simulations, M0 (Q(f)=1e), M1 (3e), M2 (5e), with a set of sodium conduction bands L0 (0.5e), L1 (1.5e), thereby obtaining a completed pattern of conduction and selectivity bands vs Q(f) for the sodium-calcium channels family. An increase of Q(f) leads to an increase of calcium selectivity: L0 (sodium-selective, nonblocking channel) → M0 (nonselective channel) → L1 (sodium-selective channel with divalent block) → M1 (calcium-selective channel exhibiting the anomalous mole fraction effect). We create a consistent identification scheme where the L0 band is putatively identified with the eukaryotic sodium channel The scheme created is able to account for the experimentally observed mutation-induced transformations between nonselective channels, sodium-selective channels, and calcium-selective channels, which we interpret as transitions between different rows of the identification table. By considering the potential energy changes during permeation, we show explicitly that the multi-ion conduction bands of calcium and sodium channels arise as the result of resonant barrierless conduction. The pattern of periodic conduction bands is explained on the basis of sequential neutralization taking account of self-energy, as Q(f)(z,i)=ze(1/2+i), where i is the order of the band and z is the valence of the ion. Our results confirm the crucial influence of electrostatic interactions on conduction and on the Ca(2+)/Na(+) valence selectivity of calcium and sodium ion channels. The model and results could be also applicable to biomimetic nanopores with charged walls.

  6. Calcium channel agonists and antagonists regulate protein phosphorylation in intact synaptosomes

    International Nuclear Information System (INIS)

    Robinson, P.J.; Lovenberg, Walter

    1986-01-01

    Protein phosphorylation in intact synaptosomes is highly sensitive to alterations in calcium fluxes and was used to probe the possible mechanism of action of the calcium channel agonist BAY K 8644 and antagonists verapamil and nifedipine. These agents (at 1μM) all increased the basal phosphorylation of a specific set of 4 synaptosomal phosphoproteins termed P139, P124, P96 and P60, but did not alter depolarization-dependent protein phosphorylation. The increases could not be explained by a direct stimulation of protein kinases and appears unrelated to the known effects of these + drugs on K + -stimulated neuro-transmitter release. This finding may reveal a possible new mechanism of action for drugs which interact with calcium channels. (Author)

  7. Two-photon activation of endogenous store-operated calcium channels without optogenetics

    Science.gov (United States)

    Cheng, Pan; Tang, Wanyi; He, Hao

    2018-02-01

    Store-operated calcium (SOC) channels, regulated by intracellular Ca2+ store, are the essential pathway of calcium signaling and participate in a wide variety of cellular activities such as gene expression, secretion and immune response1. However, our understanding and regulation of SOC channels are mainly based on pharmacological methods. Considering the unique advantages of optical control, optogenetic control of SOC channels has been developed2. However, the process of genetic engineering to express exogenous light-sensitive protein is complicated, which arouses concerns about ethic difficulties in some research of animal and applications in human. In this report, we demonstrate rapid, robust and reproducible two-photon activation of endogenous SOC channels by femtosecond laser without optogenetics. We present that the short-duration two-photon scanning on subcellular microregion induces slow Ca2+ influx from extracellular medium, which can be eliminated by removing extracellular Ca2+. Block of SOC channels using various pharmacological inhibitors or knockdown of SOC channels by RNA interference reduce the probability of two-photon activated Ca2+ influx. On the contrary, overexpression of SOC channels can increase the probability of Ca2+ influx by two-photon scanning. These results collectively indicate Ca2+ influx through two-photon activated SOC channels. Different from classical pathway of SOC entry activated by Ca2+ store depletion, STIM1, the sensor protein of Ca2+ level in endoplasmic reticulum, does not show any aggregation or migration in this two-photon activated Ca2+ influx, which rules out the possibility of intracellular Ca2+ store depletion. Thereby, we propose this all-optical method of two-photon activation of SOC channels is of great potential to be widely applied in the research of cell calcium signaling and related biological research.

  8. TRP channels in calcium homeostasis: from hormonal control to structure-function relationship of TRPV5 and TRPV6.

    Science.gov (United States)

    van Goor, Mark K C; Hoenderop, Joost G J; van der Wijst, Jenny

    2017-06-01

    Maintaining plasma calcium levels within a narrow range is of vital importance for many physiological functions. Therefore, calcium transport processes in the intestine, bone and kidney are tightly regulated to fine-tune the rate of absorption, storage and excretion. The TRPV5 and TRPV6 calcium channels are viewed as the gatekeepers of epithelial calcium transport. Several calciotropic hormones control the channels at the level of transcription, membrane expression, and function. Recent technological advances have provided the first near-atomic resolution structural models of several TRPV channels, allowing insight into their architecture. While this field is still in its infancy, it has increased our understanding of molecular channel regulation and holds great promise for future structure-function studies of these ion channels. This review will summarize the mechanisms that control the systemic calcium balance, as well as extrapolate structural views to the molecular functioning of TRPV5/6 channels in epithelial calcium transport. Copyright © 2016. Published by Elsevier B.V.

  9. Neurotoxicity Induced by Bupivacaine via T-Type Calcium Channels in SH-SY5Y Cells

    Science.gov (United States)

    Wen, Xianjie; Xu, Shiyuan; Liu, Hongzhen; Zhang, Quinguo; Liang, Hua; Yang, Chenxiang; Wang, Hanbing

    2013-01-01

    There is concern regarding neurotoxicity induced by the use of local anesthetics. A previous study showed that an overload of intracellular calcium is involved in the neurotoxic effect of some anesthetics. T-type calcium channels, which lower the threshold of action potentials, can regulate the influx of calcium ions. We hypothesized that T-type calcium channels are involved in bupivacaine-induced neurotoxicity. In this study, we first investigated the effects of different concentrations of bupivacaine on SH-SY5Y cell viability, and established a cell injury model with 1 mM bupivacaine. The cell viability of SH-SY5Y cells was measured following treatment with 1 mM bupivacaine and/or different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride, an antagonist of T-type calcium channels for 24 h. In addition, we monitored the release of lactate dehydrogenase, cytosolic Ca2+ ([Ca2+]i), cell apoptosis and caspase-3 expression. SH-SY5Y cells pretreated with different dosages (10, 50, or 100 µM) of NNC 55-0396 dihydrochloride improved cell viability, reduced lactate dehydrogenase release, inhibited apoptosis, and reduced caspase-3 expression following bupivacaine exposure. However, the protective effect of NNC 55-0396 dihydrochloride plateaued. Overall, our results suggest that T-type calcium channels may be involved in bupivacaine neurotoxicity. However, identification of the specific subtype of T calcium channels involved requires further investigation. PMID:23658789

  10. Neurotoxicity induced by bupivacaine via T-type calcium channels in SH-SY5Y cells.

    Directory of Open Access Journals (Sweden)

    Xianjie Wen

    Full Text Available There is concern regarding neurotoxicity induced by the use of local anesthetics. A previous study showed that an overload of intracellular calcium is involved in the neurotoxic effect of some anesthetics. T-type calcium channels, which lower the threshold of action potentials, can regulate the influx of calcium ions. We hypothesized that T-type calcium channels are involved in bupivacaine-induced neurotoxicity. In this study, we first investigated the effects of different concentrations of bupivacaine on SH-SY5Y cell viability, and established a cell injury model with 1 mM bupivacaine. The cell viability of SH-SY5Y cells was measured following treatment with 1 mM bupivacaine and/or different dosages (10, 50, or 100 µM of NNC 55-0396 dihydrochloride, an antagonist of T-type calcium channels for 24 h. In addition, we monitored the release of lactate dehydrogenase, cytosolic Ca(2+ ([Ca2+]i, cell apoptosis and caspase-3 expression. SH-SY5Y cells pretreated with different dosages (10, 50, or 100 µM of NNC 55-0396 dihydrochloride improved cell viability, reduced lactate dehydrogenase release, inhibited apoptosis, and reduced caspase-3 expression following bupivacaine exposure. However, the protective effect of NNC 55-0396 dihydrochloride plateaued. Overall, our results suggest that T-type calcium channels may be involved in bupivacaine neurotoxicity. However, identification of the specific subtype of T calcium channels involved requires further investigation.

  11. A Common Structural Component for β-Subunit Mediated Modulation of Slow Inactivation in Different KV Channels

    Directory of Open Access Journals (Sweden)

    Nathalie Strutz-Seebohm

    2013-06-01

    Full Text Available Background/Aims: Potassium channels are tetrameric proteins providing potassium selective passage through lipid embedded proteinaceous pores with highest fidelity. The selectivity results from binding to discrete potassium binding sites and stabilization of a hydrated potassium ion in a central internal cavity. The four potassium binding sites, generated by the conserved TTxGYGD signature sequence are formed by the backbone carbonyls of the amino acids TXGYG. Residues KV1.5-Val481, KV4.3-Leu368 and KV7.1- Ile 313 represent the amino acids in the X position of the respective channels. Methods: Here, we study the impact of these residues on ion selectivity, permeation and inactivation kinetics as well as the modulation by β-subunits using site-specific mutagenesis, electrophysiological analyses and molecular dynamics simulations. Results: We identify this position as key in modulation of slow inactivation by structurally dissimilar β-subunits in different KV channels. Conclusion: We propose a model in which structural changes accompanying activation and β-subunit modulation allosterically constrain the backbone carbonyl oxygen atoms via the side chain of the respective X-residue in the signature sequence to reduce conductance during slow inactivation.

  12. Inhibitory effect of calcium channel blockers on proliferation of human glioma cells in vitro

    International Nuclear Information System (INIS)

    Kunert-Radek, J.; Stepien, H.; Lyson, K.; Pawlikowski, M.; Radek, A.

    1989-01-01

    The effects of 2 specific calcium channel blockers, verapamil and nimodipine, on the proliferation of human glioma tumour cells were investigated in vitro. Tumour tissues for primary cell cultures were obtained bioptically from 3 patients with the histopathological diagnosis of glioblastoma. The [ 3 H]-thymidine incorporation into glioma tumour cells DNA was used as a sensitive index of the cell proliferation. It was found that varapamil (10 4 -10 5 M) and nimodipine (10 4 -10 6 M) significantly inhibited the [ 3 H]-thymidine uptake in a dose-related manner. The inhibitory effect of both calcium channel antagonists was reversed by stimultancous addition of calcium chloride (5x10 3 M). These results indicate that verapamil and nimodipine may exert an antiproliferative effect on glioma cells growth acting through a blokade of specific voltage-dependent calcium channels. (author)

  13. T-type voltage-gated calcium channels regulate the tone of mouse efferent arterioles

    DEFF Research Database (Denmark)

    Poulsen, Christian B; Al-Mashhadi, Rozh H; Cribbs, Leanne L

    2011-01-01

    Voltage-gated calcium channels are important for the regulation of renal blood flow and the glomerular filtration rate. Excitation-contraction coupling in afferent arterioles is known to require activation of these channels and we studied their role in the regulation of cortical efferent arteriolar...... tone. We used microdissected perfused mouse efferent arterioles and found a transient vasoconstriction in response to depolarization with potassium; an effect abolished by removal of extracellular calcium. The T-type voltage-gated calcium channel antagonists mibefradil and nickel blocked this potassium...... by immunocytochemistry to be located in mouse efferent arterioles, human pre- and postglomerular vasculature, and Ca(v)3.2 in rat glomerular arterioles. Inhibition of endothelial nitric oxide synthase by L-NAME or its deletion by gene knockout changed the potassium-elicited transient constriction to a sustained response...

  14. Calcium homeostasis modulator (CALHM) ion channels.

    Science.gov (United States)

    Ma, Zhongming; Tanis, Jessica E; Taruno, Akiyuki; Foskett, J Kevin

    2016-03-01

    Calcium homeostasis modulator 1 (CALHM1), formerly known as FAM26C, was recently identified as a physiologically important plasma membrane ion channel. CALHM1 and its Caenorhabditis elegans homolog, CLHM-1, are regulated by membrane voltage and extracellular Ca(2+) concentration ([Ca(2+)]o). In the presence of physiological [Ca(2+)]o (∼1.5 mM), CALHM1 and CLHM-1 are closed at resting membrane potentials but can be opened by strong depolarizations. Reducing [Ca(2+)]o increases channel open probability, enabling channel activation at negative membrane potentials. Together, voltage and Ca(2+) o allosterically regulate CALHM channel gating. Through convergent evolution, CALHM has structural features that are reminiscent of connexins and pannexins/innexins/LRRC8 (volume-regulated anion channel (VRAC)) gene families, including four transmembrane helices with cytoplasmic amino and carboxyl termini. A CALHM1 channel is a hexamer of CALHM1 monomers with a functional pore diameter of ∼14 Å. CALHM channels discriminate poorly among cations and anions, with signaling molecules including Ca(2+) and ATP able to permeate through its pore. CALHM1 is expressed in the brain where it plays an important role in cortical neuron excitability induced by low [Ca(2+)]o and in type II taste bud cells in the tongue that sense sweet, bitter, and umami tastes where it functions as an essential ATP release channel to mediate nonsynaptic neurotransmitter release. CLHM-1 is expressed in C. elegans sensory neurons and body wall muscles, and its genetic deletion causes locomotion defects. Thus, CALHM is a voltage- and Ca(2+) o-gated ion channel, permeable to large cations and anions, that plays important roles in physiology.

  15. David J. Triggle: Medicinal chemistry, to pharmacology, calcium channels, and beyond.

    Science.gov (United States)

    Walker, Michael J A

    2015-11-15

    David Triggle's scientific career began as a chemist, went through medicinal chemistry into pharmacology, and finally on to somewhat more philosophical interests in later years. It was a career marked by many contributions to all of those aspects of science. Chief amongst his many contributions, in addition to those in medicinal chemistry, was his work on the drugs known as calcium ion channel blockers or (calcium antagonists). In the calcium ion channel field he was a particularly instrumental figure in sorting out the mechanisms, actions and roles of the class of calcium channel blockers, known chemical and pharmacologically as the dihydropyridines (DHPs) in particular, as well as other calcium blockers of diverse structures. During the course of a long career, and extensive journeys into medicinal chemistry and pharmacology, he published voluminously in terms of papers, reviews, conference proceedings and books. Notably, many of his papers often had limited authorship where, as senior author it reflected his deep involvement in all aspects of the reported work. His work always helped clarify the field while his incisive reviews, together with his role in coordinating and running scientific meetings, were a great help in clarifying and organizing various fields of study. He has had a long and illustrious career, and is wellknown in the world of biomedical science; his contributions are appreciated, and well recognized everywhere. The following article attempts to chart a path through his work and contributions to medicinal chemistry, pharmacology, science, academia and students. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Inactivation of Mechanically Activated Piezo1 Ion Channels Is Determined by the C-Terminal Extracellular Domain and the Inner Pore Helix

    Directory of Open Access Journals (Sweden)

    Jason Wu

    2017-11-01

    Full Text Available Piezo proteins form mechanically activated ion channels that are responsible for our sense of light touch, proprioception, and vascular blood flow. Upon activation by mechanical stimuli, Piezo channels rapidly inactivate in a voltage-dependent manner through an unknown mechanism. Inactivation of Piezo channels is physiologically important, as it modulates overall mechanical sensitivity, gives rise to frequency filtering of repetitive mechanical stimuli, and is itself the target of numerous human disease-related channelopathies that are not well understood mechanistically. Here, we identify the globular C-terminal extracellular domain as a structure that is sufficient to confer the time course of inactivation and a single positively charged lysine residue at the adjacent inner pore helix as being required for its voltage dependence. Our results are consistent with a mechanism for inactivation that is mediated through voltage-dependent conformations of the inner pore helix and allosteric coupling with the C-terminal extracellular domain.

  17. International Union of Basic and Clinical Pharmacology. C. Nomenclature and Properties of Calcium-Activated and Sodium-Activated Potassium Channels.

    Science.gov (United States)

    Kaczmarek, Leonard K; Aldrich, Richard W; Chandy, K George; Grissmer, Stephan; Wei, Aguan D; Wulff, Heike

    2017-01-01

    A subset of potassium channels is regulated primarily by changes in the cytoplasmic concentration of ions, including calcium, sodium, chloride, and protons. The eight members of this subfamily were originally all designated as calcium-activated channels. More recent studies have clarified the gating mechanisms for these channels and have documented that not all members are sensitive to calcium. This article describes the molecular relationships between these channels and provides an introduction to their functional properties. It also introduces a new nomenclature that differentiates between calcium- and sodium-activated potassium channels. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

  18. Purification and reconstitution of the calcium antagonist receptor of the voltage-sensitive calcium channel

    International Nuclear Information System (INIS)

    Curtis, B.M.

    1986-01-01

    Treatment with digitonin solubilized the calcium antagonist receptor as a stable complex with [ 3 H]nitrendipine from rat brain membranes. The solubilized complex retains allosteric coupling to binding sites for diltiazem, verapamil, and inorganic calcium antagonist sites. The calcium antagonist receptor from cardiac sarcolemma and the transverse-tubule membrane of skeletal muscle is also efficiently solubilized with digitonin and the receptor in all three tissues is a large glycoprotein with a sedimentation coefficient of 20 S. The T-tubule calcium antagonist receptor complex was extensively purified by a combination of chromatography on WGA-Sepharose, ion exchange chromatography, and sedimentation on sucrose gradients to yield preparations estimated to be 41% homogeneous by specific activity and 63% homogeneous by SDS gel electrophoresis. Analysis of SDS gels detect three polypeptides termed α(Mr 135,000), β(Mr 50,000), and γ(Mr 32,000) as noncovalently associated subunits of the calcium antagonist receptor. The α and γ subunits are glycosylated polypeptides, and the molecular weight of the core polypeptides are 108,000 and 24,000 respectively. The calcium antagonist receptor was reconstituted into a phospholipid bilayer by adding CHAPS and exogeneous lipid to the purified receptor followed by rapid detergent removal. This procedure resulted in the incorporation of 45% of the calcium antagonist receptor into closed phospholipid vesicles. Data suggests that the α, β, and γ subunits of the T-tubule calcium antagonist receptor are sufficient to form a functional calcium channel

  19. Effect of propionyl-L-carnitine on L-type calcium channels in human heart sarcolemma

    International Nuclear Information System (INIS)

    Bevilacqua, M.; Vago, T.; Norbiato, G.

    1991-01-01

    Propionyl-L-carnitine (PC) protects perfused rat hearts against damage by ischemia-reperfusion. Activation of L-type calcium channel play a role on ischemia-reperfusion damage. Therefore, we studied the effect of PC on some properties of L-type calcium channels in an in vitro preparation from human myocardium sarcolemma (from patients with idiopathic dilated cardiomyopathy). Binding of the L-type calcium channel blockers isradipine [ 3 H]-PN 200-110 (PN) to plasma membrane preparations revealed a single population of binding sites (total number: Bmax = 213 +/- 34 fM/mg protein and affinity: Kd = 152 +/- 19 nM; n = 6). The characteristics of these binding sites were evaluated in the presence and in the absence of Ca 2+ and of calcium blockers (D-888, a verapamillike drug, and diltiazem). Incubation in a Ca 2+ -containing buffer increased the affinity of PN binding sites. Binding sites for PN were modulated by organic calcium channel blockers; in competition isotherms at 37 degree C, D-888 (desmethoxyverapamil) decreased the PN binding, whereas diltiazem increased it. These results strongly suggest that the site labelled by PN is the voltage-operated calcium channel of the human myocardium. The addition of PC (1 mM) to plasma membranes labelled with PN at 37 degree C decreased the affinity of the binding; this effect was counteracted by the addition of Ca 2+ to the medium. This result was consistent with a competition between Ca 2+ and PC. The effect of PC incubation at 4 degree C was the opposite; at this temperature PC increased the affinity of the binding sites and the effect was obscured by Ca 2+

  20. Follicle-stimulating hormone receptor-mediated uptake of 45Ca2+ by proteoliposomes and cultured rat sertoli cells: Evidence for involvement of voltage-activated and voltage-independent calcium channels

    International Nuclear Information System (INIS)

    Grasso, P.; Reichert, L.E. Jr.

    1989-01-01

    We have previously reported incorporation into liposomes of Triton X-100-solubilized FSH receptor-G-protein complexes derived from purified bovine calf testis membranes. In the present study we have used this model system to show that FSH induces flux of 45Ca2+ into such proteoliposomes in a hormone-specific concentration-dependent manner. FSH, inactivated by boiling, had no stimulatory effect on 45Ca2+ flux, nor did isolated alpha- or beta-subunits of FSH. Addition of GTP (or its analogs 5'-guanylylimidodiphosphate and guanosine-5'-O-[3-thiotriphosphate]) or sodium fluoride (in the presence or absence of GTP or its analogs) failed to induce 45Ca2+ flux into proteoliposomes, suggesting that the uptake of 45Ca2+ was receptor, and not G-protein, related. Voltage-independent (ruthenium red and gadolinium chloride) and voltage-activated (methyoxyverapamil and nifedipine) calcium channel-blocking agents reduced FSH-stimulated 45Ca2+ flux into proteoliposomes to control levels. FSH also induced uptake of 45Ca2+ by cultured rat Sertoli cells. Ruthenium red and gadolinium chloride had no effect on basal levels of 45Ca2+ uptake or estradiol secretion by cultured rat Sertoli cells, nor did methoxyverapamil or nifedipine. All four calcium channel blockers, however, were able to reduce FSH-induced 45Ca2+ uptake to basal levels and FSH-stimulated conversion of androstenedione to estradiol by up to 50%, indicating an involvement of Ca2+ in FSH-stimulated steroidogenesis. Our results suggest that the well documented changes in intracellular calcium levels consequent to FSH binding may be due, at least in part, to an influx of calcium through FSH receptor-regulated calcium channels

  1. Beta-Estradiol Regulates Voltage-Gated Calcium Channels and Estrogen Receptors in Telocytes from Human Myometrium

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    Adela Banciu

    2018-05-01

    Full Text Available Voltage-gated calcium channels and estrogen receptors are essential players in uterine physiology, and their association with different calcium signaling pathways contributes to healthy and pathological conditions of the uterine myometrium. Among the properties of the various cell subtypes present in human uterine myometrium, there is increasing evidence that calcium oscillations in telocytes (TCs contribute to contractile activity and pregnancy. Our study aimed to evaluate the effects of beta-estradiol on voltage-gated calcium channels and estrogen receptors in TCs from human uterine myometrium and to understand their role in pregnancy. For this purpose, we employed patch-clamp recordings, ratiometric Fura-2-based calcium imaging analysis, and qRT-PCR techniques for the analysis of cultured human myometrial TCs derived from pregnant and non-pregnant uterine samples. In human myometrial TCs from both non-pregnant and pregnant uterus, we evidenced by qRT-PCR the presence of genes encoding for voltage-gated calcium channels (Cav3.1, Ca3.2, Cav3.3, Cav2.1, estrogen receptors (ESR1, ESR2, GPR30, and nuclear receptor coactivator 3 (NCOA3. Pregnancy significantly upregulated Cav3.1 and downregulated Cav3.2, Cav3.3, ESR1, ESR2, and NCOA3, compared to the non-pregnant condition. Beta-estradiol treatment (24 h, 10, 100, 1000 nM downregulated Cav3.2, Cav3.3, Cav1.2, ESR1, ESR2, GRP30, and NCOA3 in TCs from human pregnant uterine myometrium. We also confirmed the functional expression of voltage-gated calcium channels by patch-clamp recordings and calcium imaging analysis of TCs from pregnant human myometrium by perfusing with BAY K8644, which induced calcium influx through these channels. Additionally, we demonstrated that beta-estradiol (1000 nM antagonized the effect of BAY K8644 (2.5 or 5 µM in the same preparations. In conclusion, we evidenced the presence of voltage-gated calcium channels and estrogen receptors in TCs from non-pregnant and pregnant

  2. Beta-Estradiol Regulates Voltage-Gated Calcium Channels and Estrogen Receptors in Telocytes from Human Myometrium.

    Science.gov (United States)

    Banciu, Adela; Banciu, Daniel Dumitru; Mustaciosu, Cosmin Catalin; Radu, Mihai; Cretoiu, Dragos; Xiao, Junjie; Cretoiu, Sanda Maria; Suciu, Nicolae; Radu, Beatrice Mihaela

    2018-05-09

    Voltage-gated calcium channels and estrogen receptors are essential players in uterine physiology, and their association with different calcium signaling pathways contributes to healthy and pathological conditions of the uterine myometrium. Among the properties of the various cell subtypes present in human uterine myometrium, there is increasing evidence that calcium oscillations in telocytes (TCs) contribute to contractile activity and pregnancy. Our study aimed to evaluate the effects of beta-estradiol on voltage-gated calcium channels and estrogen receptors in TCs from human uterine myometrium and to understand their role in pregnancy. For this purpose, we employed patch-clamp recordings, ratiometric Fura-2-based calcium imaging analysis, and qRT-PCR techniques for the analysis of cultured human myometrial TCs derived from pregnant and non-pregnant uterine samples. In human myometrial TCs from both non-pregnant and pregnant uterus, we evidenced by qRT-PCR the presence of genes encoding for voltage-gated calcium channels (Cav3.1, Ca3.2, Cav3.3, Cav2.1), estrogen receptors (ESR1, ESR2, GPR30), and nuclear receptor coactivator 3 (NCOA3). Pregnancy significantly upregulated Cav3.1 and downregulated Cav3.2, Cav3.3, ESR1, ESR2, and NCOA3, compared to the non-pregnant condition. Beta-estradiol treatment (24 h, 10, 100, 1000 nM) downregulated Cav3.2, Cav3.3, Cav1.2, ESR1, ESR2, GRP30, and NCOA3 in TCs from human pregnant uterine myometrium. We also confirmed the functional expression of voltage-gated calcium channels by patch-clamp recordings and calcium imaging analysis of TCs from pregnant human myometrium by perfusing with BAY K8644, which induced calcium influx through these channels. Additionally, we demonstrated that beta-estradiol (1000 nM) antagonized the effect of BAY K8644 (2.5 or 5 µM) in the same preparations. In conclusion, we evidenced the presence of voltage-gated calcium channels and estrogen receptors in TCs from non-pregnant and pregnant human uterine

  3. Role of T-type calcium channels in myogenic tone of skeletal muscle resistance arteries

    DEFF Research Database (Denmark)

    VanBavel, Ed; Sorop, Oana; Andreasen, Ditte

    2002-01-01

    T-type calcium channels may be involved in the maintenance of myogenic tone. We tested their role in isolated rat cremaster arterioles obtained after CO(2) anesthesia and decapitation. Total RNA was analyzed by RT-PCR and Southern blotting for calcium channel expression. We observed expression...... of voltage-operated calcium (Ca(V)) channels Ca(V)3.1 (T-type), Ca(V)3.2 (T-type), and Ca(V)1.2 (L-type) in cremaster arterioles (n = 3 rats). Amplification products were observed only in the presence of reverse transcriptase and cDNA. Concentration-response curves of the relatively specific L-type blocker......); K(+) -5.4 +/- 0.3 (n = 4); all log(IC(50)) P maintenance of myogenic tone in rat cremaster muscle arterioles....

  4. Structure-function of proteins interacting with the alpha1 pore-forming subunit of high voltage-activated calcium channel

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    Alan eNeely

    2014-06-01

    Full Text Available Openings of high-voltage-activated calcium channels lead to a transient increase in calcium concentration that in turn activate a plethora of cellular functions, including muscle contraction, secretion and gene transcription. To coordinate all these responses calcium channels form supramolecular assemblies containing effectors and regulatory proteins that couple calcium influx to the downstream signal cascades and to feedback elements. According to the original biochemical characterization of skeletal muscle Dihydropyridine receptors, high-voltage-activated calcium channels are multi-subunit protein complexes consisting of a pore-forming subunit (α1 associated with four additional polypeptide chains β, α2, δ and γ, often referred to as accessory subunits. Twenty-five years after the first purification of a high-voltage calcium channel, the concept of a flexible stoichiometry to expand the repertoire of mechanisms that regulate calcium channel influx has emerged. Several other proteins have been identified that associate directly with the α1-subunit, including calmodulin and multiple members of the small and large GTPase family. Some of these proteins only interact with a subset of α1-subunits and during specific stages of biogenesis. More strikingly, most of the α1-subunit interacting proteins, such as the β-subunit and small GTPases, regulate both gating and trafficking through a variety of mechanisms. Modulation of channel activity covers almost all biophysical properties of the channel. Likewise, regulation of the number of channels in the plasma membrane is performed by altering the release of the α1-subunit from the endoplasmic reticulum, by reducing its degradation or enhancing its recycling back to the cell surface. In this review, we discuss the structural basis, interplay and functional role of selected proteins that interact with the central pore-forming subunit of high-voltage-activated calcium channels.

  5. Structure-function of proteins interacting with the α1 pore-forming subunit of high-voltage-activated calcium channels

    Science.gov (United States)

    Neely, Alan; Hidalgo, Patricia

    2014-01-01

    Openings of high-voltage-activated (HVA) calcium channels lead to a transient increase in calcium concentration that in turn activate a plethora of cellular functions, including muscle contraction, secretion and gene transcription. To coordinate all these responses calcium channels form supramolecular assemblies containing effectors and regulatory proteins that couple calcium influx to the downstream signal cascades and to feedback elements. According to the original biochemical characterization of skeletal muscle Dihydropyridine receptors, HVA calcium channels are multi-subunit protein complexes consisting of a pore-forming subunit (α1) associated with four additional polypeptide chains β, α2, δ, and γ, often referred to as accessory subunits. Twenty-five years after the first purification of a high-voltage calcium channel, the concept of a flexible stoichiometry to expand the repertoire of mechanisms that regulate calcium channel influx has emerged. Several other proteins have been identified that associate directly with the α1-subunit, including calmodulin and multiple members of the small and large GTPase family. Some of these proteins only interact with a subset of α1-subunits and during specific stages of biogenesis. More strikingly, most of the α1-subunit interacting proteins, such as the β-subunit and small GTPases, regulate both gating and trafficking through a variety of mechanisms. Modulation of channel activity covers almost all biophysical properties of the channel. Likewise, regulation of the number of channels in the plasma membrane is performed by altering the release of the α1-subunit from the endoplasmic reticulum, by reducing its degradation or enhancing its recycling back to the cell surface. In this review, we discuss the structural basis, interplay and functional role of selected proteins that interact with the central pore-forming subunit of HVA calcium channels. PMID:24917826

  6. Active Dendrites and Differential Distribution of Calcium Channels Enable Functional Compartmentalization of Golgi Cells.

    Science.gov (United States)

    Rudolph, Stephanie; Hull, Court; Regehr, Wade G

    2015-11-25

    Interneurons are essential to controlling excitability, timing, and synaptic integration in neuronal networks. Golgi cells (GoCs) serve these roles at the input layer of the cerebellar cortex by releasing GABA to inhibit granule cells (grcs). GoCs are excited by mossy fibers (MFs) and grcs and provide feedforward and feedback inhibition to grcs. Here we investigate two important aspects of GoC physiology: the properties of GoC dendrites and the role of calcium signaling in regulating GoC spontaneous activity. Although GoC dendrites are extensive, previous studies concluded they are devoid of voltage-gated ion channels. Hence, the current view holds that somatic voltage signals decay passively within GoC dendrites, and grc synapses onto distal dendrites are not amplified and are therefore ineffective at firing GoCs because of strong passive attenuation. Using whole-cell recording and calcium imaging in rat slices, we find that dendritic voltage-gated sodium channels allow somatic action potentials to activate voltage-gated calcium channels (VGCCs) along the entire dendritic length, with R-type and T-type VGCCs preferentially located distally. We show that R- and T-type VGCCs located in the dendrites can boost distal synaptic inputs and promote burst firing. Active dendrites are thus critical to the regulation of GoC activity, and consequently, to the processing of input to the cerebellar cortex. In contrast, we find that N-type channels are preferentially located near the soma, and control the frequency and pattern of spontaneous firing through their close association with calcium-activated potassium (KCa) channels. Thus, VGCC types are differentially distributed and serve specialized functions within GoCs. Interneurons are essential to neural processing because they modulate excitability, timing, and synaptic integration within circuits. At the input layer of the cerebellar cortex, a single type of interneuron, the Golgi cell (GoC), carries these functions. The

  7. Anoctamin Calcium-Activated Chloride Channels May Modulate Inhibitory Transmission in the Cerebellar Cortex.

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    Weiping Zhang

    Full Text Available Calcium-activated chloride channels of the anoctamin (alias TMEM16 protein family fulfill critical functions in epithelial fluid transport, smooth muscle contraction and sensory signal processing. Little is known, however, about their contribution to information processing in the central nervous system. Here we examined the recent finding that a calcium-dependent chloride conductance impacts on GABAergic synaptic inhibition in Purkinje cells of the cerebellum. We asked whether anoctamin channels may underlie this chloride conductance. We identified two anoctamin channel proteins, ANO1 and ANO2, in the cerebellar cortex. ANO1 was expressed in inhibitory interneurons of the molecular layer and the granule cell layer. Both channels were expressed in Purkinje cells but, while ANO1 appeared to be retained in the cell body, ANO2 was targeted to the dendritic tree. Functional studies confirmed that ANO2 was involved in a calcium-dependent mode of ionic plasticity that reduces the efficacy of GABAergic synapses. ANO2 channels attenuated GABAergic transmission by increasing the postsynaptic chloride concentration, hence reducing the driving force for chloride influx. Our data suggest that ANO2 channels are involved in a Ca2+-dependent regulation of synaptic weight in GABAergic inhibition. Thus, in balance with the chloride extrusion mechanism via the co-transporter KCC2, ANO2 appears to regulate ionic plasticity in the cerebellum.

  8. The mechanosensory calcium-selective ion channel: key component of a plasmalemmal control centre?

    Science.gov (United States)

    Pickard, B. G.; Ding, J. P.

    1993-01-01

    Mechanosensory calcium-selective ion channels probably serve to detect not only mechanical stress but also electrical, thermal, and diverse chemical stimuli. Because all stimuli result in a common output, most notably a shift in second messenger calcium concentration, the channels are presumed to serve as signal integrators. Further, insofar as second messenger calcium in turn gives rise to mechanical, electrical, and diverse chemical changes, the channels are postulated to initiate regulatory feedbacks. It is proposed that the channels and the feedback loops play a wide range of roles in regulating normal plant function, as well as in mediating disturbance of normal function by environmental stressors and various pathogens. In developing evidence for the physiological performance of the channel, a model for a cluster of regulatory plasmalemmal proteins and cytoskeletal elements grouped around a set of wall-to-membrane and transmembrane linkers has proved useful. An illustration of how the model might operate is presented. It is founded on the demonstration that several xenobiotics interfere both with normal channel behaviour and with gravitropic reception. Accordingly, the first part of the illustration deals with how the channels and the control system within which they putatively operate might initiate gravitropism. Assuming that gravitropism is an asymmetric expression of growth, the activities of the channels and the plasmalemmal control system are extrapolated to account for regulation of both rate and allometry of cell expansion. Finally, it is discussed how light, hormones, redox agents and herbicides could in principle affect growth via the putative plasmalemmal control cluster or centre.

  9. Divergent biophysical properties, gating mechanisms, and possible functions of the two skeletal muscle Ca(V)1.1 calcium channel splice variants.

    Science.gov (United States)

    Tuluc, Petronel; Flucher, Bernhard E

    2011-12-01

    Voltage-gated calcium channels are multi-subunit protein complexes that specifically allow calcium ions to enter the cell in response to membrane depolarization. But, for many years it seemed that the skeletal muscle calcium channel Ca(V)1.1 is the exception. The classical splice variant Ca(V)1.1a activates slowly, has a very small current amplitude and poor voltage sensitivity. In fact adult muscle fibers work perfectly well even in the absence of calcium influx. Recently a new splice variant of the skeletal muscle calcium channel Ca(V)1.1e has been characterized. The lack of the 19 amino acid exon 29 in this splice variant results in a rapidly activating calcium channel with high current amplitude and good voltage sensitivity. Ca(V)1.1e is the dominant channel in embryonic muscle, where the expression of this high calcium-conducting Ca(V)1.1 isoform readily explains developmental processes depending on L-type calcium currents. Moreover, the availability of these two structurally similar but functionally distinct channel variants facilitates the analysis of the molecular mechanisms underlying the unique current properties of the classical Ca(V)1.1a channel.

  10. Dendritic calcium channels and their activation by synaptic signals in auditory coincidence detector neurons.

    Science.gov (United States)

    Blackmer, Trillium; Kuo, Sidney P; Bender, Kevin J; Apostolides, Pierre F; Trussell, Laurence O

    2009-08-01

    The avian nucleus laminaris (NL) encodes the azimuthal location of low-frequency sound sources by detecting the coincidence of binaural signals. Accurate coincidence detection requires precise developmental regulation of the lengths of the fine, bitufted dendrites that characterize neurons in NL. Such regulation has been suggested to be driven by local, synaptically mediated, dendritic signals such as Ca(2+). We examined Ca(2+) signaling through patch clamp and ion imaging experiments in slices containing nucleus laminaris from embryonic chicks. Voltage-clamp recordings of neurons located in the NL showed the presence of large Ca(2+) currents of two types, a low voltage-activated, fast inactivating Ni(2+) sensitive channel resembling mammalian T-type channels, and a high voltage-activated, slowly inactivating Cd(2+) sensitive channel. Two-photon Ca(2+) imaging showed that both channel types were concentrated on dendrites, even at their distal tips. Single action potentials triggered synaptically or by somatic current injection immediately elevated Ca(2+) throughout the entire cell. Ca(2+) signals triggered by subthreshold synaptic activity were highly localized. Thus when electrical activity is suprathreshold, Ca(2+) channels ensure that Ca(2+) rises in all dendrites, even those that are synaptically inactive.

  11. Molecular Dynamics Simulations of Orai Reveal How the Third Transmembrane Segment Contributes to Hydration and Ca2+ Selectivity in Calcium Release-Activated Calcium Channels.

    Science.gov (United States)

    Alavizargar, Azadeh; Berti, Claudio; Ejtehadi, Mohammad Reza; Furini, Simone

    2018-04-26

    Calcium release-activated calcium (CRAC) channels open upon depletion of Ca 2+ from the endoplasmic reticulum, and when open, they are permeable to a selective flux of calcium ions. The atomic structure of Orai, the pore domain of CRAC channels, from Drosophila melanogaster has revealed many details about conduction and selectivity in this family of ion channels. However, it is still unclear how residues on the third transmembrane helix can affect the conduction properties of the channel. Here, molecular dynamics and Brownian dynamics simulations were employed to analyze how a conserved glutamate residue on the third transmembrane helix (E262) contributes to selectivity. The comparison between the wild-type and mutated channels revealed a severe impact of the mutation on the hydration pattern of the pore domain and on the dynamics of residues K270, and Brownian dynamics simulations proved that the altered configuration of residues K270 in the mutated channel impairs selectivity to Ca 2+ over Na + . The crevices of water molecules, revealed by molecular dynamics simulations, are perfectly located to contribute to the dynamics of the hydrophobic gate and the basic gate, suggesting a possible role in channel opening and in selectivity function.

  12. High affinity complexes of pannexin channels and L-type calcium channel splice-variants in human lung: Possible role in clevidipine-induced dyspnea relief in acute heart failure

    Directory of Open Access Journals (Sweden)

    Gerhard P. Dahl

    2016-08-01

    Research in Context: Clevidipine lowers blood pressure by inhibiting calcium channels in vascular smooth muscle. In patients with acute heart failure, clevidipine was shown to relieve breathing problems. This was only partially related to the blood pressure lowering actions of clevidipine and not conferred by another calcium channel inhibitor. We here found calcium channel variants in human lung that are more selectively inhibited by clevidipine, especially when associated with pannexin channels. This study gives a possible mechanism for clevidipine's relief of breathing problems and supports future clinical trials testing the role of clevidipine in the treatment of acute heart failure.

  13. Biphasic voltage-dependent inactivation of human NaV 1.3, 1.6 and 1.7 Na+ channels expressed in rodent insulin-secreting cells.

    Science.gov (United States)

    Godazgar, Mahdieh; Zhang, Quan; Chibalina, Margarita V; Rorsman, Patrik

    2018-05-01

    Na + current inactivation is biphasic in insulin-secreting cells, proceeding with two voltage dependences that are half-maximal at ∼-100 mV and -60 mV. Inactivation of voltage-gated Na + (Na V ) channels occurs at ∼30 mV more negative voltages in insulin-secreting Ins1 and primary β-cells than in HEK, CHO or glucagon-secreting αTC1-6 cells. The difference in inactivation between Ins1 and non-β-cells persists in the inside-out patch configuration, discounting an involvement of a diffusible factor. In Ins1 cells and primary β-cells, but not in HEK cells, inactivation of a single Na V subtype is biphasic and follows two voltage dependences separated by 30-40 mV. We propose that Na V channels adopt different inactivation behaviours depending on the local membrane environment. Pancreatic β-cells are equipped with voltage-gated Na + channels that undergo biphasic voltage-dependent steady-state inactivation. A small Na + current component (10-15%) inactivates over physiological membrane potentials and contributes to action potential firing. However, the major Na + channel component is completely inactivated at -90 to -80 mV and is therefore inactive in the β-cell. It has been proposed that the biphasic inactivation reflects the contribution of different Na V α-subunits. We tested this possibility by expression of TTX-resistant variants of the Na V subunits found in β-cells (Na V 1.3, Na V 1.6 and Na V 1.7) in insulin-secreting Ins1 cells and in non-β-cells (including HEK and CHO cells). We found that all Na V subunits inactivated at 20-30 mV more negative membrane potentials in Ins1 cells than in HEK or CHO cells. The more negative inactivation in Ins1 cells does not involve a diffusible intracellular factor because the difference between Ins1 and CHO persisted after excision of the membrane. Na V 1.7 inactivated at 15--20 mV more negative membrane potentials than Na V 1.3 and Na V 1.6 in Ins1 cells but this small difference is insufficient to solely

  14. Cardiac voltage gated calcium channels and their regulation by β-adrenergic signaling.

    Science.gov (United States)

    Kumari, Neema; Gaur, Himanshu; Bhargava, Anamika

    2018-02-01

    Voltage-gated calcium channels (VGCCs) are the predominant source of calcium influx in the heart leading to calcium-induced calcium release and ultimately excitation-contraction coupling. In the heart, VGCCs are modulated by the β-adrenergic signaling. Signaling through β-adrenergic receptors (βARs) and modulation of VGCCs by β-adrenergic signaling in the heart are critical signaling and changes to these have been significantly implicated in heart failure. However, data related to calcium channel dysfunction in heart failure is divergent and contradictory ranging from reduced function to no change in the calcium current. Many recent studies have highlighted the importance of functional and spatial microdomains in the heart and that may be the key to answer several puzzling questions. In this review, we have briefly discussed the types of VGCCs found in heart tissues, their structure, and significance in the normal and pathological condition of the heart. More importantly, we have reviewed the modulation of VGCCs by βARs in normal and pathological conditions incorporating functional and structural aspects. There are different types of βARs, each having their own significance in the functioning of the heart. Finally, we emphasize the importance of location of proteins as it relates to their function and modulation by co-signaling molecules. Its implication on the studies of heart failure is speculated. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Long-Term Blocking of Calcium Channels in mdx Mice Results in Differential Effects on Heart and Skeletal Muscle

    DEFF Research Database (Denmark)

    Jørgensen, Louise Helskov; Blain, Alison; Greally, Elizabeth

    2011-01-01

    in older mice. However, streptomycin treatment did not show positive effects in diaphragm or heart muscle, and heart pathology was worsened. Thus, blocking calcium channels even before disease onset does not prevent dystrophy, making this an unlikely treatment for DMD. These findings highlight......The disease mechanisms underlying dystrophin-deficient muscular dystrophy are complex, involving not only muscle membrane fragility, but also dysregulated calcium homeostasis. Specifically, it has been proposed that calcium channels directly initiate a cascade of pathological events by allowing...... calcium ions to enter the cell. The objective of this study was to investigate the effect of chronically blocking calcium channels with the aminoglycoside antibiotic streptomycin from onset of disease in the mdx mouse model of Duchenne muscular dystrophy (DMD). Treatment in utero onwards delayed onset...

  16. Preparation and preclinical evaluation of 68Ga-DOTA-amlodipine for L-type calcium channel imaging

    Science.gov (United States)

    Firuzyar, Tahereh; Jalilian, Amir Reza; Aboudzadeh, Mohammad Reza; Sadeghpour, Hossein; Shafiee-Ardestani, Mahdi; Khalaj, Ali

    2016-01-01

    Aim: In order to develop a possible tracer for L-type calcium channel imaging, we here report the development of a Ga-68 amlodipine derivative for possible PET imaging. Materials and Methods: Amlodipine DOTA conjugate was synthesized, characterized and went through calcium channel blockade, toxicity, apoptosis/necrosis tests. [68Ga] DOTA AMLO was prepared at optimized conditions followed by stability tests, partition coefficient determination and biodistribution studies using tissue counting and co incidence imaging up to 2 h. Results: [68Ga] DOTA AMLO was prepared at pH 4–5 in 7–10 min at 95°C in high radiochemical purity (>99%, radio thin layer chromatography; specific activity: 1.9–2.1 GBq/mmol) and was stable up to 4 h with a log P of −0.94. Calcium channel rich tissues including myocardium, and tissues with smooth muscle cells such as colon, intestine, and lungs demonstrated significant uptake. Co incidence images supported the biodistribution data up to 2 h. Conclusions: The complex can be a candidate for further positron emission tomography imaging for L type calcium channels. PMID:27833311

  17. Effects of Calcium Channel Blockers on Antidepressant Action of Alprazolam and Imipramine

    Directory of Open Access Journals (Sweden)

    Gorash ZM

    2007-01-01

    Full Text Available Alprazolam is effective as an anxiolytic and in the adjunct treatment of depression. In this study, the effects of calcium channel antagonists on the antidepressant action of alprazolam and imipramine were investigated. A forced swimming maze was used to study behavioral despair in albino mice. Mice were divided into nine groups (n = 7 per group. One group received a single dose of 1% Tween 80; two groups each received a single dose of the antidepressant alone (alprazolam or imipramine; two groups each received a single dose of the calcium channel blocker (nifedipine or verapamil; four groups each received a single dose of the calcium channel blocker followed by a single dose of the antidepressant (with same doses used for either in the previous four groups. Drug administration was performed concurrently on the nine groups. Our data confirmed the antidepressant action of alprazolam and imipramine. Both nifedipine and verapamil produced a significant antidepressant effect (delay the onset of immobility when administered separately. Verapamil augmented the antidepressant effects of alprazolam and imipramine (additive antidepressant effect. This may be due to the possibility that verapamil might have antidepressant-like effect through different mechanism. Nifedipine and imipramine combined led to a delay in the onset of immobility greater than their single use but less than the sum of their independent administration. This may be due to the fact that nifedipine on its own might act as an antidepressant but blocks one imipramine mechanism that depends on L-type calcium channel activation. Combining nifedipine with alprazolam produced additional antidepressant effects, which indicates that they exert antidepressant effects through different mechanisms.

  18. Tx1, from Phoneutria nigriventer spider venom, interacts with dihydropyridine sensitive-calcium channels in GH3 cells

    International Nuclear Information System (INIS)

    Gouvea dos Santos, R.; Soares, M.A.; Pimenta, A.M.; De Lima, M.E.; ICB, UFMG, Belo Horizonte

    2006-01-01

    The aim of this work was to use the binding assay of tritiated-dihydropyridine and radioiodinated Tx1, isolated from the Phoneutria nigriventer venom, in order to show the presence of Ca v 1 calcium channels on pituitary tumour cell (GH3). We showed that GH3 cells have specific sites for 125 I-Tx1, which are sensitive to nifedipine (∼20%). Reverse competition assay with 3 H-PN200-110 (40% inhibition) and electrophysiological data (50% inhibition) suggest that Ca v 1 calcium channels are target sites for this toxin. To summarize, Tx1 binds to specific sites on GH3 cells and this interaction results in Ca v 1 calcium channel blockade. 3 H-PN200-110 and 125 I-Tx1 binding assays proved to be useful tools to show the presence of calcium channels on GH3 cells. (author)

  19. 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

  20. Identification of a Novel EF-Loop in the N-terminus of TRPM2 Channel Involved in Calcium Sensitivity

    Directory of Open Access Journals (Sweden)

    Yuhuan Luo

    2018-06-01

    Full Text Available As an oxidative stress sensor, transient receptor potential melastatin 2 (TRPM2 channel is involved in many physiological and pathological processes including warmth sensing, ischemia injury, inflammatory diseases and diabetes. Intracellular calcium is critical for TRPM2 channel activation and the IQ-like motif in the N-terminus has been shown to be important by mediating calmodulin binding. Sequence analysis predicted two potential EF-loops in the N-terminus of TRPM2. Site-directed mutagenesis combining with functional assay showed that substitution with alanine of several residues, most of which are conserved in the typical EF-loop, including D267, D278, D288, and E298 dramatically reduced TRPM2 channel currents. By further changing the charges or side chain length of these conserved residues, our results indicate that the negative charge of D267 and the side chain length of D278 are critical for calcium-induced TRPM2 channel activation. G272I mutation also dramatically reduced the channel currents, suggesting that this site is critical for calcium-induced TRPM2 channel activation. Furthermore, D267A mutant dramatically reduced the currents induced by calcium alone compared with that by ADPR, indicating that D267 residue in D267–D278 motif is the most important site for calcium sensitivity of TRPM2. In addition, inside-out recordings showed that mutations at D267, G272, D278, and E298 had no effect on single-channel conductance. Taken together, our data indicate that D267–D278 motif in the N-terminus as a novel EF-loop is critical for calcium-induced TRPM2 channel activation.

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

    DEFF Research Database (Denmark)

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

    2012-01-01

    Small-conductance calcium-activated potassium (SK3) channels have been detected in human myometrium and we have previously shown a functional role of SK channels in human myometrium in vitro. The aims of this study were to identify the precise localization of SK3 channels and to quantify SK3 m....... This is the first report to provide evidence for a possible role of SK3 channels in human uterine telocytes....

  2. Activity of Palythoa caribaeorum Venom on Voltage-Gated Ion Channels in Mammalian Superior Cervical Ganglion Neurons.

    Science.gov (United States)

    Lazcano-Pérez, Fernando; Castro, Héctor; Arenas, Isabel; García, David E; González-Muñoz, Ricardo; Arreguín-Espinosa, Roberto

    2016-05-05

    The Zoanthids are an order of cnidarians whose venoms and toxins have been poorly studied. Palythoa caribaeorum is a zoanthid commonly found around the Mexican coastline. In this study, we tested the activity of P. caribaeorum venom on voltage-gated sodium channel (NaV1.7), voltage-gated calcium channel (CaV2.2), the A-type transient outward (IA) and delayed rectifier (IDR) currents of KV channels of the superior cervical ganglion (SCG) neurons of the rat. These results showed that the venom reversibly delays the inactivation process of voltage-gated sodium channels and inhibits voltage-gated calcium and potassium channels in this mammalian model. The compounds responsible for these effects seem to be low molecular weight peptides. Together, these results provide evidence for the potential use of zoanthids as a novel source of cnidarian toxins active on voltage-gated ion channels.

  3. Protein structure and ionic selectivity in calcium channels: Selectivity filter size, not shape, matters

    OpenAIRE

    Malasics, Attila; Gillespie, Dirk; Nonner, Wolfgang; Henderson, Douglas; Eisenberg, Bob; Boda, Dezső

    2009-01-01

    Calcium channels have highly charged selectivity filters (4 COO− groups) that attract cations in to balance this charge and minimize free energy, forcing the cations (Na+ and Ca2+) to compete for space in the filter. A reduced model was developed to better understand the mechanism of ion selectivity in calcium channels. The charge/space competition (CSC) mechanism implies that Ca2+ is more efficient in balancing the charge of the filter because it provides twice the charge as Na+ while occupy...

  4. Sigma-1 Receptor Plays a Negative Modulation on N-type Calcium Channel

    Directory of Open Access Journals (Sweden)

    Kang Zhang

    2017-05-01

    Full Text Available The sigma-1 receptor is a 223 amino acids molecular chaperone with a single transmembrane domain. It is resident to eukaryotic mitochondrial-associated endoplasmic reticulum and plasma membranes. By chaperone-mediated interactions with ion channels, G-protein coupled receptors and cell-signaling molecules, the sigma-1 receptor performs broad physiological and pharmacological functions. Despite sigma-1 receptors have been confirmed to regulate various types of ion channels, the relationship between the sigma-1 receptor and N-type Ca2+ channel is still unclear. Considering both sigma-1 receptors and N-type Ca2+ channels are involved in intracellular calcium homeostasis and neurotransmission, we undertake studies to explore the possible interaction between these two proteins. In the experiment, we confirmed the expression of the sigma-1 receptors and the N-type calcium channels in the cholinergic interneurons (ChIs in rat striatum by using single-cell reverse transcription-polymerase chain reaction (scRT-PCR and immunofluorescence staining. N-type Ca2+ currents recorded from ChIs in the brain slice of rat striatum was depressed when sigma-1 receptor agonists (SKF-10047 and Pre-084 were administrated. The inhibition was completely abolished by sigma-1 receptor antagonist (BD-1063. Co-expression of the sigma-1 receptors and the N-type calcium channels in Xenopus oocytes presented a decrease of N-type Ca2+ current amplitude with an increase of sigma-1 receptor expression. SKF-10047 could further depress N-type Ca2+ currents recorded from oocytes. The fluorescence resonance energy transfer (FRET assays and co-immunoprecipitation (Co-IP demonstrated that sigma-1 receptors and N-type Ca2+ channels formed a protein complex when they were co-expressed in HEK-293T (Human Embryonic Kidney -293T cells. Our results revealed that the sigma-1 receptors played a negative modulation on N-type Ca2+ channels. The mechanism for the inhibition of sigma-1 receptors on

  5. Conduction velocity is regulated by sodium channel inactivation in unmyelinated axons innervating the rat cranial meninges.

    Science.gov (United States)

    De Col, Roberto; Messlinger, Karl; Carr, Richard W

    2008-02-15

    Axonal conduction velocity varies according to the level of preceding impulse activity. In unmyelinated axons this typically results in a slowing of conduction velocity and a parallel increase in threshold. It is currently held that Na(+)-K(+)-ATPase-dependent axonal hyperpolarization is responsible for this slowing but this has long been equivocal. We therefore examined conduction velocity changes during repetitive activation of single unmyelinated axons innervating the rat cranial meninges. In direct contradiction to the currently accepted postulate, Na(+)-K(+)-ATPase blockade actually enhanced activity-induced conduction velocity slowing, while the degree of velocity slowing was curtailed in the presence of lidocaine (10-300 microm) and carbamazepine (30-500 microm) but not tetrodotoxin (TTX, 10-80 nm). This suggests that a change in the number of available sodium channels is the most prominent factor responsible for activity-induced changes in conduction velocity in unmyelinated axons. At moderate stimulus frequencies, axonal conduction velocity is determined by an interaction between residual sodium channel inactivation following each impulse and the retrieval of channels from inactivation by a concomitant Na(+)-K(+)-ATPase-mediated hyperpolarization. Since the process is primarily dependent upon sodium channel availability, tracking conduction velocity provides a means of accessing relative changes in the excitability of nociceptive neurons.

  6. Conotoxins as Tools to Understand the Physiological Function of Voltage-Gated Calcium (CaV Channels

    Directory of Open Access Journals (Sweden)

    David Ramírez

    2017-10-01

    Full Text Available Voltage-gated calcium (CaV channels are widely expressed and are essential for the completion of multiple physiological processes. Close regulation of their activity by specific inhibitors and agonists become fundamental to understand their role in cellular homeostasis as well as in human tissues and organs. CaV channels are divided into two groups depending on the membrane potential required to activate them: High-voltage activated (HVA, CaV1.1–1.4; CaV2.1–2.3 and Low-voltage activated (LVA, CaV3.1–3.3. HVA channels are highly expressed in brain (neurons, heart, and adrenal medulla (chromaffin cells, among others, and are also classified into subtypes which can be distinguished using pharmacological approaches. Cone snails are marine gastropods that capture their prey by injecting venom, “conopeptides”, which cause paralysis in a few seconds. A subset of conopeptides called conotoxins are relatively small polypeptides, rich in disulfide bonds, that target ion channels, transporters and receptors localized at the neuromuscular system of the animal target. In this review, we describe the structure and properties of conotoxins that selectively block HVA calcium channels. We compare their potency on several HVA channel subtypes, emphasizing neuronal calcium channels. Lastly, we analyze recent advances in the therapeutic use of conotoxins for medical treatments.

  7. Activity of the anticonvulsant lacosamide in experimental and human epilepsy via selective effects on slow Na+ channel inactivation.

    Science.gov (United States)

    Holtkamp, Dominik; Opitz, Thoralf; Niespodziany, Isabelle; Wolff, Christian; Beck, Heinz

    2017-01-01

    In human epilepsy, pharmacoresistance to antiepileptic drug therapy is a major problem affecting ~30% of patients with epilepsy. Many classical antiepileptic drugs target voltage-gated sodium channels, and their potent activity in inhibiting high-frequency firing has been attributed to their strong use-dependent blocking action. In chronic epilepsy, a loss of use-dependent block has emerged as a potential cellular mechanism of pharmacoresistance for anticonvulsants acting on voltage-gated sodium channels. The anticonvulsant drug lacosamide (LCM) also targets sodium channels, but has been shown to preferentially affect sodium channel slow inactivation processes, in contrast to most other anticonvulsants. We used whole-cell voltage clamp recordings in acutely isolated cells to investigate the effects of LCM on transient Na + currents. Furthermore, we used whole-cell current clamp recordings to assess effects on repetitive action potential firing in hippocampal slices. We show here that LCM exerts its effects primarily via shifting the slow inactivation voltage dependence to more hyperpolarized potentials in hippocampal dentate granule cells from control and epileptic rats, and from patients with epilepsy. It is important to note that this activity of LCM was maintained in chronic experimental and human epilepsy. Furthermore, we demonstrate that the efficacy of LCM in inhibiting high-frequency firing is undiminished in chronic experimental and human epilepsy. Taken together, these results show that LCM exhibits maintained efficacy in chronic epilepsy, in contrast to conventional use-dependent sodium channel blockers such as carbamazepine. They also establish that targeting slow inactivation may be a promising strategy for overcoming target mechanisms of pharmacoresistance. Wiley Periodicals, Inc. © 2016 International League Against Epilepsy.

  8. Distribution of calcium channel Ca(V)1.3 immunoreactivity in the rat spinal cord and brain stem.

    Science.gov (United States)

    Sukiasyan, N; Hultborn, H; Zhang, M

    2009-03-03

    The function of local networks in the CNS depends upon both the connectivity between neurons and their intrinsic properties. An intrinsic property of spinal motoneurons is the presence of persistent inward currents (PICs), which are mediated by non-inactivating calcium (mainly Ca(V)1.3) and/or sodium channels and serve to amplify neuronal input signals. It is of fundamental importance for the prediction of network function to determine the distribution of neurons possessing the ion channels that produce PICs. Although the distribution pattern of Ca(V)1.3 immunoreactivity (Ca(V)1.3-IR) has been studied in some specific central nervous regions in some species, so far no systematic investigations have been performed in both the rat spinal cord and brain stem. In the present study this issue was investigated by immunohistochemistry. The results indicated that the Ca(V)1.3-IR neurons were widely distributed across different parts of the spinal cord and the brain stem although with variable labeling intensities. In the spinal gray matter large neurons in the ventral horn (presumably motoneurons) tended to display higher levels of immunoreactivity than smaller neurons in the dorsal horn. In the white matter, a subset of glial cells labeled by an oligodendrocyte marker was also Ca(V)1.3-positive. In the brain stem, neurons in the motor nuclei appeared to have higher levels of immunoreactivity than those in the sensory nuclei. Moreover, a number of nuclei containing monoaminergic cells, for example the locus coeruleus, were also strongly immunoreactive. Ca(V)1.3-IR was consistently detected in the neuronal perikarya regardless of the neuronal type. However, in the large neurons in the spinal ventral horn and the cranial motor nuclei the Ca(V)1.3-IR was clearly detectable in first and second order dendrites. These results indicate that in the rat spinal cord and brain stem Ca(V)1.3 is probably a common calcium channel used by many kinds of neurons to facilitate the neuronal

  9. Calcium in plant cells

    Directory of Open Access Journals (Sweden)

    V. V. Schwartau

    2014-04-01

    Full Text Available The paper gives the review on the role of calcium in many physiological processes of plant organisms, including growth and development, protection from pathogenic influences, response to changing environmental factors, and many other aspects of plant physiology. Initial intake of calcium ions is carried out by Ca2+-channels of plasma membrane and they are further transported by the xylem owing to auxins’ attractive ability. The level of intake and selectivity of calcium transport to ove-ground parts of the plant is controlled by a symplast. Ca2+enters to the cytoplasm of endoderm cells through calcium channels on the cortical side of Kaspary bands, and is redistributed inside the stele by the symplast, with the use of Ca2+-АТPases and Ca2+/Н+-antiports. Owing to regulated expression and activity of these calcium transporters, calclum can be selectively delivered to the xylem. Important role in supporting calcium homeostasis is given to the vacuole which is the largest depo of calcium. Regulated quantity of calcium movement through the tonoplast is provided by a number of potential-, ligand-gated active transporters and channels, like Ca2+-ATPase and Ca2+/H+ exchanger. They are actively involved in the inactivation of the calcium signal by pumping Ca2+ to the depo of cells. Calcium ATPases are high affinity pumps that efficiently transfer calcium ions against the concentration gradient in their presence in the solution in nanomolar concentrations. Calcium exchangers are low affinity, high capacity Ca2+ transporters that are effectively transporting calcium after raising its concentration in the cell cytosol through the use of protons gradients. Maintaining constant concentration and participation in the response to stimuli of different types also involves EPR, plastids, mitochondria, and cell wall. Calcium binding proteins contain several conserved sequences that provide sensitivity to changes in the concentration of Ca2+ and when you

  10. Use of calcium channel blockers in hypertension.

    Science.gov (United States)

    Conlin, P R; Williams, G H

    1998-01-01

    During the past 20 years the number of subclasses of calcium channel blockers has increased from one to four. Three classes have only a single clinically approved compound: verapamil, diltiazem, and mibefradil. The fourth class, dihydropyridines, contains numerous compounds. All agents are effective in lowering blood pressure in short-term studies, and side effects that trouble the patient are infrequent. Long-term studies in hypertensive patients are limited. Short-acting agents such as nifedipine have been associated with an increased cardiovascular risk in some, but not all studies. These agents also probably create a compliance problem for hypertensive patients because of the need for multiple daily doses and their unpleasant side effects, e.g., ankle edema, palpitations, and flushing. Therefore, they are not useful or indicated for the treatment of hypertensive patients. No data have suggested that long-acting dihydropyridines or nondihydropyridine calcium channel blockers share the same fate. Indeed, several lines of evidence suggest the opposite: they have a cardioprotective effect. However, definitive information will require the completion of several long-term trials, including ALLHAT, CONVINCE, HOT, INSIGHT and NORDIL. Finally, it is important to reflect on the lessons learned from the controversy associated with the potential risks of calcium channel blockers. First, disagreements are common when one uses case-controlled studies and are reflective of the poor precision of the methods used. What is statistically relevant in one study may not hold true for another and may have no clinical relevance, particularly if the relative risk is less than 2. Investigators need to temper their enthusiasm to reflect this reality. Second, at the cutting edge of science there is probably relatively little agreement about what is correct among equally competent scientists. All have bias in their positions and should both recognize and admit so to themselves and their

  11. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

    DEFF Research Database (Denmark)

    Jørgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne

    2003-01-01

    The propagation of mechanically induced intercellular calcium waves (ICW) among osteoblastic cells occurs both by activation of P2Y (purinergic) receptors by extracellular nucleotides, resulting in "fast" ICW, and by gap junctional communication in cells that express connexin43 (Cx43), resulting...... in "slow" ICW. Human osteoblastic cells transmit intercellular calcium signals by both of these mechanisms. In the current studies we have examined the mechanism of slow gap junction-dependent ICW in osteoblastic cells. In ROS rat osteoblastic cells, gap junction-dependent ICW were inhibited by removal...... of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx...

  12. Activity of Palythoa caribaeorum Venom on Voltage-Gated Ion Channels in Mammalian Superior Cervical Ganglion Neurons

    Directory of Open Access Journals (Sweden)

    Fernando Lazcano-Pérez

    2016-05-01

    Full Text Available The Zoanthids are an order of cnidarians whose venoms and toxins have been poorly studied. Palythoa caribaeorum is a zoanthid commonly found around the Mexican coastline. In this study, we tested the activity of P. caribaeorum venom on voltage-gated sodium channel (NaV1.7, voltage-gated calcium channel (CaV2.2, the A-type transient outward (IA and delayed rectifier (IDR currents of KV channels of the superior cervical ganglion (SCG neurons of the rat. These results showed that the venom reversibly delays the inactivation process of voltage-gated sodium channels and inhibits voltage-gated calcium and potassium channels in this mammalian model. The compounds responsible for these effects seem to be low molecular weight peptides. Together, these results provide evidence for the potential use of zoanthids as a novel source of cnidarian toxins active on voltage-gated ion channels.

  13. [Distribution diversity of integrins and calcium channels on major human and mouse host cells of Leptospira species].

    Science.gov (United States)

    Li, Cheng-xue; Zhao, Xin; Qian, Jing; Yan, Jie

    2012-07-01

    To determine the distribution of integrins and calcium channels on major human and mouse host cells of Leptospira species. The expression of β1, β2 and β3 integrins was detected with immunofluorescence assay on the surface of human monocyte line THP-1, mouse mononuclear-macrophage-like cell line J774A.1, human vascular endothelial cell line HUVEC, mouse vascular endothelial cell EOMA, human hepatocyte line L-02, mouse hepatocyte line Hepa1-6, human renal tubular epithelial cell line HEK-293, mouse glomerular membrane epithelial cell line SV40-MES13, mouse collagen blast line NIH/3T3, human and mouse platelets. The distribution of voltage gate control calcium channels Cav3.1, Cav3.2, Cav3.3 and Cav2.3, and receptor gate calcium channels P(2)X(1), P(2)2X(2), P(2)X(3), P(2)X(4), P(2)X(5), P(2)X(6) and P(2)X(7) were determined with Western blot assay. β1 integrin proteins were positively expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, L-02, Hepa1-6 and HEK-239 cells as well as human and mouse platelets. β2 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, and NIH/3T3 cells. β3 integrin proteins were expressed on the membrane surface of J774A.1, THP-1, HUVEC, EOMA, Hepa1-6, HEK-239 and NIH/3T3 cells as well as human and mouse platelets. P(2)X(1) receptor gate calcium channel was expressed on the membrane surface of human and mouse platelets, while P(2)X(5) receptor gate calcium channel was expressed on the membrane surface of J774A.1, THP-1, L-02, Hepa1-6, HEK-239 and HUVEC cells. However, the other calcium channels were not detected on the tested cell lines or platelets. There is a large distribution diversity of integrins and calcium channel proteins on the major human and mouse host cells of Leptospira species, which may be associated with the differences of leptospira-induced injury in different host cells.

  14. Thermodynamic coupling between activation and inactivation gating in potassium channels revealed by free energy molecular dynamics simulations.

    Science.gov (United States)

    Pan, Albert C; Cuello, Luis G; Perozo, Eduardo; Roux, Benoît

    2011-12-01

    The amount of ionic current flowing through K(+) channels is determined by the interplay between two separate time-dependent processes: activation and inactivation gating. Activation is concerned with the stimulus-dependent opening of the main intracellular gate, whereas inactivation is a spontaneous conformational transition of the selectivity filter toward a nonconductive state occurring on a variety of timescales. A recent analysis of multiple x-ray structures of open and partially open KcsA channels revealed the mechanism by which movements of the inner activation gate, formed by the inner helices from the four subunits of the pore domain, bias the conformational changes at the selectivity filter toward a nonconductive inactivated state. This analysis highlighted the important role of Phe103, a residue located along the inner helix, near the hinge position associated with the opening of the intracellular gate. In the present study, we use free energy perturbation molecular dynamics simulations (FEP/MD) to quantitatively elucidate the thermodynamic basis for the coupling between the intracellular gate and the selectivity filter. The results of the FEP/MD calculations are in good agreement with experiments, and further analysis of the repulsive, van der Waals dispersive, and electrostatic free energy contributions reveals that the energetic basis underlying the absence of inactivation in the F103A mutation in KcsA is the absence of the unfavorable steric interaction occurring with the large Ile100 side chain in a neighboring subunit when the intracellular gate is open and the selectivity filter is in a conductive conformation. Macroscopic current analysis shows that the I100A mutant indeed relieves inactivation in KcsA, but to a lesser extent than the F103A mutant.

  15. Evaluation Effects of Verapamil as a Calcium Channel Blocker on Acquisition, Consolidation and Retrieval of Memory in Mice

    OpenAIRE

    Nooshin Masoudian; Nahid Masoudian; Ali Rashidy Pour; Abbas Ali Vafaiee; Sasan Andalib; Golnaz Vaseghi

    2015-01-01

    Many factors are involved in learning and memory processes including brain nuclei, neurotransmitter systems, and the activity of ion channels. Studies showed inconsistent effects of calcium channel blockers on learning process, especially memory consolidation; however, little is known about their effect on memory acquisition and retrieval. Accordingly, the present study aimed to determine the effects of verapamil calcium channel antagonist as a representative of the phenylalkylamine group on ...

  16. The α2δ subunit and absence epilepsy: Beyond calcium channels?

    NARCIS (Netherlands)

    Celli, R.; Santolini, I.; Guiducci, M.; Luijtelaar, E.L.J.M. van; Parisi, P.; Striano, P.; Gradini, R.; Battaglia, G.; Ngomba, R.T.; Nicoletti, F.

    2017-01-01

    Spike-wave discharges, underlying absence seizures, are generated within a cortico-thalamo-cortical network that involves the somatosensory cortex, the reticular thalamic nucleus, and the ventrobasal thalamic nuclei. Activation of T-type voltage-sensitive calcium channels (VSCCs) contributes to the

  17. 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...... on vascular diameter in the afferent arteriole. We conclude that voltage-dependent L- and T-type calcium channels are expressed and of functional significance in renal cortical preglomerular vessels, in juxtamedullary efferent arterioles, and in outer medullary vasa recta, but not in cortical efferent...

  18. An expert protocol for immunofluorescent detection of calcium channels in tsA-201 cells.

    Science.gov (United States)

    Koch, Peter; Herzig, Stefan; Matthes, Jan

    Pore-forming subunits of voltage gated calcium channels (VGCC) are large membrane proteins (260kDa) containing 24 transmembrane domains. Despite transfection with viral promoter driven vectors, biochemical analysis of VGCC is often hampered by rather low expression levels in heterologous systems rendering VGCC challenging targets. Especially in immunofluorescent detection, calcium channels are demanding proteins. We provide an expert step-by-step protocol with adapted conditions for handling procedures (tsA-201 cell culture, transient transfection, incubation time and temperature at 28°C or 37°C and immunostaining) to address the L-type calcium-channel pore Ca v 1.2 in an immunofluorescent approach. We performed immunocytochemical analysis of Ca v 1.2 expression at single-cell level in combination with detection of different markers for cellular organelles. We show confluency levels and shapes of tsA-201 cells at different time points during an experiment. Our experiments reveal sufficient levels of Ca v 1.2 protein and a correct Ca v 1.2 expression pattern in polygonal shaped cells already 12h after transfection. A sequence of elaborated protocol modifications allows subcellular localization analysis of Ca v 1.2 in an immunocytochemical approach. We provide a protocol that may be used to achieve insights into physiological and pathophysiological processes involving voltage gated calcium channels. Our protocol may be used for expression analysis of other challenging proteins and efficient overexpression may be exploited in related biochemical techniques requiring immunolabels. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Homogeneous distribution of large-conductance calcium-dependent potassium channels on soma and apical dendrite of rat neocortical layer 5 pyramidal neurons.

    Science.gov (United States)

    Benhassine, Narimane; Berger, Thomas

    2005-02-01

    Voltage-gated conductances on dendrites of layer 5 pyramidal neurons participate in synaptic integration and output generation. We investigated the properties and the distribution of large-conductance calcium-activated potassium channels (BK channels) in this cell type using excised patches in acute slice preparations of rat somatosensory cortex. BK channels were characterized by their large conductance and sensitivity to the specific blockers paxilline and iberiotoxin. BK channels showed a pronounced calcium-dependence with a maximal opening probability of 0.69 at 10 microm and 0.42 at 3 microm free calcium. Their opening probability and transition time constants between open and closed states are voltage-dependent. At depolarized potentials, BK channel gating is described by two open and one closed states. Depolarization increases the opening probability due to a prolongation of the open time constant and a shortening of the closed time constant. Calcium-dependence and biophysical properties of somatic and dendritic BK channels were identical. The presence of BK channels on the apical dendrite of layer 5 pyramidal neurons was shown by immunofluorescence. Patch-clamp recordings revealed a homogeneous density of BK channels on the soma and along the apical dendrite up to 850 microm with a mean density of 1.9 channels per microm(2). BK channels are expressed either isolated or in clusters containing up to four channels. This study shows the presence of BK channels on dendrites. Their activation might modulate the shape of sodium and calcium action potentials, their propagation along the dendrite, and thereby the electrotonic distance between the somatic and dendritic action potential initiation zones.

  20. Dynamic transition on the seizure-like neuronal activity by astrocytic calcium channel block

    International Nuclear Information System (INIS)

    Li, Jiajia; Wang, Rong; Du, Mengmeng; Tang, Jun; Wu, Ying

    2016-01-01

    The involvement of astrocytes in neuronal firing dynamics is becoming increasingly evident. In this study, we used a classical hippocampal tripartite synapse model consisting of soma-dendrite coupled neuron models and a Hodgkin–Huxley-like astrocyte model, to investigate the seizure-like firing in the somatic neuron induced by the over-expressed neuronal N-methyl-d-aspartate (NMDA) receptors. Based on this model, we further investigated the effect of the astrocytic channel block on the neuronal firing through a bifurcation analysis. Results show that blocking inositol-1,4,5-triphosphate(IP3)-dependent calcium channel in astrocytes efficiently suppresses the astrocytic calcium oscillation, which in turn suppresses the seizure-like firing in the neuron.

  1. LRRK2 regulates voltage-gated calcium channel function.

    Directory of Open Access Journals (Sweden)

    Cade eBedford

    2016-05-01

    Full Text Available Voltage-gated Ca2+ (CaV channels enable Ca2+ influx in response to membrane depolarization. CaV2.1 channels are localized to the presynaptic membrane of many types of neurons where they are involved in triggering neurotransmitter release. Several signaling proteins have been identified as important CaV2.1 regulators including protein kinases, G-proteins and Ca2+ binding proteins. Recently, we discovered that leucine rich repeat kinase 2 (LRRK2, a protein associated with inherited Parkinson’s disease, interacts with specific synaptic proteins and influences synaptic transmission. Since synaptic proteins functionally interact with CaV2.1 channels and synaptic transmission is triggered by Ca2+ entry via CaV2.1, we investigated whether LRRK2 could impact CaV2.1 channel function. CaV2.1 channel properties were measured using whole cell patch clamp electrophysiology in HEK293 cells transfected with CaV2.1 subunits and various LRRK2 constructs. Our results demonstrate that both wild type LRRK2 and the G2019S LRRK2 mutant caused a significant increase in whole cell Ca2+ current density compared to cells expressing only the CaV2.1 channel complex. In addition, LRRK2 expression caused a significant hyperpolarizing shift in voltage-dependent activation while having no significant effect on inactivation properties. These functional changes in CaV2.1 activity are likely due to a direct action of LRRK2 as we detected a physical interaction between LRRK2 and the β3 CaV channel subunit via coimmunoprecipitation. Furthermore, effects on CaV2.1 channel function are dependent on LRRK2 kinase activity as these could be reversed via treatment with a LRRK2 inhibitor. Interestingly, LRRK2 also augmented endogenous voltage-gated Ca2+ channel function in PC12 cells suggesting other CaV channels could also be regulated by LRRK2. Overall, our findings support a novel physiological role for LRRK2 in regulating CaV2.1 function that could have implications for how

  2. [G-protein potentiates the activation of TNF-alpha on calcium-activated potassium channel in ECV304].

    Science.gov (United States)

    Lin, L; Zheng, Y; Qu, J; Bao, G

    2000-06-01

    Observe the effect of tumor necrosis factor-alpha (TNF-alpha) on calcium-activated potassium channel in ECV304 and the possible involvement of G-protein mediation in the action of TNF-alpha. Using the cell-attached configuration of patch clamp technique. (1) the activity of high-conductance calcium-activated potassium channel (BKca) was recorded. Its conductance is (202.54 +/- 16.62) pS; (2) the activity of BKca was potentiated by 200 U/ml TNF-alpha; (3) G-protein would intensify this TNF-alpha activation. TNF-alpha acted on vascular endothelial cell ECV304 could rapidly activate the activity of BKca. Opening of BKca resulted in membrane hyper-polarization which could increase electro-chemical gradient for the resting Ca2+ influx and open leakage calcium channel, thus resting cytoplasmic free Ca2+ concentration could be elevated. G-protein may exert an important regulation in this process.

  3. G Protein Regulation of Neuronal Calcium Channels: Back to the Future

    Czech Academy of Sciences Publication Activity Database

    Proft, Juliane; Weiss, Norbert

    2015-01-01

    Roč. 87, č. 6 (2015), s. 890-906 ISSN 0026-895X R&D Projects: GA ČR GA15-13556S Institutional support: RVO:61388963 Keywords : voltage gated calcium channels Cav * G proteins * GPCR Subject RIV: CE - Biochemistry Impact factor: 3.931, year: 2015

  4. Calcium channel blockers as the treatment of choice for hypertension in renal transplant recipients: fact or fiction.

    Science.gov (United States)

    Baroletti, Steven A; Gabardi, Steven; Magee, Colm C; Milford, Edgar L

    2003-06-01

    Posttransplantation hypertension has been identified as an independent risk factor for chronic allograft dysfunction and loss. Based on available morbidity and mortality data, posttransplantation hypertension must be identified and managed appropriately. During the past decade, calcium channel blockers have been recommended by some as the antihypertensive agents of choice in this population, because it was theorized that their vasodilatory effects would counteract the vasoconstrictive effects of the calcineurin inhibitors. With increasing data becoming available, reexamining the use of traditional antihypertensive agents, including diuretics and beta-blockers, or the newer agents, angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers, may be beneficial. Transplant clinicians must choose antihypertensive agents that will provide their patients with maximum benefit, from both a renal and a cardiovascular perspective. Beta-blockers, diuretics, and ACE inhibitors have all demonstrated significant benefit on morbidity and mortality in patients with cardiovascular disease. Calcium channel blockers have been shown to possess the ability to counteract cyclosporine-induced nephrotoxicity. When compared with beta-blockers, diuretics, and ACE inhibitors, however, the relative risk of cardiovascular events is increased with calcium channel blockers. With the long-term benefits of calcium channel blockers on the kidney unknown and a negative cardiovascular profile, these agents are best reserved as adjunctive therapy to beta-blockers, diuretics, and ACE inhibitors.

  5. High potency inhibition of hERG potassium channels by the sodium–calcium exchange inhibitor KB-R7943

    Science.gov (United States)

    Cheng, Hongwei; Zhang, Yihong; Du, Chunyun; Dempsey, Christopher E; Hancox, Jules C

    2012-01-01

    BACKGROUND AND PURPOSE KB-R7943 is an isothiourea derivative that is used widely as a pharmacological inhibitor of sodium–calcium exchange (NCX) in experiments on cardiac and other tissue types. This study investigated KB-R7943 inhibition of hERG (human ether-à-go-go-related gene) K+ channels that underpin the cardiac rapid delayed rectifier potassium current, IKr. EXPERIMENTAL APPROACH Whole-cell patch-clamp measurements were made of hERG current (IhERG) carried by wild-type or mutant hERG channels and of native rabbit ventricular IKr. Docking simulations utilized a hERG homology model built on a MthK-based template. KEY RESULTS KB-R7943 inhibited both IhERG and native IKr rapidly on membrane depolarization with IC50 values of ∼89 and ∼120 nM, respectively, for current tails at −40 mV following depolarizing voltage commands to +20 mV. Marked IhERG inhibition also occurred under ventricular action potential voltage clamp. IhERG inhibition by KB-R7943 exhibited both time- and voltage-dependence but showed no preference for inactivated over activated channels. Results of alanine mutagenesis and docking simulations indicate that KB-R7943 can bind to a pocket formed of the side chains of aromatic residues Y652 and F656, with the compound's nitrobenzyl group orientated towards the cytoplasmic side of the channel pore. The structurally related NCX inhibitor SN-6 also inhibited IhERG, but with a markedly reduced potency. CONCLUSIONS AND IMPLICATIONS KB-R7943 inhibits IhERG/IKr with a potency that exceeds that reported previously for acute cardiac NCX inhibition. Our results also support the feasibility of benzyloxyphenyl-containing NCX inhibitors with reduced potential, in comparison with KB-R7943, to inhibit hERG. PMID:21950687

  6. 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

    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...

  7. Computational model based approach to analysis ventricular arrhythmias: Effects of dysfunction calcium channels

    Science.gov (United States)

    Gulothungan, G.; Malathi, R.

    2018-04-01

    Disturbed sodium (Na+) and calcium (Ca2+) handling is known to be a major predisposing factor for life-threatening cardiac arrhythmias. Cardiac contractility in ventricular tissue is prominent by Ca2+ channels like voltage dependent Ca2+ channels, sodium-calcium exchanger (Na+-Ca2+x) and sacroplasmicrecticulum (SR) Ca2+ pump and leakage channels. Experimental and clinical possibilities for studying cardiac arrhythmias in human ventricular myocardium are very limited. Therefore, the use of alternative methods such as computer simulations is of great importance. Our aim of this article is to study the impact on action potential (AP) generation and propagation in single ventricular myocyte and ventricular tissue under different dysfunction Ca2+ channels condition. In enhanced activity of Na+-Ca2+x, single myocyte produces AP duration (APD90) and APD50 is significantly smaller (266 ms and 235 ms). Its Na+-Ca2+x current at depolarization is increases 60% from its normal level and repolarization current goes more negative (nonfailing= -0.28 pA/pF and failing= -0.47 pA/pF). Similarly, same enhanced activity of Na+-Ca2+x in 10 mm region of ventricular sheet, raises the plateau potential abruptly, which ultimately affects the diastolic repolarization. Compare with normal ventricular sheet region of 10 mm, 10% of ventricular sheet resting state is reduces and ventricular sheet at time 250 ms is goes to resting state very early. In hypertrophy condition, single myocyte produces APD90 and APD50 is worthy of attention smaller (232 mS and 198 ms). Its sodium-potassium (Na+-K+) pump current is 75% reduces from its control conditions (0.13 pA/pF). Hypertrophy condition, 50% of ventricular sheet is reduces to minimum plateau potential state, that starts the repolarization process very early and reduces the APD. In a single failing SR Ca2+ channels myocyte, recovery of Ca2+ concentration level in SR reduces upto 15% from its control myocytes. At time 290 ms, 70% of ventricular sheet

  8. The concentration of adrenaline and noradrenaline in the serum of dogs under the influence of calcium channels blockers

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    Milanović Tamara

    2015-01-01

    Full Text Available The most important characteristic of calcium channels is selective regulation of slow incoming stream of calcium into the cell tissue providing the slow increasement of action potential. Such tissues include smooth muscles of blood vessels, cardiocytes and heart noduses (AV and SA node. Different calcium antagonists have different effects on previous tissues due to their different chemical formula. Verapamile, Nifedipin and Diltiazem are the most frequently used of all. Their commonest characteristic is blocking the calcium channels causing vasodilatation of blood vessels as well as negative inotropic and chronotropic influence. By blocking the incoming calcium through slow channels of myofibrils of smooth muscles, the antagonists of calcium decrease the quantity of available calcium for contraction which causes vasodilatation. The most famous and most frequently used calcium antagonist is Verapamile. In terms of electrophysiology, Verapamile inhibits action potentials of heart noduses, especially the AV node, where the slow incoming of calcium is the most important for depolarization. Prolongation of the efective refractory period of SA node causes the heart frequency decreasement while prolongation of the effective refractory period of AV node slows down the work of chambers in case of flater and fibrillation of atriums. The molecules of calcium-bonding protein called kalmodulin are located in synaptic endings. Each kalmodulin can bond four calcium ions providing transfer into active calcium-kalmodulin complex which activates the kinase protein. Activated kinase protein starts the exocytosis of neurotransmitters into synaptic gap. Apart from activating kinase protein, calcium-kalmodulin complex also starts the activity of calcium pump presynaptic membrane which pumps calcium out of presynaptic ending stopping the further exocytosis of neurotransmitters into synaptic gap. Taking into consideration the fact that opening the calcium channels on

  9. A new candidate of calcium channel blocker in silico from Tectona grandis for treatment of gestational hypertension

    Science.gov (United States)

    Azizah, A.; Suselo, Y. H.; Muthmainah, M.; Indarto, D.

    2018-05-01

    Gestational Hypertension is one of the three main causes of maternal mortality in Indonesia. Nifedipine which blockes the Cav1.2 calcium channel has frequently been used to treat gestational hypertension. However the efficacy of nifedipine has not been established yet and the prevalence of gestational hypertension is still high (27.1 %). Indonesian herbal plants have potential to be developed as natural drugs. Molecular docking, a computational method, is very often used to depict interaction between molecules and target receptor This study was therefore to identify Indonesian herbal plants that could inhibit the calcium channel in silico. This was a bioinformatics study with molecular docking approach. Three-dimensional structure of human calcium channel Cav1.2 was determined by modelling with rabbit calcium channel (ID:5GJW) as template and using the SWISS MODEL software. Nifedipine was used as a standard ligand and obtained from ZINC database with the access code ZINC19594578. Active compounds of Indonesian herbal plants were registered in HerbalDB database and their molecular structure was obtained from PubChem. Binding affinity of human Cav1.2 model-ligand complexes were assesed using AutoDock Vina 1.1.2 software and visualization of molecular conformation used Chimera 1.10 and PyMol 1.3 softwares. The Lipinsky’s rules of five were used to determine active compounds which fullfilled drug criteria. The human Cav1-2 model had 72.35% sequence identity with rabbit Cav1.1. Nifedipine bound to the human Cav1.2 model with -2.1 kcal/mol binding affinity and had binding sites at Gln1060, Phe1129, Ser1132, and Ile1173 residues. A lower binding affinity was observed in 8 phytochemicals but only obtusifolin 2-glucoside (-2.2 kcal/mol) had similar binding sites as nifedipin did. In addition, obtusifolin 2-glucoside met the Lipinsky criteria and the molecule conformation was similar with nifedipine. From the HerbalDB database, obtusifolin 2-glucoside is found in Tectona

  10. Cooperative roles of glucose and asparagine-linked glycosylation in T-type calcium channel expression

    Czech Academy of Sciences Publication Activity Database

    Lazniewska, Joanna; Rzhepetskyy, Yuriy; Zhang, F. X.; Zamponi, G. W.; Weiss, Norbert

    2016-01-01

    Roč. 468, 11/12 (2016), s. 1837-1851 ISSN 0031-6768 R&D Projects: GA ČR GA15-13556S; GA MŠk 7AMB15FR015 Institutional support: RVO:61388963 Keywords : calcium channel * T-type channel * Ca(v)3.2 * glucose * N-glycosylation * trafficking Subject RIV: CE - Biochemistry Impact factor: 3.156, year: 2016

  11. Aluminium and hydrogen ions inhibit a mechanosensory calcium-selective cation channel

    Science.gov (United States)

    Ding, J. P.; Pickard, B. G.

    1993-01-01

    The tension-dependent activity of mechanosensory calcium-selective cation channels in excised plasmalemmal patches from onion bulb scale epidermis is modulated by pH in the physiologically meaningful range between 4.5 and 7.2. It is rapidly lowered by lowering pH and rapidly raised by raising pH. Channel activity is effectively inhibited by low levels of aluminium ions and activity can be partially restored by washing for a few minutes. We suggest that under normal conditions the sensitivity of the mechanosensory channels to pH of the wall free space plays important roles in regulation of plant activities such as growth. We further suggest that, when levels of acid and aluminium ions in the soil solution are high, they might inhibit similar sensory channels in cells of the root tip, thus contributing critically to the acid soil syndrome.

  12. Support for calcium channel gene defects in autism spectrum disorders

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    Lu Ake Tzu-Hui

    2012-12-01

    Full Text Available Abstract Background Alternation of synaptic homeostasis is a biological process whose disruption might predispose children to autism spectrum disorders (ASD. Calcium channel genes (CCG contribute to modulating neuronal function and evidence implicating CCG in ASD has been accumulating. We conducted a targeted association analysis of CCG using existing genome-wide association study (GWAS data and imputation methods in a combined sample of parent/affected child trios from two ASD family collections to explore this hypothesis. Methods A total of 2,176 single-nucleotide polymorphisms (SNP (703 genotyped and 1,473 imputed covering the genes that encode the α1 subunit proteins of 10 calcium channels were tested for association with ASD in a combined sample of 2,781 parent/affected child trios from 543 multiplex Caucasian ASD families from the Autism Genetics Resource Exchange (AGRE and 1,651 multiplex and simplex Caucasian ASD families from the Autism Genome Project (AGP. SNP imputation using IMPUTE2 and a combined reference panel from the HapMap3 and the 1,000 Genomes Project increased coverage density of the CCG. Family-based association was tested using the FBAT software which controls for population stratification and accounts for the non-independence of siblings within multiplex families. The level of significance for association was set at 2.3E-05, providing a Bonferroni correction for this targeted 10-gene panel. Results Four SNPs in three CCGs were associated with ASD. One, rs10848653, is located in CACNA1C, a gene in which rare de novo mutations are responsible for Timothy syndrome, a Mendelian disorder that features ASD. Two others, rs198538 and rs198545, located in CACN1G, and a fourth, rs5750860, located in CACNA1I, are in CCGs that encode T-type calcium channels, genes with previous ASD associations. Conclusions These associations support a role for common CCG SNPs in ASD.

  13. Functional importance of T-type voltage-gated calcium channels in the cardiovascular and renal system

    DEFF Research Database (Denmark)

    Hansen, Pernille B L

    2015-01-01

    Over the years, it has been discussed whether T-type calcium channels Cav3 play a role in the cardiovascular and renal system. T-type channels have been reported to play an important role in renal hemodynamics, contractility of resistance vessels, and pacemaker activity in the heart. However...

  14. Clofazimine inhibits human Kv1.3 potassium channel by perturbing calcium oscillation in T lymphocytes.

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    Yunzhao R Ren

    Full Text Available The Kv1.3 potassium channel plays an essential role in effector memory T cells and has been implicated in several important autoimmune diseases including multiple sclerosis, psoriasis and type 1 diabetes. A number of potent small molecule inhibitors of Kv1.3 channel have been reported, some of which were found to be effective in various animal models of autoimmune diseases. We report herein the identification of clofazimine, a known anti-mycobacterial drug, as a novel inhibitor of human Kv1.3. Clofazimine was initially identified as an inhibitor of intracellular T cell receptor-mediated signaling leading to the transcriptional activation of human interleukin-2 gene in T cells from a screen of the Johns Hopkins Drug Library. A systematic mechanistic deconvolution revealed that clofazimine selectively blocked the Kv1.3 channel activity, perturbing the oscillation frequency of the calcium-release activated calcium channel, which in turn led to the inhibition of the calcineurin-NFAT signaling pathway. These effects of clofazimine provide the first line of experimental evidence in support of a causal relationship between Kv1.3 and calcium oscillation in human T cells. Furthermore, clofazimine was found to be effective in blocking human T cell-mediated skin graft rejection in an animal model in vivo. Together, these results suggest that clofazimine is a promising immunomodulatory drug candidate for treating a variety of autoimmune disorders.

  15. Calcium channel modulation as a target in chronic pain control.

    Science.gov (United States)

    Patel, Ryan; Montagut-Bordas, Carlota; Dickenson, Anthony H

    2018-06-01

    Neuropathic pain remains poorly treated for large numbers of patients, and little progress has been made in developing novel classes of analgesics. To redress this issue, ziconotide (Prialt™) was developed and approved as a first-in-class synthetic version of ω-conotoxin MVIIA, a peptide blocker of Ca v 2.2 channels. Unfortunately, the impracticalities of intrathecal delivery, low therapeutic index and severe neurological side effects associated with ziconotide have restricted its use to exceptional circumstances. Ziconotide exhibits no state or use-dependent block of Ca v 2.2 channels; activation state-dependent blockers were hypothesized to circumvent the side effects of state-independent blockers by selectively targeting high-frequency firing of nociceptive neurones in chronic pain states, thus alleviating aberrant pain but not affecting normal sensory transduction. Unfortunately, numerous drugs, including state-dependent calcium channel blockers, have displayed efficacy in preclinical models but have subsequently been disappointing in clinical trials. In recent years, it has become more widely acknowledged that trans-aetiological sensory profiles exist amongst chronic pain patients and may indicate similar underlying mechanisms and drug sensitivities. Heterogeneity amongst patients, a reliance on stimulus-evoked endpoints in preclinical studies and a failure to utilize translatable endpoints, all are likely to have contributed to negative clinical trial results. We provide an overview of how electrophysiological and operant-based assays provide insight into sensory and affective aspects of pain in animal models and how these may relate to chronic pain patients in order to improve the bench-to-bedside translation of calcium channel modulators. This article is part of a themed section on Recent Advances in Targeting Ion Channels to Treat Chronic Pain. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175

  16. Oxidized Low-density Lipoprotein (ox-LDL) Cholesterol Induces the Expression of miRNA-223 and L-type Calcium Channel Protein in Atrial Fibrillation

    Science.gov (United States)

    He, Fengping; Xu, Xin; Yuan, Shuguo; Tan, Liangqiu; Gao, Lingjun; Ma, Shaochun; Zhang, Shebin; Ma, Zhanzhong; Jiang, Wei; Liu, Fenglian; Chen, Baofeng; Zhang, Beibei; Pang, Jungang; Huang, Xiuyan; Weng, Jiaqiang

    2016-08-01

    Atrial fibrillation (AF) is the most common sustained arrhythmia causing high morbidity and mortality. While changing of the cellular calcium homeostasis plays a critical role in AF, the L-type calcium channel α1c protein has suggested as an important regulator of reentrant spiral dynamics and is a major component of AF-related electrical remodeling. Our computational modeling predicted that miRNA-223 may regulate the CACNA1C gene which encodes the cardiac L-type calcium channel α1c subunit. We found that oxidized low-density lipoprotein (ox-LDL) cholesterol significantly up-regulates both the expression of miRNA-223 and L-type calcium channel protein. In contrast, knockdown of miRNA-223 reduced L-type calcium channel protein expression, while genetic knockdown of endogenous miRNA-223 dampened AF vulnerability. Transfection of miRNA-223 by adenovirus-mediated expression enhanced L-type calcium currents and promoted AF in mice while co-injection of a CACNA1C-specific miR-mimic counteracted the effect. Taken together, ox-LDL, as a known factor in AF-associated remodeling, positively regulates miRNA-223 transcription and L-type calcium channel protein expression. Our results implicate a new molecular mechanism for AF in which miRNA-223 can be used as an biomarker of AF rheumatic heart disease.

  17. Fragile X mental retardation protein controls synaptic vesicle exocytosis by modulating N-type calcium channel density

    Science.gov (United States)

    Ferron, Laurent; Nieto-Rostro, Manuela; Cassidy, John S.; Dolphin, Annette C.

    2014-04-01

    Fragile X syndrome (FXS), the most common heritable form of mental retardation, is characterized by synaptic dysfunction. Synaptic transmission depends critically on presynaptic calcium entry via voltage-gated calcium (CaV) channels. Here we show that the functional expression of neuronal N-type CaV channels (CaV2.2) is regulated by fragile X mental retardation protein (FMRP). We find that FMRP knockdown in dorsal root ganglion neurons increases CaV channel density in somata and in presynaptic terminals. We then show that FMRP controls CaV2.2 surface expression by targeting the channels to the proteasome for degradation. The interaction between FMRP and CaV2.2 occurs between the carboxy-terminal domain of FMRP and domains of CaV2.2 known to interact with the neurotransmitter release machinery. Finally, we show that FMRP controls synaptic exocytosis via CaV2.2 channels. Our data indicate that FMRP is a potent regulator of presynaptic activity, and its loss is likely to contribute to synaptic dysfunction in FXS.

  18. New Conotoxin SO-3 Targeting N-type Voltage-Sensitive Calcium Channels

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    Lei Wen

    2006-04-01

    Full Text Available Selective blockers of the N-type voltage-sensitive calcium (CaV channels are useful in the management of severe chronic pain. Here, the structure and function characteristics of a novel N-type CaV channel blocker, SO-3, are reviewed. SO-3 is a 25-amino acid conopeptide originally derived from the venom of Conus striatus, and contains the same 4-loop, 6-cysteine framework (C-C-CC-C-C as O-superfamily conotoxins. The synthetic SO-3 has high analgesic activity similar to ω-conotoxin MVIIA (MVIIA, a selective N-type CaV channel blocker approved in the USA and Europe for the alleviation of persistent pain states. In electrophysiological studies, SO-3 shows more selectivity towards the N-type CaV channels than MVIIA. The dissimilarity between SO-3 and MVIIA in the primary and tertiary structures is further discussed in an attempt to illustrate the difference in selectivity of SO-3 and MVIIA towards N-type CaV channels.

  19. Direct evidence that scorpion α-toxins (site-3 modulate sodium channel inactivation by hindrance of voltage-sensor movements.

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    Zhongming Ma

    Full Text Available The position of the voltage-sensing transmembrane segment, S4, in voltage-gated ion channels as a function of voltage remains incompletely elucidated. Site-3 toxins bind primarily to the extracellular loops connecting transmembrane helical segments S1-S2 and S3-S4 in Domain 4 (D4 and S5-S6 in Domain 1 (D1 and slow fast-inactivation of voltage-gated sodium channels. As S4 of the human skeletal muscle voltage-gated sodium channel, hNav1.4, moves in response to depolarization from the resting to the inactivated state, two D4S4 reporters (R2C and R3C, Arg1451Cys and Arg1454Cys, respectively move from internal to external positions as deduced by reactivity to internally or externally applied sulfhydryl group reagents, methane thiosulfonates (MTS. The changes in reporter reactivity, when cycling rapidly between hyperpolarized and depolarized voltages, enabled determination of the positions of the D4 voltage-sensor and of its rate of movement. Scorpion α-toxin binding impedes D4S4 segment movement during inactivation since the modification rates of R3C in hNav1.4 with methanethiosulfonate (CH3SO2SCH2CH2R, where R = -N(CH33 (+ trimethylammonium, MTSET and benzophenone-4-carboxamidocysteine methanethiosulfonate (BPMTS were slowed ~10-fold in toxin-modified channels. Based upon the different size, hydrophobicity and charge of the two reagents it is unlikely that the change in reactivity is due to direct or indirect blockage of access of this site to reagent in the presence of toxin (Tx, but rather is the result of inability of this segment to move outward to the normal extent and at the normal rate in the toxin-modified channel. Measurements of availability of R3C to internally applied reagent show decreased access (slower rates of thiol reaction providing further evidence for encumbered D4S4 movement in the presence of toxins consistent with the assignment of at least part of the toxin binding site to the region of D4S4 region of the voltage

  20. TRPP2 and TRPV4 form an EGF-activated calcium permeable channel at the apical membrane of renal collecting duct cells.

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    Zhi-Ren Zhang

    Full Text Available Regulation of apical calcium entry is important for the function of principal cells of the collecting duct. However, the molecular identity and the regulators of the transporter/channel, which is responsible for apical calcium entry and what factors regulate the calcium conduction remain unclear.We report that endogenous TRPP2 and TRPV4 assemble to form a 23-pS divalent cation-permeable non-selective ion channel at the apical membrane of renal principal cells of the collecting duct. TRPP2\\TRPV4 channel complex was identified by patch-clamp, immunofluorescence and co-immunprecipitation studies in both principal cells that either possess normal cilia (cilia (+ or in which cilia are absent (cilia (-. This channel has distinct biophysical and pharmacological and regulatory profiles compared to either TRPP2 or TRPV4 channels. The rate of occurrence detected by patch clamp was higher in cilia (- compared to cilia (+ cells. In addition, shRNA knockdown of TRPP2 increased the prevalence of TRPV4 channel activity while knockdown of TRPV4 resulted in TRPP2 activity and knockdown of both proteins vastly decreased the 23-pS channel activity. Epidermal growth factor (EGF stimulated TRPP2\\TRPV4 channel through the EGF receptor (EGFR tyrosine kinase-dependent signaling. With loss of cilia, apical EGF treatment resulted in 64-fold increase in channel activity in cilia (- but not cilia (+ cells. In addition EGF increased cell proliferation in cilia (- cell that was dependent upon TRPP2\\TRPV4 channel mediated increase in intracellular calcium.We conclude that in the absence of cilia, an EGF activated TRPP2\\TRPV4 channel may play an important role in increased cell proliferation and cystogenesis.

  1. Voltage-Gated Calcium Channel Antagonists and Traumatic Brain Injury

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    Bruce Lyeth

    2013-06-01

    Full Text Available Traumatic brain injury (TBI is a leading cause of death and disability in the United States. Despite more than 30 years of research, no pharmacological agents have been identified that improve neurological function following TBI. However, several lines of research described in this review provide support for further development of voltage gated calcium channel (VGCC antagonists as potential therapeutic agents. Following TBI, neurons and astrocytes experience a rapid and sometimes enduring increase in intracellular calcium ([Ca2+]i. These fluxes in [Ca2+]i drive not only apoptotic and necrotic cell death, but also can lead to long-term cell dysfunction in surviving cells. In a limited number of in vitro experiments, both L-type and N-type VGCC antagonists successfully reduced calcium loads as well as neuronal and astrocytic cell death following mechanical injury. In rodent models of TBI, administration of VGCC antagonists reduced cell death and improved cognitive function. It is clear that there is a critical need to find effective therapeutics and rational drug delivery strategies for the management and treatment of TBI, and we believe that further investigation of VGCC antagonists should be pursued before ruling out the possibility of successful translation to the clinic.

  2. Modulation of Ca(v)3.2 T-type calcium channel permeability by asparagine-linked glycosylation

    Czech Academy of Sciences Publication Activity Database

    Ondáčová, K.; Karmažínová, M.; Lazniewska, Joanna; Weiss, Norbert; Lacinová, L.

    2016-01-01

    Roč. 10, č. 3 (2016), s. 175-184 ISSN 1933-6950 R&D Projects: GA ČR GA15-13556S; GA MŠk 7AMB15FR015 Institutional support: RVO:61388963 Keywords : calcium channel * Ca(v)3.2 * gating * glycosylation * T-type channel Subject RIV: CE - Biochemistry Impact factor: 2.042, year: 2016

  3. Surfen is a broad-spectrum calcium channel inhibitor with analgesic properties in mouse models of acute and chronic inflammatory pain

    Czech Academy of Sciences Publication Activity Database

    Rivas-Ramirez, Paula; Gadotti, V. M.; Zamponi, G. W.; Weiss, Norbert

    2017-01-01

    Roč. 469, č. 10 (2017), s. 1325-1334 ISSN 0031-6768 R&D Projects: GA ČR GA15-13556S; GA MŠk 7AMB15FR015 Institutional support: RVO:61388963 Keywords : calcium channel * pain * inflammatory pain * calcium channel blocker * surfen * DRG neuron Subject RIV: ED - Physiology OBOR OECD: Physiology (including cytology) Impact factor: 3.156, year: 2016

  4. Molecular, biophysical, and pharmacological properties of calcium-activated chloride channels.

    Science.gov (United States)

    Kamaleddin, Mohammad Amin

    2018-02-01

    Calcium-activated chloride channels (CaCCs) are a family of anionic transmembrane ion channels. They are mainly responsible for the movement of Cl - and other anions across the biological membranes, and they are widely expressed in different tissues. Since the Cl - flow into or out of the cell plays a crucial role in hyperpolarizing or depolarizing the cells, respectively, the impact of intracellular Ca 2+ concentration on these channels is attracting a lot of attentions. After summarizing the molecular, biophysical, and pharmacological properties of CaCCs, the role of CaCCs in normal cellular functions will be discussed, and I will emphasize how dysregulation of CaCCs in pathological conditions can account for different diseases. A better understanding of CaCCs and a pivotal regulatory role of Ca 2+ can shed more light on the therapeutic strategies for different neurological disorders that arise from chloride dysregulation, such as asthma, cystic fibrosis, and neuropathic pain. © 2017 Wiley Periodicals, Inc.

  5. A Recurrent Mutation in CACNA1G Alters Cav3.1 T-Type Calcium-Channel Conduction and Causes Autosomal-Dominant Cerebellar Ataxia

    Science.gov (United States)

    Coutelier, Marie; Blesneac, Iulia; Monteil, Arnaud; Monin, Marie-Lorraine; Ando, Kunie; Mundwiller, Emeline; Brusco, Alfredo; Le Ber, Isabelle; Anheim, Mathieu; Castrioto, Anna; Duyckaerts, Charles; Brice, Alexis; Durr, Alexandra; Lory, Philippe; Stevanin, Giovanni

    2015-01-01

    Hereditary cerebellar ataxias (CAs) are neurodegenerative disorders clinically characterized by a cerebellar syndrome, often accompanied by other neurological or non-neurological signs. All transmission modes have been described. In autosomal-dominant CA (ADCA), mutations in more than 30 genes are implicated, but the molecular diagnosis remains unknown in about 40% of cases. Implication of ion channels has long been an ongoing topic in the genetics of CA, and mutations in several channel genes have been recently connected to ADCA. In a large family affected by ADCA and mild pyramidal signs, we searched for the causative variant by combining linkage analysis and whole-exome sequencing. In CACNA1G, we identified a c.5144G>A mutation, causing an arginine-to-histidine (p.Arg1715His) change in the voltage sensor S4 segment of the T-type channel protein Cav3.1. Two out of 479 index subjects screened subsequently harbored the same mutation. We performed electrophysiological experiments in HEK293T cells to compare the properties of the p.Arg1715His and wild-type Cav3.1 channels. The current-voltage and the steady-state activation curves of the p.Arg1715His channel were shifted positively, whereas the inactivation curve had a higher slope factor. Computer modeling in deep cerebellar nuclei (DCN) neurons suggested that the mutation results in decreased neuronal excitability. Taken together, these data establish CACNA1G, which is highly expressed in the cerebellum, as a gene whose mutations can cause ADCA. This is consistent with the neuropathological examination, which showed severe Purkinje cell loss. Our study further extends our knowledge of the link between calcium channelopathies and CAs. PMID:26456284

  6. L-type calcium channels refine the neural population code of sound level

    Science.gov (United States)

    Grimsley, Calum Alex; Green, David Brian

    2016-01-01

    The coding of sound level by ensembles of neurons improves the accuracy with which listeners identify how loud a sound is. In the auditory system, the rate at which neurons fire in response to changes in sound level is shaped by local networks. Voltage-gated conductances alter local output by regulating neuronal firing, but their role in modulating responses to sound level is unclear. We tested the effects of L-type calcium channels (CaL: CaV1.1–1.4) on sound-level coding in the central nucleus of the inferior colliculus (ICC) in the auditory midbrain. We characterized the contribution of CaL to the total calcium current in brain slices and then examined its effects on rate-level functions (RLFs) in vivo using single-unit recordings in awake mice. CaL is a high-threshold current and comprises ∼50% of the total calcium current in ICC neurons. In vivo, CaL activates at sound levels that evoke high firing rates. In RLFs that increase monotonically with sound level, CaL boosts spike rates at high sound levels and increases the maximum firing rate achieved. In different populations of RLFs that change nonmonotonically with sound level, CaL either suppresses or enhances firing at sound levels that evoke maximum firing. CaL multiplies the gain of monotonic RLFs with dynamic range and divides the gain of nonmonotonic RLFs with the width of the RLF. These results suggest that a single broad class of calcium channels activates enhancing and suppressing local circuits to regulate the sensitivity of neuronal populations to sound level. PMID:27605536

  7. CNTF-ACM promotes mitochondrial respiration and oxidative stress in cortical neurons through upregulating L-type calcium channel activity.

    Science.gov (United States)

    Sun, Meiqun; Liu, Hongli; Xu, Huanbai; Wang, Hongtao; Wang, Xiaojing

    2016-09-01

    A specialized culture medium termed ciliary neurotrophic factor-treated astrocyte-conditioned medium (CNTF-ACM) allows investigators to assess the peripheral effects of CNTF-induced activated astrocytes upon cultured neurons. CNTF-ACM has been shown to upregulate neuronal L-type calcium channel current activity, which has been previously linked to changes in mitochondrial respiration and oxidative stress. Therefore, the aim of this study was to evaluate CNTF-ACM's effects upon mitochondrial respiration and oxidative stress in rat cortical neurons. Cortical neurons, CNTF-ACM, and untreated control astrocyte-conditioned medium (UC-ACM) were prepared from neonatal Sprague-Dawley rat cortical tissue. Neurons were cultured in either CNTF-ACM or UC-ACM for a 48-h period. Changes in the following parameters before and after treatment with the L-type calcium channel blocker isradipine were assessed: (i) intracellular calcium levels, (ii) mitochondrial membrane potential (ΔΨm), (iii) oxygen consumption rate (OCR) and adenosine triphosphate (ATP) formation, (iv) intracellular nitric oxide (NO) levels, (v) mitochondrial reactive oxygen species (ROS) production, and (vi) susceptibility to the mitochondrial complex I toxin rotenone. CNTF-ACM neurons displayed the following significant changes relative to UC-ACM neurons: (i) increased intracellular calcium levels (p ACM (p ACM promotes mitochondrial respiration and oxidative stress in cortical neurons through elevating L-type calcium channel activity.

  8. Citalopram inhibits L-type calcium channel current in rat cardiomyocytes in culture

    Czech Academy of Sciences Publication Activity Database

    Hamplová-Peichlová, J.; Krůšek, Jan; Paclt, I.; Slavíček, J.; Lisá, Věra; Vyskočil, František

    2002-01-01

    Roč. 51, č. 3 (2002), s. 317-321 ISSN 0862-8408 R&D Projects: GA AV ČR IAA7011902; GA ČR GA305/02/1333 Institutional research plan: CEZ:AV0Z5011922 Keywords : citalopram * amitriptyline * L-type calcium channel current Subject RIV: ED - Physiology Impact factor: 0.984, year: 2002

  9. The role of L-type calcium channels in the vascular effect of Trigonella foenum-graecum L. in diabetic rats

    Directory of Open Access Journals (Sweden)

    Mehrdad Roghani

    2006-03-01

    Full Text Available Some ion channels like voltage-operated calcium channels (VOCC within the plasma membrane of vascular muscle cells from the walls of resistance arteries and arterioles play a central role in the regulation of vascular tone. On the basis of reports about the beneficial attenuating effect of fenugreek (Trigonella foenum-graecum L.; TFG on the contractile reactivity of aortic rings of diabetic rats, this study was carried out to evaluate the possible involvement of L-type voltage-operated calcium channels in the vascular effect of this medicinal plant. For this purpose, male Wistar rats were made diabetic using streptozotocin (STZ, 60 mg/Kg, i.p. The extract-treated control and diabetic rats received aqueous leaf extract of TFG (200 mg/Kg, i.p. every other day for two months. At the end of the study, contractile response of isolated aortic rings to KCl and noreadrenaline (NA was determined in the absence and presence of the calcium channel blocker nifedipine. The results showed that aortic rings from diabetic rats are more responsive to the effect of KCl and NA than those of controls, TFG extract treatment could attenuate the enhanced contractile response of aortic rings of diabetic rats, and nifedipine pretreatment could partially neutralize the beneficial effect of this extract. It is concluded that TFG extract attenuates the enhanced vascular reactivity in chronic diabetic rats and voltage-operated calcium channels are in part responsible for this effect of TFG extract.

  10. A combined role of calcium channel blockers and angiotensin receptor blockers in stroke prevention

    Directory of Open Access Journals (Sweden)

    Ji-Guang Wang

    2009-07-01

    Full Text Available Ji-Guang WangCentre for Epidemiological Studies and Clinical Trials, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, ChinaAbstract: Stroke is a leading cause of death and disability worldwide. The importance of lowering blood pressure for reducing the risk of stroke is well established. However, not all the benefits of antihypertensive treatments in stroke can be accounted for by reductions in BP and there may be differences between antihypertensive classes as to which provides optimal protection. Dihydropyridine calcium channel blockers, such as amlodipine, and angiotensin receptor blockers, such as valsartan, represent the two antihypertensive drug classes with the strongest supportive data for the prevention of stroke. Therefore, when combination therapy is required, a combination of these two antihypertensive classes represents a logical approach.Keywords: stroke, angiotensin, calcium channel, cerebrovascular, hypertension, blood pressure

  11. Clinical features of neuromuscular disorders in patients with N-type voltage-gated calcium channel antibodies

    Directory of Open Access Journals (Sweden)

    Andreas Totzeck

    2016-09-01

    Full Text Available Neuromuscular junction disorders affect the pre- or postsynaptic nerve to muscle transmission due to autoimmune antibodies. Members of the group like myasthenia gravis and Lambert-Eaton syndrome have pathophysiologically distinct characteristics. However, in practice, distinction may be difficult. We present a series of three patients with a myasthenic syndrome, dropped-head syndrome, bulbar and respiratory muscle weakness and positive testing for anti-N-type voltage-gated calcium channel antibodies. In two cases anti-acetylcholin receptor antibodies were elevated, anti-P/Q-type voltage-gated calcium channel antibodies were negative. All patients initially responded to pyridostigmine with a non-response in the course of the disease. While one patient recovered well after treatment with intravenous immunoglobulins, 3,4-diaminopyridine, steroids and later on immunosuppression with mycophenolate mofetil, a second died after restriction of treatment due to unfavorable cancer diagnosis, the third patient declined treatment. Although new antibodies causing neuromuscular disorders were discovered, clinical distinction has not yet been made. Our patients showed features of pre- and postsynaptic myasthenic syndrome as well as severe dropped-head syndrome and bulbar and axial muscle weakness, but only anti-N-type voltage-gated calcium channel antibodies were positive. When administered, one patient benefited from 3,4-diaminopyridine. We suggest that this overlap-syndrome should be considered especially in patients with assumed seronegative myasthenia gravis and lack of improvement under standard therapy.

  12. 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

  13. The genetic background affects the vascular response in T-type calcium channels 3.2 deficient mice

    DEFF Research Database (Denmark)

    Svenningsen, Per; Hansen, Pernille B L

    2016-01-01

    -type channels are the dominant Ca(2+) entry pathway in vascular smooth muscle cells, however, T-type calcium channels are also expressed in the cardiovascular system where they play a functional role in the regulation of both contraction and vasodilation in (Chen et al. 2003; Hansen et al. 2001). This article...... is protected by copyright. All rights reserved....

  14. Association of calcium sensing receptor polymorphisms at rs1801725 with circulating calcium in breast cancer patients.

    Science.gov (United States)

    Wang, Li; Widatalla, Sarrah E; Whalen, Diva S; Ochieng, Josiah; Sakwe, Amos M

    2017-08-02

    Breast cancer (BC) patients with late-stage and/or rapidly growing tumors are prone to develop high serum calcium levels which have been shown to be associated with larger and aggressive breast tumors in post and premenopausal women respectively. Given the pivotal role of the calcium sensing receptor (CaSR) in calcium homeostasis, we evaluated whether polymorphisms of the CASR gene at rs1801725 and rs1801726 SNPs in exon 7, are associated with circulating calcium levels in African American and Caucasian control subjects and BC cases. In this retrospective case-control study, we assessed the mean circulating calcium levels, the distribution of two inactivating CaSR SNPs at rs1801725 and rs1801726 in 199 cases and 384 age-matched controls, and used multivariable regression analysis to determine whether these SNPs are associated with circulating calcium in control subjects and BC cases. We found that the mean circulating calcium levels in African American subjects were higher than those in Caucasian subjects (p calcium levels were higher in BC cases compared to control subjects (p calcium levels in BC patients were independent of race. We also show that in BC cases and control subjects, the major alleles at rs1801725 (G/T, A986S) and at rs1801726 (C/G, Q1011E) were common among Caucasians and African Americans respectively. Compared to the wild type alleles, polymorphisms at the rs1801725 SNP were associated with higher calcium levels (p = 0.006) while those at rs1801726 were not. Using multivariable linear mixed-effects models and adjusting for age and race, we show that circulating calcium levels in BC cases were associated with tumor grade (p = 0.009), clinical stage (p = 0.003) and more importantly, with inactivating mutations of the CASR at the rs1801725 SNP (p = 0.038). These data suggest that decreased sensitivity of the CaSR to calcium due to inactivating polymorphisms at rs1801725, may predispose up to 20% of BC cases to high circulating calcium

  15. Interplay of Plasma Membrane and Vacuolar Ion Channels, Together with BAK1, Elicits Rapid Cytosolic Calcium Elevations in Arabidopsis during Aphid Feeding[OPEN

    Science.gov (United States)

    Vincent, Thomas R.; Avramova, Marieta; Canham, James; Higgins, Peter; Bilkey, Natasha; Mugford, Sam T.; Pitino, Marco; Toyota, Masatsugu

    2017-01-01

    A transient rise in cytosolic calcium ion concentration is one of the main signals used by plants in perception of their environment. The role of calcium in the detection of abiotic stress is well documented; however, its role during biotic interactions remains unclear. Here, we use a fluorescent calcium biosensor (GCaMP3) in combination with the green peach aphid (Myzus persicae) as a tool to study Arabidopsis thaliana calcium dynamics in vivo and in real time during a live biotic interaction. We demonstrate rapid and highly localized plant calcium elevations around the feeding sites of M. persicae, and by monitoring aphid feeding behavior electrophysiologically, we demonstrate that these elevations correlate with aphid probing of epidermal and mesophyll cells. Furthermore, we dissect the molecular mechanisms involved, showing that interplay between the plant defense coreceptor BRASSINOSTEROID INSENSITIVE-ASSOCIATED KINASE1 (BAK1), the plasma membrane ion channels GLUTAMATE RECEPTOR-LIKE 3.3 and 3.6 (GLR3.3 and GLR3.6), and the vacuolar ion channel TWO-PORE CHANNEL1 (TPC1) mediate these calcium elevations. Consequently, we identify a link between plant perception of biotic threats by BAK1, cellular calcium entry mediated by GLRs, and intracellular calcium release by TPC1 during a biologically relevant interaction. PMID:28559475

  16. Calcium-Activated Cl- Channel: Insights on the Molecular Identity in Epithelial Tissues.

    Science.gov (United States)

    Rottgen, Trey S; Nickerson, Andrew J; Rajendran, Vazhaikkurichi M

    2018-05-10

    Calcium-activated chloride secretion in epithelial tissues has been described for many years. However, the molecular identity of the channel responsible for the Ca 2+ -activated Cl − secretion in epithelial tissues has remained a mystery. More recently, TMEM16A has been identified as a new putative Ca 2+ -activated Cl − channel (CaCC). The primary goal of this article will be to review the characterization of TMEM16A, as it relates to the physical structure of the channel, as well as important residues that confer voltage and Ca 2+ -sensitivity of the channel. This review will also discuss the role of TMEM16A in epithelial physiology and potential associated-pathophysiology. This will include discussion of developed knockout models that have provided much needed insight on the functional localization of TMEM16A in several epithelial tissues. Finally, this review will examine the implications of the identification of TMEM16A as it pertains to potential novel therapies in several pathologies.

  17. Spontaneous and CRH-Induced Excitability and Calcium Signaling in Mice Corticotrophs Involves Sodium, Calcium, and Cation-Conducting Channels

    Czech Academy of Sciences Publication Activity Database

    Zemková, Hana; Tomič, M.; Kučka, M.; Aguilera, G.; Stojilkovic, S. S.

    2016-01-01

    Roč. 157, č. 4 (2016), s. 1576-1589 ISSN 0013-7227 R&D Projects: GA ČR(CZ) GBP304/12/G069; GA MŠk(CZ) LQ1604; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:67985823 Keywords : action potential * background sodium conductance * bursting activity * cation -conducting channels * cytosolic calcium concentration * resting membrane potential Subject RIV: FB - Endocrinology, Diabetology, Metabolism, Nutrition Impact factor: 4.286, year: 2016

  18. MarkoLAB: A simulator to study ionic channel's stochastic behavior.

    Science.gov (United States)

    da Silva, Robson Rodrigues; Goroso, Daniel Gustavo; Bers, Donald M; Puglisi, José Luis

    2017-08-01

    Mathematical models of the cardiac cell have started to include markovian representations of the ionic channels instead of the traditional Hodgkin & Huxley formulations. There are many reasons for this: Markov models are not restricted to the idea of independent gates defining the channel, they allow more complex description with specific transitions between open, closed or inactivated states, and more importantly those states can be closely related to the underlying channel structure and conformational changes. We used the LabVIEW ® and MATLAB ® programs to implement the simulator MarkoLAB that allow a dynamical 3D representation of the markovian model of the channel. The Monte Carlo simulation was used to implement the stochastic transitions among states. The user can specify the voltage protocol by setting the holding potential, the step-to voltage and the duration of the stimuli. The most studied feature of a channel is the current flowing through it. This happens when the channel stays in the open state, but most of the time, as revealed by the low open probability values, the channel remains on the inactive or closed states. By focusing only when the channel enters or leaves the open state we are missing most of its activity. MarkoLAB proved to be quite useful to visualize the whole behavior of the channel and not only when the channel produces a current. Such dynamic representation provides more complete information about channel kinetics and will be a powerful tool to demonstrate the effect of gene mutations or drugs on the channel function. MarkoLAB provides an original way of visualizing the stochastic behavior of a channel. It clarifies concepts, such as recovery from inactivation, calcium- versus voltage-dependent inactivation, and tail currents. It is not restricted to ionic channels only but it can be extended to other transporters, such as exchangers and pumps. This program is intended as a didactical tool to illustrate the dynamical behavior of a

  19. Receptor model for the molecular basis of tissue selectivity of 1,4-dihydropyridine calcium channel drugs

    Science.gov (United States)

    Langs, David A.; Strong, Phyllis D.; Triggle, David J.

    1990-09-01

    Our analysis of the solid state conformations of nifedipine [dimethyl 1,4-dihydro-2,6-dimethyl-4-(2-nitrophenyl)-3,5-pyridinecarboxylate] and its 1,4-dihydropyridine (1,4-DHP) analogues produced a cartoon description of the important interactions between these drugs and their voltage-dependent calcium channel receptor. In the present study a molecular-level detailed model of the 1,4-DHP receptor binding site has been built from the published amino acid sequence of the 215-1 subunit of the voltage-dependent calcium channel isolated from rabbit skeletal muscle transverse tubule membranes. The voltage-sensing component of the channel described in this work differs from others reported for the homologous sodium channel in that it incorporates a water structure and a staggered, rather than eclipsed, hydrogen bonded S4 helix conformation. The major recognition surfaces of the receptor lie in helical grooves on the S4 or voltagesensing α-helix that is positioned in the center of the bundle of transmembrane helices that define each of the four calcium channel domains. Multiple binding clefts defined by Arg-X-X-Arg-P-X-X-S `reading frames' exist on the S4 strand. The tissue selectivity of nifedipine and its analogues may arise, in part, from conservative changes in the amino acid residues at the P and S positions of the reading frame that define the ester-binding regions of receptors from different tissues. The crystal structures of two tissue-selective nifedipine analogues, nimodipine [isopropyl (2-methoxyethyl) 1,4-dihydro-2,6- dimethyl-4-(3-nitrophenyl)-3,5-pyridinecarboxylate] and nitrendipine [ethyl methyl 1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinecarboxylate] are reported. Nimodipine was observed to have an unusual ester side chain conformation that enhances the fit to the proposed ester-sensing region of the receptor.

  20. Anti-Epileptic Drugs Delay Age-Related Loss of Spiral Ganglion Neurons via T-type Calcium Channel

    Science.gov (United States)

    Lei, Debin; Gao, Xia; Perez, Philip; Ohlemiller, Kevin K; Chen, Chien-Chang; Campbell, Kevin P.; Hood, Aizhen Yang; Bao, Jianxin

    2011-01-01

    Loss of spiral ganglion neurons is a major cause of age-related hearing loss (presbycusis). Despite being the third most prevalent condition afflicting elderly persons, there are no known medications to prevent presbycusis. Because calcium signaling has long been implicated in age-related neuronal death, we investigated T-type calcium channels. This family is comprised of three members (Cav3.1, Cav3.2, and Cav3.3), based on their respective main pore-forming alpha subunits: α1G, α1H, and α1I. In the present study, we report a significant delay of age-related loss of cochlear function and preservation of spiral ganglion neurons in α1H null and heterozygous mice, clearly demonstrating an important role for Cav3.2 in age-related neuronal loss. Furthermore, we show that anticonvulsant drugs from a family of T-type calcium channel blockers can significantly preserve spiral ganglion neurons during aging. To our knowledge, this is the first report of drugs capable of diminishing age-related loss of spiral ganglion neurons. PMID:21640179

  1. It takes two T to shape immunity: emerging role for T-type calcium channels in immune cells

    Czech Academy of Sciences Publication Activity Database

    Lacinová, L.; Weiss, Norbert

    2016-01-01

    Roč. 35, č. 4 (2016), s. 393-396 ISSN 0231-5882 R&D Projects: GA ČR GA15-13556S; GA MŠk 7AMB15FR015 Institutional support: RVO:61388963 Keywords : calcium channel * T-type channel * Ca(v)3.1 * immune cells Subject RIV: CE - Biochemistry Impact factor: 1.170, year: 2016

  2. Vasodilatory effect of asafoetida essential oil on rat aorta rings: The role of nitric oxide, prostacyclin, and calcium channels.

    Science.gov (United States)

    Esmaeili, Hassan; Sharifi, Mozhdeh; Esmailidehaj, Mansour; Rezvani, Mohammad Ebrahim; Hafizibarjin, Zeynab

    2017-12-01

    Asafoetida is an oleo-gum resin mainly obtained from Ferula assa-foetida L. species in the apiaceae family. Previous studies have shown that it has antispasmodic effects on rat's and pig's ileums. The main goals of this study were to assess the vasodilatory effect of asafoetida essential oil (AEO) on the contractile response of rat's aorta rings and to find the role of nitric oxide, cyclooxygenase, and calcium channels. Thoracic aorta rings were stretched under a steady-state tension of 1 g in an organ bath apparatus for 1 h and then precontracted by KCl (80 mM) in the presence and absence of AEO. L-NAME (blocker of nitric oxide synthase) and indomethacin (blocker of cyclooxygenase) were used to assess the role of nitric oxide (NO) and prostacyclin in the vasodilatory effect of AEO. Also, the effect of AEO on the influx of calcium through the cell membrane calcium channels was determined. Data showed that AEO had vasodilatory effects on aorta rings with both intact (IC 50  = 1.6 µl/l) or denuded endothelium (IC 50  = 19.2 µl/l) with a significantly higher potency in intact endothelium rings. The vasodilatory effects of AEO were reduced, but not completely inhibited, in the presence of L-NAME or indomethacin. Adding AEO to the free-calcium medium also significantly reduced the CaCl 2 -induced contractions. The results indicated that AEO has a potent vasodilatory effect that is endothelium-dependent and endothelium-independent. Also, it reduced the influx of calcium into the cell through plasma membrane calcium channels. Copyright © 2017 Elsevier GmbH. All rights reserved.

  3. Active zone protein Bassoon co-localizes with presynaptic calcium channel, modifies channel function, and recovers from aging related loss by exercise.

    Science.gov (United States)

    Nishimune, Hiroshi; Numata, Tomohiro; Chen, Jie; Aoki, Yudai; Wang, Yonghong; Starr, Miranda P; Mori, Yasuo; Stanford, John A

    2012-01-01

    The P/Q-type voltage-dependent calcium channels (VDCCs) are essential for synaptic transmission at adult mammalian neuromuscular junctions (NMJs); however, the subsynaptic location of VDCCs relative to active zones in rodent NMJs, and the functional modification of VDCCs by the interaction with active zone protein Bassoon remain unknown. Here, we show that P/Q-type VDCCs distribute in a punctate pattern within the NMJ presynaptic terminals and align in three dimensions with Bassoon. This distribution pattern of P/Q-type VDCCs and Bassoon in NMJs is consistent with our previous study demonstrating the binding of VDCCs and Bassoon. In addition, we now show that the interaction between P/Q-type VDCCs and Bassoon significantly suppressed the inactivation property of P/Q-type VDCCs, suggesting that the Ca(2+) influx may be augmented by Bassoon for efficient synaptic transmission at NMJs. However, presynaptic Bassoon level was significantly attenuated in aged rat NMJs, which suggests an attenuation of VDCC function due to a lack of this interaction between VDCC and Bassoon. Importantly, the decreased Bassoon level in aged NMJs was ameliorated by isometric strength training of muscles for two months. The training increased Bassoon immunoreactivity in NMJs without affecting synapse size. These results demonstrated that the P/Q-type VDCCs preferentially accumulate at NMJ active zones and play essential role in synaptic transmission in conjunction with the active zone protein Bassoon. This molecular mechanism becomes impaired by aging, which suggests altered synaptic function in aged NMJs. However, Bassoon level in aged NMJs can be improved by muscle exercise.

  4. Protein structure and ionic selectivity in calcium channels: selectivity filter size, not shape, matters.

    Science.gov (United States)

    Malasics, Attila; Gillespie, Dirk; Nonner, Wolfgang; Henderson, Douglas; Eisenberg, Bob; Boda, Dezso

    2009-12-01

    Calcium channels have highly charged selectivity filters (4 COO(-) groups) that attract cations in to balance this charge and minimize free energy, forcing the cations (Na(+) and Ca(2+)) to compete for space in the filter. A reduced model was developed to better understand the mechanism of ion selectivity in calcium channels. The charge/space competition (CSC) mechanism implies that Ca(2+) is more efficient in balancing the charge of the filter because it provides twice the charge as Na(+) while occupying the same space. The CSC mechanism further implies that the main determinant of Ca(2+) versus Na(+) selectivity is the density of charged particles in the selectivity filter, i.e., the volume of the filter (after fixing the number of charged groups in the filter). In this paper we test this hypothesis by changing filter length and/or radius (shape) of the cylindrical selectivity filter of our reduced model. We show that varying volume and shape together has substantially stronger effects than varying shape alone with volume fixed. Our simulations show the importance of depletion zones of ions in determining channel conductance calculated with the integrated Nernst-Planck equation. We show that confining the protein side chains with soft or hard walls does not influence selectivity.

  5. Calcium Channel Genes Associated with Bipolar Disorder Modulate Lithium's Amplification of Circadian Rhythms

    Science.gov (United States)

    McCarthy, Michael J.; LeRoux, Melissa; Wei, Heather; Beesley, Stephen; Kelsoe, John R.; Welsh, David K.

    2015-01-01

    Bipolar disorder (BD) is associated with mood episodes and low amplitude circadian rhythms. Previously, we demonstrated that fibroblasts grown from BD patients show weaker amplification of circadian rhythms by lithium compared to control cells. Since calcium signals impact upon the circadian clock, and L-type calcium channels (LTCC) have emerged as genetic risk factors for BD, we examined whether loss of function in LTCCs accounts for the attenuated response to lithium in BD cells. We used fluorescent dyes to measure Ca2+ changes in BD and control fibroblasts after lithium treatment, and bioluminescent reporters to measure Per2∷luc rhythms in fibroblasts from BD patients, human controls, and mice while pharmacologically or genetically manipulating calcium channels. Longitudinal expression of LTCC genes (CACNA1C, CACNA1D and CACNB3) was then measured over 12-24 hr in BD and control cells. Our results indicate that independently of LTCCs, lithium stimulated intracellular Ca2+ less effectively in BD vs. control fibroblasts. In longitudinal studies, pharmacological inhibition of LTCCs or knockdown of CACNA1A, CACNA1C, CACNA1D and CACNB3 altered circadian rhythm amplitude. Diltiazem and knockdown of CACNA1C or CACNA1D eliminated lithium's ability to amplify rhythms. Knockdown of CACNA1A or CACNB3 altered baseline rhythms, but did not affect rhythm amplification by lithium. In human fibroblasts, CACNA1C genotype predicted the amplitude response to lithium, and the expression profiles of CACNA1C, CACNA1D and CACNB3 were altered in BD vs. controls. We conclude that in cells from BD patients, calcium signaling is abnormal, and that LTCCs underlie the failure of lithium to amplify circadian rhythms. PMID:26476274

  6. Copper-induced activation of TRP channels promotes extracellular calcium entry and activation of CaMs and CDPKs leading to copper entry and membrane depolarization in Ulva compressa

    Directory of Open Access Journals (Sweden)

    Melissa eGómez

    2015-03-01

    Full Text Available In order to identify channels involved in membrane depolarization, Ulva compressa was incubated with agonists of TRP channels C5, A1 and V1 and the level of intracellular calcium was detected. Agonists of TRPC5, A1 and V1 induced increases in intracellular calcium at 4, 9 and 12 min of exposure, respectively, and antagonists of TRPC5, A1 and V1 corresponding to SKF-96365 (SKF, HC-030031 (HC and capsazepin (CPZ, respectively, inhibited calcium increases indicating that functional TRPs exist in U. compressa. In addition, copper excess induced increases in intracellular calcium at 4, 9 and 12 min which were inhibited by SKF, HC and CPZ, respectively, indicating that copper activate TRPC5, A1 and V1 channels. Moreover, copper-induced calcium increases were inhibited by EGTA, a non-permeable calcium chelating agent, but not by thapsigargin, an inhibitor of endoplasmic reticulum (ER calcium ATPase, indicating that activation of TRPs leads to extracellular calcium entry. Furthermore, copper-induced calcium increases were not inhibited by W-7, an inhibitor of CaMs, and staurosporine, an inhibitor of CDPKs, indicating that extracellular calcium entry did not require CaMs and CDPKs activation. In addition, copper induced membrane depolarization events at 4, 8 and 11 min and these events were inhibited by SKF, HC, CPZ and bathocuproine, a specific copper chelating agent, indicating copper entry through TRP channels leading to membrane depolarization. Moreover, membrane depolarization events were inhibited by W-7 and staurosporine, indicating that CaMs and CDPKs are required in order to activate TRPs to allow copper entry. Thus, light-dependent copper-induced activation TRPC5, A1 and V1 promotes extracellular calcium entry leading to activation of CaMs and CDPKs which, in turn, promotes copper entry through these TRP channels leading to membrane depolarization.

  7. Postcountershock myocardial damage after pretreatment with adrenergic and calcium channel antagonists in halothane-anesthetized dogs

    Energy Technology Data Exchange (ETDEWEB)

    Gaba, D.M.; Metz, S.; Maze, M.

    1985-05-01

    Transthoracic electric countershock can cause necrotic myocardial lesions in humans as well as experimental animals. The authors investigated the effect on postcountershock myocardial damage of pretreatment with prazosin, an alpha-1 antagonist; L-metoprolol, a beta-1 antagonist, and verapamil, a calcium channel-blocking agent. Twenty dogs were anesthetized with halothane and given two transthoracic countershocks of 295 delivered joules each after drug or vehicle treatment. Myocardial injury was quantitated 24 h following countershock by measuring the uptake of technetium-99m pyrophosphate in the myocardium. Elevated technetium-99m pyrophosphate uptake occurred in visible lesions in most dogs regardless of drug treatment. For each of four parameters of myocardial damage there was no statistically significant difference between control animals and those treated with prazosin, metoprolol, or verapamil. These data suggest that adrenergic or calcium channel-mediated mechanisms are not involved in the pathogenesis of postcountershock myocardial damage.

  8. N-Acetylcysteine-induced vasodilatation is modulated by KATP channels, Na+/K+-ATPase activity and intracellular calcium concentration: An in vitro study.

    Science.gov (United States)

    Vezir, Özden; Çömelekoğlu, Ülkü; Sucu, Nehir; Yalın, Ali Erdinç; Yılmaz, Şakir Necat; Yalın, Serap; Söğüt, Fatma; Yaman, Selma; Kibar, Kezban; Akkapulu, Merih; Koç, Meryem İlkay; Seçer, Didem

    2017-08-01

    In this study, we aimed to investigate the role of ATP-sensitive potassium (K ATP ) channel, Na + /K + -ATPase activity, and intracellular calcium levels on the vasodilatory effect of N-acetylcysteine (NAC) in thoracic aorta by using electrophysiological and molecular techniques. Rat thoracic aorta ring preparations and cultured thoracic aorta cells were divided into four groups as control, 2mM NAC, 5mM NAC, and 10mM NAC. Thoracic aorta rings were isolated from rats for measurements of relaxation responses and Na + /K + -ATPase activity. In the cultured thoracic aorta cells, we measured the currents of K ATP channel, the concentration of intracellular calcium and mRNA expression level of K ATP channel subunits (KCNJ8, KCNJ11, ABCC8 and ABCC9). The relaxation rate significantly increased in all NAC groups compared to control. Similarly, Na + /K + - ATPase activity also significantly decreased in NAC groups. Outward K ATP channel current significantly increased in all NAC groups compared to the control group. Intracellular calcium concentration decreased significantly in all groups with compared control. mRNA expression level of ABCC8 subunit significantly increased in all NAC groups compared to the control group. Pearson correlation analysis showed that relaxation rate was significantly associated with K ATP current, intracellular calcium concentration, Na + /K + -ATPase activity and mRNA expression level of ABCC8 subunit. Our findings suggest that NAC relaxes vascular smooth muscle cells through a direct effect on K ATP channels, by increasing outward K+ flux, partly by increasing mRNA expression of K ATP subunit ABCC8, by decreasing in intracellular calcium and by decreasing in Na + /K + -ATPase activity. Copyright © 2017 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  9. The tarantula toxins ProTx-II and huwentoxin-IV differentially interact with human Nav1.7 voltage sensors to inhibit channel activation and inactivation.

    Science.gov (United States)

    Xiao, Yucheng; Blumenthal, Kenneth; Jackson, James O; Liang, Songping; Cummins, Theodore R

    2010-12-01

    The voltage-gated sodium channel Na(v)1.7 plays a crucial role in pain, and drugs that inhibit hNa(v)1.7 may have tremendous therapeutic potential. ProTx-II and huwentoxin-IV (HWTX-IV), cystine knot peptides from tarantula venoms, preferentially block hNa(v)1.7. Understanding the interactions of these toxins with sodium channels could aid the development of novel pain therapeutics. Whereas both ProTx-II and HWTX-IV have been proposed to preferentially block hNa(v)1.7 activation by trapping the domain II voltage-sensor in the resting configuration, we show that specific residues in the voltage-sensor paddle of domain II play substantially different roles in determining the affinities of these toxins to hNa(v)1.7. The mutation E818C increases ProTx-II's and HWTX-IV's IC(50) for block of hNa(v)1.7 currents by 4- and 400-fold, respectively. In contrast, the mutation F813G decreases ProTx-II affinity by 9-fold but has no effect on HWTX-IV affinity. It is noteworthy that we also show that ProTx-II, but not HWTX-IV, preferentially interacts with hNa(v)1.7 to impede fast inactivation by trapping the domain IV voltage-sensor in the resting configuration. Mutations E1589Q and T1590K in domain IV each decreased ProTx-II's IC(50) for impairment of fast inactivation by ~6-fold. In contrast mutations D1586A and F1592A in domain-IV increased ProTx-II's IC(50) for impairment of fast inactivation by ~4-fold. Our results show that whereas ProTx-II and HWTX-IV binding determinants on domain-II may overlap, domain II plays a much more crucial role for HWTX-IV, and contrary to what has been proposed to be a guiding principle of sodium channel pharmacology, molecules do not have to exclusively target the domain IV voltage-sensor to influence sodium channel inactivation.

  10. Orientation of the calcium channel beta relative to the alpha(12.2 subunit is critical for its regulation of channel activity.

    Directory of Open Access Journals (Sweden)

    Iuliia Vitko

    Full Text Available BACKGROUND: The Ca(vbeta subunits of high voltage-activated Ca(2+ channels control the trafficking and biophysical properties of the alpha(1 subunit. The Ca(vbeta-alpha(1 interaction site has been mapped by crystallographic studies. Nevertheless, how this interaction leads to channel regulation has not been determined. One hypothesis is that betas regulate channel gating by modulating movements of IS6. A key requirement for this direct-coupling model is that the linker connecting IS6 to the alpha-interaction domain (AID be a rigid structure. METHODOLOGY/PRINCIPAL FINDINGS: The present study tests this hypothesis by altering the flexibility and orientation of this region in alpha(12.2, then testing for Ca(vbeta regulation using whole cell patch clamp electrophysiology. Flexibility was induced by replacement of the middle six amino acids of the IS6-AID linker with glycine (PG6. This mutation abolished beta2a and beta3 subunits ability to shift the voltage dependence of activation and inactivation, and the ability of beta2a to produce non-inactivating currents. Orientation of Ca(vbeta with respect to alpha(12.2 was altered by deletion of 1, 2, or 3 amino acids from the IS6-AID linker (Bdel1, Bdel2, Bdel3, respectively. Again, the ability of Ca(vbeta subunits to regulate these biophysical properties were totally abolished in the Bdel1 and Bdel3 mutants. Functional regulation by Ca(vbeta subunits was rescued in the Bdel2 mutant, indicating that this part of the linker forms beta-sheet. The orientation of beta with respect to alpha was confirmed by the bimolecular fluorescence complementation assay. CONCLUSIONS/SIGNIFICANCE: These results show that the orientation of the Ca(vbeta subunit relative to the alpha(12.2 subunit is critical, and suggests additional points of contact between these subunits are required for Ca(vbeta to regulate channel activity.

  11. Synergistic Effect of Fluconazole and Calcium Channel Blockers against Resistant Candida albicans.

    Science.gov (United States)

    Liu, Shuyuan; Yue, Longtao; Gu, Wenrui; Li, Xiuyun; Zhang, Liuping; Sun, Shujuan

    2016-01-01

    Candidiasis has increased significantly recently that threatens patients with low immunity. However, the number of antifungal drugs on the market is limited in comparison to the number of available antibacterial drugs. This fact, coupled with the increased frequency of fungal resistance, makes it necessary to develop new therapeutic strategies. Combination drug therapy is one of the most widely used and effective strategy to alleviate this problem. In this paper, we were aimed to evaluate the combined antifungal effects of four CCBs (calcium channel blockers), amlodipine (AML), nifedipine (NIF), benidipine (BEN) and flunarizine (FNZ) with fluconazole against C. albicans by checkerboard and time-killing method. In addition, we determined gene (CCH1, MID1, CNA1, CNB1, YVC1, CDR1, CDR2 and MDR1) expression by quantitative PCR and investigated the efflux pump activity of resistant candida albicans by rhodamine 6G assay to reveal the potential mechanisms. Finally, we concluded that there was a synergy when fluconazole combined with the four tested CCBs against resistant strains, with fractional inhibitory concentration index (FICI) <0.5, but no interaction against sensitive strains (FICI = 0.56 ~ 2). The mechanism studies revealed that fluconazole plus amlodipine caused down-regulating of CNA1, CNB1 (encoding calcineurin) and YVC1 (encoding calcium channel protein in vacuole membrane).

  12. Impaired control of L-type voltage-dependent calcium channels in experimental hypertension

    Czech Academy of Sciences Publication Activity Database

    Pintérová, Mária; Líšková, Silvia; Dobešová, Zdenka; Behuliak, M.; Kuneš, Jaroslav; Zicha, Josef

    2009-01-01

    Roč. 58, Suppl.2 (2009), S43-S54 ISSN 0862-8408 R&D Projects: GA ČR(CZ) GA305/08/0139; GA ČR(CZ) GA305/09/0336; GA AV ČR(CZ) IAA500110902; GA MŠk(CZ) 1M0510 Institutional research plan: CEZ:AV0Z50110509 Keywords : calcium -activated K+ and Cl- channels * vasoactive systems * EDCF Subject RIV: ED - Physiology Impact factor: 1.430, year: 2009

  13. [Cognitive Function and Calcium. Structures and functions of Ca2+-permeable channels].

    Science.gov (United States)

    Kaneko, Shuji

    2015-02-01

    Calcium is essential for living organisms where the increase in intracellular Ca2+ concentration functions as a second messenger for many cellular processes including synaptic transmission and neural plasticity. The cytosolic concentration of Ca2+ is finely controlled by many Ca2+-permeable ion channels and transporters. The comprehensive view of their expression, function, and regulation will advance our understanding of neural and cognitive functions of Ca2+, which leads to the future drug discovery.

  14. Transmembrane proteoglycans control stretch-activated channels to set cytosolic calcium levels

    DEFF Research Database (Denmark)

    Gopal, Sandeep; Søgaard, Pernille; Multhaupt, Hinke A B

    2015-01-01

    show that syndecans regulate transient receptor potential canonical (TRPCs) channels to control cytosolic calcium equilibria and consequent cell behavior. In fibroblasts, ligand interactions with heparan sulfate of syndecan-4 recruit cytoplasmic protein kinase C to target serine714 of TRPC7...... with subsequent control of the cytoskeleton and the myofibroblast phenotype. In epidermal keratinocytes a syndecan-TRPC4 complex controls adhesion, adherens junction composition, and early differentiation in vivo and in vitro. In Caenorhabditis elegans, the TRPC orthologues TRP-1 and -2 genetically complement...

  15. The T-type calcium channel antagonist Z944 rescues impairments in crossmodal and visual recognition memory in Genetic Absence Epilepsy Rats from Strasbourg.

    Science.gov (United States)

    Marks, Wendie N; Cain, Stuart M; Snutch, Terrance P; Howland, John G

    2016-10-01

    Childhood absence epilepsy (CAE) is often comorbid with behavioral and cognitive symptoms, including impaired visual memory. Genetic Absence Epilepsy Rats from Strasbourg (GAERS) is an animal model closely resembling CAE; however, cognition in GAERS is poorly understood. Crossmodal object recognition (CMOR) is a recently developed memory task that examines not only purely visual and tactile memory, but also requires rodents to integrate sensory information about objects gained from tactile exploration to enable visual recognition. Both the visual and crossmodal variations of the CMOR task rely on the perirhinal cortex, an area with dense expression of T-type calcium channels. GAERS express a gain-in-function missense mutation in the Cav3.2 T-type calcium channel gene. Therefore, we tested whether the T-type calcium channel blocker Z944 dose dependently (1, 3, 10mg/kg; i.p.) altered CMOR memory in GAERS compared to the non-epileptic control (NEC) strain. GAERS demonstrated recognition memory deficits in the visual and crossmodal variations of the CMOR task that were reversed by the highest dose of Z944. Electroencephalogram recordings determined that deficits in CMOR memory in GAERS were not the result of seizures during task performance. In contrast, NEC showed a decrease in CMOR memory following Z944 treatment. These findings suggest that T-type calcium channels mediate CMOR in both the GAERS and NEC strains. Future research into the therapeutic potential of T-type calcium channel regulation may be particularly fruitful for the treatment of CAE and other disorders characterized by visual memory deficits. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Neuroprotective effect of gadolinium: a stretch-activated calcium channel blocker in mouse model of ischemia-reperfusion injury.

    Science.gov (United States)

    Gulati, Puja; Muthuraman, Arunachalam; Jaggi, Amteshwar S; Singh, Nirmal

    2013-03-01

    The present study was designed to investigate the potential of gadolinium, a stretch-activated calcium channel blocker in ischemic reperfusion (I/R)-induced brain injury in mice. Bilateral carotid artery occlusion of 12 min followed by reperfusion for 24 h was given to induce cerebral injury in male Swiss mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was assessed using Morris water maze test and motor incoordination was evaluated using rota-rod, lateral push, and inclined beam walking tests. In addition, total calcium, thiobarbituric acid reactive substance (TBARS), reduced glutathione (GSH), and acetylcholinesterase (AChE) activity were also estimated in brain tissue. I/R injury produced a significant increase in cerebral infarct size. A significant loss of memory along with impairment of motor performance was also noted. Furthermore, I/R injury also produced a significant increase in levels of TBARS, total calcium, AChE activity, and a decrease in GSH levels. Pretreatment of gadolinium significantly attenuated I/R-induced infarct size, behavioral and biochemical changes. On the basis of the present findings, we can suggest that opening of stretch-activated calcium channel may play a critical role in ischemic reperfusion-induced brain injury and that gadolinium has neuroprotective potential in I/R-induced injury.

  17. Effects of low-dose ionising radiation on pituitary adenoma: is there a role for L-type calcium channel?

    Energy Technology Data Exchange (ETDEWEB)

    Soares, Marcella Araugio; Santos, Raquel Gouvea dos [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN), Belo Horizonte, MG (Brazil). Lab. de Radiobiologia]. E-mail: santosr@cdtn.br

    2005-10-15

    Pituitary adenomas constitute about 6-18% of brain tumours in adults. Activation of voltage gated calcium currents can account for growth hormone over secretion in some GH-secreting pituitary adenomas that produce an acromegaly appearance and increase mortality. Ca{sup 2+} ions, as mediators of intracellular signalling, are crucial for the development of apoptosis. However, the role of [Ca{sup 2+}] in the development of apoptosis is ambiguous. In this study, the effects of low-dose ionising gamma radiation ({sup 60} Co) on rat pituitary adenoma cells survival and proliferation and the role of calcium channels on the apoptosis radio-induced were evaluated. Doses as low as 3 Gy were found to inhibit GH3 cell proliferation. Even though there was a significant number of live cells,168 hours following irradiation, they were not able to proliferate. The results indicate that the blockade of extracellular calcium influx through these channels does not interfere in the radiation-induced apoptosis in GH3 cells. (author)

  18. Investigation of the role of non-selective calcium channel blocker (flunarizine) on cerebral ischemic-reperfusion associated cognitive dysfunction in aged mice.

    Science.gov (United States)

    Gulati, Puja; Muthuraman, Arunachalam; Kaur, Parneet

    2015-04-01

    The present study was designed to investigate the role of flunarizine (a non-selective calcium channel blocker) on cerebral ischemic-reperfusion associated cognitive dysfunction in aged mice. Bilateral carotid artery occlusion of 12min followed by reperfusion for 24h was given to induce cerebral injury in male Swiss mice. The assessment of learning & memory was performed by Morris water maze test; motor in-coordination was evaluated by rota rod, lateral push and inclined beam walking tests; cerebral infarct size was quantified by triphenyltetrazolium chloride staining. In addition, reduced glutathione (GSH), total calcium and acetylcholinesterase (AChE) activity were also estimated in aged brain tissue. Donepezil treated group served as a positive control in this study. Ischemia reperfusion (I/R) injury produced significant increase in cerebral infarct size. A significant loss of memory along with impairment of motor performance was also noted. Further, I/R injury also produced significant increase in levels of total calcium, AChE activity and decrease in GSH levels. Pretreatment of flunarizine significantly attenuated I/R induced infarct size, behavioral and biochemical changes. Hence, it may be concluded that, a non-selective calcium channel blocker can be useful in I/R associated cognitive dysfunction due to its anti-oxidant, anti-infarct and modulatory actions of neurotransmitters & calcium channels. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. The Role of L-type Calcium Channels in Olfactory Learning and Its Modulation by Norepinephrine

    Directory of Open Access Journals (Sweden)

    Abhinaba Ghosh

    2017-12-01

    Full Text Available L type calcium channels (LTCCs are prevalent in different systems and hold immense importance for maintaining/performing selective functions. In the nervous system, CaV1.2 and CaV1.3 are emerging as critical modulators of neuronal functions. Although the general role of these calcium channels in modulating synaptic plasticity and memory has been explored, their role in olfactory learning is not well understood. In this review article we first discuss the role of LTCCs in olfactory learning especially focusing on early odor preference learning in neonate rodents, presenting evidence that while NMDARs initiate stimulus-specific learning, LTCCs promote protein-synthesis dependent long-term memory (LTM. Norepinephrine (NE release from the locus coeruleus (LC is essential for early olfactory learning, thus noradrenergic modulation of LTCC function and its implication in olfactory learning is discussed here. We then address the differential roles of LTCCs in adult learning and learning in aged animals.

  20. Design, synthesis, and biological testing of thiosalicylamides as a novel class of calcium channel blockers.

    Science.gov (United States)

    Mehanna, Ahmed S; Kim, Jin Yung

    2005-07-01

    The current research aimed to investigate the importance of the heterocyclic ring system in the structure of the cardiovascular drug diltiazem for its calcium channel blocking activity. The manuscript describes the design, synthesis, and biological testing of a total of 10 S-(p-methoxybenzyl), N-substituted thiosalicylamides as a series of non-cyclic compounds derived from diltiazem's structure. The new compounds maintained all diltiazem pharmacophores except the thiazepine ring system. In vitro evaluation of the new series for calcium channel blocking effects revealed moderate activities with IC50 values in the range of 4.8-56.0 microM. The data suggest that the ring system is not essential for activity; however, its absence leads to a considerable drop of activity relative to that of diltiazem (IC50=0.3 microM). Compounds of the current series showed optimum activity when the aliphatic alkyl chain on the salicylamide nitrogen is part of a piperidine or piperazine ring system substituted at the terminal nitrogen with a benzyl group.

  1. Functional role of voltage gated Ca2+ channels in heart automaticity

    Directory of Open Access Journals (Sweden)

    Pietro eMesirca

    2015-02-01

    Full Text Available Pacemaker activity of automatic cardiac myocytes controls the heartbeat in everyday life. Cardiac automaticity is under the control of several neurotransmitters and hormones and is constantly regulated by the autonomic nervous system to match the physiological needs of the organism. Several classes of ion channels and proteins involved in intracellular Ca2+ dynamics contribute to pacemaker activity. The functional role of voltage-gated calcium channels (VGCCs in heart automaticity and impulse conduction has been matter of debate for 30 years. However, growing evidence shows that VGCCs are important regulators of the pacemaker mechanisms and play also a major role in atrio-ventricular impulse conduction. Incidentally, studies performed in genetically modified mice lacking L-type Cav1.3 (Cav1.3-/- or T-type Cav3.1 (Cav3.1-/- channels show that genetic inactivation of these channels strongly impacts pacemaking. In cardiac pacemaker cells, VGCCs activate at negative voltages at the beginning of the diastolic depolarization and importantly contribute to this phase by supplying inward current. Loss-of-function of these channels also impairs atrio-ventricular conduction. Furthermore, inactivation of Cav1.3 channels promotes also atrial fibrillation and flutter in knockout mice suggesting that these channels can play a role in stabilizing atrial rhythm. Genomic analysis demonstrated that Cav1.3 and Cav3.1 channels are widely expressed in pacemaker tissue of mice, rabbits and humans. Importantly, human diseases of pacemaker activity such as congenital bradycardia and heart block have been attributed to loss-of-function of Cav1.3 and Cav3.1 channels. In this article, we will review the current knowledge on the role of VGCCs in the generation and regulation of heart rate and rhythm. We will discuss also how loss of Ca2+ entry through VGCCs could influence intracellular Ca2+ handling and promote atrial arrhythmias.

  2. The role of solvation in the binding selectivity of the L-type calcium channel.

    Science.gov (United States)

    Boda, Dezső; Henderson, Douglas; Gillespie, Dirk

    2013-08-07

    We present grand canonical Monte Carlo simulation results for a reduced model of the L-type calcium channel. While charged residues of the protein amino acids in the selectivity filter are treated explicitly, most of the degrees of freedom (including the rest of the protein and the solvent) are represented by their dielectric response, i.e., dielectric continua. The new aspect of this paper is that the dielectric coefficient in the channel is different from that in the baths. The ions entering the channel, thus, cross a dielectric boundary at the entrance of the channel. Simulating this case has been made possible by our recent methodological development [D. Boda, D. Henderson, B. Eisenberg, and D. Gillespie, J. Chem. Phys. 135, 064105 (2011)]. Our main focus is on the effect of solvation energy (represented by the Born energy) on monovalent vs. divalent ion selectivity in the channel. We find no significant change in selectivity by changing the dielectric coefficient in the channel because the larger solvation penalty is counterbalanced by the enhanced Coulomb attraction inside the channel as soon as we use the Born radii (fitted to experimental hydration energies) to compute the solvation penalty from the Born equation.

  3. Synergistic Effect of Fluconazole and Calcium Channel Blockers against Resistant Candida albicans.

    Directory of Open Access Journals (Sweden)

    Shuyuan Liu

    Full Text Available Candidiasis has increased significantly recently that threatens patients with low immunity. However, the number of antifungal drugs on the market is limited in comparison to the number of available antibacterial drugs. This fact, coupled with the increased frequency of fungal resistance, makes it necessary to develop new therapeutic strategies. Combination drug therapy is one of the most widely used and effective strategy to alleviate this problem. In this paper, we were aimed to evaluate the combined antifungal effects of four CCBs (calcium channel blockers, amlodipine (AML, nifedipine (NIF, benidipine (BEN and flunarizine (FNZ with fluconazole against C. albicans by checkerboard and time-killing method. In addition, we determined gene (CCH1, MID1, CNA1, CNB1, YVC1, CDR1, CDR2 and MDR1 expression by quantitative PCR and investigated the efflux pump activity of resistant candida albicans by rhodamine 6G assay to reveal the potential mechanisms. Finally, we concluded that there was a synergy when fluconazole combined with the four tested CCBs against resistant strains, with fractional inhibitory concentration index (FICI <0.5, but no interaction against sensitive strains (FICI = 0.56 ~ 2. The mechanism studies revealed that fluconazole plus amlodipine caused down-regulating of CNA1, CNB1 (encoding calcineurin and YVC1 (encoding calcium channel protein in vacuole membrane.

  4. Chronic deficit in nitric oxide elicits oxidative stress and augments T-type calcium-channel contribution to vascular tone of rodent arteries and arterioles

    DEFF Research Database (Denmark)

    Howitt, Lauren; Kuo, Ivana Y; Ellis, Anthie

    2013-01-01

    arteries in vitro and skeletal muscle arterioles in vivo to study the contribution of L-type (1 µmol/L nifedipine) and T-type (1 µmol/L mibefradil, 3 µmol/L NNC 55-0396) calcium channels to vascular tone, following acute or chronic inhibition of nitric oxide. Acute inhibition with l-NAME (10 µmol...... was reversed by acute scavenging of superoxide with tempol (1 mmol/L), or inhibition of NADPH oxidase with apocynin (500 µmol/L) or DPI (5 µmol/L). CONCLUSION: We conclude that nitric oxide deficit produces a significant increase in the contribution of Cav3.1 and Cav3.2 T-type calcium channels to vascular tone......, by regulating the bioavailability of reactive oxygen species produced by NADPH oxidase. Our data provide evidence for a novel causal link between nitric oxide deficit, oxidative stress, and T-type calcium channel function....

  5. Subthreshold membrane potential oscillations in inferior olive neurons are dynamically regulated by P/Q- and T-type calcium channels: a study in mutant mice.

    Science.gov (United States)

    Choi, Soonwook; Yu, Eunah; Kim, Daesoo; Urbano, Francisco J; Makarenko, Vladimir; Shin, Hee-Sup; Llinás, Rodolfo R

    2010-08-15

    The role of P/Q- and T-type calcium channels in the rhythmic oscillatory behaviour of inferior olive (IO) neurons was investigated in mutant mice. Mice lacking either the CaV2.1 gene of the pore-forming alpha1A subunit for P/Q-type calcium channel, or the CaV3.1 gene of the pore-forming alpha1G subunit for T-type calcium channel were used. In vitro intracellular recording from IO neurons reveals that the amplitude and frequency of sinusoidal subthreshold oscillations (SSTOs) were reduced in the CaV2.1-/- mice. In the CaV3.1-/- mice, IO neurons also showed altered patterns of SSTOs and the probability of SSTO generation was significantly lower (15%, 5 of 34 neurons) than that of wild-type (78%, 31 of 40 neurons) or CaV2.1-/- mice (73%, 22 of 30 neurons). In addition, the low-threshold calcium spike and the sustained endogenous oscillation following rebound potentials were absent in IO neurons from CaV3.1-/- mice. Moreover, the phase-reset dynamics of oscillatory properties of single neurons and neuronal clusters in IO were remarkably altered in both CaV2.1-/- and CaV3.1-/- mice. These results suggest that both alpha1A P/Q- and alpha1G T-type calcium channels are required for the dynamic control of neuronal oscillations in the IO. These findings were supported by results from a mathematical IO neuronal model that incorporated T and P/Q channel kinetics.

  6. Lack of direct evidence for a functional role of voltage-operated calcium channels in juxtaglomerular cells

    DEFF Research Database (Denmark)

    Kurtz, A; Skott, O; Chegini, S

    1990-01-01

    in patch-clamped nor in intact Furaester-loaded cells. Moreover, basal renin secretion from a preparation enriched in mouse juxtaglomerular cells and from rat glomeruli with attached juxtaglomerular cells was not inhibited when extracellular potassium was isoosmotically increased to 56 mmol/l. In mouse...... kidney slices, however, depolarizing potassium concentrations caused a delayed inhibition at 56 mmol/l and a delayed stimulation of renin secretion at 110 mmol/l. Taken together, our study does not provide direct evidence for a role of voltage-activated calcium channels in the regulation of calcium...

  7. Calcium waves.

    Science.gov (United States)

    Jaffe, Lionel F

    2008-04-12

    Waves through living systems are best characterized by their speeds at 20 degrees C. These speeds vary from those of calcium action potentials to those of ultraslow ones which move at 1-10 and/or 10-20 nm s(-1). All such waves are known or inferred to be calcium waves. The two classes of calcium waves which include ones with important morphogenetic effects are slow waves that move at 0.2-2 microm s(-1) and ultraslow ones. Both may be propagated by cycles in which the entry of calcium through the plasma membrane induces subsurface contraction. This contraction opens nearby stretch-sensitive calcium channels. Calcium entry through these channels propagates the calcium wave. Many slow waves are seen as waves of indentation. Some are considered to act via cellular peristalsis; for example, those which seem to drive the germ plasm to the vegetal pole of the Xenopus egg. Other good examples of morphogenetic slow waves are ones through fertilizing maize eggs, through developing barnacle eggs and through axolotl embryos during neural induction. Good examples of ultraslow morphogenetic waves are ones during inversion in developing Volvox embryos and across developing Drosophila eye discs. Morphogenetic waves may be best pursued by imaging their calcium with aequorins.

  8. Eye movements of the murine P/Q calcium channel mutant Tottering, and the impact of aging

    NARCIS (Netherlands)

    J.S. Stahl (J.); R.A. James (Robert); B.S. Oommen (Brian); F.E. Hoebeek (Freek); C.I. de Zeeuw (Chris)

    2006-01-01

    textabstractMice carrying mutations of the gene encoding the ion pore of the P/Q calcium channel (Cacna1a) are an instance in which cerebellar dysfunction may be attributable to altered electrophysiology and thus provide an opportunity to study how neuronal intrinsic properties dictate signal

  9. POSITIONS OF CALCIUM CHANNEL BLOCKER LERCANIDIPINE ACCORDING TO EVIDENCE BASED CARDIOLOGY

    Directory of Open Access Journals (Sweden)

    Yu. V. Lukina

    2010-01-01

    Full Text Available Data of evidence based cardiology including results of international clinical trials on efficacy and safety of the modern calcium channel blocker (CCB, lercanidipine, are presented. Results of these trials show the firm position of lercanidipine in the modern cardiology and confirm that treatment with lercanidipine leads to significant reduction of systolic and diastolic blood pressure (BP with no effect on heart rate (HR. Peripheral edema (the common side effect of CCBs occurs rarer with lercanidipine treatment than this with any other CCB treatment. Lercanidipine can be recommended to patients with concomitant diseases due to its additional features.

  10. Effects of endothelin, calcium channel blockade and EDRF inhibition on the contractility of human uteroplacental arteries.

    Science.gov (United States)

    Fried, G; Liu, Y A

    1994-08-01

    In order to examine the possibility that endothelin might be important in the regulation of placental blood flow, human uteroplacental vessels were superfused in vitro to study the contractile effect of endothelin as compared with a known strong contractor of placental blood vessels, serotonin (5-HT). The contractile responses were compared in the presence and absence of calcium channel blocking agents, as well as in the presence of L-NMA, an inhibitor of EDRF/nitric oxide. Endothelin (ET, 10(-10)-10(-6) M) and 5-HT (10(-8)-10(-4) M) induced contractions in the vessels. Maximal contractions in the presence of endothelin were elicited at 10(-7) M, whereas 5-HT elicited maximal contractions at 10(-5) M. At 10(-7) M, ET was more potent than 5-HT. The calcium-channel blocking agents nifedipine, diltiazem and NiCl2 relaxed the vessels by 5-15% from baseline. The contractile response to ET in the presence of nifedipine or diltiazem was reduced by 55 and 67%, respectively. The response of 5-HT in the presence of nifedipine was reduced by 58%. The contractile response to 5-HT as well as ET in the presence of both nifedipine and NiCl2 was not significantly lower than in the presence of nifedipine only. The EDRF-inhibiting agent L-NMA caused a small contractile response at concentrations of 10(-6)-10(-5) M. ET as well as 5-HT added after pretreatment with L-NMA produced a larger contractile response than ET or 5-HT alone. The results show that ET has a strong contractile effect on placental blood vessels at concentrations likely to occur during labor and delivery. The mechanism whereby ET as well as 5-HT contracts placental vessel smooth muscle appears to partly involve nifedipine- and diltiazem-sensitive calcium channels, but almost half of the response depends on mobilization of calcium through other means.

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

    DEFF Research Database (Denmark)

    Dalsgaard, Thomas; Kroigaard, Christel; Bek, Toke

    2009-01-01

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

  12. Cellular uptake of 99mTcN-NOET in human leukaemic HL-60 cells is related to calcium channel activation and cell proliferation

    International Nuclear Information System (INIS)

    Guillermet, Stephanie; Vuillez, Jean-Philippe; Caravel, Jean-Pierre; Marti-Batlle, Daniele; Fagret, Daniel; Fontaine, Eric; Pasqualini, Roberto

    2006-01-01

    A major goal of nuclear oncology is the development of new radiolabelled tracers as proliferation markers. Intracellular calcium waves play a fundamental role in the course of the cell cycle. These waves occur in non-excitable tumour cells via store-operated calcium channels (SOCCs). Bis(N-ethoxy, N-ethyldithiocarbamato) nitrido technetium (V)-99m ( 99m TcN-NOET) has been shown to interact with L-type voltage-operated calcium channels (VOCCs) in cultured cardiomyocytes. Considering the analogy between VOCCs and SOCCs, we sought to determine whether 99m TcN-NOET also binds to activated SOCCs in tumour cells in order to clarify the potential value of this tracer as a proliferation marker. Uptake kinetics of 99m TcN-NOET were measured in human leukaemic HL-60 cells over 60 min and the effect of several calcium channel modulators on 1-min tracer uptake was studied. The uptake kinetics of 99m TcN-NOET were compared both with the variations of cytosolic free calcium concentration measured by indo-1/AM and with the variations in the SG 2 M cellular proliferation index. All calcium channel inhibitors significantly decreased the cellular uptake of 99m TcN-NOET whereas the activator thapsigargin induced a significant 10% increase. In parallel, SOCC activation by thapsigargin, as measured using the indo-1/AM probe, was inhibited by nicardipine. These results indicate that the uptake of 99m TcN-NOET is related to the activation of SOCCs. Finally, a correlation was observed between the tracer uptake and variations in the proliferation index SG 2 M. The uptake of 99m TcN-NOET seems to be related to SOCC activation and to cell proliferation in HL-60 cells. These results indicate that 99m TcN-NOET might be a marker of cell proliferation. (orig.)

  13. Omega-conotoxin- and nifedipine-insensitive voltage-operated calcium channels mediate K(+)-induced release of pro-thyrotropin-releasing hormone-connecting peptides Ps4 and Ps5 from perifused rat hypothalamic slices.

    Science.gov (United States)

    Valentijn, K; Tranchand Bunel, D; Vaudry, H

    1992-07-01

    The rat thyrotropin-releasing hormone (TRH) precursor (prepro-TRH) contains five copies of the TRH progenitor sequence linked together by intervening sequences. Recently, we have shown that the connecting peptides prepro-TRH-(160-169) (Ps4) and prepro-TRH-(178-199) (Ps5) are released from rat hypothalamic neurones in response to elevated potassium concentrations, in a calcium-dependent manner. In the present study, the role of voltage-operated calcium channels in potassium-induced release of Ps4 and Ps5 was investigated, using a perifusion system for rat hypothalamic slices. The release of Ps4 and Ps5 stimulated by potassium (70 mM) was blocked by the inorganic ions Co2+ (2.6 mM) and Ni2+ (5 mM). In contrast, the stimulatory effect of KCl was insensitive to Cd2+ (100 microM). The dihydropyridine antagonist nifedipine (10 microM) had no effect on K(+)-evoked release of Ps4 and Ps5. Furthermore, the response to KCl was not affected by nifedipine (10 microM) in combination with diltiazem (1 microM), a benzothiazepine which increases the affinity of dihydropyridine antagonists for their receptor. The dihydropyridine agonist BAY K 8644, at concentrations as high as 1 mM, did not stimulate the basal secretion of Ps4 and Ps5. In addition, BAY K 8644 had no potentiating effect on K(+)-induced release of Ps4 and Ps5. The marine cone snail toxin omega-conotoxin, a blocker of both L- and N-type calcium channels had no effect on the release of Ps4 and Ps5 stimulated by potassium. Similarly, the omega-conopeptide SNX-111, a selective blocker of N-type calcium channels, did not inhibit the stimulatory effect of potassium. The release of Ps4 and Ps5 evoked by high K+ was insensitive to the non-selective calcium channel blocker verapamil (20 microM). Amiloride (1 microM), a putative blocker of T-type calcium channels, did not affect KCl-induced secretion of the two connecting peptides. Taken together, these results indicate that two connecting peptides derived from the pro-TRH, Ps

  14. 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

  15. Potassium conductances mediate bidirectional state-dependent modulation of action potential evoked dendritic calcium signals in dentate gyrus granule cells

    Directory of Open Access Journals (Sweden)

    János Brunner

    2014-03-01

    Full Text Available Backpropagating action potentials (bAPs and local calcium signals that they trigger are fundamental for dendritic functions. Here we addressed the question what extent the changes of local dendritic membrane properties can contribute to the shaping of the coupling between dendritic action potentials and the local calcium responses. Using a combination of in vitro electrophysiological and confocal imaging techniques we found that activation of dendritic GIRK channels via mGlu2 or GABAB receptors enhanced the bAP¬-triggered calcium signals in the dendrites of dentate gyrus granule cells (GCs. The enhancement of calcium signals was significant only in those dendritic regions, where these receptors are predominantly expressed. Similarly to GIRK channel activation, somatic hyperpolarization by DC current injection (from -64 mV to -77 mV, significantly increased bAP-associated calcium signals in the proximal dendrites. The hyperpolarization was associated with a decrease in the input resistance due to the rectification of the membrane potential of GCs. The effect of hyperpolarization on the calcium signals was maintained when T-type calcium currents were blocked but it decreased when GIRK channels were inhibited. Simultaneous dual somato-dendritic recordings from GCs showed that somatic hyperpolarization accelerated the repolarization phase of dendritic bAP in the proximal region whereas the rising phase and peak amplitude was not affected. We hypothesize that the larger driving force for calcium ions during the faster repolarization can contribute to the increasing in calcium signals. Employment of previously recorded dendritic bAP waveforms from hyperpolarized membrane potential as voltage command evoked larger calcium currents in nucleated patches compared to bAP waveform from the same recording at depolarized membrane potential. Furthermore, addition of native, high-voltage activated, inactivating potassium conductance by somatic dynamic clamp

  16. Reversal of Neuropathic Pain in Diabetes by Targeting Glycosylation of Cav3.2 T-Type Calcium Channels

    OpenAIRE

    Orestes, Peihan; Osuru, Hari Prasad; McIntire, William E.; Jacus, Megan O.; Salajegheh, Reza; Jagodic, Miljen M.; Choe, WonJoo; Lee, JeongHan; Lee, Sang-Soo; Rose, Kirstin E.; Poiro, Nathan; DiGruccio, Michael R.; Krishnan, Katiresan; Covey, Douglas F.; Lee, Jung-Ha

    2013-01-01

    It has been established that CaV3.2 T-type voltage-gated calcium channels (T-channels) play a key role in the sensitized (hyperexcitable) state of nociceptive sensory neurons (nociceptors) in response to hyperglycemia associated with diabetes, which in turn can be a basis for painful symptoms of peripheral diabetic neuropathy (PDN). Unfortunately, current treatment for painful PDN has been limited by nonspecific systemic drugs with significant side effects or potential for abuse. We studied i...

  17. GABA(A) Increases Calcium in Subventricular Zone Astrocyte-Like Cells Through L- and T-Type Voltage-Gated Calcium Channels

    DEFF Research Database (Denmark)

    Young, Stephanie Z; Platel, Jean-Claude; Nielsen, Jakob V

    2010-01-01

    In the adult neurogenic subventricular zone (SVZ), the behavior of astrocyte-like cells and some of their functions depend on changes in intracellular Ca(2+) levels and tonic GABA(A) receptor activation. However, it is unknown whether, and if so how, GABA(A) receptor activity regulates...... intracellular Ca(2+) dynamics in SVZ astrocytes. To monitor Ca(2+) activity selectively in astrocyte-like cells, we used two lines of transgenic mice expressing either GFP fused to a Gq-coupled receptor or DsRed under the human glial fibrillary acidic protein (hGFAP) promoter. GABA(A) receptor activation...... induced Ca(2+) increases in 40-50% of SVZ astrocytes. GABA(A)-induced Ca(2+) increases were prevented with nifedipine and mibefradil, blockers of L- and T-type voltage-gated calcium channels (VGCC). The L-type Ca(2+) channel activator BayK 8644 increased the percentage of GABA(A)-responding astrocyte...

  18. Electroconvulsive stimulations prevent chronic stress-induced increases in L-type calcium channel mRNAs in the hippocampus and basolateral amygdala

    DEFF Research Database (Denmark)

    Maigaard, Katrine; Pedersen, Ida Hageman; Jørgensen, Anders

    2012-01-01

    Although affective disorders have high prevalence, morbidity and mortality, we do not fully understand disease etiopathology, nor have we determined the exact mechanisms by which treatment works. Recent research indicates that intracellular calcium ion dysfunction might be involved. Here we use...... the chronic restraint stress model of affective disorder (6 h restraint per day for 21 days) in combination with electroconvulsive stimulations to examine the effects of stress and an effective antidepressive treatment modality on L-type voltage gated calcium channel subunit mRNA expression patterns...... in the brain. We find that stress tended to upregulate Ca(v)1.2 and Ca(v)1.3 channels in a brain region specific manner, while ECS tended to normalise this effect. This was more pronounced for Ca(v)1.2 channels, where stress clearly increased expression in both the basolateral amygdala, dentate gyrus and CA3...

  19. Calcium Regulation of Hemorrhagic Fever Virus Budding: Mechanistic Implications for Host-Oriented Therapeutic Intervention.

    Directory of Open Access Journals (Sweden)

    Ziying Han

    2015-10-01

    Full Text Available Hemorrhagic fever viruses, including the filoviruses (Ebola and Marburg and arenaviruses (Lassa and Junín viruses, are serious human pathogens for which there are currently no FDA approved therapeutics or vaccines. Importantly, transmission of these viruses, and specifically late steps of budding, critically depend upon host cell machinery. Consequently, strategies which target these mechanisms represent potential targets for broad spectrum host oriented therapeutics. An important cellular signal implicated previously in EBOV budding is calcium. Indeed, host cell calcium signals are increasingly being recognized to play a role in steps of entry, replication, and transmission for a range of viruses, but if and how filoviruses and arenaviruses mobilize calcium and the precise stage of virus transmission regulated by calcium have not been defined. Here we demonstrate that expression of matrix proteins from both filoviruses and arenaviruses triggers an increase in host cytoplasmic Ca2+ concentration by a mechanism that requires host Orai1 channels. Furthermore, we demonstrate that Orai1 regulates both VLP and infectious filovirus and arenavirus production and spread. Notably, suppression of the protein that triggers Orai activation (Stromal Interaction Molecule 1, STIM1 and genetic inactivation or pharmacological blockade of Orai1 channels inhibits VLP and infectious virus egress. These findings are highly significant as they expand our understanding of host mechanisms that may broadly control enveloped RNA virus budding, and they establish Orai and STIM1 as novel targets for broad-spectrum host-oriented therapeutics to combat these emerging BSL-4 pathogens and potentially other enveloped RNA viruses that bud via similar mechanisms.

  20. Familial hypocalciuric hypercalcemia and calcium sensing receptor

    DEFF Research Database (Denmark)

    Mrgan, Monija; Nielsen, Sanne; Brixen, Kim

    2014-01-01

    Familial hypocalciuric hypercalcemia (FHH) is a lifelong, benign autosomal dominant disease characterized by hypercalcemia, normal to increased parathyroid hormone level, and a relatively low renal calcium excretion. Inactivation of the calcium-sensing receptor in heterozygous patients results...... in FHH, while in homozygous patients as well as in compound heterozygous or dominant negative heterozygous patients, it may result in neonatal severe hyperparathyroidism (NSHPT). Parathyroid surgery is not indicated in FHH and does not lower plasma calcium unless total parathyroidectomy is performed...

  1. Mean field strategies induce unrealistic nonlinearities in calcium puffs

    Directory of Open Access Journals (Sweden)

    Guillermo eSolovey

    2011-08-01

    Full Text Available Mean field models are often useful approximations to biological systems, but sometimes, they can yield misleading results. In this work, we compare mean field approaches with stochastic models of intracellular calcium release. In particular, we concentrate on calcium signals generated by the concerted opening of several clustered channels (calcium puffs. To this end we simulate calcium puffs numerically and then try to reproduce features of the resulting calcium distribution using mean field models were all the channels open and close simultaneously. We show that an unrealistic nonlinear relationship between the current and the number of open channels is needed to reproduce the simulated puffs. Furthermore, a single channel current which is five times smaller than the one of the stochastic simulations is also needed. Our study sheds light on the importance of the stochastic kinetics of the calcium release channel activity to estimate the release fluxes.

  2. [Nonuniform distribution and contribution of the P- and P/Q-type calcium channels to short-term inhibitory synaptic transmission in cultured hippocampal neurons].

    Science.gov (United States)

    Mizerna, O P; Fedulova, S A; Veselovs'kyĭ, M S

    2010-01-01

    In the present study, we investigated the sensitivity of GABAergic short-term plasticity to the selective P- and P/Q-type calcium channels blocker omega-agatoxin-IVA. To block the P-type channels we used 30 nM of this toxin and 200 nM of the toxin was used to block the P/Q channel types. The evoked inhibitory postsynaptic currents (eIPSC) were studied using patch-clamp technique in whole-cell configuration in postsynaptic neuron and local extracellular stimulation of single presynaptic axon by rectangular pulse. The present data show that the contribution of P- and P/Q-types channels to GABAergic synaptic transmission in cultured hippocampal neurons are 30% and 45%, respectively. It was shown that the mediate contribution of the P- and P/Q-types channels to the amplitudes of eIPSC is different to every discovered neuron. It means that distribution of these channels is non-uniform. To study the short-term plasticity of inhibitory synaptic transmission, axons of presynaptic neurons were paired-pulse stimulated with the interpulse interval of 150 ms. Neurons demonstrated both the depression and facilitation. The application of 30 nM and 200 nM of the blocker decreased the depression and increased facilitation to 8% and 11%, respectively. In addition, we found that the mediate contribution of the P- and P/Q-types channels to realization of synaptic transmission after the second stimuli is 4% less compared to that after the first one. Therefore, blocking of both P- and P/Q-types calcium channels can change the efficiency of synaptic transmission. In this instance it facilitates realization of the transmission via decreased depression or increased facilitation. These results confirm that the P- and P/Q-types calcium channels are involved in regulation of the short-term inhibitory synaptic plasticity in cultured hippocampal neurons.

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  4. 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).…

  5. Depletion of intracellular calcium stores facilitates the influx of extracellular calcium in platelet derived growth factor stimulated A172 glioblastoma cells.

    Science.gov (United States)

    Vereb, G; Szöllösi, J; Mátyus, L; Balázs, M; Hyun, W C; Feuerstein, B G

    1996-05-01

    Calcium signaling in non-excitable cells is the consequence of calcium release from intracellular stores, at times followed by entry of extracellular calcium through the plasma membrane. To study whether entry of calcium depends upon the level of saturation of intracellular stores, we measured calcium channel opening in the plasma membrane of single confluent A172 glioblastoma cells stimulated with platelet derived growth factor (PDGF) and/or bradykinin (BK). We monitored the entry of extracellular calcium by measuring manganese quenching of Indo-1 fluorescence. PDGF raised intracellular calcium concentration ([Ca2+]i) after a dose-dependent delay (tdel) and then opened calcium channels after a dose-independent delay (tch). At higher doses (> 3 nM), BK increased [Ca2+]i after a tdel approximately 0 s, and tch decreased inversely with both dose and peak [Ca2+]i. Experiments with thapsigargin (TG), BK, and PDGF indicated that BK and PDGF share intracellular Ca2+ pools that are sensitive to TG. When these stores were depleted by treatment with BK and intracellular BAPTA, tdel did not change, but tch fell to almost 0 s in PDGF stimulated cells, indicating that depletion of calcium stores affects calcium channel opening in the plasma membrane. Our data support the capacitative model for calcium channel opening and the steady-state model describing quantal Ca2+ release from intracellular stores.

  6. Antischistosomal activity of a calcium channel antagonist on schistosomula and adult Schistosoma mansoni worms

    Directory of Open Access Journals (Sweden)

    Vanessa Silva-Moraes

    2013-08-01

    Full Text Available Current schistosomiasis control strategies are largely based on chemotherapeutic agents and a limited number of drugs are available today. Praziquantel (PZQ is the only drug currently used in schistosomiasis control programs. Unfortunately, this drug shows poor efficacy in patients during the earliest infection phases. The effects of PZQ appear to operate on the voltage-operated Ca2+channels, which are located on the external Schistosoma mansoni membrane. Because some Ca2+channels have dihydropyridine drug class (a class that includes nifedipine sensitivity, an in vitro analysis using a calcium channel antagonist (clinically used for cardiovascular hypertension was performed to determine the antischistosomal effects of nifedipine on schistosomula and adult worm cultures. Nifedipine demonstrated antischistosomal activity against schistosomula and significantly reduced viability at all of the concentrations used alone or in combination with PZQ. In contrast, PZQ did not show significant efficacy when used alone. Adult worms were also affected by nifedipine after a 24 h incubation and exhibited impaired motility, several lesions on the tegument and intense contractility. These data support the idea of Ca2+channels subunits as drug targets and favour alternative therapeutic schemes when drug resistance has been reported. In this paper, strong arguments encouraging drug research are presented, with a focus on exploring schistosomal Ca2+channels.

  7. Calcium Channel Blockers and Esophageal Sclerosis: Should We Expect Exacerbation of Interstitial Lung Disease

    Directory of Open Access Journals (Sweden)

    Charalampos Seretis

    2012-01-01

    Full Text Available Esophageal sclerosis is the most common visceral manifestation of systemic sclerosis, resulting in impaired esophageal clearance and retention of ingested food; in addition, co-existence of lung fibrosis with esophageal scleroderma is not uncommon. Both the progression of generalized connective tissue disorders and the damaging effect of chronic aspiration due to esophageal dysmotility appear to be involved in this procedure of interstitial fibrosis. Nifedipine is a widely prescribed calcium antagonist in a significant percentage of rheumatologic patients suffering from Raynaud syndrome, in order to inhibit peripheral vasospasm. Nevertheless, blocking calcium channels has proven to contribute to exacerbation of gastroesophageal reflux, which consequently can lead to chronic aspiration. We describe the case of severe exacerbation of interstitial lung disease in a 76-year-old female with esophageal sclerosis who was treated with oral nifedipine for Raynaud syndrome.

  8. The analgesic effect of trans-resveratrol is regulated by calcium channels in the hippocampus of mice.

    Science.gov (United States)

    Wang, Weijie; Yu, Yingcong; Li, Jing; Wang, Lin; Li, Zhi; Zhang, Chong; Zhen, Linlin; Ding, Lianshu; Wang, Gang; Sun, Xiaoyang; Xu, Ying

    2017-08-01

    Resveratrol has been widely studied in terms of it's potential to slow the progression of many diseases. But little is known about the mechanism of action in neuropathic pain. Neuropathic pain is the main type of chronic pain associated with tissue injury. Calcium channels and calcium/caffeine-sensitive pools are associated with analgesic pathway involving neuropathic pain. Our previous study suggested that the antinociceptive effect of resveratrol was involved in Ca 2+ /calmodulin-dependent signaling in the spinal cord of mice. The aim of this study was to explore the involvement of Ca 2+ in analgesic effects of trans-resveratrol in neuropathic pain and signal pathway in hippocampus. Hot plate test was used to assess antinociceptive response when mice were treated with trans-resveratrol alone or in combination with Mk 801, nimodipine, CaCl 2 , ryanodine or EGTA. The effects of trans-resveratrol and the combination on Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) and BDNF (brain-derived neurotrophic factor) expression in hippocampus were also investigated. The results showed that trans-resveratrol increased paw withdraw latency in the hot plate test. The effect of resveratrol was enhanced by Mk 801 and nimodipine. Central administration of Ca 2+ , however, abolished the antinociceptive effects of resveratrol. In contrast, centrally administered EGTA or ryanodine improved trans-resveratrol induced antinociception. There was a significant increase in p-CaMKII and BDNF expression in the hippocampus when resveratrol were combined with Mk 801, nimodipine, ryanodine and EGTA. Administration of CaCl 2 blocked changes in p-CaMKII and BDNF levels in the hippocampus. These findings suggest that trans-resveratrol exerts the effects of antinociception through regulation of calcium channels and calcium/caffeine-sensitive pools.

  9. Modeling pathogenesis and treatment response in childhood absence epilepsy.

    Science.gov (United States)

    Knox, Andrew T; Glauser, Tracy; Tenney, Jeffrey; Lytton, William W; Holland, Katherine

    2018-01-01

    Childhood absence epilepsy (CAE) is a genetic generalized epilepsy syndrome with polygenic inheritance, with genes for γ-aminobutyric acid (GABA) receptors and T-type calcium channels implicated in the disorder. Previous studies of T-type calcium channel electrophysiology have shown genetic changes and medications have multiple effects. The aim of this study was to use an established thalamocortical computer model to determine how T-type calcium channels work in concert with cortical excitability to contribute to pathogenesis and treatment response in CAE. The model is comprised of cortical pyramidal, cortical inhibitory, thalamocortical relay, and thalamic reticular single-compartment neurons, implemented with Hodgkin-Huxley model ion channels and connected by AMPA, GABA A , and GABA B synapses. Network behavior was simulated for different combinations of T-type calcium channel conductance, inactivation time, steady state activation/inactivation shift, and cortical GABA A conductance. Decreasing cortical GABA A conductance and increasing T-type calcium channel conductance converted spindle to spike and wave oscillations; smaller changes were required if both were changed in concert. In contrast, left shift of steady state voltage activation/inactivation did not lead to spike and wave oscillations, whereas right shift reduced network propensity for oscillations of any type. These results provide a window into mechanisms underlying polygenic inheritance in CAE, as well as a mechanism for treatment effects and failures mediated by these channels. Although the model is a simplification of the human thalamocortical network, it serves as a useful starting point for predicting the implications of ion channel electrophysiology in polygenic epilepsy such as CAE. Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.

  10. Stereoselective inhibition of thromboxane-induced coronary vasoconstriction by 1,4-dihydropyridine calcium channel antagonists

    International Nuclear Information System (INIS)

    Eltze, M.; Boer, R.; Sanders, K.H.; Boss, H.; Ulrich, W.R.; Flockerzi, D.

    1990-01-01

    The biological activity of the (+)-S- and (-)-R-enantiomers of niguldipine, of the (-)-S- and (+)-R-enantiomers of felodipine and nitrendipine, and of rac-nisoldipine and rac-nimodipine was investigated in vitro and in vivo. Inhibition of coronary vasoconstriction due to the thromboxane A2 (TxA2)-mimetic U-46619 in guinea pig Langendorff hearts, displacement of (+)-[ 3 H]isradipine from calcium channel binding sites of guinea pig skeletal muscle T-tubule membranes, and blood pressure reduction in spontaneously hypertensive rats were determined. The enantiomers were obtained by stereoselective synthesis. Cross-contamination was less than 0.5% for both S- and R-enantiomers of niguldipine and nitrendipine and less than 1% for those of felodipine. From the doses necessary for a 50% inhibition of coronary vasoconstriction, stereoselectivity ratios for (+)-(S)-/(-)-(R)-niguldipine, (-)-(S)-/(+)-(R)-felodipine, and (-)-(S)-/(+)-(R)-nitrendipine of 28, 13, and 7, respectively, were calculated. The potency ratio rac-nisoldipine/rac-nimodipine was 3.5. Ratios obtained from binding experiments and antihypertensive activity were (+)-(S)-/(-)-(R)-niguldipine = 45 and 35, (-)-(S)-/(+)-(R)-felodipine = 12 and 13, (-)-(S)-/(+)-(R)-nitrendipine = 8 and 8, and rac-nisoldipine/rac-nimodipine = 8 and 7, respectively. Highly significant correlations were found between the in vitro potency of the substances to prevent U-46619-induced coronary vasoconstriction and their affinity for calcium channel binding sites as well as their antihypertensive activity

  11. Pharmacology of the Nav1.1 domain IV voltage sensor reveals coupling between inactivation gating processes.

    Science.gov (United States)

    Osteen, Jeremiah D; Sampson, Kevin; Iyer, Vivek; Julius, David; Bosmans, Frank

    2017-06-27

    The Na v 1.1 voltage-gated sodium channel is a critical contributor to excitability in the brain, where pathological loss of function leads to such disorders as epilepsy, Alzheimer's disease, and autism. This voltage-gated sodium (Na v ) channel subtype also plays an important role in mechanical pain signaling by primary afferent somatosensory neurons. Therefore, pharmacologic modulation of Na v 1.1 represents a potential strategy for treating excitability disorders of the brain and periphery. Inactivation is a complex aspect of Na v channel gating and consists of fast and slow components, each of which may involve a contribution from one or more voltage-sensing domains. Here, we exploit the Hm1a spider toxin, a Na v 1.1-selective modulator, to better understand the relationship between these temporally distinct modes of inactivation and ask whether they can be distinguished pharmacologically. We show that Hm1a inhibits the gating movement of the domain IV voltage sensor (VSDIV), hindering both fast and slow inactivation and leading to an increase in Na v 1.1 availability during high-frequency stimulation. In contrast, ICA-121431, a small-molecule Na v 1.1 inhibitor, accelerates a subsequent VSDIV gating transition to accelerate entry into the slow inactivated state, resulting in use-dependent block. Further evidence for functional coupling between fast and slow inactivation is provided by a Na v 1.1 mutant in which fast inactivation removal has complex effects on slow inactivation. Taken together, our data substantiate the key role of VSDIV in Na v channel fast and slow inactivation and demonstrate that these gating processes are sequential and coupled through VSDIV. These findings provide insight into a pharmacophore on VSDIV through which modulation of inactivation gating can inhibit or facilitate Na v 1.1 function.

  12. Cellular uptake of {sup 99m}TcN-NOET in human leukaemic HL-60 cells is related to calcium channel activation and cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Guillermet, Stephanie; Vuillez, Jean-Philippe; Caravel, Jean-Pierre; Marti-Batlle, Daniele; Fagret, Daniel [Universite de Grenoble, Radiopharmaceutiques Biocliniques, La Tronche (France); Fontaine, Eric [Universite de Grenoble, Laboratoire de Bioenergetique Fondamentale et Appliquee, Grenoble (France); Pasqualini, Roberto [Cis Bio International Schering SA, Gif-sur-Yvette (France)

    2006-01-01

    A major goal of nuclear oncology is the development of new radiolabelled tracers as proliferation markers. Intracellular calcium waves play a fundamental role in the course of the cell cycle. These waves occur in non-excitable tumour cells via store-operated calcium channels (SOCCs). Bis(N-ethoxy, N-ethyldithiocarbamato) nitrido technetium (V)-99m ({sup 99m}TcN-NOET) has been shown to interact with L-type voltage-operated calcium channels (VOCCs) in cultured cardiomyocytes. Considering the analogy between VOCCs and SOCCs, we sought to determine whether {sup 99m}TcN-NOET also binds to activated SOCCs in tumour cells in order to clarify the potential value of this tracer as a proliferation marker. Uptake kinetics of {sup 99m}TcN-NOET were measured in human leukaemic HL-60 cells over 60 min and the effect of several calcium channel modulators on 1-min tracer uptake was studied. The uptake kinetics of {sup 99m}TcN-NOET were compared both with the variations of cytosolic free calcium concentration measured by indo-1/AM and with the variations in the SG{sub 2}M cellular proliferation index. All calcium channel inhibitors significantly decreased the cellular uptake of {sup 99m}TcN-NOET whereas the activator thapsigargin induced a significant 10% increase. In parallel, SOCC activation by thapsigargin, as measured using the indo-1/AM probe, was inhibited by nicardipine. These results indicate that the uptake of {sup 99m}TcN-NOET is related to the activation of SOCCs. Finally, a correlation was observed between the tracer uptake and variations in the proliferation index SG{sub 2}M. The uptake of {sup 99m}TcN-NOET seems to be related to SOCC activation and to cell proliferation in HL-60 cells. These results indicate that {sup 99m}TcN-NOET might be a marker of cell proliferation. (orig.)

  13. Calcium channel blockers ameliorate iron overload-associated hepatic fibrosis by altering iron transport and stellate cell apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Ying [Department of Pharmacology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Department of Pathology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Hebei Key Laboratory of Chinese Medicine Research on Cardio-Cerebrovascular Disease, Shijiazhuang 050200, Hebei (China); Zhao, Xin [Department of Hepatobiliary Surgery, The Third Hospital of Hebei Medical University, Shijiazhuang 050051, Hebei (China); Chang, Yanzhong [Laboratory of Molecular Iron Metabolism, College of Life Science, Hebei Normal University, Shijiazhuang 050024, Hebei (China); Zhang, Yuanyuan [Department of Pharmacology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Chu, Xi [Department of Pharmacy, The Forth Hospital of Hebei Medical University, Shijiazhuang 050011, Hebei (China); Zhang, Xuan [Department of Pharmacology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Liu, Zhenyi; Guo, Hui [Department of Medicinal Chemistry, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Wang, Na [Department of Physiology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Gao, Yonggang [Department of Pharmacology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Zhang, Jianping, E-mail: zhangjianping14@126.com [Department of Pharmacology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Chu, Li, E-mail: chuli0614@126.com [Department of Pharmacology, Hebei University of Chinese Medicine, Shijiazhuang 050200, Hebei (China); Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns, Shijiazhuang 050200, Hebei (China)

    2016-06-15

    Liver fibrosis is the principal cause of morbidity and mortality in patients with iron overload. Calcium channel blockers (CCBs) can antagonize divalent cation entry into renal and myocardial cells and inhibit fibrogenic gene expression. We investigated the potential of CCBs to resolve iron overload-associated hepatic fibrosis. Kunming mice were assigned to nine groups (n = 8 per group): control, iron overload, deferoxamine, high and low dose verapamil, high and low dose nimodipine, and high and low dose diltiazem. Iron deposition and hepatic fibrosis were measured in mouse livers. Expression levels of molecules associated with transmembrane iron transport were determined by molecular biology approaches. In vitro HSC-T6 cells were randomized into nine groups (the same groups as the mice). Changes in proliferation, apoptosis, and metalloproteinase expression in cells were detected to assess the anti-fibrotic effects of CCBs during iron overload conditions. We found that CCBs reduced hepatic iron content, intracellular iron deposition, the number of hepatic fibrotic areas, collagen expression levels, and hydroxyproline content. CCBs rescued abnormal expression of α1C protein in L-type voltage-dependent calcium channel (LVDCC) and down-regulated divalent metal transporter-1 (DMT-1) expression in mouse livers. In iron-overloaded HSC-T6 cells, CCBs reduced iron deposition, inhibited proliferation, induced apoptosis, and elevated expression of matrix metalloproteinase-13 (MMP-13) and tissue inhibitor of metalloproteinase-1 (TIMP-1). CCBs are potential therapeutic agents that can be used to address hepatic fibrosis during iron overload. They resolve hepatic fibrosis probably correlated with regulating transmembrane iron transport and inhibiting HSC growth. - Highlights: • Calcium channel blockers (CCBs) reduced hepatic iron content. • CCBs decreased hepatic fibrotic areas and collagen expression levels. • CCBs resolve fibrosis by regulating iron transport and

  14. Synthetic peptides corresponding to human follicle-stimulating hormone (hFSH)-beta-(1-15) and hFSH-beta-(51-65) induce uptake of 45Ca++ by liposomes: evidence for calcium-conducting transmembrane channel formation

    Energy Technology Data Exchange (ETDEWEB)

    Grasso, P.; Santa-Coloma, T.A.; Reichert, L.E. Jr. (Department of Biochemistry, Albany Medical College, New York, NY (USA))

    1991-06-01

    We have previously described FSH receptor-mediated influx of 45Ca++ in cultured Sertoli cells from immature rats and receptor-enriched proteoliposomes via activation of voltage-sensitive and voltage-independent calcium channels. We have further shown that this effect of FSH does not require cholera toxin- or pertussis toxin-sensitive guanine nucleotide binding protein or activation of adenylate cyclase. In the present study, we have identified regions of human FSH-beta-subunit which appear to be involved in mediating calcium influx. We screened 11 overlapping peptide amides representing the entire primary structure of hFSH-beta-subunit for their effects on 45Ca++ flux in FSH receptor-enriched proteoliposomes. hFSH-beta-(1-15) and hFSH-beta-(51-65) induced uptake of 45Ca++ in a concentration-related manner. This effect of hFSH-beta-(1-15) and hFSH-beta-(51-65) was also observed in liposomes lacking incorporated FSH receptor. Reducing membrane fluidity by incubating liposomes (containing no receptor) with hFSH-beta-(1-15) or hFSH-beta-(51-65) at temperatures lower than the transition temperatures of their constituent phospholipids resulted in no significant (P greater than 0.05) difference in 45Ca++ uptake. The effectiveness of the calcium ionophore A23187, however, was abolished. Ruthenium red, a voltage-independent calcium channel antagonist, was able to completely block uptake of 45Ca++ induced by hFSH-beta-(1-15) and hFSH-beta-(51-65) whereas nifedipine, a calcium channel blocker specific for L-type voltage-sensitive calcium channels, was without effect. These results suggest that in addition to its effect on voltage-sensitive calcium channel activity, interaction of FSH with its receptor may induce formation of transmembrane aqueous channels which also facilitate influx of extracellular calcium.

  15. Rational use of calcium-channel antagonists in Raynaud's phenomenon.

    Science.gov (United States)

    Sturgill, M G; Seibold, J R

    1998-11-01

    Raynaud's phenomenon (RP) is a peripheral circulatory disorder characterized by sudden episodes of digital artery spasm, often precipitated by cold temperature or emotional stress. Although the cause of RP is not fully known, it appears to involve inappropriate adrenergic response to cold stimuli. Treatment of RP is conservative in most patients, but in patients with severe disease includes the use of agents that promote digital vasodilation. The calcium-channel antagonists, particularly the dihydropyridine derivative nifedipine, are the most thoroughly studied drug class for the treatment of RP. Approximately two thirds of patients respond favorably, with significant reductions in the frequency and severity of vasospastic attacks. Nifedipine use is often limited by the appearance of adverse vasodilatory effects such as headache or peripheral edema. The newer second-generation dihydropyridines such as amlodipine, isradipine, nicardipine, and felodipine also appear to be effective in patients with RP and may be associated with fewer adverse effects.

  16. Calcium channel blocker prevents stress-induced activation of renin and aldosterone in conscious pig

    International Nuclear Information System (INIS)

    Ceremuzynski, L.K.; Klos, J.; Barcikowski, B.; Herbaczynska-Cedro, K.

    1991-01-01

    A considerable amount of data suggest the involvement of calcium-mediated processes in the activation of the renin-angiotensin-aldosterone (RAA) cascade. To investigate the effect of calcium-channel inhibition on the RAA system, the authors studied 21 conscious pigs. Blood renin and aldosterone levels increased by subjecting animals to 24 hours of immobilization stress. Renin and aldosterone levels were repeatedly measured by radioimmunoassay in blood samples taken periodically over 24 hours from a chronically implanted arterial cannula. Pretreatment of the animals (N = 11) with nisoldipine, 2 x 20 mg p.o. daily for 2 days before and on the day of immobilization, transiently attenuated the stress-induced increase of plasma renin activity and completely prevented the rise of aldosterone, as compared to nontreated controls (N = 10). The finding that nisoldipine suppresses RAA activation induced by a nonpharmacologic stimulus in the conscious intact animal may have clinical implications

  17. Pertussis toxin-sensitive alpha-adrenergic modulation of voltage - dependent calcium channels in spontaneously hypertensive rats (SHR)

    Czech Academy of Sciences Publication Activity Database

    Zicha, Josef; Pintérová, Mária; Dobešová, Zdenka; Líšková, Silvia; Kuneš, Jaroslav

    2006-01-01

    Roč. 24, č. S6 (2006), s. 34-34 ISSN 0263-6352. [Scientific Meeting of the International Society of Hypertension /21./. 15.10.2006-19.10.2006, Fukuoka] R&D Projects: GA MZd(CZ) NR7786 Institutional research plan: CEZ:AV0Z50110509 Keywords : pertussis toxin * alpha adrenergic vasoconstriction * voltage-dependent calcium channels * SHR rat Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery

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

    NARCIS (Netherlands)

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

    2017-01-01

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

  19. Mathematical investigation of IP3-dependent calcium dynamics in astrocytes.

    Science.gov (United States)

    Handy, Gregory; Taheri, Marsa; White, John A; Borisyuk, Alla

    2017-06-01

    We study evoked calcium dynamics in astrocytes, a major cell type in the mammalian brain. Experimental evidence has shown that such dynamics are highly variable between different trials, cells, and cell subcompartments. Here we present a qualitative analysis of a recent mathematical model of astrocyte calcium responses. We show how the major response types are generated in the model as a result of the underlying bifurcation structure. By varying key channel parameters, mimicking blockers used by experimentalists, we manipulate this underlying bifurcation structure and predict how the distributions of responses can change. We find that store-operated calcium channels, plasma membrane bound channels with little activity during calcium transients, have a surprisingly strong effect, underscoring the importance of considering these channels in both experiments and mathematical settings. Variation in the maximum flow in different calcium channels is also shown to determine the range of stable oscillations, as well as set the range of frequencies of the oscillations. Further, by conducting a randomized search through the parameter space and recording the resulting calcium responses, we create a database that can be used by experimentalists to help estimate the underlying channel distribution of their cells.

  20. First direct electron microscopic visualization of a tight spatial coupling between GABAA-receptors and voltage-sensitive calcium channels

    DEFF Research Database (Denmark)

    Hansen, Gert Helge; Belhage, B; Schousboe, A

    1992-01-01

    Using cerebellar granule neurons in culture it was demonstrated that exposure of the cells to the GABAA receptor agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) leads to an increase in the number of voltage-gated calcium channels as revealed by quantitative preembedding indirect imm...

  1. The gating cycle of a K+ channel at atomic resolution

    Energy Technology Data Exchange (ETDEWEB)

    Cuello, Luis G. [Center for Membrane Protein Research, Department of Cell Physiology and Molecular Biophysics, Texas Tech University Health Sciences Center, Lubbock, United States; Cortes, D. Marien [Center for Membrane Protein Research, Department of Cell Physiology and Molecular Biophysics, Texas Tech University Health Sciences Center, Lubbock, United States; Perozo, Eduardo [Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, United States

    2017-11-22

    C-type inactivation in potassium channels helps fine-tune long-term channel activity through conformational changes at the selectivity filter. Here, through the use of cross-linked constitutively open constructs, we determined the structures of KcsA’s mutants that stabilize the selectivity filter in its conductive (E71A, at 2.25 Å) and deep C-type inactivated (Y82A at 2.4 Å) conformations. These structural snapshots represent KcsA’s transient open-conductive (O/O) and the stable open deep C-type inactivated states (O/I), respectively. The present structures provide an unprecedented view of the selectivity filter backbone in its collapsed deep C-type inactivated conformation, highlighting the close interactions with structural waters and the local allosteric interactions that couple activation and inactivation gating. Together with the structures associated with the closed-inactivated state (C/I) and in the well-known closed conductive state (C/O), this work recapitulates, at atomic resolution, the key conformational changes of a potassium channel pore domain as it progresses along its gating cycle.

  2. CO, Pb++ and SO2 effects on L-type calcium channel and action potential in human atrial myocytes. In silico study

    Directory of Open Access Journals (Sweden)

    Diana C. Pachajoa

    2017-09-01

    Full Text Available Exposure to air pollutants like carbon monoxide (CO, lead (Pb++ and sulfur dioxide (SO2 promotes the occurrence of cardiovascular diseases. Experimental studies have shown that CO, Pb++ and SO2 block L-type calcium channels, reducing the calcium current (ICaL and the action potential duration (APD, which favors the initiation of atrial arrhythmias. The goal is to study the effects of CO, Pb++ and SO2 at different concentrations on ICaL and action potential using computational simulation. For this purpose, models of the effects of the air pollutants on the atrial L-type calcium channel were developed and were incorporated into a mathematical model of a human atrial cell. The results suggest that CO, Pb++ and SO2 block the ICaL current in a fraction that increases along with the concentration, generating an APD shortening. These results are consistent with experimental studies. The combined effect of the three air pollutants produced an APD shortening, which is considered to be a pro-arrhythmic effect.

  3. Phencyclidine block of calcium current in isolated guinea-pig hippocampal neurones.

    Science.gov (United States)

    Ffrench-Mullen, J M; Rogawski, M A

    1992-10-01

    1. Phencyclidine (PCP) block of Ca2+ channel current in enzymatically dissociated neurones from the CA1 region of the adult guinea-pig hippocampus was studied using whole-cell voltage clamp techniques. Ca2+ channel current was recorded with 3 mM-Ba2+ as the charge carrier. Na+ currents were blocked with tetrodotoxin and K+ currents were eliminated by using tetraethylammonium and N-methyl-D-glucamine as the predominant extracellular and intracellular cations, respectively. 2. Peak Ca2+ channel current evoked by depolarization from -80 to -10 mV was reduced in a use-dependent fashion by PCP. The apparent forward and reverse rate constants for block at the depolarized voltage were 10(6) s-1 M-1 and 11-14 s-1, respectively. These values were at least 60 times faster than the corresponding rates at the resting voltage. The steady-state block produced by PCP increased in a concentration-dependent fashion with an IC50 of 7 microM. Other dissociative anaesthetic drugs were substantially weaker inhibitors of the current (tiletamine > dizocilpine (MK-801) > ketamine). 3. The Ca2+ channel current recorded under identical conditions in rat dorsal root ganglion neurones was less sensitive to blockade by PCP (IC50, 90 microM). 4. PCP block of the hippocampal Ca2+ channel current occurred in a voltage-dependent fashion with the fractional block decreasing at positive membrane potentials. Analysis indicated that the PCP blocking site senses 56% of the transmembrane electric field. 5. Analysis of tail currents recorded at -80 mV demonstrated that PCP does not affect the voltage-dependent or time-dependent activation or deactivation of the Ca2+ channel current. 6. The rate and extent of inactivation of the Ca2+ channel current was maximal at -10 mV and diminished at more positive potentials. Experiments with Ba(2+)-free external solution demonstrated that inactivation of the Ca2+ channels is largely voltage-dependent and is not affected by Ba2+ influx. 7. PCP markedly increased the

  4. Discrete-State Stochastic Models of Calcium-Regulated Calcium Influx and Subspace Dynamics Are Not Well-Approximated by ODEs That Neglect Concentration Fluctuations

    Science.gov (United States)

    Weinberg, Seth H.; Smith, Gregory D.

    2012-01-01

    Cardiac myocyte calcium signaling is often modeled using deterministic ordinary differential equations (ODEs) and mass-action kinetics. However, spatially restricted “domains” associated with calcium influx are small enough (e.g., 10−17 liters) that local signaling may involve 1–100 calcium ions. Is it appropriate to model the dynamics of subspace calcium using deterministic ODEs or, alternatively, do we require stochastic descriptions that account for the fundamentally discrete nature of these local calcium signals? To address this question, we constructed a minimal Markov model of a calcium-regulated calcium channel and associated subspace. We compared the expected value of fluctuating subspace calcium concentration (a result that accounts for the small subspace volume) with the corresponding deterministic model (an approximation that assumes large system size). When subspace calcium did not regulate calcium influx, the deterministic and stochastic descriptions agreed. However, when calcium binding altered channel activity in the model, the continuous deterministic description often deviated significantly from the discrete stochastic model, unless the subspace volume is unrealistically large and/or the kinetics of the calcium binding are sufficiently fast. This principle was also demonstrated using a physiologically realistic model of calmodulin regulation of L-type calcium channels introduced by Yue and coworkers. PMID:23509597

  5. The calcium channel β2 (CACNB2 subunit repertoire in teleosts

    Directory of Open Access Journals (Sweden)

    Mueller Rachel

    2008-04-01

    Full Text Available Abstract Background Cardiomyocyte contraction is initiated by influx of extracellular calcium through voltage-gated calcium channels. These oligomeric channels utilize auxiliary β subunits to chaperone the pore-forming α subunit to the plasma membrane, and to modulate channel electrophysiology 1. Several β subunit family members are detected by RT-PCR in the embryonic heart. Null mutations in mouse β2, but not in the other three β family members, are embryonic lethal at E10.5 due to defects in cardiac contractility 2. However, a drawback of the mouse model is that embryonic heart rhythm is difficult to study in live embryos due to their intra-uterine development. Moreover, phenotypes may be obscured by secondary effects of hypoxia. As a first step towards developing a model for contributions of β subunits to the onset of embryonic heart rhythm, we characterized the structure and expression of β2 subunits in zebrafish and other teleosts. Results Cloning of two zebrafish β2 subunit genes (β2.1 and β2.2 indicated they are membrane-associated guanylate kinase (MAGUK-family genes. Zebrafish β2 genes show high conservation with mammals within the SH3 and guanylate kinase domains that comprise the "core" of MAGUK proteins, but β2.2 is much more divergent in sequence than β2.1. Alternative splicing occurs at the N-terminus and within the internal HOOK domain. In both β2 genes, alternative short ATG-containing first exons are separated by some of the largest introns in the genome, suggesting that individual transcript variants could be subject to independent cis-regulatory control. In the Tetraodon nigrovidis and Fugu rubripes genomes, we identified single β2 subunit gene loci. Comparative analysis of the teleost and human β2 loci indicates that the short 5' exon sequences are highly conserved. A subset of 5' exons appear to be unique to teleost genomes, while others are shared with mammals. Alternative splicing is temporally and

  6. Contribution of S4 segments and S4-S5 linkers to the low-voltage activation properties of T-type CaV3.3 channels.

    Directory of Open Access Journals (Sweden)

    Ana Laura Sanchez-Sandoval

    Full Text Available Voltage-gated calcium channels contain four highly conserved transmembrane helices known as S4 segments that exhibit a positively charged residue every third position, and play the role of voltage sensing. Nonetheless, the activation range between high-voltage (HVA and low-voltage (LVA activated calcium channels is around 30-40 mV apart, despite the high level of amino acid similarity within their S4 segments. To investigate the contribution of S4 voltage sensors for the low-voltage activation characteristics of CaV3.3 channels we constructed chimeras by swapping S4 segments between this LVA channel and the HVA CaV1.2 channel. The substitution of S4 segment of Domain II in CaV3.3 by that of CaV1.2 (chimera IIS4C induced a ~35 mV shift in the voltage-dependence of activation towards positive potentials, showing an I-V curve that almost overlaps with that of CaV1.2 channel. This HVA behavior induced by IIS4C chimera was accompanied by a 2-fold decrease in the voltage-dependence of channel gating. The IVS4 segment had also a strong effect in the voltage sensing of activation, while substitution of segments IS4 and IIIS4 moved the activation curve of CaV3.3 to more negative potentials. Swapping of IIS4 voltage sensor influenced additional properties of this channel such as steady-state inactivation, current decay, and deactivation. Notably, Domain I voltage sensor played a major role in preventing CaV3.3 channels to inactivate from closed states at extreme hyperpolarized potentials. Finally, site-directed mutagenesis in the CaV3.3 channel revealed a partial contribution of the S4-S5 linker of Domain II to LVA behavior, with synergic effects observed in double and triple mutations. These findings indicate that IIS4 and, to a lesser degree IVS4, voltage sensors are crucial in determining the LVA properties of CaV3.3 channels, although the accomplishment of this function involves the participation of other structural elements like S4-S5 linkers.

  7. Contribution of S4 segments and S4-S5 linkers to the low-voltage activation properties of T-type CaV3.3 channels

    Science.gov (United States)

    Sanchez-Sandoval, Ana Laura; Herrera Carrillo, Zazil; Díaz Velásquez, Clara Estela; Delgadillo, Dulce María; Rivera, Heriberto Manuel

    2018-01-01

    Voltage-gated calcium channels contain four highly conserved transmembrane helices known as S4 segments that exhibit a positively charged residue every third position, and play the role of voltage sensing. Nonetheless, the activation range between high-voltage (HVA) and low-voltage (LVA) activated calcium channels is around 30–40 mV apart, despite the high level of amino acid similarity within their S4 segments. To investigate the contribution of S4 voltage sensors for the low-voltage activation characteristics of CaV3.3 channels we constructed chimeras by swapping S4 segments between this LVA channel and the HVA CaV1.2 channel. The substitution of S4 segment of Domain II in CaV3.3 by that of CaV1.2 (chimera IIS4C) induced a ~35 mV shift in the voltage-dependence of activation towards positive potentials, showing an I-V curve that almost overlaps with that of CaV1.2 channel. This HVA behavior induced by IIS4C chimera was accompanied by a 2-fold decrease in the voltage-dependence of channel gating. The IVS4 segment had also a strong effect in the voltage sensing of activation, while substitution of segments IS4 and IIIS4 moved the activation curve of CaV3.3 to more negative potentials. Swapping of IIS4 voltage sensor influenced additional properties of this channel such as steady-state inactivation, current decay, and deactivation. Notably, Domain I voltage sensor played a major role in preventing CaV3.3 channels to inactivate from closed states at extreme hyperpolarized potentials. Finally, site-directed mutagenesis in the CaV3.3 channel revealed a partial contribution of the S4-S5 linker of Domain II to LVA behavior, with synergic effects observed in double and triple mutations. These findings indicate that IIS4 and, to a lesser degree IVS4, voltage sensors are crucial in determining the LVA properties of CaV3.3 channels, although the accomplishment of this function involves the participation of other structural elements like S4-S5 linkers. PMID:29474447

  8. Contribution of S4 segments and S4-S5 linkers to the low-voltage activation properties of T-type CaV3.3 channels.

    Science.gov (United States)

    Sanchez-Sandoval, Ana Laura; Herrera Carrillo, Zazil; Díaz Velásquez, Clara Estela; Delgadillo, Dulce María; Rivera, Heriberto Manuel; Gomora, Juan Carlos

    2018-01-01

    Voltage-gated calcium channels contain four highly conserved transmembrane helices known as S4 segments that exhibit a positively charged residue every third position, and play the role of voltage sensing. Nonetheless, the activation range between high-voltage (HVA) and low-voltage (LVA) activated calcium channels is around 30-40 mV apart, despite the high level of amino acid similarity within their S4 segments. To investigate the contribution of S4 voltage sensors for the low-voltage activation characteristics of CaV3.3 channels we constructed chimeras by swapping S4 segments between this LVA channel and the HVA CaV1.2 channel. The substitution of S4 segment of Domain II in CaV3.3 by that of CaV1.2 (chimera IIS4C) induced a ~35 mV shift in the voltage-dependence of activation towards positive potentials, showing an I-V curve that almost overlaps with that of CaV1.2 channel. This HVA behavior induced by IIS4C chimera was accompanied by a 2-fold decrease in the voltage-dependence of channel gating. The IVS4 segment had also a strong effect in the voltage sensing of activation, while substitution of segments IS4 and IIIS4 moved the activation curve of CaV3.3 to more negative potentials. Swapping of IIS4 voltage sensor influenced additional properties of this channel such as steady-state inactivation, current decay, and deactivation. Notably, Domain I voltage sensor played a major role in preventing CaV3.3 channels to inactivate from closed states at extreme hyperpolarized potentials. Finally, site-directed mutagenesis in the CaV3.3 channel revealed a partial contribution of the S4-S5 linker of Domain II to LVA behavior, with synergic effects observed in double and triple mutations. These findings indicate that IIS4 and, to a lesser degree IVS4, voltage sensors are crucial in determining the LVA properties of CaV3.3 channels, although the accomplishment of this function involves the participation of other structural elements like S4-S5 linkers.

  9. 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

  10. Procaine rapidly inactivates acetylcholine receptors from Torpedo and competes with agonist for inhibition sites

    International Nuclear Information System (INIS)

    Forman, S.A.; Miller, K.W.

    1989-01-01

    The relationship between the high-affinity procaine channel inhibition site and the agonist self-inhibition site on acetylcholine receptors (AChRs) from Torpedo electroplaque was investigated by using rapid 86 Rb + quenched-flux assays at 4 degree C in native AChR-rich vesicles on which 50-60% of ACh activation sites were blocked with α-bungarotoxin (α-BTX). In the presence of channel-activating acetylcholine (ACh) concentrations alone, AChR undergoes one phase of inactivation in under a second. Addition of procaine produces two-phase inactivation similar to that seen with self-inhibiting ACh concentrations rapid inactivation complete in 30-75 ms is followed by fast desensitization at the same k d observed without procaine. The dependence of k r on [procaine] is consistent with a bimolecular association between procaine and its AChR site. Inhibition of AChR function by mixtures of procaine plus self-inhibiting concentrations of ACh or suberyldicholine was studied by reducing the level of α-BTX block in vesicles. The data support a mechanism where procaine binds preferentially to the open-channel AChR state, since no procaine-induced inactivation is observed without agonist and k r 's dependence on [ACh] in channel-activating range closely parallels that of 86 Rb + flux response to ACh

  11. The Kinetics and the Permeation Properties of Piezo Channels.

    Science.gov (United States)

    Gnanasambandam, R; Gottlieb, P A; Sachs, F

    2017-01-01

    Piezo channels are eukaryotic, cation-selective mechanosensitive channels (MSCs), which show rapid activation and voltage-dependent inactivation. The kinetics of these channels are largely consistent across multiple cell types and different stimulation paradigms with some minor variability. No accessory subunits that associate with Piezo channels have been reported. They are homotrimers and each ∼300kD monomer has an N-terminal propeller blade-like mechanosensing module, which can confer mechanosensing capabilities on ASIC-1 (the trimeric non-MSC, acid-sensing ion channel-1) and a C-terminal pore module, which influences conductance, selectivity, and channel inactivation. Repeated stimulation can cause domain fracture and diffusion of these channels leading to synchronous loss of inactivation. The reconstituted channels spontaneously open only in asymmetric bilayers but lack inactivation. Mutations that cause hereditary xerocytosis alter PIEZO1 kinetics. The kinetics of the wild-type PIEZO1 and alterations thereof in mutants (M2225R, R2456K, and DhPIEZO1) are summarized in the form of a quantitative model and hosted online. The pore is permeable to alkali ions although Li + permeates poorly. Divalent cations, notably Ca 2+ , traverse the channel and inhibit the flux of monovalents. The large monovalent organic cations such as tetramethyl ammonium and tetraethyl ammonium can traverse the channel, but slowly, suggesting a pore diameter of ∼8Å, and the estimated in-plane area change upon opening is around 6-20nm 2 . Ruthenium red can enter the channel only from the extracellular side and seems to bind in a pocket close to residue 2496. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Is Shock Index a Valid Predictor of Mortality in Emergency Department Patients With Hypertension, Diabetes, High Age, or Receipt of β- or Calcium Channel Blockers?

    DEFF Research Database (Denmark)

    Kristensen, Anders K B; Holler, Jon G; Hallas, Jesper

    2016-01-01

    STUDY OBJECTIVE: Shock index is a widely reported tool to identify patients at risk for circulatory collapse. We hypothesize that old age, diabetes, hypertension, and β- or calcium channel blockers weaken the association between shock index and mortality. METHODS: This was a cohort study of all...... first-time emergency department (ED) visits between 1995 and 2011 (n=111,019). We examined whether age 65 years or older, diabetes, hypertension, and use of β- or calcium channel blockers modified the association between shock index and 30-day mortality. RESULTS: The 30-day mortality was 3.0%. For all...... than or equal to 1 in patients aged 65 years or older was 8.2 (95% CI 7.2 to 9.4) compared with 18.9 (95% CI 15.6 to 23.0) in younger patients. β- Or calcium channel-blocked patients had an OR of 6.4 (95% CI 4.9 to 8.3) versus 12.3 (95% CI 11.0 to 13.8) in nonusers and hypertensive patients had...

  13. Anti-Convulsant Activity of Boerhaavia diffusa: Plausible Role of Calcium Channel Antagonism

    Directory of Open Access Journals (Sweden)

    Mandeep Kaur

    2011-01-01

    Full Text Available “Ethnopharmacological” use of roots of Boerhaavia diffusa (B. diffusa in the treatment of epilepsy in Nigerian folk medicine and reports showing the presence of a calcium channel antagonistic compound “liriodendrin” in its roots, led us to undertake the present study. The study was designed to investigate the methanolic root extract of B. diffusa and its different fractions including liriodendrin-rich fraction for exploring the possible role of liriodendrin in its anti-convulsant activity. Air-dried roots of B. diffusa were extracted with methanol by cold maceration. The methanol soluble fraction of extract thus obtained was successively extracted to obtain liriodendrin-rich fraction and two side fractions, that is, chloroform fraction and phenolic compound fraction. Anti-convulsant activity of methanolic extract (1000, 1500 and 2000 mg kg-1, intraperitoneally (i.p. and its different fractions, that is, liriodendrin-rich fraction (10, 20 and 40 mg kg-1, i.p., chloroform fraction (20 mg kg-1, i.p. and phenolic compound fraction (1 mg kg-1, i.p. were studied in pentylenetetrazol (PTZ-induced seizures (75 mg kg-1, i.p.. The crude methanolic extract of B. diffusa and only its liriodendrin-rich fraction showed a dose-dependent protection against PTZ-induced convulsions. The liriodendrin-rich fraction also showed significant protection against seizures induced by BAY k-8644. These findings reiterated the anti-convulsant activity of methanolic extract of B. diffusa roots. Furthermore, it can be concluded that the observed anti-convulsant activity was due to its calcium channel antagonistic action as this activity was retained only in the liodendrin-rich fraction, which has additionally been confirmed by significant anti-convulsant activity of liriodendrin-rich fraction in BAY k-8644-induced seizures.

  14. β1 subunit stabilises sodium channel Nav1.7 against mechanical stress.

    Science.gov (United States)

    Körner, Jannis; Meents, Jannis; Machtens, Jan-Philipp; Lampert, Angelika

    2018-06-01

    The voltage-gated sodium channel Nav1.7 is a key player in neuronal excitability and pain signalling. In addition to voltage sensing, the channel is also modulated by mechanical stress. Using whole-cell patch-clamp experiments, we discovered that the sodium channel subunit β1 is able to prevent the impact of mechanical stress on Nav1.7. An intramolecular disulfide bond of β1 was identified to be essential for stabilisation of inactivation, but not activation, against mechanical stress using molecular dynamics simulations, homology modelling and site-directed mutagenesis. Our results highlight the role of segment 6 of domain IV in fast inactivation. We present a candidate mechanism for sodium channel stabilisation against mechanical stress, ensuring reliable channel functionality in living systems. Voltage-gated sodium channels are key players in neuronal excitability and pain signalling. Precise gating of these channels is crucial as even small functional alterations can lead to pathological phenotypes such as pain or heart failure. Mechanical stress has been shown to affect sodium channel activation and inactivation. This suggests that stabilising components are necessary to ensure precise channel gating in living organisms. Here, we show that mechanical shear stress affects voltage dependence of activation and fast inactivation of the Nav1.7 channel. Co-expression of the β1 subunit, however, protects both gating modes of Nav1.7 against mechanical shear stress. Using molecular dynamics simulation, homology modelling and site-directed mutagenesis, we identify an intramolecular disulfide bond of β1 (Cys21-Cys43) which is partially involved in this process: the β1-C43A mutant prevents mechanical modulation of voltage dependence of activation, but not of fast inactivation. Our data emphasise the unique role of segment 6 of domain IV for sodium channel fast inactivation and confirm previous reports that the intracellular process of fast inactivation can be

  15. A sensor for calcium uptake

    Science.gov (United States)

    Collins, Sean; Meyer, Tobias

    2011-01-01

    Mitochondria — the cell’s power plants — increase their energy production in response to calcium signals in the cytoplasm. A regulator of the elusive mitochondrial calcium channel has now been identified. PMID:20844529

  16. H2O2 augments cytosolic calcium in nucleus tractus solitarii neurons via multiple voltage-gated calcium channels.

    Science.gov (United States)

    Ostrowski, Tim D; Dantzler, Heather A; Polo-Parada, Luis; Kline, David D

    2017-05-01

    Reactive oxygen species (ROS) play a profound role in cardiorespiratory function under normal physiological conditions and disease states. ROS can influence neuronal activity by altering various ion channels and transporters. Within the nucleus tractus solitarii (nTS), a vital brainstem area for cardiorespiratory control, hydrogen peroxide (H 2 O 2 ) induces sustained hyperexcitability following an initial depression of neuronal activity. The mechanism(s) associated with the delayed hyperexcitability are unknown. Here we evaluate the effect(s) of H 2 O 2 on cytosolic Ca 2+ (via fura-2 imaging) and voltage-dependent calcium currents in dissociated rat nTS neurons. H 2 O 2 perfusion (200 µM; 1 min) induced a delayed, slow, and moderate increase (~27%) in intracellular Ca 2+ concentration ([Ca 2+ ] i ). The H 2 O 2 -mediated increase in [Ca 2+ ] i prevailed during thapsigargin, excluding the endoplasmic reticulum as a Ca 2+ source. The effect, however, was abolished by removal of extracellular Ca 2+ or the addition of cadmium to the bath solution, suggesting voltage-gated Ca 2+ channels (VGCCs) as targets for H 2 O 2 modulation. Recording of the total voltage-dependent Ca 2+ current confirmed H 2 O 2 enhanced Ca 2+ entry. Blocking VGCC L, N, and P/Q subtypes decreased the number of cells and their calcium currents that respond to H 2 O 2 The number of responder cells to H 2 O 2 also decreased in the presence of dithiothreitol, suggesting the actions of H 2 O 2 were dependent on sulfhydryl oxidation. In summary, here, we have shown that H 2 O 2 increases [Ca 2+ ] i and its Ca 2+ currents, which is dependent on multiple VGCCs likely by oxidation of sulfhydryl groups. These processes presumably contribute to the previously observed delayed hyperexcitability of nTS neurons in in vitro brainstem slices. Copyright © 2017 the American Physiological Society.

  17. Autoimmune encephalitis associated with voltage-gated potassium channels-complex and leucine-rich glioma-inactivated 1 antibodies

    DEFF Research Database (Denmark)

    Celicanin, Marko; Blaabjerg, M; Maersk-Moller, C

    2017-01-01

    BACKGROUND AND PURPOSE: The aim of this study was to describe clinical and paraclinical characteristics of all Danish patients who tested positive for anti-voltage-gated potassium channels (VGKC)-complex, anti-leucine-rich glioma-inactivated 1 (LGI1) and anti-contactin-associated protein-2......, electroencephalography and (18) F-fluorodeoxyglucose positron emission tomography scans were re-evaluated by experts in the field. RESULTS: A total of 28/192 patients tested positive for VGKC-complex antibodies by radioimmunoassay and indirect immunofluorescence; 17 had antibodies to LGI1 and 6/7 of the available....... CONCLUSIONS: Patients diagnosed with anti-LGI1 autoimmune encephalitis increased significantly from 2009 to 2014, probably due to increased awareness. In contrast to seropositive anti-VGKC-complex patients, all anti-LGI1-positive patients presented with a classical limbic encephalitis. The majority...

  18. Biological evaluation of [18F]-nifedipine as a novel PET tracer for L-type calcium channel imaging

    International Nuclear Information System (INIS)

    Sadeghpour, H.; Jalilian, A.R.; Akhlaghi, M.; Mirzaii, M.; Saddadi, F.; Shafiee, A.; Miri, R.

    2008-01-01

    Due to interesting role of dihydropyridines in cardiovascular diseases and drug resistance studies and lack of a fluorine-18 labeled imaging agent for L-type calcium channel studies, this study was designed. [ 18 F] Dimethyl 2 - (fluoromethyl) - 6 - methyl - 4 - (2 - nitrophenyl) - 1,4 - dihydropyridine - 3,5 - dicarboxylate 2 was prepared in no-carrier-added (n.c.a.) form from a starting brominated compound in one step at 80 o C in Kryptofix2.2.2/[ 18 F]. Compound 2 was administered to normal rats via their tail veins for preliminary biodistribution studies and the ID/g % of the labeled compound was determined up to 3 h post injections. Coincidence images were obtained in rats 5 to 120 min. Radiofluorination on bromo precursor gave a fluorinated compound in 95 % radiochemical purity and a 8% yield shown by RTLC and HPLC. Biodistribution studies showed that the tracer is accumulated in the heart in the first few minutes, followed by metabolism resulting in very soluble 18 F-containing metabolites eliminated through the urinary tract. In coincidence images, the target organ was shown to be the heart. Lung had high accumulation possibly due to the presence of Ca 2+ channels and/or hydrolyzing enzymes showing a significant myocardial uptake at 120 min. The data demonstrates a significant agreement with the reported L-type calcium channels throughout the animal body. To our knowledge, this is the first example of 18 F-DHPs in the literature. (authors)

  19. Antibodies to voltage-gated potassium and calcium channels in epilepsy.

    Science.gov (United States)

    Majoie, H J Marian; de Baets, Mark; Renier, Willy; Lang, Bethan; Vincent, Angela

    2006-10-01

    To determine the prevalence of antibodies to ion channels in patients with long standing epilepsy. Although the CNS is thought to be protected from circulating antibodies by the blood brain barrier, glutamate receptor antibodies have been reported in Rasmussen's encephalitis, glutamic acid decarboxylase (GAD) antibodies have been found in a few patients with epilepsy, and antibodies to voltage-gated potassium channels (VGKC) have been found in a non-paraneoplastic form of limbic encephalitis (with amnesia and seizures) that responds to immunosuppressive therapy. We retrospectively screened sera from female epilepsy patients (n=106) for autoantibodies to VGKC (Kv 1.1, 1.2 or 1.6), voltage-gated calcium channels (VGCC) (P/Q-type), and GAD. All positive results, based on the values of control data [McKnight, K., Jiang, Y., et al. (2005). Serum antibodies in epilepsy and seizure-associated disorders. Neurology 65, 1730-1735], were retested at lower serum concentrations, and results compared with previously published control data. Demographics, medical history, and epilepsy related information was gathered. The studied group consisted predominantly of patients with long standing drug resistant epilepsy. VGKC antibodies were raised (>100 pM) in six patients. VGCC antibodies (>45 pM) were slightly raised in only one patient. GAD antibodies were VGKC antibodies differed from previously described patients with limbic encephalitis-like syndrome, and were not different with respect to seizure type, age at first seizure, duration of epilepsy, or use of anti-epileptic drugs from the VGKC antibody negative patients. The results demonstrate that antibodies to VGKC are present in 6% of patients with typical long-standing epilepsy, but whether these antibodies are pathogenic or secondary to the primary disease process needs to be determined.

  20. Calcium regulation and Alzheimer’s disease

    Directory of Open Access Journals (Sweden)

    Deepthi Rapaka

    2014-09-01

    Full Text Available Activation of the neuron induces transient fluctuations in [Ca2+]i. This transient rise in [Ca2+]i is dependent on calcium entry via calcium channels and release of calcium from intracellular stores, finally resulting in increase in calcium levels, which activates calcium regulatory proteins to restore the resting calcium levels by binding to the calcium-binding proteins, sequestration into the endoplasmic reticulum and the mitochondria, and finally extrusion of calcium spike potential from the cell by adenosine triphosphate-driven Ca2+ pumps and the Na+/Ca2+ exchanger. Improper regulation of calcium signaling, sequentially, likely contributes to synaptic dysfunction and excitotoxic and/or apoptotic death of the vulnerable neuronal populations. The cognitive decline associated with normal aging is not only due to neuronal loss, but is fairly the result of synaptic connectivity. Many evidences support that Ca2+ dyshomeostasis is implicated in normal brain aging. Thus the chief factor associated with Alzheimer’s disease was found to be increase in the levels of free intracellular calcium, demonstrating that the excessive levels might lead to cell death, which provides a key target for the calcium channel blockers might be used as the neuroprotective agents in Alzheimer’s disease.

  1. Role of volume-regulated and calcium-activated anion channels in cell volume homeostasis, cancer and drug resistance

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay; Sørensen, Belinda Halling; Sauter, Daniel Rafael Peter

    2015-01-01

    to be an essential component of both VRAC and VSOAC. Reduced VRAC and VSOAC activities are seen in drug resistant cancer cells. ANO1 is a calcium-activated chloride channel expressed on the plasma membrane of e.g. secretory epithelia. ANO1 is amplified and highly expressed in a large number of carcinomas. The gene...... functions as well as their role in cancer and drug resistance....

  2. Polarized axonal surface expression of neuronal KCNQ potassium channels is regulated by calmodulin interaction with KCNQ2 subunit.

    Directory of Open Access Journals (Sweden)

    John P Cavaretta

    Full Text Available KCNQ potassium channels composed of KCNQ2 and KCNQ3 subunits give rise to the M-current, a slow-activating and non-inactivating voltage-dependent potassium current that limits repetitive firing of action potentials. KCNQ channels are enriched at the surface of axons and axonal initial segments, the sites for action potential generation and modulation. Their enrichment at the axonal surface is impaired by mutations in KCNQ2 carboxy-terminal tail that cause benign familial neonatal convulsion and myokymia, suggesting that their correct surface distribution and density at the axon is crucial for control of neuronal excitability. However, the molecular mechanisms responsible for regulating enrichment of KCNQ channels at the neuronal axon remain elusive. Here, we show that enrichment of KCNQ channels at the axonal surface of dissociated rat hippocampal cultured neurons is regulated by ubiquitous calcium sensor calmodulin. Using immunocytochemistry and the cluster of differentiation 4 (CD4 membrane protein as a trafficking reporter, we demonstrate that fusion of KCNQ2 carboxy-terminal tail is sufficient to target CD4 protein to the axonal surface whereas inhibition of calmodulin binding to KCNQ2 abolishes axonal surface expression of CD4 fusion proteins by retaining them in the endoplasmic reticulum. Disruption of calmodulin binding to KCNQ2 also impairs enrichment of heteromeric KCNQ2/KCNQ3 channels at the axonal surface by blocking their trafficking from the endoplasmic reticulum to the axon. Consistently, hippocampal neuronal excitability is dampened by transient expression of wild-type KCNQ2 but not mutant KCNQ2 deficient in calmodulin binding. Furthermore, coexpression of mutant calmodulin, which can interact with KCNQ2/KCNQ3 channels but not calcium, reduces but does not abolish their enrichment at the axonal surface, suggesting that apo calmodulin but not calcium-bound calmodulin is necessary for their preferential targeting to the axonal

  3. Calcium regulates caveolin-1 expression at the transcriptional level

    International Nuclear Information System (INIS)

    Yang, Xiao-Yan; Huang, Cheng-Cheng; Kan, Qi-Ming; Li, Yan; Liu, Dan; Zhang, Xue-Cheng; Sato, Toshinori; Yamagata, Sadako; Yamagata, Tatsuya

    2012-01-01

    Highlights: ► Caveolin-1 expression is regulated by calcium signaling at the transcriptional level. ► An inhibitor of or siRNA to L-type calcium channel suppressed caveolin-1 expression. ► Cyclosporine A or an NFAT inhibitor markedly reduced caveolin-1 expression. ► 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 2+ /calcineurin/NFAT.

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

    Science.gov (United States)

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

    2009-06-01

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

  5. Modulatory effects of the fruits of Tribulus terrestris L. on the function of atopic dermatitis-related calcium channels, Orai1 and TRPV3

    Directory of Open Access Journals (Sweden)

    Joo Hyun Nam

    2016-07-01

    Conclusions: Our results suggest that T. terrestris extract may have a therapeutic potential for recovery of abnormal skin barrier pathologies in atopic dermatitis through modulating the activities of calcium ion channels, Orai1 and TRPV3. This is the first study to report the modulatory effect of a medicinal plant on the function of ion channels in skin barrier.

  6. MacMARCKS interacts with the metabotropic glutamate receptor type 7 and modulates G protein-mediated constitutive inhibition of calcium channels

    Czech Academy of Sciences Publication Activity Database

    Bertaso, F.; Lill, Y.; Airas, J.M.; Espeut, J.; Blahoš, Jaroslav; Bockaert, J.; Fagni, L.; Betz, H.; Far, O.E.

    2006-01-01

    Roč. 99, - (2006), s. 288-298 ISSN 0022-3042 R&D Projects: GA ČR GA204/05/0920 Institutional research plan: CEZ:AV0Z50390512 Keywords : calmodulin * metabotropic glutamate receptor * calcium channel Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.260, year: 2006

  7. Modulation of CaV1.2 calcium channel by neuropeptide W regulates vascular myogenic tone via G protein-coupled receptor 7.

    Science.gov (United States)

    Ji, Li; Zhu, Huayuan; Chen, Hong; Fan, Wenyong; Chen, Junjie; Chen, Jing; Zhu, Guoqing; Wang, Juejin

    2015-12-01

    Neuropeptide W (NPW), an endogenous ligand for the G protein-coupled receptor 7 (GPR7), was first found to make important roles in central nerve system. In periphery, NPW was also present and regulated intracellular calcium homeostasis by L-type calcium channels. This study was designed to discover the effects of NPW-GPR7 on the function of CaV1.2 calcium channels in the vascular smooth muscle cells (VSMCs) and vasotone of arterial vessels. By whole-cell patch clamp, we studied the effects of NPW-23, the active form of NPW, on the CaV1.2 channels in the heterologously transfected human embryonic kidney 293 cells and VSMCs isolated from rat. Living system was used to explore the physiological function of NPW-23 in arterial myogenic tone. To investigate the pathological relevance, NPW mRNA level of mesenteric arteries was measured in the hypertensive and normotensive rats. NPW's receptor GPR7 was coexpressed with CaV1.2 channels in arterial smooth muscle. NPW-23 increased the ICa,L in transfected human embryonic kidney 293 cells and VSMCs via GPR7, which could be abrogated by phospholipase C (PLC)/protein kinase C (PKC) inhibitors, not protein kinase A or protein kinase G inhibitor. After NPW-23 application, the expression of pan phospho-PKC was increased; moreover, intracellular diacylglycerol level, the second messenger catalyzed by PLC, was increased 1.5-2-fold. Application with NPW-23 increased pressure-induced vasotone of the rat mesenteric arteries. Importantly, the expression of NPW was decreased in the hypertensive rats. NPW-23 regulates ICa,L via GPR7, which is mediated by PLC/PKC signaling, and such a mechanism plays a role in modulating vascular myogenic tone, which may involve in the development of vascular hypertension.

  8. Opening of small and intermediate calcium-activated potassium channels induces relaxation mainly mediated by nitric-oxide release in large arteries and endothelium-derived hyperpolarizing factor in small arteries from rat

    DEFF Research Database (Denmark)

    Stankevicius, Edgaras; Dalsgaard, Thomas; Kroigaard, Christel

    2011-01-01

    This study was designed to investigate whether calcium-activated potassium channels of small (SK(Ca) or K(Ca)2) and intermediate (IK(Ca) or K(Ca)3.1) conductance activated by 6,7-dichloro-1H-indole-2,3-dione 3-oxime (NS309) are involved in both nitric oxide (NO) and endothelium-derived hyperpolar......This study was designed to investigate whether calcium-activated potassium channels of small (SK(Ca) or K(Ca)2) and intermediate (IK(Ca) or K(Ca)3.1) conductance activated by 6,7-dichloro-1H-indole-2,3-dione 3-oxime (NS309) are involved in both nitric oxide (NO) and endothelium...... in human umbilical vein endothelial cells (HUVECs), and calcium concentrations were investigated in both HUVECs and mesenteric arterial endothelial cells. In both superior (∼1093 μm) and small mesenteric (∼300 μm) arteries, NS309 evoked endothelium- and concentration-dependent relaxations. In superior....... In small mesenteric arteries, NS309 relaxations were reduced slightly by ADMA, whereas apamin plus an IK(Ca) channel blocker almost abolished relaxation. Iberiotoxin did not change NS309 relaxation. HUVECs expressed mRNA for SK(Ca) and IK(Ca) channels, and NS309 induced increases in calcium, outward...

  9. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    OpenAIRE

    Oyku Gulmez; Ilyas Atar; Bülent Ozin; Mehmet Emin Korkmaz; Asli Atar; et al

    2008-01-01

    Oyku Gulmez, Ilyas Atar, Bülent Ozin, Mehmet Emin Korkmaz, Asli Atar, Alp Aydinalp, Aylin Yildirir, Haldun MuderrisogluBaskent University Faculty of Medicine, Department of Cardiology, Ankara, TurkeyBackground: Use of intracoronary calcium channel blockers (CCBs) during percutaneous coronary intervention (PCI) has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB) levels in pat...

  10. Inactivation of high concentration of pathogens in land-applied food ...

    African Journals Online (AJOL)

    In Mexico, as in other developing countries, the most important pollution and management problems of food-processing sludge are the high levels of pathogen microorganisms within the sludge and the lack of sites for its disposal. The aims of this study were to evaluate the effect of calcium oxide in the inactivation of ...

  11. Cryo-EM structures of the TMEM16A calcium-activated chloride channel.

    Science.gov (United States)

    Dang, Shangyu; Feng, Shengjie; Tien, Jason; Peters, Christian J; Bulkley, David; Lolicato, Marco; Zhao, Jianhua; Zuberbühler, Kathrin; Ye, Wenlei; Qi, Lijun; Chen, Tingxu; Craik, Charles S; Jan, Yuh Nung; Minor, Daniel L; Cheng, Yifan; Jan, Lily Yeh

    2017-12-21

    Calcium-activated chloride channels (CaCCs) encoded by TMEM16A control neuronal signalling, smooth muscle contraction, airway and exocrine gland secretion, and rhythmic movements of the gastrointestinal system. To understand how CaCCs mediate and control anion permeation to fulfil these physiological functions, knowledge of the mammalian TMEM16A structure and identification of its pore-lining residues are essential. TMEM16A forms a dimer with two pores. Previous CaCC structural analyses have relied on homology modelling of a homologue (nhTMEM16) from the fungus Nectria haematococca that functions primarily as a lipid scramblase, as well as subnanometre-resolution electron cryo-microscopy. Here we present de novo atomic structures of the transmembrane domains of mouse TMEM16A in nanodiscs and in lauryl maltose neopentyl glycol as determined by single-particle electron cryo-microscopy. These structures reveal the ion permeation pore and represent different functional states. The structure in lauryl maltose neopentyl glycol has one Ca 2+ ion resolved within each monomer with a constricted pore; this is likely to correspond to a closed state, because a CaCC with a single Ca 2+ occupancy requires membrane depolarization in order to open (C.J.P. et al., manuscript submitted). The structure in nanodiscs has two Ca 2+ ions per monomer and its pore is in a closed conformation; this probably reflects channel rundown, which is the gradual loss of channel activity that follows prolonged CaCC activation in 1 mM Ca 2+ . Our mutagenesis and electrophysiological studies, prompted by analyses of the structures, identified ten residues distributed along the pore that interact with permeant anions and affect anion selectivity, as well as seven pore-lining residues that cluster near pore constrictions and regulate channel gating. Together, these results clarify the basis of CaCC anion conduction.

  12. Optimization of TRPV6 Calcium Channel Inhibitors Using a 3D Ligand-Based Virtual Screening Method.

    OpenAIRE

    Simonin Céline; Awale Mahendra; Brand Michael; van Deursen Ruud; Schwartz Julian; Fine Michael; Kovacs Gergely; Häfliger Pascal; Gyimesi Gergely; Sithampari Abilashan; Charles Roch-Philippe; Hediger Matthias A; Reymond Jean-Louis

    2015-01-01

    Herein we report the discovery of the first potent and selective inhibitor of TRPV6 a calcium channel overexpressed in breast and prostate cancer and its use to test the effect of blocking TRPV6 mediated Ca(2+) influx on cell growth. The inhibitor was discovered through a computational method xLOS a 3D shape and pharmacophore similarity algorithm a type of ligand based virtual screening (LBVS) method described briefly here. Starting with a single weakly active seed molecule two successive rou...

  13. Ion Channels of Pituitary Gonadotrophs and Their Roles in Signaling and Secretion

    Directory of Open Access Journals (Sweden)

    Stanko S. Stojilkovic

    2017-06-01

    Full Text Available Gonadotrophs are basophilic cells of the anterior pituitary gland specialized to secrete gonadotropins in response to elevation in intracellular calcium concentration. These cells fire action potentials (APs spontaneously, coupled with voltage-gated calcium influx of insufficient amplitude to trigger gonadotropin release. The spontaneous excitability of gonadotrophs reflects the expression of voltage-gated sodium, calcium, potassium, non-selective cation-conducting, and chloride channels at their plasma membrane (PM. These cells also express the hyperpolarization-activated and cyclic nucleotide-gated cation channels at the PM, as well as GABAA, nicotinic, and purinergic P2X channels gated by γ-aminobutyric acid (GABA, acetylcholine (ACh, and ATP, respectively. Activation of these channels leads to initiation or amplification of the pacemaking activity, facilitation of calcium influx, and activation of the exocytic pathway. Gonadotrophs also express calcium-conducting channels at the endoplasmic reticulum membranes gated by inositol trisphosphate and intracellular calcium. These channels are activated potently by hypothalamic gonadotropin-releasing hormone (GnRH and less potently by several paracrine calcium-mobilizing agonists, including pituitary adenylate cyclase-activating peptides, endothelins, ACh, vasopressin, and oxytocin. Activation of these channels causes oscillatory calcium release and a rapid gonadotropin release, accompanied with a shift from tonic firing of single APs to periodic bursting type of electrical activity, which accounts for a sustained calcium signaling and gonadotropin secretion. This review summarizes our current understanding of ion channels as signaling molecules in gonadotrophs, the role of GnRH and paracrine agonists in their gating, and the cross talk among channels.

  14. Rhynchophylline from Uncaria rhynchophylla functionally turns delayed rectifiers into A-Type K+ channels.

    Science.gov (United States)

    Chou, Chun-Hsiao; Gong, Chi-Li; Chao, Chia-Chia; Lin, Chia-Huei; Kwan, Chiu-Yin; Hsieh, Ching-Liang; Leung, Yuk-Man

    2009-05-22

    Rhynchophylline (1), a neuroprotective agent isolated from the traditional Chinese medicinal herb Uncaria rhynchophylla, was shown to affect voltage-gated K(+) (Kv) channel slow inactivation in mouse neuroblastoma N2A cells. Extracellular 1 (30 microM) accelerated the slow decay of Kv currents and shifted the steady-state inactivation curve to the left. Intracellular dialysis of 1 did not accelerate the slow current decay, suggesting that this compound acts extracellularly. In addition, the percent blockage of Kv currents by this substance was independent of the degree of depolarization and the intracellular K(+) concentration. Therefore, 1 did not appear to directly block the outer channel pore, with the results obtained suggesting that it drastically accelerated Kv channel slow inactivation. Interestingly, 1 also shifted the activation curve to the left. This alkaloid also strongly accelerated slow inactivation and caused a left shift of the activation curve of Kv1.2 channels heterologously expressed in HEK293 cells. Thus, this compound functionally turned delayed rectifiers into A-type K(+) channels.

  15. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    OpenAIRE

    Gulmez, Oyku

    2008-01-01

    Oyku Gulmez, Ilyas Atar, Bülent Ozin, Mehmet Emin Korkmaz, Aslı Atar, Alp Aydinalp, Aylin Yildirir, Haldun MuderrisogluBaskent University Faculty of Medicine, Department of Cardiology, Ankara, TurkeyBackground: Use of intracoronary calcium channel blockers (CCBs) during percutaneous coronary intervention (PCI) has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB) levels in pa...

  16. Facilitation of plateau potentials in turtle motoneurones by a pathway dependent on calcium and calmodulin

    DEFF Research Database (Denmark)

    Perrier, J F; Mejia-Gervacio, S; Hounsgaard, J

    2000-01-01

    1. The involvement of intracellular calcium and calmodulin in the modulation of plateau potentials in motoneurones was investigated using intracellular recordings from a spinal cord slice preparation. 2. Chelation of intracellular calcium with BAPTA-AM or inactivation of calmodulin with W-7 or tr...

  17. Sodium channels as targets for volatile anesthetics

    Directory of Open Access Journals (Sweden)

    Karl F. Herold

    2012-03-01

    Full Text Available The molecular mechanisms of modern inhaled anesthetics although widely used in clinical settings are still poorly understood. Considerable evidence supports effects on membrane proteins such as ligand- and voltage-gated ion channels of excitable cells. Na+ channels are crucial to action potential initiation and propagation, and represent potential targets for volatile anesthetics. Inhibition of presynaptic Na+ channels leads to reduced neurotransmitter release at the synapse and could therefore contribute to the mechanisms by which volatile anesthetics produce their characteristic effects: amnesia, unconsciousness, and immobility. Early studies on crayfish and squid giant axon showed inhibition of Na+ currents by volatile anesthetics. Subsequent studies using native neuronal preparations and heterologous expression systems with various mammalian Na+ channel isoforms implicated inhibition of presynaptic Na+ channels in anesthetic actions. Volatile anesthetics reduce peak Na+ current and shift the voltage of half-maximal steady-state inactivation towards more negative potentials, thus stabilizing the fast-inactivated state. Furthermore recovery from fast-inactivation is slowed together with an enhanced use-dependent block during pulse train protocols. These effects can reduce neurotransmitter release by depressing presynaptic excitability, depolarization and Ca entry, and ultimately transmitter release. This reduction in transmitter release is more portent for glutamatergic vs. GABAergic terminals. Involvement of Na+ channel inhibition in mediating the immobility caused by volatile anesthetics has been demonstrated in animal studies, in which intrathecal infusion of the Na+ channel blocker tetrodotoxin increases volatile anesthetic potency, whereas infusion of the Na+ channels agonist veratridine reduces anesthetic potency. These studies indicate that inhibition of presynaptic Na+ channels by volatile anesthetics is involved in mediating some of

  18. Spontaneous calcium transients in human neural progenitor cells mediated by transient receptor potential channels.

    Science.gov (United States)

    Morgan, Peter J; Hübner, Rayk; Rolfs, Arndt; Frech, Moritz J

    2013-09-15

    Calcium signals affect many developmental processes, including proliferation, migration, survival, and apoptosis, processes that are of particular importance in stem cells intended for cell replacement therapies. The mechanisms underlying Ca(2+) signals, therefore, have a role in determining how stem cells respond to their environment, and how these responses might be controlled in vitro. In this study, we examined the spontaneous Ca(2+) activity in human neural progenitor cells during proliferation and differentiation. Pharmacological characterization indicates that in proliferating cells, most activity is the result of transient receptor potential (TRP) channels that are sensitive to Gd(3+) and La(3+), with the more subtype selective antagonist Ruthenium red also reducing activity, suggesting the involvement of transient receptor potential vanilloid (TRPV) channels. In differentiating cells, Gd(3+) and La(3+)-sensitive TRP channels also appear to underlie the spontaneous activity; however, no sub-type-specific antagonists had any effect. Protein levels of TRPV2 and TRPV3 decreased in differentiated cells, which is demonstrated by western blot. Thus, it appears that TRP channels represent the main route of Ca(2+) entry in human neural progenitor cells (hNPCs), but the responsible channel types are subject to substitution under differentiating conditions. The level of spontaneous activity could be increased and decreased by lowering and raising the extracellular K(+) concentration. Proliferating cells in low K(+) slowed the cell cycle, with a disproportionate increased percentage of cells in G1 phase and a reduction in S phase. Taken together, these results suggest a link between external K(+) concentration, spontaneous Ca(2+) transients, and cell cycle distribution, which is able to influence the fate of stem and progenitor cells.

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

    Science.gov (United States)

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

    1993-03-01

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

  20. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    OpenAIRE

    Gulmez, Oyku; Atar, Ilyas; Ozin, B?lent; Korkmaz, Mehmet Emin; Atar, Asli; Aydinalp, Alp; Yildirir, Aylin; Muderrisoglu, Haldun

    2008-01-01

    Background: Use of intracoronary calcium channel blockers (CCBs) during percutaneous coronary intervention (PCI) has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB) levels in patients undergoing elective PCI. Methods: A total of 570 patients who underwent PCI were evaluated for CK-MB elevation. Patients who were on CCB therapy when admitted to the hospital constituted the CCB group. ...

  1. Differential calcium sensitivity in NaV 1.5 mixed syndrome mutants.

    Science.gov (United States)

    Abdelsayed, Mena; Baruteau, Alban-Elouen; Gibbs, Karen; Sanatani, Shubhayan; Krahn, Andrew D; Probst, Vincent; Ruben, Peter C

    2017-09-15

    SCN5a mutations may express gain-of-function (Long QT Syndrome-3), loss-of-function (Brugada Syndrome 1) or both (mixed syndromes), depending on the mutation and environmental triggers. One such trigger may be an increase in cytosolic calcium, accompanying exercise. Many mixed syndromes mutants, including ∆KPQ, E1784K, 1795insD and Q1909R, are found in calcium-sensitive regions. Elevated cytosolic calcium attenuates gain-of-function properties in ∆KPQ, 1795insD and Q1909R, but not in E1784K. By contrast, elevated cytosolic calcium further exacerbates gain-of-function in E1784K by destabilizing slow inactivation. Action potential modelling, using a modified O'Hara Rudy model, suggests that elevated heart rate rescues action potential duration in ∆KPQ, 1795insD and Q1909R, but not in E1784K. Action potential simulations suggest that E1784K carriers have an increased intracellular sodium-to-calcium ratio under bradycardia and tachycardia conditions. Elevated cytosolic calcium, which is common during high heart rates, ameliorates or exacerbates the mixed syndrome phenotype depending on the genetic signature. Inherited arrhythmias may arise from mutations in the gene for SCN5a, which encodes the cardiac voltage-gated sodium channel, Na V 1.5. Mutants in Na V 1.5 result in Brugada Syndrome (BrS1), Long-QT Syndrome (LQT3) or mixed syndromes (an overlap of BrS1/LQT3). Exercise is a potential arrhythmogenic trigger in mixed syndromes. We aimed to determine the effects of elevated cytosolic calcium, which is common during exercise, in mixed syndrome Na V 1.5 mutants. We used whole-cell patch clamp to assess the biophysical properties of Na V 1.5 wild-type (WT), ∆KPQ, E1784K, 1795insD and Q1909R mutants in human embryonic kidney 293 cells transiently transfected with the Na V 1.5 α subunit (WT or mutants), β1 subunit and enhanced green fluorescent protein. Voltage-dependence and kinetics were measured at cytosolic calcium levels of approximately 0, 500 and 2500

  2. Evaluation Effects of Verapamil as a Calcium Channel Blocker on Acquisition, Consolidation and Retrieval of Memory in Mice

    Directory of Open Access Journals (Sweden)

    Nooshin Masoudian

    2015-04-01

    Full Text Available Many factors are involved in learning and memory processes including brain nuclei, neurotransmitter systems, and the activity of ion channels. Studies showed inconsistent effects of calcium channel blockers on learning process, especially memory consolidation; however, little is known about their effect on memory acquisition and retrieval. Accordingly, the present study aimed to determine the effects of verapamil calcium channel antagonist as a representative of the phenylalkylamine group on different stages of memory and learning processes including acquisition, consolidation and retrieval in mice. In this experimental study, 150 male albino mice with a mean weight of 30 g were used. The mice were trained in a passive avoidance-learning task (1 mA shock for 2 seconds for evaluation of memory acquisition and consolidation and 3 seconds for evaluation of memory retrieval. The effect of verapamil (1, 2.5, 5, 10, and 20 mg/kg on memory consolidation and the most effective dose of consolidation phase on memory acquisition and retrieval was assessed. For the evaluation of memory consolidation, the animals received the drug intraperitoneally immediately after training, while for evaluation of memory acquisition and retrieval, the drug was injected one hour before training. Memory retrieval test was performed 48 hours after training (the length of time it took the animal to enter the dark part of the device. The results showed that verapamil injection exerted no effect on memory acquisition and consolidation; nevertheless, it was capable to disrupt memory retrieval in 10 and 20 mg doses. These results indicate that as a phenylalkylamine calcium channel antagonist, high doses of verapamil can impair memory. Normal 0 false false false EN-US X-NONE AR-SA /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso

  3. Voltage-gated calcium channels of Paramecium cilia.

    Science.gov (United States)

    Lodh, Sukanya; Yano, Junji; Valentine, Megan S; Van Houten, Judith L

    2016-10-01

    Paramecium cells swim by beating their cilia, and make turns by transiently reversing their power stroke. Reversal is caused by Ca 2+ entering the cilium through voltage-gated Ca 2+ (Ca V ) channels that are found exclusively in the cilia. As ciliary Ca 2+ levels return to normal, the cell pivots and swims forward in a new direction. Thus, the activation of the Ca V channels causes cells to make a turn in their swimming paths. For 45 years, the physiological characteristics of the Paramecium ciliary Ca V channels have been known, but the proteins were not identified until recently, when the P. tetraurelia ciliary membrane proteome was determined. Three Ca V α1 subunits that were identified among the proteins were cloned and confirmed to be expressed in the cilia. We demonstrate using RNA interference that these channels function as the ciliary Ca V channels that are responsible for the reversal of ciliary beating. Furthermore, we show that Pawn (pw) mutants of Paramecium that cannot swim backward for lack of Ca V channel activity do not express any of the three Ca V 1 channels in their ciliary membrane, until they are rescued from the mutant phenotype by expression of the wild-type PW gene. These results reinforce the correlation of the three Ca V channels with backward swimming through ciliary reversal. The PwB protein, found in endoplasmic reticulum fractions, co-immunoprecipitates with the Ca V 1c channel and perhaps functions in trafficking. The PwA protein does not appear to have an interaction with the channel proteins but affects their appearance in the cilia. © 2016. Published by The Company of Biologists Ltd.

  4. Diffusive spatio-temporal noise in a first-passage time model for intracellular calcium release

    KAUST Repository

    Flegg, Mark B.

    2013-01-01

    The intracellular release of calcium from the endoplasmic reticulum is controlled by ion channels. The resulting calcium signals exhibit a rich spatio-temporal signature, which originates at least partly from microscopic fluctuations. While stochasticity in the gating transition of ion channels has been incorporated into many models, the distribution of calcium is usually described by deterministic reaction-diffusion equations. Here we test the validity of the latter modeling approach by using two different models to calculate the frequency of localized calcium signals (calcium puffs) from clustered IP3 receptor channels. The complexity of the full calcium system is here limited to the basic opening mechanism of the ion channels and, in the mathematical reduction simplifies to the calculation of a first passage time. Two models are then studied: (i) a hybrid model, where channel gating is treated stochastically, while calcium concentration is deterministic and (ii) a fully stochastic model with noisy channel gating and Brownian calcium ion motion. The second model utilises the recently developed two-regime method [M. B. Flegg, S. J. Chapman, and R. Erban, "The two-regime method for optimizing stochastic reaction-diffusion simulations," J. R. Soc., Interface 9, 859-868 (2012)] in order to simulate a large domain with precision required only near the Ca2+ absorbing channels. The expected time for a first channel opening that results in a calcium puff event is calculated. It is found that for a large diffusion constant, predictions of the interpuff time are significantly overestimated using the model (i) with a deterministic non-spatial calcium variable. It is thus demonstrated that the presence of diffusive noise in local concentrations of intracellular Ca2+ ions can substantially influence the occurrence of calcium signals. The presented approach and results may also be relevant for other cell-physiological first-passage time problems with small ligand concentration

  5. Noradrenergic mechanisms and high blood pressure maintenance in genetic hypertension: The role of Gi proteins and voltage-dependent calcium channels

    Czech Academy of Sciences Publication Activity Database

    Zicha, Josef; Pintérová, Mária; Líšková, Silvia; Dobešová, Zdenka; Kuneš, Jaroslav

    2007-01-01

    Roč. 29, č. 4 (2007), s. 229-229 ISSN 1064-1963. [International symposium on SHR /12./. 20.10.2006-21.10.2006, Kyoto] R&D Projects: GA MZd(CZ) NR7786 Institutional research plan: CEZ:AV0Z50110509 Keywords : genetic hypertension * noradrenergic mechanisms * Gi proteins * voltage-dependent calcium channels Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery

  6. Effects of calcium antagonists on isolated bovine cerebral arteries: inhibition of constriction and calcium-45 uptake induced by potassium or serotonin

    International Nuclear Information System (INIS)

    Wendling, W.W.; Harakal, C.

    1987-01-01

    The purpose of this study was to determine the mechanisms by which organic calcium channel blockers inhibit cerebral vasoconstriction. Isolated bovine middle cerebral arteries were cut into rings to measure contractility or into strips to measure radioactive calcium ( 45 Ca) influx and efflux. Calcium channel blockers (10(-5) M verapamil or 3.3 X 10(-7) M nifedipine) and calcium-deficient solutions all produced near-maximal inhibition of both potassium- and serotonin-induced constriction. In calcium-deficient solutions containing potassium or serotonin, verapamil and nifedipine each blocked subsequent calcium-induced constriction in a competitive manner. Potassium and serotonin significantly increased 45 Ca uptake into cerebral artery strips during 5 minutes of 45 Ca loading; for potassium 45 Ca uptake increased from 62 to 188 nmol/g, and for serotonin from 65 to 102 nmol/g. Verapamil or nifedipine had no effect on basal 45 Ca uptake but significantly blocked the increase in 45 Ca uptake induced by potassium or serotonin. Potassium, and to a lesser extent serotonin, each induced a brief increase in the rate of 45 Ca efflux into calcium-deficient solutions. Verapamil or nifedipine had no effect on basal or potassium-stimulated 45 Ca efflux. The results demonstrate that verapamil and nifedipine block 45 Ca uptake through both potential-operated (potassium) and receptor-operated (serotonin) channels in bovine middle cerebral arteries

  7. Congenital heart block maternal sera autoantibodies target an extracellular epitope on the α1G T-type calcium channel in human fetal hearts.

    Directory of Open Access Journals (Sweden)

    Linn S Strandberg

    Full Text Available Congenital heart block (CHB is a transplacentally acquired autoimmune disease associated with anti-Ro/SSA and anti-La/SSB maternal autoantibodies and is characterized primarily by atrioventricular (AV block of the fetal heart. This study aims to investigate whether the T-type calcium channel subunit α1G may be a fetal target of maternal sera autoantibodies in CHB.We demonstrate differential mRNA expression of the T-type calcium channel CACNA1G (α1G gene in the AV junction of human fetal hearts compared to the apex (18-22.6 weeks gestation. Using human fetal hearts (20-22 wks gestation, our immunoprecipitation (IP, Western blot analysis and immunofluorescence (IF staining results, taken together, demonstrate accessibility of the α1G epitope on the surfaces of cardiomyocytes as well as reactivity of maternal serum from CHB affected pregnancies to the α1G protein. By ELISA we demonstrated maternal sera reactivity to α1G was significantly higher in CHB maternal sera compared to controls, and reactivity was epitope mapped to a peptide designated as p305 (corresponding to aa305-319 of the extracellular loop linking transmembrane segments S5-S6 in α1G repeat I. Maternal sera from CHB affected pregnancies also reacted more weakly to the homologous region (7/15 amino acids conserved of the α1H channel. Electrophysiology experiments with single-cell patch-clamp also demonstrated effects of CHB maternal sera on T-type current in mouse sinoatrial node (SAN cells.Taken together, these results indicate that CHB maternal sera antibodies readily target an extracellular epitope of α1G T-type calcium channels in human fetal cardiomyocytes. CHB maternal sera also show reactivity for α1H suggesting that autoantibodies can target multiple fetal targets.

  8. Simultaneous allergen inactivation and detoxification of castor bean cake by treatment with calcium compounds

    International Nuclear Information System (INIS)

    Fernandes, K.V.; Deus-de-Oliveira, N.; Godoy, M.G.; Guimarães, Z.A.S.; Nascimento, V.V.; Melo, E.J.T. de; Freire, D.M.G.; Dansa-Petretski, M.; Machado, O.L.T.

    2012-01-01

    Ricinus communis L. is of great economic importance due to the oil extracted from its seeds. Castor oil has been used for pharmaceutical and industrial applications, as a lubricant or coating agent, as a component of plastic products, as a fungicide or in the synthesis of biodiesel fuels. After oil extraction, a castor cake with a large amount of protein is obtained. However, this by-product cannot be used as animal feed due to the presence of toxic (ricin) and allergenic (2S albumin) proteins. Here, we propose two processes for detoxification and allergen inactivation of the castor cake. In addition, we establish a biological test to detect ricin and validate these detoxification processes. In this test, Vero cells were treated with ricin, and cell death was assessed by cell counting and measurement of lactate dehydrogenase activity. The limit of detection of the Vero cell assay was 10 ng/mL using a concentration of 1.6 × 10 5 cells/well. Solid-state fermentation (SSF) and treatment with calcium compounds were used as cake detoxification processes. For SSF, Aspergillus niger was grown using a castor cake as a substrate, and this cake was analyzed after 24, 48, 72, and 96 h of SSF. Ricin was eliminated after 24 h of SSF treatment. The cake was treated with 4 or 8% Ca(OH) 2 or CaO, and both the toxicity and the allergenic properties were entirely abolished. A by-product free of toxicity and allergens was obtained

  9. Simultaneous allergen inactivation and detoxification of castor bean cake by treatment with calcium compounds

    Directory of Open Access Journals (Sweden)

    K.V. Fernandes

    2012-11-01

    Full Text Available Ricinus communis L. is of great economic importance due to the oil extracted from its seeds. Castor oil has been used for pharmaceutical and industrial applications, as a lubricant or coating agent, as a component of plastic products, as a fungicide or in the synthesis of biodiesel fuels. After oil extraction, a castor cake with a large amount of protein is obtained. However, this by-product cannot be used as animal feed due to the presence of toxic (ricin and allergenic (2S albumin proteins. Here, we propose two processes for detoxification and allergen inactivation of the castor cake. In addition, we establish a biological test to detect ricin and validate these detoxification processes. In this test, Vero cells were treated with ricin, and cell death was assessed by cell counting and measurement of lactate dehydrogenase activity. The limit of detection of the Vero cell assay was 10 ng/mL using a concentration of 1.6 x 10(5 cells/well. Solid-state fermentation (SSF and treatment with calcium compounds were used as cake detoxification processes. For SSF, Aspergillus niger was grown using a castor cake as a substrate, and this cake was analyzed after 24, 48, 72, and 96 h of SSF. Ricin was eliminated after 24 h of SSF treatment. The cake was treated with 4 or 8% Ca(OH2 or CaO, and both the toxicity and the allergenic properties were entirely abolished. A by-product free of toxicity and allergens was obtained.

  10. Simultaneous allergen inactivation and detoxification of castor bean cake by treatment with calcium compounds

    Energy Technology Data Exchange (ETDEWEB)

    Fernandes, K.V.; Deus-de-Oliveira, N. [Laboratório de Química e Função de Proteínas e Peptídeos, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Rio de Janeiro, RJ (Brazil); Godoy, M.G. [Laboratório de Biotecnologia Microbiana, Instituto de Química, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Guimarães, Z.A.S. [Laboratório de Biologia Celular e Tecidual, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Rio de Janeiro, RJ (Brazil); Nascimento, V.V. [Laboratório de Química e Função de Proteínas e Peptídeos, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Rio de Janeiro, RJ (Brazil); Melo, E.J.T. de [Laboratório de Biologia Celular e Tecidual, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Rio de Janeiro, RJ (Brazil); Freire, D.M.G. [Laboratório de Biotecnologia Microbiana, Instituto de Química, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Dansa-Petretski, M.; Machado, O.L.T. [Laboratório de Química e Função de Proteínas e Peptídeos, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense Darcy Ribeiro, Rio de Janeiro, RJ (Brazil)

    2012-08-24

    Ricinus communis L. is of great economic importance due to the oil extracted from its seeds. Castor oil has been used for pharmaceutical and industrial applications, as a lubricant or coating agent, as a component of plastic products, as a fungicide or in the synthesis of biodiesel fuels. After oil extraction, a castor cake with a large amount of protein is obtained. However, this by-product cannot be used as animal feed due to the presence of toxic (ricin) and allergenic (2S albumin) proteins. Here, we propose two processes for detoxification and allergen inactivation of the castor cake. In addition, we establish a biological test to detect ricin and validate these detoxification processes. In this test, Vero cells were treated with ricin, and cell death was assessed by cell counting and measurement of lactate dehydrogenase activity. The limit of detection of the Vero cell assay was 10 ng/mL using a concentration of 1.6 × 10{sup 5} cells/well. Solid-state fermentation (SSF) and treatment with calcium compounds were used as cake detoxification processes. For SSF, Aspergillus niger was grown using a castor cake as a substrate, and this cake was analyzed after 24, 48, 72, and 96 h of SSF. Ricin was eliminated after 24 h of SSF treatment. The cake was treated with 4 or 8% Ca(OH){sub 2} or CaO, and both the toxicity and the allergenic properties were entirely abolished. A by-product free of toxicity and allergens was obtained.

  11. Turtle Flexion Reflex Motor Patterns Show Windup, Mediated Partly by L-type Calcium Channels

    Directory of Open Access Journals (Sweden)

    Keith P. Johnson

    2017-10-01

    Full Text Available Windup is a form of multisecond temporal summation in which identical stimuli, delivered seconds apart, trigger increasingly strong neuronal responses. L-type Ca2+ channels have been shown to play an important role in the production of windup of spinal cord neuronal responses, initially in studies of turtle spinal cord and later in studies of mammalian spinal cord. L-type Ca2+ channels have also been shown to contribute to windup of limb withdrawal reflex (flexion reflex in rats, but flexion reflex windup has not previously been described in turtles and its cellular mechanisms have not been studied. We studied windup of flexion reflex motor patterns, evoked with weak mechanical and electrical stimulation of the dorsal hindlimb foot skin and assessed via a hip flexor (HF nerve recording, in spinal cord-transected and immobilized turtles in vivo. We found that an L-type Ca2+ channel antagonist, nifedipine, applied at concentrations of 50 μM or 100 μM to the hindlimb enlargement spinal cord, significantly reduced windup of flexion reflex motor patterns, while lower concentrations of nifedipine had no such effect. Nifedipine similarly reduced the amplitude of an individual flexion reflex motor pattern evoked by a stronger mechanical stimulus, in a dose-dependent manner, suggesting that L-type Ca2+ channels contribute to each flexion reflex as well as to multisecond summation of flexion reflex responses in turtles. We also found that we could elicit flexion reflex windup consistently using a 4-g von Frey filament, which is not usually considered a nociceptive stimulus. Thus, it may be that windup can be evoked by a wide range of tactile stimuli and that L-type calcium channels contribute to multisecond temporal summation of diverse tactile stimuli across vertebrates.

  12. Gynura procumbens Merr. decreases blood pressure in rats by vasodilatation via inhibition of calcium channels

    Directory of Open Access Journals (Sweden)

    See-Ziau Hoe

    2011-01-01

    Full Text Available INTRODUCTION: Gynura procumbens has been shown to decrease blood pressure via inhibition of the angiotensinconverting enzyme. However, other mechanisms that may contribute to the hypotensive effect have not been studied. OBJECTIVES: To investigate the cardiovascular effects of a butanolic fraction of Gynura procumbens in rats. METHODS: Anaesthetized rats were given intravenous bolus injections of butanolic fraction at doses of 2.5-20 mg/kg in vivo. The effect of butanolic fraction on vascular reactivity was recorded in isolated rat aortic rings in vitro. RESULTS: Intravenous administrations of butanolic fraction elicited significant (p<0.001 and dose-dependent decreases in the mean arterial pressure. However, a significant (p<0.05 decrease in the heart rate was observed only at the higher doses (10 and 20 mg/kg. In isolated preparations of rat aortic rings, phenylephrine (1×10-6 M- or potassium chloride (8×10-2 M-precontracted endothelium-intact and -denuded tissue; butanolic fraction (1×10-6-1×10-1 g/ml induced similar concentration-dependent relaxation of the vessels. In the presence of 2.5×10-3 and 5.0×10-3 g/ml butanolic fraction, the contractions induced by phenylephrine (1×10-9-3×10-5 M and potassium chloride (1×10-2-8×10-2 M were significantly antagonized. The calcium-induced vasocontractions (1×10-4-1×10-2 M were antagonized by butanolic fraction concentration-dependently in calcium-free and high potassium (6×10-2 M medium, as well as in calcium- and potassium-free medium containing 1×10-6 M phenylephrine. However, the contractions induced by noradrenaline (1×10-6 M and caffeine (4.5×10-2 M were not affected by butanolic fraction. CONCLUSION: Butanolic fraction contains putative hypotensive compounds that appear to inhibit calcium influx via receptor-operated and/or voltage-dependent calcium channels to cause vasodilation and a consequent fall in blood pressure.

  13. Bimodal voltage dependence of TRPA1: mutations of a key pore helix residue reveal strong intrinsic voltage-dependent inactivation.

    Science.gov (United States)

    Wan, Xia; Lu, Yungang; Chen, Xueqin; Xiong, Jian; Zhou, Yuanda; Li, Ping; Xia, Bingqing; Li, Min; Zhu, Michael X; Gao, Zhaobing

    2014-07-01

    Transient receptor potential A1 (TRPA1) is implicated in somatosensory processing and pathological pain sensation. Although not strictly voltage-gated, ionic currents of TRPA1 typically rectify outwardly, indicating channel activation at depolarized membrane potentials. However, some reports also showed TRPA1 inactivation at high positive potentials, implicating voltage-dependent inactivation. Here we report a conserved leucine residue, L906, in the putative pore helix, which strongly impacts the voltage dependency of TRPA1. Mutation of the leucine to cysteine (L906C) converted the channel from outward to inward rectification independent of divalent cations and irrespective to stimulation by allyl isothiocyanate. The mutant, but not the wild-type channel, displayed exclusively voltage-dependent inactivation at positive potentials. The L906C mutation also exhibited reduced sensitivity to inhibition by TRPA1 blockers, HC030031 and ruthenium red. Further mutagenesis of the leucine to all natural amino acids individually revealed that most substitutions at L906 (15/19) resulted in inward rectification, with exceptions of three amino acids that dramatically reduced channel activity and one, methionine, which mimicked the wild-type channel. Our data are plausibly explained by a bimodal gating model involving both voltage-dependent activation and inactivation of TRPA1. We propose that the key pore helix residue, L906, plays an essential role in responding to the voltage-dependent gating.

  14. EFFECTS OF PYRETHROIDS ON VOLTAGE-SENSITIVE CALCIUM CHANNELS: A CRITICAL EVALUATION OF STRENGTHS, WEAKNESSES, DATA NEEDS, AND RELATIONSHIP TO ASSESSMENT OF CUMULATIVE NEUROTOXICITY.

    Science.gov (United States)

    A recently published review (Soderlund et al., 2002, Toxicology 171, 3-59.) of the mechanisms of acute neurotoxicity of pyrethroid compounds postulated that voltage-sensitive calcium channels (VSCC) may be a target of some pyrethroid compounds and that effects on VSCC may contrib...

  15. Parathyroid Hormone Induces Bone Cell Motility and Loss of Mature Osteocyte Phenotype through L-Calcium Channel Dependent and Independent Mechanisms.

    Directory of Open Access Journals (Sweden)

    Matthew Prideaux

    Full Text Available Parathyroid Hormone (PTH can exert both anabolic and catabolic effects on the skeleton, potentially through expression of the PTH type1 receptor (PTH1R, which is highly expressed in osteocytes. To determine the cellular and molecular mechanisms responsible, we examined the effects of PTH on osteoblast to osteocyte differentiation using primary osteocytes and the IDG-SW3 murine cell line, which differentiate from osteoblast to osteocyte-like cells in vitro and express GFP under control of the dentin matrix 1 (Dmp1 promoter. PTH treatment resulted in an increase in some osteoblast and early osteocyte markers and a decrease in mature osteocyte marker expression. The gene expression profile of PTH-treated Day 28 IDG-SW3 cells was similar to PTH treated primary osteocytes. PTH treatment induced striking changes in the morphology of the Dmp1-GFP positive cells in IDG-SW3 cultures and primary cells from Dmp1-GFP transgenic mice. The cells changed from a more dendritic to an elongated morphology and showed increased cell motility. E11/gp38 has been shown to be important for cell migration, however, deletion of the E11/gp38/podoplanin gene had no effect on PTH-induced motility. The effects of PTH on motility were reproduced using cAMP, but not with protein kinase A (PKA, exchange proteins activated by cAMP (Epac, protein kinase C (PKC or phosphatidylinositol-4,5-bisphosphonate 3-kinase (Pi3K agonists nor were they blocked by their antagonists. However, the effects of PTH were mediated through calcium signaling, specifically through L-type channels normally expressed in osteoblasts but decreased in osteocytes. PTH was shown to increase expression of this channel, but decrease the T-type channel that is normally more highly expressed in osteocytes. Inhibition of L-type calcium channel activity attenuated the effects of PTH on cell morphology and motility but did not prevent the downregulation of mature osteocyte marker expression. Taken together, these

  16. Major Channels Involved In Neuropsychiatric Disorders And Therapeutic Perspectives

    Directory of Open Access Journals (Sweden)

    Paola eImbrici

    2013-05-01

    Full Text Available Voltage-gated ion channels are important mediators of physiological functions in the central nervous system. The cyclic activation of these channels influences neurotransmitter release, neuron excitability, gene transcription and plasticity, providing distinct brain areas with unique physiological and pharmacological response. A growing body of data has implicated ion channels in the susceptibility or pathogenesis of psychiatric diseases. Indeed, population studies support the association of polymorphisms in calcium and potassium channels with the genetic risk for bipolar disorders or schizophrenia. Moreover, point mutations in calcium, sodium and potassium channel genes have been identified in some childhood developmental disorders. Finally, antibodies against potassium channel complexes occur in a series of autoimmune psychiatric diseases. Here we report recent studies assessing the role of calcium, sodium and potassium channels in bipolar disorder, schizophrenia and autism spectrum disorders, and briefly summarize promising pharmacological strategies targeted on ion channels for the therapy of mental illness and related genetic tests.

  17. Antiapolipoprotein A-1 IgG chronotropic effects require nongenomic action of aldosterone on L-type calcium channels.

    Science.gov (United States)

    Rossier, Michel F; Pagano, Sabrina; Python, Magaly; Maturana, Andres D; James, Richard W; Mach, François; Roux-Lombard, Pascale; Vuilleumier, Nicolas

    2012-03-01

    Autoantibodies to apolipoprotein A-1 (antiapoA-1 IgG) have been shown to be associated with higher resting heart rate and morbidity in myocardial infarction patients and to behave as a chronotropic agent in the presence of aldosterone on isolated neonatal rat ventricular cardiomyocytes (NRVC). We aimed at identifying the pathways accounting for this aldosterone-dependent antiapoA-1 IgG-positive chronotropic effect on NRVC. The rate of regular spontaneous contractions was determined on NRVC in the presence of different steroid hormones and antagonists. AntiapoA-1 IgG chronotropic response was maximal within 20 min and observed only in aldosterone-pretreated cells but not in those exposed to other steroids. The positive antiapoA-1 IgG chronotropic effect was already significant after 5 min aldosterone preincubation, was dependent on 3-kinase and protein kinase A activities, was not inhibited by actinomycin D, and was fully abrogated by eplerenone (but not by spironolactone), demonstrating the dependence on a nongenomic action of aldosterone elicited through the mineralocorticoid receptor (MR). Under oxidative conditions (but not under normal redox state), corticosterone mimicked the permissive action of aldosterone on the antiapoA-1 IgG chronotropic response. Pharmacological and patch-clamp studies identified L-type calcium channels as crucial effectors of antiapoA-1 IgG chronotropic action, involving two converging pathways that increase the channel activity. The first one involves the rapid, nongenomic activation of the phosphatidylinositol 3-kinase enzyme by MR, and the second one requires a constitutive basal protein kinase A activity. In conclusion, our results indicate that, on NRVC, the aldosterone-dependent chronotropic effects of antiapoA-1 IgG involve the nongenomic activation of L-type calcium channels.

  18. Dual Regulation of Voltage-Sensitive Ion Channels by PIP2

    Directory of Open Access Journals (Sweden)

    Aldo A Rodríguez Menchaca

    2012-09-01

    Full Text Available Over the past 16 years, there has been an impressive number of ion channels shown to be sensitive to the major phosphoinositide in the plasma membrane, phosphatidilinositol 4,5-bisphosphate (PIP2. Among them are voltage-gated channels, which are crucial for both neuronal and cardiac excitability. Voltage-gated calcium (Cav channels were shown to be regulated bidirectionally by PIP2. On one hand, PIP2 stabilized their activity by reducing current rundown but on the other hand it produced a voltage-dependent inhibition by shifting the activation curve to more positive voltages. For voltage-gated potassium (Kv channels PIP2 was first shown to prevent N-type inactivation. Careful examination of the effects of PIP2 on the activation mechanism of Kv1.2 has shown a similar bidirectional regulation as in the Cav channels. The two effects could be distinguished kinetically, in terms of their sensitivities to PIP2 and by distinct molecular determinants. The rightward shift of the Kv1.2 voltage dependence implicated basic residues in the S4-S5 linker and was consistent with stabilization of the inactive state of the voltage sensor. A third type of a voltage-gated ion channel modulated by PIP2 is the hyperpolarization-activated cyclic nucleotide-gated (HCN channel. PIP2 has been shown to enhance the opening of HCN channels by shifting their voltage-dependent activation toward depolarized potentials. The sea urchin HCN channel, SpIH, showed again a PIP2-mediated bidirectional effect but in reverse order than the depolarization-activated Cav and Kv channels: a voltage-dependent potentiation, like the mammalian HCN channels, but also an inhibition of the cGMP-induced current activation. Just like the Kv1.2 channels, distinct molecular determinants underlied the PIP2 dual effects on SpIH channels. The dual regulation of these very different ion channels, all of which are voltage dependent, points to conserved mechanisms of regulation of these channels by PIP2.

  19. The metabolic impact of β-hydroxybutyrate on neurotransmission: Reduced glycolysis mediates changes in calcium responses and KATP channel receptor sensitivity.

    Science.gov (United States)

    Lund, Trine M; Ploug, Kenneth B; Iversen, Anne; Jensen, Anders A; Jansen-Olesen, Inger

    2015-03-01

    Glucose is the main energy substrate for neurons, and ketone bodies are known to be alternative substrates. However, the capacity of ketone bodies to support different neuronal functions is still unknown. Thus, a change in energy substrate from glucose alone to a combination of glucose and β-hydroxybutyrate might change neuronal function as there is a known coupling between metabolism and neurotransmission. The purpose of this study was to shed light on the effects of the ketone body β-hydroxybutyrate on glycolysis and neurotransmission in cultured murine glutamatergic neurons. Previous studies have shown an effect of β-hydroxybutyrate on glucose metabolism, and the present study further specified this by showing attenuation of glycolysis when β-hydroxybutyrate was present in these neurons. In addition, the NMDA receptor-induced calcium responses in the neurons were diminished in the presence of β-hydroxybutyrate, whereas a direct effect of the ketone body on transmitter release was absent. However, the presence of β-hydroxybutyrate augmented transmitter release induced by the KATP channel blocker glibenclamide, thus giving an indirect indication of the involvement of KATP channels in the effects of ketone bodies on transmitter release. Energy metabolism and neurotransmission are linked and involve ATP-sensitive potassium (KATP ) channels. However, it is still unclear how and to what degree available energy substrate affects this link. We investigated the effect of changing energy substrate from only glucose to a combination of glucose and R-β-hydroxybutyrate in cultured neurons. Using the latter combination, glycolysis was diminished, NMDA receptor-induced calcium responses were lower, and the KATP channel blocker glibenclamide caused a higher transmitter release. © 2014 International Society for Neurochemistry.

  20. Calcium-Induced calcium release during action potential firing in developing inner hair cells.

    Science.gov (United States)

    Iosub, Radu; Avitabile, Daniele; Grant, Lisa; Tsaneva-Atanasova, Krasimira; Kennedy, Helen J

    2015-03-10

    In the mature auditory system, inner hair cells (IHCs) convert sound-induced vibrations into electrical signals that are relayed to the central nervous system via auditory afferents. Before the cochlea can respond to normal sound levels, developing IHCs fire calcium-based action potentials that disappear close to the onset of hearing. Action potential firing triggers transmitter release from the immature IHC that in turn generates experience-independent firing in auditory neurons. These early signaling events are thought to be essential for the organization and development of the auditory system and hair cells. A critical component of the action potential is the rise in intracellular calcium that activates both small conductance potassium channels essential during membrane repolarization, and triggers transmitter release from the cell. Whether this calcium signal is generated by calcium influx or requires calcium-induced calcium release (CICR) is not yet known. IHCs can generate CICR, but to date its physiological role has remained unclear. Here, we used high and low concentrations of ryanodine to block or enhance CICR to determine whether calcium release from intracellular stores affected action potential waveform, interspike interval, or changes in membrane capacitance during development of mouse IHCs. Blocking CICR resulted in mixed action potential waveforms with both brief and prolonged oscillations in membrane potential and intracellular calcium. This mixed behavior is captured well by our mathematical model of IHC electrical activity. We perform two-parameter bifurcation analysis of the model that predicts the dependence of IHCs firing patterns on the level of activation of two parameters, the SK2 channels activation and CICR rate. Our data show that CICR forms an important component of the calcium signal that shapes action potentials and regulates firing patterns, but is not involved directly in triggering exocytosis. These data provide important insights

  1. History-dependent dynamics in a generic model of ion channels - an analytic study

    Directory of Open Access Journals (Sweden)

    Daniel Soudry

    2010-04-01

    Full Text Available Recent experiments have demonstrated that the timescale of adaptation of single neurons and ion channel populations to stimuli slows down as the length of stimulation increases; in fact, no upper bound on temporal time-scales seems to exist in such systems. Furthermore, patch clamp experiments on single ion channels have hinted at the existence of large, mostly unobservable, inactivation state spaces within a single ion channel. This raises the question of the relation between this multitude of inactivation states and the observed behavior. In this work we propose a minimal model for ion channel dynamics which does not assume any specific structure of the inactivation state space. The model is simple enough to render an analytical study possible. This leads to a clear and concise explanation of the experimentally observed exponential history-dependent relaxation in sodium channels in a voltage clamp setting, and shows that their recovery rate from slow inactivation must be voltage dependent. Furthermore, we predict that history-dependent relaxation cannot be created by overly sparse spiking activity. While the model was created with ion channel populations in mind, its simplicity and genericalness render it a good starting point for modeling similar effects in other systems, and for scaling up to higher levels such as single neurons which are also known to exhibit multiple time scales.

  2. Lipid Rescue Therapy and High-Dose insulin Euglycemic Therapy are Effective for Severe Refractory Calcium Channel Blocker Overdose: Case Report and Review of Literature

    Directory of Open Access Journals (Sweden)

    Niko Bekjarovski

    2013-09-01

    How to cite this article: Bekjarovski NG. Lipid Rescue Therapy and High-Dose insulin Euglycemic Therapy are Effective for Severe Refractory Calcium Channel Blocker Overdose: Case Report and Review of Literature. Asia Pac J Med Toxicol 2013;2:114-6.

  3. Propylparaben reduces the excitability of hippocampal neurons by blocking sodium channels.

    Science.gov (United States)

    Lara-Valderrábano, Leonardo; Rocha, Luisa; Galván, Emilio J

    2016-12-01

    Propylparaben (PPB) is an antimicrobial preservative widely used in food, cosmetics, and pharmaceutics. Virtual screening methodologies predicted anticonvulsant activity of PPB that was confirmed in vivo. Thus, we explored the effects of PPB on the excitability of hippocampal neurons by using standard patch clamp techniques. Bath perfusion of PPB reduced the fast-inactivating sodium current (I Na ) amplitude, causing a hyperpolarizing shift in the inactivation curve of the I Na, and markedly delayed the sodium channel recovery from the inactivation state. Also, PPB effectively suppressed the riluzole-sensitive, persistent sodium current (I NaP ). PPB perfusion also modified the action potential kinetics, and higher concentrations of PPB suppressed the spike activity. Nevertheless, the modulatory effects of PPB did not occur when PPB was internally applied by whole-cell dialysis. These results indicate that PPB reduces the excitability of CA1 pyramidal neurons by modulating voltage-dependent sodium channels. The mechanistic basis of this effect is a marked delay in the recovery from inactivation state of the voltage-sensitive sodium channels. Our results indicate that similar to local anesthetics and anticonvulsant drugs that act on sodium channels, PPB acts in a use-dependent manner. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. DIHYDROPYRIDINE CALCIUM- CHANNELBLOCKERSFOR ...

    African Journals Online (AJOL)

    Furthermore, the controversy over the role of calci~-channel blockers as first-line ..... group trials while fully accounting for placebo effects as well as interindividual ..... Reducing calcium overload in the ischemic brain. N Engl JMed. 1999; 341 ...

  5. Effects of the L/N-type calcium channel antagonist cilnidipine on morning blood pressure control and peripheral edema formation.

    Science.gov (United States)

    Narita, Sumito; Yoshioka, Yasuko; Ide, Atsumi; Kadokami, Toshiaki; Momii, Hidetoshi; Yoshida, Masayoshi; Ando, Shin-ichi

    2011-01-01

    The L/N-type calcium channel blocker cilnidipine has unique effects including sympathetic nerve suppression and the balanced vasodilatation of arteries and veins that may alleviate morning hypertension (MHT) or peripheral edema caused by calcium channel antagonists. We used ambulatory blood pressure monitoring (ABPM) and a unique peripheral edema measurement to evaluate the effect of morning and bedtime cilnidipine in patients with MHT. Forty-three patients with MHT (60 ± 12 years) were randomly assigned to a morning or bedtime cilnidipine (10-20 mg/day). MHT was defined as a mean systolic blood pressure (SBP) ≥ 135 mm Hg by ABPM within 2 hours after awaking. After 3 months, greater SBP reductions were observed in the bedtime administration group (versus the morning administration group) at 3:30-6:00 AM (-24 ± 20 mm Hg vs. -10 ± 4 mm Hg; P < .05) and at 6:30-9:00 AM (-26 ± 15 mm Hg vs. -14 ± 17 mm Hg; P < .05). Although physical examinations showed leg edema in 16% of the patients, quantitative evaluations did not reveal significant volume gains. Cilnidipine had a greater effect on MHT, without causing significant leg edema, when administered at bedtime. Copyright © 2011 American Society of Hypertension. Published by Elsevier Inc. All rights reserved.

  6. Molecular and kinetic determinants of local anaesthetic action on sodium channels.

    Science.gov (United States)

    French, R J; Zamponi, G W; Sierralta, I E

    1998-11-23

    (1) Local anaesthetics (LA) rely for their clinical actions on state-dependent inhibition of voltage-dependent sodium channels. (2) Single, batrachoxin-modified sodium channels in planar lipid bilayers allow direct observation of drug-channel interactions. Two modes of inhibition of single-channel current are observed: fast block of the open channels and prolongation of a long-lived closed state, some of whose properties resemble those of the inactivated state of unmodified channels. (3) Analogues of different parts of the LA molecule separately mimic each blocking mode: amines--fast block, and water-soluble aromatics--closed state prolongation. (4) Interaction between a mu-conotoxin derivative and diethylammonium indicate an intrapore site of fast, open-state block. (5) Site-directed mutagenesis studies suggest that hydrophobic residues in transmembrane segment 6 of repeat domain 4 of sodium channels are critical for both LA binding and stabilization of the inactivated state.

  7. Calcium Signaling in Taste Cells

    Science.gov (United States)

    Medler, Kathryn F.

    2014-01-01

    The sense of taste is a common ability shared by all organisms and is used to detect nutrients as well as potentially harmful compounds. Thus taste is critical to survival. Despite its importance, surprisingly little is known about the mechanisms generating and regulating responses to taste stimuli. All taste responses depend on calcium signals to generate appropriate responses which are relayed to the brain. Some taste cells have conventional synapses and rely on calcium influx through voltage-gated calcium channels. Other taste cells lack these synapses and depend on calcium release to formulate an output signal through a hemichannel. Beyond establishing these characteristics, few studies have focused on understanding how these calcium signals are formed. We identified multiple calcium clearance mechanisms that regulate calcium levels in taste cells as well as a calcium influx that contributes to maintaining appropriate calcium homeostasis in these cells. Multiple factors regulate the evoked taste signals with varying roles in different cell populations. Clearly, calcium signaling is a dynamic process in taste cells and is more complex than has previously been appreciated. PMID:25450977

  8. Acute Treatment with T-Type Calcium Channel Enhancer SAK3 Reduces Cognitive Impairments Caused by Methimazole-Induced Hypothyroidism Via Activation of Cholinergic Signaling.

    Science.gov (United States)

    Husain, Noreen; Yabuki, Yasushi; Shinoda, Yasuharu; Fukunaga, Kohji

    2018-01-01

    Hypothyroidism is a common disorder that is associated with psychological disturbances such as dementia, depression, and psychomotor disorders. We recently found that chronic treatment with the T-type calcium channel enhancer SAK3 prevents the cholinergic neurodegeneration induced by a single intraperitoneal (i.p.) injection of methimazole (MMI; 75 mg/kg), thereby improving cognition. Here, we evaluated the acute effect of SAK3 on cognitive impairments and its mechanism of action following the induction of hypothyroidism. Hypothyroidism was induced by 2 injections of MMI (75 mg/kg, i.p.) administered once per week. Four weeks after the final MMI treatment, MMI-treated mice showed reduced serum thyroxine (T4) levels and cognitive impairments without depression-like behaviors. Although acute SAK3 (1.0 mg/kg, p.o.) administration failed to ameliorate the decreased T4 levels and histochemical destruction of the glomerular structure, acute SAK3 (1.0 mg/kg, p.o.) administration significantly reduced cognitive impairments in MMI-treated mice. Importantly, the α7 nicotinic acetylcholine receptor (nAChR)-selective inhibitor methyllycaconitine (MLA; 12 mg/kg, i.p.) and T-type calcium channel-specific blocker NNC 55-0396 (25 mg/kg, i.p.) antagonized the acute effect of SAK3 on memory deficits in MMI-treated mice. We also confirmed that acute SAK3 administration does not rescue reduced olfactory marker protein or choline acetyltransferase immunoreactivity levels in the olfactory bulb or medial septum. Taken together, these results suggest that SAK3 has the ability to improve the cognitive decline caused by hypothyroidism directly through activation of nAChR signaling and T-type calcium channels. © 2018 S. Karger AG, Basel.

  9. Membrane potential and cation channels in rat juxtaglomerular cells

    DEFF Research Database (Denmark)

    Friis, U G; Jørgensen, F; Andreasen, D

    2004-01-01

    The relationship between membrane potential and cation channels in juxtaglomerular (JG) cells is not well understood. Here we review electrophysiological and molecular studies of JG cells demonstrating the presence of large voltage-sensitive, calcium-activated potassium channels (BK(Ca)) of the Z......The relationship between membrane potential and cation channels in juxtaglomerular (JG) cells is not well understood. Here we review electrophysiological and molecular studies of JG cells demonstrating the presence of large voltage-sensitive, calcium-activated potassium channels (BK...

  10. Calcium influx through L-type channels attenuates skeletal muscle contraction via inhibition of adenylyl cyclases.

    Science.gov (United States)

    Menezes-Rodrigues, Francisco Sandro; Pires-Oliveira, Marcelo; Duarte, Thiago; Paredes-Gamero, Edgar Julian; Chiavegatti, Tiago; Godinho, Rosely Oliveira

    2013-11-15

    Skeletal muscle contraction is triggered by acetylcholine induced release of Ca(2+) from sarcoplasmic reticulum. Although this signaling pathway is independent of extracellular Ca(2+), L-type voltage-gated calcium channel (Cav) blockers have inotropic effects on frog skeletal muscles which occur by an unknown mechanism. Taking into account that skeletal muscle fiber expresses Ca(+2)-sensitive adenylyl cyclase (AC) isoforms and that cAMP is able to increase skeletal muscle contraction force, we investigated the role of Ca(2+) influx on mouse skeletal muscle contraction and the putative crosstalk between extracellular Ca(2+) and intracellular cAMP signaling pathways. The effects of Cav blockers (verapamil and nifedipine) and extracellular Ca(2+) chelator EGTA were evaluated on isometric contractility of mouse diaphragm muscle under direct electrical stimulus (supramaximal voltage, 2 ms, 0.1 Hz). Production of cAMP was evaluated by radiometric assay while Ca(2+) transients were assessed by confocal microscopy using L6 cells loaded with fluo-4/AM. Ca(2+) channel blockers verapamil and nifedipine had positive inotropic effect, which was mimicked by removal of extracellular Ca(+2) with EGTA or Ca(2+)-free Tyrode. While phosphodiesterase inhibitor IBMX potentiates verapamil positive inotropic effect, it was abolished by AC inhibitors SQ22536 and NYK80. Finally, the inotropic effect of verapamil was associated with increased intracellular cAMP content and mobilization of intracellular Ca(2+), indicating that positive inotropic effects of Ca(2+) blockers depend on cAMP formation. Together, our results show that extracellular Ca(2+) modulates skeletal muscle contraction, through inhibition of Ca(2+)-sensitive AC. The cross-talk between extracellular calcium and cAMP-dependent signaling pathways appears to regulate the extent of skeletal muscle contraction responses. © 2013 Published by Elsevier B.V.

  11. An overview of techniques for the measurement of calcium distribution, calcium fluxes, and cytosolic free calcium in mammalian cells

    International Nuclear Information System (INIS)

    Borle, A.B.

    1990-01-01

    An array of techniques can be used to study cell calcium metabolism that comprises several calcium compartments and many types of transport systems such as ion channels, ATP-dependent pumps, and antiporters. The measurement of total call calcium brings little information of value since 60 to 80% of total cell calcium is actually bound to the extracellular glycocalyx. Cell fractionation and differential centrifugation have been used to study intracellular Ca 2+ compartmentalization, but the methods suffer from the possibility of Ca 2+ loss or redistribution among cell fractions. Steady-state kinetic analyses of 45 Ca uptake or desaturation curves have been used to study the distribution of Ca 2+ among various kinetic pools in living cells and their rate of Ca 2+ exchange, but the analyses are constrained by many limitations. Nonsteady-state tracer studies can provide information about rapid changes in calcium influx or efflux in and out of the cell. Zero-time kinetics of 45 Ca uptake can detect instantaneous changes in calcium influx, while 45 Ca fractional efflux ratio, can detect rapid stimulations or inhibitions of calcium efflux out of cells. The best strategy to study cell calcium metabolism is to use several different methods that focus on a specific problem from widely different angles

  12. Calcium Domains around Single and Clustered IP3 Receptors and Their Modulation by Buffers

    Science.gov (United States)

    Rüdiger, S.; Nagaiah, Ch.; Warnecke, G.; Shuai, J.W.

    2010-01-01

    Abstract We study Ca2+ release through single and clustered IP3 receptor channels on the ER membrane under presence of buffer proteins. Our computational scheme couples reaction-diffusion equations and a Markovian channel model and allows our investigating the effects of buffer proteins on local calcium concentrations and channel gating. We find transient and stationary elevations of calcium concentrations around active channels and show how they determine release amplitude. Transient calcium domains occur after closing of isolated channels and constitute an important part of the channel's feedback. They cause repeated openings (bursts) and mediate increased release due to Ca2+ buffering by immobile proteins. Stationary domains occur during prolonged activity of clustered channels, where the spatial proximity of IP3Rs produces a distinct [Ca2+] scale (0.5–10 μM), which is smaller than channel pore concentrations (>100 μM) but larger than transient levels. While immobile buffer affects transient levels only, mobile buffers in general reduce both transient and stationary domains, giving rise to Ca2+ evacuation and biphasic modulation of release amplitude. Our findings explain recent experiments in oocytes and provide a general framework for the understanding of calcium signals. PMID:20655827

  13. Vitamin E isomer δ-tocopherol enhances the efficiency of neural stem cell differentiation via L-type calcium channel.

    Science.gov (United States)

    Deng, Sihao; Hou, Guoqiang; Xue, Zhiqin; Zhang, Longmei; Zhou, Yuye; Liu, Chao; Liu, Yanqing; Li, Zhiyuan

    2015-01-12

    The effects of the vitamin E isomer δ-tocopherol on neural stem cell (NSC) differentiation have not been investigated until now. Here we investigated the effects of δ-tocopherol on NSC neural differentiation, maturation and its possible mechanisms. Neonatal rat NSCs were grown in suspended neurosphere cultures, and were identified by their expression of nestin protein and their capacity for self-renewal. Treatment with a low concentration of δ-tocopherol induced a significant increase in the percentage of β-III-tubulin-positive cells. δ-Tocopherol also stimulated morphological maturation of neurons in culture. We further observed that δ-tocopherol stimulation increased the expression of voltage-dependent Ca(2+) channels. Moreover, a L-type specific Ca(2+) channel blocker verapamil reduced the percentage of differentiated neurons after δ-tocopherol treatment, and blocked the effects of δ-tocopherol on NSC differentiation into neurons. Together, our study demonstrates that δ-tocopherol may act through elevation of L-type calcium channel activity to increase neuronal differentiation. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  14. T-type Ca2+ channels. New players in the aging brain

    Czech Academy of Sciences Publication Activity Database

    Proft, Juliane; Weiss, Norbert

    2014-01-01

    Roč. 7, č. 2 (2014), e28424/1-e28424/4 ISSN 1942-0889 Institutional support: RVO:61388963 Keywords : Alzheimer's disease * Amyloid beta * calcium channel * calcium signaling * T-type channel Subject RIV: CE - Biochemistry

  15. Depolarization of the conductance-voltage relationship in the NaV1.5 mutant, E1784K, is due to altered fast inactivation

    Science.gov (United States)

    Yu, Alec; Zhu, Wandi; Silva, Jonathan R.; Ruben, Peter C.

    2017-01-01

    E1784K is the most common mixed long QT syndrome/Brugada syndrome mutant in the cardiac voltage-gated sodium channel NaV1.5. E1784K shifts the midpoint of the channel conductance-voltage relationship to more depolarized membrane potentials and accelerates the rate of channel fast inactivation. The depolarizing shift in the midpoint of the conductance curve in E1784K is exacerbated by low extracellular pH. We tested whether the E1784K mutant shifts the channel conductance curve to more depolarized membrane potentials by affecting the channel voltage-sensors. We measured ionic currents and gating currents at pH 7.4 and pH 6.0 in Xenopus laevis oocytes. Contrary to our expectation, the movement of gating charges is shifted to more hyperpolarized membrane potentials by E1784K. Voltage-clamp fluorimetry experiments show that this gating charge shift is due to the movement of the DIVS4 voltage-sensor being shifted to more hyperpolarized membrane potentials. Using a model and experiments on fast inactivation-deficient channels, we show that changes to the rate and voltage-dependence of fast inactivation are sufficient to shift the conductance curve in E1784K. Our results localize the effects of E1784K to DIVS4, and provide novel insight into the role of the DIV-VSD in regulating the voltage-dependencies of activation and fast inactivation. PMID:28898267

  16. Depolarization of the conductance-voltage relationship in the NaV1.5 mutant, E1784K, is due to altered fast inactivation.

    Directory of Open Access Journals (Sweden)

    Colin H Peters

    Full Text Available E1784K is the most common mixed long QT syndrome/Brugada syndrome mutant in the cardiac voltage-gated sodium channel NaV1.5. E1784K shifts the midpoint of the channel conductance-voltage relationship to more depolarized membrane potentials and accelerates the rate of channel fast inactivation. The depolarizing shift in the midpoint of the conductance curve in E1784K is exacerbated by low extracellular pH. We tested whether the E1784K mutant shifts the channel conductance curve to more depolarized membrane potentials by affecting the channel voltage-sensors. We measured ionic currents and gating currents at pH 7.4 and pH 6.0 in Xenopus laevis oocytes. Contrary to our expectation, the movement of gating charges is shifted to more hyperpolarized membrane potentials by E1784K. Voltage-clamp fluorimetry experiments show that this gating charge shift is due to the movement of the DIVS4 voltage-sensor being shifted to more hyperpolarized membrane potentials. Using a model and experiments on fast inactivation-deficient channels, we show that changes to the rate and voltage-dependence of fast inactivation are sufficient to shift the conductance curve in E1784K. Our results localize the effects of E1784K to DIVS4, and provide novel insight into the role of the DIV-VSD in regulating the voltage-dependencies of activation and fast inactivation.

  17. Regulation of cardiomyocyte autophagy by calcium.

    Science.gov (United States)

    Shaikh, Soni; Troncoso, Rodrigo; Criollo, Alfredo; Bravo-Sagua, Roberto; García, Lorena; Morselli, Eugenia; Cifuentes, Mariana; Quest, Andrew F G; Hill, Joseph A; Lavandero, Sergio

    2016-04-15

    Calcium signaling plays a crucial role in a multitude of events within the cardiomyocyte, including cell cycle control, growth, apoptosis, and autophagy. With respect to calcium-dependent regulation of autophagy, ion channels and exchangers, receptors, and intracellular mediators play fundamental roles. In this review, we discuss calcium-dependent regulation of cardiomyocyte autophagy, a lysosomal mechanism that is often cytoprotective, serving to defend against disease-related stress and nutrient insufficiency. We also highlight the importance of the subcellular distribution of calcium and related proteins, interorganelle communication, and other key signaling events that govern cardiomyocyte autophagy. Copyright © 2016 the American Physiological Society.

  18. Azadirachtin blocks the calcium channel and modulates the cholinergic miniature synaptic current in the central nervous system of Drosophila.

    Science.gov (United States)

    Qiao, Jingda; Zou, Xiaolu; Lai, Duo; Yan, Ying; Wang, Qi; Li, Weicong; Deng, Shengwen; Xu, Hanhong; Gu, Huaiyu

    2014-07-01

    Azadirachtin is a botanical pesticide, which possesses conspicuous biological actions such as insecticidal, anthelmintic, antifeedancy, antimalarial effects as well as insect growth regulation. Deterrent for chemoreceptor functions appears to be the main mechanism involved in the potent biological actions of Azadirachtin, although the cytotoxicity and subtle changes to skeletal muscle physiology may also contribute to its insecticide responses. In order to discover the effects of Azadirachtin on the central nervous system (CNS), patch-clamp recording was applied to Drosophila melanogaster, which has been widely used in neurological research. Here, we describe the electrophysiological properties of a local neuron located in the suboesophageal ganglion region of D. melanogaster using the whole brain. The patch-clamp recordings suggested that Azadirachtin modulates the properties of cholinergic miniature excitatory postsynaptic current (mEPSC) and calcium currents, which play important roles in neural activity of the CNS. The frequency of mEPSC and the peak amplitude of the calcium currents significantly decreased after application of Azadirachtin. Our study indicates that Azadirachtin can interfere with the insect's CNS via inhibition of excitatory cholinergic transmission and partly blocking the calcium channel. © 2013 Society of Chemical Industry.

  19. The effects of prior calcium channel blocker therapy on creatine kinase-MB levels after percutaneous coronary interventions

    Directory of Open Access Journals (Sweden)

    Oyku Gulmez

    2008-12-01

    Full Text Available Oyku Gulmez, Ilyas Atar, Bülent Ozin, Mehmet Emin Korkmaz, Aslı Atar, Alp Aydinalp, Aylin Yildirir, Haldun MuderrisogluBaskent University Faculty of Medicine, Department of Cardiology, Ankara, TurkeyBackground: Use of intracoronary calcium channel blockers (CCBs during percutaneous coronary intervention (PCI has been shown to have favorable effects on coronary blood flow. We aimed to investigate the effects of CCBs administrated perorally on creatine kinase-MB (CK-MB levels in patients undergoing elective PCI.Methods: A total of 570 patients who underwent PCI were evaluated for CK-MB elevation. Patients who were on CCB therapy when admitted to the hospital constituted the CCB group. No CCBs were given to the rest of the patients during the periprocedural period and these patients served as the control group. Blood samples for CK-MB were obtained before and at 6 h, 24 h, and 36 h after the procedure.Results: 217 patients were in the CCB group (mean age 60.2 ± 9.3 years, 162 males, and 353 were in the control group (mean age 60.0 ± 10.1 years, 262 males. CK-MB levels increased above the normal values in 41 patients (18.9% of the CCBs group and in 97 patients (27.5% of the control group (p = 0.02. Median CK-MB levels were significantly higher in the control group for all studied hours (for all p < 0.05.Conclusions: Prior oral CCB therapy may have favorable effects in preventing myocyte necrosis after elective PCI.Keywords: calcium channel blockers, myonecrosis, percutaneous coronary interventions

  20. The metabolic impact of β-hydroxybutyrate on neurotransmission: Reduced glycolysis mediates changes in calcium responses and KATP channel receptor sensitivity

    DEFF Research Database (Denmark)

    Lund, Trine Meldgaard; Ploug, K.B.; Iversen, Anne

    2015-01-01

    -hydroxybutyrate might change neuronal function as there is a known coupling between metabolism and neurotransmission. The purpose of this study was to shed light on the effects of the ketone body β-hydroxybutyrate on glycolysis and neurotransmission in cultured murine glutamatergic neurons. Previous studies have shown...... an effect of β-hydroxybutyrate on glucose metabolism, and the present study further specified this by showing attenuation of glycolysis when β-hydroxybutyrate was present in these neurons. In addition, the NMDA receptor-induced calcium responses in the neurons were diminished in the presence of β...... to a combination of glucose and R-β-hydroxybutyrate in cultured neurons. Using the latter combination, glycolysis was diminished, NMDA receptor-induced calcium responses were lower, and the KATP channel blocker glibenclamide caused a higher transmitter release....

  1. Radiosynthesis of dimethyl-2-[{sup 18}F]-(fluoromethyl)-6-methyl-4-(2-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylate for L-type calcium channel imaging

    Energy Technology Data Exchange (ETDEWEB)

    Sadeghpour, H. [Nuclear Medicine Research Group, Agricultural, Medical and Industrial Research School (AMIRS), Karaj (Iran); Faculty of Pharmacy and Pharmaceutical Sciences Research Center, Tehran Univ. of Medical Sciences, Tehran (Iran); Jalilian, A.R.; Akhlaghi, M.; Mirzaei, M. [Nuclear Medicine Research Group, Agricultural, Medical and Industrial Research School (AMIRS), Karaj (Iran); Shafiee, A. [Faculty of Pharmacy and Pharmaceutical Sciences Research Center, Tehran Univ. of Medical Sciences, Tehran (Iran); Miri, R. [Medicinal and Natural Products Chemistry Research Center, Shiraz Univ. of Medical Sciences, Shiraz (Iran)

    2008-07-01

    Dimethyl 2-(fluoromethyl)-6-methyl-4-(2-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylate 4a, a fluorinated nifedipine analog, has been shown to elicit significant calcium channel blocker activity using a guinea pig ileal longitudinal smooth muscle model. In order to perform biological studies for detection of L-type calcium channel distribution, we decided to prepare the [{sup 18}F]-labeled compound. The latter compound was prepared in no-carrier-added (n.c.a.) form from dimethyl 2-(bromomethyl)-6-methyl-4-(2-nitrophenyl)-1,4-dihydropyridine-3,5-dicarboxylate 2 in one step at 80 C in Kryptofix[222]/K[{sup 18}F]F and acetonitrile as a solvent in 15 min. Column chromatography afforded the radiochemically pure compound in 20 min. Radiochemical purity of the {sup 18}F-nifedipine was determined by RTLC and HPLC (> 98%) and specific activity of 21-48 GBq/{mu}mol (EOB). (orig.)

  2. Effect of Topical Calcium Channel Blockers on Intraocular Pressure in Steroid-induced Glaucoma.

    Science.gov (United States)

    Ganekal, Sunil; Dorairaj, Syril; Jhanji, Vishal; Kudlu, Krishnaprasad

    2014-01-01

    To evaluate the effect of 0.125% verapamil and 0.5% diltiazem eye drops on intraocular pressure (IOP) in steroid-induced glaucoma in rabbit eyes. A total of 18 rabbits with steroid-induced glaucoma were divided into three groups (A, B and C; n = 6 each). Right eyes in groups A, B and C received 0.5% diltiazem, 0.125% verapamil and 0.5% timolol eye drops twice daily for 12 days, respectively; whereas, left eyes received distilled water. IOP was measured with Tono-pen XL at baseline, day 4, day 8, and day 12 of treatment. Both 0.5% diltiazem and 0.125% verapamil eye drops significantly reduced IOP compared to control eyes (p cite this article: Ganekal S, Dorairaj S, Jhanji V, Kudlu K. Effect of Topical Calcium Channel Blockers on Intraocular Pressure in Steroid-induced Glaucoma. J Current Glau Prac 2014;8(1):15-19.

  3. Discovery of a Potent, Selective T-type Calcium Channel Blocker as a Drug Candidate for the Treatment of Generalized Epilepsies.

    Science.gov (United States)

    Bezençon, Olivier; Heidmann, Bibia; Siegrist, Romain; Stamm, Simon; Richard, Sylvia; Pozzi, Davide; Corminboeuf, Olivier; Roch, Catherine; Kessler, Melanie; Ertel, Eric A; Reymond, Isabelle; Pfeifer, Thomas; de Kanter, Ruben; Toeroek-Schafroth, Michael; Moccia, Luca G; Mawet, Jacques; Moon, Richard; Rey, Markus; Capeleto, Bruno; Fournier, Elvire

    2017-12-14

    We report here the discovery and pharmacological characterization of N-(1-benzyl-1H-pyrazol-3-yl)-2-phenylacetamide derivatives as potent, selective, brain-penetrating T-type calcium channel blockers. Optimization focused mainly on solubility, brain penetration, and the search for an aminopyrazole metabolite that would be negative in an Ames test. This resulted in the preparation and complete characterization of compound 66b (ACT-709478), which has been selected as a clinical candidate.

  4. The interplay between HIF-1 and calcium signalling in cancer.

    Science.gov (United States)

    Azimi, Iman

    2018-04-01

    The interplay between hypoxia-inducible factor-1 (HIF-1) and calcium in cancer has begun to be unravelled with recent findings demonstrating the relationships between the two in different cancer types. This is an area of significance considering the crucial roles of both HIF-1 and calcium signalling in cancer progression and metastasis. This review summarises the experimental evidence of the crosstalk between HIF-1 and specific calcium channels, pumps and regulators in the context of cancer. HIF-1 as a master regulator of hypoxic transcriptional responses, mediates transcription of several calcium modulators. On the other hand, specific calcium channels and pumps regulate HIF-1 activity through controlling its transcription, translation, stabilisation, or nuclear translocation. Identifying the interplay between HIF-1 and components of the calcium signal will give new insights into mechanisms underlying cellular responses to physiological and pathophysiological cues, and may provide novel and more efficient therapeutic strategies for the control of cancer progression. Copyright © 2018 Elsevier Ltd. All rights reserved.

  5. T-type calcium channel antagonism decreases motivation for nicotine and blocks nicotine- and cue-induced reinstatement for a response previously reinforced with nicotine.

    Science.gov (United States)

    Uslaner, Jason M; Vardigan, Joshua D; Drott, Jason M; Uebele, Victor N; Renger, John J; Lee, Ariel; Li, Zhaoxia; Lê, A D; Hutson, Pete H

    2010-10-15

    Recent evidence suggests an involvement of T-type calcium channels in the effects of drugs of abuse. We examined the influence of the novel, potent, and selective T-type calcium channel antagonist [2-(4-cyclopropylphenyl)-N-((1R)-1-{5-[2,2,2-trifluoroethyl]oxo}pyridine-2-yl)ethyl]acetamide] (TTA-A2) (.3, 1, or 3 mg/kg) on motivation for nicotine, as measured by nicotine self-administration on a progressive ratio (PR) schedule, and nicotine- and cue-induced reinstatement for a response previously reinforced with nicotine delivery (n = 11 or 12 Long Evans rats/group). Furthermore, we examined the specificity of the TTA-A2 effects by characterizing its influence on PR responding for food (in the absence or presence of nicotine-potentiated responding), food- versus nicotine-induced cue-potentiated reinstatement for a response previously reinforced by food administration (n = 11 or 12 Wistar Hannover rats/group), and its ability to induce a conditioned place aversion. TTA-A2 dose-dependently decreased self-administration of nicotine on a PR schedule and the ability of both nicotine and a cue paired with nicotine to reinstate responding. The effects were specific for nicotine's incentive motivational properties, as TTA-A2 did not influence responding for food on a PR schedule but did attenuate the ability of nicotine to potentiate responding for food. Likewise, TTA-A2 did not alter food-induced cue-potentiated reinstatement for a response previously reinforced by food but did decrease nicotine-induced cue-potentiated reinstatement. Finally, TTA-A2 did not produce an aversive state, as indicated by a lack of ability to induce conditioned place aversion. These data suggest that T-type calcium channel antagonists have potential for alleviating nicotine addiction by selectively decreasing the incentive motivational properties of nicotine. Copyright © 2010 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

  6. Autoimmune encephalitis associated with voltage-gated potassium channels-complex and leucine-rich glioma-inactivated 1 antibodies - a national cohort study

    DEFF Research Database (Denmark)

    Celicanin, M; Blaabjerg, Morten; Maersk-Moller, C

    2017-01-01

    BACKGROUND AND PURPOSE: The aim of this study was to describe clinical and paraclinical characteristics of all Danish patients who tested positive for anti-voltage-gated potassium channels (VGKC)-complex, anti-leucine-rich glioma-inactivated 1 (LGI1) and anti-contactin-associated protein-2...... antibodies in the serum/cerebrospinal fluid between 2009 and 2013 with follow-up interviews in 2015 and 2016. METHODS: We evaluated antibody status, symptoms leading to testing, course of disease, suspected diagnosis and time of admission as well as diagnosis and treatment. All magnetic resonance imaging......-Barré syndrome, Creutzfeldt-Jakob disease, neuromyotonia and anti-N-methyl-D-aspartate receptor encephalitis. Magnetic resonance imaging abnormalities were demonstrated in 69% of the LGI1-positive patients. Two patients with normal magnetic resonance imaging demonstrated temporal lobe hypermetabolism using (18...

  7. Activation of endothelial and epithelial K(Ca) 2.3 calcium-activated potassium channels by NS309 relaxes human small pulmonary arteries and bronchioles

    DEFF Research Database (Denmark)

    Kroigaard, Christel; Dalsgaard, Thomas; Nielsen, Gorm

    2012-01-01

    BACKGROUND AND PURPOSE: Small (K(Ca) 2) and intermediate (K(Ca) 3.1) conductance calcium-activated potassium channels (K(Ca) ) may contribute to both epithelium- and endothelium-dependent relaxations, but this has not been established in human pulmonary arteries and bronchioles. Therefore, we inv...... targets for treatment of pulmonary hypertension and chronic obstructive pulmonary disease....

  8. Allopregnanolone-induced rise in intracellular calcium in embryonic hippocampal neurons parallels their proliferative potential

    Directory of Open Access Journals (Sweden)

    Brinton Roberta

    2008-12-01

    Full Text Available Abstract Background Factors that regulate intracellular calcium concentration are known to play a critical role in brain function and neural development, including neural plasticity and neurogenesis. We previously demonstrated that the neurosteroid allopregnanolone (APα; 5α-pregnan-3α-ol-20-one promotes neural progenitor proliferation in vitro in cultures of rodent hippocampal and human cortical neural progenitors, and in vivo in triple transgenic Alzheimer's disease mice dentate gyrus. We also found that APα-induced proliferation of neural progenitors is abolished by a calcium channel blocker, nifedipine, indicating a calcium dependent mechanism for the proliferation. Methods In the present study, we investigated the effect of APα on the regulation of intracellular calcium concentration in E18 rat hippocampal neurons using ratiometric Fura2-AM imaging. Results Results indicate that APα rapidly increased intracellular calcium concentration in a dose-dependent and developmentally regulated manner, with an EC50 of 110 ± 15 nM and a maximal response occurring at three days in vitro. The stereoisomers 3β-hydroxy-5α-hydroxy-pregnan-20-one, and 3β-hydroxy-5β-hydroxy-pregnan-20-one, as well as progesterone, were without significant effect. APα-induced intracellular calcium concentration increase was not observed in calcium depleted medium and was blocked in the presence of the broad spectrum calcium channel blocker La3+, or the L-type calcium channel blocker nifedipine. Furthermore, the GABAA receptor blockers bicuculline and picrotoxin abolished APα-induced intracellular calcium concentration rise. Conclusion Collectively, these data indicate that APα promotes a rapid, dose-dependent, stereo-specific, and developmentally regulated increase of intracellular calcium concentration in rat embryonic hippocampal neurons via a mechanism that requires both the GABAA receptor and L-type calcium channel. These data suggest that AP

  9. SK2 channels regulate mitochondrial respiration and mitochondrial Ca2+ uptake

    NARCIS (Netherlands)

    Honrath, Birgit; Matschke, Lina; Meyer, Tammo; Magerhans, Lena; Perocchi, Fabiana; Ganjam, Goutham K; Zischka, Hans; Krasel, Cornelius; Gerding, Albert; Bakker, Barbara M; Bünemann, Moritz; Strack, Stefan; Decher, Niels; Culmsee, Carsten; Dolga, Amalia M

    Mitochondrial calcium ([Ca(2+)]m) overload and changes in mitochondrial metabolism are key players in neuronal death. Small conductance calcium-activated potassium (SK) channels provide protection in different paradigms of neuronal cell death. Recently, SK channels were identified at the inner

  10. No apparent role for T-type Ca2+ channels in renal autoregulation

    DEFF Research Database (Denmark)

    Frandsen, Rasmus Hassing; Salomonsson, Max; Hansen, Pernille B. Lærkegaard

    2016-01-01

    -type and CaV3.1 knockout mice were assessed. Autoregulation of renal blood flow was examined during acute increases in RPP in normo- and hypertensive rats under pharmacological blockade of T- and L-type calcium channels using mibefradil (0.1 μM) and nifedipine (1 μM). In contrast to the results from previous......Renal autoregulation protects glomerular capillaries against increases in renal perfusion pressure (RPP). In the mesentery, both L- and T-type calcium channels are involved in autoregulation. L-type calcium channels participate in renal autoregulation, but the role of T-type channels is not fully...... pharmacological studies, genetic deletion of T-type channels CaV3.1 did not affect renal autoregulation. Pharmacological blockade of T-type channels using concentrations of mibefradil which specifically blocks T-type channels also had no effect in wild-type or knockout mice. Blockade of L-type channels...

  11. Ion channel recordings on an injection-molded polymer chip

    DEFF Research Database (Denmark)

    Tanzi, Simone; Matteucci, Marco; Christiansen, Thomas Lehrmann

    2013-01-01

    state-of-the-art system for automated ion channel recordings. These experiments considered current–voltage (IV) relationships for activation and inactivation of the Nav1.7 channels and their sensitivity to a local anesthetic, lidocaine. Both IVs and lidocaine dose–response curves obtained from...

  12. The L-Type Voltage-Gated Calcium Channel Ca [subscript V] 1.2 Mediates Fear Extinction and Modulates Synaptic Tone in the Lateral Amygdala

    Science.gov (United States)

    Temme, Stephanie J.; Murphy, Geoffrey G.

    2017-01-01

    L-type voltage-gated calcium channels (LVGCCs) have been implicated in both the formation and the reduction of fear through Pavlovian fear conditioning and extinction. Despite the implication of LVGCCs in fear learning and extinction, studies of the individual LVGCC subtypes, Ca[subscript V]1.2 and Ca[subscript V] 1.3, using transgenic mice have…

  13. Presynaptic muscarinic receptors, calcium channels, and protein kinase C modulate the functional disconnection of weak inputs at polyinnervated neonatal neuromuscular synapses.

    Science.gov (United States)

    Santafe, M M; Garcia, N; Lanuza, M A; Tomàs, M; Besalduch, N; Tomàs, J

    2009-04-01

    We studied the relation among calcium inflows, voltage-dependent calcium channels (VDCC), presynaptic muscarinic acetylcholine receptors (mAChRs), and protein kinase C (PKC) activity in the modulation of synapse elimination. We used intracellular recording to determine the synaptic efficacy in dually innervated endplates of the levator auris longus muscle of newborn rats during axonal competition in the postnatal synaptic elimination period. In these dual junctions, the weak nerve terminal was potentiated by partially reducing calcium entry (P/Q-, N-, or L-type VDCC-specific block or 500 muM magnesium ions), M1- or M4-type selective mAChR block, or PKC block. Moreover, reducing calcium entry or blocking PKC or mAChRs results in unmasking functionally silent nerve endings that now recover neurotransmitter release. Our results show interactions between these molecules and indicate that there is a release inhibition mechanism based on an mAChR-PKC-VDCC intracellular cascade. When it is fully active in certain weak motor axons, it can depress ACh release and even disconnect synapses. We suggest that this mechanism plays a central role in the elimination of redundant neonatal synapses, because functional axonal withdrawal can indeed be reversed by mAChRs, VDCCs, or PKC block.

  14. Presynaptic calcium signalling in cerebellar mossy fibres

    DEFF Research Database (Denmark)

    Thomsen, Louiza Bohn; Jörntell, Henrik; Midtgaard, Jens

    2010-01-01

    Whole-cell recordings were obtained from mossy fibre terminals in adult turtles in order to characterize the basic membrane properties. Calcium imaging of presynaptic calcium signals was carried out in order to analyse calcium dynamics and presynaptic GABA B inhibition. A tetrodotoxin (TTX......)-sensitive fast Na(+) spike faithfully followed repetitive depolarizing pulses with little change in spike duration or amplitude, while a strong outward rectification dominated responses to long-lasting depolarizations. High-threshold calcium spikes were uncovered following addition of potassium channel blockers....... Calcium imaging using Calcium-Green dextran revealed a stimulus-evoked all-or-none TTX-sensitive calcium signal in simple and complex rosettes. All compartments of a complex rosette were activated during electrical activation of the mossy fibre, while individual simple and complex rosettes along an axon...

  15. Modulation of T-type Ca2+ channels by Lavender and Rosemary extracts.

    Directory of Open Access Journals (Sweden)

    Chaymae El Alaoui

    Full Text Available Medicinal plants represent a significant reservoir of unexplored substances for early-stage drug discovery. Of interest, two flowering Mediterranean plants have been used for thousands of years for their beneficial effects on nervous disorders, including anxiety and mood. However, the therapeutic potential of these plants regarding their ability to target ion channels and neuronal excitability remains largely unknown. Towards this goal, we have investigated the ability of Lavender and Rosemary to modulate T-type calcium channels (TTCCs. TTCCs play important roles in neuronal excitability, neuroprotection, sensory processes and sleep. These channels are also involved in epilepsy and pain. Using the whole-cell patch-clamp technique, we have characterized how Lavender and Rosemary extracts, as well as their major active compounds Linalool and Rosmarinic acid, modulate the electrophysiological properties of recombinant TTCCs (CaV3.2 expressed in HEK-293T cells. Both the methanolic and essential oil extracts as well as the active compounds of these plants inhibit Cav3.2 current in a concentration-dependent manner. In addition, these products also induce a negative shift of the steady-state inactivation of CaV3.2 current with no change in the activation properties. Taken together, our findings reveal that TTCCs are a molecular target of the Lavender and Rosemary compounds, suggesting that inhibition of TTCCs could contribute to the anxiolytic and the neuroprotective effects of these plants.

  16. Adrenomedullin increases the short-circuit current in the rat prostate: Receptors, chloride channels, the effects of cAMP and calcium ions and implications on fluid secretion.

    Science.gov (United States)

    Liao, S B; Cheung, K H; Cheung, M P L; Wong, P F; O, W S; Tang, F

    2014-05-01

    In this study, we have investigated the effects of adrenomedullin on chloride and fluid secretion in the rat prostate. The presence of adrenomedullin (ADM) in rat prostate was confirmed using immunostaining, and the molecular species was determined using gel filtration chromatography coupled with an enzyme-linked assay for ADM. The effects of ADM on fluid secretion were studied by short-circuit current technique in a whole mount preparation of the prostate in an Ussing chamber. The results indicated that the ADM level was higher in the ventral than the dorso-lateral prostate and the major molecular species was the active peptide. ADM increased the short-circuit current through both the cAMP- and calcium-activated chloride channels in the ventral lobe, but only through the calcium-activated channels in the dorso-lateral lobe. These stimulatory effects were blocked by the calcitonin gene-related peptide (CGRP) receptor antagonist, hCGRP8-37. We conclude that ADM may regulate prostatic fluid secretion through the chloride channels, which may affect the composition of the seminal plasma bathing the spermatozoa and hence fertility. © 2014 American Society of Andrology and European Academy of Andrology.

  17. Imaging large cohorts of single ion channels and their activity

    Directory of Open Access Journals (Sweden)

    Katia eHiersemenzel

    2013-09-01

    Full Text Available As calcium is the most important signaling molecule in neurons and secretory cells, amongst many other cell types, it follows that an understanding of calcium channels and their regulation of exocytosis is of vital importance. Calcium imaging using calcium dyes such as Fluo3, or FRET-based dyes that have been used widely has provided invaluable information, which combined with modeling has estimated the sub-types of channels responsible for triggering the exocytotic machinery as well as inferences about the relative distances away from vesicle fusion sites these molecules adopt. Importantly, new super-resolution microscopy techniques, combined with novel Ca2+ indicators and imaginative imaging approaches can now define directly the nanoscale locations of very large cohorts of single channel molecules in relation to single vesicles. With combinations of these techniques the activity of individual channels can be visualized and quantified using novel Ca2+ indicators. Fluorescently labeled specific channel toxins can also be used to localize endogenous assembled channel tetramers. Fluorescence lifetime imaging microscopy and other single-photon-resolution spectroscopic approaches offer the possibility to quantify protein-protein interactions between populations of channels and the SNARE protein machinery for the first time. Together with simultaneous electrophysiology, this battery of quantitative imaging techniques has the potential to provide unprecedented detail describing the locations, dynamic behaviours, interactions and conductance activities of many thousands of channel molecules and vesicles in living cells.

  18. Polyaniline-graphene oxide nanocomposite sensor for quantification of calcium channel blocker levamlodipine.

    Science.gov (United States)

    Jain, Rajeev; Sinha, Ankita; Khan, Ab Lateef

    2016-08-01

    A novel polyaniline-graphene oxide nanocomposite (PANI/GO/GCE) sensor has been fabricated for quantification of a calcium channel blocker drug levamlodipine (LAMP). Fabricated sensor has been characterized by electrochemical impedance spectroscopy, square wave and cyclic voltammetry, Raman spectroscopy and Fourier transform infrared (FTIR) spectroscopy. The developed PANI/GO/GCE sensor has excellent analytical performance towards electrocatalytic oxidation as compared to PANI/GCE, GO/GCE and bare GCE. Under optimized experimental conditions, the fabricated sensor exhibits a linear response for LAMP for its oxidation over a concentration range from 1.25μgmL(-1) to 13.25μgmL(-1) with correlation coefficient of 0.9950 (r(2)), detection limit of 1.07ngmL(-1) and quantification limit of 3.57ngmL(-1). The sensor shows an excellent performance for detecting LAMP with reproducibility of 2.78% relative standard deviation (RSD). The proposed method has been successfully applied for LAMP determination in pharmaceutical formulation with a recovery from 99.88% to 101.75%. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. 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

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

    Science.gov (United States)

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

    2014-01-01

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

  1. Movement of calcium signals and calcium-binding proteins: firewalls, traps and tunnels.

    Science.gov (United States)

    Barrow, S L; Sherwood, M W; Dolman, N J; Gerasimenko, O V; Voronina, S G; Tepikin, A V

    2006-06-01

    In the board game 'Snakes and Ladders', placed on the image of a pancreatic acinar cell, calcium ions have to move from release sites in the secretory region to the nucleus. There is another important contraflow - from calcium entry channels in the basal part of the cell to ER (endoplasmic reticulum) terminals in the secretory granule region. Both transport routes are perilous as the messenger can disappear in any place on the game board. It can be grabbed by calcium ATPases of the ER (masquerading as a snake but functioning like a ladder) and tunnelled through its low buffering environment, it can be lured into the whirlpools of mitochondria uniporters and forced to regulate the tricarboxylic acid cycle, and it can be permanently placed inside the matrix of secretory granules and released only outside the cell. The organelles could trade calcium (e.g. from the ER to mitochondria and vice versa) almost depriving this ion the light of the cytosol and noble company of cytosolic calcium buffers. Altogether it is a rich and colourful story.

  2. A membrane model for cytosolic calcium oscillations. A study using Xenopus oocytes.

    OpenAIRE

    Jafri, M S; Vajda, S; Pasik, P; Gillo, B

    1992-01-01

    Cytosolic calcium oscillations occur in a wide variety of cells and are involved in different cellular functions. We describe these calcium oscillations by a mathematical model based on the putative electrophysiological properties of the endoplasmic reticulum (ER) membrane. The salient features of our membrane model are calcium-dependent calcium channels and calcium pumps in the ER membrane, constant entry of calcium into the cytosol, calcium dependent removal from the cytosol, and buffering ...

  3. Modulation of the transient outward current (Ito) in rat cardiac myocytes and human Kv4.3 channels by mefloquine

    International Nuclear Information System (INIS)

    Perez-Cortes, E.J.; Islas, A.A.; Arevalo, J.P.; Mancilla, C.; Monjaraz, E.; Salinas-Stefanon, E.M.

    2015-01-01

    The antimalarial drug mefloquine, is known to be a potassium channel blocker, although its mechanism of action has not being elucidated and its effects on the transient outward current (I to ) and the molecular correlate, the K v 4.3 channel has not being studied. Here, we describe the mefloquine-induced inhibition of the rat ventricular I to and of CHO cells co-transfected with human K v 4.3 and its accessory subunit hKChIP2C by whole-cell voltage-clamp. Mefloquine inhibited rat I to and hK v 4.3 + KChIP2C currents in a concentration-dependent manner with a limited voltage dependence and similar potencies (IC 50 = 8.9 μM and 10.5 μM for cardiac myocytes and K v 4.3 channels, respectively). In addition, mefloquine did not affect the activation of either current but significantly modified the hK v 4.3 steady-state inactivation and recovery from inactivation. The effects of this drug was compared with that of 4-aminopyridine (4-AP), a well-known potassium channel blocker and its binding site does not seem to overlap with that of 4-AP. - Highlights: • Mefloquine inhibited ventricular I to and hK v 4.3 channels. IC 50 = 8.9 and 10.5 μM. • Inactivation and recovery from inactivation in the hK v 4.3 channels were modified by mefloquine. • Mefloquine displayed a higher affinity for the inactivated state. • The binding site for mefloquine may be located in the extracellular side of the channel.

  4. Comparison of the efficacy of dihydropyridine calcium channel blockers in African American patients with hypertension. ISHIB Investigators Group. International Society on Hypertension in Blacks.

    Science.gov (United States)

    Hall, W D; Reed, J W; Flack, J M; Yunis, C; Preisser, J

    1998-10-12

    Hypertension is a prevalent disease among African Americans, and successful treatment rates are low. Since calcium channel blockers are well-tolerated and efficacious in African Americans, we undertook this study to compare the efficacy, safety, and tolerability of 3 commonly prescribed calcium channel blockers: amlodipine besylate (Norvasc), nifedipine coat core (CC) (Adalat CC), and nifedipine gastrointestinal therapeutic system (GITS) (Procardia XL). One hundred ninety-two hypertensive patients across 10 study centers were randomly assigned to double-blind monotherapy with amlodipine besylate (5 mg/d), nifedipine CC (30 mg/d), or nifedipine GITS (30 mg/d) for 8 weeks. Patients not achieving therapeutic response after 4 weeks had their dose doubled for the next 4 weeks. The primary end point was a comparison of the average reduction (week 8 minus baseline) in 24-hour ambulatory diastolic blood pressure (DBP). Secondary end points included a comparison of average 24-hour ambulatory systolic blood pressure (SBP), office SBP or DBP reduction, responder rates, safety, and tolerability. One hundred sixty-three patients were evaluable for efficacy after 8 weeks. There was no significant difference in the average 24-hour ambulatory DBP (-8.5, -9.0, and -6.1 mm Hg, respectively) or SBP (-14.3, -15.7, and -11.8 mm Hg, respectively) reduction. Average office SBP and DBP were reduced to a comparable degree (19-22 mm Hg [P =.50] and 12-14 mm Hg [P =.51], respectively). Responder rates (DBP or = 10 mm Hg) were similar (P = .38). Discontinuation rates and adverse event frequency were distributed similarly across the 3 treatment groups. The efficacy, safety, and tolerability of the 3 dihydropyridine calcium channel blockers are equivalent in African Americans with stages 1 and 2 hypertension.

  5. Vitamin K3 inhibits mouse uterine contraction in vitro via interference with the calcium transfer and the potassium channels.

    Science.gov (United States)

    Zhang, Xian-Xia; Lu, Li-Min; Wang, Li

    2016-08-05

    Previous studies have demonstrated vitamin K3 had a great relief to smooth muscle spastic disorders, but no researches have yet pinpointed its possible anti-contractile activity in the uterus. Here, we evaluated the effect of vitamin K3 on myometrial contractility and explored the possible mechanisms of vitamin K3 action. Myograph apparatus were used to record the changes in contractility of isolated mouse uterine strips in a tissue bath. Uterine strips were exposed to vitamin K3 or vehicle. Vitamin K3 suppressed spontaneous contractions in a concentration dependent manner. It significantly decreased the contractile frequency induced by PGF2ɑ but not their amplitude (expect 58.0 μM). Prior incubation with vitamin K3 reduced the effectiveness of PGF2ɑ-induced contraction. The antispasmodic effect of vitamin K3 was also sensitive to potassium channel blockers, such as tetraethylammonium, 4-aminopyridine, iberiotoxin) but not to the nitric oxide related pathway blockers. High concentrations (29.0, 58.0 μM) of vitamin K3 weakened the Ca(2+) dose response and inhibited phase 1 contraction (intracellular stored calcium release). These dates suggest that vitamin K3 specifically suppresses myometrial contractility by affecting calcium and potassium channels; thus, this approach has potential therapy for uterine contractile activity related disorders. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Intracellular sphingosine releases calcium from lysosomes.

    Science.gov (United States)

    Höglinger, Doris; Haberkant, Per; Aguilera-Romero, Auxiliadora; Riezman, Howard; Porter, Forbes D; Platt, Frances M; Galione, Antony; Schultz, Carsten

    2015-11-27

    To elucidate new functions of sphingosine (Sph), we demonstrate that the spontaneous elevation of intracellular Sph levels via caged Sph leads to a significant and transient calcium release from acidic stores that is independent of sphingosine 1-phosphate, extracellular and ER calcium levels. This photo-induced Sph-driven calcium release requires the two-pore channel 1 (TPC1) residing on endosomes and lysosomes. Further, uncaging of Sph leads to the translocation of the autophagy-relevant transcription factor EB (TFEB) to the nucleus specifically after lysosomal calcium release. We confirm that Sph accumulates in late endosomes and lysosomes of cells derived from Niemann-Pick disease type C (NPC) patients and demonstrate a greatly reduced calcium release upon Sph uncaging. We conclude that sphingosine is a positive regulator of calcium release from acidic stores and that understanding the interplay between Sph homeostasis, calcium signaling and autophagy will be crucial in developing new therapies for lipid storage disorders such as NPC.

  7. Pharmacological modulation of SK3 channels

    DEFF Research Database (Denmark)

    Grunnet, M; Jespersen, Thomas; Angelo, K

    2001-01-01

    Small-conductance, calcium-activated K+ channels (SK channels) are voltage-insensitive channels that have been identified molecularly within the last few years. As SK channels play a fundamental role in most excitable cells and participate in afterhyperpolarization (AHP) and spike-frequency adapt...... at concentrations of 3 microM and above. Amitriptyline, a tricyclic antidepressive widely used clinically, inhibits SK3 channels with an IC50 of 39.1 +/- 10 microM (n=6)....

  8. Resveratrol Protects Against Ultraviolet A-Mediated Inhibition of the Phagocytic Function of Human Retinal Pigment Epithelial Cells Via Large-Conductance Calcium-Activated Potassium Channels

    Directory of Open Access Journals (Sweden)

    Shwu-Jiuan Sheu

    2009-07-01

    Full Text Available This study was undertaken to examine the protective effect of resveratrol on human retinal pigment epithelial (RPE cell phagocytosis against ultraviolet irradiation damage. Cultured RPE cells were exposed to ultraviolet A (UVA, 20 minutes irradiation, and treated with meclofenamic acid (30μM, 20 minutes, paxilline (100 μM, 20 minutes or resveratrol (10μM, 20 minutes. Meclofenamic acid and resveratrol were given after exposure to UVA. Pretreatment with meclofenamic acid, resveratrol or paxilline before UVA irradiation was also performed. Fluorescent latex beads were then fed for 4 hours and the phagocytotic function was assessed by flow cytometry. UVA irradiation inhibited the phagocytic function of human RPE cells. The large-conductance calcium-activated potassium channel activator meclofenamic acid ameliorated the damage caused by UVA irradiation. Pretreatment with resveratrol acid also provided protection against damage caused by UVA. Posttreatment with meclofenamic acid offered mild protection, whereas resveratrol did not. In conclusion, the red wine flavonoid resveratrol ameliorated UVA-mediated inhibition of human RPE phagocytosis. The underlying mechanism might involve the large-conductance calcium-activated potassium channels.

  9. Neuronal calcium channel antagonists. Discrimination between calcium channel subtypes using omega-conotoxin from Conus magus venom

    International Nuclear Information System (INIS)

    Olivera, B.M.; Cruz, L.J.; de Santos, V.

    1987-01-01

    The omega-conotoxins from the venom of fish-hunting cone snails are probably the most useful of presently available ligands for neuronal Ca channels from vertebrates. Two of these peptide toxins, omega-conotoxins MVIIA and MVIIB from the venom of Conus magus, were purified. The amino acid sequences show significant differences from omega-conotoxins from Conus geographus. Total synthesis of omega-conotoxin MVIIA was achieved, and biologically active radiolabeled toxin was produced by iodination. Although omega-conotoxins from C. geographus (GVIA) and C. magus (MVIIA) appear to compete for the same sites in mammalian brain, in amphibian brain the high-affinity binding of omega-conotoxin MVIIA has narrower specificity. In this system, it is demonstrated that a combination of two omega-conotoxins can be used for biochemically defining receptor subtypes and suggested that these correspond to subtypes of neutronal Ca 2+ channels

  10. Substituted N-(biphenyl-4'-yl)methyl (R)-2-acetamido-3-methoxypropionamides: potent anticonvulsants that affect frequency (use) dependence and slow inactivation of sodium channels.

    Science.gov (United States)

    Lee, Hyosung; Park, Ki Duk; Torregrosa, Robert; Yang, Xiao-Fang; Dustrude, Erik T; Wang, Yuying; Wilson, Sarah M; Barbosa, Cindy; Xiao, Yucheng; Cummins, Theodore R; Khanna, Rajesh; Kohn, Harold

    2014-07-24

    We prepared 13 derivatives of N-(biphenyl-4'-yl)methyl (R)-2-acetamido-3-methoxypropionamide that differed in type and placement of a R-substituent in the terminal aryl unit. We demonstrated that the R-substituent impacted the compound's whole animal and cellular pharmacological activities. In rodents, select compounds exhibited excellent anticonvulsant activities and protective indices (PI=TD50/ED50) that compared favorably with clinical antiseizure drugs. Compounds with a polar, aprotic R-substituent potently promoted Na+ channel slow inactivation and displayed frequency (use) inhibition of Na+ currents at low micromolar concentrations. The possible advantage of affecting these two pathways to decrease neurological hyperexcitability is discussed.

  11. The alpha2-delta protein: an auxiliary subunit of voltage-dependent calcium channels as a recognized drug target.

    Science.gov (United States)

    Thorpe, Andrew J; Offord, James

    2010-07-01

    Currently, there are two drugs on the market, gabapentin (Neurontin) and pregabalin (Lyrica), that are proposed to exert their therapeutic effect through binding to the alpha2-delta subunit of voltage-sensitive calcium channels. This activity was unexpected, as the alpha2-delta subunit had previously been considered not to be a pharmacological target. In this review, the role of the alpha2-delta subunits is discussed and the mechanism of action of the alpha2-delta ligands in vitro and in vivo is summarized. Finally, new insights into the mechanism of drugs that bind to this protein are discussed.

  12. Glucose decouples intracellular Ca2+ activity from glucagon secretion in mouse pancreatic islet alpha-cells.

    Directory of Open Access Journals (Sweden)

    Sylvain J Le Marchand

    Full Text Available The mechanisms of glucagon secretion and its suppression by glucose are presently unknown. This study investigates the relationship between intracellular calcium levels ([Ca(2+](i and hormone secretion under low and high glucose conditions. We examined the effects of modulating ion channel activities on [Ca(2+](i and hormone secretion from ex vivo mouse pancreatic islets. Glucagon-secreting α-cells were unambiguously identified by cell specific expression of fluorescent proteins. We found that activation of L-type voltage-gated calcium channels is critical for α-cell calcium oscillations and glucagon secretion at low glucose levels. Calcium channel activation depends on K(ATP channel activity but not on tetrodotoxin-sensitive Na(+ channels. The use of glucagon secretagogues reveals a positive correlation between α-cell [Ca(2+](i and secretion at low glucose levels. Glucose elevation suppresses glucagon secretion even after treatment with secretagogues. Importantly, this inhibition is not mediated by K(ATP channel activity or reduction in α-cell [Ca(2+](i. Our results demonstrate that glucose uncouples the positive relationship between [Ca(2+](i and secretory activity. We conclude that glucose suppression of glucagon secretion is not mediated by inactivation of calcium channels, but instead, it requires a calcium-independent inhibitory pathway.

  13. The electrically silent Kv6.4 subunit confers hyperpolarized gating charge movement in Kv2.1/Kv6.4 heterotetrameric channels.

    Directory of Open Access Journals (Sweden)

    Elke Bocksteins

    Full Text Available The voltage-gated K(+ (Kv channel subunit Kv6.4 does not form functional homotetrameric channels but co-assembles with Kv2.1 to form functional Kv2.1/Kv6.4 heterotetrameric channels. Compared to Kv2.1 homotetramers, Kv6.4 exerts a ~40 mV hyperpolarizing shift in the voltage-dependence of Kv2.1/Kv6.4 channel inactivation, without a significant effect on activation gating. However, the underlying mechanism of this Kv6.4-induced modulation of Kv2.1 channel inactivation, and whether the Kv6.4 subunit participates in the voltage-dependent gating of heterotetrameric channels is not well understood. Here we report distinct gating charge movement of Kv2.1/Kv6.4 heterotetrameric channels, compared to Kv2.1 homotetramers, as revealed by gating current recordings from mammalian cells expressing these channels. The gating charge movement of Kv2.1/Kv6.4 heterotetrameric channels displayed an extra component around the physiological K(+ equilibrium potential, characterized by a second sigmoidal relationship of the voltage-dependence of gating charge movement. This distinct gating charge displacement reflects movement of the Kv6.4 voltage-sensing domain and has a voltage-dependency that matches the hyperpolarizing shift in Kv2.1/Kv6.4 channel inactivation. These results provide a mechanistic basis for the modulation of Kv2.1 channel inactivation gating kinetics by silent Kv6.4 subunits.

  14. Coassembly of big conductance Ca2+-activated K+ channels and L-type voltage-gated Ca2+ channels in rat brain

    DEFF Research Database (Denmark)

    Grunnet, Morten; Kaufmann, Walter A

    2004-01-01

    Based on electrophysiological studies, Ca(2+)-activated K(+) channels and voltage-gated Ca(2+) channels appear to be located in close proximity in neurons. Such colocalization would ensure selective and rapid activation of K(+) channels by local increases in the cytosolic calcium concentration...

  15. Calcium Activated K+ Channels in The Electroreceptor of the Skate Confirmed by Cloning. Details of Subunits and Splicing

    Science.gov (United States)

    King, Benjamin L.; Shi, Ling Fang; Kao, Peter; Clusin, William T.

    2015-01-01

    Elasmobranchs detect small potentials using excitable cells of the ampulla of Lorenzini which have calcium-activated K+ channels, first described in l974. A distinctive feature of the outward current in voltage clamped ampullae is its apparent insensitivity to voltage. The sequence of a BK channel α isoform expressed in the ampulla of the skate was characterized. A signal peptide is present at the beginning of the gene. When compared to human isoform 1 (the canonical sequence), the largest difference was absence of a 59 amino acid region from the S8-S9 intracellular linker that contains the strex regulatory domain. The ampulla isoform was also compared with the isoform predicted˜ in late skate embryos where strex was also absent. The BK voltage sensors were conserved in both skate isoforms. Differences between the skate and human BK channel included alternative splicing. Alternative splicing occurs at seven previously defined sites that are characteristic for BK channels in general and hair cells in particular. Skate BK sequences were highly similar to the Australian ghost shark and several other vertebrate species. Based on alignment of known BK sequences with the skate genome and transcriptome, there are at least two isoforms of Kcnma1α expressed in the skate. One of the β subunits (β4), which is known to decrease voltage sensitivity, was also identified in the skate genome and transcriptome and in the ampulla. These studies advance our knowledge of BK channels and suggest further studies in the ampulla and other excitable tissues. PMID:26687710

  16. Establishing homology between mitochondrial calcium uniporters, prokaryotic magnesium channels and chlamydial IncA proteins.

    Science.gov (United States)

    Lee, Andre; Vastermark, Ake; Saier, Milton H

    2014-08-01

    Mitochondrial calcium uniporters (MCUs) (TC no. 1.A.77) are oligomeric channel proteins found in the mitochondrial inner membrane. MCUs have two well-conserved transmembrane segments (TMSs), connected by a linker, similar to bacterial MCU homologues. These proteins and chlamydial IncA proteins (of unknown function; TC no. 9.B.159) are homologous to prokaryotic Mg(2+) transporters, AtpI and AtpZ, based on comparison scores of up to 14.5 sds. A phylogenetic tree containing all of these proteins showed that the AtpZ proteins cluster coherently as a subset within the large and diverse AtpI cluster, which branches separately from the MCUs and IncAs, both of which cluster coherently. The MCUs and AtpZs share the same two TMS topology, but the AtpIs have four TMSs, and IncAs can have either two (most frequent) or four (less frequent) TMSs. Binary alignments, comparison scores and motif analyses showed that TMSs 1 and 2 align with TMSs 3 and 4 of the AtpIs, suggesting that the four TMS AtpI proteins arose via an intragenic duplication event. These findings establish an evolutionary link interconnecting eukaryotic and prokaryotic Ca(2+) and Mg(2+) transporters with chlamydial IncAs, and lead us to suggest that all members of the MCU superfamily, including IncAs, function as divalent cation channels. © 2014 The Authors.

  17. Stretch induced endothelin-1 secretion by adult rat astrocytes involves calcium influx via stretch-activated ion channels (SACs)

    International Nuclear Information System (INIS)

    Ostrow, Lyle W.; Suchyna, Thomas M.; Sachs, Frederick

    2011-01-01

    Highlights: → Endothelin-1 expression by adult rat astrocytes correlates with cell proliferation. → Stretch-induced ET-1 is inhibited by GsMtx-4, a specific inhibitor of Ca 2+ permeant SACs. → The less specific SAC inhibitor streptomycin also inhibits ET-1 secretion. → Stretch-induced ET-1 production depends on a calcium influx. → SAC pharmacology may provide a new class of therapeutic agents for CNS pathology. -- Abstract: The expression of endothelins (ETs) and ET-receptors is often upregulated in brain pathology. ET-1, a potent vasoconstrictor, also inhibits the expression of astrocyte glutamate transporters and is mitogenic for astrocytes, glioma cells, neurons, and brain capillary endothelia. We have previously shown that mechanical stress stimulates ET-1 production by adult rat astrocytes. We now show in adult astrocytes that ET-1 production is driven by calcium influx through stretch-activated ion channels (SACs) and the ET-1 production correlates with cell proliferation. Mechanical stimulation using biaxial stretch ( 2+ threshold. This coupling of mechanical stress to the astrocyte endothelin system through SACs has treatment implications, since all pathology deforms the surrounding parenchyma.

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

    Directory of Open Access Journals (Sweden)

    Amalia M Dolga

    2012-11-01

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

  19. L-type calcium channels play a critical role in maintaining lens transparency by regulating phosphorylation of aquaporin-0 and myosin light chain and expression of connexins.

    Science.gov (United States)

    Maddala, Rupalatha; Nagendran, Tharkika; de Ridder, Gustaaf G; Schey, Kevin L; Rao, Ponugoti Vasantha

    2013-01-01

    Homeostasis of intracellular calcium is crucial for lens cytoarchitecture and transparency, however, the identity of specific channel proteins regulating calcium influx within the lens is not completely understood. Here we examined the expression and distribution profiles of L-type calcium channels (LTCCs) and explored their role in morphological integrity and transparency of the mouse lens, using cDNA microarray, RT-PCR, immunoblot, pharmacological inhibitors and immunofluorescence analyses. The results revealed that Ca (V) 1.2 and 1.3 channels are expressed and distributed in both the epithelium and cortical fiber cells in mouse lens. Inhibition of LTCCs with felodipine or nifedipine induces progressive cortical cataract formation with time, in association with decreased lens weight in ex-vivo mouse lenses. Histological analyses of felodipine treated lenses revealed extensive disorganization and swelling of cortical fiber cells resembling the phenotype reported for altered aquaporin-0 activity without detectable cytotoxic effects. Analysis of both soluble and membrane rich fractions from felodipine treated lenses by SDS-PAGE in conjunction with mass spectrometry and immunoblot analyses revealed decreases in β-B1-crystallin, Hsp-90, spectrin and filensin. Significantly, loss of transparency in the felodipine treated lenses was preceded by an increase in aquaporin-0 serine-235 phosphorylation and levels of connexin-50, together with decreases in myosin light chain phosphorylation and the levels of 14-3-3ε, a phosphoprotein-binding regulatory protein. Felodipine treatment led to a significant increase in gene expression of connexin-50 and 46 in the mouse lens. Additionally, felodipine inhibition of LTCCs in primary cultures of mouse lens epithelial cells resulted in decreased intracellular calcium, and decreased actin stress fibers and myosin light chain phosphorylation, without detectable cytotoxic response. Taken together, these observations reveal a crucial

  20. Inactivation of Caliciviruses

    Directory of Open Access Journals (Sweden)

    Raymond Nims

    2013-03-01

    Full Text Available The Caliciviridae family of viruses contains clinically important human and animal pathogens, as well as vesivirus 2117, a known contaminant of biopharmaceutical manufacturing processes employing Chinese hamster cells. An extensive literature exists for inactivation of various animal caliciviruses, especially feline calicivirus and murine norovirus. The caliciviruses are susceptible to wet heat inactivation at temperatures in excess of 60 °C with contact times of 30 min or greater, to UV-C inactivation at fluence ≥30 mJ/cm2, to high pressure processing >200 MPa for >5 min at 4 °C, and to certain photodynamic inactivation approaches. The enteric caliciviruses (e.g.; noroviruses display resistance to inactivation by low pH, while the non-enteric species (e.g.; feline calicivirus are much more susceptible. The caliciviruses are inactivated by a variety of chemicals, including alcohols, oxidizing agents, aldehydes, and β-propiolactone. As with inactivation of viruses in general, inactivation of caliciviruses by the various approaches may be matrix-, temperature-, and/or contact time-dependent. The susceptibilities of the caliciviruses to the various physical and chemical inactivation approaches are generally similar to those displayed by other small, non-enveloped viruses, with the exception that the parvoviruses and circoviruses may require higher temperatures for inactivation, while these families appear to be more susceptible to UV-C inactivation than are the caliciviruses.

  1. When lithium does not help: the use of anticonvulsants and calcium channel blockers in the treatment of bipolar disorder in the older person.

    Science.gov (United States)

    Masters, J C

    1996-01-01

    Although anticonvulsant agents and calcium channel blockers do not have any clear advantages over lithium, they do offer patients who cannot (or will not) take lithium another treatment option. It is not yet clear from the literature who will respond best to which drug or combination of drugs. The nurse should be supportive to the patients and family, in what may be a drawn out process, to find the best treatment. Optimism is justified because a lack of response to one drug is not indicative of nonresponse to other drugs. It is important to actively treat bipolar disorder because each episode of mania increases the risk of progression of the illness, with increasingly severe episodes occurring closer together. Bipolar disorder has high social costs (legal, financial, and relationship problems) that make improvements in treatment important for the patient and society. Anticonvulsant agents and calcium channel blockers may also be useful in treating depression. The number of people whose depressive symptoms respond is far less (25% to 30%) than the number who respond to the anti-manic effects, but this is an option when antidepressants and electroconvulsive therapy are not effective.

  2. Cortisone Dissociates the Shaker Family K Channels from their Beta Subunit

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Y.; Weng, J; Kabaleeswaran, V; Li, H; Cao, Y; Bholse, R; Zhou, M

    2008-01-01

    The Shaker family voltage-dependent potassium channels (Kv1) are expressed in a wide variety of cells and are 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 (Kv{Beta}s), and certain Kv{Beta}s, for example Kv{Beta}1, have an N-terminal segment that closes the channel by the N-type inactivation mechanism. In principle, dissociation of Kv{Beta}1, although never reported, should eliminate inactivation and thus potentiate Kv1 current. We found that cortisone increases rat Kv1 channel activity by binding to Kv{Beta}1. A crystal structure of the K{Beta}v-cortisone complex was solved to 1.82-{angstrom}resolution and revealed novel cortisone binding sites. Further studies demonstrated that cortisone promotes dissociation of Kv{Beta}. The new mode of channel modulation may be explored by native or synthetic ligands to fine-tune cellular excitability.

  3. Differential Modulation of Rhythmic Brain Activity in Healthy Adults by a T-Type Calcium Channel Blocker: An MEG Study.

    Science.gov (United States)

    Walton, Kerry D; Maillet, Emeline L; Garcia, John; Cardozo, Timothy; Galatzer-Levy, Isaac; Llinás, Rodolfo R

    2017-01-01

    1-octanol is a therapeutic candidate for disorders involving the abnormal activation of the T-type calcium current since it blocks this current specifically. Such disorders include essential tremor and a group of neurological and psychiatric disorders resulting from thalamocortical dysrhythmia (TCD). For example, clinically, the observable phenotype in essential tremor is the tremor itself. The differential diagnostic of TCD is not based only on clinical signs and symptoms. Rather, TCD incorporates an electromagnetic biomarker, the presence of abnormal thalamocortical low frequency brain oscillations. The effect of 1-octanol on brain activity has not been tested. As a preliminary step to such a TCD study, we examined the short-term effects of a single dose of 1-octanol on resting brain activity in 32 healthy adults using magnetoencephalograpy. Visual inspection of baseline power spectra revealed that the subjects fell into those with strong low frequency activity (set 2, n = 11) and those without such activity, but dominated by an alpha peak (set 1, n = 22). Cross-validated linear discriminant analysis, using mean spectral density (MSD) in nine frequency bands as predictors, found overall that 82.5% of the subjects were classified as determined by visual inspection. The effect of 1-octanol on the MSD in narrow frequency bands differed between the two subject groups. In set 1 subjects the MSD increased in the 4.5-6.5Hz and 6.5-8.5 Hz bands. This was consistent with a widening of the alpha peak toward lower frequencies. In the set two subjects the MSD decrease in the 2.5-4.5 Hz and 4.5-6.5 Hz bands. This decreased power is consistent with the blocking effect of 1-octanol on T-type calcium channels. The subjects reported no adverse effects of the 1-octanol. Since stronger low frequency activity is characteristic of patients with TCD, 1-octanol and other T-type calcium channel blockers are good candidates for treatment of this group of disorders following a placebo

  4. Differential Modulation of Rhythmic Brain Activity in Healthy Adults by a T-Type Calcium Channel Blocker: An MEG Study

    Science.gov (United States)

    Walton, Kerry D.; Maillet, Emeline L.; Garcia, John; Cardozo, Timothy; Galatzer-Levy, Isaac; Llinás, Rodolfo R.

    2017-01-01

    1-octanol is a therapeutic candidate for disorders involving the abnormal activation of the T-type calcium current since it blocks this current specifically. Such disorders include essential tremor and a group of neurological and psychiatric disorders resulting from thalamocortical dysrhythmia (TCD). For example, clinically, the observable phenotype in essential tremor is the tremor itself. The differential diagnostic of TCD is not based only on clinical signs and symptoms. Rather, TCD incorporates an electromagnetic biomarker, the presence of abnormal thalamocortical low frequency brain oscillations. The effect of 1-octanol on brain activity has not been tested. As a preliminary step to such a TCD study, we examined the short-term effects of a single dose of 1-octanol on resting brain activity in 32 healthy adults using magnetoencephalograpy. Visual inspection of baseline power spectra revealed that the subjects fell into those with strong low frequency activity (set 2, n = 11) and those without such activity, but dominated by an alpha peak (set 1, n = 22). Cross-validated linear discriminant analysis, using mean spectral density (MSD) in nine frequency bands as predictors, found overall that 82.5% of the subjects were classified as determined by visual inspection. The effect of 1-octanol on the MSD in narrow frequency bands differed between the two subject groups. In set 1 subjects the MSD increased in the 4.5-6.5Hz and 6.5–8.5 Hz bands. This was consistent with a widening of the alpha peak toward lower frequencies. In the set two subjects the MSD decrease in the 2.5–4.5 Hz and 4.5–6.5 Hz bands. This decreased power is consistent with the blocking effect of 1-octanol on T-type calcium channels. The subjects reported no adverse effects of the 1-octanol. Since stronger low frequency activity is characteristic of patients with TCD, 1-octanol and other T-type calcium channel blockers are good candidates for treatment of this group of disorders following a

  5. Delayed rectifier K channels contribute to contrast adaptation in mammalian retinal ganglion cells

    Science.gov (United States)

    Weick, Michael; Demb, Jonathan B.

    2011-01-01

    SUMMARY Retinal ganglion cells adapt by reducing their sensitivity during periods of high contrast. Contrast adaptation in the firing response depends on both presynaptic and intrinsic mechanisms. Here, we investigated intrinsic mechanisms for contrast adaptation in OFF Alpha ganglion cells in the in vitro guinea pig retina. Using either visual stimulation or current injection, we show that brief depolarization evoked spiking and suppressed firing during subsequent depolarization. The suppression could be explained by Na channel inactivation, as shown in salamander cells. However, brief hyperpolarization in the physiological range (5–10 mV) also suppressed firing during subsequent depolarization. This suppression was sensitive selectively to blockers of delayed-rectifier K channels (KDR). Somatic membrane patches showed TEA-sensitive KDR currents with activation near −25 mV and removal of inactivation at voltages negative to Vrest. Brief periods of hyperpolarization apparently remove KDR inactivation and thereby increase the channel pool available to suppress excitability during subsequent depolarization. PMID:21745646

  6. A deleterious gene-by-environment interaction imposed by calcium channel blockers in Marfan syndrome.

    Science.gov (United States)

    Doyle, Jefferson J; Doyle, Alexander J; Wilson, Nicole K; Habashi, Jennifer P; Bedja, Djahida; Whitworth, Ryan E; Lindsay, Mark E; Schoenhoff, Florian; Myers, Loretha; Huso, Nick; Bachir, Suha; Squires, Oliver; Rusholme, Benjamin; Ehsan, Hamid; Huso, David; Thomas, Craig J; Caulfield, Mark J; Van Eyk, Jennifer E; Judge, Daniel P; Dietz, Harry C

    2015-10-27

    Calcium channel blockers (CCBs) are prescribed to patients with Marfan syndrome for prophylaxis against aortic aneurysm progression, despite limited evidence for their efficacy and safety in the disorder. Unexpectedly, Marfan mice treated with CCBs show accelerated aneurysm expansion, rupture, and premature lethality. This effect is both extracellular signal-regulated kinase (ERK1/2) dependent and angiotensin-II type 1 receptor (AT1R) dependent. We have identified protein kinase C beta (PKCβ) as a critical mediator of this pathway and demonstrate that the PKCβ inhibitor enzastaurin, and the clinically available anti-hypertensive agent hydralazine, both normalize aortic growth in Marfan mice, in association with reduced PKCβ and ERK1/2 activation. Furthermore, patients with Marfan syndrome and other forms of inherited thoracic aortic aneurysm taking CCBs display increased risk of aortic dissection and need for aortic surgery, compared to patients on other antihypertensive agents.

  7. Physiological studies in heterozygous calcium sensing receptor (CaSR gene-ablated mice confirm that the CaSR regulates calcitonin release in vivo

    Directory of Open Access Journals (Sweden)

    Kovacs Christopher S

    2004-04-01

    Full Text Available Abstract Background The calcium sensing receptor (CaSR regulates serum calcium by suppressing secretion of parathyroid hormone; it also regulates renal tubular calcium excretion. Inactivating mutations of CaSR raise serum calcium and reduce urine calcium excretion. Thyroid C-cells (which make calcitonin express CaSR and may, therefore, be regulated by it. Since calcium stimulates release of calcitonin, the higher blood calcium caused by inactivation of CaSR should increase serum calcitonin, unless CaSR mutations alter the responsiveness of calcitonin to calcium. To demonstrate regulatory effects of CaSR on calcitonin release, we studied calcitonin responsiveness to calcium in normal and CaSR heterozygous-ablated (Casr+/- mice. Casr+/- mice have hypercalcemia and hypocalciuria, and live normal life spans. Each mouse received either 500 μl of normal saline or one of two doses of elemental calcium (500 μmol/kg or 5 mmol/kg by intraperitoneal injection. Ionized calcium was measured at baseline and 10 minutes, and serum calcitonin was measured on the 10 minute sample. Results At baseline, Casr+/- mice had a higher blood calcium, and in response to the two doses of elemental calcium, had greater increments and peak levels of ionized calcium than their wild type littermates. Despite significantly higher ionized calcium levels, the calcitonin levels of Casr+/- mice were consistently lower than wild type at any ionized calcium level, indicating that the dose-response curve of calcitonin to increases in ionized calcium had been significantly blunted or shifted to the right in Casr+/- mice. Conclusions These results confirm that the CaSR is a physiological regulator of calcitonin; therefore, in response to increases in ionized calcium, the CaSR inhibits parathyroid hormone secretion and stimulates calcitonin secretion.

  8. A negative charge in transmembrane segment 1 of domain II of the cockroach sodium channel is critical for channel gating and action of pyrethroid insecticides

    International Nuclear Information System (INIS)

    Du Yuzhe; Song Weizhong; Groome, James R.; Nomura, Yoshiko; Luo Ningguang; Dong Ke

    2010-01-01

    Voltage-gated sodium channels are the primary target of pyrethroids, an important class of synthetic insecticides. Pyrethroids bind to a distinct receptor site on sodium channels and prolong the open state by inhibiting channel deactivation and inactivation. Recent studies have begun to reveal sodium channel residues important for pyrethroid binding. However, how pyrethroid binding leads to inhibition of sodium channel deactivation and inactivation remains elusive. In this study, we show that a negatively charged aspartic acid residue at position 802 (D802) located in the extracellular end of transmembrane segment 1 of domain II (IIS1) is critical for both the action of pyrethroids and the voltage dependence of channel activation. Charge-reversing or -neutralizing substitutions (K, G, or A) of D802 shifted the voltage dependence of activation in the depolarizing direction and reduced channel sensitivity to deltamethrin, a pyrethroid insecticide. The charge-reversing mutation D802K also accelerated open-state deactivation, which may have counteracted the inhibition of sodium channel deactivation by deltamethrin. In contrast, the D802G substitution slowed open-state deactivation, suggesting an additional mechanism for neutralizing the action of deltamethrin. Importantly, Schild analysis showed that D802 is not involved in pyrethroid binding. Thus, we have identified a sodium channel residue that is critical for regulating the action of pyrethroids on the sodium channel without affecting the receptor site of pyrethroids.

  9. A molecular switch between the outer and the inner vestibule of the voltage-gated Na+ channel

    International Nuclear Information System (INIS)

    Zarrabi, T.

    2010-01-01

    Na+ channels permit rapid transmission of depolarizing impulses throughout cells and cell networks, and are essential to the proper function of skeletal muscle, the heart and the nervous system. The selectivity filter of the channel is considered to be formed by the amino acids D400, E755, K1237, and A1529 ('DEKA' motif) which are located at the innermost turn of the P-loops connecting S5 and S6 segments of each domain. The inner vestibule is believed to be lined by four S6 helices, one from each domain. Comparison of crystal structures of K+ channels in open and closed states as well as electron paramagnetic resonance spectroscopic studies suggest that the activation gate of voltage-gated ion channels is located at the inner part of the S6 segments. This may also hold true for voltage-gated Na+ channels because mutations in S6 segments alter activation gating. The gate for fast inactivation of the channel has been mapped to the intracellular linker between domains III and IV. This intracellular loop is currently considered to produce channel inactivation by transiently occluding the intracellular vestibule of the channel. The time constants of entry into and recovery from fast inactivation are on the order of milliseconds. Apart from 'fast inactivation' a number of slower inactivated states have been described. During very long depolarizations, on the order of several minutes, rNaV1.4 channels enter a very stable inactivated state which we refer to as 'ultra-slow' inactivation (IUS). In these channels the likelihood of entry into IUS is substantially increased by a mutation in the selectivity filter, K1237E. IUS can be modulated by molecules binding to the outer vestibule, suggesting that a conformational change of the outer vestibule gives rise to this kinetic state. On the other hand, the local anesthetic drug lidocaine, which binds to the internal part of the channel pore, inhibits entry into IUS by a 'foot-in-the-door' mechanism indicating that a

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

    International Nuclear Information System (INIS)

    Long, Yan; Lin, Zuoxian; Xia, Menghang; Zheng, Wei; Li, Zhiyuan

    2013-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-03-01

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

  12. [Ion channels that are sensitive to the extracellular concentration of protons: their structure, function, pharmacology and pathophysiology].

    Science.gov (United States)

    Mercado, F; Vega, R; Soto, E

    Acid sensing ion channels (ASIC) members of the ENaC degenerine channel family, have been shown to participate in various sensorial pathways including nociception, also they have been shown to participate in synaptic transmission, learning and memory processes and in the physiopathology of the ischemic stroke. The proton concentration in the organism is strictly regulated by distinct buffer systems. Drastic changes of pH are generated only by pathological conditions as is the ischemia; however, some physiological processes may produce local changes in the extracellular pH. Recently, a new family of proton receptors known as ASIC has been cloned. These are ionic channels inactivated at physiological pH (7.4) and activated with a pH fall (increase in H+ concentration). ASICs are permeable to sodium ions and in a lesser degree to calcium ions, activation of these channels leads to an increase in cell excitability. The ASICs are distributed widely in the central and peripheral nervous system, and in specialized epithelia. In the past few years they have become a focus of interest due to its role in nociception, taste perception, long term potentation and the physiopathology of ischemic stroke. In this review we address the most relevant molecular, physiological and pharmacological aspects of the ASICs, its participation in some pathological process, and the perspectives of basic and clinic investigation in this arising research field.

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

    Science.gov (United States)

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

    2017-08-01

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

  14. 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.

  15. Charge immobilization of the voltage sensor in domain IV is independent of sodium current inactivation.

    Science.gov (United States)

    Sheets, Michael F; Hanck, Dorothy A

    2005-02-15

    Recovery from fast inactivation in voltage-dependent Na+ channels is associated with a slow component in the time course of gating charge during repolarization (i.e. charge immobilization), which results from the slow movement of the S4 segments in domains III and IV (S4-DIII and S4-DIV). Previous studies have shown that the non-specific removal of fast inactivation by the proteolytic enzyme pronase eliminated charge immobilization, while the specific removal of fast inactivation (by intracellular MTSET modification of a cysteine substituted for the phenylalanine in the IFM motif, ICMMTSET, in the inactivation particle formed by the linker between domains III and IV) only reduced the amount of charge immobilization by nearly one-half. To investigate the molecular origin of the remaining slow component of charge immobilization we studied the human cardiac Na+ channel (hH1a) in which the outermost arginine in the S4-DIV, which contributes approximately 20% to total gating charge (Qmax), was mutated to a cysteine (R1C-DIV). Gating charge could be fully restored in R1C-DIV by exposure to extracellular MTSEA, a positively charged methanethiosulphonate reagent. The RIC-DIV mutation was combined with ICMMTSET to remove fast inactivation, and the gating currents of R1C-DIV-ICM(MTSET) were recorded before and after modification with MTSEAo. Prior to MTSEAo, the time course of the gating charge during repolarization (off-charge) was best described by a single fast time constant. After MTSEA, the off-charge had both fast and slow components, with the slow component accounting for nearly 35% of Qmax. These results demonstrate that the slow movement of the S4-DIV during repolarization is not dependent upon the normal binding of the inactivation particle.

  16. Activation of TRPV1-dependent calcium oscillation exacerbates seawater inhalation-induced acute lung injury.

    Science.gov (United States)

    Li, Congcong; Bo, Liyan; Liu, Qingqing; Liu, Wei; Chen, Xiangjun; Xu, Dunquan; Jin, Faguang

    2016-03-01

    Calcium is an important second messenger and it is widely recognized that acute lung injury (ALI) is often caused by oscillations of cytosolic free Ca2+. Previous studies have indicated that the activation of transient receptor potential‑vanilloid (TRPV) channels and subsequent Ca2+ entry initiates an acute calcium‑dependent permeability increase during ALI. However, whether seawater exposure induces such an effect through the activation of TRPV channels remains unknown. In the current study, the effect of calcium, a component of seawater, on the inflammatory reactions that occur during seawater drowning‑induced ALI, was examined. The results demonstrated that a high concentration of calcium ions in seawater increased lung tissue myeloperoxidase activity and the secretion of inflammatory mediators, such as tumor necrosis factor‑α (TNF‑α) and interleukin (IL)‑1β and IL‑6. Further study demonstrated that the seawater challenge elevated cytosolic Ca2+ concentration, indicated by [Ca2+]c, by inducing calcium influx from the extracellular medium via TRPV1 channels. The elevated [Ca2+c] may have resulted in the increased release of TNF‑α and IL‑1β via increased phosphorylation of nuclear factor‑κB (NF‑κB). It was concluded that a high concentration of calcium in seawater exacerbated lung injury, and TRPV1 channels were notable mediators of the calcium increase initiated by the seawater challenge. Calcium influx through TRPV1 may have led to greater phosphorylation of NF‑κB and increased release of TNF‑α and IL‑1β.

  17. Properties of an intermediate-duration inactivation process of the voltage-gated sodium conductance in rat hippocampal CA1 neurons.

    Science.gov (United States)

    French, Christopher R; Zeng, Zhen; Williams, David A; Hill-Yardin, Elisa L; O'Brien, Terence J

    2016-02-01

    Rapid transmembrane flow of sodium ions produces the depolarizing phase of action potentials (APs) in most excitable tissue through voltage-gated sodium channels (NaV). Macroscopic currents display rapid activation followed by fast inactivation (IF) within milliseconds. Slow inactivation (IS) has been subsequently observed in several preparations including neuronal tissues. IS serves important physiological functions, but the kinetic properties are incompletely characterized, especially the operative timescales. Here we present evidence for an "intermediate inactivation" (II) process in rat hippocampal CA1 neurons with time constants of the order of 100 ms. The half-inactivation potentials (V0.5) of steady-state inactivation curves were hyperpolarized by increasing conditioning pulse duration from 50 to 500 ms and could be described by a sum of Boltzmann relations. II state transitions were observed after opening as well as subthreshold potentials. Entry into II after opening was relatively insensitive to membrane potential, and recovery of II became more rapid at hyperpolarized potentials. Removal of fast inactivation with cytoplasmic papaine revealed time constants of INa decay corresponding to II and IS with long depolarizations. Dynamic clamp revealed attenuation of trains of APs over the 10(2)-ms timescale, suggesting a functional role of II in repetitive firing accommodation. These experimental findings could be reproduced with a five-state Markov model. It is likely that II affects important aspects of hippocampal neuron response and may provide a drug target for sodium channel modulation. Copyright © 2016 the American Physiological Society.

  18. Calcium activated K⁺ channels in the electroreceptor of the skate confirmed by cloning. Details of subunits and splicing.

    Science.gov (United States)

    King, Benjamin L; Shi, Ling Fang; Kao, Peter; Clusin, William T

    2016-03-01

    Elasmobranchs detect small potentials using excitable cells of the ampulla of Lorenzini which have calcium-activated K(+) channels, first described in 1974. A distinctive feature of the outward current in voltage clamped ampullae is its apparent insensitivity to voltage. The sequence of a BK channel α isoform expressed in the ampulla of the skate was characterized. A signal peptide is present at the beginning of the gene. When compared to human isoform 1 (the canonical sequence), the largest difference was absence of a 59 amino acid region from the S8-S9 intra-cellular linker that contains the strex regulatory domain. The ampulla isoform was also compared with the isoform predicted in late skate embryos where strex was also absent. The BK voltage sensors were conserved in both skate isoforms. Differences between the skate and human BK channel included alternative splicing. Alternative splicing occurs at seven previously defined sites that are characteristic for BK channels in general and hair cells in particular. Skate BK sequences were highly similar to the Australian ghost shark and several other vertebrate species. Based on alignment of known BK sequences with the skate genome and transcriptome, there are at least two isoforms of Kcnma1α expressed in the skate. One of the β subunits (β4), which is known to decrease voltage sensitivity, was also identified in the skate genome and transcriptome and in the ampulla. These studies advance our knowledge of BK channels and suggest further studies in the ampulla and other excitable tissues. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Inhibition of large conductance calcium-dependent potassium ...

    African Journals Online (AJOL)

    conductance, calcium and voltage- dependent potassium (BKCa) channels thereby promoting vasoconstriction. Our results show that the Rho-kinase inhibitor, Y-27632, induced concentration-dependent relaxation in rat mesenteric artery.

  20. NMDA receptors in mouse anterior piriform cortex initialize early odor preference learning and L-type calcium channels engage for long-term memory.

    Science.gov (United States)

    Mukherjee, Bandhan; Yuan, Qi

    2016-10-14

    The interactions of L-type calcium channels (LTCCs) and NMDA receptors (NMDARs) in memories are poorly understood. Here we investigated the specific roles of anterior piriform cortex (aPC) LTCCs and NMDARs in early odor preference memory in mice. Using calcium imaging in aPC slices, LTCC activation was shown to be dependent on NMDAR activation. Either D-APV (NMDAR antagonist) or nifedipine (LTCC antagonist) reduced somatic calcium transients in pyramidal cells evoked by lateral olfactory tract stimulation. However, nifedipine did not further reduce calcium in the presence of D-APV. In mice that underwent early odor preference training, blocking NMDARs in the aPC prevented short-term (3 hr) and long-term (24 hr) odor preference memory, and both memories were rescued when BayK-8644 (LTCC agonist) was co-infused. However, activating LTCCs in the absence of NMDARs resulted in loss of discrimination between the conditioned odor and a similar odor mixture at 3 hr. Elevated synaptic AMPAR expression at 3 hr was prevented by D-APV infusion but restored when LTCCs were directly activated, mirroring the behavioral outcomes. Blocking LTCCs prevented 24 hr memory and spared 3 hr memory. These results suggest that NMDARs mediate stimulus-specific encoding of odor memory while LTCCs mediate intracellular signaling leading to long-term memory.

  1. Differential effects of ciguatoxin and maitotoxin in primary cultures of cortical neurons.

    Science.gov (United States)

    Martin, Victor; Vale, Carmen; Antelo, Alvaro; Hirama, Masahiro; Yamashita, Shuji; Vieytes, Mercedes R; Botana, Luis M

    2014-08-18

    Ciguatoxins (CTXs) and maitotoxins (MTXs) are polyether ladder shaped toxins derived from the dinoflagellate Gambierdiscus toxicus. Despite the fact that MTXs are 3 times larger than CTXs, part of the structure of MTXs resembles that of CTXs. To date, the synthetic ciguatoxin, CTX 3C has been reported to activate voltage-gated sodium channels, whereas the main effect of MTX is inducing calcium influx into the cell leading to cell death. However, there is a lack of information regarding the effects of these toxins in a common cellular model. Here, in order to have an overview of the main effects of these toxins in mice cortical neurons, we examined the effects of MTX and the synthetic ciguatoxin CTX 3C on the main voltage dependent ion channels in neurons, sodium, potassium, and calcium channels as well as on membrane potential, cytosolic calcium concentration ([Ca(2+)]c), intracellular pH (pHi), and neuronal viability. Regarding voltage-gated ion channels, neither CTX 3C nor MTX affected voltage-gated calcium or potassium channels, but while CTX 3C had a large effect on voltage-gated sodium channels (VGSC) by shifting the activation and inactivation curves to more hyperpolarized potentials and decreasing peak sodium channel amplitude, MTX, at 5 nM, had no effect on VGSC activation and inactivation but decreased peak sodium current amplitude. Other major differences between both toxins were the massive calcium influx and intracellular acidification produced by MTX but not by CTX 3C. Indeed, the novel finding that MTX produces acidosis supports a pathway recently described in which MTX produces calcium influx via the sodium-hydrogen exchanger (NHX). For the first time, we found that VGSC blockers partially blocked the MTX-induced calcium influx, intracellular acidification, and protected against the short-term MTX-induced cytotoxicity. The results presented here provide the first report that shows the comparative effects of two prototypical ciguatera toxins, CTX 3C

  2. Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

    Directory of Open Access Journals (Sweden)

    Piotr Koprowski

    Full Text Available Bacterial mechano-sensitive (MS channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

  3. Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

    Science.gov (United States)

    Koprowski, Piotr; Grajkowski, Wojciech; Balcerzak, Marcin; Filipiuk, Iwona; Fabczak, Hanna; Kubalski, Andrzej

    2015-01-01

    Bacterial mechano-sensitive (MS) channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS) family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

  4. Delayed-rectifier K channels contribute to contrast adaptation in mammalian retinal ganglion cells.

    Science.gov (United States)

    Weick, Michael; Demb, Jonathan B

    2011-07-14

    Retinal ganglion cells adapt by reducing their sensitivity during periods of high contrast. Contrast adaptation in the firing response depends on both presynaptic and intrinsic mechanisms. Here, we investigated intrinsic mechanisms for contrast adaptation in OFF Alpha ganglion cells in the in vitro guinea pig retina. Using either visual stimulation or current injection, we show that brief depolarization evoked spiking and suppressed firing during subsequent depolarization. The suppression could be explained by Na channel inactivation, as shown in salamander cells. However, brief hyperpolarization in the physiological range (5-10 mV) also suppressed firing during subsequent depolarization. This suppression was selectively sensitive to blockers of delayed-rectifier K channels (K(DR)). In somatic membrane patches, we observed tetraethylammonium-sensitive K(DR) currents that activated near -25 mV. Recovery from inactivation occurred at potentials hyperpolarized to V(rest). Brief periods of hyperpolarization apparently remove K(DR) inactivation and thereby increase the channel pool available to suppress excitability during subsequent depolarization. Copyright © 2011 Elsevier Inc. All rights reserved.

  5. A New Negative Allosteric Modulator AP14145 for the Study of Small Conductance Calcium-Activated Potassium Channels

    DEFF Research Database (Denmark)

    Simo Vicens, Rafel; Kirchhoff, Jeppe Egedal; Dolce, Bernardo

    2017-01-01

    ) prolongation in anaesthetised rats and a beam walk test was performed in mice to determine acute CNS related effects of the drug. Key results: AP14145 was found to be an equipotent negative allosteric modulator of KCa2.2 and KCa2.3 channels (IC50 = 1.1 ± 0.3 μM L-1). The presence of AP14145 (10 μM L-1......) increased the EC50 of Ca2+ on KCa2.3 from 0.36 ± 0.02 μM L-1 to 1.2 ± 0.1 μM L-1. The inhibitory effect strongly depended on two amino acids, S508 and A533. AP14145 concentration-dependently prolonged AERP in rats. Moreover, AP14145 (10 mg kg-1) did not trigger any apparent CNS effects in mice. Conclusion...... and implications: AP14145 is a negative allosteric modulator of KCa2.2 and KCa2.3 that shifts the calcium dependence of channel activation, an effect strongly dependent on two identified amino acids. AP14145 prolongs AERP in rats and does not trigger any acute CNS effects in mice. The understanding of how KCa2...

  6. The human TRPV6 channel protein is associated with cyclophilin B in human placenta.

    Science.gov (United States)

    Stumpf, Tobias; Zhang, Qi; Hirnet, Daniela; Lewandrowski, Urs; Sickmann, Albert; Wissenbach, Ulrich; Dörr, Janka; Lohr, Christian; Deitmer, Joachim W; Fecher-Trost, Claudia

    2008-06-27

    Transcellular calcium transport in the kidney, pancreas, small intestine, and placenta is partly mediated by transient receptor potential (TRP) channels. The highly selective TRPV6 calcium channel protein is most likely important for the calcium transfer in different specialized epithelial cells. In the human placenta the protein is expressed in trophoblast tissue, where it is implicated in the transepithelial calcium transfer from mother to the fetus. We enriched the TRPV6 channel protein endogenously expressed in placenta together with annexin A2 and cyclophilin B (CypB), which is a member of the huge immunophilin family. In the human placenta TRPV6 and CypB are mainly located intracellularly in the syncytiotrophoblast layer, but a small amount of the mature glycosylated TRPV6 channel protein and CypB is also expressed in microvilli apical membranes, the fetomaternal barrier. To understand the role of CypB on the TRPV6 channel function, we evaluated the effect of CypB co-expression on TRPV6-mediated calcium uptake into Xenopus laevis oocytes expressing TRPV6. A significant increase of TRPV6-mediated calcium uptake was observed after CypB/TRPV6 co-expression. This stimulatory effect of CypB was reversed by the immunosuppressive drug cyclosporin A, which inhibits the enzymatic activity of CypB. Cyclosporin A had no significant effect on TRPV6 and CypB protein expression levels in the oocytes. In summary, our results establish CypB as a new TRPV6 accessory protein with potential involvement in TRPV6 channel activation through its peptidyl-prolyl cis/trans isomerase activity.

  7. Hyperthyroidism enhances 5-HT-induced contraction of the rat pulmonary artery: role of calcium-activated chloride channel activation.

    Science.gov (United States)

    Oriowo, Mabayoje A; Oommen, Elsie; Khan, Islam

    2011-11-01

    Experimentally-induced hyperthyroidism in rodents is associated with signs and symptoms of pulmonary hypertension. The main objective of the present study was to investigate the effect of thyroxine-induced pulmonary hypertension on the contractile response of the pulmonary artery to 5-HT and the possible underlying signaling pathway. 5-HT concentration-dependently contracted artery segments from control and thyroxine-treated rats with pD(2) values of 5.04 ± 0.19 and 5.34 ± 0.14, respectively. The maximum response was significantly greater in artery segments from thyroxine-treated rats. Neither BW 723C86 (5-HT(2B)-receptor agonist) nor CP 93129 (5-HT(1B)-receptor agonist) contracted ring segments of the pulmonary artery from control and thyroxine-treated rats at concentrations up to 10(-4)M. There was no significant difference in the level of expression of 5-HT(2A)-receptor protein between the two groups. Ketanserin (3 × 10(-8)M) produced a rightward shift of the concentration-response curve to 5-HT in both groups with equal potency (-logK(B) values were 8.1 ± 0.2 and 7.9 ± 0.1 in control and thyroxine-treated rats, respectively). Nifedipine (10(-6)M) inhibited 5-HT-induced contractions in artery segments from control and thyroxine-treated rats and was more effective against 5-HT-induced contraction in artery segments for thyroxine-treated rats. The calcium-activated chloride channel blocker, niflumic acid (10(-4)M) also inhibited 5-HT-induced contractions in artery segments from control and thyroxine-treated rats and was more effective against 5-HT-induced contraction in artery segments for thyroxine-treated rats. It was concluded that hyperthyroidism enhanced 5-HT-induced contractions of the rat pulmonary artery by a mechanism involving increased activity of calcium-activated chloride channels. Copyright © 2011 Elsevier B.V. All rights reserved.

  8. A voltage-gated calcium channel regulates lysosomal fusion with endosomes and autophagosomes and is required for neuronal homeostasis.

    Directory of Open Access Journals (Sweden)

    Xuejun Tian

    2015-03-01

    Full Text Available Autophagy helps deliver sequestered intracellular cargo to lysosomes for proteolytic degradation and thereby maintains cellular homeostasis by preventing accumulation of toxic substances in cells. In a forward mosaic screen in Drosophila designed to identify genes required for neuronal function and maintenance, we identified multiple cacophony (cac mutant alleles. They exhibit an age-dependent accumulation of autophagic vacuoles (AVs in photoreceptor terminals and eventually a degeneration of the terminals and surrounding glia. cac encodes an α1 subunit of a Drosophila voltage-gated calcium channel (VGCC that is required for synaptic vesicle fusion with the plasma membrane and neurotransmitter release. Here, we show that cac mutant photoreceptor terminals accumulate AV-lysosomal fusion intermediates, suggesting that Cac is necessary for the fusion of AVs with lysosomes, a poorly defined process. Loss of another subunit of the VGCC, α2δ or straightjacket (stj, causes phenotypes very similar to those caused by the loss of cac, indicating that the VGCC is required for AV-lysosomal fusion. The role of VGCC in AV-lysosomal fusion is evolutionarily conserved, as the loss of the mouse homologues, Cacna1a and Cacna2d2, also leads to autophagic defects in mice. Moreover, we find that CACNA1A is localized to the lysosomes and that loss of lysosomal Cacna1a in cerebellar cultured neurons leads to a failure of lysosomes to fuse with endosomes and autophagosomes. Finally, we show that the lysosomal CACNA1A but not the plasma-membrane resident CACNA1A is required for lysosomal fusion. In summary, we present a model in which the VGCC plays a role in autophagy by regulating the fusion of AVs with lysosomes through its calcium channel activity and hence functions in maintaining neuronal homeostasis.

  9. Autosomal dominant hypercalciuria in a mouse model due to a mutation of the epithelial calcium channel, TRPV5.

    Directory of Open Access Journals (Sweden)

    Nellie Y Loh

    Full Text Available Hypercalciuria is a major cause of nephrolithiasis, and is a common and complex disorder involving genetic and environmental factors. Identification of genetic factors for monogenic forms of hypercalciuria is hampered by the limited availability of large families, and to facilitate such studies, we screened for hypercalciuria in mice from an N-ethyl-N-nitrosourea mutagenesis programme. We identified a mouse with autosomal dominant hypercalciuria (HCALC1. Linkage studies mapped the Hcalc1 locus to a 11.94 Mb region on chromosome 6 containing the transient receptor potential cation channel, subfamily V, members 5 (Trpv5 and 6 (Trpv6 genes. DNA sequence analysis of coding regions, intron-exon boundaries and promoters of Trpv5 and Trpv6 identified a novel T to C transition in codon 682 of TRPV5, mutating a conserved serine to a proline (S682P. Compared to wild-type littermates, heterozygous (Trpv5(682P/+ and homozygous (Trpv5(682P/682P mutant mice had hypercalciuria, polyuria, hyperphosphaturia and a more acidic urine, and ∼10% of males developed tubulointerstitial nephritis. Trpv5(682P/682P mice also had normal plasma parathyroid hormone but increased 1,25-dihydroxyvitamin D(3 concentrations without increased bone resorption, consistent with a renal defect for the hypercalciuria. Expression of the S682P mutation in human embryonic kidney cells revealed that TRPV5-S682P-expressing cells had a lower baseline intracellular calcium concentration than wild-type TRPV5-expressing cells, suggesting an altered calcium permeability. Immunohistological studies revealed a selective decrease in TRPV5-expression from the renal distal convoluted tubules of Trpv5(682P/+ and Trpv5(682P/682P mice consistent with a trafficking defect. In addition, Trpv5(682P/682P mice had a reduction in renal expression of the intracellular calcium-binding protein, calbindin-D(28K, consistent with a specific defect in TRPV5-mediated renal calcium reabsorption. Thus, our findings

  10. Preventing effect of L-type calcium channel blockade on electrophysiological alterations in dentate gyrus granule cells induced by entorhinal amyloid pathology.

    Directory of Open Access Journals (Sweden)

    Hamid Gholami Pourbadie

    Full Text Available The entorhinal cortex (EC is one of the earliest affected brain regions in Alzheimer's disease (AD. EC-amyloid pathology induces synaptic failure in the dentate gyrus (DG with resultant behavioral impairment, but there is little known about its impact on neuronal properties in the DG. It is believed that calcium dyshomeostasis plays a pivotal role in the etiology of AD. Here, the effect of the EC amyloid pathogenesis on cellular properties of DG granule cells and also possible neuroprotective role of L-type calcium channel blockers (CCBs, nimodipine and isradipine, were investigated. The amyloid beta (Aβ 1-42 was injected bilaterally into the EC of male rats and one week later, electrophysiological properties of DG granule cells were assessed. Voltage clamp recording revealed appearance of giant sIPSC in combination with a decrease in sEPSC frequency which was partially reversed by CCBs in granule cells from Aβ treated rats. EC amyloid pathogenesis induced a significant reduction of input resistance (Rin accompanied by a profound decreased excitability in the DG granule cells. However, daily administration of CCBs, isradipine or nimodipine (i.c.v. for 6 days, almost preserved the normal excitability against Aβ. In conclusion, lower tendency to fire AP along with reduced Rin suggest that DG granule cells might undergo an alteration in the membrane ion channel activities which finally lead to the behavioral deficits observed in animal models and patients with early-stage Alzheimer's disease.

  11. Escitalopram block of hERG potassium channels.

    Science.gov (United States)

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

    2014-01-01

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

  12. Anion-sensitive regions of L-type CaV1.2 calcium channels expressed in HEK293 cells.

    Directory of Open Access Journals (Sweden)

    Norbert Babai

    2010-01-01

    Full Text Available L-type calcium currents (I(Ca are influenced by changes in extracellular chloride, but sites of anion effects have not been identified. Our experiments showed that CaV1.2 currents expressed in HEK293 cells are strongly inhibited by replacing extracellular chloride with gluconate or perchlorate. Variance-mean analysis of I(Ca and cell-attached patch single channel recordings indicate that gluconate-induced inhibition is due to intracellular anion effects on Ca(2+ channel open probability, not conductance. Inhibition of CaV1.2 currents produced by replacing chloride with gluconate was reduced from approximately 75%-80% to approximately 50% by omitting beta subunits but unaffected by omitting alpha(2delta subunits. Similarly, gluconate inhibition was reduced to approximately 50% by deleting an alpha1 subunit N-terminal region of 15 residues critical for beta subunit interactions regulating open probability. Omitting beta subunits with this mutant alpha1 subunit did not further diminish inhibition. Gluconate inhibition was unchanged with expression of different beta subunits. Truncating the C terminus at AA1665 reduced gluconate inhibition from approximately 75%-80% to approximately 50% whereas truncating it at AA1700 had no effect. Neutralizing arginines at AA1696 and 1697 by replacement with glutamines reduced gluconate inhibition to approximately 60% indicating these residues are particularly important for anion effects. Expressing CaV1.2 channels that lacked both N and C termini reduced gluconate inhibition to approximately 25% consistent with additive interactions between the two tail regions. Our results suggest that modest changes in intracellular anion concentration can produce significant effects on CaV1.2 currents mediated by changes in channel open probability involving beta subunit interactions with the N terminus and a short C terminal region.

  13. Molecular dynamics of CYP2D6 polymorphisms in the absence and presence of a mechanism-based inactivator reveals changes in local flexibility and dominant substrate access channels.

    Directory of Open Access Journals (Sweden)

    Parker W de Waal

    Full Text Available Cytochrome P450 enzymes (CYPs represent an important enzyme superfamily involved in metabolism of many endogenous and exogenous small molecules. CYP2D6 is responsible for ∼ 15% of CYP-mediated drug metabolism and exhibits large phenotypic diversity within CYPs with over 100 different allelic variants. Many of these variants lead to functional changes in enzyme activity and substrate selectivity. Herein, a molecular dynamics comparative analysis of four different variants of CYP2D6 was performed. The comparative analysis included simulations with and without SCH 66712, a ligand that is also a mechanism-based inactivator, in order to investigate the possible structural basis of CYP2D6 inactivation. Analysis of protein stability highlighted significantly altered flexibility in both proximal and distal residues from the variant residues. In the absence of SCH 66712, *34, *17-2, and *17-3 displayed more flexibility than *1, and *53 displayed more rigidity. SCH 66712 binding reversed flexibility in *17-2 and *17-3, through *53 remained largely rigid. Throughout simulations with docked SCH 66712, ligand orientation within the heme-binding pocket was consistent with previously identified sites of metabolism and measured binding energies. Subsequent tunnel analysis of substrate access, egress, and solvent channels displayed varied bottle-neck radii. Taken together, our results indicate that SCH 66712 should inactivate these allelic variants, although varied flexibility and substrate binding-pocket accessibility may alter its interaction abilities.

  14. L-type calcium channel CaV 1.2 in transgenic mice overexpressing human AbetaPP751 with the London (V717I) and Swedish (K670M/N671L) mutations.

    Science.gov (United States)

    Willis, Michael; Kaufmann, Walter A; Wietzorrek, Georg; Hutter-Paier, Birgit; Moosmang, Sven; Humpel, Christian; Hofmann, Franz; Windisch, Manfred; Knaus, Hans-Günther; Marksteiner, Josef

    2010-01-01

    Cumulative evidence indicates that amyloid-beta peptides exert some of their neurodegenerative effects through modulation of L-type voltage gated calcium channels, which play key roles in a diverse range of CNS functions. In this study we examined the expression of CaV1.2 L-type voltage gated calcium channels in transgenic mice overexpressing human AbetaPP751 with the London (V717I) and Swedish (K670M/N671L) mutations by immunohistochemistry in light and electron microscopy. In hippocampal layers of wild type and transgenic mice, CaV1.2 channels were predominantly localized to somato-dendritic domains of neurons, and to astrocytic profiles with an age-dependent increase in labeling density. In transgenic animals, CaV1.2-like immunoreactive clusters were found in neuronal profiles in association with amyloid-beta plaques. Both the number and density of these clusters depended upon age of animals and number of plaques. The most striking difference between wild type and transgenic mice was the age-dependent expression of CaV1.2 channels in reactive astrocytes. At the age of 6 month, CaV1.2 channels were rarely detected in reactive astrocytes of transgenic mice, but an incremental number of CaV1.2 expressing reactive astrocytes was found with increasing age of animals and number of amyloid-beta plaques. This study demonstrates that CaV1.2 channels are highly expressed in reactive astrocytes of 12-months of age transgenic mice, which might be a consequence of the increasing amyloid burden. Further studies should clarify which functional implications are associated with the higher availability of CaV1.2 channels in late stage Alzheimer's disease.

  15. Transmitter modulation of spike-evoked calcium transients in arousal related neurons

    DEFF Research Database (Denmark)

    Kohlmeier, Kristi Anne; Leonard, Christopher S

    2006-01-01

    Nitric oxide synthase (NOS)-containing cholinergic neurons in the laterodorsal tegmentum (LDT) influence behavioral and motivational states through their projections to the thalamus, ventral tegmental area and a brainstem 'rapid eye movement (REM)-induction' site. Action potential-evoked intracel......Nitric oxide synthase (NOS)-containing cholinergic neurons in the laterodorsal tegmentum (LDT) influence behavioral and motivational states through their projections to the thalamus, ventral tegmental area and a brainstem 'rapid eye movement (REM)-induction' site. Action potential......-evoked intracellular calcium transients dampen excitability and stimulate NO production in these neurons. In this study, we investigated the action of several arousal-related neurotransmitters and the role of specific calcium channels in these LDT Ca(2+)-transients by simultaneous whole-cell recording and calcium...... of cholinergic LDT neurons and that inhibition of spike-evoked Ca(2+)-transients is a common action of neurotransmitters that also activate GIRK channels in these neurons. Because spike-evoked calcium influx dampens excitability, our findings suggest that these 'inhibitory' transmitters could boost firing rate...

  16. Derived (mutated)-types of TRPV6 channels elicit greater Ca²+ influx into the cells than ancestral-types of TRPV6: evidence from Xenopus oocytes and mammalian cell expression system.

    Science.gov (United States)

    Sudo, Yuka; Matsuo, Kiyotaka; Tetsuo, Tomoyuki; Tsutsumi, Satoshi; Ohkura, Masamichi; Nakai, Junichi; Uezono, Yasuhito

    2010-01-01

    The frequency of the allele containing three derived nonsynonymous SNPs (157C, 378M, 681M) of the gene encoding calcium permeable TRPV6 channels expressed in the intestine has been increased by positive selection in non-African populations. To understand the nature of these SNPs, we compared the properties of Ca²+ influx of ancestral (in African populations) and derived-TRPV6 (in non-African populations) channels with electrophysiological, Ca²+-imaging, and morphological methods using both the Xenopus oocyte and mammalian cell expression systems. Functional electrophysiological and Ca²+-imaging analyses indicated that the derived-TRPV6 elicited more Ca²+ influx than the ancestral one in TRPV6-expressing cells where both channels were equally expressed in the cells. Ca²+-inactivation properties in the ancestral- and derived-TRPV6 were almost the same. Furthermore, fluorescence resonance energy transfer (FRET) analysis showed that both channels have similar multimeric formation properties, suggesting that derived-TRPV6 itself could cause higher Ca²+ influx. These findings suggest that populations having derived-TRPV6 in non-African areas may absorb higher Ca²+ from the intestine than ancestral-TRPV6 in the African area.

  17. A mathematical model of T lymphocyte calcium dynamics derived from single transmembrane protein properties

    Directory of Open Access Journals (Sweden)

    Christine Dorothee Schmeitz

    2013-09-01

    Full Text Available Fate decision processes of T lymphocytes are crucial for health and disease. Whether a T lymphocyte is activated, divides, gets anergic or initiates apoptosis depends on extracellular triggers and intracellular signalling. Free cytosolic calcium dynamics plays an important role in this context. The relative contributions of store-derived calcium entry and calcium entry from extracellular space to T lymphocyte activation are still a matter of debate. Here we develop a quantitative mathematical model of T lymphocyte calcium dynamics in order to establish a tool which allows to disentangle cause-effect relationships between ion fluxes and observed calcium time courses. The model is based on single transmembrane protein characteristics which have been determined in independent experiments. This reduces the number of unknown parameters in the model to a minimum and ensures the predictive power of the model. Simulation results are subsequently used for an analysis of whole cell calcium dynamics measured under various experimental conditions. The model accounts for a variety of these conditions, which supports the suitability of the modelling approach. The simulation results suggest a model in which calcium dynamics dominantly relies on the opening of channels in calcium stores while calcium entry through calcium-release activated channels (CRAC is more associated with the maintenance of the T lymphocyte calcium levels and prevents the cell from calcium depletion. Our findings indicate that CRAC guarantees a long-term stable calcium level which is required for cell survival and sustained calcium enhancement.

  18. Structural dynamics of the cell nucleus: basis for morphology modulation of nuclear calcium signaling and gene transcription.

    Science.gov (United States)

    Queisser, Gillian; Wiegert, Simon; Bading, Hilmar

    2011-01-01

    Neuronal morphology plays an essential role in signal processing in the brain. Individual neurons can undergo use-dependent changes in their shape and connectivity, which affects how intracellular processes are regulated and how signals are transferred from one cell to another in a neuronal network. Calcium is one of the most important intracellular second messengers regulating cellular morphologies and functions. In neurons, intracellular calcium levels are controlled by ion channels in the plasma membrane such as NMDA receptors (NMDARs), voltage-gated calcium channels (VGCCs) and certain α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) as well as by calcium exchange pathways between the cytosol and internal calcium stores including the endoplasmic reticulum and mitochondria. Synaptic activity and the subsequent opening of ligand and/or voltage-gated calcium channels can initiate cytosolic calcium transients which propagate towards the cell soma and enter the nucleus via its nuclear pore complexes (NPCs) embedded in the nuclear envelope. We recently described the discovery that in hippocampal neurons the morphology of the nucleus affects the calcium dynamics within the nucleus. Here we propose that nuclear infoldings determine whether a nucleus functions as an integrator or detector of oscillating calcium signals. We outline possible ties between nuclear mophology and transcriptional activity and discuss the importance of extending the approach to whole cell calcium signal modeling in order to understand synapse-to-nucleus communication in healthy and dysfunctional neurons.

  19. Dose calcium channel blocker verapamil decrease urinary VMA levels in sympathoadrenal hyperactive patients with posttraumatic stress disorder?

    Institute of Scientific and Technical Information of China (English)

    Munawar Alam Ansari; Shahida PAhmed; Zahida Memon

    2008-01-01

    Objective:The majority of the patients with posttraumatic stress disorders (PTSD)embrace augmented urina-ry flow of Vanillylmandelic Acid (VMA)than normal subjects owing to superior sympathetic doings,which steer to cardiovascular catastrophe.Urinary flow of VMA was evaluated as sympathoadrenal bustle marker in patients with posttraumatic stress disorder.Calcium ion shows a noteworthy dependability in nervousness owing to its special effects on brain synaptosomes.So this study was conducted to explore the effects of Verapamil on sympathoadrenal motion in patients with PTSD.Methods:Placebo controlled clinical tryout was conducted. At first hundred (100)PTSD patients were chosen and enrolled in the study,from department of Psychological Medicine Dow University of Health Sciences,Karachi.Verapamil 120 mg/day was specified in divided doses to group-I (n =50)patients and group-II (n =50)patients received placebo therapy on a daily basis for nine weeks.Each and every patient was monitored weekly,all the way through extent of study.Results:Under-neath the posttraumatic stress disorder,urinary excretion of VMA was greater.Calcium channel blocker vera-pamil additionally abolished the embellished retort in urinary flow of VMA appreciably in patients with PTSD. Conclusion:Verapamil was experiential to be exceedingly effectual treatment.It reduces VMA levels in u-rine,and on the whole cardiovascular threat in PTSD patients.

  20. Blockade of T-type calcium channels prevents tonic-clonic seizures in a maximal electroshock seizure model.

    Science.gov (United States)

    Sakkaki, Sophie; Gangarossa, Giuseppe; Lerat, Benoit; Françon, Dominique; Forichon, Luc; Chemin, Jean; Valjent, Emmanuel; Lerner-Natoli, Mireille; Lory, Philippe

    2016-02-01

    T-type (Cav3) calcium channels play important roles in neuronal excitability, both in normal and pathological activities of the brain. In particular, they contribute to hyper-excitability disorders such as epilepsy. Here we have characterized the anticonvulsant properties of TTA-A2, a selective T-type channel blocker, in mouse. Using the maximal electroshock seizure (MES) as a model of tonic-clonic generalized seizures, we report that mice treated with TTA-A2 (0.3 mg/kg and higher doses) were significantly protected against tonic seizures. Although no major change in Local Field Potential (LFP) pattern was observed during the MES seizure, analysis of the late post-ictal period revealed a significant increase in the delta frequency power in animals treated with TTA-A2. Similar results were obtained for Cav3.1-/- mice, which were less prone to develop tonic seizures in the MES test, but not for Cav3.2-/- mice. Analysis of extracellular signal-regulated kinase 1/2 (ERK) phosphorylation and c-Fos expression revealed a rapid and elevated neuronal activation in the hippocampus following MES clonic seizures, which was unchanged in TTA-A2 treated animals. Overall, our data indicate that TTA-A2 is a potent anticonvulsant and that the Cav3.1 isoform plays a prominent role in mediating TTA-A2 tonic seizure protection. Copyright © 2015. Published by Elsevier Ltd.

  1. Caffeine-Induced Suppression of GABAergic Inhibition and Calcium-Independent Metaplasticity

    Directory of Open Access Journals (Sweden)

    Masako Isokawa

    2016-01-01

    Full Text Available GABAergic inhibition plays a critical role in the regulation of neuron excitability; thus, it is subject to modulations by many factors. Recent evidence suggests the elevation of intracellular calcium ([Ca2+]i and calcium-dependent signaling molecules underlie the modulations. Caffeine induces a release of calcium from intracellular stores. We tested whether caffeine modulated GABAergic transmission by increasing [Ca2+]i. A brief local puff-application of caffeine to hippocampal CA1 pyramidal cells transiently suppressed GABAergic inhibitory postsynaptic currents (IPSCs by 73.2 ± 6.98%. Time course of suppression and the subsequent recovery of IPSCs resembled DSI (depolarization-induced suppression of inhibition, mediated by endogenous cannabinoids that require a [Ca2+]i rise. However, unlike DSI, caffeine-induced suppression of IPSCs (CSI persisted in the absence of a [Ca2+]i rise. Intracellular applications of BAPTA and ryanodine (which blocks caffeine-induced calcium release from intracellular stores failed to prevent the generation of CSI. Surprisingly, ruthenium red, an inhibitor of multiple calcium permeable/release channels including those of stores, induced metaplasticity by amplifying the magnitude of CSI independently of calcium. This metaplasticity was accompanied with the generation of a large inward current. Although ionic basis of this inward current is undetermined, the present result demonstrates that caffeine has a robust Ca2+-independent inhibitory action on GABAergic inhibition and causes metaplasticity by opening plasma membrane channels.

  2. Diffusive spatio-temporal noise in a first-passage time model for intracellular calcium release

    KAUST Repository

    Flegg, Mark B.; Rüdiger, Sten; Erban, Radek

    2013-01-01

    The intracellular release of calcium from the endoplasmic reticulum is controlled by ion channels. The resulting calcium signals exhibit a rich spatio-temporal signature, which originates at least partly from microscopic fluctuations. While

  3. A Single Nucleotide Polymorphism (rs4236480 in TRPV5 Calcium Channel Gene Is Associated with Stone Multiplicity in Calcium Nephrolithiasis Patients

    Directory of Open Access Journals (Sweden)

    Anas Khaleel

    2015-01-01

    Full Text Available Nephrolithiasis is characterized by calcification of stones in the kidneys from an unknown cause. Animal models demonstrated the functional roles of the transient receptor potential vanilloid member 5 (TRPV5 gene in calcium renal reabsorption and hypercalciuria. Therefore, TRPV5 was suggested to be involved in calcium homeostasis. However, whether genetic polymorphisms of TRPV5 are associated with kidney stone multiplicity or recurrence is unclear. In this study, 365 Taiwanese kidney-stone patients were recruited. Both biochemical data and DNA samples were collected. Genotyping was performed by a TaqMan allelic discrimination assay. We found that a TRPV5 polymorphism (rs4236480 was observed to be associated with stone multiplicity of calcium nephrolithiasis, as the risk of stone multiplicity was higher in patients with the TT+CT genotype than in patients with the CC genotype (p=0.0271. In summary, despite the complexity of nephrolithiasis and the potential association of numerous calcium homeostatic absorption/reabsorption factors, TRPV5 plays an important role in the pathogenesis of calcium nephrolithiasis.

  4. Canonical transient receptor potential channel 2 (TRPC2): old name-new games. Importance in regulating of rat thyroid cell physiology.

    Science.gov (United States)

    Törnquist, Kid; Sukumaran, Pramod; Kemppainen, Kati; Löf, Christoffer; Viitanen, Tero

    2014-11-01

    In addition to the TSH-cyclic AMP signalling pathway, calcium signalling is of crucial importance in thyroid cells. Although the importance of calcium signalling has been thoroughly investigated for several decades, the nature of the calcium channels involved in signalling is unknown. In a recent series of investigations using the well-studied rat thyroid FRTL-5 cell line, we showed that these cells exclusively express the transient receptor potential canonical 2 (TRPC2) channel. Our results suggested that the TRPC2 channel is of significant importance in regulating thyroid cell function. These investigations were the first to show that thyroid cells express a member of the TRPC family of ion channels. In this review, we will describe the importance of the TRPC2 channel in regulating TSH receptor expression, thyroglobulin maturation, intracellular calcium and iodide homeostasis and that the channel also regulates thyroid cell proliferation.

  5. Glu¹⁰⁶ in the Orai1 pore contributes to fast Ca²⁺-dependent inactivation and pH dependence of Ca²⁺ release-activated Ca²⁺ (CRAC) current.

    Science.gov (United States)

    Scrimgeour, Nathan R; Wilson, David P; Rychkov, Grigori Y

    2012-01-15

    FCDI (fast Ca²⁺-dependent inactivation) is a mechanism that limits Ca²⁺ entry through Ca²⁺ channels, including CRAC (Ca²⁺ release-activated Ca²⁺) channels. This phenomenon occurs when the Ca²⁺ concentration rises beyond a certain level in the vicinity of the intracellular mouth of the channel pore. In CRAC channels, several regions of the pore-forming protein Orai1, and STIM1 (stromal interaction molecule 1), the sarcoplasmic/endoplasmic reticulum Ca²⁺ sensor that communicates the Ca²⁺ load of the intracellular stores to Orai1, have been shown to regulate fast Ca²⁺-dependent inactivation. Although significant advances in unravelling the mechanisms of CRAC channel gating have occurred, the mechanisms regulating fast Ca²⁺-dependent inactivation in this channel are not well understood. We have identified that a pore mutation, E106D Orai1, changes the kinetics and voltage dependence of the ICRAC (CRAC current), and the selectivity of the Ca²⁺-binding site that regulates fast Ca²⁺-dependent inactivation, whereas the V102I and E190Q mutants when expressed at appropriate ratios with STIM1 have fast Ca²⁺-dependent inactivation similar to that of WT (wild-type) Orai1. Unexpectedly, the E106D mutation also changes the pH dependence of ICRAC. Unlike WT ICRAC, E106D-mediated current is not inhibited at low pH, but instead the block of Na⁺ permeation through the E106D Orai1 pore by Ca²⁺ is diminished. These results suggest that Glu¹⁰⁶ inside the CRAC channel pore is involved in co-ordinating the Ca²⁺-binding site that mediates fast Ca²⁺-dependent inactivation.

  6. Lipid nanocapsules containing the non-ionic surfactant Solutol HS15 inhibit the transport of calcium through hyperforin-activated channels in neuronal cells.

    Science.gov (United States)

    Chauvet, Sylvain; Barras, Alexandre; Boukherroub, Rabah; Bouron, Alexandre

    2015-12-01

    Hyperforin is described as a natural antidepressant inhibiting the reuptake of neurotransmitters and also activating cation channels. However the blood-brain barrier limits the access to the brain of this biomolecule. To circumvent this problem it was envisaged to encapsulate hyperforin into biomimetic lipid nano-carriers like lipid nanocapsules (LNCs). When testing the safety of 25 nm LNCs it appeared that they strongly blocked hyperforin-activated Ca2+ channels of cultured cortical neurons. This inhibition was due to one of their main component: solutol HS15 (polyoxyethylene-660-12-hydroxy stearate), a non-ionic soluble surfactant. Solutol HS15 rapidly depresses in a concentration-dependent manner the entry of Ca2+ through hyperforin-activated channels without influencing store-operated channels. This effect is mimicked by Brij58 but not by PEG600, indicating that the lipid chain of Solutol HS15 is important in determining its effects on the channels. The inhibition of the Ca2+ fluxes depends on the cellular cholesterol content; it is stronger after depleting cholesterol with methyl-β-cyclodextrin and is nearly absent on cells cultured in a cholesterol-rich medium. When chronically applied for 24 h, Solutol HS15 slightly up-regulates the entry of Ca2+ through hyperforin-activated channels. Similar observations were made when testing 25 nm lipid nanocapsules containing the surfactant Solutol HS15. Altogether, this study shows that Solutol HS15 perturbs in a cholesterol-dependent manner the activity of some neuronal channels. This is the first demonstration that LNCs containing this surfactant can influence cellular calcium signaling in the brain, a finding that can have important clinical implications. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Potential impact of renin-angiotensin system inhibitors and calcium channel blockers on plasma high-molecular-weight adiponectin levels in hemodialysis patients

    International Nuclear Information System (INIS)

    Nakagawa, Naoki; Yao, Naoyuki; Hirayama, Tomoya

    2011-01-01

    Although metabolic syndrome confers an increased risk of cardiovascular disease in the general population, little is known about the alteration of abdominal adiposity and its association with adipocytokines in hemodialysis patients. We investigated the plasma high-molecular-weight (HMW) adiponectin level and its relationship to visceral fat area (VFA) and various markers of atherosclerosis in hemodialysis patients. In a cross-sectional study, conventional cardiovascular risk factors, plasma total and HMW adiponectin, the number of components of the metabolic syndrome and, using computed tomography, the distribution of abdominal adiposity were assessed in 144 hemodialysis patients (90 men and 54 women; mean age, 60.7 years) and 30 age- and sex-matched patients with chronic kidney disease (CKD). Plasma HMW adiponectin levels in hemodialysis patients were significantly higher than those in patients with CKD, negatively associated with VFA and serum triglycerides and positively associated with plasma total adiponectin, as well as the HMW-to-total adiponectin ratio in men and women (all P<0.05) in a simple regression analysis. In a multiple regression analysis, VFA was a significant determinant of HMW adiponectin in hemodialysis patients. Furthermore, after adjustment for classical risk factors, HMW adiponectin levels were significantly higher in patients undergoing treatment with renin-angiotensin system inhibitors or calcium channel blockers compared with patients not undergoing such treatment. This study shows that plasma HMW adiponectin levels were negatively associated with VFA and positively associated with treatment with blockade of the renin-angiotensin system and of the calcium channel. Therefore, these drugs might be effective for improving adipocytokine-related metabolic abnormalities in hemodialysis patients. (author)

  8. The human Nav1.5 F1486 deletion associated with long QT syndrome leads to impaired sodium channel inactivation and reduced lidocaine sensitivity

    Science.gov (United States)

    Song, Weihua; Xiao, Yucheng; Chen, Hanying; Ashpole, Nicole M; Piekarz, Andrew D; Ma, Peilin; Hudmon, Andy; Cummins, Theodore R; Shou, Weinian

    2012-01-01

    The deletion of phenylalanine 1486 (F1486del) in the human cardiac voltage-gated sodium channel (hNav1.5) is associated with fatal long QT (LQT) syndrome. In this study we determined how F1486del impairs the functional properties of hNav1.5 and alters action potential firing in heterologous expression systems (human embryonic kidney (HEK) 293 cells) and their native cardiomyocyte background. Cells expressing hNav1.5-F1486del exhibited a loss-of-function alteration, reflected by an 80% reduction of peak current density, and several gain-of-function alterations, including reduced channel inactivation, enlarged window current, substantial augmentation of persistent late sodium current and an increase in ramp current. We also observed substantial action potential duration (APD) prolongation and prominent early afterdepolarizations (EADs) in neonatal cardiomyocytes expressing the F1486del channels, as well as in computer simulations of myocyte activity. In addition, lidocaine sensitivity was dramatically reduced, which probably contributed to the poor therapeutic outcome observed in the patient carrying the hNav1.5-F1486del mutation. Therefore, despite the significant reduction in peak current density, the F1486del mutation also leads to substantial gain-of-function alterations that are sufficient to cause APD prolongation and EADs, the predominant characteristic of LQTs. These data demonstrate that hNav1.5 mutations can have complex functional consequences and highlight the importance of identifying the specific molecular defect when evaluating potential treatments for individuals with prolonged QT intervals. PMID:22826127

  9. Voltage-gated sodium channels: pharmaceutical targets via anticonvulsants to treat epileptic syndromes.

    Science.gov (United States)

    Abdelsayed, Mena; Sokolov, Stanislav

    2013-01-01

    Epilepsy is a brain disorder characterized by seizures and convulsions. The basis of epilepsy is an increase in neuronal excitability that, in some cases, may be caused by functional defects in neuronal voltage gated sodium channels, Nav1.1 and Nav1.2. The effects of antiepileptic drugs (AEDs) as effective therapies for epilepsy have been characterized by extensive research. Most of the classic AEDs targeting Nav share a common mechanism of action by stabilizing the channel's fast-inactivated state. In contrast, novel AEDs, such as lacosamide, stabilize the slow-inactivated state in neuronal Nav1.1 and Nav1.7 isoforms. This paper reviews the different mechanisms by which this stabilization occurs to determine new methods for treatment.

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

    DEFF Research Database (Denmark)

    Lundby, Alicia; Olesen, Søren-Peter

    2006-01-01

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

  11. Kv1 channels and neural processing in vestibular calyx afferents

    Directory of Open Access Journals (Sweden)

    Frances L Meredith

    2015-06-01

    Full Text Available Potassium-selective ion channels are important for accurate transmission of signals from auditory and vestibular sensory end organs to their targets in the central nervous system. During different gravity conditions, astronauts experience altered input signals from the peripheral vestibular system resulting in sensorimotor dysfunction. Adaptation to altered sensory input occurs, but it is not explicitly known whether this involves synaptic modifications within the vestibular epithelia. Future investigations of such potential plasticity require a better understanding of the electrophysiological mechanisms underlying the known heterogeneity of afferent discharge under normal conditions. This study advances this understanding by examining the role of the Kv1 potassium channel family in mediating action potentials in specialized vestibular afferent calyx endings in the gerbil crista and utricle. Pharmacological agents selective for different sub-types of Kv1 channels were tested on membrane responses in whole cell recordings in the crista. Kv1 channels sensitive to α-dendrotoxin and dendrotoxin-K were found to prevail in the central regions, whereas K+ channels sensitive to margatoxin, which blocks Kv1.3 and 1.6 channels, were more prominent in peripheral regions. Margatoxin-sensitive currents showed voltage-dependent inactivation. Dendrotoxin-sensitive currents showed no inactivation and dampened excitability in calyces in central neuroepithelial regions. The differential distribution of Kv1 potassium channels in vestibular afferents supports their importance in accurately relaying gravitational and head movement signals through specialized lines to the central nervous system. Pharmacological modulation of specific groups of K+ channels could help alleviate vestibular dysfunction on earth and in space.

  12. Kv1 channels and neural processing in vestibular calyx afferents.

    Science.gov (United States)

    Meredith, Frances L; Kirk, Matthew E; Rennie, Katherine J

    2015-01-01

    Potassium-selective ion channels are important for accurate transmission of signals from auditory and vestibular sensory end organs to their targets in the central nervous system. During different gravity conditions, astronauts experience altered input signals from the peripheral vestibular system resulting in sensorimotor dysfunction. Adaptation to altered sensory input occurs, but it is not explicitly known whether this involves synaptic modifications within the vestibular epithelia. Future investigations of such potential plasticity require a better understanding of the electrophysiological mechanisms underlying the known heterogeneity of afferent discharge under normal conditions. This study advances this understanding by examining the role of the Kv1 potassium channel family in mediating action potentials in specialized vestibular afferent calyx endings in the gerbil crista and utricle. Pharmacological agents selective for different sub-types of Kv1 channels were tested on membrane responses in whole cell recordings in the crista. Kv1 channels sensitive to α-dendrotoxin and dendrotoxin-K were found to prevail in the central regions, whereas K(+) channels sensitive to margatoxin, which blocks Kv1.3 and 1.6 channels, were more prominent in peripheral regions. Margatoxin-sensitive currents showed voltage-dependent inactivation. Dendrotoxin-sensitive currents showed no inactivation and dampened excitability in calyces in central neuroepithelial regions. The differential distribution of Kv1 potassium channels in vestibular afferents supports their importance in accurately relaying gravitational and head movement signals through specialized lines to the central nervous system. Pharmacological modulation of specific groups of K(+) channels could help alleviate vestibular dysfunction on earth and in space.

  13. Altered expression of the voltage-gated calcium channel subunit alpha(2)delta-1: A comparison between two experimental models of epilepsy and a sensory nerve ligation model of neuropathic pain

    Czech Academy of Sciences Publication Activity Database

    Nieto-Rostro, M.; Sandhu, G.; Bauer, C. S.; Jiruška, Přemysl; Jefferys, J. G. R.; Dolphin, A. C.

    2014-01-01

    Roč. 283, Dec (2014), s. 124-137 ISSN 0306-4522 R&D Projects: GA MZd(CZ) NT14489 Institutional support: RVO:67985823 Keywords : calcium channel * dorsal root ganglion (DRG) * alpha2delta subunit * epilepsy * neuropathic pain * reactive gliosis Subject RIV: FH - Neurology Impact factor: 3.357, year: 2014

  14. Mechanisms of pyrethroid insecticide-induced stimulation of calcium influx in neocortical neurons

    Science.gov (United States)

    Pyrethroid insecticides bind to voltage-gated sodium channels (VGSCs) and modify their gating kinetics, thereby disrupting neuronal function. Pyrethroids have also been reported to alter the function of other channel types, including activation of voltage-gated Ca2+ calcium chann...

  15. ATP Release Channels

    Directory of Open Access Journals (Sweden)

    Akiyuki Taruno

    2018-03-01

    Full Text Available Adenosine triphosphate (ATP has been well established as an important extracellular ligand of autocrine signaling, intercellular communication, and neurotransmission with numerous physiological and pathophysiological roles. In addition to the classical exocytosis, non-vesicular mechanisms of cellular ATP release have been demonstrated in many cell types. Although large and negatively charged ATP molecules cannot diffuse across the lipid bilayer of the plasma membrane, conductive ATP release from the cytosol into the extracellular space is possible through ATP-permeable channels. Such channels must possess two minimum qualifications for ATP permeation: anion permeability and a large ion-conducting pore. Currently, five groups of channels are acknowledged as ATP-release channels: connexin hemichannels, pannexin 1, calcium homeostasis modulator 1 (CALHM1, volume-regulated anion channels (VRACs, also known as volume-sensitive outwardly rectifying (VSOR anion channels, and maxi-anion channels (MACs. Recently, major breakthroughs have been made in the field by molecular identification of CALHM1 as the action potential-dependent ATP-release channel in taste bud cells, LRRC8s as components of VRACs, and SLCO2A1 as a core subunit of MACs. Here, the function and physiological roles of these five groups of ATP-release channels are summarized, along with a discussion on the future implications of understanding these channels.

  16. Role of T-type channels in vasomotor function

    DEFF Research Database (Denmark)

    Kuo, Ivana Y-T; Howitt, Lauren; Sandow, Shaun L

    2014-01-01

    Low-voltage-activated T-type calcium channels play an important role in regulating cellular excitability and are implicated in conditions, such as epilepsy and neuropathic pain. T-type channels, especially Cav3.1 and Cav3.2, are also expressed in the vasculature, although patch clamp studies of i...

  17. PKA Controls Calcium Influx into Motor Neurons during a Rhythmic Behavior

    Science.gov (United States)

    Wang, Han; Sieburth, Derek

    2013-01-01

    Cyclic adenosine monophosphate (cAMP) has been implicated in the execution of diverse rhythmic behaviors, but how cAMP functions in neurons to generate behavioral outputs remains unclear. During the defecation motor program in C. elegans, a peptide released from the pacemaker (the intestine) rhythmically excites the GABAergic neurons that control enteric muscle contractions by activating a G protein-coupled receptor (GPCR) signaling pathway that is dependent on cAMP. Here, we show that the C. elegans PKA catalytic subunit, KIN-1, is the sole cAMP target in this pathway and that PKA is essential for enteric muscle contractions. Genetic analysis using cell-specific expression of dominant negative or constitutively active PKA transgenes reveals that knockdown of PKA activity in the GABAergic neurons blocks enteric muscle contractions, whereas constitutive PKA activation restores enteric muscle contractions to mutants defective in the peptidergic signaling pathway. Using real-time, in vivo calcium imaging, we find that PKA activity in the GABAergic neurons is essential for the generation of synaptic calcium transients that drive GABA release. In addition, constitutively active PKA increases the duration of calcium transients and causes ectopic calcium transients that can trigger out-of-phase enteric muscle contractions. Finally, we show that the voltage-gated calcium channels UNC-2 and EGL-19, but not CCA-1 function downstream of PKA to promote enteric muscle contractions and rhythmic calcium influx in the GABAergic neurons. Thus, our results suggest that PKA activates neurons during a rhythmic behavior by promoting presynaptic calcium influx through specific voltage-gated calcium channels. PMID:24086161

  18. PKA controls calcium influx into motor neurons during a rhythmic behavior.

    Directory of Open Access Journals (Sweden)

    Han Wang

    Full Text Available Cyclic adenosine monophosphate (cAMP has been implicated in the execution of diverse rhythmic behaviors, but how cAMP functions in neurons to generate behavioral outputs remains unclear. During the defecation motor program in C. elegans, a peptide released from the pacemaker (the intestine rhythmically excites the GABAergic neurons that control enteric muscle contractions by activating a G protein-coupled receptor (GPCR signaling pathway that is dependent on cAMP. Here, we show that the C. elegans PKA catalytic subunit, KIN-1, is the sole cAMP target in this pathway and that PKA is essential for enteric muscle contractions. Genetic analysis using cell-specific expression of dominant negative or constitutively active PKA transgenes reveals that knockdown of PKA activity in the GABAergic neurons blocks enteric muscle contractions, whereas constitutive PKA activation restores enteric muscle contractions to mutants defective in the peptidergic signaling pathway. Using real-time, in vivo calcium imaging, we find that PKA activity in the GABAergic neurons is essential for the generation of synaptic calcium transients that drive GABA release. In addition, constitutively active PKA increases the duration of calcium transients and causes ectopic calcium transients that can trigger out-of-phase enteric muscle contractions. Finally, we show that the voltage-gated calcium channels UNC-2 and EGL-19, but not CCA-1 function downstream of PKA to promote enteric muscle contractions and rhythmic calcium influx in the GABAergic neurons. Thus, our results suggest that PKA activates neurons during a rhythmic behavior by promoting presynaptic calcium influx through specific voltage-gated calcium channels.

  19. Macrophage activation by a vanadyl-aspirin complex is dependent on L-type calcium channel and the generation of nitric oxide

    International Nuclear Information System (INIS)

    Molinuevo, Maria Silvina; Etcheverry, Susana Beatriz; Cortizo, Ana Maria

    2005-01-01

    Bone homeostasis is the result of a tight balance between bone resorption and bone formation where macrophage activation is believed to contribute to bone resorption. We have previously shown that a vanadyl(IV)-aspirin complex (VOAspi) regulates cell proliferation and differentiation of osteoblasts in culture. In this study, we assessed VOAspi and VO effects and their possible mechanism of action on a mouse macrophage cell line RAW 264.7. Both vanadium compounds inhibited cell proliferation in a dose-dependent manner. Nifedipine completely reversed the VOAspi-induced macrophage cytotoxicity, while it could not block the effect of VO. VOAspi also stimulated nitric oxide (NO) production, the oxidation of dihydrorhodamine 123 (DHR-123) and enhanced the expression of both constitutive and inducible isoforms of nitric oxide syntases (NOS). All these effects were abolished by nifedipine. Althogether our finding give evidence that VOAspi-induced macrophage cytotoxicity is dependent on L-type calcium channel and the generation of NO though the induction of eNOS and iNOS. Contrary, the parent compound VO exerted a cytotoxic effect by mechanisms independent of a calcium entry and the NO/NOS activation

  20. Ion channels in glioblastoma.

    Science.gov (United States)

    Molenaar, Remco J

    2011-01-01

    Glioblastoma is the most common primary brain tumor with the most dismal prognosis. It is characterized by extensive invasion, migration, and angiogenesis. Median survival is only 15 months due to this behavior, rendering focal surgical resection ineffective and adequate radiotherapy impossible. At this moment, several ion channels have been implicated in glioblastoma proliferation, migration, and invasion. This paper summarizes studies on potassium, sodium, chloride, and calcium channels of glioblastoma. It provides an up-to-date overview of the literature that could ultimately lead to new therapeutic targets.

  1. Effects of the β1 auxiliary subunit on modification of Rat Na{sub v}1.6 sodium channels expressed in HEK293 cells by the pyrethroid insecticides tefluthrin and deltamethrin

    Energy Technology Data Exchange (ETDEWEB)

    He, Bingjun [College of Life Sciences, Nankai University, Tianjin 300071 (China); Soderlund, David M., E-mail: dms6@cornell.edu [Department of Entomology, Cornell University, Geneva, NY 14456 (United States)

    2016-01-15

    We expressed rat Na{sub v}1.6 sodium channels with or without the rat β1 subunit in human embryonic kidney (HEK293) cells and evaluated the effects of the pyrethroid insecticides tefluthrin and deltamethrin on whole-cell sodium currents. In assays with the Na{sub v}1.6 α subunit alone, both pyrethroids prolonged channel inactivation and deactivation and shifted the voltage dependence of channel activation and steady-state inactivation toward hyperpolarization. Maximal shifts in activation were ~ 18 mV for tefluthrin and ~ 24 mV for deltamethrin. These compounds also caused hyperpolarizing shifts of ~ 10–14 mV in the voltage dependence of steady-state inactivation and increased in the fraction of sodium current that was resistant to inactivation. The effects of pyrethroids on the voltage-dependent gating greatly increased the size of sodium window currents compared to unmodified channels; modified channels exhibited increased probability of spontaneous opening at membrane potentials more negative than the normal threshold for channel activation and incomplete channel inactivation. Coexpression of Na{sub v}1.6 with the β1 subunit had no effect on the kinetic behavior of pyrethroid-modified channels but had divergent effects on the voltage-dependent gating of tefluthrin- or deltamethrin-modified channels, increasing the size of tefluthrin-induced window currents but decreasing the size of corresponding deltamethrin-induced currents. Unexpectedly, the β1 subunit did not confer sensitivity to use-dependent channel modification by either tefluthrin or deltamethrin. We conclude from these results that functional reconstitution of channels in vitro requires careful attention to the subunit composition of channel complexes to ensure that channels in vitro are faithful functional and pharmacological models of channels in neurons. - Highlights: • We expressed Na{sub v}1.6 sodium channels with or without β1 subunits in HEK293 cells. • Tefluthrin and deltamethrin

  2. Fragile X mental retardation protein controls ion channel expression and activity.

    Science.gov (United States)

    Ferron, Laurent

    2016-10-15

    Fragile X-associated disorders are a family of genetic conditions resulting from the partial or complete loss of fragile X mental retardation protein (FMRP). Among these disorders is fragile X syndrome, the most common cause of inherited intellectual disability and autism. FMRP is an RNA-binding protein involved in the control of local translation, which has pleiotropic effects, in particular on synaptic function. Analysis of the brain FMRP transcriptome has revealed hundreds of potential mRNA targets encoding postsynaptic and presynaptic proteins, including a number of ion channels. FMRP has been confirmed to bind voltage-gated potassium channels (K v 3.1 and K v 4.2) mRNAs and regulates their expression in somatodendritic compartments of neurons. Recent studies have uncovered a number of additional roles for FMRP besides RNA regulation. FMRP was shown to directly interact with, and modulate, a number of ion channel complexes. The sodium-activated potassium (Slack) channel was the first ion channel shown to directly interact with FMRP; this interaction alters the single-channel properties of the Slack channel. FMRP was also shown to interact with the auxiliary β4 subunit of the calcium-activated potassium (BK) channel; this interaction increases calcium-dependent activation of the BK channel. More recently, FMRP was shown to directly interact with the voltage-gated calcium channel, Ca v 2.2, and reduce its trafficking to the plasma membrane. Studies performed on animal models of fragile X syndrome have revealed links between modifications of ion channel activity and changes in neuronal excitability, suggesting that these modifications could contribute to the phenotypes observed in patients with fragile X-associated disorders. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

  3. Calcium fluxes across the plasma membrane of Commelina communis L. assayed in a cell-free system

    International Nuclear Information System (INIS)

    Siebers, B.; Graef, P.; Weiler, E.W.

    1990-01-01

    The inside-out fraction of plasma membrane-rich vesicles prepared from leaves of Commelina communis L. by aqueous two-phase partitioning was loaded with 45 Ca 2+ through the action of the plasma membrane Ca 2+ -ATPase. Results suggest the presence of a Ca 2+ channel in the plasma membrane of C. communis. The channel is obtained in a Ca 2+ -inactivated state after preparation and Ca 2+ -loading of the vesicles. The inactivation is removed by TFP [trifluoperazine] or W-7 [N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide], presumably due to the Ca 2+ -mobilizing effect of these compounds. The activated Ca 2+ channel is La 3+ sensitive and, in the cell, would allow for passage of Ca 2+ into the cell. The possibility that TFP or W-7 act independent of CM, or through CM tightly associated with the plasma membrane, is discussed

  4. Synthesis and evaluation of calcium channel antagonist activity of new 1, 4-dihydropyridines containing phenylamineimidazolyl substitute in guinea-pig ileal smooth muscle

    Directory of Open Access Journals (Sweden)

    A Fassihi

    2004-02-01

    Full Text Available Background: 1,4-dihydropyridines are a class of drugs which are used in the treatment of some cardiovascular disorders. The prototype, Nifedipine, does not have optimal pharmacokinetic and pharmacodynamic properties. Several new derivatives of 1, 4-dihydropyridine have been produced and pharmacologically evaluated in order to find drugs with better pharmacological properties. Among them, those with a substituted heteroaromatic ring in the C4 position of the 1, 4-dihydropyridine ring, instead of the phenyl ring in Nifedipine, are most considered. In this study, eight novel derivatives of this class with “2-methylthio-1-(phenylaminoimidazole-5-yl” in the C4, C3 and C5 positions were prepared and evaluated as calcium channel antagonist agents. Methods: To prepare these compounds, Hantzsch method for the synthesis of 1, 4-dihydropyridine derivatives was deployed. An aldehyde was reacted with appropriate acetoacetate ester and ammonium acetate. This aldehyde was prepared in three steps. Cumulative doses were applied to determine the relaxing effect of the compounds on the longitudinal smooth muscle of male albino guinea pigs. Results: Chemical structures of the compounds were characterized by 1H nuclear magnetic resonance, infrared and mass spectroscopy. The IC50 of each compound was graphically determined from the concentration-response curves. Conclusions: Two compounds were more active than Nifedipine. Both had lipophilic ester groups with low steric hindrance that met the merits of a better receptor binding of 1, 4-dihydropyridines. These derivatives have high potential for further study. Keywords: 1, 4-dihydropyridine, Calcium channel antagonist, Phenylamineimidazolyl, Cardiovascular disorder

  5. TRPV5: an ingeniously controlled calcium channel.

    NARCIS (Netherlands)

    Groot, T. de; Bindels, R.J.M.; Hoenderop, J.G.J.

    2008-01-01

    Body Ca(2+) homeostasis is tightly controlled and slight disturbances in renal Ca(2+) reabsorption can lead to excessive urine Ca(2+) excretion and promote kidney stone formation. The epithelial Ca(2+) channel TRPV5 constitutes the rate-limiting step of active Ca(2+) reabsorption in the kidney.

  6. Mechanisms of the gabapentinoids and α 2 δ-1 calcium channel subunit in neuropathic pain.

    Science.gov (United States)

    Patel, Ryan; Dickenson, Anthony H

    2016-04-01

    The gabapentinoid drugs gabapentin and pregabalin are key front-line therapies for various neuropathies of peripheral and central origin. Originally designed as analogs of GABA, the gabapentinoids bind to the α 2 δ-1 and α 2 δ-2 auxiliary subunits of calcium channels, though only the former has been implicated in the development of neuropathy in animal models. Transgenic approaches also identify α 2 δ-1 as key in mediating the analgesic effects of gabapentinoids, however the precise molecular mechanisms remain unclear. Here we review the current understanding of the pathophysiological role of the α 2 δ-1 subunit, the mechanisms of analgesic action of gabapentinoid drugs and implications for efficacy in the clinic. Despite widespread use, the number needed to treat for gabapentin and pregabalin averages from 3 to 8 across neuropathies. The failure to treat large numbers of patients adequately necessitates a novel approach to treatment selection. Stratifying patients by sensory profiles may imply common underlying mechanisms, and a greater understanding of these mechanisms could lead to more direct targeting of gabapentinoids.

  7. Inactivation Data.xlsx

    Data.gov (United States)

    U.S. Environmental Protection Agency — The data set is a spreadsheet that contains results of inactivation experiments that were conducted to to determine the effectiveness of chlorine in inactivating B....

  8. SK2 channels regulate mitochondrial respiration and mitochondrial Ca2+ uptake.

    Science.gov (United States)

    Honrath, Birgit; Matschke, Lina; Meyer, Tammo; Magerhans, Lena; Perocchi, Fabiana; Ganjam, Goutham K; Zischka, Hans; Krasel, Cornelius; Gerding, Albert; Bakker, Barbara M; Bünemann, Moritz; Strack, Stefan; Decher, Niels; Culmsee, Carsten; Dolga, Amalia M

    2017-05-01

    Mitochondrial calcium ([Ca 2+ ] m ) overload and changes in mitochondrial metabolism are key players in neuronal death. Small conductance calcium-activated potassium (SK) channels provide protection in different paradigms of neuronal cell death. Recently, SK channels were identified at the inner mitochondrial membrane, however, their particular role in the observed neuroprotection remains unclear. Here, we show a potential neuroprotective mechanism that involves attenuation of [Ca 2+ ] m uptake upon SK channel activation as detected by time lapse mitochondrial Ca 2+ measurements with the Ca 2+ -binding mitochondria-targeted aequorin and FRET-based [Ca 2+ ] m probes. High-resolution respirometry revealed a reduction in mitochondrial respiration and complex I activity upon pharmacological activation and overexpression of mitochondrial SK2 channels resulting in reduced mitochondrial ROS formation. Overexpression of mitochondria-targeted SK2 channels enhanced mitochondrial resilience against neuronal death, and this effect was inhibited by overexpression of a mitochondria-targeted dominant-negative SK2 channel. These findings suggest that SK channels provide neuroprotection by reducing [Ca 2+ ] m uptake and mitochondrial respiration in conditions, where sustained mitochondrial damage determines progressive neuronal death.

  9. Changes in the distribution of lens calcium during development of x-ray cataract

    International Nuclear Information System (INIS)

    Hightower, K.R.; Giblin, F.J.; Reddy, V.N.

    1983-01-01

    The present study was designed to examine the possible role of calcium in the opacification of x-ray-induced cataract in rabbit. The results demonstrate that the concentration of calcium in x-rayed lenses, just prior to lens hydration (7.5 weeks postirradiation), was twice that present in contralateral control lenses. At this stage of immature cataract, the lens nucleus remained transparent and maintained a normal level of calcium, but the lens cortex, containing regions of subcapsular opacification, accumulated a level of calcium that was twice that of the control. In the completely opaque mature cataract, (8-9 weeks post x-ray), both the cortex and nucleus had gained significant amounts of calcium. As the concentration of total calcium increased in the immature x-ray cataract, the amount of the cation bound to membranes and insoluble proteins of the cytosol also increased comparably. However, the relative proportion of calcium in the various fractions remained unaltered in the immature cataract; in both control lenses and immature cataracts, 20% of the total calcium remained in the membrane pellet and 70% was located in the soluble protein fraction. Only in the mature stage of cataract was a shift in the distribution of calcium apparent, as the proportion of calcium in the soluble protein fraction increased to 90%. Although only 7% of the total calcium in a mature cataract was bound to membrane, the amount represented a fivefold increase over the control. The results of this study demonstrate that an elevation in lens calcium accompanies the opacification process in x-ray cataract. The work also suggests that changes in calcium levels are not likely to result from inactivation of Ca-ATPase

  10. T-type channels: release a brake, engage a gear

    Czech Academy of Sciences Publication Activity Database

    Weiss, Norbert; Lacinová, L.

    2016-01-01

    Roč. 10, č. 2 (2016), s. 78-80 ISSN 1933-6950 Institutional support: RVO:61388963 Keywords : gating brake * pore opening * Ca(V)3.3 * channel gating * Ca(V)3.1 * low-voltage activated calcium channels Subject RIV: CE - Biochemistry Impact factor: 2.042, year: 2016

  11. Effects of the small molecule HERG activator NS1643 on Kv11.3 channels.

    Directory of Open Access Journals (Sweden)

    Arne Bilet

    Full Text Available NS1643 is one of the small molecule HERG (Kv11.1 channel activators and has also been found to increase erg2 (Kv11.2 currents. We now investigated whether NS1643 is also able to act as an activator of Kv11.3 (erg3 channels expressed in CHO cells. Activation of rat Kv11.3 current occurred in a dose-dependent manner and maximal current increasing effects were obtained with 10 µM NS1643. At this concentration, steady-state outward current increased by about 80% and the current increase was associated with a significant shift in the voltage dependence of activation to more negative potentials by about 15 mV. In addition, activation kinetics were accelerated, whereas deactivation was slowed. There was no significant effect on the kinetics of inactivation and recovery from inactivation. The strong current-activating agonistic effect of NS1643 did not result from a shift in the voltage dependence of Kv11.3 channel inactivation and was independent from external Na(+ or Ca(2+. At the higher concentration of 20 µM, NS1643 induced clearly less current increase. The left shift in the voltage dependence of activation reversed and the voltage sensitivity of activation dramatically decreased along with a slowing of Kv11.3 channel activation. These data show that, in comparison to other Kv11 family members, NS1643 exerts distinct effects on Kv11.3 channels with especially pronounced partial antagonistic effects at higher concentration.

  12. One Dimensional Finite Element Method Approach to Study Effect of Ryanodine Receptor and Serca Pump on Calcium Distribution in Oocytes

    Science.gov (United States)

    Naik, Parvaiz Ahmad; Pardasani, Kamal Raj

    2013-11-01

    Oocyte is a female gametocyte or germ cell involved in reproduction. Calcium ions (Ca2+) impact nearly all aspects of cellular life as they play an important role in a variety of cellular functions. Calcium ions contributes to egg activation upon fertilization. Since it is the internal stores which provide most of the calcium signal, much attention has been focused on the intracellular channels. There are mainly two types of calcium channels which release calcium from the internal stores to the cytoplasm in many cell types. These channels are IP3-Receptor and Ryanodine Receptor (RyR). Further it is essential to maintain low cytosolic calcium concentration, the cell engages the Serco/Endoplasmic reticulum Ca2+ ATPases (SERCA) present on the ER or SR membrane for the re-uptake of cytosolic calcium at the expense of ATP hydrolysis. In view of above an attempt has been made to study the effect of the Ryanodine receptor (RyR) and the SERCA pump on the calcium distribution in oocytes. The main aim of this paper is to study the calcium concentration in absence and presence of these parameters. The FEM is used to solve the proposed Mathematical model under appreciate initial and boundary conditions. The program has been developed in MATLAB 7.10 for the entire problem to get numerical results.

  13. T-type Ca(2+) channels and Autoregulation of Local Blood Flow

    DEFF Research Database (Denmark)

    Jensen, Lars Jørn; Nielsen, Morten Schak; Salomonsson, Max

    2017-01-01

    L-type voltage gated Ca(2+) channels are considered to be the primary source of calcium influx during the myogenic response. However, many vascular beds also express T-type voltage gated Ca(2+) channels. Recent studies suggest that these channels may also play a role in autoregulation. At low pre...

  14. Voltage gating of mechanosensitive PIEZO channels.

    Science.gov (United States)

    Moroni, Mirko; Servin-Vences, M Rocio; Fleischer, Raluca; Sánchez-Carranza, Oscar; Lewin, Gary R

    2018-03-15

    Mechanosensitive PIEZO ion channels are evolutionarily conserved proteins whose presence is critical for normal physiology in multicellular organisms. Here we show that, in addition to mechanical stimuli, PIEZO channels are also powerfully modulated by voltage and can even switch to a purely voltage-gated mode. Mutations that cause human diseases, such as xerocytosis, profoundly shift voltage sensitivity of PIEZO1 channels toward the resting membrane potential and strongly promote voltage gating. Voltage modulation may be explained by the presence of an inactivation gate in the pore, the opening of which is promoted by outward permeation. Older invertebrate (fly) and vertebrate (fish) PIEZO proteins are also voltage sensitive, but voltage gating is a much more prominent feature of these older channels. We propose that the voltage sensitivity of PIEZO channels is a deep property co-opted to add a regulatory mechanism for PIEZO activation in widely different cellular contexts.

  15. Inactivation of pathogenic bacteria in food matrices: high pressure processing, photodynamic inactivation and pressure-assisted photodynamic inactivation

    Science.gov (United States)

    Cunha, A.; Couceiro, J.; Bonifácio, D.; Martins, C.; Almeida, A.; Neves, M. G. P. M. S.; Faustino, M. A. F.; Saraiva, J. A.

    2017-09-01

    Traditional food processing methods frequently depend on the application of high temperature. However, heat may cause undesirable changes in food properties and often has a negative impact on nutritional value and organoleptic characteristics. Therefore, reducing the microbial load without compromising the desirable properties of food products is still a technological challenge. High-pressure processing (HPP) can be classified as a cold pasteurization technique, since it is a non-thermal food preservation method that uses hydrostatic pressure to inactivate spoilage microorganisms. At the same time, it increases shelf life and retains the original features of food. Photodynamic inactivation (PDI) is also regarded as promising approach for the decontamination of food matrices. In this case, the inactivation of bacterial cells is achieved by the cytotoxic effects of reactive oxygens species (ROS) produced from the combined interaction of a photosensitizer molecule, light and oxygen. This short review examines some recent developments on the application of HPP and PDI with food-grade photosensitizers for the inactivation of listeriae, taken as a food pathogen model. The results of a proof-of-concept trial of the use of high-pressure as a coadjutant to increase the efficiency of photodynamic inactivation of bacterial endospores is also addressed.

  16. Arctigenin, a Potential Anti-Arrhythmic Agent, Inhibits Aconitine-Induced Arrhythmia by Regulating Multi-Ion Channels

    Directory of Open Access Journals (Sweden)

    Zhenying Zhao

    2013-11-01

    Full Text Available Background/Aims: Arctigenin possesses biological activities, but its underlying mechanisms at the cellular and ion channel levels are not completely understood. Therefore, the present study was designed to identify the anti-arrhythmia effect of arctigenin in vivo, as well as its cellular targets and mechanisms. Methods: A rat arrhythmia model was established via continuous aconitine infusion, and the onset times of ventricular premature contraction, ventricular tachycardia and death were recorded. The Action Potential Duration (APD, sodium current (INa, L-type calcium current (ICa, L and transient outward potassium current (Ito were measured and analysed using a patch-clamp recording technique in normal rat cardiomyocytes and myocytes of arrhythmia aconitine-induced by. Results: Arctigenin significantly delayed the arrhythmia onset in the aconitine-induced rat model. The 50% and 90% repolarisations (APD50 and APD90 were shortened by 100 µM arctigenin; the arctigenin dose also inhibited the prolongation of APD50 and APD90 caused by 1 µM aconitine. Arctigenin inhibited INa and ICa,L and attenuated the aconitine-increased INa and ICa,L by accelerating the activation process and delaying the inactivation process. Arctigenin enhanced Ito by facilitating the activation process and delaying the inactivation process, and recoverd the decreased Ito induced by aconitine. Conclusions: Arctigenin has displayed anti-arrhythmia effects, both in vivo and in vitro. In the context of electrophysiology, INa, ICa, L, and Ito may be multiple targets of arctigenin, leading to its antiarrhythmic effect.

  17. Effect of Skeletal Muscle Na+ Channel Delivered Via a Cell Platform on Cardiac Conduction and Arrhythmia Induction

    NARCIS (Netherlands)

    Boink, Gerard J. J.; Lu, Jia; Driessen, Helen E.; Duan, Lian; Sosunov, Eugene A.; Anyukhovsky, Evgeny P.; Shlapakova, Iryna N.; Lau, David H.; Rosen, Tove S.; Danilo, Peter; Jia, Zhiheng; Ozgen, Nazira; Bobkov, Yevgeniy; Guo, Yuanjian; Brink, Peter R.; Kryukova, Yelena; Robinson, Richard B.; Entcheva, Emilia; Cohen, Ira S.; Rosen, Michael R.

    2012-01-01

    Background-In depolarized myocardial infarct epicardial border zones, the cardiac sodium channel is largely inactivated, contributing to slow conduction and reentry. We have demonstrated that adenoviral delivery of the skeletal muscle Na+ channel (SkM1) to epicardial border zones normalizes

  18. Heavy metal cations permeate the TRPV6 epithelial cation channel.

    Science.gov (United States)

    Kovacs, Gergely; Danko, Tamas; Bergeron, Marc J; Balazs, Bernadett; Suzuki, Yoshiro; Zsembery, Akos; Hediger, Matthias A

    2011-01-01

    TRPV6 belongs to the vanilloid family of the transient receptor potential channel (TRP) superfamily. This calcium-selective channel is highly expressed in the duodenum and the placenta, being responsible for calcium absorption in the body and fetus. Previous observations have suggested that TRPV6 is not only permeable to calcium but also to other divalent cations in epithelial tissues. In this study, we tested whether TRPV6 is indeed also permeable to cations such as zinc and cadmium. We found that the basal intracellular calcium concentration was higher in HEK293 cells transfected with hTRPV6 than in non-transfected cells, and that this difference almost disappeared in nominally calcium-free solution. Live cell imaging experiments with Fura-2 and NewPort Green DCF showed that overexpression of human TRPV6 increased the permeability for Ca(2+), Ba(2+), Sr(2+), Mn(2+), Zn(2+), Cd(2+), and interestingly also for La(3+) and Gd(3+). These results were confirmed using the patch clamp technique. (45)Ca uptake experiments showed that cadmium, lanthanum and gadolinium were also highly efficient inhibitors of TRPV6-mediated calcium influx at higher micromolar concentrations. Our results suggest that TRPV6 is not only involved in calcium transport but also in the transport of other divalent cations, including heavy metal ions, which may have toxicological implications. Copyright © 2010 Elsevier Ltd. All rights reserved.

  19. Studies of alpha-helicity and intersegmental interactions in voltage-gated Na+ channels: S2D4.

    Directory of Open Access Journals (Sweden)

    Zhongming Ma

    2009-11-01

    Full Text Available Much data, including crystallographic, support structural models of sodium and potassium channels consisting of S1-S4 transmembrane segments (the "voltage-sensing domain" clustered around a central pore-forming region (S5-S6 segments and the intervening loop. Voltage gated sodium channels have four non-identical domains which differentiates them from the homotetrameric potassium channels that form the basis for current structural models. Since potassium and sodium channels also exhibit many different functional characteristics and the fourth domain (D4 of sodium channels differs in function from other domains (D1-D3, we have explored its structure in order to determine whether segments in D4 of sodium channels differ significantly from that determined for potassium channels. We have probed the secondary and tertiary structure and the role of the individual amino acid residues of the S2D4 of Na(v1.4 by employing cysteine-scanning mutagenesis (with tryptophan and glutamine substituted for native cysteine. A Fourier transform power spectrum of perturbations in free energy of steady-state inactivation gating (using midpoint potentials and slopes of Boltzmann equation fits of channel availability, h(infinity-V plots indicates a substantial amount of alpha-helical structure in S2D4 (peak at 106 degrees, alpha-Periodicity Index (alpha-PI of 3.10, This conclusion is supported by alpha-PI values of 3.28 and 2.84 for the perturbations in rate constants of entry into (beta and exit from (alpha fast inactivation at 0 mV for mutant channels relative to WT channels assuming a simple two-state model for transition from the open to inactivated state. The results of cysteine substitution at the two most sensitive sites of the S2D4 alpha-helix (N1382 and E1392C support the existence of electrostatic network interactions between S2 and other transmembrane segments within Na(v1.4D4 similar to but not identical to those proposed for K+ channels.

  20. Playing with Opening and Closing of Heterocycles: Using the Cusmano-Ruccia Reaction to Develop a Novel Class of Oxadiazolothiazinones, Active as Calcium Channel Modulators and P-Glycoprotein Inhibitors

    Directory of Open Access Journals (Sweden)

    Domenico Spinelli

    2014-10-01

    Full Text Available As a result of the ring-into-ring conversion of nitrosoimidazole derivatives, we obtained a molecular scaffold that, when properly decorated, is able to decrease inotropy by blocking L-type calcium channels. Previously, we used this scaffold to develop a quantitative structure-activity relationship (QSAR model, and we used the most potent oxadiazolothiazinone as a template for ligand-based virtual screening. Here, we enlarge the diversity of chemical decorations, present the synthesis and in vitro data for 11 new derivatives, and develop a new 3D-QSAR model with recent in silico techniques. We observed a key role played by the oxadiazolone moiety: given the presence of positively charged calcium ions in the transmembrane channel protein, we hypothesize the formation of a ternary complex between the oxadiazolothiazinone, the Ca2+ ion and the protein. We have supported this hypothesis by means of pharmacophore generation and through the docking of the pharmacophore into a homology model of the protein. We also studied with docking experiments the interaction with a homology model of P-glycoprotein, which is inhibited by this series of molecules, and provided further evidence toward the relevance of this scaffold in biological interactions.

  1. Metaflumizone is a novel sodium channel blocker insecticide.

    Science.gov (United States)

    Salgado, V L; Hayashi, J H

    2007-12-15

    Metaflumizone is a novel semicarbazone insecticide, derived chemically from the pyrazoline sodium channel blocker insecticides (SCBIs) discovered at Philips-Duphar in the early 1970s, but with greatly improved mammalian safety. This paper describes studies confirming that the insecticidal action of metaflumizone is due to the state-dependent blockage of sodium channels. Larvae of the moth Spodoptera eridania injected with metaflumizone became paralyzed, concomitant with blockage of all nerve activity. Furthermore, tonic firing of abdominal stretch receptor organs from Spodoptera frugiperda was blocked by metaflumizone applied in the bath, consistent with the block of voltage-dependent sodium channels. Studies on native sodium channels, in primary-cultured neurons isolated from the CNS of the larvae of the moth Manduca sexta and on Para/TipE sodium channels heterologously expressed in Xenopus (African clawed frog) oocytes, confirmed that metaflumizone blocks sodium channels by binding selectively to the slow-inactivated state, which is characteristic of the SCBIs. The results confirm that metaflumizone is a novel sodium channel blocker insecticide.

  2. Cytoplasmic Domains and Voltage-Dependent Potassium Channel Gating

    Science.gov (United States)

    Barros, Francisco; Domínguez, Pedro; de la Peña, Pilar

    2012-01-01

    The basic architecture of the voltage-dependent K+ channels (Kv channels) corresponds to a transmembrane protein core in which the permeation pore, the voltage-sensing components and the gating machinery (cytoplasmic facing gate and sensor–gate coupler) reside. Usually, large protein tails are attached to this core, hanging toward the inside of the cell. These cytoplasmic regions are essential for normal channel function and, due to their accessibility to the cytoplasmic environment, constitute obvious targets for cell-physiological control of channel behavior. Here we review the present knowledge about the molecular organization of these intracellular channel regions and their role in both setting and controlling Kv voltage-dependent gating properties. This includes the influence that they exert on Kv rapid/N-type inactivation and on activation/deactivation gating of Shaker-like and eag-type Kv channels. Some illustrative examples about the relevance of these cytoplasmic domains determining the possibilities for modulation of Kv channel gating by cellular components are also considered. PMID:22470342

  3. Heparin/heparan sulfates bind to and modulate neuronal L-type (Cav1.2) voltage-dependent Ca2+ channels

    DEFF Research Database (Denmark)

    Garau, Gianpiero; Magotti, Paola; Heine, Martin

    2015-01-01

    Our previous studies revealed that L-type voltage-dependent Ca2+ 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......M), 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...

  4. Calcium and Nuclear Signaling in Prostate Cancer

    Directory of Open Access Journals (Sweden)

    Ivan V. Maly

    2018-04-01

    Full Text Available Recently, there have been a number of developments in the fields of calcium and nuclear signaling that point to new avenues for a more effective diagnosis and treatment of prostate cancer. An example is the discovery of new classes of molecules involved in calcium-regulated nuclear import and nuclear calcium signaling, from the G protein-coupled receptor (GPCR and myosin families. This review surveys the new state of the calcium and nuclear signaling fields with the aim of identifying the unifying themes that hold out promise in the context of the problems presented by prostate cancer. Genomic perturbations, kinase cascades, developmental pathways, and channels and transporters are covered, with an emphasis on nuclear transport and functions. Special attention is paid to the molecular mechanisms behind prostate cancer progression to the malignant forms and the unfavorable response to anti-androgen treatment. The survey leads to some new hypotheses that connect heretofore disparate results and may present a translational interest.

  5. Calcium-channel blockers do not alter the clinical efficacy of clopidogrel after myocardial infarction: a nationwide cohort study

    DEFF Research Database (Denmark)

    Olesen, Jonas B; Gislason, Gunnar H; Charlot, Mette G

    2011-01-01

    Objectives The purpose of this study was to determine the risk of adverse cardiovascular events associated with concomitant use of clopidogrel and calcium-channel blockers (CCBs) in patients with myocardial infarction (MI). Background CCBs inhibit a variety of cytochrome P-450 enzymes, some...... patients treated and not treated with clopidogrel, with a hazard ratio of 1.15 (95% confidence interval [CI]: 1.07 to 1.24) and 1.05 (95% CI: 1.01 to 1.11), respectively. The increased risk was independent of clopidogrel use; the hazard rate ratio was 1.08 (95% CI: 0.99 to 1.18). Analyses of all additional...... adverse end points and propensity score–matched models provided similar results. Conclusions The clinical efficacy of clopidogrel in patients with a recent MI is not modified by concomitant CCB treatment. This potential drug interaction is unlikely to have clinical significance....

  6. A Tour de Force: The Discovery, Properties, and Function of Piezo Channels.

    Science.gov (United States)

    Gottlieb, P A

    2017-01-01

    Mechanical transducers appear throughout cell biology and are used to convert mechanical stress into chemical or electrical signals that allow the cell to respond to environmental changes. In the past six years, a eukaryotic mechanical channel family with two members, Piezo1 and Piezo2, has been identified. Piezo1 was shown to be a cation-selective channel that does not require ancillary proteins for activity. Mouse Piezo1 is large, with over 2500 amino acids, and is not homologous to other ion channels. Both piezo channels have rapid voltage-dependent inactivation with a reversal potential near 0mV. The CryoEm structure of Piezo1 at 4.8Å shows trimer stoichiometry. Since the discovery of the piezo channels, their roles in the physiological response of cells have started to emerge. Significant progress has been made in understanding the intrinsic properties of the channels and how these properties are modulated by cytoskeletal elements. Specific diseases, such as hereditary xerocytosis affecting red blood cells, have mutations in Piezo1 that alter the cell's response to force, typically slowing inactivation and introducing a latency for activation. A number of physiological functions for piezo channels have been identified. These range from sensing the stiffness of surrounding substrate, to the response to light touch, to serotonin release from the gut. This review provides a general overview of the properties and roles of Piezo1 and Piezo2 in eukaryotic mechanotransduction. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Achievement report for fiscal 1988 on research and development of basic technologies for next-generation industry. Researches into organisms for biological device (information processing in organisms); 1988 nendo bio soshi seibutsu chosa kenkyu seika hokokusho. Seibutsu no joho shori

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1998-09-01

    Researches into organisms for their information processing, conducted in fiscal 1988 as part of a 'biological device research and development' project under the research and development system for basic technologies for the next-generation industry, are compiled into an article entitled 'information processing in neurons, and calcium ions.' The level of neuronal activity depends on changes in the level of nerve-membrane ion channel activity. Some kinds of ion channels are chemically activated (or inactivated) within the membrane. The phosphorylation of a protein ion channel is a typical case where an ion channel level is regulated chemically. In the muscarinic response of a neuron, a chemically operated ion channel plays an important roll relative to neuronal functions. In this response, a second messenger system in the neuron transmits extracellular information to the chemically operated ion channel. In this system, a calcium ion is one of important second messengers. (NEDO)

  8. Efficacy and safety of a therapeutic interchange from high-dose calcium channel blockers to a fixed-dose combination of amlodipine/benazepril in patients with moderate-to-severe hypertension.

    Science.gov (United States)

    Hilleman, D E; Reyes, A P; Wurdeman, R L; Faulkner, M

    2001-08-01

    Recent hypertension trials have demonstrated the importance of achieving goal blood pressures to reduce the risk of target organ damage. In patients with moderate to severe hypertension, the use of high-dose monotherapy and/or combinations of drugs are necessary to achieve these goals. Fixed-dose combination products may be useful in these patients by reducing the number of daily doses required to control blood pressure. The objective of the present study was to evaluate the efficacy and safety of a therapeutic interchange between high-dose calcium channel blocker therapy and a fixed-dose combination of amlodipine/ benazepril (Lotrel; Novartis Pharmaceuticals, USA) in patients with moderate to severe hypertension. A total of 75 patients were switched from amlodipine (n = 25), felodipine (n = 25), and nifedipine-GITS (n = 25) to amlodipine/benazepril. Twenty-eight of the 75 patients (37%) were taking either a beta-blocker or a diuretic in addition to the high-dose calcium channel blocker prior to the switch. Blood pressure control, side effects and the cost of the therapeutic interchange were evaluated in the year following the therapeutic interchange. Sixty-six of the 75 (88%) patients were successfully switched with maintenance of blood pressure control and without the development of new dose-limiting side effects. Reasons for treatment failure after the therapeutic interchange included loss of blood pressure control in five patients and the development of new dose-limiting side effects in four patients. These side effects included cough in three patients and rash in one patient. After accounting for differences in drug acquisition cost and costs related to the switch (clinic and emergency room and laboratory tests), a cost savings of $16030 for all 75 patients was realised in the first year. The per patient-per year cost savings was $214. Our data indicate that a therapeutic interchange from selected high-dose calcium channel blockers to a fixed-dose combination

  9. Identification of sodium channel isoforms that mediate action potential firing in lamina I/II spinal cord neurons

    Directory of Open Access Journals (Sweden)

    Smith Paula L

    2011-09-01

    Full Text Available Abstract Background Voltage-gated sodium channels play key roles in acute and chronic pain processing. The molecular, biophysical, and pharmacological properties of sodium channel currents have been extensively studied for peripheral nociceptors while the properties of sodium channel currents in dorsal horn spinal cord neurons remain incompletely understood. Thus far, investigations into the roles of sodium channel function in nociceptive signaling have primarily focused on recombinant channels or peripheral nociceptors. Here, we utilize recordings from lamina I/II neurons withdrawn from the surface of spinal cord slices to systematically determine the functional properties of sodium channels expressed within the superficial dorsal horn. Results Sodium channel currents within lamina I/II neurons exhibited relatively hyperpolarized voltage-dependent properties and fast kinetics of both inactivation and recovery from inactivation, enabling small changes in neuronal membrane potentials to have large effects on intrinsic excitability. By combining biophysical and pharmacological channel properties with quantitative real-time PCR results, we demonstrate that functional sodium channel currents within lamina I/II neurons are predominantly composed of the NaV1.2 and NaV1.3 isoforms. Conclusions Overall, lamina I/II neurons express a unique combination of functional sodium channels that are highly divergent from the sodium channel isoforms found within peripheral nociceptors, creating potentially complementary or distinct ion channel targets for future pain therapeutics.

  10. Activation of calcium-sensing receptor increases TRPC3 expression in rat cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Feng, Shan-Li [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Sun, Ming-Rui [Department of Pharmacology, Qiqihaer Medical College, Qiqihaer 160001 (China); Li, Ting-Ting; Yin, Xin [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Xu, Chang-Qing [Department of Pathophysiology, Harbin Medical University, Harbin 150086 (China); Sun, Yi-Hua, E-mail: syh200415@126.com [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China)

    2011-03-11

    Research highlights: {yields} Calcium-sensing receptor (CaR) activation stimulates TRP channels. {yields} CaR promoted transient receptor potential C3 (TRPC3) expression. {yields} Adult rat ventricular myocytes display capacitative calcium entry (CCE), which was operated by TRPCs. {yields} TRPC channels activation induced by CaR activator sustained the increased [Ca{sup 2+}]{sub i} to evoke cardiomyocytes apoptosis. -- Abstract: Transient receptor potential (TRP) channels are expressed in cardiomyocytes, which gate a type of influx of extracellular calcium, the capacitative calcium entry. TRP channels play a role in mediating Ca{sup 2+} overload in the heart. Calcium-sensing receptors (CaR) are also expressed in rat cardiac tissue and promote the apoptosis of cardiomyocytes by Ca{sup 2+} overload. However, data about the link between CaR and TRP channels in rat heart are few. In this study, reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting were used to examine the expression of the TRP canonical proteins TRPC1 and TRPC3 in adult and neonatal rat cardiomyocytes. Laser scan confocal microscopy was used to detect intracellular [Ca{sup 2+}]{sub i} levels in isolated adult rat ventricular myocytes. The results showed that, in adult rat cardiomyocytes, the depletion of Ca{sup 2+} stores in the endoplasmic/sarcoplasmic reticulum (ER/SR) by thapsigargin induced a transient increase in [Ca{sup 2+}]{sub i} in the absence of [Ca{sup 2+}]{sub o} and the subsequent restoration of [Ca{sup 2+}]{sub o} sustained the increased [Ca{sup 2+}]{sub i} for a few minutes, whereas, the persisting elevation of [Ca{sup 2+}]{sub i} was reduced in the presence of the TRPC inhibitor SKF96365. The stimulation of CaR by its activator gadolinium chloride (GdCl{sub 3}) or spermine also resulted in the same effect and the duration of [Ca{sup 2+}]{sub i} increase was also shortened in the absence of [Ca{sup 2+}]{sub o}. In adult and neonatal rat cardiomyocytes, GdCl{sub 3

  11. Cav2.3 (R-Type Calcium Channels are Critical for Mediating Anticonvulsive and Neuroprotective Properties of Lamotrigine In Vivo

    Directory of Open Access Journals (Sweden)

    Maxine Dibué-Adjei

    2017-11-01

    Full Text Available Background/Aims: Lamotrigine (LTG is a popular modern antiepileptic drug (AED, however, its mechanism of action has yet to be fully understood, as it is known to modulate many members of several ion channel families. In heterologous systems, LTG inhibits Cav2.3 (R-type calcium currents, which contribute to kainic-acid- (KA induced epilepsy in vivo. To gain insight into the role of R-type currents in LTG drug action in vivo, we compared the effects of LTG to topiramate and lacosamide in Cav2.3-deficient mice and controls on KA-induced seizures. Methods: Behavioral seizure rating and quantitative electrocorticography were performed after injection of 20 mg/kg [and 30 mg/kg] KA. One hour before KA injection, mice were pretreated with either 30 mg/kg LTG, 50 mg/kg topiramate (TPM or 30 mg/kg lacosamide (LSM. Results: Ablation of Cav2.3 reduced total seizure scores by 28.6% (p=0.0012 and pretreatment with LTG reduced seizure activity of control mice by 23.2% (p=0.02. In Cav2.3-deficient mice LTG pretreatment increased seizure activity by 22.1% (p=0.018 and increased the percentage of degenerated CA1 pyramidal neurons (p=0.02. All three tested AEDs reduced seizure activity in control mice, however only the non-calcium channel modulating AED, LSM had an anticonvulsive effect in Cav2.3-deficient mice. Furthermore LTG altered electrocorticographic parameters differently in the two genotypes, decreasing relative power of ictal spikes in control mice compared to Cav2.3-defcient mice. Conclusion: These findings give first in vivo evidence for an essential role for Cav2.3 in LTG pharmacology and shed light on a paradoxical effect of LTG in their absence. Furthermore, LTG appears to promote ictal activity in Cav2.3-deficient mice resulting in increased neurotoxicity in the CA1 region. This paradoxical mechanism, possibly reflecting rebound hyperexcitation of pyramidal CA1 neurons after increased inhibition, may be key in understanding LTG-induced seizure