Sample records for cav2 calcium channel

  1. RGS12 interacts with the SNARE-binding region of the Cav2.2 calcium channel. (United States)

    Richman, Ryan W; Strock, Jesse; Hains, Melinda D; Cabanilla, Nory Jun; Lau, King-Kei; Siderovski, David P; Diversé-Pierluissi, María


    Activation of GABAB receptors in chick dorsal root ganglion (DRG) neurons inhibits the Cav2.2 calcium channel in both a voltage-dependent and voltage-independent manner. The voltage-independent inhibition requires activation of a tyrosine kinase that phosphorylates the alpha1 subunit of the channel and thereby recruits RGS12, a member of the "regulator of G protein signaling" (RGS) proteins. Here we report that RGS12 binds to the SNARE-binding or "synprint" region (amino acids 726-985) in loop II-III of the calcium channel alpha1 subunit. A recombinant protein encompassing the N-terminal PTB domain of RGS12 binds to the synprint region in protein overlay and surface plasmon resonance binding assays; this interaction is dependent on tyrosine phosphorylation and yet is within a sequence that differs from the canonical NPXY motif targeted by other PTB domains. In electrophysiological experiments, microinjection of DRG neurons with synprint-derived peptides containing the tyrosine residue Tyr-804 altered the rate of desensitization of neurotransmitter-mediated inhibition of the Cav2.2 calcium channel, whereas peptides centered about a second tyrosine residue, Tyr-815, were without effect. RGS12 from a DRG neuron lysate was precipitated using synprint peptides containing phosphorylated Tyr-804. The high degree of conservation of Tyr-804 in the SNARE-binding region of Cav2.1 and Cav2.2 calcium channels suggests that this region, in addition to the binding of SNARE proteins, is also important for determining the time course of the modulation of calcium current via tyrosine phosphorylation.

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

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    Lei Zhu

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

  3. Cav2-type calcium channels encoded by cac regulate AP-independent neurotransmitter release at cholinergic synapses in adult Drosophila brain. (United States)

    Gu, Huaiyu; Jiang, Shaojuan Amy; Campusano, Jorge M; Iniguez, Jorge; Su, Hailing; Hoang, Andy An; Lavian, Monica; Sun, Xicui; O'Dowd, Diane K


    Voltage-gated calcium channels containing alpha1 subunits encoded by Ca(v)2 family genes are critical in regulating release of neurotransmitter at chemical synapses. In Drosophila, cac is the only Ca(v)2-type gene. Cacophony (CAC) channels are localized in motor neuron terminals where they have been shown to mediate evoked, but not AP-independent, release of glutamate at the larval neuromuscular junction (NMJ). Cultured embryonic neurons also express CAC channels, but there is no information about the properties of CAC-mediated currents in adult brain nor how these channels regulate transmission in central neural circuits where fast excitatory synaptic transmission is predominantly cholinergic. Here we report that wild-type neurons cultured from late stage pupal brains and antennal lobe projection neurons (PNs) examined in adult brains, express calcium currents with two components: a slow-inactivating current sensitive to the spider toxin Plectreurys toxin II (PLTXII) and a fast-inactivating PLTXII-resistant component. CAC channels are the major contributors to the slow-inactivating PLTXII-sensitive current based on selective reduction of this component in hypomorphic cac mutants (NT27 and TS3). Another characteristic of cac mutant neurons both in culture and in whole brain recordings is a reduced cholinergic miniature excitatory postsynaptic current frequency that is mimicked in wild-type neurons by acute application of PLTXII. These data demonstrate that cac encoded Ca(v)2-type calcium channels regulate action potential (AP)-independent release of neurotransmitter at excitatory cholinergic synapses in the adult brain, a function not predicted from studies at the larval NMJ.

  4. Protein Kinase Cα and P-Type Ca2+ Channel CaV2.1 in Red Blood Cell Calcium Signalling

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    Lisa Wagner-Britz


    Full Text Available Background/Aims: Protein kinase Cα (PKCα is activated by an increase in cytosolic Ca2+ in red blood cells (RBCs. Previous work has suggested that PKCα directly stimulates the CaV2.1 channel, whereas other studies revealed that CaV2.1 is insensitive to activation by PKC. The aim of this study was to resolve this discrepancy. Methods: We performed experiments based on a single cell read-out of the intracellular Ca2+ concentration in terms of Fluo-4 fluorescence intensity and phosphatidylserine exposure to the external membrane leaflet. Measurement modalities included flow cytometry and live cell imaging. Results: Treatment of RBCs with phorbol 12-myristate 13-acetate (PMA led to two distinct populations of cells with an increase in intracellular Ca2+: a weak-responding and a strong-responding population. The EC50 of PMA for the number of cells with Ca2+ elevation was 2.7±1.2 µM; for phosphatidylserine exposure to the external membrane surface, it was 2.8±0.5 µM; and for RBC haemolysis, it was 2.9±0.5 µM. Using pharmacological manipulation with the CaV2.1 inhibitor ω-agatoxin TK and the broad protein kinase C inhibitor Gö6983, we are able to show that there are two independent PMA-activated Ca2+ entry processes: the first is independent of CaV2.1 and directly PKCα-activated, while the second is associated with a likely indirect activation of CaV2.1. Further studies using lysophosphatidic acid (LPA as a stimulation agent have provided additional evidence that PKCα and CaV2.1 are not directly interconnected in a signalling chain. Conclusion: Although we provide evidence for a lack of interaction between PKCα and CaV2.1 in RBCs, further studies are required to decipher the signalling relationship between LPA, PKCα and CaV2.1.

  5. A novel CaV2.2 channel inhibition by piracetam in peripheral and central neurons. (United States)

    Bravo-Martínez, Jorge; Arenas, Isabel; Vivas, Oscar; Rebolledo-Antúnez, Santiago; Vázquez-García, Mario; Larrazolo, Arturo; García, David E


    No mechanistic actions for piracetam have been documented to support its nootropic effects. Voltage-gated calcium channels have been proposed as a promising pharmacological target of nootropic drugs. In this study, we investigated the effect of piracetam on Ca(V)2.2 channels in peripheral neurons, using patch-clamp recordings from cultured superior cervical ganglion neurons. In addition, we tested if Ca(V)2.2 channel inhibition could be related with the effects of piracetam on central neurons. We found that piracetam inhibited native Ca(V)2.2 channels in superior cervical ganglion neurons in a dose-dependent manner, with an IC(50) of 3.4 μmol/L and a Hill coefficient of 1.1. GDPβS dialysis did not prevent piracetam-induced inhibition of Ca(V)2.2 channels and G-protein-coupled receptor activation by noradrenaline did not occlude the piracetam effect. Piracetam altered the biophysical characteristics of Ca(V)2.2 channel such as facilitation ratio. In hippocampal slices, piracetam and ω-conotoxin GVIA diminished the frequency of excitatory postsynaptic potentials and action potentials. Our results provide evidence of piracetam's actions on Ca(V)2.2 channels in peripheral neurons, which might explain some of its nootropic effects in central neurons.

  6. Voltage-dependent regulation of CaV2.2 channels by Gq-coupled receptor is facilitated by membrane-localized β subunit. (United States)

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


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

  7. Familial hemiplegic migraine type 1 mutations W1684R and V1696I alter G protein-mediated regulation of Ca(V)2.1 voltage-gated calcium channels. (United States)

    Garza-López, Edgar; Sandoval, Alejandro; González-Ramírez, Ricardo; Gandini, María A; Van den Maagdenberg, Arn; De Waard, Michel; Felix, Ricardo


    Familial hemiplegic migraine type 1 (FHM-1) is a monogenic form of migraine with aura that is characterized by recurrent attacks of a typical migraine headache with transient hemiparesis during the aura phase. In a subset of patients, additional symptoms such as epilepsy and cerebellar ataxia are part of the clinical phenotype. FHM-1 is caused by missense mutations in the CACNA1A gene that encodes the pore-forming subunit of Ca(V)2.1 voltage-gated Ca(2+) channels. Although the functional effects of an increasing number of FHM-1 mutations have been characterized, knowledge on the influence of most of these mutations on G protein regulation of channel function is lacking. Here, we explored the effects of G protein-dependent modulation on mutations W1684R and V1696I which cause FHM-1 with and without cerebellar ataxia, respectively. Both mutations were introduced into the human Ca(V)2.1α(1) subunit and their functional consequences investigated after heterologous expression in human embryonic kidney 293 (HEK-293) cells using patch-clamp recordings. When co-expressed along with the human μ-opioid receptor, application of the agonist [d-Ala2, N-MePhe4, Gly-ol]-enkephalin (DAMGO) inhibited currents through both wild-type (WT) and mutant Ca(V)2.1 channels, which is consistent with the known modulation of these channels by G protein-coupled receptors. Prepulse facilitation, which is a way to characterize the relief of direct voltage-dependent G protein regulation, was reduced by both FHM-1 mutations. Moreover, the kinetic analysis of the onset and decay of facilitation showed that the W1684R and V1696I mutations affect the apparent dissociation and reassociation rates of the Gβγ dimer from the channel complex, suggesting that the G protein-Ca(2+) channel affinity may be altered by the mutations. These biophysical studies may shed new light on the pathophysiology underlying FHM-1.

  8. Purkinje cell-specific ablation of Cav2.1 channels is sufficient to cause cerebellar ataxia in mice. (United States)

    Todorov, Boyan; Kros, Lieke; Shyti, Reinald; Plak, Petra; Haasdijk, Elize D; Raike, Robert S; Frants, Rune R; Hess, Ellen J; Hoebeek, Freek E; De Zeeuw, Chris I; van den Maagdenberg, Arn M J M


    The Cacna1a gene encodes the α(1A) subunit of voltage-gated Ca(V)2.1 Ca(2+) channels that are involved in neurotransmission at central synapses. Ca(V)2.1-α(1)-knockout (α1KO) mice, which lack Ca(V)2.1 channels in all neurons, have a very severe phenotype of cerebellar ataxia and dystonia, and usually die around postnatal day 20. This early lethality, combined with the wide expression of Ca(V)2.1 channels throughout the cerebellar cortex and nuclei, prohibited determination of the contribution of particular cerebellar cell types to the development of the severe neurobiological phenotype in Cacna1a mutant mice. Here, we crossed conditional Cacna1a mice with transgenic mice expressing Cre recombinase, driven by the Purkinje cell-specific Pcp2 promoter, to specifically ablate the Ca(V)2.1-α(1A) subunit and thereby Ca(V)2.1 channels in Purkinje cells. Purkinje cell Ca(V)2.1-α(1A)-knockout (PCα1KO) mice aged without difficulties, rescuing the lethal phenotype seen in α1KO mice. PCα1KO mice exhibited cerebellar ataxia starting around P12, much earlier than the first signs of progressive Purkinje cell loss, which appears in these mice between P30 and P45. Secondary cell loss was observed in the granular and molecular layers of the cerebellum and the volume of all individual cerebellar nuclei was reduced. In this mouse model with a cell type-specific ablation of Ca(V)2.1 channels, we show that ablation of Ca(V)2.1 channels restricted to Purkinje cells is sufficient to cause cerebellar ataxia. We demonstrate that spatial ablation of Ca(V)2.1 channels may help in unraveling mechanisms of human disease.

  9. Suppression of peripheral pain by blockade of Cav2.2 channels in nociceptors induces RANKL and impairs recovery from inflammatory arthritis (United States)

    Baddack, Uta; Frahm, Silke; Antolin-Fontes, Beatriz; Grobe, Jenny; Lipp, Martin; Müller, Gerd; Ibañez-Tallon, Ines


    Objective A hallmark of rheumatoid arthritis (RA) is the chronic pain that accompanies the inflammation and joint deformation. Patients with RA rate pain relief with highest priority, however, few studies have addressed the efficacy and safety of therapies directed specifically towards pain pathways. The conotoxin MVIIA (Prialt/Ziconotide) is used in humans to alleviate persistent pain syndromes because it specifically blocks the CaV2.2 voltage-gated calcium channel, which mediates the release of neurotransmitters and proinflammatory mediators from peripheral nociceptor nerve terminals. The purpose of this study was to investigate whether block of CaV2.2 can suppress arthritic pain, and to examine the progression of induced arthritis during persistent CaV2.2 blockade. Methods Transgenic mice (Tg-MVIIA) expressing a membrane-tethered form of the ω-conotoxin MVIIA, under the control of a nociceptor-specific gene, were employed. These mice were subjected to unilateral induction of joint inflammation using the Antigen- and Collagen-Induced Arthritis (ACIA) model. Results We observed that CaV2.2-blockade mediated by t-MVIIA effectively suppressed arthritis-induced pain; however, in contrast to their wild-type littermates, which ultimately regained use of their injured joint as inflammation subsides, Tg-MVIIA mice showed continued inflammation with an up-regulation of the osteoclast activator RANKL and concomitant joint and bone destruction. Conclusion Altogether, our results indicate that alleviation of peripheral pain by blockade of CaV2.2- mediated calcium influx and signaling in nociceptor sensory neurons, impairs recovery from induced arthritis and point to the potentially devastating effects of using CaV2.2 channel blockers as analgesics during inflammation. PMID:25733371

  10. Apamin Boosting of Synaptic Potentials in CaV2.3 R-Type Ca2+ Channel Null Mice. (United States)

    Wang, Kang; Kelley, Melissa H; Wu, Wendy W; Adelman, John P; Maylie, James


    SK2- and KV4.2-containing K+ channels modulate evoked synaptic potentials in CA1 pyramidal neurons. Each is coupled to a distinct Ca2+ source that provides Ca2+-dependent feedback regulation to limit AMPA receptor (AMPAR)- and NMDA receptor (NMDAR)-mediated postsynaptic depolarization. SK2-containing channels are activated by Ca2+ entry through NMDARs, whereas KV4.2-containing channel availability is increased by Ca2+ entry through SNX-482 (SNX) sensitive CaV2.3 R-type Ca2+ channels. Recent studies have challenged the functional coupling between NMDARs and SK2-containing channels, suggesting that synaptic SK2-containing channels are instead activated by Ca2+ entry through R-type Ca2+ channels. Furthermore, SNX has been implicated to have off target affects, which would challenge the proposed coupling between R-type Ca2+ channels and KV4.2-containing K+ channels. To reconcile these conflicting results, we evaluated the effect of SK channel blocker apamin and R-type Ca2+ channel blocker SNX on evoked excitatory postsynaptic potentials (EPSPs) in CA1 pyramidal neurons from CaV2.3 null mice. The results show that in the absence of CaV2.3 channels, apamin application still boosted EPSPs. The boosting effect of CaV2.3 channel blockers on EPSPs observed in neurons from wild type mice was not observed in neurons from CaV2.3 null mice. These data are consistent with a model in which SK2-containing channels are functionally coupled to NMDARs and KV4.2-containing channels to CaV2.3 channels to provide negative feedback regulation of EPSPs in the spines of CA1 pyramidal neurons.

  11. N-type calcium current, Cav2.2, is enhanced in small-diameter sensory neurons isolated from Nf1+/- mice. (United States)

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


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

  12. Ablation of Ca(V2.1 voltage-gated Ca²⁺ channels in mouse forebrain generates multiple cognitive impairments.

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    Robert Theodor Mallmann

    Full Text Available Voltage-gated Ca(V2.1 (P/Q-type Ca²⁺ channels located at the presynaptic membrane are known to control a multitude of Ca²⁺-dependent cellular processes such as neurotransmitter release and synaptic plasticity. Our knowledge about their contributions to complex cognitive functions, however, is restricted by the limited adequacy of existing transgenic Ca(V2.1 mouse models. Global Ca(V2.1 knock-out mice lacking the α1 subunit Cacna1a gene product exhibit early postnatal lethality which makes them unsuitable to analyse the relevance of Ca(V2.1 Ca²⁺ channels for complex behaviour in adult mice. Consequently we established a forebrain specific Ca(V2.1 knock-out model by crossing mice with a floxed Cacna1a gene with mice expressing Cre-recombinase under the control of the NEX promoter. This novel mouse model enabled us to investigate the contribution of Ca(V2.1 to complex cognitive functions, particularly learning and memory. Electrophysiological analysis allowed us to test the specificity of our conditional knock-out model and revealed an impaired synaptic transmission at hippocampal glutamatergic synapses. At the behavioural level, the forebrain-specific Ca(V2.1 knock-out resulted in deficits in spatial learning and reference memory, reduced recognition memory, increased exploratory behaviour and a strong attenuation of circadian rhythmicity. In summary, we present a novel conditional Ca(V2.1 knock-out model that is most suitable for analysing the in vivo functions of Ca(V2.1 in the adult murine forebrain.

  13. Ca(V)1 and Ca(V)2 channels engage distinct modes of Ca(2+) signaling to control CREB-dependent gene expression. (United States)

    Wheeler, Damian G; Groth, Rachel D; Ma, Huan; Barrett, Curtis F; Owen, Scott F; Safa, Parsa; Tsien, Richard W


    Activity-dependent gene expression triggered by Ca(2+) entry into neurons is critical for learning and memory, but whether specific sources of Ca(2+) act distinctly or merely supply Ca(2+) to a common pool remains uncertain. Here, we report that both signaling modes coexist and pertain to Ca(V)1 and Ca(V)2 channels, respectively, coupling membrane depolarization to CREB phosphorylation and gene expression. Ca(V)1 channels are advantaged in their voltage-dependent gating and use nanodomain Ca(2+) to drive local CaMKII aggregation and trigger communication with the nucleus. In contrast, Ca(V)2 channels must elevate [Ca(2+)](i) microns away and promote CaMKII aggregation at Ca(V)1 channels. Consequently, Ca(V)2 channels are ~10-fold less effective in signaling to the nucleus than are Ca(V)1 channels for the same bulk [Ca(2+)](i) increase. Furthermore, Ca(V)2-mediated Ca(2+) rises are preferentially curbed by uptake into the endoplasmic reticulum and mitochondria. This source-biased buffering limits the spatial spread of Ca(2+), further attenuating Ca(V)2-mediated gene expression.

  14. Calcium channels, neuromuscular synaptic transmission and neurological diseases. (United States)

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


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

  15. Voltage-independent inhibition of Cav2.2 channels is delimited to a specific region of the membrane potential in rat SCG neurons

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    Oscar Vivas; Isabel Arenas; David E.García


    Neurotransmitters and hormones regulate Cav2.2 channels through a voltage-independent pathway which is not well understood.It has been suggested that this voltageindependent inhibition is constant at all membrane voltages.However,changes in the percent of voltageindependent inhibition of Cav2.2 have not been tested within a physiological voltage range.Here,we used a double-pulse protocol to isolate the voltage-independent inhibition of Cav2.2 channels induced by noradrenaline in rat superior cervical ganglion neurons.To assess changes in the percent of the voltage-independent inhibition,the activation voltage of the channels was tested between -40 and +40 mV.We found that the percent of voltage-independent inhibition induced by noradrenaline changed with the activation voltage used.In addition,voltage-independent inhibition induced by oxo-M,a muscarinic agonist,exhibited thesame dependence on activation voltage,which supports that this pattern is not exclusive for adrenergic activation.Our results suggested that voltage-independent inhibition of Cav2.2 channels depends on the activation voltage of the channel in a physiological voltage range.This may have relevant implications in the understanding of the mechanism involved in voltage-independent inhibition.

  16. Regulation of neuronal high-voltage activated Ca(V)2 Ca(2+) channels by the small GTPase RhoA. (United States)

    Rousset, Matthieu; Cens, Thierry; Menard, Claudine; Bowerman, Melissa; Bellis, Michel; Brusés, Juan; Raoul, Cedric; Scamps, Frédérique; Charnet, Pierre


    High-Voltage-Activated (HVA) Ca(2+) channels are known regulators of synapse formation and transmission and play fundamental roles in neuronal pathophysiology. Small GTPases of Rho and RGK families, via their action on both cytoskeleton and Ca(2+) channels are key molecules for these processes. While the effects of RGK GTPases on neuronal HVA Ca(2+) channels have been widely studied, the effects of RhoA on the HVA channels remains however elusive. Using heterologous expression in Xenopus laevis oocytes, we show that RhoA activity reduces Ba(2+) currents through CaV2.1, CaV2.2 and CaV2.3 Ca(2+) channels independently of CaVβ subunit. This inhibition occurs independently of RGKs activity and without modification of biophysical properties and global level of expression of the channel subunit. Instead, we observed a marked decrease in the number of active channels at the plasma membrane. Pharmacological and expression studies suggest that channel expression at the plasma membrane is impaired via a ROCK-sensitive pathway. Expression of constitutively active RhoA in primary culture of spinal motoneurons also drastically reduced HVA Ca(2+) current amplitude. Altogether our data revealed that HVA Ca(2+) channels regulation by RhoA might govern synaptic transmission during development and potentially contribute to pathophysiological processes when axon regeneration and growth cone kinetics are impaired.

  17. N-type calcium channel blockers: novel therapeutics for the treatment of pain. (United States)

    Schroeder, C I; Doering, C J; Zamponi, G W; Lewis, R J


    Highly selective Ca(v)2.2 voltage-gated calcium channel (VGCC) inhibitors have emerged as a new class of therapeutics for the treatment of chronic and neuropathic pain. Cone snail venoms provided the first drug in class with FDA approval granted in 2005 to Prialt (omega-conotoxin MVIIA, Elan) for the treatment of neuropathic pain. Since this pioneering work, major efforts underway to develop alternative small molecule inhibitors of Ca(v)2.2 calcium channel have met with varied success. This review focuses on the properties of the Ca(v)2.2 calcium channel in different pain states, the action of omega-conotoxins GVIA, MVIIA and CVID, describing their structure-activity relationships and potential as leads for the design of improved Ca(v)2.2 calcium channel therapeutics, and finally the development of small molecules for the treatment of chronic pain.

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

    DEFF Research Database (Denmark)

    Hansen, Pernille B L


    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 o...

  19. A Single Amino Acid Deletion (ΔF1502 in the S6 Segment of CaV2.1 Domain III Associated with Congenital Ataxia Increases Channel Activity and Promotes Ca2+ Influx.

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    Maria Isabel Bahamonde

    Full Text Available Mutations in the CACNA1A gene, encoding the pore-forming CaV2.1 (P/Q-type channel α1A subunit, result in heterogeneous human neurological disorders, including familial and sporadic hemiplegic migraine along with episodic and progressive forms of ataxia. Hemiplegic Migraine (HM mutations induce gain-of-channel function, mainly by shifting channel activation to lower voltages, whereas ataxia mutations mostly produce loss-of-channel function. However, some HM-linked gain-of-function mutations are also associated to congenital ataxia and/or cerebellar atrophy, including the deletion of a highly conserved phenylalanine located at the S6 pore region of α1A domain III (ΔF1502. Functional studies of ΔF1502 CaV2.1 channels, expressed in Xenopus oocytes, using the non-physiological Ba2+ as the charge carrier have only revealed discrete alterations in channel function of unclear pathophysiological relevance. Here, we report a second case of congenital ataxia linked to the ΔF1502 α1A mutation, detected by whole-exome sequencing, and analyze its functional consequences on CaV2.1 human channels heterologously expressed in mammalian tsA-201 HEK cells, using the physiological permeant ion Ca2+. ΔF1502 strongly decreases the voltage threshold for channel activation (by ~ 21 mV, allowing significantly higher Ca2+ current densities in a range of depolarized voltages with physiological relevance in neurons, even though maximal Ca2+ current density through ΔF1502 CaV2.1 channels is 60% lower than through wild-type channels. ΔF1502 accelerates activation kinetics and slows deactivation kinetics of CaV2.1 within a wide range of voltage depolarization. ΔF1502 also slowed CaV2.1 inactivation kinetic and shifted the inactivation curve to hyperpolarized potentials (by ~ 28 mV. ΔF1502 effects on CaV2.1 activation and deactivation properties seem to be of high physiological relevance. Thus, ΔF1502 strongly promotes Ca2+ influx in response to either single or

  20. Inhibition of N-Type Calcium Channels by Fluorophenoxyanilide Derivatives

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    Ellen C. Gleeson


    Full Text Available A set of fluorophenoxyanilides, designed to be simplified analogues of previously reported ω-conotoxin GVIA mimetics, were prepared and tested for N-type calcium channel inhibition in a SH-SY5Y neuroblastoma FLIPR assay. N-type or Cav2.2 channel is a validated target for the treatment of refractory chronic pain. Despite being significantly less complex than the originally designed mimetics, up to a seven-fold improvement in activity was observed.

  1. Voltage-Gated Calcium Channels in Nociception (United States)

    Yasuda, Takahiro; Adams, David J.

    Voltage-gated calcium channels (VGCCs) are a large and functionally diverse group of membrane ion channels ubiquitously expressed throughout the central and peripheral nervous systems. VGCCs contribute to various physiological processes and transduce electrical activity into other cellular functions. This chapter provides an overview of biophysical properties of VGCCs, including regulation by auxiliary subunits, and their physiological role in neuronal functions. Subsequently, then we focus on N-type calcium (Cav2.2) channels, in particular their diversity and specific antagonists. We also discuss the role of N-type calcium channels in nociception and pain transmission through primary sensory dorsal root ganglion neurons (nociceptors). It has been shown that these channels are expressed predominantly in nerve terminals of the nociceptors and that they control neurotransmitter release. To date, important roles of N-type calcium channels in pain sensation have been elucidated genetically and pharmacologically, indicating that specific N-type calcium channel antagonists or modulators are particularly useful as therapeutic drugs targeting chronic and neuropathic pain.

  2. CACNA1B (Cav2.2 Overexpression and Its Association with Clinicopathologic Characteristics and Unfavorable Prognosis in Non-Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Xiaoyu Zhou


    Full Text Available CACNA1B (Cav2.2 encodes an N-type voltage-gated calcium channel (VGCC ubiquitously expressed in brain and peripheral nervous system that is important for regulating neuropathic pain. Because intracellular calcium concentration is a key player in cell proliferation and apoptosis, VGCCs are implicated in tumorigenesis. Recent studies have identified CACNA1B (Cav2.2 being overexpressed in prostate and breast cancer tissues when compared to adjacent normal tissues; however, its role in non-small cell lung cancer (NSCLC has not been investigated. In this study, we determined the mRNA and protein expression of CACNA1B (Cav2.2 in NSCLC tumorous and adjacent nontumorous tissues by quantitative reverse transcription PCR (qRT-PCR and tissue microarray immunohistochemistry analysis (TMA-IHC, respectively. CACNA1B (Cav2.2 protein expressions in tumorous tissues were correlated with NSCLC patients’ clinical characteristics and overall survival. CACNA1B (Cav2.2 mRNA and protein expression levels were higher in NSCLC tumorous tissues than in nontumorous tissues. High CACNA1B (Cav2.2 protein expression was associated with higher TNM stages, and CACNA1B (Cav2.2 protein expression is an independent prognostic marker in NSCLC. Based on our results, we conclude that CACNA1B (Cav2.2 plays a role in NSCLC development and progression. Elucidating the underlying mechanism may help design novel treatment by specifically targeting the calcium regulation pathway for NSCLC, a devastating disease with increasing incidence and mortality in China.

  3. Alternative splicing generates a smaller assortment of CaV2.1 transcripts in cerebellar Purkinje cells than in the cerebellum. (United States)

    Kanumilli, Srinivasan; Tringham, Elizabeth W; Payne, C Elizabeth; Dupere, Jonathan R B; Venkateswarlu, Kanamarlapudi; Usowicz, Maria M


    P/Q-type calcium channels control many calcium-driven functions in the brain. The CACNA1A gene encoding the pore-forming CaV2.1 (alpha1A) subunit of P/Q-type channels undergoes alternative splicing at multiple loci. This results in channel variants with different phenotypes. However, the combinatorial patterns of alternative splice events at two or more loci, and hence the diversity of CaV2.1 transcripts, are incompletely defined for specific brain regions and types of brain neurons. Using RT-PCR and splice variant-specific primers, we have identified multiple CaV2.1 transcript variants defined by different pairs of splice events in the cerebellum of adult rat. We have uncovered new splice variations between exons 28 and 34 (some of which predict a premature stop codon) and a new variation in exon 47 (which predicts a novel extended COOH-terminus). Single cell RT-PCR reveals that each individual cerebellar Purkinje neuron also expresses multiple alternative CaV2.1 transcripts, but the assortment is smaller than in the cerebellum. Two of these variants encode different extended COOH-termini which are not the same as those previously reported in Purkinje cells of the mouse. Our patch-clamp recordings show that calcium channel currents in the soma and dendrites of Purkinje cells are largely inhibited by a concentration of omega-agatoxin IVA selective for P-type over Q-type channels, suggesting that the different transcripts may form phenotypic variants of P-type calcium channels in Purkinje cells. These results expand the known diversity of CaV2.1 transcripts in cerebellar Purkinje cells, and propose the selective expression of distinct assortments of CaV2.1 transcripts in different brain neurons and species.

  4. Voltage-Gated R-Type Calcium Channel Inhibition via Human μ-, δ-, and κ-opioid Receptors Is Voltage-Independently Mediated by Gβγ Protein Subunits. (United States)

    Berecki, Géza; Motin, Leonid; Adams, David J


    Elucidating the mechanisms that modulate calcium channels via opioid receptor activation is fundamental to our understanding of both pain perception and how opioids modulate pain. Neuronal voltage-gated N-type calcium channels (Cav2.2) are inhibited by activation of G protein-coupled opioid receptors (ORs). However, inhibition of R-type (Cav2.3) channels by μ- or κ-ORs is poorly defined and has not been reported for δ-ORs. To investigate such interactions, we coexpressed human μ-, δ-, or κ-ORs with human Cav2.3 or Cav2.2 in human embryonic kidney 293 cells and measured depolarization-activated Ba(2+) currents (IBa). Selective agonists of μ-, δ-, and κ-ORs inhibited IBa through Cav2.3 channels by 35%. Cav2.2 channels were inhibited to a similar extent by κ-ORs, but more potently (60%) via μ- and δ-ORs. Antagonists of δ- and κ-ORs potentiated IBa amplitude mediated by Cav2.3 and Cav2.2 channels. Consistent with G protein βγ (Gβγ) interaction, modulation of Cav2.2 was primarily voltage-dependent and transiently relieved by depolarizing prepulses. In contrast, Cav2.3 modulation was voltage-independent and unaffected by depolarizing prepulses. However, Cav2.3 inhibition was sensitive to pertussis toxin and to intracellular application of guanosine 5'-[β-thio]diphosphate trilithium salt and guanosine 5'-[γ-thio]triphosphate tetralithium salt. Coexpression of Gβγ-specific scavengers-namely, the carboxyl terminus of the G protein-coupled receptor kinase 2 or membrane-targeted myristoylated-phosducin-attenuated or abolished Cav2.3 modulation. Our study reveals the diversity of OR-mediated signaling at Cav2 channels and identifies neuronal Cav2.3 channels as potential targets for opioid analgesics. Their novel modulation is dependent on pre-existing OR activity and mediated by membrane-delimited Gβγ subunits in a voltage-independent manner.

  5. Voltage-gated calcium channels and their auxiliary subunits: physiology and pathophysiology and pharmacology. (United States)

    Dolphin, Annette C


    Voltage-gated calcium channels are essential players in many physiological processes in excitable cells. There are three main subdivisions of calcium channel, defined by the pore-forming α1 subunit, the CaV 1, CaV 2 and CaV 3 channels. For all the subtypes of voltage-gated calcium channel, their gating properties are key for the precise control of neurotransmitter release, muscle contraction and cell excitability, among many other processes. For the CaV 1 and CaV 2 channels, their ability to reach their required destinations in the cell membrane, their activation and the fine tuning of their biophysical properties are all dramatically influenced by the auxiliary subunits that associate with them. Furthermore, there are many diseases, both genetic and acquired, involving voltage-gated calcium channels. This review will provide a general introduction and then concentrate particularly on the role of auxiliary α2 δ subunits in both physiological and pathological processes involving calcium channels, and as a therapeutic target.

  6. Calcium ion channel and epilepsy

    Institute of Scientific and Technical Information of China (English)

    Yudan Lü; Weihong Lin; Dihui Ma


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

  7. 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


    in kidney function. It was hypothesized that human renal vascular excitation-contraction coupling involves different subtypes of channels. In human renal artery and dissected intrarenal blood vessels from nephrectomies, PCR analysis showed expression of L-type (Ca(v) 1.2), P/Q-type (Ca(v) 2.1), and T-type......, and L- and P/Q-type channels are of functional importance for the depolarization-induced vasoconstriction. The contribution of P/Q-type channels to contraction in the human vasculature is a novel mechanism for the regulation of renal blood flow and suggests that clinical treatment with calcium blockers......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...

  8. In vivo impact of presynaptic calcium channel dysfunction on motor axons in episodic ataxia type 2. (United States)

    Tomlinson, Susan E; Tan, S Veronica; Burke, David; Labrum, Robyn W; Haworth, Andrea; Gibbons, Vaneesha S; Sweeney, Mary G; Griggs, Robert C; Kullmann, Dimitri M; Bostock, Hugh; Hanna, Michael G


    Ion channel dysfunction causes a range of neurological disorders by altering transmembrane ion fluxes, neuronal or muscle excitability, and neurotransmitter release. Genetic neuronal channelopathies affecting peripheral axons provide a unique opportunity to examine the impact of dysfunction of a single channel subtype in detail in vivo. Episodic ataxia type 2 is caused by mutations in CACNA1A, which encodes the pore-forming subunit of the neuronal voltage-gated calcium channel Cav2.1. In peripheral motor axons, this channel is highly expressed at the presynaptic neuromuscular junction where it contributes to action potential-evoked neurotransmitter release, but it is not expressed mid-axon or thought to contribute to action potential generation. Eight patients from five families with genetically confirmed episodic ataxia type 2 underwent neurophysiological assessment to determine whether axonal excitability was normal and, if not, whether changes could be explained by Cav2.1 dysfunction. New mutations in the CACNA1A gene were identified in two families. Nerve conduction studies were normal, but increased jitter in single-fibre EMG studies indicated unstable neuromuscular transmission in two patients. Excitability properties of median motor axons were compared with those in 30 age-matched healthy control subjects. All patients had similar excitability abnormalities, including a high electrical threshold and increased responses to hyperpolarizing (P ataxia type 2 thus has unexpected effects on axon excitability, which may reflect an indirect effect of abnormal calcium current fluxes during development.

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

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


    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.

  10. Fetal calcium regulates branching morphogenesis in the developing human and mouse lung: involvement of voltage-gated calcium channels. (United States)

    Brennan, Sarah C; Finney, Brenda A; Lazarou, Maria; Rosser, Anne E; Scherf, Caroline; Adriaensen, Dirk; Kemp, Paul J; Riccardi, Daniela


    Airway branching morphogenesis in utero is essential for optimal postnatal lung function. In the fetus, branching morphogenesis occurs during the pseudoglandular stage (weeks 9-17 of human gestation, embryonic days (E)11.5-16.5 in mouse) in a hypercalcaemic environment (~1.7 in the fetus vs. ~1.1-1.3 mM for an adult). Previously we have shown that fetal hypercalcemia exerts an inhibitory brake on branching morphogenesis via the calcium-sensing receptor. In addition, earlier studies have shown that nifedipine, a selective blocker of L-type voltage-gated Ca(2+) channels (VGCC), inhibits fetal lung growth, suggesting a role for VGCC in lung development. The aim of this work was to investigate the expression of VGCC in the pseudoglandular human and mouse lung, and their role in branching morphogenesis. Expression of L-type (CaV1.2 and CaV1.3), P/Q type (CaV2.1), N-type (CaV2.2), R-type (CaV2.3), and T-type (CaV3.2 and CaV3.3) VGCC was investigated in paraffin sections from week 9 human fetal lungs and E12.5 mouse embryos. Here we show, for the first time, that Cav1.2 and Cav1.3 are expressed in both the smooth muscle and epithelium of the developing human and mouse lung. Additionally, Cav2.3 was expressed in the lung epithelium of both species. Incubating E12.5 mouse lung rudiments in the presence of nifedipine doubled the amount of branching, an effect which was partly mimicked by the Cav2.3 inhibitor, SNX-482. Direct measurements of changes in epithelial cell membrane potential, using the voltage-sensitive fluorescent dye DiSBAC2(3), demonstrated that cyclic depolarisations occur within the developing epithelium and coincide with rhythmic occlusions of the lumen, driven by the naturally occurring airway peristalsis. We conclude that VGCC are expressed and functional in the fetal human and mouse lung, where they play a role in branching morphogenesis. Furthermore, rhythmic epithelial depolarisations evoked by airway peristalsis would allow for branching to match

  11. Atypical properties of a conventional calcium channel β subunit from the platyhelminth Schistosoma mansoni

    Directory of Open Access Journals (Sweden)

    Schneider Toni


    Full Text Available Abstract Background The function of voltage-gated calcium (Cav channels greatly depends on coupling to cytoplasmic accessory β subunits, which not only promote surface expression, but also modulate gating and kinetic properties of the α1 subunit. Schistosomes, parasitic platyhelminths that cause schistosomiasis, express two β subunit subtypes: a structurally conventional β subunit and a variant β subunit with unusual functional properties. We have previously characterized the functional properties of the variant Cavβ subunit. Here, we focus on the modulatory phenotype of the conventional Cavβ subunit (SmCavβ using the human Cav2.3 channel as the substrate for SmCavβ and the whole-cell patch-clamp technique. Results The conventional Schistosoma mansoni Cavβ subunit markedly increases Cav2.3 currents, slows macroscopic inactivation and shifts steady state inactivation in the hyperpolarizing direction. However, currents produced by Cav2.3 in the presence of SmCavβ run-down to approximately 75% of their initial amplitudes within two minutes of establishing the whole-cell configuration. This suppressive effect was independent of Ca2+, but dependent on intracellular Mg2+-ATP. Additional experiments revealed that SmCavβ lends the Cav2.3/SmCavβ complex sensitivity to Na+ ions. A mutant version of the Cavβ subunit lacking the first forty-six amino acids, including a string of twenty-two acidic residues, no longer conferred sensitivity to intracellular Mg2+-ATP and Na+ ions, while continuing to show wild type modulation of current amplitude and inactivation of Cav2.3. Conclusion The data presented in this article provide insights into novel mechanisms employed by platyhelminth Cavβ subunits to modulate voltage-gated Ca2+ currents that indicate interactions between the Ca2+ channel complex and chelated forms of ATP as well as Na+ ions. These results have potentially important implications for understanding previously unknown mechanisms by

  12. SLO BK Potassium Channels Couple Gap Junctions to Inhibition of Calcium Signaling in Olfactory Neuron Diversification. (United States)

    Alqadah, Amel; Hsieh, Yi-Wen; Schumacher, Jennifer A; Wang, Xiaohong; Merrill, Sean A; Millington, Grethel; Bayne, Brittany; Jorgensen, Erik M; Chuang, Chiou-Fen


    The C. elegans AWC olfactory neuron pair communicates to specify asymmetric subtypes AWCOFF and AWCON in a stochastic manner. Intercellular communication between AWC and other neurons in a transient NSY-5 gap junction network antagonizes voltage-activated calcium channels, UNC-2 (CaV2) and EGL-19 (CaV1), in the AWCON cell, but how calcium signaling is downregulated by NSY-5 is only partly understood. Here, we show that voltage- and calcium-activated SLO BK potassium channels mediate gap junction signaling to inhibit calcium pathways for asymmetric AWC differentiation. Activation of vertebrate SLO-1 channels causes transient membrane hyperpolarization, which makes it an important negative feedback system for calcium entry through voltage-activated calcium channels. Consistent with the physiological roles of SLO-1, our genetic results suggest that slo-1 BK channels act downstream of NSY-5 gap junctions to inhibit calcium channel-mediated signaling in the specification of AWCON. We also show for the first time that slo-2 BK channels are important for AWC asymmetry and act redundantly with slo-1 to inhibit calcium signaling. In addition, nsy-5-dependent asymmetric expression of slo-1 and slo-2 in the AWCON neuron is necessary and sufficient for AWC asymmetry. SLO-1 and SLO-2 localize close to UNC-2 and EGL-19 in AWC, suggesting a role of possible functional coupling between SLO BK channels and voltage-activated calcium channels in AWC asymmetry. Furthermore, slo-1 and slo-2 regulate the localization of synaptic markers, UNC-2 and RAB-3, in AWC neurons to control AWC asymmetry. We also identify the requirement of bkip-1, which encodes a previously identified auxiliary subunit of SLO-1, for slo-1 and slo-2 function in AWC asymmetry. Together, these results provide an unprecedented molecular link between gap junctions and calcium pathways for terminal differentiation of olfactory neurons.

  13. Store-Operated Calcium Channels. (United States)

    Prakriya, Murali; Lewis, Richard S


    Store-operated calcium channels (SOCs) are a major pathway for calcium signaling in virtually all metozoan cells and serve a wide variety of functions ranging from gene expression, motility, and secretion to tissue and organ development and the immune response. SOCs are activated by the depletion of Ca(2+) from the endoplasmic reticulum (ER), triggered physiologically through stimulation of a diverse set of surface receptors. Over 15 years after the first characterization of SOCs through electrophysiology, the identification of the STIM proteins as ER Ca(2+) sensors and the Orai proteins as store-operated channels has enabled rapid progress in understanding the unique mechanism of store-operate calcium entry (SOCE). Depletion of Ca(2+) from the ER causes STIM to accumulate at ER-plasma membrane (PM) junctions where it traps and activates Orai channels diffusing in the closely apposed PM. Mutagenesis studies combined with recent structural insights about STIM and Orai proteins are now beginning to reveal the molecular underpinnings of these choreographic events. This review describes the major experimental advances underlying our current understanding of how ER Ca(2+) depletion is coupled to the activation of SOCs. Particular emphasis is placed on the molecular mechanisms of STIM and Orai activation, Orai channel properties, modulation of STIM and Orai function, pharmacological inhibitors of SOCE, and the functions of STIM and Orai in physiology and disease.

  14. RNAi silencing of P/Q-type calcium channels in Purkinje neurons of adult mouse leads to episodic ataxia type 2. (United States)

    Salvi, Julie; Bertaso, Federica; Mausset-Bonnefont, Anne-Laure; Metz, Alexandra; Lemmers, Céline; Ango, Fabrice; Fagni, Laurent; Lory, Philippe; Mezghrani, Alexandre


    Episodic ataxia type-2 (EA2) is a dominantly inherited human neurological disorder caused by loss of function mutations in the CACNA1A gene, which encodes the CaV2.1 subunit of P/Q-type voltage-gated calcium channels. It remains however unknown whether the deficit of cerebellar CaV2.1 in adult is in direct link with the disease. To address this issue, we have used lentiviral based-vector RNA interference (RNAi) to knock-down CaV2.1 expression in the cerebellum of adult mice. We show that suppression of the P/Q-type channels in Purkinje neurons induced motor abnormalities, such as imbalance and ataxic gait. Interestingly, moderate channel suppression caused no basal ataxia, while β-adrenergic activation and exercise mimicked stress induced motor disorders. Moreover, stress-induced ataxia was stable, non-progressive and totally abolished by acetazolamide, a carbonic anhydrase inhibitor used to treat EA2. Altogether, these data reveal that P/Q-type channel suppression in adult mice supports the episodic status of EA2 disease.

  15. Calcium channel blockers and Alzheimer's disease

    Institute of Scientific and Technical Information of China (English)

    Yi Tan; Yulin Deng; Hong Qing


    Alzheimer's disease is characterized by two pathological hallmarks: amyloid plaques and neurofi-brillary tangles. In addition, calcium homeostasis is disrupted in the course of human aging. Recent research shows that dense plaques can cause functional alteration of calcium signals in mice with Alzheimer's disease. Calcium channel blockers are effective therapeutics for treating Alzheimer's disease. This review provides an overview of the current research of calcium channel blockers in-volved in Alzheimer's disease therapy.

  16. Calcium signalling and calcium channels: evolution and general principles. (United States)

    Verkhratsky, Alexei; Parpura, Vladimir


    Calcium as a divalent cation was selected early in evolution as a signaling molecule to be used by both prokaryotes and eukaryotes. Its low cytosolic concentration likely reflects the initial concentration of this ion in the primordial soup/ocean as unicellular organisms were formed. As the concentration of calcium in the ocean subsequently increased, so did the diversity of homeostatic molecules handling calcium. This includes the plasma membrane channels that allowed the calcium entry, as well as extrusion mechanisms, i.e., exchangers and pumps. Further diversification occurred with the evolution of intracellular organelles, in particular the endoplasmic reticulum and mitochondria, which also contain channels, exchanger(s) and pumps to handle the homeostasis of calcium ions. Calcium signalling system, based around coordinated interactions of the above molecular entities, can be activated by the opening of voltage-gated channels, neurotransmitters, second messengers and/or mechanical stimulation, and as such is all-pervading pathway in physiology and pathophysiology of organisms.

  17. Disease causing mutations of calcium channels. (United States)

    Lorenzon, Nancy M; Beam, Kurt G


    Calcium ions play an important role in the electrical excitability of nerve and muscle, as well as serving as a critical second messenger for diverse cellular functions. As a result, mutations of genes encoding calcium channels may have subtle affects on channel function yet strongly perturb cellular behavior. This review discusses the effects of calcium channel mutations on channel function, the pathological consequences for cellular physiology, and possible links between altered channel function and disease. Many cellular functions are directly or indirectly regulated by the free cytosolic calcium concentration. Thus, calcium levels must be very tightly regulated in time and space. Intracellular calcium ions are essential second messengers and play a role in many functions including, action potential generation, neurotransmitter and hormone release, muscle contraction, neurite outgrowth, synaptogenesis, calcium-dependent gene expression, synaptic plasticity and cell death. Calcium ions that control cell activity can be supplied to the cell cytosol from two major sources: the extracellular space or intracellular stores. Voltage-gated and ligand-gated channels are the primary way in which Ca(2+) ions enter from the extracellular space. The sarcoplasm reticulum (SR) in muscle and the endoplasmic reticulum in non-muscle cells are the main intracellular Ca(2+) stores: the ryanodine receptor (RyR) and inositol-triphosphate receptor channels are the major contributors of calcium release from internal stores.

  18. Calcium channel as a potential anticancer agent. (United States)

    Kriazhev, L


    Anticancer treatment in modern clinical practices includes chemotherapy and radiation therapy with or without surgical interventions. Efficiency of both methods varies greatly depending on cancer types and stages. Besides, chemo- and radiotherapy are toxic and damaging that causes serious side effects. This fact prompts the search for alternative methods of antitumor therapy. It is well known that prolonged or high increase of intracellular calcium concentration inevitably leads to the cell death via apoptosis or necrosis. However, stimulation of cell calcium level by chemical agents is hardly achievable because cells have very sophisticated machinery for maintaining intracellular calcium in physiological ranges. This obstacle can be overridden, nevertheless. It was found that calcium channels in so called calcium cells in land snails are directly regulated by extracellular calcium concentration. The higher the concentration the higher the calcium intake is through the channels. Bearing in mind that extracellular/intracellular calcium concentration ratio in human beings is 10,000-12,000 fold the insertion of the channel into cancer cells would lead to fast and uncontrollable by the cells calcium intake and cell death. Proteins composing the channel may be extracted from plasma membrane of calcium cells and sequenced by mass-spectrometry or N-terminal sequencing. Either proteins or corresponding genes could be used for targeted delivery into cancer cells.

  19. Calcium binding protein-mediated regulation of voltage-gated calcium channels linked to human diseases

    Institute of Scientific and Technical Information of China (English)

    Nasrin NFJATBAKHSH; Zhong-ping FENG


    Calcium ion entry through voltage-gated calcium channels is essential for cellular signalling in a wide variety of cells and multiple physiological processes. Perturbations of voltage-gated calcium channel function can lead to pathophysiological consequences. Calcium binding proteins serve as calcium sensors and regulate the calcium channel properties via feedback mechanisms. This review highlights the current evidences of calcium binding protein-mediated channel regulation in human diseases.

  20. Gene splicing of an invertebrate beta subunit (LCavβ in the N-terminal and HOOK domains and its regulation of LCav1 and LCav2 calcium channels.

    Directory of Open Access Journals (Sweden)

    Taylor F Dawson

    Full Text Available The accessory beta subunit (Ca(vβ of calcium channels first appear in the same genome as Ca(v1 L-type calcium channels in single-celled coanoflagellates. The complexity of this relationship expanded in vertebrates to include four different possible Ca(vβ subunits (β1, β2, β3, β4 which associate with four Ca(v1 channel isoforms (Ca(v1.1 to Ca(v1.4 and three Ca(v2 channel isoforms (Ca(v2.1 to Ca(v2.3. Here we assess the fundamentally-shared features of the Ca(vβ subunit in an invertebrate model (pond snail Lymnaea stagnalis that bears only three homologous genes: (LCa(v1, LCa(v2, and LCa(vβ. Invertebrate Ca(vβ subunits (in flatworms, snails, squid and honeybees slow the inactivation kinetics of Ca(v2 channels, and they do so with variable N-termini and lacking the canonical palmitoylation residues of the vertebrate β2a subunit. Alternative splicing of exon 7 of the HOOK domain is a primary determinant of a slow inactivation kinetics imparted by the invertebrate LCa(vβ subunit. LCa(vβ will also slow the inactivation kinetics of LCa(v3 T-type channels, but this is likely not physiologically relevant in vivo. Variable N-termini have little influence on the voltage-dependent inactivation kinetics of differing invertebrate Ca(vβ subunits, but the expression pattern of N-terminal splice isoforms appears to be highly tissue specific. Molluscan LCa(vβ subunits have an N-terminal "A" isoform (coded by exons: 1a and 1b that structurally resembles the muscle specific variant of vertebrate β1a subunit, and has a broad mRNA expression profile in brain, heart, muscle and glands. A more variable "B" N-terminus (exon 2 in the exon position of mammalian β3 and has a more brain-centric mRNA expression pattern. Lastly, we suggest that the facilitation of closed-state inactivation (e.g. observed in Ca(v2.2 and Ca(vβ3 subunit combinations is a specialization in vertebrates, because neither snail subunit (LCa(v2 nor LCa(vβ appears to be compatible

  1. Molecular Basis of Regulating High Voltage-Activated Calcium Channels by S-Nitrosylation. (United States)

    Zhou, Meng-Hua; Bavencoffe, Alexis; Pan, Hui-Lin


    Nitric oxide (NO) is involved in a variety of physiological processes, such as vasoregulation and neurotransmission, and has a complex role in the regulation of pain transduction and synaptic transmission. We have shown previously that NO inhibits high voltage-activated Ca(2+) channels in primary sensory neurons and excitatory synaptic transmission in the spinal dorsal horn. However, the molecular mechanism involved in this inhibitory action remains unclear. In this study, we investigated the role of S-nitrosylation in the NO regulation of high voltage-activated Ca(2+) channels. The NO donor S-nitroso-N-acetyl-DL-penicillamine (SNAP) rapidly reduced N-type currents when Cav2.2 was coexpressed with the Cavβ1 or Cavβ3 subunits in HEK293 cells. In contrast, SNAP only slightly inhibited P/Q-type and L-type currents reconstituted with various Cavβ subunits. SNAP caused a depolarizing shift in voltage-dependent N-type channel activation, but it had no effect on Cav2.2 protein levels on the membrane surface. The inhibitory effect of SNAP on N-type currents was blocked by the sulfhydryl-specific modifying reagent methanethiosulfonate ethylammonium. Furthermore, the consensus motifs of S-nitrosylation were much more abundant in Cav2.2 than in Cav1.2 and Cav2.1. Site-directed mutagenesis studies showed that Cys-805, Cys-930, and Cys-1045 in the II-III intracellular loop, Cys-1835 and Cys-2145 in the C terminus of Cav2.2, and Cys-346 in the Cavβ3 subunit were nitrosylation sites mediating NO sensitivity of N-type channels. Our findings demonstrate that the consensus motifs of S-nitrosylation in cytoplasmically accessible sites are critically involved in post-translational regulation of N-type Ca(2+) channels by NO. S-Nitrosylation mediates the feedback regulation of N-type channels by NO.

  2. Calcium channel antagonists in hypertension. (United States)

    Ambrosioni, E; Borghi, C


    The clinical usefulness of calcium entry-blockers for the treatment of high blood pressure is related to their capacity to act upon the primary hemodynamic derangement in hypertension: the increased peripheral vascular resistance. They can be used alone or in combination with other antihypertensive agents for the treatment of various forms of hypertensive disease. The calcium entry-blockers appear to be the most useful agents for the treatment of hypertension in the elderly and for the treatment of hypertension associated with ischemic heart disease, pulmonary obstructive disease, peripheral vascular disease, and supraventricular arrhythmias. They are effective in reducing blood pressure in pregnancy-associated hypertension and must be considered as first-line therapy for the treatment of hypertensive crisis.

  3. Syntaxin-3 Binds and Regulates Both R- and L-Type Calcium Channels in Insulin-Secreting INS-1 832/13 Cells.

    Directory of Open Access Journals (Sweden)

    Li Xie

    Full Text Available Syntaxin (Syn-1A mediates exocytosis of predocked insulin-containing secretory granules (SGs during first-phase glucose-stimulated insulin secretion (GSIS in part via its interaction with plasma membrane (PM-bound L-type voltage-gated calcium channels (Cav. In contrast, Syn-3 mediates exocytosis of newcomer SGs that accounts for second-phase GSIS. We now hypothesize that the newcomer SG Syn-3 preferentially binds and modulates R-type Cav opening, which was postulated to mediate second-phase GSIS. Indeed, glucose-stimulation of pancreatic islet β-cell line INS-1 induced a predominant increase in interaction between Syn-3 and Cavα1 pore-forming subunits of R-type Cav2.3 and to lesser extent L-type Cavs, while confirming the preferential interactions between Syn-1A with L-type (Cav1.2, Cav1.3 Cavs. Consistently, direct binding studies employing heterologous HEK cells confirmed that Syn-3 preferentially binds Cav2.3, whereas Syn-1A prefers L-type Cavs. We then used siRNA knockdown (KD of Syn-3 in INS-1 to study the endogenous modulatory actions of Syn-3 on Cav channels. Syn-3 KD enhanced Ca2+ currents by 46% attributed mostly to R- and L-type Cavs. Interestingly, while the transmembrane domain of Syn-1A is the putative functional domain modulating Cav activity, it is the cytoplasmic domain of Syn-3 that appears to modulate Cav activity. We conclude that Syn-3 may mimic Syn-1A in the ability to bind and modulate Cavs, but preferring Cav2.3 to perhaps participate in triggering fusion of newcomer insulin SGs during second-phase GSIS.

  4. Congenital ataxia and hemiplegic migraine with cerebral edema associated with a novel gain of function mutation in the calcium channel CACNA1A. (United States)

    García Segarra, Nuria; Gautschi, Ivan; Mittaz-Crettol, Laureane; Kallay Zetchi, Christine; Al-Qusairi, Lama; Van Bemmelen, Miguel Xavier; Maeder, Philippe; Bonafé, Luisa; Schild, Laurent; Roulet-Perez, Eliane


    Mutations in the CACNA1A gene, encoding the α1 subunit of the voltage-gated calcium channel Ca(V)2.1 (P/Q-type), have been associated with three neurological phenotypes: familial and sporadic hemiplegic migraine type 1 (FHM1, SHM1), episodic ataxia type 2 (EA2), and spinocerebellar ataxia type 6 (SCA6). We report a child with congenital ataxia, abnormal eye movements and developmental delay who presented severe attacks of hemiplegic migraine triggered by minor head traumas and associated with hemispheric swelling and seizures. Progressive cerebellar atrophy was also observed. Remission of the attacks was obtained with acetazolamide. A de novo 3 bp deletion was found in heterozygosity causing loss of a phenylalanine residue at position 1502, in one of the critical transmembrane domains of the protein contributing to the inner part of the pore. We characterized the electrophysiology of this mutant in a Xenopus oocyte in vitro system and showed that it causes gain of function of the channel. The mutant Ca(V)2.1 activates at lower voltage threshold than the wild type. These findings provide further evidence of this molecular mechanism as causative of FHM1 and expand the phenotypic spectrum of CACNA1A mutations with a child exhibiting severe SHM1 and non-episodic ataxia of congenital onset.

  5. Archaerhodopsin voltage imaging: synaptic calcium and BK channels stabilize action potential repolarization at the Drosophila neuromuscular junction. (United States)

    Ford, Kevin J; Davis, Graeme W


    The strength and dynamics of synaptic transmission are determined, in part, by the presynaptic action potential (AP) waveform at the nerve terminal. The ion channels that shape the synaptic AP waveform remain essentially unknown for all but a few large synapses amenable to electrophysiological interrogation. The Drosophila neuromuscular junction (NMJ) is a powerful system for studying synaptic biology, but it is not amenable to presynaptic electrophysiology. Here, we demonstrate that Archaerhodopsin can be used to quantitatively image AP waveforms at the Drosophila NMJ without disrupting baseline synaptic transmission or neuromuscular development. It is established that Shaker mutations cause a dramatic increase in neurotransmitter release, suggesting that Shaker is predominantly responsible for AP repolarization. Here we demonstrate that this effect is caused by a concomitant loss of both Shaker and slowpoke (slo) channel activity because of the low extracellular calcium concentrations (0.2-0.5 mM) used typically to assess synaptic transmission in Shaker. In contrast, at physiological extracellular calcium (1.5 mM), the role of Shaker during AP repolarization is limited. We then provide evidence that calcium influx through synaptic CaV2.1 channels and subsequent recruitment of Slo channel activity is important, in concert with Shaker, to ensure proper AP repolarization. Finally, we show that Slo assumes a dominant repolarizing role during repetitive nerve stimulation. During repetitive stimulation, Slo effectively compensates for Shaker channel inactivation, stabilizing AP repolarization and limiting neurotransmitter release. Thus, we have defined an essential role for Slo channels during synaptic AP repolarization and have revised our understanding of Shaker channels at this model synapse.

  6. Calcium channel blockers in cardiovascular pharmacotherapy. (United States)

    Godfraind, Theophile


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

  7. Expression and pharmacology of endogenous Cav channels in SH-SY5Y human neuroblastoma cells.

    Directory of Open Access Journals (Sweden)

    Silmara R Sousa

    Full Text Available SH-SY5Y human neuroblastoma cells provide a useful in vitro model to study the mechanisms underlying neurotransmission and nociception. These cells are derived from human sympathetic neuronal tissue and thus, express a number of the Cav channel subtypes essential for regulation of important physiological functions, such as heart contraction and nociception, including the clinically validated pain target Cav2.2. We have detected mRNA transcripts for a range of endogenous expressed subtypes Cav1.3, Cav2.2 (including two Cav1.3, and three Cav2.2 splice variant isoforms and Cav3.1 in SH-SY5Y cells; as well as Cav auxiliary subunits α2δ1-3, β1, β3, β4, γ1, γ4-5, and γ7. Both high- and low-voltage activated Cav channels generated calcium signals in SH-SY5Y cells. Pharmacological characterisation using ω-conotoxins CVID and MVIIA revealed significantly (∼ 10-fold higher affinity at human versus rat Cav2.2, while GVIA, which interacts with Cav2.2 through a distinct pharmacophore had similar affinity for both species. CVID, GVIA and MVIIA affinity was higher for SH-SY5Y membranes vs whole cells in the binding assays and functional assays, suggesting auxiliary subunits expressed endogenously in native systems can strongly influence Cav2.2 channels pharmacology. These results may have implications for strategies used to identify therapeutic leads at Cav2.2 channels.

  8. Analytical models of calcium binding in a calcium channel

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jinn-Liang [Department of Applied Mathematics, National Hsinchu University of Education, Hsinchu 300, Taiwan (China); Eisenberg, Bob [Department of Molecular Biophysics and Physiology, Rush University, Chicago, Illinois 60612 (United States)


    The anomalous mole fraction effect of L-type calcium channels is analyzed using a Fermi like distribution with the experimental data of Almers and McCleskey [J. Physiol. 353, 585 (1984)] and the atomic resolution model of Lipkind and Fozzard [Biochemistry 40, 6786 (2001)] of the selectivity filter of the channel. Much of the analysis is algebraic, independent of differential equations. The Fermi distribution is derived from the configuration entropy of ions and water molecules with different sizes, different valences, and interstitial voids between particles. It allows us to calculate potentials and distances (between the binding ion and the oxygen ions of the glutamate side chains) directly from the experimental data using algebraic formulas. The spatial resolution of these results is comparable with those of molecular models, but of course the accuracy is no better than that implied by the experimental data. The glutamate side chains in our model are flexible enough to accommodate different types of binding ions in different bath conditions. The binding curves of Na{sup +} and Ca{sup 2+} for [CaCl{sub 2}] ranging from 10{sup −8} to 10{sup −2} M with a fixed 32 mM background [NaCl] are shown to agree with published Monte Carlo simulations. The Poisson-Fermi differential equation—that includes both steric and correlation effects—is then used to obtain the spatial profiles of energy, concentration, and dielectric coefficient from the solvent region to the filter. The energy profiles of ions are shown to depend sensitively on the steric energy that is not taken into account in the classical rate theory. We improve the rate theory by introducing a steric energy that lumps the effects of excluded volumes of all ions and water molecules and empty spaces between particles created by Lennard-Jones type and electrostatic forces. We show that the energy landscape varies significantly with bath concentrations. The energy landscape is not constant.

  9. Voltage-gated Calcium Channels and Autism Spectrum Disorders. (United States)

    Breitenkamp, Alexandra F; Matthes, Jan; Herzig, Stefan


    Autism spectrum disorder is a complex-genetic disease and its etiology is unknown for the majority of cases. So far, more than one hundred different susceptibility genes were detected. Voltage-gated calcium channels are among the candidates linked to autism spectrum disorder by results of genetic studies. Mutations of nearly all pore-forming and some auxiliary subunits of voltage gated calcium channels have been revealed from investigations of autism spectrum disorder patients and populations. Though there are only few electrophysiological characterizations of voltage-gated calcium channel mutations found in autistic patients these studies suggest their functional relevance. In summary, both genetic and functional data suggest a potential role of voltage-gated calcium channels in autism spectrum disorder. Future studies require refinement of the clinical and systems biological concepts of autism spectrum disorder and an appropriate holistic approach at the molecular level, e.g. regarding all facets of calcium channel functions.

  10. STIM and calcium channel complexes in cancer. (United States)

    Jardin, Isaac; Rosado, Juan A


    The ion Ca(2+) is a ubiquitous second messenger that mediates a variety of cellular functions. Dysfunction of the mechanisms involved in Ca(2+) homeostasis underlies a number of pathological processes, including cancer. Store-operated Ca(2+) entry (SOCE) is a major mechanism for Ca(2+) entry modulated by the intracellular Ca(2+) stores. The Ca(2+)-selective store-operated current (ICRAC) is mediated by the endoplasmic reticulum (ER) Ca(2+) sensor STIM1 and the store-operated Ca(2+) (SOC) channel Orai1, while other non-selective cation currents (ISOC) involves the participation of members of the canonical transient receptor potential (TRPC) channel family, including TRPC1. Distinct isoforms of the key components of SOCE have been described in mammalian cells, STIM1 and 2, Orai1-3 and TRPC1-7. In cancer cells, SOCE has been reported to play an important role in cell cycle progression and proliferation, migration, metastasis and evasion of apoptosis. Changes in the expression of the key elements of SOCE and Ca(2+) homeostasis remodeling have been account to play important roles in the phenotypic changes observed in transformed cells. Despite there are differences in the expression level of the molecular components of SOCE, as well as in the relevance of the STIM, Orai and TRPC isoforms in SOCE and tumorigenesis among cancer cell types, there is a body of evidence supporting an important role for SOCE underlying the phenotypic modifications of cancer cells that propose STIM and the SOC channels as suitable candidate targets for future prognostic or therapeutic strategies. This article is part of a Special Issue entitled: Calcium and Cell Fate. Guest Editors: Jacques Haiech, Claus Heizmann, Joachim Krebs, Thierry Capiod and Olivier Mignen.

  11. Drosophila mushroom body Kenyon cells generate spontaneous calcium transients mediated by PLTX-sensitive calcium channels. (United States)

    Jiang, Shaojuan Amy; Campusano, Jorge M; Su, Hailing; O'Dowd, Diane K


    Spontaneous calcium oscillations in mushroom bodies of late stage pupal and adult Drosophila brains have been implicated in memory consolidation during olfactory associative learning. This study explores the cellular mechanisms regulating calcium dynamics in Kenyon cells, principal neurons in mushroom bodies. Fura-2 imaging shows that Kenyon cells cultured from late stage Drosophila pupae generate spontaneous calcium transients in a cell autonomous fashion, at a frequency similar to calcium oscillations in vivo (10-20/h). The expression of calcium transients is up regulated during pupal development. Although the ability to generate transients is a property intrinsic to Kenyon cells, transients can be modulated by bath application of nicotine and GABA. Calcium transients are blocked, and baseline calcium levels reduced, by removal of external calcium, addition of cobalt, or addition of Plectreurys toxin (PLTX), an insect-specific calcium channel antagonist. Transients do not require calcium release from intracellular stores. Whole cell recordings reveal that the majority of voltage-gated calcium channels in Kenyon cells are PLTX-sensitive. Together these data show that influx of calcium through PLTX-sensitive voltage-gated calcium channels mediates spontaneous calcium transients and regulates basal calcium levels in cultured Kenyon cells. The data also suggest that these calcium transients represent cellular events underlying calcium oscillations in the intact mushroom bodies. However, spontaneous calcium transients are not unique to Kenyon cells as they are present in approximately 60% of all cultured central brain neurons. This suggests the calcium transients play a more general role in maturation or function of adult brain neurons.

  12. Renal vascular effects of calcium channel blockers in hypertension. (United States)

    Benstein, J A; Dworkin, L D


    Recent evidence suggests that calcium channel blockers have specific effects on renal hemodynamics in patients with hypertension and may also slow the progression of chronic renal failure. When these agents are studied in vitro, their predominant effect is to reverse afferent arteriolar vasoconstriction induced by catecholamines or angiotensin II. Because efferent resistance may remain high, glomerular filtration rate rises while renal blood flow remains low. The effects in vivo are less consistent. In human hypertension, calcium channel blockers lower renal resistance and may raise both renal blood flow and glomerular filtration rate. In experimental models of chronic renal disease, calcium channel blockers slow the progression of renal damage; however, variable effects on renal hemodynamics have been found. Other factors implicated in the progression of renal damage, including compensatory renal hypertrophy, platelet aggregation, and calcium deposition, may also be favorably influenced by these agents. Recent studies suggest that calcium channel blockers may have similar protective effects in patients with hypertension and chronic renal disease.

  13. NALCN ion channels have alternative selectivity filters resembling calcium channels or sodium channels.

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    Adriano Senatore

    Full Text Available NALCN is a member of the family of ion channels with four homologous, repeat domains that include voltage-gated calcium and sodium channels. NALCN is a highly conserved gene from simple, extant multicellular organisms without nervous systems such as sponges and placozoans and mostly remains a single gene compared to the calcium and sodium channels which diversified into twenty genes in humans. The single NALCN gene has alternatively-spliced exons at exons 15 or exon 31 that splices in novel selectivity filter residues that resemble calcium channels (EEEE or sodium channels (EKEE or EEKE. NALCN channels with alternative calcium, (EEEE and sodium, (EKEE or EEKE -selective pores are conserved in simple bilaterally symmetrical animals like flatworms to non-chordate deuterostomes. The single NALCN gene is limited as a sodium channel with a lysine (K-containing pore in vertebrates, but originally NALCN was a calcium-like channel, and evolved to operate as both a calcium channel and sodium channel for different roles in many invertebrates. Expression patterns of NALCN-EKEE in pond snail, Lymnaea stagnalis suggest roles for NALCN in secretion, with an abundant expression in brain, and an up-regulation in secretory organs of sexually-mature adults such as albumen gland and prostate. NALCN-EEEE is equally abundant as NALCN-EKEE in snails, but is greater expressed in heart and other muscle tissue, and 50% less expressed in the brain than NALCN-EKEE. Transfected snail NALCN-EEEE and NALCN-EKEE channel isoforms express in HEK-293T cells. We were not able to distinguish potential NALCN currents from background, non-selective leak conductances in HEK293T cells. Native leak currents without expressing NALCN genes in HEK-293T cells are NMDG(+ impermeant and blockable with 10 µM Gd(3+ ions and are indistinguishable from the hallmark currents ascribed to mammalian NALCN currents expressed in vitro by Lu et al. in Cell. 2007 Apr 20;129(2:371-83.

  14. 43. Calmodulin regulating calcium sensitivity of Na channels

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    R. Vegiraju


    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

  15. Heart failure drug digitoxin induces calcium uptake into cells by forming transmembrane calcium channels



    Digitoxin and other cardiac glycosides are important, centuries-old drugs for treating congestive heart failure. However, the mechanism of action of these compounds is still being elucidated. Calcium is known to potentiate the toxicity of these drugs, and we have hypothesized that digitoxin might mediate calcium entry into cells. We report here that digitoxin molecules mediate calcium entry into intact cells. Multimers of digitoxin molecules also are able to form calcium channels in pure plan...

  16. Osteoblasts detect pericellular calcium concentration increase via neomycin-sensitive voltage gated calcium channels. (United States)

    Sun, Xuanhao; Kishore, Vipuil; Fites, Kateri; Akkus, Ozan


    The mechanisms underlying the detection of critically loaded or micro-damaged regions of bone by bone cells are still a matter of debate. Our previous studies showed that calcium efflux originates from pre-failure regions of bone matrix and MC3T3-E1 osteoblasts respond to such efflux by an increase in the intracellular calcium concentration. The mechanisms by which the intracellular calcium concentration increases in response to an increase in the pericellular calcium concentration are unknown. Elevation of the intracellular calcium may occur via release from the internal calcium stores of the cell and/or via the membrane bound channels. The current study applied a wide range of pharmaceutical inhibitors to identify the calcium entry pathways involved in the process: internal calcium release from endoplasmic reticulum (ER, inhibited by thapsigargin and TMB-8), calcium receptor (CaSR, inhibited by calhex), stretch-activated calcium channel (SACC, inhibited by gadolinium), voltage-gated calcium channels (VGCC, inhibited by nifedipine, verapamil, neomycin, and ω-conotoxin), and calcium-induced-calcium-release channel (CICRC, inhibited by ryanodine and dantrolene). These inhibitors were screened for their effectiveness to block intracellular calcium increase by using a concentration gradient induced calcium efflux model which mimics calcium diffusion from the basal aspect of cells. The inhibitor(s) which reduced the intracellular calcium response was further tested on osteoblasts seeded on mechanically loaded notched cortical bone wafers undergoing damage. The results showed that only neomycin reduced the intracellular calcium response in osteoblasts, by 27%, upon extracellular calcium stimulus induced by concentration gradient. The inhibitory effect of neomycin was more pronounced (75% reduction in maximum fluorescence) for osteoblasts seeded on notched cortical bone wafers loaded mechanically to damaging load levels. These results imply that the increase in

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

    Goldberg, Joshua A; Wilson, Charles J


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

  18. Calcium signaling and T-type calcium channels in cancer cell cycling

    Institute of Scientific and Technical Information of China (English)

    James T Taylor; Xiang-Bin Zeng; Jonathan E Pottle; Kevin Lee; Alun R Wang; Stephenie G Yi; Jennifer A S Scruggs; Suresh S Sikka; Ming Li


    Regulation of intracellular calcium is an important signaling mechanism for cell proliferation in both normal and cancerous cells. In normal epithelial cells,free calcium concentration is essential for cells to enter and accomplish the S phase and the M phase of the cell cycle. In contrast, cancerous cells can pass these phases of the cell cycle with much lower cytoplasmic free calcium concentrations, indicating an alternative mechanism has developed for fulfilling the intracellular calcium requirement for an increased rate of DNA synthesis and mitosis of fast replicating cancerous cells. The detailed mechanism underlying the altered calcium loading pathway remains unclear;however, there is a growing body of evidence that suggests the T-type Ca2+ channel is abnormally expressed in cancerous cells and that blockade of these channels may reduce cell proliferation in addition to inducing apoptosis. Recent studies also show that the expression of T-type Ca2+ channels in breast cancer cells is proliferation state dependent, i.e. the channels are expressed at higher levels during the fast-replication period, and once the cells are in a non-proliferation state, expression of this channel isminimal. Therefore, selectively blocking calcium entry into cancerous cells may be a valuable approach for preventing tumor growth. Since T-type Ca2+ channels are not expressed in epithelial cells, selective T-type Ca2+ channel blockers may be useful in the treatment of certain types of cancers.

  19. Up-regulation of L-type high voltage-gated calcium channel subunits by sustained exposure to 1,4- and 1,5-benzodiazepines in cerebrocortical neurons. (United States)

    Katsura, Masashi; Shibasaki, Masahiro; Kurokawa, Kazuhiro; Tsujimura, Atsushi; Ohkuma, Seitaro


    The aim of this study is to examine how sustained exposure to two 1,4-benzodiazepines (BZDs) with different action period, diazepam and brotizolam, and a 1,5-BZD, clobazam, affects L-type high voltage-gated calcium channel (HVCC) functions and its mechanisms using primary cultures of mouse cerebral cortical neurons. The sustained exposure to these three BZDs increased [(45)Ca2+] influx, which was due to the enhanced [(45)Ca2+] entry through L-type HVCCs but not through of Cav2.1 and Cav2.2. Increase in [(3)H]diltiazem binding after the exposure to these three BZDs was due to the increase in the binding sites of [(3)H]diltiazem. Western blot analysis showed increase of Cav1.2 and Cav1.3 in association with the increased expression of alpha2/delta1 subunit. Similar changes in [(3)H]diltiazem binding and L-type HVCC subunit expression were found in the cerebral cortex from mouse with BZD physical dependence. These results indicate that BZDs examined here have the potential to increase L-type HVCC functions mediated via the enhanced expression of not only Cav1.2 and Cav1.3 but also alpha2/delta1 subunit after their sustained exposure, which may participate in the development of physical dependence by these BZDs.

  20. Heart failure drug digitoxin induces calcium uptake into cells by forming transmembrane calcium channels. (United States)

    Arispe, Nelson; Diaz, Juan Carlos; Simakova, Olga; Pollard, Harvey B


    Digitoxin and other cardiac glycosides are important, centuries-old drugs for treating congestive heart failure. However, the mechanism of action of these compounds is still being elucidated. Calcium is known to potentiate the toxicity of these drugs, and we have hypothesized that digitoxin might mediate calcium entry into cells. We report here that digitoxin molecules mediate calcium entry into intact cells. Multimers of digitoxin molecules also are able to form calcium channels in pure planar phospholipid bilayers. These digitoxin channels are blocked by Al(3+) and La(3+) but not by Mg(2+) or the classical l-type calcium channel blocker, nitrendipine. In bilayers, we find that the chemistry of the lipid affects the kinetics of the digitoxin channel activity, but not the cation selectivity. Antibodies against digitoxin promptly neutralize digitoxin channels in both cells and bilayers. We propose that these digitoxin calcium channels may be part of the mechanism by which digitoxin and other active cardiac glycosides, such as digoxin, exert system-wide actions at and above the therapeutic concentration range.

  1. T-type calcium channel: a privileged gate for calcium entry and control of adrenal steroidogenesis

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    Michel Florian Rossier


    Full Text Available Intracellular calcium plays a crucial role in modulating a variety of functions such as muscle contraction, hormone secretion, gene expression or cell growth. Calcium signaling has been however shown to be more complex than initially thought. Indeed, it is confined within cell microdomains and different calcium channels are associated with different functions, as shown by various channelopathies.Sporadic mutations on voltage-operated L-type calcium channels in adrenal glomerulosa cells have been shown recently to be the second most prevalent genetic abnormalities present in human aldosterone-producing adenoma. The observed modification of the threshold of activation of the mutated channels not only provides an explanation for this gain of function but reminds us on the importance of maintaining adequate electrophysiological characteristics to make channels able to exert specific cellular functions. Indeed, the contribution to steroid production of the various calcium channels expressed in adrenocortical cells is not equal and the reason has been investigated for a long time. Given the very negative resting potential of these cells, and the small membrane depolarization induced by their physiological agonists, low threshold T-type calcium channels are particularly well suited for responding under these conditions and conveying calcium into the cell, at the right place for controlling steroidogenesis. In contrast, high threshold L-type channels are normally activated by much stronger cell depolarizations. The fact that dihydropyridine calcium antagonists, specific for L-type channels, are poorly efficient for reducing aldosterone secretion either in vivo or in vitro, strongly supports the view that these two types of channels differently affect steroid biosynthesis.Whether a similar analysis is transposable to fasciculata cells and cortisol secretion is one of the questions addressed in the present review. No similar mutations on L-type or T

  2. The molecular choreography of a store-operated calcium channel. (United States)

    Lewis, Richard S


    Store-operated calcium channels (SOCs) serve essential functions from secretion and motility to gene expression and cell growth. A fundamental mystery is how the depletion of Ca2+ from the endoplasmic reticulum (ER) activates Ca2+ entry through SOCs in the plasma membrane. Recent studies using genetic approaches have identified genes encoding the ER Ca2+ sensor and a prototypic SOC, the Ca2+-release-activated Ca2+ (CRAC) channel. New findings reveal a unique mechanism for channel activation, in which the CRAC channel and its sensor migrate independently to closely apposed sites of interaction in the ER and the plasma membrane.

  3. Store-operated calcium channels and pro-inflammatory signals

    Institute of Scientific and Technical Information of China (English)

    Wei-chiao CHANG


    In non-excitable cells such as T lymphocytes,hepatocytes,mast cells,endothelia and epithelia,the major pathway for calcium(Ca2+)entry is through store-operated Ca2+ channels in the plasma membrane.These channels are activated by the emptying of intracellular Ca2+ stores,however,neither the gating mechanism nor the downstream targets of these channels has been clear established.Here,I review some of the proposed gating mechanisms of store-operated Ca2+ channels and the functional implications in regulating pro-inflammatory signals.

  4. Activity-Dependent Calcium, Oxygen, and Vascular Responses in a Mouse Model of Familial Hemiplegic Migraine Type 1

    DEFF Research Database (Denmark)

    Khennouf, Lila; Gesslein, Bodil; Lind, Barbara Lykke;


    Objective: Familial hemiplegic migraine type 1 (FHM1) is a subtype of migraine with aura caused by a gain-of-function mutation in the pore-forming α1 subunit of CaV2.1 (P/Q-type) calcium channels. However, the mechanisms underlying how the disease is brought about and the prolonged aura remain...... explain impaired neurovascular responses in the mutant, and these alterations could contribute to brain frailty in FHM1 patients...

  5. Regulation of voltage gated calcium channels by GPCRs and post-translational modification. (United States)

    Huang, Junting; Zamponi, Gerald W


    Calcium entry via voltage gated calcium channels mediates a wide range of physiological functions, whereas calcium channel dysregulation has been associated with numerous pathophysiological conditions. There are myriad cell signaling pathways that act on voltage gated calcium channels to fine tune their activities and to regulate their cell surface expression. These regulatory mechanisms include the activation of G protein-coupled receptors and downstream phosphorylation events, and their control over calcium channel trafficking through direct physical interactions. Calcium channels also undergo post-translational modifications that alter both function and density of the channels in the plasma membrane. Here we focus on select aspects of these regulatory mechanisms and highlight recent developments.

  6. Diltiazem and verapamil preferentially block inactivated cardiac calcium channels. (United States)

    Kanaya, S; Arlock, P; Katzung, B G; Hondeghem, L M


    Diltiazem has been proposed to act by blocking calcium channels of cardiac and smooth muscle since it has pharmacological [12-14] and clinical [10] effects that resemble those of verapamil, an agent that has been shown to block these channels [3]. However, block of the slow inward current by diltiazem has not been directly demonstrated. In fact, it has been suggested that diltiazem has an entirely different mechanism of action [7]. We therefore studied the blocking effects of diltiazem and verapamil on cardiac calcium channels by measuring the slow inward current in voltage-clamped ferret myocardium. Both drugs blocked the slow inward current in a use-dependent fashion, i.e. the block was enhanced by increased frequency of activating clamps and by more positive holding potentials. However, we found that short single activating clamps resulted in minimal block, whereas prolonging the clamp step progressively enhanced the blockade. Thus, a single long clamp caused as much blockade as a train of shorter pulses. These results demonstrate that diltiazem and verapamil block the slow inward current by binding to calcium channels in a state-dependent fashion, i.e. inactivated channels have a high affinity for the drugs, while rested and open channels have a lower affinity.

  7. Effects of calcium ion, calpains, and calcium channel blockers on retinitis pigmentosa. (United States)

    Nakazawa, Mitsuru


    Recent advances in molecular genetic studies have revealed many of the causative genes of retinitis pigmentosa (RP). These achievements have provided clues to the mechanisms of photoreceptor degeneration in RP. Apoptosis is known to be a final common pathway in RP and, therefore, a possible therapeutic target for photoreceptor rescue. However, apoptosis is not a single molecular cascade, but consists of many different reactions such as caspase-dependent and caspase-independent pathways commonly leading to DNA fractionation and cell death. The intracellular concentration of calcium ions is also known to increase in apoptosis. These findings suggest that calpains, one of the calcium-dependent proteinases, play some roles in the process of photoreceptor apoptosis and that calcium channel antagonists may potentially inhibit photoreceptor apoptosis. Herein, the effects of calpains and calcium channel antagonists on photoreceptor degeneration are reviewed.

  8. Effects of Calcium Ion, Calpains, and Calcium Channel Blockers on Retinitis Pigmentosa

    Directory of Open Access Journals (Sweden)

    Mitsuru Nakazawa


    Full Text Available Recent advances in molecular genetic studies have revealed many of the causative genes of retinitis pigmentosa (RP. These achievements have provided clues to the mechanisms of photoreceptor degeneration in RP. Apoptosis is known to be a final common pathway in RP and, therefore, a possible therapeutic target for photoreceptor rescue. However, apoptosis is not a single molecular cascade, but consists of many different reactions such as caspase-dependent and caspase-independent pathways commonly leading to DNA fractionation and cell death. The intracellular concentration of calcium ions is also known to increase in apoptosis. These findings suggest that calpains, one of the calcium-dependent proteinases, play some roles in the process of photoreceptor apoptosis and that calcium channel antagonists may potentially inhibit photoreceptor apoptosis. Herein, the effects of calpains and calcium channel antagonists on photoreceptor degeneration are reviewed.

  9. Oxidative Stress and Maxi Calcium-Activated Potassium (BK Channels

    Directory of Open Access Journals (Sweden)

    Anton Hermann


    Full Text Available All cells contain ion channels in their outer (plasma and inner (organelle membranes. Ion channels, similar to other proteins, are targets of oxidative impact, which modulates ion fluxes across membranes. Subsequently, these ion currents affect electrical excitability, such as action potential discharge (in neurons, muscle, and receptor cells, alteration of the membrane resting potential, synaptic transmission, hormone secretion, muscle contraction or coordination of the cell cycle. In this chapter we summarize effects of oxidative stress and redox mechanisms on some ion channels, in particular on maxi calcium-activated potassium (BK channels which play an outstanding role in a plethora of physiological and pathophysiological functions in almost all cells and tissues. We first elaborate on some general features of ion channel structure and function and then summarize effects of oxidative alterations of ion channels and their functional consequences.

  10. Heterogeneity of Calcium Channel/cAMP-Dependent Transcriptional Activation. (United States)

    Kobrinsky, Evgeny


    The major function of the voltage-gated calcium channels is to provide the Ca(2+) flux into the cell. L-type voltage-gated calcium channels (Cav1) serve as voltage sensors that couple membrane depolarization to many intracellular processes. Electrical activity in excitable cells affects gene expression through signaling pathways involved in the excitation-transcription (E-T) coupling. E-T coupling starts with activation of the Cav1 channel and results in initiation of the cAMP-response element binding protein (CREB)-dependent transcription. In this review we discuss the new quantitative approaches to measuring E-T signaling events. We describe the use of wavelet transform to detect heterogeneity of transcriptional activation in nuclei. Furthermore, we discuss the properties of discovered microdomains of nuclear signaling associated with the E-T coupling and the basis of the frequency-dependent transcriptional regulation.

  11. A calcium channel mutant mouse model of hypokalemic periodic paralysis


    Wu, Fenfen; Mi, Wentao; Hernández-Ochoa, Erick O.; Burns, Dennis K.; Fu, Yu; Gray, Hillery F; Struyk, Arie F.; Martin F Schneider; Cannon, Stephen C.


    Hypokalemic periodic paralysis (HypoPP) is a familial skeletal muscle disorder that presents with recurrent episodes of severe weakness lasting hours to days associated with reduced serum potassium (K+). HypoPP is genetically heterogeneous, with missense mutations of a calcium channel (CaV1.1) or a sodium channel (NaV1.4) accounting for 60% and 20% of cases, respectively. The mechanistic link between CaV1.1 mutations and the ictal loss of muscle excitability during an attack of weakness in Hy...

  12. Calcium-permeable ion channels in the kidney. (United States)

    Zhou, Yiming; Greka, Anna


    Calcium ions (Ca(2+)) are crucial for a variety of cellular functions. The extracellular and intracellular Ca(2+) concentrations are thus tightly regulated to maintain Ca(2+) homeostasis. The kidney, one of the major organs of the excretory system, regulates Ca(2+) homeostasis by filtration and reabsorption. Approximately 60% of the Ca(2+) in plasma is filtered, and 99% of that is reabsorbed by the kidney tubules. Ca(2+) is also a critical signaling molecule in kidney development, in all kidney cellular functions, and in the emergence of kidney diseases. Recently, studies using genetic and molecular biological approaches have identified several Ca(2+)-permeable ion channel families as important regulators of Ca(2+) homeostasis in kidney. These ion channel families include transient receptor potential channels (TRP), voltage-gated calcium channels, and others. In this review, we provide a brief and systematic summary of the expression, function, and pathological contribution for each of these Ca(2+)-permeable ion channels. Moreover, we discuss their potential as future therapeutic targets.

  13. Physiology and Regulation of Calcium Channels in Stomatal Guard Cells

    Energy Technology Data Exchange (ETDEWEB)

    Schroeder, Julian I.


    Stomatal pores in the epidermis of leaves regulate the diffusion of CO2 into leaves for photosynthetic carbon fixation and control water loss of plants during drought periods. Guard cells sense CO2, water status, light and other environmental conditions to regulate stomatal apertures for optimization of CO2 intake and plant growth under drought stress. The cytosolic second messenger calcium contributes to stomatal movements by transducing signals and regulating ion channels in guard cells. Studies suggest that both plasma membrane Ca2+ influx channels and vacuolar/organellar Ca2+ release channels contribute to ABA-induced Ca2+ elevations in guard cells. Recent research in the P.I.'s laboratory has led to identification of a novel major cation-selective Ca2+-permeable influx channel (Ica) in the plasma membrane of Arabidopsis guard cells. These advances will allow detailed characterization of Ica plasma membrane Ca2+ influx channels in guard cells. The long term goal of this research project is to gain a first detailed characterization of these novel plasma membrane Ca2+-permeable channel currents in Arabidopsis guard cells. The proposed research will investigate the hypothesis that Ica represents an important Ca2+ influx pathway for ABA and CO2 signal transduction in Arabidopsis guard cells. These studies will lead to elucidation of key signal transduction mechanisms by which plants balance CO2 influx into leaves and transpirational water loss and may contribute to future strategies for manipulating gas exchange for improved growth of crop plants and for biomass production.

  14. Magnesium: Effect on ocular health as a calcium channel antagonist

    Directory of Open Access Journals (Sweden)

    Şafak Korkmaz


    Full Text Available Magnesium is the physiologic calcium channel blocker,involving in many different metabolic processes by maintainingcell membrane function, modulating smooth musclecontraction and influencing enzymatic activities. Magnesiumhas been shown to increase blood flow to tissuesby modifying endothelial function via endothelin-1 (ET-1and nitric Oxide (NO pathways. Magnesium also exhibitsneuroprotective role by blocking N-methyl-D-aspartate(NMDA receptor related calcium influx and by inhibitingthe release of glutamate, hence protects the cell againstoxidative stress and apoptosis. Both increase in bloodflow and its neuroprotective effect make magnesium agood candidate for glaucoma studies. Magnesium hasbeen shown to decrease oxidative stress and apoptosisin retinal tissue and to have retinal ganglion cell sparingeffect. A series of studies has been conducted aboutmagnesium could decrease insulin resistance in diabeticpatients, ease glycemia control and prevent diabetic retinopathy.Magnesium is found to be critically important inmaintaining normal ionic homeostasis of lens. Magnesiumdeficiency has been shown to cause increased lenticularoxidative stress and ionic imbalance in the lens so triggercataractogenesis. J Clin Exp Invest 2013; 4 (2: 244-251Key words: Magnesium, calcium channel blockage,glaucoma, neuroprotection, diabetic retinopathy, cataract

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

    Directory of Open Access Journals (Sweden)

    Sulayma A Albarwani


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

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

    Directory of Open Access Journals (Sweden)

    Lu Ake Tzu-Hui


    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.

  17. Inhibition of voltage-gated calcium channels by sequestration of beta subunits. (United States)

    Cuchillo-Ibañez, Inmaculada; Aldea, Marcos; Brocard, Jacques; Albillos, Almudena; Weiss, Norbert; Garcia, Antonio G; De Waard, Michel


    The auxiliary Ca(v)beta subunit is essential for functional expression of high-voltage activated Ca(2+) channels. Here, we describe a lure sequence designed to sequester the Ca(v)beta subunits in transfected bovine chromaffin cells. This sequence is composed of the extracellular and transmembrane domains of the alpha chain of the human CD8, the I-II loop of Ca(v)2.1 subunit, and EGFP. We showed that expressing the CD8-I-II-EGFP sequence in chromaffin cells led to a >50% decrease in overall Ca(2+) current density. Although this decrease involved all the Ca(2+) channel types (L, N, P/Q, R), the proportion of each type supporting the remaining current was altered. A similar effect was observed after transfection when measuring the functional role of Ca(2+) channels in catecholamine release by chromaffin cells: global decrease of release and change of balance between the different channel types supporting it. Possible explanations for this apparent discrepancy are further discussed.

  18. Stimulation of protein kinase C recruits covert calcium channels in Aplysia bag cell neurons. (United States)

    Strong, J A; Fox, A P; Tsien, R W; Kaczmarek, L K

    The modulation of voltage-activated calcium currents by protein kinases provides excitable cells with a mechanism for regulating their electrical behaviour. At the single channel level, modulation of calcium current has, to date, been characterized only in cardiac muscle, where beta-adrenergic agonists, acting through cyclic AMP-dependent protein kinase, enhance the calcium current by increasing channel availability and opening. We now report that enhancement of calcium current in the peptidergic bag cell neurons of Aplysia by protein kinase C occurs through a different mechanism, the recruitment of a previously covert class of calcium channel. Under control conditions, bag cell neurons contain only one class of voltage-activated calcium channel with a conductance of approximately 12 pS. After exposure to agents that activate protein kinase C, these neurons also express a second class of calcium channel with a different unitary conductance (approximately 24 pS) that is never seen in untreated cells.

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

    Directory of Open Access Journals (Sweden)

    Bruce Lyeth


    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.

  20. End organ protection by calcium-channel blockers. (United States)

    Tzivoni, D


    In recent years, much attention has been given to end organ protection by antihypertensive, anti-heart failure, and anti-ischemic medications. This review describes the available information on end organ protection by calcium-channel blockers (CCBs). In normotensive patients and patients with hypertension treated with long-acting dihydropyridines, medial thickness was thinner than in patients treated with atenolol or in untreated hypertensive patients. Long-term treatment was associated with significant reduction in left ventricular mass. Calcium-channel blockers also improved endothelial-dependent relaxation and reversed the vasoconstrictive response to nitric oxide inhibitors. In diabetic patients, CCBs were effective in preserving kidney function and microalbuminurea. The combination of angiotensin-converting enzyme (ACE) inhibitors and CCBs was more effective than ACE inhibitors alone in preserving kidney function. In animal experiments, CCBs prevented development of coronary atheroschlerosis; however, in humans only limited data are available on their antiatherogenic effect. Some studies suggest that CCBs exert antiplatelets properties and may therefore be beneficial in patients with coronary artery disease.

  1. Newer calcium channel antagonists and the treatment of hypertension. (United States)

    Cummins, D F


    Calcium channel antagonists have become popular medications for the management of hypertension. These agents belong to the diphenylalkylamine, benzothiazepine, dihydropyridine, or tetralol chemical classes. Although the medications share a common pharmacological mechanism in reducing peripheral vascular resistance, clinical differences between the sub-classes can be linked to structural profiles. This heterogeneity is manifested by differences in vascular selectivity, effects on cardiac conduction and adverse events. The lack of differentiation between calcium channel antagonists in clinical trials has contributed to uncertainty associated with their impact on morbidity and mortality. Data from more recent studies in specific patient populations underscores the importance of investigating these antihypertensives as individual agents. A proposed therapeutic classification system suggests that newer agents should share the slow onset and long-acting antihypertensive effect of amlodipine. Additionally, a favourable trough-to-peak ratio has been recommended as an objective measurement of efficacy. The newer drugs, barnidipine and lacidipine, have a therapeutic profile similar to amlodipine, but trough-to-peak ratios are not substantially greater than the recommended minimum of 0.50. Aranidipine, cilnidipine and efonidipine have unique pharmacological properties that distinguish them from traditional dihydropyridines. Although clinical significance is unconfirmed, these newer options may be beneficial for patients with co-morbid conditions that preclude use of older antagonists.

  2. Glucocorticoids specifically enhance L-type calcium current amplitude and affect calcium channel subunit expression in the mouse hippocampus. (United States)

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


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

  3. Differential calcium signaling mediated by voltage-gated calcium channels in rat retinal ganglion cells and their unmyelinated axons.

    Directory of Open Access Journals (Sweden)

    Allison Sargoy

    Full Text Available Aberrant calcium regulation has been implicated as a causative factor in the degeneration of retinal ganglion cells (RGCs in numerous injury models of optic neuropathy. Since calcium has dual roles in maintaining homeostasis and triggering apoptotic pathways in healthy and injured cells, respectively, investigation of voltage-gated Ca channel (VGCC regulation as a potential strategy to reduce the loss of RGCs is warranted. The accessibility and structure of the retina provide advantages for the investigation of the mechanisms of calcium signalling in both the somata of ganglion cells as well as their unmyelinated axons. The goal of the present study was to determine the distribution of VGCC subtypes in the cell bodies and axons of ganglion cells in the normal retina and to define their contribution to calcium signals in these cellular compartments. We report L-type Ca channel α1C and α1D subunit immunoreactivity in rat RGC somata and axons. The N-type Ca channel α1B subunit was in RGC somata and axons, while the P/Q-type Ca channel α1A subunit was only in the RGC somata. We patch clamped isolated ganglion cells and biophysically identified T-type Ca channels. Calcium imaging studies of RGCs in wholemounted retinas showed that selective Ca channel antagonists reduced depolarization-evoked calcium signals mediated by L-, N-, P/Q- and T-type Ca channels in the cell bodies but only by L-type Ca channels in the axons. This differential contribution of VGCC subtypes to calcium signals in RGC somata and their axons may provide insight into the development of target-specific strategies to spare the loss of RGCs and their axons following injury.

  4. Role of L-type calcium-channel modulation in nonconvulsive epilepsy in rats

    NARCIS (Netherlands)

    Luijtelaar, E.L.J.M. van; Ates, N.; Coenen, A.M.L.


    Old male Wistar rats spontaneously showing hundreds of spike-wave discharges daily were used to investigate the role of calcium ions in nonconvulsive epilepsy. The effects of the L-type calcium channel blocker nimodipine and the L-type channel opener BAY K 8644 on number and duration of these spike-

  5. CaV1.2 calcium channels: just cut out to be regulated? (United States)

    Groth, Rachel D; Tirko, Natasha N; Tsien, Richard W


    Tight regulation of calcium entry through the L-type calcium channel CaV1.2 ensures optimal excitation-response coupling. In this issue of Neuron, Michailidis et al. (2014) demonstrate that CaV1.2 activity triggers negative feedback regulation through proteolytic cleavage of the channel within the core of the pore-forming subunit.


    Directory of Open Access Journals (Sweden)

    O. D. Ostroumova


    Full Text Available Classification, modes of action and clinical effects of calcium channel blockers are presented. Advantages of the third generation of dihydropyridine calcium channel blockers are considered. Clinical pharmacology, studies on the efficacy, safety and prevention of hypertensive complications with lercanidipine are detailed.

  7. Calcium channel structural determinants of synaptic transmission between identified invertebrate neurons. (United States)

    Spafford, J David; Munno, David W; Van Nierop, Pim; Feng, Zhong-Ping; Jarvis, Scott E; Gallin, Warren J; Smit, August B; Zamponi, Gerald W; Syed, Naweed I


    We report here that unlike what was suggested for many vertebrate neurons, synaptic transmission in Lymnaea stagnalis occurs independent of a physical interaction between presynaptic calcium channels and a functional complement of SNARE proteins. Instead, synaptic transmission in Lymnaea requires the expression of a C-terminal splice variant of the Lymnaea homolog to mammalian N- and P/Q-type calcium channels. We show that the alternately spliced region physically interacts with the scaffolding proteins Mint1 and CASK, and that synaptic transmission is abolished following RNA interference knockdown of CASK or after the injection of peptide sequences designed to disrupt the calcium channel-Mint1 interactions. Our data suggest that Mint1 and CASK may serve to localize the non-L-type channels at the active zone and that synaptic transmission in invertebrate neurons utilizes a mechanism for optimizing calcium entry, which occurs independently of a physical association between calcium channels and SNARE proteins.

  8. The cardiac L-type calcium channel distal carboxy terminus autoinhibition is regulated by calcium. (United States)

    Crump, Shawn M; Andres, Douglas A; Sievert, Gail; Satin, Jonathan


    The L-type calcium channel (LTCC) provides trigger Ca(2+) for sarcoplasmic reticulum Ca-release, and LTCC function is influenced by interacting proteins including the LTCC distal COOH terminus (DCT) and calmodulin. DCT is proteolytically cleaved and reassociates with the LTCC complex to regulate calcium channel function. DCT reduces LTCC barium current (I(Ba,L)) in reconstituted channel complexes, yet the contribution of DCT to LTCC Ca(2+) current (I(Ca,L)) in cardiomyocyte systems is unexplored. This study tests the hypothesis that DCT attenuates cardiomyocyte I(Ca,L). We measured LTCC current and Ca(2+) transients with DCT coexpressed in murine cardiomyocytes. We also heterologously coexpressed DCT and Ca(V)1.2 constructs with truncations corresponding to the predicted proteolytic cleavage site, Ca(V)1.2Δ1801, and a shorter deletion corresponding to well-studied construct, Ca(V)1.2Δ1733. DCT inhibited I(Ba,L) in cardiomyocytes, and in human embryonic kidney (HEK) 293 cells expressing Ca(V)1.2Δ1801 and Ca(V)1.2Δ1733. Ca(2+)-CaM relieved DCT block in cardiomyocytes and HEK cells. The selective block of I(Ba,L) combined with Ca(2+)-CaM effects suggested that DCT-mediated blockade may be relieved under conditions of elevated Ca(2+). We therefore tested the hypothesis that DCT block is dynamic, increasing under relatively low Ca(2+), and show that DCT reduced diastolic Ca(2+) at low stimulation frequencies but spared high frequency Ca(2+) entry. DCT reduction of diastolic Ca(2+) and relief of block at high pacing frequencies and under conditions of supraphysiological bath Ca(2+) suggests that a physiological function of DCT is to increase the dynamic range of Ca(2+) transients in response to elevated pacing frequencies. Our data motivate the new hypothesis that DCT is a native reverse use-dependent inhibitor of LTCC current.

  9. Effects of calcium channel on 3-morpholinosydnonimine-induced rat hippocampal neuronal apoptosis

    Institute of Scientific and Technical Information of China (English)

    Quanzhong Chang; Shuling Zhang; Yuanyin Zheng; Lijuan Xu; Jinbao Yin; Shining Cai


    Previous studies have demonstrated that increased chloride channel activity plays a role in nitric oxide-induced neuronal apoptosis in the rat hippocampus.The present study investigated the effects of the broad-spectrum calcium channel blocker CdC12 on survival rate, percentage of apoptosis, and morphological changes in hippocampal neurons cultured in vitro, as well as the effects of calcium channels on neuronal apoptosis.The chloride channel blockers 4-acetamido-4'-isothiocyanatostilbene-2, 2'-disulfonic acid (SITS) or 4, 4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) increased the survival rate of 3-morpholinosydnonimine (SIN-1)-treated neurons and suppressed SIN-1-induced neuronal apoptosis.The calcium channel blocker CdC12 did not increase the survival rate of neurons and did not affect SIN-1-induced apoptosis or SITS- or DIDS-suppressed neuronal apoptosis.Results demonstrated that calcium channels did not significantly affect neuronal apoptosis.

  10. Aging Reduces L-Type Calcium Channel Current and the Vasodilatory Response of Small Mesenteric Arteries to Calcium Channel Blockers (United States)

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


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

  11. Reversible block of the calcium release channel/ryanodine receptor by protamine, a heparin antidote. (United States)

    Koulen, P; Ehrlich, B E


    Channel activity of the calcium release channel from skeletal muscle, ryanodine receptor type 1, was measured in the presence and absence of protamine sulfate on the cytoplasmic side of the channel. Single-channel activity was measured after incorporating channels into planar lipid bilayers. Optimally and suboptimally calcium-activated calcium release channels were inactivated by the application of protamine to the cytoplasmic side of the channel. Recovery of channel activity was not observed while protamine was present. The addition of protamine bound to agarose beads did not change channel activity, implying that the mechanism of action involves an interaction with the ryanodine receptor rather than changes in the bulk calcium concentration of the medium. The block of channel activity by protamine could be reversed either by removal by perfusion with buffer or by the addition of heparin to the cytoplasmic side of the channel. Microinjection of protamine into differentiated C(2)C(12) mouse muscle cells prevented caffeine-induced intracellular calcium release. The results suggest that protamine acts on the ryanodine receptor in a similar but opposite manner from heparin and that protamine can be used as a potent, reversible inhibitor of ryanodine receptor activity.

  12. Large-conductance calcium-activated potassium channels facilitate transmitter release in salamander rod synapse. (United States)

    Xu, Jian Wei; Slaughter, Malcolm M


    Large-conductance calcium-activated potassium (BK) channels are colocalized with calcium channels at sites of exocytosis at the presynaptic terminals throughout the nervous system. It is expected that their activation would provide negative feedback to transmitter release, but the opposite is sometimes observed. Attempts to resolve this apparent paradox based on alterations in action potential waveform have been ambiguous. In an alternative approach, we investigated the influence of this channel on neurotransmitter release in a nonspiking neuron, the salamander rod photoreceptors. Surprisingly, the BK channel facilitates calcium-mediated transmitter release from rods. The two presynaptic channels form a positive coupled loop. Calcium influx activates the BK channel current, leading to potassium efflux that increases the calcium current. The normal physiological voltage range of the rod is well matched to the dynamics of this positive loop. When the rod is further depolarized, then the hyperpolarizing BK channel current exceeds its facilitatory effect, causing truncation of transmitter release. Thus, the calcium channel-BK channel linkage performs two functions at the synapse: nonlinear potentiator and safety brake.

  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


    Voltage-gated calcium channels (Cav ) are important regulators of vascular tone and are attractive targets for pharmacological treatment of hypertension. The clinical used calcium blockers are often not selective for one channel but affect several types of calcium channels (Hansen 2015). L...

  14. mRNA Expression of Ion Channels in GnRH Neurons: Subtype-Specific Regulation by 17β-Estradiol (United States)

    Bosch, Martha A.; Tonsfeldt, Karen J.; Rønnekleiv, Oline K.


    Burst firing of neurons optimizes neurotransmitter release. GnRH neurons exhibit burst firing activity and T-type calcium channels, which are vital for burst firing activity, are regulated by 17β-estradiol (E2) in GnRH neurons. To further elucidate ion channel expression and E2 regulation during positive and negative feedback on GnRH neurosecretion, we used single cell RT-PCR and real-time qPCR to quantify channel mRNA expression in GnRH neurons. GFP-GnRH neurons expressed numerous ion channels important for burst firing activity. E2-treatment sufficient to induce an LH surge increased mRNA expression of HCN1 channels, which underlie the pacemaker current, the calcium-permeable CaV1.3, CaV2.2, CaV2.3 channels, and TRPC4 channels, which mediate the kisspeptin excitatory response. E2 also decreased mRNA expression of SK3 channels underlying the medium AHP current. Therefore, E2 exerts fundamental changes in ion channel expression in GnRH neurons, to prime them to respond to incoming stimuli with increased excitability at the time of the surge. PMID:23305677

  15. 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


    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.

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

    DEFF Research Database (Denmark)

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


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

    DEFF Research Database (Denmark)

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


    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 mRNA...

  18. Signal processing by T-type calcium channel interactions in the cerebellum (United States)

    Engbers, Jordan D. T.; Anderson, Dustin; Zamponi, Gerald W.; Turner, Ray W.


    T-type calcium channels of the Cav3 family are unique among voltage-gated calcium channels due to their low activation voltage, rapid inactivation, and small single channel conductance. These special properties allow Cav3 calcium channels to regulate neuronal processing in the subthreshold voltage range. Here, we review two different subthreshold ion channel interactions involving Cav3 channels and explore the ability of these interactions to expand the functional roles of Cav3 channels. In cerebellar Purkinje cells, Cav3 and intermediate conductance calcium-activated potassium (IKCa) channels form a novel complex which creates a low voltage-activated, transient outward current capable of suppressing temporal summation of excitatory postsynaptic potentials (EPSPs). In large diameter neurons of the deep cerebellar nuclei, Cav3-mediated calcium current (IT) and hyperpolarization-activated cation current (IH) are activated during trains of inhibitory postsynaptic potentials. These currents have distinct, and yet synergistic, roles in the subthreshold domain with IT generating a rebound burst and IH controlling first spike latency and rebound spike precision. However, by shortening the membrane time constant the membrane returns towards resting value at a faster rate, allowing IH to increase the efficacy of IT and increase the range of burst frequencies that can be generated. The net effect of Cav3 channels thus depends on the channels with which they are paired. When expressed in a complex with a KCa channel, Cav3 channels reduce excitability when processing excitatory inputs. If functionally coupled with an HCN channel, the depolarizing effect of Cav3 channels is accentuated, allowing for efficient inversion of inhibitory inputs to generate a rebound burst output. Therefore, signal processing relies not only on the activity of individual subtypes of channels but also on complex interactions between ion channels whether based on a physical complex or by indirect

  19. Signal processing by T-type calcium channel interactions in the cerebellum

    Directory of Open Access Journals (Sweden)

    Jordan D.T. Engbers


    Full Text Available T-type calcium channels of the Cav3 family are unique among voltage-gated calcium channels due to their low activation voltage, rapid inactivation, and small single channel conductance. These special properties allow Cav3 calcium channels to regulate neuronal processing in the subthreshold voltage range. Here, we review two different subthreshold ion channel interactions involving Cav3 channels and explore the ability of these interactions to expand the functional roles of Cav3 channels. In cerebellar Purkinje cells, Cav3 and intermediate conductance calcium-activated potassium (IKCa channels form a novel complex which creates a low voltage-activated, transient outward current capable of suppressing temporal summation of excitatory postsynaptic potentials (EPSPs. In large diameter neurons of the deep cerebellar nuclei, Cav3-mediated calcium current (IT and hyperpolarization-activated cation current (IH are activated during trains of IPSPs. These currents have distinct, and yet synergistic, roles in the subthreshold domain with IT generating a rebound burst and IH controlling first spike latency and rebound spike precision. However, by shortening the membrane time constant the membrane returns towards resting value at a faster rate, allowing IH to increase the efficacy of IT, and increase the range of burst frequencies that can be generated. The net effect of Cav3 channels thus depends on the channels with which they are paired. When expressed in a complex with a KCa channel, Cav3 channels reduce excitability when processing excitatory inputs. If functionally coupled with an HCN channel, the depolarizing effect of Cav3 channels is accentuated, allowing for efficient inversion of inhibitory inputs to generate a rebound burst output. Therefore, signal processing relies not only on the activity of individual subtypes of channels but also on complex interactions between ion channels whether based on a physical complex or by indirect effects on

  20. Signal processing by T-type calcium channel interactions in the cerebellum. (United States)

    Engbers, Jordan D T; Anderson, Dustin; Zamponi, Gerald W; Turner, Ray W


    T-type calcium channels of the Cav3 family are unique among voltage-gated calcium channels due to their low activation voltage, rapid inactivation, and small single channel conductance. These special properties allow Cav3 calcium channels to regulate neuronal processing in the subthreshold voltage range. Here, we review two different subthreshold ion channel interactions involving Cav3 channels and explore the ability of these interactions to expand the functional roles of Cav3 channels. In cerebellar Purkinje cells, Cav3 and intermediate conductance calcium-activated potassium (IKCa) channels form a novel complex which creates a low voltage-activated, transient outward current capable of suppressing temporal summation of excitatory postsynaptic potentials (EPSPs). In large diameter neurons of the deep cerebellar nuclei, Cav3-mediated calcium current (I T) and hyperpolarization-activated cation current (I H) are activated during trains of inhibitory postsynaptic potentials. These currents have distinct, and yet synergistic, roles in the subthreshold domain with I T generating a rebound burst and I H controlling first spike latency and rebound spike precision. However, by shortening the membrane time constant the membrane returns towards resting value at a faster rate, allowing I H to increase the efficacy of I T and increase the range of burst frequencies that can be generated. The net effect of Cav3 channels thus depends on the channels with which they are paired. When expressed in a complex with a KCa channel, Cav3 channels reduce excitability when processing excitatory inputs. If functionally coupled with an HCN channel, the depolarizing effect of Cav3 channels is accentuated, allowing for efficient inversion of inhibitory inputs to generate a rebound burst output. Therefore, signal processing relies not only on the activity of individual subtypes of channels but also on complex interactions between ion channels whether based on a physical complex or by indirect

  1. Calcium-permeable ion channels in control of autophagy and cancer. (United States)

    Kondratskyi, Artem; Yassine, Maya; Kondratska, Kateryna; Skryma, Roman; Slomianny, Christian; Prevarskaya, Natalia


    Autophagy, or cellular self-eating, is a tightly regulated cellular pathway the main purpose of which is lysosomal degradation and subsequent recycling of cytoplasmic material to maintain normal cellular homeostasis. Defects in autophagy are linked to a variety of pathological states, including cancer. Cancer is the disease associated with abnormal tissue growth following an alteration in such fundamental cellular processes as apoptosis, proliferation, differentiation, migration and autophagy. The role of autophagy in cancer is complex, as it can promote both tumor prevention and survival/treatment resistance. It's now clear that modulation of autophagy has a great potential in cancer diagnosis and treatment. Recent findings identified intracellular calcium as an important regulator of both basal and induced autophagy. Calcium is a ubiquitous secondary messenger which regulates plethora of physiological and pathological processes such as aging, neurodegeneration and cancer. The role of calcium and calcium-permeable channels in cancer is well-established, whereas the information about molecular nature of channels regulating autophagy and the mechanisms of this regulation is still limited. Here we review existing mechanisms of autophagy regulation by calcium and calcium-permeable ion channels. Furthermore, we will also discuss some calcium-permeable channels as the potential new candidates for autophagy regulation. Finally we will propose the possible link between calcium permeable channels, autophagy and cancer progression and therapeutic response.

  2. (S)-Lacosamide Binding to Collapsin Response Mediator Protein 2 (CRMP2) Regulates CaV2.2 Activity by Subverting Its Phosphorylation by Cdk5. (United States)

    Moutal, Aubin; François-Moutal, Liberty; Perez-Miller, Samantha; Cottier, Karissa; Chew, Lindsey Anne; Yeon, Seul Ki; Dai, Jixun; Park, Ki Duk; Khanna, May; Khanna, Rajesh


    The neuronal circuit remodels during development as well as in human neuropathologies such as epilepsy. Neurite outgrowth is an obligatory step in these events. We recently reported that alterations in the phosphorylation state of an axon specification/guidance protein, the collapsin response mediator protein 2 (CRMP2), play a major role in the activity-dependent regulation of neurite outgrowth. We also identified (S)-LCM, an inactive stereoisomer of the clinically used antiepileptic drug (R)-LCM (Vimpat®), as a novel tool for preferentially targeting CRMP2-mediated neurite outgrowth. Here, we investigated the mechanism by which (S)-LCM affects CRMP2 phosphorylation by two key kinases, cyclin-dependent kinase 5 (Cdk5) and glycogen synthase kinase 3β (GSK-3β). (S)-LCM application to embryonic cortical neurons resulted in reduced levels of Cdk5- and GSK-3β-phosphorylated CRMP2. Mechanistically, (S)-LCM increased CRMP2 binding to both Cdk5- and GSK-3β without affecting binding of CRMP2 to its canonical partner tubulin. Saturation transfer difference nuclear magnetic resonance (STD NMR) and differential scanning fluorimetry (DSF) experiments demonstrated direct binding of (S)-LCM to CRMP2. Using an in vitro luminescent kinase assay, we observed that (S)-LCM specifically inhibited Cdk5-mediated phosphorylation of CRMP2. Cross-linking experiments and analytical ultracentrifugation showed no effect of (S)-LCM on the oligomerization state of CRMP2. The increased association between Cdk5-phosphorylated CRMP2 and CaV2.2 was reduced by (S)-LCM in vitro and in vivo. This reduction translated into a decrease of calcium influx via CaV2.2 in (S)-LCM-treated neurons compared to controls. (S)-LCM, to our knowledge, is the first molecule described to directly inhibit CRMP2 phosphorylation and may be useful for delineating CRMP2-facilitated functions.

  3. Emerging roles of L-type voltage gated and other calcium channels in T lymphocytes.

    Directory of Open Access Journals (Sweden)

    Abdallah eBadou


    Full Text Available In T lymphocytes, calcium ion controls a variety of biological processes including development, survival, proliferation, and effector functions. These distinct and specific roles are regulated by different calcium signals, which are generated by various plasma membrane calcium channels. The repertoire of calcium-conducting proteins in T lymphocytes includes store-operated CRAC channels, transient receptor potential (TRP channels, P2X channels, and L-type voltage-gated calcium (Cav1 channels. In this paper, we will focus mainly on the role of the Cav1 channels found expressed by T lymphocytes, where these channels appear to operate in a TCR stimulation-dependent and voltage-sensor independent manner. We will review their expression profile at various differentiation stages of CD4 and CD8 T lymphocytes. Then, we will present crucial genetic evidence in favor of a role of these Cav1 channels and related regulatory proteins in both CD4 and CD8 T cell functions such as proliferation, survival, cytokine production and cytolysis. Finally, we will provide evidence and speculate on how these voltage-gated channels might function in the T lymphocyte, a non-excitable cell.

  4. Ryanodine Receptors Selectively Interact with L Type Calcium Channels in Mouse Taste Cells.

    Directory of Open Access Journals (Sweden)

    Michelle R Rebello

    Full Text Available WE REPORTED THAT RYANODINE RECEPTORS ARE EXPRESSED IN TWO DIFFERENT TYPES OF MAMMALIAN PERIPHERAL TASTE RECEPTOR CELLS: Type II and Type III cells. Type II cells lack voltage-gated calcium channels (VGCCs and chemical synapses. In these cells, ryanodine receptors contribute to the taste-evoked calcium signals that are initiated by opening inositol trisphosphate receptors located on internal calcium stores. In Type III cells that do have VGCCs and chemical synapses, ryanodine receptors contribute to the depolarization-dependent calcium influx.The goal of this study was to establish if there was selectivity in the type of VGCC that is associated with the ryanodine receptor in the Type III taste cells or if the ryanodine receptor opens irrespective of the calcium channels involved. We also wished to determine if the ryanodine receptors and VGCCs require a physical linkage to interact or are simply functionally associated with each other. Using calcium imaging and pharmacological inhibitors, we found that ryanodine receptors are selectively associated with L type VGCCs but likely not through a physical linkage.Taste cells are able to undergo calcium induced calcium release through ryanodine receptors to increase the initial calcium influx signal and provide a larger calcium response than would otherwise occur when L type channels are activated in Type III taste cells.

  5. Exploring the dominant role of Cav1 channels in signalling to the nucleus. (United States)

    Ma, Huan; Cohen, Samuel; Li, Boxing; Tsien, Richard W


    Calcium is important in controlling nuclear gene expression through the activation of multiple signal-transduction pathways in neurons. Compared with other voltage-gated calcium channels, Ca(V)1 channels demonstrate a considerable advantage in signalling to the nucleus. In this review, we summarize the recent progress in elucidating the mechanisms involved. Ca(V)1 channels, already advantaged in their responsiveness to depolarization, trigger communication with the nucleus by attracting colocalized clusters of activated CaMKII (Ca(2+)/calmodulin-dependent protein kinase II). Ca(V)2 channels lack this ability, but must work at a distance of >1 μm from the Ca(V)1-CaMKII co-clusters, which hampers their relative efficiency for a given rise in bulk [Ca(2+)](i) (intracellular [Ca(2+)]). Moreover, Ca(2+) influx from Ca(V)2 channels is preferentially buffered by the ER (endoplasmic reticulum) and mitochondria, further attenuating their effectiveness in signalling to the nucleus.

  6. Exploring the dominant role of Cav1 channels in signalling to the nucleus (United States)

    Ma, Huan; Cohen, Samuel; Li, Boxing; Tsien, Richard W.


    Calcium is important in controlling nuclear gene expression through the activation of multiple signal-transduction pathways in neurons. Compared with other voltage-gated calcium channels, CaV1 channels demonstrate a considerable advantage in signalling to the nucleus. In this review, we summarize the recent progress in elucidating the mechanisms involved. CaV1 channels, already advantaged in their responsiveness to depolarization, trigger communication with the nucleus by attracting colocalized clusters of activated CaMKII (Ca2+/calmodulin-dependent protein kinase II). CaV2 channels lack this ability, but must work at a distance of >1 μm from the CaV1-CaMKII co-clusters, which hampers their relative efficiency for a given rise in bulk [Ca2+]i (intracellular [Ca2+]). Moreover, Ca2+ influx from CaV2 channels is preferentially buffered by the ER (endoplasmic reticulum) and mitochondria, further attenuating their effectiveness in signalling to the nucleus. PMID:23088728

  7. The coupling of acetylcholine-induced BK channel and calcium channel in guinea pig saccular type II vestibular hair cells. (United States)

    Kong, Wei-Jia; Guo, Chang-Kai; Zhang, Xiao-Wen; Chen, Xiong; Zhang, Song; Li, Guan-Qiao; Li, Zhi-Wang; Van Cauwenberge, Paul


    Molecular biological studies and electrophysiological data have demonstrated that acetylcholine (ACh) is the principal cochlear and vestibular efferent neurotransmitter among mammalians. However, the functional roles of ACh in type II vestibular hair cells (VHCs II) among mammalians are still unclear, with the exception of the well-known alpha9-containing nicotinic ACh receptor (alpha9-containing nAChR)-activated small conductance, calcium-dependent potassium current (SK) in cochlear hair cells and frog saccular hair cells. The activation of SK current was necessary for the calcium influx through the alpha9-containing nAChR. Recently, we have demonstrated that ACh-induced big conductance, calcium-dependent potassium current (BK) was present in VHCs II of the vestibular end-organ of guinea pig. In this study, the nature of calcium influx for the activation of ACh-induced BK current in saccular VHCs II of guinea pig was investigated. Following extracellular perfusion of ACh, saccular VHCs II displayed a sustained outward current, which was sensitive to iberiotoxin (IBTX). High concentration of apamin failed to inhibit the current amplitude of ACh-induced outward current. Intracellular application of Cs(+) completely abolished the current evoked by ACh. ACh-induced current was potently inhibited by nifedipine, nimodipine, Cd(2+) and Ni(2+), respectively. The inhibition potency of these four calcium channel antagonists was nimodipine>nifedipine>cadmium>nickel. The L-type Ca(2+) channels agonist, (-)-Bay-K 8644 mimicked the effect of ACh and activated an IBTX-sensitive current. In addition, partial VHCs II displayed a biphasic waveform. In conclusion, the present data showed that in the guinea pig saccular VHCs II, ACh-induced BK channel was coupled with the calcium channel, but not the receptor. The perfusion of ACh will drive the opening of calcium channels; the influx of calcium ions will then activate the BK current.

  8. Spontaneous and CRH-Induced Excitability and Calcium Signaling in Mice Corticotrophs Involves Sodium, Calcium, and Cation-Conducting Channels. (United States)

    Zemkova, Hana; Tomić, Melanija; Kucka, Marek; Aguilera, Greti; Stojilkovic, Stanko S


    Transgenic mice expressing the tdimer2(12) form of Discosoma red fluorescent protein under control of the proopiomelanocortin gene's regulatory elements are a useful model for studying corticotrophs. Using these mice, we studied the ion channels and mechanisms controlling corticotroph excitability. Corticotrophs were either quiescent or electrically active, with a 22-mV difference in the resting membrane potential (RMP) between the 2 groups. In quiescent cells, CRH depolarized the membrane, leading to initial single spiking and sustained bursting; in active cells, CRH further facilitated or inhibited electrical activity and calcium spiking, depending on the initial activity pattern and CRH concentration. The stimulatory but not inhibitory action of CRH on electrical activity was mimicked by cAMP independently of the presence or absence of arachidonic acid. Removal of bath sodium silenced spiking and hyperpolarized the majority of cells; in contrast, the removal of bath calcium did not affect RMP but reduced CRH-induced depolarization, which abolished bursting electrical activity and decreased the spiking frequency but not the amplitude of single spikes. Corticotrophs with inhibited voltage-gated sodium channels fired calcium-dependent action potentials, whereas cells with inhibited L-type calcium channels fired sodium-dependent spikes; blockade of both channels abolished spiking without affecting the RMP. These results indicate that the background voltage-insensitive sodium conductance influences RMP, the CRH-depolarization current is driven by a cationic conductance, and the interplay between voltage-gated sodium and calcium channels plays a critical role in determining the status and pattern of electrical activity and calcium signaling.

  9. 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;


    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....... Low concentrations of nickel, an agent that blocks Ca(v)3.2, had a similar effect. Thus, T-type voltage-gated calcium channels are functionally important for depolarization-induced vasoconstriction and subsequent dilatation in mouse cortical efferent arterioles.Kidney International advance online...

  10. Tryptophan hydroxylase is modulated by L-type calcium channels in the rat pineal gland. (United States)

    Barbosa, Roseli; Scialfa, Julieta Helena; Terra, Ilza Mingarini; Cipolla-Neto, José; Simonneaux, Valérie; Afeche, Solange Castro


    Calcium is an important second messenger in the rat pineal gland, as well as cAMP. They both contribute to melatonin synthesis mediated by the three main enzymes of the melatonin synthesis pathway: tryptophan hydroxylase, arylalkylamine N-acetyltransferase and hydroxyindole-O-methyltransferase. The cytosolic calcium is elevated in pinealocytes following alpha(1)-adrenergic stimulation, through IP(3)-and membrane calcium channels activation. Nifedipine, an L-type calcium channel blocker, reduces melatonin synthesis in rat pineal glands in vitro. With the purpose of investigating the mechanisms involved in melatonin synthesis regulation by the L-type calcium channel, we studied the effects of nifedipine on noradrenergic stimulated cultured rat pineal glands. Tryptophan hydroxylase, arylalkylamine N-acetyltransferase and hydroxyindole-O-methyltransferase activities were quantified by radiometric assays and 5-hydroxytryptophan, serotonin, N-acetylserotonin and melatonin contents were quantified by HPLC with electrochemical detection. The data showed that calcium influx blockaded by nifedipine caused a decrease in tryptophan hydroxylase activity, but did not change either arylalkylamine N-acetyltransferase or hydroxyindole-O-methyltransferase activities. Moreover, there was a reduction of 5-hydroxytryptophan, serotonin, N-acetylserotonin and melatonin intracellular content, as well as a reduction of serotonin and melatonin secretion. Thus, it seems that the calcium influx through L-type high voltage-activated calcium channels is essential for the full activation of tryptophan hydroxylase leading to melatonin synthesis in the pineal gland.

  11. Dihydropyridine type calcium channel blocker-induced turbid dialysate in patients undergoing peritoneal dialysis. (United States)

    Yoshimoto, K; Saima, S; Nakamura, Y; Nakayama, M; Kubo, H; Kawaguchi, Y; Nishitani, H; Nakamura, Y; Yasui, A; Yokoyama, K; Kuriyama, S; Shirai, D; Kugiyama, A; Hayano, K; Fukui, H; Horigome, I; Amagasaki, Y; Tsubakihara, Y; Kamekawa, T; Ando, R; Tomura, S; Okamoto, R; Miwa, S; Koyama, T; Echizen, H


    We previously reported that manidipine, a new dihydropyridine type calcium channel blocker, produced chylous peritoneal dialysate being visually indistinguishable from infective peritonitis in 5 patients undergoing continuous ambulatory peritoneal dialysis (CAPD) [Yoshimoto et al. 1993]. To study whether such an adverse drug reaction would also be elicited by other commonly prescribed calcium channel blockers in CAPD patients, we have conducted postal inquiry to 15 collaborating hospitals and an institutional survey in International Medical Center of Japan as to the possible occurrence of calcium channel blocker-associated non-infective, turbid peritoneal dialysate in CAPD patients. Our diagnostic criteria for drug-induced turbidity of dialysate as a) it developed within 48 h after the administration of a newly introduced calcium channel blocker to the therapeutic regimen, b) absence of clinical symptoms of peritoneal inflammation (i.e., pyrexia, abdominal pain, nausea or vomiting), c) the fluid containing normal leukocyte counts and being negative for bacterial and fungal culture of the fluid, and d) it disappeared shortly after the withdrawal of the assumed causative agent. Results showed that 19 out of 251 CAPD patients given one of the calcium channel blockers developed non-infective turbid peritoneal dialysis that fulfilled all the above criteria. Four calcium channel blockers were suspected to be associated with the events: benidipine [2 out of 2 (100%) patients given the drug], manidipine [15 out of 36 (42%) patients], nisoldipine [1 out of 11 (9%) patients] and nifedipine [1 out of 159 (0.6%)] in descending order of frequency. None of the patients who received nicardipine, nilvadipine, nitrendipine, barnidipine and diltiazem (25, 7, 2, 1 and 8 patients, respectively) exhibited turbid dialysate. In conclusion, we consider that certain dihydropyridine type calcium channel blockers would cause turbid peritoneal dialysate being similar to that observed in

  12. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells (United States)

    Jorgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne; Civitelli, Roberto; Sorensen, Ole Helmer; Steinberg, Thomas H.


    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 Cx43 (UMR/Cx43) we confirmed that nifedipine sensitivity of ICW required Cx43 expression. In human osteoblastic cells, gap junction-dependent ICW also required activation of L-type calcium channels and influx of extracellular calcium.

  13. T-type calcium channel expression in cultured human neuroblastoma cells

    Institute of Scientific and Technical Information of China (English)

    Xianjie Wen; Shiyuan Xu; Lingling Wang; Hua Liang; Chengxiang Yang; Hanbing Wang; Hongzhen Liu


    Human neuroblastoma cells (SH-SY5Y) have similar structures and functions as neural cells and have been frequently used for cell culture studies of neural cell functions. Previous studies have revealed Land N-type calcium channels in SH-SY5Y cells. However, the distribution of the low -voltage activated calcium channel (namely called T-type calcium channel, including Cav3.1, Cav3.2, and Cav3.3) in SH-SY5Y cells remains poorly understood. The present study detected mRNA and protein expression of the T-type calcium channel (Cav3.1, Cav3.2, and Cav3.3) in cultured SH-SY5Y cells using real-time polymerase chain reaction (PCR) and western blot analysis. Results revealed mRNA and protein expression from all three T-type calcium channel subtypes in SH-SY5Y cells. Moreover,Cav3.1 was the predominant T-type calcium channel subtype in SH-SY5Y cells.

  14. Osteoclast cytosolic calcium, regulated by voltage-gated calcium channels and extracellular calcium, controls podosome assembly and bone resorption (United States)

    Miyauchi, A.; Hruska, K. A.; Greenfield, E. M.; Duncan, R.; Alvarez, J.; Barattolo, R.; Colucci, S.; Zambonin-Zallone, A.; Teitelbaum, S. L.; Teti, A.


    The mechanisms of Ca2+ entry and their effects on cell function were investigated in cultured chicken osteoclasts and putative osteoclasts produced by fusion of mononuclear cell precursors. Voltage-gated Ca2+ channels (VGCC) were detected by the effects of membrane depolarization with K+, BAY K 8644, and dihydropyridine antagonists. K+ produced dose-dependent increases of cytosolic calcium ([Ca2+]i) in osteoclasts on glass coverslips. Half-maximal effects were achieved at 70 mM K+. The effects of K+ were completely inhibited by dihydropyridine derivative Ca2+ channel blocking agents. BAY K 8644 (5 X 10(-6) M), a VGCC agonist, stimulated Ca2+ entry which was inhibited by nicardipine. VGCCs were inactivated by the attachment of osteoclasts to bone, indicating a rapid phenotypic change in Ca2+ entry mechanisms associated with adhesion of osteoclasts to their resorption substrate. Increasing extracellular Ca2+ ([Ca2+]e) induced Ca2+ release from intracellular stores and Ca2+ influx. The Ca2+ release was blocked by dantrolene (10(-5) M), and the influx by La3+. The effects of [Ca2+]e on [Ca2+]i suggests the presence of a Ca2+ receptor on the osteoclast cell membrane that could be coupled to mechanisms regulating cell function. Expression of the [Ca2+]e effect on [Ca2+]i was similar in the presence or absence of bone matrix substrate. Each of the mechanisms producing increases in [Ca2+]i, (membrane depolarization, BAY K 8644, and [Ca2+]e) reduced expression of the osteoclast-specific adhesion structure, the podosome. The decrease in podosome expression was mirrored by a 50% decrease in bone resorptive activity. Thus, stimulated increases of osteoclast [Ca2+]i lead to cytoskeletal changes affecting cell adhesion and decreasing bone resorptive activity.

  15. 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


    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...

  16. Differential CaMKII regulation by voltage-gated calcium channels in the striatum. (United States)

    Pasek, Johanna G; Wang, Xiaohan; Colbran, Roger J


    Calcium signaling regulates synaptic plasticity and many other functions in striatal medium spiny neurons to modulate basal ganglia function. Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is a major calcium-dependent signaling protein that couples calcium entry to diverse cellular changes. CaMKII activation results in autophosphorylation at Thr286 and sustained calcium-independent CaMKII activity after calcium signals dissipate. However, little is known about the mechanisms regulating striatal CaMKII. To address this, mouse brain slices were treated with pharmacological modulators of calcium channels and punches of dorsal striatum were immunoblotted for CaMKII Thr286 autophosphorylation as an index of CaMKII activation. KCl depolarization increased levels of CaMKII autophosphorylation ~2-fold; this increase was blocked by an LTCC antagonist and was mimicked by treatment with pharmacological LTCC activators. The chelation of extracellular calcium robustly decreased basal CaMKII autophosphorylation within 5min and increased levels of total CaMKII in cytosolic fractions, in addition to decreasing the phosphorylation of CaMKII sites in the GluN2B subunit of NMDA receptors and the GluA1 subunit of AMPA receptors. We also found that the maintenance of basal levels of CaMKII autophosphorylation requires low-voltage gated T-type calcium channels, but not LTCCs or R-type calcium channels. Our findings indicate that CaMKII activity is dynamically regulated by multiple calcium channels in the striatum thus coupling calcium entry to key downstream substrates.

  17. Multi-ion conduction bands in a simple model of calcium ion channels

    CERN Document Server

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


    We report self-consistent Brownian dynamics simulations of a simple electrostatic model of the selectivity filters (SF) of calcium ion channels. They reveal regular structure in the conductance and selectivity as functions of the fixed negative charge Qf at the SF. This structure comprises distinct regions of high conductance (conduction bands) M0, M1, M2 separated by regions of zero-conductance (stop-bands). Two of these conduction bands, M1 and M2, demonstrate high calcium selectivity and prominent anomalous mole fraction effects and can be identified with the L-type and RyR calcium channels.

  18. A different dihydropyridine calcium channel blocker in hypertensive patients who developed pedal edema on dihydropyridine calcium channel blocker therapy

    Directory of Open Access Journals (Sweden)

    Ayşe Yüksel


    Full Text Available Abstract Aim. Dihydropyridine calcium channel blockers (CCB are widely preferred for the treatment of hypertension for their efficacy, metabolic neutrality and low side effect profile. However pedal edema formation limits their usage. The aim of the present study is to evaluate the incidence of pedal edema formation with a different dihydropyridine CCB in hypertensive patients who developed pedal edema during a dihydropyridine CCB therapy. Method. Fifty-eight hypertensive patients (34 female, 24 male, mean age: 65.3±10.5 in whom pedal edema developed during treatment with a dihydropyridine CCB (amlodipine 10mg/day in 40 patients, amlodipine 5mg/day in 14 patients, nifedipine GITS 30mg/day in 4 patients were enrolled. CCB which caused pedal edema was withdrawn and a different CCB (felodipine or lacidipine were initiated after the resolution of the pedal edema. CCB therapy was continued as long as the patient tolerated pedal edema. Results. At the end of one year, 44 out of 58 patients (36 [81.8%] free of pedal edema, 8 [19.2%] with pedal edema continued CCB therapy. Eleven (37.9% patients in the felodipine group and 9 (31.0% patients in the lacidipine group developed pedal edema. In 7 patients in felodipine group and in 5 patients in the lacidipine group the study drug was withdrawn due to pedal edema. In two patients, study drug was withdrawn due to intractable headache (felodipine group or due to flushing (lacidipine group. Conclusion. A different group of dihydropyridine CCB be used as an alternative therapy for hypertension whenever pedal edema develops during treatment with a dihydropyridine CCB.

  19. Synthesis, QSAR and calcium channel modulator activity of new hexahydroquinoline derivatives containing nitroimidazole. (United States)

    Miri, Ramin; Javidnia, Katayoun; Mirkhani, Hossein; Hemmateenejad, Bahram; Sepeher, Zahra; Zalpour, Masomeh; Behzad, Taherh; Khoshneviszadeh, Mehdi; Edraki, Najmeh; Mehdipour, Ahmad R


    The discovery that 1,4-dihydropyridine class of calcium channel antagonists inhibit Ca2+ influx represented a major therapeutic advance in the treatment of cardiovascular disease. In contrast to the effects of known calcium channel blockers of the Nifedipine-type, the so-called calcium channel agonists, such as Bay K8644 and CGP 28392, increase calcium influx by binding at the same receptor regions. Our goal was to discover a dual cardioselective Ca2+-channel agonist/vascular selective smooth muscle Ca2+ channel antagonist third-generation 1,4-dihydropyridine drug which would have a suitable therapeutic profile for treating congestive heart failure (CHF) patients. A series of unsymmetrical alkyl, cycloalkyl and aryl ester analogues of 2-methyl-4-(1-methyl)-5-nitro-2-imidazolyl-5-oxo-1,4,5,6,7, 8-hexahydroquinolin-3-arboxylate were synthesized using modified Hantzsch reaction. All compounds show calcium antagonist activity on guinea-pig ileum longitudinal smooth muscle and some of them show agonist effect activity on guinea-pig auricle. Effect of structural parameters on the Ca2+ channel agonist/antagonist was evaluated by quantitative structure-activity relationship analysis. These compounds could be considered as a synthon for developing a suitable drug for treating CHF patients.

  20. 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 Brings; Aalkjær, Christian; Nilsson, Holger;


    waves sweeping through the cytoplasm when the sarcoplasmic reticulum (SR) is stimulated to release calcium. A rise in 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 a uniform opening of L-type calcium...

  1. The effects of inorganic lead on voltage-sensitive calcium channels differ among cell types and among channel subtypes. (United States)

    Audesirk, G; Audesirk, T


    The whole-cell version of patch clamping was used to compare the effects of acute in vitro exposure to inorganic lead (Pb2+) on voltage-sensitive calcium channels in cultured N1E-115 mouse neuroblastoma cells and E18 rat hippocampal neurons. Free Pb2+ concentrations in salines with a high lead-buffering capacity were measured with a calibrated Pb(2+)-selective electrode. Previously, we found that N1E-115 neurons contain low voltage activated, rapidly inactivating (T) channels and high voltage activated, slowly inactivating (L) channels. Pb2+ inhibits both channel subtypes in N1E-115 cells, with some selectivity against L-type channels (IC50 approximately 700 nM free Pb2+ for L-type channels, 1300 nM free Pb2+ for T-type channels; Audesirk and Audesirk, 1991). In addition to T-type and L-type channels, cultured E18 rat hippocampal neurons have been reported to contain high voltage-activated, rapidly inactivating (N) channels. In our experiments with 5 to 20 day old cultures, almost all neurons showed substantial L-type current, approximately half showed significant N-type current, and fewer than 5% showed significant T-type current. We found that Pb2+ is somewhat selective against L-type channels (IC50 approximately 30 nM free Pb2+ in 10 mM Ba2+ as the charge carrier, 55 nM in 50 mM Ba2+) compared to N-channels (IC50 approximately 80 nM free Pb2+ in 10 mM Ba2+, 200 nM in 50 mM Ba2+). These results suggest that the effects of Pb2+ on calcium channels of vertebrate neurons vary both among cell types and among channel subtypes.

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


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


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

  3. Transcriptional Response of Human Neurospheres to Helper-Dependent CAV-2 Vectors Involves the Modulation of DNA Damage Response, Microtubule and Centromere Gene Groups.

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    Stefania Piersanti

    Full Text Available Brain gene transfer using viral vectors will likely become a therapeutic option for several disorders. Helper-dependent (HD canine adenovirus type 2 vectors (CAV-2 are well suited for this goal. These vectors are poorly immunogenic, efficiently transduce neurons, are retrogradely transported to afferent structures in the brain and lead to long-term transgene expression. CAV-2 vectors are being exploited to unravel behavior, cognition, neural networks, axonal transport and therapy for orphan diseases. With the goal of better understanding and characterizing HD-CAV-2 for brain therapy, we analyzed the transcriptomic modulation induced by HD-CAV-2 in human differentiated neurospheres derived from midbrain progenitors. This 3D model system mimics several aspects of the dynamic nature of human brain. We found that differentiated neurospheres are readily transduced by HD-CAV-2 and that transduction generates two main transcriptional responses: a DNA damage response and alteration of centromeric and microtubule probes. Future investigations on the biochemistry of processes highlighted by probe modulations will help defining the implication of HD-CAV-2 and CAR receptor binding in enchaining these functional pathways. We suggest here that the modulation of DNA damage genes is related to viral DNA, while the alteration of centromeric and microtubule probes is possibly enchained by the interaction of the HD-CAV-2 fibre with CAR.

  4. Voltage gated calcium channels negatively regulate protective immunity to Mycobacterium tuberculosis.

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    Shashank Gupta

    Full Text Available Mycobacterium tuberculosis modulates levels and activity of key intracellular second messengers to evade protective immune responses. Calcium release from voltage gated calcium channels (VGCC regulates immune responses to pathogens. In this study, we investigated the roles of VGCC in regulating protective immunity to mycobacteria in vitro and in vivo. Inhibiting L-type or R-type VGCC in dendritic cells (DCs either using antibodies or by siRNA increased calcium influx in an inositol 1,4,5-phosphate and calcium release calcium activated channel dependent mechanism that resulted in increased expression of genes favoring pro-inflammatory responses. Further, VGCC-blocked DCs activated T cells that in turn mediated killing of M. tuberculosis inside macrophages. Likewise, inhibiting VGCC in infected macrophages and PBMCs induced calcium influx, upregulated the expression of pro-inflammatory genes and resulted in enhanced killing of intracellular M. tuberculosis. Importantly, compared to healthy controls, PBMCs of tuberculosis patients expressed higher levels of both VGCC, which were significantly reduced following chemotherapy. Finally, blocking VGCC in vivo in M. tuberculosis infected mice using specific antibodies increased intracellular calcium and significantly reduced bacterial loads. These results indicate that L-type and R-type VGCC play a negative role in M. tuberculosis infection by regulating calcium mobilization in cells that determine protective immunity.


    Ju, Weina; Wu, Jiang; Pritz, Michael B.; Khanna, Rajesh


    Vertebrate brains share many features in common. Early in development, both the hindbrain and diencephalon are built similarly. Only later in time do differences in morphology occur. Factors that could potentially influence such changes include certain physiological properties of neurons. As an initial step to investigate this problem, embryonic Alligator brain neurons were cultured and calcium responses were characterized. The present report is the first to document culture of Alligator brain neurons in artificial cerebrospinal fluid (ACSF) as well as in standard mammalian tissue culture medium supplemented with growth factors. Alligator brain neuron cultures were viable for at least 1 week with unipolar neurites emerging by 24 hours. Employing Fura-2 AM, robust depolarization-induced calcium influx, was observed in these neurons. Using selective blockers of the voltage-gated calcium channels, the contributions of N-, P/Q-, R-, T-, and L-type channels in these neurons were assessed and their presence documented. Lastly, Alligator brain neurons were challenged with an excitotoxic stimulus (glutamate + glycine) where delayed calcium deregulation could be prevented by a classical NMDA receptor antagonist. PMID:24260711

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

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    Jean-Yves eTano


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

  7. Calcium homeostasis and cone signaling are regulated by interactions between calcium stores and plasma membrane ion channels.

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    Tamas Szikra

    Full Text Available Calcium is a messenger ion that controls all aspects of cone photoreceptor function, including synaptic release. The dynamic range of the cone output extends beyond the activation threshold for voltage-operated calcium entry, suggesting another calcium influx mechanism operates in cones hyperpolarized by light. We have used optical imaging and whole-cell voltage clamp to measure the contribution of store-operated Ca(2+ entry (SOCE to Ca(2+ homeostasis and its role in regulation of neurotransmission at cone synapses. Mn(2+ quenching of Fura-2 revealed sustained divalent cation entry in hyperpolarized cones. Ca(2+ influx into cone inner segments was potentiated by hyperpolarization, facilitated by depletion of intracellular Ca(2+ stores, unaffected by pharmacological manipulation of voltage-operated or cyclic nucleotide-gated Ca(2+ channels and suppressed by lanthanides, 2-APB, MRS 1845 and SKF 96365. However, cation influx through store-operated channels crossed the threshold for activation of voltage-operated Ca(2+ entry in a subset of cones, indicating that the operating range of inner segment signals is set by interactions between store- and voltage-operated Ca(2+ channels. Exposure to MRS 1845 resulted in approximately 40% reduction of light-evoked postsynaptic currents in photopic horizontal cells without affecting the light responses or voltage-operated Ca(2+ currents in simultaneously recorded cones. The spatial pattern of store-operated calcium entry in cones matched immunolocalization of the store-operated sensor STIM1. These findings show that store-operated channels regulate spatial and temporal properties of Ca(2+ homeostasis in vertebrate cones and demonstrate their role in generation of sustained excitatory signals across the first retinal synapse.

  8. T Cell Receptor Mediated Calcium Entry Requires Alternatively Spliced Cav1.1 Channels.

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    Didi Matza

    Full Text Available The process of calcium entry in T cells is a multichannel and multi-step process. We have studied the requirement for L-type calcium channels (Cav1.1 α1S subunits during calcium entry after TCR stimulation. High expression levels of Cav1.1 channels were detected in activated T cells. Sequencing and cloning of Cav1.1 channel cDNA from T cells revealed that a single splice variant is expressed. This variant lacks exon 29, which encodes the linker region adjacent to the voltage sensor, but contains five new N-terminal exons that substitute for exons 1 and 2, which are found in the Cav1.1 muscle counterpart. Overexpression studies using cloned T cell Cav1.1 in 293HEK cells (that lack TCR suggest that the gating of these channels was altered. Knockdown of Cav1.1 channels in T cells abrogated calcium entry after TCR stimulation, suggesting that Cav1.1 channels are controlled by TCR signaling.

  9. Comparative pharmacodynamics of eight calcium channel blocking agents in Japanese essential hypertensive patients. (United States)

    Shimada, S; Nakajima, Y; Yamamoto, K; Sawada, Y; Iga, T


    The relationships between plasma drug concentration and antihypertensive effect of eight calcium channel antagonists (nicardipine, nifedipine, nilvadipine, benidipine, manidipine, barnidipine, nitrendipine and efonidipine) in Japanese essential hypertensive patients were analyzed. Based on the effect compartment model, we could explain the long duration of the pharmacological effect, and there was significant correlation (r = 0.876, p < 0.05) between estimated EC50 values and the dissociation constants (Kd) obtained from in vitro binding studies. We also developed the ion-channel binding model to understand the pharmacodynamics of long acting calcium antagonists. The model was also well fitted to antihypertensive effect data. A significant correlation between the apparent in vivo dissociation constants and in vitro Kd values was observed with a slope of 1.45 (r = 0.913), suggesting that the mechanism of long-lasting antihypertensive effect of newer developed calcium antagonists is due to their high binding affinity at ion-channel sites.

  10. 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;


    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....

  11. Voltage-dependent calcium channels from brain incorporated into planar lipid bilayers (United States)

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


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

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

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    Akira Katoh

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

  13. Calcium-activated potassium channels sustain calcium signaling in T lymphocytes. Selective blockers and manipulated channel expression levels. (United States)

    Fanger, C M; Rauer, H; Neben, A L; Miller, M J; Rauer, H; Wulff, H; Rosa, J C; Ganellin, C R; Chandy, K G; Cahalan, M D


    To maintain Ca(2+) entry during T lymphocyte activation, a balancing efflux of cations is necessary. Using three approaches, we demonstrate that this cation efflux is mediated by Ca(2+)-activated K(+) (K(Ca)) channels, hSKCa2 in the human leukemic T cell line Jurkat and hIKCa1 in mitogen-activated human T cells. First, several recently developed, selective and potent pharmacological inhibitors of K(Ca) channels but not K(V) channels reduce Ca(2+) entry in Jurkat and in mitogen-activated human T cells. Second, dominant-negative suppression of the native K(Ca) channel in Jurkat T cells by overexpression of a truncated fragment of the cloned hSKCa2 channel decreases Ca(2+) influx. Finally, introduction of the hIKCa1 channel into Jurkat T cells maintains rapid Ca(2+) entry despite pharmacological inhibition of the native small conductance K(Ca) channel. Thus, K(Ca) channels play a vital role in T cell Ca(2+) signaling.

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

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    Masahiro Kawahara


    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.

  15. Microdamage induced calcium efflux from bone matrix activates intracellular calcium signaling in osteoblasts via L-type and T-type voltage-gated calcium channels. (United States)

    Jung, Hyungjin; Best, Makenzie; Akkus, Ozan


    Mechanisms by which bone microdamage triggers repair response are not completely understood. It has been shown that calcium efflux ([Ca(2+)]E) occurs from regions of bone undergoing microdamage. Such efflux has also been shown to trigger intracellular calcium signaling ([Ca(2+)]I) in MC3T3-E1 cells local to damaged regions. Voltage-gated calcium channels (VGCCs) are implicated in the entry of [Ca(2+)]E to the cytoplasm. We investigated the involvement of VGCC in the extracellular calcium induced intracellular calcium response (ECIICR). MC3T3-E1 cells were subjected to one dimensional calcium efflux from their basal aspect which results in an increase in [Ca(2+)]I. This increase was concomitant with membrane depolarization and it was significantly reduced in the presence of Bepridil, a non-selective VGCC inhibitor. To identify specific type(s) of VGCC in ECIICR, the cells were treated with selective inhibitors for different types of VGCC. Significant changes in the peak intensity and the number of [Ca(2+)]I oscillations were observed when L-type and T-type specific VGCC inhibitors (Verapamil and NNC55-0396, respectively) were used. So as to confirm the involvement of L- and T-type VGCC in the context of microdamage, cells were seeded on devitalized notched bone specimen, which were loaded to induce microdamage in the presence and absence of Verapamil and NNC55-0396. The results showed significant decrease in [Ca(2+)]I activity of cells in the microdamaged regions of bone when L- and T-type blockers were applied. This study demonstrated that extracellular calcium increase in association with damage depolarizes the cell membrane and the calcium ions enter the cell cytoplasm by L- and T-type VGCCs.

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

    Energy Technology Data Exchange (ETDEWEB)

    Bevilacqua, M.; Vago, T.; Norbiato, G. (Servizio di Endocrinologia, Milano, (Italy))


    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 ({sup 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{sup 2}{sup +} and of calcium blockers (D-888, a verapamillike drug, and diltiazem). Incubation in a Ca{sup 2}{sup +}-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{sup 2}{sup +} to the medium. This result was consistent with a competition between Ca{sup 2}{sup +} 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{sup 2}{sup +}.

  17. David J. Triggle: Medicinal chemistry, to pharmacology, calcium channels, and beyond. (United States)

    Walker, Michael J A


    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.

  18. CRMP-2 peptide mediated decrease of high and low voltage-activated calcium channels, attenuation of nociceptor excitability, and anti-nociception in a model of AIDS therapy-induced painful peripheral neuropathy

    Directory of Open Access Journals (Sweden)

    Piekarz Andrew D


    Full Text Available Abstract Background The ubiquity of protein-protein interactions in biological signaling offers ample opportunities for therapeutic intervention. We previously identified a peptide, designated CBD3, that suppressed inflammatory and neuropathic behavioral hypersensitivity in rodents by inhibiting the ability of collapsin response mediator protein 2 (CRMP-2 to bind to N-type voltage-activated calcium channels (CaV2.2 [Brittain et al. Nature Medicine 17:822–829 (2011]. Results and discussion Here, we utilized SPOTScan analysis to identify an optimized variation of the CBD3 peptide (CBD3A6K that bound with greater affinity to Ca2+ channels. Molecular dynamics simulations demonstrated that the CBD3A6K peptide was more stable and less prone to the unfolding observed with the parent CBD3 peptide. This mutant peptide, conjugated to the cell penetrating motif of the HIV transduction domain protein TAT, exhibited greater anti-nociception in a rodent model of AIDS therapy-induced peripheral neuropathy when compared to the parent TAT-CBD3 peptide. Remarkably, intraperitoneal administration of TAT-CBD3A6K produced none of the minor side effects (i.e. tail kinking, body contortion observed with the parent peptide. Interestingly, excitability of dissociated small diameter sensory neurons isolated from rats was also reduced by TAT-CBD3A6K peptide suggesting that suppression of excitability may be due to inhibition of T- and R-type Ca2+ channels. TAT-CBD3A6K had no effect on depolarization-evoked calcitonin gene related peptide (CGRP release compared to vehicle control. Conclusions Collectively, these results establish TAT-CBD3A6K as a peptide therapeutic with greater efficacy in an AIDS therapy-induced model of peripheral neuropathy than its parent peptide, TAT-CBD3. Structural modifications of the CBD3 scaffold peptide may result in peptides with selectivity against a particular subset of voltage-gated calcium channels resulting in a multipharmacology of

  19. [Discovering L-type calcium channels inhibitors of antihypertensive drugs based on drug repositioning]. (United States)

    Liang, Ying-xi; He, Yu-su; Jiang, Lu-di; Yue, Qiao-xin; Cui, Shuai; Bin, Li; Ye, Xiao-tong; Zhang, Xiao-hua; Zhang, Yang-ling


    This study was amid to construct the pharmacophore model of L-type calcium channel antagonist in the application of screening Drugbank and TCMD. This paper repositions the approved drugs resulting from virtual screening and discusses the relocation-based drug discovery methods, screening antihypertensive drugs with L-type calcium channel function from TCMD. Qualitative hypotheses wre generated by HipHop separately on the basis of 12 compounds with antagonistic action on L-type calcium channel expressed in rabbit cardiac muscle. Datebase searching method was used to evaluate the generated hypotheses. The optimum hypothesis was used to search Drugbank and TCMD. This paper repositions the approved drugs and evaluates the antihypertensive effect of the chemical constituent of traditional Chinese medicine resulting from virtual screening by the matching score and literature. The results showed that optimum qualitative hypothesis is with six features, which were two hydrogen-bond acceptors, four hydrophobic groups, and the CAI value of 2.78. Screening Drugbank achieves 93 approved drugs. Screening TCMD achieves 285 chemical constituents of traditional Chinese medicine. It was concluded that the hypothesis is reliable and can be used to screen datebase. The approved drugs resulting from virtual screening, such as pravastatin, are potentially L-type calcium channels inhibitors. The chemical constituents of traditional Chinese medicine, such as Arctigenin III and Arctigenin are potentially antihypertensive drugs. It indicates that Drug Repositioning based on hypothesis is possible.

  20. L-type Voltage-Gated Calcium Channels in Conditioned Fear: A Genetic and Pharmacological Analysis (United States)

    McKinney, Brandon C.; Sze, Wilson; White, Jessica A.; Murphy, Geoffrey G.


    Using pharmacological approaches, others have suggested that L-type voltage-gated calcium channels (L-VGCCs) mediate both consolidation and extinction of conditioned fear. In the absence of L-VGCC isoform-specific antagonists, we have begun to investigate the subtype-specific role of LVGCCs in consolidation and extinction of conditioned fear…

  1. Calcium channel blockers and cancer : A risk analysis using the UK Clinical Practice Research Datalink (CPRD)

    NARCIS (Netherlands)

    Grimaldi-Bensouda, Lamiae; Klungel, Olaf; Kurz, Xavier; De Groot, Mark C H; Afonso, Ana S Maciel; De Bruin, Marie L.; Reynolds, Robert; Rossignol, Michel


    OBJECTIVE: The evidence of an association between calcium channel blockers (CCBs) and cancer is conflicting. The objective of the present study was to evaluate the risk of cancer (all, breast, prostate and colon cancers) in association with exposure to CCB. METHODS: This is a population-based cohort

  2. [Influence of rifampicin on antihypertensive effects of dihydropiridine calcium-channel blockers in four elderly patients]. (United States)

    Yoshimoto, H; Takahashi, M; Saima, S


    Rifamicin, an antituberculosis agent, is one of the most potent inducers of hepatic drug-oxidation enzymes. Rifampicin can reduce the efficacy of several therapeutically important drugs (including verapamil and diltiazem) by accelerating systemic elimination or by increasing hepatic first-pass metabolism. Because dihydropyridine calcium-channel blockers are mainly metabolized by the liver, rifampicin may also increase the extraction of these drugs and thereby reduce their antihypertensive effects. Here we report four possible cases of interaction between rifampicin and dihydropiridine calcium-channel blockers. Rifampicin was given to treat tuberculosis in four elderly hypertensive patients whose blood pressure was well-controlled by one or more dihydropiridine calcium-channel blockers (nisoldipine, nifedipine, or barnidipine and manidipine), shortly after the start of antituberculosis therapy, their blood pressures rose. Either much greater doses of dihydropyridines or additional antihypertensive agents had to be given to keep blood pressure under control. After withdrawal of rifampicin, blood pressure fell in all patients and the doses of the antihypertensive agents had to be reduced. These findings indicate that rifampicin may lessen the antihypertensive effects of dihydropiridine calcium-channel blockers.

  3. Calcium channel antagonists increase morphine-induced analgesia and antagonize morphine tolerance. (United States)

    Contreras, E; Tamayo, L; Amigo, M


    The influence of calcium channel blockers on morphine-induced analgesia and on tolerance to the chronic administration of the opiate was investigated in mice. The effects of a test dose of morphine were significantly increased by the administration of diltiazem, flunarizine, nicardipine and verapamil. In contrast, nifedipine induced an antagonistic effect. The calcium channel antagonists did not change the reaction time to thermal stimulation in mice (hot plate test). The administration of nifedipine, flunarizine and verapamil reduced the intensity of the tolerance induced by a single dose of morphine administered in a slow release preparation. Diltiazem induced a non-significant decrease of the process. The present results are in accordance with the known interaction of acute and chronic morphine administration with the intracellular calcium concentration in neurones of the central nervous system.

  4. Activity-dependent regulation of T-type calcium channels by submembrane calcium ions. (United States)

    Cazade, Magali; Bidaud, Isabelle; Lory, Philippe; Chemin, Jean


    Voltage-gated Ca(2+) channels are involved in numerous physiological functions and various mechanisms finely tune their activity, including the Ca(2+) ion itself. This is well exemplified by the Ca(2+)-dependent inactivation of L-type Ca(2+) channels, whose alteration contributes to the dramatic disease Timothy Syndrome. For T-type Ca(2+) channels, a long-held view is that they are not regulated by intracellular Ca(2+). Here we challenge this notion by using dedicated electrophysiological protocols on both native and expressed T-type Ca(2+) channels. We demonstrate that a rise in submembrane Ca(2+) induces a large decrease in T-type current amplitude due to a hyperpolarizing shift in the steady-state inactivation. Activation of most representative Ca(2+)-permeable ionotropic receptors similarly regulate T-type current properties. Altogether, our data clearly establish that Ca(2+) entry exerts a feedback control on T-type channel activity, by modulating the channel availability, a mechanism that critically links cellular properties of T-type Ca(2+) channels to their physiological roles.

  5. Calcium Channel Expression and Applicability as Targeted Therapies in Melanoma

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    A. Macià


    Full Text Available The remodeling of Ca2+ signaling is a common finding in cancer pathophysiology serving the purpose of facilitating proliferation, migration, or survival of cancer cells subjected to stressful conditions. One particular facet of these adaptive changes is the alteration of Ca2+ fluxes through the plasma membrane, as described in several studies. In this review, we summarize the current knowledge about the expression of different Ca2+ channels in the plasma membrane of melanoma cells and its impact on oncogenic Ca2+ signaling. In the last few years, new molecular components of Ca2+ influx pathways have been identified in melanoma cells. In addition, new links between Ca2+ homeostasis and specific cell processes important in melanoma tumor progression have been unveiled. Thus, not only do Ca2+ channels appear to have a potential as prognostic markers, but their pharmacological blockade or gene silencing is hinted as interesting therapeutic approaches.


    Institute of Scientific and Technical Information of China (English)

    ZHANG Ming-mei; XIE Ji-min; CHEN Min; ZHANG Yan


    Objective: To study the effect of electroacupuncture (EA) and Verapamil and Nifedipine (calcium channel inhibitors) on free calcium concentrations of cells and intrasynaptosomes in hypothalamus (HT), periaqueductual grey matter (PAG) and hippocampus (HIP) of mice. Methods: The female ICR mice were randomly divided into control, EA, CaCl2 and CaCl2+EA groups (n=8 in each group). Pain threshold was detected by using radiation-heat irradiation-induced tail flick method. EA (8 Hz, a suitable stimulating strength, dense-sparse waves and duration of 30 min) was applied to"Shuigou" (水沟 GV 26) and "Chengjiang" (承浆CV 24). CaCl2 (10 μL, 0.2 μmol/L) was injected into the lateral cerebral ventricle of mice after EA. The concentrations of cytosolic free calcium ([Ca2+]i) in HIP, PAG, HT cell suspension specimen and hippocampal intrasynaptosome suspension of mice were determined by the fluorescent calcium indicator Fura-2-AM and a spectrofluorometer. Results: During EA analgesia, the intracellular free [Ca2+]i in HT and PAG specimens and intrsynaptosomal [Ca2+]i of the 3 cerebral regions decreased considerably (P<0.05~0.01), but that in hippocampal cell suspension increased significantly (P<0.01) in comparison with control group. The concentrations of hippocampal intrasynaptosomal free [Ca2+]i decreased significantly after adding Verapamil and Nifedipine to the extracted hippocampal intrasynaptosomal specimen. Microinjection of CaCl2 into lateral ventricle had no apparent influence on degree of analgesia (DA)% and intracellular and intrasynapsotomal [Ca2+]i, but significantly lower DA% and reduce changes of cytosolic and intrasynaptosomal [Ca2+]i induced by EA stimulation. Conclusion: Calcium ion in the neurons and intrasynaptosome of HT, PAG and HIP is involved in electroacupuncture analgesia.

  7. Barnidipine: a new calcium channel blocker for hypertension treatment. (United States)

    Liau, Chiau-Suong


    Although it is commonly agreed that all antihypertensive medications have similar efficacy, there are important differences related to safety, tolerability, patient adherence, cost effectiveness and effects on the prevention or retardation of associated disease progression. It is desirable for antihypertensives to have a long duration of action so that once-daily dosing is possible. In addition, antihypertensive medication must be able to be administered concomitantly with other drugs likely to be taken by the patients. This is particularly critical in the elderly population. Barnidipine, a novel, long-acting calcium antagonist, has met these challenges of modern pharmacotherapy. Its once-daily dosing, good tolerability and durable antihypertensive effect contribute to excellent patient adherence and make this drug a valuable addition to the antihypertensive formulary.

  8. Enhanced excitatory transmission at cortical synapses as the basis for facilitated spreading depression in Ca(v)2.1 knockin migraine mice. (United States)

    Tottene, Angelita; Conti, Rossella; Fabbro, Alessandra; Vecchia, Dania; Shapovalova, Maryna; Santello, Mirko; van den Maagdenberg, Arn M J M; Ferrari, Michel D; Pietrobon, Daniela


    Migraine is a common disabling brain disorder. A subtype of migraine with aura (familial hemiplegic migraine type 1: FHM1) is caused by mutations in Ca(V)2.1 (P/Q-type) Ca(2+) channels. Knockin mice carrying a FHM1 mutation show increased neuronal P/Q-type current and facilitation of induction and propagation of cortical spreading depression (CSD), the phenomenon that underlies migraine aura and may activate migraine headache mechanisms. We studied cortical neurotransmission in neuronal microcultures and brain slices of FHM1 mice. We show gain of function of excitatory neurotransmission due to increased action-potential-evoked Ca(2+) influx and increased probability of glutamate release at pyramidal cell synapses but unaltered inhibitory neurotransmission at fast-spiking interneuron synapses. Using an in vitro model of CSD, we show a causative link between enhanced glutamate release and CSD facilitation. The synapse-specific effect of FHM1 mutations points to disruption of excitation-inhibition balance and neuronal hyperactivity as the basis for episodic vulnerability to CSD ignition in migraine.

  9. Boosting of synaptic potentials and spine Ca transients by the peptide toxin SNX-482 requires alpha-1E-encoded voltage-gated Ca channels.

    Directory of Open Access Journals (Sweden)

    Andrew J Giessel

    Full Text Available The majority of glutamatergic synapses formed onto principal neurons of the mammalian central nervous system are associated with dendritic spines. Spines are tiny protuberances that house the proteins that mediate the response of the postsynaptic cell to the presynaptic release of glutamate. Postsynaptic signals are regulated by an ion channel signaling cascade that is active in individual dendritic spines and involves voltage-gated calcium (Ca channels, small conductance (SK-type Ca-activated potassium channels, and NMDA-type glutamate receptors. Pharmacological studies using the toxin SNX-482 indicated that the voltage-gated Ca channels that signal within spines to open SK channels belong to the class Ca(V2.3, which is encoded by the Alpha-1E pore-forming subunit. In order to specifically test this conclusion, we examined the effects of SNX-482 on synaptic signals in acute hippocampal slices from knock-out mice lacking the Alpha-1E gene. We find that in these mice, application of SNX-482 has no effect on glutamate-uncaging evoked synaptic potentials and Ca influx, indicating that that SNX-482 indeed acts via the Alpha-1E-encoded Ca(V2.3 channel.

  10. P-type calcium channels are blocked by the alkaloid daurisoline. (United States)

    Lu, Y M; Fröstl, W; Dreessen, J; Knöpfel, T


    IN looking for a structurally defined non-peptide P-channel blocker we have tested the alkaloid daurisoline which has been isolated from traditional Chinese medicinal herb (Menispermum dauricum) used for the treatment of epilepsy, hypertension and asthma. We have found that daurisoline is an inhibitor of omega-Aga-IVA sensitive barium currents in cerebellar Purkinje cells and of excitatory postsynaptic potentials evoked in Purkinje cells by stimulating parallel fibres in acutely prepared cerebellar slices. Daurisoline did not significantly affect omega-Aga-IVA-insensitive barium currents recorded from granule cells freshly isolated from rat cerebellum. Daurisoline passes the blood-brain barrier and will, therefore, facilitate the functional characterization of brain calcium channels as well as the exploration of P-type calcium channels as possible drug targets.

  11. New 1,4-dihydropyridines endowed with NO-donor and calcium channel agonist properties. (United States)

    Visentin, Sonja; Rolando, Barbara; Di Stilo, Antonella; Fruttero, Roberta; Novara, Monica; Carbone, Emilio; Roussel, Christian; Vanthuyne, Nicolas; Gasco, Alberto


    A new series of calcium channel agonists structurally related to Bay K8644, containing NO donor furoxans and the related furazans unable to release NO, is described. The racemic mixtures were studied for their action on L-type Ca(2+) channels expressed in cultured rat insulinoma RINm5F cells. All the products proved to be potent calcium channel agonists. All the racemic mixtures, with the only exception of the carbamoyl derivatives 9, 12 endowed with scanty solubility, were separated by chiral chromatography into the corresponding enantiomers; the (+) enantiomers were found to be potent agonists while the (-) ones were feeble antagonists. The racemic mixtures were also assessed for their positive inotropic activity on electrically stimulated rat papillary muscle and for their ability to increase Ca(2+) entry into the vascular smooth muscle of rat aorta strips. The cyanofuroxan 8 proved to be an interesting product with dual Ca(2+)-dependent positive inotropic and NO-dependent vasodilating activity.

  12. Inhibition of collagen synthesis by select calcium and sodium channel blockers can be mitigated by ascorbic acid and ascorbyl palmitate


    Ivanov, Vadim; Ivanova, Svetlana; KALINOVSKY, TATIANA; NIEDZWIECKI, ALEKSANDRA; Rath, Matthias


    Calcium, sodium and potassium channel blockers are widely prescribed medications for a variety of health problems, most frequently for cardiac arrhythmias, hypertension, angina pectoris and other disorders. However, chronic application of channel blockers is associated with numerous side effects, including worsening cardiac pathology. For example, nifedipine, a calcium-channel blocker was found to be associated with increased mortality and increased risk for myocardial infarction. In addition...

  13. Putative calcium-binding domains of the Caenorhabditis elegans BK channel are dispensable for intoxication and ethanol activation. (United States)

    Davis, S J; Scott, L L; Ordemann, G; Philpo, A; Cohn, J; Pierce-Shimomura, J T


    Alcohol modulates the highly conserved, voltage- and calcium-activated potassium (BK) channel, which contributes to alcohol-mediated behaviors in species from worms to humans. Previous studies have shown that the calcium-sensitive domains, RCK1 and the Ca(2+) bowl, are required for ethanol activation of the mammalian BK channel in vitro. In the nematode Caenorhabditis elegans, ethanol activates the BK channel in vivo, and deletion of the worm BK channel, SLO-1, confers strong resistance to intoxication. To determine if the conserved RCK1 and calcium bowl domains were also critical for intoxication and basal BK channel-dependent behaviors in C. elegans, we generated transgenic worms that express mutated SLO-1 channels predicted to have the RCK1, Ca(2+) bowl or both domains rendered insensitive to calcium. As expected, mutating these domains inhibited basal function of SLO-1 in vivo as neck and body curvature of these mutants mimicked that of the BK null mutant. Unexpectedly, however, mutating these domains singly or together in SLO-1 had no effect on intoxication in C. elegans. Consistent with these behavioral results, we found that ethanol activated the SLO-1 channel in vitro with or without these domains. By contrast, in agreement with previous in vitro findings, C. elegans harboring a human BK channel with mutated calcium-sensing domains displayed resistance to intoxication. Thus, for the worm SLO-1 channel, the putative calcium-sensitive domains are critical for basal in vivo function but unnecessary for in vivo ethanol action.

  14. Single-channel Analysis and Calcium Imaging in the Podocytes of the Freshly Isolated Glomeruli. (United States)

    Ilatovskaya, Daria V; Palygin, Oleg; Levchenko, Vladislav; Staruschenko, Alexander


    Podocytes (renal glomerular epithelial cells) are known to regulate glomerular permeability and maintain glomerular structure; a key role for these cells in the pathogenesis of various renal diseases has been established since podocyte injury leads to proteinuria and foot process effacement. It was previously reported that various endogenous agents may cause a dramatic overload in intracellular Ca(2+) concentration in podocytes, presumably leading to albuminuria, and this likely occurs via calcium-conducting ion channels. Therefore, it appeared important to study calcium handling in the podocytes both under normal conditions and in various pathological states. However, available experimental approaches have remained somewhat limited to cultured and transfected cells. Although they represent a good basic model for such studies, they are essentially extracted from the native environment of the glomerulus. Here we describe the methodology of studying podocytes as a part of the freshly isolated whole glomerulus. This preparation retains the functional potential of the podocytes, which are still attached to the capillaries; therefore, podocytes remain in the environment that conserves the major parts of the glomeruli filtration apparatus. The present manuscript elaborates on two experimental approaches that allow 1) real-time detection of calcium concentration changes with the help of ratiometric confocal fluorescence microscopy, and 2) the recording of the single ion channels activity in the podocytes of the freshly isolated glomeruli. These methodologies utilize the advantages of the native environment of the glomerulus that enable researchers to resolve acute changes in the intracellular calcium handling in response to applications of various agents, measure basal concentration of calcium within the cells (for instance, to evaluate disease progression), and assess and manipulate calcium conductance at the level of single ion channels.

  15. Enhanced currents through L-type calcium channels in cardiomyocytes disturb the electrophysiology of the dystrophic heart. (United States)

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


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

  16. Novel tacrine derivatives that block neuronal calcium channels. (United States)

    de los Ríos, Cristóbal; Marco, José L; Carreiras, María D C; Chinchón, P M; García, Antonio G; Villarroya, Mercedes


    A new series of tacrine (9-amino-1,2,3,4-tetrahydroacridine) derivatives were synthesized and their effects on 45Ca(2+) entry into bovine adrenal chromaffin cells stimulated with dimethylphenylpiperazinium (DMPP) or K(+), studied. At 3 microM, compound 1 did not affect (45)Ca(2+) uptake evoked by DMPP. Compounds 14, 15 and 17 inhibited the effects of DMPP by 30%. Compounds 3, 9 and tacrine blocked the DMPP signal by about 50%. Compounds 5 and 12 were the most potent blockers of DMPP-stimulated 45Ca(2+) entry (90%); the rest of the compounds inhibited the effects of DMPP by 70-80%. Compounds 1, 3, 4, 8, 10, 11, 13, 16, 17 and tacrine inhibited 45Ca(2+) uptake induced by K(+) about 20%. Compounds 6, 14 and 15 inhibited the K(+) effects by 10% or less. Compounds 7, 9, 12 and 18 blocked the K(+) signal by 30% and, finally, compounds 2 and 5 inhibited the K(+)-induced 45Ca(2+) entry by 50%. None of the new compounds was as effective as diltiazem (IC(50)=0.03 microM) in causing relaxation of the rat aorta precontracted with 35 mM K(+); the most potent was compound 7 (IC(50)=0.3 microM). Compounds 5, 6, 8, 9, 10 and 13 had IC(50)s around 10 microM and compounds 3, 4, 11 and 12 around 20 microM. Blockade of Ca(2+) entry through neuronal voltage-dependent Ca(2+) channels, without concomitant blockade of vascular Ca(2+) channels, suggests that some of these compounds might exhibit neuroprotectant effects but not undesirable hemodynamic effects.

  17. Iron overload and apoptosis of HL-1 cardiomyocytes: effects of calcium channel blockade.

    Directory of Open Access Journals (Sweden)

    Mei-pian Chen

    Full Text Available Iron overload cardiomyopathy that prevails in some forms of hemosiderosis is caused by excessive deposition of iron into the heart tissue and ensuing damage caused by a raise in labile cell iron. The underlying mechanisms of iron uptake into cardiomyocytes in iron overload condition are still under investigation. Both L-type calcium channels (LTCC and T-type calcium channels (TTCC have been proposed to be the main portals of non-transferrinic iron into heart cells, but controversies remain. Here, we investigated the roles of LTCC and TTCC as mediators of cardiac iron overload and cellular damage by using specific Calcium channel blockers as potential suppressors of labile Fe(II and Fe(III ingress in cultured cardiomyocytes and ensuing apoptosis.Fe(II and Fe(III uptake was assessed by exposing HL-1 cardiomyocytes to iron sources and quantitative real-time fluorescence imaging of cytosolic labile iron with the fluorescent iron sensor calcein while iron-induced apoptosis was quantitatively measured by flow cytometry analysis with Annexin V. The role of calcium channels as routes of iron uptake was assessed by cell pretreatment with specific blockers of LTCC and TTCC.Iron entered HL-1 cardiomyocytes in a time- and dose-dependent manner and induced cardiac apoptosis via mitochondria-mediated caspase-3 dependent pathways. Blockade of LTCC but not of TTCC demonstrably inhibited the uptake of ferric but not of ferrous iron. However, neither channel blocker conferred cardiomyocytes with protection from iron-induced apoptosis.Our study implicates LTCC as major mediators of Fe(III uptake into cardiomyocytes exposed to ferric salts but not necessarily as contributors to ensuing apoptosis. Thus, to the extent that apoptosis can be considered a biological indicator of damage, the etiopathology of cardiosiderotic damage that accompanies some forms of hemosiderosis would seem to be unrelated to LTCC or TTCC, but rather to other routes of iron ingress present in

  18. Role for voltage gated calcium channels in calcitonin gene-related peptide release in the rat trigeminovascular system

    DEFF Research Database (Denmark)

    Amrutkar, D V; Ploug, K B; Olesen, J


    Clinical and genetic studies have suggested a role for voltage gated calcium channels (VGCCs) in the pathogenesis of migraine. Release of calcitonin gene-related peptide (CGRP) from trigeminal neurons has also been implicated in migraine. The VGCCs are located presynaptically on neurons and are i...... releases CGRP, and the release is regulated by Ca2+ ions and voltage-gated calcium channels.......Clinical and genetic studies have suggested a role for voltage gated calcium channels (VGCCs) in the pathogenesis of migraine. Release of calcitonin gene-related peptide (CGRP) from trigeminal neurons has also been implicated in migraine. The VGCCs are located presynaptically on neurons...

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

    Energy Technology Data Exchange (ETDEWEB)

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


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

  20. A role for L-type calcium channels in the maturation of parvalbumin-containing hippocampal interneurons. (United States)

    Jiang, M; Swann, J W


    While inhibitory interneurons are well recognized to play critical roles in the brain, relatively little is know about the molecular events that regulate their growth and differentiation. Calcium ions are thought to be important in neuronal development and L-type voltage gated Ca(+2) channels have been implicated in activity-dependent mechanisms of early-life. However, few studies have examined the role of these channels in the maturation of interneurons. The studies reported here were conducted in hippocampal slice cultures and indicate that the L-type Ca(+2) channel agonists and antagonists accelerate and suppress respectively the growth of parvalbumin-containing interneurons. The effects of channel blockade were reversible suggesting they are not the result of interneuronal cell death. Results from immunoblotting showed that these drugs have similar effects on the expression of the GABA synthetic enzymes, glutamic acid decarboxylase65, glutamic acid decarboxylase67 and the vesicular GABA transporter. This suggests that L-type Ca(+2) channels regulate not only parvalbumin expression but also interneuron development. These effects are likely mediated by actions on the interneurons themselves since the alpha subunits of L-type channels, voltage-gated calcium channel subunit 1.2 and voltage-gated calcium channel subunit 1.3 were found to be highly expressed in neonatal mouse hippocampus and co-localized with parvalbumin in interneurons. Results also showed that while these interneurons can contain either subunit, voltage-gated calcium channel subunit 1.3 was more widely expressed. Taken together results suggest that an important subset of developing interneurons expresses L-type Ca(+2) channels alpha subunits, voltage-gated calcium channel subunit 1.2 and especially voltage-gated calcium channel subunit 1.3 and that these channels likely regulate the development of these interneurons in an activity-dependent manner.

  1. Aluminium and hydrogen ions inhibit a mechanosensory calcium-selective cation channel (United States)

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


    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.

  2. Role of Calcium Channels in the Protective Effect of Hydrogen Sulfide in Rat Cardiomyoblasts

    Directory of Open Access Journals (Sweden)

    Daniele Avanzato


    Full Text Available Background: Hydrogen sulfide contributes to the reduction of oxidative stress-related injury in cardiomyocytes but the underlying mechanism is still unclear. Aims: Here we investigated the role of voltage-operated calcium channels (VOCCs as mediators of the beneficial effect of H2S against oxidative stress in cultured rat cardiomyoblasts (H9c2. Methods: Intracellular calcium signals were measured by fluorimetric live cell imaging and cell viability by colorimetric assay. Results: Treatment with H2S donor (NaHS 10 µM or Nifedipine (10 µM decreased resting intracellular calcium concentration [Ca]i, suggesting that L-type VOCCs are negatively modulated by H2S. In the presence of Nifedipine H2S was still able to lower [Ca]i, while co-incubation with Nifedipine and Ni2+ 100 µM completely prevented H2S-dependent [Ca]i decrease, suggesting that both L-type and T-type VOCCs are inhibited by H2S. In addition, in the same experimental conditions, H2S triggered a slow increase of [Ca]i whose molecular nature remains to be clarified. Pretreatment of H9c2 with NaHS (10 µM significantly prevented cell death induced by H2O2. This effect was mimicked by pretreatment with L-Type calcium channel inhibitor Nifedipine (10 µM. Conclusions: The data provide the first evidence that H2S protects rat cardiomyoblasts against oxidative challenge through the inhibition of L-type calcium channels.

  3. Experts Consensus Recommendations for the Management of Calcium Channel Blocker Poisoning in Adults (United States)

    Anseeuw, Kurt; Cantrell, Frank Lee; Gilchrist, Ian C.; Hantson, Philippe; Bailey, Benoit; Lavergne, Valéry; Gosselin, Sophie; Kerns, William; Laliberté, Martin; Lavonas, Eric J.; Juurlink, David N.; Muscedere, John; Yang, Chen-Chang; Sinuff, Tasnim; Rieder, Michael; Mégarbane, Bruno


    Objective: To provide a management approach for adults with calcium channel blocker poisoning. Data Sources, Study Selection, and Data Extraction: Following the Appraisal of Guidelines for Research & Evaluation II instrument, initial voting statements were constructed based on summaries outlining the evidence, risks, and benefits. Data Synthesis: We recommend 1) for asymptomatic patients, observation and consideration of decontamination following a potentially toxic calcium channel blocker ingestion (1D); 2) as first-line therapies (prioritized based on desired effect), IV calcium (1D), high-dose insulin therapy (1D–2D), and norepinephrine and/or epinephrine (1D). We also suggest dobutamine or epinephrine in the presence of cardiogenic shock (2D) and atropine in the presence of symptomatic bradycardia or conduction disturbance (2D); 3) in patients refractory to the first-line treatments, we suggest incremental doses of high-dose insulin therapy if myocardial dysfunction is present (2D), IV lipid-emulsion therapy (2D), and using a pacemaker in the presence of unstable bradycardia or high-grade arteriovenous block without significant alteration in cardiac inotropism (2D); 4) in patients with refractory shock or who are periarrest, we recommend incremental doses of high-dose insulin (1D) and IV lipid-emulsion therapy (1D) if not already tried. We suggest venoarterial extracorporeal membrane oxygenation, if available, when refractory shock has a significant cardiogenic component (2D), and using pacemaker in the presence of unstable bradycardia or high-grade arteriovenous block in the absence of myocardial dysfunction (2D) if not already tried; 5) in patients with cardiac arrest, we recommend IV calcium in addition to the standard advanced cardiac life-support (1D), lipid-emulsion therapy (1D), and we suggest venoarterial extracorporeal membrane oxygenation if available (2D). Conclusion: We offer recommendations for the stepwise management of calcium channel blocker

  4. 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.


    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.

  5. Synaptic Ribbons Require Ribeye for Electron Density, Proper Synaptic Localization, and Recruitment of Calcium Channels

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    Caixia Lv


    Full Text Available Synaptic ribbons are structures made largely of the protein Ribeye that hold synaptic vesicles near release sites in non-spiking cells in some sensory systems. Here, we introduce frameshift mutations in the two zebrafish genes encoding for Ribeye and thus remove Ribeye protein from neuromast hair cells. Despite Ribeye depletion, vesicles collect around ribbon-like structures that lack electron density, which we term “ghost ribbons.” Ghost ribbons are smaller in size but possess a similar number of smaller vesicles and are poorly localized to synapses and calcium channels. These hair cells exhibit enhanced exocytosis, as measured by capacitance, and recordings from afferent neurons post-synaptic to hair cells show no significant difference in spike rates. Our results suggest that Ribeye makes up most of the synaptic ribbon density in neuromast hair cells and is necessary for proper localization of calcium channels and synaptic ribbons.

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

    Directory of Open Access Journals (Sweden)

    Ji-Guang Wang


    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

  7. Channel properties of the splicing isoforms of the olfactory calcium-activated chloride channel Anoctamin 2. (United States)

    Ponissery Saidu, Samsudeen; Stephan, Aaron B; Talaga, Anna K; Zhao, Haiqing; Reisert, Johannes


    Anoctamin (ANO)2 (or TMEM16B) forms a cell membrane Ca(2+)-activated Cl(-) channel that is present in cilia of olfactory receptor neurons, vomeronasal microvilli, and photoreceptor synaptic terminals. Alternative splicing of Ano2 transcripts generates multiple variants with the olfactory variants skipping exon 14 and having alternative splicing of exon 4. In the present study, 5' rapid amplification of cDNA ends analysis was conducted to characterize the 5' end of olfactory Ano2 transcripts, which showed that the most abundant Ano2 transcripts in the olfactory epithelium contain a novel starting exon that encodes a translation initiation site, whereas transcripts of the publically available sequence variant, which has an alternative and longer 5' end, were present in lower abundance. With two alternative starting exons and alternative splicing of exon 4, four olfactory ANO2 isoforms are thus possible. Patch-clamp experiments in transfected HEK293T cells expressing these isoforms showed that N-terminal sequences affect Ca(2+) sensitivity and that the exon 4-encoded sequence is required to form functional channels. Coexpression of the two predominant isoforms, one with and one without the exon 4 sequence, as well as coexpression of the two rarer isoforms showed alterations in channel properties, indicating that different isoforms interact with each other. Furthermore, channel properties observed from the coexpression of the predominant isoforms better recapitulated the native channel properties, suggesting that the native channel may be composed of two or more splicing isoforms acting as subunits that together shape the channel properties.

  8. Direct recording and molecular identification of the calcium channel of primary cilia (United States)

    Decaen, Paul G.; Delling, Markus; Vien, Thuy N.; Clapham, David E.


    A primary cilium is a solitary, slender, non-motile protuberance of structured microtubules (9+0) enclosed by plasma membrane. Housing components of the cell division apparatus between cell divisions, primary cilia also serve as specialized compartments for calcium signalling and hedgehog signalling pathways. Specialized sensory cilia such as retinal photoreceptors and olfactory cilia use diverse ion channels. An ion current has been measured from primary cilia of kidney cells, but the responsible genes have not been identified. The polycystin proteins (PC and PKD), identified in linkage studies of polycystic kidney disease, are candidate channels divided into two structural classes: 11-transmembrane proteins (PKD1, PKD1L1 and PKD1L2) remarkable for a large extracellular amino terminus of putative cell adhesion domains and a G-protein-coupled receptor proteolytic site, and the 6-transmembrane channel proteins (PKD2, PKD2L1 and PKD2L2; TRPPs). Evidence indicates that the PKD1 proteins associate with the PKD2 proteins via coiled-coil domains. Here we use a transgenic mouse in which only cilia express a fluorophore and use it to record directly from primary cilia, and demonstrate that PKD1L1 and PKD2L1 form ion channels at high densities in several cell types. In conjunction with an accompanying manuscript, we show that the PKD1L1-PKD2L1 heteromeric channel establishes the cilia as a unique calcium compartment within cells that modulates established hedgehog pathways.

  9. Applying Theoretical Approach for Predicting the Selective Calcium Channel Blockers Pharmacological Parameter by Biopartitioning Micellar Chromatography

    Institute of Scientific and Technical Information of China (English)

    WANG Su-Min; YANG Geng-Liang; LI Zhi-Wei; LIU Hai-Yan; GUO Hui-Juan


    The usefulness of biopartitioning micellar chromatography (BMC) for predicting oral drug acute toxicity and apparent bioavailability was demonstrated. A logarithmic model (an LD50 model) and the second order polynomial models (apparent bioavailability model) have been obtained using the retention data of the selective calcium channel blockers to predict pharmacological properties of compounds. The use of BMC is simple, reproducible and can provide key information about the acute toxicity and transport properties of new compounds during the drug discovery process.

  10. Calcium-channel blockers for the prevention of stroke: from scientific evidences to the clinical practice

    Directory of Open Access Journals (Sweden)

    S. Taddei


    Full Text Available AIM OF THE REVIEW The present review aims to analyze the role of calcium-channel blockers, and particularly newer molecules, as first-line therapy for cerebrovascular disease. BACKGROUND Stroke is the leading cause of disability in the general population. Among traditional cardiovascular risk factors, hypertension has a key role in the genesis of both hemorrhagic and ischemic stroke and a direct correlation exists between blood pressure values and the risk of stroke. Moreover, blood pressure reduction has been demonstrated to be the most important route to reduce stroke incidence and recurrence. However, the mere reduction of blood pressure values does not normalize the cardiovascular risk of the hypertensive patient. It is therefore necessary to use drug classes that beyond their blood pressure-lowering effect have also an additional effect in terms of organ protection. Among these, calcium-channel blockers have a crucial profile. Firstly, they are effective in inducing left ventricular hypertrophy regression, with a strength at least equal to that of ACE-inhibitors. Secondly, they have an antithrombotic and an endothelium-protecting effect, mediated by their antioxidant activity. Finally, calcium-channel blockers are the most powerful drugs in preventing vascular remodeling. For these reasons this drug class has probably the strongest antiatherosclerotic effect, and it is the first-choice treatment mainly for cerebrovascular disease. Among different available calcium-channel blockers, the newer ones seem to possess pharmacokinetic characteristics allowing a more homogeneous 24 hours coverage as compared to older molecules, and preliminary data seem to suggest a greater beneficial effect also on left ventricular hypertrophy and lower incidence of side effects. CONCLUSIONS Although blood pressure reduction is the main tool to reduce cerebrovascular risk in hypertensive patients, some drug classes, such as calciumchannel blockers, seem to provide

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


    Mandeep Kaur; Rajesh Kumar Goel


    “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. A...

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



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

  13. Expression of the apoptotic calcium channel P2X7 in the glandular epithelium. (United States)

    Slater, Michael; Danieletto, Suzanne; Barden, Julian A


    In the current study, expression of the apoptotic calcium channel receptor P2X(7) and prostate-specific antigen (PSA) levels were studied in biopsy cores from 174 patients as well as 20 radical prostatectomy cases. In clinical biopsies, we have previously demonstrated that P2X(1 )and P2X(2) calcium channel receptors are absent from normal prostate epithelium that does not progress to prostate cancer within 5 years. In cases that did progress to prostate cancer however, P2X(1 )and P2X(2) labeling was observed in a stage-specific manner first in the nucleus, then the cytoplasm and finally on the apical epithelium, as prostate cancer developed. These markers were present up to 5 years before cancer was detectable by the usual morphological criteria (Gleason grading) as determined by H and E staining. In the current study, the apoptotic calcium channel receptor P2X(7) yielded similar results to that of P2X(1) and P2X(2). Using radical prostatectomy tissue sections as well as biopsies, these changes in calcium channel metabolism were noted throughout the prostate, indicating a field effect. This finding suggests that the presence of a prostate tumor could be detected without the need for direct sampling of tumor tissue, leading to detection of false negative cases missed by H or E stain. The reliability of PSA levels as a prognostic indicator has been questioned in recent years. In the current study, PSA levels were correlated with the P2X(7) labeling results. All patients who exhibited no P2X(7) labeling had a prostatic serum antigen (PSA) level of 2. This finding suggests that increasing PSA may be an accurate indicator of cancer development.

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

    Directory of Open Access Journals (Sweden)

    Gorash ZM


    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.

  15. Deltamethrin Inhibits the Human T-type Voltage-Sensitive Calcium Channel (Cav3.2

    Directory of Open Access Journals (Sweden)

    Steven B. Symington


    Full Text Available The goal of this study was to determine the effect of deltamethrin, a pyrethroid insecticide, on CaV3.2, a human T-type voltage-sensitive calcium channel expressed in Xenopus laevis (X.laevis oocytes. Cav3.2 cDNA was transcribed into cRNA; the cRNA was then injected into X.laevis oocytes and electrophysiologically characterized using the two-electrode voltage clamp technique with Ba2+ as a charge carrier. Deltamethrin (10-7 M reduced peak current in a nonreversible manner compared to the untreated control, but had no effect on the voltagedependent activation and inactivation kinetics. These findings confirm that human CaV3.2 is a target for deltamethrin and quite possibly other pyrethroid insecticides. These studies provide insight into the molecular mechanisms of the effect that pyrethroids have on voltage-sensitive calcium channels in general. This information will allow a more complete understanding of the molecular and cellular nature of pyrethroid-induced toxicity and expand our knowledge of the structure-activity relationships of pyrethroids with regard to their action on voltage-sensitive calcium channels.

  16. Quantitative Structure-Activity Relationship Studies of 4-Imidazolyl- 1,4-dihydropyridines as Calcium Channel Blockers

    Directory of Open Access Journals (Sweden)

    Farzin Hadizadeh


    Conclusion: The predictive ability of the model was found to be satisfactory and could be used for designing a similar group of 1,4- dihydropyridines , based on a pyridine structure core which can block calcium channels.

  17. Comparison of electrophysiological effects of calcium channel blockers on cardiac repolarization. (United States)

    Lee, Hyang-Ae; Hyun, Sung-Ae; Park, Sung-Gurl; Kim, Ki-Suk; Kim, Sung Joon


    Dihydropyridine (DHP) calcium channel blockers (CCBs) have been widely used to treat of several cardiovascular diseases. An excessive shortening of action potential duration (APD) due to the reduction of Ca(2+) channel current (I Ca) might increase the risk of arrhythmia. In this study we investigated the electrophysiological effects of nicardipine (NIC), isradipine (ISR), and amlodipine (AML) on the cardiac APD in rabbit Purkinje fibers, voltage-gated K(+) channel currents (I Kr, I Ks) and voltage-gated Na(+) channel current (I Na). The concentration-dependent inhibition of Ca(2+) channel currents (I Ca) was examined in rat cardiomyocytes; these CCBs have similar potency on I Ca channel blocking with IC50 (the half-maximum inhibiting concentration) values of 0.142, 0.229, and 0.227 nM on NIC, ISR, and AML, respectively. However, ISR shortened both APD50 and APD90 already at 1 µM whereas NIC and AML shortened APD50 but not APD90 up to 30 µM. According to ion channel studies, NIC and AML concentration-dependently inhibited I Kr and I Ks while ISR had only partial inhibitory effects (NIC and AML could compensate for the AP shortening effects due to the block of I Ca.

  18. 电压门控钙通道在三硝基苯磺酸诱导的炎症性内脏高敏中作用机制的研究%The study on the mechanism of voltage gated calcium channels on 2, 4, 6-trinitrobenzene sulfonic acid induced inflammatory visceral hypersensitivity

    Institute of Scientific and Technical Information of China (English)

    钱爱华; 宋丹丹; 李勇; 姚玮艳; 孙菁; 袁耀宗


    目的 研究炎症性内脏高敏大鼠背根神经节(DRG)基因表达的变化及电压门控钙离子通道(VGCC)在炎症性内脏高敏感性中的作用.方法 雄性清洁级SD大鼠180只,体重200~300 g,三硝基苯磺酸(TNBS)模型组(n=90)缓慢注入2.0%TNBS,剂量为100 mg/kg;正常对照组(n=90)仅灌注等量0.9%氯化钠溶液.造模后第4天,大鼠的cDNA表达谱基因芯片检测大鼠1.6S2节段DRG基因表达谱变化.通过实时定量逆转录聚合酶链反应(RT-PCR)和Western印迹检测进行验证.胞内钙检测及全细胞膜片钳记录分析胞内钙及电压门控钙电流变化.另取SD大鼠54只,均分为6组,分别予以TNBS造模及脊髓鞘内给予特异性钙通道阻滞剂,观察大鼠内脏敏感性的变化.腹部撤离反射(AWR)检测大鼠内脏敏感性.结果 TNBS模型组L6~s2节段DRG172个基因的表达出现显著变化,包括离子通道、膜受体及胞内第二信使等.其中,L-型钙通道(Cav 1.2)和R-型钙通道(Cav2.3)显著上调,RT-PCR及Western印迹检测结果均验证了芯片结果.胞内钙检测显示,TNBS模型组结肠特异感觉神经元的静息胞内钙含量与正常对照组相比差异无统计学意义(P>0.05);而胞内钙瞬变量较正常对照组显著增加(P<0.05).全细胞膜片钳记录显示TNBS模型组结肠初级感觉神经元L-型和R-型钙电流较正常对照组明显增加(P<0.05).脊髓鞘内给予尼莫地平和SNX-482后可显著降低炎症性内脏高敏大鼠的内脏敏感性(P<0.05).结论 Cav1.2和Cav2.3表达的上调在炎症性内脏高敏中起了非常重要的作用,很可能是炎症性内脏高敏治疗的潜在靶点.%Objective To screen the difference of gene expression in dorsal root ganglia (DRG)of inflammatory visceral hypersensitivity rats and to explore the role of voltage gated calcium channel (VGCC) in inflammatory visceral hypersensitivity. Methods Total 180 male Sprague-Dawley rats were in this study,the weight varied from

  19. The involvement of the Mid1/Cch1/Yvc1 calcium channels in Aspergillus fumigatus virulence.

    Directory of Open Access Journals (Sweden)

    Patrícia Alves de Castro

    Full Text Available Aspergillus fumigatus is a major opportunistic pathogen and allergen of mammals. Calcium homeostasis and signaling is essential for numerous biological processes and also influences A. fumigatus pathogenicity. The presented study characterized the function of the A. fumigatus homologues of three Saccharomyces cerevisiae calcium channels, voltage-gated Cch1, stretch-activated Mid1 and vacuolar Yvc1. The A. fumigatus calcium channels cchA, midA and yvcA were regulated at transcriptional level by increased calcium levels. The YvcA::GFP fusion protein localized to the vacuoles. Both ΔcchA and ΔmidA mutant strains showed reduced radial growth rate in nutrient-poor minimal media. Interestingly, this growth defect in the ΔcchA strain was rescued by the exogenous addition of CaCl2. The ΔcchA, ΔmidA, and ΔcchA ΔmidA strains were also sensitive to the oxidative stress inducer, paraquat. Restriction of external Ca(2+ through the addition of the Ca(2+-chelator EGTA impacted upon the growth of the ΔcchA and ΔmidA strains. All the A. fumigatus ΔcchA, ΔmidA, and ΔyvcA strains demonstrated attenuated virulence in a neutropenic murine model of invasive pulmonary aspergillosis. Infection with the parental strain resulted in a 100% mortality rate at 15 days post-infection, while the mortality rate of the ΔcchA, ΔmidA, and ΔyvcA strains after 15 days post-infection was only 25%. Collectively, this investigation strongly indicates that CchA, MidA, and YvcA play a role in A. fumigatus calcium homeostasis and virulence.

  20. Inhibitors of arachidonate-regulated calcium channel signaling suppress triggered activity induced by the late sodium current. (United States)

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


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

  1. Role of low voltage activated calcium channels in neuritogenesis and active migration of embryonic neural progenitor cells. (United States)

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


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

  2. Correlated ions in a calcium channel model: a Poisson-Fermi theory. (United States)

    Liu, Jinn-Liang; Eisenberg, Bob


    We derive a continuum model, called the Poisson-Fermi equation, of biological calcium channels (of cardiac muscle, for example) designed to deal with crowded systems in which ionic species and side chains nearly fill space. The model is evaluated in three dimensions. It includes steric and correlation effects and is derived from classical hard-sphere lattice models of configurational entropy of finite size ions and solvent molecules. The maximum allowable close packing (saturation) condition is satisfied by all ionic species with different sizes and valences in a channel system, as shown theoretically and numerically. Unphysical overcrowding ("divergence") predicted by the Gouy-Chapman diffuse model (produced by a Boltzmann distribution of point charges with large potentials) does not occur with the Fermi-like distribution. Using probability theory, we also provide an analytical description of the implicit dielectric model of ionic solutions that gives rise to a global and a local formula for the chemical potential. In this primitive model, ions are treated as hard spheres and solvent molecules are described as a dielectric medium. The Poisson-Fermi equation is a local formula dealing with different correlations at different places. The correlation effects are apparent in our numerical results. Our results show variations of dielectric permittivity from bath to channel pore described by a new dielectric function derived as an output from the Poisson-Fermi analysis. The results are consistent with existing theoretical and experimental results. The binding curve of Poisson-Fermi is shown to match Monte Carlo data and illustrates the anomalous mole fraction effect of calcium channels, an effective blockage of permeation of sodium ions by a tiny concentration (or number) of calcium ions.

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

    Institute of Scientific and Technical Information of China (English)

    林先明; 李真; 胡本容; 夏国瑾; 姚伟星; 向继洲


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

  4. 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;


    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...... 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...

  5. Effects of glycoprotein Ⅱb/Ⅲa antagonists and chloride channel blockers on platelet cytoplasmic free calcium

    Institute of Scientific and Technical Information of China (English)

    YIN Song-mei; XIE Shuang-feng; NIE Da-nian; LI Yi-qing; LI Hai-ming; MA Li-ping; WANG Xiu-ju; WU Yu-dan; FENG Jian-hong


    @@ Platelet activation plays an important role in thrombosis. Platelet glycoprotein Ⅱb/Ⅲa (GP Ⅱb/Ⅲa) is the receptor of fibrinogen. Platelet cross-linking with fibrinogen through GPⅡb/Ⅲa is the process of thrombosis. Ca2+ is an important intracellular second messenger in platelet activation. It has been reported that GPⅡb/Ⅲa receptors were involved in the calcium influx of activated platelet, and GPⅡb/Ⅲa receptor had characteristics of calcium channel or an adjacent calcium channel.

  6. Formaldehyde increases intracellular calcium concentration in primary cultured hippocampal neurons partly through NMDA receptors and T-type calcium channels

    Institute of Scientific and Technical Information of China (English)

    Ye-Nan Chi; Xu Zhang; Jie Cai; Feng-Yu Liu; Guo-Gang Xing; You Wan


    Objective Formaldehyde at high concentrations is a contributor to air pollution.It is also an endogenous metabolic product in cells,and when beyond physiological concentrations,has pathological effects on neurons.Formaldehyde induces mis-folding and aggregation of neuronal tau protein,hippocampal neuronal apoptosis,cognitive impairment and loss of memory functions,as well as excitation of peripheral nociceptive neurons in cancer pain models.Intracellular calcium ([Ca2+]i) is an important intracellular messenger,and plays a key role in many pathological processes.The present study aimed to investigate the effect of formaldehyde on [Ca2+]i and the possible involvement of N-methyl-D-aspartate receptors (NMDARs) and T-type Ca2+ channels on the cell membrane.Methods Using primary cultured hippocampal neurons as a model,changes of [Ca2+]i in the presence of formaldehyde at a low concentration were detected by confocal laser scanning microscopy.Results Formaldehyde at 1 mmol/L approximately doubled [Ca2+]i.(2R)-amino-5-phosphonopentanoate (AP5,25 μtmol/L,an NMDAR antagonist) and mibefradil (MIB,1 μtmol/L,a T-type Ca2+ channel blocker),given 5 min after formaldehyde perfusion,each partly inhibited the formaldehyde-induced increase of [Ca2+]i,and this inhibitory effect was reinforced by combined application of AP5 and MIB.When applied 3 min before formaldehyde perfusion,AP5 (even at 50 μmol/L) did not inhibit the formaldehyde-induced increase of [Ca2+]i,but MIB (1 μmol/L) significantly inhibited this increase by 70%.Conclusion These results suggest that formaldehyde at a low concentration increases [Ca2+]i in cultured hippocampal neurons; NMDARs and T-type Ca2+ channels may be involved in this process.

  7. Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density. (United States)

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


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

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

    CERN Document Server

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


    We use Brownian dynamics simulations to study the permeation properties of a generic electrostatic model of a biological ion channel as a function of the fixed charge Q_f at its selectivity filter. We reconcile the recently-discovered discrete calcium conduction bands M0 (Q_f=1e), M1 (3e), M2 (5e) with the set of sodium conduction bands L0 (0.5-0.7e), L1 (1.5-2e) thereby obtaining a completed pattern of conduction and selectivity bands v Q_f for the sodium-calcium channels family. An increase of Q_f leads to an increase of calcium selectivity: L0 (sodium selective, non-blocking channel) -> M0 (non-selective 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 L1 band is identified with the eukaryotic (DEKA) sodium channel, and L0 (speculatively) with the bacterial NaChBac channel. The scheme created is able to account for the experimentally observed mutation-induced ...

  9. Distinct regions that control ion selectivity and calcium-dependent activation in the bestrophin ion channel. (United States)

    Vaisey, George; Miller, Alexandria N; Long, Stephen B


    Cytoplasmic calcium (Ca(2+)) activates the bestrophin anion channel, allowing chloride ions to flow down their electrochemical gradient. Mutations in bestrophin 1 (BEST1) cause macular degenerative disorders. Previously, we determined an X-ray structure of chicken BEST1 that revealed the architecture of the channel. Here, we present electrophysiological studies of purified wild-type and mutant BEST1 channels and an X-ray structure of a Ca(2+)-independent mutant. From these experiments, we identify regions of BEST1 responsible for Ca(2+) activation and ion selectivity. A "Ca(2+) clasp" within the channel's intracellular region acts as a sensor of cytoplasmic Ca(2+). Alanine substitutions within a hydrophobic "neck" of the pore, which widen it, cause the channel to be constitutively active, irrespective of Ca(2+). We conclude that the primary function of the neck is as a "gate" that controls chloride permeation in a Ca(2+)-dependent manner. In contrast to what others have proposed, we find that the neck is not a major contributor to the channel's ion selectivity. We find that mutation of a cytosolic "aperture" of the pore does not perturb the Ca(2+) dependence of the channel or its preference for anions over cations, but its mutation dramatically alters relative permeabilities among anions. The data suggest that the aperture functions as a size-selective filter that permits the passage of small entities such as partially dehydrated chloride ions while excluding larger molecules such as amino acids. Thus, unlike ion channels that have a single "selectivity filter," in bestrophin, distinct regions of the pore govern anion-vs.-cation selectivity and the relative permeabilities among anions.

  10. Antioxidant effect of T-type calcium channel blockers in gastric injury. (United States)

    Bilici, Dilek; Banoğlu, Z Nur; Kiziltunç, Ahmet; Avci, Bahattin; Ciftçioğlu, Akif; Bilici, Sefa


    It is known that calcium ion has an important role in the cellular function. For this reason, calcium channel blockers may have a protective action against gastric injury which is induced by various stimuli. In this study, the influence of mibefradil on ethanol-induced gastric injury was investigated in rats. Mibefradil was given at a dose 50 mg/kg intraperitoneally 30 min before administration of 1 ml absolute ethanol given by gavage. We compared this effect of mibefradil with that of omeprazol. Ethanol-induced mucosal damage was evaluated using three different approaches: analysis of biochemical parameters and pathologic and macroscopic investigation. It was found that pretreatment with mibefradil significantly reduced ethanol-induced macroscopic, pathologic, and biochemical changes in the gastric mucosa. In conclusion, it is speculated that this findings may prove important in the development of new and improved therapies for the treatment and prevention of gastric ulcers in humans.

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

    Directory of Open Access Journals (Sweden)

    Charalampos Seretis


    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Ceremuzynski, L.K.; Klos, J.; Barcikowski, B.; Herbaczynska-Cedro, K. (Department of Cardiology, Postgraduate Medical School, Warsaw (Poland))


    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 {times} 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.

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

    Directory of Open Access Journals (Sweden)

    Paula Patricia Perissinotti


    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.

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

    Directory of Open Access Journals (Sweden)

    Lei Ray Zhong

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

  15. Responses of channel catfish (Ictalurus punctatus) swim-up fry to dietary calcium in soft and hard water. (United States)

    Scarpa, J; Gatlin, D M


    1. Responses of channel catfish (Ictalurus punctatus) swim-up fry to dietary calcium in soft ( 100 mg/l as CaCO3) water were determined by feeding purified egg-white diets containing 0, 0.5, 1.0, or 2.0% calcium from CaCO3 for 8 weeks. 2. Catfish fry fed the basal diet (0.03% Ca) in hard and soft water had lower whole-body ash and whole-body calcium concentrations but higher weight gain and survival than those fed calcium-supplemented diets. 3. Fry in soft water generally had lower whole-body ash, whole-body calcium, and survival, as well as a higher incidence of spinal deformities than fry in hard water. 4. Feeding higher levels of calcium to fry reared in soft water did not increase whole-body calcium levels or decrease spinal deformities to the levels observed for fry reared in hard water and fed supplemental calcium. 5. These data indicate that calcium derived solely from dietary or environmental sources was not sufficient for optimum health of channel catfish fry.

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

    Directory of Open Access Journals (Sweden)

    Vanessa Silva-Moraes


    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.


    Directory of Open Access Journals (Sweden)



    Full Text Available In the present study, 3-D QSAR analysis was performed on the previously synthesized and evaluated derivatives of novel 2-arylthiazolidinones as selective analgesic N-type calcium channel blockers. Calcium Channel blockers is the molecular target responsible for the treatment of neuropathic and inflammatory pain. The 3D-QSAR study based on the principle of the alignment of pharmacophoric features by PHASE module of Schrodinger suite has been carried out on the same set of calcium channel blockers. Statistically significant 3-D QSAR model (R2=0.9288 were generated using 21 molecules in the training set. The predictive ability of model was determined using a randomly chosen test set of 6 molecules which gave predictive correlation coefficients (R2pred of 0.946 for 3-D models, indicating good predictive power. PHASE pharmacophore hypothesis AAHR.13 may correspond very closely to the interactions recorded in the active site of the ligand bound complex. These studies produced models with high correlation coefficient and good predictive abilities. Docking studies were also carried out wherein these analogues were docked into the active sites of COX-2 to analyze the receptor-ligand interactions that confer selectivity for COX-2. Compound 2 have the highest dock score (-7.28. In the active site, there are some strong hydrogen-bonding interactions observed between residues GLU67, ALA103, ASP96, SER184 and ASP22. Additionally a correlation of the quantitative structure –activity relationship data and the docking results is found to validate each other and suggest the importance of the binding step in overall drug action.

  18. The role of L-type calcium channels in the development and expression of behavioral sensitization to ethanol. (United States)

    Broadbent, Julie


    Behavioral sensitization is thought to play a significant role in drug addiction. L-type calcium channels have been implicated in sensitization to stimulant and opiate drugs but it is unclear if these channels also contribute to sensitization to ethanol. The effects of three L-type calcium channel blockers, nifedipine (1-7.5 mg/kg), diltiazem (12.5-50 mg/kg), and verapamil (12.5 and 25 mg/kg), on sensitization to ethanol (2 g/kg) were examined in DBA/2J mice. All three blockers reduced but did not prevent expression of sensitization. Only nifedipine blocked acquisition of sensitization. Nifedipine and verapamil decreased blood ethanol levels. The current findings suggest L-type calcium channels do not play a substantial role in sensitization to ethanol and that the neural mechanisms underlying sensitization to ethanol are distinct from those mediating sensitization to stimulants and opiates.


    Directory of Open Access Journals (Sweden)

    Yu. V. Lukina


    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.

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

    DEFF Research Database (Denmark)

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


    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...

  1. The omega-atracotoxins: selective blockers of insect M-LVA and HVA calcium channels. (United States)

    Chong, Youmie; Hayes, Jessica L; Sollod, Brianna; Wen, Suping; Wilson, David T; Hains, Peter G; Hodgson, Wayne C; Broady, Kevin W; King, Glenn F; Nicholson, Graham M


    The omega-atracotoxins (omega-ACTX) are a family of arthropod-selective peptide neurotoxins from Australian funnel-web spider venoms (Hexathelidae: Atracinae) that are candidates for development as biopesticides. We isolated a 37-residue insect-selective neurotoxin, omega-ACTX-Ar1a, from the venom of the Sydney funnel-web spider Atrax robustus, with high homology to several previously characterized members of the omega-ACTX-1 family. The peptide induced potent excitatory symptoms, followed by flaccid paralysis leading to death, in acute toxicity tests in house crickets. Using isolated smooth and skeletal nerve-muscle preparations, the toxin was shown to lack overt vertebrate toxicity at concentrations up to 1 microM. To further characterize the target of the omega-ACTXs, voltage-clamp analysis using the whole-cell patch-clamp technique was undertaken using cockroach dorsal unpaired median neurons. It is shown here for the first time that omega-ACTX-Ar1a, and its homolog omega-ACTX-Hv1a from Hadronyche versuta, reversibly block both mid-low- (M-LVA) and high-voltage-activated (HVA) insect calcium channel (Ca(v)) currents. This block occurred in the absence of alterations in the voltage-dependence of Ca(v) channel activation, and was voltage-independent, suggesting that omega-ACTX-1 family toxins are pore blockers rather than gating modifiers. At a concentration of 1 microM omega-ACTX-Ar1a failed to significantly affect global K(v) channel currents. However, 1 microM omega-ACTX-Ar1a caused a modest 18% block of insect Na(v) channel currents, similar to the minor block of Na(v) channels reported for other insect Ca(v) channel blockers such as omega-agatoxin IVA. These findings validate both M-LVA and HVA Ca(v) channels as potential targets for insecticides.

  2. Developmental mapping of small-conductance calcium-activated potassium channel expression in the rat nervous system. (United States)

    Gymnopoulos, Marco; Cingolani, Lorenzo A; Pedarzani, Paola; Stocker, Martin


    Early electrical activity and calcium influx regulate crucial aspects of neuronal development. Small-conductance calcium-activated potassium (SK) channels regulate action potential firing and shape calcium influx through feedback regulation in mature neurons. These functions, observed in the adult nervous system, make them ideal candidates to regulate activity- and calcium-dependent processes in neurodevelopment. However, to date little is known about the onset of expression and regions expressing SK channel subunits in the embryonic and postnatal development of the central nervous system (CNS). To allow studies on the contribution of SK channels to different phases of development of single neurons and networks, we have performed a detailed in situ hybridization mapping study, providing comprehensive distribution profiles of all three SK subunits (SK1, SK2, and SK3) in the rat CNS during embryonic and postnatal development. SK channel transcripts are expressed at early stages of prenatal CNS development. The three SK channel subunits display different developmental expression gradients in distinct CNS regions, with time points of expression and up- or downregulation that can be associated with a range of diverse developmental events. Their early expression in embryonic development suggests an involvement of SK channels in the regulation of developmental processes. Additionally, this study shows how the postnatal ontogenetic patterns lead to the adult expression map for each SK channel subunit and how their coexpression in the same regions or neurons varies throughout development.

  3. Voltage and Calcium Dual Channel Optical Mapping of Cultured HL-1 Atrial Myocyte Monolayer (United States)

    Zhao, Weiwei; Fast, Vladimir G.; Ye, Tong; Ai, Xun


    Optical mapping has proven to be a valuable technique to detect cardiac electrical activity on both intact ex vivo hearts and in cultured myocyte monolayers. HL-1 cells have been widely used as a 2-Dimensional cellular model for studying diverse aspects of cardiac physiology. However, it has been a great challenge to optically map calcium (Ca) transients and action potentials simultaneously from the same field of view in a cultured HL-1 atrial cell monolayer. This is because special handling and care is required to prepare healthy cells that can be electrically captured and optically mapped. Therefore, we have developed an optimal working protocol for dual channel optical mapping. In this manuscript, we have described in detail how to perform the dual channel optical mapping experiment. This protocol is a useful tool to enhance the understanding of action potential propagation and Ca kinetics in arrhythmia development. PMID:25867896

  4. L-type calcium channels and calcium/calmodulin-dependent kinase II differentially mediate behaviors associated with nicotine withdrawal in mice. (United States)

    Jackson, K J; Damaj, M I


    Smoking is a widespread health problem. Because the nicotine withdrawal syndrome is a major contributor to continued smoking and relapse, it is important to understand the molecular and behavioral mechanisms of nicotine withdrawal to generate more effective smoking cessation therapies. Studies suggest a role for calcium-dependent mechanisms, such as L-type calcium channels and calcium/calmodulin-dependent protein kinase II (CaMKII), in the effects of nicotine dependence; however, the role of these mechanisms in nicotine-mediated behaviors is unclear. Thus, the goal of this study was to elucidate the role of L-type calcium channels and CaMKII in nicotine withdrawal behaviors. Using both pharmacological and genetic methods, our results show that L-type calcium channels are involved in physical, but not affective, nicotine withdrawal behaviors. Although our data do provide evidence of a role for CaMKII in nicotine withdrawal behaviors, our pharmacological and genetic assessments yielded different results concerning the specific role of the kinase. Pharmacological data suggest that CaMKII is involved in somatic signs and affective nicotine withdrawal, and activity level is decreased after nicotine withdrawal, whereas the genetic assessments yielded results suggesting that CaMKII is involved only in the anxiety-related response, yet the kinase activity may be increased after nicotine withdrawal; thus, future studies are necessary to clarify the precise behavioral specifics of the relevance of CaMKII in nicotine withdrawal behaviors. Overall, our data show that L-type calcium channels and CaMKII are relevant in nicotine withdrawal and differentially mediate nicotine withdrawal behaviors.

  5. T-type calcium channels consolidate tonic action potential output of thalamic neurons to neocortex. (United States)

    Deleuze, Charlotte; David, François; Béhuret, Sébastien; Sadoc, Gérard; Shin, Hee-Sup; Uebele, Victor N; Renger, John J; Lambert, Régis C; Leresche, Nathalie; Bal, Thierry


    The thalamic output during different behavioral states is strictly controlled by the firing modes of thalamocortical neurons. During sleep, their hyperpolarized membrane potential allows activation of the T-type calcium channels, promoting rhythmic high-frequency burst firing that reduces sensory information transfer. In contrast, in the waking state thalamic neurons mostly exhibit action potentials at low frequency (i.e., tonic firing), enabling the reliable transfer of incoming sensory inputs to cortex. Because of their nearly complete inactivation at the depolarized potentials that are experienced during the wake state, T-channels are not believed to modulate tonic action potential discharges. Here, we demonstrate using mice brain slices that activation of T-channels in thalamocortical neurons maintained in the depolarized/wake-like state is critical for the reliable expression of tonic firing, securing their excitability over changes in membrane potential that occur in the depolarized state. Our results establish a novel mechanism for the integration of sensory information by thalamocortical neurons and point to an unexpected role for T-channels in the early stage of information processing.

  6. Molecular mechanism underlying β1 regulation in voltage- and calcium-activated potassium (BK) channels. (United States)

    Castillo, Karen; Contreras, Gustavo F; Pupo, Amaury; Torres, Yolima P; Neely, Alan; González, Carlos; Latorre, Ramon


    Being activated by depolarizing voltages and increases in cytoplasmic Ca(2+), voltage- and calcium-activated potassium (BK) channels and their modulatory β-subunits are able to dampen or stop excitatory stimuli in a wide range of cellular types, including both neuronal and nonneuronal tissues. Minimal alterations in BK channel function may contribute to the pathophysiology of several diseases, including hypertension, asthma, cancer, epilepsy, and diabetes. Several gating processes, allosterically coupled to each other, control BK channel activity and are potential targets for regulation by auxiliary β-subunits that are expressed together with the α (BK)-subunit in almost every tissue type where they are found. By measuring gating currents in BK channels coexpressed with chimeras between β1 and β3 or β2 auxiliary subunits, we were able to identify that the cytoplasmic regions of β1 are responsible for the modulation of the voltage sensors. In addition, we narrowed down the structural determinants to the N terminus of β1, which contains two lysine residues (i.e., K3 and K4), which upon substitution virtually abolished the effects of β1 on charge movement. The mechanism by which K3 and K4 stabilize the voltage sensor is not electrostatic but specific, and the α (BK)-residues involved remain to be identified. This is the first report, to our knowledge, where the regulatory effects of the β1-subunit have been clearly assigned to a particular segment, with two pivotal amino acids being responsible for this modulation.

  7. Mechanism underlying blockade of voltage-gated calcium channels by agmatine in cultured rat hippocampal neurons

    Institute of Scientific and Technical Information of China (English)

    Jian-quan ZHENG; Xie-chuan WENG; Xiao-dan GAI; Jin LI; Wen-bin XIAO


    AIM: To investigate whether agmatine could selectively block a given type of the voltage-gated calcium channels (VGCC) and whether related receptors are involved in the blocking effect of agmatine on VGCC. METHODS: The whole-cell patch recording technique was performed to record VGCC currents in the cultured neonatal rat hippocampal neurons. RESULTS: Verapamil (100 μmol/L), a selective blocker of L-type calcium channel, significantly inhibited VGCC current by 80 %± 7 %. Agmatine (100 μmol/L) could further depress the remained currents by 25 %±6 %. The α2-adrenoceptor antagonist yohimbine (10 μmol/L) and the I2 imidazoline receptor antagonist idazoxon (10 and 40 μmol/L) had no significant effect on VGCC currents when used respectively. When the mixture of yohimbine and agmatine was applied, VGCC currents were still depressed remarkably. However, the blocking effect of agmatine was decreased by 29 %± 8 % in the presence of idazoxon (10 μmol/L). The effect of idazoxon did not increase at a higher concentration (40 μmol/L). CONCLUSION: Agmatine could block the L- and other types of VGCC currents in the cultured rat hippocampal neurons. Blocking effect of agmatine on VGCC was partially related to I2 imidazoline receptor and had no relationship with α2-adrenoceptors.

  8. Synergistic Effect of Fluconazole and Calcium Channel Blockers against Resistant Candida albicans. (United States)

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


    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) fluconazole plus amlodipine caused down-regulating of CNA1, CNB1 (encoding calcineurin) and YVC1 (encoding calcium channel protein in vacuole membrane).

  9. 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.

  10. Synergism of ochratoxin B and calcium-channel antagonist verapamil caused mitochondrial dysfunction. (United States)

    Chatopadhyay, Pronobesh; Tariang, Banlumlang; Agnihotri, Amit; Veer, Vijay


    We examined the mechanism by which the ochratoxin B induced interaction with calcium-channel antagonist verapamil and mitochondrial dysfunction of the rat trachea in vitro experiment. The tracheas were cut into 2-3 mm wide rings and suspended in a tissue bath. Isometric tension was continuously measured with an isometric force transducer connected to a computer-based data acquisition system. Verapamil (1 × 10(-6) M) produced a concentration-dependent contraction response in rat's tracheal rings pre-contracted by acetylcholine. Incubation of rat's tracheal rings with the ochratoxin B significantly potentiated the contraction responses of verapamil. Verapamil and OTB accelerate the overloading of Ca(2+) in tracheal smooth muscle contributes the tissue toxicity as shown in electron microscopy and mitochondrial enzymes, through a mechanism that could involve perturbations of Ca(2+) homeostasis. These results proved that ochratoxin B is a potential vasoconstrictor mycotoxin with the presence of calcium-channel antagonist. In conclusion, disturbance of Ca(2+) homeostasis caused by OTA and plays a significant role in produces toxicity through mitochondrial enzyme inhibition.

  11. Calcium-channel blockers and other factors influencing delayed function in renal allografts. (United States)

    Ferguson, C J; Hillis, A N; Williams, J D; Griffin, P J; Salaman, J R


    A retrospective analysis was undertaken to examine the influence of calcium-channel blocking drugs on early renal allograft function. Delayed function was defined as the need for dialysis or a reduction in serum creatinine of less than 15% within 4 days of transplantation. The drug histories of 172 patients were examined. After exclusions, the data from 138 patients were analysed. No patient was taking any calcium-channel blocking drug other than nifedipine. Thirty-one patients were taking nifedipine at the time of transplantation and these had a delayed function rate of 16% compared with 40% for 107 patients not taking nifedipine (chi 2, P less than 0.05). Delayed function occurred in 61% of cases when the donor age was over 50 years compared with 29% with younger donors (chi 2, P less than 0.05). A total ischaemic time of longer than 24 h and administration of inotropic support to the donor were associated with delayed function (chi 2, P less than 0.05). Administration to the donor of mannitol, steroids, phenoxybenzamine and heparin had no effect on the rate of delayed function. Recipients treated with low-dose dopamine in the perioperative period had no advantage. Elevated trough whole blood concentrations of cyclosporin in the first week after transplant were associated with delayed function (Mann-Whitney U, P less than 0.05).

  12. High-dose insulin therapy in beta-blocker and calcium channel-blocker poisoning. (United States)

    Engebretsen, Kristin M; Kaczmarek, Kathleen M; Morgan, Jenifer; Holger, Joel S


    INTRODUCTION. High-dose insulin therapy, along with glucose supplementation, has emerged as an effective treatment for severe beta-blocker and calcium channel-blocker poisoning. We review the experimental data and clinical experience that suggests high-dose insulin is superior to conventional therapies for these poisonings. PRESENTATION AND GENERAL MANAGEMENT. Hypotension, bradycardia, decreased systemic vascular resistance (SVR), and cardiogenic shock are characteristic features of beta-blocker and calcium-channel blocker poisoning. Initial treatment is primarily supportive and includes saline fluid resuscitation which is essential to correct vasodilation and low cardiac filling pressures. Conventional therapies such as atropine, glucagon and calcium often fail to improve hemodynamic status in severely poisoned patients. Catecholamines can increase blood pressure and heart rate, but they also increase SVR which may result in decreases in cardiac output and perfusion of vascular beds. The increased myocardial oxygen demand that results from catecholamines and vasopressors may be deleterious in the setting of hypotension and decreased coronary perfusion. METHODS. The Medline, Embase, Toxnet, and Google Scholar databases were searched for the years 1975-2010 using the terms: high-dose insulin, hyperinsulinemia-euglycemia, beta-blocker, calcium-channel blocker, toxicology, poisoning, antidote, toxin-induced cardiovascular shock, and overdose. In addition, a manual search of the Abstracts of the North American Congress of Clinical Toxicology and the Congress of the European Association of Poisons Centres and Clinical Toxicologists published in Clinical Toxicology for the years 1996-2010 was undertaken. These searches identified 485 articles of which 72 were considered relevant. MECHANISMS OF HIGH-DOSE INSULIN BENEFIT. There are three main mechanisms of benefit: increased inotropy, increased intracellular glucose transport, and vascular dilatation. EFFICACY OF HIGH

  13. Calcium regulation of HCN channels supports persistent activity in a multiscale model of neocortex. (United States)

    Neymotin, S A; McDougal, R A; Bulanova, A S; Zeki, M; Lakatos, P; Terman, D; Hines, M L; Lytton, W W


    Neuronal persistent activity has been primarily assessed in terms of electrical mechanisms, without attention to the complex array of molecular events that also control cell excitability. We developed a multiscale neocortical model proceeding from the molecular to the network level to assess the contributions of calcium (Ca(2+)) regulation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in providing additional and complementary support of continuing activation in the network. The network contained 776 compartmental neurons arranged in the cortical layers, connected using synapses containing AMPA/NMDA/GABAA/GABAB receptors. Metabotropic glutamate receptors (mGluR) produced inositol triphosphate (IP3) which caused the release of Ca(2+) from endoplasmic reticulum (ER) stores, with reuptake by sarco/ER Ca(2+)-ATP-ase pumps (SERCA), and influence on HCN channels. Stimulus-induced depolarization led to Ca(2+) influx via NMDA and voltage-gated Ca(2+) channels (VGCCs). After a delay, mGluR activation led to ER Ca(2+) release via IP3 receptors. These factors increased HCN channel conductance and produced firing lasting for ∼1min. The model displayed inter-scale synergies among synaptic weights, excitation/inhibition balance, firing rates, membrane depolarization, Ca(2+) levels, regulation of HCN channels, and induction of persistent activity. The interaction between inhibition and Ca(2+) at the HCN channel nexus determined a limited range of inhibition strengths for which intracellular Ca(2+) could prepare population-specific persistent activity. Interactions between metabotropic and ionotropic inputs to the neuron demonstrated how multiple pathways could contribute in a complementary manner to persistent activity. Such redundancy and complementarity via multiple pathways is a critical feature of biological systems. Mediation of activation at different time scales, and through different pathways, would be expected to protect against disruption, in

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

    Institute of Scientific and Technical Information of China (English)

    LI JunYing


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

  15. Effects of acute and chronic nicotine on elevated plus maze in mice: involvement of calcium channels. (United States)

    Biala, Grazyna; Budzynska, Barbara


    The current experiments examined the anxiety-related effects of acute and repeated nicotine administration using the elevated plus maze test in mice. Nicotine (0.1 mg/kg s.c., 5 and 30 min after injection; 0.5 mg/kg, s.c., 5 min after injection) had an anxiogenic effect, shown by specific decreases in the percentage of time spent on the open arms and in the percentage of open arm entries. Tolerance developed to this anxiogenic action after 6 days of daily nicotine administration (0.1 mg/kg, s.c.). Five minutes after the seventh injection, an anxiolytic effect was observed, i.e., specific increases in the percentage of time spent on the open arms and in the percentage of open arm entries. L-type voltage-dependent calcium channel antagonists nimodipine (5 and 10 mg/kg, i.p.), flunarizine (5 and 10 mg/kg, i.p.), verapamil (5, 10, 20 mg/kg) and diltiazem (5, 10, 20 mg/kg, i.p.) were also injected prior to an acute low dose of nicotine or to each injection of chronic nicotine. Our results revealed that calcium channel blockers dose-dependently attenuated both an anxiogenic effect of nicotine as well as the development of tolerance to this effect. Our results suggest that neural calcium-dependent mechanisms are involved in the anxiety-related responses to acute and chronic nicotine injection that may ultimately lead to addiction and smoking relapse in human smokers.

  16. Changes of neuronal calcium channel following brain damage induced by injection of pertussis bacilli in rats

    Institute of Scientific and Technical Information of China (English)

    陈立华; 于嘉; 刘丽旭; 曹美鸿


    To explore changes of neuronal calcium channel following brain damage induced by injection of pertussis bacilli in rats, and to investigate the relationship between cytosolic free calcium concentration ( [ Ca2 + ] i ) in the synaptosome and Ca2 + -ATPase activities of mitochondria. Methods: The level of [ Ca2+ ]i in the synaptosome and Ca2+ -ATPase activities of mitochondria in the acute brain damage induced by injection of pertussis bacilli (PB)in rat was determined and nimodipine was administrated to show its effects on [ Ca2+ ]i in the synaptosome and on alteration of Ca2+ -ATPase activity in the mitochondria.Seventy-three rats were randomly divided into four groups,ie, normal control group (Group A ), sham-operation control group (Group B), PB group (Group C) and nimodipine treatment group (Group D). Results: The level of [ Ca2+ ]i was significantly increased in the PB-injected cerebral hemisphere in the Group C as compared with that in the Group A and the Group B at 30 minutes after injection of PB. The level of [ Ca2+ ]i was kept higher in the 4 hours and 24 hours subgroups after the injection in the Group C ( P < 0.05).In contrast, the Ca2+ -ATPase activities were decreased remarkably among all of the subgroups in the Group C.Nimodipine, which was administered after injection of PB,could significantly decrease the [ Ca2+ ]i and increase the activity of Ca2 + -ATPase ( P < 0.05 ). Conclusions: The neuronal calcium channel is opened after injection of PB. There is a negative correlation between activities of Ca2 +-ATPase and [ Ca2 + ]i.Nimodipine can reduce brain damage through stimulating the activities of Ca2+ -ATPase in the mitochondria, and decrease the level of [ Ca2+ ]i in the synaptosome.Treatment with nimodipine dramatically reduces the effects of brain damage induced by injection of PB.

  17. Lavender oil-potent anxiolytic properties via modulating voltage dependent calcium channels. (United States)

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


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

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

    Directory of Open Access Journals (Sweden)

    Anita M Schuwald

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

  19. Neuroprotective activity of stiripentol with a possible involvement of voltage-dependent calcium and sodium channels. (United States)

    Verleye, Marc; Buttigieg, Dorothée; Steinschneider, Rémy


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

  20. Possible roles of exceptionally conserved residues around the selectivity filters of sodium and calcium channels. (United States)

    Tikhonov, Denis B; Zhorov, Boris S


    In the absence of x-ray structures of sodium and calcium channels their homology models are used to rationalize experimental data and design new experiments. A challenge is to model the outer-pore region that folds differently from potassium channels. Here we report a new model of the outer-pore region of the NaV1.4 channel, which suggests roles of highly conserved residues around the selectivity filter. The model takes from our previous study (Tikhonov, D. B., and Zhorov, B. S. (2005) Biophys. J. 88, 184-197) the general disposition of the P-helices, selectivity filter residues, and the outer carboxylates, but proposes new intra- and inter-domain contacts that support structural stability of the outer pore. Glycine residues downstream from the selectivity filter are proposed to participate in knob-into-hole contacts with the P-helices and S6s. These contacts explain the adapted tetrodotoxin resistance of snakes that feed on toxic prey through valine substitution of isoleucine in the P-helix of repeat IV. Polar residues five positions upstream from the selectivity filter residues form H-bonds with the ascending-limb backbones. Exceptionally conserved tryptophans are engaged in inter-repeat H-bonds to form a ring whose π-electrons would facilitate passage of ions from the outer carboxylates to the selectivity filter. The outer-pore model of CaV1.2 derived from the NaV1.4 model is also stabilized by the ring of exceptionally conservative tryptophans and H-bonds between the P-helices and ascending limbs. In this model, the exceptionally conserved aspartate downstream from the selectivity-filter glutamate in repeat II facilitates passage of calcium ions to the selectivity-filter ring through the tryptophan ring. Available experimental data are discussed in view of the models.

  1. TRPC1 regulates calcium-activated chloride channels in salivary gland cells. (United States)

    Sun, Yuyang; Birnbaumer, Lutz; Singh, Brij B


    Calcium-activated chloride channel (CaCC) plays an important role in modulating epithelial secretion. It has been suggested that in salivary tissues, sustained fluid secretion is dependent on Ca(2+) influx that activates ion channels such as CaCC to initiate Cl(-) efflux. However direct evidence as well as the molecular identity of the Ca(2+) channel responsible for activating CaCC in salivary tissues is not yet identified. Here we provide evidence that in human salivary cells, an outward rectifying Cl(-) current was activated by increasing [Ca(2+)]i, which was inhibited by the addition of pharmacological agents niflumic acid (NFA), an antagonist of CaCC, or T16Ainh-A01, a specific TMEM16a inhibitor. Addition of thapsigargin (Tg), that induces store-depletion and activates TRPC1-mediated Ca(2+) entry, potentiated the Cl(-) current, which was inhibited by the addition of a non-specific TRPC channel blocker SKF96365 or removal of external Ca(2+). Stimulation with Tg also increased plasma membrane expression of TMEM16a protein, which was also dependent on Ca(2+) entry. Importantly, in salivary cells, TRPC1 silencing, but not that of TRPC3, inhibited CaCC especially upon store depletion. Moreover, primary acinar cells isolated from submandibular gland also showed outward rectifying Cl(-) currents upon increasing [Ca(2+)]i. These Cl(-) currents were again potentiated with the addition of Tg, but inhibited in the presence of T16Ainh-A01. Finally, acinar cells isolated from the submandibular glands of TRPC1 knockout mice showed significant inhibition of the outward Cl(-) currents without decreasing TMEM16a expression. Together the data suggests that Ca(2+) entry via the TRPC1 channels is essential for the activation of CaCC.

  2. Differential regulation of voltage- and calcium-activated potassium channels in human B lymphocytes. (United States)

    Partiseti, M; Choquet, D; Diu, A; Korn, H


    The expression and characteristics of K+ channels of human B lymphocytes were studied by using single and whole-cell patch-clamp recordings. They were gated by depolarization (voltage-gated potassium current, IKv, 11-20 pS) and by an increase in intracellular Ca2+ concentration (calcium-activated potassium current, IKCa, 26 pS), respectively. The level of expression of these channels was correlated with the activational status of the cell. Both conductances are blocked by tetraethylammonium, verapamil, and charybdotoxin, and are insensitive to apamin; 4-aminopyridine blocks IK, preferentially. We used a protein kinase C activator (PMA) or antibodies to membrane Ig (anti-mu) to activate resting splenocytes in culture. Although IKv was recorded in the majority of the resting lymphocytic population, less than 20% of the activated cells expressed this conductance. However, in this subset the magnitude of IKv was 20-fold larger than in resting cells. On the other hand, IKCa was detected in nearly one half of the resting cells, whereas all activated cells expressed this current. The magnitude of IKCa was, on average, 30 times larger in activated than in nonactivated cells. These results probably reflect that during the course of activation 1) the number of voltage-dependent K+ channels per cell decreases and increases in a small subset and 2) the number of Ca(2+)-dependent K+ channels per cell increases in all cells. We suggest that the expression of functional Ca(2+)- and voltage-activated K+ channels are under the control of different regulatory signals.

  3. First-principles investigation of exchange interactions in quasi-one-dimensional antiferromagnet CaV2O4 (United States)

    Pchelkina, Z. V.; Solovyev, I. V.


    The effects of orbital degrees of freedom on the exchange interactions in a quasi-one-dimensional spin-1 antiferromagnet CaV2O4 are systematically studied. For this purpose a realistic low-energy electron model with the parameters derived from the first-principles calculations is constructed in the Wannier basis for the t2g bands. The exchange interactions are calculated using both the theory of infinitesimal spin rotations near the mean-field ground state and the superexchange model, which provide a consistent description. The obtained behaviour of exchange interactions differs substantially from the previously proposed phenomenological picture based on magnetic measurements and structural considerations, namely: (i) despite the quasi-one-dimensional character of the crystal structure, consisting of the zigzag chains of the edge-sharing VO6 octahedra, the electronic structure is essentially three-dimensional, that leads to finite interactions between the chains; (ii) the exchange interactions along the legs of the chains appear to dominate; and (iii) there is a substantial difference in exchange interactions in two crystallographically inequivalent chains. The combination of these three factors successfully reproduces the behaviour of experimental magnetic susceptibility.

  4. [Effect of calcium channel blockers on developing nervous syndrome of high pressure and nitrogen narcosis in mice]. (United States)

    Sledkov, A I


    In the experiments conducted on mice which prior to compression in a heliox environment have been injected the blockers of various types of calcium channels (flunarezine, verapramil and nifedipine) as well as bemethyl (actoprotector) and oxymethacye (antioxidant) there escaped detection of noticeable effect of these drugs on developing the high pressure nervous syndrome (HPNS). On exposure to the hyperbaric nitrogen-oxygen environment verapromil (phenylalkulamine blocker of L-type calcium channels) had a protection effect with respect to a convulsive component of the nitrogen narcosis.

  5. The large conductance calcium-activated K(+) channel interacts with the small GTPase Rab11b. (United States)

    Sokolowski, Sophia; Harvey, Margaret; Sakai, Yoshihisa; Jordan, Amy; Sokolowski, Bernd


    The transduction of sound by the receptor or hair cells of the cochlea leads to the activation of ion channels found in the basal and lateral regions of these cells. Thus, the processing of these transduced signals to the central nervous system is tied to the regulation of baso-lateral ion channels. The large conductance calcium-activated potassium or BK channel was revealed to interact with the small GTPase, Rab11b, which is one of many Rabs found in various endosomal pathways. Immunoelectron microscopy showed the colocalization of these two proteins in receptor cells and auditory neurons. Using Chinese hamster ovary cells as a heterologous expression system, Rab11b increased or decreased BK expression, depending on the overexpression or RNAi knockdown of Rab, respectively. Additional mutation analyses, using a yeast two-hybrid assay, suggested that this GTPase moderately interacts within a region of BK exclusive of the N- or C-terminal tails. These data suggest that this small GTPase regulates BK in a slow recycling process through the endocytic compartment and to the plasmalemma.

  6. R-type calcium channels are crucial for semaphorin 3A-induced DRG axon growth cone collapse.

    Directory of Open Access Journals (Sweden)

    Rimantas Treinys

    Full Text Available Semaphorin 3A (Sema3A is a secreted protein involved in axon path-finding during nervous system development. Calcium signaling plays an important role during axonal growth in response to different guidance cues; however it remains unclear whether this is also the case for Sema3A. In this study we used intracellular calcium imaging to figure out whether Sema3A-induced growth cone collapse is a Ca2+ dependent process. Intracellular Ca2+ imaging results using Fura-2 AM showed Ca2+ increase in E15 mice dorsal root ganglia neurons upon Sema3A treatment. Consequently we analyzed Sema3A effect on growth cones after blocking or modifying intracellular and extracellular Ca2+ channels that are expressed in E15 mouse embryos. Our results demonstrate that Sema3A increased growth cone collapse rate is blocked by the non-selective R- and T- type Ca2+ channel blocker NiCl2 and by the selective R-type Ca2+ channel blocker SNX482. These Ca2+ channel blockers consistently decreased the Sema3A-induced intracellular Ca2+ concentration elevation. Overall, our results demonstrate that Sema3A-induced growth cone collapses are intimately related with increase in intracellular calcium concentration mediated by R-type calcium channels.

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

    Directory of Open Access Journals (Sweden)

    See-Ziau Hoe


    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.

  8. Exposure to extremely low-frequency electromagnetic fields inhibits T-type calcium channels via AA/LTE4 signaling pathway. (United States)

    Cui, Yujie; Liu, Xiaoyu; Yang, Tingting; Mei, Yan-Ai; Hu, Changlong


    Extremely low-frequency electromagnetic fields (ELF-EMF) causes various biological effects through altering intracellular calcium homeostasis. The role of high voltage-gated (HVA) calcium channels in ELF-EMF induced effects has been extensively studied. However, the effect of ELF-EMF on low-voltage-gated (LVA) T-type calcium channels has not been reported. In this study, we test the effect of ELF-EMF (50Hz) on human T-type calcium channels transfected in HEK293 cells. Conversely to its stimulant effects on HVA channels, ELF-EMF exposure inhibited all T-type (Cav3.1, Cav3.2 and Cav3.3) channels. Neither the protein expression nor the steady-state activation and inactivation kinetics of Cav3.2 channels were altered by ELF-EMF (50Hz, 0.2mT) exposure. Exposure to ELF-EMF increased both arachidonic acid (AA) and leukotriene E4 (LTE4) levels in HEK293 cells. CAY10502 and bestatin, which block the increase of AA and LTE4 respectively, abrogated the ELF-EMF inhibitory effect on Cav3.2 channels. Exogenous LTE4 mimicked the ELF-EMF inhibition of T-type calcium channels. ELF-EMF (50Hz) inhibits native T-type calcium channels in primary cultured mouse cortical neurons via LTE4. We conclude that 50Hz ELF-EMF inhibits T-type calcium channels through AA/LTE4 signaling pathway.

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

    NARCIS (Netherlands)

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


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

  10. A deleterious gene-by-environment interaction imposed by calcium channel blockers in Marfan syndrome. (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


    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.

  11. Use of clopidogrel and calcium channel blockers and risk of major adverse cardiovascular events

    DEFF Research Database (Denmark)

    Schmidt, Morten; Johansen, Martin B; Robertson, Douglas J


    Eur J Clin Invest 2011 ABSTRACT: Background  The CYP3A4 inhibition by calcium channel blockers (CCBs) may attenuate the effectiveness of clopidogrel. Using time-varying drug exposure ascertainment, we examined whether CCB use modified the association between clopidogrel use and major adverse......-month follow-up, we tracked the use of clopidogrel and CCBs and the rate of MACE (composite of myocardial infarction, ischaemic stroke, stent thrombosis, target lesion revascularization, or cardiac death). We used Cox regression to compute hazard ratios, controlling for potential confounders. Results......  Overall, the 12-month risk for MACE was 14·5%. The rate was 130 per 1000 person years for concomitant clopidogrel and CCB use, 106 for clopidogrel without CCB use, 213 for CCB without clopidogrel use, and 248 for no use of either drug. The adjusted hazard ratio for MACE comparing clopidogrel use...

  12. Mechanism of Action of Novel Glibenclamide Derivatives on Potassium and Calcium Channels for Insulin Secretion. (United States)

    Frederico, Marisa Jádson Silva; Castro, Allisson Jhonatan Gomes; Menegaz, Danusa; De Bernardis Murat, Cahuê; Mendes, Camila Pires; Mascarello, Alessandra; Nunes, Ricardo José; Silva, Fátima Regina Mena Barreto


    Glibenclamide is widely used and remains a cornerstone and an effective antihyperglycemic drug. After the casual discovery of its hypoglycemic potential, this compound was introduced for diabetes treatment. However, the long-term side effects reveal that glibenclamide should be replaced by new molecules able to maintain the health of β-cells, protecting them from hyperstimulation/hyperexcitability, hyperinsulinemia, functional failure and cell death. The aim of this review was to highlight the main mechanism of action of glibenclamide and the influence of its derivatives, such as acyl-hydrazones, sulfonamides and sulfonylthioureas on β-cells potassium and calcium channels for insulin secretion as well as the contribution of these new compounds to restore glucose homeostasis. Furthermore, the role of glibenclamide-based novel structures that promise less excitability of β-cell in a long-term treatment with effectiveness and safety for diabetes therapy was discussed.

  13. 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


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

  14. The mutant Moonwalker TRPC3 channel links calcium signaling to lipid metabolism in the developing cerebellum. (United States)

    Dulneva, Anna; Lee, Sheena; Oliver, Peter L; Di Gleria, Katalin; Kessler, Benedikt M; Davies, Kay E; Becker, Esther B E


    The Moonwalker (Mwk) mouse is a model of dominantly inherited cerebellar ataxia caused by a gain-of-function mutation in the transient receptor potential (TRP) channel TRPC3. Here, we report impairments in dendritic growth and synapse formation early on during Purkinje cell development in the Mwk cerebellum that are accompanied by alterations in calcium signaling. To elucidate the molecular effector pathways that regulate Purkinje cell dendritic arborization downstream of mutant TRPC3, we employed transcriptomic analysis of developing Purkinje cells isolated by laser-capture microdissection. We identified significant gene and protein expression changes in molecules involved in lipid metabolism. Consistently, lipid homeostasis in the Mwk cerebellum was found to be disturbed, and treatment of organotypic cerebellar slices with ceramide significantly improved dendritic outgrowth of Mwk Purkinje cells. These findings provide the first mechanistic insights into the TRPC3-dependent mechanisms, by which activated calcium signaling is coupled to lipid metabolism and the regulation of Purkinje cell development in the Mwk cerebellum.

  15. A key role for STIM1 in store operated calcium channel activation in airway smooth muscle

    Directory of Open Access Journals (Sweden)

    Peel Samantha E


    Full Text Available Abstract Background Control of cytosolic calcium plays a key role in airway myocyte function. Changes in intracellular Ca2+ stores can modulate contractile responses, modulate proliferation and regulate synthetic activity. Influx of Ca2+ in non excitable smooth muscle is believed to be predominantly through store operated channels (SOC or receptor operated channels (ROC. Whereas agonists can activate both SOC and ROC in a range of smooth muscle types, the specific trigger for SOC activation is depletion of the sarcoplasmic reticulum Ca2+ stores. The mechanism underlying SOC activation following depletion of intracellular Ca2+ stores in smooth muscle has not been identified. Methods To investigate the roles of the STIM homologues in SOC activation in airway myocytes, specific siRNA sequences were utilised to target and selectively suppress both STIM1 and STIM2. Quantitative real time PCR was employed to assess the efficiency and the specificity of the siRNA mediated knockdown of mRNA. Activation of SOC was investigated by both whole cell patch clamp electrophysiology and a fluorescence based calcium assay. Results Transfection of 20 nM siRNA specific for STIM1 or 2 resulted in robust decreases (>70% of the relevant mRNA. siRNA targeted at STIM1 resulted in a reduction of SOC associated Ca2+ influx in response to store depletion by cyclopiazonic acid (60% or histamine but not bradykinin. siRNA to STIM2 had no effect on these responses. In addition STIM1 suppression resulted in a more or less complete abrogation of SOC associated inward currents assessed by whole cell patch clamp. Conclusion Here we show that STIM1 acts as a key signal for SOC activation following intracellular Ca2+ store depletion or following agonist stimulation with histamine in human airway myocytes. These are the first data demonstrating a role for STIM1 in a physiologically relevant, non-transformed endogenous expression cell model.

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

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    Mandeep Kaur


    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.

  17. Anti-Convulsant Activity of Boerhaavia diffusa: Plausible Role of Calcium Channel Antagonism. (United States)

    Kaur, Mandeep; Goel, Rajesh Kumar


    "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.

  18. Calcium influx through L-type channels attenuates skeletal muscle contraction via inhibition of adenylyl cyclases. (United States)

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


    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.

  19. Calmodulin modulates the delay period between release of calcium from internal stores and activation of calcium influx via endogenous TRP1 channels. (United States)

    Vaca, Luis; Sampieri, Alicia


    In the present study we have explored the role of calmodulin (CaM) and inositol 1,4,5-trisphosphate receptor (IP(3)R) in the communication process activated after the release of calcium from the endoplasmic reticulum (ER) and the activation of calcium influx via endogenous TRP1 channels from Chinese hamster ovary cells. Experiments using combined rapid confocal calcium and electrophysiology measurements uncovered a consistent delay of around 900 ms between the first detectable calcium released from the ER and the activation of the calcium current. This delay was evident with two different methods used to release calcium from the ER: either the blockade of the microsomal calcium ATPase with thapsigargin or activation of bradykinin receptors linked to the IP(3) cascade. Direct application of IP(3) or a peptide from the NH(2)-terminal region of the IP(3)R activated store operated calcium, reducing the delay period. Introduction of CaM into the cell via the patch pipette increased the delay period from 900 +/- 100 ms to 10 +/- 2.1 s (n = 18). Furthermore, the use of selective CaM antagonists W7 and trifluoperazine maleate resulted in a substantial reduction of the delay period to 200 +/- 100 ms with 5 microm trifluoperazine maleate (n = 16) and 150 +/- 50 ms with 500 nm W7 (n = 22). CaM reduced also the current density activated by thapsigargin or brandykinin to about 60% from control. The CaM antagonists did not affect significantly the current density. The results presented here are consistent with an antagonistic effect of IP(3)R and CaM for the activation of store operated calcium after depletion of the ER. The functional competition between the activating effect of IP(3)R and the inhibiting effect of CaM may modulate the delay period between the release of calcium from the ER and the activation of calcium influx observed in different cells, as well as the amount of current activated after depletion of the ER.

  20. CNTF-ACM promotes mitochondrial respiration and oxidative stress in cortical neurons through upregulating L-type calcium channel activity. (United States)

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


    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.

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

    Institute of Scientific and Technical Information of China (English)

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


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

  2. 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


    -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...

  3. Efficacy and safety of calcium channel blockers in heart failure : Focus on recent trials with second-generation dihydropyridines

    NARCIS (Netherlands)

    de Vries, RJM; van Veldhuisen, DJ; Dunselman, PHJM


    Background Chronic heart failure (CHF) has high morbidity and mortality rates despite treatment with angiotensin-converting-enzyme inhibitors, diuretics, and digoxin. Adjunctive-vasodilation through calcium channel blockade has been suggested as potentially useful, However, the first-generation calc

  4. L—type calcium channel blockers inhibit the development but not the expression of sensitization to morphine in mice

    Institute of Scientific and Technical Information of China (English)

    ZhanQ; ZhenJW


    The relationship between opioid actions and L-type calcium channel blockers has been well documented.However,there is no report relevant to L-type calcium channel blockers and morphinesensitization,which is suggested to be an analog of behaviors that are the characteristics of drug addiction.Here the effects of three L-type calcium channel blockers,nimodipine,nifedipine and verapamil,on morphine-induced locomotor activity,the development and the expression of sensitization to morphine were studied systematically.The results showed that both nimodipine and verapamil attenuated,while nifedipine had only a tendency to decrease morphine-induced locomotor activity.All the three drugs inhibited the development of sensitization to morphine.However,none of them showed any effects on the expression of morphine sensitization.These results indicate that blocking L-tpye calcium channel attenuates the locomotor stimulating effects of morphine and inhibits the development but not the expression of morphine-sensitization.

  5. Dopamine Induces LTP Differentially in Apical and Basal Dendrites through BDNF and Voltage-Dependent Calcium Channels (United States)

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


    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).…

  6. Role for voltage gated calcium channels in calcitonin gene-related peptide release in the rat trigeminovascular system

    DEFF Research Database (Denmark)

    Amrutkar, D V; Ploug, K B; Olesen, J;


    Clinical and genetic studies have suggested a role for voltage gated calcium channels (VGCCs) in the pathogenesis of migraine. Release of calcitonin gene-related peptide (CGRP) from trigeminal neurons has also been implicated in migraine. The VGCCs are located presynaptically on neurons and are i...

  7. Increased pressure-induced tone in rat parenchymal arterioles vs. middle cerebral arteries: role of ion channels and calcium sensitivity. (United States)

    Cipolla, Marilyn J; Sweet, Julie; Chan, Siu-Lung; Tavares, Matthew J; Gokina, Natalia; Brayden, Joseph E


    Brain parenchymal arterioles (PAs) are high-resistance vessels that branch off pial arteries and perfuse the brain parenchyma. PAs are the target of cerebral small vessel disease and have been shown to have greater pressure-induced tone at lower pressures than pial arteries. We investigated mechanisms by which brain PAs have increased myogenic tone compared with middle cerebral arteries (MCAs), focusing on differences in vascular smooth muscle (VSM) calcium and ion channel function. The amount of myogenic tone and VSM calcium was measured using Fura 2 in isolated and pressurized PAs and MCAs. Increases in intraluminal pressure caused larger increases in tone and cytosolic calcium in PAs compared with MCAs. At 50 mmHg, myogenic tone was 37 ± 5% for PAs vs. 6.5 ± 4% for MCAs (P channel (VDCC) inhibitor nifedipine than MCAs (EC50 for PAs was 3.5 ± 0.4 vs. 82.1 ± 2.1 nmol/l for MCAs;P channel inhibitor iberiotoxin, whereas MCAs constricted ∼15%. Thus increased myogenic tone in PAs appears related to differences in ion channel activity that promotes VSM membrane depolarization but not to a direct sensitization of the contractile apparatus to calcium.

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

    DEFF Research Database (Denmark)

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


    The calcium-activated potassium channel KCa3.1 controls different cellular processes such as proliferation and volume homeostasis. We investigated the role of KCa3.1 in experimental and human liver fibrosis. KCa3.1 gene expression was investigated in healthy and injured human and rodent liver. Ef...

  9. Increased Expression of the Calcium-Activated Chloride Channel in Hclca1 in Airways of Patients with Obstructive Chronic Bronchitis

    Directory of Open Access Journals (Sweden)

    Hans-Peter Hauber


    Full Text Available BACKGROUND: Interleukin (IL-9 and its effect on enhancing the human calcium-activated chloride channel 1 (hCLCA1 expression have been shown to induce mucin production. Increased expression of hCLCA1 may, in turn, contribute to mucus overproduction in chronic obstructive pulmonary disease (COPD with a chronic bronchitis (CB phenotype.

  10. Immunogenicity of P/Q-type calcium channel in small cell lung cancer: investigation of alpha1 subunit polyglutamine expansion. (United States)

    Black, J L; Nelson, T R; Snow, K; Lennon, V A


    The ectopic expression of neuronal P/Q-type voltage-gated calcium channels in small cell lung carcinoma (SCLC) is thought to induce antisynaptic autoimmunity in the paraneoplastic Lambert-Eaton myasthenic syndrome. The gene CACNL1A4, encoding the principal (alpha1A) subunit of this calcium channel, is mutated in several inherited neurological disorders. One of these disorders (spinocerebellar ataxia, type 6, or SCA-6) involves the expansion of a trinucleotide (CAG) repeat unit. We hypothesized that a somatic CAG repeat instability of this gene in neoplastic cells might generate a non-self epitope capable of initiating autoimmunity to P/Q-type calcium channels. We therefore analyzed the CACNL1A4 gene in SCLC lines established from metastases derived from seven individual patients (four associated with Lambert-Eaton myasthenic syndrome, one associated with myasthenia gravis, and two not associated with neurological autoimmunity). We compared their CAG repeat numbers (determined by polymerase chain reaction (PCR) amplification followed by separation of products on a 6% polyacrylamide/8M urea gel) to published norms and to DNA from a patient with SCA-6. The number of CAG repeats in SCLC DNA fell within a normal range whether or not the neoplasm was complicated by neurological autoimmunity. Therefore, it is unlikely that somatically unstable CAG repeat units in the gene encoding the P/Q-type voltage-gated calcium channel account for this tumor protein's immunogenicity in the Lambert-Eaton myasthenic syndrome.

  11. 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;


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

  12. Defining the role of calcium channel antagonists in heart failure due to systolic dysfunction. (United States)

    Mahé, Isabelle; Chassany, Olivier; Grenard, Anne-Sophie; Caulin, Charles; Bergmann, Jean-François


    Calcium channel antagonists (CCAs) may either be divided into the dihydropyridines (e.g. amlodipine, felodipine, isradipine, lacidipine, nilvadipine, nifedipine, nicardipine etc.), the phenylalkylamines (e.g. verapamil) and the benzothiazepines (e.g. diltiazem) according to their chemical structure, or into first generation agents (nifedipine, verapamil and diltiazem) and second generation agents (subsequently developed dihydropyridine-derivatives). Second generation CCAs are characterized by greater selectivity for calcium channels in vascular smooth muscle cells than the myocardium, a longer duration of action and a small trough-to-peak variation in plasma concentrations. Heart failure is characterized by decreased cardiac output resulting in inadequate oxygen delivery to peripheral tissues. Although the accompanying neurohormonal activation, leading to vasoconstriction and increased blood pressure, is initially beneficial in increasing tissue perfusion, prolonged activation is detrimental because it increases afterload and further reduces cardiac output. At the level of the myocyte, heart failure is associated with increased intracellular calcium levels which are thought to impair diastolic function. These changes indicate that the CCAs would be beneficial in patients with heart failure. There has been a strong interest and increasing experience in the use of CCAs in patients with heart failure. Despite potential beneficial effects in initial small trials, findings from larger trials suggest that CCA may have detrimental effects upon survival and cardiovascular events. However, this may not necessarily be a 'class b' effect of the CCAs as there is considerable heterogeneity in the chemical structure of individual agents. Clinical experience with different CCAs in patients with heart failure includes trials that evaluated their effects on hemodynamic parameters, exercise tolerance and on symptomatology. However, the most relevant results are those from randomized

  13. BIN1 localizes the L-type calcium channel to cardiac T-tubules.

    Directory of Open Access Journals (Sweden)

    Ting-Ting Hong


    Full Text Available The BAR domain protein superfamily is involved in membrane invagination and endocytosis, but its role in organizing membrane proteins has not been explored. In particular, the membrane scaffolding protein BIN1 functions to initiate T-tubule genesis in skeletal muscle cells. Constitutive knockdown of BIN1 in mice is perinatal lethal, which is associated with an induced dilated hypertrophic cardiomyopathy. However, the functional role of BIN1 in cardiomyocytes is not known. An important function of cardiac T-tubules is to allow L-type calcium channels (Cav1.2 to be in close proximity to sarcoplasmic reticulum-based ryanodine receptors to initiate the intracellular calcium transient. Efficient excitation-contraction (EC coupling and normal cardiac contractility depend upon Cav1.2 localization to T-tubules. We hypothesized that BIN1 not only exists at cardiac T-tubules, but it also localizes Cav1.2 to these membrane structures. We report that BIN1 localizes to cardiac T-tubules and clusters there with Cav1.2. Studies involve freshly acquired human and mouse adult cardiomyocytes using complementary immunocytochemistry, electron microscopy with dual immunogold labeling, and co-immunoprecipitation. Furthermore, we use surface biotinylation and live cell confocal and total internal fluorescence microscopy imaging in cardiomyocytes and cell lines to explore delivery of Cav1.2 to BIN1 structures. We find visually and quantitatively that dynamic microtubules are tethered to membrane scaffolded by BIN1, allowing targeted delivery of Cav1.2 from the microtubules to the associated membrane. Since Cav1.2 delivery to BIN1 occurs in reductionist non-myocyte cell lines, we find that other myocyte-specific structures are not essential and there is an intrinsic relationship between microtubule-based Cav1.2 delivery and its BIN1 scaffold. In differentiated mouse cardiomyocytes, knockdown of BIN1 reduces surface Cav1.2 and delays development of the calcium transient

  14. Calcium-dependent modulation and plasma membrane targeting of the AKT2 potassium channel by the CBL4/ CIPK6 calcium sensor/protein kinase complex

    Institute of Scientific and Technical Information of China (English)

    Katrin Held; Jean-Baptiste Thibaud; J(o)rg Kudla; Francois Pascaud; Christian Eckert; Pawel Gajdanowicz; Kenji Hashimoto; Claire Corratgé-Faillie; Jan Niklas Offenborn; Beno(i)t Lacombe; Ingo Dreyer


    Potassium (K+) channel function is fundamental to many physiological processes. However, components and mechanisms regulating the activity of plant K+ channels remain poorly understood. Here, we show that the calcium (Ca2+)sensor CBL4 together with the interacting protein kinase CIPK6 modulates the activity and plasma membrane (PM)targeting of the K+ channel AKT2 from Arabidopsis thaliana by mediating translocation of AKT2 to the PM in plant cells and enhancing AKT2 activity in oocytes. Accordingly, akt2, cbl4 and cipk6 mutants share similar developmental and delayed flowering pheuotypes. Moreover, the isolated regulatory C-terminal domain of CIPK6 is sufficient for mediating CBL4- and Ca2+-dependent channel translocation from the endoplasmic reticulum membrane to the PM by a novel targeting pathway that is dependent on dual lipid modifications of CBL4 by myristoylation and palmitoylation. Thus, we describe a critical mechanism of ion-channel regulation where a Ca2+ sensor modulates K+ channel activity by promoting a kinase interaction-dependent but phosphorylation-independent translocation of the channel to the PM.

  15. Calcium-activated chloride channel TMEM16A modulates mucin secretion and airway smooth muscle contraction (United States)

    Huang, Fen; Zhang, Hongkang; Wu, Meng; Yang, Huanghe; Kudo, Makoto; Peters, Christian J.; Woodruff, Prescott G.; Solberg, Owen D.; Donne, Matthew L.; Huang, Xiaozhu; Sheppard, Dean; Fahy, John V.; Wolters, Paul J.; Hogan, Brigid L. M.; Finkbeiner, Walter E.; Li, Min; Jan, Yuh-Nung; Jan, Lily Yeh; Rock, Jason R.


    Mucous cell hyperplasia and airway smooth muscle (ASM) hyperresponsiveness are hallmark features of inflammatory airway diseases, including asthma. Here, we show that the recently identified calcium-activated chloride channel (CaCC) TMEM16A is expressed in the adult airway surface epithelium and ASM. The epithelial expression is increased in asthmatics, particularly in secretory cells. Based on this and the proposed functions of CaCC, we hypothesized that TMEM16A inhibitors would negatively regulate both epithelial mucin secretion and ASM contraction. We used a high-throughput screen to identify small-molecule blockers of TMEM16A-CaCC channels. We show that inhibition of TMEM16A-CaCC significantly impairs mucus secretion in primary human airway surface epithelial cells. Furthermore, inhibition of TMEM16A-CaCC significantly reduces mouse and human ASM contraction in response to cholinergic agonists. TMEM16A-CaCC blockers, including those identified here, may positively impact multiple causes of asthma symptoms. PMID:22988107

  16. Establishing homology between mitochondrial calcium uniporters, prokaryotic magnesium channels and chlamydial IncA proteins. (United States)

    Lee, Andre; Vastermark, Ake; Saier, Milton H


    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.

  17. BARP suppresses voltage-gated calcium channel activity and Ca2+-evoked exocytosis. (United States)

    Béguin, Pascal; Nagashima, Kazuaki; Mahalakshmi, Ramasubbu N; Vigot, Réjan; Matsunaga, Atsuko; Miki, Takafumi; Ng, Mei Yong; Ng, Yu Jin Alvin; Lim, Chiaw Hwee; Tay, Hock Soon; Hwang, Le-Ann; Firsov, Dmitri; Tang, Bor Luen; Inagaki, Nobuya; Mori, Yasuo; Seino, Susumu; Launey, Thomas; Hunziker, Walter


    Voltage-gated calcium channels (VGCCs) are key regulators of cell signaling and Ca(2+)-dependent release of neurotransmitters and hormones. Understanding the mechanisms that inactivate VGCCs to prevent intracellular Ca(2+) overload and govern their specific subcellular localization is of critical importance. We report the identification and functional characterization of VGCC β-anchoring and -regulatory protein (BARP), a previously uncharacterized integral membrane glycoprotein expressed in neuroendocrine cells and neurons. BARP interacts via two cytosolic domains (I and II) with all Cavβ subunit isoforms, affecting their subcellular localization and suppressing VGCC activity. Domain I interacts at the α1 interaction domain-binding pocket in Cavβ and interferes with the association between Cavβ and Cavα1. In the absence of domain I binding, BARP can form a ternary complex with Cavα1 and Cavβ via domain II. BARP does not affect cell surface expression of Cavα1 but inhibits Ca(2+) channel activity at the plasma membrane, resulting in the inhibition of Ca(2+)-evoked exocytosis. Thus, BARP can modulate the localization of Cavβ and its association with the Cavα1 subunit to negatively regulate VGCC activity.

  18. Inhibition of nitrite-induced toxicity in channel catfish by calcium chloride and sodium chloride (United States)

    Tommasso J.R., Wright; Simco, B.A.; Davis, K.B.


    Environmental chloride has been shown to inhibit methemoglobin formation in fish, thereby offering a protective effect against nitrite toxicity. Channel catfish (Ictalurus punctatus) were simultaneously exposed to various environmental nitrite and chloride levels (as either CaCl2 or NaCl) in dechlorinated tap water (40 mg/L total hardness, 47 mg/L alkalinity, 4 mg/L chloride, pH = 6.9-7.1, and temperature 21-24°C). Methemoglobin levels in fish simultaneously exposed to 2.5 mg/L nitrite and up to 30 mg/L chloride as either CaCl2 or NaCl were similar but significantly lower than in unprotected fish. Exposure to 10 mg/L nitrite and 60 mg/L chloride resulted in methemoglobin levels similar to those of the controls; most unprotected fish died. Fish exposed to 10 mg/L nitrite had significantly lower methemoglobin levels when protected with 15.0 mg/L chloride as CaCl2 than with NaCl. Fish exposed to nitrite in the presence of 60 mg/L chloride (as either CaCl2 or NaCl) had similar 24-h LC50 values that were significantly elevated above those obtained in the absence of chloride. Calcium had little effect on tolerance to nitrite toxicity in channel catfish in contrast to its large effect reported in steelhead trout (Salmo gairdneri).

  19. Expression and cellular localization of the voltage-gated calcium channel α2δ3 in the rodent retina. (United States)

    Pérez de Sevilla Müller, Luis; Sargoy, Allison; Fernández-Sánchez, Laura; Rodriguez, Allen; Liu, Janelle; Cuenca, Nicolás; Brecha, Nicholas


    High-voltage-activated calcium channels are hetero-oligomeric protein complexes that mediate multiple cellular processes, including the influx of extracellular Ca(2+), neurotransmitter release, gene transcription, and synaptic plasticity. These channels consist of a primary α(1) pore-forming subunit, which is associated with an extracellular α(2)δ subunit and an intracellular β auxiliary subunit, which alter the gating properties and trafficking of the calcium channel. The cellular localization of the α(2)δ(3) subunit in the mouse and rat retina is unknown. In this study using RT-PCR, a single band at ∼ 305 bp corresponding to the predicted size of the α(2)δ(3) subunit fragment was found in mouse and rat retina and brain homogenates. Western blotting of rodent retina and brain homogenates showed a single 123-kDa band. Immunohistochemistry with an affinity-purified antibody to the α(2)δ(3) subunit revealed immunoreactive cell bodies in the ganglion cell layer and inner nuclear layer and immunoreactive processes in the inner plexiform layer and the outer plexiform layer. α(2)δ(3) immunoreactivity was localized to multiple cell types, including ganglion, amacrine, and bipolar cells and photoreceptors, but not horizontal cells. The expression of the α(2)δ(3) calcium channel subunit to multiple cell types suggests that this subunit participates widely in Ca-channel-mediated signaling in the retina.

  20. Effects of inorganic lead on voltage-sensitive calcium channels in N1E-115 neuroblastoma cells. (United States)

    Audesirk, G; Audesirk, T


    N1E-115 mouse neuroblastoma cells have been reported to possess two types of voltage-sensitive calcium channels: Low voltage activated, rapidly inactivating T-type (type I) and high voltage activated, slowly inactivating L-type (type II). We studied the effects of acute in vitro exposure to inorganic lead on these calcium channels, using the whole-cell variant of patch clamping. Using salines with a high lead-buffering capacity, we found that both T-type and L-type channels are reversibly inhibited in a dose-dependent manner at free Pb2+ concentrations ranging from 20 nM to 14 microM. L-type channels are somewhat more sensitive to Pb2+ than T-type channels are (L-type: IC50 approx. 0.7 microM; T-type: IC50 approx. 1.3 microM). Both channels show small but significant inhibition (approx. 10%) at 20 nM free Pb2+. Pb2+ affects neither activation nor inactivation of T-type channels, but enhances inactivation of L-type channels at holding potentials around -60 to -40 mV. A peculiar phenomenon was observed in cells exposed to 2.3 microM free Pb2+. T-type channels were inhibited in all 20 cells studied. In 15 cells, L-type channels were also inhibited, but in the remaining 5 cells, current flow through L-type channels was enhanced by Pb2+ exposure.

  1. 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


    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.

  2. Diseases of the nervous system associated with calcium channelopathies

    NARCIS (Netherlands)

    Todorov, Boyan Bogdanov


    The aim of the studies described in this thesis was to investigate how abnormal CaV2.1 channel function can cause disease, in particular motor coordination dysfunction. The chapters illustrate how various neuronal cell types in the periphery (peripheral nervous system) and the central nervous system

  3. Water Channels Are Involved in Stomatal Oscillations Encoded by Parameter-Specific Cytosolic Calcium Oscillations

    Institute of Scientific and Technical Information of China (English)


    Earlier studies have shown that various stimuli can induce specific cytosolic calcium ([Ca2+]cyt) oscillations in guard cells and various oscillations in stomatal apertures. Exactly how [Ca2+]cyt oscillation signaling functions in stomatal oscillation is not known. In the present study, the epidermis of broad bean (Vicia faba L.)was used and a rapid ion-exchange treatment with two shifting buffers differing in K+ and Ca2+ concentrations was applied. The treatment for five transients at a 10-min transient period induced clear and regular stomatal oscillation. However, for other transient numbers and periods, the treatments induced some irregular oscillations or even no obvious oscillations in stomatal aperture. The results indicate that stomatal oscillation is encoded by parameter-specific [Ca2+]cyt oscillation: the parameters of [Ca2+]cyt oscillation affected the occurrence rate and the parameters of stomatal oscillation. The water channel inhibitor HgCl2 completely inhibited stomatal oscillation and the inhibitory effect could be partially reversed by β-mercaptoethanol (an agent capable of reversing water channel inhibition by HgCl2). Other inhibitory treatments against ion transport (i.e. the application of LaCl3, EGTA, or tetraethylammonium chloride (TEACl))weakly impaired stomatal oscillation when the compounds were added after rapid ion-exchange treatment.If these compounds were added before rapid-ion exchange treatment, the inhibitory effect was much more apparent (except in the case of TEACI). The results of the present study suggest that water channels are involved in stomatal oscillation as a downstream element of [Ca2+]cyt oscillation signaling.

  4. Sustained calcium entry through P2X nucleotide receptor channels in human airway epithelial cells. (United States)

    Zsembery, Akos; Boyce, Amanda T; Liang, Lihua; Peti-Peterdi, János; Bell, P Darwin; Schwiebert, Erik M


    Purinergic receptor stimulation has potential therapeutic effects for cystic fibrosis (CF). Thus, we explored roles for P2Y and P2X receptors in stably increasing [Ca(2+)](i) in human CF (IB3-1) and non-CF (16HBE14o(-)) airway epithelial cells. Cytosolic Ca(2+) was measured by fluorospectrometry using the fluorescent dye Fura-2/AM. Expression of P2X receptor (P2XR) subtypes was assessed by immunoblotting and biotinylation. In IB3-1 cells, ATP and other P2Y agonists caused only a transient increase in [Ca(2+)](i) derived from intracellular stores in a Na(+)-rich environment. In contrast, ATP induced an increase in [Ca(2+)](i) that had transient and sustained components in a Na(+)-free medium; the sustained plateau was potentiated by zinc or increasing extracellular pH. Benzoyl-benzoyl-ATP, a P2XR-selective agonist, increased [Ca(2+)](i) only in Na(+)-free medium, suggesting competition between Na(+) and Ca(2+) through P2XRs. Biochemical evidence showed that the P2X(4) receptor is the major subtype shared by these airway epithelial cells. A role for store-operated Ca(2+) channels, voltage-dependent Ca(2+) channels, or Na(+)/Ca(2+) exchanger in the ATP-induced sustained Ca(2+) signal was ruled out. In conclusion, these data show that epithelial P2X(4) receptors serve as ATP-gated calcium entry channels that induce a sustained increase in [Ca(2+)](i). In airway epithelia, a P2XR-mediated Ca(2+) signal may have therapeutic benefit for CF.

  5. Mapping of dihydropyridine binding residues in a less sensitive invertebrate L-type calcium channel (LCa v 1). (United States)

    Senatore, Adriano; Boone, Adrienne; Lam, Stanley; Dawson, Taylor F; Zhorov, Boris; Spafford, J David


    Invertebrate L-type calcium channel, LCa(v) 1, isolated from the pond snail Lymnaea stagnalis is nearly indistinguishable from mammalian Ca(v) 1.2 (α1C) calcium channel in biophysical characteristics observed in vitro. These L-type channels are likely constrained within a narrow range of biophysical parameters to perform similar functions in the snail and mammalian cardiovascular systems. What distinguishes snail and mammalian L-type channels is a difference in dihydropyridine sensitivity: 100 nM isradipine exhibits a significant block of mammalian Ca(v) 1.2 currents without effect on snail LCa(v)1 currents. The native snail channel serves as a valuable surrogate for validating key residue differences identified from previous experimental and molecular modeling work. As predicted, three residue changes in LCa(v)1 (N_3o18, F_3i10, and I_4i12) replaced with DHP-sensing residues in respective positions of Ca(v) 1.2, (Q_3o18, Y_3i10, and M_4i12) raises the potency of isradipine block of LCa(v)1 channels to that of mammalian Ca(v) 1.2. Interestingly, the single N_3o18_Q mutation in LCa(v) 1 channels lowers DHP sensitivity even further and the triple mutation bearing enhanced isradipine sensitivity, still retains a reduced potency of agonist, (S)-Bay K8644.

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

    Institute of Scientific and Technical Information of China (English)

    Wei Zhang; Liu-Min Fan


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

  7. A cell model study of calcium influx mechanism regulated by calcium-dependent potassium channels in Purkinje cell dendrites. (United States)

    Chono, Koji; Takagi, Hiroshi; Koyama, Shozo; Suzuki, Hideo; Ito, Etsuro


    The present study was designed to elucidate the roles of dendritic voltage-gated K+ channels in Ca2+ influx mechanism of a rat Purkinje cell using a computer simulation program. First, we improved the channel descriptions and the maximum conductance in the Purkinje cell model to mimic both the kinetics of ion channels and the Ca2+ spikes, which had failed in previous studies. Our cell model is, therefore, much more authentic than those in previous studies. Second, synaptic inputs that mimic stimulation of parallel fibers and induce sub-threshold excitability were simultaneously applied to the spiny dendrites. As a result, transient Ca2+ responses were observed in the stimulation points and they decreased with the faster decay rate in the cell model including high-threshold Ca2+-dependent K+ channels than in those excluding these channels. Third, when a single synaptic input was applied into a spiny dendrite, Ca2+-dependent K+ channels suppressed Ca2+ increases at stimulation and recording points. Finally, Ca2+-dependent K+ channels were also found to suppress the time to peak Ca2+ values in the recording points. These results suggest that the opening of Ca2+-dependent K+ channels by Ca2+ influx through voltage-gated Ca2+ channels hyperpolarizes the membrane potentials and deactivates these Ca2+ channels in a negative feedback manner, resulting in local, weak Ca2+ responses in spiny dendrites of Purkinje cells.

  8. Block by a putative antiarrhythmic agent of a calcium-dependent potassium channel in cultured hippocampal neurons. (United States)

    McLarnon, J G


    The actions of a new, putative antiarrhythmic drug, KC-8851 on single channel currents in hippocampal CA1 neurons have been studied. A calcium-dependent potassium current IK(Ca) was activated in the cultured neurons when a solution containing 140 mM K+ and 0.2 mM Ca2+ was applied to inside-out patches. Addition of the compound KC-8851, at concentrations between 1-50 microM, resulted in significant, dose-dependent, decreases in the mean open times of the K channel. The onward (blocking) rate constant was determined from a simple channel blockade scheme and was 5 x 10(7) M-1s-1; this rate constant was not dependent on voltage. Addition of KC-8851 to the solution bath with outside-out patches also caused significant decreases in the mean open times of the IK(Ca) channel consistent with channel blockade by the drug.

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

    Directory of Open Access Journals (Sweden)

    Alexandra F S Breitenkamp

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

  10. L-Type Calcium Channels Do Not Play a Critical Role in Chest Blow Induced Ventricular Fibrillation: Commotio Cordis

    Directory of Open Access Journals (Sweden)

    Christopher Madias


    Full Text Available Background. In a commotio cordis swine model, ventricular fibrillation (VF can be induced by a ball blow to the chest believed secondary to activation of mechanosensitive ion channels. The purpose of the current study is to evaluate whether stretch induced activation of the L-type calcium channel may cause intracellular calcium overload and underlie the VF in commotio cordis. Method and Results. Anesthetized juvenile swine received 6 chest wall strikes with a 17.9 m/s lacrosse ball timed to the vulnerable period for VF induction. Animals were randomized to IV verapamil (n=6 or placebo (n=6. There was no difference in the observed frequency of VF between verapamil (19/26: 73% and placebo (20/36: 56% treated animals (p=0.16. There was also no significant difference in the combined endpoint of VF or nonsustained VF (21/26: 81% in verapamil versus 24/36: 67% in controls, p=0.22. Conclusions. In this experimental model of commotio cordis, verapamil did not prevent VF induction. Thus, in commotio cordis it is unlikely that stretch activation of the L-type calcium channel with resultant intracellular calcium overload plays a prominent role.

  11. Calcium channel antagonist and beta-blocker overdose: antidotes and adjunct therapies. (United States)

    Graudins, Andis; Lee, Hwee Min; Druda, Dino


    Management of cardiovascular instability resulting from calcium channel antagonist (CCB) or beta-adrenergic receptor antagonist (BB) poisoning follows similar principles. Significant myocardial depression, bradycardia and hypotension result in both cases. CCBs can also produce vasodilatory shock. Additionally, CCBs, such as verapamil and diltiazem, are commonly ingested in sustained-release formulations. This can also be the case for some BBs. Peak toxicity can be delayed by several hours. Provision of early gastrointestinal decontamination with activated charcoal and whole-bowel irrigation might mitigate this. Treatment of shock requires a multimodal approach to inotropic therapy that can be guided by echocardiographic or invasive haemodynamic assessment of myocardial function. High-dose insulin euglycaemia is commonly recommended as a first-line treatment in these poisonings, to improve myocardial contractility, and should be instituted early when myocardial dysfunction is suspected. Catecholamine infusions are complementary to this therapy for both inotropic and chronotropic support. Catecholamine vasopressors and vasopressin are used in the treatment of vasodilatory shock. Optimizing serum calcium concentration can confer some benefit to improving myocardial function and vascular tone after CCB poisoning. High-dose glucagon infusions have provided moderate chronotropic and inotropic benefits in BB poisoning. Phosphodiesterase inhibitors and levosimendan have positive inotropic effects but also produce peripheral vasodilation, which can limit blood pressure improvement. In cases of severe cardiogenic shock and/or cardiac arrest post-poisoning, extracorporeal cardiac assist devices have resulted in successful recovery. Other treatments used in refractory hypotension include intravenous lipid emulsion for lipophilic CCB and BB poisoning and methylene blue for refractory vasodilatory shock.

  12. Phorbol Ester Modulation of Ca2+ Channels Mediates Nociceptive Transmission in Dorsal Horn Neurones

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    Gary J. Stephens


    Full Text Available Phorbol esters are analogues of diacylglycerol which activate C1 domain proteins, such as protein kinase C (PKC. Phorbol ester/PKC pathways have been proposed as potential therapeutic targets for chronic pain states, potentially by phosphorylating proteins involved in nociception, such as voltage-dependent Ca2+ channels (VDCCs. In this brief report, we investigate the potential involvement of CaV2 VDCC subtypes in functional effects of the phorbol ester, phorbol 12-myristate 13-acetate (PMA on nociceptive transmission in the spinal cord. Effects of PMA and of selective pharmacological blockers of CaV2 VDCC subtypes on nociceptive transmission at laminae II dorsal horn neurones were examined in mouse spinal cord slices. Experiments were extended to CaV2.3(−/− mice to complement pharmacological studies. PMA increased the mean frequency of spontaneous postsynaptic currents (sPSCs in dorsal horn neurones, without an effect on event amplitude or half-width. sPSC frequency was reduced by selective VDCC blockers, w-agatoxin-IVA (AgTX; CaV2.1, w-conotoxin-GVIA (CTX; CaV2.2 or SNX-482 (CaV2.3. PMA effects were attenuated in the presence of each VDCC blocker and, also, in CaV2.3(−/− mice. These initial data demonstrate that PMA increases nociceptive transmission at dorsal horn neurones via actions on different CaV2 subtypes suggesting potential anti-nociceptive targets in this system.

  13. PIP2 in pancreatic β-cells regulates voltage-gated calcium channels by a voltage-independent pathway. (United States)

    de la Cruz, Lizbeth; Puente, Erika I; Reyes-Vaca, Arturo; Arenas, Isabel; Garduño, Julieta; Bravo-Martínez, Jorge; Garcia, David E


    Phosphatidylinositol-4,5-bisphosphate (PIP2) is a membrane phosphoinositide that regulates the activity of many ion channels. Influx of calcium primarily through voltage-gated calcium (CaV) channels promotes insulin secretion in pancreatic β-cells. However, whether CaV channels are regulated by PIP2, as is the case for some non-insulin-secreting cells, is unknown. The purpose of this study was to investigate whether CaV channels are regulated by PIP2 depletion in pancreatic β-cells through activation of a muscarinic pathway induced by oxotremorine methiodide (Oxo-M). CaV channel currents were recorded by the patch-clamp technique. The CaV current amplitude was reduced by activation of the muscarinic receptor 1 (M1R) in the absence of kinetic changes. The Oxo-M-induced inhibition exhibited the hallmarks of voltage-independent regulation and did not involve PKC activation. A small fraction of the Oxo-M-induced CaV inhibition was diminished by a high concentration of Ca(2+) chelator, whereas ≥50% of this inhibition was prevented by diC8-PIP2 dialysis. Localization of PIP2 in the plasma membrane was examined by transfecting INS-1 cells with PH-PLCδ1, which revealed a close temporal association between PIP2 hydrolysis and CaV channel inhibition. Furthermore, the depletion of PIP2 by a voltage-sensitive phosphatase reduced CaV currents in a way similar to that observed following M1R activation. These results indicate that activation of the M1R pathway inhibits the CaV channel via PIP2 depletion by a Ca(2+)-dependent mechanism in pancreatic β- and INS-1 cells and thereby support the hypothesis that membrane phospholipids regulate ion channel activity by interacting with ion channels.

  14. Raised activity of L-type calcium channels renders neurons prone to form paroxysmal depolarization shifts. (United States)

    Rubi, Lena; Schandl, Ulla; Lagler, Michael; Geier, Petra; Spies, Daniel; Gupta, Kuheli Das; Boehm, Stefan; Kubista, Helmut


    Neuronal L-type voltage-gated calcium channels (LTCCs) are involved in several physiological functions, but increased activity of LTCCs has been linked to pathology. Due to the coupling of LTCC-mediated Ca(2+) influx to Ca(2+)-dependent conductances, such as KCa or non-specific cation channels, LTCCs act as important regulators of neuronal excitability. Augmentation of after-hyperpolarizations may be one mechanism that shows how elevated LTCC activity can lead to neurological malfunctions. However, little is known about other impacts on electrical discharge activity. We used pharmacological up-regulation of LTCCs to address this issue on primary rat hippocampal neurons. Potentiation of LTCCs with Bay K8644 enhanced excitatory postsynaptic potentials to various degrees and eventually resulted in paroxysmal depolarization shifts (PDS). Under conditions of disturbed Ca(2+) homeostasis, PDS were evoked frequently upon LTCC potentiation. Exposing the neurons to oxidative stress using hydrogen peroxide also induced LTCC-dependent PDS. Hence, raising LTCC activity had unidirectional effects on brief electrical signals and increased the likeliness of epileptiform events. However, long-lasting seizure-like activity induced by various pharmacological means was affected by Bay K8644 in a bimodal manner, with increases in one group of neurons and decreases in another group. In each group, isradipine exerted the opposite effect. This suggests that therapeutic reduction in LTCC activity may have little beneficial or even adverse effects on long-lasting abnormal discharge activities. However, our data identify enhanced activity of LTCCs as one precipitating cause of PDS. Because evidence is continuously accumulating that PDS represent important elements in neuropathogenesis, LTCCs may provide valuable targets for neuroprophylactic therapy.

  15. Hypothyroid state reduces calcium channel function in 18-day pregnant rat uterus. (United States)

    Parija, S C; Mishra, S K; Raviprakash, V


    Hypothyroidism significantly reduced the mean amplitude and increased the mean frequency of spontaneous rhythmic contractions in 18 day pregnant rat uterus. Nifedipine (10(-12)-10(-9) M) and diltiazem (10(-10)-10(-6) M) caused concentration related inhibition of the myogenic responses of the uterine strips obtained from both pregnant and hypothyroid state. However, nifedipine was less potent (IC50:2.11 x 10(-11) M) in pregnant hypothyroid state as compared to pregnant control (IC50: 3.1 x 10(-12) M). Similarly, diltiazem was less potent (IC50: 3.72 x 10(-9) M) in inhibiting the uterine spontaneous contractions in hypothyroid than in pregnant rat uterus (IC50:5.37 x 10(-10) M). A similar decrease in the sensitivity to nifedipine and diltiazem for reversal of K+ (100 mM)-induced tonic contraction and K(+)-stimulated 45Ca2+ influx was observed with these calcium channel antagonists in uterus obtained from hypothyroid pregnant rats compared to the controls. Nifedipine-sensitive influx of 45Ca(2+)-stimulated either by K+ (100 mM) or by Bay K8644 (1,4-dihydro-2,6-methyl-5-nitro-4-[2'-(trifluromethyl)phenyl]-3-pyridine carboxylic acid methyl ester) (10(-9) M) was significantly less in uterine strips from hypothyroid rats compared to controls. The results suggest that the inhibition of uterine rhythmic contractions may be attributable to a reduction in rat myometrial Ca2+ channel function in the hypothyroid state.

  16. Ion channels to inactivate neurons in Drosophila

    Directory of Open Access Journals (Sweden)

    James J L Hodge


    Full Text Available Ion channels are the determinants of excitability; therefore, manipulation of their levels and properties provides an opportunity for the investigator to modulate neuronal and circuit function. There are a number of ways to suppress electrical activity in Drosophila neurons, for instance, over-expression of potassium channels (i.e. Shaker Kv1, Shaw Kv3, Kir2.1 and DORK that are open at resting membrane potential. This will result in increased potassium efflux and membrane hyperpolarisation setting resting membrane potential below the threshold required to fire action potentials. Alternatively over-expression of other channels, pumps or co-transporters that result in a hyperpolarised membrane potential will also prevent firing. Lastly, neurons can be inactivated by, disrupting or reducing the level of functional voltage-gated sodium (Nav1 paralytic or calcium (Cav2 cacophony channels that mediate the depolarisation phase of action potentials. Similarly, strategies involving the opposite channel manipulation should allow net depolarisation and hyperexcitation in a given neuron. These changes in ion channel expression can be brought about by the versatile transgenic (i.e. Gal4/UAS based systems available in Drosophila allowing fine temporal and spatial control of (channel transgene expression. These systems are making it possible to electrically inactivate (or hyperexcite any neuron or neural circuit in the fly brain, and much like an exquisite lesion experiment, potentially elucidate whatever interesting behaviour or phenotype each network mediates. These techniques are now being used in Drosophila to reprogram electrical activity of well-defined circuits and bring about robust and easily quantifiable changes in behaviour, allowing different models and hypotheses to be rapidly tested.

  17. The role of calcium, calcium-activated K+ channels, and tyrosine/kinase in psoralen-evoked responses in human melanoma cells

    Directory of Open Access Journals (Sweden)

    Isoldi M.C.


    Full Text Available 8-Methoxy psoralen (8-MOP exerts a short-term (24 h mitogenic action, and a long-term (48-72 h anti-proliferative and melanogenic action on two human melanoma cell lines, SK-Mel 28 and C32TG. An increase of intracellular calcium concentration was observed by spectrofluorometry immediately after the addition of 0.1 mM 8-MOP to both cell lines, previously incubated with calcium probe fluo-3 AM (5 µM. The intracellular Ca2+ chelator BAPTA/AM (1 µM blocked both early (mitogenic and late (anti-proliferative and melanogenic 8-MOP effects on both cell lines, thus revealing the importance of the calcium signal in both short- and long-term 8-MOP-evoked responses. Long-term biological assays with 5 and 10 mM tetraethylammonium chloride (TEA, an inhibitor of Ca2+-dependent K+ channels did not affect the responses to psoralen; however, in 24-h assays 10 mM TEA blocked the proliferative peak, indicating a modulation of Ca2+-dependent K+ channels by 8-MOP. No alteration of cAMP basal levels or forskolin-stimulated cAMP levels was promoted by 8-MOP in SK-Mel 28 cells, as determined by radioimmunoassay. However, in C32TG cells forskolin-stimulated cAMP levels were further increased in the presence of 8-MOP. In addition, assays with 1 µM protein kinase C and calcium/calmodulin-dependent kinase inhibitors, Ro 31-8220 and KN-93, respectively, excluded the participation of these kinases in the responses evoked by 8-MOP. Western blot with antibodies anti-phosphotyrosine indicated a 92% increase of the phosphorylated state of a 43-kDa band, suggesting that the phosphorylation of this protein is a component of the cascade that leads to the increase of tyrosinase activity.

  18. Short-term exposure to L-type calcium channel blocker, verapamil, alters the expression pattern of calcium-binding proteins in the brain of goldfish, Carassius auratus. (United States)

    Palande, Nikhil V; Bhoyar, Rahul C; Biswas, Saikat P; Jadhao, Arun G


    The influx of calcium ions (Ca(2+)) is responsible for various physiological events including neurotransmitter release and synaptic modulation. The L-type voltage dependent calcium channels (L-type VDCCs) transport Ca(2+) across the membrane. Calcium-binding proteins (CaBPs) bind free cytosolic Ca(2+) and prevent excitotoxicity caused by sudden increase in cytoplasmic Ca(2+). The present study was aimed to understand the regulation of expression of neuronal CaBPs, namely, calretinin (CR) and parvalbumin (PV) following blockade of L-type VDCCs in the CNS of Carassius auratus. Verapamil (VRP), a potent L-type VDCC blocker, selectively blocks Ca(2+) entry at the plasma membrane level. VRP present in the aquatic environment at a very low residual concentration has shown ecotoxicological effects on aquatic animals. Following acute exposure for 96h, median lethal concentration (LC50) for VRP was found to be 1.22mg/L for goldfish. At various doses of VRP, the behavioral alterations were observed in the form of respiratory difficulty and loss of body balance confirming the cardiovascular toxicity caused by VRP at higher doses. In addition to affecting the cardiovascular system, VRP also showed effects on the nervous system in the form of altered expression of PV. When compared with controls, the pattern of CR expression did not show any variations, while PV expression showed significant alterations in few neuronal populations such as the pretectal nucleus, inferior lobes, and the rostral corpus cerebellum. Our result suggests possible regulatory effect of calcium channel blockers on the expression of PV.

  19. Degradation kinetics and pathways of three calcium channel blockers under UV irradiation. (United States)

    Zhu, Bing; Zonja, Bozo; Gonzalez, Oscar; Sans, Carme; Pérez, Sandra; Barceló, Damia; Esplugas, Santiago; Xu, Ke; Qiang, Zhimin


    Calcium channel blockers (CCBs) are a group of pharmaceuticals widely prescribed to lower blood pressure and treat heart diseases. They have been frequently detected in wastewater treatment plant (WWTP) effluents and downstream river waters, thus inducing a potential risk to aquatic ecosystems. However, little is known about the behavior and fate of CCBs under UV irradiation, which has been adopted as a primary disinfection method for WWTP effluents. This study investigated the degradation kinetics and pathways of three commonly-used CCBs, including amlodipine (AML), diltiazem (DIL), and verapamil (VER), under UV (254 nm) irradiation. The chemical structures of transformation byproducts (TBPs) were first identified to assess the potential ecological hazards. On that basis, a generic solid-phase extraction method, which simultaneously used four different cartridges, was adopted to extract and enrich the TBPs. Thereafter, the photo-degradation of target CCBs was performed under UV fluences typical for WWTP effluent disinfection. The degradation of all three CCBs conformed to the pseudo-first-order kinetics, with rate constants of 0.031, 0.044 and 0.011 min(-1) for AML, DIL and VER, respectively. By comparing the MS(2) fragments and the evolution (i.e., formation or decay) trends of identified TBPs, the degradation pathways were proposed. In the WWTP effluent, although the target CCBs could be degraded, several TBPs still contained the functional pharmacophores and reached peak concentrations under UV fluences of 40-100 mJ cm(-2).

  20. Efficacy and safety of calcium channel blockers in hypertensive patients with concomitant left ventricular dysfunction. (United States)

    Parmley, W W


    The use of calcium channel blockers (CCBs) in the treatment of hypertension and concomitant left ventricular dysfunction is reviewed. Some CCBs, particularly second-generation dihydropyridine agents such as felodipine, isradipine, nicardipine, nimodipine, and nitrendipine, have properties that enhance their usefulness in these patients. All CCBs have a similar mechanism of action. Differences in their selective action at various tissue sites determine which are most appropriate for patients with concomitant hypertension and left ventricular dysfunction. Most CCBs do not produce reflex stimulation of the heart or induce intravascular expansion. While all CCBs produce arteriolar dilation, all local beds and regional circulations in target organs are not affected equally. Most CCBs can decrease cardiac mass, and second-generation CCBs tend to have little or no negative inotropic effects at therapeutic dosages. In addition, they increase blood flow and reduce myocardial oxygen requirements. Because of differences in functional and electrophysiologic effects, specific CCBs may not be appropriate for all patients. Since second-generation dihydropyridine CCBs lack clinically relevant negative inotropic effects, and have been shown to improve exercise tolerance and coronary artery perfusion, they are appropriate for hypertensive patients with left ventricular dysfunction, angina, and coronary heart disease. Second-generation CCBs tend to lack cardiodepressant side effects and are less likely to react with digoxin than are first-generation CCBs.

  1. Eugenol dilates rat cerebral arteries by inhibiting smooth muscle cell voltage-dependent calcium channels. (United States)

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


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

  2. The Low-Threshold Calcium Channel Cav3.2 Determines Low-Threshold Mechanoreceptor Function

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    Amaury François


    Full Text Available The T-type calcium channel Cav3.2 emerges as a key regulator of sensory functions, but its expression pattern within primary afferent neurons and its contribution to modality-specific signaling remain obscure. Here, we elucidate this issue using a unique knockin/flox mouse strain wherein Cav3.2 is replaced by a functional Cav3.2-surface-ecliptic GFP fusion. We demonstrate that Cav3.2 is a selective marker of two major low-threshold mechanoreceptors (LTMRs, Aδ- and C-LTMRs, innervating the most abundant skin hair follicles. The presence of Cav3.2 along LTMR-fiber trajectories is consistent with critical roles at multiple sites, setting their strong excitability. Strikingly, the C-LTMR-specific knockout uncovers that Cav3.2 regulates light-touch perception and noxious mechanical cold and chemical sensations and is essential to build up that debilitates allodynic symptoms of neuropathic pain, a mechanism thought to be entirely A-LTMR specific. Collectively, our findings support a fundamental role for Cav3.2 in touch/pain pathophysiology, validating their critic pharmacological relevance to relieve mechanical and cold allodynia.

  3. S-acylation dependent post-translational cross-talk regulates large conductance calcium- and voltage- activated potassium (BK channels

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    Michael J Shipston


    Full Text Available Mechanisms that control surface expression and/or activity of large conductance calcium-activated potassium (BK channels are important determinants of their (pathophysiological function. Indeed, BK channel dysfunction is associated with major human disorders ranging from epilepsy to hypertension and obesity. S-acylation (S-palmitoylation represents a major reversible, post-translational modification controlling the properties and function of many proteins including ion channels. Recent evidence reveals that both pore-forming and regulatory subunits of BK channels are S-acylated and control channel trafficking and regulation by AGC-family protein kinases. The pore-forming α-subunit is S-acylated at two distinct sites within the N- and C-terminus, each site being regulated by different palmitoyl acyl transferases (zDHHCs and acyl thioesterases. (APTs. S-acylation of the N-terminus controls channel trafficking and surface expression whereas S-acylation of the C-terminal domain determines regulation of channel activity by AGC-family protein kinases. S-acylation of the regulatory β4-subunit controls ER exit and surface expression of BK channels but does not affect ion channel kinetics at the plasma membrane. Furthermore, a significant number of previously identified BK-channel interacting proteins have been shown, or are predicted to be, S-acylated. Thus, the BK channel multi-molecular signalling complex may be dynamically regulated by this fundamental post-translational modification and thus S-acylation likely represents an important determinant of BK channel physiology in health and disease.

  4. TRPV4 and AQP4 Channels Synergistically Regulate Cell Volume and Calcium Homeostasis in Retinal Müller Glia

    DEFF Research Database (Denmark)

    Jo, Andrew O; Ryskamp, Daniel A; Phuong, Tam T T


    and facilitates the time course and amplitude of hypotonicity-induced swelling and regulatory volume decrease. We confirm the crucial facets of the signaling mechanism in heterologously expressing oocytes. These results identify the molecular mechanism that contributes to dynamic regulation of glial volume...... through TRPV4 channels reciprocally modulates volume regulation, swelling, and Aqp4 gene expression. Therefore, TRPV4-AQP4 interactions constitute a molecular system that fine-tunes astroglial volume regulation by integrating osmosensing, calcium signaling, and water transport and, when overactivated...... set of mechanisms involving reciprocal interactions at the level of glial gene expression, calcium homeostasis, swelling, and volume regulation. Specifically, water influx through AQP4 drives calcium influx via TRPV4 in the glial end foot, which regulates expression of Aqp4 and Kir4.1 genes...

  5. 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


    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...... immunogold labelling using a monoclonal antibody specific for phenylalkylamine and dihydropyridine sensitive Ca2+ channels. Using the same technique and a monoclonal antibody (bd-17) to the beta 2/beta 3-subunit of the GABAA-receptor, double labelling of Ca2+ channels and GABAA-receptors with gold particles...... of THIP-treated cultures. This suggests that primarily low affinity GABAA-receptors are closely associated with Ca2+ channels and this may be important for the ability of these receptors to mediate an inhibitory action on transmitter release even under extreme depolarizing conditions....

  6. Calcium-dependent expression of transient receptor potential canonical type 3 channels in patients with chronic kidney disease

    DEFF Research Database (Denmark)

    Liu, Ying; Krueger, Katharina; Hovsepian, Anahit;


    It is unknown whether extracellular calcium may regulate the expression of transient receptor potential canonical type 3 (TRPC3) channels in patients with chronic kidney disease. Using quantitative in-cell Western assay we compared the expression of TRPC3 channel protein in monocytes from 20...... patients with chronic kidney disease and 19 age- and sex-matched healthy control subjects. TRPC3 channels were identified by immunoblotting using specific antibodies and TRPC3 protein was further confirmed by mass spectrometry. We observed a significant increase of TRPC3 channel protein expression...... in patients with chronic kidney disease compared to healthy control subjects (normalized expression, 0.42±0.06 vs. 0.19±0.03; p...

  7. Calcium Channel Blockade and Peroxisome Proliferator Activated Receptor γ Agonism Diminish Cognitive Loss and Preserve Endothelial Function During Diabetes Mellitus. (United States)

    Jain, Swati; Sharma, B M; Sharma, Bhupesh


    Diabetes mellitus is considered as a main risk factor for vascular dementia. In the past, we have reported the induction of vascular dementia (VaD) by experimental diabetes. This study investigates the efficacy of a nifedipine, a calcium channel blocker and pioglitazone in the pharmacological interdiction of pancreatectomy diabetes (PaD) induced vascular endothelial dysfunction and subsequent VaD in rats. Attentional set shifting (ASST) and Morris water-maze (MWM) test were used for assessment of learning and memory. Vascular endothelial function, blood brain barrier permeability, serum glucose, serum nitrite/nitrate, oxidative stress (viz. aortic superoxide anion, brain thiobarbituric acid reactive species and brain glutathione), brain calcium and inflammation (myeloperoxidase) were also estimated. PaD rats have shown impairment of endothelial function, blood brain barrier permeability, learning and memory along with an increase in brain inflammation, oxidative stress and calcium. Administration of nifedipine and pioglitazone significantly attenuated PaD induced impairment of learning, memory, blood brain barrier permeability, endothelial function and biochemical parameters. It may be concluded that nifedipine, a calcium channel blocker may be considered as a potent pharmacological agent for the management of PaD induced endothelial dysfunction and subsequent VaD.

  8. Neuroprotective effect of gadolinium: a stretch-activated calcium channel blocker in mouse model of ischemia-reperfusion injury. (United States)

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


    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.

  9. Synthesis and Calcium Channel Blocking Activity of 1, 4-Dihydropyridine Derivatives Containing Ester Substitute and Phenyl Carbamoyl Group

    Directory of Open Access Journals (Sweden)

    Bassem Sadek


    Full Text Available Problem statement: Several studies on the synthesis of new nifedipine analogs have been carried out, but the literature reveled that no study on the synthesis and calcium channel blocking activity of the substituted ester with an amide (5-phenylcarbamoyl moiety has been reported. Approach: Six new derivatives of m-nifedipine have been successfully synthesized by substituting an ester moiety with an amide (5-phenylcarbamoyl moiety, using a modified Hantzsch reactions and tested for their pharmacological activities. The nifedipine analogs 1-6 were characterized and confirmed using elemental analysis, Infrared spectroscopy (IR, Nuclear Magnetic Resonance (1H NMR and Mass spectroscopy. The purity of the compounds was ascertained by melting point and TLC. The in vitro calcium channel blocking activities were evaluated using the high K+ concentration of Porcine Coronary Artery Smooth Muscles (PCASM assay. Results: The compounds (1-2 failed to exhibit any blocking activity (IC50 = 10−7 to 10−5 M range, while the compounds 3-6 relaxed precontracted porcine coronary artery smooth muscles with pEC50 values ranging between 4.37±0.10 (compound 3 and 6.46±0.07 (compound 5, indicating that compounds 3-6 exhibit comparable potencies in blocking calcium channels to reference drug varapamil (6.97±0.15 and m-nifedipine (6.48±0.05. Conclusion: The results of this study showed that some of the developed new compounds possess maximal calcium channel blocking effects comparable to m-nifedipine. The developed compounds in the present study will predicatively show an increased metabolic stability and consequently longer duration of actions compared to m-nifedipine and could be, therefore, suitable candidates for further optimization to be evaluated as a new class of antihypertensive drugs.

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

    Directory of Open Access Journals (Sweden)

    Alan eNeely


    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.

  11. Synthesis and Effects of Novel Dihydropyridines as Dual Calcium Channel Blocker and Angiotensin Antagonist on Isolated Rat Aorta

    Directory of Open Access Journals (Sweden)

    Farzin Hadizadeh


    Full Text Available Four novel losartan analogues 5a-d were synthesized by connecting a dihydropyridine nucleus to imidazole ring. The effects of 5a and 5b on angiotensin receptors (AT1 and L-type calcium channels were investigated on isolated rat aorta. Materials and MethodsAortic rings were pre-contracted with 1 µM Angiotensin II or 80 mM KCl and relaxant effects of losartan, nifedipine, 5a and 5b were evaluated by cumulative addition of these drugs to the bath solution.ResultsThe results showed that compounds 5a and 5b have both L-type calcium channel and AT1 receptor blocking activity. Their effects on AT1 receptors are 1000 and 100,000 times more than losartan respectively. The activity of compound 5b on L-type calcium channel is significantly less than nifedipine but compound 5a has comparable effect with nifedipine. ConclusionFinally we concluded that these two new Compounds can be potential candidates to be used as effective antihypertensive agents.

  12. Effects of calcium channel antagonists on the induction of nitric oxide synthase in cultured cells by immunostimulants. (United States)

    Hattori, Y; Kasai, K; So, S; Hattori, S; Banba, N; Shimoda, S


    We investigated whether calcium channel antagonists would alter the induction of nitric oxide (NO) synthesis by bacterial lipopolysaccharide (LPS) alone or in combination with interferon-gamma (IFN gamma) in cultured J774 macrophages, rat vascular smooth muscle cells, rat renal mesangial cells, and rat cardiac myocytes. The induction of NO synthesis was determined by measuring nitrite, the stable end-product. The dihydropyridine calcium channel antagonists, nifedipine, manidipine, nitrendipine, benidipine, barnidipine, perdipine, and nilvadipine all reduced the LPS-induced nitrite production in a dose-dependent manner, each with a differing half-maximal inhibitory concentration, in cultured J774 macrophages. Nifedipine also inhibited nitrite production in vascular smooth muscle cells, mesangial cells, and cardiac myocytes. The half-maximal inhibitory concentrations of nifedipine were ranked as follows: smooth muscle cells < mesangial cells < cardiac myocytes. Diltiazem, at nontoxic concentrations, had no effect on the nitrite formation in the three cell types. Verapamil markedly increased the formation of nitrite in cardiac myocytes in response to LPS and IFN gamma, but not in vascular smooth muscle or mesangial cells. Exposure of cardiac myocytes to LPS and IFN gamma caused the expression of NO synthase mRNA that was significantly increased by verapamil. Thus, certain calcium channel antagonists modulate NO synthesis by altering the induction of NO synthase.

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

    Directory of Open Access Journals (Sweden)

    Andreas Totzeck


    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.

  14. Dopamine midbrain neurons in health and Parkinson's disease: emerging roles of voltage-gated calcium channels and ATP-sensitive potassium channels. (United States)

    Dragicevic, E; Schiemann, J; Liss, B


    Dopamine (DA) releasing midbrain neurons are essential for multiple brain functions, such as voluntary movement, working memory, emotion and cognition. DA midbrain neurons within the substantia nigra (SN) and the ventral tegmental area (VTA) exhibit a variety of distinct axonal projections and cellular properties, and are differentially affected in diseases like schizophrenia, attention deficit hyperactivity disorder, and Parkinson's disease (PD). Apart from having diverse functions in health and disease states, DA midbrain neurons display distinct electrical activity patterns, crucial for DA release. These activity patterns are generated and modulated by specific sets of ion channels. Recently, two ion channels have been identified, not only contributing to these activity patterns and to functional properties of DA midbrain neurons, but also seem to render SN DA neurons particularly vulnerable to degeneration in PD and its animal models: L-type calcium channels (LTCCs) and ATP-sensitive potassium channels (K-ATPs). In this review, we focus on the emerging physiological and pathophysiological roles of these two ion channels (and their complex interplay with other ion channels), particularly in highly vulnerable SN DA neurons, as selective degeneration of these neurons causes the major motor symptoms of PD.

  15. Amino acid substitutions in the FXYD motif enhance phospholemman-induced modulation of cardiac L-type calcium channels. (United States)

    Guo, Kai; Wang, Xianming; Gao, Guofeng; Huang, Congxin; Elmslie, Keith S; Peterson, Blaise Z


    We have found that phospholemman (PLM) associates with and modulates the gating of cardiac L-type calcium channels (Wang et al., Biophys J 98: 1149-1159, 2010). The short 17 amino acid extracellular NH(2)-terminal domain of PLM contains a highly conserved PFTYD sequence that defines it as a member of the FXYD family of ion transport regulators. Although we have learned a great deal about PLM-dependent changes in calcium channel gating, little is known regarding the molecular mechanisms underlying the observed changes. Therefore, we investigated the role of the PFTYD segment in the modulation of cardiac calcium channels by individually replacing Pro-8, Phe-9, Thr-10, Tyr-11, and Asp-12 with alanine (P8A, F9A, T10A, Y11A, D12A). In addition, Asp-12 was changed to lysine (D12K) and cysteine (D12C). As expected, wild-type PLM significantly slows channel activation and deactivation and enhances voltage-dependent inactivation (VDI). We were surprised to find that amino acid substitutions at Thr-10 and Asp-12 significantly enhanced the ability of PLM to modulate Ca(V)1.2 gating. T10A exhibited a twofold enhancement of PLM-induced slowing of activation, whereas D12K and D12C dramatically enhanced PLM-induced increase of VDI. The PLM-induced slowing of channel closing was abrogated by D12A and D12C, whereas D12K and T10A failed to impact this effect. These studies demonstrate that the PFXYD motif is not necessary for the association of PLM with Ca(V)1.2. Instead, since altering the chemical and/or physical properties of the PFXYD segment alters the relative magnitudes of opposing PLM-induced effects on Ca(V)1.2 channel gating, PLM appears to play an important role in fine tuning the gating kinetics of cardiac calcium channels and likely plays an important role in shaping the cardiac action potential and regulating Ca(2+) dynamics in the heart.

  16. Voltage-gated potassium channel Kvl.3 in rabbit ciliary epithelium regulates the membrane potential via coupling intracellular calcium

    Institute of Scientific and Technical Information of China (English)

    LI Yan-feng; ZHUO Ye-hong; BI Wei-na; BAI Yu-jing; LI Yan-na; WANG Zhi-jian


    Background The cell layer of the ciliary epithelium is responsible for aqueous humor secretion and maintenance.Ion channels play an important role in these processes.The main aim of this study was to determine whether the well-characterized members of the Kvl family (Kv1.3) contribute to the Kv currents in ciliary epithelium.Methods New Zealand White rabbits were maintained in a 12 hours light/dark cycle.Ciliary epithelium samples were isolated from the rabbits.We used Western blotting and immunocytochemistry to identify the expression and location of a voltage-gated potassium channel Kvl.3 in ciliary body epithelium.Membrane potential change after adding of Kv1.3 inhibitor margatoxin (MgTX) was observed with a fluorescence method.Results Western blotting and immunocytochemical studies showed that the Kv1.3 protein expressed in pigment ciliary epithelium and nonpigment ciliary epithelium,however it seemed to express more in the apical membrane of the nonpigmented epithelial cells.One nmol/L margatoxin,a specific inhibitor of Kv1.3 channels caused depolarization of the cultured nonpigmented epithelium (NPE) membrane potential.The cytosotic calcium increased after NPE cell depolarization,this increase of cytosolic calcium was partially blocked by 12.5 μmol/L dantrolene and 10 μmol/L nifedipine.These observations suggest that Kv1.3 channels modulate ciliary epithelium potential and effect calcium dependent mechanisms.Conclusion Kv1.3 channels contribute to K+ efflux at the membrane of rabbit ciliary epithelium.

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

    DEFF Research Database (Denmark)

    Hansen, Pernille B L


    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...... to the conclusion that Cav3.1 and Cav3.2 channels have important, but different, functions in mice. T-type Cav3.1 channels affect heart rate, whereas Cav3.2 channels are involved in cardiac hypertrophy. In the vascular system, Cav3.2 activation leads to dilation of blood vessels, whereas Cav3.1 channels are mainly.......2, are expressed in blood vessels, the kidney, and the heart. Studies with gene-deficient mice have provided a way to investigate the Cav3.1 and Cav3.2 channels and their role in the cardiovascular system. This review discusses the results from these knockout mice. Evaluation of the literature leads...

  18. Calcium Transients in Dendrites of Neocortical Neurons Evoked by Single Subthreshold Excitatory Postsynaptic Potentials via Low-Voltage-Activated Calcium Channels (United States)

    Markram, Henry; Sakmann, Bert


    Simultaneous recordings of membrane voltage and concentration of intracellular Ca2+ ([Ca2+]_i) were made in apical dendrites of layer 5 pyramidal cells of rat neocortex after filling dendrites with the fluorescent Ca2+ indicator Calcium Green-1. Subthreshold excitatory postsynaptic potentials (EPSPs), mediated by the activation of glutamate receptor channels, caused a brief increase in dendritic [Ca2+]_i. This rise in dendritic [Ca2+]_i was mediated by the opening of low-voltage-activated Ca2+ channels in the dendritic membrane. The results provide direct evidence that dendrites do not function as passive cables even at low-frequency synaptic activity; rather, a single subthreshold EPSP changes the dendritic membrane conductance by opening Ca2+ channels and generating a [Ca2+]_i transient that may propagate towards the soma. The activation of these Ca2+ channels at a low-voltage threshold is likely to influence the way in which dendritic EPSPs contribute to the electrical activity of the neuron.

  19. T-type calcium channels promote predictive homeostasis of input-output relations in thalamocortical neurons of lateral geniculate nucleus. (United States)

    Hong, Su Z; Kim, Haram R; Fiorillo, Christopher D


    A general theory views the function of all neurons as prediction, and one component of this theory is that of "predictive homeostasis" or "prediction error." It is well established that sensory systems adapt so that neuronal output maintains sensitivity to sensory input, in accord with information theory. Predictive homeostasis applies the same principle at the cellular level, where the challenge is to maintain membrane excitability at the optimal homeostatic level so that spike generation is maximally sensitive to small gradations in synaptic drive. Negative feedback is a hallmark of homeostatic mechanisms, as exemplified by depolarization-activated potassium channels. In contrast, T-type calcium channels exhibit positive feedback that appears at odds with the theory. In thalamocortical neurons of lateral geniculate nucleus (LGN), T-type channels are capable of causing bursts of spikes with an all-or-none character in response to excitation from a hyperpolarized potential. This "burst mode" would partially uncouple visual input from spike output and reduce the information spikes convey about gradations in visual input. However, past observations of T-type-driven bursts may have resulted from unnaturally high membrane excitability. Here we have mimicked within rat brain slices the patterns of synaptic conductance that occur naturally during vision. In support of the theory of predictive homeostasis, we found that T-type channels restored excitability toward its homeostatic level during periods of hyperpolarization. Thus, activation of T-type channels allowed two retinal input spikes to cause one output spike on average, and we observed almost no instances in which output count exceeded input count (a "burst"). T-type calcium channels therefore help to maintain a single optimal mode of transmission rather than creating a second mode. More fundamentally our results support the general theory, which seeks to predict the properties of a neuron's ion channels and

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


    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.

  1. CNTF-Treated Astrocyte Conditioned Medium Enhances Large-Conductance Calcium-Activated Potassium Channel Activity in Rat Cortical Neurons. (United States)

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


    Seizure activity is linked to astrocyte activation as well as dysfunctional cortical neuron excitability produced from changes in calcium-activated potassium (KCa) channel function. Ciliary neurotrophic factor-treated astrocyte conditioned medium (CNTF-ACM) can be used to investigate the peripheral effects of activated astrocytes upon cortical neurons. However, CNTF-ACM's effect upon KCa channel activity in cultured cortical neurons has not yet been investigated. Whole-cell patch clamp recordings were performed in rat cortical neurons to evaluate CNTF-ACM's effects upon charybdotoxin-sensitive large-conductance KCa (BK) channel currents and apamin-sensitive small-conductance KCa (SK) channel current. Biotinylation and RT-PCR were applied to assess CNTF-ACM's effects upon the protein and mRNA expression, respectively, of the SK channel subunits SK2 and SK3 and the BK channel subunits BKα1 and BKβ3. An anti-fibroblast growth factor-2 (FGF-2) monoclonal neutralizing antibody was used to assess the effects of the FGF-2 component of CNTF-ACM. CNTF-ACM significantly increased KCa channel current density, which was predominantly attributable to gains in BK channel activity (p ACM produced a significant increase in BKα1 and BKβ3 expression (p  0.05). Blocking FGF-2 produced significant reductions in KCa channel current density (p > 0.05) as well as BKα1 and BKβ3 expression in CNTF-ACM-treated neurons (p > 0.05). CNTF-ACM significantly enhances BK channel activity in rat cortical neurons and that FGF-2 is partially responsible for these effects. CNTF-induced astrocyte activation results in secretion of neuroactive factors which may affect neuronal excitability and resultant seizure activity in mammalian cortical neurons.

  2. Potential L-Type Voltage-Operated Calcium Channel Blocking Effect of Drotaverine on Functional Models. (United States)

    Patai, Zoltán; Guttman, András; Mikus, Endre G


    Drotaverine is considered an inhibitor of cyclic-3',5'-nucleotide-phophodiesterase (PDE) enzymes; however, published receptor binding data also support the potential L-type voltage- operated calcium channel (L-VOCC) blocking effect of drotaverine. Hence, in this work, we focus on the potential L-VOCC blocking effect of drotaverine by using L-VOCC-associated functional in vitro models. Accordingly, drotaverine and reference agents were tested on KCl-induced guinea pig tracheal contraction. Drotaverine, like the L-VOCC blockers nifedipine or diltiazem, inhibited the KCl-induced inward Ca(2+)- induced contraction in a concentration- dependent fashion. The PDE inhibitor theophylline had no effect on the KCl-evoked contractions, indicating its lack of inhibition on inward Ca(2+) flow. Drotaverine was also tested on the L-VOCC-mediated resting Ca(2+) refill model. In this model, the extracellular Ca(2+) enters the cells to replenish the emptied intracellular Ca(2+) stores. Drotaverine and L-VOCC blocker reference molecules inhibited Ca(2+) replenishment of Ca(2+)-depleted preparations detected by agonist-induced contractions in post-Ca(2+) replenishment Ca(2+)-free medium. Theophylline did not modify the Ca(2+) store replenishment after contraction. It seems that drotaverine, but not theophylline, inhibits inward Ca(2+) flux. The addition of CaCl2 to Ca(2+)-free medium containing the agonist induced inward Ca(2+) flow and subsequent contraction of Ca(2+)-depleted tracheal preparations. Drotaverine, similar to the L-VOCC blockers, inhibited inward Ca(2+) flow and blunted the slope of CaCl2-induced contraction in agonist containing Ca(2+)-free medium with Ca(2+)-depleted tracheal preparations. These results show that drotaverine behaves like L-VOCC blockers but, unlike PDE inhibitors using L-VOCC associated in vitro experimental models.

  3. Effects of calcium channel blockers on proteinuria in patients with diabetic nephropathy. (United States)

    Toto, Robert D; Tian, Min; Fakouhi, Kaffa; Champion, Annette; Bacher, Peter


    Diabetic nephropathy management should include the use of an angiotensin-converting enzyme inhibitor (ACEI) or an angiotensin receptor blocker with additional antihypertensive medications to reduce proteinuria and cardiovascular events. Some studies suggest that adding a nondihydropyridine rather than a dihydropyridine calcium channel blocker (CCB) may more effectively lower proteinuria. We hypothesized that a trandolapril/verapamil SR (T/V) fixed-dose combination (FDC) was superior to a benazepril/amlodipine (B/A) FDC for reducing albuminuria in 304 hypertensive diabetic nephropathy patients when treated for 36 weeks. No statistically significant differences were observed between groups in the primary end point; adjusted percentage change in urinary albumin/creatinine ratio (UACR), which increased (mean T/V, 29.29%; mean B/A, 8.49%; difference, 20.80%; P=.34); or in change in absolute UACR, which decreased (mean [g/g] T/V, -0.11; mean [g/g] B/A, -0.08; difference -0.03; P=.78). There were significant reductions in log UACR (mean change in T/V, -0.28; P<.01; mean change in B/A, -0.31; P<.001) and diastolic blood pressure in both groups and in systolic blood pressure in the B/A group. T/V was not superior to B/A for reducing UACR. Both ACEI/CCB FDCs may reduce albuminuria; in the case of T/V, this appears to be independent of systolic blood pressure reduction in patients who had previously been treated and had baseline blood pressure levels of 142/77 mm Hg.

  4. Calcium-sensing receptor activation contributed to apoptosis stimulates TRPC6 channel in rat neonatal ventricular myocytes

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Yi-hua [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Li, Yong-quan [Harbin Medical University, Harbin 150086 (China); Feng, Shan-li [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Li, Bao-xin; Pan, Zhen-wei [Department of Pharmacology, Harbin Medical University, Harbin 150086 (China); Xu, Chang-qing [Department of Pathophysiology, Harbin Medical University, Harbin 150086 (China); Li, Ting-ting [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Yang, Bao-feng, E-mail: [Department of Pharmacology, Harbin Medical University, Harbin 150086 (China)


    Capacitative calcium entry (CCE) refers to the influx of calcium through plasma membrane channels activated on depletion of endoplasmic sarcoplasmic/reticulum (ER/SR) Ca{sup 2+} stores, which is performed mainly by the transient receptor potential (TRP) channels. TRP channels are expressed in cardiomyocytes. Calcium-sensing receptor (CaR) is also expressed in rat cardiac tissue and plays an important role in mediating cardiomyocyte apoptosis. However, there are no data regarding the link between CaR and TRP channels in rat heart. In this study, in rat neonatal myocytes, by Ca{sup 2+} imaging, we found that the depletion of ER/SR Ca{sup 2+} stores by thapsigargin (TG) elicited a transient rise in cytoplasmic Ca{sup 2+} ([Ca{sup 2+}]{sub i}), followed by sustained increase depending on extracellular Ca{sup 2+}. But, TRP channels inhibitor (SKF96365), not L-type channels or the Na{sup +}/Ca{sup 2+} exchanger inhibitors, inhibited [Ca{sup 2+}]{sub i} relatively high. Then, we found that the stimulation of CaR with its activator gadolinium chloride (GdCl{sub 3}) or by an increased extracellular Ca{sup 2+}([Ca{sup 2+}]{sub o}) increased the concentration of intracelluar Ca{sup 2+}, whereas, the sustained elevation of [Ca{sup 2+}]{sub i} was reduced in the presence of SKF96365. Similarly, the duration of [Ca{sup 2+}]{sub i} increase was also shortened in the absence of extracellular Ca{sup 2+}. Western blot analysis showed that GdCl{sub 3} increased the expression of TRPC6, which was reversed by SKF96365. Additionally, SKF96365 reduced cardiomyocyte apoptosis induced by GdCl{sub 3}. Our results suggested that CCE exhibited in rat neonatal myocytes and CaR activation induced Ca{sup 2+}-permeable cationic channels TRPCs to gate the CCE, for which TRPC6 was one of the most likely candidates. TRPC6 channel was functionally coupled with CaR to enhance the cardiomyocyte apoptosis.

  5. Membrane coordination of receptors and channels mediating the inhibition of neuronal ion currents by ADP. (United States)

    Gafar, Hend; Dominguez Rodriguez, Manuel; Chandaka, Giri K; Salzer, Isabella; Boehm, Stefan; Schicker, Klaus


    ADP and other nucleotides control ion currents in the nervous system via various P2Y receptors. In this respect, Cav2 and Kv7 channels have been investigated most frequently. The fine tuning of neuronal ion channel gating via G protein coupled receptors frequently relies on the formation of higher order protein complexes that are organized by scaffolding proteins and harbor receptors and channels together with interposed signaling components. However, ion channel complexes containing P2Y receptors have not been described. Therefore, the regulation of Cav2.2 and Kv7.2/7.3 channels via P2Y1 and P2Y12 receptors and the coordination of these ion channels and receptors in the plasma membranes of tsA 201 cells have been investigated here. ADP inhibited currents through Cav2.2 channels via both P2Y1 and P2Y12 receptors with phospholipase C and pertussis toxin-sensitive G proteins being involved, respectively. The nucleotide controlled the gating of Kv7 channels only via P2Y1 and phospholipase C. In fluorescence energy transfer assays using conventional as well as total internal reflection (TIRF) microscopy, both P2Y1 and P2Y12 receptors were found juxtaposed to Cav2.2 channels, but only P2Y1, and not P2Y12, was in close proximity to Kv7 channels. Using fluorescence recovery after photobleaching in TIRF microscopy, evidence for a physical interaction was obtained for the pair P2Y12/Cav2.2, but not for any other receptor/channel combination. These results reveal a membrane juxtaposition of P2Y receptors and ion channels in parallel with the control of neuronal ion currents by ADP. This juxtaposition may even result in apparent physical interactions between receptors and channels.

  6. Tolperisone-type drugs inhibit spinal reflexes via blockade of voltage-gated sodium and calcium channels. (United States)

    Kocsis, Pál; Farkas, Sándor; Fodor, László; Bielik, Norbert; Thán, Márta; Kolok, Sándor; Gere, Anikó; Csejtei, Mónika; Tarnawa, István


    The spinal reflex depressant mechanism of tolperisone and some of its structural analogs with central muscle relaxant action was investigated. Tolperisone (50-400 microM), eperisone, lanperisone, inaperisone, and silperisone (25-200 microM) dose dependently depressed the ventral root potential of isolated hemisected spinal cord of 6-day-old rats. The local anesthetic lidocaine (100-800 microM) produced qualitatively similar depression of spinal functions in the hemicord preparation, whereas its blocking effect on afferent nerve conduction was clearly stronger. In vivo, tolperisone and silperisone as well as lidocaine (10 mg/kg intravenously) depressed ventral root reflexes and excitability of motoneurons. However, in contrast with lidocaine, the muscle relaxant drugs seemed to have a more pronounced action on the synaptic responses than on the excitability of motoneurons. Whole-cell measurements in dorsal root ganglion cells revealed that tolperisone and silperisone depressed voltage-gated sodium channel conductance at concentrations that inhibited spinal reflexes. Results obtained with tolperisone and its analogs in the [3H]batrachotoxinin A 20-alpha-benzoate binding in cortical neurons and in a fluorimetric membrane potential assay in cerebellar neurons further supported the view that blockade of sodium channels may be a major component of the action of tolperisone-type centrally acting muscle relaxant drugs. Furthermore, tolperisone, eperisone, and especially silperisone had a marked effect on voltage-gated calcium channels, whereas calcium currents were hardly influenced by lidocaine. These data suggest that tolperisone-type muscle relaxants exert their spinal reflex inhibitory action predominantly via a presynaptic inhibition of the transmitter release from the primary afferent endings via a combined action on voltage-gated sodium and calcium channels.

  7. Release of glutamate and CGRP from trigeminal ganglion neurons: Role of calcium channels and 5-HT1 receptor signaling

    Directory of Open Access Journals (Sweden)

    Hurley Joyce H


    Full Text Available Abstract Background The aberrant release of the neurotransmitters, glutamate and calcitonin-gene related peptide (CGRP, from trigeminal neurons has been implicated in migraine. The voltage-gated P/Q-type calcium channel has a critical role in controlling neurotransmitter release and has been linked to Familial Hemiplegic Migraine. Therefore, we examined the importance of voltage-dependent calcium channels in controlling release of glutamate and CGRP from trigeminal ganglion neurons isolated from male and female rats and grown in culture. Serotonergic pathways are likely involved in migraine, as triptans, a class of 5-HT1 receptor agonists, are effective in the treatment of migraine and their effectiveness may be due to inhibiting neurotransmitter release from trigeminal neurons. We also studied the effect of serotonin receptor activation on release of glutamate and CGRP from trigeminal neurons grown in culture. Results P/Q-, N- and L-type channels each mediate a significant fraction of potassium-stimulated release of glutamate and CGRP. We determined that 5-HT significantly inhibits potassium-stimulated release of both glutamate and CGRP. Serotonergic inhibition of both CGRP and glutamate release can be blocked by pertussis toxin and NAS-181, a 5-HT1B/1D antagonist. Stimulated release of CGRP is unaffected by Y-25130, a 5-HT3 antagonist and SB 200646, a 5-HT2B/2C antagonist. Conclusion These data suggest that release of both glutamate and CGRP from trigeminal neurons is controlled by calcium channels and modulated by 5-HT signaling in a pertussis-toxin dependent manner and probably via 5-HT1 receptor signaling. This is the first characterization of glutamate release from trigeminal neurons grown in culture.

  8. Seeing the forest through the trees: towards a unified view on physiological calcium regulation of voltage-gated sodium channels. (United States)

    Van Petegem, Filip; Lobo, Paolo A; Ahern, Christopher A


    Voltage-gated sodium channels (Na(V)s) underlie the upstroke of the action potential in the excitable tissues of nerve and muscle. After opening, Na(V)s rapidly undergo inactivation, a crucial process through which sodium conductance is negatively regulated. Disruption of inactivation by inherited mutations is an established cause of lethal cardiac arrhythmia, epilepsy, or painful syndromes. Intracellular calcium ions (Ca(2+)) modulate sodium channel inactivation, and multiple players have been suggested in this process, including the cytoplasmic Na(V) C-terminal region including two EF-hands and an IQ motif, the Na(V) domain III-IV linker, and calmodulin. Calmodulin can bind to the IQ domain in both Ca(2+)-bound and Ca(2+)-free conditions, but only to the DIII-IV linker in a Ca(2+)-loaded state. The mechanism of Ca(2+) regulation, and its composite effect(s) on channel gating, has been shrouded in much controversy owing to numerous apparent experimental inconsistencies. Herein, we attempt to summarize these disparate data and propose a novel, to our knowledge, physiological mechanism whereby calcium ions promote sodium current facilitation due to Ca(2+) memory at high-action-potential frequencies where Ca(2+) levels may accumulate. The available data suggest that this phenomenon may be disrupted in diseases where cytoplasmic calcium ion levels are chronically high and where targeted phosphorylation may decouple the Ca(2+) regulatory machinery. Many Na(V) disease mutations associated with electrical dysfunction are located in the Ca(2+)-sensing machinery and misregulation of Ca(2+)-dependent channel modulation is likely to contribute to disease phenotypes.

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

    Directory of Open Access Journals (Sweden)

    Ong John M


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

  10. Physiology and evolution of voltage-gated calcium channels in early diverging animal phyla: Cnidaria, Placozoa, Porifera and Ctenophora

    Directory of Open Access Journals (Sweden)

    Adriano Senatore


    Full Text Available Voltage-gated calcium (Cav channels serve dual roles in the cell, where they can both depolarize the membrane potential for electrical excitability, and activate transient cytoplasmic Ca2+ signals. In animals, Cav channels play crucial roles including driving muscle contraction (excitation-contraction coupling, gene expression (excitation-transcription coupling, pre-synaptic and neuroendocrine exocytosis (excitation-secretion coupling, regulation of flagellar/ciliary beating, and regulation of cellular excitability, either directly or through modulation of other Ca2+-sensitive ion channels. In recent years, genome sequencing has provided significant insights into the molecular evolution of Cav channels. Furthermore, expanded gene datasets have permitted improved inference of the species phylogeny at the base of Metazoa, providing clearer insights into the evolution of complex animal traits which involve Cav channels, including the nervous system. For the various types of metazoan Cav channels, key properties that determine their cellular contribution include: ion selectivity, pore gating, and, importantly, cytoplasmic protein-protein interactions that direct sub-cellular localization and functional complexing. It is unclear when many of these defining features, many of which are essential for nervous system function, evolved. In this review, we highlight some experimental observations that implicate Cav channels in the physiology and behavior of the most early-diverging animals from the phyla Cnidaria, Placozoa, Porifera and Ctenophora. Given our limited understanding of the molecular biology of Cav channels in these basal animal lineages, we infer insights from better-studied vertebrate and invertebrate animals. We also highlight some apparently conserved cellular functions of Cav channels, which might have emerged very early on during metazoan evolution, or perhaps predated it.

  11. Physiology and Evolution of Voltage-Gated Calcium Channels in Early Diverging Animal Phyla: Cnidaria, Placozoa, Porifera and Ctenophora (United States)

    Senatore, Adriano; Raiss, Hamad; Le, Phuong


    Voltage-gated calcium (Cav) channels serve dual roles in the cell, where they can both depolarize the membrane potential for electrical excitability, and activate transient cytoplasmic Ca2+ signals. In animals, Cav channels play crucial roles including driving muscle contraction (excitation-contraction coupling), gene expression (excitation-transcription coupling), pre-synaptic and neuroendocrine exocytosis (excitation-secretion coupling), regulation of flagellar/ciliary beating, and regulation of cellular excitability, either directly or through modulation of other Ca2+-sensitive ion channels. In recent years, genome sequencing has provided significant insights into the molecular evolution of Cav channels. Furthermore, expanded gene datasets have permitted improved inference of the species phylogeny at the base of Metazoa, providing clearer insights into the evolution of complex animal traits which involve Cav channels, including the nervous system. For the various types of metazoan Cav channels, key properties that determine their cellular contribution include: Ion selectivity, pore gating, and, importantly, cytoplasmic protein-protein interactions that direct sub-cellular localization and functional complexing. It is unclear when these defining features, many of which are essential for nervous system function, evolved. In this review, we highlight some experimental observations that implicate Cav channels in the physiology and behavior of the most early-diverging animals from the phyla Cnidaria, Placozoa, Porifera, and Ctenophora. Given our limited understanding of the molecular biology of Cav channels in these basal animal lineages, we infer insights from better-studied vertebrate and invertebrate animals. We also highlight some apparently conserved cellular functions of Cav channels, which might have emerged very early on during metazoan evolution, or perhaps predated it. PMID:27867359

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


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


    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. ...

  13. 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:


    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)

  14. [Clinical efficacy of calcium channel blockers slow the third generation of lercanidipine in the treatment of patients with arterial hypertension and metabolic disorders (review)]. (United States)

    Tabidze, G A; Gezeli, T D; Tsibadze, T A; Dolidze, N M


    Arterial hypertension is the most common risk factor in patients with metabolic disorders. In the selection of antihypertensive therapy it is necessary to consider not only the anti-hypertensive and organoprotective effects of drugs and their metabolic effects, which has prognostic value. Calcium antaginists, along. Lercanidipine related to the third generation dihydripyridine calcium antagonist, has been much more selective for the so-called slow calcium channels of vascular smooth muscle cells, which is associated with a good hypertensive, organo and metabolic action. Combination therapy with an ACE inhibitor and a calcium channel blocker is also a justified tactic for the management of patients with high-risk cardiovascular and metabolic disorders. Attention is paid new fixed combinations, including angiotensin converting enzyme inhibitors and calcium antagonists.

  15. Pharmacologic inhibition of small-conductance calcium-activated potassium (SK) channels by NS8593 reveals atrial antiarrhythmic potential in horses

    DEFF Research Database (Denmark)

    Haugaard, Maria Mathilde; Hesselkilde, Eva Zander; Pehrson, Steen Michael


    BACKGROUND: Small-conductance calcium-activated potassium (SK) channels have been found to play an important role in atrial repolarization and atrial fibrillation (AF). OBJECTIVE: The purpose of this study was to investigate the existence and functional role of SK channels in the equine heart...

  16. Aberrant Splicing Promotes Proteasomal Degradation of L-type Ca v 1.2 Calcium Channels by Competitive Binding for CaV β Subunits in Cardiac Hypertrophy

    NARCIS (Netherlands)

    Hu, Zhenyu; Wang, Jiong Wei; Yu, Dejie; Soon, Jia Lin; De Kleijn, Dominique P V; Foo, Roger; Liao, Ping; Colecraft, Henry M.; Soong, Tuck Wah


    Decreased expression and activity of Ca V1.2 calcium channels has been reported in pressure overload-induced cardiac hypertrophy and heart failure. However, the underlying mechanisms remain unknown. Here we identified in rodents a splice variant of Ca V1.2 channel, named Ca V1.2 e21+22, that contain

  17. 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


    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.

  18. High glucose enhances transient receptor potential channel canonical type 6-dependent calcium influx in human platelets via phosphatidylinositol 3-kinase-dependent pathway

    DEFF Research Database (Denmark)

    Liu, Daoyan; Maier, Alexandra; Scholze, Alexandra;


    Transient receptor potential canonical type 6 (TRPC6) channels mediating 1-oleoyl-2-acetyl-sn-glycerol (OAG)-induced calcium entry have been identified on human platelets. In the present study we tested the hypothesis that hyperglycemia increases the expression of TRPC6 channels....

  19. Presynaptic calcium channels and α3-integrins are complexed with synaptic cleft laminins, cytoskeletal elements and active zone components. (United States)

    Carlson, Steven S; Valdez, Gregorio; Sanes, Joshua R


    At chemical synapses, synaptic cleft components interact with elements of the nerve terminal membrane to promote differentiation and regulate function. Laminins containing the β2 subunit are key cleft components, and they act in part by binding the pore-forming subunit of a pre-synaptic voltage-gated calcium channel (Ca(v)α) (Nishimune et al. 2004). In this study, we identify Ca(v)α-associated intracellular proteins that may couple channel-anchoring to assembly or stabilization of neurotransmitter release sites called active zones. Using Ca(v)α-antibodies, we isolated a protein complex from Torpedo electric organ synapses, which resemble neuromuscular junctions but are easier to isolate in bulk. We identified 10 components of the complex: six cytoskeletal proteins (α2/β2 spectrins, plectin 1, AHNAK/desmoyokin, dystrophin, and myosin 1), two active zone components (bassoon and piccolo), synaptic laminin, and a calcium channel β subunit. Immunocytochemistry confirmed these proteins in electric organ synapses, and PCR analysis revealed their expression by developing mammalian motor neurons. Finally, we show that synaptic laminins also interact with pre-synaptic integrins containing the α3 subunit. Together with our previous finding that a distinct synaptic laminin interacts with SV2 on nerve terminals (Son et al. 2000), our results identify three paths by which synaptic cleft laminins can send developmentally important signals to nerve terminals.

  20. Potentiation of Opioid-Induced Analgesia by L-Type Calcium Channel Blockers: Need for Clinical Trial in Cancer Pain

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    S Basu Ray


    Full Text Available Previous reports indicate that the analgesic effect of opioids is due to both closure of specific voltage-gated calcium channels (N- and P/Q-types and opening of G protein-coupled inwardly rectifying potassium channels (GIRKs in neurons concerned with transmission of pain. However, administration of opioids leads to unacceptable levels of side effects, particularly at high doses. Thus, current research is directed towards simultaneously targeting other voltage-gated calcium channels (VGCCs like the L-type VGCCs or even other cell signaling mechanisms, which would aug-ment opioid-mediated analgesic effect without a concurrent increase in the side effects. Unfortunately, the results of these studies are often conflicting considering the different experimental paradigms (variable drug selection and their doses and also the specific pain test used for studying analgesia adopted by researchers. The present review focuses on some of the interesting findings regarding the analgesic effect of Opioids + L-VGCC blockers and suggests that time has come for a clinical trial of this combination of drugs in the treatment of cancer pain.

  1. Science Signaling Podcast for 24 January 2017: Tissue-specific regulation of L-type calcium channels. (United States)

    Hell, Johannes W; Navedo, Manuel F; VanHook, Annalisa M


    This Podcast features an interview with Johannes Hell and Manuel Navedo, senior authors of two Research Articles that appear in the 24 January 2017 issue of Science Signaling, about tissue-specific regulation of the L-type calcium channel CaV1.2. This channel is present in many tissues, including the heart, vasculature, and brain, and allows calcium to flow into cells when it is activated. Signaling through the β-adrenergic receptor (βAR) stimulates CaV1.2 activity in heart cells and neurons to accelerate heart rate and increase neuronal excitability, respectively. Using mouse models, Qian et al found that βAR-mediated enhancement of CaV1.2 activity in the brain required phosphorylation of Ser(1928), whereas βAR-mediated enhancement of CaV1.2 activity in the heart did not require phosphorylation of this residue. In a related study, Nystoriak et al demonstrated that phosphorylation of Ser(1928) in arterial myocytes was required for vasoconstriction during acute hyperglycemia and in diabetic mice. These findings demonstrate tissue-specific differences in CaV1.2 regulation and suggest that it may be possible to design therapies to target this channel in specific tissues.Listen to Podcast.

  2. Antihypertensive and vasorelaxant activities of Laelia autumnalis are mainly through calcium channel blockade. (United States)

    Vergara-Galicia, Jorge; Ortiz-Andrade, Rolffy; Castillo-España, Patricia; Ibarra-Barajas, Maximiliano; Gallardo-Ortiz, Itzell; Villalobos-Molina, Rafael; Estrada-Soto, Samuel


    The aim of the present study was to evaluate the possible mechanism of the vasorelaxant action of methanol extract from Laelia autumnalis (MELa) in isolated rat aortic rings, and to establish its antihypertensive activity in vivo. MELa (0.15-->50 microg/mL) induced relaxation in aortic rings pre-contracted with KCl (80 mM), showing an IC50 value of 34.61+/-1.41 microg/mL and E max value of 85.0+/-4.38% (in endothelium-intact rings) and an IC50 value of 45.11+/-4.17 microg/mL and E max value of 80.0+/-12.1% (in endothelium-denuded rings). Serotonin (5-HT, 1 x 10(-4) M) provoked sustained contraction, which was markedly inhibited by MELa (0.15-->50 microg/mL) in a concentration-dependent and endothelium-independent manner. Pretreatment with MELa (15, 46, 150, 300 and 1500 microg/mL) also inhibited contractile responses to norepinephrine (NE 1 x 10(-11) M to 1 x 10(-5.5) M). In endothelium-denuded rings, the vasorelaxant effect of MELa was reduced partially by ODQ (1 microM), but not by tetraethylammonium (5 microM), glibenclamide (10 microM), and 2-aminopyridine (100 microM). The extract also reduced NE-induced transient contraction in Ca2+-free solution, and inhibited contraction induced by increasing external calcium in Ca2+-free medium plus high KCl (80 mM). The antihypertensive effect of MELa was determined in spontaneously hypertensive rats (SHR). A single oral administration of the extract (100 mg/kg) exhibited a significant decrease in systolic and diastolic blood pressure and heart rate (p<0.05) in SHR rats. Our results suggest that MELa induces relaxation in rat aortic rings through an endothelium-independent pathway, involving blockade of Ca2+ channels and a possible cGMP enhanced concentrations and also causes an antihypertensive effect.

  3. Effects of calcium channel antagonists on the motivational effects of nicotine and morphine in conditioned place aversion paradigm. (United States)

    Budzynska, Barbara; Polak, Piotr; Biala, Grazyna


    The motivational component of drug withdrawal may contribute to drug seeking and relapse through the negative reinforcement-related process; thus, it is important to understand the mechanisms that mediate affective withdrawal behaviors. The present study was undertaken to examine the calcium-dependent mechanism of negative motivational symptoms of nicotine and morphine withdrawal using the conditioned place aversion (CPA) paradigm. Rats were chronically treated with nicotine (1.168 mg/kg, free base, s.c., 11 days, three times daily) or morphine (10 mg/kg,s.c., 11 days, twice daily). Then, during conditioning, rats pre-treated with nicotine or morphine received a nicotinic receptor antagonist mecamylamine (3.5 mg/kg) or an opioid receptor antagonist naloxone (1 mg/kg) to precipitate withdrawal in their initially preferred compartment, or saline in their non-preferred compartment. Our results demonstrated that after three conditioning sessions, mecamylamine induced a clear place aversion in rats that had previously received nicotine injections, and naloxone induced a significant place aversion in rats that had previously received morphine injections. Further, the major findings showed that calcium channel antagonists, i.e., nimodipine, verapamil and flunarizine (5 and 10 mg/kg, i.p.), injected before the administration of mecamylamine or naloxone, attenuated nicotine or morphine place aversion. As an outcome, these findings support the hypothesis that similar calcium-dependent mechanisms are involved in aversive motivational component associated with nicotine a morphine withdrawal. We can suggest that calcium channel blockers have potential for alleviating nicotine and morphine addiction by selectively decreasing the incentive motivational properties of both drugs, and may be beneficial as smoking cessation or opioid dependence pharmacotherapies.

  4. Lung adenocarcinoma with Lambert–Eaton myasthenic syndrome indicated by voltage-gated calcium channel: a case report

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    Arai Hiromasa


    Full Text Available Abstract Introduction Lambert–Eaton myasthenic syndrome is a rare disorder and it is known as a paraneoplastic neurological syndrome. Small cell lung cancer often accompanies this syndrome. Lambert–Eaton myasthenic syndrome associated with lung adenocarcinoma is extremely rare; there are only a few reported cases worldwide. Case presentation A 75-year-old Japanese man with a past history of chronic rheumatoid arthritis and Sjögren syndrome was diagnosed with Lambert–Eaton myasthenic syndrome by electromyography and serum anti-P/Q-type voltage-gated calcium channel antibody level preceding the diagnosis of lung cancer. A chest computed tomography to screen for malignant lesions revealed an abnormal shadow in the lung. Although a histopathological examination by bronchoscopic study could not reveal the malignancy, lung cancer was mostly suspected after the results of a chest computed tomography and [18F]-fluorodeoxyglucose positron emission tomography. An intraoperative diagnosis based on the frozen section obtained by tumor biopsy was adenocarcinoma so the patient underwent a lobectomy of the right lower lobe and lymph node dissection with video-assisted thoracoscopic surgery. The permanent pathological examination was the same as the frozen diagnosis (pT2aN1M0: Stage IIa: TNM staging 7th edition. Immunohistochemistry revealed that most of the cancer cells were positive for P/Q-type voltage-gated calcium channel. Conclusions Our case is a rare combination of Lambert–Eaton myasthenic syndrome associated with lung adenocarcinoma, rheumatoid arthritis and Sjögren syndrome, and to the best of our knowledge it is the first report that indicates the presence of voltage-gated calcium channel in lung adenocarcinoma by immunostaining.

  5. Hydrogen sulfide-induced itch requires activation of Cav3.2 T-type calcium channel in mice (United States)

    Wang, Xue-Long; Tian, Bin; Huang, Ya; Peng, Xiao-Yan; Chen, Li-Hua; Li, Jun-Cheng; Liu, Tong


    The contributions of gasotransmitters to itch sensation are largely unknown. In this study, we aimed to investigate the roles of hydrogen sulfide (H2S), a ubiquitous gasotransmitter, in itch signaling. We found that intradermal injection of H2S donors NaHS or Na2S, but not GYY4137 (a slow-releasing H2S donor), dose-dependently induced scratching behavior in a μ-opioid receptor-dependent and histamine-independent manner in mice. Interestingly, NaHS induced itch via unique mechanisms that involved capsaicin-insensitive A-fibers, but not TRPV1-expressing C-fibers that are traditionally considered for mediating itch, revealed by depletion of TRPV1-expressing C-fibers by systemic resiniferatoxin treatment. Moreover, local application of capsaizapine (TRPV1 blocker) or HC-030031 (TRPA1 blocker) had no effects on NaHS-evoked scratching. Strikingly, pharmacological blockade and silencing of Cav3.2 T-type calcium channel by mibefradil, ascorbic acid, zinc chloride or Cav3.2 siRNA dramatically decreased NaHS-evoked scratching. NaHS induced robust alloknesis (touch-evoked itch), which was inhibited by T-type calcium channels blocker mibefradil. Compound 48/80-induced itch was enhanced by an endogenous precursor of H2S (L-cysteine) but attenuated by inhibitors of H2S-producing enzymes cystathionine γ-lyase and cystathionine β-synthase. These results indicated that H2S, as a novel nonhistaminergic itch mediator, may activates Cav3.2 T-type calcium channel, probably located at A-fibers, to induce scratching and alloknesis in mice. PMID:26602811

  6. P/Q-type and T-type voltage-gated calcium channels are involved in the contraction of mammary and brain blood vessels from hypertensive patients

    DEFF Research Database (Denmark)

    Thuesen, A D; Lyngsø, K S; Rasmussen, L


    AIM: Calcium channel blockers are widely used in cardiovascular diseases. Besides L-type channels, T- and P/Q-type calcium channels are involved in the contraction of human renal blood vessels. It was hypothesized that T- and P/Q-type channels are involved in the contraction of human brain...... and mammary blood vessels. METHODS: Internal mammary arteries from bypass surgery patients and cerebral arterioles from patients with brain tumours with and without hypertension were tested in a myograph and perfusion set-up. PCR and immunohistochemistry were performed on isolated blood vessels. RESULTS......: The P/Q-type antagonist ω-agatoxin IVA (10(-8) mol L(-1) ) and the T-type calcium blocker mibefradil (10(-7) mol L(-1) ) inhibited KCl depolarization-induced contraction in mammary arteries from hypertensive patients with no effect on blood vessels from normotensive patients. ω-Agatoxin IVA decreased...

  7. Emerging roles of calcium-activated K channels and TRPV4 channels in lung oedema and pulmonary circulatory collapse

    DEFF Research Database (Denmark)

    Simonsen, Ulf; Wandall-Frostholm, Christine; Oliván-Viguera, Aida;


    endothelial/epithelial barrier functions and vascular integrity, while KCa3.1 channels provide the driving force required for Cl(-) and water transport in some cells and most secretory epithelia. The three conditions, increased pulmonary venous pressure caused by left heart disease, high inflation pressure......, fluid extravasation, hemorrhage, pulmonary circulatory collapse, and cardiac arrest in vivo. These data identify KCa3.1 channels as crucial molecular components in downstream TRPV4-signal transduction and as a potential target for the prevention of undesired fluid extravasation, vasodilatation...

  8. The mystery is solved-CatSper is the principal calcium channel activated by progesterone in human spermatozoa

    Institute of Scientific and Technical Information of China (English)

    Christopher LR Barratt


    @@ Aremarkable advance in sperm physiology has recently been published in Nature.Two groups using patch clamping techniques on human sperm have solved a mystery about the sperm cell that has puzzled both andrologists and those involved in non-genomic cellular signalling for over 20 years.In these papers, Lishko1 and Strunker2 independently demonstrate that the universal characteristic effect of progesterone on sperm-a rapid influx of calcium-is via a sperm-specific channel CatSper.

  9. 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;


    pathophysiology. Here we study the expression and localization of BK(Ca) channels and CGRP in the rat trigeminal ganglion (TG) and the trigeminal nucleus caudalis (TNC) as these structures are involved in migraine pain. Also the effect of the BK(Ca) channel blocker iberiotoxin and the BK(Ca) channel opener NS......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 migraine...

  10. Activation of TRPV2 and BKCa channels by the LL-37 enantiomers stimulates calcium entry and migration of cancer cells (United States)

    Guéguinou, Maxime; Chourpa, Igor; Fromont, Gaëlle; Bouchet, Ana Maria; Burlaud-Gaillard, Julien; Potier-Cartereau, Marie; Roger, Sébastien; Aucagne, Vincent; Chevalier, Stéphan; Vandier, Christophe


    Expression of the antimicrobial peptide hCAP18/LL-37 is associated to malignancy in various cancer forms, stimulating cell migration and metastasis. We report that LL-37 induces migration of three cancer cell lines by activating the TRPV2 calcium-permeable channel and recruiting it to pseudopodia through activation of the PI3K/AKT pathway. Ca2+ entry through TRPV2 cooperated with a K+ efflux through the BKCa channel. In a panel of human breast tumors, the expression of TRPV2 and LL-37 was found to be positively correlated. The D-enantiomer of LL-37 showed identical effects as the L-peptide, suggesting that no binding to a specific receptor was involved. LL-37 attached to caveolae and pseudopodia membranes and decreased membrane fluidity, suggesting that a modification of the physical properties of the lipid membrane bilayer was the underlying mechanism of its effects. PMID:26993604

  11. Sensitization of voltage activated calcium channel currents for capsaicin in nociceptive neurons by tumor-necrosis-factor-alpha. (United States)

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


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

  12. PKC enhances the capacity for secretion by rapidly recruiting covert voltage-gated Ca2+ channels to the membrane. (United States)

    Groten, Christopher J; Magoski, Neil S


    It is unknown whether neurons can dynamically control the capacity for secretion by promptly changing the number of plasma membrane voltage-gated Ca(2+) channels. To address this, we studied peptide release from the bag cell neurons of Aplysia californica, which initiate reproduction by secreting hormone during an afterdischarge. This burst engages protein kinase C (PKC) to trigger the insertion of a covert Ca(2+) channel, Apl Cav2, alongside a basal channel, Apl Cav1. The significance of Apl Cav2 recruitment to secretion remains undetermined; therefore, we used capacitance tracking to assay secretion, along with Ca(2+) imaging and Ca(2+) current measurements, from cultured bag cell neurons under whole-cell voltage-clamp. Activating PKC with the phorbol ester, PMA, enhanced Ca(2+) entry, and potentiated stimulus-evoked secretion. This relied on channel insertion, as it was occluded by preventing Apl Cav2 engagement with prior whole-cell dialysis or the cytoskeletal toxin, latrunculin B. Channel insertion reduced the stimulus duration and/or frequency required to initiate secretion and strengthened excitation-secretion coupling, indicating that Apl Cav2 accesses peptide release more readily than Apl Cav1. The coupling of Apl Cav2 to secretion also changed with behavioral state, as Apl Cav2 failed to evoke secretion in silent neurons from reproductively inactive animals. Finally, PKC also acted secondarily to enhance prolonged exocytosis triggered by mitochondrial Ca(2+) release. Collectively, our results suggest that bag cell neurons dynamically elevate Ca(2+) channel abundance in the membrane to ensure adequate secretion during the afterdischarge.

  13. GABAA increases calcium in subventricular zone astrocyte-like cells through L- and T-type voltage-gated calcium channels

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    Stephanie Z Young


    Full Text Available In the adult neurogenic subventricular zone (SVZ, the behavior of astrocyte-like cells and some of their functions depend on changes in intracellular Ca2+ levels and tonic GABAA receptor activation. However, it is unknown whether, and if so how, GABAA receptor activity regulates intracellular Ca2+ dynamics in SVZ astrocytes. To monitor Ca2+ 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. GABAA receptor activation induced Ca2+ increases in 40-50% of SVZ astrocytes. GABAA-induced Ca2+ increases were prevented with nifedipine and mibefradil, blockers of L- and T-type voltage-gated calcium channels (VGCC. The L-type Ca2+ channel activator BayK 8644 increased the percentage of GABAA-responding astrocyte-like cells to 75%, suggesting that the majority of SVZ astrocytes express functional VGCCs. SVZ astrocytes also displayed spontaneous Ca2+ activity, the frequency of which was regulated by tonic GABAA receptor activation. These data support a role for ambient GABA in tonically regulating intracellular Ca2+ dynamics through GABAA receptors and VGCC in a subpopulation of astrocyte-like cells in the postnatal SVZ.

  14. Types of voltage—dependent calcium channels involved in high potassium depolarization—induced amylase secretion in the exocrine pancreatic tumour cell line AR4—2J

    Institute of Scientific and Technical Information of China (English)



    In the perifused fura-2 loaded exocrine pancreatic acinar cell line AR4-2J pulses of high potassium induced repetitive increases in intracellular calcium,Attached cells when stimulated with high potassium secreted large amount of amylase.High potassium-induced secretion was dependent both on the concentration of potassium and duration of stimulation.High potassium induced increases in intracellular calcium were inhibited by voltage-dependent calcium channel anatagonists with an order of potency as follows:nifedipine>ω-agatoxin IVA>ω-conotoxin GVIA.In contrast,the L-type calcium channel anatagonist nifedipine almost completely inhibited potassium-induced amylase secretion,whereas the N-type channel antagonist ω-conotoxin GVIA was without effect.The P-type channel antagonist ω-agatoxin IVA had a small inhibitory effect,but this inhibition was not significant at the level of amylase secretion.In conclusion,the AR4-2J cell line posesses different voltage-dependent calcium channels(L,P,N)with the L-type predominantly involved in depolarization induced amylase secretion.

  15. Negative inotropic action of denbufylline through interfering with the calcium channel independently of its PDE IV inhibitory activity in guinea pig ventricle papillary muscles. (United States)

    Sanae, F; Ohmae, S; Kobayashi, D; Takag, K; Miyamoto, K


    The inotropic actions of xanthine derivatives with long alkyl chains were investigated in guinea pig ventricular papillary muscle. A potent and nonselective phosphodiesterase (PDE) inhibitor, 3-isobutyl-1-methylxanthine, elicited a positive inotropy and inhibited the negative inotropic effects of calcium channel inhibitors, as did a selective PDE III inhibitor, amrinone, and these effects were canceled by a protein kinase inhibitor, N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89). However, 1,3-di-n-butyl-7-(2'oxopropyl)xanthine (denbufylline) and 1-n-butyl-3-n-propylxanthine (XT-044), which have potent and selective PDE IV-inhibitory activities, showed negative inotropic actions that became more potent in the presence of H-89. Denbufylline abolished the late restoration phase induced by ryanodine. This xanthine derivative attenuated the effects of both the calcium channel acting agents Bay K 8644 and verapamil, without interaction with caffeine and dihydropyridine calcium channel inhibitors, and denbufylline had little direct influence on the specific binding of [(3)H]azidopine and [(3)H]desmethoxyverapamil to cardiac membranes. A nonxanthine PDE IV inhibitor, Ro 20-1724, did not affect the inotropic actions of calcium channel inhibitors. The attenuation by denbufylline or XT-044 of the negative inotropic action of verapamil was not influenced by treatment with H-89. These results suggest that in the ventricular papillary muscle, these xanthine derivatives elicit negative inotropy by acting on a verapamil-sensitive site of the calcium channel without involving their PDE-inhibitory activity.

  16. Intermediate-Conductance-Ca2-Activated K Channel Intermediate-Conductance Calcium-Activated Potassium Channel (IKCa1) is Upregulated and Promotes Cell Proliferation in Cervical Cancer (United States)

    Liu, Ling; Zhan, Ping; Nie, Dan; Fan, Lingye; Lin, Hairui; Gao, Lanyang; Mao, Xiguang


    Background Accumulating data point to intermediate-conductance calcium-activated potassium channel (IKCa1) as a key player in controlling cell cycle progression and proliferation of human cancer cells. However, the role that IKCa1 plays in the growth of human cervical cancer cells is largely unexplored. Material/Methods In this study, Western blot analysis, immunohistochemical staining, and RT-PCR were first used for IKCa1protein and gene expression assays in cervical cancer tissues and HeLa cells. Then, IKCa1 channel blocker and siRNA were employed to inhibit the functionality of IKCa1 and downregulate gene expression in HeLa cells, respectively. After these treatments, we examined the level of cell proliferation by MTT method and measured IKCa1 currents by conventional whole-cell patch clamp technique. Cell apoptosis was assessed using the Annexin V-FITC/Propidium Iodide (PI) double-staining apoptosis detection kit. Results We demonstrated that IKCa1 mRNA and protein are preferentially expressed in cervical cancer tissues and HeLa cells. We also showed that the IKCa1 channel blocker, clotrimazole, and IKCa1 channel siRNA can be used to suppress cervical cancer cell proliferation and decrease IKCa1 channel current. IKCa1 downregulation by specific siRNAs induced a significant increase in the proportion of apoptotic cells in HeLa cells. Conclusions IKCa1 is overexpressed in cervical cancer tissues, and IKCa1 upregulation in cervical cancer cell linea enhances cell proliferation, partly by reducing the proportion of apoptotic cells. PMID:28280257

  17. Towards a Unified Theory of Calmodulin Regulation (Calmodulation) of Voltage-Gated Calcium and Sodium Channels. (United States)

    Ben-Johny, Manu; Dick, Ivy E; Sang, Lingjie; Limpitikul, Worawan B; Kang, Po Wei; Niu, Jacqueline; Banerjee, Rahul; Yang, Wanjun; Babich, Jennifer S; Issa, John B; Lee, Shin Rong; Namkung, Ho; Li, Jiangyu; Zhang, Manning; Yang, Philemon S; Bazzazi, Hojjat; Adams, Paul J; Joshi-Mukherjee, Rosy; Yue, Daniel N; Yue, David T


    Voltage-gated Na and Ca(2+) channels represent two major ion channel families that enable myriad biological functions including the generation of action potentials and the coupling of electrical and chemical signaling in cells. Calmodulin regulation (calmodulation) of these ion channels comprises a vital feedback mechanism with distinct physiological implications. Though long-sought, a shared understanding of the channel families remained elusive for two decades as the functional manifestations and the structural underpinnings of this modulation often appeared to diverge. Here, we review recent advancements in the understanding of calmodulation of Ca(2+) and Na channels that suggest a remarkable similarity in their regulatory scheme. This interrelation between the two channel families now paves the way towards a unified mechanistic framework to understand vital calmodulin-dependent feedback and offers shared principles to approach related channelopathic diseases. An exciting era of synergistic study now looms.

  18. Inhibition of collagen synthesis by select calcium and sodium channel blockers can be mitigated by ascorbic acid and ascorbyl palmitate. (United States)

    Ivanov, Vadim; Ivanova, Svetlana; Kalinovsky, Tatiana; Niedzwiecki, Aleksandra; Rath, Matthias


    Calcium, sodium and potassium channel blockers are widely prescribed medications for a variety of health problems, most frequently for cardiac arrhythmias, hypertension, angina pectoris and other disorders. However, chronic application of channel blockers is associated with numerous side effects, including worsening cardiac pathology. For example, nifedipine, a calcium-channel blocker was found to be associated with increased mortality and increased risk for myocardial infarction. In addition to the side effects mentioned above by different channel blockers, these drugs can cause arterial wall damage, thereby contributing to vascular wall structure destabilization and promoting events facilitating rupture of plaques. Collagen synthesis is regulated by ascorbic acid, which is also essential for its optimum structure as a cofactor in lysine and proline hydroxylation, a precondition for optimum crosslinking of collagen and elastin. Therefore, the main objective in this study was to evaluate effects of various types of channel blockers on intracellular accumulation and cellular functions of ascorbate, specifically in relation to formation and extracellular deposition of major collagen types relevant for vascular function. Effects of select Na- and Ca- channel blockers on collagen synthesis and deposition were evaluated in cultured human dermal fibroblasts and aortic smooth muscle cells by immunoassay. All channel blockers tested demonstrated inhibitory effects on collagen type I deposition to the ECM by fibroblasts, each to a different degree. Ascorbic acid significantly increased collagen I ECM deposition. Nifedipine (50 µM), a representative of channel blockers tested, significantly reduced ascorbic acid and ascorbyl palmitate-dependent ECM deposition of collagen type l and collagen type lV by cultured aortic smooth muscle cells. In addition, nifedipine (50 µM) significantly reduced ascorbate-dependent collagen type l and type lV synthesis by cultured aortic smooth

  19. Three dimensional neuronal cell cultures more accurately model voltage gated calcium channel functionality in freshly dissected nerve tissue.

    Directory of Open Access Journals (Sweden)

    Yinzhi Lai

    Full Text Available It has been demonstrated that neuronal cells cultured on traditional flat surfaces may exhibit exaggerated voltage gated calcium channel (VGCC functionality. To gain a better understanding of this phenomenon, primary neuronal cells harvested from mice superior cervical ganglion (SCG were cultured on two dimensional (2D flat surfaces and in three dimensional (3D synthetic poly-L-lactic acid (PLLA and polystyrene (PS polymer scaffolds. These 2D- and 3D-cultured cells were compared to cells in freshly dissected SCG tissues, with respect to intracellular calcium increase in response to high K(+ depolarization. The calcium increases were identical for 3D-cultured and freshly dissected, but significantly higher for 2D-cultured cells. This finding established the physiological relevance of 3D-cultured cells. To shed light on the mechanism behind the exaggerated 2D-cultured cells' functionality, transcriptase expression and related membrane protein distributions (caveolin-1 were obtained. Our results support the view that exaggerated VGCC functionality from 2D cultured SCG cells is possibly due to differences in membrane architecture, characterized by uniquely organized caveolar lipid rafts. The practical implication of use of 3D-cultured cells in preclinical drug discovery studies is that such platforms would be more effective in eliminating false positive hits and as such improve the overall yield from screening campaigns.

  20. Regulation of L-type Voltage Gated Calcium Channel CACNA1S in Macrophages upon Mycobacterium tuberculosis Infection. (United States)

    Antony, Cecil; Mehto, Subhash; Tiwari, Brijendra K; Singh, Yogendra; Natarajan, Krishnamurthy


    We demonstrated earlier the inhibitory role played by Voltage Gated Calcium Channels (VGCCs) in regulating Mycobacterium tuberculosis (M. tb) survival and pathogenesis. In this report, we investigated mechanisms and key players that regulate the surface expression of VGCC-CACNA1S by Rv2463 and M. tb infection in macrophages. Our earlier work identified Rv2463 to be expressed at early times post infection in macrophages that induced suppressor responses to dendritic cells and macrophages. Our results in this study demonstrate a role of MyD88 independent TLR pathway in mediating CACNA1S expression. Dissecting the role for second messengers, we show that calcium homeostasis plays a key role in CACNA1S expression during M. tb infection. Using siRNAs against molecular sensors of calcium regulation, we show an involvement of ER associated Stromal Interaction Molecules 1 and 2 (STIM1 and STIM2), and transcription factor pCREB, towards CACNA1S expression that also involved the MyD88 independent pathway. Interestingly, reactive oxygen species played a negative role in M. tb mediated CACNA1S expression. Further, a cross-regulation of ROS and pCREB was noted that governed CACNA1S expression. Characterizing the mechanisms governing CACNA1S expression would improve our understanding of the regulation of VGCC expression and its role in M. tb pathogenesis during M. tb infection.

  1. Reciprocal regulation of reactive oxygen species and phospho-CREB regulates voltage gated calcium channel expression during Mycobacterium tuberculosis infection.

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    Arti Selvakumar

    Full Text Available Our previous work has demonstrated the roles played by L-type Voltage Gated Calcium Channels (VGCC in regulating Mycobacterium tuberculosis (M. tb survival and pathogenesis. Here we decipher mechanisms and pathways engaged by the pathogen to regulate VGCC expression in macrophages. We show that M. tb and its antigen Rv3416 use phospho-CREB (pCREB, Reactive Oxygen Species (ROS, Protein Kinase C (PKC and Mitogen Activated Protein Kinase (MAPK to modulate VGCC expression in macrophages. siRNA mediated knockdown of MyD88, IRAK1, IRAK2 or TRAF6 significantly inhibited antigen mediated VGCC expression. Inhibiting Protein Kinase C (PKC or MEK-ERK1/2 further increased VGCC expression. Interestingly, inhibiting intracellular calcium release upregulated antigen mediated VGCC expression, while inhibiting extracellular calcium influx had no significant effect. siRNA mediated knockdown of transcription factors c-Jun, SOX5 and CREB significantly inhibited Rv3416 mediated VGCC expression. A dynamic reciprocal cross-regulation between ROS and pCREB was observed that in turn governed VGCC expression with ROS playing a limiting role in the process. Further dissection of the mechanisms such as the interplay between ROS and pCREB would improve our understanding of the regulation of VGCC expression during M. tb infection.

  2. Hypericum perforatum modulates apoptosis and calcium mobilization through voltage-gated and TRPM2 calcium channels in neutrophil of patients with Behcet's disease. (United States)

    Nazıroğlu, Mustafa; Sahin, Mehmet; Ciğ, Bilal; Aykur, Mehmet; Erturan, Ijlal; Ugan, Yunus


    Behcet's disease (BD) is a chronic, inflammatory, and multisystemic condition although its pathogenesis is uncertain. Main component of St. John's wort (Hypericum perforatum, HP) is hyperforin and induces antiinflammatory and antioxidant properties. We aimed to investigate effects of HP on oxidative stress, apoptosis, and cytosolic-free Ca²⁺ [Ca²⁺](i) concentration in neutrophil of BD patients. Nine new-diagnosed active patients with BD and nine control subjects were included in the study. Disease activity was considered by clinical findings. Neutrophil samples were obtained from the patients and controls. The neutrophils from patients were divided into three subgroups and were incubated with HP, voltage-gated calcium channel (VGCC) blockers, (verapamil+dilitiazem) and non-specific TRPM2 channel blocker (2-aminoethyl diphenylborinate, 2-APB), respectively. The neutrophils were stimulated by fMLP as a Ca²⁺-concentration agonist and oxidative stress former. Caspase-3, caspase-9, apoptosis, lipid peroxidation, and [Ca²⁺](i) values were high in the patient groups, although cell viability, glutathione (GSH), and glutathione peroxidase (GSH-Px) values were low in patient group. However, the [Ca²⁺](i), caspase-3, and caspase-9 values decreased markedly in patient+HP group although GSH and GSH-Px values increased in the group. The [Ca²⁺](i) concentration was also decreased in the patient group by V+D, 2-APB, and HP incubations. In conclusion, we observed the importance of neutrophil Ca²⁺ entry, apoptosis, and oxidative stress through gating VGCC and TRPM2 channels in the neutrophils in the pathogenesis and activation of the patients with BD. HP induced protective effects on oxidative stress by modulating Ca²⁺ influx in BD patients.

  3. Optical modulation of neurotransmission using calcium photocurrents through the ion channel LiGluR

    Directory of Open Access Journals (Sweden)

    Mercè eIzquierdo-Serra


    Full Text Available A wide range of light-activated molecules (photoswitches and phototriggers have been used to the study of computational properties of an isolated neuron by acting pre and postsynaptically. However, new tools are being pursued to elicit a presynaptic calcium influx that triggers the release of neurotransmitters, most of them based in calcium-permeable Channelrhodopsin-2 mutants. Here we describe a method to control exocytosis of synaptic vesicles through the use of a light-gated glutamate receptor (LiGluR, which has recently been demonstrated that supports secretion by means of calcium influx in chromaffin cells. Expression of LiGluR in hippocampal neurons enables reversible control of neurotransmission with light, and allows modulating the firing rate of the postsynaptic neuron with the wavelength of illumination. This method may be useful for the determination of the complex transfer function of individual synapses.

  4. Genetic Evidence for Possible Involvement of the Calcium Channel Gene CACNA1A in Autism Pathogenesis in Chinese Han Population.

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    Jun Li

    Full Text Available Autism spectrum disorders (ASD are a group of neurodevelopmental disorders. Recent studies suggested that calcium channel genes might be involved in the genetic etiology of ASD. CACNA1A, encoding an alpha-1 subunit of voltage-gated calcium channel, has been reported to play an important role in neural development. Previous study detected that a single nucleotide polymorphism (SNP in CACNA1A confers risk to ASD in Central European population. However, the genetic relationship between autism and CACNA1A in Chinese Han population remains unclear. To explore the association of CACNA1A with autism, we performed a family-based association study. First, we carried out a family-based association test between twelve tagged SNPs and autism in 239 trios. To further confirm the association, the sample size was expanded to 553 trios by recruiting 314 additional trios. In a total of 553 trios, we identified association of rs7249246 and rs12609735 with autism though this would not survive after Bonferroni correction. Our findings suggest that CACNA1A might play a role in the etiology of autism.

  5. Calpains, cleaved mini-dysferlinC72, and L-type channels underpin calcium-dependent muscle membrane repair. (United States)

    Lek, Angela; Evesson, Frances J; Lemckert, Frances A; Redpath, Gregory M I; Lueders, Ann-Katrin; Turnbull, Lynne; Whitchurch, Cynthia B; North, Kathryn N; Cooper, Sandra T


    Dysferlin is proposed as a key mediator of calcium-dependent muscle membrane repair, although its precise role has remained elusive. Dysferlin interacts with a new membrane repair protein, mitsugumin 53 (MG53), an E3 ubiquitin ligase that shows rapid recruitment to injury sites. Using a novel ballistics assay in primary human myotubes, we show it is not full-length dysferlin recruited to sites of membrane injury but an injury-specific calpain-cleavage product, mini-dysferlinC72. Mini-dysferlinC72-rich vesicles are rapidly recruited to injury sites and fuse with plasma membrane compartments decorated by MG53 in a process coordinated by L-type calcium channels. Collective interplay between activated calpains, dysferlin, and L-type channels explains how muscle cells sense a membrane injury and mount a specialized response in the unique local environment of a membrane injury. Mini-dysferlinC72 and MG53 form an intricate lattice that intensely labels exposed phospholipids of injury sites, then infiltrates and stabilizes the membrane lesion during repair. Our results extend functional parallels between ferlins and synaptotagmins. Whereas otoferlin exists as long and short splice isoforms, dysferlin is subject to enzymatic cleavage releasing a synaptotagmin-like fragment with a specialized protein- or phospholipid-binding role for muscle membrane repair.

  6. A Putative Calcium-Permeable Cyclic Nucleotide-Gated Channel, CNGC18, Regulates Polarized Pollen Tube Growth

    Institute of Scientific and Technical Information of China (English)


    A tip-focused Ca2+ gradient is tightly coupled to polarized pollen tube growth, and tip-localized influxes of extracellular Ca2+ are required for this process. However the molecular identity and regulation of the potential Ca2+ channels remains elusive.The present study has implicated CNGC18 (cyclic nucleotide-gated channel 18) in polarized pollen tube growth, because its overexpression induced wider and shorter pollen tubes. Moreover, CNGC18 overexpression induced depolarization of pollen tube growth was suppressed by lower extracellular calcium ([Ca2+]ex). CNGC18-yellow fluorescence protein (YFP)was preferentially localized to the apparent post-Golgi vesicles and the plasma membrane (PM) in the apex of pollen tubes.The PM localization was affected by tip-localized ROP1 signaling. Expression of wild type ROP1 or an active form of ROP1enhanced CNGC18-YFP localization to the apical region of the PM, whereas expression of RopGAP1 (a ROP1 deactivator)blocked the PM localization. These results support a role for PM-localized CNGC18 in the regulation of polarized pollen tube growth through Its potential function in the modulation of calcium influxes.

  7. Cloning and characterization of a novel calcium channel toxin-like gene BmCa1 from Chinese scorpion Mesobuthus martensii Karsch. (United States)

    Zhijian, Cao; Yun, Xie; Chao, Dai; Shunyi, Zhu; Shijin, Yin; Yingliang, Wu; Wenxin, Li


    Many studies have been carried on peptides and genes encoding scorpion toxins from the venom of Mesobuthus martensii Karsch (synonym: Buthus martensii Karsch, BmK), such as Na+, K+ and Cl- channel modulators. In this study, a novel calcium channel toxin-like gene BmCa1 was isolated and characterized from the venom of Mesobuthus martensii Karsch. First, a partial cDNA sequence of the Ca2+ channel toxin-like gene was identified by random sequencing method from a venomous gland cDNA library of Mesobuthus martensii Karsch. The full-length sequence of BmCa1 was then obtained by 5'RACE technique. The peptide deduced from BmCa1 precursor nucleotide sequence contains a 27-residue signal peptide and a 37-residue mature peptide. Although BmCa1 and other scorpion toxins are different at the gene and protein primary structure levels, BmCa1 has the same precursor nucleotide organization and cysteine arrangement as that of the first subfamily members of calcium channel scorpion toxins. Genomic DNA sequence of BmCa1 was also cloned by PCR. Sequence analysis showed that BmCa1 gene consists of three exons separated by two introns of 72 bp and 1076 bp in length, respectively. BmCa1 is the first calcium channel toxin-like gene cloned from the venom of Mesobuthus martensii Karsch and potentially represents a novel class of calcium channel toxins in scorpion venoms.

  8. Splice variants of the CaV1.3 L-type calcium channel regulate dendritic spine morphology (United States)

    Stanika, Ruslan; Campiglio, Marta; Pinggera, Alexandra; Lee, Amy; Striessnig, Jörg; Flucher, Bernhard E.; Obermair, Gerald J.


    Dendritic spines are the postsynaptic compartments of glutamatergic synapses in the brain. Their number and shape are subject to change in synaptic plasticity and neurological disorders including autism spectrum disorders and Parkinson’s disease. The L-type calcium channel CaV1.3 constitutes an important calcium entry pathway implicated in the regulation of spine morphology. Here we investigated the importance of full-length CaV1.3L and two C-terminally truncated splice variants (CaV1.342A and CaV1.343S) and their modulation by densin-180 and shank1b for the morphology of dendritic spines of cultured hippocampal neurons. Live-cell immunofluorescence and super-resolution microscopy of epitope-tagged CaV1.3L revealed its localization at the base-, neck-, and head-region of dendritic spines. Expression of the short splice variants or deletion of the C-terminal PDZ-binding motif in CaV1.3L induced aberrant dendritic spine elongation. Similar morphological alterations were induced by co-expression of densin-180 or shank1b with CaV1.3L and correlated with increased CaV1.3 currents and dendritic calcium signals in transfected neurons. Together, our findings suggest a key role of CaV1.3 in regulating dendritic spine structure. Under physiological conditions it may contribute to the structural plasticity of glutamatergic synapses. Conversely, altered regulation of CaV1.3 channels may provide an important mechanism in the development of postsynaptic aberrations associated with neurodegenerative disorders. PMID:27708393

  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


    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. The Epithelial Calcium Channel TRPV5 Is Regulated Differentially by Klotho and Sialidase.

    NARCIS (Netherlands)

    Leunissen, E.H.P.; Nair, A.V.; Büll, C.; Lefeber, D.J.; Delft, F.L. van; Bindels, R.J.M.; Hoenderop, J.G.J.


    The transient receptor potential vanilloid type 5 (TRPV5) Ca(2+) channel facilitates transcellular Ca(2+) transport in the distal convoluted tubule (DCT) of the kidney. The channel is glycosylated with a complex type N-glycan and it has been postulated that hydrolysis of the terminal sialic acid(s)

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

    DEFF Research Database (Denmark)

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


    Small-conductance Ca2+-activated potassium (SK) channels are relative newcomers within the field of cardiac electrophysiology. In recent years, an increased focus has been given to these channels since they might constitute a relatively atrial selective target. The present review will give...

  12. Anion channels in Chara corallina tonoplast membrane: Calcium dependence and rectification

    NARCIS (Netherlands)

    Berecki, G.; Varga, Z.; Iren, F. van; Duijn, B. van


    Tonoplast K+ channels of Chara corallina are well characterized but only a few reports mention anion channels, which are likely to play an important role in the tonoplast action potential and osmoregulation of this plant. For experiments internodal cells were isolated. Cytoplasmic droplets were form

  13. 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;


    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......, while stress only upregulated Ca(v)1.3 channel expression significantly in the dentate gyrus. ECS effects on Ca(v)1.2 channel expression were generally specific to stressed animals. Our findings are consistent with and extent previous studies on the involvement of intracellular calcium ion dysfunction...

  14. Interactions of drugs and toxins with permeant ions in potassium, sodium, and calcium channels. (United States)

    Zhorov, B S


    Ion channels in cell membranes are targets for a multitude of ligands including naturally occurring toxins, illicit drugs, and medications used to manage pain and treat cardiovascular, neurological, autoimmune, and other health disorders. In the past decade, the x-ray crystallography revealed 3D structures of several ion channels in their open, closed, and inactivated states, shedding light on mechanisms of channel gating, ion permeation and selectivity. However, atomistic mechanisms of the channel modulation by ligands are poorly understood. Increasing evidence suggest that cationophilic groups in ion channels and in some ligands may simultaneously coordinate permeant cations, which form indispensible (but underappreciated) components of respective receptors. This review describes ternary ligand-metal-channel complexes predicted by means of computer-based molecular modeling. The models rationalize a large body of experimental data including paradoxes in structure-activity relationships, effects of mutations on the ligand action, sensitivity of the ligand action to the nature of current-carrying cations, and action of ligands that bind in the ion-permeation pathway but increase rather than decrease the current. Recent mutational and ligand-binding experiments designed to test the models have confirmed the ternary-complex concept providing new knowledge on physiological roles of metal ions and atomistic mechanisms of action of ion channel ligands.

  15. Calmodulin mediates calcium-dependent activation of the intermediate conductance KCa channel, IKCa1. (United States)

    Fanger, C M; Ghanshani, S; Logsdon, N J; Rauer, H; Kalman, K; Zhou, J; Beckingham, K; Chandy, K G; Cahalan, M D; Aiyar, J


    Small and intermediate conductance Ca2+-activated K+ channels play a crucial role in hyperpolarizing the membrane potential of excitable and nonexcitable cells. These channels are exquisitely sensitive to cytoplasmic Ca2+, yet their protein-coding regions do not contain consensus Ca2+-binding motifs. We investigated the involvement of an accessory protein in the Ca2+-dependent gating of hIKCa1, a human intermediate conductance channel expressed in peripheral tissues. Cal- modulin was found to interact strongly with the cytoplasmic carboxyl (C)-tail of hIKCa1 in a yeast two-hybrid system. Deletion analyses defined a requirement for the first 62 amino acids of the C-tail, and the binding of calmodulin to this region did not require Ca2+. The C-tail of hSKCa3, a human neuronal small conductance channel, also bound calmodulin, whereas that of a voltage-gated K+ channel, mKv1.3, did not. Calmodulin co-precipitated with the channel in cell lines transfected with hIKCa1, but not with mKv1. 3-transfected lines. A mutant calmodulin, defective in Ca2+ sensing but retaining binding to the channel, dramatically reduced current amplitudes when co-expressed with hIKCa1 in mammalian cells. Co-expression with varying amounts of wild-type and mutant calmodulin resulted in a dominant-negative suppression of current, consistent with four calmodulin molecules being associated with the channel. Taken together, our results suggest that Ca2+-calmodulin-induced conformational changes in all four subunits are necessary for the channel to open.

  16. Co-localization of putative calcium channels (phenylalkylamine-binding sites) on oil bodies in protoplasts from dark-grown sunflower seedling cotyledons. (United States)

    Vandana, Shweta; Bhatla, Satish C


    Oil bodies are spherical entities containing a triacylglycerol (TAG) matrix encased by a phospholipid monolayer, which is stabilized by oil body-specific proteins, principally oleosins. Biochemical investigations in the recent past have also demonstrated the expression of calcium-binding proteins, called caleosins, as a component of oil body membranes during seed germination. Using DM-Bodipy-phenylalkylamine (PAA; a fluorescent derivative of phenylalkylamine)-a fluorescent probe known to bind L-type calcium channel proteins, present investigations provide the first report on the localization and preferential accumulation of putative calcium channel proteins on/around oil bodies during peak lipolytic phase in protoplasts derived from dark-grown sunflower (Helianthus annuus L. cv Morden) seedling cotyledons. Specificity of DM-Bodipy-PAA labeling was confirmed by using bepridil, a non-fluorescent competitor of PAA while non-specific dye accumulation has been ruled out by using Bodipy-FL as control. Co-localization of fluorescence from DM-Bodipy-PAA binding sites (ex: 504 nm; em: 511 nm) and nile red fluorescing oil bodies (ex: 552 nm; em: 636 nm) has been undertaken by epifluorescence and confocal laser scanning microscopy (CLSM). It revealed the affinity of PAA-sensitive ion channels for the oil body surface. Findings from the current investigations highlight the significance of calcium and calcium channel proteins during oil body mobilization in sunflower.

  17. Effects of in vitro and in vivo lead exposure on voltage-dependent calcium channels in central neurons of Lymnaea stagnalis. (United States)

    Audesirk, G


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

  18. Kinetics of low threshold calcium channels of relay cells in cat lateral geniculate nucleus

    Institute of Scientific and Technical Information of China (English)

    罗茀荪; S.M.Sherman


    Kinetics of the low threshold T-type Ca2+ channel is studied with single electrode voltage damp technique on brain slices of the cat lateral geniculate nucleus (LGN). Space damp is dramatically improved by blocking various K+ and Na+ channels, decreasing Ca2+ current and selecting proper holding potentials. Results from this study are similar to those obtained from acutely dissociated LGN neurons of the rat, indicating that the kinetics of T-Ca2+ channels of the cat LGN neurons is the same as that of the rat LGN. The result reported previously on the cat LGN may result from a defect in space damp.

  19. External bioenergy-induced increases in intracellular free calcium concentrations are mediated by Na+/Ca2+ exchanger and L-type calcium channel. (United States)

    Kiang, Juliann G; Ives, John A; Jonas, Wayne B


    External bioenergy (EBE, energy emitted from a human body) has been shown to increase intracellular calcium concentration ([Ca2+]i, an important factor in signal transduction) and regulate the cellular response to heat stress in cultured human lymphoid Jurkat T cells. In this study, we wanted to elucidate the underlying mechanisms. A bioenergy specialist emitted bioenergy sequentially toward tubes of cultured Jurkat T cells for one 15-minute period in buffers containing different ion compositions or different concentrations of inhibitors. [Ca2+], was measured spectrofluorometrically using the fluorescent probe fura-2. The resting [Ca2+]i in Jurkat T cells was 70 +/- 3 nM (n = 130) in the normal buffer. Removal of external calcium decreased the resting [Ca2+]i to 52 +/- 2 nM (n = 23), indicating that Ca2+ entry from the external source is important for maintaining the basal level of [Ca2+]i. Treatment of Jurkat T cells with EBE for 15 min increased [Ca2+]i by 30 +/- 5% (P EBE did not attenuate [Ca2+]i responsiveness to EBE. Removal of external Ca2+ or Na+, but not Mg2+, inhibited the EBE-induced increase in [Ca2+]i. Dichlorobenzamil, an inhibitor of Na+/Ca2+ exchangers, also inhibited the EBE-induced increase in [Ca2+]i in a concentration-dependent manner with an IC50 of 0.11 +/- 0.02 nM. When external [K+] was increased from 4.5 mM to 25 mM, EBE decreased [Ca2+]i. The EBE-induced increase was also blocked by verapamil, an L-type voltage-gated Ca2+ channel blocker. These results suggest that the EBE-induced [Ca2+]i increase may serve as an objective means for assessing and validating bioenergy effects and those specialists claiming bioenergy capability. The increase in [Ca2+]i is mediated by activation of Na+/Ca2+ exchangers and opening of L-type voltage-gated Ca2+ channels.

  20. Cav1.4 L-Type Calcium Channels Contribute to Calpain Activation in Degenerating Photoreceptors of rd1 Mice.

    Directory of Open Access Journals (Sweden)

    Christian Schön

    Full Text Available Retinitis pigmentosa is an inherited blinding disorder characterized by progressive degeneration and loss of photoreceptors. The exact mechanism of degeneration and cell death of photoreceptors is not known, but is thought to involve disturbed Ca2+-signaling. Ca2+ can enter the photoreceptor cell via outer segment cyclic nucleotide-gated (CNG channels or synaptic Cav1.4 L-type voltage-gated calcium channels (VGCC. Previously, we have shown that genetic ablation of the Cngb1 gene encoding the B subunit of the rod CNG channel delays the fast progressing degeneration in the rd1 mutant mouse model of retinitis pigmentosa. In this study, we crossbred rd1 mice with the Cacna1f-deficient mouse lacking the Cav1.4 α1 subunit of the L-type VGCC. Longitudinal in vivo examinations of photoreceptor layer thickness by optical coherence tomography revealed a significant, but not sustained delay of retinal degeneration in Cacna1f x rd1 double mutant mice compared to rd1 mice. This was accompanied by a reduction of TUNEL positive cells in the early phase of rod degeneration. Remarkably, Cacna1f x rd1 double mutant mice displayed a strong decrease in the activation of the Ca2+-dependent protease calpain during photoreceptor loss. Our results show that genetic deletion of the synaptic Cav1.4 L-type VGCCs impairs calpain activation and leads to a short-term preservation of photoreceptors in the rd1 mouse.

  1. Current view on regulation of voltage-gated sodium channels by calcium and auxiliary proteins. (United States)

    Pitt, Geoffrey S; Lee, Seok-Yong


    In cardiac and skeletal myocytes, and in most neurons, the opening of voltage-gated Na(+) channels (NaV channels) triggers action potentials, a process that is regulated via the interactions of the channels' intercellular C-termini with auxiliary proteins and/or Ca(2+) . The molecular and structural details for how Ca(2+) and/or auxiliary proteins modulate NaV channel function, however, have eluded a concise mechanistic explanation and details have been shrouded for the last decade behind controversy about whether Ca(2+) acts directly upon the NaV channel or through interacting proteins, such as the Ca(2+) binding protein calmodulin (CaM). Here, we review recent advances in defining the structure of NaV intracellular C-termini and associated proteins such as CaM or fibroblast growth factor homologous factors (FHFs) to reveal new insights into how Ca(2+) affects NaV function, and how altered Ca(2+) -dependent or FHF-mediated regulation of NaV channels is perturbed in various disease states through mutations that disrupt CaM or FHF interaction.

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

    Institute of Scientific and Technical Information of China (English)

    Joo Hyun Nam; Hyo Won Jung; Young-Won Chin; Woo Kyung Kim; Hyo Sang Bae


    Objective: To examine the effects of Tribulus terrestris L. (T. terrestris) extract on the modulation of calcium channels to evaluate its use in topical agents for treatment of atopic dermatitis. Methods: The 70% methanol extract of T. terrestris was prepared. Human HEK293T cells with over-expressed calcium release-activated calcium channel protein 1 (Orai1), transient receptor potential vanilloid 1, or transient receptor potential vanilloid 3 (TRPV3) were treated with T. terrestris extract. Modulation of ion channels was measured using a conventional whole-cell patch-clamp technique. Results: T. terrestris extract (100 mg/mL) significantly inhibited Orai1 activity in Orai1-stromal interaction molecule 1 co-overexpressed HEK293T cells. In addition, T. terrestris extract significantly increased the TRPV3 activity compared with 2-Aminoethyl diphe-nylborinate (100 mmol/L), which induces the full activation of TRPV3. Conclusions: Our results suggest that T. terrestris extract may have a therapeutic po-tential 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.

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

    Institute of Scientific and Technical Information of China (English)

    Joo Hyun Nam; Hyo Won Jung; Young-Won Chin; Woo Kyung Kim; Hyo Sang Bae


    Objective: To examine the effects of Tribulus terrestris L.(T. terrestris) extract on the modulation of calcium channels to evaluate its use in topical agents for treatment of atopic dermatitis.Methods: The 70% methanol extract of T. terrestris was prepared. Human HEK293 T cells with over-expressed calcium release-activated calcium channel protein 1(Orai1),transient receptor potential vanilloid 1, or transient receptor potential vanilloid 3(TRPV3)were treated with T. terrestris extract. Modulation of ion channels was measured using a conventional whole-cell patch-clamp technique.Results: T. terrestris extract(100 mg/m L) significantly inhibited Orai1 activity in Orai1-stromal interaction molecule 1 co-overexpressed HEK293 T cells. In addition, T. terrestris extract significantly increased the TRPV3 activity compared with 2-Aminoethyl diphenylborinate(100 mmol/L), which induces the full activation of TRPV3.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.

  4. Effects of in vitro lead exposure on voltage-sensitive calcium channels differ among cell types in central neurons of Lymnaea stagnalis. (United States)

    Audesirk, G; Audesirk, T


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

  5. 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

  6. Nitric oxide-induced calcium release: activation of type 1 ryanodine receptor, a calcium release channel, through non-enzymatic posttranslational modification by nitric oxide

    Directory of Open Access Journals (Sweden)

    Sho eKakizawa


    Full Text Available Nitric oxide (NO is a typical gaseous messenger involved in a wide range of biological processes. In our classical knowledge, effects of NO are largely achieved by activation of soluble guanylyl cyclase to form cyclic guanosine-3’, 5’-monophosphate. However, emerging evidences have suggested another signaling mechanism mediated by NO: S-nitrosylation of target proteins.S-nitrosylation is a covalent addition of an NO group to a cysteine thiol/sulfhydryl (RSH, and categorized into non-enzymatic posttranslational modification of proteins, contrasted to enzymatic posttranslational modification of proteins, such as phosphorylation mediated by various protein kinases.Very recently, we found novel intracellular calcium (Ca2+ mobilizing mechanism, NO-induced Ca2+ release (NICR in cerebellar Purkinje cells. NICR is mediated by type 1 ryanodine receptor (RyR1, a Ca2+ release channel expressed in endoplasmic-reticular membrane. Furthermore, NICR is indicated to be dependent on S-nitrosylation of RyR1, and involved in synaptic plasticity in the cerebellum. In this review, molecular mechanisms and functional significance of NICR, as well as non-enzymatic posttranslational modification of proteins by gaseous signals, are described.

  7. Sodium-calcium exchanger and multiple sodium channel isoforms in intra-epidermal nerve terminals

    Directory of Open Access Journals (Sweden)

    Gasser Andreas


    Full Text Available Abstract Background Nociception requires transduction and impulse electrogenesis in nerve fibers which innervate the body surface, including the skin. However, the molecular substrates for transduction and action potential initiation in nociceptors are incompletely understood. In this study, we examined the expression and distribution of Na+/Ca2+ exchanger (NCX and voltage-gated sodium channel isoforms in intra-epidermal free nerve terminals. Results Small diameter DRG neurons exhibited robust NCX2, but not NCX1 or NCX3 immunolabeling, and virtually all PGP 9.5-positive intra-epidermal free nerve terminals displayed NCX2 immunoreactivity. Sodium channel NaV1.1 was not detectable in free nerve endings. In contrast, the majority of nerve terminals displayed detectable levels of expression of NaV1.6, NaV1.7, NaV1.8 and NaV1.9. Sodium channel immunoreactivity in the free nerve endings extended from the dermal boundary to the terminal tip. A similar pattern of NCX and sodium channel immunolabeling was observed in DRG neurons in vitro. Conclusions NCX2, as well as NaV1.6, NaV1.7, NaV1.8 and NaV1.9, are present in most intra-epidermal free nerve endings. The presence of NCX2, together with multiple sodium channel isoforms, in free nerve endings may have important functional implications.

  8. Vasorelaxant and antihypertensive effects of methanolic extract from roots of Laelia anceps are mediated by calcium-channel antagonism. (United States)

    Vergara-Galicia, Jorge; Ortiz-Andrade, Rolffy; Rivera-Leyva, Julio; Castillo-España, Patricia; Villalobos-Molina, Rafael; Ibarra-Barajas, Maximiliano; Gallardo-Ortiz, Itzell; Estrada-Soto, Samuel


    RMELanc-induced relaxation in aortic rings precontracted with NE, 5-HT and KCl. It also reduced NE-induced transient contraction in Ca(2+)-free solution and inhibited contraction induced by increasing external calcium. Nevertheless, the vasorelaxant effect of RMELanc was not reduced by ODQ, 1-alprenolol, TEA, glibenclamide, and 2-AP. Oral administration of 100 mg/kg of RMELanc exhibited a significant decrease in systolic and diastolic blood pressures in SHR rats. HPLC analysis allowed us to detect the presence of 2,7-dihydroxy-3,4,9-trimethoxyphenantrene (1), which induced a significant relaxation effect. Therefore, our results suggest that RMELanc induces vasorelaxant and antihypertensive effects by blockade of Ca(2+) channels.

  9. 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


    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...... of which contribute to clopidogrel metabolic activation. This interaction may diminish the efficacy of clopidogrel. Methods All patients surviving 30 days after a first-time MI in the period 2000 to 2006 in Denmark were identified by individual-level linkage of nationwide administrative registers....... The cohort was divided into patients treated with and without clopidogrel and followed for 1 year after discharge. The risk of a composite of cardiovascular death, MI, or stroke and the risk of the individual components of the composite end point and all-cause death associated with CCBs were analyzed...

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

    Directory of Open Access Journals (Sweden)

    Ben eAviner


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

  11. The secret life of CFTR as a calcium-activated chloride channel. (United States)

    Billet, Arnaud; Hanrahan, John W


    cAMP-stimulated anion conductance is defective in cystic fibrosis (CF). The regulatory domain of CFTR, the anion channel protein encoded by the CF gene, possesses an unusually high density of consensus sequences for phosphorylation by protein kinase A (14 in a stretch of CFTR is viewed primarily as a cAMP-stimulated anion channel, and most studies have focused on this mode of activation. However, there is growing evidence that CFTR also responds to Ca(2+)-mobilizing secretagogues and contributes substantially to cholinergic and purinergic responses in native tissues. G protein-coupled receptors that signal through Gαq can stimulate CFTR channels by activating Ca(2+)-dependent adenylyl cyclase and tyrosine kinases, and also by inhibiting protein phosphatase type 2A. Here we review evidence for these novel mechanisms of CFTR activation and discuss how they may help explain previous observations.

  12. Inhibition of voltage-gated calcium channels as common mode of action for (mixtures of) distinct classes of insecticides. (United States)

    Meijer, Marieke; Dingemans, Milou M L; van den Berg, Martin; Westerink, Remco H S


    Humans are exposed to distinct structural classes of insecticides with different neurotoxic modes of action. Because calcium homeostasis is essential for proper neuronal function and development, we investigated the effects of insecticides from different classes (pyrethroid: (α-)cypermethrin; organophosphate: chlorpyrifos; organochlorine: endosulfan; neonicotinoid: imidacloprid) and mixtures thereof on the intracellular calcium concentration ([Ca(2+)]i). Effects of acute (20 min) exposure to (mixtures of) insecticides on basal and depolarization-evoked [Ca(2+)]i were studied in vitro with Fura-2-loaded PC12 cells and high resolution single-cell fluorescence microscopy. The data demonstrate that cypermethrin, α-cypermethrin, endosulfan, and chlorpyrifos concentration-dependently decreased depolarization-evoked [Ca(2+)]i, with 50% (IC50) at 78nM, 239nM, 250nM, and 899nM, respectively. Additionally, acute exposure to chlorpyrifos or endosulfan (10μM) induced a modest increase in basal [Ca(2+)]i, amounting to 68 ± 8nM and 53 ± 8nM, respectively. Imidacloprid did not disturb basal or depolarization-evoked [Ca(2+)]i at 10μM. Following exposure to binary mixtures, effects on depolarization-evoked [Ca(2+)]i were within the expected effect additivity range, whereas the effect of the tertiary mixture was less than this expected additivity effect range. These results demonstrate that different types of insecticides inhibit depolarization-evoked [Ca(2+)]i in PC12 cells by inhibiting voltage-gated calcium channels (VGCCs) in vitro at concentrations comparable with human occupational exposure levels. Moreover, the effective concentrations in this study are below those for earlier described modes of action. Because inhibition of VGCCs appears to be a common and potentially additive mode of action of several classes of insecticides, this target should be considered in neurotoxicity risk assessment studies.

  13. Presynaptic Localization and Possible Function of Calcium-Activated Chloride Channel Anoctamin 1 in the Mammalian Retina.

    Directory of Open Access Journals (Sweden)

    Ji Hyun Jeon

    Full Text Available Calcium (Ca(2+-activated chloride (Cl(- channels (CaCCs play a role in the modulation of action potentials and synaptic responses in the somatodendritic regions of central neurons. In the vertebrate retina, large Ca(2+-activated Cl(- currents (ICl(Ca regulate synaptic transmission at photoreceptor terminals; however, the molecular identity of CaCCs that mediate ICl(Ca remains unclear. The transmembrane protein, TMEM16A, also called anoctamin 1 (ANO1, has been recently validated as a CaCC and is widely expressed in various secretory epithelia and nervous tissues. Despite the fact that tmem16a was first cloned in the retina, there is little information on its cellular localization and function in the mammalian retina. In this study, we found that ANO1 was abundantly expressed as puncta in 2 synaptic layers. More specifically, ANO1 immunoreactivity was observed in the presynaptic terminals of various retinal neurons, including photoreceptors. ICl(Ca was first detected in dissociated rod bipolar cells expressing ANO1. ICl(Ca was abolished by treatment with the Ca(2+ channel blocker Co(2+, the L-type Ca(2+ channel blocker nifedipine, and the Cl(- channel blockers 5-nitro-2-(3-phenylpropylamino benzoic acid (NPPB and niflumic acid (NFA. More specifically, a recently discovered ANO1-selective inhibitor, T16Ainh-A01, and a neutralizing antibody against ANO1 inhibited ICl(Ca in rod bipolar cells. Under a current-clamping mode, the suppression of ICl(Ca by using NPPB and T16Ainh-A01 caused a prolonged Ca(2+ spike-like depolarization evoked by current injection in dissociated rod bipolar cells. These results suggest that ANO1 confers ICl(Ca in retinal neurons and acts as an intrinsic regulator of the presynaptic membrane potential during synaptic transmission.

  14. Chronic fluoxetine treatment increases NO bioavailability and calcium-sensitive potassium channels activation in rat mesenteric resistance arteries. (United States)

    Pereira, Camila A; Ferreira, Nathanne S; Mestriner, Fabiola L; Antunes-Rodrigues, José; Evora, Paulo R B; Resstel, Leonardo B M; Carneiro, Fernando S; Tostes, Rita C


    Fluoxetine, a selective serotonin reuptake inhibitor (SSRI), has effects beyond its antidepressant properties, altering, e.g., mechanisms involved in blood pressure and vasomotor tone control. Although many studies have addressed the acute impact of fluoxetine on the cardiovascular system, there is a paucity of information on the chronic vascular effects of this SSRI. We tested the hypothesis that chronic fluoxetine treatment enhances the vascular reactivity to vasodilator stimuli by increasing nitric oxide (NO) signaling and activation of potassium (K+) channels. Wistar rats were divided into two groups: (I) vehicle (water for 21 days) or (II) chronic fluoxetine (10 mg/kg/day in the drinking water for 21 days). Fluoxetine treatment increased endothelium-dependent and independent vasorelaxation (analyzed by mesenteric resistance arteries reactivity) as well as constitutive NO synthase (NOS) activity, phosphorylation of eNOS at Serine1177 and NO production, determined by western blot and fluorescence. On the other hand, fluoxetine treatment did not alter vascular expression of neuronal and inducible NOS or guanylyl cyclase (GC). Arteries from fluoxetine-treated rats exhibited increased relaxation to pinacidil. Increased acetylcholine vasorelaxation was abolished by a calcium-activated K+ channel (KCa) blocker, but not by an inhibitor of KATP channels. On the other hand, vascular responses to Bay 41-2272 and 8-bromo-cGMP were similar between the groups. In conclusion, chronic fluoxetine treatment increases endothelium-dependent and independent relaxation of mesenteric resistance arteries by mechanisms that involve increased eNOS activity, NO generation, and KCa channels activation. These effects may contribute to the cardiovascular effects associated with chronic fluoxetine treatment.

  15. Lipid Storage Disorders Block Lysosomal Trafficking By Inhibiting TRP Channel and Calcium Release



    Lysosomal lipid accumulation, defects in membrane trafficking, and altered Ca2+ homeostasis are common features in many lysosomal storage diseases. Mucolipin TRP channel 1 (TRPML1) is the principle Ca2+ channel in the lysosome. Here we show that TRPML1-mediated lysosomal Ca2+ release, measured using a genetically-encoded Ca2+ indicator (GCaMP3) attached directly to TRPML1 and elicited by a potent membrane-permeable synthetic agonist, is dramatically reduced in Niemann-Pick (NP) disease cells....

  16. Effect of subclinical, clinical and supraclinical doses of calcium channel blockers on models of drug-induced hepatotoxicity in rats. (United States)

    Okwa, Iniviefien B; Akindele, Abidemi J; Agbaje, Esther O; Oshinuga, Oladoyin T; Anunobi, Chidozie C; Adeyemi, Olufunmilayo O


    Drug-related hepatotoxicity is the leading cause of acute liver failure, and hepatic problems are responsible for a significant number of liver transplantations and deaths worldwide. Calcium has been associated with various metabolic processes that lead to cell death and apoptosis, and increased cytosolic Ca(2+) has been implicated in hepatotoxicity. This study was designed to investigate the effects of calcium channel blockers (CCBs) on isoniazid-rifampicin, zidovudine and erythromycin-induced hepatotoxicity in rats. Treatment groups comprised control, hepatotoxicant, hepatotoxicant along with each of silymarin, nifedipine, verapamil and diltiazem at subclinical, clinical and supraclinical doses. A day to the end of treatment for each model, rats were subjected to the hexobarbitone-induced hypnosis test. On the last days of treatment, blood samples were collected and serum was analyzed for relevant biochemical parameters. Animals were sacrificed after blood collection and livers were harvested, and samples obtained for in vivo antioxidant indices assay and histopathology. The hepatotoxicants significantly increased serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP), as well as duration of sleep in the hypnosis test. These drugs significantly reduced the hepatic levels of reduced glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and increased the level of malondialdehyde (MDA). The CCBs at the various doses significantly reversed the effects of isoniazid-rifampicin, zidovudine and erythromycin. The results obtained in this study suggest that the CCBs possess hepatoprotective activity in drug-induced hepatotoxicity and may be beneficial at the subclinical and clinical doses.

  17. Effects and mechanisms of store-operated calcium channel blockade on hepatic ischemia-reperfusion injury in rats

    Institute of Scientific and Technical Information of China (English)

    Li-Jie Pan; Zi-Chao Zhang; Zhen-Ya Zhang; Wen-Jun Wang; Yue Xu; Zong-Ming Zhang


    AIM: To further investigate the important role of storeoperated calcium channels (SOCs) in rat hepatocytes and to explore the effects of SOC blockers on hepatic ischemia-reperfusion injury (HIRI). METHODS: Using freshly isolated hepatocytes from a rat model of HIRI (and controls), we measured cytosolic free Ca2+ concentration (by calcium imaging), net Ca2+ fluxes (by a non-invasive micro-test technique), the SOC current (ISOC; by whole-cell patch-clamp recording), and taurocholate secretion [by high-performance liquid chromatography and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays]. RESULTS: Ca2+ oscillations and net Ca2+ fluxes mediated by Ca2+ entry via SOCs were observed in rat hepatocytes. ISOC was significantly higher in HIRI groups than in controls (57.0 ± 7.5 pA vs 31.6 ± 2.7 pA, P <0.05) and was inhibited by La3+. Taurocholate secretion by hepatocytes into culture supernatant was distinctly lower in HIRI hepatocytes than in controls, an effect reversed by SOC blockers. CONCLUSION: SOCs are pivotal in HIRI. SOC blockers protected against HIRI and assisted the recovery of secretory function in hepatocytes. Thus, they are likely to become a novel class of effective drugs for prevention or therapy of HIRI patients in the future.

  18. A systematic review of calcium channel antagonists in bipolar disorder and some considerations for their future development (United States)

    Cipriani, A; Saunders, K; Attenburrow, M-J; Stefaniak, J; Panchal, P; Stockton, S; Lane, T A; Tunbridge, E M; Geddes, J R; Harrison, P J


    l-type calcium channel (LTCC) antagonists have been used in bipolar disorder for over 30 years, without becoming an established therapeutic approach. Interest in this class of drugs has been rekindled by the discovery that LTCC genes are part of the genetic aetiology of bipolar disorder and related phenotypes. We have therefore conducted a systematic review of LTCC antagonists in the treatment and prophylaxis of bipolar disorder. We identified 23 eligible studies, with six randomised, double-blind, controlled clinical trials, all of which investigated verapamil in acute mania, and finding no evidence that it is effective. Data for other LTCC antagonists (diltiazem, nimodipine, nifedipine, methyoxyverapamil and isradipine) and for other phases of the illness are limited to observational studies, and therefore no robust conclusions can be drawn. Given the increasingly strong evidence for calcium signalling dysfunction in bipolar disorder, the therapeutic candidacy of this class of drugs has become stronger, and hence we also discuss issues relevant to their future development and evaluation. In particular, we consider how genetic, molecular and pharmacological data can be used to improve the selectivity, efficacy and tolerability of LTCC antagonists. We suggest that a renewed focus on LTCCs as targets, and the development of ‘brain-selective' LTCC ligands, could be one fruitful approach to innovative pharmacotherapy for bipolar disorder and related phenotypes. PMID:27240535

  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


    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.


    Directory of Open Access Journals (Sweden)

    J Nandhakumar


    Full Text Available The role of PDE-4 inhibitor etazolate, was evaluated in the presence of PDE-7 inhibitor, BRL-50481, in animal models of epilepsy. Seizures were induced in the animals by subjecting them to injection of chemical convulsants, Pilocarpine, Kainic acid (KA and maximal electroshock (MES. The combination of etazolate and BRL50481 treated mice showed a significant (P<0.001 quick onset of action, jerky movements and convulsion when compared to gabapentin. The combination of etazolate and sGC inhibitor, methylene blue (MB treated mice showed a significant (P<0.001 delay in onset of action, jerky movements and convulsion when compare to gabapentin as well as against the combination of etazolate with BRL 50481.The present study mainly highlights the individual effects of etazolate and combination with BRL-50481 potentiates (P<0.001 the onset of seizure activity against all models of convulsion. The study mainly comprises the onset of seizures, mortality/recovery, percentage of prevention of seizures (anticonvulsant and total duration of convulsive time. The total convulsive time was prolonged significantly (P<0.05 and P<0.01 in combination of methylene blue with etazolate treated (28.59% and 35.15 % groups, compared to DMSO received group (100% in the MES model. In the same way, the combination of calcium channel modulator (CCM and calcium channel blocker (CCB amiodarone and nifedipine respectively, with etazolate showed a significant (P<0.001 delay in onset of seizures, compared to DMSO and etazolate treated groups in all models of epilepsy. This confirms that both CCM and CCB possess anticonvulsant activity. Finally, the study reveals that identification of new cAMP mediated phosphodiesterases family members offers a potential new therapy for epilepsy management in future.

  1. Design, Synthesis and Structure-activity of N-Glycosyl-1-pyridyl-1H-pyrazole-5-carboxamide as Inhibitors of Calcium Channels

    Institute of Scientific and Technical Information of China (English)

    ZHOU Yun-yun; LI Yu-xin; LI Yi-ming; YANG Xiao-ping; MAO Ming-zhen; LI Zheng-ming


    Carbohydrates,with broad-spectrum structures and biological functions,are key organic compounds in nature,along with nucleic acids and proteins.As part of our ongoing efforts to develop a new class of pesticides with novel mechanism of action,a series of novel N-glycosyl-l-pyridyl-lH-pyrazole-5-carboxamide was designed and synthesized via the reactions of glycosyl methanamides and pyridyl-pyrazole acid.The compounds were characterized by 1H NMR and 13C NMR.The bioassay results indicate that some of these compounds exhibit moderate insecticidal activities and assessed as potential inhibitors of calcium channels.The modulation of voltage-gated calcium channels by compounds 4a and 5a in the central neurons isolated from the third instar larvae of Spodoptera exigua was studied by whole-cell patch-clamp technique.In addition,compound 5a inhibits the recorded calcium currents reversible on washout.Experimental results also indicate that compound 5a did not release stored calcium from the Endoplasmic Reticulum.The present work demonstrates that N-glycosyl-l-pyridyl-lH-pyrazole-5-carboxamides cannot be used as possible inhibitors of calcium channels for developing novel pesticides.

  2. Inhibitory effects of calcium channel blockers on thyroid hormone uptake in neonatal rat cardiomyocytes

    NARCIS (Netherlands)

    F.A. Verhoeven; E.P.C.M. Moerings (Ellis); J.M.J. Lamers (Jos); G. Hennemann; T.J. Visser (Theo); M.E. Everts (Maria)


    textabstractThe effects of the Ca2+ channel blockers verapamil, nifedipine, and diltiazem on triiodothyronine (T3) and thyroxine (T4) uptake were tested in cultured cardiomyocytes from 2-day-old rats. Experiments were performed at 37 degrees C in medium with 0.5% BSA for [125I]T3 (

  3. Shikonin inhibits intestinal calcium-activated chloride channels and prevents rotaviral diarrhea

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    Yu Jiang


    Full Text Available Secretory diarrhea remains a global health burden and causes major mortality in children. There have been some focuses on antidiarrheal therapies that may reduce fluid losses and intestinal motility in diarrheal diseases. In the present study, we identified shikonin as an inhibitor of TMEM16A chloride channel activity using cell-based fluorescent-quenching assay. The IC50 value of shikonin was 6.5 μM. Short-circuit current measurements demonstrated that shikonin inhibited Eact-induced Cl current in a dose-dependent manner, with IC50 value of 1.5 μM. Short-circuit current measurement showed that shikonin exhibited inhibitory effect against CCh-induced Cl currents in mouse colonic epithelia but did not affect cytoplasmic Ca2+ concentration as well as the other major enterocyte chloride channel CFTR. Characterization study found that shikonin inhibited basolateral K+ channel activity without affecting Na+/K+-ATPase activities. In-vivo studies revealed that shikonin significantly delayed intestinal motility in mice and reduced stool water content in a neonatal mice model of rotaviral diarrhea without affecting the viral infection process in-vivo. Taken together, the results suggested that shikonin inhibited enterocyte CaCCs, the inhibitory effect was partially through inhbition of basolateral K+ channel acitivty, and shikonin could be a lead compound in the treatment of rotaviral secretory diarrhea.

  4. Clusters of calcium release channels harness the Ising phase transition to confine their elementary intracellular signals

    CERN Document Server

    Maltsev, Anna; Stern, Michael


    Intracellular Ca signals represent a universal mechanism of cell function. Messages carried by Ca are local, rapid, and powerful enough to be delivered over the thermal noise. A higher signal to noise ratio is achieved by a cooperative action of Ca release channels such as IP3 receptors or ryanodine receptors arranged in clusters or release units containing a few to several hundred release channels. The release channels synchronize their openings via Ca-induced-Ca-release, generating high-amplitude local Ca signals known as puffs in neurons or sparks in muscle cells. Despite the high release amplitude and positive feedback nature of the activation, Ca signals are strictly confined in time and space by an unexplained termination mechanism. Here we show that the collective transition of release channels from an open to a closed state is identical to the phase transition associated with the reversal of magnetic field in an Ising ferromagnet. We demonstrate this mechanism using numerical model simulations of Ca s...

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

    NARCIS (Netherlands)

    Majoie, H.J.; Baets, M.H.V. de; Renier, W.O.; Lang, B.; Vincent, A.


    OBJECTIVE: To determine the prevalence of antibodies to ion channels in patients with long standing epilepsy. BACKGROUND: 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,

  6. Calcium Channels: Structure and Function (Annals of the New York Academy of Sciences. Volume 560) (United States)


    8033 Planegg, Federal Republic of Germany c Dipartimento di Anatomia e Fisiologia Umana Corso Raffaello 30 1-10125 Torino, Italy INTRODUCTION In...4313-4317. I Modulation of Ca Channels in Peripheral Neuronsa E. CARBONE Dipardmento di Anatomia e Fisiologia Umana Corso Raffaello 30 1-10125 Torino

  7. Nitric oxide and L-type calcium channel influences the changes in arterial blood pressure and heart rate induced by central angiotesin II

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    Guarda Ismael FMS


    Full Text Available Abstract We study the voltage dependent calcium channels and nitric oxide involvement in angiotensin II-induced pressor effect. The antipressor action of L-Type calcium channel antagonist, nifedipine, has been studied when it was injected into the third ventricle prior to angiotensin II. The influence of nitric oxide on nifedipine antipressor action has also been studied by utilizing NW-nitro-L-arginine methyl ester (LNAME (40 μg/0.2 μl a nitric oxide synthase inhibitor and L-arginine (20 μg/0.2 μl, a nitric oxide donor agent. Adult male Holtzman rats weighting 200–250 g, with cannulae implanted into the third ventricle were injected with angiotensin II. Angiotensin II produced an elevation in mean arterial pressure and a decreased in heart rate. Such effects were potentiated by the prior injection of LNAME. L-arginine and nifedipine blocked the effects of angiotensin II. These data showed the involvement of L-Type calcium channel and a free radical gas nitric oxide in the central control of angiotensin II-induced pressor effect. This suggested that L-Type calcium channel of the circunventricular structures of central nervous system participated in both short and long term neuronal actions of ANG II with the influence of nitrergic system.

  8. The L-Type Voltage-Gated Calcium Channel Ca[subscript v]1.3 Mediates Consolidation, but Not Extinction, of Contextually Conditioned Fear in Mice (United States)

    McKinney, Brandon C.; Murphy, Geoffrey G.


    Using pharmacological techniques, it has been demonstrated that both consolidation and extinction of Pavlovian fear conditioning are dependent to some extent upon L-type voltage-gated calcium channels (LVGCCs). Although these studies have successfully implicated LVGCCs in Pavlovian fear conditioning, they do not provide information about the…

  9. Inhibition of voltage-gated calcium channels after subchronic and repeated exposure of PC12 cells to different classes of insecticides

    NARCIS (Netherlands)

    Meijer, Marieke; Brandsema, Joske A R; Nieuwenhuis, Desirée; Wijnolts, Fiona M J; Dingemans, Milou M L; Westerink, Remco H S


    We previously demonstrated that acute inhibition of voltage-gated calcium channels (VGCCs) is a common mode of action for (sub)micromolar concentrations of chemicals, including insecticides. However, since human exposure to chemicals is usually chronic and repeated, we investigated if selected insec

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

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    Niko Bekjarovski


    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.

  11. Like Extinction, Latent Inhibition of Conditioned Fear in Mice Is Blocked by Systemic Inhibition of L-Type Voltage-Gated Calcium Channels (United States)

    Blouin, Ashley M.; Cain, Chris K.; Barad, Mike


    Having recently shown that extinction of conditioned fear depends on L-type voltage-gated calcium channels (LVGCCs), we have been seeking other protocols that require this unusual induction mechanism. We tested latent inhibition (LI) of fear, because LI resembles extinction except that cue exposures precede, rather than follow, cue-shock pairing.…

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

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


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

  13. Calcium competes with zinc for a channel mechanism on the brush border membrane of piglet intestine. (United States)

    Bertolo, R F.P.; Bettger, W J.; Atkinson, S A.


    Interactions between Ca(+2) and Zn(+2) at the intestinal brush border membrane occur via unclear mechanisms. We hypothesized that Zn(+2) and Ca(+2) are transported across the brush border membrane via a multidivalent metal channel. Using brush border membrane vesicles (BBMV) prepared from intestines of 8 sow-fed piglets, we sought to determine whether Ca(+2) competes with Zn(+2) for uptake. Extravesicular Zn(+2) was removed with ethylenediamine-tetraacetic acid. Time curves of Zn(+2) and Ca(+2) uptake by BBMV were conducted with increasing concentrations of Ca(+2) and Zn(+2), respectively. Saturation curves compared kinetic parameters of Zn(+2) uptake with and without Ca(+2). In addition, Zn(+2) uptake was measured in the presence of various classical Ca(+2) channel modulators. Over 20 min, a 0.4x concentration of Zn(+2) lowered Ca(+2) uptake by vesicles, whereas a 30x concentration of Ca(+2) was necessary to lower Zn(+2) uptake. These data suggest that Ca(+2) has lower affinity than Zn(+2) for a brush border membrane transport protein. Kinetic parameters showed higher K(m) values with 4 or 15 mM Ca(+2) but unchanged J(max), suggesting competitive inhibition. The Ca(+2) channel blocking agents, La(+3), Ba(+2), verapamil, and diltiazem, inhibited Zn(+2) uptake, whereas calcitriol, trans 1,2 cyclohexanediol, cis/trans 1,3 cyclohexanediol, and the L-type Ca(+2) channel agonist, Bay K8644, induced Zn(+2) uptake. These data were consistent with competition for a common transport mechanism on the brush border membrane, possibly a novel multimetal channel. Copyright 2001 Elsevier Science Inc.

  14. Expression of calcium-activated chloride channels Ano1 and Ano2 in mouse taste cells. (United States)

    Cherkashin, Alexander P; Kolesnikova, Alisa S; Tarasov, Michail V; Romanov, Roman A; Rogachevskaja, Olga A; Bystrova, Marina F; Kolesnikov, Stanislav S


    Specialized Ca(2+)-dependent ion channels ubiquitously couple intracellular Ca(2+) signals to a change in cell polarization. The existing physiological evidence suggests that Ca(2+)-activated Cl(-) channels (CaCCs) are functional in taste cells. Because Ano1 and Ano2 encode channel proteins that form CaCCs in a variety of cells, we analyzed their expression in mouse taste cells. Transcripts for Ano1 and Ano2 were detected in circumvallate (CV) papillae, and their expression in taste cells was confirmed using immunohistochemistry. When dialyzed with CsCl, taste cells of the type III exhibited no ion currents dependent on cytosolic Ca(2+). Large Ca(2+)-gated currents mediated by TRPM5 were elicited in type II cells by Ca(2+) uncaging. When TRPM5 was inhibited by triphenylphosphine oxide (TPPO), ionomycin stimulated a small but resolvable inward current that was eliminated by anion channel blockers, including T16Ainh-A01 (T16), a specific Ano1 antagonist. This suggests that CaCCs, including Ano1-like channels, are functional in type II cells. In type I cells, CaCCs were prominently active, blockable with the CaCC antagonist CaCCinh-A01 but insensitive to T16. By profiling Ano1 and Ano2 expressions in individual taste cells, we revealed Ano1 transcripts in type II cells only, while Ano2 transcripts were detected in both type I and type II cells. P2Y agonists stimulated Ca(2+)-gated Cl(-) currents in type I cells. Thus, CaCCs, possibly formed by Ano2, serve as effectors downstream of P2Y receptors in type I cells. While the role for TRPM5 in taste transduction is well established, the physiological significance of expression of CaCCs in type II cells remains to be elucidated.

  15. Calcium release channel RyR2 regulates insulin release and glucose homeostasis. (United States)

    Santulli, Gaetano; Pagano, Gennaro; Sardu, Celestino; Xie, Wenjun; Reiken, Steven; D'Ascia, Salvatore Luca; Cannone, Michele; Marziliano, Nicola; Trimarco, Bruno; Guise, Theresa A; Lacampagne, Alain; Marks, Andrew R


    The type 2 ryanodine receptor (RyR2) is a Ca2+ release channel on the endoplasmic reticulum (ER) of several types of cells, including cardiomyocytes and pancreatic β cells. In cardiomyocytes, RyR2-dependent Ca2+ release is critical for excitation-contraction coupling; however, a functional role for RyR2 in β cell insulin secretion and diabetes mellitus remains controversial. Here, we took advantage of rare RyR2 mutations that were identified in patients with a genetic form of exercise-induced sudden death (catecholaminergic polymorphic ventricular tachycardia [CPVT]). As these mutations result in a "leaky" RyR2 channel, we exploited them to assess RyR2 channel function in β cell dynamics. We discovered that CPVT patients with mutant leaky RyR2 present with glucose intolerance, which was heretofore unappreciated. In mice, transgenic expression of CPVT-associated RyR2 resulted in impaired glucose homeostasis, and an in-depth evaluation of pancreatic islets and β cells from these animals revealed intracellular Ca2+ leak via oxidized and nitrosylated RyR2 channels, activated ER stress response, mitochondrial dysfunction, and decreased fuel-stimulated insulin release. Additionally, we verified the effects of the pharmacological inhibition of intracellular Ca2+ leak in CPVT-associated RyR2-expressing mice, in human islets from diabetic patients, and in an established murine model of type 2 diabetes mellitus. Taken together, our data indicate that RyR2 channels play a crucial role in the regulation of insulin secretion and glucose homeostasis.

  16. Cyclic-AMP regulation of calcium-dependent K channels in an insect central neurone. (United States)

    David, J A; Pitman, R M


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

  17. Parathyroid Hormone and the Use of Diuretics and Calcium-Channel Blockers: The Multi-Ethnic Study of Atherosclerosis. (United States)

    Zaheer, Sarah; de Boer, Ian; Allison, Matthew; Brown, Jenifer M; Psaty, Bruce M; Robinson-Cohen, Cassianne; Ix, Joachim H; Kestenbaum, Bryan; Siscovick, David; Vaidya, Anand


    Thiazide diuretic (TZ) use is associated with higher bone mineral density, whereas loop diuretic (LD) use is associated with lower bone density and incident fracture. Dihydropyridine-sensitive calcium channels are expressed on parathyroid cells and may play a role in parathyroid hormone (PTH) regulation. The potential for diuretics and calcium-channel blockers (CCBs) to modulate PTH and calcium homeostasis may represent a mechanism by which they influence skeletal outcomes. We hypothesized that the use of LD and dihydropyridine CCBs is associated with higher PTH, and TZ use is associated with lower PTH. We conducted cross-sectional analyses of participants treated for hypertension in the Multi-Ethnic Study of Atherosclerosis who did not have primary hyperparathyroidism or chronic kidney disease (n = 1888). We used adjusted regression models to evaluate the independent association between TZ, LD, and CCB medication classes and PTH. TZ use was associated with lower PTH when compared with non-TZ use (44.4 versus 46.9 pg/mL, p = 0.02), whereas the use of LD and CCBs was associated with higher PTH when compared with non-users of each medication class (LD: 60.7 versus 45.5 pg/mL, p < 0.0001; CCB: 49.5 versus. 44.4 pg/mL, p < 0.0001). Adjusted regression models confirmed independent associations between TZ use and lower PTH (β = -3.2 pg/mL, p = 0.0007), and LD or CCB use and higher PTH (LD: β = +12.0 pg/mL, p < 0.0001; CCB: +3.7 pg/mL, p < 0.0001). Among CCB users, the use of dihydropyridines was independently associated with higher PTH (β = +5.0 pg/mL, p < 0.0001), whereas non-dihydropyridine use was not (β = +0.58 pg/mL, p = 0.68). We conclude that in a large community-based cohort with normal kidney function, TZ use is associated with lower PTH, whereas LD and dihydropyridine CCB use is associated with higher PTH. These associations may provide a mechanistic explanation linking use of these

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

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    Marcella Araugio Soares


    Full Text Available Pituitary adenomas constitute about 6-18% of brain tumours in adults. Activation of voltage gated calcium currents can account for growth hormone oversecretion in some GH-secreting pituitary adenomas that produce an acromegaly appearance and increase mortality. Ca2+ ions, as mediators of intracellular signalling, are crucial for the development of apoptosis. However, the role of [Ca2+] in the development of apoptosis is ambiguous. In this study, the effects of low-dose ionising gamma radiation (60Co 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.Adenomas de pituitária constituem cerca de 6-18% dos tumores cerebrais em adultos. A ativação de correntes de cálcio dependentes de voltagem podem levar à super-excreção de hormônio do crescimento produzindo acromegalia e aumentando a mortalidade. Íons Ca2+ como mediadores de sinalização intracelular são cruciais no desenvolvimento da apoptose. No entanto, o papel da [Ca 2+] no desenvolvimento da apoptose é ambíguo. Neste estudo nós avaliamos os efeitos de baixas doses de radiação gama (60Co na sobrevivência e proliferação de células de adenoma de pituitária de rato e o papel do cálcio na apoptose radio-induzida. Nossos resultados mostraram que a dose de 3Gy foi suficiente para inibir a proliferação das células GH3. Apesar de existir um número significativo de células vivas após 168 horas do tratamento com radiação, elas não estavam aptas a proliferar. Nossos resultados também indicaram que bloqueio do influxo de cálcio extracelular n

  19. Molecular pharmacology of the calcium channel: evidence for subtypes, multiple drug-receptor sites, channel subunits, and the development of a radioiodinated 1,4-dihydropyridine calcium channel label, (/sup 125/I)iodipine

    Energy Technology Data Exchange (ETDEWEB)

    Glossmann, H.; Ferry, D.R.; Goll, A.; Rombusch, M.


    Radiolabeled Ca2+ antagonists (1,4-dihydropyridines, verapamil, and D-cis-diltiazem) were used to study voltage-operated Ca2+ channels in different excitable tissues. The concept of three subtypes of Ca2+ channels, represented by brain, heart, and skeletal-muscle isoreceptors for 1,4-dihydropyridines, is developed. The three subtypes are characterized by a variety of criteria. Despite the biochemical differences between the subtypes, they have the same Mr in situ by target-size analysis (Mr approximately equal to 180,000, when evaluated by (/sub 3/H)nimodipine). The concept of the metalloprotein nature of the channel and the interaction of channel drugs with the Me2+ binding sites of the ionic pore is demonstrated. Distinct but interacting drug-receptor sites of the Ca2+ channel are found by direct labeling as well as indirectly by drug competition studies. The authors distinguish between the 1,4-dihydropyridine site, the verapamil site, and the D-cis-diltiazem site. Each receptor site can exist in high and low-affinity state; the distribution of receptor sites in these states is regulated by temperature, ions, and drugs. The concept of intrinsic activity of drugs to stabilize the high-affinity state is exemplified for the 1,4-dihydropyridines. A change in the channel architecture is induced by binding of D-cis-diltiazem to its drug receptor site. This is proven by target-size analysis of the channel in situ. Partially purified t-tubule membranes from skeletal muscle are an extremely rich source of Ca2+ channel drug-receptor sites. The stoichiometry was determined in this preparation and found to be four verapamil:two 1,4-dihydropyridine:one D-cis-diltiazem site. A novel Ca2+ channel probe, (/sup 125/I)iodipine (2,200 Ci/mmol), was synthetized, and the properties of this ligand are presented.

  20. 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)


    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.)

  1. Calcium-permeable ion channels involved in glutamate receptor-independent ischemic brain injury

    Institute of Scientific and Technical Information of China (English)

    Ming-hua LI; Koichi INOUE; Hong-fang SI; Zhi-gang XIONG


    Brain ischemia is a leading cause of death and long-term disabilities worldwide. Unfortunately, current treatment is limited to thrombolysis, which has limited success and a potential side effect of intracerebral hemorrhage. Searching for new cell injury mechanisms and therapeutic interventions has become a major challenge in the field. It has been recognized for many years that intracellular Ca2+overload in neurons is essential for neuronal injury associated with brain ischemia. However, the exact pathway(s) underlying the toxic Ca2+ loading remained elusive. This review discusses the role of two Ca2+-permeable cation channels, TRPM7 and acid-sensing channels, in glutamate-independent Ca2+ toxicity associated with brain ischemia.

  2. Transient receptor potential melastatin 4 channel controls calcium signals and dental follicle stem cell differentiation. (United States)

    Nelson, Piper; Ngoc Tran, Tran Doan; Zhang, Hanjie; Zolochevska, Olga; Figueiredo, Marxa; Feng, Ji-Ming; Gutierrez, Dina L; Xiao, Rui; Yao, Shaomian; Penn, Arthur; Yang, Li-Jun; Cheng, Henrique


    Elevations in the intracellular Ca(2+) concentration are a phenomena commonly observed during stem cell differentiation but cease after the process is complete. The transient receptor potential melastatin 4 (TRPM4) is an ion channel that controls Ca(2+) signals in excitable and nonexcitable cells. However, its role in stem cells remains unknown. The aim of this study was to characterize TRPM4 in rat dental follicle stem cells (DFSCs) and to determine its impact on Ca(2+) signaling and the differentiation process. We identified TRPM4 gene expression in DFSCs, but not TRPM5, a closely related channel with similar function. Perfusion of cells with increasing buffered Ca(2+) resulted in a concentration-dependent activation of currents typical for TRPM4, which were also voltage-dependent and had Na(+) conductivity. Molecular suppression with shRNA decreased channel activity and cell proliferation during osteogenesis but not adipogenesis. As a result, enhanced mineralization and phosphatase enzyme activity were observed during osteoblast formation, although DFSCs failed to differentiate into adipocytes. Furthermore, the normal agonist-induced first and secondary phases of Ca(2+) signals were transformed into a gradual and sustained increase which confirmed the channels' ability to control Ca(2+) signaling. Using whole genome microarray analysis, we identified several genes impacted by TRPM4 during DFSC differentiation. These findings suggest an inhibitory role for TRPM4 on osteogenesis while it appears to be required for adipogenesis. The data also provide a potential link between the Ca(2+) signaling pattern and gene expression during stem cell differentiation.

  3. Micromolar-Affinity Benzodiazepine Receptors Regulate Voltage-Sensitive Calcium Channels in Nerve Terminal Preparations (United States)

    Taft, William C.; Delorenzo, Robert J.


    Benzodiazepines in micromolar concentrations significantly inhibit depolarization-sensitive Ca2+ uptake in intact nerve-terminal preparations. Benzodiazepine inhibition of Ca2+ uptake is concentration dependent and stereospecific. Micromolar-affinity benzodiazepine receptors have been identified and characterized in brain membrane and shown to be distinct from nanomolar-affinity benzodiazepine receptors. Evidence is presented that micromolar, and not nanomolar, benzodiazepine binding sites mediate benzodiazepine inhibition of Ca2+ uptake. Irreversible binding to micromolar benzodiazepine binding sites also irreversibly blocked depolarization-dependent Ca2+ uptake in synaptosomes, indicating that these compounds may represent a useful marker for identifying the molecular components of Ca2+ channels in brain. Characterization of benzodiazepine inhibition of Ca2+ uptake demonstrates that these drugs function as Ca2+ channel antagonists, because benzodiazepines effectively blocked voltage-sensitive Ca2+ uptake inhibited by Mn2+, Co2+, verapamil, nitrendipine, and nimodipine. These results indicate that micromolar benzodiazepine binding sites regulate voltage-sensitive Ca2+ channels in brain membrane and suggest that some of the neuronal stabilizing effects of micromolar benzodiazepine receptors may be mediated by the regulation of Ca2+ conductance.

  4. Lipid storage disorders block lysosomal trafficking by inhibiting a TRP channel and lysosomal calcium release. (United States)

    Shen, Dongbiao; Wang, Xiang; Li, Xinran; Zhang, Xiaoli; Yao, Zepeng; Dibble, Shannon; Dong, Xian-ping; Yu, Ting; Lieberman, Andrew P; Showalter, Hollis D; Xu, Haoxing


    Lysosomal lipid accumulation, defects in membrane trafficking and altered Ca(2+) homoeostasis are common features in many lysosomal storage diseases. Mucolipin transient receptor potential channel 1 (TRPML1) is the principle Ca(2+) channel in the lysosome. Here we show that TRPML1-mediated lysosomal Ca(2+) release, measured using a genetically encoded Ca(2+) indicator (GCaMP3) attached directly to TRPML1 and elicited by a potent membrane-permeable synthetic agonist, is dramatically reduced in Niemann-Pick (NP) disease cells. Sphingomyelins (SMs) are plasma membrane lipids that undergo sphingomyelinase (SMase)-mediated hydrolysis in the lysosomes of normal cells, but accumulate distinctively in lysosomes of NP cells. Patch-clamp analyses revealed that TRPML1 channel activity is inhibited by SMs, but potentiated by SMases. In NP-type C cells, increasing TRPML1's expression or activity was sufficient to correct the trafficking defects and reduce lysosome storage and cholesterol accumulation. We propose that abnormal accumulation of luminal lipids causes secondary lysosome storage by blocking TRPML1- and Ca(2+)-dependent lysosomal trafficking.

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

    Parkash, Jai


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

  6. Rapid changes in skeletal muscle calcium uptake induced in vitro by 1,25-dihydroxyvitamin D/sub 3/ are suppressed by calcium channel blockers

    Energy Technology Data Exchange (ETDEWEB)

    de Boland, A.R.; Boland, R.L.


    Previous investigations have shown that 1,25-dihydroxyvitamin D/sub 3/ (1,25-(OH)/sub 2/D/sub 3/) stimulates muscle Ca uptake through a nuclear mechanism. The possibility that 1,25-(OH)/sub 2/D/sub 3/ would induce rapid changes in muscle Ca fluxes independent of de novo protein synthesis was investigated in the present work. In vitro preparations of soleus muscles obtained from vitamin D-deficient chicks were used. A significant increase in /sup 45/Ca labeling of the tissue was already observed after 3-min treatment with 2.4 X 10(-10) M 1,25-(OH)/sub 2/D/sub 3/. This early stimulation in muscle Ca uptake became maximal at 10-15 min. Cycloheximide (50 microM) did not block the effect of the metabolite at 15 and 30 min. However, the antibiotic effectively blocked the increase in Ca uptake induced by 1,25-(OH)/sub 2/D/sub 3/ after 1-h treatment. The rapid 1,25-(OH)/sub 2/D/sub 3/-dependent stimulation of /sup 45/Ca labeling of soleus muscle was not associated to changes in lipid synthesis as assessed by measurements of /sup 3/H-glycerol incorporation into the tissue lipids. However, the calcium antagonists verapamil and nifedipine (50 microM) abolished the stimulation in Ca uptake produced by 1,25-(OH)/sub 2/D/sub 3/ in 5 min. These results suggest that 1,25-(OH)/sub 2/D/sub 3/ can act directly at the muscle membrane level affecting Ca fluxes through Ca channels.

  7. Converging roles of ion channels, calcium, metabolic stress, and activity pattern of Substantia nigra dopaminergic neurons in health and Parkinson's disease. (United States)

    Duda, Johanna; Pötschke, Christina; Liss, Birgit


    Dopamine-releasing neurons within the Substantia nigra (SN DA) are particularly vulnerable to degeneration compared to other dopaminergic neurons. The age-dependent, progressive loss of these neurons is a pathological hallmark of Parkinson's disease (PD), as the resulting loss of striatal dopamine causes its major movement-related symptoms. SN DA neurons release dopamine from their axonal terminals within the dorsal striatum, and also from their cell bodies and dendrites within the midbrain in a calcium- and activity-dependent manner. Their intrinsically generated and metabolically challenging activity is created and modulated by the orchestrated function of different ion channels and dopamine D2-autoreceptors. Here, we review increasing evidence that the mechanisms that control activity patterns and calcium homeostasis of SN DA neurons are not only crucial for their dopamine release within a physiological range but also modulate their mitochondrial and lysosomal activity, their metabolic stress levels, and their vulnerability to degeneration in PD. Indeed, impaired calcium homeostasis, lysosomal and mitochondrial dysfunction, and metabolic stress in SN DA neurons represent central converging trigger factors for idiopathic and familial PD. We summarize double-edged roles of ion channels, activity patterns, calcium homeostasis, and related feedback/feed-forward signaling mechanisms in SN DA neurons for maintaining and modulating their physiological function, but also for contributing to their vulnerability in PD-paradigms. We focus on the emerging roles of maintained neuronal activity and calcium homeostasis within a physiological bandwidth, and its modulation by PD-triggers, as well as on bidirectional functions of voltage-gated L-type calcium channels and metabolically gated ATP-sensitive potassium (K-ATP) channels, and their probable interplay in health and PD. We propose that SN DA neurons possess several feedback and feed-forward mechanisms to protect and adapt

  8. COOH-terminal association of human smooth muscle calcium channel Ca(v)1.2b with Src kinase protein binding domains: effect of nitrotyrosylation. (United States)

    Kang, Minho; Ross, Gracious R; Akbarali, Hamid I


    The carboxyl terminus of the calcium channel plays an important role in the regulation of calcium entry, signal transduction, and gene expression. Potential protein-protein interaction sites within the COOH terminus of the L-type calcium channel include those for the SH3 and SH2 binding domains of c-Src kinase that regulates calcium currents in smooth muscle. In this study, we examined the binding sites involved in Src kinase-mediated phosphorylation of the human voltage-gated calcium channel (Ca(v)) 1.2b (hCav1.2b) and the effect of nitrotyrosylation. Cotransfection of human embryonic kidney (HEK)-293 cells with hCa(v)1.2b and c-Src resulted in tyrosine phosphorylation of the calcium channel, which was prevented by nitration of tyrosine residues by peroxynitrite. Whole cell calcium currents were reduced by 58 + 5% by the Src kinase inhibitor PP2 and 64 + 6% by peroxynitrite. Nitrotyrosylation prevented Src-mediated regulation of the currents. Glutathione S-transferase fusion protein of the distal COOH terminus of hCa(v)1.2b (1809-2138) bound to SH2 domain of Src following tyrosine phosphorylation, while binding to SH3 required the presence of the proline-rich motif. Site-directed mutation of Y(2134) prevented SH2 binding and resulted in reduced phosphorylation of hCa(v)1.2b. Within the distal COOH terminus, single, double, or triple mutations of Y(1837), Y(1861), and Y(2134) were constructed and expressed in HEK-293 cells. The inhibitory effects of PP2 and peroxynitrite on calcium currents were significantly reduced in the double mutant Y(1837-2134F). These data demonstrate that the COOH terminus of hCa(v)1.2b contains sites for the SH2 and SH3 binding of Src kinase. Nitrotyrosylation of these sites prevents Src kinase regulation and may be importantly involved in calcium influx regulation during inflammation.

  9. 17β-estradiol regulation of T-type calcium channels in gonadotropin-releasing hormone neurons (United States)

    Zhang, Chunguang; Bosch, Martha A.; Rick, Elizabeth A.; Kelly, Martin J.; Ronnekleiv, Oline K.


    T-type calcium channels are responsible for generating low-threshold spikes that facilitate burst firing and neurotransmitter release in neurons. GnRH neurons exhibit burst firing, but the underlying conductances are not known. Previously, we have found that 17β-estradiol (E2) increases T-type channel expression and excitability of hypothalamic arcuate nucleus neurons. Therefore, we used ovariectomized oil- or E2-treated EGFP-GnRH mice to explore the expression and E2-regulation of T-type channels in GnRH neurons. Based on single cell RT-PCR and real-time PCR quantification of the T-type channel α1-subunits, we found that all three subunits were expressed in GnRH neurons with Cav3.3≥Cav3.2>Cav3.1. The mRNA expression of the three subunits was increased with surge-inducing levels of E2 during the morning. During the afternoon, Cav3.3 mRNA expression remained elevated, whereas Cav3.1 and Cav3.2 were decreased. The membrane estrogen receptor agonist STX increased the expression of Cav3.3, but not Cav3.2 in GnRH neurons. Whole-cell patch recordings in GnRH neurons revealed that E2 treatment significantly augmented T-type current density at both time-points, and increased the rebound excitation during the afternoon. Although E2 regulated the mRNA expression of all three subunits in GnRH neurons, the increased expression combined with the slower inactivation kinetics of the T-type current indicates that Cav3.3 may be the most important for bursting activity associated with the GnRH/LH surge. The E2-induced increase in mRNA expression, which depends in part on membrane-initiated signaling, leads to increased channel function and neuronal excitability, and could be a mechanism by which E2 facilitates burst firing and cyclic GnRH neurosecretion. PMID:19710308

  10. Calcium-dependent potassium channels play a critical role for burst termination in the locomotor network in lamprey. (United States)

    el Manira, A; Tegnér, J; Grillner, S


    1. The possible involvement of calcium-dependent potassium channels (KCa) in the termination of locomotor bursts was investigated by administration of a specific blocker, apamin, in the lamprey spinal cord in vitro. The effects were examined by recording the efferent activity in ventral roots and by intracellular recording from interneurons and motoneurons. During fictive locomotion induced by N-methyl-D-aspartate (NMDA), apamin was found to affect both the frequency of bursting and the regularity of the locomotor pattern. 2. At the single cell level, NMDA can induce pacemaker-like membrane potential oscillations in individual neurons after administration of tetrodotoxin. Apamin (2.5 microM) produced a marked increase of the duration of the depolarizing plateau phase occurring during these NMDA-induced oscillations; this shows that the repolarization of the plateau is initiated by a progressive activation of apamin-sensitive KCa-channels. 3. The action potential is followed by an afterhyperpolarization (AHP) with a fast and a slow phase (sAHP). The latter is known to be caused by apamin-sensitive KCa-channels. During repetitive firing, the interspike interval is dependent on the amplitude and the duration of the sAHP. Apamin caused a reduction of the spike frequency adaptation with a concomitant increase in the firing frequency. In some cells, apamin in addition reduced the threshold for the action potential. Apamin-sensitive KCa-channels thus will be involved in controlling both the onset and the duration of neuronal firing in the lamprey spinal cord. 4. During fictive locomotion induced by NMDA (40-200 microM), a blockade of KCa-channels by apamin produced an increase of the coefficient of variation (mean = 167%, n = 26), which was statistically significant in 21 out of 26 experiments. At 40-150 microM NMDA, an average increase in cycle duration was 77% and statistically significant in 15 out of 20 preparations. At 200 microM NMDA (corresponding to higher burst

  11. Calcium-release channels in paramecium. Genomic expansion, differential positioning and partial transcriptional elimination.

    Directory of Open Access Journals (Sweden)

    Eva-Maria Ladenburger

    Full Text Available The release of Ca²⁺ from internal stores is a major source of signal Ca²⁺ in almost all cell types. The internal Ca²⁺ pools are activated via two main families of intracellular Ca²⁺-release channels, the ryanodine and the inositol 1,4,5-trisphosphate (InsP₃ receptors. Among multicellular organisms these channel types are ubiquitous, whereas in most unicellular eukaryotes the identification of orthologs is impaired probably due to evolutionary sequence divergence. However, the ciliated protozoan Paramecium allowed us to prognosticate six groups, with a total of 34 genes, encoding proteins with characteristics typical of InsP₃ and ryanodine receptors by BLAST search of the Paramecium database. We here report that these Ca²⁺-release channels may display all or only some of the characteristics of canonical InsP₃ and ryanodine receptors. In all cases, prediction methods indicate the presence of six trans-membrane regions in the C-terminal domains, thus corresponding to canonical InsP₃ receptors, while a sequence homologous to the InsP₃-binding domain is present only in some types. Only two types have been analyzed in detail previously. We now show, by using antibodies and eventually by green fluorescent protein labeling, that the members of all six groups localize to distinct organelles known to participate in vesicle trafficking and, thus, may provide Ca²⁺ for local membrane-membrane interactions. Whole genome duplication can explain radiation within the six groups. Comparative and evolutionary evaluation suggests derivation from a common ancestor of canonical InsP₃ and ryanodine receptors. With one group we could ascertain, to our knowledge for the first time, aberrant splicing in one thoroughly analyzed Paramecium gene. This yields truncated forms and, thus, may indicate a way to pseudogene formation. No comparable analysis is available for any other, free-living or parasitic/pathogenic protozoan.

  12. The effects of calcium channel blockade on agouti-induced obesity

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jung Han; Moustaid, N.; Zemel, M.B. [Univ. of Tennessee, Knoxville, TN (United States)] [and others


    We have previously observed that obese viable yellow (A{sup vy}/a) mice exhibit increased intracellular Ca{sup 2+} ([Ca{sup 2+}]i) and fatty acid synthase (FAS) gene expression; further, recombinant agouti protein increases in cultured adipocytes and these effects are inhibited by Ca{sup 2+} channel blockade. Accordingly, we determined the effect of Ca{sup 2+} channel blockade (nifedipine for 4 wk) on FAS and obesity in transgenic mice expressing the agouti gene in a ubiquitous manner. The transgenic mice initially were significantly heavier (30.5 {+-} 0.6 vs. 27.3 {+-} 0.3 g; P<0.001) and exhibited a 0.81{degrees}C lower initial core temperature (P<0.0005), an approximately twofold increase in fat pad weights (P=0.002), a sevenfold increase in adipose FAS activity (P=0.009), and a twofold increase in plasma insulin level (P<0.05) compared to control mice. Nifedipine treatment resulted in an 18% decrease in fat pad weights (P<0.007) and a 74% decrease in adipose FAS activity (P=0.03), normalized circulating insulin levels and insulin sensitivity (P,0.05), and transiently elevated core temperature in the transgenic mice, but was without effect in the control mice. These data suggest that agouti regulates FAS, fat storage, and possibly thermogenesis, at least partially, via a [Ca{sup 2+}]{sub i}-dependent mechanism, and that Ca{sup 2+} channel blockade may partially attenuate agouti-induced obesity. 42 refs., 4 figs., 1 tab.

  13. Transient Receptor Potential Melastatin 4 channel controls calcium signals and dental follicle stem cell differentiation



    Elevations in the intracellular Ca2+ concentration are a phenomena commonly observed during stem cell differentiation but cease after the process is complete. The Transient Receptor Potential Melastatin 4 (TRPM4) is an ion channel that controls Ca2+ signals in excitable and non-excitable cells. However, its role in stem cells remains unknown. The aim of this study was to characterize TRPM4 in rat dental follicle stem cells (DFSCs) and to determine its impact on Ca2+ signaling and the differen...

  14. Calcium release channel RyR2 regulates insulin release and glucose homeostasis


    Santulli, Gaetano; Pagano, Gennaro; Sardu, Celestino; Xie, Wenjun; Reiken, Steven; D’Ascia, Salvatore Luca; Cannone, Michele; Marziliano, Nicola; Trimarco, Bruno; Guise, Theresa A.; Lacampagne, Alain; Marks, Andrew R.


    The type 2 ryanodine receptor (RyR2) is a Ca2+ release channel on the endoplasmic reticulum (ER) of several types of cells, including cardiomyocytes and pancreatic β cells. In cardiomyocytes, RyR2-dependent Ca2+ release is critical for excitation-contraction coupling; however, a functional role for RyR2 in β cell insulin secretion and diabetes mellitus remains controversial. Here, we took advantage of rare RyR2 mutations that were identified in patients with a genetic form of exercise-induced...

  15. Two distinct pools of large-conductance calcium-activated potassium channels in the somatic plasma membrane of central principal neurons (United States)

    Kaufmann, W.A.; Kasugai, Y.; Ferraguti, F.; Storm, J.F.


    Although nerve cell membranes are often assumed to be uniform with respect to electrical properties, there is increasing evidence for compartmentalization into subdomains with heterogeneous impacts on the overall cell function. Such microdomains are characterized by specific sets of proteins determining their functional properties. Recently, clustering of large-conductance calcium-activated potassium (BKCa) channels was shown at sites of subsurface membrane cisterns in cerebellar Purkinje cells (PC), where they likely participate in building a subcellular signaling unit, the 'PLasmERosome'. By applying SDS-digested freeze-fracture replica labeling (SDS-FRL) and postembedding immunogold electron microscopy, we have now studied the spatial organization of somatic BKCa channels in neocortical layer 5 pyramidal neurons, principal neurons of the central and basolateral amygdaloid nuclei, hippocampal pyramidal neurons and dentate gyrus (DG) granule cells to establish whether there is a common organizational principle in the distribution of BKCa channels in central principal neurons. In all cell types analyzed, somatic BKCa channels were found to be non-homogenously distributed in the plasma membrane, forming two pools of channels with one pool consisting of clustered channels and the other of scattered channels in the extrasynaptic membrane. Quantitative analysis by means of SDS-FRL revealed that about two-thirds of BKCa channels belong to the scattered pool and about one-third to the clustered pool in principal cell somata. Overall densities of channels in both pools differed in the different cell types analyzed, although being considerably lower compared to cerebellar PC. Postembedding immunogold labeling revealed association of clustered channels with subsurface membrane cisterns and confirmed extrasynaptic localization of scattered channels. This study indicates a common organizational principle for somatic BKCa channels in central principal neurons with the

  16. 生物活性肽促钙吸收机制与钙离子通道%Mechanism of promoting calcium absorption by bioactive peptide and calcium ion channel

    Institute of Scientific and Technical Information of China (English)

    郭丹郡; 侯焘; 何慧


    ABSTRACT:In recent years, the fourth generation of calcium supplements-calcium binding peptide (such as casein phosphopeptide) has become a research hotspot, which plays an important role in promoting calcium absorption through the calcium ion channels. The calcium ion channels are a group of transmembrane proteins, which control the process of Ca2+ into the cell. TRPV6 and Cav1.3 are the 2 kinds of important calcium ion channels in intestinal calcium absorption. TRPV6 is a high selectivity of Ca2+ transport channel in TRPV ion channel family, which is regulated by Calbindin-D9k and vitamin D. Cav1.3 is effective in promoting the absorption of calcium under the condition of depolarization, which does not depend on the regulation of Calbindin-D9k and vitamin D. PepT1 is a kind of co-transporter with low affinity and high capacity. It can not only eliminate the competition among amino acids, accelerate protein synthesis, but also promote the absorption and utilization of minerals. This paper summarized the interaction between bioactive peptides and 2 kinds of common calcium ion channels (TRPV6 and Cav1.3), and peptide transporter system (PepT1), so as to clarify the mechanism of promoting calcium uptake by bioactive peptide.%近年来,第4代补钙产品——促钙吸收肽(如酪蛋白磷酸肽)的研究已成为热点,其主要是通过钙离子通道发挥促钙吸收作用。钙离子通道是一组跨细胞膜的蛋白质,它控制着 Ca2+进入细胞的过程, TRPV6和Cav1.3是钙在肠道跨膜吸收的2种重要钙离子通道。TRPV6是TRPV离子通道家族成员中高选择性的Ca2+转运通道,由钙结合蛋白D9k(Calbindin-D9k)和维生素D共同参与调节。Cav1.3在去极化条件下被激活,从而发挥重要的促钙吸收作用,但其不依赖于 Calbindin-D9k和维生素 D 的调控。小肽转运蛋白是一种低亲和力、高容量的协同转运载体,其不仅可以消除氨基酸之间的相互竞争,加速蛋白合

  17. Preadmission Use of Calcium Channel Blockers and Outcomes After Hospitalization With Pneumonia: A Retrospective Propensity-Matched Cohort Study. (United States)

    Zheng, Lin; Hunter, Krystal; Gaughan, John; Poddar, Sameer

    In sepsis, an overwhelming immune response, as mediated by the release of various inflammatory mediators, can lead to shock, multiple organ damage, and even death. Pneumonia is the leading cause of sepsis. In animal septic models, sepsis could induce uncontrolled calcium (Ca) leaking, raising cytosolic Ca to a toxic level, causing irreversible cellular injuries and organ failure. All types of calcium channel blockers (CCBs), by inhibiting Ca influx, have been shown to decrease overall mortality in various septic animal models. However, to our best knowledge, no clinical study had been conducted to investigate the beneficial effect(s) of CCBs in sepsis. We conducted a retrospective propensity-matched cohort study after screening 2214 patients hospitalized for pneumonia from year 2012 to 2014 at our institution. We identified 387 preadmission CCB users and 387 nonusers by propensity score matching. Logistic regression analysis was then used to determine the association between preadmission CCB use and outcomes in pneumonia. Our study showed that the odds for development of severe sepsis was significantly lower in the CCB user group [odds ratio (OR), 0.466; 95% confidence interval (CI), 0.311-0.697; P = 0.002]. Preadmission CCB use was associated with a lower risk of contracting bacteremia (OR, 0.498; 95% CI, 0.262-0.99; P = 0.0327), lower risk of acute respiratory insufficiency (OR, 0.573; 95% CI, 0.412-0.798; P = 0.001), lower risk of intensive care unit admission (OR, 0.602; 95% CI, 0.432-0.840; P = 0.0028). In conclusion, our study suggested preadmission CCB use was associated with a reduction in the risks of development of respiratory insufficiency, bacteremia, and severe sepsis in patients admitted to the hospital with pneumonia.

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

    Directory of Open Access Journals (Sweden)

    Sarah McDavid

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

  19. Butanol isomers exert distinct effects on voltage-gated calcium channel currents and thus catecholamine secretion in adrenal chromaffin cells. (United States)

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


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

  20. The calcium channel blocker amlodipine exerts its anti-proliferative action via p21(Waf1/Cip1) gene activation. (United States)

    Ziesche, Rolf; Petkov, Ventzislav; Lambers, Christopher; Erne, Paul; Block, Lutz-Henning


    Proliferation of vascular smooth muscle cells (VSMC) contributes to the progression of atherosclerotic plaques. Calcium channel blockers have been shown to reduce VSMC proliferation, but the underlying molecular mechanism remains unclear. p21(Waf1/Cip1) is a potent inhibitor of cell cycle progression. Here, we demonstrate that amlodipine (10(-6) to 10(-8) M) activates de novo synthesis of p21(Waf1/Cip1) in vitro. We show that amlodipine-dependent activation of p21(Waf1/Cip1) involves the action of the glucocorticoid receptor (GR) and C/EBP-alpha. The underlying pathway apparently involves the action of mitogen-activated protein kinase or protein kinase C, but not of extracellular signal-related kinase or changes of intracellular calcium. Amlodipine-induced p21(Waf1/Cip1) promoter activity and expression were abrogated by C/EBP-alpha antisense oligonucleotide or by the GR antagonist RU486. Amlodipine-dependent inhibition of cell proliferation was partially reversed by RU486 at 10(-8) M (58%+/-29%), antisense oligonucleotides targeting C/EBP-alpha (91%+/-26%), or antisense mRNAs targeting p21(Waf1/Cip1) (96%+/-32%, n=6); scrambled antisense oligonucleotides or those directed against C/EBP-beta were ineffective. The data suggest that the anti-proliferative action of amlodipine is achieved by induction of the p21 (Waf1/Cip1) gene, which may explain beneficial covert effects of this widely used cardiovascular therapeutic drug beyond a more limited role as a vascular relaxant.

  1. Pharmacokinetics of barnidipine hydrochloride, a new dihydropyridine calcium channel blocker, in the rat, dog and human. (United States)

    Teramura, T; Watanabe, T; Higuchi, S; Hashimoto, K


    1. The pharmacokinetics of a new calcium antagonist barnidipine hydrochloride, a stereochemically pure enantiomer, was studied after intravenous and oral dosing to the rat and dog, and oral to man. 2. After intravenous dosing, plasma concentrations of barnidipine hydrochloride declined bi-exponentially with the terminal half-lives of 0.6 h in the rat and 4.1 h in the dog. The blood clearance was 5.2 l/h/kg in the rat and 3.3 l/h/kg in the dog, and was comparable with hepatic blood flow in both species. 3. After oral dosing, plasma concentrations of barnidipine hydrochloride peaked rapidly (0.3-0.4 h in the rat and dog, 1.0-1.6 h in man). Cmax and AUC rose non-linearly with increasing doses in all three species. 4. The absolute bioavailability was low (11-18% in the rat and 6-9% in the dog), suggesting a marked first-pass metabolism.

  2. Synapsin II desynchronizes neurotransmitter release at inhibitory synapses by interacting with presynaptic calcium channels. (United States)

    Medrihan, Lucian; Cesca, Fabrizia; Raimondi, Andrea; Lignani, Gabriele; Baldelli, Pietro; Benfenati, Fabio


    In the central nervous system, most synapses show a fast mode of neurotransmitter release known as synchronous release followed by a phase of asynchronous release, which extends over tens of milliseconds to seconds. Synapsin II (SYN2) is a member of the multigene synapsin family (SYN1/2/3) of synaptic vesicle phosphoproteins that modulate synaptic transmission and plasticity, and are mutated in epileptic patients. Here we report that inhibitory synapses of the dentate gyrus of Syn II knockout mice display an upregulation of synchronous neurotransmitter release and a concomitant loss of delayed asynchronous release. Syn II promotes γ-aminobutyric acid asynchronous release in a Ca(2+)-dependent manner by a functional interaction with presynaptic Ca(2+) channels, revealing a new role in synaptic transmission for synapsins.

  3. L-type calcium channel blockers enhance 5-HTP-induced antinociception in mice

    Institute of Scientific and Technical Information of China (English)

    Jian-hui LIANG; Jun-xu LI; Xu-hua WANG; Bi CHEN; Ying LU; Pan ZHANG; Rong HAN; Xiang-feng YE


    AIM: To investigate the involvement of L-type Ca2+ channels in antinociceptive action induced by the 5-HT precursor,5-hydroxytryptophan (5-HTP). METHODS: Female Kunming mice were treated with either 5-HTP (20-80 mg/kg,ip) alone, or the combination of 5-HTP and fluoxetine (2-8 mg/kg, ip), pargyline (15-60 mg/kg, ip), nimodipine (2.5-10 mg/kg, ip), nifedipine (2.5-10 mg/kg, ip), verapamil (2.5-10 mg/kg, ip), CaC12 (5-20 mmol/L, icv), or EGTA (0.5-3 mmol/L, icv) prior to the hot-plate test (55 ℃, hind-paw licking latency). In addition, locomotor activity in mice treated with 5-HTP alone was measured using an ambulometer with five activity boxes. RESULTS: Ip injection of 5-HTP alone had no influence on the spontaneous locomotor activity, whereas dose-dependently increased the latency to licking hind-paw in the hot-plate test in mice. The inhibitory effects of 5-HTP on nociceptive response were significantly enhanced by fluoxetine in the mouse hot-plate test. At a sub-effective dose, pargyline could cause a leftward shift in the dose-response curve of 5-HTP-induced antinociception. Co-administration with 5-HTP and nimodipine, nifedipine, or verapamil obviously potentiated the antinociceptive effects elicited by 5-HTP.Interestingly, 5-HTP-induced antinociception was antagonized by CaC12 and enhanced by EGTA injected icv in the mouse hot-plate test. CONCLUSION: These findings suggest that systemic administration of 5-HTP may yield the antinociceptive effects, which are related to Ca2+ influx from extracellular fluid through L-type Ca2+ channels.

  4. TMEM16A is associated with voltage-gated calcium channels in mouse retina and its function is disrupted upon mutation of the auxiliary α2δ4 subunit (United States)

    Caputo, Antonella; Piano, Ilaria; Demontis, Gian Carlo; Bacchi, Niccolò; Casarosa, Simona; Santina, Luca Della; Gargini, Claudia


    Photoreceptors rely upon highly specialized synapses to efficiently transmit signals to multiple postsynaptic targets. Calcium influx in the presynaptic terminal is mediated by voltage-gated calcium channels (VGCC). This event triggers neurotransmitter release, but also gates calcium-activated chloride channels (TMEM), which in turn regulate VGCC activity. In order to investigate the relationship between VGCC and TMEM channels, we analyzed the retina of wild type (WT) and Cacna2d4 mutant mice, in which the VGCC auxiliary α2δ4 subunit carries a nonsense mutation, disrupting the normal channel function. Synaptic terminals of mutant photoreceptors are disarranged and synaptic proteins as well as TMEM16A channels lose their characteristic localization. In parallel, calcium-activated chloride currents are impaired in rods, despite unaltered TMEM16A protein levels. Co-immunoprecipitation revealed the interaction between VGCC and TMEM16A channels in the retina. Heterologous expression of these channels in tsA-201 cells showed that TMEM16A associates with the CaV1.4 subunit, and the association persists upon expression of the mutant α2δ4 subunit. Collectively, our experiments show association between TMEM16A and the α1 subunit of VGCC. Close proximity of these channels allows optimal function of the photoreceptor synaptic terminal under physiological conditions, but also makes TMEM16A channels susceptible to changes occurring to calcium channels. PMID:26557056

  5. Muscarinic cholinergic regulation of L-type calcium channel in heart of embryonic mice at different developmental stages

    Institute of Scientific and Technical Information of China (English)

    Hua-min LIANG; Su-yun LI; Ling-ling LAI; Juergen HESCHELER; Ming TANG; Chang-jin LIU; Hong-yan LUO; Yuan-long SONG; Xin-wu HU; Jiao-ya XI; Lin-lin GAO; Bin NIE


    AIM: To investigate the muscarinic regulation of L-type calcium current (ICa-L) during development. METHODS:The whole cell patch-clamp technique was used to record Ica- L in mice embryonic cardiomyocytes at different stages (the early developmental stage, EDS; the intermediate developmental stage, IDS; and the late developmental stage, LDS). Carbachol (CCh) was used to stimulate M-receptor in the embryonic cardiomyocytes of mice.RESULTS: The expression of Ica-L density did not change in different developmental stages (P>0.05). There was no difference in the sensitivity of ICa-L to CCh during development (P>0.05). This inhibitory action of CCh was mediated by inhibition of cyclic AMP since 8-bromo-cAMP completely reversed the muscarinic inhibitory action.IBMX, a non-selective inhibitor of phosphodiesterase (PDE), reversed the inhibitory action of M-receptor on ICa-Lcurrent by 71.2 %±9.2 % (n=8) and 11.3 %±2.5 % (n=9) in EDS and LDS respectively. However forskolin, an agonist of adenylyl cyclase (AC), reversed the action of CCh by 14.5 %±3.5 % (n=5) and 82.7 %±10.4 % (n=7) in EDS and LDS respectively. CONCLUSION: The inhibitory action of CCh on ICa-L current was mediated in different pathways: in EDS, the inhibitory action of M-receptor on ICa-L channel mainly depended on the stimulation of PDE. However, in LDS, the regulation by M-receptor on ICa-L channel mainly depended on the inactivation of AC.

  6. Muscarinic cholinergic regulation of L-type calcium channel in heart of embryonic mice at different developmental stages

    Institute of Scientific and Technical Information of China (English)

    Hua-minLIANG; MingTANG; Chang-jinLIU; Hong-yanLUO; Yuan-longSONG; Xin-wuHU; Jiao-yaXI; Lin-linGAO; BinNIE; Su-yunLI; Ling-lingLAI; JuergenHESCHELER


    AIM: To investigate the muscarinic regulation of L-type calcium current (ICa-L) during development. METHODS:The whole cell patch-clamp technique was used to record ICa-L in mice embryonic cardiomyocytes at different stages (the early developmental stage, EDS; the intermediate developmental stage, IDS; and the late developmental stage, LDS). Carbachol (CCh) was used to stimulate M-receptor in the embryonic cardiomyocytes of mice.RESULTS: The expression of lCa.L density did not change in different developmental stages (P>0.05). There was no difference in the sensitivity of ICa-L to CCh during development (P>0.05). This inhibitory action of CCh was mediated by inhibition of cyclic AMP since 8-bromo-cAMP completely reversed the muscarinic inhibitory action. IBMX, a non-selective inhibitor of phosphodiesterase (PDE), reversed the inhibitory action of M-receptor on ICa-L current by 71.2 %±9.2% (n=8) and 11.3%±2.5% (n=9) in EDS and LDS respectively. However forskolin, an agonist of adenylyl cyclase (AC), reversed the action of CCh by 14.5%±3.5% (n=5) and 82.7%± 10.4% (n=7) in EDS and LDS respectively. CONCLUSION: The inhibitory action of CCh on lca.L current was mediated in different pathways: in EDS, the inhibitory action of M-receptor on ICa-L channel mainly depended on the stimulation of PDE. However, in LDS, the regulation by M-receptor on lCa.L channel mainly depended on the inactivation of AC.

  7. Circadian profiles in the embryonic chick heart: L-type voltage-gated calcium channels and signaling pathways. (United States)

    Ko, Michael L; Shi, Liheng; Grushin, Kirill; Nigussie, Fikru; Ko, Gladys Y-P


    Circadian clocks exist in the heart tissue and modulate multiple physiological events, from cardiac metabolism to contractile function and expression of circadian oscillator and metabolic-related genes. Ample evidence has demonstrated that there are endogenous circadian oscillators in adult mammalian cardiomyocytes. However, mammalian embryos cannot be entrained independently to light-dark (LD) cycles in vivo without any maternal influence, but circadian genes are well expressed and able to oscillate in embryonic stages. The authors took advantage of using chick embryos that are independent of maternal influences to investigate whether embryonic hearts could be entrained under LD cycles in ovo. The authors found circadian regulation of L-type voltage-gated calcium channels (L-VGCCs), the ion channels responsible for the production of cardiac muscle contraction in embryonic chick hearts. The mRNA levels and protein expression of VGCCα1C and VGCCα1D are under circadian control, and the average L-VGCC current density is significantly larger when cardiomyocytes are recorded during the night than day. The phosphorylation states of several kinases involved in insulin signaling and cardiac metabolism, including extracellular signal-regulated kinase (Erk), stress-activated protein kinase (p38), protein kinase B (Akt), and glycogen synthase kinase-3β (GSK-3β), are also under circadian control. Both Erk and p38 have been implicated in regulating cardiac contractility and in the development of various pathological states, such as cardiac hypertrophy and heart failure. Even though both Erk and phosphoinositide 3-kinase (PI3K)-Akt signaling pathways participate in complex cellular processes regarding physiological or pathological states of cardiomyocytes, the circadian oscillators in the heart regulate these pathways independently, and both pathways contribute to the circadian regulation of L-VGCCs.

  8. Electromagnetic fields act via activation of voltage-gated calcium channels to produce beneficial or adverse effects. (United States)

    Pall, Martin L


    The direct targets of extremely low and microwave frequency range electromagnetic fields (EMFs) in producing non-thermal effects have not been clearly established. However, studies in the literature, reviewed here, provide substantial support for such direct targets. Twenty-three studies have shown that voltage-gated calcium channels (VGCCs) produce these and other EMF effects, such that the L-type or other VGCC blockers block or greatly lower diverse EMF effects. Furthermore, the voltage-gated properties of these channels may provide biophysically plausible mechanisms for EMF biological effects. Downstream responses of such EMF exposures may be mediated through Ca(2+) /calmodulin stimulation of nitric oxide synthesis. Potentially, physiological/therapeutic responses may be largely as a result of nitric oxide-cGMP-protein kinase G pathway stimulation. A well-studied example of such an apparent therapeutic response, EMF stimulation of bone growth, appears to work along this pathway. However, pathophysiological responses to EMFs may be as a result of nitric oxide-peroxynitrite-oxidative stress pathway of action. A single such well-documented example, EMF induction of DNA single-strand breaks in cells, as measured by alkaline comet assays, is reviewed here. Such single-strand breaks are known to be produced through the action of this pathway. Data on the mechanism of EMF induction of such breaks are limited; what data are available support this proposed mechanism. Other Ca(2+) -mediated regulatory changes, independent of nitric oxide, may also have roles. This article reviews, then, a substantially supported set of targets, VGCCs, whose stimulation produces non-thermal EMF responses by humans/higher animals with downstream effects involving Ca(2+) /calmodulin-dependent nitric oxide increases, which may explain therapeutic and pathophysiological effects.

  9. A genetic screen for dihydropyridine (DHP-resistant worms reveals new residues required for DHP-blockage of mammalian calcium channels.

    Directory of Open Access Journals (Sweden)

    Trevor C Y Kwok


    Full Text Available Dihydropyridines (DHPs are L-type calcium channel (Ca(v1 blockers prescribed to treat several diseases including hypertension. Ca(v1 channels normally exist in three states: a resting closed state, an open state that is triggered by membrane depolarization, followed by a non-conducting inactivated state that is triggered by the influx of calcium ions, and a rapid change in voltage. DHP binding is thought to alter the conformation of the channel, possibly by engaging a mechanism similar to voltage dependent inactivation, and locking a calcium ion in the pore, thereby blocking channel conductance. As a Ca(v1 channel crystal structure is lacking, the current model of DHP action has largely been achieved by investigating the role of candidate Ca(v1 residues in mediating DHP-sensitivity. To better understand DHP-block and identify additional Ca(v1 residues important for DHP-sensitivity, we screened 440,000 randomly mutated Caenorhabditis elegans genomes for worms resistant to DHP-induced growth defects. We identified 30 missense mutations in the worm Ca(v1 pore-forming (alpha(1 subunit, including eleven in conserved residues known to be necessary for DHP-binding. The remaining polymorphisms are in eight conserved residues not previously associated with DHP-sensitivity. Intriguingly, all of the worm mutants that we analyzed phenotypically exhibited increased channel activity. We also created orthologous mutations in the rat alpha(1C subunit and examined the DHP-block of current through the mutant channels in culture. Six of the seven mutant channels examined either decreased the DHP-sensitivity of the channel and/or exhibited significant residual current at DHP concentrations sufficient to block wild-type channels. Our results further support the idea that DHP-block is intimately associated with voltage dependent inactivation and underscores the utility of C. elegans as a screening tool to identify residues important for DHP interaction with mammalian

  10. Reduced Calcium Channel Function in Drosophila Disrupts Associative Learning in Larva and Behavior in Adults

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    Robin L. Cooper


    Full Text Available The temperature sensitive nature of a mutation in the Cacophony gene, which codes for the alpha subunit in the voltage-gated Ca2+ channel, reduces Ca2+ influx when exposed to non-permissive temperatures. We investigated the subtle nature in the impact for this mutation on whole animal function, in regards to learning and memory, in larvae and adults. The effects in acutely reducing evoked Ca2+ influx in nerve terminals during various behavioural assays greatly decreased the ability of larval Drosophila to learn, as demonstrated in associative learning assays. These assays are based on olfaction and gustation with association to light or dark environments with negative reinforces. Adult flies also showed defects in olfaction and sense of light when the animal is acutely depressed in normal Ca2+ influx within the nervous system. We demonstrated that this particular mutation does not alter cardiac function acutely. Thus, implying that the alpha 1 subunit mutation which retards neuronal function is not relevant for the pace maker and cardiac contractility as indexed by heart rate.

  11. ω-Tbo-IT1-New Inhibitor of Insect Calcium Channels Isolated from Spider Venom. (United States)

    Mikov, Alexander N; Fedorova, Irina M; Potapieva, Natalia N; Maleeva, Ekaterina E; Andreev, Yaroslav A; Zaitsev, Alexey V; Kim, Kira K; Bocharov, Eduard V; Bozin, Timur N; Altukhov, Dmitry A; Lipkin, Alexey V; Kozlov, Sergey A; Tikhonov, Denis B; Grishin, Eugene V


    Novel disulfide-containing polypeptide toxin was discovered in the venom of the Tibellus oblongus spider. We report on isolation, spatial structure determination and electrophysiological characterization of this 41-residue toxin, called ω-Tbo-IT1. It has an insect-toxic effect with LD50 19 μg/g in experiments on house fly Musca domestica larvae and with LD50 20 μg/g on juvenile Gromphadorhina portentosa cockroaches. Electrophysiological experiments revealed a reversible inhibition of evoked excitatory postsynaptic currents in blow fly Calliphora vicina neuromuscular junctions, while parameters of spontaneous ones were not affected. The inhibition was concentration dependent, with IC50 value 40 ± 10 nM and Hill coefficient 3.4 ± 0.3. The toxin did not affect frog neuromuscular junctions or glutamatergic and GABAergic transmission in rat brains. Ca(2+) currents in Calliphora vicina muscle were not inhibited, whereas in Periplaneta americana cockroach neurons at least one type of voltage gated Ca(2+) current was inhibited by ω-Tbo-IT1. Thus, the toxin apparently acts as an inhibitor of presynaptic insect Ca(2+) channels. Spatial structure analysis of the recombinant ω-Tbo-IT1 by NMR spectroscopy in aqueous solution revealed that the toxin comprises the conventional ICK fold containing an extended β-hairpin loop and short β-hairpin loop which are capable of making "scissors-like mutual motions".

  12. Characterisation of marrubenol, a diterpene extracted from Marrubium vulgare, as an L-type calcium channel blocker. (United States)

    El-Bardai, Sanae; Wibo, Maurice; Hamaide, Marie-Christine; Lyoussi, Badiaa; Quetin-Leclercq, Joelle; Morel, Nicole


    1. The objective of the present study was to investigate the mechanism of the relaxant activity of marrubenol, a diterpenoid extracted from Marrubium vulgare. In rat aorta, marrubenol was a more potent inhibitor of the contraction evoked by 100 mM KCl (IC50: 11.8+/-0.3 microM, maximum relaxation: 93+/-0.6%) than of the contraction evoked by noradrenaline (maximum relaxation: 30+/-1.5%). 2. In fura-2-loaded aorta, marrubenol simultaneously inhibited the Ca2+ signal and the contraction evoked by 100 mM KCl, and decreased the quenching rate of fura-2 fluorescence by Mn2+. 3. Patch-clamp data obtained in aortic smooth muscle cells (A7r5) indicated that marrubenol inhibited Ba2+ inward current in a voltage-dependent manner (KD: 8+/-2 and 40+/-6 microM at holding potentials of -50 and -100 mV, respectively). 4. These results showed that marrubenol inhibits smooth muscle contraction by blocking L-type calcium channels.

  13. Troponin T3 regulates nuclear localization of the calcium channel Cavβ1a subunit in skeletal muscle. (United States)

    Zhang, Tan; Taylor, Jackson; Jiang, Yang; Pereyra, Andrea S; Messi, Maria Laura; Wang, Zhong-Min; Hereñú, Claudia; Delbono, Osvaldo


    The voltage-gated calcium channel (Cav) β1a subunit (Cavβ1a) plays an important role in excitation-contraction coupling (ECC), a process in the myoplasm that leads to muscle-force generation. Recently, we discovered that the Cavβ1a subunit travels to the nucleus of skeletal muscle cells where it helps to regulate gene transcription. To determine how it travels to the nucleus, we performed a yeast two-hybrid screening of the mouse fast skeletal muscle cDNA library and identified an interaction with troponin T3 (TnT3), which we subsequently confirmed by co-immunoprecipitation and co-localization assays in mouse skeletal muscle in vivo and in cultured C2C12 muscle cells. Interacting domains were mapped to the leucine zipper domain in TnT3 COOH-terminus (160-244 aa) and Cavβ1a NH2-terminus (1-99 aa), respectively. The double fluorescence assay in C2C12 cells co-expressing TnT3/DsRed and Cavβ1a/YFP shows that TnT3 facilitates Cavβ1a nuclear recruitment, suggesting that the two proteins play a heretofore unknown role during early muscle differentiation in addition to their classical role in ECC regulation.

  14. Metabotropic glutamate receptor 6 signaling enhances TRPM1 calcium channel function and increases melanin content in human melanocytes. (United States)

    Devi, Sulochana; Markandeya, Yogananda; Maddodi, Nityanand; Dhingra, Anuradha; Vardi, Noga; Balijepalli, Ravi C; Setaluri, Vijayasaradhi


    Mutations in TRPM1, a calcium channel expressed in retinal bipolar cells and epidermal melanocytes, cause complete congenital stationary night blindness with no discernible skin phenotype. In the retina, TRPM1 activity is negatively coupled to metabotropic glutamate receptor 6 (mGluR6) signaling through Gαo and TRPM1 mutations result in the loss of responsiveness of TRPM1 to mGluR6 signaling. Here, we show that human melanocytes express mGluR6, and treatment of melanocytes with L-AP4, a type III mGluR-selective agonist, enhances Ca(2+) uptake. Knockdown of TRPM1 or mGluR6 by shRNA abolished L-AP4-induced Ca(2+) influx and TRPM1 currents, showing that TRPM1 activity in melanocytes is positively coupled to mGluR6 signaling. Gαo protein is absent in melanocytes. However, forced expression of Gαo restored negative coupling of TRPM1 to mGluR6 signaling, but treatment with pertussis toxin, an inhibitor of Gi /Go proteins, did not affect basal or mGluR6-induced Ca(2+) uptake. Additionally, chronic stimulation of mGluR6 altered melanocyte morphology and increased melanin content. These data suggest differences in coupling of TRPM1 function to mGluR6 signaling explain different cellular responses to glutamate in the retina and the skin.

  15. Investigation of Calcium Channel Blockers as Antiprotozoal Agents and Their Interference in the Metabolism of Leishmania (L. infantum

    Directory of Open Access Journals (Sweden)

    Juliana Quero Reimão


    Full Text Available Leishmaniasis and Chagas disease are neglected parasitic diseases endemic in developing countries; efforts to find new therapies remain a priority. Calcium channel blockers (CCBs are drugs in clinical use for hypertension and other heart pathologies. Based on previous reports about the antileishmanial activity of dihydropyridine-CCBs, this work aimed to investigate whether the in vitro anti-Leishmania infantum and anti-Trypanosoma cruzi activities of this therapeutic class would be shared by other non-dihydropyridine-CCBs. Except for amrinone, our results demonstrated antiprotozoal activity for fendiline, mibefradil, and lidoflazine, with IC50 values in a range between 2 and 16 μM and Selectivity Index between 4 and 10. Fendiline demonstrated depolarization of mitochondrial membrane potential, with increased reactive oxygen species production in amlodipine and fendiline treated Leishmania, but without plasma membrane disruption. Finally, in vitro combinations of amphotericin B, miltefosine, and pentamidine against L. infantum showed in isobolograms an additive interaction when these drugs were combined with fendiline, resulting in overall mean sum of fractional inhibitory concentrations between 0.99 and 1.10. These data demonstrated that non-dihydropyridine-CCBs present antiprotozoal activity and could be useful candidates for future in vivo efficacy studies against Leishmaniasis and Chagas’ disease.

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

    Directory of Open Access Journals (Sweden)

    Juliana Quero Reimão


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

  17. Commitment of Satellite Cells Expressing the Calcium Channel α2δ1 Subunit to the Muscle Lineage

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    Tammy Tamayo


    Full Text Available Satellite cells can maintain or repair muscle because they possess stem cell properties, making them a valuable option for cell therapy. However, cell transplants into skeletal muscle of patients with muscular dystrophy are limited by donor cell attachment, migration, and survival in the host tissue. Cells used for therapy are selected based on specific markers present in the plasma membrane. Although many markers have been identified, there is a need to find a marker that is expressed at different states in satellite cells, activated, quiescent, or differentiated cell. Furthermore, the marker has to be present in human tissue. Recently we reported that the plasma membrane α2δ1 protein is involved in cell attachment and migration in myoblasts. The α2δ1 subunit forms a part of the L-type voltage-dependent calcium channel in adult skeletal muscle. We found that the α2δ1 subunit is expressed in the majority of newly isolated satellite cells and that it appears earlier than the α1 subunits and at higher levels than the β or γ subunits. We also found that those cells that expressed α2δ1 would differentiate into muscle cells. This evidence indicates that the α2δ1 may be used as a marker of satellite cells that will differentiate into muscle.

  18. Cost containment for treating hypertension in African Americans: impact of a combined ACE inhibitor-calcium channel blocker. (United States)

    Kountz, D S


    The use of calcium channel blockers (CCBs) and angiotensin-converting enzyme (ACE) inhibitors has increased dramatically over the last 10 years and now accounts for 60% to 70% of all new antihypertensive prescriptions. Even though these two classes are efficacious, they are costly. Combined ACE inhibitor/CCB therapy (amlodipine-benazepril) was introduced in 1995. An analysis was done to assess the potential financial impact of substituting this agent for patients being treated with on ACE inhibitor/CCB combination. A pharmaceutical profile review of prescriptions during October 1995 was performed on 219 randomly selected patients enrolled in a Medicaid managed care program. Eighty-four profiles were analyzed; 24% of patients were on a combination ACE inhibitor/CCB regimen with an average monthly cost of $135. If the single agent amlodipine-benazepril with an average monthly cost of $45 (all strengths) was substituted, the savings would be considerable: $1080 per patient per year and $1,080,000 annualized for the calculated number of hypertensives on combination therapy in our network of 15,000 patients. Therapeutic substitution is one method of achieving cost containment in managed care. The cost differential between separately prescribed CCBs and ACE inhibitors and amlodipine-benazepril is significant. Compliance also should be enhanced as the patient would need to take only one pill daily. Once a patient has been maintained on a stable dose of a CCB/ACE inhibitor, substitution with amlodipine-benazepril should be considered.


    Directory of Open Access Journals (Sweden)

    M. A. Maksimova


    Full Text Available Aim. To study efficacy and safety of calcium channel blocker, S-amlodipine, in combination with β-blocker, atenolol, in patients with arterial hypertension (HT 1-2 degree com- pared to fixed combination of racemic amlodipine and atenolol.Material and methods. Patients (n=31, 7 men and 24 women with HT 1–2 degree were included into the study. The patients were randomized into two groups by the com- binations sequence. Treatment with each combination lasted 4 weeks. Office blood pressure (BP was assessed at baseline and at the end of the treatment periods, possible side effects were registered.Results. All patients completed the study. Both combination of S-amlodipine+atenolol and fixed combination of racemic amlodipine+atenolol reduced systolic (in average, -15.9 and -12.7 mm Hg, respectively and diastolic (in average, -7.3 and -5.3 mmHg, respectively BP significantly. Heart rate also decreased during therapy (in average, -3 and -4 bt/min, respectively. The differences between combinations BP and heart rate effects were not significant. 8 and 16 adverse events were registered during S-amlodipine+atenolol and racemic amlodipine+atenolol therapies, respectively Conclusion. Combination of S-amlodipine+atenolol, as well as combination of racemic amlodipine+atenolol are effective in the treatment of patients with HT 1-2 degree, however combination with S-amlodipine has less number of adverse events.

  20. Effects of the long-acting calcium channel blocker barnidipine hydrochloride on 24-h ambulatory blood pressure. (United States)

    Kuwajima, Iwao; Abe, Keishi


    The effect of the long acting calcium channel blocker, barnidipine hydrochloride (barnidipine) on 24-h ambulatory blood pressure (ABP) was evaluated in J-MUBA (Japanese Multicentre Study on Barnidipine with Ambulatory Blood Pressure Monitoring). Following an observation period of two weeks, antihypertensive treatment with barnidipine was continued for at least six months. At the end of each period, ABP were measured. The patients were divided into high- and low-range groups based on ABP measurement. Throughout the 24 h, barnidipine exerted an excellent antihypertensive effect in the high-range group, but not in the low-range group. Barnidipine had comparable effects in the daytime and nighttime in inverted dippers and non-dippers, but it was more effective on daytime ABP than on nighttime ABP in dippers and in extreme dippers. Morning blood pressure before and after waking was evaluated before and after barnidipine administration in 233 patients. Barnidipine inhibited increases in blood pressure before and after waking, especially in surge-type patients whose blood pressure increased rapidly after waking. A positive correlation among 24-h ABP, daytime and night time ABP, morning blood pressure, and clinic blood pressure during the observation period and the antihypertensive effect of barnidipine was observed, with barnidipine exhibiting stronger antihypertensive effects in patients with persistently high blood pressure. It was concluded that the antihypertensive effects of barnidipine are maintained for 24 h but it has no excessive hypotensive effects on lower blood pressure and is thus a safe antihypertensive agent.

  1. Serum response factor regulates smooth muscle contractility via myotonic dystrophy protein kinases and L-type calcium channels (United States)

    Lee, Moon Young; Park, Chanjae; Ha, Se Eun; Park, Paul J.; Berent, Robyn M.; Jorgensen, Brian G.; Corrigan, Robert D.; Grainger, Nathan; Blair, Peter J.; Slivano, Orazio J.; Miano, Joseph M.; Ward, Sean M.; Smith, Terence K.; Sanders, Kenton M.


    Serum response factor (SRF) transcriptionally regulates expression of contractile genes in smooth muscle cells (SMC). Lack or decrease of SRF is directly linked to a phenotypic change of SMC, leading to hypomotility of smooth muscle in the gastrointestinal (GI) tract. However, the molecular mechanism behind SRF-induced hypomotility in GI smooth muscle is largely unknown. We describe here how SRF plays a functional role in the regulation of the SMC contractility via myotonic dystrophy protein kinase (DMPK) and L-type calcium channel CACNA1C. GI SMC expressed Dmpk and Cacna1c genes into multiple alternative transcriptional isoforms. Deficiency of SRF in SMC of Srf knockout (KO) mice led to reduction of SRF-dependent DMPK, which down-regulated the expression of CACNA1C. Reduction of CACNA1C in KO SMC not only decreased intracellular Ca2+ spikes but also disrupted their coupling between cells resulting in decreased contractility. The role of SRF in the regulation of SMC phenotype and function provides new insight into how SMC lose their contractility leading to hypomotility in pathophysiological conditions within the GI tract. PMID:28152551

  2. Modulation of MAA-induced apoptosis in male germ cells: role of Sertoli cell P/Q-type calcium channels

    Directory of Open Access Journals (Sweden)

    Aguanno Salvatore


    Full Text Available Abstract Spontaneous germ cell death by apoptosis occurs during normal spermatogenesis in mammals and is thought to play a role in the physiological mechanism limiting the clonal expansion of such cell population in the male gonad. In the prepubertal rat testis, the most conspicuous dying cells are pachytene spermatocytes, which are also the primary target of the apoptosis experimentally induced by the methoxyacetic acid (MAA. Since we have recently reported that Sertoli cells, the somatic component of the seminiferous epithelium, regulate not only germ cell viability and differentiation but also their death, we have further investigated the mechanism involved in such a control. In this paper we have used the protein clusterin, produced by Sertoli cells and associated with tissue damage or injury, as indicator of germ cell apoptosis in rat seminiferous tubules treated with MAA in the presence or in the absence of omega-agatoxin, a specific inhibitor of P/Q type voltage-operated calcium channels (VOCC's. We performed both a qualitative analysis of clusterin content and germ cell apoptosis by immunofluorescence experiments and a quantitative analysis by in situ end labelling of apoptotic germ cells followed by flow cytometry. The results obtained demonstrate that Sertoli cells modulate germ cell apoptosis induced by methoxyacetic acid also throughout the P/Q-type VOCC's.

  3. Steroid hormone regulation of the voltage-gated, calcium-activated potassium channel expression in developing muscular and neural systems. (United States)

    Garrison, Sheldon L; Witten, Jane L


    A precise organization of gene expression is required for developing neural and muscular systems. Steroid hormones can control the expression of genes that are critical for development. In this study we test the hypothesis that the steroid hormone ecdysone regulates gene expression of the voltage-gated calcium-activated potassium ion channel, Slowpoke or KCNMA1. Late in adult development of the tobacco hawkmoth Manduca sexta, slowpoke (msslo) levels increased contributing to the maturation of the dorsal longitudinal flight muscles (DLMs) and CNS. We show that critical components of ecdysteroid gene regulation were present during upreglation of msslo in late adult DLM and CNS development. Ecdysteroid receptor complex heterodimeric partner proteins, the ecdysteroid receptor (EcR) and ultraspiracle (USP), and the ecdysone-induced early gene, msE75B, were expressed at key developmental time points, suggesting that ecdysteroids direct aspects of gene expression in the DLMs during these late developmental stages. We provide evidence that ecdysteroids suppress msslo transcription in the DLMs; when titers decline msslo transcript levels increase. These results are consistent with msslo being a downstream gene in an ecdysteroid-mediated gene cascade during DLM development. We also show that the ecdysteroids regulate msslo transcript levels in the developing CNS. These results will contribute to our understanding of how the spatiotemporal regulation of slowpoke transcription contributes to tailoring cell excitability to the differing physiological and behavioral demands during development.

  4. Functional importance of T-type voltage-gated calcium channels in the cardiovascular and renal system: news from the world of knockout mice. (United States)

    Hansen, Pernille B L


    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, the lack of highly specific blockers cast doubt on the conclusions. As new T-type channel antagonists are being designed, the roles of T-type channels in cardiovascular and renal pathology need to be elucidated before T-type blockers can be clinically useful. Two types of T-type channels, Cav3.1 and Cav3.2, are expressed in blood vessels, the kidney, and the heart. Studies with gene-deficient mice have provided a way to investigate the Cav3.1 and Cav3.2 channels and their role in the cardiovascular system. This review discusses the results from these knockout mice. Evaluation of the literature leads to the conclusion that Cav3.1 and Cav3.2 channels have important, but different, functions in mice. T-type Cav3.1 channels affect heart rate, whereas Cav3.2 channels are involved in cardiac hypertrophy. In the vascular system, Cav3.2 activation leads to dilation of blood vessels, whereas Cav3.1 channels are mainly suggested to affect constriction. The Cav3.1 channel is also involved in neointima formation following vascular damage. In the kidney, Cav3.1 regulates plasma flow and Cav3.2 plays a role setting glomerular filtration rate. In conclusion, Cav3.1 and Cav3.2 are new therapeutic targets in several cardiovascular pathologies, but the use of T-type blockers should be specifically directed to the disease and to the channel subtype.

  5. The Involvement of Ser1898 of the Human L-Type Calcium Channel in Evoked Secretion

    Directory of Open Access Journals (Sweden)

    Niv Bachnoff


    Full Text Available A PKA consensus phosphorylation site S1928 at the α11.2 subunit of the rabbit cardiac L-type channel, CaV1.2, is involved in the regulation of CaV1.2 kinetics and affects catecholamine secretion. This mutation does not alter basal CaV1.2 current properties or regulation of CaV1.2 current by PKA and the beta-adrenergic receptor, but abolishes CaV1.2 phosphorylation by PKA. Here, we test the contribution of the corresponding PKA phosphorylation site of the human α11.2 subunit S1898, to the regulation of catecholamine secretion in bovine chromaffin cells. Chromaffin cells were infected with a Semliki-Forest viral vector containing either the human wt or a mutated S1898A α11.2 subunit. Both subunits harbor a T1036Y mutation conferring nifedipine insensitivity. Secretion evoked by depolarization in the presence of nifedipine was monitored by amperometry. Depolarization-triggered secretion in cells infected with either the wt α11.2 or α11.2/S1898A mutated subunit was elevated to a similar extent by forskolin. Forskolin, known to directly activate adenylyl-cyclase, increased the rate of secretion in a manner that is largely independent of the presence of S1898. Our results are consistent with the involvement of additional PKA regulatory site(s at the C-tail of α11.2, the pore forming subunit of CaV1.2.

  6. Age-dependent impact of CaV3.2 T-type calcium channel deletion on myogenic tone and flow-mediated vasodilatation in small arteries

    DEFF Research Database (Denmark)

    Mikkelsen, Miriam F.; Björling, Karl; Jensen, Lars Jørn


    .2-dependent and -independent effects. No changes in mRNA expression of several important K(+) and Ca(2+) channel genes were induced by CaV3.2 knock-out. However, the expression of the other T-type channel isoform (CaV3.1) was reduced at the mRNA and protein level in mature adult compared to young WT arteries......The myogenic response and flow-mediated vasodilatation are important regulators of local blood perfusion and total peripheral resistance, and are known to entail a calcium influx into vascular smooth muscle cells (VSMCs) and endothelial cells (ECs), respectively. CaV3.2 T-type calcium channels...... are expressed in both VSMCs and ECs of small arteries. The T-type channels are important drug targets but due to the lack of specific antagonists our understanding of the role of CaV3.2 channels in vasomotor tone at various ages is scarce. We evaluated the myogenic response, flow-mediated vasodilatation...

  7. Non-cardiogenic pulmonary edema and life-threatening shock due to calcium channel blocker overdose: a case report and clinical review. (United States)

    Siddiqi, Tauseef Afaq; Hill, Jennifer; Huckleberry, Yvonne; Parthasarathy, Sairam


    Calcium channel blockers (CCBs) overdose can be life-threatening when manifest as catastrophic shock and non-cardiogenic pulmonary edema. We describe a case of massive overdose of multiple medications, including sustained-release verapamil, which was resistant to conventional support. Initial treatment for CCB overdose is primarily supportive, and includes fluid resuscitation. The mechanism of non-cardiogenic pulmonary edema is not well known, and reported cases have been successfully treated with mechanical ventilation. Circulatory shock may fail to respond to atropine, glucagon, and calcium in severely poisoned patients, and vasopressors are usually required. Attempting to overcome calcium-channel antagonism with the supra-therapeutic doses of calcium salts is clinically indicated to reverse hypotension and bradycardia. There is evidence that hyperinsulinemia-euglycemia therapy is superior to other therapies for CCB poisoning, and the mechanism is thought to be the insulin-mediated active transport of glucose into the cells, which counters the CCB-induced intra-cellular carbohydrate-deficient state. Conventional decontamination measures are ineffective in accelerating clearance of CCB. Experience with intravenous lipid emulsion for lipophilic drug overdose, such as verapamil, is limited, but has been proposed as a rescue therapy and might improve cardiac inotropy through intravascular sequestration of the lipophilic CCB.

  8. 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;


    AIMS: As cardiovascular disease is characterized by reduced nitric oxide bioavailability, our aim was to determine the impact of this change on the mechanism underlying vascular tone of pressurized arteries in vitro and in vivo. METHODS AND RESULTS: We used pressurized cerebral and mesenteric......, 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....

  9. Two distinct pools of large-conductance calcium-activated potassium channels in the somatic plasma membrane of central principal neurons


    Kaufmann, W.A.; Kasugai, Y.; Ferraguti, F.; Storm, J F


    Although nerve cell membranes are often assumed to be uniform with respect to electrical properties, there is increasing evidence for compartmentalization into subdomains with heterogeneous impacts on the overall cell function. Such microdomains are characterized by specific sets of proteins determining their functional properties. Recently, clustering of large-conductance calcium-activated potassium (BKCa) channels was shown at sites of subsurface membrane cisterns in cerebellar Purkinje cel...

  10. Differential effect of T-type voltage-gated calcium channel disruption on renal plasma flow and glomerular filtration rate in vivo

    DEFF Research Database (Denmark)

    Thuesen, Anne D; Andersen, Henrik; Cardel, Majken


    Voltage-gated calcium channels (Cav) play an essential role in regulation of renal blood flow and GFR. Because T-type Cavs are differentially expressed in pre- and postglomerular vessels it was hypothesized that they impact renal blood flow and GFR differentially. The question was addressed by us...... contribute to renal vascular resistance. It is suggested that endothelial and nerve localization of Cav 3.2 and Cav 3.1, respectively, may account for the observed effects....

  11. The nociception role of Cav3.2 calcium channel%Cav3.2通道在伤害性感受中的作用

    Institute of Scientific and Technical Information of China (English)

    苏昊; 徐世元


    Background Some of the earliest detailed descriptions of biophysical properties of low voltage-activated or transient (T) type Calcium channels were done using in vitro preparation of primary sensory or dorsal root ganglion (DRG) neurons that are known for their functional role in processing pain signals.However, in spite of these early discoveries, T-type calcium channels were not implicated in sensory transmission in general and pain processing (nociception) in particular until recently.Objective To summarize the important role of peripheral T-type Calcium channels in boosting nociceptive transmission.Content T-type calcium channels in peripheral sensory neurons play important role under physiological conditions (e.g.acute nociceptive pain) and to pain processing under pathological conditions.Trend peripheral T-type Calcium channels in nociception and the value of these channels as cellular targets for potential drug developments.%背景 疼痛是危害人类健康,影响人类生存质量的重要因素,但目前可用于临床治疗的手段尚不能满足患者的无痛要求,且存在不同程度的副作用,其主要原因之一是目前对疼痛的产生机制尚未完全明了.最近的研究表明,T型钙通道Cav3.2亚型在疼痛信号转导中起重要作用. 目的 综述T型钙通道Cav3.2亚型在痛觉传导中的作用. 内容 阐述位于背根神经节(dorsal root ganglion,DRG)的T型钙通道参与传递生理性疼痛(疼痛感知)和病理性疼痛(神经病理性疼痛)伤害性刺激信号. 趋向 T型钙离子通道可调控疼痛伤害性刺激信号转导,并可作为可能的疼痛治疗靶点.

  12. Fibronectin-induced VEGF receptor and calcium channel transactivation stimulate GLUT-1 synthesis and trafficking through PPARγ and TC10 in mouse embryonic stem cells. (United States)

    Suh, Han Na; Han, Ho Jae


    Extracellular matrix (ECM) mediates interactions between integrin and growth factor receptor (GFR) or ion channel. Although this crosstalk promotes integration of the downstream signal pathways and then regulates cellular function, the effect of ECM on glucose transporter (GLUT) in stem cells has not been elucidated. Therefore, we examined the effect of fibronectin on GLUT-1 expression, trafficking, and its related signal pathways in mouse embryonic stem cells (mESCs). Fibronectin increased 2-deoxyglucose (DG) uptake and GLUT-1 protein expression that were blocked by transcription or translation inhibitors. Integrin α5β1-bound fibronectin increased 2-DG uptake through cluster formation with vascular endothelial growth factor receptor (VEGFR) 2, and then activated Ras and PI3K/Akt. In another pathway, integrin α5β1 displayed structural and functional interactions with calcium channels, and stimulated 2-DG uptake through calcium influx and PKC activation. Akt and PKC-induced PPARγ phosphorylation enhanced the decreased expression of PPARγ protein, and subsequently increased GLUT-1 protein synthesis and 2-DG uptake. Fibronectin stimulated TC10 activity and cytoskeleton (F-actin) rearrangement, followed by GLUT-1 trafficking. In conclusion, integrin-bound fibronectin stimulates GLUT-1 synthesis through VEGFR2/Ras/PI3K/Akt and calcium channel/Ca(2+)/PKC, which are merged at PPARγ and GLUT-1 trafficking through TC10 and F-actin.

  13. Genetic contribution to iron status: SNPs related to iron deficiency anaemia and fine mapping of CACNA2D3 calcium channel subunit. (United States)

    Baeza-Richer, Carlos; Arroyo-Pardo, Eduardo; Blanco-Rojo, Ruth; Toxqui, Laura; Remacha, Angel; Vaquero, M Pilar; López-Parra, Ana M


    Numerous studies associate genetic markers with iron- and erythrocyte-related parameters, but few relate them to iron-clinical phenotypes. Novel SNP rs1375515, located in a subunit of the calcium channel gene CACNA2D3, is associated with a higher risk of anaemia. The aim of this study is to further investigate the association of this SNP with iron-related parameters and iron-clinical phenotypes, and to explore the potential role of calcium channel subunit region in iron regulation. Furthermore, we aim to replicate the association of other SNPs reported previously in our population. We tested 45 SNPs selected via systematic review and fine mapping of CACNA2D3 region, with haematological and biochemical traits in 358 women of reproductive age. Multivariate analyses include back-step logistic regression and decision trees. The results replicate the association of SNPs with iron-related traits, and also confirm the protective effect of both A allele of rs1800562 (HFE) and G allele of rs4895441 (HBS1L-MYB). The risk of developing anaemia is increased in reproductive age women carriers of A allele of rs18