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Sample records for specific trpc6 channel

  1. TRPC6 is the endothelial calcium channel that regulates leukocyte transendothelial migration during the inflammatory response.

    PubMed

    Weber, Evan W; Han, Fei; Tauseef, Mohammad; Birnbaumer, Lutz; Mehta, Dolly; Muller, William A

    2015-10-19

    Leukocyte transendothelial migration (TEM) is a tightly regulated, multistep process that is critical to the inflammatory response. A transient increase in endothelial cytosolic free calcium ion concentration (↑[Ca(2+)]i) is required for TEM. However, the mechanism by which endothelial ↑[Ca(2+)]i regulates TEM and the channels mediating this ↑[Ca(2+)]i are unknown. Buffering ↑[Ca(2+)]i in endothelial cells does not affect leukocyte adhesion or locomotion but selectively blocks TEM, suggesting a role for ↑[Ca(2+)]i specifically for this step. Transient receptor potential canonical 6 (TRPC6), a Ca(2+) channel expressed in endothelial cells, colocalizes with platelet/endothelial cell adhesion molecule-1 (PECAM) to surround leukocytes during TEM and clusters when endothelial PECAM is engaged. Expression of dominant-negative TRPC6 or shRNA knockdown in endothelial cells arrests neutrophils apically over the junction, similar to when PECAM is blocked. Selectively activating endothelial TRPC6 rescues TEM during an ongoing PECAM blockade, indicating that TRPC6 functions downstream of PECAM. Furthermore, endothelial TRPC6 is required for trafficking of lateral border recycling compartment membrane, which facilitates TEM. Finally, mice lacking TRPC6 in the nonmyeloid compartment (i.e., endothelium) exhibit a profound defect in neutrophil TEM with no effect on leukocyte trafficking. Our findings identify endothelial TRPC6 as the calcium channel mediating the ↑[Ca(2+)]i required for TEM at a step downstream of PECAM homophilic interactions. PMID:26392222

  2. TRPC6 is the endothelial calcium channel that regulates leukocyte transendothelial migration during the inflammatory response

    PubMed Central

    Weber, Evan W.; Han, Fei; Tauseef, Mohammad; Birnbaumer, Lutz; Mehta, Dolly

    2015-01-01

    Leukocyte transendothelial migration (TEM) is a tightly regulated, multistep process that is critical to the inflammatory response. A transient increase in endothelial cytosolic free calcium ion concentration (↑[Ca2+]i) is required for TEM. However, the mechanism by which endothelial ↑[Ca2+]i regulates TEM and the channels mediating this ↑[Ca2+]i are unknown. Buffering ↑[Ca2+]i in endothelial cells does not affect leukocyte adhesion or locomotion but selectively blocks TEM, suggesting a role for ↑[Ca2+]i specifically for this step. Transient receptor potential canonical 6 (TRPC6), a Ca2+ channel expressed in endothelial cells, colocalizes with platelet/endothelial cell adhesion molecule-1 (PECAM) to surround leukocytes during TEM and clusters when endothelial PECAM is engaged. Expression of dominant-negative TRPC6 or shRNA knockdown in endothelial cells arrests neutrophils apically over the junction, similar to when PECAM is blocked. Selectively activating endothelial TRPC6 rescues TEM during an ongoing PECAM blockade, indicating that TRPC6 functions downstream of PECAM. Furthermore, endothelial TRPC6 is required for trafficking of lateral border recycling compartment membrane, which facilitates TEM. Finally, mice lacking TRPC6 in the nonmyeloid compartment (i.e., endothelium) exhibit a profound defect in neutrophil TEM with no effect on leukocyte trafficking. Our findings identify endothelial TRPC6 as the calcium channel mediating the ↑[Ca2+]i required for TEM at a step downstream of PECAM homophilic interactions. PMID:26392222

  3. Molecular determinants for cardiovascular TRPC6 channel regulation by Ca2+/calmodulin-dependent kinase II

    PubMed Central

    Shi, Juan; Geshi, Naomi; Takahashi, Shinichi; Kiyonaka, Shigeki; Ichikawa, Jun; Hu, Yaopeng; Mori, Yasuo; Ito, Yushi; Inoue, Ryuji

    2013-01-01

    The molecular mechanism underlying Ca2+/calmodulin (CaM)-dependent kinase II (CaMKII)-mediated regulation of the mouse transient receptor potential channel TRPC6 was explored by chimera, deletion and site-directed mutagenesis approaches. Induction of currents (ICCh) in TRPC6-expressing HEK293 cells by a muscarinic agonist carbachol (CCh; 100 μm) was strongly attenuated by a CaMKII-specific peptide, autocamtide-2-related inhibitory peptide (AIP; 10 μm). TRPC6/C7 chimera experiments showed that the TRPC6 C-terminal sequence is indispensable for ICCh to be sensitive to AIP-induced CaMKII inhibition. Further, deletion of a distal region (Gln855–Glu877) of the C-terminal CaM/inositol-1,4,5-trisphosphate receptor binding domain (CIRB) of TRPC6 was sufficient to abolish ICCh. Systematic alanine scanning for potential CaMKII phosphorylation sites revealed that Thr487 was solely responsible for the activation of the TRPC6 channel by receptor stimulation. The abrogating effect of the alanine mutation of Thr487 (T487A) was reproduced with other non-polar amino acids, namely glutamine or asparagine, while being partially rescued by phosphomimetic mutations with glutamate or aspartate. The cellular expression and distribution of TRPC6 channels did not significantly change with these mutations. Electrophysiological and immunocytochemical data with the Myc-tagged TRPC6 channel indicated that Thr487 is most likely located at the intracellular side of the cell membrane. Overexpression of T487A caused significant reduction of endogenous TRPC6-like current induced by Arg8-vasopressin in A7r5 aortic myocytes. Based on these results, we propose that the optimal spatial arrangement of a C-terminal domain (presumably the distal CIRB region) around a single CaMKII phosphorylation site Thr487 may be essential for CaMKII-mediated regulation of TRPC6 channels. This mechanism may be of physiological significance in a native environment such as in vascular smooth muscle cells. PMID

  4. TRPC6 channel translocation into phagosomal membrane augments phagosomal function

    PubMed Central

    Riazanski, Vladimir; Gabdoulkhakova, Aida G.; Boynton, Lin S.; Eguchi, Raphael R.; Deriy, Ludmila V.; Hogarth, D. Kyle; Loaëc, Nadège; Oumata, Nassima; Galons, Hervé; Brown, Mary E.; Shevchenko, Pavel; Gallan, Alexander J.; Yoo, Sang Gune; Naren, Anjaparavanda P.; Villereal, Mitchel L.; Beacham, Daniel W.; Bindokas, Vytautas P.; Birnbaumer, Lutz; Meijer, Laurent; Nelson, Deborah J.

    2015-01-01

    Defects in the innate immune system in the lung with attendant bacterial infections contribute to lung tissue damage, respiratory insufficiency, and ultimately death in the pathogenesis of cystic fibrosis (CF). Professional phagocytes, including alveolar macrophages (AMs), have specialized pathways that ensure efficient killing of pathogens in phagosomes. Phagosomal acidification facilitates the optimal functioning of degradative enzymes, ultimately contributing to bacterial killing. Generation of low organellar pH is primarily driven by the V-ATPases, proton pumps that use cytoplasmic ATP to load H+ into the organelle. Critical to phagosomal acidification are various channels derived from the plasma membrane, including the anion channel cystic fibrosis transmembrane conductance regulator, which shunt the transmembrane potential generated by movement of protons. Here we show that the transient receptor potential canonical-6 (TRPC6) calcium-permeable channel in the AM also functions to shunt the transmembrane potential generated by proton pumping and is capable of restoring microbicidal function to compromised AMs in CF and enhancement of function in non-CF cells. TRPC6 channel activity is enhanced via translocation to the cell surface (and then ultimately to the phagosome during phagocytosis) in response to G-protein signaling activated by the small molecule (R)-roscovitine and its derivatives. These data show that enhancing vesicular insertion of the TRPC6 channel to the plasma membrane may represent a general mechanism for restoring phagosome activity in conditions, where it is lost or impaired. PMID:26604306

  5. TRPC6 channel translocation into phagosomal membrane augments phagosomal function.

    PubMed

    Riazanski, Vladimir; Gabdoulkhakova, Aida G; Boynton, Lin S; Eguchi, Raphael R; Deriy, Ludmila V; Hogarth, D Kyle; Loaëc, Nadège; Oumata, Nassima; Galons, Hervé; Brown, Mary E; Shevchenko, Pavel; Gallan, Alexander J; Yoo, Sang Gune; Naren, Anjaparavanda P; Villereal, Mitchel L; Beacham, Daniel W; Bindokas, Vytautas P; Birnbaumer, Lutz; Meijer, Laurent; Nelson, Deborah J

    2015-11-24

    Defects in the innate immune system in the lung with attendant bacterial infections contribute to lung tissue damage, respiratory insufficiency, and ultimately death in the pathogenesis of cystic fibrosis (CF). Professional phagocytes, including alveolar macrophages (AMs), have specialized pathways that ensure efficient killing of pathogens in phagosomes. Phagosomal acidification facilitates the optimal functioning of degradative enzymes, ultimately contributing to bacterial killing. Generation of low organellar pH is primarily driven by the V-ATPases, proton pumps that use cytoplasmic ATP to load H(+) into the organelle. Critical to phagosomal acidification are various channels derived from the plasma membrane, including the anion channel cystic fibrosis transmembrane conductance regulator, which shunt the transmembrane potential generated by movement of protons. Here we show that the transient receptor potential canonical-6 (TRPC6) calcium-permeable channel in the AM also functions to shunt the transmembrane potential generated by proton pumping and is capable of restoring microbicidal function to compromised AMs in CF and enhancement of function in non-CF cells. TRPC6 channel activity is enhanced via translocation to the cell surface (and then ultimately to the phagosome during phagocytosis) in response to G-protein signaling activated by the small molecule (R)-roscovitine and its derivatives. These data show that enhancing vesicular insertion of the TRPC6 channel to the plasma membrane may represent a general mechanism for restoring phagosome activity in conditions, where it is lost or impaired. PMID:26604306

  6. Cardioprotection by Klotho through downregulation of TRPC6 channels in the mouse heart

    PubMed Central

    Xie, Jian; Cha, Seung-Kuy; An, Sung-Wan; Kuro-o, Makoto; Birnbaumer, Lutz; Huang, Chou-Long

    2012-01-01

    Klotho is a membrane protein predominantly produced in the kidney that exerts some anti-ageing effects. Ageing is associated with an increased risk of heart failure; whether Klotho is cardioprotective is unknown. Here we show that Klotho-deficient mice have no baseline cardiac abnormalities but develop exaggerated pathological cardiac hypertrophy and remodeling in response to stress. Cardioprotection by Klotho in normal mice is mediated by downregulation of TRPC6 channels in the heart. We demonstrate that deletion of Trpc6 prevents stress-induced exaggerated cardiac remodeling in Klotho-deficient mice. Furthermore, mice with heart-specific overexpression of TRPC6 develop spontaneous cardiac hypertrophy and remodeling. Klotho overexpression ameliorates cardiac pathologies in these mice and improves their long-term survival. Soluble Klotho present in the systemic circulation inhibits TRPC6 currents in cardiomyocytes by blocking phosphoinositide-3-kinase-dependent exocytosis of TRPC6 channels. These results provide a new perspective on the pathogenesis of cardiomyopathies and open new avenues for treatment of the disease. PMID:23212367

  7. Activation of TRPC6 channels is essential for lung ischaemia–reperfusion induced oedema in mice

    PubMed Central

    Weissmann, Norbert; Sydykov, Akylbek; Kalwa, Hermann; Storch, Ursula; Fuchs, Beate; Schnitzler, Michael Mederos y; Brandes, Ralf P.; Grimminger, Friedrich; Meissner, Marcel; Freichel, Marc; Offermanns, Stefan; Veit, Florian; Pak, Oleg; Krause, Karl-Heinz; Schermuly, Ralph T.; Brewer, Alison C; Schmidt, Harald H.H.W.; Seeger, Werner; Shah, Ajay M.; Gudermann, Thomas; Ghofrani, Hossein A.; Dietrich, Alexander

    2012-01-01

    Lung ischaemia–reperfusion-induced oedema (LIRE) is a life-threatening condition that causes pulmonary oedema induced by endothelial dysfunction. Here we show that lungs from mice lacking nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox2y/−) or the classical transient receptor potential channel 6 (TRPC6−/−) are protected from LIR-induced oedema (LIRE). Generation of chimeric mice by bone marrow cell transplantation and endothelial-specific Nox2 deletion showed that endothelial Nox2, but not leukocytic Nox2 or TRPC6, are responsible for LIRE. Lung endothelial cells from Nox2- or TRPC6-deficient mice showed attenuated ischaemia-induced Ca2+ influx, cellular shape changes and impaired barrier function. Production of reactive oxygen species was completely abolished in Nox2y/− cells. A novel mechanistic model comprising endothelial Nox2-derived production of superoxide, activation of phospholipase C-γ, inhibition of diacylglycerol (DAG) kinase, DAG-mediated activation of TRPC6 and ensuing LIRE is supported by pharmacological and molecular evidence. This mechanism highlights novel pharmacological targets for the treatment of LIRE. PMID:22337127

  8. Podocyte-Specific Overexpression of Wild Type or Mutant Trpc6 in Mice Is Sufficient to Cause Glomerular Disease

    PubMed Central

    Kairath, Pamela; Carmona-Mora, Paulina; Molina, Jessica; Carpio, J. Daniel; Ruiz, Phillip; Mezzano, Sergio A.; Li, Jing; Wei, Changli; Reiser, Jochen; Young, Juan I.; Walz, Katherina

    2010-01-01

    Mutations in the TRPC6 calcium channel (Transient receptor potential channel 6) gene have been associated with familiar forms of Focal and Segmental Glomerulosclerosis (FSGS) affecting children and adults. In addition, acquired glomerular diseases are associated with increased expression levels of TRPC6. However, the exact role of TRPC6 in the pathogenesis of FSGS remains to be elucidated. In this work we describe the generation and phenotypic characterization of three different transgenic mouse lines with podocyte-specific overexpression of the wild type or any of two mutant forms of Trpc6 (P111Q and E896K) previously related to FSGS. Consistent with the human phenotype a non-nephrotic range of albuminuria was detectable in almost all transgenic lines. The histological analysis demonstrated that the transgenic mice developed a kidney disease similar to human FSGS. Differences of 2–3 folds in the presence of glomerular lesions were found between the non transgenic and transgenic mice expressing Trpc6 in its wild type or mutant forms specifically in podocytes. Electron microscopy of glomerulus from transgenic mice showed extensive podocyte foot process effacement. We conclude that overexpression of Trpc6 (wild type or mutated) in podocytes is sufficient to cause a kidney disease consistent with FSGS. Our results contribute to reinforce the central role of podocytes in the etiology of FSGS. These mice constitute an important new model in which to study future therapies and outcomes of this complex disease. PMID:20877463

  9. Receptor channel TRPC6 orchestrate the activation of human hepatic stellate cell under hypoxia condition

    SciTech Connect

    Iyer, Soumya C; Kannan, Anbarasu; Gopal, Ashidha; Devaraj, Niranjali; Halagowder, Devaraj

    2015-08-01

    Hepatic stellate cells (HSCs), a specialized stromal cytotype have a great impact on the biological behaviors of liver diseases. Despite this fact, the underlying mechanism that regulates HSC still remains poorly understood. The aim of the present study was to understand the role of TRPC6 signaling in regulating the molecular mechanism of HSCs in response to hypoxia. In the present study we showed that under hypoxia condition, the upregulated Hypoxia Inducible Factor 1α (HIF1α) increases NICD activation, which in turn induces the expression of transient receptor potential channel 6 (TRPC6) in HSC line lx-2. TRPC6 causes a sustained elevation of intracellular calcium which is coupled with the activation of the calcineurin-nuclear factor of activated T-cell (NFAT) pathway which activates the synthesis of extracellular matrix proteins. TRPC6 also activates SMAD2/3 dependent TGF-β signaling in facilitating upregulated expression of αSMA and collagen. As activated HSCs may be a suitable target for HCC therapy and targeting these cells rather than the HCC cells may result in a greater response. Collectively, our studies indicate for the first time the detailed mechanism of activation of HSC through TRPC6 signaling and thus being a promising therapeutic target. - Highlights: • HIF1α increases NICD, induces TRPC6 in lx2 cells. • TRPC6 a novel regulator in the activation of HSC. • HSCs as target for HCC therapy.

  10. Classical transient receptor potential channel 6 (TRPC6) is essential for hypoxic pulmonary vasoconstriction and alveolar gas exchange

    PubMed Central

    Weissmann, Norbert; Dietrich, Alexander; Fuchs, Beate; Kalwa, Hermann; Ay, Mahmut; Dumitrascu, Rio; Olschewski, Andrea; Storch, Ursula; Mederos y Schnitzler, Michael; Ghofrani, Hossein Ardeschir; Schermuly, Ralph Theo; Pinkenburg, Olaf; Seeger, Werner; Grimminger, Friedrich; Gudermann, Thomas

    2006-01-01

    Regional alveolar hypoxia causes local vasoconstriction in the lung, shifting blood flow from hypoxic to normoxic areas, thereby maintaining gas exchange. This mechanism is known as hypoxic pulmonary vasoconstriction (HPV). Disturbances in HPV can cause life-threatening hypoxemia whereas chronic hypoxia triggers lung vascular remodeling and pulmonary hypertension. The signaling cascade of this vitally important mechanism is still unresolved. Using transient receptor potential channel 6 (TRPC6)-deficient mice, we show that this channel is a key regulator of acute HPV as this regulatory mechanism was absent in TRPC6−/− mice whereas the pulmonary vasoconstrictor response to the thromboxane mimetic U46619 was unchanged. Accordingly, induction of regional hypoventilation resulted in severe arterial hypoxemia in TRPC6−/− but not in WT mice. This effect was mirrored by a lack of hypoxia-induced cation influx and currents in smooth-muscle cells from precapillary pulmonary arteries (PASMC) of TRPC6−/− mice. In both WT and TRPC6−/− PASMC hypoxia caused diacylglycerol (DAG) accumulation. DAG seems to exert its action via TRPC6, as DAG kinase inhibition provoked a cation influx only in WT but not in TRPC6−/− PASMC. Notably, chronic hypoxia-induced pulmonary hypertension was independent of TRPC6 activity. We conclude that TRPC6 plays a unique and indispensable role in acute hypoxic pulmonary vasoconstriction. Manipulation of TRPC6 function may thus offer a therapeutic strategy for the control of pulmonary hemodynamics and gas exchange. PMID:17142322

  11. 20-Hydroxyeicosatetraenoic Acid (20-HETE) Modulates Canonical Transient Receptor Potential-6 (TRPC6) Channels in Podocytes

    PubMed Central

    Roshanravan, Hila; Kim, Eun Y.; Dryer, Stuart E.

    2016-01-01

    The arachidonic acid metabolite 20-hydroxyeicosatetraenoic acid (20-HETE) regulates renal function, including changes in glomerular function evoked during tubuloglomerular feedback (TGF). This study describes the cellular actions of 20-HETE on cultured podocytes, assessed by whole-cell recordings from cultured podocytes combined with pharmacological and cell-biological manipulations of cells. Bath superfusion of 20-HETE activates cationic currents that are blocked by the pan-TRP blocker SKF-96365 and by 50 μM La3+, and which are attenuated after siRNA knockdown of TRPC6 subunits. Similar currents are evoked by a membrane-permeable analog of diacylgycerol (OAG), but OAG does not occlude responses to maximally-activating concentrations of 20-HETE (20 μM). Exposure to 20-HETE also increased steady-state surface abundance of TRPC6 subunits in podocytes as assessed by cell-surface biotinylation assays, and increased cytosolic concentrations of reactive oxygen species (ROS). TRPC6 activation by 20-HETE was eliminated in cells pretreated with TEMPOL, a membrane-permeable superoxide dismutase mimic. Activation of TRPC6 by 20-HETE was also blocked when whole-cell recording pipettes contained GDP-βS, indicating a role for either small or heterotrimeric G proteins in the transduction cascade. Responses to 20-HETE were eliminated by siRNA knockdown of podocin, a protein that organizes NADPH oxidase complexes with TRPC6 subunits in this cell type. In summary, modulation of ionic channels in podocytes may contribute to glomerular actions of 20-HETE.

  12. The TRPC6 channel activator hyperforin induces the release of zinc and calcium from mitochondria.

    PubMed

    Tu, Peng; Gibon, Julien; Bouron, Alexandre

    2010-01-01

    Hyperforin, an extract of the medicinal plant hypericum perforatum (also named St John's wort), possesses antidepressant properties. Recent data showed that it elevates the intracellular concentration of Ca(2+) by activating diacylglycerol-sensitive C-class of transient receptor potential (TRPC6) channels without activating the other isoforms (TRPC1, TRPC3, TRPC4, TRPC5, and TRPC7). This study was undertaken to further characterize the cellular neuronal responses induced by hyperforin. Experiments conducted on cortical neurons in primary culture and loaded with fluorescent probes for Ca(2+) (Fluo-4) and Zn(2+) (FluoZin-3) showed that it not only controls the activity of plasma membrane channels but it also mobilizes these two cations from internal pools. Experiments conducted on isolated brain mitochondria indicated that hyperforin, like the inhibitor of oxidative phosphorylation, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), collapses the mitochondrial membrane potential. Furthermore, it promotes the release of Ca(2+) and Zn(2+) from these organelles via a ruthenium red-sensitive transporter. In fact, hyperforin exerts complex actions on CNS neurons. This antidepressant not only triggers the entry of cations via plasma membrane TRPC6 channels but it displays protonophore-like properties. As hyperforin is now use to probe the functions of native TRPC6 channels, our data indicate that caution is required when interpreting results obtained with this antidepressant. PMID:19845832

  13. Role of endogenous TRPC6 channels in Ca2+ signal generation in A7r5 smooth muscle cells.

    PubMed

    Soboloff, Jonathan; Spassova, Maria; Xu, Wen; He, Li-Ping; Cuesta, Natalia; Gill, Donald L

    2005-12-01

    The ubiquitously expressed canonical transient receptor potential (TRPC) ion channels are considered important in Ca2+ signal generation, but their mechanisms of activation and roles remain elusive. Whereas most studies have examined overexpressed TRPC channels, we used molecular, biochemical, and electrophysiological approaches to assess the expression and function of endogenous TRPC channels in A7r5 smooth muscle cells. Real time PCR and Western analyses reveal TRPC6 as the only member of the diacylglycerol-responsive TRPC3/6/7 subfamily of channels expressed at significant levels in A7r5 cells. TRPC1, TRPC4, and TRPC5 were also abundant. An outwardly rectifying, nonselective cation current was activated by phospholipase C-coupled vasopressin receptor activation or by the diacylglycerol analogue, oleoyl-2-acetyl-sn-glycerol (OAG). Introduction of TRPC6 small interfering RNA sequences into A7r5 cells by electroporation led to 90% reduction of TRPC6 transcript and 80% reduction of TRPC6 protein without any detectable compensatory changes in the expression of other TRPC channels. The OAG-activated nonselective cation current was similarly reduced by TRPC6 RNA interference. Intracellular Ca2+ measurements using fura-2 revealed that thapsigargin-induced store-operated Ca2+ entry was unaffected by TRPC6 knockdown, whereas vasopressin-induced Ca2+ entry was suppressed by more than 50%. In contrast, OAG-induced Ca2+ transients were unaffected by TRPC6 knockdown. Nevertheless, OAG-induced Ca2+ entry bore the hallmarks of TRPC6 function; it was inhibited by protein kinase C and blocked by the Src-kinase inhibitor, 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2). Importantly, OAG-induced Ca2+ entry was blocked by the potent L-type Ca2+ channel inhibitor, *nimodipine. Thus, TRPC6 activation probably results primarily in Na ion entry and depolarization, leading to activation of L-type channels as the mediators of Ca2+ entry. Calculations reveal that even

  14. Functional consequences of the over-expression of TRPC6 channels in HEK cells: impact on the homeostasis of zinc.

    PubMed

    Chevallet, Mireille; Jarvis, Louis; Harel, Amélie; Luche, Sylvie; Degot, Sébastien; Chapuis, Violaine; Boulay, Guylain; Rabilloud, Thierry; Bouron, Alexandre

    2014-07-01

    The canonical transient receptor potential 6 (TRPC6) protein is a non-selective cation channel able to transport essential trace elements like iron (Fe) and zinc (Zn) through the plasma membrane. Its over-expression in HEK-293 cells causes an intracellular accumulation of Zn, indicating that it could be involved in Zn transport. This finding prompted us to better understand the role played by TRPC6 in Zn homeostasis. Experiments done using the fluorescent probe FluoZin-3 showed that HEK cells possess an intracellular pool of mobilisable Zn present in compartments sensitive to the vesicular proton pump inhibitor Baf-A, which affects endo/lysosomes. TRPC6 over-expression facilitates the basal uptake of Zn and enhances the size of the pool of Zn sensitive to Baf-A. Quantitative RT-PCR experiments showed that TRPC6 over-expression does not affect the mRNA expression of Zn transporters (ZnT-1, ZnT-5, ZnT-6, ZnT-7, ZnT-9, Zip1, Zip6, Zip7, and Zip14); however it up-regulates the mRNA expression of metallothionein-I and -II. This alters the Zn buffering capacities of the cells as illustrated by the experiments done using the Zn ionophore Na pyrithione. In addition, HEK cells over-expressing TRPC6 grow slower than their parental HEK cells. This feature can be mimicked by growing HEK cells in a culture medium supplemented with 5 μM of Zn acetate. Finally, a proteomic analysis revealed that TRPC6 up-regulates the expression of the actin-associated proteins ezrin and cofilin-1, and changes the organisation of the actin cytoskeleton without changing the cellular actin content. Altogether, these data indicate that TRPC6 is participating in the transport of Zn and influences the Zn storage and buffering capacities of the cells. PMID:24733507

  15. TRPC6 specifically interacts with APP to inhibit its cleavage by γ-secretase and reduce Aβ production

    PubMed Central

    Wang, Junfeng; Lu, Rui; Yang, Jian; Li, Hongyu; He, Zhuohao; Jing, Naihe; Wang, Xiaomin; Wang, Yizheng

    2015-01-01

    Generation of β-amyloid (Aβ) peptide in Alzheimer's disease involves cleavage of amyloid precursor protein (APP) by γ-secretase, a protease known to cleave several substrates, including Notch. Finding specific modulators for γ-secretase could be a potential avenue to treat the disease. Here, we report that transient receptor potential canonical (TRPC) 6 specifically interacts with APP leading to inhibition of its cleavage by γ-secretase and reduction in Aβ production. TRPC6 interacts with APP (C99), but not with Notch, and prevents C99 interaction with presenilin 1 (PS1). A fusion peptide derived from TRPC6 also reduces Aβ levels without effect on Notch cleavage. Crossing APP/PS1 mice with TRPC6 transgenic mice leads to a marked reduction in both plaque load and Aβ levels, and improvement in structural and behavioural impairment. Thus, TRPC6 specifically modulates γ-secretase cleavage of APP and preventing APP (C99) interaction with PS1 via TRPC6 could be a novel strategy to reduce Aβ formation. PMID:26581893

  16. TRPC6 specifically interacts with APP to inhibit its cleavage by γ-secretase and reduce Aβ production.

    PubMed

    Wang, Junfeng; Lu, Rui; Yang, Jian; Li, Hongyu; He, Zhuohao; Jing, Naihe; Wang, Xiaomin; Wang, Yizheng

    2015-01-01

    Generation of β-amyloid (Aβ) peptide in Alzheimer's disease involves cleavage of amyloid precursor protein (APP) by γ-secretase, a protease known to cleave several substrates, including Notch. Finding specific modulators for γ-secretase could be a potential avenue to treat the disease. Here, we report that transient receptor potential canonical (TRPC) 6 specifically interacts with APP leading to inhibition of its cleavage by γ-secretase and reduction in Aβ production. TRPC6 interacts with APP (C99), but not with Notch, and prevents C99 interaction with presenilin 1 (PS1). A fusion peptide derived from TRPC6 also reduces Aβ levels without effect on Notch cleavage. Crossing APP/PS1 mice with TRPC6 transgenic mice leads to a marked reduction in both plaque load and Aβ levels, and improvement in structural and behavioural impairment. Thus, TRPC6 specifically modulates γ-secretase cleavage of APP and preventing APP (C99) interaction with PS1 via TRPC6 could be a novel strategy to reduce Aβ formation. PMID:26581893

  17. Membrane translocation of TRPC6 channels and endothelial migration are regulated by calmodulin and PI3 kinase activation.

    PubMed

    Chaudhuri, Pinaki; Rosenbaum, Michael A; Sinharoy, Pritam; Damron, Derek S; Birnbaumer, Lutz; Graham, Linda M

    2016-02-23

    Lipid oxidation products, including lysophosphatidylcholine (lysoPC), activate canonical transient receptor potential 6 (TRPC6) channels leading to inhibition of endothelial cell (EC) migration in vitro and delayed EC healing of arterial injuries in vivo. The precise mechanism through which lysoPC activates TRPC6 channels is not known, but calmodulin (CaM) contributes to the regulation of TRPC channels. Using site-directed mutagenesis, cDNAs were generated in which Tyr(99) or Tyr(138) of CaM was replaced with Phe, generating mutant CaM, Phe(99)-CaM, or Phe(138)-CaM, respectively. In ECs transiently transfected with pcDNA3.1-myc-His-Phe(99)-CaM, but not in ECs transfected with pcDNA3.1-myc-His-Phe(138)-CaM, the lysoPC-induced TRPC6-CaM dissociation and TRPC6 externalization was disrupted. Also, the lysoPC-induced increase in intracellular calcium concentration was inhibited in ECs transiently transfected with pcDNA3.1-myc-His-Phe(99)-CaM. Blocking phosphorylation of CaM at Tyr(99) also reduced CaM association with the p85 subunit and subsequent activation of phosphatidylinositol 3-kinase (PI3K). This prevented the increase in phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and the translocation of TRPC6 to the cell membrane and reduced the inhibition of EC migration by lysoPC. These findings suggest that lysoPC induces CaM phosphorylation at Tyr(99) by a Src family kinase and that phosphorylated CaM activates PI3K to produce PIP3, which promotes TRPC6 translocation to the cell membrane. PMID:26858457

  18. Membrane translocation of TRPC6 channels and endothelial migration are regulated by calmodulin and PI3 kinase activation

    PubMed Central

    Chaudhuri, Pinaki; Rosenbaum, Michael A.; Sinharoy, Pritam; Damron, Derek S.; Birnbaumer, Lutz; Graham, Linda M.

    2016-01-01

    Lipid oxidation products, including lysophosphatidylcholine (lysoPC), activate canonical transient receptor potential 6 (TRPC6) channels leading to inhibition of endothelial cell (EC) migration in vitro and delayed EC healing of arterial injuries in vivo. The precise mechanism through which lysoPC activates TRPC6 channels is not known, but calmodulin (CaM) contributes to the regulation of TRPC channels. Using site-directed mutagenesis, cDNAs were generated in which Tyr99 or Tyr138 of CaM was replaced with Phe, generating mutant CaM, Phe99-CaM, or Phe138-CaM, respectively. In ECs transiently transfected with pcDNA3.1-myc-His-Phe99-CaM, but not in ECs transfected with pcDNA3.1-myc-His-Phe138-CaM, the lysoPC-induced TRPC6-CaM dissociation and TRPC6 externalization was disrupted. Also, the lysoPC-induced increase in intracellular calcium concentration was inhibited in ECs transiently transfected with pcDNA3.1-myc-His-Phe99-CaM. Blocking phosphorylation of CaM at Tyr99 also reduced CaM association with the p85 subunit and subsequent activation of phosphatidylinositol 3-kinase (PI3K). This prevented the increase in phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and the translocation of TRPC6 to the cell membrane and reduced the inhibition of EC migration by lysoPC. These findings suggest that lysoPC induces CaM phosphorylation at Tyr99 by a Src family kinase and that phosphorylated CaM activates PI3K to produce PIP3, which promotes TRPC6 translocation to the cell membrane. PMID:26858457

  19. A mutation in TRPC6 channels abolishes their activation by hypoosmotic stretch but does not affect activation by diacylglycerol or G protein signaling cascades.

    PubMed

    Wilson, Cory; Dryer, Stuart E

    2014-05-01

    Canonical transient receptor potential-6 (TRPC6) channels have been implicated in the pathogenesis of kidney disease and in the regulation of vascular smooth muscle tone, podocyte function, and a variety of processes in other cell types. The question of whether their gating is intrinsically mechanosensitive has been controversial. In this study we have examined activation of two alleles of TRPC6 transiently expressed in CHO-K1 cells: the wild-type human TRPC6 channel, and TRPC6-N143S, an allele originally identified in a family with autosomal dominant familial focal and segmental glomerulosclerosis (FSGS). We observed that both channel variants carried robust cationic currents that could be evoked by application of membrane-permeable analogs of diacylglycerol (DAG) or by the P2Y receptor agonist ATP. The amplitudes and characteristics of currents evoked by the DAG analog or ATP were indistinguishable in cells expressing the two TRPC6 alleles. By contrast, hypoosmotic stretch evoked robust currents in wild-type TRPC6 channels but had no discernible effect on currents in cells expressing TRPC6-N143S, indicating that the mutant form lacks mechanosensitivity. Coexpression of TRPC6-N143S with wild-type TRPC6 or TRPC3 channels did not alter stretch-evoked responses compared with when TRPC3 channels were expressed by themselves, indicating that TRPC6-N143S does not function as a dominant-negative. These data indicate that mechanical activation and activation evoked by DAG or ATP occur through fundamentally distinct biophysical mechanisms, and they provide support for the hypothesis that protein complexes containing wild-type TRPC6 subunits can be intrinsically mechanosensitive. PMID:24598806

  20. TRPC1 and TRPC6 channels cooperate with TRPV4 to mediate mechanical hyperalgesia and nociceptor sensitization

    PubMed Central

    Alessandri-Haber, Nicole; Dina, Olayinka A.; Chen, Xiaoje; Levine, Jon D.

    2009-01-01

    The transient receptor potential vanilloid 4 (TRPV4) contributes to mechanical hyperalgesia of diverse etiologies, presumably as part of a mechanoreceptor signaling complex (Alessandri-Haber et al., 2008). To investigate the hypothesis that a functional interaction between TRPV4 and stretch-activated ion channels (SACs) is involved in this mechanical transduction mechanism, we used a selective SACs inhibitor, GsMTx-4. Intradermal injection of GsMTx-4 in the rat hind paw reversed the mechanical hyperalgesia induced by intradermal injection of inflammatory mediators. In vivo single fiber recordings showed that GsMTx-4 reversed inflammatory mediator-induced decrease in mechanical threshold in half of sensitized C-fibers. Furthermore, GsMTx-4 reduced hyperalgesia to both mechanical and hypotonic stimuli in different models of inflammatory and neuropathic pain while it had no effect on baseline mechanical nociceptive thresholds. TRPC1 and TRPC6, two GsMTx-4-sensitive SACs are expressed in dorsal root ganglion neurons (DRG). Single-cell RT-PCR showed that messenger RNAs for TRPV4, TRPC1 and TRPC6 are frequently co-expressed in DRG neurons. Spinal intrathecal administration of oligodeoxynucleotides antisense to TRPC1 and TRPC6, like that to TRPV4, reversed the hyperalgesia to mechanical and hypotonic stimuli induced by inflammatory mediators without affecting baseline mechanical nociceptive threshold. However, antisense to TRPC6, but not to TRPC1, reversed the mechanical hyperalgesia induced by a thermal injury or the TRPV4 selective agonist 4α-PDD. We conclude that TRPC1 and TRPC6 channels cooperate with TRPV4 channels to mediate mechanical hyperalgesia and primary afferent nociceptor sensitization although they may have distinctive roles. PMID:19439599

  1. Increased Vascular Smooth Muscle Contractility in TRPC6−/− Mice

    PubMed Central

    Dietrich, Alexander; Mederos y Schnitzler, Michael; Gollasch, Maik; Gross, Volkmar; Storch, Ursula; Dubrovska, Galyna; Obst, Michael; Yildirim, Eda; Salanova, Birgit; Kalwa, Hermann; Essin, Kirill; Pinkenburg, Olaf; Luft, Friedrich C.; Gudermann, Thomas; Birnbaumer, Lutz

    2005-01-01

    Among the TRPC subfamily of TRP (classical transient receptor potential) channels, TRPC3, -6, and -7 are gated by signal transduction pathways that activate C-type phospholipases as well as by direct exposure to diacylglycerols. Since TRPC6 is highly expressed in pulmonary and vascular smooth muscle cells, it represents a likely molecular candidate for receptor-operated cation entry. To define the physiological role of TRPC6, we have developed a TRPC6-deficient mouse model. These mice showed an elevated blood pressure and enhanced agonist-induced contractility of isolated aortic rings as well as cerebral arteries. Smooth muscle cells of TRPC6-deficient mice have higher basal cation entry, increased TRPC-carried cation currents, and more depolarized membrane potentials. This higher basal cation entry, however, was completely abolished by the expression of a TRPC3-specific small interference RNA in primary TRPC6−/− smooth muscle cells. Along these lines, the expression of TRPC3 in wild-type cells resulted in increased basal activity, while TRPC6 expression in TRPC6−/− smooth muscle cells reduced basal cation influx. These findings imply that constitutively active TRPC3-type channels, which are up-regulated in TRPC6-deficient smooth muscle cells, are not able to functionally replace TRPC6. Thus, TRPC6 has distinct nonredundant roles in the control of vascular smooth muscle tone. PMID:16055711

  2. Ins(1,4,5)P3 interacts with PIP2 to regulate activation of TRPC6/C7 channels by diacylglycerol in native vascular myocytes

    PubMed Central

    Ju, Min; Shi, Jian; Saleh, Sohag N; Albert, Anthony P; Large, William A

    2010-01-01

    We investigated synergism between inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) and diacylglycerol (DAG) on TRPC6-like channel activity in rabbit portal vein myocytes using single channel recording and immunoprecipitation techniques. Ins(1,4,5)P3 at 10 μm increased 3-fold TRPC6-like activity induced by 10 μm 1-oleoyl-2-acetyl-sn-glycerol (OAG), a DAG analogue. Ins(1,4,5)P3 had no effect on OAG-induced TRPC6 activity in mesenteric artery myocytes. Anti-TRPC6 and anti-TRPC7 antibodies blocked channel activity in portal vein but only anti-TRPC6 inhibited activity in mesenteric artery. TRPC6 and TRPC7 proteins strongly associated in portal vein but only weakly associated in mesenteric artery tissue lysates. Therefore in portal vein the conductance consists of TRPC6/C7 subunits, while OAG activates a homomeric TRPC6 channel in mesenteric artery myocytes. Wortmannin at 20 μm reduced phosphatidylinositol 4,5-bisphosphate (PIP2) association with TRPC6 and TRPC7, and produced a 40-fold increase in OAG-induced TRPC6/C7 activity. Anti-PIP2 antibodies evoked TRPC6/C7 activity, which was blocked by U73122, a phospholipase C inhibitor. DiC8-PIP2, a water-soluble PIP2 analogue, inhibited OAG-induced TRPC6/C7 activity with an IC50 of 0.74 μm. Ins(1,4,5)P3 rescued OAG-induced TRPC6/C7 activity from inhibition by diC8-PIP2 in portal vein myocytes, and this was not prevented by the Ins(1,4,5)P3 receptor antagonist heparin. In contrast, Ins(1,4,5)P3 did not overcome diC8-PIP2-induced inhibition of TRPC6 activity in mesenteric artery myocytes. 2,3,6-Tri-O-butyryl-Ins(1,4,5)P3/AM (6-Ins(1,4,5)P3), a cell-permeant analogue of Ins(1,4,5)P3, at 10 μm increased TRPC6/C7 activity in portal vein and reduced association between TRPC7 and PIP2, but not TRPC6 and PIP2. In contrast, 10 μm OAG reduced association between TRPC6 and PIP2, but not between TRPC7 and PIP2. The present work provides the first evidence that Ins(1,4,5)P3 modulates native TRPC channel activity through removal of the

  3. TRPC6 channel activation promotes neonatal glomerular mesangial cell apoptosis via calcineurin/NFAT and FasL/Fas signaling pathways

    PubMed Central

    Soni, Hitesh; Adebiyi, Adebowale

    2016-01-01

    Glomerular mesangial cell (GMC) proliferation and death are involved in the pathogenesis of glomerular disorders. The mechanisms that control GMC survival are poorly understood, but may include signal transduction pathways that are modulated by changes in intracellular Ca2+ ([Ca2+]i) concentration. In this study, we investigated whether activation of the canonical transient receptor potential (TRPC) 6 channels and successive [Ca2+]i elevation alter neonatal GMC survival. Hyperforin (HF)-induced TRPC6 channel activation increased [Ca2+]i concentration, inhibited proliferation, and triggered apoptotic cell death in primary neonatal pig GMCs. HF-induced neonatal GMC apoptosis was not associated with oxidative stress. However, HF-induced TRPC6 channel activation stimulated nuclear translocation of the nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1). HF also increased cell death surface receptor Fas ligand (FasL) level and caspase-8 activity in the cells; effects mitigated by [Ca2+]i chelator BAPTA, calcineurin/NFAT inhibitor VIVIT, and TRPC6 channel knockdown. Accordingly, HF-induced neonatal GMC apoptosis was attenuated by BAPTA, VIVIT, Fas blocking antibody, and a caspase-3/7 inhibitor. These findings suggest that TRPC6 channel-dependent [Ca2+]i elevation and the ensuing induction of the calcineurin/NFAT, FasL/Fas, and caspase signaling cascades promote neonatal pig GMC apoptosis. PMID:27383564

  4. Gq signaling causes glomerular injury by activating TRPC6

    PubMed Central

    Wang, Liming; Jirka, Grant; Rosenberg, Paul B.; Buckley, Anne F.; Gomez, Jose A.; Fields, Timothy A.; Winn, Michelle P.; Spurney, Robert F.

    2015-01-01

    Familial forms of focal segmental glomerulosclerosis (FSGS) have been linked to gain-of-function mutations in the gene encoding the transient receptor potential channel C6 (TRPC6). GPCRs coupled to Gq signaling activate TRPC6, suggesting that Gq-dependent TRPC6 activation underlies glomerular diseases. Here, we developed a murine model in which a constitutively active Gq α subunit (GqQ209L, referred to herein as GqQ>L) is specifically expressed in podocytes and examined the effects of this mutation in response to puromycin aminonucleoside (PAN) nephrosis. We found that compared with control animals, animals expressing GqQ>L exhibited robust albuminuria, structural features of FSGS, and reduced numbers of glomerular podocytes. Gq activation stimulated calcineurin (CN) activity, resulting in CN-dependent upregulation of TRPC6 in murine kidneys. Deletion of TRPC6 in GqQ>L-expressing mice prevented FSGS development and inhibited both tubular damage and podocyte loss induced by PAN nephrosis. Similarly, administration of the CN inhibitor FK506 reduced proteinuria and tubular injury but had more modest effects on glomerular pathology and podocyte numbers in animals with constitutive Gq activation. Moreover, these Gq-dependent effects on podocyte injury were generalizable to diabetic kidney disease, as expression of GqQ>L promoted albuminuria, mesangial expansion, and increased glomerular basement membrane width in diabetic mice. Together, these results suggest that targeting Gq/TRPC6 signaling may have therapeutic benefits for the treatment of glomerular diseases. PMID:25844902

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

    SciTech Connect

    Sun, Yi-hua; Li, Yong-quan; Feng, Shan-li; Li, Bao-xin; Pan, Zhen-wei; Xu, Chang-qing; Li, Ting-ting; Yang, Bao-feng

    2010-04-16

    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.

  6. Phospholipase C Epsilon (PLCε) Induced TRPC6 Activation: A Common but Redundant Mechanism in Primary Podocytes

    PubMed Central

    Kalwa, Hermann; Storch, Ursula; Demleitner, Jana; Fiedler, Susanne; Mayer, Tim; Kannler, Martina; Fahlbusch, Meike; Barth, Holger; Smrcka, Alan; Hildebrandt, Friedhelm; Gudermann, Thomas; Dietrich, Alexander

    2016-01-01

    In eukaryotic cells, activation of phospholipase C (PLC)-coupled membrane receptors by hormones leads to an increase in the intracellular Ca2+ concentration [Ca2+]i. Catalytic activity of PLCs results in the hydrolysis of phosphatidylinositol 4,5-bisphosphate to generate inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG) which opens DAG-sensitive classical transient receptor channels 3, 6, and 7 (TRPC3/6/7), initiating Ca2+ influx from the extracellular space. Patients with focal segmental glomerulosclerosis (FSGS) express gain-of-function mutants of TRPC6, while others carry loss-of-function mutants of PLCε, raising the intriguing possibility that both proteins interact and might work in the same signalling pathway. While TRPC6 activation by PLCβ and PLCγ isozymes was extensively studied, the role of PLCε in TRPC6 activation remains elusive. TRPC6 was co-immunoprecipitated with PLCε in a heterologous overexpression system in HEK293 cells as well as in freshly isolated murine podocytes. Receptor-operated TRPC6 currents in HEK293 cells expressing TRPC6 were reduced by a specific PLCε siRNA and by a PLCε loss-of-function mutant isolated from a patient with FSGS. PLCε-induced TRPC6 activation was also identified in murine embryonic fibroblasts (MEFs) lacking Gαq/11 proteins. Further analysis of the signal transduction pathway revealed a Gα12/13 Rho-GEF activation which induced Rho-mediated PLCε stimulation. Therefore, we identified a new pathway for TRPC6 activation by PLCε. PLCε-/- podocytes however, were undistinguishable from WT podocytes in their angiotensin II-induced formation of actin stress fibers and their GTPγS-induced TRPC6 activation, pointing to a redundant role of PLCε-mediated TRPC6 activation at least in podocytes. PMID:25521631

  7. Modeling non-syndromic autism and the impact of TRPC6 disruption in human neurons

    PubMed Central

    Griesi-Oliveira, Karina; Acab, Allan; Gupta, Abha R.; Sunaga, Daniele Yumi; Chailangkarn, Thanathom; Nicol, Xavier; Nunez, Yanelli; Walker, Michael F.; Murdoch, John D.; Sanders, Stephan J.; Fernandez, Thomas V.; Ji, Weizhen; Lifton, Richard P.; Vadasz, Estevão; Dietrich, Alexander; Pradhan, Dennis; Song, Hongjun; Ming, Guo-li; Guoe, Xiang; Haddad, Gabriel; Marchetto, Maria C. N.; Spitzer, Nicholas; Passos-Bueno, Maria Rita; State, Matthew W.; Muotri, Alysson R.

    2014-01-01

    An increasing number of genetic variants have been implicated in autism spectrum disorders (ASD), and the functional study of such variants will be critical for the elucidation of autism pathophysiology. Here, we report a de novo balanced translocation disruption of TRPC6, a cation channel, in a non-syndromic autistic individual. Using multiple models, such as dental pulp cells, iPSC-derived neuronal cells and mouse models, we demonstrate that TRPC6 reduction or haploinsufficiency leads to altered neuronal development, morphology, and function. The observed neuronal phenotypes could then be rescued by TRPC6 complementation and by treatment with IGF1 or hyperforin, a TRPC6-specific agonist, suggesting that ASD individuals with alterations in this pathway might benefit from these drugs. We also demonstrate that MeCP2 levels affect TRPC6 expression. Mutations in MeCP2 cause Rett syndrome, revealing common pathways among ASDs. Genetic sequencing of TRPC6 in 1041 ASD individuals and 2872 controls revealed significantly more nonsynonymous mutations in the ASD population, and identified loss-of-function mutations with incomplete penetrance in two patients. Taken together, these findings suggest that TRPC6 is a novel predisposing gene for ASD that may act in a multiple-hit model. This is the first study to use iPSC-derived human neurons to model non-syndromic ASD and illustrate the potential of modeling genetically complex sporadic diseases using such cells. PMID:25385366

  8. Second Messenger-Operated Calcium Entry Through TRPC6.

    PubMed

    Bouron, Alexandre; Chauvet, Sylvain; Dryer, Stuart; Rosado, Juan A

    2016-01-01

    Canonical transient receptor potential 6 (TRPC6) proteins assemble into heteromultimeric structures forming non-selective cation channels. In addition, many TRPC6-interacting proteins have been identified like some enzymes, channels, pumps, cytoskeleton-associated proteins, immunophilins, or cholesterol-binding proteins, indicating that TRPC6 are engaged into macromolecular complexes. Depending on the cell type and the experimental conditions used, TRPC6 activity has been reported to be controlled by diverse modalities. For instance, the second messenger diacylglycerol, store-depletion, the plant extract hyperforin or H2O2 have all been shown to trigger the opening of TRPC6 channels. A well-characterized consequence of TRPC6 activation is the elevation of the cytosolic concentration of Ca(2+). This latter response can reflect the entry of Ca(2+) through open TRPC6 channels but it can also be due to the Na(+)/Ca(2+) exchanger (operating in its reverse mode) or voltage-gated Ca(2+) channels (recruited in response to a TRPC6-mediated depolarization). Although TRPC6 controls a diverse array of biological functions in many tissues and cell types, its pathophysiological functions are far from being fully understood. This chapter covers some key features of TRPC6, with a special emphasis on their biological significance in kidney and blood cells. PMID:27161231

  9. Modeling non-syndromic autism and the impact of TRPC6 disruption in human neurons.

    PubMed

    Griesi-Oliveira, K; Acab, A; Gupta, A R; Sunaga, D Y; Chailangkarn, T; Nicol, X; Nunez, Y; Walker, M F; Murdoch, J D; Sanders, S J; Fernandez, T V; Ji, W; Lifton, R P; Vadasz, E; Dietrich, A; Pradhan, D; Song, H; Ming, G-L; Gu, X; Haddad, G; Marchetto, M C N; Spitzer, N; Passos-Bueno, M R; State, M W; Muotri, A R

    2015-11-01

    An increasing number of genetic variants have been implicated in autism spectrum disorders (ASDs), and the functional study of such variants will be critical for the elucidation of autism pathophysiology. Here, we report a de novo balanced translocation disruption of TRPC6, a cation channel, in a non-syndromic autistic individual. Using multiple models, such as dental pulp cells, induced pluripotent stem cell (iPSC)-derived neuronal cells and mouse models, we demonstrate that TRPC6 reduction or haploinsufficiency leads to altered neuronal development, morphology and function. The observed neuronal phenotypes could then be rescued by TRPC6 complementation and by treatment with insulin-like growth factor-1 or hyperforin, a TRPC6-specific agonist, suggesting that ASD individuals with alterations in this pathway may benefit from these drugs. We also demonstrate that methyl CpG binding protein-2 (MeCP2) levels affect TRPC6 expression. Mutations in MeCP2 cause Rett syndrome, revealing common pathways among ASDs. Genetic sequencing of TRPC6 in 1041 ASD individuals and 2872 controls revealed significantly more nonsynonymous mutations in the ASD population, and identified loss-of-function mutations with incomplete penetrance in two patients. Taken together, these findings suggest that TRPC6 is a novel predisposing gene for ASD that may act in a multiple-hit model. This is the first study to use iPSC-derived human neurons to model non-syndromic ASD and illustrate the potential of modeling genetically complex sporadic diseases using such cells. PMID:25385366

  10. Immunohistochemical localization of TRPC6 in the rat substantia nigra.

    PubMed

    Giampà, Carmela; DeMarch, Zena; Patassini, Stefano; Bernardi, Giorgio; Fusco, Francesca R

    2007-09-13

    Transient receptor potential channels (TRPC) are plasma membrane, nonselective cationic channels and have been proposed as candidates involved in the regulation of cellular Ca2+ influx [D.E. Clapham, L.W. Runnels, C., Strubing, The TRP ion channel family, Nat. Rev. Neurosci. 2 (2001) 387-396; A. Martorana, C. Giampa, Z. DeMarch, M.T. Viscomi, S. Patassini, G. Sancesario, G. Bernardi, F.R. Fusco, Distribution of TRPC1 receptors in dendrites of rat substantia nigra: a confocal and electron microscopy study, Eur. J. Neurosci. 24 (2006) 732-738]. Studies on regional localization patterns of TRPCs are necessary to provide helpful guidelines for correlating current types with particular channels. In this study, we examined the distribution of one particular member of TRPC superfamily, namely, TRPC6, in the substantia nigra of normal rat brain. Single and double label immunohistochemistry were employed to perform both light and confocal microscopy observations. Our single label studies showed that, in the substantia nigra, TRPC6 labeled the perikarya with a diffuse and intense immunoreaction product distributed throughout cell cytoplasm whereas only a light immunostaining was observed in the cell nuclei. No labeling of axon or terminals was observed, although TRPC6 was evenly distributed in the neuropil. Our dual label studies showed a TRPC6 immunoreactivity pattern that was localized into the proximal dendrites and axon hillock of the large dopaminergic neurons identified by TH immunoreaction. Furthermore, our double label immunofluorescence study for TRPC6 and mGluR1 showed a complete co-localization of the two markers in the substantia nigra. Moreover, TRPC6 did not co-localize with synaptophysin. Thus, our study shows the postsynaptic localization of TRPC6 and its association with mGluR1 in the midbrain dopamine neurons. PMID:17723267

  11. Identification and Validation of Larixyl Acetate as a Potent TRPC6 Inhibitor.

    PubMed

    Urban, Nicole; Wang, Liming; Kwiek, Sandra; Rademann, Jörg; Kuebler, Wolfgang M; Schaefer, Michael

    2016-01-01

    Classical or canonical transient receptor potential 6 (TRPC6), a nonselective and Ca(2+)-permeable cation channel, mediates pathophysiological responses within pulmonary and renal diseases that are still poorly controlled by current medication. Thus, controlling TRPC6 activity may provide a promising and challenging pharmacological approach. Recently identified chemical entities have demonstrated that TRPC6 is pharmacologically targetable. However, isotype-selectivity with regard to its closest relative, TRPC3, is difficult to achieve. Reasoning that balsams, essential oils, or incense materials that are traditionally used for inhalation may contain biologic activities to block TRPC6 activity, we embarked on a natural compound strategy to identify new TRPC6-blocking chemical entities. Within several preparations of plant extracts, a strong TRPC6-inhibitory activity was found in conifer balsams. The biologic activity was associated with nonvolatile resins, but not with essential oils. Of various conifers, the larch balsam was unique in displaying a marked TRPC6-prevalent mode of action. By testing the main constituents of larch resin, we identified larixol and larixyl acetate as blockers of Ca(2+) entry and ionic currents through diacylglycerol- or receptor-activated recombinant TRPC6 channels, exhibiting approximately 12- and 5-fold selectivity compared with its closest relatives TRPC3 and TRPC7, respectively. No significant inhibition of more distantly related TRPV or TRPM channels was seen. The potent inhibition of recombinant TRPC6 by larixyl acetate (IC50 = 0.1-0.6 µM) was confirmed for native TRPC6-like [Ca(2+)]i signals in diacylglycerol-stimulated rat pulmonary artery smooth muscle cells. In isolated mouse lungs, larix-6-yl monoacetate (CAS 4608-49-5; larixyl acetate; 5 µM) prevented acute hypoxia-induced vasoconstriction. We conclude that larch-derived labdane-type diterpenes are TRPC6-selective inhibitors and may represent a starting point for

  12. TRPC6 G757D Loss-of-Function Mutation Associates with FSGS.

    PubMed

    Riehle, Marc; Büscher, Anja K; Gohlke, Björn-Oliver; Kaßmann, Mario; Kolatsi-Joannou, Maria; Bräsen, Jan H; Nagel, Mato; Becker, Jan U; Winyard, Paul; Hoyer, Peter F; Preissner, Robert; Krautwurst, Dietmar; Gollasch, Maik; Weber, Stefanie; Harteneck, Christian

    2016-09-01

    FSGS is a CKD with heavy proteinuria that eventually progresses to ESRD. Hereditary forms of FSGS have been linked to mutations in the transient receptor potential cation channel, subfamily C, member 6 (TRPC6) gene encoding a nonselective cation channel. Most of these TRPC6 mutations cause a gain-of-function phenotype, leading to calcium-triggered podocyte cell death, but the underlying molecular mechanisms are unclear. We studied the molecular effect of disease-related mutations using tridimensional in silico modeling of tetrameric TRPC6. Our results indicated that G757 is localized in a domain forming a TRPC6-TRPC6 interface and predicted that the amino acid exchange G757D causes local steric hindrance and disruption of the channel complex. Notably, functional characterization of model interface domain mutants suggested a loss-of-function phenotype. We then characterized 19 human FSGS-related TRPC6 mutations, the majority of which caused gain-of-function mutations. However, five mutations (N125S, L395A, G757D, L780P, and R895L) caused a loss-of-function phenotype. Coexpression of wild-type TRPC6 and TRPC6 G757D, mimicking heterozygosity observed in patients, revealed a dominant negative effect of TRPC6 G757D. Our comprehensive analysis of human disease-causing TRPC6 mutations reveals loss of TRPC6 function as an additional concept of hereditary FSGS and provides molecular insights into the mechanism responsible for the loss-of-function phenotype of TRPC6 G757D in humans. PMID:26892346

  13. Expression of TRPC6 and BDNF in Cortical Lesions From Patients With Focal Cortical Dysplasia

    PubMed Central

    Zheng, Da-Hai; Guo, Wei; Sun, Fei-Ji; Xu, Guang-Zhen; Zang, Zhen-Le; Shu, Hai-Feng

    2016-01-01

    Focal cortical dysplasia (FCD) likely results from abnormal migration of neural progenitor cells originating from the subventricular zone. To elucidate the roles in molecules that are involved in neural migration pathway abnormalities in FCDs, we investigated the expression patterns of transient receptor potential canonical channel 6 (TRPC6) and brain-derived neurotrophic factor (BDNF) in cortical lesions from FCD patients and in samples of normal control cortex. TRPC6 and BDNF mRNA and protein levels were increased in FCD lesions. By immunohistochemistry, they were strongly expressed in microcolumns, heterotopic neurons, dysmorphic neurons, and balloon cells (BCs). Colocalization assays revealed that most of the misshapen TRPC6-positive or heterotopic cells had a neuronal lineage with the exception of TRPC6-positive FCDiib patient BCs, which had both neuronal and glial features. Most TRPC6-positive cells were glutamatergic neurons. There was also greater expression of calmodulin-dependent kinase IV (CaMKIV), the downstream factor of TRPC6, in FCD lesions, suggesting that TRPC6 expression promoted dendritic growth and the development of dendritic spines and excitatory synapses via the CaMKIV-CREB pathway in FCD. Thus, overexpression of BDNF and TRPC6 and activation of the TRPC6 signal transduction pathway in cortical lesions of FCD patients may contribute to FC pathogenesis and epileptogenesis. PMID:27288906

  14. Post-transcriptional silencing of TRPC1 ion channel gene by RNA interference upregulates TRPC6 expression and store-operated Ca2+ entry in A7r5 vascular smooth muscle cells.

    PubMed

    Selli, Cigdem; Erac, Yasemin; Kosova, Buket; Tosun, Metiner

    2009-01-01

    This study investigates functional consequences of TRPC1 ion channel downregulation observed in aging rat aorta by employing RNA interference in cultured vascular smooth muscle cells. For this purpose, A7r5 aortic smooth muscle cells were used in quantitative gene and protein expression as well as in functional analyses. According to quantitative RT-PCR results, TRPC3, TRPC4 and TRPC5 mRNAs were not at detectable levels. In siTRPC1-transfected cells, TRPC1 mRNA and protein levels were decreased by 40% and 64%; however, those of TRPC6 were drastically increased by 100% and 200%, respectively. In fura-2-loaded TRPC1 knockdown cells, despite the decreased TRPC1 levels, cyclopiazonic acid-induced Ca2+ entry and store-operated Ca2+ entry following Ca2+ addition were elevated by 77% and 135%, respectively. Results suggest that decrease in TRPC1 may be compensated by upregulated TRPC6 that possibly takes part in store-operated Ca2+ entry in vascular smooth muscle cells. PMID:19386284

  15. Functional and structural analysis of mice TRPC6 with human analogue through homology modelling.

    PubMed

    Chigurupati, Soumya; Bhasin, Arnima; Inampudi, Krishna Kishore; Asuthkar, Swapna; Madarampalli, Bhanupriya; Kammili, Ramana Kumar; Velpula, Kiran Kumar

    2014-01-01

    Homology models are increasingly used to determine structural and functional relationships of genes and proteins in biomedical research. In the current study, for the first time, we compared the TRPC6 gene in mouse and human. The protein encoded by this gene forms a receptor activated calcium channel in cell membrane. Defects in this gene have been implicated in a wide range of diseases including glioblastomas. To determine the structural similarities in mouse and human TRPC6, we used standard bioinformatics tools such as fold prediction to identify the protein 3D structure, sequence-structure comparison, and prediction of template and protein structure. We also used glioblastoma cell line U373MG and human glioblastoma tumour tissues to study the expression of TRPC6 in disease conditions to implicate this gene in pathological ailment. Based on the results we conclude that human TRPC6 contains 90% identity and 93% similarity with mouse TRPC6, suggesting that this protein is well conserved in these two species. These isoforms likely demonstrate similar mechanisms in regulating gene expression; thus TRPC6 studies in mice may be extrapolated to humans. PMID:24589838

  16. Astragaloside IV prevents high glucose-induced podocyte apoptosis via downregulation of TRPC6.

    PubMed

    Yao, Xing-Mei; Liu, Yu-Jun; Wang, Yun-Man; Wang, Hao; Zhu, Bing-Bing; Liang, Yong-Ping; Yao, Wei-Guo; Yu, Hui; Wang, Nian-Song; Zhang, Xue-Mei; Peng, Wen

    2016-06-01

    Diabetic nephropathy (DN) is one of the most important causes of end‑stage renal disease. Astragaloside IV (AS-IV) is a saponin isolated from Astragalus membranaceus, which possesses various pharmacological activities. AS‑IV prevents podocyte apoptosis and ameliorates renal injury in DN; however, few studies have focused on its effects on ion channels. The transient receptor potential channel 6 (TRPC6) is an important Ca2+‑permeable ion channel in podocytes, which is involved in high glucose (HG)-induced podocyte apoptosis. The aim of the present study was to investigate whether AS‑IV prevented HG‑induced podocyte apoptosis via TRPC6. Cultured podocytes were pre‑treated with 10, 20 or 40 µM AS‑IV for 1 h prior to HG exposure for 24 h. Apoptosis, cell viability, expression of TRPC6, nuclear factor of activated T cells (NFAT2) and B‑cell lymphoma 2‑associated X protein (Bax), as well as the intracellular Ca2+ concentration were subsequently analyzed. The results indicated that HG induced podocyte apoptosis and upregulation of TRPC6, and increased intracellular Ca2+. Furthermore, enhanced NFAT2 and Bax expression was detected. Conversely, AS‑IV protected HG‑induced podocyte apoptosis, downregulated TRPC6 expression and suppressed intracellular Ca2+ in HG-stimulated podocytes. AS‑IV also suppressed NFAT2 and Bax expression. These results suggest that AS‑IV may prevent HG-induced podocyte apoptosis via downregulation of TRPC6, which is possibly mediated via the calcineurin/NFAT signaling pathway. PMID:27109610

  17. Regulation of Multi-drug Resistance in hepatocellular carcinoma cells is TRPC6/Calcium Dependent

    PubMed Central

    Wen, Liang; Liang, Chao; Chen, Enjiang; Chen, Wei; Liang, Feng; Zhi, Xiao; Wei, Tao; Xue, Fei; Li, Guogang; Yang, Qi; Gong, Weihua; Feng, Xinhua; Bai, Xueli; Liang, Tingbo

    2016-01-01

    Hepatocellular carcinoma (HCC) is notoriously refractory to chemotherapy because of its tendency to develop multi-drug resistance (MDR), whose various underlying mechanisms make it difficult to target. The calcium signalling pathway is associated with many cellular biological activities, and is also a critical player in cancer. However, its role in modulating tumour MDR remains unclear. In this study, stimulation by doxorubicin, hypoxia and ionizing radiation was used to induce MDR in HCC cells. A sustained aggregation of intracellular calcium was observed upon these stimuli, while inhibition of calcium signalling enhanced the cells’ sensitivity to various drugs by attenuating epithelial-mesenchymal transition (EMT), Hif1-α signalling and DNA damage repair. The effect of calcium signalling is mediated via transient receptor potential canonical 6 (TRPC6), a subtype of calcium-permeable channel. An in vivo xenograft model of HCC further confirmed that inhibiting TRPC6 enhanced the efficacy of doxorubicin. In addition, we deduced that STAT3 activation is a downstream signalling pathway in MDR. Collectively, this study demonstrated that the various mechanisms regulating MDR in HCC cells are calcium dependent through the TRPC6/calcium/STAT3 pathway. We propose that targeting TRPC6 in HCC may be a novel antineoplastic strategy, especially combined with chemotherapy. PMID:27011063

  18. Infantile hypertrophic pyloric stenosis: evaluation of three positional candidate genes, TRPC1, TRPC5 and TRPC6, by association analysis and re-sequencing.

    PubMed

    Everett, Kate V; Chioza, Barry A; Georgoula, Christina; Reece, Ashley; Gardiner, R Mark; Chung, Eddie M K

    2009-12-01

    Infantile hypertrophic pyloric stenosis (IHPS) is the most common inherited form of gastrointestinal obstruction in infancy with a striking male preponderance. Infants present with vomiting due to gastric outlet obstruction caused by hypertrophy of the smooth muscle of the pylorus. Two loci specific to extended pedigrees displaying autosomal dominant inheritance have been identified. A genome scan identified loci on chromosomes 11q14-q22 and Xq23-q24 which are predicted to be responsible for a subset of smaller families with IHPS demonstrating non-Mendelian inheritance. The two linked chromosomal regions both harbour functional candidate genes which are members of the canonical transient receptor potential (TRPC) family of ion channels. Both TRPC5 (Xq23-q24) and TRPC6 (11q14-q22) have a potential role in smooth muscle control and hypertrophy. Here, we report suggestive evidence for a third locus on chromosome 3q12-q25 (Zmax = 2.7, p < 0.004), a region which harbours a third TRPC gene, TRPC1. Fine mapping of all three genes using a tagSNP approach and re-sequencing identified a SNP in the promoter region of TRPC6 and a missense variant in exon 4 of TRPC6 which may be putative causal variants. PMID:19701773

  19. Phosphodiesterase 5 inhibition ameliorates angiontensin II-induced podocyte dysmotility via the protein kinase G-mediated downregulation of TRPC6 activity.

    PubMed

    Hall, Gentzon; Rowell, Janelle; Farinelli, Federica; Gbadegesin, Rasheed A; Lavin, Peter; Wu, Guanghong; Homstad, Alison; Malone, Andrew; Lindsey, Thomas; Jiang, Ruiji; Spurney, Robert; Tomaselli, Gordon F; Kass, David A; Winn, Michelle P

    2014-06-15

    The emerging role of the transient receptor potential cation channel isotype 6 (TRPC6) as a central contributor to various pathological processes affecting podocytes has generated interest in the development of therapeutics to modulate its function. Recent insights into the regulation of TRPC6 have revealed PKG as a potent negative modulator of TRPC6 conductance and associated signaling via its phosphorylation at two highly conserved amino acid residues: Thr(69)/Thr(70) (Thr(69) in mice and Thr(70) in humans) and Ser(321)/Ser(322) (Ser(321) in mice and Ser(322) in humans). Here, we tested the role of PKG in modulating TRPC6-dependent responses in primary and conditionally immortalized mouse podocytes. TRPC6 was phosphorylated at Thr(69) in nonstimulated podocytes, but this declined upon ANG II stimulation or overexpression of constitutively active calcineurin phosphatase. ANG II induced podocyte motility in an in vitro wound assay, and this was reduced 30-60% in cells overexpressing a phosphomimetic mutant TRPC6 (TRPC6T70E/S322E) or activated PKG (P < 0.05). Pretreatment of podocytes with the PKG agonists S-nitroso-N-acetyl-dl-penicillamine (nitric oxide donor), 8-bromo-cGMP, Bay 41-2772 (soluble guanylate cyclase activator), or phosphodiesterase 5 (PDE5) inhibitor 4-{[3',4'-(methylenedioxy)benzyl]amino}[7]-6-methoxyquinazoline attenuated ANG II-induced Thr(69) dephosphorylation and also inhibited TRPC6-dependent podocyte motility by 30-60%. These data reveal that PKG activation strategies, including PDE5 inhibition, ameliorate ANG II-induced podocyte dysmotility by targeting TRPC6 in podocytes, highlighting the potential therapeutic utility of these approaches to treat hyperactive TRPC6-dependent glomerular disease. PMID:24740790

  20. The contribution of TRPC1, TRPC3, TRPC5 and TRPC6 to touch and hearing

    PubMed Central

    Sexton, Jane E.; Desmonds, Terri; Quick, Kathryn; Taylor, Ruth; Abramowitz, Joel; Forge, Andy; Kros, Corné J.; Birnbaumer, Lutz; Wood, John N.

    2016-01-01

    Transient receptor potential channels have diverse roles in mechanosensation. Evidence is accumulating that members of the canonical subfamily of TRP channels (TRPC) are involved in touch and hearing. Characteristic features of TRP channels include their high structural homology and their propensity to form heteromeric complexes which suggests potential functional redundancy. We previously showed that TRPC3 and TRPC6 double knockout animals have deficits in light touch and hearing whilst single knockouts were apparently normal. We have extended these studies to analyse deficits in global quadruple TRPC1, 3, 5 and 6 null mutant mice. We examined both touch and hearing in behavioural and electrophysiological assays, and provide evidence that the quadruple knockout mice have larger deficits than the TRPC3 TRPC6 double knockouts. Mechano-electrical transducer currents of cochlear outer hair cells were however normal. This suggests that TRPC1, TRPC3, TRPC5 and TRPC6 channels contribute to cutaneous and auditory mechanosensation in a combinatorial manner, but have no direct role in cochlear mechanotransduction. PMID:26520460

  1. The contribution of TRPC1, TRPC3, TRPC5 and TRPC6 to touch and hearing.

    PubMed

    Sexton, Jane E; Desmonds, Terri; Quick, Kathryn; Taylor, Ruth; Abramowitz, Joel; Forge, Andy; Kros, Corné J; Birnbaumer, Lutz; Wood, John N

    2016-01-01

    Transient receptor potential channels have diverse roles in mechanosensation. Evidence is accumulating that members of the canonical subfamily of TRP channels (TRPC) are involved in touch and hearing. Characteristic features of TRP channels include their high structural homology and their propensity to form heteromeric complexes which suggests potential functional redundancy. We previously showed that TRPC3 and TRPC6 double knockout animals have deficits in light touch and hearing whilst single knockouts were apparently normal. We have extended these studies to analyse deficits in global quadruple TRPC1, 3, 5 and 6 null mutant mice. We examined both touch and hearing in behavioural and electrophysiological assays, and provide evidence that the quadruple knockout mice have larger deficits than the TRPC3 TRPC6 double knockouts. Mechano-electrical transducer currents of cochlear outer hair cells were however normal. This suggests that TRPC1, TRPC3, TRPC5 and TRPC6 channels contribute to cutaneous and auditory mechanosensation in a combinatorial manner, but have no direct role in cochlear mechanotransduction. PMID:26520460

  2. Mechanosensitive ATP release from hemichannels and Ca²⁺ influx through TRPC6 accelerate wound closure in keratinocytes.

    PubMed

    Takada, Hiroya; Furuya, Kishio; Sokabe, Masahiro

    2014-10-01

    Cutaneous wound healing is accelerated by exogenous mechanical forces and is impaired in TRPC6-knockout mice. Therefore, we designed experiments to determine how mechanical force and TRPC6 channels contribute to wound healing using HaCaT keratinocytes. HaCaT cells were pretreated with hyperforin, a major component of a traditional herbal medicine for wound healing and also a TRPC6 activator, and cultured in an elastic chamber. At 3 h after scratching the confluent cell layer, the ATP release and intracellular Ca(2+) increases in response to stretching (20%) were live-imaged. ATP release was observed only in cells at the frontier facing the scar. The diffusion of released ATP caused intercellular Ca(2+) waves that propagated towards the rear cells in a P2Y-receptor-dependent manner. The Ca(2+) response and wound healing were inhibited by ATP diphosphohydrolase apyrase, the P2Y antagonist suramin, the hemichannel blocker CBX and the TRPC6 inhibitor diC8-PIP2. Finally, the hemichannel-permeable dye calcein was taken up only by ATP-releasing cells. These results suggest that stretch-accelerated wound closure is due to the ATP release through mechanosensitive hemichannels from the foremost cells and the subsequent Ca(2+) waves mediated by P2Y and TRPC6 activation. PMID:25097230

  3. Proteomic analysis of TRPC5- and TRPC6-binding partners reveals interaction with the plasmalemmal Na(+)/K(+)-ATPase.

    PubMed

    Goel, Monu; Sinkins, William; Keightley, Andrew; Kinter, Michael; Schilling, William P

    2005-10-01

    Mammalian transient receptor potential canonical (TRPC) genes encode a family of nonselective cation channels that are activated following stimulation of G-protein-coupled membrane receptors linked to phospholipase C. In Drosophila photoreceptor cells, TRP channels are found in large, multimolecular signaling complexes in association with the PDZ-containing scaffolding protein, INAD. A similar mammalian TRPC "signalplex" has been proposed, but has yet to be defined. In the present study, affinity-purified polyclonal antibodies against TRPC5 and TRPC6 were used to immunoprecipitate signalplex components from rat brain lysates. Immunoprecipitated proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, digested with trypsin, and sequenced by mass spectrometry. Proteins identified in the immunoprecipitates included cytoskeletal proteins spectrin, myosin, actin, drebrin, tubulin, and neurabin; endocytic vesicle-associated proteins clathrin, dynamin and AP-2; and the plasmalemmal Na(+)/K(+)-ATPase (NKA) pump. Several of these interactions were confirmed by reciprocal immunoprecipitation followed by Western blot analysis. In lysates from rat kidney, TRPC6, but not TRPC3, was found to coimmunoprecipitate with the NKA pump. Likewise, TRPC6, stably expressed in human embryonic kidney (HEK) cells, coimmunoprecipitated with endogenous NKA and colocalized with the pump to the plasmalemma when examined by immunofluorescence microscopy. Cell surface biotinylation experiments in intact HEK cells, confirmed that both the Na(+) pump and TRPC6 were present in the surface membrane and appeared to interact. Lastly, TRPC6 coimmunoprecipitated with the NKA pump when the proteins were coexpressed in Spodoptera frugiperda insect cells using recombinant baculoviruses. These observations suggest that TRPC6 and the Na(+) pump are part of a functional complex that may be involved in ion transport and homeostasis in both the brain and kidney. PMID:16025302

  4. In silico analysis of functional nsSNPs in human TRPC6 gene associated with steroid resistant nephrotic syndrome.

    PubMed

    Joshi, Bhoomi B; Koringa, Prakash G; Mistry, Kinnari N; Patel, Amrut K; Gang, Sishir; Joshi, Chaitanya G

    2015-11-01

    The aim of the present study is to identify functional non-synonymous SNPs of TRPC6 gene using various in silico approaches. These SNPs are believed to have a direct impact on protein stability through conformation changes. Transient receptor potential cation channel-6 (TRPC6) is one of the proteins that plays a key role causing focal segmental glomerulosclerosis (FSGS) associated with the steroid-resistant nephritic syndrome (SRNS). Data of TRPC6 was collected from dbSNP and further used to investigate a damaging effect using SIFT, PolyPhen, PROVEAN, and PANTHER. The comparative analysis predicted that two functional SNPs "rs35857503 at position N157T and rs36111323 at position A404V" showed a damaging effect (score of 0.096-1.00).We modeled the 3D structure of TRPC6 using a SWISS-MODEL workspace and validated it via PROCHECK to get a Ramachandran plot (83.0% residues in the most favored region, 12.7% in additionally allowed regions, 2.3% in a generously allowed region and 2.0% were in a disallowed region). QMEAN (0.311) and MUSTER (10.06) scores were under acceptable limits. Putative functional SNPs that may possibly undergo post-translation modifications were also identified in TRPC6 protein. It was found that mutation at N157T can lead to alteration in glycation whereas mutation at A404V was present at a ligand binding site. Additionally, I-Mutant showed a decrease in stability for these nsSNPs upon mutation, thus suggesting that the N157T and A404V variants of TRPC6 could directly or indirectly destabilize the amino acid interactions causing functional deviations of protein to some extent. PMID:26127002

  5. Acute ethanol induces apoptosis by stimulating TRPC6 via elevation of superoxide in oxygenated podocytes.

    PubMed

    Lu, Xiao-Yu; Liu, Bing-Chen; Wang, Li-Hua; Yang, Li-Li; Bao, Qing; Zhai, Yu-Jia; Alli, Abdel A; Thai, Tiffany L; Eaton, Douglas C; Wang, Wei-Zhi; Ma, He-Ping

    2015-05-01

    Our recent studies indicate that hydrogen peroxide (H2O2) only at high concentrations can cause oxidative stress in renal epithelial cells and induce apoptosis of podocytes. Consistently, the present study shows that H2O2, even at 1 mM, failed to induce intracellular oxidative stress and apoptosis of the podocytes due to efficient activity of catalase, an enzyme which degrades H2O2 to produce water and oxygen (O2). However, H2O2 acted as a source of O2 to allow acute ethanol to induce superoxide production and cause apoptosis of the podocytes. In contrast, acute ethanol alone did not elevate intracellular superoxide, even though it stimulates expression and translocation of p47phox to the plasma membrane. Inhibition of catalase abolished not only O2 production from H2O2 degradation, but also NOX2-dependent superoxide production in the podocytes challenged by both H2O2 and acute ethanol. In parallel, acute ethanol in the presence of H2O2, but neither ethanol nor H2O2 alone, stimulated transient receptor potential canonical 6 (TRPC6) channels and caused TRPC6-dependent elevation of intracellular Ca2+. These data suggest that exogenous H2O2 does not induce oxidative stress due to rapid degradation to produce O2 in the podocytes, but the oxygenated podocytes become sensitive to acute ethanol challenge and undergo apoptosis via a TRPC6-dependent elevation of intracellular Ca2+. Since cultured podocytes are considered in hypoxic conditions, H2O2 may be used as a source of O2 to establish an ischemia-reperfusion model in some type of cultured cells in which H2O2 does not directly induce intracellular oxidative stress. PMID:25601712

  6. Balancing Calcium Signals through TRPC5 and TRPC6 in Podocytes

    PubMed Central

    2011-01-01

    Calcium (Ca2+) ions are important mediators of cellular homeostasis owing to their ability to elicit a dynamic, transient, and tightly regulated range of biochemical responses. More than a decade ago, a nonselective, Ca2+-permeable, cationic conductance was identified in podocytes downstream of angiotensin II (Ang II) signaling, but its molecular structure remained elusive. Six years ago, transient receptor potential canonical 6 (TRPC6) mutations were found in families with hereditary FSGS, and TRPC5 and TRPC6 channels are now known as the Ca2+ influx pathways for this previously described, nonselective, cationic current in podocytes. Ang II activation engages this Ca2+ influx to modulate the actin cytoskeleton in podocytes. These discoveries dovetail with previously described regulation of actin dynamics by the Ca2+-activated phosphatase, calcineurin, and the emergence of Rho GTPases as critical regulators of podocyte function in health and disease. Understanding the interconnected signaling regulated by Ca2+ currents offers potential new therapeutic targets and highlights the notion that synergistic therapies targeting multiple levels of biochemistry may be useful in treating proteinuric kidney disease. PMID:21980113

  7. A Critical Role for the Transient Receptor Potential Channel Type 6 in Human Platelet Activation

    PubMed Central

    Conlon, Christine; Khasawneh, Fadi T.

    2015-01-01

    While calcium signaling is known to play vital roles in platelet function, the mechanisms underlying its receptor-operated calcium entry component (ROCE) remain poorly understood. It has been proposed, but never proven in platelets, that the canonical transient receptor potential channel-6 (TRPC6) mediates ROCE. Nonetheless, we have previously shown that the mouse TRPC6 regulates hemostasis, thrombogenesis by regulating platelet aggregation. In the present studies, we used a pharmacological approach to characterize the role of TRPC6 in human platelet biology. Thus, interestingly, we observed that a TRPC6 inhibitor exerted significant inhibitory effects on human platelet aggregation in a thromboxane receptor (TPR)-selective manner; no additional inhibition was observed in the presence of the calcium chelator BAPTA. This inhibitor also significantly inhibited human platelet secretion (dense and alpha granules), integrin IIb-IIIa, Akt and ERK phosphorylation, again, in a TPR-selective manner; no effects were observed in response to ADP receptor stimulation. Furthermore, there was a causal relationship between these inhibitory effects, and the capacity of the TRPC6 inhibitor to abrogate elevation in intracellular calcium, that was again found to be TPR-specific. This effect was not found to be due to antagonism of TPR, as the TRPC6 inhibitor did not displace the radiolabeled antagonist [3H]SQ29,548 from its binding sites. Finally, our studies also revealed that TRPC6 regulates human clot retraction, as well as physiological hemostasis and thrombus formation, in mice. Taken together, our findings demonstrate, for the first time, that TRPC6 directly regulates TPR-dependent ROCE and platelet function. Moreover, these data highlight TRPC6 as a novel promising therapeutic strategy for managing thrombotic disorders. PMID:25928636

  8. The canonical transient receptor potential 6 channel as a putative phosphatidylinositol 3,4,5-trisphosphate-sensitive calcium entry system.

    PubMed

    Tseng, Ping-Hui; Lin, Ho-Pi; Hu, Hongzhen; Wang, Chunbo; Zhu, Michael Xi; Chen, Ching-Shih

    2004-09-21

    We previously reported that phosphatidylinositol 3,4,5-trisphosphate (PIP(3)), a lipid product of phosphoinositide 3-kinase (PI3K), induced Ca(2+) influx via a noncapacitative pathway in platelets, Jurkat T cells, and RBL-2H3 mast cells. The identity of this Ca(2+) influx system, however, remains unclear. Here, we investigate a potential link between PIP(3)-sensitive Ca(2+) entry and the canonical transient receptor potential (TRPC) channels by developing stable human embryonic kidney (HEK) 293 cell lines expressing TRPC1, TRPC3, TRPC5, and TRPC6. Two lines of evidence support TRPC6 as a putative target by which PIP(3) induces Ca(2+) influx. First, Fura-2 fluorometric Ca(2+) analysis shows the ability of PIP(3) to selectively stimulate [Ca(2+)](i) increase in TRPC6-expressing cells. Second, pull-down analysis indicates specific interactions between biotin-PIP(3) and TRPC6 protein. Our data indicate that PIP(3) activates store-independent Ca(2+) entry in TRPC6 cells via a nonselective cation channel. Although the activating effect of PIP(3) on TRPC6 is reminiscent to that of 1-oleoyl-2-acetyl-sn-glycerol, this activation is not attributable to the diacylglycerol substructure of PIP(3) since other phosphoinositides failed to trigger Ca(2+) responses. The PIP(3)-activated Ca(2+) entry is inhibited by known TRPC6 inhibitors such as Gd(3+) and SKF96365 and is independent of IP(3) production. Furthermore, we demonstrated that TRPC6 overexpression or antisense downregulation significantly alters the amplitude of PIP(3)- and anti-CD3-activated Ca(2+) responses in Jurkat T cells. Consequently, the link between TRPC6 and PIP(3)-mediated Ca(2+) entry provides a framework to account for an intimate relationship between PI3K and PLCgamma in initiating Ca(2+) response to agonist stimulation in T lymphocytes. PMID:15362854

  9. Genetic Interactions Between TRPC6 and NPHS1 Variants Affect Posttransplant Risk of Recurrent Focal Segmental Glomerulosclerosis.

    PubMed

    Sun, Z J; Ng, K H; Liao, P; Zhang, Y; Ng, J L; Liu, I D; Tan, P H; Chong, S S C; Chan, Y H; Liu, J; Davila, S; Heng, C K; Jordan, S C; Soong, T W; Yap, H K

    2015-12-01

    Individuals with TRPC6 mutations have variable phenotypes, ranging from healthy carrier to focal segmental glomerulosclerosis (FSGS) leading to renal failure. Here, we describe a family where six members had a novel TRPC6 p.R68W (c.202C>T) mutation, two of whom had renal failure from FSGS, and one had proteinuria. One healthy carrier donated a kidney to her sister. Both donor and recipient had no proteinuria at 20 years posttransplant. Two synonymous NPHS1 polymorphisms, rs2285450 (c.294C>T) and rs437168 (c.2289C>T) segregated with renal failure in this family. These variants had higher allele frequencies in 97 unrelated patients with nephrotic syndrome or FSGS compared to 224 controls. Using patch-clamp experiments in HEK293 and podocytes, we showed that the p.R68W mutation increased TRPC6 current amplitudes, which may be explained by enhanced TRPC6 surface expression. Additionally, while wild-type nephrin suppressed TRPC6 currents, this ability was lost in the presence of NPHS1 c.294C>T polymorphism. When cells were transfected according to combined TRPC6 and NPHS1 genotypes in the family, those representing the donor had lower TRPC6 currents than cells representing the recipient, suggesting that interactions between TRPC6 and NPHS1 variants could possibly account for the variable penetrance of TRPC6 mutations and the absence of recurrence in the graft. PMID:26147534

  10. TRPC6 is required for hypoxia-induced basal intracellular calcium concentration elevation, and for the proliferation and migration of rat distal pulmonary venous smooth muscle cells.

    PubMed

    Wang, Qingjie; Wang, Dong; Yan, Gaoling; Sun, Ling; Tang, Chengchun

    2016-02-01

    Hypoxia induces pulmonary vasoconstriction and reconstruction in the pulmonary arteries and pulmonary veins (PVs), and elevation of intracellular calcium concentration ([Ca2+]i) is a primary factor of these processes. In the present study, the role of transient receptor potential cation channels (TRPCs) in mediating the hypoxia-induced elevation of [Ca2+]i in rat distal pulmonary venous smooth muscle cells (PVSMCs) was investigated. Rats with chronic hypoxic pulmonary hypertension (CHPH) were used for in vivo experiments, and PVSMCs were isolated for in vitro experiments. [Ca2+]i was measured using fura-2-based fluorescence calcium imaging. Reverse transcription-quantitative polymerase chain reaction and western blotting were used to detect the mRNA and protein expression levels of TRPCs. Methyl thiazolyl tetrazolium and Transwell assays were used to investigate the proliferation and migration of PVSMCs, respectively. The results of the present study demonstrated that TRPC6 was increased in the distal PVs of CHPH rats, and in PVSMCs exposed to hypoxic conditions (4% O2, 72 h); however, TRPC1 was not. The 1-oleoyl-2-acetyl-sn-glycerol-induced [Ca2+]i elevation was increased in PVSMCs isolated from CHPH rats and in PVSMCs cultured under hypoxic conditions (4% O2, 72 h). Hypoxia induced PVSMC [Ca2+]i elevation, proliferation and migration. These alterations were inhibited following TRPC6 knockdown. Results from the present study suggest that TRPC6 expression is increased during chronic hypoxia, which contributes to Ca2+ entry into the cell, thus promoting proliferation and migration of PVSMCs. PMID:26718737

  11. TRPC6 is required for hypoxia-induced basal intracellular calcium concentration elevation, and for the proliferation and migration of rat distal pulmonary venous smooth muscle cells

    PubMed Central

    WANG, QINGJIE; WANG, DONG; YAN, GAOLING; SUN, LING; TANG, CHENGCHUN

    2016-01-01

    Hypoxia induces pulmonary vasoconstriction and reconstruction in the pulmonary arteries and pulmonary veins (PVs), and elevation of intracellular calcium concentration ([Ca2+]i) is a primary factor of these processes. In the present study, the role of transient receptor potential cation channels (TRPCs) in mediating the hypoxia-induced elevation of [Ca2+]i in rat distal pulmonary venous smooth muscle cells (PVSMCs) was investigated. Rats with chronic hypoxic pulmonary hypertension (CHPH) were used for in vivo experiments, and PVSMCs were isolated for in vitro experiments. [Ca2+]i was measured using fura-2-based fluorescence calcium imaging. Reverse transcription-quantitative polymerase chain reaction and western blotting were used to detect the mRNA and protein expression levels of TRPCs. Methyl thiazolyl tetrazolium and Transwell assays were used to investigate the proliferation and migration of PVSMCs, respectively. The results of the present study demonstrated that TRPC6 was increased in the distal PVs of CHPH rats, and in PVSMCs exposed to hypoxic conditions (4% O2, 72 h); however, TRPC1 was not. The 1-oleoyl-2-acetyl-sn-glycerol-induced [Ca2+]i elevation was increased in PVSMCs isolated from CHPH rats and in PVSMCs cultured under hypoxic conditions (4% O2, 72 h). Hypoxia induced PVSMC [Ca2+]i elevation, proliferation and migration. These alterations were inhibited following TRPC6 knockdown. Results from the present study suggest that TRPC6 expression is increased during chronic hypoxia, which contributes to Ca2+ entry into the cell, thus promoting proliferation and migration of PVSMCs. PMID:26718737

  12. MicroRNA-26a prevents endothelial cell apoptosis by directly targeting TRPC6 in the setting of atherosclerosis

    PubMed Central

    Zhang, Yong; Qin, Wei; Zhang, Longyin; Wu, Xianxian; Du, Ning; Hu, Yingying; Li, Xiaoguang; Shen, Nannan; Xiao, Dan; Zhang, Haiying; Li, Zhange; Zhang, Yue; Yang, Huan; Gao, Feng; Du, Zhimin; Xu, Chaoqian; Yang, Baofeng

    2015-01-01

    Atherosclerosis, a chronic inflammatory disease, is the major cause of life-threatening complications such as myocardial infarction and stroke. Endothelial apoptosis plays a vital role in the initiation and progression of atherosclerotic lesions. Although a subset of microRNAs (miRs) have been identified as critical regulators of atherosclerosis, studies on their participation in endothelial apoptosis in atherosclerosis have been limited. In our study, we found that miR-26a expression was substantially reduced in the aortic intima of ApoE−/− mice fed with a high-fat diet (HFD). Treatment of human aortic endothelial cells (HAECs) with oxidized low-density lipoprotein (ox-LDL) suppressed miR-26a expression. Forced expression of miR-26a inhibited endothelial apoptosis as evidenced by MTT assay and TUNEL staining results. Further analysis identified TRPC6 as a target of miR-26a, and TRPC6 overexpression abolished the anti-apoptotic effect of miR-26a. Moreover, the cytosolic calcium and the mitochondrial apoptotic pathway were found to mediate the beneficial effects of miR-26a on endothelial apoptosis. Taken together, our study reveals a novel role of miR-26a in endothelial apoptosis and indicates a therapeutic potential of miR-26a for atherosclerosis associated with apoptotic cell death. PMID:25801675

  13. MicroRNA-26a prevents endothelial cell apoptosis by directly targeting TRPC6 in the setting of atherosclerosis

    NASA Astrophysics Data System (ADS)

    Zhang, Yong; Qin, Wei; Zhang, Longyin; Wu, Xianxian; Du, Ning; Hu, Yingying; Li, Xiaoguang; Shen, Nannan; Xiao, Dan; Zhang, Haiying; Li, Zhange; Zhang, Yue; Yang, Huan; Gao, Feng; Du, Zhimin; Xu, Chaoqian; Yang, Baofeng

    2015-03-01

    Atherosclerosis, a chronic inflammatory disease, is the major cause of life-threatening complications such as myocardial infarction and stroke. Endothelial apoptosis plays a vital role in the initiation and progression of atherosclerotic lesions. Although a subset of microRNAs (miRs) have been identified as critical regulators of atherosclerosis, studies on their participation in endothelial apoptosis in atherosclerosis have been limited. In our study, we found that miR-26a expression was substantially reduced in the aortic intima of ApoE-/- mice fed with a high-fat diet (HFD). Treatment of human aortic endothelial cells (HAECs) with oxidized low-density lipoprotein (ox-LDL) suppressed miR-26a expression. Forced expression of miR-26a inhibited endothelial apoptosis as evidenced by MTT assay and TUNEL staining results. Further analysis identified TRPC6 as a target of miR-26a, and TRPC6 overexpression abolished the anti-apoptotic effect of miR-26a. Moreover, the cytosolic calcium and the mitochondrial apoptotic pathway were found to mediate the beneficial effects of miR-26a on endothelial apoptosis. Taken together, our study reveals a novel role of miR-26a in endothelial apoptosis and indicates a therapeutic potential of miR-26a for atherosclerosis associated with apoptotic cell death.

  14. Role of transient receptor potential canonical 6 (TRPC6) in non-transferrin-bound iron uptake in neuronal phenotype PC12 cells.

    PubMed Central

    Mwanjewe, James; Grover, Ashok K

    2004-01-01

    Cells take up transferrin-bound iron or NTBI (non-transferrin-bound iron). After treatment with NGF (nerve growth factor), PC12 cells exhibited a neuronal phenotype and an increase in the NTBI uptake (55Fe2+ or 55Fe3+). We loaded the cells with the dye calcein, whose fluorescence increases in the presence of Ca2+ but is quenched with Fe2+ or Fe3+. When examined using calcein fluorescence or radioactive iron, DAG (diacylglycerol)-stimulated NTBI entry was more in NGF-treated PC12 cells compared with untreated cells. All experiments were performed at 1.5 mM extracellular Ca2+. Nramp2 (natural-resistance-associated macrophage protein 2) mRNA expression did not change after the NGF treatment. Expression of the bivalent cation entry protein TRPC6 (transient receptor potential canonical 6) was detected only in the NGF-treated cells. To verify that increased NTBI uptake depended on TRPC6, we examined whether transfecting HEK-293 (human embryonic kidney 293) cells with TRPC6 also increased the NTBI (55Fe) uptake. We also cotransfected HEK-293 cells with two plasmids, one expressing TRPC6 and the other expressing the fluorescent protein DsRED2 to identify the transfected cells. Challenging the calcein-loaded HEK-293 cells (which intrinsically express the a1-adrenergic receptors) with phenylephrine or a cell-permeant DAG increased the fluorescence signal more rapidly in transfected cells compared with untransfected cells. However, when iron (Fe2+ and Fe3+) was added before adding phenylephrine or DAG, the fluorescence intensity decreased more rapidly in transfected cells compared with untransfected cells, thereby indicating a greater stimulation of the NTBI uptake in cells expressing TRPC6. We postulate that the increase in the NTBI entry into neuronal PC12 cells is through TRPC6, a pathway that is unique since it is receptor-stimulated. Since neuronal cells express TRPC6, this pathway may have a role in neurotoxicity. PMID:14640978

  15. High glucose modifies transient receptor potential canonical type 6 channels via increased oxidative stress and syndecan-4 in human podocytes.

    PubMed

    Thilo, Florian; Lee, Marlene; Xia, Shengqiang; Zakrzewicz, Andreas; Tepel, Martin

    2014-07-18

    Transient receptor potential canonical (TRPC) channels type 6 play an important role in the function of human podocytes. Diabetic nephropathy is characterized by altered TRPC6 expression and functions of podocytes. Thus, we hypothesized that high glucose modifies TRPC6 channels via increased oxidative stress and syndecan-4 (SDC-4) in human podocytes. Human podocytes were exposed to control conditions (5.6 mmol/L D-glucose), high glucose (30 mmol/L D-glucose or L-glucose), 100 μmol/L peroxynitrite, or high glucose and the superoxide dismutase mimetic tempol (100 μmol/L). TRPC6 and SDC-4 transcripts and protein expression were measured using RT-PCR and in-cell Western assay. Intracellular reactive oxygen species (ROS) and cytosolic calcium were measured using fluorescent dye techniques. High D-glucose increased TRPC6 transcripts to 8.66±4.08 (p<0.05) and TRPC6 protein expression to 1.44±0.07 (p<0.05) without altering SDC-4 transcripts or protein expression. The D-glucose induced increase of TRPC6 expression was blocked by tempol. Increased oxidative stress using peroxynitrite significantly increased TRPC6 transcripts to 4.29±1.26 (p<0.05) and TRPC6 protein expression to 1.28±0.05 (p<0.05) without altering SDC-4 transcripts or protein expression. In human podocytes transfected with scrambled siRNA, high D-glucose increased ROS after 90 min to 3.55±0.08 arbitrary units while 5.6 mmol/L D-glucose increased ROS to 2.49±0.09 (p<0.001) only. The increase in ROS was inhibited by tempol and by SDC-4 knockdown. High glucose modifies TRPC6 channels and ROS production via SDC-4 in human podocytes. PMID:24942878

  16. Co-expression of non-selective cation channels of the transient receptor potential canonical family in central aminergic neurones.

    PubMed

    Sergeeva, Olga A; Korotkova, Tatiana M; Scherer, Annette; Brown, Ritchie E; Haas, Helmut L

    2003-06-01

    The mammalian transient receptor potential canonical (TRPC) group of channels is a family of Ca2+-permeable cation channels that are activated following receptor-mediated stimulation of different isoforms of phospholipase C. In vitro TRPC proteins can form hetero- or homo-oligomeric channels. We performed single-cell RT-PCR analysis to reveal the co-expression of seven TRPC channels in identified rat aminergic neurones. All serotonergic neurones of the dorsal raphe (DR), the majority of histaminergic (tuberomamillary nucleus; TMN) and dopaminergic cells of the ventral tegmental area (VTA), as well as some GABAergic neurones from the VTA, expressed at least one variant of TRPC channels. No TRPC channel expression was found in the locus coeruleus. In raphe neurones TRPC6 and TRPC5 mRNAs occurred most frequently. In VTA and TMN co-expression of TRPC4 with TRPC5 and TRPC6 with TRPC7 was not found in individual neurones (in contrast to the whole-brain regions). Their co-expression in non-neuronal cells could not be excluded. The neonatal TRPC3 subunit was rarely seen. In DR, but not in the other nuclei studied, the expression of orexin receptors correlated with the expression of TRPC channels. We conclude that several TRPC channel populations exist in individual neurones and that their subunit co-expression pattern is region and cell-type specific. PMID:12787073

  17. Podocyte Purinergic P2X4 Channels Are Mechanotransducers That Mediate Cytoskeletal Disorganization.

    PubMed

    Forst, Anna-Lena; Olteanu, Vlad Sorin; Mollet, Géraldine; Wlodkowski, Tanja; Schaefer, Franz; Dietrich, Alexander; Reiser, Jochen; Gudermann, Thomas; Mederos y Schnitzler, Michael; Storch, Ursula

    2016-03-01

    Podocytes are specialized, highly differentiated epithelial cells in the kidney glomerulus that are exposed to glomerular capillary pressure and possible increases in mechanical load. The proteins sensing mechanical forces in podocytes are unconfirmed, but the classic transient receptor potential channel 6 (TRPC6) interacting with the MEC-2 homolog podocin may form a mechanosensitive ion channel complex in podocytes. Here, we observed that podocytes respond to mechanical stimulation with increased intracellular calcium concentrations and increased inward cation currents. However, TRPC6-deficient podocytes responded in a manner similar to that of control podocytes, and mechanically induced currents were unaffected by genetic inactivation of TRPC1/3/6 or administration of the broad-range TRPC blocker SKF-96365. Instead, mechanically induced currents were significantly decreased by the specific P2X purinoceptor 4 (P2X4) blocker 5-BDBD. Moreover, mechanical P2X4 channel activation depended on cholesterol and podocin and was inhibited by stabilization of the actin cytoskeleton. Because P2X4 channels are not intrinsically mechanosensitive, we investigated whether podocytes release ATP upon mechanical stimulation using a fluorometric approach. Indeed, mechanically induced ATP release from podocytes was observed. Furthermore, 5-BDBD attenuated mechanically induced reorganization of the actin cytoskeleton. Altogether, our findings reveal a TRPC channel-independent role of P2X4 channels as mechanotransducers in podocytes. PMID:26160898

  18. Function and regulation of endothelin type A receptor-operated transient receptor potential canonical channels.

    PubMed

    Horinouchi, Takahiro; Terada, Koji; Higa, Tsunaki; Aoyagi, Hiroyuki; Nishiya, Tadashi; Suzuki, Hiroyuki; Miwa, Soichi

    2011-01-01

    The purpose of this study is to identify transient receptor potential canonical (TRPC) channels responsible for receptor-operated Ca(2+) entry (ROCE) triggered by activation of endothelin type A receptor (ET(A)R) and to clarify the importance of calmodulin (CaM) / inositol 1,4,5-trisphosphate (IP(3)) receptor binding (CIRB) domain at the C terminus of TRPC channels in ET(A)R-activated channel regulation. In HEK293 cells coexpressing ET(A)R and one of seven TRPC isoforms, ET(A)R stimulation induced ROCE through TRPC3, TRPC5, TRPC6, and TRPC7. The TRPC3- and TRPC6-mediated ROCE was inhibited by selective inhibitors of G(q) protein, phospholipase C (PLC), and CaM. The CIRB domain deletion mutants of TRPC3 and TRPC6 failed to induce ET(A)R-mediated ROCE. Either deletion of the CIRB domain or pharmacological inhibition of CaM did not inhibit the targeting of these channels to the plasma membrane. These results suggest that 1) TRPC3, TRPC5, TRPC6, and TRPC7 can function as ET(A)R-operated Ca(2+) channels; 2) G(q) protein, PLC, and CaM are involved in TRPC3- and TRPC6-mediated ROCE; 3) ET(A)R-mediated activation of TRPC3 and TRPC6 requires the CIRB domain; and 4) abolition of ET(A)R-induced ROCE by CIRB domain deletion and CaM inhibition is due to loss of CaM binding to the channels but not loss of cell surface TRPC3 and TRPC6. PMID:22129540

  19. Identification of TRPC6 as a possible candidate target gene within an amplicon at 11q21-q22.2 for migratory capacity in head and neck squamous cell carcinomas

    PubMed Central

    2013-01-01

    Background Cytogenetic and gene expression analyses in head and neck squamous cell carcinomas (HNSCC) have allowed identification of genomic aberrations that may contribute to cancer pathophysiology. Nevertheless, the molecular consequences of numerous genetic alterations still remain unclear. Methods To identify novel genes implicated in HNSCC pathogenesis, we analyzed the genomic alterations present in five HNSCC-derived cell lines by array CGH, and compared high level focal gene amplifications with gene expression levels to identify genes whose expression is directly impacted by these genetic events. Next, we knocked down TRPC6, one of the most highly amplified and over-expressed genes, to characterize the biological roles of TRPC6 in carcinogenesis. Finally, real time PCR was performed to determine TRPC6 gene dosage and mRNA levels in normal mucosa and human HNSCC tissues. Results The data showed that the HNSCC-derived cell lines carry most of the recurrent genomic abnormalities previously described in primary tumors. High-level genomic amplifications were found at four chromosomal sites (11q21-q22.2, 18p11.31-p11.21, 19p13.2-p13.13, and 21q11) with associated gene expression changes in selective candidate genes suggesting that they may play an important role in the malignant behavior of HNSCC. One of the most dramatic alterations of gene transcription involved the TRPC6 gene (located at 11q21-q22.2) which has been recently implicated in tumour invasiveness. siRNA-induced knockdown of TRPC6 expression in HNSCC-derived cells dramatically inhibited HNSCC-cell invasion but did not significantly alter cell proliferation. Importantly, amplification and concomitant overexpression of TRPC6 was also found in HNSCC tumour samples. Conclusions Altogether, these data show that TRPC6 is likely to be a target for 11q21–22.2 amplification that confers enhanced invasive behavior to HNSCC cells. Therefore, TRPC6 may be a promising therapeutic target in the treatment of HNSCC

  20. Screening of Transient Receptor Potential Canonical Channel Activators Identifies Novel Neurotrophic Piperazine Compounds.

    PubMed

    Sawamura, Seishiro; Hatano, Masahiko; Takada, Yoshinori; Hino, Kyosuke; Kawamura, Tetsuya; Tanikawa, Jun; Nakagawa, Hiroshi; Hase, Hideharu; Nakao, Akito; Hirano, Mitsuru; Rotrattanadumrong, Rachapun; Kiyonaka, Shigeki; Mori, Masayuki X; Nishida, Motohiro; Hu, Yaopeng; Inoue, Ryuji; Nagata, Ryu; Mori, Yasuo

    2016-03-01

    Transient receptor potential canonical (TRPC) proteins form Ca(2+)-permeable cation channels activated upon stimulation of metabotropic receptors coupled to phospholipase C. Among the TRPC subfamily, TRPC3 and TRPC6 channels activated directly by diacylglycerol (DAG) play important roles in brain-derived neurotrophic factor (BDNF) signaling, promoting neuronal development and survival. In various disease models, BDNF restores neurologic deficits, but its therapeutic potential is limited by its poor pharmacokinetic profile. Elucidation of a framework for designing small molecules, which elicit BDNF-like activity via TRPC3 and TRPC6, establishes a solid basis to overcome this limitation. We discovered, through library screening, a group of piperazine-derived compounds that activate DAG-activated TRPC3/TRPC6/TRPC7 channels. The compounds [4-(5-chloro-2-methylphenyl)piperazin-1-yl](3-fluorophenyl)methanone (PPZ1) and 2-[4-(2,3-dimethylphenyl)piperazin-1-yl]-N-(2-ethoxyphenyl)acetamide (PPZ2) activated, in a dose-dependent manner, recombinant TRPC3/TRPC6/TRPC7 channels, but not other TRPCs, in human embryonic kidney cells. PPZ2 activated native TRPC6-like channels in smooth muscle cells isolated from rabbit portal vein. Also, PPZ2 evoked cation currents and Ca(2+) influx in rat cultured central neurons. Strikingly, both compounds induced BDNF-like neurite growth and neuroprotection, which were abolished by a knockdown or inhibition of TRPC3/TRPC6/TRPC7 in cultured neurons. Inhibitors of Ca(2+) signaling pathways, except calcineurin, impaired neurite outgrowth promotion induced by PPZ compounds. PPZ2 increased activation of the Ca(2+)-dependent transcription factor, cAMP response element-binding protein. These findings suggest that Ca(2+) signaling mediated by activation of DAG-activated TRPC channels underlies neurotrophic effects of PPZ compounds. Thus, piperazine-derived activators of DAG-activated TRPC channels provide important insights for future development of a

  1. Association of immunophilins with mammalian TRPC channels.

    PubMed

    Sinkins, William G; Goel, Monu; Estacion, Mark; Schilling, William P

    2004-08-13

    Drosophila photoreceptor channels TRP and TRPL are held in a large signalplex by the scaffolding protein, INAD. Immunophilin FKBP59, another member of the signalplex, binds to both INAD and TRPL. Mutation P702Q or P709Q in the highly conserved TRPL sequence (701)LPPPFNVLP(709), eliminates TRPL interaction with FKBP59. The first leucylprolyl (LP) dipeptide in this region is conserved in mammalian TRPC channel proteins. However, the second LP is changed to isoleucylprolyl (IP) in TRPC1, -C4, and -C5, and valylprolyl (VP) in TRPC3, -C6, and -C7. The purpose of the present study was to determine if mammalian FKBP12 or FKBP52 interact with TRPC channel proteins. Using TRPC-specific antibodies, immunoprecipitations from Sf9 cells individually co-expressing each of the TRPC proteins along with the immunophilins showed that TRPC3, -C6, and -C7 interact with FKBP12, whereas TRPC1, -C4, and -C5 interact with FKBP52. The binding of FKBP12 and FKBP52 was specific and could be displaced by the immunosuppressant drug FK506, at concentrations of 0.5 and 10 microm, respectively. To evaluate TRPC-immunophilin interactions in vivo, immunoprecipitations were performed using membrane lysates of rat cerebral cortex. FKBP12 co-immunoprecipitated with TRPC3, -C6, and -C7 from rat brain, whereas FKBP52 was found to associate with TRPC1, -C4, and -C5. The association of immunophilins with the TRPC channels in rat brain lysates could be displaced by FK506. Receptor-mediated activation of TRPC6, stably expressed in HEK cells, was significantly inhibited by FK506, which also disrupted interaction between TRPC6 and the endogenous immunophilin found in HEK cells. Pro to Gln mutations in the first LP dipeptide in the putative FKBP binding domain eliminated FKBP12 and FKBP52 interaction with TRPC3 and -C6, and TRPC1 and -C4, respectively. However, mutual swap of VP and IP in TRPC3 and TRPC5 did not alter the association or the selectivity of the channels for their respective immunophilin binding

  2. STIM1 heteromultimerizes TRPC channels to determine their function as store-operated channels.

    PubMed

    Yuan, Joseph P; Zeng, Weizhong; Huang, Guo N; Worley, Paul F; Muallem, Shmuel

    2007-06-01

    Stromal interacting molecule 1 (STIM1) is a Ca(2+) sensor that conveys the Ca(2+) load of the endoplasmic reticulum to store-operated channels (SOCs) at the plasma membrane. Here, we report that STIM1 binds TRPC1, TRPC4 and TRPC5 and determines their function as SOCs. Inhibition of STIM1 function inhibits activation of TRPC5 by receptor stimulation, but not by La(3+), suggesting that STIM1 is obligatory for activation of TRPC channels by agonists, but STIM1 is not essential for channel function. Through a distinct mechanism, STIM1 also regulates TRPC3 and TRPC6. STIM1 does not bind TRPC3 and TRPC6, and regulates their function indirectly by mediating the heteromultimerization of TRPC3 with TRPC1 and TRPC6 with TRPC4. TRPC7 is not regulated by STIM1. We propose a new definition of SOCs, as channels that are regulated by STIM1 and require the store depletion-mediated clustering of STIM1. By this definition, all TRPC channels, except TRPC7, function as SOCs. PMID:17486119

  3. TRPC channels function independently of STIM1 and Orai1.

    PubMed

    DeHaven, Wayne I; Jones, Bertina F; Petranka, John G; Smyth, Jeremy T; Tomita, Takuro; Bird, Gary S; Putney, James W

    2009-05-15

    Recent studies have defined roles for STIM1 and Orai1 as calcium sensor and calcium channel, respectively, for Ca(2+)-release activated Ca(2+) (CRAC) channels, channels underlying store-operated Ca(2+) entry (SOCE). In addition, these proteins have been suggested to function in signalling and constructing other channels with biophysical properties distinct from the CRAC channels. Using the human kidney cell line, HEK293, we examined the hypothesis that STIM1 can interact with and regulate members of a family of non-selective cation channels (TRPC) which have been suggested to also function in SOCE pathways under certain conditions. Our data reveal no role for either STIM1 or Orai1 in signalling of TRPC channels. Specifically, Ca(2+) entry seen after carbachol treatment in cells transiently expressing TRPC1, TRPC3, TRPC5 or TRPC6 was not enhanced by the co-expression of STIM1. Further, knockdown of STIM1 in cells expressing TRPC5 did not reduce TRPC5 activity, in contrast to one published report. We previously reported in stable TRPC7 cells a Ca(2+) entry which was dependent on TRPC7 and appeared store-operated. However, we show here that this TRPC7-mediated entry was also not dependent on either STIM1 or Orai1, as determined by RNA interference (RNAi) and expression of a constitutively active mutant of STIM1. Further, we determined that this entry was not actually store-operated, but instead TRPC7 activity which appears to be regulated by SERCA. Importantly, endogenous TRPC activity was also not regulated by STIM1. In vascular smooth muscle cells, arginine-vasopressin (AVP) activated non-selective cation currents associated with TRPC6 activity were not affected by RNAi knockdown of STIM1, while SOCE was largely inhibited. Finally, disruption of lipid rafts significantly attenuated TRPC3 activity, while having no effect on STIM1 localization or the development of I(CRAC). Also, STIM1 punctae were found to localize in regions distinct from lipid rafts. This suggests

  4. Distribution of TRPC channels in a visceral sensory pathway.

    PubMed

    Buniel, Maria; Wisnoskey, Brian; Glazebrook, Patricia A; Schilling, William P; Kunze, Diana L

    2004-01-01

    Until recently most of the published studies addressing the mechanisms of activation of TRPC channels have been carried out in heterologous expression systems. Lack of specific antagonists for the TRPC channels has hampered functional studies of endogenous channels. We approached the role of TRPC channels in native tissue with a study of the distribution of the channel proteins in the carotid chemosensory pathway in the rat. In a previous report we showed that TRPC3/4/5/6 and TRPC7 were present in neurons throughout the petrosal ganglion while TRPC1 was expressed in only a subpopulation of petrosal neurons, at least half of which projected to the carotid body. The TRPC proteins were differentially distributed to the branches of the axons that project centrally to the nucleus of the solitary tract and peripherally to the carotid body. The smallest unmyelinated sensory fibres projecting to the carotid body contained TRPC1/3/4/5 or TRPC6 but not TRPC7. TRPC1 and TRPC3 were concentrated in the larger diameter fibres. Interestingly, only TRPC1 and TRPC4 could be demonstrated in the final terminal endings within glomus cell clusters of the carotid body. In the central axon of the sensory neurons, both TRPC4 and TRPC5 were demonstrated in fibres exiting the solitary tract and projecting to the secondary relay neurons the nucleus of the solitary tract. PMID:15104186

  5. The role of transient receptor potential channels in metabolic syndrome.

    PubMed

    Liu, Daoyan; Zhu, Zhiming; Tepel, Martin

    2008-11-01

    Metabolic syndrome is correlated with increased cardiovascular risk and characterized by several factors, including visceral obesity, hypertension, insulin resistance, and dyslipidemia. Several members of a large family of nonselective cation entry channels, e.g., transient receptor potential (TRP) canonical (TRPC), vanilloid (TRPV), and melastatin (TRPM) channels, have been associated with the development of cardiovascular diseases. Thus, disruption of TRP channel expression or function may account for the observed increased cardiovascular risk in metabolic syndrome patients. TRPV1 regulates adipogenesis and inflammation in adipose tissues, whereas TRPC3, TRPC5, TRPC6, TRPV1, and TRPM7 are involved in vasoconstriction and regulation of blood pressure. Other members of the TRP family are involved in regulation of insulin secretion, lipid composition, and atherosclerosis. Although there is no evidence that a single TRP channelopathy may be the cause of all metabolic syndrome characteristics, further studies will help to clarify the role of specific TRP channels involved in the metabolic syndrome. (Hypertens Res 2008; 31: 1989-1995). PMID:19098369

  6. Substrate Specificity within a Family of Outer Membrane Carboxylate Channels

    SciTech Connect

    Eren, Elif; Vijayaraghavan, Jagamya; Liu, Jiaming; Cheneke, Belete R.; Touw, Debra S.; Lepore, Bryan W.; Indic, Mridhu; Movileanu, Liviu; van den Berg, Bert; Dutzler, Raimund

    2012-01-17

    Many Gram-negative bacteria, including human pathogens such as Pseudomonas aeruginosa, do not have large-channel porins. This results in an outer membrane (OM) that is highly impermeable to small polar molecules, making the bacteria intrinsically resistant towards many antibiotics. In such microorganisms, the majority of small molecules are taken up by members of the OprD outer membrane protein family. Here we show that OprD channels require a carboxyl group in the substrate for efficient transport, and based on this we have renamed the family Occ, for outer membrane carboxylate channels. We further show that Occ channels can be divided into two subfamilies, based on their very different substrate specificities. Our results rationalize how certain bacteria can efficiently take up a variety of substrates under nutrient-poor conditions without compromising membrane permeability. In addition, they explain how channel inactivation in response to antibiotics can cause resistance but does not lead to decreased fitness.

  7. Formation of novel TRPC channels by complex subunit interactions in embryonic brain.

    PubMed

    Strübing, Carsten; Krapivinsky, Grigory; Krapivinsky, Luba; Clapham, David E

    2003-10-01

    Mammalian short TRP channels (TRPCs) are putative receptor- and store-operated cation channels that play a fundamental role in the regulation of cellular Ca2+ homeostasis. Assembly of the seven TRPC homologs (TRPC1-7) into homo- and heteromers can create a large variety of different channels. However, the compositions as well as the functional properties of native TRPC complexes are largely undefined. We performed a systematic biochemical study of TRPC interactions in mammalian brain and identified previously unrecognized channel heteromers composed of TRPC1, TRPC4, or TRPC5 and the diacylglycerol-activated TRPC3 or TRPC6 subunits. The novel TRPC heteromers were found exclusively in embryonic brain. In heterologous systems, we demonstrated that assembly of these novel heteromers required the combination of TRPC1 plus TRPC4 or TRPC5 subunits along with diacylglycerol-sensitive subunits in the channel complexes. Functional interaction of the TRPC subunits was verified using a dominant negative TRPC5 mutant (TRPC5DN). Co-expression of TRPC5DN suppressed currents through TRPC5- and TRPC4-containing complexes; TRPC3-associated currents were unaffected by TRPC5DN unless TRPC1 was also co-expressed. This complex assembly mechanism increases the diversity of TRPC channels in mammalian brain and may generate novel heteromers that have specific roles in the developing brain. PMID:12857742

  8. Vascular TRP Channels: Performing Under Pressure and Going with the Flow

    PubMed Central

    Hill-Eubanks, David C.; Gonzales, Albert L.; Sonkusare, Swapnil K.

    2014-01-01

    Endothelial cells and smooth muscle cells of resistance arteries mediate opposing responses to mechanical forces acting on the vasculature, promoting dilation in response to flow and constriction in response to pressure, respectively. In this review, we explore the role of TRP channels, particularly endothelial TRPV4 and smooth muscle TRPC6 and TRPM4 channels, in vascular mechanosensing circuits, placing their putative mechanosensitivity in context with other proposed upstream and downstream signaling pathways. PMID:25180264

  9. Isoform-selective physical coupling of TRPC3 channels to IP3 receptors in smooth muscle cells regulates arterial contractility

    PubMed Central

    Adebiyi, Adebowale; Zhao, Guiling; Narayanan, Damodaran; Thomas, Candice M.; Bannister, John P.; Jaggar, Jonathan H.

    2010-01-01

    Rationale Inositol 1,4,5-trisphosphate (IP3)-induced vasoconstriction can occur independently of intracellular Ca2+ release and via IP3 receptor (IP3R) and canonical transient receptor potential (TRPC) channel activation, but functional signaling mechanisms mediating this effect are unclear. Objectives Study mechanisms by which IP3Rs stimulate TRPC channels in myocytes of resistance-size cerebral arteries. Methods and Results Immunofluorescence resonance energy transfer (immuno-FRET) microscopy using isoform-selective antibodies indicated that endogenous type 1 IP3Rs (IP3R1) are in close spatial proximity to TRPC3, but distant from TRPC6 or TRPM4 channels in arterial myocytes. Endothelin-1 (ET-1), a phospholipase C-coupled receptor agonist, elevated immuno-FRET between IP3R1 and TRPC3, but not between IP3R1 and TRPC6 or TRPM4. TRPC3, but not TRPC6, co-immunoprecipitated with IP3R1. TRPC3 and TRPC6 antibodies selectively inhibited recombinant channels, but only the TRPC3 antibody blocked IP3-induced non-selective cation current (ICat) in myocytes. TRPC3 knockdown attenuated immuno-FRET between IP3R1 and TRPC3, IP3-induced ICat activation, and ET-1 and IP3-induced vasoconstriction, whereas TRPC6 channel knockdown had no effect. ET-1 did not alter total or plasma membrane-localized TRPC3, as determined using surface biotinylation. RT-PCR demonstrated that C-terminal calmodulin and IP3R binding (CIRB) domains are present in myocyte TRPC3 and TRPC6 channels. A peptide corresponding to the IP3R N-terminal region that can interact with TRPC channels activated ICat. A TRPC3 CIRB domain peptide attenuated IP3- and ET-1-induced ICat activation and vasoconstriction. Conclusions IP3 stimulates direct coupling between IP3R1 and membrane-resident TRPC3 channels in arterial myocytes, leading to ICat activation and vasoconstriction. Close spatial proximity between IP3R1 and TRPC3 establishes this isoform-selective functional interaction. PMID:20378853

  10. Specific inhibition of the plasmodial surface anion channel by dantrolene.

    PubMed

    Lisk, Godfrey; Kang, Myungsa; Cohn, Jamieson V; Desai, Sanjay A

    2006-11-01

    The plasmodial surface anion channel (PSAC), induced on human erythrocytes by the malaria parasite Plasmodium falciparum, is an important target for antimalarial drug development because it may contribute to parasite nutrient acquisition. However, known antagonists of this channel are quite nonspecific, inhibiting many other channels and carriers. This lack of specificity not only complicates drug development but also raises doubts about the exact role of PSAC in the well-known parasite-induced permeability changes. We recently identified a family of new PSAC antagonists structurally related to dantrolene, an antagonist of muscle Ca++ release channels. Here, we explored the mechanism of dantrolene's actions on parasite-induced permeability changes. We found that dantrolene inhibits the increased permeabilities of sorbitol, two amino acids, an organic cation, and hypoxanthine, suggesting a common pathway shared by these diverse solutes. It also produced parallel reductions in PSAC single-channel and whole-cell Cl- currents. In contrast to its effect on parasite-induced permeabilities, dantrolene had no measurable effect on five other classes of anion channels, allaying concerns of poor specificity inherent to other known antagonists. Our studies indicate that dantrolene binds PSAC at an extracellular site distinct from the pore, where it inhibits the conformational changes required for channel gating. Its affinity for this site depends on ionic strength, implicating electrostatic interactions in dantrolene binding. In addition to the potential therapeutic applications of its derivatives, dantrolene's specificity and its defined mechanism of action on PSAC make it a useful tool for transport studies of infected erythrocytes. PMID:16950925

  11. Chitoporin from Vibrio harveyi, a channel with exceptional sugar specificity.

    PubMed

    Suginta, Wipa; Chumjan, Watcharin; Mahendran, Kozhinjampara R; Schulte, Albert; Winterhalter, Mathias

    2013-04-19

    Chitoporin (VhChiP) is a sugar-specific channel responsible for the transport of chitooligosaccharides through the outer membrane of the marine bacterium Vibrio harveyi. Single channel reconstitution into black lipid membrane allowed single chitosugar binding events in the channel to be resolved. VhChiP has an exceptionally high substrate affinity, with a binding constant of K = 5.0 × 10(6) M(-1) for its best substrate (chitohexaose). The on-rates of chitosugars depend on applied voltages, as well as the side of the sugar addition, clearly indicating the inherent asymmetry of the VhChiP lumen. The binding affinity of VhChiP for chitohexaose is 1-5 orders of magnitude larger than that of other known sugar-specific porins for their preferred substrates. Thus, VhChiP is the most potent sugar-specific channel reported to date, with its high efficiency presumably reflecting the need for the bacterium to take up chitin-containing nutrients promptly under turbulent aquatic conditions to exploit them efficiently as its sole source of energy. PMID:23447539

  12. Chitoporin from Vibrio harveyi, a Channel with Exceptional Sugar Specificity

    PubMed Central

    Suginta, Wipa; Chumjan, Watcharin; Mahendran, Kozhinjampara R.; Schulte, Albert; Winterhalter, Mathias

    2013-01-01

    Chitoporin (VhChiP) is a sugar-specific channel responsible for the transport of chitooligosaccharides through the outer membrane of the marine bacterium Vibrio harveyi. Single channel reconstitution into black lipid membrane allowed single chitosugar binding events in the channel to be resolved. VhChiP has an exceptionally high substrate affinity, with a binding constant of K = 5.0 × 106 m−1 for its best substrate (chitohexaose). The on-rates of chitosugars depend on applied voltages, as well as the side of the sugar addition, clearly indicating the inherent asymmetry of the VhChiP lumen. The binding affinity of VhChiP for chitohexaose is 1–5 orders of magnitude larger than that of other known sugar-specific porins for their preferred substrates. Thus, VhChiP is the most potent sugar-specific channel reported to date, with its high efficiency presumably reflecting the need for the bacterium to take up chitin-containing nutrients promptly under turbulent aquatic conditions to exploit them efficiently as its sole source of energy. PMID:23447539

  13. Transient Receptor Potential Canonical 7 (TRPC7): A Diacylglycerol-Activated Non-Selective Cation Channel

    PubMed Central

    Zhang, Xuexin

    2016-01-01

    Transient receptor potential canonical 7 (TRPC7) channel is the seventh member of the mammalian TRPC channel family. TRPC7 mRNA, protein and channel activity have been detected in many tissues and organs from mouse, rat and human. TRPC7 has high sequence homology with TRPC3 and TRPC6 and all three channels are activated by membrane receptors that couple to isoforms of phospholipase C (PLC) and mediate non-selective cation currents. TRPC7, along with TRPC3 and TRPC6 can be activated by direct exogenous application of diacylglycerol (DAG) analogs and by pharmacological maneuvers that increase endogenous DAG in cells. TRPC7 shows distinct properties of activation, such as constitutive activity, susceptibility to negative regulation by extracellular Ca2+ and by protein kinase C. TRPC7 can form heteromultimers with TRPC3 and TRPC6. Although TRPC7 remains one of the least studied TRPC channel, its role in various cell types and physiological and pathophysiological conditions is begining to emerge. PMID:24756707

  14. Large-Conductance Calcium-Activated Potassium Channels in Glomerulus: From Cell Signal Integration to Disease.

    PubMed

    Tao, Jie; Lan, Zhen; Wang, Yunman; Hei, Hongya; Tian, Lulu; Pan, Wanma; Zhang, Xuemei; Peng, Wen

    2016-01-01

    Large-conductance calcium-activated potassium (BK) channels are currently considered as vital players in a variety of renal physiological processes. In podocytes, BK channels become active in response to stimuli that increase local cytosolic Ca(2+), possibly secondary to activation of slit diaphragm TRPC6 channels by chemical or mechanical stimuli. Insulin increases filtration barrier permeability through mobilization of BK channels. In mesangial cells, BK channels co-expressed with β1 subunits act as a major component of the counteractive response to contraction in order to regulate glomerular filtration. This review aims to highlight recent discoveries on the localization, physiological and pathological roles of BK channels in glomerulus. PMID:27445840

  15. Large-Conductance Calcium-Activated Potassium Channels in Glomerulus: From Cell Signal Integration to Disease

    PubMed Central

    Tao, Jie; Lan, Zhen; Wang, Yunman; Hei, Hongya; Tian, Lulu; Pan, Wanma; Zhang, Xuemei; Peng, Wen

    2016-01-01

    Large-conductance calcium-activated potassium (BK) channels are currently considered as vital players in a variety of renal physiological processes. In podocytes, BK channels become active in response to stimuli that increase local cytosolic Ca2+, possibly secondary to activation of slit diaphragm TRPC6 channels by chemical or mechanical stimuli. Insulin increases filtration barrier permeability through mobilization of BK channels. In mesangial cells, BK channels co-expressed with β1 subunits act as a major component of the counteractive response to contraction in order to regulate glomerular filtration. This review aims to highlight recent discoveries on the localization, physiological and pathological roles of BK channels in glomerulus.

  16. Non-specific activation of the epithelial sodium channel by the CFTR chloride channel

    PubMed Central

    Nagel, Georg; Szellas, Tanjef; Riordan, John R.; Friedrich, Thomas; Hartung, Klaus

    2001-01-01

    The genetic disease cystic fibrosis is caused by mutation of the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR). Controversial studies reported regulation of the epithelial sodium channel (ENaC) by CFTR. We found that uptake of 22Na+ through ENaC is modulated by activation of CFTR in oocytes, coexpressing CFTR and ENaC, depending on extracellular chloride concentration. Furthermore we found that the effect of CFTR activation could be mimicked by other chloride channels. Voltage– and patch–clamp measurements, however, showed neither stimulation nor inhibition of ENaC-mediated conductance by activated CFTR. We conclude that the observed modulation of 22Na+ uptake by activated CFTR is due to the effect of CFTR-mediated chloride conductance on the membrane potential. These findings argue against the notion of a specific influence of CFTR on ENaC and emphasize the chloride channel function of CFTR. PMID:11266369

  17. Non-specific activation of the epithelial sodium channel by the CFTR chloride channel.

    PubMed

    Nagel, G; Szellas, T; Riordan, J R; Friedrich, T; Hartung, K

    2001-03-01

    The genetic disease cystic fibrosis is caused by mutation of the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR). Controversial studies reported regulation of the epithelial sodium channel (ENaC) by CFTR. We found that uptake of (22)Na(+) through ENaC is modulated by activation of CFTR in oocytes, coexpressing CFTR and ENaC, depending on extracellular chloride concentration. Furthermore we found that the effect of CFTR activation could be mimicked by other chloride channels. Voltage- and patch-clamp measurements, however, showed neither stimulation nor inhibition of ENaC-mediated conductance by activated CFTR. We conclude that the observed modulation of (22)Na(+) uptake by activated CFTR is due to the effect of CFTR-mediated chloride conductance on the membrane potential. These findings argue against the notion of a specific influence of CFTR on ENaC and emphasize the chloride channel function of CFTR. PMID:11266369

  18. Specification for wide channel bandwidth one-inch video tape

    NASA Technical Reports Server (NTRS)

    Perry, Jimmy L.

    1988-01-01

    Standards and controls are established for the procurement of wide channel bandwidth one inch video magnetic recording tapes for Very Long Base Interferometer (VLBI) system applications. The Magnetic Tape Certification Facility (MTCF) currently maintains three specifications for the Quality Products List (QPL) and acceptance testing of magnetic tapes. NASA-TM-79724 is used for the QPL and acceptance testing of new analog tapes; NASA-TM-80599 is used for QPL and acceptance testing of new digital tapes; and NASA-TM-100702 is used for the QPL and acceptance testing of new IBM/IBM compatible 3480 magnetic tape cartridges. This specification will be used for the QPL and acceptance testing of new wide channel bandwidth one inch video magnetic recording tapes. The one inch video tapes used by the Jet Propulsion Lab., the Deep Space Network and the Haystack Observatory will be covered by this specification. These NASA stations will use the video tapes for their VLBI system applications. The VLBI system is used for the tracking of quasars and the support of interplanetary exploration.

  19. Activation of TRPC cationic channels by mercurial compounds confers the cytotoxicity of mercury exposure.

    PubMed

    Xu, Shang-Zhong; Zeng, Bo; Daskoulidou, Nikoleta; Chen, Gui-Lan; Atkin, Stephen L; Lukhele, Bhekithemba

    2012-01-01

    Mercury is an established worldwide environmental pollutant with well-known toxicity affecting neurodevelopment in humans, but the molecular basis of cytotoxicity and the detoxification procedure are still unclear. Here we examined the involvement of the canonical transient receptor potential (TRPC) channel in the mercury-induced cytotoxicity and the potential detoxification strategy. Whole-cell and excised patches, Ca(2+) imaging, and site-directed mutagenesis were used to determine the mechanism of action of mercurial compounds on TRPC channels overexpressed in HEK293 cells, and cytotoxicity and preventive effect were investigated in cell culture models using small interfering RNA and pharmacological blockers. Mercury potently activates TRPC4 and TRPC5 channels. The extracellular cysteine residues (C(553) and C(558)) near the channel pore region of TRPC5 are the molecular targets for channel activation by mercury. The sensitivity of mercury to TRPC5 is presumed to be specific because other divalent heavy metal pollutants, such as Cd(2+), Ni(2+), and Zn(2+), had no stimulating effect, and TRPC3, TRPC6, TRPV1, and TRPM2 were resistant to mercurial compounds. The channel activity of TRPC5, as well as TRPC4, induced by mercury, was prevented by 2-aminoethoxydiphenyl borate and modified by a reducing environment. The inhibition of TRPC5 channels by specific TRPC5 pore-blocking antibody or by SKF-96365 alleviated the cytotoxicity, whereas the mercury chelator, meso-2,3-dimercaptosuccinic acid, showed nonselective prevention of cell survival. Silencing of the TRPC5 gene reduced the mercury-induced neuronal damage. These results indicate that mercurial compounds are activators for TRPC5 and TRPC4 channels. Blockade of TRPC channels could be a novel strategy for preventing mercury-induced cytotoxicity and neurodevelopment impairment. PMID:21984481

  20. Specific features of steam condensation inside tubes and channels

    NASA Astrophysics Data System (ADS)

    Mil'man, O. O.; Fedorov, V. A.; Kondrat'ev, A. V.; Ptakhin, A. V.

    2015-04-01

    The results of theoretical and experimental investigations revealed a dependence between the parameters of steam condensation process in tubes and channels on the coolant flow pattern: counter flow, forward flow, and cross flow. The loss of total steam pressure in case of counter flow is higher than it is in case of using a forward or cross flow pattern. During condensation, superheated steam retains its superheated state over the entire tube length if the steam quality at the outlet x 1 > 0, and the superheating temperature depends on the coolant flow pattern. A method for calculating the pressure loss to steam temperature ratio as a function of coolant flow pattern is developed, which is confirmed by experimental data. The ratio or its versions (where c and G are the specific heat capacity and flow rate of coolant) should be regarded as the main parameter in generalizing data.

  1. Transient receptor potential (TRP) channels, vascular tone and autoregulation of cerebral blood flow.

    PubMed

    Brayden, Joseph E; Earley, Scott; Nelson, Mark T; Reading, Stacey

    2008-09-01

    Members of the transient receptor potential (TRP) channel superfamily are present in vascular smooth muscle cells and play important roles in the regulation of vascular contractility. The TRPC3 and TRPC6 channels are activated by stimulation of several excitatory receptors in vascular smooth muscle cells. Activation of these channels leads to myocyte depolarization, which stimulates Ca2+ entry via voltage-dependent Ca2+ channels (VDCC), leading to vasoconstriction. The TRPV4 channels in arterial myocytes are activated by epoxyeicosatrienoic acids, and activation of the channels enhances Ca2+ spark and transient Ca2+-sensitive K+ channel activity, thereby hyperpolarizing and relaxing vascular smooth muscle cells. The TRPC6 and TRPM4 channels are activated by mechanical stimulation of cerebral artery myocytes. Subsequent depolarization and activation of VDCC Ca2+ entry is directly linked to the development of myogenic tone in vitro and to autoregulation of cerebral blood flow in vivo. These findings imply a fundamental importance of TRP channels in the regulation of vascular smooth muscle tone and suggest that TRP channels could be important targets for drug therapy under conditions in which vascular contractility is disturbed (e.g. hypertension, stroke, vasospasm). PMID:18215190

  2. TRPC channels in smooth muscle cells.

    PubMed

    Gonzalez-Cobos, Jose C; Trebak, Mohamed

    2010-01-01

    Transient receptor potential canonical (TRPC) proteins constitute a family of seven (TRPC1-7) nonselective cation channels within the wider TRP superfamily. TRPC1, TRPC3, TRPC4, TRPC5 and TRPC6 channels are expressed in vascular smooth muscle cells from human vessels of all calibers and in smooth muscle from organs such as the uterus and the gastrointestinal tract. TRPC channels have recently emerged as important players in the control of smooth muscle function. This review will focus on the retrospective analysis of studies proposing contributions of TRPC channels to native calcium entry pathways in smooth muscle and to physiological and pathophysiological responses with emphasis on the vascular system. PMID:20515740

  3. The role of classical transient receptor potential channels in the regulation of hypoxic pulmonary vasoconstriction.

    PubMed

    Fuchs, B; Dietrich, A; Gudermann, T; Kalwa, H; Grimminger, F; Weissmann, N

    2010-01-01

    Hypoxic pulmonary vasoconstriction (HPV) is an essential mechanism of the lung matching blood perfusion to ventilation during local alveolar hypoxia. HPV thus optimizes pulmonary gas exchange. In contrast chronic and generalized hypoxia leads to pulmonary vascular remodeling with subsequent pulmonary hypertension and right heart hypertrophy. Among other non-selective cation channels, the family of classical transient receptor potential channels (TRPC) has been shown to be expressed in pulmonary arterial smooth muscle cells. Among this family, TRPC6 is essential for the regulation of acute HPV in mice. Against this background, in this chapter we give an overview about the TRPC family and their role in HPV. PMID:20204731

  4. Role of Src in C3 transient receptor potential channel function and evidence for a heterogeneous makeup of receptor- and store-operated Ca2+ entry channels.

    PubMed

    Kawasaki, Brian T; Liao, Yanhong; Birnbaumer, Lutz

    2006-01-10

    Receptor-operated Ca2+ entry (ROCE) and store-operated Ca2+ entry (SOCE) are known to be inhibited by tyrosine kinase inhibitors and activation of C-type transient receptor potential channel (TRPC) isoform 3 (TRPC3), a cation channel thought to be involved in SOCE and/or ROCE, was recently shown to depend on src tyrosine kinase activity. What is not known is the step at which src acts on TRPC3 and whether the role for tyrosine kinases in ROCE or SOCE is a general phenomenon. Using in vitro and in cell protein-protein interaction assays we now report that src phosphorylates TRPC3 at Y226 and that formation of phospho-Y226 is essential for TRPC3 activation. This requirement is unique for TRPC3 because (i) mutation of the cognate tyrosines of the closely related TRPC6 and TRPC7 had no effect; (ii) TRPC6 and TRPC7 were activated in src-, yes-, and fyn-deficient cells; and (iii) src, but not yes or fyn, rescued TRPC3 activation in src-, yes-, and fyn-deficient cells. The Src homology 2 domain of src was found to interact with either the N or the C termini of all TRPCs, suggesting that other tyrosine kinases may play a role in ion fluxes mediated by TRPCs other than TRPC3. A side-by-side comparison of the effects of genistein (a general tyrosine kinase inhibitor) on endogenous ROCE and SOCE in mouse fibroblasts, HEK and COS-7 cells, and ROCE in HEK cells mediated by TRPC3, TRPC6, TRPC7, and TRPC5 showed differences that argue for ROCE and SOCE channels to be heterogeneous. PMID:16407161

  5. Donor's position-specific channel interference in substituted biphenyl molecules.

    PubMed

    Mehboob Alam, Md

    2015-07-21

    The channel interference mechanism describes the two-photon absorption process in terms of the relative orientations of different transition dipole moment vectors (TDMVs) in a molecule. Controlling these interferences is a difficult but important task, because it can provide new information about such absorption/emission processes. The present contribution aims to explain the reversal in the nature of these interferences with the relative positions of the donor-acceptor groups in substituted biphenyl molecules. The results obtained from the linear and quadratic response theories indicate that having the donor at the para position leads to a constructive interference whereas having it at any of the other two positions results in a destructive interference. This is explained by a systematic analysis of the relative orientations of all the TDMVs involved in a three-state model. PMID:26088458

  6. Calcium, TRPC channels, and regulation of the actin cytoskeleton in podocytes: towards a future of targeted therapies.

    PubMed

    Wieder, Nicolas; Greka, Anna

    2016-07-01

    With more than 6,000 new pediatric patients with treatment-resistant nephrotic syndrome in the US each year alone, the unmet need for novel, podocyte-specific therapies is substantial. Recently, the established therapeutic benefit of angiotensin-converting enzyme (ACE) inhibitors and angiotensin receptor blockers (ARB) was used as a starting point to gain insight into the pathomechanism of primary podocytopathies. A calcium (Ca(2+))-mediated pathway has been identified that connects the angiotensin type 1 receptor (AT1R) to podocyte cytoskeletal dynamics, essential for a functioning glomerular filtration barrier. This discovery provided an important missing piece in our understanding of the pathomechanism of filter barrier damage, revealing Ca(2+) signaling as critical for podocyte health and disease. The identification of the two Ca(2+) permeant channels TRPC5 and TRPC6 as mediators of this pathway not only bolstered the importance of podocyte cytoskeleton dynamics but also revealed promising drug targets for treatment-resistant nephrotic syndrome. This review will focus on this novel signaling pathway in primary podocytopathies and its implications for next-generation therapies for glomerular disease. PMID:26490951

  7. ANTIARRHYTHMIC DRUG-INDUCED INTERNALIZATION OF THE ATRIAL SPECIFIC K+ CHANNEL, Kv1.5

    PubMed Central

    Schumacher, Sarah M.; McEwen, Dyke P.; Zhang, Lian; Arendt, Kristin L.; Van Genderen, Kristin M.; Martens, Jeffrey R.

    2009-01-01

    Conventional antiarrhythmic drugs target the ion permeability of channels, but increasing evidence suggests that functional ion channel density can also be modified pharmacologically. Kv1.5 mediates the ultrarapid potassium current (IKur) that controls atrial action potential duration. Given the atrial specific expression of Kv1.5 and its alterations in human atrial fibrillation, significant effort has been made to identify novel channel blockers. In this study, treatment of HL-1 atrial myocytes expressing Kv1.5-GFP with the class I antiarrhythmic agent quinidine, resulted in a dose-, and temperature-dependent internalization of Kv1.5, concomitant with channel block. This quinidine-induced channel internalization was confirmed in acutely dissociated neonatal myocytes. Channel internalization was subunit-dependent, activity-independent, stereospecific, and blocked by pharmacologic disruption of the endocytic machinery. Pore block and channel internalization partially overlap in the structural requirements for drug binding. Surprisingly, quinidine-induced endocytosis was calcium-dependent and therefore unrecognized by previous biophysical studies focused on isolating channel-drug interactions. Importantly, while acute quinidine-induced internalization was reversible, chronic treatment led to channel degradation. Together, these data reveal a novel mechanism of antiarrhythmic drug action and highlight the possibility for new agents that selectively modulate the stability of channel protein in the membrane as an approach for treating cardiac arrhythmias. PMID:19443837

  8. High expression of calcium channel subtypes in uterine fibroid of patients

    PubMed Central

    Ke, Xiaoping; Cheng, Zhongping; Qu, Xiaoyan; Dai, Hong; Zhang, Wenchao; Chen, Zi-Jiang

    2014-01-01

    Aim: To investigate the expression of calcium channel protein in uterine fibroids, and to explore the relationship between calcium signaling pathway and the pathogenesis of uterine fibroids. Methods: Uterine fibroid tissues (UFC) and adjacent healthy uterine smooth muscle tissues (SMC) were collected from 30 cases of uterine fibroids. Real-time quantitative PCR and western blot were used to detect cell membrane calcium channel protein subtypes: TRPC1, TRPC3, TRPC4, TRPC6, TRPM6 and TRPM7. The effects of genes exhibiting most-notable differences on cell proliferation were examined using gene interference techniques. Results: We found that calcium channel protein subtypes expressed differently in fibroids and the surrounding smooth muscles. The mRNA and protein expressions of TRPC1 and TRPM7 were higher in uterine fibroid tissues than in smooth muscle (P < 0.05), while no obvious difference was found in terms of other subtypes (TRPC3, TRPC4, TRPC6 and TRPM6). In cultured uterine leiomyoma cells, modifying the expressions of TRPC1 and TRPM7 significantly affected the proliferation rate of uterine fibroids. Conclusion: Calcium channel subtypes TRPC1 and TRPM7 exhibit different expression patterns in uterine fibroids and surrounding smooth muscles, suggesting that calcium signaling pathway regulated by these calcium channel proteins may be associated with the incidence of uterine fibroids. PMID:24995090

  9. The non-specific ion channel in Torpedo ocellata fused synaptic vesicles.

    PubMed Central

    Yakir, N; Rahamimoff, R

    1995-01-01

    1. Synaptic vesicles were isolated and fused into large structures with a diameter of more than 20 microns to characterize their ionic channels. The 'cell'-attached and inside-out configurations of the patch clamp technique were used. 2. Two types of ion channels were most frequently observed: a low conductance chloride channel and a high conductance non-specific channel. 3. The non-specific channel has a main conducting state and a substate. The main conducting state has a slope conductance of 246 +/- 15 pS (+/- S.E.M., n = 15), in the presence of different combinations of KCl and potassium glutamate. 4. From the reversal potentials of the current-voltage (I-V) relation, it was concluded that this channel conducts both Cl- and K+. 5. The non-specific channel is highly voltage dependent: under steady-state voltages it has a high open probability near 0 mV and does not inactivate; when the membrane is hyperpolarized (pipette side more positive), the open probability decreases dramatically. 6. Voltage pulses showed that upon hyperpolarization (from holding potentials between -20 and + 20 mV), the channels deactivated; when the membrane was stepped back to the holding potential, the channels reactivated rapidly. 7. In a number of experiments, when the pipette side was made more negative than the bath, the open probability also decreased. 8. Frequently, a substate with a conductance of about 44 +/- 4% (+/- S.E.M., n = 3) of the main state was detected. 9. We speculate that this non-specific ion channel may have different roles at the various stages of the life cycle of the synaptic vesicle. When the synaptic vesicle is an intracellular structure, it might help its transmitter-concentrating capacity by dissipating the polarization. After fusion with the surface membrane, it might constitute an additional conductance pathway, taking part in frequency modulation of synaptic transmission. PMID:7562610

  10. Immunohistochemical study on the distribution of TRPC channels in the rat hippocampus.

    PubMed

    Chung, Yoon Hee; Sun Ahn, Hyang; Kim, Daejin; Hoon Shin, Dong; Su Kim, Sung; Yong Kim, Kyung; Bok Lee, Won; Ik Cha, Choong

    2006-04-26

    In the present study, we performed immunohistochemistry using antibodies directed against TRPCs to study the localizations of these channels in rat hippocampus. The pyramidal cell bodies of CA1-3 areas and the granule cell bodies of the dentate gyrus were immunoreactive for TRPC1, TRPC3, TRPC4 and TRPC5. On the other hand, TRPC6 exhibited the cloud-like neuropil staining only in the molecular layer of the dentate gyrus. As a whole, the present study has clearly shown the localization of TRPCs in rat hippocampus and may provide useful data for the future investigations on the structural and functional properties of TRPCs. PMID:16580647

  11. Molecular determinants on the insect sodium channel for the specific action of type II pyrethroid insecticides

    SciTech Connect

    Du Yuzhe; Nomura, Yoshiko; Luo Ningguang; Liu Zhiqi; Lee, Jung-Eun; Khambay, Bhupinder; Dong Ke

    2009-01-15

    Pyrethroid insecticides are classified as type I or type II based on their distinct symptomology and effects on sodium channel gating. Structurally, type II pyrethroids possess an {alpha}-cyano group at the phenylbenzyl alcohol position, which is lacking in type I pyrethroids. Both type I and type II pyrethroids inhibit deactivation consequently prolonging the opening of sodium channels. However, type II pyrethroids inhibit the deactivation of sodium channels to a greater extent than type I pyrethroids inducing much slower decaying of tail currents upon repolarization. The molecular basis of a type II-specific action, however, is not known. Here we report the identification of a residue G{sup 1111} and two positively charged lysines immediately downstream of G{sup 1111} in the intracellular linker connecting domains II and III of the cockroach sodium channel that are specifically involved in the action of type II pyrethroids, but not in the action of type I pyrethroids. Deletion of G{sup 1111}, a consequence of alternative splicing, reduced the sodium channel sensitivity to type II pyrethroids, but had no effect on channel sensitivity to type I pyrethroids. Interestingly, charge neutralization or charge reversal of two positively charged lysines (Ks) downstream of G{sup 1111} had a similar effect. These results provide the molecular insight into the type II-specific interaction of pyrethroids with the sodium channel at the molecular level.

  12. Molecular determinants on the insect sodium channel for the specific action of type II pyrethroid insecticides.

    PubMed

    Du, Yuzhe; Nomura, Yoshiko; Luo, Ningguang; Liu, Zhiqi; Lee, Jung-Eun; Khambay, Bhupinder; Dong, Ke

    2009-01-15

    Pyrethroid insecticides are classified as type I or type II based on their distinct symptomology and effects on sodium channel gating. Structurally, type II pyrethroids possess an alpha-cyano group at the phenylbenzyl alcohol position, which is lacking in type I pyrethroids. Both type I and type II pyrethroids inhibit deactivation consequently prolonging the opening of sodium channels. However, type II pyrethroids inhibit the deactivation of sodium channels to a greater extent than type I pyrethroids inducing much slower decaying of tail currents upon repolarization. The molecular basis of a type II-specific action, however, is not known. Here we report the identification of a residue G(1111) and two positively charged lysines immediately downstream of G(1111) in the intracellular linker connecting domains II and III of the cockroach sodium channel that are specifically involved in the action of type II pyrethroids, but not in the action of type I pyrethroids. Deletion of G(1111), a consequence of alternative splicing, reduced the sodium channel sensitivity to type II pyrethroids, but had no effect on channel sensitivity to type I pyrethroids. Interestingly, charge neutralization or charge reversal of two positively charged lysines (Ks) downstream of G(1111) had a similar effect. These results provide the molecular insight into the type II-specific interaction of pyrethroids with the sodium channel at the molecular level. PMID:19022275

  13. The Na+/Ca2+ exchange inhibitor KB-R7943 potently blocks TRPC channels.

    PubMed

    Kraft, Robert

    2007-09-14

    Na(+)/Ca(2+) exchangers (NCXs) and members of the canonical transient receptor potential (TRPC) channels play an important role in Ca(2+) homeostasis in heart and brain. With respect to their overlapping expression and their role as physiological Ca(2+) influx pathways a functional discrimination of both mechanisms seems to be necessary. Here, the effect of the reverse-mode NCX inhibitor KB-R7943 was investigated on different TRPC channels heterologously expressed in HEK293 cells. In patch-clamp recordings KB-R7943 potently blocked currents through TRPC3 (IC(50)=0.46 microM), TRPC6 (IC(50)=0.71 microM), and TRPC5 (IC(50)=1.38 microM). 1-Oleoyl-2-acetyl-sn-glycerol-induced Ca(2+) entry was nearly completely suppressed by 10 microM KB-R7943 in TRPC6-transfected cells. Thus, KB-R7943 is able to block receptor-operated TRP channels at concentrations which are equal or below those required to inhibit reverse-mode NCX activity. These data further suggest that the protective effects of KB-R7943 in ischemic tissue may, at least partly, be due to inhibition of TRPC channels. PMID:17658472

  14. Transient receptor potential cation channels in normal and dystrophic mdx muscle.

    PubMed

    Krüger, Jana; Kunert-Keil, Christiane; Bisping, Frederike; Brinkmeier, Heinrich

    2008-06-01

    To investigate the defective calcium regulation of dystrophin-deficient muscle fibres we studied gene expression and localization of non-voltage gated cation channels in normal and mdx mouse skeletal muscle. We found TRPC3, TRPC6, TRPV4, TRPM4 and TRPM7 to be the most abundant isoforms. Immunofluorescent staining of muscle cross-sections with antibodies against TRP proteins showed sarcolemmal localization of TRPC6 and TRPM7, both, for mdx and control. TRPV4 was found only in a fraction of fibres at the sarcolemma and around myonuclei, while TRPC3 staining revealed intracellular patches, preferentially in mdx muscle. Transcripts of low abundance coding for TRPC5, TRPA1 and TRPM1 channels were increased in mdx skeletal muscle at certain stages. The increased Ca(2+)-influx into dystrophin-deficient mdx fibres cannot be explained by increased gene expression of major TRP channels. However, a constant TRP channel expression in combination with the well described weaker Ca(2+)-handling system of mdx fibres may indicate an imbalance between Ca(2+)-influx and cellular Ca(2+)-control. PMID:18504127

  15. Interactions of cryptosin with mammalian cardiac dihydropyridine-specific calcium channels

    SciTech Connect

    Rao, V.R.; Banning, J.W. )

    1990-01-01

    Cryptosin, a new cardenolide, was found to be a potent inhibitor of cardiac Na{sup +} and K{sup +} dependent Adenosinetri-phosphatase. In experiments with dog heart ex vivo, development of inotropic and toxic effect correlated with changes in the cardiac dihydropyridine-specific calcium channels as measured by the binding of {sup 3}(H)PN 200-110. A significant change in the PN 200-110 binding was observed when guinea pig and dog heart sarcolemmal membranes were pre-incubated with cryptosin in vitro. Binding analysis of {sup 3}(H)PN 200-110 (Isradipine), a 1,4-dihydropyridine analog with very specific calcium channel binding properties, in both in vitro and ex vivo studies were consistent and indicated a non-specific type of interaction of cryptosin with mammalian cardiac 1,4-dihydropyridine-specific calcium channels.

  16. A novel slow-inactivation-specific ion channel modulator attenuates neuropathic pain.

    PubMed

    Hildebrand, Michael E; Smith, Paula L; Bladen, Chris; Eduljee, Cyrus; Xie, Jennifer Y; Chen, Lina; Fee-Maki, Molly; Doering, Clint J; Mezeyova, Janette; Zhu, Yongbao; Belardetti, Francesco; Pajouhesh, Hassan; Parker, David; Arneric, Stephen P; Parmar, Manjeet; Porreca, Frank; Tringham, Elizabeth; Zamponi, Gerald W; Snutch, Terrance P

    2011-04-01

    Voltage-gated ion channels are implicated in pain sensation and transmission signaling mechanisms within both peripheral nociceptors and the spinal cord. Genetic knockdown and knockout experiments have shown that specific channel isoforms, including Na(V)1.7 and Na(V)1.8 sodium channels and Ca(V)3.2 T-type calcium channels, play distinct pronociceptive roles. We have rationally designed and synthesized a novel small organic compound (Z123212) that modulates both recombinant and native sodium and calcium channel currents by selectively stabilizing channels in their slow-inactivated state. Slow inactivation of voltage-gated channels can function as a brake during periods of neuronal hyperexcitability, and Z123212 was found to reduce the excitability of both peripheral nociceptors and lamina I/II spinal cord neurons in a state-dependent manner. In vivo experiments demonstrate that oral administration of Z123212 is efficacious in reversing thermal hyperalgesia and tactile allodynia in the rat spinal nerve ligation model of neuropathic pain and also produces acute antinociception in the hot-plate test. At therapeutically relevant concentrations, Z123212 did not cause significant motor or cardiovascular adverse effects. Taken together, the state-dependent inhibition of sodium and calcium channels in both the peripheral and central pain signaling pathways may provide a synergistic mechanism toward the development of a novel class of pain therapeutics. PMID:21349638

  17. Identification of Specific Sensory Neuron Populations for Study of Expressed Ion Channels

    PubMed Central

    Ramachandra, Renuka; McGrew, Stephanie; Elmslie, Keith

    2013-01-01

    Sensory neurons transmit signals from various parts of the body to the central nervous system. The soma for these neurons are located in the dorsal root ganglia that line the spinal column. Understanding the receptors and channels expressed by these sensory afferent neurons could lead to novel therapies for disease. The initial step is to identify the specific subset of sensory neurons of interest. Here we describe a method to identify afferent neurons innervating the muscles by retrograde labeling using a fluorescent dye DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate). Understanding the contribution of ion channels to excitation of muscle afferents could help to better control excessive excitability induced by certain disease states such as peripheral vascular disease or heart failure. We used two approaches to identify the voltage dependent ion channels expressed by these neurons, patch clamp electrophysiology and immunocytochemistry. While electrophysiology plus pharmacological blockers can identify functional ion channel types, we used immunocytochemistry to identify channels for which specific blockers were unavailable and to better understand the ion channel distribution pattern in the cell population. These techniques can be applied to other areas of the nervous system to study specific neuronal groups. PMID:24430510

  18. Designing a C84 fullerene as a specific voltage-gated sodium channel blocker

    NASA Astrophysics Data System (ADS)

    Hilder, Tamsyn A.; Chung, Shin-Ho

    2013-07-01

    Fullerene derivatives demonstrate considerable potential for numerous biological applications, such as the effective inhibition of HIV protease. Recently, they were identified for their ability to indiscriminately block biological ion channels. A fullerene derivative which specifically blocks a particular ion channel could lead to a new set of drug leads for the treatment of various ion channel-related diseases. Here, we demonstrate their extraordinary potential by designing a fullerene which mimics some of the functions of μ-conotoxin, a peptide derived from cone snail venom which potently binds to the bacterial voltage-gated sodium channel (NavAb). We show, using molecular dynamics simulations, that the C84 fullerene with six lysine derivatives uniformly attached to its surface is selective to NavAb over a voltage-gated potassium channel (Kv1.3). The side chain of one of the lysine residues protrudes into the selectivity filter of the channel, while the methionine residues located just outside of the channel form hydrophobic contacts with the carbon atoms of the fullerene. The modified C84 fullerene strongly binds to the NavAb channel with an affinity of 46 nM but binds weakly to Kv1.3 with an affinity of 3 mM. This potent blocker of NavAb may serve as a structural template from which potent compounds can be designed for the targeting of mammalian Nav channels. There is a genuine need to target mammalian Nav channels as a form of treatment of various diseases which have been linked to their malfunction, such as epilepsy and chronic pain.

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

    PubMed Central

    Bergeron, Zachary L.; Bingham, Jon-Paul

    2012-01-01

    Scorpion toxins have been central to the investigation and understanding of the physiological role of potassium (K+) channels and their expansive function in membrane biophysics. As highly specific probes, toxins have revealed a great deal about channel structure and the correlation between mutations, altered regulation and a number of human pathologies. Radio- and fluorescently-labeled toxin isoforms have contributed to localization studies of channel subtypes in expressing cells, and have been further used in competitive displacement assays for the identification of additional novel ligands for use in research and medicine. Chimeric toxins have been designed from multiple peptide scaffolds to probe channel isoform specificity, while advanced epitope chimerization has aided in the development of novel molecular therapeutics. Peptide backbone cyclization has been utilized to enhance therapeutic efficiency by augmenting serum stability and toxin half-life in vivo as a number of K+-channel isoforms have been identified with essential roles in disease states ranging from HIV, T-cell mediated autoimmune disease and hypertension to various cardiac arrhythmias and Malaria. Bioengineered scorpion toxins have been monumental to the evolution of channel science, and are now serving as templates for the development of invaluable experimental molecular therapeutics. PMID:23202307

  20. TRPC4 forms store-operated Ca2+ channels in mouse mesangial cells.

    PubMed

    Wang, Xiaoxia; Pluznick, Jennifer L; Wei, Peilin; Padanilam, Babu J; Sansom, Steven C

    2004-08-01

    Studies were performed to identify the molecular component responsible for store-operated Ca(2+) entry in murine mesangial cells (MMC). Because the canonical transient receptor potential (TRPC) family of proteins was previously shown to comprise Ca(2+)-selective and -nonselective cation channels in a variety of cells, we screened TRPC1-TRPC7 with the use of molecular methods and the fura 2 method to determine their participation as components of the mesangial store-operated Ca(2+) (SOC) channel. Using TRPC-specific primers and RT-PCR, we found that cultured MMC contained mRNA for TRPC1 and TRPC4 but not for TRPC2, TRPC3, TRPC5, TRPC6, and TRPC7. Immunocytochemical staining of MMC revealed predominantly cytoplasmic expression of TRPC1 and plasmalemmal expression of TRPC4. The role of TRPC4 in SOC was determined with TRPC4 antisense and fura 2 ratiometric measurements of intracellular Ca(2+) concentration ([Ca(2+)](i)). SOC was measured as the increase in [Ca(2+)](i) after extracellular Ca(2+) was increased from <10 nM to 1 mM in the continued presence of thapsigargin. We found that TRPC4 antisense, which reduced plasmalemmal expression of TRPC4, inhibited SOC by 83%. Incubation with scrambled TRPC4 oligonucleotides did not affect SOC. Immunohistochemical staining identified expressed TRPC4 in the glomeruli of mouse renal sections. The results of RT-PCR performed to distinguish between TRPC4-alpha and TRPC4-beta were consistent with expression of both isoforms in brain but with only TRPC4-alpha expression in MMC. These studies show that TRPC4-alpha may form the homotetrameric SOC in mouse mesangial cells. PMID:15044151

  1. Evidence of a Role for Fibroblast Transient Receptor Potential Canonical 3 Ca2+ Channel in Renal Fibrosis.

    PubMed

    Saliba, Youakim; Karam, Ralph; Smayra, Viviane; Aftimos, Georges; Abramowitz, Joel; Birnbaumer, Lutz; Farès, Nassim

    2015-08-01

    Transient receptor potential canonical (TRPC) Ca(2+)-permeant channels, especially TRPC3, are increasingly implicated in cardiorenal diseases. We studied the possible role of fibroblast TRPC3 in the development of renal fibrosis. In vitro, a macromolecular complex formed by TRPC1/TRPC3/TRPC6 existed in isolated cultured rat renal fibroblasts. However, specific blockade of TRPC3 with the pharmacologic inhibitor pyr3 was sufficient to inhibit both angiotensin II- and 1-oleoyl-2-acetyl-sn-glycerol-induced Ca(2+) entry in these cells, which was detected by fura-2 Ca(2+) imaging. TRPC3 blockade or Ca(2+) removal inhibited fibroblast proliferation and myofibroblast differentiation by suppressing the phosphorylation of extracellular signal-regulated kinase (ERK1/2). In addition, pyr3 inhibited fibrosis and inflammation-associated markers in a noncytotoxic manner. Furthermore, TRPC3 knockdown by siRNA confirmed these pharmacologic findings. In adult male Wistar rats or wild-type mice subjected to unilateral ureteral obstruction, TRPC3 expression increased in the fibroblasts of obstructed kidneys and was associated with increased Ca(2+) entry, ERK1/2 phosphorylation, and fibroblast proliferation. Both TRPC3 blockade in rats and TRPC3 knockout in mice inhibited ERK1/2 phosphorylation and fibroblast activation as well as myofibroblast differentiation and extracellular matrix remodeling in obstructed kidneys, thus ameliorating tubulointerstitial damage and renal fibrosis. In conclusion, TRPC3 channels are present in renal fibroblasts and control fibroblast proliferation, differentiation, and activation through Ca(2+)-mediated ERK signaling. TRPC3 channels might constitute important therapeutic targets for improving renal remodeling in kidney disease. PMID:25479966

  2. Specificity of sites within eight-membered ring zeolite channels for carbonylation of methyls to acetyls.

    PubMed

    Bhan, Aditya; Allian, Ayman D; Sunley, Glenn J; Law, David J; Iglesia, Enrique

    2007-04-25

    The acid-catalyzed formation of carbon-carbon bonds from C1 precursors via CO insertion into chemisorbed methyl groups occurs selectively within eight-membered ring (8-MR) zeolite channels. This elementary step controls catalytic carbonylation rates of dimethyl ether (DME) to methyl acetate. The number of O-H groups within 8-MR channels was measured by rigorous deconvolution of the infrared bands for O-H groups in cation-exchanged and acid forms of mordenite (M,H-MOR) and ferrierite (H-FER) after adsorption of basic probe molecules of varying size. DME carbonylation rates are proportional to the number of O-H groups within 8-MR channels. Na+ cations selectively replaced protons within 8-MR channels and led to a disproportionate decrease in carbonylation turnover rates (per total H+). These conclusions are consistent with the low or undetectable rates of carbonylation on zeolites without 8-MR channels (H-BEA, H-FAU, H-MFI). Such specificity of methyl reactivity upon confinement within small channels appears to be unprecedented in catalysis by microporous solids, which typically select reactions by size exclusion of bulkier transition states. PMID:17397162

  3. Sodium-dependent inhibition of the epithelial sodium channel by an arginyl-specific reagent

    SciTech Connect

    Garty, H.; Yeger, O.; Asher, C.

    1988-04-25

    Effects of the arginyl- and lysyl-specific reagent phenylglyoxal (PGO) on the epithelial Na+ channel were evaluated by measuring the amiloride-blockable /sup 22/Na+ fluxes in membrane vesicles derived from the toad bladder epithelium. Incubating whole cells or isolated membranes with PGO readily and irreversibly blocked the channel-mediated tracer flux. Na+ ions present during the interaction of membranes with PGO could protect channels from inactivation by PGO. This effect required the presence of Na+ at the luminal side of the membrane and was characterized by an IC50 of 79 mM Na+. Amiloride, too, could desensitize channels to PGO, but its effect was significant only when whole cells were interacted with the protein-modifying reagent. The data are compatible with a model in which the conductive path of the channel contains a functional arginine, possibly forming a salt bridge with a carboxylic group, which is involved in Na+ translocation and amiloride binding. It was also shown that the augmentation of transport induced by incubating whole cells in Ca2+-free solution involves the activation or recruitment of channels that are not vulnerable to PGO prior to incubation.

  4. The Validation of Nematode-Specific Acetylcholine-Gated Chloride Channels as Potential Anthelmintic Drug Targets

    PubMed Central

    Wever, Claudia M.; Farrington, Danielle; Dent, Joseph A.

    2015-01-01

    New compounds are needed to treat parasitic nematode infections in humans, livestock and plants. Small molecule anthelmintics are the primary means of nematode parasite control in animals; however, widespread resistance to the currently available drug classes means control will be impossible without the introduction of new compounds. Adverse environmental effects associated with nematocides used to control plant parasitic species are also motivating the search for safer, more effective compounds. Discovery of new anthelmintic drugs in particular has been a serious challenge due to the difficulty of obtaining and culturing target parasites for high-throughput screens and the lack of functional genomic techniques to validate potential drug targets in these pathogens. We present here a novel strategy for target validation that employs the free-living nematode Caenorhabditis elegans to demonstrate the value of new ligand-gated ion channels as targets for anthelmintic discovery. Many successful anthelmintics, including ivermectin, levamisole and monepantel, are agonists of pentameric ligand-gated ion channels, suggesting that the unexploited pentameric ion channels encoded in parasite genomes may be suitable drug targets. We validated five members of the nematode-specific family of acetylcholine-gated chloride channels as targets of agonists with anthelmintic properties by ectopically expressing an ivermectin-gated chloride channel, AVR-15, in tissues that endogenously express the acetylcholine-gated chloride channels and using the effects of ivermectin to predict the effects of an acetylcholine-gated chloride channel agonist. In principle, our strategy can be applied to validate any ion channel as a putative anti-parasitic drug target. PMID:26393923

  5. TRPC channels as STIM1-regulated store-operated channels.

    PubMed

    Worley, Paul F; Zeng, Weizhong; Huang, Guo N; Yuan, Joseph P; Kim, Joo Young; Lee, Min Goo; Muallem, Shmuel

    2007-08-01

    Receptor-activated Ca(2+) influx is mediated largely by store-operated channels (SOCs). TRPC channels mediate a significant portion of the receptor-activated Ca(2+) influx. However, whether any of the TRPC channels function as a SOC remains controversial. Our understanding of the regulation of TRPC channels and their function as SOCs is being reshaped with the discovery of the role of STIM1 in the regulation of Ca(2+) influx channels. The findings that STIM1 is an ER resident Ca(2+) binding protein that regulates SOCs allow an expanded and molecular definition of SOCs. SOCs can be considered as channels that are regulated by STIM1 and require the clustering of STIM1 in response to depletion of the ER Ca(2+) stores and its translocation towards the plasma membrane. TRPC1 and other TRPC channels fulfill these criteria. STIM1 binds to TRPC1, TRPC2, TRPC4 and TRPC5 but not to TRPC3, TRPC6 and TRPC7, and STIM1 regulates TRPC1 channel activity. Structure-function analysis reveals that the C-terminus of STIM1 contains the binding and gating function of STIM1. The ERM domain of STIM1 binds to TRPC channels and a lysine-rich region participates in the gating of SOCs and TRPC1. Knock-down of STIM1 by siRNA and prevention of its translocation to the plasma membrane inhibit the activity of native SOCs and TRPC1. These findings support the conclusion that TRPC1 is a SOC. Similar studies with other TRPC channels demonstrate their regulation by STIM1 and indicate that all TRPC channels, except TRPC7, function as SOCs. PMID:17517433

  6. Activation of Plant Plasma Membrane Ca2+-Permeable Channels by Race-Specific Fungal Elicitors.

    PubMed Central

    Gelli, A.; Higgins, V. J.; Blumwald, E.

    1997-01-01

    The response of plant cells to invading pathogens is regulated by fluctuations in cytosolic Ca2+ levels that are mediated by Ca2+-permeable channels located at the plasma membrane of the host cell. The mechanisms by which fungal elicitors can induce Ca2+ uptake by the host cell were examined by the application of conventional patch-clamp techniques. Whole-cell and single-channel experiments on tomato (Lycopersicon esculentum L.) protoplasts revealed a race-specific fungal elicitor-induced activation of a plasma membrane Ca2+-permeable channel. The presence of the fungal elicitor resulted in a greater probability of channel opening. Guanosine 5[prime]-[[beta]-thio]diphosphate, a GDP analog that locks heterotrimeric G-proteins into their inactivated state, abolished the channel activation induced by the fungal elicitor, whereas guanosine 5[prime][[gamma]-thio]triphosphate, a nonhydrolyzable GTP analog that locks heterotrimeric G-proteins into their activated state, produced an effect similar to that observed with the fungal elicitor. Mastoparan, which stimulates GTPase activity, mimicked the effect of GTP[[gamma

  7. Spermatogonial stem cells specific marker identification in channel catfish, Ictalurus punctatus and blue catfish, I. furcatus.

    PubMed

    Shang, Mei; Su, Baofeng; Lipke, Elizabeth A; Perera, Dayan A; Li, Chao; Qin, Zhenkui; Li, Yun; Dunn, David A; Cek, Sehriban; Peatman, Eric; Dunham, Rex A

    2015-12-01

    Testicular germ cells of channel catfish, Ictalurus punctatus, and blue catfish, I. furcatus were separated into four layers with Percoll density gradient centrifugation, containing different cell types (40% in the first layer were spermatogonial stem cells, SSCs). Expression of seventeen genes was analyzed for cells from different layers by real-time quantitative PCR. Pfkfb4, Urod, Plzf, Integrin6, IntegrinV, Thy1 and Cdh1 genes showed the same expression change pattern in both channel and blue catfish as these genes were down-regulated in the spermatocytes and even more so in spermatids. Plzf and Integrin6 had especially high expression in SSCs and can be used as SSCs specific markers. Sox2 gene was up-regulated in spermatocytes and even more highly up-regulated in spermatids, which indicated it could be a spermatid marker. In contrast to channel catfish, Id4, Smad5 and Prdm14 gene expressions were strongly down-regulated in spermatocyte cells, but up-regulated in spermatid cells in blue catfish. Smad5 gene was down-regulated in spermatocytes, but up-regulated in both spermatogonia and spermatids, allowing identification as a marker for spermatocytes in blue catfish. Oct4, Id4, Gfrα2, Pum2 and Prdm14 genes showed different expression patterns in the testicular germ cells of channel and blue catfish. This may be a partial explanation to the differing responses of channel catfish and blue catfish to induced spawning technologies. The SSCs specific markers can be used for further SSCs labeling, which can increase the SSCs sorting efficiency and be applied in various studies involving SSCs and other germ cells. PMID:26251285

  8. Bioinformatic Characterization of the Trimeric Intracellular Cation-Specific Channel Protein Family

    PubMed Central

    Silverio, Abe L. F.

    2014-01-01

    Trimeric intracellular cation-specific (TRIC) channels are integral to muscle excitation–contraction coupling. TRIC channels provide counter-ionic flux when calcium is rapidly transported from intracellular stores to the cell cytoplasm. Until recently, knowledge of the presence of these proteins was limited to animals. We analyzed the TRIC family and identified a profusion of prokaryotic family members with topologies and motifs similar to those of their eukaryotic counterparts. Prokaryotic members far outnumber eukaryotic members, and although none has been functionally characterized, the evidence suggests that they function as secondary carriers. The presence of fused N- or C-terminal domains of known biochemical functions as well as genomic context analyses provide clues about the functions of these prokaryotic homologs. They are proposed to function in metabolite (e.g., amino acid/ nucleotide) efflux. Phylogenetic analysis revealed that TRIC channel homologs diverged relatively early during evolutionary history and that horizontal gene transfer was frequent in prokaryotes but not in eukaryotes. Topological analyses of TRIC channels revealed that these proteins possess seven putative transmembrane segments (TMSs), which arose by intragenic duplication of a three-TMS polypeptide-encoding genetic element followed by addition of a seventh TMS at the C terminus to give the precursor of all current TRIC family homologs. We propose that this family arose in prokaryotes. PMID:21519847

  9. Synthesis and Evaluation of Potent KCNQ2/3-Specific Channel Activators.

    PubMed

    Kumar, Manoj; Reed, Nicholas; Liu, Ruiting; Aizenman, Elias; Wipf, Peter; Tzounopoulos, Thanos

    2016-06-01

    KQT-like subfamily (KCNQ) channels are voltage-gated, noninactivating potassium ion channels, and their down-regulation has been implicated in several hyperexcitability-related disorders, including epilepsy, neuropathic pain, and tinnitus. Activators of these channels reduce the excitability of central and peripheral neurons, and, as such, have therapeutic utility. Here, we synthetically modified several moieties of the KCNQ2-5 channel activator retigabine, an anticonvulsant approved by the U.S. Food and Drug Administration. By introducing a CF3-group at the 4-position of the benzylamine moiety, combined with a fluorine atom at the 3-position of the aniline ring, we generated Ethyl (2-amino-3-fluoro-4-((4-(trifluoromethyl)benzyl)amino)phenyl)carbamate (RL648_81), a new KCNQ2/3-specific activator that is >15 times more potent and also more selective than retigabine. We suggest that RL648_81 is a promising clinical candidate for treating or preventing neurologic disorders associated with neuronal hyperexcitability. PMID:27005699

  10. Calcium Channel CaVα₁ Splice Isoforms - Tissue Specificity and Drug Action.

    PubMed

    Lipscombe, Diane; Andrade, Arturo

    2015-01-01

    Voltage-gated calcium ion channels are essential for numerous biological functions of excitable cells and there is wide spread appreciation of their importance as drug targets in the treatment of many disorders including those of cardiovascular and nervous systems. Each Cacna1 gene has the potential to generate a number of structurally, functionally, and in some cases pharmacologically unique CaVα1 subunits through alternative pre-mRNA splicing and the use of alternate promoters. Analyses of rapidly emerging deep sequencing data for a range of human tissue transcriptomes contain information to quantify tissue-specific and alternative exon usage patterns for Cacna1 genes. Cellspecific actions of nuclear DNA and RNA binding proteins control the use of alternate promoters and the selection of alternate exons during pre-mRNA splicing, and they determine the spectrum of protein isoforms expressed within different types of cells. Amino acid compositions within discrete protein domains can differ substantially among CaV isoforms expressed in different tissues, and such differences may be greater than those that exist across CaV channel homologs of closely related species. Here we highlight examples of CaV isoforms that have unique expression patterns and that exhibit different pharmacological sensitivities. Knowledge of expression patterns of CaV isoforms in different human tissues, cell populations, ages, and disease states should inform strategies aimed at developing the next generation of CaV channel inhibitors and agonists with improved tissue-specificity. PMID:25966698

  11. Sildenafil prevents the up-regulation of transient receptor potential canonical channels in the development of cardiomyocyte hypertrophy

    SciTech Connect

    Kiso, Hironori; Ohba, Takayoshi; Iino, Kenji; Sato, Kazuhiro; Terata, Yutaka; Murakami, Manabu; Ono, Kyoichi; Watanabe, Hiroyuki; Ito, Hiroshi

    2013-07-05

    Highlights: •Transient receptor potential canonical (TRPC1, 3 and 6) are up-regulated by ET-1. •Sildenafil inhibited hypertrophic responses (BNP, Ca entry, NFAT activation). •Sildenafil suppressed TRPC1, 3 and 6 expression. -- Abstract: Background: Transient receptor potential canonical (TRPCs) channels are up-regulated in the development of cardiac hypertrophy. Sildenafil inhibits TRPC6 activation and expression, leading to the prevention of cardiac hypertrophy. However, the effects of sildenafil on the expression of other TRPCs remain unknown. We hypothesized that in addition to its effects of TRPC6, sildenafil blocks the up-regulation of other TRPC channels to suppress cardiomyocyte hypertrophy. Methods and results: In cultured neonatal rat cardiomyocytes, a 48 h treatment with 10 nM endothelin (ET)-1 induced hypertrophic responses characterized by nuclear factor of activated T cells activation and enhancement of brain natriuretic peptide expression and cell surface area. Co-treatment with sildenafil (1 μM, 48 h) inhibited these ET-1-induced hypertrophic responses. Although ET-1 enhanced the gene expression of TRPCs, sildenafil inhibited the enhanced gene expression of TRPC1, C3 and C6. Moreover, co-treatment with sildenafil abolished the augmentation of SOCE in the hypertrophied cardiomyocytes. Conclusions: These results suggest that sildenafil inhibits cardiomyocyte hypertrophy by suppressing the up-regulation of TRPC expression.

  12. Evolutionary Genomics Reveals Lineage-Specific Gene Loss and Rapid Evolution of a Sperm-Specific Ion Channel Complex: CatSpers and CatSperβ

    PubMed Central

    Cai, Xinjiang; Clapham, David E.

    2008-01-01

    The mammalian CatSper ion channel family consists of four sperm-specific voltage-gated Ca2+ channels that are crucial for sperm hyperactivation and male fertility. All four CatSper subunits are believed to assemble into a heteromultimeric channel complex, together with an auxiliary subunit, CatSperβ. Here, we report a comprehensive comparative genomics study and evolutionary analysis of CatSpers and CatSperβ, with important correlation to physiological significance of molecular evolution of the CatSper channel complex. The development of the CatSper channel complex with four CatSpers and CatSperβ originated as early as primitive metazoans such as the Cnidarian Nematostella vectensis. Comparative genomics revealed extensive lineage-specific gene loss of all four CatSpers and CatSperβ through metazoan evolution, especially in vertebrates. The CatSper channel complex underwent rapid evolution and functional divergence, while distinct evolutionary constraints appear to have acted on different domains and specific sites of the four CatSper genes. These results reveal unique evolutionary characteristics of sperm-specific Ca2+ channels and their adaptation to sperm biology through metazoan evolution. PMID:18974790

  13. Understanding the Cellular Function of TRPV2 Channel through Generation of Specific Monoclonal Antibodies

    PubMed Central

    Cohen, Matthew R.; Huynh, Kevin W.; Cawley, Daniel; Moiseenkova-Bell, Vera Y.

    2013-01-01

    Transient receptor potential vanilloid 2 (TRPV2) is a Ca2+-permeable nonselective cation channel proposed to play a critical role in a wide array of cellular processes. Although TRPV2 surface expression was originally determined to be sensitive to growth factor signaling, regulated trafficking of TRPV2 has remained controversial. TRPV2 has proven difficult to study due to the lack of specific pharmacological tools to modulate channel activity; therefore, most studies of the cellular function of TRPV2 rely on immuno-detection techniques. Polyclonal antibodies against TRPV2 have not been properly validated and characterized, which may contribute to conflicting results regarding its function in the cell. Here, we developed monoclonal antibodies using full-length TRPV2 as an antigen. Extensive characterization of these antibodies and comparison to commonly used commercially available TRPV2 antibodies revealed that while monoclonal antibodies generated in our laboratory were suitable for detection of endogenous TRPV2 by western blot, immunoprecipitation and immunocytochemistry, the commercially available polyclonal antibodies we tested were not able to recognize endogenous TRPV2. We used our newly generated and validated TRPV2 antibodies to determine the effects of insulin-like growth factor 1 (IGF-1) on TRPV2 surface expression in heterologous and endogenous expression systems. We found that IGF-1 had little to no effect on trafficking and plasma membrane expression of TRPV2. Overall, these new TRPV2 monoclonal antibodies served to dispel the controversy of the effects of IGF-1 on TRPV2 plasma membrane expression and will clarify the role TRPV2 plays in cellular function. Furthermore, our strategy of using full-length tetrameric TRP channels may allow for the generation of antibodies against other TRP channels of unclear function. PMID:24392006

  14. Upregulation of canonical transient receptor potential channel in the pulmonary arterial smooth muscle of a chronic thromboembolic pulmonary hypertension rat model.

    PubMed

    Yun, Xin; Chen, Yuqin; Yang, Kai; Wang, Sabrina; Lu, Wenju; Wang, Jian

    2015-12-01

    Chronic ligation of the left main pulmonary artery (PA) results in pulmonary vascular remodeling and sustained vasoconstriction. This method has been used to generate a postobstructive pulmonary vasculopathy model to mimic severe chronic thromboembolic pulmonary hypertension (CTEPH). The aim of this study was to examine the cellular and molecular mechanisms underlying CTEPH and to provide evidence for potential treatments. The CTEPH rat model was induced by surgical left PA ligation (LPAL). Right ventricular systolic pressure (RVSP), lung histochemistry and plasma D-dimer measurements were carried out to evaluate the model. A fluorescence microscope was used to measure the basal intracellular Ca(2+) concentration ([Ca(2+)]i) and store-operated Ca(2+) entry (SOCE) in rat distal pulmonary arterial smooth muscle cells through a Fura-2 fluorescence-based method. The expression of the canonical transient receptor potential channel 1 (TRPC1) and TRPC6 was determined by western blotting and real-time quantitative PCR in isolated distal pulmonary arteries (PA). At the time points of 2 and 5 weeks postsurgery, the RVSP showed significant increases in the LPAL groups in comparison with the respective control groups. LPAL also led to right ventricular hypertrophy (RVH), distal pulmonary arterial remodeling in unobstructed territories and persistently higher plasma D-dimer levels. Increases of the basal [Ca(2+)]i and SOCE in LPAL were associated with a clear upregulation of TRPC1 and TRPC6 expression in the distal PA. Our study demonstrated that LPAL successfully reproduced the vascular tone changes that mimic CTEPH pathogenesis. In this model, the increased RVSP and RVH are likely related to enhanced SOCE and upregulated TRPC1 and TRPC6 expression levels in the distal PA. PMID:26155749

  15. Open conformation of hERG channel turrets revealed by a specific scorpion toxin BmKKx2

    PubMed Central

    2014-01-01

    Background The human ether-a-go-go-related gene potassium channel (hERG) has an unusual long turret, whose role in recognizing scorpion toxins remains controversial. Here, BmKKx2, the first specific blocker of hERG channel derived from scorpion Mesobuthus martensii, was identified and the turret role of hERG channel was re-investigated using BmKKx2 as a molecular probe. Results BmKKx2 was found to block hERG channel with an IC50 of 6.7 ± 1.7 nM and share similar functional surface with the known hERG channel inhibitor BeKm-1. The alanine-scanning mutagenesis data indicate that different residue substitutions on hERG channel by alanine decreased the affinities of toxin BmKKx2 by about 10-fold compared with that of wild-type hERG channel, which reveals that channel turrets play a secondary role in toxin binding. Different from channel turret, the pore region of hERG channel was found to exert the conserved and essential function for toxin binding because the mutant hERG-S631A channel remarkably decreased toxin BmKKx2 affinity by about 104-fold. Conclusions Our results not only revealed that channel turrets of hERG channel formed an open conformation in scorpion toxin binding, but also enriched the diversity of structure-function relationships among the different potassium channel turrets. PMID:24725272

  16. Multiple genes, tissue specificity, and expression-dependent modulationcontribute to the functional diversity of potassium channels in Arabidopsis thaliana.

    PubMed Central

    Cao, Y; Ward, J M; Kelly, W B; Ichida, A M; Gaber, R F; Anderson, J A; Uozumi, N; Schroeder, J I; Crawford, N M

    1995-01-01

    K+ channels play diverse roles in mediating K+ transport and in modulating the membrane potential in higher plant cells during growth and development. Some of the diversity in K+ channel functions may arise from the regulated expression of multiple genes encoding different K+ channel polypeptides. Here we report the isolation of a novel Arabidopsis thaliana cDNA (AKT2) that is highly homologous to the two previously identified K+ channel genes, KAT1 and AKT1. This cDNA mapped to the center of chromosome 4 by restriction fragment length polymorphism analysis and was highly expressed in leaves, whereas AKT1 was mainly expressed in roots. In addition, we show that diversity in K+ channel function may be attributable to differences in expression levels. Increasing KAT1 expression in Xenopus oocytes by polyadenylation of the KAT1 mRNA increased the current amplitude and led to higher levels of KAT1 protein, as assayed in western blots. The increase in KAT1 expression in oocytes produced shifts in the threshold potential for activation to more positive membrane potentials and decreased half-activation times. These results suggest that different levels of expression and tissue-specific expression of different K+ channel isoforms can contribute to the functional diversity of plant K+ channels. The identification of a highly expressed, leaf-specific K+ channel homolog in plants should allow further molecular characterization of K+ channel functions for physiological K+ transport processes in leaves. PMID:8552711

  17. Specific profiles of ion channels and ionotropic receptors define adipose- and bone marrow derived stromal cells.

    PubMed

    Forostyak, Oksana; Butenko, Olena; Anderova, Miroslava; Forostyak, Serhiy; Sykova, Eva; Verkhratsky, Alexei; Dayanithi, Govindan

    2016-05-01

    Adherent, fibroblastic cells from different tissues are thought to contain subsets of tissue-specific stem/progenitor cells (often called mesenchymal stem cells). These cells display similar cell surface characteristics based on their fibroblastic nature, but also exhibit differences in molecular phenotype, growth rate, and their ability to differentiate into various cell phenotypes. The mechanisms underlying these differences remain poorly understood. We analyzed Ca(2+) signals and membrane properties in rat adipose-derived stromal cells (ADSCs) and bone marrow stromal cells (BMSCs) in basal conditions, and then following a switch into medium that contains factors known to modify their character. Modified ADSCs (mADSCs) expressed L-type Ca(2+) channels whereas both L- and P/Q- channels were operational in mBMSCs. Both mADSCs and mBMSCs possessed functional endoplasmic reticulum Ca(2+) stores, expressed ryanodine receptor-1 and -3, and exhibited spontaneous [Ca(2+)]i oscillations. The mBMSCs expressed P2X7 purinoceptors; the mADSCs expressed both P2X (but not P2X7) and P2Y (but not P2Y1) receptors. Both types of stromal cells exhibited [Ca(2+)]i responses to vasopressin (AVP) and expressed V1 type receptors. Functional oxytocin (OT) receptors were, in contrast, expressed only in modified ADSCs and BMSCs. AVP and OT-induced [Ca(2+)]i responses were dose-dependent and were blocked by their respective specific receptor antagonists. Electrophysiological data revealed that passive ion currents dominated the membrane conductance in ADSCs and BMSCs. Medium modification led to a significant shift in the reversal potential of passive currents from -40 to -50mV in cells in basal to -80mV in modified cells. Hence membrane conductance was mediated by non-selective channels in cells in basal conditions, whereas in modified medium conditions, it was associated with K(+)-selective channels. Our results indicate that modification of ADSCs and BMSCs by alteration in medium

  18. Kv8.1, a new neuronal potassium channel subunit with specific inhibitory properties towards Shab and Shaw channels.

    PubMed Central

    Hugnot, J P; Salinas, M; Lesage, F; Guillemare, E; de Weille, J; Heurteaux, C; Mattéi, M G; Lazdunski, M

    1996-01-01

    Outward rectifier K+ channels have a characteristic structure with six transmembrane segments and one pore region. A new member of this family of transmembrane proteins has been cloned and called Kv8.1. Kv8.1 is essentially present in the brain where it is located mainly in layers II, IV and VI of the cerebral cortex, in hippocampus, in CA1-CA4 pyramidal cell layer as well in granule cells of the dentate gyrus, in the granule cell layer and in the Purkinje cell layer of the cerebellum. The Kv8.1 gene is in the 8q22.3-8q24.1 region of the human genome. Although Kv8.1 has the hallmarks of functional subunits of outward rectifier K+ channels, injection of its cRNA in Xenopus oocytes does not produce K+ currents. However Kv8.1 abolishes the functional expression of members of the Kv2 and Kv3 subfamilies, suggesting that the functional role of Kv8.1 might be to inhibit the function of a particular class of outward rectifier K+ channel types. Immunoprecipitation studies have demonstrated that inhibition occurs by formation of heteropolymeric channels, and results obtained with Kv8.1 chimeras have indicated that association of Kv8.1 with other types of subunits is via its N-terminal domain. Images PMID:8670833

  19. [The relation of the relative specificity of point to channel lines or spinal segments].

    PubMed

    Liu, X; Zhang, S; Wei, B

    1995-01-01

    This work is to study whut the analgesia of EA is related with specificity of channel point or innervation of point. The discharges of 28 NRM neurons and their nociceptive responses induced by stimulation of dental pulp or tail tip in rats were recorded. The effects of EA of "Shangguan" and "Yanglingquan" of gall bladder channel of foot shaoyang on NRM neurons and their nociceptive responses were compared. and with that of "Xiaguan" and "Zusanli" of the stomach channel of foot Yangming. "Shangguan" and "Xiaguan" were located in the depressions at the upper and lower border of zygomatic arch respectively, and innervated by the n. trigeminus. The hind leg points "Yanglingquan" is located in the depression anterior and inferior to capitulum fibulae above "Zusanli" and both were innervated by n. peroneus communis. The results showed that EA of "Shangguan" or "Xiaguan" could activate NRM, increasing unit discharges about 70-120% for 10-15 minutes (P < 0.05-0.001) and inhibit both nociceptive responses, but the inhibition on the nociceptive response induced by dental pulp (about -60% for 5-10 minutes, P < 0.05-0.001) was stronger than that by tail tip (no effect or about -30% occasionally). "Yanglingquan" or "Zusanli" EA could also activate NRM and inhibit the nociceptive responses, but the inhibition on the nociceptive response of tail stimulation about -60-(-)40% for 30 minutes, P < 0.05-0.001) was stronger and longer than that of dental pulp (about -40% for 5-10 minutes, P < 0.05-0.001).(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7648703

  20. Phosphorylation regulates the water channel activity of the seed-specific aquaporin alpha-TIP.

    PubMed

    Maurel, C; Kado, R T; Guern, J; Chrispeels, M J

    1995-07-01

    The vacuolar membrane protein alpha-TIP is a seed-specific protein of the Major Intrinsic Protein family. Expression of alpha-TIP in Xenopus oocytes conferred a 4- to 8-fold increase in the osmotic water permeability (Pf) of the oocyte plasma membrane, showing that alpha-TIP forms water channels and is thus a new aquaporin. alpha-TIP has three putative phosphorylation sites on the cytoplasmic side of the membrane (Ser7, Ser23 and Ser99), one of which (Ser7) has been shown to be phosphorylated. We present several lines of evidence that the activity of this aquaporin is regulated by phosphorylation. First, mutation of the putative phosphorylation sites in alpha-TIP (Ser7Ala, Ser23Ala and Ser99Ala) reduced the apparent water transport activity of alpha-TIP in oocytes, suggesting that phosphorylation of alpha-TIP occurs in the oocytes and participates in the control of water channel activity. Second, exposure of oocytes to the cAMP agonists 8-bromoadenosine 3',5'-cyclic monophosphate, forskolin and 3-isobutyl-1-methylxanthine, which stimulate endogenous protein kinase A (PKA), increased the water transport activity of alpha-TIP by 80-100% after 60 min. That the protein can be phosphorylated by PKA was demonstrated by phosphorylating alpha-TIP in isolated oocyte membranes with the bovine PKA catalytic subunit. Third, the integrity of the three sites at positions 7, 23 and 99 was necessary for the cAMP-dependent increase in the Pf of oocytes expressing alpha-TIP, as well as for in vitro phosphorylation of alpha-TIP. These findings demonstrate that the alpha-TIP water channel can be modulated via phosphorylation of Ser7, Ser23 and Ser99.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7542585

  1. Isoform-specific Prolongation of Kv7 (KCNQ) Potassium Channel Opening Mediated by New Molecular Determinants for Drug-Channel Interactions*

    PubMed Central

    Gao, Zhaobing; Zhang, Tangzhi; Wu, Meng; Xiong, Qiaojie; Sun, Haiyan; Zhang, Yinan; Zu, Liansuo; Wang, Wei; Li, Min

    2010-01-01

    Kv7 channels, especially Kv7.2 (KCNQ2) and Kv7.3 (KCNQ3), are key determinants for membrane excitability in the brain. Some chemical modulators of KCNQ channels are in development for use as anti-epileptic drugs, such as retigabine (D-23129, N-(2-amino-4-(4-fluorobenzylamino)-phenyl)), which was recently approved for clinical use. In addition, several other compounds were also reported to potentiate activity of the Kv7 channels. It is therefore of interest to investigate compound-channel interactions, so that more insights may be gained to aid future development of therapeutics. We have conducted a screen of 20,000 compounds for KCNQ2 potentiators using rubidium flux combined with atomic absorption spectrometry. Here, we report the characterization of a series of new structures that display isoform specificity and induce a marked reduction of deactivation distinct from that of retigabine. Furthermore, KCNQ2(W236L), a previously reported mutation that abolishes sensitivity to retigabine, remains fully sensitive to these compounds. This result, together with mutagenesis and other studies, suggests that the reported compounds confer a unique mode of action and involve new molecular determinants on the channel protein, consistent with the idea of recognizing a new site on channel protein. PMID:20584905

  2. Channel

    NASA Technical Reports Server (NTRS)

    2006-01-01

    [figure removed for brevity, see original site] Context image for PIA03693 Channel

    This channel is located south of Iani Chaos.

    Image information: VIS instrument. Latitude -10.9N, Longitude 345.5E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  3. Understanding transporter specificity and the discrete appearance of channel-like gating domains in transporters

    PubMed Central

    Diallinas, George

    2014-01-01

    Transporters are ubiquitous proteins mediating the translocation of solutes across cell membranes, a biological process involved in nutrition, signaling, neurotransmission, cell communication and drug uptake or efflux. Similarly to enzymes, most transporters have a single substrate binding-site and thus their activity follows Michaelis-Menten kinetics. Substrate binding elicits a series of structural changes, which produce a transporter conformer open toward the side opposite to the one from where the substrate was originally bound. This mechanism, involving alternate outward- and inward-facing transporter conformers, has gained significant support from structural, genetic, biochemical and biophysical approaches. Most transporters are specific for a given substrate or a group of substrates with similar chemical structure, but substrate specificity and/or affinity can vary dramatically, even among members of a transporter family that show high overall amino acid sequence and structural similarity. The current view is that transporter substrate affinity or specificity is determined by a small number of interactions a given solute can make within a specific binding site. However, genetic, biochemical and in silico modeling studies with the purine transporter UapA of the filamentous ascomycete Aspergillus nidulans have challenged this dogma. This review highlights results leading to a novel concept, stating that substrate specificity, but also transport kinetics and transporter turnover, are determined by subtle intramolecular interactions between a major substrate binding site and independent outward- or cytoplasmically-facing gating domains, analogous to those present in channels. This concept is supported by recent structural evidence from several, phylogenetically and functionally distinct transporter families. The significance of this concept is discussed in relationship to the role and potential exploitation of transporters in drug action. PMID:25309439

  4. TRP channels: sensors and transducers of gasotransmitter signals

    PubMed Central

    Takahashi, Nobuaki; Kozai, Daisuke; Mori, Yasuo

    2012-01-01

    The transient receptor potential (trp) gene superfamily encodes cation channels that act as multimodal sensors for a wide variety of stimuli from outside and inside the cell. Upon sensing, they transduce electrical and Ca2+ signals via their cation channel activities. These functional features of TRP channels allow the body to react and adapt to different forms of environmental changes. Indeed, members of one class of TRP channels have emerged as sensors of gaseous messenger molecules that control various cellular processes. Nitric oxide (NO), a vasoactive gaseous molecule, regulates TRP channels directly via cysteine (Cys) S-nitrosylation or indirectly via cyclic GMP (cGMP)/protein kinase G (PKG)-dependent phosphorylation. Recent studies have revealed that changes in the availability of molecular oxygen (O2) also control the activation of TRP channels. Anoxia induced by O2-glucose deprivation and severe hypoxia (1% O2) activates TRPM7 and TRPC6, respectively, whereas TRPA1 has recently been identified as a novel sensor of hyperoxia and mild hypoxia (15% O2) in vagal and sensory neurons. TRPA1 also detects other gaseous molecules such as hydrogen sulfide (H2S) and carbon dioxide (CO2). In this review, we focus on how signaling by gaseous molecules is sensed and integrated by TRP channels. PMID:22934072

  5. Specific serum antibody responses in channel catfish (Ictalurus punctatus) provide limited protection against Streptococcus ictaluri challenge.

    PubMed

    Pasnik, David J; Evans, Joyce J; Klesius, Phillip H

    2011-11-15

    Passive immunization studies were conducted to determine the role of specific antibodies in immunity to Streptococcus ictaluri. Adult channel catfish (Ictalurus punctatus) were injected i.p. with tryptic soy broth as control or with 1.5 × 10(7)colony-forming units (cfu) S. ictaluri/fish at 0, 30, and 60 d, and serum was collected 90 d after the original challenge. Fish were passively immunized by i.p. injection with serum from the tryptic soy broth (TSB) control group, anti-S. ictaluri serum from fish immunized three times and sampled at 90 d (SSI), or heat-inactivated anti-S. ictaluri serum from fish immunized three times and sampled at 90 d (HISSI). These passively immunized fish were then challenged 72 h later with 1.5 × 10(8)cfu S. ictaluri/fish. Over 21 d, the mean cumulative percent survival was 43.3 (TSB), 63.3 (SSI), and 50.0 (HISSI). A significant difference in cumulative percent survival was noted between the TSB and the HISSI groups, and significant differences were noted between these groups and the SSI group. Serum obtained from immunized fish 72 h after passive immunization exhibited increased anti-S. ictaluri antibody levels. Twenty-one days after the challenge, the HISSI and SSI group antibody levels significantly increased above their corresponding pre-challenge levels. No significant (r(2)=0.0806; P<0.5985) correlation between increased pre-challenge specific serum antibody levels and survival after challenge was demonstrated when analyzing the control and passive immunization groups. The results indicate that both specific anti-S. ictaluri antibodies and non-specific immune responses are important for protection against S. ictaluri. PMID:21962634

  6. Characterization of the first honeybee Ca²⁺ channel subunit reveals two novel species- and splicing-specific modes of regulation of channel inactivation.

    PubMed

    Cens, Thierry; Rousset, Matthieu; Collet, Claude; Raymond, Valérie; Démares, Fabien; Quintavalle, Annabelle; Bellis, Michel; Le Conte, Yves; Chahine, Mohamed; Charnet, Pierre

    2013-07-01

    The honeybee is a model system to study learning and memory, and Ca(2+) signals play a key role in these processes. We have cloned, expressed, and characterized the first honeybee Ca(2+) channel subunit. We identified two splice variants of the Apis CaVβ Ca(2+) channel subunit (Am-CaVβ) and demonstrated expression in muscle and neurons. Although AmCaVβ shares with vertebrate CaVβ subunits the SH3 and GK domains, it beholds a unique N terminus that is alternatively spliced in the first exon to produce a long (a) and short (b) variant. When expressed with the CaV2 channels both, AmCaVβa and AmCaVβb, increase current amplitude, shift the voltage-sensitivity of the channel, and slow channel inactivation as the vertebrate CaVβ2a subunit does. However, as opposed to CaVβ2a, slow inactivation induced by Am-CaVβa was insensitive to palmitoylation but displayed a unique PI3K sensitivity. Inactivation produced by the b variant was PI3K-insensitive but staurosporine/H89-sensitive. Deletion of the first exon suppressed the sensitivity to PI3K inhibitors, staurosporine, or H89. Recording of Ba(2+) currents in Apis neurons or muscle cells evidenced a sensitivity to PI3K inhibitors and H89, suggesting that both AmCaVβ variants may be important to couple cell signaling to Ca(2+) entry in vivo. Functional interactions with phospho-inositide and identification of phosphorylation sites in AmCaVβa and AmCaVβb N termini, respectively, suggest that AmCaVβ splicing promoted two novel and alternative modes of regulation of channel activity with specific signaling pathways. This is the first description of a splicing-dependent kinase switch in the regulation of Ca(2+) channel activity by CaVβ subunit. PMID:23588376

  7. Properties of a potassium channel in cultured human gastric cells (HGT-1) possessing specific omeprazole binding sites.

    PubMed Central

    Sandle, G I; Fraser, G; Fogg, K; Warhurst, G

    1993-01-01

    The HGT-1 human gastric cell line is similar to acid secreting parietal cells in that it possesses H2 receptors, histamine sensitive adenyl cyclase, and Cl- channels, which are activated by histamine by a cyclic adenosine monophosphate (cAMP) dependent mechanism. To discover if HGT-1 cells have additional properties found in parietal cells, [3H]omeprazole and patch clamp recording techniques were used to evaluate specific omeprazole binding sites and K+ channels in the plasma membrane. HGT-1 cells exhibited [3H]omeprazole binding in the non-stimulated state, which increased 100% in the presence of 1 mM histamine. High conductance (about 155 pS) K+ channels were active spontaneously in 17% of cell attached or excised inside out patches in non-stimulated subconfluent HGT-1 cells. In inside out patches, channel activity increased fivefold during depolarisation, ion substitution experiments confirmed that the channels were highly selective for K+, and channel activity was almost abolished by removal of Ca2+ or addition of 5 mM Ba2+. In quiescent cell attached patches, 0.1 mM dibutyryl cAMP failed to activate K+ channels. In contrast, 6.7 microM A23187 (a Ca2+ ionophore) increased intracellular Ca2+ concentration from mean (SEM) 14 (3) nM to 248 (30) nM and activated K+ channels in 21% of patches. It is concluded that the plasma membrane of HGT-1 cells possesses (a) specific 3H-omeprazole binding sites, which may reflect the omeprazole sensitive H+,K(+)-ATPase present in gastric parietal cells; and (b) Ca(2+)-activated K+ channels, which may be located in the basolateral membrane of human gastric parietal cells and play a part in acid secretion triggered by Ca(2+)-mediated secretory agonists. PMID:8244097

  8. Hair-Cell Mechanotransduction Persists in TRP Channel Knockout Mice

    PubMed Central

    Niksch, Paul D.; Webber, Roxanna M.; Garcia-Gonzalez, Miguel; Watnick, Terry; Zhou, Jing; Vollrath, Melissa A.; Corey, David P.

    2016-01-01

    Members of the TRP superfamily of ion channels mediate mechanosensation in some organisms, and have been suggested as candidates for the mechanotransduction channel in vertebrate hair cells. Some TRP channels can be ruled out based on lack of an inner ear phenotype in knockout animals or pore properties not similar to the hair-cell channel. Such studies have excluded Trpv4, Trpa1, Trpml3, Trpm1, Trpm3, Trpc1, Trpc3, Trpc5, and Trpc6. However, others remain reasonable candidates. We used data from an RNA-seq analysis of gene expression in hair cells as well as data on TRP channel conductance to narrow the candidate group. We then characterized mice lacking functional Trpm2, Pkd2, Pkd2l1, Pkd2l2 and Pkd1l3, using scanning electron microscopy, auditory brainstem response, permeant dye accumulation, and single-cell electrophysiology. In all of these TRP-deficient mice, and in double and triple knockouts, mechanotransduction persisted. Together with published studies, these results argue against the participation of any of the 33 mouse TRP channels in hair cell transduction. PMID:27196058

  9. Hair-Cell Mechanotransduction Persists in TRP Channel Knockout Mice.

    PubMed

    Wu, Xudong; Indzhykulian, Artur A; Niksch, Paul D; Webber, Roxanna M; Garcia-Gonzalez, Miguel; Watnick, Terry; Zhou, Jing; Vollrath, Melissa A; Corey, David P

    2016-01-01

    Members of the TRP superfamily of ion channels mediate mechanosensation in some organisms, and have been suggested as candidates for the mechanotransduction channel in vertebrate hair cells. Some TRP channels can be ruled out based on lack of an inner ear phenotype in knockout animals or pore properties not similar to the hair-cell channel. Such studies have excluded Trpv4, Trpa1, Trpml3, Trpm1, Trpm3, Trpc1, Trpc3, Trpc5, and Trpc6. However, others remain reasonable candidates. We used data from an RNA-seq analysis of gene expression in hair cells as well as data on TRP channel conductance to narrow the candidate group. We then characterized mice lacking functional Trpm2, Pkd2, Pkd2l1, Pkd2l2 and Pkd1l3, using scanning electron microscopy, auditory brainstem response, permeant dye accumulation, and single-cell electrophysiology. In all of these TRP-deficient mice, and in double and triple knockouts, mechanotransduction persisted. Together with published studies, these results argue against the participation of any of the 33 mouse TRP channels in hair cell transduction. PMID:27196058

  10. Distribution of transient receptor potential channels in the rat carotid chemosensory pathway.

    PubMed

    Buniel, Maria C F; Schilling, William P; Kunze, Diana L

    2003-09-22

    Glomus cells in the carotid body respond to decreases in oxygen tension of the blood and transmit this sensory information in the carotid sinus nerve to the brain via neurons in the petrosal ganglion. G-protein-coupled membrane receptors linked to phospholipase C may play an important role in this response through the activation of the cation channels formed by the transient receptor potential (TRP) proteins. In the present study, expression of TRPC proteins in the rat carotid body and petrosal ganglion was examined using immunohistochemical techniques. TRPC3, TRPC4, TRPC5, TRPC6, and TRPC7 were present in neurons throughout the ganglion. TRPC1 was expressed in only 28% of petrosal neurons, and of this population, 45% were tyrosine hydroxylase (TH)-positive, accounting for essentially all the TH-expressing neurons in the ganglion. Because TH-positive neurons project to the carotid body, this result suggests that TRPC1 is selectively associated with the chemosensory pathway. Confocal images through the carotid body showed that TRPC1/3/4/5/6 proteins localize to the carotid sinus nerve fibers, some of which were immunoreactive to an anti-neurofilament (NF) antibody cocktail. TRPC1 and TRPC3 were present in both NF-positive and NF-negative fibers, whereas TPRC4, TRPC5, and TRPC6 expression was primarily localized to NF-negative fibers. Only TRPC1 and TRPC4 were localized in the afferent nerve terminals that encircle individual glomus cells. TRPC7 was not expressed in sensory fibers. All the TRPC proteins studied were present in type I glomus cells. Although their role as receptor-activated cation channels in the chemosensory pathway is yet to be established, the presence of TRPC channels in glomus cells and sensory nerves of the carotid body suggests a role in facilitating and/or sustaining the hypoxic response. PMID:12900933

  11. Calciseptine, a peptide isolated from black mamba venom, is a specific blocker of the L-type calcium channel.

    PubMed Central

    de Weille, J R; Schweitz, H; Maes, P; Tartar, A; Lazdunski, M

    1991-01-01

    The venom of the black mamba contains a 60-amino acid peptide called calciseptine. The peptide has been fully sequenced. It is a smooth muscle relaxant and an inhibitor of cardiac contractions. Its physiological action resembles that of drugs, such as the 1,4-dihydropyridines, which are important in the treatment of cardiovascular diseases. Calciseptine, like the 1,4-dihydropyridines, selectively blocks L-type Ca2+ channels and is totally inactive on other voltage-dependent Ca2+ channels such as N-type and T-type channels. To our knowledge, it is the only natural polypeptide that has been shown to be a specific inhibitor of L-type Ca2+ channels. Images PMID:1848702

  12. Cartilage-Specific Knockout of the Mechanosensory Ion Channel TRPV4 Decreases Age-Related Osteoarthritis

    PubMed Central

    O’Conor, Christopher J.; Ramalingam, Sendhilnathan; Zelenski, Nicole A.; Benefield, Halei C.; Rigo, Isaura; Little, Dianne; Wu, Chia-Lung; Chen, Di; Liedtke, Wolfgang; McNulty, Amy L.; Guilak, Farshid

    2016-01-01

    Osteoarthritis (OA) is a progressive degenerative disease of articular cartilage and surrounding tissues, and is associated with both advanced age and joint injury. Biomechanical factors play a critical role in the onset and progression of OA, yet the mechanisms through which physiologic or pathologic mechanical signals are transduced into a cellular response are not well understood. Defining the role of mechanosensory pathways in cartilage during OA pathogenesis may yield novel strategies or targets for the treatment of OA. The transient receptor potential vanilloid 4 (TRPV4) ion channel transduces mechanical loading of articular cartilage via the generation of intracellular calcium ion transients. Using tissue-specific, inducible Trpv4 gene-targeted mice, we demonstrate that loss of TRPV4-mediated cartilage mechanotransduction in adulthood reduces the severity of aging-associated OA. However, loss of chondrocyte TRPV4 did not prevent OA development following destabilization of the medial meniscus (DMM). These results highlight potentially distinct roles of TRPV4-mediated cartilage mechanotransduction in age-related and post-traumatic OA, and point to a novel disease-modifying strategy to therapeutically target the TRPV4-mediated mechanotransduction pathway for the treatment of aging-associated OA. PMID:27388701

  13. Rare cell chemiluminescence detection based on aptamer-specific capture in microfluidic channels.

    PubMed

    Liu, Wu; Wei, Huibin; Lin, Zhen; Mao, Sifeng; Lin, Jin-Ming

    2011-10-15

    An aptamer-based "sandwich" approach combined with the chemiluminescence (CL) analysis was developed for the capture and detection of rare cells on a microfluidic chip. Aptamers were immobilized on microfluidic channels to achieve capture and isolation of the specific cells from a cell mixture. The capture efficiency for target cells was more than 70% with the purity greater than 97%, when the content of the target cells was between 0.5% and 10% in the initial cell mixture. Gold nanoparticles (Au NPs) modified with aptamers were then added in to bind on the cells and trigger a CL reaction. A satisfactory linearity of the log/log calibration curve between the CL intensity and the number of target cells was observed with a low detection limit of 30 target cells in a 3 μL cell mixture. Spiked whole blood samples were also used to verify the practicality of the present method. This work demonstrated the potential application of the cheap and rapid CL detection into the early diagnosis of cancers. PMID:21856143

  14. Quaternary ammonium compounds as water channel blockers. Specificity, potency, and site of action.

    PubMed

    Detmers, Frank J M; de Groot, Bert L; Müller, E Matthias; Hinton, Andrew; Konings, Irene B M; Sze, Mozes; Flitsch, Sabine L; Grubmüller, Helmut; Deen, Peter M T

    2006-05-19

    Excessive water uptake through Aquaporins (AQP) can be life-threatening and reversible AQP inhibitors are needed. Here, we determined the specificity, potency, and binding site of tetraethylammonium (TEA) to block Aquaporin water permeability. Using oocytes, externally applied TEA blocked AQP1/AQP2/AQP4 with IC50 values of 1.4, 6.2, and 9.8 microM, respectively. Related tetraammonium compounds yielded some (propyl) or no (methyl, butyl, or pentyl) inhibition. TEA inhibition was lost upon a Tyr to Phe amino acid switch in the external water pore of AQP1/AQP2/AQP4, whereas the water permeability of AQP3 and AQP5, which lack a corresponding Tyr, was not blocked by TEA. Consistent with experimental data, multi-nanosecond molecular dynamics simulations showed one stable binding site for TEA, but not tetramethyl (TMA), in AQP1, resulting in a nearly 50% water permeability inhibition, which was reduced in AQP1-Y186F due to effects on the TEA inhibitory binding region. Moreover, in the simulation TEA interacted with charged residues in the C (Asp128) and E (Asp185) loop, and the A(Tyr37-Asn42-Thr44) loop of the neighboring monomer, but not directly with Tyr186. The loss of TEA inhibition in oocytes expressing properly folded AQP1-N42A or -T44A is in line with the computationally predicted binding mode. Our data reveal that the molecular interaction of TEA with AQP1 differs and is about 1000-fold more effective on AQPs than on potassium channels. Moreover, the observed experimental and simulated similarities open the way for rational design and virtual screening for AQP-specific inhibitors, with quaternary ammonium compounds in general, and TEA in particular as a lead compound. PMID:16551622

  15. Immunohistochemical study on the distribution of canonical transient receptor potential channels in rat basal ganglia.

    PubMed

    Chung, Yoon Hee; Kim, Daejin; Moon, Nam Joo; Oh, Chang Seok; Lee, Eunju; Shin, Dong Hoon; Kim, Sung Su; Lee, Won Bok; Lee, Jun-Young; Cha, Choong Ik

    2007-07-01

    In the present study, we examined the localizations of canonical transient receptor potential channels (TRPCs) in rat basal ganglia. The dot-like staining pattern of TRPC5 was observed through the globus pallidus (GP) and caudate-putamen. TRPC7 had a strikingly high level of expression in the neuropil in the GP. In the subthalamic nucleus, strong staining for TRPC5 was observed in the cell bodies, while moderate to high immunoreactivies for TRPC1, TRPC3, TRPC4 and TRPC7 were found in the cell bodies and surrounding neuropil. In the substantia nigra, immunoreactivities for TRPC3 and TRPC7 were prominent in the cell bodies and several processes in the pars compacta and pars reticulata. TRPC6 was expressed in the neuropil, not in the cell bodies. This study may provide useful data for the future investigations on the structural and functional properties of TRPCs. PMID:17590510

  16. Sequence and structure-specific elements of HERG mRNA determine channel synthesis and trafficking efficiency

    PubMed Central

    Sroubek, Jakub; Krishnan, Yamini; McDonald, Thomas V.

    2013-01-01

    Human ether-á-gogo-related gene (HERG) encodes a potassium channel that is highly susceptible to deleterious mutations resulting in susceptibility to fatal cardiac arrhythmias. Most mutations adversely affect HERG channel assembly and trafficking. Why the channel is so vulnerable to missense mutations is not well understood. Since nothing is known of how mRNA structural elements factor in channel processing, we synthesized a codon-modified HERG cDNA (HERG-CM) where the codons were synonymously changed to reduce GC content, secondary structure, and rare codon usage. HERG-CM produced typical IKr-like currents; however, channel synthesis and processing were markedly different. Translation efficiency was reduced for HERG-CM, as determined by heterologous expression, in vitro translation, and polysomal profiling. Trafficking efficiency to the cell surface was greatly enhanced, as assayed by immunofluorescence, subcellular fractionation, and surface labeling. Chimeras of HERG-NT/CM indicated that trafficking efficiency was largely dependent on 5′ sequences, while translation efficiency involved multiple areas. These results suggest that HERG translation and trafficking rates are independently governed by noncoding information in various regions of the mRNA molecule. Noncoding information embedded within the mRNA may play a role in the pathogenesis of hereditary arrhythmia syndromes and could provide an avenue for targeted therapeutics.—Sroubek, J., Krishnan, Y., McDonald, T V. Sequence- and structure-specific elements of HERG mRNA determine channel synthesis and trafficking efficiency. PMID:23608144

  17. Cell-type-specific tuning of Cav1.3 Ca2+-channels by a C-terminal automodulatory domain

    PubMed Central

    Scharinger, Anja; Eckrich, Stephanie; Vandael, David H.; Schönig, Kai; Koschak, Alexandra; Hecker, Dietmar; Kaur, Gurjot; Lee, Amy; Sah, Anupam; Bartsch, Dusan; Benedetti, Bruno; Lieb, Andreas; Schick, Bernhard; Singewald, Nicolas; Sinnegger-Brauns, Martina J.; Carbone, Emilio; Engel, Jutta; Striessnig, Jörg

    2015-01-01

    Cav1.3 L-type Ca2+-channel function is regulated by a C-terminal automodulatory domain (CTM). It affects channel binding of calmodulin and thereby tunes channel activity by interfering with Ca2+- and voltage-dependent gating. Alternative splicing generates short C-terminal channel variants lacking the CTM resulting in enhanced Ca2+-dependent inactivation and stronger voltage-sensitivity upon heterologous expression. However, the role of this modulatory domain for channel function in its native environment is unkown. To determine its functional significance in vivo, we interrupted the CTM with a hemagglutinin tag in mutant mice (Cav1.3DCRDHA/HA). Using these mice we provide biochemical evidence for the existence of long (CTM-containing) and short (CTM-deficient) Cav1.3 α1-subunits in brain. The long (HA-labeled) Cav1.3 isoform was present in all ribbon synapses of cochlear inner hair cells. CTM-elimination impaired Ca2+-dependent inactivation of Ca2+-currents in hair cells but increased it in chromaffin cells, resulting in hyperpolarized resting potentials and reduced pacemaking. CTM disruption did not affect hearing thresholds. We show that the modulatory function of the CTM is affected by its native environment in different cells and thus occurs in a cell-type specific manner in vivo. It stabilizes gating properties of Cav1.3 channels required for normal electrical excitability. PMID:26379493

  18. G-protein beta-subunit specificity in the fast membrane-delimited inhibition of Ca2+ channels.

    PubMed

    García, D E; Li, B; García-Ferreiro, R E; Hernández-Ochoa, E O; Yan, K; Gautam, N; Catterall, W A; Mackie, K; Hille, B

    1998-11-15

    We investigated which subtypes of G-protein beta subunits participate in voltage-dependent modulation of N-type calcium channels. Calcium currents were recorded from cultured rat superior cervical ganglion neurons injected intranuclearly with DNA encoding five different G-protein beta subunits. Gbeta1 and Gbeta2 strongly mimicked the fast voltage-dependent inhibition of calcium channels produced by many G-protein-coupled receptors. The Gbeta5 subunit produced much weaker effects than Gbeta1 and Gbeta2, whereas Gbeta3 and Gbeta4 were nearly inactive in these electrophysiological studies. The specificity implied by these results was confirmed and extended using the yeast two-hybrid system to test for protein-protein interactions. Here, Gbeta1 or Gbeta2 coupled to the GAL4-activation domain interacted strongly with a channel sequence corresponding to the intracellular loop connecting domains I and II of a alpha1 subunit of the class B calcium channel fused to the GAL4 DNA-binding domain. In this assay, the Gbeta5 subunit interacted weakly, and Gbeta3 and Gbeta4 failed to interact. Together, these results suggest that Gbeta1 and/or Gbeta2 subunits account for most of the voltage-dependent inhibition of N-type calcium channels and that the linker between domains I and II of the calcium channel alpha1 subunit is a principal receptor for this inhibition. PMID:9801356

  19. Homeostasis or channelopathy? Acquired cell type-specific ion channel changes in temporal lobe epilepsy and their antiepileptic potential

    PubMed Central

    Wolfart, Jakob; Laker, Debora

    2015-01-01

    Neurons continuously adapt the expression and functionality of their ion channels. For example, exposed to chronic excitotoxicity, neurons homeostatically downscale their intrinsic excitability. In contrast, the “acquired channelopathy” hypothesis suggests that proepileptic channel characteristics develop during epilepsy. We review cell type-specific channel alterations under different epileptic conditions and discuss the potential of channels that undergo homeostatic adaptations, as targets for antiepileptic drugs (AEDs). Most of the relevant studies have been performed on temporal lobe epilepsy (TLE), a widespread AED-refractory, focal epilepsy. The TLE patients, who undergo epilepsy surgery, frequently display hippocampal sclerosis (HS), which is associated with degeneration of cornu ammonis subfield 1 pyramidal cells (CA1 PCs). Although the resected human tissue offers insights, controlled data largely stem from animal models simulating different aspects of TLE and other epilepsies. Most of the cell type-specific information is available for CA1 PCs and dentate gyrus granule cells (DG GCs). Between these two cell types, a dichotomy can be observed: while DG GCs acquire properties decreasing the intrinsic excitability (in TLE models and patients with HS), CA1 PCs develop channel characteristics increasing intrinsic excitability (in TLE models without HS only). However, thorough examination of data on these and other cell types reveals the coexistence of protective and permissive intrinsic plasticity within neurons. These mechanisms appear differentially regulated, depending on the cell type and seizure condition. Interestingly, the same channel molecules that are upregulated in DG GCs during HS-related TLE, appear as promising targets for future AEDs and gene therapies. Hence, GCs provide an example of homeostatic ion channel adaptation which can serve as a primer when designing novel anti-epileptic strategies. PMID:26124723

  20. Canonical transient receptor potential channels expression is elevated in a porcine model of metabolic syndrome.

    PubMed

    Hu, Guoqing; Oboukhova, Elena A; Kumar, Sanjay; Sturek, Michael; Obukhov, Alexander G

    2009-05-01

    Plasma epinephrine and heart rate are elevated in metabolic syndrome, suggesting enhanced catecholamine secretion from the adrenal medulla. Canonical transient receptor potential (TRPC) channels are implicated in mediating hormone-induced Ca(2+) influx and catecholamine secretion in adrenomedullary chromaffin cells. We studied the pattern of TRPC expression in the pig adrenal medulla and investigated whether adrenal TRPC expression is altered in prediabetic metabolic syndrome Ossabaw miniature pigs. We used a combination of molecular biological, biochemical, and fluorescence imaging techniques. We determined the sequence of pig TRPC1 and TRPC3-7 channels. We found that the pig adrenal medulla expressed predominantly TRPC1, TRPC5, and TRPC6 transcripts. The expression level of these TRPCs was significantly elevated in the adrenal medulla from pigs with metabolic syndrome. Interestingly, aldosterone, which is endogenously secreted in the adjacent adrenal cortex, increased TRPC1, TRPC5, and TRPC6 expression in adrenal chromaffin cells isolated from metabolic syndrome but not control pigs. Spironolactone, a blocker of mineralocorticoid receptors, inhibited the aldosterone effect. Dexamethasone also increased TRPC5 expression in metabolic syndrome chromaffin cells. The amplitude of hormone-induced divalent cation influx correlated with the level of TRPC expression in adrenal chromaffin cells. Orai1/Stim1 protein expression was not significantly altered in the metabolic syndrome adrenal medulla when compared with the control. We propose that in metabolic syndrome, abnormally elevated adrenal TRPC expression may underlie increased plasma epinephrine and heart rate. The excess of plasma catecholamines and increased heart rate are risk factors for cardiovascular disease. Thus, TRPCs are potential therapeutic targets in the fight against cardiovascular disease. PMID:19221052

  1. Comparative genomic analysis of bacteriophages specific to the channel catfish pathogen Edwardsiella ictaluri

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: The bacterial pathogen Edwardsiella ictaluri is a primary cause of mortality in channel catfish raised commercially in aquaculture farms. Additional treatment and diagnostic regimes are needed for this enteric pathogen, motivating the discovery and characterization of bacteriophages spe...

  2. Hypoxia-Dependent Reactive Oxygen Species Signaling in the Pulmonary Circulation: Focus on Ion Channels

    PubMed Central

    Veit, Florian; Pak, Oleg; Brandes, Ralf P.

    2015-01-01

    Abstract Significance: An acute lack of oxygen in the lung causes hypoxic pulmonary vasoconstriction, which optimizes gas exchange. In contrast, chronic hypoxia triggers a pathological vascular remodeling causing pulmonary hypertension, and ischemia can cause vascular damage culminating in lung edema. Recent Advances: Regulation of ion channel expression and gating by cellular redox state is a widely accepted mechanism; however, it remains a matter of debate whether an increase or a decrease in reactive oxygen species (ROS) occurs under hypoxic conditions. Ion channel redox regulation has been described in detail for some ion channels, such as Kv channels or TRPC6. However, in general, information on ion channel redox regulation remains scant. Critical Issues and Future Directions: In addition to the debate of increased versus decreased ROS production during hypoxia, we aim here at describing and deciphering why different oxidants, under different conditions, can cause both activation and inhibition of channel activity. While the upstream pathways affecting channel gating are often well described, we need a better understanding of redox protein modifications to be able to determine the complexity of ion channel redox regulation. Against this background, we summarize the current knowledge on hypoxia-induced ROS-mediated ion channel signaling in the pulmonary circulation. Antioxid. Redox Signal. 22, 537–552 PMID:25545236

  3. Caenorhabditis elegans TRPV Channels Function in a Modality-Specific Pathway to Regulate Response to Aberrant Sensory Signaling

    PubMed Central

    Ezak , Meredith J.; Hong , Elizabeth; Chaparro-Garcia , Angela; Ferkey , Denise M.

    2010-01-01

    Olfaction and some forms of taste (including bitter) are mediated by G protein-coupled signal transduction pathways. Olfactory and gustatory ligands bind to chemosensory G protein-coupled receptors (GPCRs) in specialized sensory cells to activate intracellular signal transduction cascades. G protein-coupled receptor kinases (GRKs) are negative regulators of signaling that specifically phosphorylate activated GPCRs to terminate signaling. Although loss of GRK function usually results in enhanced cellular signaling, Caenorhabditis elegans lacking GRK-2 function are not hypersensitive to chemosensory stimuli. Instead, grk-2 mutant animals do not chemotax toward attractive olfactory stimuli or avoid aversive tastes and smells. We show here that loss-of-function mutations in the transient receptor potential vanilloid (TRPV) channels OSM-9 and OCR-2 selectively restore grk-2 behavioral avoidance of bitter tastants, revealing modality-specific mechanisms for TRPV channel function in the regulation of C. elegans chemosensation. Additionally, a single amino acid point mutation in OCR-2 that disrupts TRPV channel-mediated gene expression, but does not decrease channel function in chemosensory primary signal transduction, also restores grk-2 bitter taste avoidance. Thus, loss of GRK-2 function may lead to changes in gene expression, via OSM-9/OCR-2, to selectively alter the levels of signaling components that transduce or regulate bitter taste responses. Our results suggest a novel mechanism and multiple modality-specific pathways that sensory cells employ in response to aberrant signal transduction. PMID:20176974

  4. Subtype-specific control of P2X receptor channel signaling by ATP and Mg2+

    PubMed Central

    Li, Mufeng; Silberberg, Shai D.; Swartz, Kenton J.

    2013-01-01

    The identity and forms of activating ligands for ion channels are fundamental to their physiological roles in rapid electrical signaling. P2X receptor channels are ATP-activated cation channels that serve important roles in sensory signaling and inflammation, yet the active forms of the nucleotide are unknown. In physiological solutions, ATP is ionized and primarily found in complex with Mg2+. Here we investigated the active forms of ATP and found that the action of MgATP2− and ATP4− differs between subtypes of P2X receptors. The slowly desensitizing P2X2 receptor can be activated by free ATP, but MgATP2− promotes opening with very low efficacy. In contrast, both free ATP and MgATP2− robustly open the rapidly desensitizing P2X3 subtype. A further distinction between these two subtypes is the ability of Mg2+ to regulate P2X3 through a distinct allosteric mechanism. Importantly, heteromeric P2X2/3 channels present in sensory neurons exhibit a hybrid phenotype, characterized by robust activation by MgATP2− and weak regulation by Mg2+. These results reveal the existence of two classes of homomeric P2X receptors with differential sensitivity to MgATP2− and regulation by Mg2+, and demonstrate that both restraining mechanisms can be disengaged in heteromeric channels to form fast and sensitive ATP signaling pathways in sensory neurons. PMID:23959888

  5. Regionally specific expression of high-voltage-activated calcium channels in thalamic nuclei of epileptic and non-epileptic rats.

    PubMed

    Kanyshkova, Tatyana; Ehling, Petra; Cerina, Manuela; Meuth, Patrick; Zobeiri, Mehrnoush; Meuth, Sven G; Pape, Hans-Christian; Budde, Thomas

    2014-07-01

    The polygenic origin of generalized absence epilepsy results in dysfunction of ion channels that allows the switch from physiological asynchronous to pathophysiological highly synchronous network activity. Evidence from rat and mouse models of absence epilepsy indicates that altered Ca(2+) channel activity contributes to cellular and network alterations that lead to seizure activity. Under physiological circumstances, high voltage-activated (HVA) Ca(2+) channels are important in determining the thalamic firing profile. Here, we investigated a possible contribution of HVA channels to the epileptic phenotype using a rodent genetic model of absence epilepsy. In this study, HVA Ca(2+) currents were recorded from neurons of three different thalamic nuclei that are involved in both sensory signal transmission and rhythmic-synchronized activity during epileptic spike-and-wave discharges (SWD), namely the dorsal part of the lateral geniculate nucleus (dLGN), the ventrobasal thalamic complex (VB) and the reticular thalamic nucleus (NRT) of epileptic Wistar Albino Glaxo rats from Rijswijk (WAG/Rij) and non-epileptic August Copenhagen Irish (ACI) rats. HVA Ca(2+) current densities in dLGN neurons were significantly increased in epileptic rats compared with non-epileptic controls while other thalamic regions revealed no differences between the strains. Application of specific channel blockers revealed that the increased current was carried by L-type Ca(2+) channels. Electrophysiological evidence of increased L-type current correlated with up-regulated mRNA and protein expression of a particular L-type channel, namely Cav1.3, in dLGN of epileptic rats. No significant changes were found for other HVA Ca(2+) channels. Moreover, pharmacological inactivation of L-type Ca(2+) channels results in altered firing profiles of thalamocortical relay (TC) neurons from non-epileptic rather than from epileptic rats. While HVA Ca(2+) channels influence tonic and burst firing in ACI and WAG

  6. Cation channels of the transient receptor potential superfamily: their role in physiological and pathophysiological processes of smooth muscle cells.

    PubMed

    Dietrich, Alexander; Chubanov, Vladimir; Kalwa, Hermann; Rost, Benjamin R; Gudermann, Thomas

    2006-12-01

    Smooth muscle cells (SMC) are essential components of many tissues of the body. Ion channels regulate their membrane potential, the intracellular Ca(2+) concentration ([Ca(2+)](i)) and their contractility. Among the ion channels expressed in SMC cation channels of the transient receptor potential (TRP) superfamily allow the entry of Na(+), Ca(2+) and Mg(2+). Members of the TRP superfamily are essential constituents of tonically active channels (TAC), receptor-operated channels (ROC), store-operated channels (SOC) and stretch-activated channels (SAC). This review focusses on TRP channels (TRPC1, TRPC3, TRPC4, TRPC5, TRPC6, TRPC7, TRPV2, TRPV4, TRPM4, TRPM7, TRPP2) whose physiological functions in SMC were dissected by downregulating channel activity in isolated tissues or by the analysis of gene-deficient mouse models. Their possible functional role and physiological regulation as homomeric or heteromeric channels in SMC are discussed. Moreover, TRP channels may also be responsible for pathophysiological processes involving SMC-like airway hyperresponsiveness and pulmonary hypertension. Therefore, they present important drug targets for future pharmacological interventions. PMID:16842858

  7. A family of fluoride-specific ion channels with dual-topology architecture

    PubMed Central

    Stockbridge, Randy B; Robertson, Janice L; Kolmakova-Partensky, Ludmila; Miller, Christopher

    2013-01-01

    Fluoride ion, ubiquitous in soil, water, and marine environments, is a chronic threat to microorganisms. Many prokaryotes, archea, unicellular eukaryotes, and plants use a recently discovered family of F− exporter proteins to lower cytoplasmic F− levels to counteract the anion’s toxicity. We show here that these ‘Fluc’ proteins, purified and reconstituted in liposomes and planar phospholipid bilayers, form constitutively open anion channels with extreme selectivity for F− over Cl−. The active channel is a dimer of identical or homologous subunits arranged in antiparallel transmembrane orientation. This dual-topology assembly has not previously been seen in ion channels but is known in multidrug transporters of the SMR family, and is suggestive of an evolutionary antecedent of the inverted repeats found within the subunits of many membrane transport proteins. DOI: http://dx.doi.org/10.7554/eLife.01084.001 PMID:23991286

  8. Mitragynine and its potential blocking effects on specific cardiac potassium channels.

    PubMed

    Tay, Yea Lu; Teah, Yi Fan; Chong, Yoong Min; Jamil, Mohd Fadzly Amar; Kollert, Sina; Adenan, Mohd Ilham; Wahab, Habibah Abdul; Döring, Frank; Wischmeyer, Erhard; Tan, Mei Lan

    2016-08-15

    Mitragyna speciosa Korth is known for its euphoric properties and is frequently used for recreational purposes. Several poisoning and fatal cases involving mitragynine have been reported but the underlying causes remain unclear. Human ether-a-go-go-related gene (hERG) encodes the cardiac IKr current which is a determinant of the duration of ventricular action potentials and QT interval. On the other hand, IK1, a Kir current mediated by Kir2.1 channel and IKACh, a receptor-activated Kir current mediated by GIRK channel are also known to be important in maintaining the cardiac function. This study investigated the effects of mitragynine on the current, mRNA and protein expression of hERG channel in hERG-transfected HEK293 cells and Xenopus oocytes. The effects on Kir2.1 and GIRK channels currents were also determined in the oocytes. The hERG tail currents following depolarization pulses were inhibited by mitragynine with an IC50 value of 1.62μM and 1.15μM in the transfected cell line and Xenopus oocytes, respectively. The S6 point mutations of Y652A and F656A attenuated the inhibitor effects of mitragynine, indicating that mitragynine interacts with these high affinity drug-binding sites in the hERG channel pore cavity which was consistent with the molecular docking simulation. Interestingly, mitragynine does not affect the hERG expression at the transcriptional level but inhibits the protein expression. Mitragynine is also found to inhibit IKACh current with an IC50 value of 3.32μM but has no significant effects on IK1. Blocking of both hERG and GIRK channels may cause additive cardiotoxicity risks. PMID:27260674

  9. A designer ligand specific for Kv1.3 channels from a scorpion neurotoxin-based library

    PubMed Central

    Takacs, Zoltan; Toups, Megan; Kollewe, Astrid; Johnson, Erik; Cuello, Luis G.; Driessens, Gregory; Biancalana, Matthew; Koide, Akiko; Ponte, Cristiano G.; Perozo, Eduardo; Gajewski, Thomas F.; Suarez-Kurtz, Guilherme; Koide, Shohei; Goldstein, Steve A. N.

    2009-01-01

    Venomous animals immobilize prey using protein toxins that act on ion channels and other targets of biological importance. Broad use of toxins for biomedical research, diagnosis, and therapy has been limited by inadequate target discrimination, for example, among ion channel subtypes. Here, a synthetic toxin is produced by a new strategy to be specific for human Kv1.3 channels, critical regulators of immune T cells. A phage display library of 11,200 de novo proteins is designed using the α-KTx scaffold of 31 scorpion toxin sequences known or predicted to bind to potassium channels. Mokatoxin-1 (moka1) is isolated by affinity selection on purified target. Moka1 blocks Kv1.3 at nanomolar levels that do not inhibit Kv1.1, Kv1.2, or KCa1.1. As a result, moka1 suppresses CD3/28-induced cytokine secretion by T cells without cross-reactive gastrointestinal hyperactivity. The 3D structure of moka1 rationalizes its specificity and validates the engineering approach, revealing a unique interaction surface supported on an α-KTx scaffold. This scaffold-based/target-biased strategy overcomes many obstacles to production of selective toxins. PMID:20007782

  10. Specific inhibition of long-lasting, L-type calcium channels by synthetic parathyroid hormone

    SciTech Connect

    Pang, P.K.T.; Wang, R.; Shan, J.; Karpinski, E.; Benishin, C.G. )

    1990-01-01

    The effect of an active synthetic N-terminal fragment of bovine parathyroid hormone (bPTH), bPTH-(1-34), on Ca{sup 2+} channels was studied in mouse neuroblastoma cells (N1E-115). With the whole-cell variation of the patch-clamp technique, T (transient) and L (long-lasting) types of Ca{sup 2+} currents were identified. Pharmacological characterization showed that the L current was amplified by the Ca{sup 2+} channel stimulator BAY K-8644, but the T current was unaffected. The administration of bPTH-(1-34) produced dose-related inhibition of the L current, which could be reversed by BAY K-8644. The peptide had no effect on the T current. In addition, use of the fluorescent indicator fura-2 showed that bPTH-(1-34) inhibited the KCl-stimulated increase in intracellular free Ca{sup 2+} in neuroblastoma cells with L channels but not in cells with T channels. An inactivated (oxidized) preparation of bPTH-(1-34) failed to affect the L current. High-affinity binding of labeled PTH analog to these neuroblastoma cells was also demonstrated. In addition, bPTH-(1-34) inhibited the L current in cultured vascular smooth muscle cells from rat tail artery. These data indicate that, in some tissues PTH can act as an endogenous blocker of Ca{sup 2+} entry.

  11. Death of Neurons following Injury Requires Conductive Neuronal Gap Junction Channels but Not a Specific Connexin.

    PubMed

    Fontes, Joseph D; Ramsey, Jon; Polk, Jeremy M; Koop, Andre; Denisova, Janna V; Belousov, Andrei B

    2015-01-01

    Pharmacological blockade or genetic knockout of neuronal connexin 36 (Cx36)-containing gap junctions reduces neuronal death caused by ischemia, traumatic brain injury and NMDA receptor (NMDAR)-mediated excitotoxicity. However, whether Cx36 gap junctions contribute to neuronal death via channel-dependent or channel-independent mechanism remains an open question. To address this, we manipulated connexin protein expression via lentiviral transduction of mouse neuronal cortical cultures and analyzed neuronal death twenty-four hours following administration of NMDA (a model of NMDAR excitotoxicity) or oxygen-glucose deprivation (a model of ischemic injury). In cultures prepared from wild-type mice, over-expression and knockdown of Cx36-containing gap junctions augmented and prevented, respectively, neuronal death from NMDAR-mediated excitotoxicity and ischemia. In cultures obtained form from Cx36 knockout mice, re-expression of functional gap junction channels, containing either neuronal Cx36 or non-neuronal Cx43 or Cx31, resulted in increased neuronal death following insult. In contrast, the expression of communication-deficient gap junctions (containing mutated connexins) did not have this effect. Finally, the absence of ethidium bromide uptake in non-transduced wild-type neurons two hours following NMDAR excitotoxicity or ischemia suggested the absence of active endogenous hemichannels in those neurons. Taken together, these results suggest a role for neuronal gap junctions in cell death via a connexin type-independent mechanism that likely relies on channel activities of gap junctional complexes among neurons. A possible contribution of gap junction channel-permeable death signals in neuronal death is discussed. PMID:26017008

  12. Death of Neurons following Injury Requires Conductive Neuronal Gap Junction Channels but Not a Specific Connexin

    PubMed Central

    Fontes, Joseph D.; Ramsey, Jon; Polk, Jeremy M; Koop, Andre; Denisova, Janna V.; Belousov, Andrei B.

    2015-01-01

    Pharmacological blockade or genetic knockout of neuronal connexin 36 (Cx36)-containing gap junctions reduces neuronal death caused by ischemia, traumatic brain injury and NMDA receptor (NMDAR)-mediated excitotoxicity. However, whether Cx36 gap junctions contribute to neuronal death via channel-dependent or channel-independent mechanism remains an open question. To address this, we manipulated connexin protein expression via lentiviral transduction of mouse neuronal cortical cultures and analyzed neuronal death twenty-four hours following administration of NMDA (a model of NMDAR excitotoxicity) or oxygen-glucose deprivation (a model of ischemic injury). In cultures prepared from wild-type mice, over-expression and knockdown of Cx36-containing gap junctions augmented and prevented, respectively, neuronal death from NMDAR-mediated excitotoxicity and ischemia. In cultures obtained form from Cx36 knockout mice, re-expression of functional gap junction channels, containing either neuronal Cx36 or non-neuronal Cx43 or Cx31, resulted in increased neuronal death following insult. In contrast, the expression of communication-deficient gap junctions (containing mutated connexins) did not have this effect. Finally, the absence of ethidium bromide uptake in non-transduced wild-type neurons two hours following NMDAR excitotoxicity or ischemia suggested the absence of active endogenous hemichannels in those neurons. Taken together, these results suggest a role for neuronal gap junctions in cell death via a connexin type-independent mechanism that likely relies on channel activities of gap junctional complexes among neurons. A possible contribution of gap junction channel-permeable death signals in neuronal death is discussed. PMID:26017008

  13. D2 dopamine receptor activation of potassium channels is selectively decoupled by Galpha-specific GoLoco motif peptides.

    PubMed

    Webb, Christina K; McCudden, Christopher R; Willard, Francis S; Kimple, Randall J; Siderovski, David P; Oxford, Gerry S

    2005-03-01

    The GoLoco motif is a short polypeptide sequence found in G-protein signaling regulators such as regulator of G-protein signaling proteins type 12 and 14 and activator of G-protein signaling protein type 3. A unique property of the GoLoco motifs from these three proteins is their preferential interaction with guanosine diphosphate (GDP)-bound Galpha(i1), Galpha(i3) and, sometimes, Galpha(i2) subunits over Galpha(o) subunits. This interaction prevents both spontaneous guanine nucleotide release and reassociation of Galpha(i)-GDP with Gbetagamma. We utilized this property of the GoLoco motif to examine dopamine (D2 and D3) and somatostatin receptor coupling to G-protein-regulated inwardly rectifying potassium (GIRK) channels in mouse AtT20 cells. GoLoco motif peptides had no effect on either basal channel activity or the initial responses to agonists, suggesting that the GoLoco motif cannot disrupt pre-formed G-protein heterotrimers. GoLoco motif peptides did, however, interfere with human D2((short)) receptor coupling to GIRK channels as demonstrated by the progressively diminished responses after repeated agonist application. This behavior is consistent with some form of compartmentalization of D2 receptors and GIRK channels such that Gbetagamma subunits, freed by local receptor activation and prevented from reforming a heterotrimeric complex, are not functionally constrained within the receptor-channel complex and thus are unable to exert a persistent activating effect. In contrast, GoLoco motif peptides had no effect on either D3 or somatostatin coupling to GIRK channels. Our results suggest that GoLoco motif-based peptides will be useful tools in examining the specificity of G-protein-coupled receptor-effector coupling. PMID:15748159

  14. Diffusion of nitric oxide across cell membranes of the vascular wall requires specific connexin-based channels.

    PubMed

    Figueroa, Xavier F; Lillo, Mauricio A; Gaete, Pablo S; Riquelme, Manuel A; Sáez, Juan C

    2013-12-01

    NO is generated within cells and frequently must be transferred to responsive neighboring cells, as occurs in the endothelium-dependent relaxation of smooth muscle cells observed in blood vessels. It is thought that NO diffuses freely across cell membranes, but it may also permeate through low resistant membrane pathways. Here, we describe the participation of connexin (Cx)-formed channels in the NO transport across cell membranes and between endothelial and smooth muscle cells. We used a water-soluble NO donor of high molecular weight (S-nitrosylated albumin, BSA-NO) that does not permeate through cell membranes or Cx-based channels and the NO-sensitive dye 4,5-diaminofluorescein diacetate to detect changes of intracellular NO concentration. We found that NO generated in the extracellular space was not detected intracellularly in Cx-deficient HeLa cells, suggesting that cell membrane represents a significant diffusion barrier for NO transfer. However, Cx-based channels provide efficient pathways for NO signaling because NO opened and permeated hemichannels expressed in HeLa cells transfected with Cx43, Cx40, or Cx37. In contrast, NO closed hemichannels of HeLa-Cx32 cells, which otherwise are permeable to NO if are opened by a divalent cation-free extracellular solution. Consistent with this, blockade of Cx-based channels abolished the myoendothelial NO transfer and associated NO-dependent vasodilation induced by acethylcholine. These results indicate that Cx-based channels play a key role in the NO-dependent tonic control of vascular function and may direct the NO signal to specific targets, which provides a novel mechanistic basis for the critical role of Cxs in cell-cell communication in the vessel wall. This article is part of the Special Issue Section entitled 'Current Pharmacology of Gap Junction Channels and Hemichannels'. PMID:23499665

  15. Inhibition of diacylglycerol-sensitive TRPC channels by synthetic and natural steroids.

    PubMed

    Miehe, Susanne; Crause, Peter; Schmidt, Thorsten; Löhn, Matthias; Kleemann, Heinz-Werner; Licher, Thomas; Dittrich, Werner; Rütten, Hartmut; Strübing, Carsten

    2012-01-01

    TRPC channels are a family of nonselective cation channels that regulate ion homeostasis and intracellular Ca(2+) signaling in numerous cell types. Important physiological functions such as vasoregulation, neuronal growth, and pheromone recognition have been assigned to this class of ion channels. Despite their physiological relevance, few selective pharmacological tools are available to study TRPC channel function. We, therefore, screened a selection of pharmacologically active compounds for TRPC modulating activity. We found that the synthetic gestagen norgestimate inhibited diacylglycerol-sensitive TRPC3 and TRPC6 with IC(50)s of 3-5 µM, while half-maximal inhibition of TRPC5 required significantly higher compound concentrations (>10 µM). Norgestimate blocked TRPC-mediated vasopressin-induced cation currents in A7r5 smooth muscle cells and caused vasorelaxation of isolated rat aorta, indicating that norgestimate could be an interesting tool for the investigation of TRP channel function in native cells and tissues. The steroid hormone progesterone, which is structurally related to norgestimate, also inhibited TRPC channel activity with IC(50)s ranging from 6 to 18 µM but showed little subtype selectivity. Thus, TRPC channel inhibition by high gestational levels of progesterone may contribute to the physiological decrease of uterine contractility and immunosuppression during pregnancy. PMID:22530015

  16. Variants in Ion Channel Genes Link Phenotypic Features of Bipolar Illness to Specific Neurobiological Process Domains

    PubMed Central

    Balaraman, Yokesh; Lahiri, Debomoy K.; Nurnberger, John I.

    2015-01-01

    Recent advances in genome-wide association studies are pointing towards a major role for voltage-gated ion channels in neuropsychiatric disorders and, in particular, bipolar disorder (BD). The phenotype of BD is complex, with symptoms during mood episodes and deficits persisting between episodes. We have tried to elucidate the common neurobiological mechanisms associated with ion channel signaling in order to provide a new perspective on the clinical symptoms and possible endophenotypes seen in BD patients. We propose a model in which the multiple variants in genes coding for ion channel proteins would perturb motivational circuits, synaptic plasticity, myelination, hypothalamic-pituitary-adrenal axis function, circadian neuronal rhythms, and energy regulation. These changes in neurobiological mechanisms would manifest in endophenotypes of aberrant reward processing, white matter hyperintensities, deficits in executive function, altered frontolimbic connectivity, increased amygdala activity, increased melatonin suppression, decreased REM latency, and aberrant myo-inositol/ATP shuttling. The endophenotypes result in behaviors of poor impulse control, motivational changes, cognitive deficits, abnormal stress response, sleep disturbances, and energy changes involving different neurobiological process domains. The hypothesis is that these disturbances start with altered neural circuitry during development, following which multiple environmental triggers may disrupt the neuronal excitability balance through an activity-dependent molecular process, resulting in clinical mood episodes.

  17. Subject-Specific Effect of Metallic Body Accessories on Path Loss of Dynamic on-Body Propagation Channels.

    PubMed

    Rahim, H A; Abdulmalek, M; Soh, P J; Rani, K A; Hisham, N; Vandenbosch, G A E

    2016-01-01

    This paper presents the investigation of path loss variation for subject-specific on-body radio propagation channels, considering the effect of metallic spectacles and loop like metallic accessories. Adding metallic items may affect the operability of Body Centric Wireless Communications (BCWC). Measurements were carried out in an RF-shielded room lined with microwave absorbing sheets for strategically placed bodyworn antennas covering the upper front torso and the lower limbs. The path loss of the on-body radio channel was characterized explicitly taking into account the body size of the subjects. For metallic loop-like accessories, the results indicate that for underweight subjects, there was a slightly higher influence, up to 2%, compared to normal and overweight subjects. Our findings indicate that a noticeable effect exists on on-body channels for dynamic movements where the metallic watch acts as a local scatterer that affects the non-line-of-sight (NLOS) signal path between transmitter and receiver for underweight subjects in comparison to normal and overweight subjects. The path loss decreases when the receiving terminal was positioned very close to the metallic item. If a loop-like metallic accessory is not appropriately considered when designing the radio channel on a subject, the reliability of the body-centric wireless system may degrade. PMID:27436496

  18. Subject-Specific Effect of Metallic Body Accessories on Path Loss of Dynamic on-Body Propagation Channels

    NASA Astrophysics Data System (ADS)

    Rahim, H. A.; Abdulmalek, M.; Soh, P. J.; Rani, K. A.; Hisham, N.; Vandenbosch, G. A. E.

    2016-07-01

    This paper presents the investigation of path loss variation for subject-specific on-body radio propagation channels, considering the effect of metallic spectacles and loop like metallic accessories. Adding metallic items may affect the operability of Body Centric Wireless Communications (BCWC). Measurements were carried out in an RF-shielded room lined with microwave absorbing sheets for strategically placed bodyworn antennas covering the upper front torso and the lower limbs. The path loss of the on-body radio channel was characterized explicitly taking into account the body size of the subjects. For metallic loop-like accessories, the results indicate that for underweight subjects, there was a slightly higher influence, up to 2%, compared to normal and overweight subjects. Our findings indicate that a noticeable effect exists on on-body channels for dynamic movements where the metallic watch acts as a local scatterer that affects the non-line-of-sight (NLOS) signal path between transmitter and receiver for underweight subjects in comparison to normal and overweight subjects. The path loss decreases when the receiving terminal was positioned very close to the metallic item. If a loop-like metallic accessory is not appropriately considered when designing the radio channel on a subject, the reliability of the body-centric wireless system may degrade.

  19. Subject-Specific Effect of Metallic Body Accessories on Path Loss of Dynamic on-Body Propagation Channels

    PubMed Central

    Rahim, H. A.; Abdulmalek, M.; Soh, P. J.; Rani, K. A.; Hisham, N.; Vandenbosch, G. A. E.

    2016-01-01

    This paper presents the investigation of path loss variation for subject-specific on-body radio propagation channels, considering the effect of metallic spectacles and loop like metallic accessories. Adding metallic items may affect the operability of Body Centric Wireless Communications (BCWC). Measurements were carried out in an RF-shielded room lined with microwave absorbing sheets for strategically placed bodyworn antennas covering the upper front torso and the lower limbs. The path loss of the on-body radio channel was characterized explicitly taking into account the body size of the subjects. For metallic loop-like accessories, the results indicate that for underweight subjects, there was a slightly higher influence, up to 2%, compared to normal and overweight subjects. Our findings indicate that a noticeable effect exists on on-body channels for dynamic movements where the metallic watch acts as a local scatterer that affects the non-line-of-sight (NLOS) signal path between transmitter and receiver for underweight subjects in comparison to normal and overweight subjects. The path loss decreases when the receiving terminal was positioned very close to the metallic item. If a loop-like metallic accessory is not appropriately considered when designing the radio channel on a subject, the reliability of the body-centric wireless system may degrade. PMID:27436496

  20. Bioinspired Protein Channel-Based Scanning Ion Conductance Microscopy (Bio-SICM) for Simultaneous Conductance and Specific Molecular Imaging.

    PubMed

    Macazo, Florika C; White, Ryan J

    2016-03-01

    The utility of stochastic single-molecule detection using protein nanopores has found widespread application in bioanalytical sensing as a result of the inherent signal amplification of the resistive pulse method. Integration of protein nanopores with high-resolution scanning ion conductance microscopy (SICM) extends the utility of SICM by enabling selective chemical imaging of specific target molecules, while simultaneously providing topographical information about the net ion flux through a pore under a concentration gradient. In this study, we describe the development of a bioinspired scanning ion conductance microscopy (bio-SICM) approach that couples the imaging ability of SICM with the sensitivity and chemical selectivity of protein channels to perform simultaneous pore imaging and specific molecule mapping. To establish the framework of the bio-SICM platform, we utilize the well-studied protein channel α-hemolysin (αHL) to map the presence of β-cyclodextrin (βCD) at a substrate pore opening. We demonstrate concurrent pore and specific molecule imaging by raster scanning an αHL-based probe over a glass membrane containing a single 25-μm-diameter glass pore while recording the lateral positions of the probe and channel activity via ionic current. We use the average channel current to create a conductance image and the raw current-time traces to determine spatial localization of βCD. With further optimization, we believe that the bio-SICM platform will provide a powerful analytical methodology that is generalizable, and thus offers significant utility in a myriad of bioanalytical applications. PMID:26848947

  1. Electrophysiological evidence for the broad distribution of specific odorant receptor molecules across the olfactory organ of the channel catfish.

    PubMed

    Chang, Q; Caprio, J

    1996-10-01

    To determine if there is a spatial segregation of responsiveness to odorants within the olfactory epithelium, microelectrode recordings were obtained from small populations of olfactory receptor neurons located across different lamellar sensory regions of the olfactory organ of the channel catfish, lctalurus punctatus. Stimuli included L-alanine, L-methionine, L-arginine hydrochloride, L-glutamic acid, ATP and a mixture of bile salts-odorants previously reported to stimulate independent receptor sites in aquatic species. The peak integrated olfactory receptor responses at each recording site were standardized to the response to L-alanine. The relative stimulatory effectiveness of the stimuli was preserved across the 10 olfactory lamellae recording sites. These data support previous molecular biological results of a broad distribution of receptor neurons that express specific receptor genes across the olfactory organ of the channel catfish. PMID:8902281

  2. Subunit-specific inhibition of acid sensing ion channels by stomatin-like protein 1

    PubMed Central

    Kozlenkov, Alexey; Lapatsina, Liudmila; Lewin, Gary R; Smith, Ewan St John

    2014-01-01

    There are five mammalian stomatin-domain genes, all of which encode peripheral membrane proteins that can modulate ion channel function. Here we examined the ability of stomatin-like protein 1 (STOML1) to modulate the proton-sensitive members of the acid-sensing ion channel (ASIC) family. STOML1 profoundly inhibits ASIC1a, but has no effect on the splice variant ASIC1b. The inactivation time constant of ASIC3 is also accelerated by STOML1. We examined STOML1 null mutant mice with a β-galactosidase-neomycin cassette gene-trap reporter driven from the STOML1 gene locus, which indicated that STOML1 is expressed in at least 50% of dorsal root ganglion (DRG) neurones. Patch clamp recordings from mouse DRG neurones identified a trend for larger proton-gated currents in neurones lacking STOML1, which was due to a contribution of effects upon both transient and sustained currents, at different pH, a finding consistent with an endogenous inhibitory function for STOML1. PMID:24247984

  3. Simulation-based scenario-specific channel modeling for WBAN cooperative transmission schemes.

    PubMed

    Naganawa, Jun-ichi; Wangchuk, Karma; Kim, Minseok; Aoyagi, Takahiro; Takada, Jun-ichi

    2015-03-01

    Wireless body area networks (WBANs) are an emerging technology for realizing efficient healthcare and remote medicine for the aging society of the future. In order to improve the reliability of WBAN systems and support its various applications, channel modeling and performance evaluation are important. This paper proposes a simulation-based channel modeling for evaluating the performance of WBAN cooperative transmission schemes. The time series of path losses among seven on-body nodes are generated by the finite-difference time-domain method for seven body motions. The statistical parameters of the path loss for all the motions are also obtained. The generated path loss is then applied to the evaluation of the two-hop decode-and-forward relaying scheme, yielding an improvement in transmit power. From the evaluation of body motion, useful insights are obtained such as which relay links are more robust than others. Finally, the proposed approach is validated through comparison with a measurement-based approach. PMID:24876134

  4. Calcium binding to calmodulin mutants having domain-specific effects on the regulation of ion channels.

    PubMed

    VanScyoc, Wendy S; Newman, Rhonda A; Sorensen, Brenda R; Shea, Madeline A

    2006-12-01

    Calmodulin (CaM) is an essential, eukaryotic protein comprised of two highly homologous domains (N and C). CaM binds four calcium ions cooperatively, regulating a wide array of target proteins. A genetic screen of Paramecia by Kung [Kung, C. et al. (1992) Cell Calcium 13, 413-425] demonstrated that the domains of CaM have separable physiological roles: "under-reactive" mutations affecting calcium-dependent sodium currents mapped to the N-domain, while "over-reactive" mutations affecting calcium-dependent potassium currents localized to the C-domain of CaM. To determine whether and how these mutations affected intrinsic calcium-binding properties of CaM domains, phenylalanine fluorescence was used to monitor calcium binding to sites I and II (N-domain) and tyrosine fluorescence was used to monitor sites III and IV (C-domain). To explore interdomain interactions, binding properties of each full-length mutant were compared to those of its corresponding domain fragments. The calcium-binding properties of six under-reactive mutants (V35I/D50N, G40E, G40E/D50N, D50G, E54K, and G59S) and one over-reactive mutant (M145V) were indistinguishable from those of wild-type CaM, despite their deleterious physiological effects on ion-channel regulation. Four over-reactive mutants (D95G, S101F, E104K, and H135R) significantly decreased the calcium affinity of the C-domain. Of these, one (E104K) also increased the calcium affinity of the N-domain, demonstrating that the magnitude and direction of wild-type interdomain coupling had been perturbed. This suggests that, while some of these mutations alter calcium-binding directly, others probably alter CaM-channel association or calcium-triggered conformational change in the context of a ternary complex with the affected ion channel. PMID:17128970

  5. In Silico Structure and Sequence Analysis of Bacterial Porins and Specific Diffusion Channels for Hydrophilic Molecules: Conservation, Multimericity and Multifunctionality.

    PubMed

    Vollan, Hilde S; Tannæs, Tone; Vriend, Gert; Bukholm, Geir

    2016-01-01

    Diffusion channels are involved in the selective uptake of nutrients and form the largest outer membrane protein (OMP) family in Gram-negative bacteria. Differences in pore size and amino acid composition contribute to the specificity. Structure-based multiple sequence alignments shed light on the structure-function relations for all eight subclasses. Entropy-variability analysis results are correlated to known structural and functional aspects, such as structural integrity, multimericity, specificity and biological niche adaptation. The high mutation rate in their surface-exposed loops is likely an important mechanism for host immune system evasion. Multiple sequence alignments for each subclass revealed conserved residue positions that are involved in substrate recognition and specificity. An analysis of monomeric protein channels revealed particular sequence patterns of amino acids that were observed in other classes at multimeric interfaces. This adds to the emerging evidence that all members of the family exist in a multimeric state. Our findings are important for understanding the role of members of this family in a wide range of bacterial processes, including bacterial food uptake, survival and adaptation mechanisms. PMID:27110766

  6. In Silico Structure and Sequence Analysis of Bacterial Porins and Specific Diffusion Channels for Hydrophilic Molecules: Conservation, Multimericity and Multifunctionality

    PubMed Central

    Vollan, Hilde S.; Tannæs, Tone; Vriend, Gert; Bukholm, Geir

    2016-01-01

    Diffusion channels are involved in the selective uptake of nutrients and form the largest outer membrane protein (OMP) family in Gram-negative bacteria. Differences in pore size and amino acid composition contribute to the specificity. Structure-based multiple sequence alignments shed light on the structure-function relations for all eight subclasses. Entropy-variability analysis results are correlated to known structural and functional aspects, such as structural integrity, multimericity, specificity and biological niche adaptation. The high mutation rate in their surface-exposed loops is likely an important mechanism for host immune system evasion. Multiple sequence alignments for each subclass revealed conserved residue positions that are involved in substrate recognition and specificity. An analysis of monomeric protein channels revealed particular sequence patterns of amino acids that were observed in other classes at multimeric interfaces. This adds to the emerging evidence that all members of the family exist in a multimeric state. Our findings are important for understanding the role of members of this family in a wide range of bacterial processes, including bacterial food uptake, survival and adaptation mechanisms. PMID:27110766

  7. Purification of charybdotoxine, a specific inhibitor of the high-conductance Ca/sup 2 +/-activated K/sup +/ channel

    SciTech Connect

    Smith, C.; Phillips, M.; Miller, C.

    1986-11-05

    Charybdotoxim is a high-affinity specific inhibitor of the high-conductance Ca/sup 2 +/-activated K/sup +/ channel found in the plasma membranes of many vertebrate cell types. Using Ca/sup 2 +/-activated K/sup +/ channels reconstituted into planar lipid bilayer membranes as an assay, the authors have purified the toxin from the venom of the scorpion Leiurus quinquestriatus by a two-step procedure involving chromatofocusing on SP-Sephadex, followed by reversed-phase high-performance liquid chromatography. Charybdotoxin is shown to be a highly basic protein with a mass of 10 kDa. Under the standard assay conditions, the purified toxin inhibits the Ca/sup 2 +/-activated K/sup +/ channel with an apparent dissociation constant of 3.5 nM. The protein is unusually stable, with inhibitory potency being insensitive to boiling or exposure to organic solvents. The toxin's activity is sensitive to chymotrypsin treatment and to acylation of lysine groups. The protein may be radioiodinated without loss of activity.

  8. Patch-Clamp Study of Hepatitis C p7 Channels Reveals Genotype-Specific Sensitivity to Inhibitors.

    PubMed

    Breitinger, Ulrike; Farag, Noha S; Ali, Nourhan K M; Breitinger, Hans-Georg A

    2016-06-01

    Hepatitis C is a major worldwide disease and health hazard, affecting ∼3% of the world population. The p7 protein of hepatitis C virus (HCV) is an intracellular ion channel and pH regulator that is involved in the viral replication cycle. It is targeted by various classical ion channel blockers. Here, we generated p7 constructs corresponding to HCV genotypes 1a, 2a, 3a, and 4a for recombinant expression in HEK293 cells, and studied p7 channels using patch-clamp recording techniques. The pH50 values for recombinant p7 channels were between 6.0 and 6.5, as expected for proton-activated channels, and current-voltage dependence did not show any differences between genotypes. Inhibition of p7-mediated currents by amantadine, however, exhibited significant, genotype-specific variation. The IC50 values of p7-1a and p7-4a were 0.7 ± 0.1 nM and 3.2 ± 1.2 nM, whereas p7-2a and p7-3a had 50- to 1000-fold lower sensitivity, with IC50 values of 2402 ± 334 nM and 344 ± 64 nM, respectively. The IC50 values for rimantadine were low across all genotypes, ranging from 0.7 ± 0.1 nM, 1.6 ± 0.6 nM, and 3.0 ± 0.8 nM for p7-1a, p7-3a, and p7-4a, respectively, to 24 ± 4 nM for p7-2a. Results from patch-clamp recordings agreed well with cellular assays of p7 activity, namely, measurements of intracellular pH and hemadsorption assays, which confirmed the much reduced amantadine sensitivity of genotypes 2a and 3a. Thus, our results establish patch-clamp studies of recombinant viroporins as a valid analytical tool that can provide quantitative information about viroporin channel properties, complementing established techniques. PMID:27276260

  9. The Sarcoglycan complex is expressed in the cerebrovascular system and is specifically regulated by astroglial Cx30 channels.

    PubMed

    Boulay, Anne-Cécile; Saubaméa, Bruno; Cisternino, Salvatore; Mignon, Virginie; Mazeraud, Aurélien; Jourdren, Laurent; Blugeon, Corinne; Cohen-Salmon, Martine

    2015-01-01

    Astrocytes, the most prominent glial cell type in the brain, send specialized processes called endfeet, around blood vessels and express a large molecular repertoire regulating the cerebrovascular system physiology. One of the most striking properties of astrocyte endfeet is their enrichment in gap junction proteins Connexin 43 and 30 (Cx43 and Cx30) allowing in particular for direct intercellular trafficking of ions and small signaling molecules through perivascular astroglial networks. In this study, we addressed the specific role of Cx30 at the gliovascular interface. Using an inactivation mouse model for Cx30 (Cx30(Δ/Δ); Δ means deleted allele) we showed that absence of Cx30 does not affect blood-brain barrier (BBB) organization and permeability. However, it results in the cerebrovascular fraction, in a strong upregulation of Sgcg encoding γ-Sarcoglycan (γ-SG), a member of the Dystrophin-associated protein complex (DAPC) connecting cytoskeleton and the extracellular matrix. The same molecular event occurs in Cx30(T5M/T5M) mutated mice, where Cx30 channels are closed, demonstrating that Sgcg regulation relied on Cx30 channel functions. We further characterized the expression of other Sarcoglycan complex (SGC) molecules in the cerebrovascular system and showed the presence of α-, β-, δ-, γ-, ε- and ζ- SG, as well as Sarcospan. Their expression was however not modified in Cx30(Δ/Δ). These results suggest that a full SGC might be present in the cerebrovascular system, and that expression of one of its member, γ-SG, depends on Cx30 channels. As described in skeletal muscles, the SGC may contribute to membrane stabilization and signal transduction in the cerebrovascular system, which may therefore be regulated by Cx30 channel-mediated functions. PMID:25698924

  10. The Sarcoglycan complex is expressed in the cerebrovascular system and is specifically regulated by astroglial Cx30 channels

    PubMed Central

    Boulay, Anne-Cécile; Saubaméa, Bruno; Cisternino, Salvatore; Mignon, Virginie; Mazeraud, Aurélien; Jourdren, Laurent; Blugeon, Corinne; Cohen-Salmon, Martine

    2015-01-01

    Astrocytes, the most prominent glial cell type in the brain, send specialized processes called endfeet, around blood vessels and express a large molecular repertoire regulating the cerebrovascular system physiology. One of the most striking properties of astrocyte endfeet is their enrichment in gap junction proteins Connexin 43 and 30 (Cx43 and Cx30) allowing in particular for direct intercellular trafficking of ions and small signaling molecules through perivascular astroglial networks. In this study, we addressed the specific role of Cx30 at the gliovascular interface. Using an inactivation mouse model for Cx30 (Cx30Δ/Δ; Δ means deleted allele) we showed that absence of Cx30 does not affect blood-brain barrier (BBB) organization and permeability. However, it results in the cerebrovascular fraction, in a strong upregulation of Sgcg encoding γ-Sarcoglycan (γ-SG), a member of the Dystrophin-associated protein complex (DAPC) connecting cytoskeleton and the extracellular matrix. The same molecular event occurs in Cx30T5M/T5M mutated mice, where Cx30 channels are closed, demonstrating that Sgcg regulation relied on Cx30 channel functions. We further characterized the expression of other Sarcoglycan complex (SGC) molecules in the cerebrovascular system and showed the presence of α-, β-, δ-, γ-, ε- and ζ- SG, as well as Sarcospan. Their expression was however not modified in Cx30Δ/Δ. These results suggest that a full SGC might be present in the cerebrovascular system, and that expression of one of its member, γ-SG, depends on Cx30 channels. As described in skeletal muscles, the SGC may contribute to membrane stabilization and signal transduction in the cerebrovascular system, which may therefore be regulated by Cx30 channel-mediated functions. PMID:25698924

  11. Channel specific rate constants relevant to the thermal decomposition of disilane.

    SciTech Connect

    Matsumoto, Keiji; Klippenstein, Stephen J.; Koshi, Mitsuo; Tonokura, Kenichi

    2005-01-01

    Rate constants for the thermal dissociation of Si{sub 2}H{sub 6} are predicted with a novel transition state model. The saddle points for dissociation on the Si{sub 2}H{sub 6} potential energy surface are lower in energy than the corresponding separated products, as confirmed by high level ab initio quantum mechanical calculations. Thus, the dissociations of Si{sub 2}H{sub 6} to produce SiH{sub 2} + SiH{sub 4} (R1) and H{sub 3}SiSiH + H{sub 2} (R2) both proceed through tight inner transition states followed by loose outer transition states. The present 'dual' transition state model couples variational phase space theory treatments of the outer transition states with ab initio based fixed harmonic vibrator treatments of the inner transition states to obtain effective numbers of states for the two transition states acting in series. It is found that, at least near room temperature, such a dual transition state model is generally required for the proper description of each of the dissociations. Only at quite high temperatures, i.e., above 2000 K for (R1) and 600 K for (R2), does a single fixed inner transition state provide an adequate description. Similarly, only at quite low temperatures (below 100 and 10 K for (R1) and (R2), respectively) does a single outer transition state provide an adequate description. Pressure dependent rate constants are obtained from solutions to the multichannel master equation. These calculations confirm that dissociation channel (R2) is negligible under conditions relevant to the thermal chemical vapor deposition (CVD) processes. Rate constants for the chemical activation reactions, SiH{sub 2} + SiH{sub 4} {yields} Si{sub 2}H{sub 6} (R-1) and SiH{sub 2} + SiH{sub 4} {yields} H{sub 3}SiSiH + H{sub 2} (R3), are also evaluated within the dual transition state model. It is found that reaction R3 is the dominant channel for low pressures and high temperatures, i.e., below 100 Torr for temperatures above 1100 K.

  12. Channel specific rate constants relevant to the thermal decomposition of disilane.

    PubMed

    Matsumoto, Keiji; Klippenstein, Stephen J; Tonokura, Kenichi; Koshi, Mitsuo

    2005-06-01

    Rate constants for the thermal dissociation of Si2H6 are predicted with a novel transition state model. The saddle points for dissociation on the Si2H6 potential energy surface are lower in energy than the corresponding separated products, as confirmed by high level ab initio quantum mechanical calculations. Thus, the dissociations of Si2H6 to produce SiH2 + SiH4 (R1) and H3SiSiH + H2 (R2) both proceed through tight inner transition states followed by loose outer transition states. The present "dual" transition state model couples variational phase space theory treatments of the outer transition states with ab initio based fixed harmonic vibrator treatments of the inner transition states to obtain effective numbers of states for the two transition states acting in series. It is found that, at least near room temperature, such a dual transition state model is generally required for the proper description of each of the dissociations. Only at quite high temperatures, i.e., above 2000 K for (R1) and 600 K for (R2), does a single fixed inner transition state provide an adequate description. Similarly, only at quite low temperatures (below 100 and 10 K for (R1) and (R2), respectively) does a single outer transition state provide an adequate description. Pressure dependent rate constants are obtained from solutions to the multichannel master equation. These calculations confirm that dissociation channel (R2) is negligible under conditions relevant to the thermal chemical vapor deposition (CVD) processes. Rate constants for the chemical activation reactions, SiH2 + SiH4 --> Si2H6 (R-1) and SiH2 + SiH4 --> H3SiSiH + H2 (R3), are also evaluated within the dual transition state model. It is found that reaction R3 is the dominant channel for low pressures and high temperatures, i.e., below 100 Torr for temperatures above 1100 K. PMID:16833838

  13. Highly specific olfactory receptor neurons for types of amino acids in the channel catfish.

    PubMed

    Nikonov, Alexander A; Caprio, John

    2007-10-01

    Odorant specificity to l-alpha-amino acids was determined electrophysiologically for 93 single catfish olfactory receptor neurons (ORNs) selected for their narrow excitatory molecular response range (EMRR) to only one type of amino acid (i.e., Group I units). These units were excited by either a basic amino acid, a neutral amino acid with a long side chain, or a neutral amino acid with a short side chain when tested at 10(-7) to 10(-5) M. Stimulus-induced inhibition, likely for contrast enhancement, was primarily observed in response to the types of amino acid stimuli different from that which activated a specific ORN. The high specificity of single Group I ORNs to type of amino acid was also previously observed for single Group I neurons in both the olfactory bulb and forebrain of the same species. These results indicate that for Group I neurons olfactory information concerning specific types of amino acids is processed from receptor neurons through mitral cells of the olfactory bulb to higher forebrain neurons without significant alteration in unit odorant specificity. PMID:17686913

  14. A 10-Gbps × 12-Channel Pluggable Parallel Optical Transceiver Based on CXP Interface Specifications

    NASA Astrophysics Data System (ADS)

    Yang, Kun; Li, Zhihua; Li, Baoxia; Gao, Wei; Liu, Fengman; Song, Jian; Wan, Lixi

    2012-06-01

    A novel 10-Gbps × 12-channel pluggable parallel optical transceiver is designed and fabricated. Compared with other optical transceivers, this transceiver emphasizes small size, high-density, low power consumption, and high transmission speed. Most importantly, its optical coupling structure is simplified to promote cost-effective large-scale production. This transceiver is electrically pluggable and consists of transmitter and receiver modules linked by parallel multi-mode fibers. Each module consists of a six-layered high-speed, high-density printed circuit board, the size of which is 30 mm × 18 mm × 1 mm, packaged with optoelectronic devices and corresponding control chips. The printed circuit boards not only provide a high-speed electrical connection between the I/O interfaces and high-speed chips, but they also supply power and ground planes to those chips. End-to-end error-free transmission at 10.3125 Gbps was obtained for a 231 - 1 non-return-to-zero pseudo-random bit sequence.

  15. Using channel-specific statistical models to detect reverberation in cochlear implant stimuli.

    PubMed

    Desmond, Jill M; Collins, Leslie M; Throckmorton, Chandra S

    2013-08-01

    Reverberation is especially detrimental for cochlear implant listeners; thus, mitigating its effects has the potential to provide significant improvements to cochlear implant communication. Efforts to model and correct for reverberation in acoustic listening scenarios can be quite complex, requiring estimation of the room transfer function and localization of the source and receiver. However, due to the limited resolution associated with cochlear implant stimulation, simpler processing for reverberation detection and mitigation may be possible for cochlear implants. This study models speech stimuli in a cochlear implant on a per-channel basis both in quiet and in reverberation, and assesses the efficacy of these models for detecting the presence of reverberation. This study was able to successfully detect reverberation in cochlear implant pulse trains, and the results appear to be robust to varying room conditions and cochlear implant stimulation parameters. Reverberant signals were detected 100% of the time for a long reverberation time of 1.2 s and 86% of the time for a shorter reverberation time of 0.5 s. PMID:23927111

  16. Transient and specific inactivation of Drosophila neurons in vivo using a native ligand-gated ion channel.

    PubMed

    Liu, Wendy W; Wilson, Rachel I

    2013-07-01

    A key tool in neuroscience is the ability to transiently inactivate specific neurons on timescales of milliseconds to minutes. In Drosophila, there are two available techniques for accomplishing this (shibire(ts) and halorhodopsin [1-3]), but both have shortcomings [4-9]. Here we describe a complementary technique using a native histamine-gated chloride channel (Ort). Ort is the receptor at the first synapse in the visual system. It forms large-conductance homomeric channels that desensitize only modestly in response to ligand [10]. Many regions of the CNS are devoid of histaminergic neurons [11, 12], raising the possibility that Ort could be used to artificially inactivate specific neurons in these regions. To test this idea, we performed in vivo whole-cell recordings from antennal lobe neurons misexpressing Ort. In these neurons, histamine produced a rapid and reversible drop in input resistance, clamping the membrane potential below spike threshold and virtually abolishing spontaneous and odor-evoked activity. Every neuron type in this brain region could be inactivated in this manner. Neurons that did not misexpress Ort showed negligible responses to histamine. Ort also performed favorably in comparison to the available alternative effector transgenes. Thus, Ort misexpression is a useful tool for probing functional connectivity among Drosophila neurons. PMID:23770187

  17. Specific serum antibody responses in channel catfish (Ictalurus punctatus) provide limited protection against Streptococcus ictaluri challenge

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Passive immunization has been shown to provide a spectrum of protection against certain piscine pathogens, and studies were conducted to determine the role of specific antibodies in immunity to Streptococcus ictaluri. Adult Nile tilapia (Oreochromis niloticus) were injected i.p. with tryptic soy br...

  18. Affinity and specificity of interactions between Nedd4 isoforms and the epithelial Na+ channel.

    PubMed

    Henry, Pauline C; Kanelis, Voula; O'Brien, M Christine; Kim, Brian; Gautschi, Ivan; Forman-Kay, Julie; Schild, Laurent; Rotin, Daniela

    2003-05-30

    The epithelial Na+ channel (alphabetagammaENaC) regulates salt and fluid homeostasis and blood pressure. Each ENaC subunit contains a PY motif (PPXY) that binds to the WW domains of Nedd4, a Hect family ubiquitin ligase containing 3-4 WW domains and usually a C2 domain. It has been proposed that Nedd4-2, but not Nedd4-1, isoforms can bind to and suppress ENaC activity. Here we challenge this notion and show that, instead, the presence of a unique WW domain (WW3*) in either Nedd4-2 or Nedd4-1 determines high affinity interactions and the ability to suppress ENaC. WW3* from either Nedd4-2 or Nedd4-1 binds ENaC-PY motifs equally well (e.g. Kd approximately 10 microm for alpha- or betaENaC, 3-6-fold higher affinity than WW4), as determined by intrinsic tryptophan fluorescence. Moreover, dNedd4-1, which naturally contains a WW3* instead of WW2, is able to suppress ENaC function equally well as Nedd4-2. Homology models of the WW3*.betaENaC-PY complex revealed that a Pro and Ala conserved in all WW3*, but not other Nedd4-WW domains, help form the binding pocket for PY motif prolines. Extensive contacts are formed between the betaENaC-PY motif and the Pro in WW3*, and the small Ala creates a large pocket to accommodate the peptide. Indeed, mutating the conserved Pro and Ala in WW3* reduces binding affinity 2-3-fold. Additionally, we demonstrate that mutations in PY motif residues that form contacts with the WW domain based on our previously solved structure either abolish or severely reduce binding affinity to the WW domain and that the extent of binding correlates with the level of ENaC suppression. Independently, we show that a peptide encompassing the PY motif of sgk1, previously proposed to bind to Nedd4-2 and alter its ability to regulate ENaC, does not bind (or binds poorly) the WW domains of Nedd4-2. Collectively, these results suggest that high affinity of WW domain-PY-motif interactions rather than affiliation with Nedd4-1/Nedd-2 is critical for ENaC suppression

  19. Syntaxin 1A inhibits CFTR chloride channels by means of domain-specific protein–protein interactions

    PubMed Central

    Naren, Anjaparavanda P.; Quick, Michael W.; Collawn, James F.; Nelson, Deborah J.; Kirk, Kevin L.

    1998-01-01

    Previously we showed that the functional activity of the epithelial chloride channel that is encoded by the cystic fibrosis gene (CFTR) is reciprocally modulated by two components of the vesicle fusion machinery, syntaxin 1A and Munc-18. Here we report that syntaxin 1A inhibits CFTR chloride channels by means of direct and domain-specific protein–protein interactions. Syntaxin 1A stoichiometrically binds to the N-terminal cytoplasmic tail of CFTR, and this binding is blocked by Munc-18. The modulation of CFTR currents by syntaxin 1A is eliminated either by deletion of this tail or by injecting this tail as a blocking peptide into coexpressing Xenopus oocytes. The CFTR binding site on syntaxin 1A maps to the third predicted helical domain (H3) of this membrane protein. Moreover, CFTR Cl− currents are effectively inhibited by a minimal syntaxin 1A construct (i.e., the membrane-anchored H3 domain) that cannot fully substitute for wild-type syntaxin 1A in membrane fusion reactions. We also show that syntaxin 1A binds to and inhibits the activities of disease-associated mutants of CFTR, and that the chloride current activity of recombinant ΔF508 CFTR (i.e., the most common cystic fibrosis mutant) can be potentiated by disrupting its interaction with syntaxin 1A in cultured epithelial cells. Our results provide evidence for a direct physical interaction between CFTR and syntaxin 1A that limits the functional activities of normal and disease-associated forms of this chloride channel. PMID:9724814

  20. MxA, a member of the dynamin superfamily, interacts with the ankyrin-like repeat domain of TRPC.

    PubMed

    Lussier, Marc P; Cayouette, Sylvie; Lepage, Pascale K; Bernier, Cynthia L; Francoeur, Nancy; St-Hilaire, Marie; Pinard, Maxime; Boulay, Guylain

    2005-05-13

    Mammalian transient receptor potential canonical channels have been proposed as the molecular entities associated with calcium entry activity in nonexcitable cells. Amino acid sequence analyses of TRPCs revealed the presence of ankyrin-like repeat domains, one of the most common protein-protein interaction motifs. Using a yeast two-hybrid interaction assay, we found that the second ankyrin-like repeat domain of TRPC6 interacted with MxA, a member of the dynamin superfamily. Using a GST pull-down and co-immunoprecipitation assay, we showed that MxA interacted with TRPC1, -3, -4, -5, -6, and -7. Overexpression of MxA in HEK293T cells slightly increased endogenous calcium entry subsequent to stimulation of G(q) protein-coupled receptors or store depletion by thapsigargin. Co-expression of MxA with TRPC6 enhanced agonist-induced or OAG-induced calcium entry activity. GTP binding-defective MxA mutants had only a minor potentiating effect on OAG-induced TRPC6 activity. However, a MxA mutant that could bind GTP but that lacked GTPase activity produced the same effect as MxA on OAG-induced TRPC6 activity. These results indicated that MxA interacted specifically with the second ankyrin-like repeat domain of TRPCs and suggested that monomeric MxA regulated the activity of TRPC6 by a mechanism requiring GTP binding. Additional results showed that an increase in the endogenous expression of MxA, induced by a treatment with interferon alpha, regulated the activity of TRPC6. The study clearly identified MxA as a new regulatory protein involved in Ca2+ signaling. PMID:15757897

  1. How much donor financing for health is channelled to global versus country-specific aid functions?

    PubMed

    Schäferhoff, Marco; Fewer, Sara; Kraus, Jessica; Richter, Emil; Summers, Lawrence H; Sundewall, Jesper; Yamey, Gavin; Jamison, Dean T

    2015-12-12

    The slow global response to the Ebola crisis in west Africa suggests that important gaps exist in donor financing for key global functions, such as support for health research and development for diseases of poverty and strengthening of outbreak preparedness. In this Health Policy, we use the International Development Statistics databases to quantify donor support for such functions. We classify donor funding for health into aid for global functions (provision of global public goods, management of cross-border externalities, and fostering of leadership and stewardship) versus country-specific aid. We use a new measure of donor funding that combines official development assistance (ODA) for health with additional donor spending on research and development (R&D) for diseases of poverty. Much R&D spending falls outside ODA--ie, the assistance that is conventionally reported through ODA databases of the Organisation for Economic Co-operation and Development. This expanded definition, which we term health ODA plus, provides a more comprehensive picture of donor support for health that could reshape how policy makers will approach their support for global health. PMID:26178405

  2. Distinct Cell- and Layer-Specific Expression Patterns and Independent Regulation of Kv2 Channel Subtypes in Cortical Pyramidal Neurons

    PubMed Central

    Bishop, Hannah I.; Guan, Dongxu; Bocksteins, Elke; Parajuli, Laxmi Kumar; Murray, Karl D.; Cobb, Melanie M.; Misonou, Hiroaki; Zito, Karen; Foehring, Robert C.

    2015-01-01

    The Kv2 family of voltage-gated potassium channel α subunits, comprising Kv2.1 and Kv2.2, mediate the bulk of the neuronal delayed rectifier K+ current in many mammalian central neurons. Kv2.1 exhibits robust expression across many neuron types and is unique in its conditional role in modulating intrinsic excitability through changes in its phosphorylation state, which affect Kv2.1 expression, localization, and function. Much less is known of the highly related Kv2.2 subunit, especially in forebrain neurons. Here, through combined use of cortical layer markers and transgenic mouse lines, we show that Kv2.1 and Kv2.2 are localized to functionally distinct cortical cell types. Kv2.1 expression is consistently high throughout all cortical layers, especially in layer (L) 5b pyramidal neurons, whereas Kv2.2 expression is primarily limited to neurons in L2 and L5a. In addition, L4 of primary somatosensory cortex is strikingly devoid of Kv2.2 immunolabeling. The restricted pattern of Kv2.2 expression persists in Kv2.1-KO mice, suggesting distinct cell- and layer-specific functions for these two highly related Kv2 subunits. Analyses of endogenous Kv2.2 in cortical neurons in situ and recombinant Kv2.2 expressed in heterologous cells reveal that Kv2.2 is largely refractory to stimuli that trigger robust, phosphorylation-dependent changes in Kv2.1 clustering and function. Immunocytochemistry and voltage-clamp recordings from outside-out macropatches reveal distinct cellular expression patterns for Kv2.1 and Kv2.2 in intratelencephalic and pyramidal tract neurons of L5, indicating circuit-specific requirements for these Kv2 paralogs. Together, these results support distinct roles for these two Kv2 channel family members in mammalian cortex. SIGNIFICANCE STATEMENT Neurons within the neocortex are arranged in a laminar architecture and contribute to the input, processing, and/or output of sensory and motor signals in a cell- and layer-specific manner. Neurons of different

  3. Astrocytes in the optic nerve head express putative mechanosensitive channels

    PubMed Central

    Choi, Hee Joo; Sun, Daniel

    2015-01-01

    Purpose To establish whether optic nerve head astrocytes express candidate molecules to sense tissue stretch. Methods We used conventional PCR, quantitative PCR, and single-cell reverse transcription PCR (RT–PCR) to assess the expression of various members of the transient receptor potential (TRP) channel family and of the recently characterized mechanosensitive channels Piezo1 and 2 in optic nerve head tissue and in single, isolated astrocytes. Results Most TRP subfamilies (TRPC, TRPM, TRPV, TRPA, and TRPP) and Piezo1 and 2 were expressed in the optic nerve head of the mouse. Quantitative real-time PCR analysis showed that TRPC1, TRPM7, TRPV2, TRPP2, and Piezo1 are the dominant isoforms in each subfamily. Single-cell RT–PCR revealed that many TRP isoforms, TRPC1–2, TRPC6, TRPV2, TRPV4, TRPM2, TRPM4, TRPM6–7, TRPP1–2, and Piezo1–2, are expressed in astrocytes of the optic nerve head, and that most astrocytes express TRPC1 and TRPP1–2. Comparisons of the TRPP and Piezo expression levels between different tissue regions showed that Piezo2 expression was higher in the optic nerve head and the optic nerve proper than in the brain and the corpus callosum. TRPP2 also showed higher expression in the optic nerve head. Conclusions Astrocytes in the optic nerve head express multiple putative mechanosensitive channels, in particular the recently identified channels Piezo1 and 2. The expression of putative mechanosensitive channels in these cells may contribute to their responsiveness to traumatic or glaucomatous injury. PMID:26236150

  4. Optogenetic determination of the myocardial requirements for extrasystoles by cell type-specific targeting of ChannelRhodopsin-2

    PubMed Central

    Zaglia, Tania; Pianca, Nicola; Borile, Giulia; Da Broi, Francesca; Richter, Claudia; Campione, Marina; Lehnart, Stephan E.; Luther, Stefan; Corrado, Domenico; Miquerol, Lucile; Mongillo, Marco

    2015-01-01

    Extrasystoles lead to several consequences, ranging from uneventful palpitations to lethal ventricular arrhythmias, in the presence of pathologies, such as myocardial ischemia. The role of working versus conducting cardiomyocytes, as well as the tissue requirements (minimal cell number) for the generation of extrasystoles, and the properties leading ectopies to become arrhythmia triggers (topology), in the normal and diseased heart, have not been determined directly in vivo. Here, we used optogenetics in transgenic mice expressing ChannelRhodopsin-2 selectively in either cardiomyocytes or the conduction system to achieve cell type-specific, noninvasive control of heart activity with high spatial and temporal resolution. By combining measurement of optogenetic tissue activation in vivo and epicardial voltage mapping in Langendorff-perfused hearts, we demonstrated that focal ectopies require, in the normal mouse heart, the simultaneous depolarization of at least 1,300–1,800 working cardiomyocytes or 90–160 Purkinje fibers. The optogenetic assay identified specific areas in the heart that were highly susceptible to forming extrasystolic foci, and such properties were correlated to the local organization of the Purkinje fiber network, which was imaged in three dimensions using optical projection tomography. Interestingly, during the acute phase of myocardial ischemia, focal ectopies arising from this location, and including both Purkinje fibers and the surrounding working cardiomyocytes, have the highest propensity to trigger sustained arrhythmias. In conclusion, we used cell-specific optogenetics to determine with high spatial resolution and cell type specificity the requirements for the generation of extrasystoles and the factors causing ectopies to be arrhythmia triggers during myocardial ischemia. PMID:26204914

  5. Using specific and adaptive arrangement of grid-type pilot in channel estimation for white-lightLED-based OFDM visible light communication system

    NASA Astrophysics Data System (ADS)

    Lin, Wan-Feng; Chow, Chi-Wai; Yeh, Chien-Hung

    2015-03-01

    Orthogonal frequency division multiplexing (OFDM) is a promising candidate for light emitting diode (LED)-based optical wireless communication (OWC); however, precise channel estimation is required for synchronization and equalization. In this work, we study and discover that the channel response of the white-lightLED-based OWC was smooth and stable. Hence we propose and demonstrate using a specific and adaptive arrangement of grid-type pilot scheme to estimate the LED OWC channel response. Experimental results show that our scheme can achieve better transmission performance and with some transmission capacity enhancement when compared with the method using training-symbol scheme (also called block-type pilot scheme).

  6. Heterogeneous distribution of TRPC proteins in the embryonic cortex.

    PubMed

    Boisseau, Sylvie; Kunert-Keil, Christiane; Lucke, Silke; Bouron, Alexandre

    2009-03-01

    The present study was initiated to gain some information about the tissue distribution of transient receptor potential proteins of C-type (TRPC), a family of voltage-independent cation channels, at the beginning of neurogenesis in the telencephalon of embryonic mice. The mRNAs of all known TRPCs (TRPC1-TRPC7) could be found in the cortex at E13. TRPC1, TRPC3 and TRPC5 were the main isoforms, whereas the mRNAs for TRPC2, TRPC4, TRPC6 and TRPC7 were less abundant. The distribution throughout the cortical wall of TRPC1, TRPC3 and TRPC6 was studied by means of immuno-histochemistry. The data collected pointed to a heterogeneous expression of the channels. Three groups were identified. The first one comprises TRPC1, specifically found in the preplate but only in some post-mitotic neurons. It was mainly observed in a subset of cells distinct from the Cajal-Retzius cells. The second group is composed of TRPC3. It was found in non-neuronal cells and also in dividing (5-bromo-2'-deoxyuridine-positive) cells, indicating that TRPC3 is present in precursor cells. The third group contains TRPC6 detected in neuronal and in dividing non-neuronal cells. Double immunostaining experiments showed that TRPC3-positive cells also express TRPC6. Collectively, this report highlights a specific TRPC expression pattern in the immature cortical wall. PMID:18989690

  7. The role of voltage-gated calcium channels in neurotransmitter phenotype specification: Coexpression and functional analysis in Xenopus laevis

    PubMed Central

    Lewis, Brittany B; Miller, Lauren E; Herbst, Wendy A; Saha, Margaret S

    2014-01-01

    Calcium activity has been implicated in many neurodevelopmental events, including the specification of neurotransmitter phenotypes. Higher levels of calcium activity lead to an increased number of inhibitory neural phenotypes, whereas lower levels of calcium activity lead to excitatory neural phenotypes. Voltage-gated calcium channels (VGCCs) allow for rapid calcium entry and are expressed during early neural stages, making them likely regulators of activity-dependent neurotransmitter phenotype specification. To test this hypothesis, multiplex fluorescent in situ hybridization was used to characterize the coexpression of eight VGCC α1 subunits with the excitatory and inhibitory neural markers xVGlut1 and xVIAAT in Xenopus laevis embryos. VGCC coexpression was higher with xVGlut1 than xVIAAT, especially in the hindbrain, spinal cord, and cranial nerves. Calcium activity was also analyzed on a single-cell level, and spike frequency was correlated with the expression of VGCC α1 subunits in cell culture. Cells expressing Cav2.1 and Cav2.2 displayed increased calcium spiking compared with cells not expressing this marker. The VGCC antagonist diltiazem and agonist (−)BayK 8644 were used to manipulate calcium activity. Diltiazem exposure increased the number of glutamatergic cells and decreased the number of γ-aminobutyric acid (GABA)ergic cells, whereas (−)BayK 8644 exposure decreased the number of glutamatergic cells without having an effect on the number of GABAergic cells. Given that the expression and functional manipulation of VGCCs are correlated with neurotransmitter phenotype in some, but not all, experiments, VGCCs likely act in combination with a variety of other signaling factors to determine neuronal phenotype specification. J. Comp. Neurol. 522:2518–2531, 2014. PMID:24477801

  8. Gene expressions of TRP channels in glioblastoma multiforme and relation with survival.

    PubMed

    Alptekin, M; Eroglu, S; Tutar, E; Sencan, S; Geyik, M A; Ulasli, M; Demiryurek, A T; Camci, C

    2015-12-01

    Glioblastoma multiforme (GBM) is one of the most lethal forms of cancer in humans, with a median survival of 10 to 12 months. Glioblastoma is highly malignant since the cells are supported by a great number of blood vessels. Although new treatments have been developed by increasing knowledge of molecular nature of the disease, surgical operation remains the standard of care. The TRP (transient receptor potential) superfamily consists of cation-selective channels that have roles in sensory physiology such as thermo- and osmosensation and in several complex diseases such as cancer, cardiovascular, and neuronal diseases. The aim of this study was to investigate the expression levels of TRP channel genes in patients with glioblastoma multiforme and to evaluate the relationship between TRP gene expressions and survival of the patients. Thirty-three patients diagnosed with glioblastoma were enrolled to the study. The expression levels of 21 TRP genes were quantified by using qRT-PCR with dynamic array 48 × 48 chip (BioMark HD System, Fluidigm, South San Francisco, CA, USA). TRPC1, TRPC6, TRPM2, TRPM3, TRPM7, TRPM8, TRPV1, and TRPV2 were found significantly higher in glioblastoma patients. Moreover, there was a significant relationship between the overexpression of TRP genes and the survival of the patients. These results demonstrate for the first time that TRP channels contribute to the progression and survival of the glioblastoma patients. PMID:26088448

  9. Roles of specific Kv channel types in repolarization of the action potential in genetically identified subclasses of pyramidal neurons in mouse neocortex.

    PubMed

    Pathak, Dhruba; Guan, Dongxu; Foehring, Robert C

    2016-05-01

    The action potential (AP) is a fundamental feature of excitable cells that serves as the basis for long-distance signaling in the nervous system. There is considerable diversity in the appearance of APs and the underlying repolarization mechanisms in different neuronal types (reviewed in Bean BP. Nat Rev Neurosci 8: 451-465, 2007), including among pyramidal cell subtypes. In the present work, we used specific pharmacological blockers to test for contributions of Kv1, Kv2, or Kv4 channels to repolarization of single APs in two genetically defined subpopulations of pyramidal cells in layer 5 of mouse somatosensory cortex (etv1 and glt) as well as pyramidal cells from layer 2/3. These three subtypes differ in AP properties (Groh A, Meyer HS, Schmidt EF, Heintz N, Sakmann B, Krieger P. Cereb Cortex 20: 826-836, 2010; Guan D, Armstrong WE, Foehring RC. J Neurophysiol 113: 2014-2032, 2015) as well as laminar position, morphology, and projection targets. We asked what the roles of Kv1, Kv2, and Kv4 channels are in AP repolarization and whether the underlying mechanisms are pyramidal cell subtype dependent. We found that Kv4 channels are critically involved in repolarizing neocortical pyramidal cells. There are also pyramidal cell subtype-specific differences in the role for Kv1 channels. Only Kv4 channels were involved in repolarizing the narrow APs of glt cells. In contrast, in etv1 cells and layer 2/3 cells, the broader APs are partially repolarized by Kv1 channels in addition to Kv4 channels. Consistent with their activation in the subthreshold range, Kv1 channels also regulate AP voltage threshold in all pyramidal cell subtypes. PMID:26864770

  10. Relationship between expression of muscle-specific uncoupling protein 2 messenger RNA and genetic selection toward growth in channel catfish

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Uncoupling protein 2 is a member of the mitochondrial channel proteins that regulate the flow of hydrogen ions and ATP generation. The relationship between UCP2 and nutrient metabolism has been well-defined in humans but unclear in fish. We hypothesized that increased muscle growth in channel catf...

  11. Subcellular compartment-specific molecular diversity of pre- and postsynaptic GABAB-activated GIRK channels in Purkinje cells

    PubMed Central

    Fernández-Alacid, Laura; Aguado, Carolina; Ciruela, Francisco; Martín, Ricardo; Colón, José; Cabañero, María José; Gassmann, Martin; Watanabe, Masahiko; Shigemoto, Ryuichi; Wickman, Kevin; Bettler, Bernhard; Sánchez-Prieto, José; Luján, Rafael

    2009-01-01

    Activation of G protein-gated inwardly-rectifying K+ (GIRK or Kir3) channels by metabotropic gamma-aminobutyric acid (B) (GABAB) receptors is an essential signalling pathway controlling neuronal excitability and synaptic transmission in the brain. To investigate the relationship between GIRK channel subunits and GABAB receptors in cerebellar Purkinje cells at post- and pre-synaptic sites, we used biochemical, functional and immunohistochemical techniques. Co-immunoprecipitation analysis demonstrated that GIRK subunits are co-assembled with GABAB receptors in the cerebellum. Immunoelectron microscopy showed that the subunit composition of GIRK channels in Purkinje cell spines is compartment-dependent. Thus, at extrasynaptic sites GIRK channels are formed by GIRK1/GIRK2/GIRK3, postsynaptic densities contain GIRK2/GIRK3 and dendritic shafts contain GIRK1/GIRK3. The postsynaptic association of GIRK subunits with GABAB receptors in Purkinje cells is supported by the subcellular regulation of the ion channel and the receptor in mutant mice. At presynaptic sites, GIRK channels localized to parallel fibre terminals are formed by GIRK1/GIRK2/GIRK3 and co-localize with GABAB receptors. Consistent with this morphological evidence we demonstrate their functional interaction at axon terminals in the cerebellum by showing that GIRK channels play a role in the inhibition of glutamate release by GABAB receptors. The association of GIRK channels and GABAB receptors with excitatory synapses at both post- and presynaptic sites indicates their intimate involvement in the modulation of glutamatergic neurotransmission in the cerebellum. PMID:19558451

  12. Potent KCNQ2/3-Specific Channel Activator Suppresses In Vivo Epileptic Activity and Prevents the Development of Tinnitus

    PubMed Central

    Kalappa, Bopanna I.; Soh, Heun; Duignan, Kevin M.; Furuya, Takeru; Edwards, Scott

    2015-01-01

    Voltage-gated Kv7 (KCNQ) channels are voltage-dependent potassium channels that are activated at resting membrane potentials and therefore provide a powerful brake on neuronal excitability. Genetic or experience-dependent reduction of KCNQ2/3 channel activity is linked with disorders that are characterized by neuronal hyperexcitability, such as epilepsy and tinnitus. Retigabine, a small molecule that activates KCNQ2–5 channels by shifting their voltage-dependent opening to more negative voltages, is an US Food and Drug Administration (FDA) approved anti-epileptic drug. However, recently identified side effects have limited its clinical use. As a result, the development of improved KCNQ2/3 channel activators is crucial for the treatment of hyperexcitability-related disorders. By incorporating a fluorine substituent in the 3-position of the tri-aminophenyl ring of retigabine, we synthesized a small-molecule activator (SF0034) with novel properties. Heterologous expression of KCNQ2/3 channels in HEK293T cells showed that SF0034 was five times more potent than retigabine at shifting the voltage dependence of KCNQ2/3 channels to more negative voltages. Moreover, unlike retigabine, SF0034 did not shift the voltage dependence of either KCNQ4 or KCNQ5 homomeric channels. Conditional deletion of Kcnq2 from cerebral cortical pyramidal neurons showed that SF0034 requires the expression of KCNQ2/3 channels for reducing the excitability of CA1 hippocampal neurons. Behavioral studies demonstrated that SF0034 was a more potent and less toxic anticonvulsant than retigabine in rodents. Furthermore, SF0034 prevented the development of tinnitus in mice. We propose that SF0034 provides, not only a powerful tool for investigating ion channel properties, but, most importantly, it provides a clinical candidate for treating epilepsy and preventing tinnitus. PMID:26063916

  13. Brain-derived neurotrophic factor activation of CaM-kinase kinase via transient receptor potential canonical channels induces the translation and synaptic incorporation of GluA1-containing calcium-permeable AMPA receptors.

    PubMed

    Fortin, Dale A; Srivastava, Taasin; Dwarakanath, Diya; Pierre, Philippe; Nygaard, Sean; Derkach, Victor A; Soderling, Thomas R

    2012-06-13

    Glutamatergic synapses in early postnatal development transiently express calcium-permeable AMPA receptors (CP-AMPARs). Although these GluA2-lacking receptors are essential and are elevated in response to brain-derived neurotrophic factor (BDNF), little is known regarding molecular mechanisms that govern their expression and synaptic insertion. Here we show that BDNF-induced GluA1 translation in rat primary hippocampal neurons requires the activation of mammalian target of rapamycin (mTOR) via calcium calmodulin-dependent protein kinase kinase (CaMKK). Specifically, BDNF-mediated phosphorylation of threonine 308 (T308) in AKT, a known substrate of CaMKK and an upstream activator of mTOR-dependent translation, was prevented by (1) pharmacological inhibition of CaMKK with STO-609, (2) overexpression of a dominant-negative CaMKK, or (3) short hairpin-mediated knockdown of CaMKK. GluA1 surface expression induced by BDNF, as assessed by immunocytochemistry using an extracellular N-terminal GluA1 antibody or by surface biotinylation, was impaired following knockdown of CaMKK or treatment with STO-609. Activation of CaMKK by BDNF requires transient receptor potential canonical (TRPC) channels as SKF-96365, but not the NMDA receptor antagonist d-APV, prevented BDNF-induced GluA1 surface expression as well as phosphorylation of CaMKI, AKT(T308), and mTOR. Using siRNA we confirmed the involvement of TRPC5 and TRPC6 subunits in BDNF-induced AKT(T308) phosphorylation. The BDNF-induced increase in mEPSC was blocked by IEM-1460, a selected antagonist of CP-AMPARs, as well as by the specific repression of acute GluA1 translation via siRNA to GluA1 but not GluA2. Together these data support the conclusion that newly synthesized GluA1 subunits, induced by BDNF, are readily incorporated into synapses where they enhance the expression of CP-AMPARs and synaptic strength. PMID:22699894

  14. Gating-pore currents demonstrate selective and specific modulation of individual sodium channel voltage-sensors by biological toxins.

    PubMed

    Xiao, Yucheng; Blumenthal, Kenneth; Cummins, Theodore R

    2014-08-01

    Voltage-gated sodium channels are critical determinants of nerve and muscle excitability. Although numerous toxins and small molecules target sodium channels, identifying the mechanisms of action is challenging. Here we used gating-pore currents selectively generated in each of the voltage-sensors from the four α-subunit domains (DI-DIV) to monitor the activity of individual voltage-sensors and to investigate the molecular determinants of sodium channel pharmacology. The tarantula toxin huwentoxin-IV (HWTX-IV), which inhibits sodium channel current, exclusively enhanced inward gating-pore currents through the DII voltage-sensor. By contrast, the tarantula toxin ProTx-II, which also inhibits sodium channel currents, altered the gating-pore currents in multiple voltage-sensors in a complex manner. Thus, whereas HWTX-IV inhibits central-pore currents by selectively trapping the DII voltage-sensor in the resting configuration, ProTx-II seems to inhibit central-pore currents by differentially altering the configuration of multiple voltage-sensors. The sea anemone toxin anthopleurin B, which impairs open-channel inactivation, exclusively enhanced inward gating-pore currents through the DIV voltage-sensor. This indicates that trapping the DIV voltage-sensor in the resting configuration selectively impairs open-channel inactivation. Furthermore, these data indicate that although activation of all four voltage-sensors is not required for central-pore current generation, activation of the DII voltage-sensor is crucial. Overall, our data demonstrate that gating-pore currents can determine the mechanism of action for sodium channel gating modifiers with high precision. We propose this approach could be adapted to identify the molecular mechanisms of action for gating modifiers of various voltage-gated ion channels. PMID:24898004

  15. Gating-Pore Currents Demonstrate Selective and Specific Modulation of Individual Sodium Channel Voltage-Sensors by Biological Toxins

    PubMed Central

    Xiao, Yucheng; Blumenthal, Kenneth

    2014-01-01

    Voltage-gated sodium channels are critical determinants of nerve and muscle excitability. Although numerous toxins and small molecules target sodium channels, identifying the mechanisms of action is challenging. Here we used gating-pore currents selectively generated in each of the voltage-sensors from the four α-subunit domains (DI–DIV) to monitor the activity of individual voltage-sensors and to investigate the molecular determinants of sodium channel pharmacology. The tarantula toxin huwentoxin-IV (HWTX-IV), which inhibits sodium channel current, exclusively enhanced inward gating-pore currents through the DII voltage-sensor. By contrast, the tarantula toxin ProTx-II, which also inhibits sodium channel currents, altered the gating-pore currents in multiple voltage-sensors in a complex manner. Thus, whereas HWTX-IV inhibits central-pore currents by selectively trapping the DII voltage-sensor in the resting configuration, ProTx-II seems to inhibit central-pore currents by differentially altering the configuration of multiple voltage-sensors. The sea anemone toxin anthopleurin B, which impairs open-channel inactivation, exclusively enhanced inward gating-pore currents through the DIV voltage-sensor. This indicates that trapping the DIV voltage-sensor in the resting configuration selectively impairs open-channel inactivation. Furthermore, these data indicate that although activation of all four voltage-sensors is not required for central-pore current generation, activation of the DII voltage-sensor is crucial. Overall, our data demonstrate that gating-pore currents can determine the mechanism of action for sodium channel gating modifiers with high precision. We propose this approach could be adapted to identify the molecular mechanisms of action for gating modifiers of various voltage-gated ion channels. PMID:24898004

  16. A Specific Subset of Transient Receptor Potential Vanilloid-Type Channel Subunits in Caenorhabditis elegans Endocrine Cells Function as Mixed Heteromers to Promote Neurotransmitter Release

    PubMed Central

    Jose, Antony M.; Bany, I. Amy; Chase, Daniel L.; Koelle, Michael R.

    2007-01-01

    Transient receptor potential (TRP) channel subunits form homotetramers that function in sensory transduction. Heteromeric channels also form, but their physiological subunit compositions and functions are largely unknown. We found a dominant-negative mutant of the C. elegans TRPV (vanilloid-type) subunit OCR-2 that apparently incorporates into and inactivates OCR-2 homomers as well as heteromers with the TRPV subunits OCR-1 and -4, resulting in a premature egg-laying defect. This defect is reproduced by knocking out all three OCR genes, but not by any single knockout. Thus a mixture of redundant heteromeric channels prevents premature egg laying. These channels, as well as the G-protein Gαo, function in neuroendocrine cells to promote release of neurotransmitters that block egg laying until eggs filling the uterus deform the neuroendocrine cells. The TRPV channel OSM-9, previously suggested to be an obligate heteromeric partner of OCR-2 in sensory neurons, is expressed in the neuroendocrine cells but has no detectable role in egg laying. Our results identify a specific set of heteromeric TRPV channels that redundantly regulate neuroendocrine function and show that a subunit combination that functions in sensory neurons is also present in neuroendocrine cells but has no detectable function in these cells. PMID:17057248

  17. Sensory neuron-specific sodium channel SNS is abnormally expressed in the brains of mice with experimental allergic encephalomyelitis and humans with multiple sclerosis

    NASA Astrophysics Data System (ADS)

    Black, Joel A.; Dib-Hajj, Sulayman; Baker, David; Newcombe, Jia; Cuzner, M. Louise; Waxman, Stephen G.

    2000-10-01

    Clinical abnormalities in multiple sclerosis (MS) have classically been considered to be caused by demyelination and/or axonal degeneration; the possibility of molecular changes in neurons, such as the deployment of abnormal repertoires of ion channels that would alter neuronal electrogenic properties, has not been considered. Sensory Neuron-Specific sodium channel SNS displays a depolarized voltage dependence, slower activation and inactivation kinetics, and more rapid recovery from inactivation than classical "fast" sodium channels. SNS is selectively expressed in spinal sensory and trigeminal ganglion neurons within the peripheral nervous system and is not expressed within the normal brain. Here we show that sodium channel SNS mRNA and protein, which are not present within the cerebellum of control mice, are expressed within cerebellar Purkinje cells in a mouse model of MS, chronic relapsing experimental allergic encephalomyelitis. We also demonstrate SNS mRNA and protein expression within Purkinje cells from tissue obtained postmortem from patients with MS, but not in control subjects with no neurological disease. These results demonstrate a change in sodium channel expression in neurons within the brain in an animal model of MS and in humans with MS and suggest that abnormal patterns of neuronal ion channel expression may contribute to clinical abnormalities such as ataxia in these disorders.

  18. Ion-Channel Genosensor for the Detection of Specific DNA Sequences Derived from Plum Pox Virus in Plant Extracts

    PubMed Central

    Malecka, Kamila; Michalczuk, Lech; Radecka, Hanna; Radecki, Jerzy

    2014-01-01

    A DNA biosensor for detection of specific oligonucleotides sequences of Plum Pox Virus (PPV) in plant extracts and buffer is proposed. The working principles of a genosensor are based on the ion-channel mechanism. The NH2-ssDNA probe was deposited onto a glassy carbon electrode surface to form an amide bond between the carboxyl group of oxidized electrode surface and amino group from ssDNA probe. The analytical signals generated as a result of hybridization were registered in Osteryoung square wave voltammetry in the presence of [Fe(CN)6]3−/4− as a redox marker. The 22-mer and 42-mer complementary ssDNA sequences derived from PPV and DNA samples from plants infected with PPV were used as targets. Similar detection limits of 2.4 pM (31.0 pg/mL) and 2.3 pM (29.5 pg/mL) in the concentration range 1–8 pM were observed in the presence of the 22-mer ssDNA and 42-mer complementary ssDNA sequences of PPV, respectively. The genosensor was capable of discriminating between samples consisting of extracts from healthy plants and leaf extracts from infected plants in the concentration range 10–50 pg/mL. The detection limit was 12.8 pg/mL. The genosensor displayed good selectivity and sensitivity. The 20-mer partially complementary DNA sequences with four complementary bases and DNA samples from healthy plants used as negative controls generated low signal. PMID:25302809

  19. Co-chaperone Specificity in Gating of the Polypeptide Conducting Channel in the Membrane of the Human Endoplasmic Reticulum*

    PubMed Central

    Schorr, Stefan; Klein, Marie-Christine; Gamayun, Igor; Melnyk, Armin; Jung, Martin; Schäuble, Nico; Wang, Qian; Hemmis, Birgit; Bochen, Florian; Greiner, Markus; Lampel, Pavel; Urban, Sabine Katharina; Hassdenteufel, Sarah; Dudek, Johanna; Chen, Xing-Zhen; Wagner, Richard; Cavalié, Adolfo; Zimmermann, Richard

    2015-01-01

    In mammalian cells, signal peptide-dependent protein transport into the endoplasmic reticulum (ER) is mediated by a dynamic polypeptide-conducting channel, the heterotrimeric Sec61 complex. Previous work has characterized the Sec61 complex as a potential ER Ca2+ leak channel in HeLa cells and identified ER lumenal molecular chaperone immunoglobulin heavy-chain-binding protein (BiP) as limiting Ca2+ leakage via the open Sec61 channel by facilitating channel closing. This BiP activity involves binding of BiP to the ER lumenal loop 7 of Sec61α in the vicinity of tyrosine 344. Of note, the Y344H mutation destroys the BiP binding site and causes pancreatic β-cell apoptosis and diabetes in mice. Here, we systematically depleted HeLa cells of the BiP co-chaperones by siRNA-mediated gene silencing and used live cell Ca2+ imaging to monitor the effects on ER Ca2+ leakage. Depletion of either one of the ER lumenal BiP co-chaperones, ERj3 and ERj6, but not the ER membrane-resident co-chaperones (such as Sec63 protein, which assists BiP in Sec61 channel opening) led to increased Ca2+ leakage via Sec6 complex, thereby phenocopying the effect of BiP depletion. Thus, BiP facilitates Sec61 channel closure (i.e. limits ER Ca2+ leakage) via the Sec61 channel with the help of ERj3 and ERj6. Interestingly, deletion of ERj6 causes pancreatic β-cell failure and diabetes in mice and humans. We suggest that co-chaperone-controlled gating of the Sec61 channel by BiP is particularly important for cells, which are highly active in protein secretion, and that breakdown of this regulatory mechanism can cause apoptosis and disease. PMID:26085089

  20. Co-chaperone Specificity in Gating of the Polypeptide Conducting Channel in the Membrane of the Human Endoplasmic Reticulum.

    PubMed

    Schorr, Stefan; Klein, Marie-Christine; Gamayun, Igor; Melnyk, Armin; Jung, Martin; Schäuble, Nico; Wang, Qian; Hemmis, Birgit; Bochen, Florian; Greiner, Markus; Lampel, Pavel; Urban, Sabine Katharina; Hassdenteufel, Sarah; Dudek, Johanna; Chen, Xing-Zhen; Wagner, Richard; Cavalié, Adolfo; Zimmermann, Richard

    2015-07-24

    In mammalian cells, signal peptide-dependent protein transport into the endoplasmic reticulum (ER) is mediated by a dynamic polypeptide-conducting channel, the heterotrimeric Sec61 complex. Previous work has characterized the Sec61 complex as a potential ER Ca(2+) leak channel in HeLa cells and identified ER lumenal molecular chaperone immunoglobulin heavy-chain-binding protein (BiP) as limiting Ca(2+) leakage via the open Sec61 channel by facilitating channel closing. This BiP activity involves binding of BiP to the ER lumenal loop 7 of Sec61α in the vicinity of tyrosine 344. Of note, the Y344H mutation destroys the BiP binding site and causes pancreatic β-cell apoptosis and diabetes in mice. Here, we systematically depleted HeLa cells of the BiP co-chaperones by siRNA-mediated gene silencing and used live cell Ca(2+) imaging to monitor the effects on ER Ca(2+) leakage. Depletion of either one of the ER lumenal BiP co-chaperones, ERj3 and ERj6, but not the ER membrane-resident co-chaperones (such as Sec63 protein, which assists BiP in Sec61 channel opening) led to increased Ca(2+) leakage via Sec6 complex, thereby phenocopying the effect of BiP depletion. Thus, BiP facilitates Sec61 channel closure (i.e. limits ER Ca(2+) leakage) via the Sec61 channel with the help of ERj3 and ERj6. Interestingly, deletion of ERj6 causes pancreatic β-cell failure and diabetes in mice and humans. We suggest that co-chaperone-controlled gating of the Sec61 channel by BiP is particularly important for cells, which are highly active in protein secretion, and that breakdown of this regulatory mechanism can cause apoptosis and disease. PMID:26085089

  1. Clemizole hydrochloride is a novel and potent inhibitor of transient receptor potential channel TRPC5.

    PubMed

    Richter, Julia M; Schaefer, Michael; Hill, Kerstin

    2014-11-01

    Canonical transient receptor potential channel 5 (TRPC5) is a nonselective, Ca(2+)-permeable cation channel that belongs to the large family of transient receptor potential channels. It is predominantly found in the central nervous system with a high expression density in the hippocampus, the amygdala, and the frontal cortex. Several studies confirm that TRPC5 channels are implicated in the regulation of neurite length and growth cone morphology. We identified clemizole as a novel inhibitor of TRPC5 channels. Clemizole efficiently blocks TRPC5 currents and Ca(2+) entry in the low micromolar range (IC50 = 1.0-1.3 µM), as determined by fluorometric intracellular free Ca(2+) concentration ([Ca(2+)]i) measurements and patch-clamp recordings. Clemizole blocks TRPC5 currents irrespectively of the mode of activation, for example, stimulation of G protein-coupled receptors, hypo-osmotic buffer conditions, or by the direct activator riluzole. Electrophysiological whole-cell recordings revealed that the block was mostly reversible. Moreover, clemizole was still effective in blocking TRPC5 single channels in excised inside-out membrane patches, hinting to a direct block of TRPC5 by clemizole. Based on fluorometric [Ca(2+)]i measurements, clemizole exhibits a sixfold selectivity for TRPC5 over TRPC4β (IC50 = 6.4 µM), the closest structural relative of TRPC5, and an almost 10-fold selectivity over TRPC3 (IC50 = 9.1 µM) and TRPC6 (IC50 = 11.3 µM). TRPM3 and M8 as well as TRPV1, V2, V3, and V4 channels were only weakly affected by markedly higher clemizole concentrations. Clemizole was not only effective in blocking heterologously expressed TRPC5 homomers but also TRPC1:TRPC5 heteromers as well as native TRPC5-like currents in the U-87 glioblastoma cell line. PMID:25140002

  2. Ligand-specific opening of a gated-porin channel in the outer membrane of living bacteria.

    PubMed

    Jiang, X; Payne, M A; Cao, Z; Foster, S B; Feix, J B; Newton, S M; Klebba, P E

    1997-05-23

    Ligand-gated membrane channels selectively facilitate the entry of iron into prokaryotic cells. The essential role of iron in metabolism makes its acquisition a determinant of bacterial pathogenesis and a target for therapeutic strategies. In Gram-negative bacteria, TonB-dependent outer membrane proteins form energized, gated pores that bind iron chelates (siderophores) and internalize them. The time-resolved operation of the Escherichia coli ferric enterobactin receptor FepA was observed in vivo with electron spin resonance spectroscopy by monitoring the mobility of covalently bound nitroxide spin labels. A ligand-binding surface loop of FepA, which normally closes its transmembrane channel, exhibited energy-dependent structural changes during iron and toxin (colicin) transport. These changes were not merely associated with ligand binding, but occurred during ligand uptake through the outer membrane bilayer. The results demonstrate by a physical method that gated-porin channels open and close during membrane transport in vivo. PMID:9157886

  3. Identification of ML204, a Novel Potent Antagonist That Selectively Modulates Native TRPC4/C5 Ion Channels*

    PubMed Central

    Miller, Melissa; Shi, Jie; Zhu, Yingmin; Kustov, Maksym; Tian, Jin-bin; Stevens, Amy; Wu, Meng; Xu, Jia; Long, Shunyou; Yang, Pu; Zholos, Alexander V.; Salovich, James M.; Weaver, C. David; Hopkins, Corey R.; Lindsley, Craig W.; McManus, Owen; Li, Min; Zhu, Michael X.

    2011-01-01

    Transient receptor potential canonical (TRPC) channels are Ca2+-permeable nonselective cation channels implicated in diverse physiological functions, including smooth muscle contractility and synaptic transmission. However, lack of potent selective pharmacological inhibitors for TRPC channels has limited delineation of the roles of these channels in physiological systems. Here we report the identification and characterization of ML204 as a novel, potent, and selective TRPC4 channel inhibitor. A high throughput fluorescent screen of 305,000 compounds of the Molecular Libraries Small Molecule Repository was performed for inhibitors that blocked intracellular Ca2+ rise in response to stimulation of mouse TRPC4β by μ-opioid receptors. ML204 inhibited TRPC4β-mediated intracellular Ca2+ rise with an IC50 value of 0.96 μm and exhibited 19-fold selectivity against muscarinic receptor-coupled TRPC6 channel activation. In whole-cell patch clamp recordings, ML204 blocked TRPC4β currents activated through either μ-opioid receptor stimulation or intracellular dialysis of guanosine 5′-3-O-(thio)triphosphate (GTPγS), suggesting a direct interaction of ML204 with TRPC4 channels rather than any interference with the signal transduction pathways. Selectivity studies showed no appreciable block by 10–20 μm ML204 of TRPV1, TRPV3, TRPA1, and TRPM8, as well as KCNQ2 and native voltage-gated sodium, potassium, and calcium channels in mouse dorsal root ganglion neurons. In isolated guinea pig ileal myocytes, ML204 blocked muscarinic cation currents activated by bath application of carbachol or intracellular infusion of GTPγS, demonstrating its effectiveness on native TRPC4 currents. Therefore, ML204 represents an excellent novel tool for investigation of TRPC4 channel function and may facilitate the development of therapeutics targeted to TRPC4. PMID:21795696

  4. The Subfamily-specific Assembly of Eag and Erg K+ Channels Is Determined by Both the Amino and the Carboxyl Recognition Domains*

    PubMed Central

    Lin, Ting-Feng; Lin, I-Wen; Chen, Shu-Ching; Wu, Hao-Han; Yang, Chi-Sheng; Fang, Hsin-Yu; Chiu, Mei-Miao; Jeng, Chung-Jiuan

    2014-01-01

    A functional voltage-gated K+ (Kv) channel comprises four pore-forming α-subunits, and only members of the same Kv channel subfamily may co-assemble to form heterotetramers. The ether-à-go-go family of Kv channels (KCNH) encompasses three distinct subfamilies: Eag (Kv10), Erg (Kv11), and Elk (Kv12). Members of different ether-à-go-go subfamilies, such as Eag and Erg, fail to form heterotetramers. Although a short stretch of amino acid sequences in the distal C-terminal section has been implicated in subfamily-specific subunit assembly, it remains unclear whether this region serves as the sole and/or principal subfamily recognition domain for Eag and Erg. Here we aim to ascertain the structural basis underlying the subfamily specificity of ether-à-go-go channels by generating various chimeric constructs between rat Eag1 and human Erg subunits. Biochemical and electrophysiological characterizations of the subunit interaction properties of a series of different chimeric and truncation constructs over the C terminus suggested that the putative C-terminal recognition domain is dispensable for subfamily-specific assembly. Further chimeric analyses over the N terminus revealed that the N-terminal region may also harbor a subfamily recognition domain. Importantly, exchanging either the N-terminal or the C-terminal domain alone led to a virtual loss of the intersubfamily assembly boundary. By contrast, simultaneously swapping both recognition domains resulted in a reversal of subfamily specificity. Our observations are consistent with the notion that both the N-terminal and the C-terminal recognition domains are required to sustain the subfamily-specific assembly of rat Eag1 and human Erg. PMID:25008323

  5. The subfamily-specific assembly of Eag and Erg K+ channels is determined by both the amino and the carboxyl recognition domains.

    PubMed

    Lin, Ting-Feng; Lin, I-Wen; Chen, Shu-Ching; Wu, Hao-Han; Yang, Chi-Sheng; Fang, Hsin-Yu; Chiu, Mei-Miao; Jeng, Chung-Jiuan

    2014-08-15

    A functional voltage-gated K(+) (Kv) channel comprises four pore-forming α-subunits, and only members of the same Kv channel subfamily may co-assemble to form heterotetramers. The ether-à-go-go family of Kv channels (KCNH) encompasses three distinct subfamilies: Eag (Kv10), Erg (Kv11), and Elk (Kv12). Members of different ether-à-go-go subfamilies, such as Eag and Erg, fail to form heterotetramers. Although a short stretch of amino acid sequences in the distal C-terminal section has been implicated in subfamily-specific subunit assembly, it remains unclear whether this region serves as the sole and/or principal subfamily recognition domain for Eag and Erg. Here we aim to ascertain the structural basis underlying the subfamily specificity of ether-à-go-go channels by generating various chimeric constructs between rat Eag1 and human Erg subunits. Biochemical and electrophysiological characterizations of the subunit interaction properties of a series of different chimeric and truncation constructs over the C terminus suggested that the putative C-terminal recognition domain is dispensable for subfamily-specific assembly. Further chimeric analyses over the N terminus revealed that the N-terminal region may also harbor a subfamily recognition domain. Importantly, exchanging either the N-terminal or the C-terminal domain alone led to a virtual loss of the intersubfamily assembly boundary. By contrast, simultaneously swapping both recognition domains resulted in a reversal of subfamily specificity. Our observations are consistent with the notion that both the N-terminal and the C-terminal recognition domains are required to sustain the subfamily-specific assembly of rat Eag1 and human Erg. PMID:25008323

  6. Role of the central arginine R133 toward the ion selectivity of the phosphate specific channel OprP: effects of charge and solvation.

    PubMed

    Modi, Niraj; Bárcena-Uribarri, Iván; Bains, Manjeet; Benz, Roland; Hancock, Robert E W; Kleinekathöfer, Ulrich

    2013-08-20

    The outer membrane porin OprP of Pseudomonas aeruginosa forms a highly specific phosphate selective channel. This channel is responsible for the high-affinity uptake of phosphate ions into the periplasmic space of the bacteria. A detailed investigation of the structure-function relationship of OprP is inevitable to decipher the anion and phosphate selectivity of this porin in particular and to broaden the present understanding of the ion selectivity of different channels. To this end we investigated the role of the central arginine of OprP, R133, in terms of its effects in selectivity and ion transport properties of the pore. Electrophysiological bilayer measurements and free-energy molecular dynamics simulations were carried out to probe the transport of different ions through various R133 mutants. For these mutants, the change in phosphate binding specificity, ion conduction, and anion selectivity was determined and compared to previous molecular dynamic calculations and electrophysiological measurements with wild-type OprP. Molecular analysis revealed a rather particular role of arginine 133 and its charge, while at the same time this residue together with the network of other residues, namely, D94 and Y114, has the ability to dehydrate the permeating ion. These very specific features govern the ion selectivity of OprP. PMID:23875754

  7. Substrate specificity, substrate channeling, and allostery in BphJ: an acylating aldehyde dehydrogenase associated with the pyruvate aldolase BphI.

    PubMed

    Baker, Perrin; Carere, Jason; Seah, Stephen Y K

    2012-06-01

    BphJ, a nonphosphorylating acylating aldehyde dehydrogenase, catalyzes the conversion of aldehydes to form acyl-coenzyme A in the presence of NAD(+) and coenzyme A (CoA). The enzyme is structurally related to the nonacylating aldehyde dehydrogenases, aspartate-β-semialdehyde dehydrogenase and phosphorylating glyceraldehyde-3-phosphate dehydrogenase. Cys-131 was identified as the catalytic thiol in BphJ, and pH profiles together with site-specific mutagenesis data demonstrated that the catalytic thiol is not activated by an aspartate residue, as previously proposed. In contrast to the wild-type enzyme that had similar specificities for two- or three-carbon aldehydes, an I195A variant was observed to have a 20-fold higher catalytic efficiency for butyraldehyde and pentaldehyde compared to the catalytic efficiency of the wild type toward its natural substrate, acetaldehyde. BphJ forms a heterotetrameric complex with the class II aldolase BphI that channels aldehydes produced in the aldol cleavage reaction to the dehydrogenase via a molecular tunnel. Replacement of Ile-171 and Ile-195 with bulkier amino acid residues resulted in no more than a 35% reduction in acetaldehyde channeling efficiency, showing that these residues are not critical in gating the exit of the channel. Likewise, the replacement of Asn-170 in BphJ with alanine and aspartate did not substantially alter aldehyde channeling efficiencies. Levels of activation of BphI by BphJ N170A, N170D, and I171A were reduced by ≥3-fold in the presence of NADH and ≥4.5-fold when BphJ was undergoing turnover, indicating that allosteric activation of the aldolase has been compromised in these variants. The results demonstrate that the dehydrogenase coordinates the catalytic activity of BphI through allostery rather than through aldehyde channeling. PMID:22574886

  8. Site-specific fluorescence spectrum detection and characterization of hASIC1a channels upon toxin mambalgin-1 binding in live mammalian cells.

    PubMed

    Wen, Ming; Guo, Xiaoqi; Sun, Peibei; Xiao, Liang; Li, Juan; Xiong, Ying; Bao, Jin; Xue, Tian; Zhang, Longhua; Tian, Changlin

    2015-05-11

    The synthesis of fluorescent unnatural amino-acid Anap was optimized and the Anap was incorporated into four sites in an acid-pocket or a transmembrane region of human acid-sensing ion channel-1a (hASIC1a). Combinational Anap fluorescence spectra and patch-clamp electrophysiology data illustrated site-specific conformational responses upon toxin mambalgin-1 binding. This combinational approach can be used to analyse conformational properties of many different eukaryotic proteins in their functional states, in a site-specific manner in live mammalian cells. PMID:25873388

  9. Second coiled-coil domain of KCNQ channel controls current expression and subfamily specific heteromultimerization by salt bridge networks.

    PubMed

    Nakajo, Koichi; Kubo, Yoshihiro

    2008-06-15

    KCNQ channels carry the slowly activating, voltage-dependent M-current in excitable cells such as neurons. Although the KCNQ2 homomultimer can form a functional voltage-gated K(+) channel, heteromultimerization with KCNQ3 produces a > 10-fold increase in current amplitude. All KCNQ channels contain double coiled-coil domains (TCC1 and TCC2, or A-domain Head and Tail), of which TCC2 (A-domain Tail) is thought to be important for subunit recognition, channel assembly and surface expression. The mechanism by which TCC2 recognizes and associates with its partner is not fully understood, however. Our aim in the present study was to elucidate the recognition mechanism by examining the phenotypes of TCC2-deletion mutants, TCC2-swapped chimeras and point mutants. Electrophysiological analysis using Xenopus oocytes under two-electrode voltage clamp revealed that homotetrameric KCNQ3 TCC2 is a negative regulator of current expression in the absence of KCNQ2 TCC2. Recent structural analysis of KCNQ4 TCC2 revealed the presence of intercoil salt bridge networks. We therefore swapped the sign of the charged residues reportedly involved in the salt bridge formation and functionally confirmed that the intercoil salt bridge network is responsible for the subunit recognition between KCNQ2 and KCNQ3. Finally, we constructed TCC2-swapped KCNQ2/KCNQ3 mutants with KCNQ1 TCC2 or GCN4-pLI, a coiled-coil domain from an unrelated protein, and found that TCC2 is substitutable and even GCN4-pLI can work as a substitute for TCC2. Our present data provide some new insights into the role played by TCC2 during current expression, and also provide functional evidence of the importance of the intercoil salt bridge network for subunit recognition and coiled-coil formation, as is suggested by recent crystallographic data. PMID:18440995

  10. CaV1.3 as pacemaker channels in adrenal chromaffin cells: specific role on exo- and endocytosis?

    PubMed

    Comunanza, Valentina; Marcantoni, Andrea; Vandael, David H; Mahapatra, Satyajit; Gavello, Daniela; Carabelli, Valentina; Carbone, Emilio

    2010-01-01

    Voltage-gated L-type calcium channels (LTCCs) are expressed in adrenal chromaffin cells. Besides shaping the action potential (AP), LTCCs are involved in the excitation-secretion coupling controlling catecholamine release and in Ca (2+) -dependent vesicle retrieval. Of the two LTCCs expressed in chromaffin cells (CaV1.2 and CaV1.3), CaV1.3 possesses the prerequisites for pacemaking spontaneously firing cells: low-threshold, steep voltage-dependence of activation and slow inactivation. By using CaV1 .3 (-/-) KO mice and the AP-clamp it has been possible to resolve the time course of CaV1.3 pacemaker currents, which is similar to that regulating substantia nigra dopaminergic neurons. In mouse chromaffin cells CaV1.3 is coupled to fast-inactivating BK channels within membrane nanodomains and controls AP repolarization. The ability to carry subthreshold Ca (2+) currents and activate BK channels confers to CaV1.3 the unique feature of driving Ca (2+) loading during long interspike intervals and, possibly, to control the Ca (2+) -dependent exocytosis and endocytosis processes that regulate catecholamine secretion and vesicle recycling. PMID:21084859

  11. Sensitivity and specificity of simultaneously acquired (dual channel) radiogallium and Tc-99m-HDP in painful hip and knee prosthetic joints

    SciTech Connect

    Skarzynski, J.J.; Sziklas, J.J.; Rosenberg, R.J.; Rich, D.A.; Spencer, R.P.

    1985-05-01

    Differentiation of prosthetic loosening from infection, by use of sequential bone and radiogallium imaging, has been discussed in the literature. The authors investigated simultaneous (2 channel) imaging of Ga-67 and Tc-99m-HDP in multiviews, in order to assess the parameter of Tc-99m-Ga-67 incongruity. Acquisition of data was carried out 2 days after 5 mCi of Ga-67 citrate IV and 2 hours after 8 mCi of Tc-99m-HDP. Dual data channels were used to insure perfect superimposition of the images and to reduce total imaging time. Normalized bone images were taken, then subtracted from those of Ga-67, by means of progressive weighting factors. A total of 68 studies were carried out on 43 patients. Exams involved both knee and hip prostheses, in population with 63% of the patients over age 60 years. Time from placement of the prosthesis to the dual radionuclide exam was within 2 years in 48% and within 5 years in 78%. Sensitivity was 0.88 and specificity 0.89. Using information on the follow-up dual channel studies, 40/43 cases were correctly identified (93%). Dual channel radionuclide imaging offers a readily available and accurate means of differentiating infection from loosening of hip or knee prostheses.

  12. Dependence of spontaneous electrical activity and basal prolactin release on nonselective cation channels in pituitary lactotrophs.

    PubMed

    Kučka, M; Kretschmannová, K; Stojilkovic, S S; Zemková, H; Tomić, M

    2012-01-01

    All secretory anterior pituitary cells fire action potentials spontaneously and exhibit a high resting cation conductance, but the channels involved in the background permeability have not been identified. In cultured lactotrophs and immortalized GH(3) cells, replacement of extracellular Na(+) with large organic cations, but not blockade of voltage-gated Na(+) influx, led to an instantaneous hyperpolarization of cell membranes that was associated with a cessation of spontaneous firing. When cells were clamped at -50 mV, which was close to the resting membrane potential in these cells, replacement of bath Na(+) with organic cations resulted in an outward-like current, reflecting an inhibition of the inward holding membrane current and indicating loss of a background-depolarizing conductance. Quantitative RT-PCR analysis revealed the high expression of mRNA transcripts for TRPC1 and much lower expression of TRPC6 in both lactotrophs and GH(3) cells. Very low expression of TRPC3, TRPC4, and TRPC5 mRNA transcripts were also present in pituitary but not GH(3) cells. 2-APB and SKF-96365, relatively selective blockers of TRPC channels, inhibited electrical activity, Ca(2+) influx and prolactin release in a concentration-dependent manner. Gd(3+), a common Ca(2+) channel blocker, and flufenamic acid, an inhibitor of non-selective cation channels, also inhibited electrical activity, Ca(2+) influx and prolactin release. These results indicate that nonselective cation channels, presumably belonging to the TRPC family, contribute to the background depolarizing conductance and firing of action potentials with consequent contribution to Ca(2+) influx and hormone release in lactotrophs and GH(3) cells. PMID:22480423

  13. Decreased expression of transient receptor potential channels in cerebral vascular tissue from patients after hypertensive intracerebral hemorrhage.

    PubMed

    Thilo, Florian; Suess, Olaf; Liu, Ying; Tepel, Martin

    2011-01-01

    Recent data indicate that transient receptor potential (TRP) cation channels play an important role in hypertension. Now, we tested the hypothesis that TRP expression is altered in human cerebral vascular tissue in patients who had experienced hypertensive intracerebral hemorrhage. TRPC1, TRPC3, TRPC5, TRPC6, TRPM4, TRPM6, and TRPM7 channels were detected in cerebral vascular tissue by quantitative real-time RT-PCR. Control cerebral vascular tissue was obtained from normotensive patients who underwent neurosurgical operation because of brain tumor. To examine a possible relation between the expression of TRP expression and hypoxic conditions caused by the intracerebral bleeding, we examined the expression of hypoxia inducible factor 1a (HIF1a). Transcripts of TRPC3, TRPC5, TRPM6, and HIF1a were significantly reduced in cerebral vascular tissue from patients after hypertensive intracerebral hemorrhage compared to controls. TRPC3 mRNA correlated well with the expression of HIF1a mRNA (r(2) = 0.59; p = 0.01). TRPC3 expression is associated with hypertension and hypoxic conditions in human cerebral vascular tissue. PMID:21957871

  14. Site- and kinase-specific phosphorylation-mediated activation of SLAC1, a guard cell anion channel stimulated by abscisic acid.

    PubMed

    Maierhofer, Tobias; Diekmann, Marion; Offenborn, Jan Niklas; Lind, Christof; Bauer, Hubert; Hashimoto, Kenji; S Al-Rasheid, Khaled A; Luan, Sheng; Kudla, Jörg; Geiger, Dietmar; Hedrich, Rainer

    2014-09-01

    Under drought stress, abscisic acid (ABA) triggers closure of leaf cell pores called stomata, which are formed by two specialized cells called guard cells in plant epidermis. Two pathways downstream of ABA stimulate phosphorylation of the S-type anion channels SLAC1 (slow anion channel associated 1) and SLAH3 (SLAC1 homolog 3), which causes these channels to open, reducing guard cell volume and triggering stomatal closure. One branch involves OST1 (open stomata 1), a calcium-independent SnRK2-type kinase, and the other branch involves calcium-dependent protein kinases of the CPK (calcium-dependent protein kinase) family. We used coexpression analyses in Xenopus oocytes to show that the calcineurin B-like (CBL) calcium sensors CBL1 and CBL9 and their interacting protein kinase CIPK23 also triggered SLAC1 and SLAH3 opening. We analyzed whether regulation of SLAC1 opening by these different families of kinases involved the same or different sites on SLAC1 by measuring channel conductance of SLAC1 with mutations in the putative phosphorylation sites in the amino or carboxyl termini coexpressed with specific kinases in Xenopus oocytes. SLAC1 mutants lacking the OST1-phosphorylated site were still activated by CPK or by CBL/CIPK complexes. Phosphorylation and activation of SLAC1 by any of the kinases were inhibited by the phosphatase ABI1 (ABA insensitive 1), which is inactivated in response to ABA signaling. These findings identified CBL/CIPK complexes as potential regulators of stomatal aperture through S-type anion channels and indicated that phosphorylation at distinct sites enables SLAC1 activation by both calcium-dependent and calcium-independent pathways downstream of ABA. PMID:25205850

  15. Catalytic mechanism and substrate specificity of the β-subunit of the voltage-gated potassium (Kv) channel

    PubMed Central

    Tipparaju, Srinivas M.; Barski, Oleg A.; Srivastava, Sanjay; Bhatnagar, Aruni

    2008-01-01

    The β-subunits of voltage-gated potassium (Kv) channels are members of aldo-keto reductase (AKR) superfamily. These proteins regulate inactivation and membrane localization of Kv1 and Kv4 channels. The Kvβ proteins bind to pyridine nucleotides with high affinity; however, their catalytic properties remain unclear. Here we report that recombinant rat Kvβ2 catalyzes the reduction of a wide range of aldehydes and ketones. The rate of catalysis was slower (0.06 to 0.2 min−1) than that of other AKRs, but displayed the expected hyperbolic dependence on substrate concentration, with no evidence of allosteric cooperativity. Catalysis was prevented by site-directed substitution of Tyr-90 with phenylalanine, indicating that the acid-base catalytic residue, identified in other AKRs, has a conserved function in Kvβ2. The protein catalyzed the reduction of a broad range of carbonyls including aromatic carbonyls, electrophilic aldehydes and prostaglandins, phospholipid and sugar aldehydes. Little or no activity was detected with carbonyl steroids. Initial velocity profiles were consistent with an ordered bi-bi rapid-equilibrium mechanism in which NADPH binding precedes carbonyl binding. Significant primary kinetic isotope effects (2.0 – 3.1) were observed under single and multiple turnover conditions, indicating that the bond-breaking chemical step is rate-limiting. Structure-activity relationships with a series of para-substituted benzaldehydes indicated that the electronic interactions predominate during substrate binding and that no significant charge develops during the transition state. These data strengthen the view that Kvβ proteins are catalytically-active AKRs that impart redox-sensitivity to Kv channels. PMID:18672894

  16. Subtype-specific Modulation of Acid-sensing Ion Channel (ASIC) Function by 2-Guanidine-4-methylquinazoline*

    PubMed Central

    Alijevic, Omar; Kellenberger, Stephan

    2012-01-01

    Acid-sensing ion channels (ASICs) are neuronal Na+-selective channels that are transiently activated by extracellular acidification. ASICs are involved in fear and anxiety, learning, neurodegeneration after ischemic stroke, and pain sensation. The small molecule 2-guanidine-4-methylquinazoline (GMQ) was recently shown to open ASIC3 at physiological pH. We have investigated the mechanisms underlying this effect and the possibility that GMQ may alter the function of other ASICs besides ASIC3. GMQ shifts the pH dependence of activation to more acidic pH in ASIC1a and ASIC1b, whereas in ASIC3 this shift goes in the opposite direction and is accompanied by a decrease in its steepness. GMQ also induces an acidic shift of the pH dependence of inactivation of ASIC1a, -1b, -2a, and -3. As a consequence, the activation and inactivation curves of ASIC3 but not other ASICs overlap in the presence of GMQ at pH 7.4, thereby creating a window current. At concentrations >1 mm, GMQ decreases maximal peak currents by reducing the unitary current amplitude. Mutation of residue Glu-79 in the palm domain of ASIC3, previously shown to be critical for channel opening by GMQ, disrupted the GMQ effects on inactivation but not activation. This suggests that this residue is involved in the consequences of GMQ binding rather than in the binding interaction itself. This study describes the mechanisms underlying the effects of a novel class of ligands that modulate the function of all ASICs as well as activate ASIC3 at physiological pH. PMID:22948146

  17. Three-dimensional solution structure of mu-conotoxin GIIIB, a specific blocker of skeletal muscle sodium channels.

    PubMed

    Hill, J M; Alewood, P F; Craik, D J

    1996-07-01

    The three-dimensional solution structure of mu-conotoxin GIIIB, a 22-residue polypeptide from the venom of the piscivorous cone snail Conus geographus, has been determined using 2D 1H NMR spectroscopy. GIIIB binds with high affinity and selectivity to skeletal muscle sodium channels and is a valuable tool for characterizing both the structure and function of these channels. Structural restraints consisting of 289 interproton distances inferred from NOEs and 9 backbone and 5 side chain dihedral angle restraints from spin-spin coupling constants were used as input for simulated annealing calculations and energy minimization in the program X-PLOR. In addition to the 1H NMR derived information, the 13C resonances of GIIIB were assigned at natural abundance, and hydroxyproline C beta and C gamma chemical shifts were used to distinguish between the cis and trans peptide bond conformations. The final set of 20 structures had mean pairwise rms differences over the whole molecule of 1.22 A for the backbone atoms and 2.48 A for all heavy atoms. For the well-defined region encompassing residues 3-21, the corresponding values were 0.74 and 2.54 A, respectively. GIIIB adopts a compact structure consisting of a distorted 310-helix, a small beta-hairpin, a cis-hydroxyproline, and several turns. The molecule is stabilized by three disulfide bonds, two of which connect the helix and the beta-sheet, forming a structural core with similarities to the CS alpha beta motif [Cornet, B., Bonmatin, J.-M., Hetru, C., Hoffmann, J. A., Ptak, M., & Vovelle, F. (1995) Structure 3, 435-448]. This motif is common to several families of small proteins including scorpion toxins and insect defensins. Other structural features of GIIIB include the presence of eight arginine and lysine side chains that project into the solvent in a radial orientation relative to the core of the molecule. These cationic side chains form potential sites of interaction with anionic sites on sodium channels. The global

  18. Expression in Pichia pastoris and characterization of APETx2, a specific inhibitor of acid sensing ion channel 3.

    PubMed

    Anangi, Raveendra; Chen, Chih-Cheng; Lin, Yi-Wen; Cheng, Yuan-Ren; Cheng, Chun-Ho; Chen, Yi-Chun; Chu, Yuan-Ping; Chuang, Woei-Jer

    2010-12-01

    Acid sensing ion channels (ASICs) are family of proteins predominantly present in the central and peripheral nervous system. They are known to play important roles in the pathophysiology of pain and ischemic stroke. APETx2 is a potent and selective inhibitor of ASIC3-containing channels and was isolated from sea anemone Anthopleura elegantissima. To facilitate the study on the molecular determinants of ASIC3-ligand interactions, we expressed recombinant APETx2 in the Pichia pastoris (P. pastoris) expression system and purified it to homogeneity. Recombinant APETx2 produced in P. pastoris inhibited the acid-evoked ASIC3 current with the IC(50) value of 37.3 nM. The potency of recombinant toxin is similar to that of native APETx2. The sequential assignment and structure analysis of APETx2 were obtained by 2D and 3D (15)N-edited NMR spectra. Our NMR data suggests that APETx2 produced in P. pastoris retained its native fold. The results presented here provide the first direct evidence that highly disulfide bonded peptide inhibitor of ASIC3, APETx2, can be expressed in P. pastoris with correct fold and high yield. We also showed that the R17A mutant exhibited a decrease in activity, suggesting the feasibility of the use of this expression system to study the interactions between APETx2 and ASIC3. These evidences may serve as the basis for understanding the selectivity and activity of APETx2. PMID:20813121

  19. Subunit composition of VRAC channels determines substrate specificity and cellular resistance to Pt-based anti-cancer drugs.

    PubMed

    Planells-Cases, Rosa; Lutter, Darius; Guyader, Charlotte; Gerhards, Nora M; Ullrich, Florian; Elger, Deborah A; Kucukosmanoglu, Asli; Xu, Guotai; Voss, Felizia K; Reincke, S Momsen; Stauber, Tobias; Blomen, Vincent A; Vis, Daniel J; Wessels, Lodewyk F; Brummelkamp, Thijn R; Borst, Piet; Rottenberg, Sven; Jentsch, Thomas J

    2015-12-14

    Although platinum-based drugs are widely used chemotherapeutics for cancer treatment, the determinants of tumor cell responsiveness remain poorly understood. We show that the loss of subunits LRRC8A and LRRC8D of the heteromeric LRRC8 volume-regulated anion channels (VRACs) increased resistance to clinically relevant cisplatin/carboplatin concentrations. Under isotonic conditions, about 50% of cisplatin uptake depended on LRRC8A and LRRC8D, but neither on LRRC8C nor on LRRC8E. Cell swelling strongly enhanced LRRC8-dependent cisplatin uptake, bolstering the notion that cisplatin enters cells through VRAC. LRRC8A disruption also suppressed drug-induced apoptosis independently from drug uptake, possibly by impairing VRAC-dependent apoptotic cell volume decrease. Hence, by mediating cisplatin uptake and facilitating apoptosis, VRAC plays a dual role in the cellular drug response. Incorporation of the LRRC8D subunit into VRAC substantially increased its permeability for cisplatin and the cellular osmolyte taurine, indicating that LRRC8 proteins form the channel pore. Our work suggests that LRRC8D-containing VRACs are crucial for cell volume regulation by an important organic osmolyte and may influence cisplatin/carboplatin responsiveness of tumors. PMID:26530471

  20. A process evaluation: Does recruitment for an exercise program through ethnically specific channels and key figures contribute to its reach and receptivity in ethnic minority mothers?

    PubMed Central

    2013-01-01

    Background Ethnic minority women from low-income countries who live in high-income countries are more physically inactive than ethnic majority women in those countries. At the same time, they can be harder to reach with health promotion programs. Targeting recruitment channels and execution to ethnic groups could increase reach and receptivity to program participation. We explored using ethnically specific channels and key figures to reach Ghanaian, Antillean, and Surinamese mothers with an invitation for an exercise program, and subsequently, to determine the mothers’ receptivity and participation. Methods We conducted a mixed methods process evaluation in Amsterdam, the Netherlands. To recruit mothers, we employed ethnically specific community organizations and ethnically matched key figures as recruiters over Dutch health educators. Reach and participation were measured using reply cards and the attendance records from the exercise programs. Observations were made of the recruitment process. We interviewed 14 key figures and 32 mothers to respond to the recruitment channel and recruiter used. Content analysis was used to analyze qualitative data. Results Recruitment through ethnically specific community channels was successful among Ghanaian mothers, but less so among Antillean and Surinamese mothers. The more close-knit an ethnic community was, retaining their own culture and having poorer comprehension of the Dutch language, the more likely we were to reach mothers through ethnically specific organizations. Furthermore, we found that using ethnically matched recruiters resulted in higher receptivity to the program and, among the Ghanaian mothers in particular, in greater participation. This was because the ethnically matched recruiter was a familiar, trusted person, a translator, and a motivator who was enthusiastic, encouraging, and able to adapt her message (targeting/tailoring). Using a health expert was preferred in order to increase the credibility and

  1. Rat hypocretin/orexin neurons are maintained in a depolarized state by TRPC channels.

    PubMed

    Cvetkovic-Lopes, Vesna; Eggermann, Emmanuel; Uschakov, Aaron; Grivel, Jeremy; Bayer, Laurence; Jones, Barbara E; Serafin, Mauro; Mühlethaler, Michel

    2010-01-01

    In a previous study we proposed that the depolarized state of the wake-promoting hypocretin/orexin (hcrt/orx) neurons was independent of synaptic inputs as it persisted in tetrodotoxin and low calcium/high magnesium solutions. Here we show first that these cells are hyperpolarized when external sodium is lowered, suggesting that non-selective cation channels (NSCCs) could be involved. As canonical transient receptor channels (TRPCs) are known to form NSCCs, we looked for TRPCs subunits using single-cell RT-PCR and found that TRPC6 mRNA was detectable in a small minority, TRPC1, TRPC3 and TRPC7 in a majority and TRPC4 and 5 in the vast majority (∼90%) of hcrt/orx neurons. Using intracellular applications of TRPC antibodies against subunits known to form NSCCs, we then found that only TRPC5 antibodies elicited an outward current, together with hyperpolarization and inhibition of the cells. These effects were blocked by co-application of a TRPC5 antigen peptide. Voltage-clamp ramps in the presence or absence of TRPC5 antibodies indicated the presence of a current with a reversal potential close to -15 mV. Application of the non-selective TRPC channel blocker, flufenamic acid, had a similar effect, which could be occluded in cells pre-loaded with TRPC5 antibodies. Finally, using the same TRPC5 antibodies we found that most hcrt/orx cells show immunostaining for the TRPC5 subunit. These results suggest that hcrt/orx neurons are endowed with a constitutively active non-selective cation current which depends on TRPC channels containing the TRPC5 subunit and which is responsible for the depolarized and active state of these cells. PMID:21179559

  2. Identification of the Sensory Neuron Specific Regulatory Region for the Mouse Gene Encoding the Voltage Gated Sodium Channel Nav1.8

    PubMed Central

    Puhl, Henry L.; Ikeda, Stephen R.

    2008-01-01

    Voltage-gated sodium channels (VGSC) are critical membrane components that participate in the electrical activity of excitable cells. The type one VGSC family includes the tetrodotoxin insensitive sodium channel, Nav1.8, encoded by the Scn10a gene. Nav1.8 expression is restricted to small and medium diameter nociceptive sensory neurons of the dorsal root (DRG) and cranial sensory ganglia. In order to understand the stringent transcriptional regulation of the Scn10a gene, the sensory neuron specific promoter was functionally identified. While identifying the mRNA 5’ end, alternative splicing within the 5’ UTR was observed to create heterogeneity in the RNA transcript. Four kilobases of upstream genomic DNA was cloned and the presence of tissue specific promoter activity was tested by microinjection and adenoviral infection of fluorescent protein reporter constructs into primary mouse and rat neurons, and cell lines. The region contained many putative transcription factor binding sites and strong homology with the predicted rat ortholog. Homology to the predicted human ortholog was limited to the proximal end and several conserved cis elements were noted. Two regulatory modules were identified by microinjection of reporter constructs into DRG and superior cervical ganglia neurons: a neuron specific proximal promoter region between −1.6 and −0.2kb of the transcription start site cluster, and a distal sensory neuron switch region beyond −1.6kb that restricted fluorescent protein expression to a subset of primary sensory neurons. PMID:18466327

  3. Role of nonselective cation channels in spontaneous and protein kinase A-stimulated calcium signaling in pituitary cells.

    PubMed

    Tomić, Melanija; Kucka, Marek; Kretschmannova, Karla; Li, Shuo; Nesterova, Maria; Stratakis, Constantine A; Stojilkovic, Stanko S

    2011-08-01

    Several receptors linked to the adenylyl cyclase signaling pathway stimulate electrical activity and calcium influx in endocrine pituitary cells, and a role for an unidentified sodium-conducting channel in this process has been proposed. Here we show that forskolin dose-dependently increases cAMP production and facilitates calcium influx in about 30% of rat and mouse pituitary cells at its maximal concentration. The stimulatory effect of forskolin on calcium influx was lost in cells with inhibited PKA (cAMP-dependent protein kinase) and in cells that were haploinsufficient for the main PKA regulatory subunit but was preserved in cells that were also haploinsufficient for the main PKA catalytic subunit. Spontaneous and forskolin-stimulated calcium influx was present in cells with inhibited voltage-gated sodium and hyperpolarization-activated cation channels but not in cells bathed in medium, in which sodium was replaced with organic cations. Consistent with the role of sodium-conducting nonselective cation channels in PKA-stimulated Ca(2+) influx, cAMP induced a slowly developing current with a reversal potential of about 0 mV. Two TRP (transient receptor potential) channel blockers, SKF96365 and 2-APB, as well as flufenamic acid, an inhibitor of nonselective cation channels, also inhibited spontaneous and forskolin-stimulated electrical activity and calcium influx. Quantitative RT-PCR analysis indicated the expression of mRNA transcripts for TRPC1 > TRPC6 > TRPC4 > TRPC5 > TRPC3 in rat pituitary cells. These experiments suggest that in pituitary cells constitutively active cation channels are stimulated further by PKA and contribute to calcium signaling indirectly by controlling the pacemaking depolarization in a sodium-dependent manner and directly by conducting calcium. PMID:21586701

  4. Isoform-specific modulation of the chemical sensitivity of conserved TRPA1 channel in the major honeybee ectoparasitic mite, Tropilaelaps mercedesae

    PubMed Central

    Dong, Xiaofeng; Kashio, Makiko; Peng, Guangda; Wang, Xinyue; Tominaga, Makoto

    2016-01-01

    We identified and characterized the TRPA1 channel of Tropilaelaps mercedesae (TmTRPA1), one of two major species of honeybee ectoparasitic mite. Three TmTRPA1 isoforms with unique N-terminal sequences were activated by heat, and the isoform highly expressed in the mite's front legs, TmTRPA1b, was also activated by 27 plant-derived compounds including electrophiles. This suggests that the heat- and electrophile-dependent gating mechanisms as nocisensitive TRPA1 channel are well conserved between arthropod species. Intriguingly, one TmTRPA1 isoform, TmTRPA1a, was activated by only six compounds compared with two other isoforms, demonstrating that the N-terminal sequences are critical determinants for the chemical sensitivity. This is the first example of isoform-specific modulation of chemical sensitivity of TRPA1 channel in one species. α-terpineol showed repellent activity towards T. mercedesae in a laboratory assay and repressed T. mercedesae entry for reproduction into the brood cells with fifth instar larvae in hives. Thus, α-terpineol could be used as the potential compound to control two major honeybee ectoparasitic mites, T. mercedesae and Varroa destructor, in the apiculture industry. PMID:27307515

  5. CPK13, a Noncanonical Ca2+-Dependent Protein Kinase, Specifically Inhibits KAT2 and KAT1 Shaker K+ Channels and Reduces Stomatal Opening1[C][W

    PubMed Central

    Ronzier, Elsa; Corratgé-Faillie, Claire; Sanchez, Frédéric; Prado, Karine; Brière, Christian; Leonhardt, Nathalie; Thibaud, Jean-Baptiste; Xiong, Tou Cheu

    2014-01-01

    Ca2+-dependent protein kinases (CPKs) form a large family of 34 genes in Arabidopsis (Arabidopsis thaliana). Based on their dependence on Ca2+, CPKs can be sorted into three types: strictly Ca2+-dependent CPKs, Ca2+-stimulated CPKs (with a significant basal activity in the absence of Ca2+), and essentially calcium-insensitive CPKs. Here, we report on the third type of CPK, CPK13, which is expressed in guard cells but whose role is still unknown. We confirm the expression of CPK13 in Arabidopsis guard cells, and we show that its overexpression inhibits light-induced stomatal opening. We combine several approaches to identify a guard cell-expressed target. We provide evidence that CPK13 (1) specifically phosphorylates peptide arrays featuring Arabidopsis K+ Channel KAT2 and KAT1 polypeptides, (2) inhibits KAT2 and/or KAT1 when expressed in Xenopus laevis oocytes, and (3) closely interacts in plant cells with KAT2 channels (Förster resonance energy transfer-fluorescence lifetime imaging microscopy). We propose that CPK13 reduces stomatal aperture through its inhibition of the guard cell-expressed KAT2 and KAT1 channels. PMID:25037208

  6. Isoform-specific modulation of the chemical sensitivity of conserved TRPA1 channel in the major honeybee ectoparasitic mite, Tropilaelaps mercedesae.

    PubMed

    Dong, Xiaofeng; Kashio, Makiko; Peng, Guangda; Wang, Xinyue; Tominaga, Makoto; Kadowaki, Tatsuhiko

    2016-06-01

    We identified and characterized the TRPA1 channel of Tropilaelaps mercedesae (TmTRPA1), one of two major species of honeybee ectoparasitic mite. Three TmTRPA1 isoforms with unique N-terminal sequences were activated by heat, and the isoform highly expressed in the mite's front legs, TmTRPA1b, was also activated by 27 plant-derived compounds including electrophiles. This suggests that the heat- and electrophile-dependent gating mechanisms as nocisensitive TRPA1 channel are well conserved between arthropod species. Intriguingly, one TmTRPA1 isoform, TmTRPA1a, was activated by only six compounds compared with two other isoforms, demonstrating that the N-terminal sequences are critical determinants for the chemical sensitivity. This is the first example of isoform-specific modulation of chemical sensitivity of TRPA1 channel in one species. α-terpineol showed repellent activity towards T. mercedesae in a laboratory assay and repressed T. mercedesae entry for reproduction into the brood cells with fifth instar larvae in hives. Thus, α-terpineol could be used as the potential compound to control two major honeybee ectoparasitic mites, T. mercedesae and Varroa destructor, in the apiculture industry. PMID:27307515

  7. CPK13, a noncanonical Ca2+-dependent protein kinase, specifically inhibits KAT2 and KAT1 shaker K+ channels and reduces stomatal opening.

    PubMed

    Ronzier, Elsa; Corratgé-Faillie, Claire; Sanchez, Frédéric; Prado, Karine; Brière, Christian; Leonhardt, Nathalie; Thibaud, Jean-Baptiste; Xiong, Tou Cheu

    2014-09-01

    Ca(2) (+)-dependent protein kinases (CPKs) form a large family of 34 genes in Arabidopsis (Arabidopsis thaliana). Based on their dependence on Ca(2+), CPKs can be sorted into three types: strictly Ca(2+)-dependent CPKs, Ca(2+)-stimulated CPKs (with a significant basal activity in the absence of Ca(2+)), and essentially calcium-insensitive CPKs. Here, we report on the third type of CPK, CPK13, which is expressed in guard cells but whose role is still unknown. We confirm the expression of CPK13 in Arabidopsis guard cells, and we show that its overexpression inhibits light-induced stomatal opening. We combine several approaches to identify a guard cell-expressed target. We provide evidence that CPK13 (1) specifically phosphorylates peptide arrays featuring Arabidopsis K(+) Channel KAT2 and KAT1 polypeptides, (2) inhibits KAT2 and/or KAT1 when expressed in Xenopus laevis oocytes, and (3) closely interacts in plant cells with KAT2 channels (Förster resonance energy transfer-fluorescence lifetime imaging microscopy). We propose that CPK13 reduces stomatal aperture through its inhibition of the guard cell-expressed KAT2 and KAT1 channels. PMID:25037208

  8. Specific functioning of Cav3.2 T-type calcium and TRPV1 channels under different types of STZ-diabetic neuropathy.

    PubMed

    Khomula, Eugen V; Viatchenko-Karpinski, Viacheslav Y; Borisyuk, Anya L; Duzhyy, Dmytro E; Belan, Pavel V; Voitenko, Nana V

    2013-05-01

    Streptozotocin (STZ)-induced type 1 diabetes in rats leads to the development of peripheral diabetic neuropathy (PDN) manifested as thermal hyperalgesia at early stages (4th week) followed by hypoalgesia after 8weeks of diabetes development. Here we found that 6-7 week STZ-diabetic rats developed either thermal hyper- (18%), hypo- (25%) or normalgesic (57%) types of PDN. These developmentally similar diabetic rats were studied in order to analyze mechanisms potentially underlying different thermal nociception. The proportion of IB4-positive capsaicin-sensitive small DRG neurons, strongly involved in thermal nociception, was not altered under different types of PDN implying differential changes at cellular and molecular level. We further focused on properties of T-type calcium and TRPV1 channels, which are known to be involved in Ca(2+) signaling and pathological nociception. Indeed, TRPV1-mediated signaling in these neurons was downregulated under hypo- and normalgesia and upregulated under hyperalgesia. A complex interplay between diabetes-induced changes in functional expression of Cav3.2 T-type calcium channels and depolarizing shift of their steady-state inactivation resulted in upregulation of these channels under hyper- and normalgesia and their downregulation under hypoalgesia. As a result, T-type window current was increased by several times under hyperalgesia partially underlying the increased resting [Ca(2+)]i observed in the hyperalgesic rats. At the same time Cav3.2-dependent Ca(2+) signaling was upregulated in all types of PDN. These findings indicate that alterations in functioning of Cav3.2 T-type and TRPV1 channels, specific for each type of PDN, may underlie the variety of pain syndromes induced by type 1 diabetes. PMID:23376589

  9. Role of an isoform-specific substrate access channel residue in CO ligand accessibilities of neuronal and inducible nitric oxide synthase isoforms.

    PubMed

    Feng, Changjian; Fan, Weihong; Ghosh, Dipak K; Tollin, Gordon

    2011-03-01

    The rates of the bimolecular CO rebinding to the oxygenase domains of inducible and neuronal NOS proteins (iNOSoxy and nNOSoxy, respectively) after photolytic dissociation have been determined by laser flash photolysis. The following mutants at the isoform-specific sites (murine iNOSoxy N115L and rat nNOSoxy L337N, L337F) have been constructed to investigate role of the residues in the CO ligand accessibilities of the NOS isoforms. These residues are in the NOS distal substrate access channel. The effect of the (6R)-5,6,7,8-tetrahydrobiopterin (H(4)B) cofactor and l-arginine (Arg) substrate on the rates of CO rebinding have also been assessed. Addition of l-Arg to the iNOSoxy N115L mutant results in much faster CO rebinding rates, compared to the wild type. The results indicate that modifications to the iNOS channel in which the hydrophilic residue N115 is replaced by leucine (to resemble its nNOS cognate) open the channel somewhat, thereby improving access to the axial heme ligand binding position. On the other hand, introduction of a hydrophilic residue (L337N) or a bulky rigid aromatic residue (L337F) in the nNOS isoform does not significantly affect the kinetics profile, suggesting that the geometry of the substrate access pocket is not greatly altered. The bimolecular CO rebinding rate data indicate that the opening of the substrate access channel in the iNOS N115L mutant may be due to more widespread structural alterations induced by the mutation. PMID:21146639

  10. Organ-Specific Splice Variants of Aquaporin Water Channel AgAQP1 in the Malaria Vector Anopheles gambiae

    PubMed Central

    Tsujimoto, Hitoshi; Liu, Kun; Linser, Paul J.; Agre, Peter; Rasgon, Jason L.

    2013-01-01

    Background Aquaporin (AQP) water channels are important for water homeostasis in all organisms. Malaria transmission is dependent on Anopheles mosquitoes. Water balance is a major factor influencing mosquito survival, which may indirectly affect pathogen transmission. Methodology/Principal Findings We obtained full-length mRNA sequences for Anopheles gambiae aquaporin 1 (AgAQP1) and identified two splice variants for the gene. In vitro expression analysis showed that both variants transported water and were inhibited by Hg2+. One splice variant (AgAQP1A) was exclusively expressed in adult female ovaries indicating a function in mosquito reproduction. The other splice variant (AgAQP1B) was expressed in the midgut, malpighian tubules and the head in adult mosquitoes. Immunolabeling showed that in malpighian tubules, AgAQP1 is expressed in principal cells in the proximal portion and in stellate cells in the distal portion. Moreover, AgAQP1 is expressed in Johnston’s organ (the “ear”), which is important for courtship behavior. Conclusions And Significance These results suggest that AgAQP1 may play roles associated with mating (courtship) and reproduction in addition to water homeostasis in this important African malaria vector. PMID:24066188

  11. Vibratory sources as compound stimuli for the octavolateralis systems: dissection of specific stimulation channels using multiple behavioral approaches.

    PubMed

    Braun, Christopher B; Coombs, Sheryl

    2010-04-01

    Underwater vibratory sources simultaneously present acoustic and hydrodynamic disturbances. Because vibratory dipole sources are poor sonic projectors, most researchers have assumed that such sources are of greatest relevance to the lateral line system. Both hydroacoustic principles and empirical studies have shown that dipole sources are also a potent stimulus to the inner ear. Responses to vibratory sources in mottled sculpin (Cottus bairdi) were assessed using unconditioned orienting, differential and nondifferential conditioning. Orienting responses are dominated by lateral line inputs and eliminated by lateral line inactivation. Simple conditioning depends on inputs from other systems and was not affected by lateral line inactivation. Differential conditioning alters behavioral control, and sculpin could be conditioned to ignore substrate-borne vibrations and respond only to hydroacoustic stimulation of the ear. The lateral line and inner ear of mottled sculpin do not necessarily exhibit range fractionation, as both systems operate over a similar distance (within 1.5 body lengths) and respond to many of the same sources. Vibratory dipole sources generate compound stimuli that simultaneously activate multiple octavolateralis systems, and sculpin make use of the channels differentially under different behavioral tasks. PMID:20384404

  12. Vibratory sources as compound stimuli for the octavolateralis systems: Dissection of specific stimulation channels using multiple behavioral approaches

    PubMed Central

    Braun, Christopher B.; Coombs, Sheryl

    2010-01-01

    An underwater vibratory source simultaneously presents acoustic and hydrodynamic disturbances. Because vibratory dipole sources are poor sonic projectors, most researchers have assumed that such sources are of greatest relevance to the lateral line system (LL). Both hydroacoustic principles and empirical studies have shown that vibratory dipole sources are also a potent stimulus to the inner ear of fishes. Responses to vibratory sources in mottled sculpin (Cottus bairdi) were assessed using unconditioned orienting, differential and non-differential-classical conditioning. Orienting responses are dominated by LL inputs and eliminated by LL pharmacological inactivation. Simple conditioning depends on inputs from other systems and was not affected by LL inactivation. Differential conditioning alters behavioral control, and sculpin could be conditioned to ignore substrate borne vibrations and respond only to hydroacoustic stimulation of the ear. The lateral line and inner ear of mottled sculpin do not necessarily exhibit range fractionation, as both systems operate over a similar distance from the animal (within 1.5 body lengths) and respond to many of the same sources. Vibratory dipole sources generate compound stimuli that simultaneously activate multiple octavolateralis systems, and animals make use of the channels differentially under different behavioral tasks. PMID:20384404

  13. Regulation of the cellular localization and function of human transient receptor potential channel 1 by other members of the TRPC family.

    PubMed

    Alfonso, Salgado; Benito, Ordaz; Alicia, Sampieri; Angélica, Zepeda; Patricia, Glazebrook; Diana, Kunze; Vaca, Luis; Luis, Vaca

    2008-04-01

    Members of the Canonical Transient Receptor Potential (TRPC) family of ionic channels are able to form homo- and heterotetrameric channels. Depending on the study, TRPC1 has been detected on both the surface and inside the cell, probably in the endoplasmic reticulum (ER). Likewise, TRPC1 has been described both as a store-operated channel and as one unable to function when forming a homotetramer. It is possible that the apparent differences in the expression and function of TRPC1 are due to its association with other proteins, possibly from the same TRPC family. In the present study we used confocal microscopy and a fluorescently tagged TRPC1 to examine the localization of this protein when co-expressed with other members of the TRPC family. Whole-cell and single channel electrophysiological recordings were conducted to study the function of TRPC1 expressed alone or co-expressed with other members of the TRPC family. A FRET-based calcium sensor fused to TRPC1 was used to assess the functionality of the intracellular TRPC1. Our results showed that TRPC4 and TRPC5 were able to increase the amount of membrane-expressed TRPC1 as evaluated by confocal microscopy and patch clamp recordings. The FRET-based calcium sensor fused to TRPC1 strongly suggests that this protein forms ER-expressed functional homotetrameric channels activated by agonists coupled to the IP(3) cascade. These results indicate that TRPC1 is a multifunctional protein able to form intracellular calcium release channels when expressed alone, and plasma membrane channels when co-expressed with TRPC4 or TRPC5, but not TRPC3 or TRPC6. Both (ER and plasma membrane) forms of the channel are activated upon addition of agonists coupled to the IP(3) cascade. PMID:17850866

  14. Voltage-gated potassium channels involved in regulation of physiological function in MrgprA3-specific itch neurons.

    PubMed

    Tang, Min; Wu, Guanyi; Wang, Zhongli; Yang, Niuniu; Shi, Hao; He, Qian; Zhu, Chan; Yang, Yan; Yu, Guang; Wang, Changming; Yuan, Xiaolin; Liu, Qin; Guan, Yun; Dong, Xinzhong; Tang, Zongxiang

    2016-04-01

    Itch is described as an unpleasant or irritating skin sensation that elicits the desire or reflex to scratch. MrgprA3, one of members of the Mrgprs family, is specifically expressed in a subpopulation of dorsal root ganglion (DRG) in the peripheral nervous system (PNS). These MrgprA3-expressing DRG neurons have been identified as itch-specific neurons. They can be activated by the compound, chloroquine, which is used as a drug to treat malaria. In the present study, we labeled these itch-specific neurons using the method of molecular genetic markers, and then studied their electrophysiological properties. We also recorded the cutaneous MrgprA3(-) neurons retrogradely labeled by Dil dye (MrgprA3(-)-Dil). We first found that MrgprA3(+) neurons have a lower excitability than MrgprA3(-) neurons (MrgprA3(-)-non-Dil and MrgprA3(-)-Dil). The number of action potential (AP) was reduced more obviously in MrgprA3(+) neurons than that of in MrgprA3(-) neurons. In most cases, MrgprA3(+) neurons only generated single AP; however, in MrgprA3(-) neurons, the same stimulation could induce multiple AP firing due to the greater voltage-gated potassium (Kv) current existence in MrgprA3(+) than in MrgprA3(-) neurons. Thus, Kv current plays an important role in the regulation of excitability in itch-specific neurons. PMID:26874069

  15. Specific interactions between alkali metal cations and the KcsA channel studied using ATR-FTIR spectroscopy

    PubMed Central

    Furutani, Yuji; Shimizu, Hirofumi; Asai, Yusuke; Oiki, Shigetoshi; Kandori, Hideki

    2015-01-01

    The X-ray structure of KcsA, a eubacterial potassium channel, displays a selectivity filter composed of four parallel peptide strands. The backbone carbonyl oxygen atoms of these strands solvate multiple K+ ions. KcsA structures show different distributions of ions within the selectivity filter in solutions containing different cations. To assess the interactions of cations with the selectivity filter, we used attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy. Ion-exchange-induced ATR-FTIR difference spectra were obtained by subtracting the spectrum of KcsA soaked in K+ solution from that obtained in Li+, Na+, Rb+, and Cs+ solutions. Large spectral changes in the amide-I and -II regions were observed upon replacing K+ with smaller-sized cations Li+ and Na+ but not with larger-sized cations Rb+ and Cs+. These results strongly suggest that the selectivity filter carbonyls coordinating Rb+ or Cs+ adopt a conformation similar to those coordinating K+ (cage configuration), but those coordinating Li+ or Na+ adopt a conformation (plane configuration) considerably different from those coordinating K+. We have identified a cation-type sensitive amide-I band at 1681 cm−1 and an insensitive amide-I band at 1659 cm−1. The bands at 1650, 1639, and 1627 cm−1 observed for Na+-coordinating carbonyls were almost identical to those observed in Li+ solution, suggesting that KcsA forms a similar filter structure in Li+ and Na+ solutions. Thus, we conclude that the filter structure adopts a collapsed conformation in Li+ solution that is similar to that in Na+ solution but is in clear contrast to the X-ray crystal structure of KcsA with Li+. PMID:27493853

  16. In pursuit of small molecule chemistry for calcium-permeable non-selective TRPC channels -- mirage or pot of gold?

    PubMed

    Bon, Robin S; Beech, David J

    2013-10-01

    The primary purpose of this review is to address the progress towards small molecule modulators of human Transient Receptor Potential Canonical proteins (TRPC1, TRPC3, TRPC4, TRPC5, TRPC6 and TRPC7). These proteins generate channels for calcium and sodium ion entry. They are relevant to many mammalian cell types including acinar gland cells, adipocytes, astrocytes, cardiac myocytes, cochlea hair cells, endothelial cells, epithelial cells, fibroblasts, hepatocytes, keratinocytes, leukocytes, mast cells, mesangial cells, neurones, osteoblasts, osteoclasts, platelets, podocytes, smooth muscle cells, skeletal muscle and tumour cells. There are broad-ranging positive roles of the channels in cell adhesion, migration, proliferation, survival and turning, vascular permeability, hypertrophy, wound-healing, hypo-adiponectinaemia, angiogenesis, neointimal hyperplasia, oedema, thrombosis, muscle endurance, lung hyper-responsiveness, glomerular filtration, gastrointestinal motility, pancreatitis, seizure, innate fear, motor coordination, saliva secretion, mast cell degranulation, cancer cell drug resistance, survival after myocardial infarction, efferocytosis, hypo-matrix metalloproteinase, vasoconstriction and vasodilatation. Known small molecule stimulators of the channels include hyperforin, genistein and rosiglitazone, but there is more progress with inhibitors, some of which have promising potency and selectivity. The inhibitors include 2-aminoethoxydiphenyl borate, 2-aminoquinolines, 2-aminothiazoles, fatty acids, isothiourea derivatives, naphthalene sulfonamides, N-phenylanthranilic acids, phenylethylimidazoles, piperazine/piperidine analogues, polyphenols, pyrazoles and steroids. A few of these agents are starting to be useful as tools for determining the physiological and pathophysiological functions of TRPC channels. We suggest that the pursuit of small molecule modulators for TRPC channels is important but that it requires substantial additional effort and

  17. Phospholipase cgamma1 is required for activation of store-operated channels in human keratinocytes.

    PubMed

    Tu, Chia-Ling; Chang, Wenhan; Bikle, Daniel D

    2005-01-01

    Store-operated calcium entry depicts the movement of extracellular Ca2+ into cells through plasma membrane Ca2+ channels activated by depletion of intracellular Ca2+ stores. The members of the canonical subfamily of transient receptor potential channels (TRPC) have been implicated as the molecular bases for store-operated channels (SOC). Here we investigate the role of phospholipase C (PLC) in regulation of native SOC and the expression of endogenous TRPC in human epidermal keratinocytes. Calcium entry in response to store depletion with thapsigargin was reversibly blocked by 2-aminoethoxydiphenyl borane, an effective SOC inhibitor, and suppressed by the diacylglycerol analoge, 1-oleoyl-2-acetyl-sn-glycerol. Inhibition of PLC with U73122 or transfection of a PLCgamma1 antisense cDNA construct completely blocked SOC activity, indicating a requirement for PLC, especially PLCgamma1, in the activation of SOC. RT-PCR and immunoblotting analyses showed that TRPC1, TRPC3, TRPC4, TRPC5, and TRPC6 are expressed in keratinocytes. Knockdown of the level of endogenous TRPC1 or TRPC4 inhibited store-operated calcium entry, indicating they are part of the native SOC. Co-immunoprecipitation studies demonstrated that TRPC1, but not TRPC4, interacts with PLCgamma1 and the inositol 1,4,5-trisphosphate receptor (IP3R). The association of TRPC1 with PLCgamma1 and IP3R decreased in keratinocytes with higher intracellular Ca2+, coinciding with a downregulation in SOC activity. Our results indicate that the activation of SOC in keratinocytes depends, at least partly, on the interaction of TRPC with PLCgamma1 and IP3R. PMID:15654973

  18. A family of transcripts encoding water channel proteins: tissue-specific expression in the common ice plant.

    PubMed Central

    Yamada, S; Katsuhara, M; Kelly, W B; Michalowski, C B; Bohnert, H J

    1995-01-01

    Seawater-strength salt stress of the ice plant (Mesembryanthemum crystallinum) initially results in wilting, but full turgor is restored within approximately 2 days. We are interested in a mechanistic explanation for this behavior and, as a requisite for in-depth biochemical studies, have begun to analyze gene expression changes in roots coincident with the onset of stress. cDNAs that suggested changes in mRNA amount under stress were found; their deduced amino acid sequences share homologies with proteins of the Mip (major intrinsic protein) gene family and potentially encode aquaporins. One transcript, MipB, was found only in root RNA, whereas two other transcripts, MipA and MipC, were detected in roots and leaves. Transcript levels of MipB were of low abundance. All transcripts declined initially during salt stress but later recovered to at least prestress level. The most drastic decline was in MipA and MipC transcripts. MipA mRNA distribution in roots detected by in situ hybridization indicated that the transcript was present in all cells in the root tip. In the expansion zone of the root where vascular bundles differentiate, MipA transcript amounts were most abundant in the endodermis. In older roots, which had undergone secondary growth, MipA was highly expressed in cell layers surrounding individual xylem strands. MipA was also localized in leaf vascular tissue and, in lower amounts, in mesophyll cells. Transcripts for MipB seemed to be present exclusively in the tip of the root, in a zone before and possibly coincident with the development of a vascular system. MipA- and MipB-encoded proteins expressed in Xenopus oocytes led to increased water permeability. mRNA fluctuations of the most highly expressed MipA and MipC coincided with turgor changes in leaves under stress. As the leaves regained turgor, transcript levels of these water channel proteins increased. PMID:7549476

  19. Nanosecond photoreduction of cytochrome p450cam by channel-specific Ru-diimine electron tunneling wires.

    PubMed

    Dunn, Alexander R; Dmochowski, Ivan J; Winkler, Jay R; Gray, Harry B

    2003-10-15

    We report the synthesis and characterization of Ru-diimine complexes designed to bind to cytochrome p450cam (CYP101). The sensitizer core has the structure [Ru(L(2))L'](2+), where L' is a perfluorinated biphenyl bridge (F(8)bp) connecting 4,4'-dimethylbipyridine to an enzyme substrate (adamantane, F(8)bp-Ad), a heme ligand (imidazole, F(8)bp-Im), or F (F(9)bp). The electron-transfer (ET) driving force (-deltaG degrees ) is varied by replacing the ancillary 2,2'-bipyridine ligands with 4,4',5,5'-tetramethylbipyridine (tmRu). The four complexes all bind p450cam tightly: Ru-F(8)bp-Ad (1, K(d) = 0.077 microM); Ru-F(8)bp-Im (2, K(d) = 3.7 microM); tmRu-F(9)bp (3, K(d) = 2.1 microM); and tmRu-F(8)bp-Im (4, K(d) = 0.48 microM). Binding is predominantly driven by hydrophobic interactions between the Ru-diimine wires and the substrate access channel. With Ru-F(8)bp wires, redox reactions can be triggered on the nanosecond time scale. Ru-wire 2, which ligates the heme iron, shows a small amount of transient heme photoreduction (ca. 30%), whereas the transient photoreduction yield for 4 is 76%. Forward ET with 4 occurs in roughly 40 ns (k(f) = 2.8 x 10(7) s(-)(1)), and back ET (Fe(II) --> Ru(III), k(b) approximately 1.7 x 10(8) s(-)(1)) is near the coupling-limited rate (k(max)). Direct photoreduction was not observed for 1 or 3. The large variation in ET rates among the Ru-diimine:p450 conjugates strongly supports a through-bond model of Ru-heme electronic coupling. PMID:14531688

  20. Durable pharmacological responses from the peptide ShK-186, a specific Kv1.3 channel inhibitor that suppresses T cell mediators of autoimmune disease.

    PubMed

    Tarcha, Eric J; Chi, Victor; Muñoz-Elías, Ernesto J; Bailey, David; Londono, Luz M; Upadhyay, Sanjeev K; Norton, Kayla; Banks, Amy; Tjong, Indra; Nguyen, Hai; Hu, Xueyou; Ruppert, Greg W; Boley, Scott E; Slauter, Richard; Sams, James; Knapp, Brian; Kentala, Dustin; Hansen, Zachary; Pennington, Michael W; Beeton, Christine; Chandy, K George; Iadonato, Shawn P

    2012-09-01

    The Kv1.3 channel is a recognized target for pharmaceutical development to treat autoimmune diseases and organ rejection. ShK-186, a specific peptide inhibitor of Kv1.3, has shown promise in animal models of multiple sclerosis and rheumatoid arthritis. Here, we describe the pharmacokinetic-pharmacodynamic relationship for ShK-186 in rats and monkeys. The pharmacokinetic profile of ShK-186 was evaluated with a validated high-performance liquid chromatography-tandem mass spectrometry method to measure the peptide's concentration in plasma. These results were compared with single-photon emission computed tomography/computed tomography data collected with an ¹¹¹In-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid-conjugate of ShK-186 to assess whole-blood pharmacokinetic parameters as well as the peptide's absorption, distribution, and excretion. Analysis of these data support a model wherein ShK-186 is absorbed slowly from the injection site, resulting in blood concentrations above the Kv1.3 channel-blocking IC₅₀ value for up to 7 days in monkeys. Pharmacodynamic studies on human peripheral blood mononuclear cells showed that brief exposure to ShK-186 resulted in sustained suppression of cytokine responses and may contribute to prolonged drug effects. In delayed-type hypersensitivity, chronic relapsing-remitting experimental autoimmune encephalomyelitis, and pristane-induced arthritis rat models, a single dose of ShK-186 every 2 to 5 days was as effective as daily administration. ShK-186's slow distribution from the injection site and its long residence time on the Kv1.3 channel contribute to the prolonged therapeutic effect of ShK-186 in animal models of autoimmune disease. PMID:22637724

  1. Binding of the proline-rich region of the epithelial Na+ channel to SH3 domains and its association with specific cellular proteins.

    PubMed Central

    McDonald, F J; Welsh, M J

    1995-01-01

    The amiloride-sensitive epithelial Na+ channel (ENaC) is an important component of the Na(+)-reabsorption pathway in many epithelia. The identification of three subunits of ENaC (alpha, beta and gamma), as well as results from a number of functional and biochemical studies, suggests that functional Na+ channels are composed of a complex of proteins. To learn about possible interactions of the channel with other proteins, we studied the alpha-subunit of rat and human ENaC. We found that the proline-rich C-terminal domains of both rat and human alpha-ENaC, expressed as glutathione S-transferase fusion proteins, bound to SH3 domains in vitro. A 116 kDa protein from a human lung adenocarcinoma cell line (H441) was specifically bound by the human alpha-ENaC C-terminal fusion protein and by a shorter 18-amino acid proline-rich peptide derived from the larger fusion protein. The 116 kDa protein was not glycosylated and was not phosphorylated on tyrosine or by cyclic AMP-dependent protein kinase (PKA). A 134 kDa protein which was also bound by the human alpha-ENaC C-terminal fusion protein was a substrate for phosphorylation by PKA. These data suggest that the proline-rich C-terminal tail of alpha-ENaC may interact with other proteins that control its function, regulation or localization. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8526861

  2. Molecular Modeling of the Structural and Dynamical Changes in Calcium Channel TRPV5 Induced by the African-Specific A563T Variation.

    PubMed

    Wang, Lingyun; Holmes, Ross P; Peng, Ji-Bin

    2016-03-01

    Transient receptor potential cation channels, vanilloid subfamily, member 5 (TRPV5) plays a key role in active Ca(2+) reabsorption in the kidney. Variations in TRPV5 occur at high frequency in African populations and may contribute to their higher efficiency of Ca(2+) reabsorption. One of the African specific variations, A563T, exhibits increased Ca(2+) transport ability. However, it is unclear how this variation influences the channel pore. On the basis of the structure of TRPV1, a TRPV5 model was generated to simulate the structural and dynamical changes induced by the A563T variation. On the basis of this model, amino acid residue 563 interacts with V540, which is one residue away from the key residue, D542, involved in Ca(2+) selectivity and Mg(2+) blockade. The A563T variation increases secondary structure stability and reduces dynamical motion of D542. In addition, the A563T variation alters the electrostatic potential of the outer surface of the pore. Differences in contact between selective filter residues and residue 563 and in electrostatic potential between the two TRPV5 variants were also observed in another model derived from an alternative alignment in the selective filters between TRPV5 and TRPV1. These findings indicate that the A563T variation induces structural, dynamical, and electrostatic changes in the TRPV5 pore, providing structural insight into the functional alterations associated with the A563T variation. PMID:26837804

  3. High-efficient photo-electron transport channel in SiC constructed by depositing cocatalysts selectively on specific surface sites for visible-light H2 production

    NASA Astrophysics Data System (ADS)

    Wang, Da; Peng, Yuan; Wang, Qi; Pan, Nanyan; Guo, Zhongnan; Yuan, Wenxia

    2016-04-01

    Control cocatalyst location on a metal-free semiconductor to promote surface charge transfer for decreasing the electron-hole recombination is crucial for enhancing solar energy conversion. Based on the findings that some metals have an affinity for bonding with the specific atoms of polar semiconductors at a heterostructure interface, we herein control Pt deposition selectively on the Si sites of a micro-SiC photocatalyst surface via in-situ photo-depositing. The Pt-Si bond forming on the interface constructs an excellent channel, which is responsible for accelerating photo-electron transfer from SiC to Pt and then reducing water under visible-light. The hydrogen production is enhanced by two orders of magnitude higher than that of bare SiC, and 2.5 times higher than that of random-depositing nano-Pt with the same loading amount.

  4. A 1.83 μJ/Classification, 8-Channel, Patient-Specific Epileptic Seizure Classification SoC Using a Non-Linear Support Vector Machine.

    PubMed

    Bin Altaf, Muhammad Awais; Yoo, Jerald

    2016-02-01

    A non-linear support vector machine (NLSVM) seizure classification SoC with 8-channel EEG data acquisition and storage for epileptic patients is presented. The proposed SoC is the first work in literature that integrates a feature extraction (FE) engine, patient specific hardware-efficient NLSVM classification engine, 96 KB SRAM for EEG data storage and low-noise, high dynamic range readout circuits. To achieve on-chip integration of the NLSVM classification engine with minimum area and energy consumption, the FE engine utilizes time division multiplexing (TDM)-BPF architecture. The implemented log-linear Gaussian basis function (LL-GBF) NLSVM classifier exploits the linearization to achieve energy consumption of 0.39 μ J/operation and reduces the area by 28.2% compared to conventional GBF implementation. The readout circuits incorporate a chopper-stabilized DC servo loop to minimize the noise level elevation and achieve noise RTI of 0.81 μ Vrms for 0.5-100 Hz bandwidth with an NEF of 4.0. The 5 × 5 mm (2) SoC is implemented in a 0.18 μm 1P6M CMOS process consuming 1.83 μ J/classification for 8-channel operation. SoC verification has been done with the Children's Hospital Boston-MIT EEG database, as well as with a specific rapid eye-blink pattern detection test, which results in an average detection rate, average false alarm rate and latency of 95.1%, 0.94% (0.27 false alarms/hour) and 2 s, respectively. PMID:25700471

  5. Region-specific changes in transient receptor potential vanilloid channel expression in the vasopressin magnocellular system in hepatic cirrhosis-induced hyponatraemia.

    PubMed

    Nedungadi, T P; Carreño, F R; Walch, J D; Bathina, C S; Cunningham, J T

    2012-04-01

    The present study aimed to measure the expression of transient receptor potential (TRP) channels in the magnocellular neurones of the paraventricular (PVN) and supraoptic nucleus (SON) in an animal model of hepatic cirrhosis associated with inappropriate vasopressin (AVP) release. In these studies, we used chronic bile duct ligation (BDL) in the rat, which is a commonly used model of hepatic cirrhosis, associated with elevated plasma AVP. The present study tested the hypothesis that changes in TRP vanilloid (TRPV) channel expression may be related to inappropriate AVP release in BDL rats. To test our hypothesis, we utilised laser capture microdissection of AVP neurones in the PVN and SON and western blot analysis from brain punches. Laser capture microdissection and quantitative reverse transcriptase-polymerase chain reaction demonstrated elevated TRPV2 mRNA in the PVN and SON of BDL compared to sham-ligated controls. AVP transcription was also increased as determined using intron specific primers to measure heteronuclear RNA. Immunohistochemistry demonstrated increased AVP and TRPV2 positive cells in both the PVN and SON after BDL. Also, there was an increased co-expression of TRPV2 and AVP cells after BDL. However, there was no change in the colocalisation counts of TRPV2 and oxytocin in both the magnocellular regions evaluated. In the SON but not the PVN, the transcription levels of TRPV4 were also significantly increased in BDL rats. Western blot analysis of punches containing the PVN and SON revealed that TRPV2 protein content was significantly increased in these brain regions in BDL rats compared to sham rats. Our data suggest that regionally specific changes in TRPV expression in the magnocellular neurosecretory cell AVP neurones could alter their osmosensing ability. PMID:22188460

  6. Enhanced capacitative calcium entry and TRPC channel gene expression in human LES smooth muscle.

    PubMed

    Wang, Jian; Laurier, Lisanne G; Sims, Stephen M; Preiksaitis, Harold G

    2003-06-01

    Transient receptor potential channel (TRPC) genes encode Ca(2+)-permeable channels mediating capacitative Ca(2+) entry (CCE), which maintains intracellular Ca(2+) stores. We compared TRPC gene expression and CCE in human esophageal body (EB) and lower esophageal sphincter (LES), because these smooth muscles have distinct contractile functions that are likely associated with different Ca(2+) regulatory mechanisms. Circular layer smooth muscle cells were grown in primary culture. Transcriptional expression of TRPC genes was compared by semiquantitative RT-PCR. CCE was measured by fura 2 Ca(2+) fluorescence after blockade of sarcoplasmic reticulum Ca(2+)-ATPase with thapsigargin. mRNA for TRPC1, TRPC3, TRPC4, TRPC5, and TRPC6 was identified in EB and LES. TRPC3 and TRPC4 were more abundant in LES than EB. Basal concentration of free intracellular Ca(2+) ([Ca(2+)](i)) was similar in cells from LES (138 +/- 8 nmol/l) and EB (110 +/- 6 nmol/l) and increased with ACh (10 micromol/l; 650 +/- 28 and 590 +/- 21 nmol/l, respectively). With zero Ca(2+) in bath, thapsigargin (2 micromol/l) increased [Ca(2+)](i) more in LES (550 +/- 22 nmol/l) than EB (250 +/- 15 nmol/l, P < 0.001). Subsequent external application of 1 mmol/l Ca(2+) increased [Ca(2+)](i) more in LES (585 +/- 35 nmol/l) than EB (295 +/- 21 nmol/l, P < 0.001), indicating enhanced CCE in LES. This demonstrates CCE and TRPC transcriptional expression in human esophageal smooth muscle. In LES cells, enhanced CCE and expression of TRPC3 and TRPC4 may contribute to the physiological characteristics that distinguish LES from EB. PMID:12736151

  7. Increased rhythmicity in hypertensive arterial smooth muscle is linked to transient receptor potential canonical channels.

    PubMed

    Chen, Xiaoping; Yang, Dachun; Ma, Shuangtao; He, Hongbo; Luo, Zhidan; Feng, Xiaoli; Cao, Tingbing; Ma, Liqun; Yan, Zhencheng; Liu, Daoyan; Tepel, Martin; Zhu, Zhiming

    2010-10-01

    Vasomotion describes oscillations of arterial vascular tone due to synchronized changes of intracellular calcium concentrations. Since increased calcium influx into vascular smooth muscle cells from spontaneously hypertensive rats (SHR) has been associated with variances of transient receptor potential canonical (TRPC) channels, in the present study we tested the hypothesis that increased vasomotion in hypertension is directly linked to increased TRPC expression. Using a small vessel myograph we observed significantly increased norepinephrine-induced vasomotion in mesenteric arterioles from SHR compared to normotensive Wistar-Kyoto (WKY) rats. Using immunoblottings we obtained significantly increased expression of TRPC1, TRPC3 and TRPC5 in mesenteric arterioles from SHR compared to WKY, whereas TRPC4 and TRPC6 showed no differences. Norepinephrine-induced vasomotion from SHR was significantly reduced in the presence of verapamil, SKF96365, 2-aminoethoxydiphenylborane (2-APB) or gadolinium. Pre-incubation of mesenteric arterioles with anti-TRPC1 and anti-TRPC3 antibodies significantly reduced norepinephrine-induced vasomotion and calcium influx. Control experiments with pre-incubation of TRPC antibodies plus their respective antigenic peptide or in the presence of anti-β-actin antibodies or random immunoglobulins not related to TRPC channels showed no inhibitory effects of norepinephrine-induced vasomotion and calcium influx. Administration of candesartan or telmisartan, but not amlodipine to SHR for 16 weeks significantly reduced either the expression of TRPC1, TRPC3 and TRPC5 as well as norepinephrine-induced vasomotion in mesenteric arterioles. In conclusion we gave experimental evidence that the increased TRPC1, TRPC3 and TRPC5 expression in mesenteric arterioles from SHR causes increased vasomotion in hypertension. PMID:19725917

  8. Use of antibodies specific to defined regions of scorpion. cap alpha. -toxin to study its interaction with its receptor site on the sodium channel

    SciTech Connect

    Ayeb, M.E.; Bahraoui, E.M.; Granier, C.; Rochat, H.

    1986-10-21

    Five antibody populations selected by immunoaffinity chromatography for the specificity toward various regions of toxin II of the scorpion Androctonus australis Hector were used to probe the interaction of this protein with its receptor site on the sodium channel. These studies indicate that two antigenic sites, one located around the disulfide bridge 12-63 and one encompassing residues 50-59, are involved in the molecular mechanisms of toxicity neutralization. Fab fragments specific to the region around disulfide bridge 12-63 inhibit binding of the /sup 125/I-labeled toxin to its receptor site. Also, these two antigenic regions are inaccessible to the antibodies when the toxin is bound to its receptor site. In contrast, the two other antigenic sites encompassing the only ..cap alpha..-helix region (residues 23-32) and a ..beta..-turn structure (residues 32-35) are accessible to the respective antibodies when the toxin is bound to its receptor. Together, these data support the recent proposal that a region made of residues that are conserved in the scorpion toxin family is involved in the binding of the toxin to the receptor.

  9. Channel catfish granzyme-like I is a highly specific serine protease with metase activity that is expressed by fish NK-like cells.

    PubMed

    Thorpe, Michael; Akula, Srinivas; Hellman, Lars

    2016-10-01

    Here we present the extended cleavage specificity of catfish granzyme-like I, previously identified in fish NK-like cells. This protease has been characterised using substrate phage display and further validated by using a panel of recombinant substrates. A strict preference for Met in the P1 (cleavage) position, indicating metase specificity was observed. A screening of potential in vivo substrates was performed based on the derived P5-P3' consensus: Arg-Val-Thr-Gly-Met(↓)Ser-Leu-Val. Channel catfish caspase 6 was one very interesting potential target identified. This site was present in an adjacent position to the classic caspase activation site (Asp179 in human caspase 6). Cleavage of this site (hence potential activation) by the catfish granzyme-like I could reveal a novel mechanism of caspase 6 activation. This poses an interesting idea that the role of granzyme-like proteases in the activation of caspase dependent apoptosis mechanisms has been conserved for over 400 million years. PMID:27216028

  10. Severe Salt-Losing Syndrome and Hyperkalemia Induced by Adult Nephron-Specific Knockout of the Epithelial Sodium Channel α-Subunit.

    PubMed

    Perrier, Romain; Boscardin, Emilie; Malsure, Sumedha; Sergi, Chloé; Maillard, Marc P; Loffing, Johannes; Loffing-Cueni, Dominique; Sørensen, Mads Vaarby; Koesters, Robert; Rossier, Bernard C; Frateschi, Simona; Hummler, Edith

    2016-08-01

    Systemic pseudohypoaldosteronism type 1 (PHA-1) is a severe salt-losing syndrome caused by loss-of-function mutations of the amiloride-sensitive epithelial sodium channel (ENaC) and characterized by neonatal life-threatening hypovolemia and hyperkalemia. The very high plasma aldosterone levels detected under hypovolemic or hyperkalemic challenge can lead to increased or decreased sodium reabsorption, respectively, through the Na(+)/Cl(-) cotransporter (NCC). However, the role of ENaC deficiency remains incompletely defined, because constitutive inactivation of individual ENaC subunits is neonatally lethal in mice. We generated adult inducible nephron-specific αENaC-knockout mice (Scnn1a(Pax8/LC1)) that exhibit hyperkalemia and body weight loss when kept on a regular-salt diet, thus mimicking PHA-1. Compared with control mice fed a regular-salt diet, knockout mice fed a regular-salt diet exhibited downregulated expression and phosphorylation of NCC protein, despite high plasma aldosterone levels. In knockout mice fed a high-sodium and reduced-potassium diet (rescue diet), although plasma aldosterone levels remained significantly increased, NCC expression returned to control levels, and body weight, plasma and urinary electrolyte concentrations, and excretion normalized. Finally, shift to a regular diet after the rescue diet reinstated the symptoms of severe PHA-1 syndrome and significantly reduced NCC phosphorylation. In conclusion, lack of ENaC-mediated sodium transport along the nephron cannot be compensated for by other sodium channels and/or transporters, only by a high-sodium and reduced-potassium diet. We further conclude that hyperkalemia becomes the determining factor in regulating NCC activity, regardless of sodium loss, in the ENaC-mediated salt-losing PHA-1 phenotype. PMID:26701978

  11. In pursuit of small molecule chemistry for calcium-permeable non-selective TRPC channels – mirage or pot of gold?

    PubMed Central

    Bon, Robin S; Beech, David J

    2013-01-01

    The primary purpose of this review is to address the progress towards small molecule modulators of human Transient Receptor Potential Canonical proteins (TRPC1, TRPC3, TRPC4, TRPC5, TRPC6 and TRPC7). These proteins generate channels for calcium and sodium ion entry. They are relevant to many mammalian cell types including acinar gland cells, adipocytes, astrocytes, cardiac myocytes, cochlea hair cells, endothelial cells, epithelial cells, fibroblasts, hepatocytes, keratinocytes, leukocytes, mast cells, mesangial cells, neurones, osteoblasts, osteoclasts, platelets, podocytes, smooth muscle cells, skeletal muscle and tumour cells. There are broad-ranging positive roles of the channels in cell adhesion, migration, proliferation, survival and turning, vascular permeability, hypertrophy, wound-healing, hypo-adiponectinaemia, angiogenesis, neointimal hyperplasia, oedema, thrombosis, muscle endurance, lung hyper-responsiveness, glomerular filtration, gastrointestinal motility, pancreatitis, seizure, innate fear, motor coordination, saliva secretion, mast cell degranulation, cancer cell drug resistance, survival after myocardial infarction, efferocytosis, hypo-matrix metalloproteinase, vasoconstriction and vasodilatation. Known small molecule stimulators of the channels include hyperforin, genistein and rosiglitazone, but there is more progress with inhibitors, some of which have promising potency and selectivity. The inhibitors include 2-aminoethoxydiphenyl borate, 2-aminoquinolines, 2-aminothiazoles, fatty acids, isothiourea derivatives, naphthalene sulfonamides, N-phenylanthranilic acids, phenylethylimidazoles, piperazine/piperidine analogues, polyphenols, pyrazoles and steroids. A few of these agents are starting to be useful as tools for determining the physiological and pathophysiological functions of TRPC channels. We suggest that the pursuit of small molecule modulators for TRPC channels is important but that it requires substantial additional effort and

  12. Gαi2- and Gαi3-Specific Regulation of Voltage-Dependent L-Type Calcium Channels in Cardiomyocytes

    PubMed Central

    Dizayee, Sara; Kaestner, Sonja; Kuck, Fabian; Hein, Peter; Klein, Christoph; Piekorz, Roland P.; Meszaros, Janos; Matthes, Jan; Nürnberg, Bernd; Herzig, Stefan

    2011-01-01

    Background Two pertussis toxin sensitive Gi proteins, Gi2 and Gi3, are expressed in cardiomyocytes and upregulated in heart failure. It has been proposed that the highly homologous Gi isoforms are functionally distinct. To test for isoform-specific functions of Gi proteins, we examined their role in the regulation of cardiac L-type voltage-dependent calcium channels (L-VDCC). Methods Ventricular tissues and isolated myocytes were obtained from mice with targeted deletion of either Gαi2 (Gαi2−/−) or Gαi3 (Gαi3−/−). mRNA levels of Gαi/o isoforms and L-VDCC subunits were quantified by real-time PCR. Gαi and Cavα1 protein levels as well as protein kinase B/Akt and extracellular signal-regulated kinases 1/2 (ERK1/2) phosphorylation levels were assessed by immunoblot analysis. L-VDCC function was assessed by whole-cell and single-channel current recordings. Results In cardiac tissue from Gαi2−/− mice, Gαi3 mRNA and protein expression was upregulated to 187±21% and 567±59%, respectively. In Gαi3−/− mouse hearts, Gαi2 mRNA (127±5%) and protein (131±10%) levels were slightly enhanced. Interestingly, L-VDCC current density in cardiomyocytes from Gαi2−/− mice was lowered (−7.9±0.6 pA/pF, n = 11, p<0.05) compared to wild-type cells (−10.7±0.5 pA/pF, n = 22), whereas it was increased in myocytes from Gαi3−/− mice (−14.3±0.8 pA/pF, n = 14, p<0.05). Steady-state inactivation was shifted to negative potentials, and recovery kinetics slowed in the absence of Gαi2 (but not of Gαi3) and following treatment with pertussis toxin in Gαi3−/−. The pore forming Cavα1 protein level was unchanged in all mouse models analyzed, similar to mRNA levels of Cavα1 and Cavβ2 subunits. Interestingly, at the cellular signalling level, phosphorylation assays revealed abolished carbachol-triggered activation of ERK1/2 in mice lacking Gαi2. Conclusion Our data provide novel evidence for an isoform-specific modulation of L-VDCC by G

  13. A recombinant fusion protein containing a spider toxin specific for the insect voltage-gated sodium ion channel shows oral toxicity towards insects of different orders

    PubMed Central

    Yang, Sheng; Pyati, Prashant; Fitches, Elaine; Gatehouse, John A.

    2014-01-01

    Recombinant fusion protein technology allows specific insecticidal protein and peptide toxins to display activity in orally-delivered biopesticides. The spider venom peptide δ-amaurobitoxin-PI1a, which targets insect voltage-gated sodium channels, was fused to the “carrier” snowdrop lectin (GNA) to confer oral toxicity. The toxin itself (PI1a) and an amaurobitoxin/GNA fusion protein (PI1a/GNA) were produced using the yeast Pichia pastoris as expression host. Although both proteins caused mortality when injected into cabbage moth (Mamestra brassicae) larvae, the PI1a/GNA fusion was approximately 6 times as effective as recombinant PI1a on a molar basis. PI1a alone was not orally active against cabbage moth larvae, but a single 30 μg dose of the PI1a/GNA fusion protein caused 100% larval mortality within 6 days when fed to 3rd instar larvae, and caused significant reductions in survival, growth and feeding in 4th – 6th instar larvae. Transport of fusion protein from gut contents to the haemolymph of cabbage moth larvae, and binding to the nerve chord, was shown by Western blotting. The PI1a/GNA fusion protein also caused mortality when delivered orally to dipteran (Musca domestica; housefly) and hemipteran (Acyrthosiphon pisum; pea aphid) insects, making it a promising candidate for development as a biopesticide. PMID:24486516

  14. Gliadin fragments and a specific gliadin 33-mer peptide close KATP channels and induce insulin secretion in INS-1E cells and rat islets of langerhans.

    PubMed

    Dall, Morten; Calloe, Kirstine; Haupt-Jorgensen, Martin; Larsen, Jesper; Schmitt, Nicole; Josefsen, Knud; Buschard, Karsten

    2013-01-01

    In non-obese diabetic (NOD) mice, diabetes incidence is reduced by a gluten-free diet. Gluten peptides, such as the compound gliadin, can cross the intestinal barrier and may directly affect pancreatic beta cells. We investigated the effects of enzymatically-digested gliadin in NOD mice, INS-1E cells and rat islets. Six injections of gliadin digest in 6-week-old NOD mice did not affect diabetes development, but increased weight gain (20% increase by day 100). In INS-1E cells, incubation with gliadin digest induced a dose-dependent increase in insulin secretion, up to 2.5-fold after 24 hours. A similar effect was observed in isolated rat islets (1.6-fold increase). In INS-1E cells, diazoxide reduced the stimulatory effect of gliadin digest. Additionally, gliadin digest was shown to decrease current through KATP-channels. A specific gliadin 33-mer had a similar effect, both on current and insulin secretion. Finally, INS-1E incubation with gliadin digest potentiated palmitate-induced insulin secretion by 13% compared to controls. Our data suggest that gliadin fragments may contribute to the beta-cell hyperactivity observed prior to the development of type 1 diabetes. PMID:23785500

  15. A recombinant fusion protein containing a spider toxin specific for the insect voltage-gated sodium ion channel shows oral toxicity towards insects of different orders.

    PubMed

    Yang, Sheng; Pyati, Prashant; Fitches, Elaine; Gatehouse, John A

    2014-04-01

    Recombinant fusion protein technology allows specific insecticidal protein and peptide toxins to display activity in orally-delivered biopesticides. The spider venom peptide δ-amaurobitoxin-PI1a, which targets insect voltage-gated sodium channels, was fused to the "carrier" snowdrop lectin (GNA) to confer oral toxicity. The toxin itself (PI1a) and an amaurobitoxin/GNA fusion protein (PI1a/GNA) were produced using the yeast Pichia pastoris as expression host. Although both proteins caused mortality when injected into cabbage moth (Mamestra brassicae) larvae, the PI1a/GNA fusion was approximately 6 times as effective as recombinant PI1a on a molar basis. PI1a alone was not orally active against cabbage moth larvae, but a single 30 μg dose of the PI1a/GNA fusion protein caused 100% larval mortality within 6 days when fed to 3rd instar larvae, and caused significant reductions in survival, growth and feeding in 4th - 6th instar larvae. Transport of fusion protein from gut contents to the haemolymph of cabbage moth larvae, and binding to the nerve chord, was shown by Western blotting. The PI1a/GNA fusion protein also caused mortality when delivered orally to dipteran (Musca domestica; housefly) and hemipteran (Acyrthosiphon pisum; pea aphid) insects, making it a promising candidate for development as a biopesticide. PMID:24486516

  16. Human autoantibodies specific for the α1A calcium channel subunit reduce both P-type and Q-type calcium currents in cerebellar neurons

    PubMed Central

    Pinto, Ashwin; Gillard, Samantha; Moss, Fraser; Whyte, Kathryn; Brust, Paul; Williams, Mark; Stauderman, Ken; Harpold, Michael; Lang, Bethan; Newsom-Davis, John; Bleakman, David; Lodge, David; Boot, John

    1998-01-01

    The pharmacological properties of voltage-dependent calcium channel (VDCC) subtypes appear mainly to be determined by the α1 pore-forming subunit but, whether P-and Q-type VDCCs are encoded by the same α1 gene presently is unresolved. To investigate this, we used IgG antibodies to presynaptic VDCCs at motor nerve terminals that underlie muscle weakness in the autoimmune Lambert–Eaton myasthenic syndrome (LEMS). We first studied their action on changes in intracellular free Ca2+ concentration [Ca2+]i in human embryonic kidney (HEK293) cell lines expressing different combinations of human recombinant VDCC subunits. Incubation for 18 h with LEMS IgG (2 mg/ml) caused a significant dose-dependent reduction in the K+-stimulated [Ca2+]i increase in the α1A cell line but not in the α1B, α1C, α1D, and α1E cell lines, establishing the α1A subunit as the target for these autoantibodies. Exploiting this specificity, we incubated cultured rat cerebellar neurones with LEMS IgG and observed a reduction in P-type current in Purkinje cells and both P- and Q-type currents in granule cells. These data are consistent with the hypothesis that the α1A gene encodes for the pore-forming subunit of both P-type and Q-type VDCCs. PMID:9653186

  17. Sequence-specific 1H-NMR assignment and secondary structure of black mamba dendrotoxin I, a highly selective blocker of voltage-gated potassium channels.

    PubMed

    Foray, M F; Lancelin, J M; Hollecker, M; Marion, D

    1993-02-01

    The secondary structure of dendrotoxin I, an important constituent of the venom of the African black mamba snake Dendroaspis polylepis polylepis, was determined in aqueous solution by two-dimensional methods. Complete sequence-specific 1H-NMR assignment was obtained with the exception of the backbone amide proton of Gly39 and Cys40. Dendrotoxin I is based on a central antiparallel beta-sheet and two small helices located at the N- and the C-terminal extremities. These secondary-structural units occur at exactly the same places in the amino acid sequence as those of bovine pancreatic trypsin inhibitor (BPTI), with which dendrotoxin I shares 33% sequence similarity. According to the disulfide-bridge positions and the long-range NOE observed these secondary-structural elements fold in a similar manner to BPTI. This similarity allows an hypothesis according to which dendrotoxin I could derive from an ancestral Künitz-type proteinase inhibitor. This ancestor would have been heavily mutated at amino acid positions not critical for gross structure. The spatial locations of the solvent-exposed amino acids concerned could therefore serve as a guideline for interpretation of the structure/activity relationship of dendrotoxin I for the blockage of voltage-sensitive potassium channels of which dendrotoxin I is a strong inhibitor. The possible connections with other polypeptide toxins that block related ion currents is discussed. PMID:7679640

  18. The Ca{sup 2+} channel TRPML3 specifically interacts with the mammalian ATG8 homologue GATE16 to regulate autophagy

    SciTech Connect

    Choi, Suzy; Kim, Hyun Jin

    2014-01-03

    Highlights: •Split-ubiquitin MY2H screen identified GATE16 as an interacting protein of TRPML3. •TRPML3 specifically binds to a mammalian ATG8 homologue GATE16, not to LC3B. •The interaction of TRPML3 with GATE16 facilitates autophagosome formation. •GATE16 is expressed in both autophagosome and extra-autophagosomal compartments. -- Abstract: TRPML3 is a Ca{sup 2+} permeable cation channel expressed in multiple intracellular compartments. Although TRPML3 is implicated in autophagy, how TRPML3 can regulate autophagy is not understood. To search interacting proteins with TRPML3 in autophagy, we performed split-ubiquitin membrane yeast two-hybrid (MY2H) screening with TRPML3-loop as a bait and identified GATE16, a mammalian ATG8 homologue. GST pull-down assay revealed that TRPML3 and TRPML3-loop specifically bind to GATE16, not to LC3B. Co-immunoprecipitation (co-IP) experiments showed that TRPML3 and TRPML3-loop pull down only the lipidated form of GATE16, indicating that the interaction occurs exclusively at the organellar membrane. The interaction of TRPML3 with GATE16 and GATE16-positive vesicle formation were increased in starvation induced autophagy, suggesting that the interaction facilitates the function of GATE16 in autophagosome formation. However, GATE16 was not required for TRPML3 trafficking to autophagosomes. Experiments using dominant-negative (DN) TRPML3(D458K) showed that GATE16 is localized not only in autophagosomes but also in extra-autophagosomal compartments, by contrast with LC3B. Since GATE16 acts at a later stage of the autophagosome biogenesis, our results suggest that TRPML3 plays a role in autophagosome maturation through the interaction with GATE16, by providing Ca{sup 2+} in the fusion process.

  19. Inhibition of Cancer Cell Growth by Exposure to a Specific Time-Varying Electromagnetic Field Involves T-Type Calcium Channels

    PubMed Central

    Buckner, Carly A.; Buckner, Alison L.; Koren, Stan A.; Persinger, Michael A.; Lafrenie, Robert M.

    2015-01-01

    Electromagnetic field (EMF) exposures affect many biological systems. The reproducibility of these effects is related to the intensity, duration, frequency, and pattern of the EMF. We have shown that exposure to a specific time-varying EMF can inhibit the growth of malignant cells. Thomas-EMF is a low-intensity, frequency-modulated (25-6 Hz) EMF pattern. Daily, 1 h, exposures to Thomas-EMF inhibited the growth of malignant cell lines including B16-BL6, MDA-MB-231, MCF-7, and HeLa cells but did not affect the growth of non-malignant cells. Thomas-EMF also inhibited B16-BL6 cell proliferation in vivo. B16-BL6 cells implanted in syngeneic C57b mice and exposed daily to Thomas-EMF produced smaller tumours than in sham-treated controls. In vitro studies showed that exposure of malignant cells to Thomas-EMF for > 15 min promoted Ca2+ influx which could be blocked by inhibitors of voltage-gated T-type Ca2+ channels. Blocking Ca2+ uptake also blocked Thomas-EMF-dependent inhibition of cell proliferation. Exposure to Thomas-EMF delayed cell cycle progression and altered cyclin expression consistent with the decrease in cell proliferation. Non-malignant cells did not show any EMF-dependent changes in Ca2+ influx or cell growth. These data confirm that exposure to a specific EMF pattern can affect cellular processes and that exposure to Thomas-EMF may provide a potential anti-cancer therapy. PMID:25875081

  20. Direct Evidence for Microdomain-Specific Localization and Remodeling of Functional L-Type Calcium Channels in Rat and Human Atrial Myocytes

    PubMed Central

    Glukhov, Alexey V.; Balycheva, Marina; Sanchez-Alonso, Jose L.; Ilkan, Zeki; Alvarez-Laviada, Anita; Bhogal, Navneet; Diakonov, Ivan; Schobesberger, Sophie; Sikkel, Markus B.; Bhargava, Anamika; Faggian, Giuseppe; Punjabi, Prakash P.; Houser, Steven R.

    2015-01-01

    Background— Distinct subpopulations of L-type calcium channels (LTCCs) with different functional properties exist in cardiomyocytes. Disruption of cellular structure may affect LTCC in a microdomain-specific manner and contribute to the pathophysiology of cardiac diseases, especially in cells lacking organized transverse tubules (T-tubules) such as atrial myocytes (AMs). Methods and Results— Isolated rat and human AMs were characterized by scanning ion conductance, confocal, and electron microscopy. Half of AMs possessed T-tubules and structured topography, proportional to cell width. A bigger proportion of myocytes in the left atrium had organized T-tubules and topography than in the right atrium. Super-resolution scanning patch clamp showed that LTCCs distribute equally in T-tubules and crest areas of the sarcolemma, whereas, in ventricular myocytes, LTCCs primarily cluster in T-tubules. Rat, but not human, T-tubule LTCCs had open probability similar to crest LTCCs, but exhibited ≈40% greater current. Optical mapping of Ca2+ transients revealed that rat AMs presented ≈3-fold as many spontaneous Ca2+ release events as ventricular myocytes. Occurrence of crest LTCCs and spontaneous Ca2+ transients were eliminated by either a caveolae-targeted LTCC antagonist or disrupting caveolae with methyl-β-cyclodextrin, with an associated ≈30% whole-cell ICa,L reduction. Heart failure (16 weeks post–myocardial infarction) in rats resulted in a T-tubule degradation (by ≈40%) and significant elevation of spontaneous Ca2+ release events. Although heart failure did not affect LTCC occurrence, it led to ≈25% decrease in T-tubule LTCC amplitude. Conclusions— We provide the first direct evidence for the existence of 2 distinct subpopulations of functional LTCCs in rat and human AMs, with their biophysical properties modulated in heart failure in a microdomain-specific manner. PMID:26450916

  1. Specific expression of the human voltage-gated proton channel Hv1 in highly metastatic breast cancer cells, promotes tumor progression and metastasis

    SciTech Connect

    Wang, Yifan; Li, Shu Jie; Pan, Juncheng; Che, Yongzhe; Yin, Jian; Zhao, Qing

    2011-08-26

    Highlights: {yields} Hv1 is specifically expressed in highly metastatic human breast tumor tissues. {yields} Hv1 regulates breast cancer cytosolic pH. {yields} Hv1 acidifies extracellular milieu. {yields} Hv1 exacerbates the migratory ability of metastatic cells. -- Abstract: The newly discovered human voltage-gated proton channel Hv1 is essential for proton transfer, which contains a voltage sensor domain (VSD) without a pore domain. We report here for the first time that Hv1 is specifically expressed in the highly metastatic human breast tumor tissues, but not in poorly metastatic breast cancer tissues, detected by immunohistochemistry. Meanwhile, real-time RT-PCR and immunocytochemistry showed that the expression levels of Hv1 have significant differences among breast cancer cell lines, MCF-7, MDA-MB-231, MDA-MB-468, MDA-MB-453, T-47D and SK-BR-3, in which Hv1 is expressed at a high level in highly metastatic human breast cancer cell line MDA-MB-231, but at a very low level in poorly metastatic human breast cancer cell line MCF-7. Inhibition of Hv1 expression in the highly metastatic MDA-MB-231 cells by small interfering RNA (siRNA) significantly decreases the invasion and migration of the cells. The intracellular pH of MDA-MB-231 cells down-regulated Hv1 expression by siRNA is obviously decreased compared with MDA-MB-231 with the scrambled siRNA. The expression of matrix metalloproteinase-2 and gelatinase activity in MDA-MB-231 cells suppressed Hv1 by siRNA were reduced. Our results strongly suggest that Hv1 regulates breast cancer intracellular pH and exacerbates the migratory ability of metastatic cells.

  2. REACH SPECIFIC CHANNEL STABILIZATION BASED ON COMPREHENSIVE EVALUATION OF VALLEY FILL HISTORY, ALLUVIAL ARCHITECTURE AND GROUNDWATER HYDROLOGY IN A MOUNTAIN STREAM IN THE CENTRAL GREAT BASIN, NEVADA

    EPA Science Inventory

    Kingston meadow, located in the Toiyabe Range, is one of many wet meadow complexes threatened by rapid channel incision in the mountain ranges of the central Great Basin. Channel incision can lower the baselevel for groundwater discharge and de-water meadow complexes resulting in...

  3. Molecular analysis of a store-operated and 2-acetyl-sn-glycerol-sensitive non-selective cation channel. Heteromeric assembly of TRPC1-TRPC3.

    PubMed

    Liu, Xibao; Bandyopadhyay, Bidhan C; Singh, Brij B; Groschner, Klaus; Ambudkar, Indu S

    2005-06-01

    We have reported that internal Ca2+ store depletion in HSY cells stimulates a nonselective cation current which is distinct from I(CRAC) in RBL cells and TRPC1-dependent I(SOC) in HSG cells (Liu, X., Groschner, K., and Ambudkar, I. S. (2004) J. Membr. Biol. 200, 93-104). Here we have analyzed the molecular composition of this channel. Both thapsigargin (Tg) and 2-acetyl-sn-glycerol (OAG) stimulated similar non-selective cation currents and Ca2+ entry in HSY cells. The effects of Tg and OAG were not additive. HSY cells endogenously expressed TRPC1, TRPC3, and TRPC4 but not TRPC5 or TRPC6. Immunoprecipitation of TRPC1 pulled down TRPC3 but not TRPC4. Conversely, TRPC1 co-immunoprecipitated with TRPC3. Expression of antisense TRPC1 decreased (i) Tg- and OAG-stimulated currents and Ca2+ entry and (ii) the level of endogenous TRPC1 but not TRPC4. Antisense TRPC3 similarly reduced Ca2+ entry and endogenous TRPC3. Yeast two-hybrid analysis revealed an interaction between NTRPC1 and NTRPC3 (CTRPC1-CTRPC3, CTRPC3-CTRPC1, or CTRPC1-NTRPC3 did not interact), which was confirmed by glutathione S-transferase (GST) pull-down assays (GST-NTRPC3 pulled down TRPC1 and vice versa). Expression of NTRPC1 or NTRPC3 induced similar dominant suppression of Tg- and OAG-stimulated Ca2+ entry. NTRPC3 did not alter surface expression of TRPC1 or TRPC3 but disrupted TRPC1-TRPC3 association. In aggregate, our data demonstrate that TRPC1 and TRPC3 co-assemble, via N-terminal interactions, to form a heteromeric store-operated non-selective cation channel in HSY cells. Thus selective association between TRPCs generate distinct store-operated channels. Diversity of store-operated channels might be related to the physiology of the different cell types. PMID:15834157

  4. A model for the generation of localized transient [Na{sup +}] elevations in vascular smooth muscle

    SciTech Connect

    Fameli, Nicola; Kuo, Kuo-Hsing; Breemen, Cornelis van

    2009-11-20

    We present a stochastic computational model to study the mechanism of signaling between a source and a target ionic transporter, both localized on the plasma membrane (PM). In general this requires a nanometer-scale cytoplasmic space, or nanodomain, between the PM and a peripheral organelle to reflect ions back towards the PM. Specifically we investigate the coupling between Na{sup +} entry via the transient receptor potential canonical channel 6 (TRPC6) and the Na{sup +}/Ca{sup 2+} exchanger (NCX), a process which is essential for reloading the sarcoplasmic reticulum (SR) via the sarco/endoplasmic reticulum Ca{sup 2+}ATPase (SERCA) and maintaining Ca{sup 2+} oscillations in activated vascular smooth muscle. Having previously modeled the flow of Ca{sup 2+} between reverse NCX and SERCA during SR refilling, this quantitative approach now allows us to model the upstream linkage of Na{sup +} entry through TRPC6 to reversal of NCX. We have implemented a random walk (RW) Monte Carlo (MC) model with simulations mimicking a diffusion process originating at the TRPC6 within PM-SR junctions. The model calculates the average Na{sup +} in the nanospace and also produces profiles as a function of distance from the source. Our results highlight the necessity of a strategic juxtaposition of the relevant ion translocators as well as other physical structures within the nanospaces to permit adequate Na{sup +} build-up to initiate NCX reversal and Ca{sup 2+} influx to refill the SR.

  5. Voltage-dependent and -independent titration of specific residues accounts for complex gating of a ClC chloride channel by extracellular protons.

    PubMed

    Niemeyer, María Isabel; Cid, L Pablo; Yusef, Yamil R; Briones, Rodolfo; Sepúlveda, Francisco V

    2009-04-01

    The ClC transport protein family comprises both Cl(-) ion channel and H(+)/Cl(-) and H(+)/NO(3)(-) exchanger members. Structural studies on a bacterial ClC transporter reveal a pore obstructed at its external opening by a glutamate side-chain which acts as a gate for Cl(-) passage and in addition serves as a staging post for H(+) exchange. This same conserved glutamate acts as a gate to regulate Cl(-) flow in ClC channels. The activity of ClC-2, a genuine Cl(-) channel, has a biphasic response to extracellular pH with activation by moderate acidification followed by abrupt channel closure at pH values lower than approximately 7. We have now investigated the molecular basis of this complex gating behaviour. First, we identify a sensor that couples extracellular acidification to complete closure of the channel. This is extracellularly-facing histidine 532 at the N-terminus of transmembrane helix Q whose neutralisation leads to channel closure in a cooperative manner. We go on to show that acidification-dependent activation of ClC-2 is voltage dependent and probably mediated by protonation of pore gate glutamate 207. Intracellular Cl(-) acts as a voltage-independent modulator, as though regulating the pK(a) of the protonatable residue. Our results suggest that voltage dependence of ClC-2 is given by hyperpolarisation-dependent penetration of protons from the extracellular side to neutralise the glutamate gate deep within the channel, which allows Cl(-) efflux. This is reminiscent of a partial exchanger cycle, suggesting that the ClC-2 channel evolved from its transporter counterparts. PMID:19153159

  6. Voltage-dependent and -independent titration of specific residues accounts for complex gating of a ClC chloride channel by extracellular protons

    PubMed Central

    Niemeyer, María Isabel; Cid, L Pablo; Yusef, Yamil R; Briones, Rodolfo; Sepúlveda, Francisco V

    2009-01-01

    The ClC transport protein family comprises both Cl− ion channel and H+/Cl− and H+/NO3− exchanger members. Structural studies on a bacterial ClC transporter reveal a pore obstructed at its external opening by a glutamate side-chain which acts as a gate for Cl− passage and in addition serves as a staging post for H+ exchange. This same conserved glutamate acts as a gate to regulate Cl− flow in ClC channels. The activity of ClC-2, a genuine Cl− channel, has a biphasic response to extracellular pH with activation by moderate acidification followed by abrupt channel closure at pH values lower than ∼7. We have now investigated the molecular basis of this complex gating behaviour. First, we identify a sensor that couples extracellular acidification to complete closure of the channel. This is extracellularly-facing histidine 532 at the N-terminus of transmembrane helix Q whose neutralisation leads to channel closure in a cooperative manner. We go on to show that acidification-dependent activation of ClC-2 is voltage dependent and probably mediated by protonation of pore gate glutamate 207. Intracellular Cl− acts as a voltage-independent modulator, as though regulating the pKa of the protonatable residue. Our results suggest that voltage dependence of ClC-2 is given by hyperpolarisation-dependent penetration of protons from the extracellular side to neutralise the glutamate gate deep within the channel, which allows Cl− efflux. This is reminiscent of a partial exchanger cycle, suggesting that the ClC-2 channel evolved from its transporter counterparts. PMID:19153159

  7. Kisspeptin depolarizes gonadotropin-releasing hormone neurons through activation of TRPC-like cationic channels.

    PubMed

    Zhang, Chunguang; Roepke, Troy A; Kelly, Martin J; Rønnekleiv, Oline K

    2008-04-23

    Kisspeptin and its cognate receptor, GPR54, are critical for reproductive development and for the regulation of gonadotropin-releasing hormone (GnRH) secretion. Although kisspeptin has been found to depolarize GnRH neurons, the underlying ionic mechanism has not been elucidated. Presently, we found that kisspeptin depolarized GnRH neurons in a concentration-dependent manner with a maximum depolarization of 22.6 +/- 0.6 mV and EC(50) of 2.8 +/- 0.2 nM. Under voltage-clamp conditions, kisspeptin induced an inward current of 18.2 +/- 1.6 pA (V(hold) = -60 mV) that reversed near -115 mV in GnRH neurons. The more negative reversal potential than E(K)(+) (-90 mV) was caused by the concurrent inhibition of barium-sensitive, inwardly rectifying (Kir) potassium channels and activation of sodium-dependent, nonselective cationic channels (NSCCs). Indeed, reducing extracellular Na(+) (to 5 mM) essentially eliminated the kisspeptin-induced inward current. The current-voltage relationships of the kisspeptin-activated NSCC currents exhibited double rectification with negative slope conductance below -40 mV in the majority of the cells. Pharmacological examination showed that the kisspeptin-induced inward currents were blocked by TRPC (canonical transient receptor potential) channel blockers 2-APB (2-aminoethyl diphenylborinate), flufenamic acid, SKF96365 (1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride), and Cd(2+), but not by lanthanum (100 microM). Furthermore, single-cell reverse transcription-PCR analysis revealed that TRPC1, TRPC3, TRPC4, TRPC5, TRPC6, and TRPC7 subunits were expressed in GnRH neurons. Therefore, it appears that kisspeptin depolarizes GnRH neurons through activating TRPC-like channels and, to a lesser extent, inhibition of Kir channels. These actions of kisspeptin contribute to the pronounced excitation of GnRH neurons that is critical for mammalian reproduction. PMID:18434521

  8. Learning Channels and Verbal Behavior

    ERIC Educational Resources Information Center

    Lin, Fan-Yu; Kubina, Richard M., Jr.

    2004-01-01

    This article reviews the basics of learning channels and how specification of stimuli can help enhance verbal behavior. This article will define learning channels and the role of the ability matrix in training verbal behavior.

  9. Bacterial Ion Channels.

    PubMed

    Compton, Emma L R; Mindell, Joseph A

    2010-09-01

    Bacterial ion channels were known, but only in special cases, such as outer membrane porins in Escherichia coli and bacterial toxins that form pores in their target (bacterial or mammalian) membranes. The exhaustive coverage provided by a decade of bacterial genome sequencing has revealed that ion channels are actually widespread in bacteria, with homologs of a broad range of mammalian channel proteins coded throughout the bacterial and archaeal kingdoms. This review discusses four groups of bacterial channels: porins, mechano-sensitive (MS) channels, channel-forming toxins, and bacterial homologs of mammalian channels. The outer membrane (OM) of gram-negative bacteria blocks access of essential nutrients; to survive, the cell needs to provide a mechanism for nutrients to penetrate the OM. Porin channels provide this access by forming large, nonspecific aqueous pores in the OM that allow ions and vital nutrients to cross it and enter the periplasm. MS channels act as emergency release valves, allowing solutes to rapidly exit the cytoplasm and to dissipate the large osmotic disparity between the internal and external environments. MS channels are remarkable in that they do this by responding to forces exerted by the membrane itself. Some bacteria produce toxic proteins that form pores in trans, attacking and killing other organisms by virtue of their pore formation. The review focuses on those bacterial toxins that kill other bacteria, specifically the class of proteins called colicins. Colicins reveal the dangers of channel formation in the plasma membrane, since they kill their targets with exactly that approach. PMID:26443789

  10. RFI channels

    NASA Technical Reports Server (NTRS)

    Mceliece, R. J.

    1980-01-01

    A class of channel models is presented which exhibit varying burst error severity much like channels encountered in practice. An information-theoretic analysis of these channel models is made, and conclusions are drawn that may aid in the design of coded communication systems for realistic noisy channels.

  11. On-resin assembly of a linkerless lanthanide(III)-based luminescence label and its application to the total synthesis of site-specifically labeled mechanosensitive channels.

    PubMed

    Becker, Christian F W; Clayton, Daniel; Shapovalov, George; Lester, Henry A; Kochendoerfer, Gerd G

    2004-01-01

    A synthesis strategy for the on-resin assembly of luminescent lanthanide chelates from commercially available compounds was developed. Advantages of the approach include the absence of spacers between the metal ion and the attachment site, and the compatibility with typical chemical protein synthesis protection schemes. Methoxycoumarin-labeled lysine and tris(tert-butyl)-DOTA were consecutively coupled with high efficiency to a free amino group in otherwise fully protected peptide segments using standard peptide synthesis methods. Addition of stoichiometric amounts of Tb(3+) to the modified, cleaved, and purified peptides yielded the desired lanthanide chelate. Incorporation of this label into a chemically synthesized, full-length mechanosensitive channel of large conductance (MscL) of E. coli and subsequent reconstitution into vesicles resulted in a functional mechanosensitive channel of comparable conductance to the wild-type channel. However, this channel required increased suction to gate. Excitation of the antenna molecule methoxycoumarin at 336 nm resulted in an emission spectrum typical for Tb(3+) and a luminescence lifetime of 0.67 ms. The location of the probe close to the backbone of this protein may provide precise information about conformational changes during channel opening from LRET studies. PMID:15366968

  12. 33 CFR 117.966 - Galveston Channel.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Texas § 117.966 Galveston Channel. Link to an... across Galveston Channel, mile 4.5 of the Galveston Channel, (GIWW mile 356.1) at Galveston, Texas,...

  13. Molecular determinants mediating gating of Transient Receptor Potential Canonical (TRPC) channels by stromal interaction molecule 1 (STIM1).

    PubMed

    Lee, Kyu Pil; Choi, Seok; Hong, Jeong Hee; Ahuja, Malini; Graham, Sarabeth; Ma, Rong; So, Insuk; Shin, Dong Min; Muallem, Shmuel; Yuan, Joseph P

    2014-03-01

    Transient receptor potential canonical (TRPC) channels mediate a critical part of the receptor-evoked Ca(2+) influx. TRPCs are gated open by the endoplasmic reticulum Ca(2+) sensor STIM1. Here we asked which stromal interaction molecule 1 (STIM1) and TRPC domains mediate the interaction between them and how this interaction is used to open the channels. We report that the STIM1 Orai1-activating region domain of STIM1 interacts with the TRPC channel coiled coil domains (CCDs) and that this interaction is essential for opening the channels by STIM1. Thus, disruption of the N-terminal (NT) CCDs by triple mutations eliminated TRPC surface localization and reduced binding of STIM1 to TRPC1 and TRPC5 while increasing binding to TRPC3 and TRPC6. Single mutations in TRPC1 NT or C-terminal (CT) CCDs reduced interaction and activation of TRPC1 by STIM1. Remarkably, single mutations in the TRPC3 NT CCD enhanced interaction and regulation by STIM1. Disruption in the TRPC3 CT CCD eliminated regulation by STIM1 and the enhanced interaction caused by NT CCD mutations. The NT CCD mutations converted TRPC3 from a TRPC1-dependent to a TRPC1-independent, STIM1-regulated channel. TRPC1 reduced the FRET between BFP-TRPC3 and TRPC3-YFP and between CFP-TRPC3-YFP upon stimulation. Accordingly, knockdown of TRPC1 made TRPC3 STIM1-independent. STIM1 dependence of TRPC3 was reconstituted by the TRPC1 CT CCD alone. Knockout of Trpc1 and Trpc3 similarly inhibited Ca(2+) influx, and inhibition of Trpc3 had no further effect on Ca(2+) influx in Trpc1(-/-) cells. Cell stimulation enhanced the formation of Trpc1-Stim1-Trpc3 complexes. These findings support a model in which the TRPC3 NT and CT CCDs interact to shield the CT CCD from interaction with STIM1. The TRPC1 CT CCD dissociates this interaction to allow the STIM1 Orai1-activating region within STIM1 access to the TRPC3 CT CCD and regulation of TRPC3 by STIM1. These studies provide evidence that the TRPC channel CCDs participate in channel

  14. Molecular Determinants Mediating Gating of Transient Receptor Potential Canonical (TRPC) Channels by Stromal Interaction Molecule 1 (STIM1)*

    PubMed Central

    Lee, Kyu Pil; Choi, Seok; Hong, Jeong Hee; Ahuja, Malini; Graham, Sarabeth; Ma, Rong; So, Insuk; Shin, Dong Min; Muallem, Shmuel; Yuan, Joseph P.

    2014-01-01

    Transient receptor potential canonical (TRPC) channels mediate a critical part of the receptor-evoked Ca2+ influx. TRPCs are gated open by the endoplasmic reticulum Ca2+ sensor STIM1. Here we asked which stromal interaction molecule 1 (STIM1) and TRPC domains mediate the interaction between them and how this interaction is used to open the channels. We report that the STIM1 Orai1-activating region domain of STIM1 interacts with the TRPC channel coiled coil domains (CCDs) and that this interaction is essential for opening the channels by STIM1. Thus, disruption of the N-terminal (NT) CCDs by triple mutations eliminated TRPC surface localization and reduced binding of STIM1 to TRPC1 and TRPC5 while increasing binding to TRPC3 and TRPC6. Single mutations in TRPC1 NT or C-terminal (CT) CCDs reduced interaction and activation of TRPC1 by STIM1. Remarkably, single mutations in the TRPC3 NT CCD enhanced interaction and regulation by STIM1. Disruption in the TRPC3 CT CCD eliminated regulation by STIM1 and the enhanced interaction caused by NT CCD mutations. The NT CCD mutations converted TRPC3 from a TRPC1-dependent to a TRPC1-independent, STIM1-regulated channel. TRPC1 reduced the FRET between BFP-TRPC3 and TRPC3-YFP and between CFP-TRPC3-YFP upon stimulation. Accordingly, knockdown of TRPC1 made TRPC3 STIM1-independent. STIM1 dependence of TRPC3 was reconstituted by the TRPC1 CT CCD alone. Knockout of Trpc1 and Trpc3 similarly inhibited Ca2+ influx, and inhibition of Trpc3 had no further effect on Ca2+ influx in Trpc1−/− cells. Cell stimulation enhanced the formation of Trpc1-Stim1-Trpc3 complexes. These findings support a model in which the TRPC3 NT and CT CCDs interact to shield the CT CCD from interaction with STIM1. The TRPC1 CT CCD dissociates this interaction to allow the STIM1 Orai1-activating region within STIM1 access to the TRPC3 CT CCD and regulation of TRPC3 by STIM1. These studies provide evidence that the TRPC channel CCDs participate in channel gating

  15. Natural killer cells and single nucleotide polymorphisms of specific ion channels and receptor genes in myalgic encephalomyelitis/chronic fatigue syndrome

    PubMed Central

    Marshall-Gradisnik, Sonya; Huth, Teilah; Chacko, Anu; Johnston, Samantha; Smith, Pete; Staines, Donald

    2016-01-01

    Aim The aim of this paper was to determine natural killer (NK) cytotoxic activity and if single nucleotide polymorphisms (SNPs) and genotypes in transient receptor potential (TRP) ion channels and acetylcholine receptors (AChRs) were present in isolated NK cells from previously identified myalgic encephalomyelitis (ME)/chronic fatigue syndrome (CFS) patients. Subjects and methods A total of 39 ME/CFS patients (51.69±2 years old) and 30 unfatigued controls (47.60±2.39 years old) were included in this study. Patients were defined according to the 1994 Centers for Disease Control and Prevention criteria. Flow cytometry protocols were used to examine NK cytotoxic activity. A total of 678 SNPs from isolated NK cells were examined for 21 mammalian TRP ion channel genes and for nine mammalian AChR genes via the Agena Bioscience iPlex Gold assay. SNP association and genotype was determined using analysis of variance and Plink software. Results ME/CFS patients had a significant reduction in NK percentage lysis of target cells (17%±4.68%) compared with the unfatigued control group (31%±6.78%). Of the 678 SNPs examined, eleven SNPs for TRP ion channel genes (TRPC4, TRPC2, TRPM3, and TRPM8) were identified in the ME/CFS group. Five of these SNPs were associated with TRPM3, while the remainder were associated with TRPM8, TRPC2, and TRPC4 (P<0.05). Fourteen SNPs were associated with nicotinic and muscarinic AChR genes: six with CHRNA3, while the remainder were associated with CHRNA2, CHRNB4, CHRNA5, and CHRNE (P<0.05). There were sixteen genotypes identified from SNPs in TRP ion channels and AChRs for TRPM3 (n=5), TRPM8 (n=2), TRPC4 (n=3), TRPC2 (n=1), CHRNE (n=1), CHRNA2 (n=2), CHRNA3 (n=1), and CHRNB4 (n=1) (P<0.05). Conclusion We identified a number of SNPs and genotypes for TRP ion channels and AChRs from isolated NK cells in patients with ME/CFS, suggesting these SNPs and genotypes may be involved in changes in NK cell function and the development of ME/CFS pathology

  16. TRP Channels

    PubMed Central

    Venkatachalam, Kartik; Montell, Craig

    2011-01-01

    The TRP (Transient Receptor Potential) superfamily of cation channels is remarkable in that it displays greater diversity in activation mechanisms and selectivities than any other group of ion channels. The domain organizations of some TRP proteins are also unusual, as they consist of linked channel and enzyme domains. A unifying theme in this group is that TRP proteins play critical roles in sensory physiology, which include contributions to vision, taste, olfaction, hearing, touch, and thermo- and osmosensation. In addition, TRP channels enable individual cells to sense changes in their local environment. Many TRP channels are activated by a variety of different stimuli and function as signal integrators. The TRP superfamily is divided into seven subfamilies: the five group 1 TRPs (TRPC, TRPV, TRPM, TRPN, and TRPA) and two group 2 subfamilies (TRPP and TRPML). TRP channels are important for human health as mutations in at least four TRP channels underlie disease. PMID:17579562

  17. Biochemical and physiological characterization of a new Na(+)-channel specific peptide from the venom of the Argentinean scorpion Tityus trivittatus.

    PubMed

    Coronas, Fredy I V; Diego-García, Elia; Restano-Cassulini, Rita; de Roodt, Adolfo R; Possani, Lourival D

    2015-06-01

    A new peptide with 61 amino acids cross-linked by 4 disulfide bridges, with molecular weight of 6938.12Da, and an amidated C-terminal amino acid residue was purified and characterized. The primary structure was obtained by direct Edman degradation and sequencing its gene. The peptide is lethal to mammals and was shown to be similar (95% identity) to toxin Ts1 (gamma toxin) from the Brazilian scorpion Tityus serrulatus; it was named Tt1g (from T. trivittatus toxin 1 gamma-like). Tt1g was assayed on several sub-types of Na(+)-channels showing displacement of the currents to more negative voltages, being the hNav1.3 the most affected channel. This toxin displays characteristics typical to the β-type sodium scorpion toxins. Lethality tests and physiological assays indicate that this peptide is probably the most important toxic component of this species of scorpion, known for causing human fatalities in the South American continent. PMID:24862827

  18. Application of a compressible flow solver and barotropic cavitation model for the evaluation of the suction head in a low specific speed centrifugal pump impeller channel

    NASA Astrophysics Data System (ADS)

    Limbach, P.; Müller, T.; Skoda, R.

    2015-12-01

    Commonly, for the simulation of cavitation in centrifugal pumps incompressible flow solvers with VOF kind cavitation models are applied. Since the source/sink terms of the void fraction transport equation are based on simplified bubble dynamics, empirical parameters may need to be adjusted to the particular pump operating point. In the present study a barotropic cavitation model, which is based solely on thermodynamic fluid properties and does not include any empirical parameters, is applied on a single flow channel of a pump impeller in combination with a time-explicit viscous compressible flow solver. The suction head curves (head drop) are compared to the results of an incompressible implicit standard industrial CFD tool and are predicted qualitatively correct by the barotropic model.

  19. Specification of skeletal muscle differentiation by repressor element-1 silencing transcription factor (REST)-regulated Kv7.4 potassium channels.

    PubMed

    Iannotti, Fabio Arturo; Barrese, Vincenzo; Formisano, Luigi; Miceli, Francesco; Taglialatela, Maurizio

    2013-02-01

    Changes in the expression of potassium (K(+)) channels is a pivotal event during skeletal muscle differentiation. In mouse C(2)C(12) cells, similarly to human skeletal muscle cells, myotube formation increased the expression of K(v)7.1, K(v)7.3, and K(v)7.4, the last showing the highest degree of regulation. In C(2)C(12) cells, K(v)7.4 silencing by RNA interference reduced the expression levels of differentiation markers (myogenin, myosin heavy chain, troponinT-1, and Pax3) and impaired myotube formation and multinucleation. In K(v)7.4-silenced cells, the differentiation-promoting effect of the K(v)7 activator N-(2-amino-4-(4-fluorobenzylamino)-phenyl)-carbamic acid ethyl ester (retigabine) was abrogated. Expression levels for the repressor element-1 silencing transcription factor (REST) declined during myotube formation. Transcript levels for K(v)7.4, as well as for myogenin, troponinT-1, and Pax3, were reduced by REST overexpression and enhanced upon REST suppression by RNA interference. Four regions containing potential REST-binding sites in the 5' untranslated region and in the first intron of the K(v)7.4 gene were identified by bioinformatic analysis. Chromatin immunoprecipitation assays showed that REST binds to these regions, exhibiting a higher efficiency in myoblasts than in myotubes. These data suggest that K(v)7.4 plays a permissive role in skeletal muscle differentiation and highlight REST as a crucial transcriptional regulator for this K(+) channel subunit. PMID:23242999

  20. TRP channels in disease.

    PubMed

    Jordt, S E; Ehrlich, B E

    2007-01-01

    The transient receptor potential (TRP) channels are a large family of proteins with six main subfamilies termed the TRPC (canonical), TRPV (vanilloid), TRPM (melastatin), TRPP (polycystin), TRPML (mucolipin), and TRPA (ankyrin) groups. The sheer number of different TRPs with distinct functions supports the statement that these channels are involved in a wide range of processes ranging from sensing of thermal and chemical signals to reloading intracellular stores after responding to an extracellular stimulus. Mutations in TRPs are linked to pathophysiology and specific diseases. An understanding of the role of TRPs in normal physiology is just beginning; the progression from mutations in TRPs to pathophysiology and disease will follow. In this review, we focus on two distinct aspects of TRP channel physiology, the role of TRP channels in intracellular Ca2+ homeostasis, and their role in the transduction of painful stimuli in sensory neurons. PMID:18193640

  1. Temperature and species-specific effects on ß3-adrenergic receptor cardiac regulation in two freshwater teleosts: Channel catfish (Ictalurus punctatus) and common carp (Cyprinus carpio).

    PubMed

    Petersen, L H; Burleson, M L; Huggett, D B

    2015-07-01

    β₃-adrenergic receptors (AR) are important in teleost cardiovascular regulation. To date, it is unknown whether temperature acclimation changes ß₃-AR functionality and consequently the involvement of this AR subtype in teleost cardiac regulation. Common carp (Cyprinus carpio) were acclimated at 12 °C or 23 °C (minimum 3 weeks) after which cardiovascular variables (cardiac output (Q), stroke volume (Sv) and heart rate (fH)) were measured upon injection of the ß₃-AR agonist, BRL(37344), and antagonist, SR(59230A). In both 12 °C and 23 °C acclimated carp, BRL(37344) induced significant increases in fH and Q whereas Sv was significantly decreased. While temperature did not affect the change (increase vs. decrease) in cardiac variables, the magnitude and on-set of responses differed. For instance, fH, Sv and Q responded significantly faster to ß₃-AR stimulation in 23 °C carp. In contrast, maximum responses of fH and Q were significantly higher in 23 °C carp whereas the maximum response of Sv was significantly greater in 12 °C carp. These findings suggest that temperature acclimation induced changes in β₃-AR receptor functionality (e.g. density and/or affinity). Stimulation of β₃-ARs in 23 °C acclimated channel catfish (Ictalurus punctatus) caused significant increases in fH, Sv and Q. The increase in Sv was opposite to the decrease observed in 23 °C acclimated common carp. SR(59230A) induced significant decreases in Sv and Q but had no effect in carp (23 °C). Results suggest species diversity in the density and affinity or structure of ß₃-ARs which may explain the different cardiac responses to ß₃-AR ligands. PMID:25882086

  2. Toxin I from the snake Dendroaspis polylepis polylepis: a highly specific blocker of one type of potassium channel in myelinated nerve fiber.

    PubMed

    Benoit, E; Dubois, J M

    1986-07-01

    Toxin I from the venom of the black mamba snake Dendroaspis polylepis polylepis specifically blocks one component of the K-current (IKf1) in the frog node of Ranvier, with a high affinity (Kd = 4 X 10(-10) M) without significantly affecting either the others components of the K current (IKf2 and IKs) or the Na-current. Moreover, for toxin concentrations corresponding to 10- or 100-fold the Kd value, the block was almost irreversible. PMID:2425902

  3. Channel catfish

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This book chapter provides a comprehensive overview of channel catfish aquaculture. Sections include fish biology; commercial culture; culture facilities; production practices; water quality management; nutrition, feeding and feed formulation; infectious diseases; harvesting and processing; and the...

  4. Cholesterol binding to ion channels

    PubMed Central

    Levitan, Irena; Singh, Dev K.; Rosenhouse-Dantsker, Avia

    2014-01-01

    Numerous studies demonstrated that membrane cholesterol is a major regulator of ion channel function. The goal of this review is to discuss significant advances that have been recently achieved in elucidating the mechanisms responsible for cholesterol regulation of ion channels. The first major insight that comes from growing number of studies that based on the sterol specificity of cholesterol effects, show that several types of ion channels (nAChR, Kir, BK, TRPV) are regulated by specific sterol-protein interactions. This conclusion is supported by demonstrating direct saturable binding of cholesterol to a bacterial Kir channel. The second major advance in the field is the identification of putative cholesterol binding sites in several types of ion channels. These include sites at locations associated with the well-known cholesterol binding motif CRAC and its reversed form CARC in nAChR, BK, and TRPV, as well as novel cholesterol binding regions in Kir channels. Notably, in the majority of these channels, cholesterol is suggested to interact mainly with hydrophobic residues in non-annular regions of the channels being embedded in between transmembrane protein helices. We also discuss how identification of putative cholesterol binding sites is an essential step to understand the mechanistic basis of cholesterol-induced channel regulation. Clearly, however, these are only the first few steps in obtaining a general understanding of cholesterol-ion channels interactions and their roles in cellular and organ functions. PMID:24616704

  5. Visual Channel Problems.

    ERIC Educational Resources Information Center

    Mann, Philip H.; Suiter, Patricia A.

    This teacher training module classifies visual channel problems into the following four main areas: visual perception, revisualization (memory), visual-motor (eye-hand coordination), and ocular-motor tasks. Specific deficits are listed under these main headings, behaviors are given to help identify the problem, and ways to improve the condition…

  6. Learning in Tactile Channels

    ERIC Educational Resources Information Center

    Gescheider, George A.; Wright, John H.

    2012-01-01

    Vibrotactile intensity-discrimination thresholds for sinusoidal stimuli applied to the thenar eminence of the hand declined as a function of practice. However, improvement was confined to the tactile information-processing channel in which learning had occurred. Specifically, improvements in performance with training within the Pacinian-corpuscle…

  7. Activation of mammalian target of rapamycin contributes to pain nociception induced in rats by BmK I, a sodium channel-specific modulator.

    PubMed

    Jiang, Feng; Hua, Li-Ming; Jiao, Yun-Lu; Ye, Pin; Fu, Jin; Cheng, Zhi-Jun; Ding, Gang; Ji, Yong-Hua

    2014-02-01

    The mammalian target of rapamycin (mTOR) pathway is essential for maintenance of the sensitivity of certain adult sensory neurons. Here, we investigated whether the mTOR cascade is involved in scorpion envenomation-induced pain hypersensitivity in rats. The results showed that intraplantar injection of a neurotoxin from Buthus martensii Karsch, BmK I (10 μg), induced the activation of mTOR, as well as its downstream molecules p70 ribosomal S6 protein kinase (p70 S6K) and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), in lumbar 5-6 dorsal root ganglia neurons on both sides in rats. The activation peaked at 2 h and recovered 1 day after injection. Compared with the control group, the ratios of p-mTOR/p-p70 S6K/p-4EBP1 in three types of neurons changed significantly. The cell typology of p-mTOR/p-p70 S6K/p-4E-BP1 immuno-reactive neurons also changed. Intrathecal administration of deforolimus, a specific inhibitor of mTOR, attenuated BmK I-induced pain responses (spontaneous flinching, paroxysmal pain-like behavior, and mechanical hypersensitivity). Together, these results imply that the mTOR signaling pathway is mobilized by and contributes to experimental scorpion sting-induced pain. PMID:24132796

  8. Measurement of the B(s)(0) lifetime in the flavor-specific decay channel B(s)(0)→D(s)(-)μ(+)νX.

    PubMed

    Abazov, V M; Abbott, B; Acharya, B S; Adams, M; Adams, T; Agnew, J P; Alexeev, G D; Alkhazov, G; Alton, A; Askew, A; Atkins, S; Augsten, K; Avila, C; Badaud, F; Bagby, L; Baldin, B; Bandurin, D V; Banerjee, S; Barberis, E; Baringer, P; Bartlett, J F; Bassler, U; Bazterra, V; Bean, A; Begalli, M; Bellantoni, L; Beri, S B; Bernardi, G; Bernhard, R; Bertram, I; Besançon, M; Beuselinck, R; Bhat, P C; Bhatia, S; Bhatnagar, V; Blazey, G; Blessing, S; Bloom, K; Boehnlein, A; Boline, D; Boos, E E; Borissov, G; Borysova, M; Brandt, A; Brandt, O; Brock, R; Bross, A; Brown, D; Bu, X B; Buehler, M; Buescher, V; Bunichev, V; Burdin, S; Buszello, C P; Camacho-Pérez, E; Casey, B C K; Castilla-Valdez, H; Caughron, S; Chakrabarti, S; Chan, K M; Chandra, A; Chapon, E; Chen, G; Cho, S W; Choi, S; Choudhary, B; Cihangir, S; Claes, D; Clutter, J; Cooke, M; Cooper, W E; Corcoran, M; Couderc, F; Cousinou, M-C; Cutts, D; Das, A; Davies, G; de Jong, S J; De La Cruz-Burelo, E; Déliot, F; Demina, R; Denisov, D; Denisov, S P; Desai, S; Deterre, C; DeVaughan, K; Diehl, H T; Diesburg, M; Ding, P F; Dominguez, A; Dubey, A; Dudko, L V; Duperrin, A; Dutt, S; Eads, M; Edmunds, D; Ellison, J; Elvira, V D; Enari, Y; Evans, H; Evdokimov, V N; Fauré, A; Feng, L; Ferbel, T; Fiedler, F; Filthaut, F; Fisher, W; Fisk, H E; Fortner, M; Fox, H; Fuess, S; Garbincius, P H; Garcia-Bellido, A; García-González, J A; Gavrilov, V; Geng, W; Gerber, C E; Gershtein, Y; Ginther, G; Gogota, O; Golovanov, G; Grannis, P D; Greder, S; Greenlee, H; Grenier, G; Gris, Ph; Grivaz, J-F; Grohsjean, A; Grünendahl, S; Grünewald, M W; Guillemin, T; Gutierrez, G; Gutierrez, P; Haley, J; Han, L; Harder, K; Harel, A; Hauptman, J M; Hays, J; Head, T; Hebbeker, T; Hedin, D; Hegab, H; Heinson, A P; Heintz, U; Hensel, C; Heredia-De La Cruz, I; Herner, K; Hesketh, G; Hildreth, M D; Hirosky, R; Hoang, T; Hobbs, J D; Hoeneisen, B; Hogan, J; Hohlfeld, M; Holzbauer, J L; Howley, I; Hubacek, Z; Hynek, V; Iashvili, I; Ilchenko, Y; Illingworth, R; Ito, A S; Jabeen, S; Jaffré, M; Jayasinghe, A; Jeong, M S; Jesik, R; Jiang, P; Johns, K; Johnson, E; Johnson, M; Jonckheere, A; Jonsson, P; Joshi, J; Jung, A W; Juste, A; Kajfasz, E; Karmanov, D; Katsanos, I; Kaur, M; Kehoe, R; Kermiche, S; Khalatyan, N; Khanov, A; Kharchilava, A; Kharzheev, Y N; Kiselevich, I; Kohli, J M; Kozelov, A V; Kraus, J; Kumar, A; Kupco, A; Kurča, T; Kuzmin, V A; Lammers, S; Lebrun, P; Lee, H S; Lee, S W; Lee, W M; Lei, X; Lellouch, J; Li, D; Li, H; Li, L; Li, Q Z; Lim, J K; Lincoln, D; Linnemann, J; Lipaev, V V; Lipton, R; Liu, H; Liu, Y; Lobodenko, A; Lokajicek, M; Lopes de Sa, R; Luna-Garcia, R; Lyon, A L; Maciel, A K A; Madar, R; Magaña-Villalba, R; Malik, S; Malyshev, V L; Mansour, J; Martínez-Ortega, J; McCarthy, R; McGivern, C L; Meijer, M M; Melnitchouk, A; Menezes, D; Mercadante, P G; Merkin, M; Meyer, A; Meyer, J; Miconi, F; Mondal, N K; Mulhearn, M; Nagy, E; Narain, M; Nayyar, R; Neal, H A; Negret, J P; Neustroev, P; Nguyen, H T; Nunnemann, T; Orduna, J; Osman, N; Osta, J; Pal, A; Parashar, N; Parihar, V; Park, S K; Partridge, R; Parua, N; Patwa, A; Penning, B; Perfilov, M; Peters, Y; Petridis, K; Petrillo, G; Pétroff, P; Pleier, M-A; Podstavkov, V M; Popov, A V; Prewitt, M; Price, D; Prokopenko, N; Qian, J; Quadt, A; Quinn, B; Ratoff, P N; Razumov, I; Ripp-Baudot, I; Rizatdinova, F; Rominsky, M; Ross, A; Royon, C; Rubinov, P; Ruchti, R; Sajot, G; Sánchez-Hernández, A; Sanders, M P; Santos, A S; Savage, G; Savitskyi, M; Sawyer, L; Scanlon, T; Schamberger, R D; Scheglov, Y; Schellman, H; Schwanenberger, C; Schwienhorst, R; Sekaric, J; Severini, H; Shabalina, E; Shary, V; Shaw, S; Shchukin, A A; Simak, V; Skubic, P; Slattery, P; Smirnov, D; Snow, G R; Snow, J; Snyder, S; Söldner-Rembold, S; Sonnenschein, L; Soustruznik, K; Stark, J; Stoyanova, D A; Strauss, M; Suter, L; Svoisky, P; Titov, M; Tokmenin, V V; Tsai, Y-T; Tsybychev, D; Tuchming, B; Tully, C; Uvarov, L; Uvarov, S; Uzunyan, S; Van Kooten, R; van Leeuwen, W M; Varelas, N; Varnes, E W; Vasilyev, I A; Verkheev, A Y; Vertogradov, L S; Verzocchi, M; Vesterinen, M; Vilanova, D; Vokac, P; Wahl, H D; Wang, M H L S; Warchol, J; Watts, G; Wayne, M; Weichert, J; Welty-Rieger, L; Williams, M R J; Wilson, G W; Wobisch, M; Wood, D R; Wyatt, T R; Xie, Y; Yamada, R; Yang, S; Yasuda, T; Yatsunenko, Y A; Ye, W; Ye, Z; Yin, H; Yip, K; Youn, S W; Yu, J M; Zennamo, J; Zhao, T G; Zhou, B; Zhu, J; Zielinski, M; Zieminska, D; Zivkovic, L

    2015-02-13

    We present an updated measurement of the B(s)(0) lifetime using the semileptonic decays B(s)(0)→D(s)(-)μ(+)νX, with D(s)(-)→ϕπ(-) and ϕ→K(+)K(-) (and the charge conjugate process). This measurement uses the full Tevatron Run II sample of proton-antiproton collisions at √[s]=1.96  TeV, comprising an integrated luminosity of 10.4  fb(-1). We find a flavor-specific lifetime τ(fs)(B(s)(0))=1.479±0.010(stat)±0.021(syst)  ps. This technique is also used to determine the B(0) lifetime using the analogous B(0)→D(-)μ(+)νX decay with D(-)→ϕπ(-) and ϕ→K(+)K(-), yielding τ(B(0))=1.534±0.019(stat)±0.021(syst)  ps. Both measurements are consistent with the current world averages, and the B(s)(0) lifetime measurement is one of the most precise to date. Taking advantage of the cancellation of systematic uncertainties, we determine the lifetime ratio τ(fs)(B(s)(0))/τ(B(0))=0.964±0.013(stat)±0.007(syst). PMID:25723207

  9. Measurement of the B0s lifetime in the flavor-specific decay channel B0s → D-sμ+νX

    DOE PAGESBeta

    Abazov, Victor Mukhamedovich

    2015-02-09

    We present an updated measurement of the B0s lifetime using the semileptonic decays B0s → D-sμ+νX, with D–s → π– and Φ → K+K– (and the charge conjugate process). This measurement uses the full Tevatron Run II sample of proton-antiproton collisions at √s = 1.96 TeV, comprising an integrated luminosity of 10.4 fb–1. We find a flavor-specific lifetime τfs(B0s) = 1.479 ± 0.010(stat) ± 0.021(syst) ps. This technique is also used to determine the B0 lifetime using the analogous B0 → D–μ+νX decay with D– → Φπ– and Φ → K+K–, yielding τ(B0) = 1.534 ± 0.019(stat) ± 0.021(syst) ps.more » Both measurements are consistent with the current world averages, and the B0s lifetime measurement is one of the most precise to date. As a result, taking advantage of the cancellation of systematic uncertainties, we determine the lifetime ratio τfs(B0s)/τ(B0) = 0.964 ± 0.013(stat) ± 0.007(syst).« less

  10. TRP channels.

    PubMed

    Benemei, Silvia; Patacchini, Riccardo; Trevisani, Marcello; Geppetti, Pierangelo

    2015-06-01

    Evidence is accumulating on the role of transient receptor potential (TRP) channels, namely TRPV1, TRPA1, TRPV4 and TRPM8, expressed by C- and Aδ-fibres primary sensory neurons, in cough mechanism. Selective stimuli for these channels have been proven to provoke and, more rarely, to inhibit cough. More importantly, cough threshold to TRP agonists is increased by proinflammatory conditions, known to favour cough. Off-target effects of various drugs, such as tiotropium or desflurane, seem to produce their protective or detrimental actions on airway irritation and cough via TRPV1 and TRPA1, respectively. Thus, TRPs appear to encode the process that initiates or potentiates cough, activated by exogenous irritants and endogenous proinflammatory mediators. More research on TRP channels may result in innovative cough medicines. PMID:25725213

  11. The channels of Mars

    NASA Technical Reports Server (NTRS)

    Baker, Victor R.

    1988-01-01

    The geomorphology of Mars is discussed, focusing on the Martian channels. The great flood channels of Mars, the processes of channel erosion, and dendritic channel networks, are examined. The topography of the Channeled Scabland region of the northwestern U.S. is described and compared to the Martian channels. The importance of water in the evolution of the channel systems is considered.

  12. Starburst Channels

    NASA Technical Reports Server (NTRS)

    2007-01-01

    [figure removed for brevity, see original site] Figure 1

    Translucent carbon dioxide ice covers the polar regions of Mars seasonally. It is warmed and sublimates (evaporates) from below, and escaping gas carves a numerous channel morphologies.

    In this example (figure 1) the channels form a 'starburst' pattern, radiating out into feathery extensions. The center of the pattern is being buried with dust and new darker dust fans ring the outer edges. This may be an example of an expanding morphology, where new channels are formed as the older ones fill and are no longer efficiently channeling the subliming gas out.

    Observation Geometry Image PSP_003443_0980 was taken by the High Resolution Imaging Science Experiment (HiRISE) camera onboard the Mars Reconnaissance Orbiter spacecraft on 21-Apr-2007. The complete image is centered at -81.8 degrees latitude, 76.2 degrees East longitude. The range to the target site was 247.1 km (154.4 miles). At this distance the image scale is 24.7 cm/pixel (with 1 x 1 binning) so objects 74 cm across are resolved. The image shown here has been map-projected to 25 cm/pixel. The image was taken at a local Mars time of 04:52 PM and the scene is illuminated from the west with a solar incidence angle of 71 degrees, thus the sun was about 19 degrees above the horizon. At a solar longitude of 223.4 degrees, the season on Mars is Northern Autumn.

  13. Ultrasound modulates ion channel currents.

    PubMed

    Kubanek, Jan; Shi, Jingyi; Marsh, Jon; Chen, Di; Deng, Cheri; Cui, Jianmin

    2016-01-01

    Transcranial focused ultrasound (US) has been demonstrated to stimulate neurons in animals and humans, but the mechanism of this effect is unknown. It has been hypothesized that US, a mechanical stimulus, may mediate cellular discharge by activating mechanosensitive ion channels embedded within cellular membranes. To test this hypothesis, we expressed potassium and sodium mechanosensitive ion channels (channels of the two-pore-domain potassium family (K2P) including TREK-1, TREK-2, TRAAK; NaV1.5) in the Xenopus oocyte system. Focused US (10 MHz, 0.3-4.9 W/cm(2)) modulated the currents flowing through the ion channels on average by up to 23%, depending on channel and stimulus intensity. The effects were reversible upon repeated stimulation and were abolished when a channel blocker (ranolazine to block NaV1.5, BaCl2 to block K2P channels) was applied to the solution. These data reveal at the single cell level that focused US modulates the activity of specific ion channels to mediate transmembrane currents. These findings open doors to investigations of the effects of  US on ion channels expressed in neurons, retinal cells, or cardiac cells, which may lead to important medical applications. The findings may also pave the way to the development of sonogenetics: a non-invasive, US-based analogue of optogenetics. PMID:27112990

  14. Ultrasound modulates ion channel currents

    PubMed Central

    Kubanek, Jan; Shi, Jingyi; Marsh, Jon; Chen, Di; Deng, Cheri; Cui, Jianmin

    2016-01-01

    Transcranial focused ultrasound (US) has been demonstrated to stimulate neurons in animals and humans, but the mechanism of this effect is unknown. It has been hypothesized that US, a mechanical stimulus, may mediate cellular discharge by activating mechanosensitive ion channels embedded within cellular membranes. To test this hypothesis, we expressed potassium and sodium mechanosensitive ion channels (channels of the two-pore-domain potassium family (K2P) including TREK-1, TREK-2, TRAAK; NaV1.5) in the Xenopus oocyte system. Focused US (10 MHz, 0.3–4.9 W/cm2) modulated the currents flowing through the ion channels on average by up to 23%, depending on channel and stimulus intensity. The effects were reversible upon repeated stimulation and were abolished when a channel blocker (ranolazine to block NaV1.5, BaCl2 to block K2P channels) was applied to the solution. These data reveal at the single cell level that focused US modulates the activity of specific ion channels to mediate transmembrane currents. These findings open doors to investigations of the effects of  US on ion channels expressed in neurons, retinal cells, or cardiac cells, which may lead to important medical applications. The findings may also pave the way to the development of sonogenetics: a non-invasive, US-based analogue of optogenetics. PMID:27112990

  15. Bone morphogenetic protein 2 decreases TRPC expression, store-operated Ca2+ entry, and basal [Ca2+]i in rat distal pulmonary arterial smooth muscle cells

    PubMed Central

    Zhang, Yi; Yang, Kai; Xu, Lei; Lai, Ning; Tian, Lichun; Jiang, Qian; Duan, Xin; Chen, Minsheng

    2013-01-01

    Recent studies indicate that multiple bone morphogenetic protein (BMP) family ligands and receptors are involved in the development of pulmonary arterial hypertension, yet the underlying mechanisms are incompletely understood. Although BMP2 and BMP4 share high homology in amino acid sequence, they appear to exert divergent effects on chronic hypoxic pulmonary hypertension (CHPH). While BMP4 promotes vascular remodeling, BMP2 prevents CHPH. We previously demonstrated that BMP4 upregulates the expression of canonical transient receptor potential channel (TRPC) proteins and, thereby, enhances store-operated Ca2+ entry (SOCE) and elevates intracellular Ca2+ concentration ([Ca2+]i) in pulmonary arterial smooth muscle cells (PASMCs). In this study, we investigated the effects of BMP2 on these variables in rat distal PASMCs. We found that treatment with BMP2 (50 ng/ml, 60 h) inhibited TRPC1, TRPC4, and TRPC6 mRNA and protein expression. Moreover, BMP2 treatment led to reduced SOCE and decreased basal [Ca2+]i in PASMCs. These alterations were associated with decreased PASMC proliferation and migration. Conversely, knockdown of BMP2 with specific small interference RNA resulted in increased cellular levels of TRPC1, TRPC4, and TRPC6 mRNA and protein, enhanced SOCE, elevated basal [Ca2+]i, and increased proliferation and migration of PASMCs. Together, these results indicate that BMP2 participates in regulating Ca2+ signaling in PASMCs by inhibiting TRPC1, TRPC4, and TRPC6 expression, thus leading to reduced SOCE and basal [Ca2+]i and inhibition of cell proliferation and migration. PMID:23447035

  16. Clofilium inhibits Slick and Slack potassium channels

    PubMed Central

    de los Angeles Tejada, Maria; Stolpe, Kathleen; Meinild, Anne-Kristine; Klaerke, Dan A

    2012-01-01

    Slick and Slack high-conductance potassium channels have been recently discovered, and are found in the central nervous system and in the heart. Both channels are activated by Na+ and Cl−, and Slick channels are also inhibited by adenosine triphospate (ATP). An important role of setting the resting membrane potential and controlling the basal excitability of neurons has been suggested for these channels. In addition, no specific blockers for these channels are known up to the present. With the purpose of studying the pharmacological characteristics of Slick and Slack channels, the effects of exposure to the antiarrhythmic compound clofilium were evaluated. Clofilium was able to modulate the activity of Slick and Slack channels effectively, with a stronger effect on Slack than Slick channels. In order to evaluate the pharmacological behavior of Slick and Slack channels further, 38 commonly used potassium channel blockers were tested. Screening of these compounds did not reveal any modulators of Slick and Slack channels, except for clofilium. The present study provides a first approach towards elucidating the pharmacological characteristics of Slick and Slack channels and could be the basis for future studies aimed at developing potent and specific blockers and activators for these channels. PMID:23271893

  17. Channel Networks

    NASA Astrophysics Data System (ADS)

    Rinaldo, Andrea; Rodriguez-Iturbe, Ignacio; Rigon, Riccardo

    This review proceeds from Luna Leopold's and Ronald Shreve's lasting accomplishments dealing with the study of random-walk and topologically random channel networks. According to the random perspective, which has had a profound influence on the interpretation of natural landforms, nature's resiliency in producing recurrent networks and landforms was interpreted to be the consequence of chance. In fact, central to models of topologically random networks is the assumption of equal likelihood of any tree-like configuration. However, a general framework of analysis exists that argues that all possible network configurations draining a fixed area are not necessarily equally likely. Rather, a probability P(s) is assigned to a particular spanning tree configuration, say s, which can be generally assumed to obey a Boltzmann distribution: P(s) % e^-H(s)/T, where T is a parameter and H(s) is a global property of the network configuration s related to energetic characters, i.e. its Hamiltonian. One extreme case is the random topology model where all trees are equally likely, i.e. the limit case for T6 4 . The other extreme case is T 6 0, and this corresponds to network configurations that tend to minimize their total energy dissipation to improve their likelihood. Networks obtained in this manner are termed optimal channel networks (OCNs). Observational evidence suggests that the characters of real river networks are reproduced extremely well by OCNs. Scaling properties of energy and entropy of OCNs suggest that large network development is likely to effectively occur at zero temperature (i.e. minimizing its Hamiltonian). We suggest a corollary of dynamic accessibility of a network configuration and speculate towards a thermodynamics of critical self-organization. We thus conclude that both chance and necessity are equally important ingredients for the dynamic origin of channel networks---and perhaps of the geometry of nature.

  18. Requirements for signaling channel authentication

    SciTech Connect

    Tarman, T.D.

    1995-12-11

    This contribution addresses requirements for ATM signaling channel authentication. Signaling channel authentication is an ATM security service that binds an ATM signaling message to its source. By creating this binding, the message recipient, and even a third party, can confidently verify that the message originated from its claimed source. This provides a useful mechanism to mitigate a number of threats. For example, a denial of service attack which attempts to tear-down an active connection by surreptitiously injecting RELEASE or DROP PARTY messages could be easily thwarted when authenticity assurances are in place for the signaling channel. Signaling channel authentication could also be used to provide the required auditing information for accurate billing which is impervious to repudiation. Finally, depending on the signaling channel authentication mechanism, end-to-end integrity of the message (or at least part of it) can be provided. None of these capabilities exist in the current specifications.

  19. Expression of the muscular dystrophy-associated caveolin-3(P104L) mutant in adult mouse skeletal muscle specifically alters the Ca(2+) channel function of the dihydropyridine receptor.

    PubMed

    Weiss, Norbert; Couchoux, Harold; Legrand, Claude; Berthier, Christine; Allard, Bruno; Jacquemond, Vincent

    2008-11-01

    Caveolins are plasma-membrane-associated proteins potentially involved in a variety of signalling pathways. Different mutations in CAV3, the gene encoding for the muscle-specific isoform caveolin-3 (Cav-3), lead to muscle diseases, but the underlying molecular mechanisms remain largely unknown. Here, we explored the functional consequences of a Cav-3 mutation (P104L) inducing the 1C type limb-girdle muscular dystrophy (LGMD 1C) in human on intracellular Ca(2+) regulation of adult skeletal muscle fibres. A YFP-tagged human Cav-3(P104L) mutant was expressed in vivo in muscle fibres from mouse. Western blot analysis revealed that expression of this mutant led to an approximately 80% drop of the level of endogenous Cav-3. The L-type Ca(2+) current density was found largely reduced in fibres expressing the Cav-3(P104L) mutant, with no change in the voltage dependence of activation and inactivation. Interestingly, the maximal density of intramembrane charge movement was unaltered in the Cav-3(P104L)-expressing fibres, suggesting no change in the total amount of functional voltage-sensing dihydropyridine receptors (DHPRs). Also, there was no obvious alteration in the properties of voltage-activated Ca(2+) transients in the Cav-3(P104L)-expressing fibres. Although the actual role of the Ca(2+) channel function of the DHPR is not clearly established in adult skeletal muscle, its specific alteration by the Cav-3(P104L) mutant suggests that it may be involved in the physiopathology of LGMD 1C. PMID:18509671

  20. An elevation in physical coupling of type 1 inositol 1,4,5-trisphosphate (IP3) receptors to transient receptor potential 3 (TRPC3) channels constricts mesenteric arteries in genetic hypertension.

    PubMed

    Adebiyi, Adebowale; Thomas-Gatewood, Candice M; Leo, M Dennis; Kidd, Michael W; Neeb, Zachary P; Jaggar, Jonathan H

    2012-11-01

    Hypertension is associated with an elevation in agonist-induced vasoconstriction, but mechanisms involved require further investigation. Many vasoconstrictors bind to phospholipase C-coupled receptors, leading to an elevation in inositol 1,4,5-trisphosphate (IP(3)) that activates sarcoplasmic reticulum IP(3) receptors. In cerebral artery myocytes, IP(3) receptors release sarcoplasmic reticulum Ca(2+) and can physically couple to canonical transient receptor potential 3 (TRPC3) channels in a caveolin-1-containing macromolecular complex, leading to cation current activation that stimulates vasoconstriction. Here, we investigated mechanisms by which IP(3) receptors control vascular contractility in systemic arteries and IP(3)R involvement in elevated agonist-induced vasoconstriction during hypertension. Total and plasma membrane-localized TRPC3 protein was ≈2.7- and 2-fold higher in mesenteric arteries of spontaneously hypertensive rats (SHRs) than in Wistar-Kyoto (WKY) rat controls, respectively. In contrast, IP(3)R1, TRPC1, TRPC6, and caveolin-1 expression was similar. TRPC3 expression was also similar in arteries of pre-SHRs and WKY rats. Control, IP(3)-induced and endothelin-1 (ET-1)-induced fluorescence resonance energy transfer between IP3R1 and TRPC3 was higher in SHR than WKY myocytes. IP3-induced cation current was ≈3-fold larger in SHR myocytes. Pyr3, a selective TRPC3 channel blocker, and calmodulin and IP(3) receptor binding domain peptide, an IP(3)R-TRP physical coupling inhibitor, reduced IP(3)-induced cation current and ET-1-induced vasoconstriction more in SHR than WKY myocytes and arteries. Thapsigargin, a sarcoplasmic reticulum Ca(2+)-ATPase blocker, did not alter ET-1-stimulated vasoconstriction in SHR or WKY arteries. These data indicate that ET-1 stimulates physical coupling of IP(3)R1 to TRPC3 channels in mesenteric artery myocytes, leading to vasoconstriction. Furthermore, an elevation in IP(3)R1 to TRPC3 channel molecular coupling augments

  1. Marine Toxins Targeting Ion Channels

    PubMed Central

    Arias, Hugo R.

    2006-01-01

    This introductory minireview points out the importance of ion channels for cell communication. The basic concepts on the structure and function of ion channels triggered by membrane voltage changes, the so-called voltage-gated ion channels (VGICs), as well as those activated by neurotransmitters, the so-called ligand-gated ion channel (LGICs), are introduced. Among the most important VGIC superfamiles, we can name the voltage-gated Na+ (NaV), Ca2+ (CaV), and K+ (KV) channels. Among the most important LGIC super families, we can include the Cys-loop or nicotinicoid, the glutamate-activated (GluR), and the ATP-activated (P2XnR) receptor superfamilies. Ion channels are transmembrane proteins that allow the passage of different ions in a specific or unspecific manner. For instance, the activation of NaV, CaV, or KV channels opens a pore that is specific for Na+, Ca2+, or K+, respectively. On the other hand, the activation of certain LGICs such as nicotinic acetylcholine receptors, GluRs, and P2XnRs allows the passage of cations (e.g., Na+, K+, and/or Ca2+), whereas the activation of other LGICs such as type A γ-butyric acid and glycine receptors allows the passage of anions (e.g., Cl− and/or HCO3−). In this regard, the activation of NaV and CaV as well as ligand-gated cation channels produce membrane depolarization, which finally leads to stimulatory effects in the cell, whereas the activation of KV as well as ligand-gated anion channels induce membrane hyperpolarization that finally leads to inhibitory effects in the cell. The importance of these ion channel superfamilies is emphasized by considering their physiological functions throughout the body as well as their pathophysiological implicance in several neuronal diseases. In this regard, natural molecules, and especially marine toxins, can be potentially used as modulators (e.g., inhibitors or prolongers) of ion channel functions to treat or to alleviate a specific ion channel-linked disease (e

  2. Dense spermatozoa in stallion ejaculates contain lower concentrations of mRNAs encoding the sperm specific calcium channel 1, ornithine decarboxylase antizyme 3, aromatase, and estrogen receptor alpha than less dense spermatozoa.

    PubMed

    Ing, N H; Forrest, D W; Love, C C; Varner, D D

    2014-07-15

    Stallions are unique among livestock in that, like men, they commonly receive medical treatment for subfertility. In both species, about 15% of individuals have normal semen parameters but are subfertile, indicating a need for novel analyses of spermatozoa function. One procedure for improving fertilizing capability of stallions and men is isolation of dense spermatozoa from an ejaculate for use in artificial insemination. In the current study, dense and less dense spermatozoa were purified by density gradient centrifugation from individual ejaculates from seven reproductively normal adult stallions. The RNA isolated from the spermatozoa seemed to be naturally fragmented to an average length of 250 bases, consistent with reports of spermatozoa RNA from other species. The DNAse treatment of RNA prepared from spermatozoa removed any genomic DNA contamination, as assessed by PCR with intron spanning primers for the protamine 1 (PRM1) gene. Concentrations of seven mRNAs in spermatozoa, correlated with the fertility of men and bulls, were quantified by reverse transcription polymerase chain reaction in dense and less dense spermatozoa. Concentrations of four mRNAs were two- to four-fold lower in dense spermatozoa compared with less dense spermatozoa: Encoding the spermatozoa-specific calcium channel (P < 0.03), ornithine decarboxylase antizyme 3 (P < 0.02), aromatase (P < 0.02), and estrogen receptor alpha (P < 0.08). In contrast, concentrations of three other mRNAs, encoding PRM1 and heat shock proteins HSPA8 and DNAJC4, were not different (P > 0.1). These results identify new differences in mRNA concentrations in populations of spermatozoa with dissimilar densities. PMID:24857629

  3. Evaporative cooling in microfluidic channels

    NASA Astrophysics Data System (ADS)

    Maltezos, George; Rajagopal, Aditya; Scherer, Axel

    2006-08-01

    Evaporative cooling is an effective and energy efficient way to rapidly remove heat from a system. Specifically, evaporative cooling in microfluidic channels can provide a cost-effective solution for the cooling of electronic devices and chemical reactors. Here we present microfluidic devices fabricated by using soft-lithography techniques to form simple fluidic junctions between channels carrying refrigerant and channels carrying N2 gas. The effects of channel geometry and delivery pressure on the performance of refrigeration through vaporization of acetone, isopropyl alcohol, and ethyl ether were characterized. By varying gas inlet pressures, refrigerants, and angles of the microfluidic junctions, optimal cooling conditions were found. Refrigeration rates in excess of 40°C/s were measured, and long lasting subzero cooling in the junction could be observed.

  4. Ion channels and migraine

    PubMed Central

    Yan, Jin; Dussor, Gregory

    2014-01-01

    Migraine is one of the most common neurological disorders. Despite its prevalence, the basic physiology of the molecules and mechanisms that contribute to migraine headache is still poorly understood, making the discovery of more effective treatments extremely difficult. The consistent presence of head-specific pain during migraine suggests an important role for activation of the peripheral nociceptors localized to the head. Accordingly, this review will cover the current understanding of the biological mechanisms leading to episodic activation and sensitization of the trigeminovascular pain pathway, focusing on recent advances regarding activation and modulation of ion channels. PMID:24697223

  5. Potassium Channels in Epilepsy.

    PubMed

    Köhling, Rüdiger; Wolfart, Jakob

    2016-01-01

    This review attempts to give a concise and up-to-date overview on the role of potassium channels in epilepsies. Their role can be defined from a genetic perspective, focusing on variants and de novo mutations identified in genetic studies or animal models with targeted, specific mutations in genes coding for a member of the large potassium channel family. In these genetic studies, a demonstrated functional link to hyperexcitability often remains elusive. However, their role can also be defined from a functional perspective, based on dynamic, aggravating, or adaptive transcriptional and posttranslational alterations. In these cases, it often remains elusive whether the alteration is causal or merely incidental. With ∼80 potassium channel types, of which ∼10% are known to be associated with epilepsies (in humans) or a seizure phenotype (in animals), if genetically mutated, a comprehensive review is a challenging endeavor. This goal may seem all the more ambitious once the data on posttranslational alterations, found both in human tissue from epilepsy patients and in chronic or acute animal models, are included. We therefore summarize the literature, and expand only on key findings, particularly regarding functional alterations found in patient brain tissue and chronic animal models. PMID:27141079

  6. Aquaglyceroporins: generalized metalloid channels

    PubMed Central

    Mukhopadhyay, Rita; Bhattacharjee, Hiranmoy; Rosen, Barry P.

    2014-01-01

    Background: Aquaporins (AQPs), members of a superfamily of transmembrane channel proteins, are ubiquitous in all domains of life. They fall into a number of branches that can be functionally categorized into two major sub-groups: i) orthodox aquaporins, which are water-specific channels, and ii) aquaglyceroporins, which allow the transport of water, non-polar solutes, such as urea or glycerol, the reactive oxygen species hydrogen peroxide, and gases such as ammonia, carbon dioxide and nitric oxide and, as described in this review, metalloids. Scope of Review: This review summarizes the key findings that AQP channels conduct bidirectional movement of metalloids into and out of cells. Major Conclusions: As(OH)3 and Sb(OH)3 behave as inorganic molecular mimics of glycerol, a property that allows their passage through AQP channels. Plant AQPs also allow the passage of boron and silicon as their hydroxyacids, boric acid (B(OH)3) and orthosilicic acid (Si(OH)4), respectively. Genetic analysis suggests that germanic acid (GeO2) is also a substrate. While As(III), Sb(III) and Ge(IV) are toxic metalloids, borate (B(III)) and silicate (Si(IV)) are essential elements in higher plants. General Significance: The uptake of environmental metalloids by aquaporins provides an understanding of (i) how toxic elements such as arsenic enter the food chain; (ii) the delivery of arsenic and antimony containing drugs in the treatment of certain forms of leukemia and chemotherapy of diseases caused by pathogenic protozoa; and (iii) the possibility that food plants such as rice could be made safer by genetically modifying them to exclude arsenic while still accumulating boron and silicon. PMID:24291688

  7. Post-Translational Modifications of TRP Channels

    PubMed Central

    Voolstra, Olaf; Huber, Armin

    2014-01-01

    Transient receptor potential (TRP) channels constitute an ancient family of cation channels that have been found in many eukaryotic organisms from yeast to human. TRP channels exert a multitude of physiological functions ranging from Ca2+ homeostasis in the kidney to pain reception and vision. These channels are activated by a wide range of stimuli and undergo covalent post-translational modifications that affect and modulate their subcellular targeting, their biophysical properties, or channel gating. These modifications include N-linked glycosylation, protein phosphorylation, and covalent attachment of chemicals that reversibly bind to specific cysteine residues. The latter modification represents an unusual activation mechanism of ligand-gated ion channels that is in contrast to the lock-and-key paradigm of receptor activation by its agonists. In this review, we summarize the post-translational modifications identified on TRP channels and, when available, explain their physiological role. PMID:24717323

  8. Quantum channels with a finite memory

    NASA Astrophysics Data System (ADS)

    Bowen, Garry; Mancini, Stefano

    2004-01-01

    In this paper we study quantum communication channels with correlated noise effects, i.e., quantum channels with memory. We derive a model for correlated noise channels that includes a channel memory state. We examine the case where the memory is finite, and derive bounds on the classical and quantum capacities. For the entanglement-assisted and unassisted classical capacities it is shown that these bounds are attainable for certain classes of channel. Also, we show that the structure of any finite-memory state is unimportant in the asymptotic limit, and specifically, for a perfect finite-memory channel where no information is lost to the environment, achieving the upper bound implies that the channel is asymptotically noiseless.

  9. M channel enhancers and physiological M channel block.

    PubMed

    Linley, John E; Pettinger, Louisa; Huang, Dongyang; Gamper, Nikita

    2012-02-15

    M-type (Kv7, KCNQ) K(+) channels control the resting membrane potential of many neurons, including peripheral nociceptive sensory neurons. Several M channel enhancers were suggested as prospective analgesics, and targeting M channels specifically in peripheral nociceptors is a plausible strategy for peripheral analgesia. However, receptor-induced inhibition of M channels in nociceptors is often observed in inflammation and may contribute to inflammatory pain. Such inhibition is predominantly mediated by phospholipase C. We investigated four M channel enhancers (retigabine, flupirtine, zinc pyrithione and H(2)O(2)) for their ability to overcome M channel inhibition via two phospholipase C-mediated mechanisms, namely depletion of membrane phosphatidylinositol 4,5-bisphosphate (PIP(2)) and a rise in intracellular Ca(2+) (an action mediated by calmodulin). Data from overexpressed Kv7.2/Kv7.3 heteromers and native M currents in dorsal root ganglion neurons suggest the following conclusions. (i) All enhancers had a dual effect on M channel activity, a negative shift in voltage dependence and an increase of the maximal current at saturating voltages. The enhancers differed in their efficacy to produce these effects. (ii) Both PIP(2) depletion and Ca(2+)/calmodulin strongly reduced the M current amplitude; however, at voltages near the threshold for M channel activation (-60 mV) all enhancers were able to restore M channel activity to a control level or above, while at saturating voltages the effects were more variable. (iii) Receptor-mediated inhibition of M current in nociceptive dorsal root ganglion neurons did not reduce the efficacy of retigabine or flupirtine to hyperpolarize the resting membrane potential. In conclusion, we show that all four M channel enhancers tested could overcome both PIP(2) and Ca(2+)-calmodulin-induced inhibition of Kv7.2/7.3 at voltages close to the threshold for action potential firing (-60 mV) but generally had reduced efficacy at a

  10. A novel Bacillus thuringiensis (PS149B1) containing a Cry34Ab1/Cry35Ab1 binary toxin specific for the western corn rootworm Diabrotica virgifera virgifera LeConte forms ion channels in lipid membranes.

    PubMed

    Masson, Luke; Schwab, George; Mazza, Alberto; Brousseau, Roland; Potvin, Lena; Schwartz, Jean-Louis

    2004-09-28

    The binary Bacillus thuringiensis PS149B1 insecticidal crystal (Cry) protein is comprised of two components, Cry34Ab1, a 14-kDa protein, and Cry35Ab1, a 44-kDa protein, the combination of which forms a novel binary toxin active on western corn rootworm larvae. The permeabilizing behavior of the native binary toxin and its two individual components expressed as recombinant proteins was studied using calcein efflux determination in liposomes and by ion channel activity measurements in planar lipid bilayers (PLBs). Data obtained with solubilized native PS149B1 binary protein revealed it to be a pore-forming toxin that can permeabilize liposomes and form ion channels ( approximately 300-900 pS) in PLBs at pH 5.5 but not pH 9.0. The 14-kDa component of the toxin also formed ion channels ( approximately 15-300 pS) at pH 5.5 but did not insert easily in PLBs. While the 44-kDa moiety did seldomly form resolvable ion channels ( approximately 15-750 pS) in PLBs, it did destabilize the membranes. It showed pH-dependent truncation to a stable 40-kDa protein. The purified 40-kDa truncated product formed channels ( approximately 10-450 pS) in PLBs at pH 5.5. At that same pH, while a 3:1 molar mixture (14:44 kDa) of the individual components of the toxin induced channel activity that resembled that of the 14-kDa component alone, the 3:1 molar mixture of the 14-kDa component and 40-kDa truncated product induced channel activity ( approximately 20-800 pS) similar to that of PS149B1 in planar lipid bilayers. We conclude that the overall membrane permeabilization process of Cry34Ab1/Cry35Ab1 is a result of ion channel formation. PMID:15379574

  11. 33 CFR 117.T966 - Pelican Island Causeway, Galveston Channel.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Texas § 117.T966 Pelican... Channel, mile 4.5 of the Galveston Channel, (GIWW mile 356.1) at Galveston, Texas, need not open for...

  12. 33 CFR 117.977 - Pelican Island Causeway, Galveston Channel.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Texas § 117.977 Pelican... Galveston Channel, mile 4.5 of the Galveston Channel, (GIWW mile 356.1) at Galveston, Texas, must open...

  13. Ion channels, channelopathies, and tooth formation.

    PubMed

    Duan, X

    2014-02-01

    The biological functions of ion channels in tooth development vary according to the nature of their gating, the species of ions passing through those gates, the number of gates, localization of channels, tissue expressing the channel, and interactions between cells and microenvironment. Ion channels feature unique and specific ion flux in ameloblasts, odontoblasts, and other tooth-specific cell lineages. Both enamel and dentin have active chemical systems orchestrating a variety of ion exchanges and demineralization and remineralization processes in a stage-dependent manner. An important role for ion channels is to regulate and maintain the calcium and pH homeostasis that are critical for proper enamel and dentin biomineralization. Specific functions of chloride channels, TRPVs, calcium channels, potassium channels, and solute carrier superfamily members in tooth formation have been gradually clarified in recent years. Mutations in these ion channels or transporters often result in disastrous changes in tooth development. The channelopathies of tooth include altered eruption (CLCN7, KCNJ2, TRPV3), root dysplasia (CLCN7, KCNJ2), amelogenesis imperfecta (KCNJ1, CFTR, AE2, CACNA1C, GJA1), dentin dysplasia (CLCN5), small teeth (CACNA1C, GJA1), tooth agenesis (CLCN7), and other impairments. The mechanisms leading to tooth channelopathies are primarily related to pH regulation, calcium homeostasis, or other alterations of the niche for tooth eruption and development. PMID:24076519

  14. Single-channel properties of α3β4, α3β4α5 and α3β4β2 nicotinic acetylcholine receptors in mice lacking specific nicotinic acetylcholine receptor subunits

    PubMed Central

    Ciuraszkiewicz, Anna; Schreibmayer, Wolfgang; Platzer, Dieter; Orr-Urtreger, Avi; Scholze, Petra; Huck, Sigismund

    2013-01-01

    Previous attempts to measure the functional properties of recombinant nicotinic acetylcholine receptors (nAChRs) composed of known receptor subunits have yielded conflicting results. The use of knockout mice that lack α5, β2, α5β2 or α5β2α7 nAChR subunits enabled us to measure the single-channel properties of distinct α3β4, α3β4α5 and α3β4β2 receptors in superior cervical ganglion (SCG) neurons. Using this approach, we found that α3β4 receptors had a principal conductance level of 32.6 ± 0.8 pS (mean ± SEM) and both higher and lower secondary conductance levels. α3β4α5 receptors had the same conductance as α3β4 receptors, but differed from α3β4 receptors by having an increased channel open time and increased burst duration. By contrast, α3β4β2 receptors differed from α3β4 and α3β4α5 receptors by having a significantly smaller conductance level (13.6 ± 0.5 pS). After dissecting the single-channel properties of these receptors using our knockout models, we then identified these properties – and hence the receptors themselves – in wild-type SCG neurons. This study is the first to identify the single-channel properties of distinct neuronal nicotinic receptors in their native environment. PMID:23613527

  15. mRNA distribution analysis of human TRPC family in CNS and peripheral tissues.

    PubMed

    Riccio, Antonio; Medhurst, Andrew D; Mattei, Cesar; Kelsell, Rosemary E; Calver, Andrew R; Randall, Andrew D; Benham, Christopher D; Pangalos, Menelas N

    2002-12-30

    The mammalian homologues of the Drosophila transient receptor potential (TRP) channel are plasma membrane proteins involved in the regulation of cellular Ca(2+) influx. These ion channels can be activated subsequent to either depletion of Ca(2+) from internal stores or through receptor-mediated processes. The mRNA expression patterns of several individual mammalian short transient receptor potential channels (TRPCs) have been described. Cross-comparisons between these data, however, are at best difficult predominantly due to the non-quantitative methods used. Furthermore there is limited data on the expression of TRPC family members in human tissues. In the present study we used a single technique, namely TaqMan real-time quantitative RT-PCR, to investigate the mRNA distribution of human TRPC1, TRPC3, TRPC4, TRPC5, TRPC6 and TRPC7 (hTRPCs) in discrete human brain areas, peripheral tissues as well as a panel of cell-lines. All hTRPCs studied were widely expressed within CNS and significant peripheral expression was often observed. Despite this, each channel exhibited a distinctive hallmark distribution profile. hTRPC1 was widely expressed in CNS and peripheral tissues, whereas hTRPC3 and hTRPC5 were predominantly expressed in tissues of CNS. hTRPC4 mRNA was detected in CNS and certain peripheral tissues such as bone, heart and prostate. hTRPC6 was homogeneously expressed throughout the CNS and peripheral tissues with the highest levels in placenta and lung. hTRPC7 mRNA was also broadly expressed in CNS as well as some peripheral tissues. The pattern of expression of the TRPCs was quite different in the various cell lines examined. TRPC3 and TRPC6 were selectively present in HEK-293 cells whilst TRPC1 was broadly distributed in the cell lines analyzed. In contrast TRPC4 and TRPC5 mRNAs were predominantly expressed in HK-2 and HEK-293 cell lines respectively. TRPC7 was selectively expressed in COS-1, COS-7 and HK-2 cell lines. These results show tissue- and cell-specific

  16. 33 CFR 117.720 - Great Channel.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Great Channel. 117.720 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.720 Great Channel. The draw of the County of Cape May bridge, mile 0.7, between Stone Harbor and Nummy Island, shall open on signal...

  17. 33 CFR 117.720 - Great Channel.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 1 2013-07-01 2013-07-01 false Great Channel. 117.720 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.720 Great Channel. The draw of the County of Cape May bridge, mile 0.7, between Stone Harbor and Nummy Island, shall open on signal...

  18. 33 CFR 117.720 - Great Channel.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 1 2014-07-01 2014-07-01 false Great Channel. 117.720 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.720 Great Channel. The draw of the County of Cape May bridge, mile 0.7, between Stone Harbor and Nummy Island, shall open on signal...

  19. 33 CFR 117.720 - Great Channel.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Great Channel. 117.720 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.720 Great Channel. The draw of the County of Cape May bridge, mile 0.7, between Stone Harbor and Nummy Island, shall open on signal...

  20. 33 CFR 117.720 - Great Channel.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Great Channel. 117.720 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.720 Great Channel. The draw of the County of Cape May bridge, mile 0.7, between Stone Harbor and Nummy Island, shall open on signal...

  1. 33 CFR 117.1005 - Chincoteague Channel.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Chincoteague Channel. 117.1005 Section 117.1005 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Virginia § 117.1005 Chincoteague Channel. The draw...

  2. 33 CFR 117.1005 - Chincoteague Channel.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 1 2014-07-01 2014-07-01 false Chincoteague Channel. 117.1005 Section 117.1005 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Virginia § 117.1005 Chincoteague Channel. The draw...

  3. 33 CFR 117.1005 - Chincoteague Channel.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Chincoteague Channel. 117.1005 Section 117.1005 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Virginia § 117.1005 Chincoteague Channel. The draw...

  4. 33 CFR 117.1005 - Chincoteague Channel.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Chincoteague Channel. 117.1005 Section 117.1005 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Virginia § 117.1005 Chincoteague Channel. The draw...

  5. Equalization in redundant channels

    NASA Technical Reports Server (NTRS)

    Tulpule, Bhalchandra R. (Inventor); Collins, Robert E. (Inventor); Cominelli, Donald F. (Inventor); O'Neill, Richard D. (Inventor)

    1988-01-01

    A miscomparison between a channel's configuration data base and a voted system configuration data base in a redundant channel system having identically operating, frame synchronous channels triggers autoequalization of the channel's historical signal data bases in a hierarchical, chronological manner with that of a correctly operating channel. After equalization, symmetrization of the channel's configuration data base with that of the system permits upgrading of the previously degraded channel to full redundancy. An externally provided equalization command, e.g., manually actuated, can also trigger equalization.

  6. [Synopsis about the hypothesis of "information channel" of channel-collateral system].

    PubMed

    Chang, Xi-Lang

    2008-10-01

    The author of the present paper founded a theorem about the "incompleteness of single channel structure" (nerve, blood vessel, lymphatic, interspace, aperture, etc.) through quantitative and qualitative analysis about the economic information channel in the human body, which eliminates the probability of single channel structure in the information channel of channel (meridian)-collateral system. After comprehensive analysis on the current researches, the author puts forward a neodoxy, i.e., the body "information channel" structure of the channel-collateral system, mainly follows the distribution regularity of systemic statistics, and is not a single specific entity; various layers of the information channel in the main stems of the channel-collaterals are composed of optimized structure tissues. Hence, the structure of this information channel of channel-collateral system is an overall-optimized, sequential and compatible systemic structure. From this neodoxy, the author brings forward a working principle of channel-collaterals, which is supported theoretically by bio-auxology. The longitudinal distribution of the main stems of meridian-collaterals is considered to result from that in the process of the animal evolution, in the animals moving forward, the microscopic complicated movement of intracorporeal information and energy molecules is related to the forward macroscopic and non-uniform movement of organism in trans-measure. Its impulse and kinetic momentum forms a main vector in the longitudinal direction of the body (the direction of the main stem of channel-collaterals). In order to adapt to and utilize natural regularities, the main stems of the channel-collaterals gradually differentiate and evolve in the living organism, forming a whole system. The "hypothesis of biological origin of channel-collateral system" and "that of information channel of the channel-collaterals in the body" constitute a relatively complete theoretical system framework. PMID

  7. Channel nut tool

    DOEpatents

    Olson, Marvin

    2016-01-12

    A method, system, and apparatus for installing channel nuts includes a shank, a handle formed on a first end of a shank, and an end piece with a threaded shaft configured to receive a channel nut formed on the second end of the shaft. The tool can be used to insert or remove a channel nut in a channel framing system and then removed from the channel nut.

  8. A Micromechanical RF Channelizer

    NASA Astrophysics Data System (ADS)

    Akgul, Mehmet

    The power consumption of a radio generally goes as the number and strength of the RF signals it must process. In particular, a radio receiver would consume much less power if the signal presented to its electronics contained only the desired signal in a tiny percent bandwidth frequency channel, rather than the typical mix of signals containing unwanted energy outside the desired channel. Unfortunately, a lack of filters capable of selecting single channel bandwidths at RF forces the front-ends of contemporary receivers to accept unwanted signals, and thus, to operate with sub-optimal efficiency. This dissertation focuses on the degree to which capacitive-gap transduced micromechanical resonators can achieve the aforementioned RF channel-selecting filters. It aims to first show theoretically that with appropriate scaling capacitive-gap transducers are strong enough to meet the needed coupling requirements; and second, to fully detail an architecture and design procedure needed to realize said filters. Finally, this dissertation provides an actual experimentally demonstrated RF channel-select filter designed using the developed procedures and confirming theoretical predictions. Specifically, this dissertation introduces four methods that make possible the design and fabrication of RF channel-select filters. The first of these introduces a small-signal equivalent circuit for parallel-plate capacitive-gap transduced micromechanical resonators that employs negative capacitance to model the dependence of resonance frequency on electrical stiffness in a way that facilitates the analysis of micromechanical circuits loaded with arbitrary electrical impedances. The new circuit model not only correctly predicts the dependence of electrical stiffness on the impedances loading the input and output electrodes of parallel-plate capacitive-gap transduced micromechanical device, but does so in a visually intuitive way that identifies current drive as most appropriate for

  9. Channel degradation in southeastern Nebraska Rivers

    USGS Publications Warehouse

    Wahl, Kenneth L.; Weiss, Linda S.

    1995-01-01

    Many stream channels in southeastern Nebraska were dredged and straightened during 1904-15. The resulting channels were both shorter and steeper than the original channels. Tests for time trends were conducted using the nonparametric Kendall tau test to see if the channels have responded to these changes. Tests were conducted on the stages associated with specific discharges and on measurement characteristics at gaging stations. Tests also were conducted on hydrologic forcing variables (annual mean precipitation, annual peak discharges, annual mean discharge, and annual mean base flows). The null hypothesis (that the data were free from trend) was rejected for stages associated with the mean of the annual discharges for 6 of 7 gaging stations in the study area, but was accepted for all 3 gages on the main stem of the Missouri River. The trends at the 6 streamflow gaging stations were for decreasing stages (degrading channels) for specific discharges. The rates of change ranged from about 0.2 to 0.5 m per decade. Mean stream bed elevations computed for individual discharge measurements at these streamflow gaging stations confirmed that the channels are degrading. However, neither the precipitation nor flow variables show evidence of trends. The tendency for the channels to degrade thus cannot be attributed to changes in runoff characteristics and are assumed to be a response to the channel modifications in the early 1900's. Indications are that the channels presently are continuing to degrade.

  10. The Origins of Transmembrane Ion Channels

    NASA Technical Reports Server (NTRS)

    Pohorille, Andrew; Wilson, Michael A.

    2012-01-01

    Even though membrane proteins that mediate transport of ions and small molecules across cell walls are among the largest and least understood biopolymers in contemporary cells, it is still possible to shed light on their origins and early evolution. The central observation is that transmembrane portions of most ion channels are simply bundles of -helices. By combining results of experimental and computer simulation studies on synthetic models and natural channels, mostly of non-genomic origin, we show that the emergence of -helical channels was protobiologically plausible, and did not require highly specific amino acid sequences. Despite their simple structure, such channels could possess properties that, at the first sight, appear to require markedly larger complexity. Specifically, we explain how the antiamoebin channels, which are made of identical helices, 16 amino acids in length, achieve efficiency comparable to that of highly evolved channels. We further show that antiamoebin channels are extremely flexible, compared to modern, genetically coded channels. On the basis of our results, we propose that channels evolved further towards high structural complexity because they needed to acquire stable rigid structures and mechanisms for precise regulation rather than improve efficiency. In general, even though architectures of membrane proteins are not nearly as diverse as those of water-soluble proteins, they are sufficiently flexible to adapt readily to the functional demands arising during evolution.

  11. The Earliest Ion Channels

    NASA Astrophysics Data System (ADS)

    Pohorille, A.; Wilson, M. A.; Wei, C.

    2009-12-01

    Supplying protocells with ions required assistance from channels spanning their membrane walls. The earliest channels were most likely short proteins that formed transmembrane helical bundles surrounding a water-filled pore. These simple aggregates were capable of transporting ions with efficiencies comparable to those of complex, contemporary ion channels. Channels with wide pores exhibited little ion selectivity but also imposed only modest constraints on amino acid sequences of channel-forming proteins. Channels with small pores could have been selective but also might have required a more precisely defined sequence of amino acids. In contrast to modern channels, their protocellular ancestors had only limited capabilities to regulate ion flux. It is postulated that subsequent evolution of ion channels progressed primarily to acquire precise regulation, and not high efficiency or selectivity. It is further proposed that channels and the surrounding membranes co-evolved.

  12. Microfluidic channel fabrication method

    DOEpatents

    Arnold, Don W.; Schoeniger, Joseph S.; Cardinale, Gregory F.

    2001-01-01

    A new channel structure for microfluidic systems and process for fabricating this structure. In contrast to the conventional practice of fabricating fluid channels as trenches or grooves in a substrate, fluid channels are fabricated as thin walled raised structures on a substrate. Microfluidic devices produced in accordance with the invention are a hybrid assembly generally consisting of three layers: 1) a substrate that can or cannot be an electrical insulator; 2) a middle layer, that is an electrically conducting material and preferably silicon, forms the channel walls whose height defines the channel height, joined to and extending from the substrate; and 3) a top layer, joined to the top of the channels, that forms a cover for the channels. The channels can be defined by photolithographic techniques and are produced by etching away the material around the channel walls.

  13. Structure of Thermally Activated TRP Channels

    PubMed Central

    Cohen, Matthew R.; Moiseenkova-Bell, Vera Y.

    2015-01-01

    Temperature sensation is important for adaptation and survival of organisms. While temperature has the potential to affect all biological macromolecules, organisms have evolved specific thermosensitive molecular detectors that are able to transduce temperature changes into physiologically relevant signals. Among these thermosensors are ion channels from the transient receptor potential (TRP) family. Prime candidates include TRPV1–4, TRPA1, and TRPM8 (the so-called “thermoTRP” channels), which are expressed in sensory neurons and gated at specific temperatures. Electrophysiological and thermodynamic approaches have been employed to determine the nature by which thermoTRPs detect temperature and couple temperature changes to channel gating. To further understand how thermoTRPs sense temperature, high-resolution structures of full-length thermoTRPs channels will be required. Here, we will discuss current progress in unraveling the structures of thermoTRP channels. PMID:25366237

  14. Gramicidin Channels: Versatile Tools

    NASA Astrophysics Data System (ADS)

    Andersen, Olaf S.; Koeppe, Roger E., II; Roux, Benoît

    Gramicidin channels are miniproteins in which two tryptophan-rich subunits associate by means of transbilayer dimerization to form the conducting channels. That is, in contrast to other ion channels, gramicidin channels do not open and close; they appear and disappear. Each subunit in the bilayer-spanning channel is tied to the bilayer/solution interface through hydrogen bonds that involve the indole NH groups as donors andwater or the phospholipid backbone as acceptors. The channel's permeability characteristics are well-defined: gramicidin channels are selective for monovalent cations, with no measurable permeability to anions or polyvalent cations; ions and water move through a pore whose wall is formed by the peptide backbone; and the single-channel conductance and cation selectivity vary when the amino acid sequence is varied, even though the permeating ions make no contact with the amino acid side chains. Given the plethora of available experimental information—for not only the wild-type channels but also for channels formed by amino acid-substituted gramicidin analogues—gramicidin channels continue to provide important insights into the microphysics of ion permeation through bilayer-spanning channels. For similar reasons, gramicidin channels constitute a system of choice for evaluating computational strategies for obtaining mechanistic insights into ion permeation through the more complex channels formed by integral membrane proteins.

  15. Energy, time, and channel evolution in catastrophically disturbed fluvial systems

    USGS Publications Warehouse

    Simon, A.

    1992-01-01

    Specific energy is shown to decrease nonlinearly with time during channel evolution and provides a measure of reductions in available energy at the channel bed. Data from two sites show convergence towards a minimum specific energy with time. Time-dependent reductions in specific energy at a point act in concert with minimization of the rate of energy dissipation over a reach during channel evolution as the fluvial systems adjust to a new equilibrium.

  16. A Micromechanical RF Channelizer

    NASA Astrophysics Data System (ADS)

    Akgul, Mehmet

    The power consumption of a radio generally goes as the number and strength of the RF signals it must process. In particular, a radio receiver would consume much less power if the signal presented to its electronics contained only the desired signal in a tiny percent bandwidth frequency channel, rather than the typical mix of signals containing unwanted energy outside the desired channel. Unfortunately, a lack of filters capable of selecting single channel bandwidths at RF forces the front-ends of contemporary receivers to accept unwanted signals, and thus, to operate with sub-optimal efficiency. This dissertation focuses on the degree to which capacitive-gap transduced micromechanical resonators can achieve the aforementioned RF channel-selecting filters. It aims to first show theoretically that with appropriate scaling capacitive-gap transducers are strong enough to meet the needed coupling requirements; and second, to fully detail an architecture and design procedure needed to realize said filters. Finally, this dissertation provides an actual experimentally demonstrated RF channel-select filter designed using the developed procedures and confirming theoretical predictions. Specifically, this dissertation introduces four methods that make possible the design and fabrication of RF channel-select filters. The first of these introduces a small-signal equivalent circuit for parallel-plate capacitive-gap transduced micromechanical resonators that employs negative capacitance to model the dependence of resonance frequency on electrical stiffness in a way that facilitates the analysis of micromechanical circuits loaded with arbitrary electrical impedances. The new circuit model not only correctly predicts the dependence of electrical stiffness on the impedances loading the input and output electrodes of parallel-plate capacitive-gap transduced micromechanical device, but does so in a visually intuitive way that identifies current drive as most appropriate for

  17. Fading channel simulator

    SciTech Connect

    Argo, P.E.; Fitzgerald, T.J.

    1991-12-31

    This invention relates to high frequency (HF) radio signal propagation through fading channels and, more particularly, to simulation of fading channels in order to characterize HF radio system performance in transmitting and receiving signals through such fading channels. Fading channel effects on a transmitted communication signal are simulated with both frequency and time variations using a channel scattering function to affect the transmitted signal. A conventional channel scattering function is converted to a series of channel realizations by multiplying the square root of the channel scattering function by a complex number of which the real and imaginary parts are each independent variables. The two-dimensional inverse-FFT of this complex-valued channel realization yields a matrix of channel coefficients that provide a complete frequency-time description of the channel. The transmitted radio signal is segmented to provide a series of transmitted signal and each segment is subject to FFT to generate a series of signal coefficient matrices. The channel coefficient matrices and signal coefficient matrices are then multiplied and subjected to inverse-FFT to output a signal representing the received affected radio signal. A variety of channel scattering functions can be used to characterize the response of a transmitter-receiver system to such atmospheric effects.

  18. 33 CFR 117.721 - Grassy Sound Channel.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 1 2014-07-01 2014-07-01 false Grassy Sound Channel. 117.721... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.721 Grassy Sound Channel. The draw of the Grassy Sound Channel Bridge, mile 1.0 in Middle Township, shall open on signal from 6 a.m. to 8...

  19. 33 CFR 117.721 - Grassy Sound Channel.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Grassy Sound Channel. 117.721... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.721 Grassy Sound Channel. The draw of the Grassy Sound Channel Bridge, mile 1.0 in Middle Township, shall open on signal from 6 a.m. to 8...

  20. 33 CFR 117.721 - Grassy Sound Channel.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 1 2013-07-01 2013-07-01 false Grassy Sound Channel. 117.721... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.721 Grassy Sound Channel. The draw of the Grassy Sound Channel Bridge, mile 1.0 in Middle Township, shall open on signal from 6 a.m. to 8...

  1. 33 CFR 117.721 - Grassy Sound Channel.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Grassy Sound Channel. 117.721... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.721 Grassy Sound Channel. The draw of the Grassy Sound Channel Bridge, mile 1.0 in Middle Township, shall open on signal from 6 a.m. to 8...

  2. 33 CFR 117.721 - Grassy Sound Channel.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Grassy Sound Channel. 117.721... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements New Jersey § 117.721 Grassy Sound Channel. The draw of the Grassy Sound Channel Bridge, mile 1.0 in Middle Township, shall open on signal from 6 a.m. to 8...

  3. Calcium channel blocker overdose

    MedlinePlus

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

  4. Optimal channels for channelized quadratic estimators.

    PubMed

    Kupinski, Meredith K; Clarkson, Eric

    2016-06-01

    We present a new method for computing optimized channels for estimation tasks that is feasible for high-dimensional image data. Maximum-likelihood (ML) parameter estimates are challenging to compute from high-dimensional likelihoods. The dimensionality reduction from M measurements to L channels is a critical advantage of channelized quadratic estimators (CQEs), since estimating likelihood moments from channelized data requires smaller sample sizes and inverting a smaller covariance matrix is easier. The channelized likelihood is then used to form ML estimates of the parameter(s). In this work we choose an imaging example in which the second-order statistics of the image data depend upon the parameter of interest: the correlation length. Correlation lengths are used to approximate background textures in many imaging applications, and in these cases an estimate of the correlation length is useful for pre-whitening. In a simulation study we compare the estimation performance, as measured by the root-mean-squared error (RMSE), of correlation length estimates from CQE and power spectral density (PSD) distribution fitting. To abide by the assumptions of the PSD method we simulate an ergodic, isotropic, stationary, and zero-mean random process. These assumptions are not part of the CQE formalism. The CQE method assumes a Gaussian channelized likelihood that can be a valid for non-Gaussian image data, since the channel outputs are formed from weighted sums of the image elements. We have shown that, for three or more channels, the RMSE of CQE estimates of correlation length is lower than conventional PSD estimates. We also show that computing CQE by using a standard nonlinear optimization method produces channels that yield RMSE within 2% of the analytic optimum. CQE estimates of anisotropic correlation length estimation are reported to demonstrate this technique on a two-parameter estimation problem. PMID:27409452

  5. Solution structure of the potassium channel inhibitor agitoxin 2: caliper for probing channel geometry.

    PubMed Central

    Krezel, A. M.; Kasibhatla, C.; Hidalgo, P.; MacKinnon, R.; Wagner, G.

    1995-01-01

    The structure of the potassium channel blocker agitoxin 2 was solved by solution NMR methods. The structure consists of a triple-stranded antiparallel beta-sheet and a single helix covering one face of the beta-sheet. The cysteine side chains connecting the beta-sheet and the helix form the core of the molecule. One edge of the beta-sheet and the adjacent face of the helix form the interface with the Shaker K+ channel. The fold of agitoxin is homologous to the previously determined folds of scorpion venom toxins. However, agitoxin 2 differs significantly from the other channel blockers in the specificity of its interactions. This study was thus focused on a precise characterization of the surface residues at the face of the protein interacting with the Shaker K+ channel. The rigid toxin molecule can be used to estimate dimensions of the potassium channel. Surface-exposed residues, Arg24, Lys27, and Arg31 of the beta-sheet, have been identified from mutagenesis studies as functionally important for blocking the Shaker K+ channel. The sequential and spatial locations of Arg24 and Arg31 are not conserved among the homologous toxins. Knowledge on the details of the channel-binding sites of agitoxin 2 formed a basis for site-directed mutagenesis studies of the toxin and the K+ channel sequences. Observed interactions between mutated toxin and channel are being used to elucidate the channel structure and mechanisms of channel-toxin interactions. PMID:8520473

  6. Channel catfish pond fertilization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The most successful aquaculture enterprise in the U.S. is channel catfish Ictalurus punctatus production. In the U.S., 257 million kg of channel catfish were sold in 2007 at a value of $455 million. Large-scale commercial channel catfish culture began in the late 1950s and expanded rapidly from 1978...

  7. Connexin Channel Modulators and their Mechanisms of Action

    PubMed Central

    Verselis, Vytas K.; Srinivas, Miduturu

    2013-01-01

    Gap junction channels and hemichannels formed by the connexin family of proteins play important roles in many aspects of tissue homeostasis in the brain and in other organs. In addition, connexin channels have been proposed as pharmacological targets in the treatment of a number of human disorders. In this review, we describe the connexin-subtype selectivity and specificity of pharmacological agents that are commonly used to modulate connexin channels. We also highlight recent progress made towards identifying new agents for connexin channels that act in a selective and specific manner. Finally, we discuss developing insights into possible mechanisms by which these agents modulate connexin channel function. PMID:23597508

  8. Caveolae, Ion Channels and Cardiac Arrhythmias

    PubMed Central

    Balijepalli, Ravi C.; Kamp, Timothy J.

    2009-01-01

    Caveolae are specialized membrane microdomains enriched in cholesterol and sphingolipids which are present in multiple cell types including cardiomyocytes. Along with the essential scaffolding protein caveolin-3, a number of different ion channels and transporters have been localized to caveolae in the heart including L-type Ca2+ channels (Cav1.2), Na+ channels (Nav1.5), pacemaker channels (HCN4), Na+/Ca2+ exchnager (NCX1) and others. Closely associated with these channels are specific macromolecular signaling complexes that provide highly localized regulation of the channels. Mutations in the caveolin-3 gene (CAV3) have been linked with the congenital long QT syndrome (LQT9), and mutations in caveolar-localized ion channels may contribute to other inherited arrhythmias. Changes in the caveolar microdomain in acquired heart disease may also lead to dysregulation and dysfunction of ion channels, altering the risk of arrhythmias in conditions such as heart failure. This review highlights the existing evidence identifying and characterizing ion channels localized to caveolae in cardiomyocytes and their role in arrhythmogenesis. PMID:19351512

  9. Connexin channel permeability to cytoplasmic molecules.

    PubMed

    Harris, Andrew L

    2007-01-01

    Connexin channels are known to be permeable to a variety of cytoplasmic molecules. The first observation of second messenger junctional permeability, made approximately 30 years ago, sparked broad interest in gap junction channels as mediators of intercellular molecular signaling. Since then, much has been learned about the diversity of connexin channels with regard to isoform diversity, tissue and developmental distribution, modes of channel regulation, assembly, expression, biochemical modification and permeability, all of which appear to be dynamically regulated. This information has expanded the potential roles of connexin channels in development, physiology and disease, and made their elucidation much more complex--30 years ago such an orchestra of junctional dynamics was unanticipated. Only recently, however, have investigators been able to directly address, in this more complex framework, the key issue: what specific biological molecules, second messengers and others, are able to permeate the various types of connexin channels, and how well? An important related issue, given the ever-growing list of connexin-related pathologies, is how these permeabilities are altered by disease-causing connexin mutations. Together, many studies show that a variety of cytoplasmic molecules can permeate the different types of connexin channels. A few studies reveal differences in permeation by different molecules through a particular type of connexin channel, and differences in permeation by a particular molecule through different types of connexin channels. This article describes and evaluates the various methods used to obtain these data, presents an annotated compilation of the results, and discusses the findings in the context of what can be inferred about mechanism of selectivity and potential relevance to signaling. The data strongly suggest that highly specific interactions take place between connexin pores and specific biological molecular permeants, and that those

  10. Injury-specific functional alteration of N-type voltage-gated calcium channels in synaptic transmission of primary afferent C-fibers in the rat spinal superficial dorsal horn.

    PubMed

    Takasu, Keiko; Ogawa, Koichi; Minami, Kazuhisa; Shinohara, Shunji; Kato, Akira

    2016-02-01

    We investigated functional alterations of voltage-gated calcium channels (VGCCs) in excitatory synaptic transmission from primary afferent A- and C-fibers after peripheral nerve injury. Patch-clamp recordings were performed on substantia gelatinosa (SG) neurons of spinal cord slices with an attached dorsal root, prepared from L5 spinal nerve-ligated (SNL) rats. The effects of neuronal VGCC blockers, ω-conotoxin GVIA (ω-CgTX) for N-type channels and ω-agatoxin IVA (ω-AgaIVA) for P/Q-type channels, on evoked excitatory postsynaptic currents (eEPSCs) by stimulation of A- or C-fibers were studied. Besides, electrophysiological assay using dorsal root ganglion (DRG) and immunohistochemistry were done. In naïve rats, ω-CgTX (0.1-1μM) reduced more effectively A-fiber eEPSCs than C-fiber ones. After nerve injury, ω-CgTX produced great inhibition of C-fiber eEPSCs in slices with the injured L5 dorsal root of SNL model rats, as compared to sham-operated rats. By contrast, in slices with the non-injured L4 one, inhibitory effects of ω-CgTX were not changed. This occurred concurrently with increased expression of N-type VGCCs in L5 spinal dorsal horn and with enhanced Ca(2+) currents through N-type VGCCs in small-sized (C-type) L5 DRG. In terms of A-fiber eEPSCs, ω-CgTX elicited similar inhibition in nerve-injured and sham-operated rats. ω-AgaIVA (0.1μM) had less effect on A- or C-fiber eEPSCs. These results indicate that N-type, but not P/Q-type, VGCCs mainly contribute to excitatory synaptic transmission from A- and C-fibers in the spinal dorsal horn. More importantly, following nerve injury, the functional contribution of N-type VGCCs to nociceptive transmission is increased in the pre-synaptic terminals of injured C-fibers. PMID:26708163

  11. Mechanically Activated Ion Channels.

    PubMed

    Ranade, Sanjeev S; Syeda, Ruhma; Patapoutian, Ardem

    2015-09-23

    Mechanotransduction, the conversion of physical forces into biochemical signals, is essential for various physiological processes such as the conscious sensations of touch and hearing, and the unconscious sensation of blood flow. Mechanically activated (MA) ion channels have been proposed as sensors of physical force, but the identity of these channels and an understanding of how mechanical force is transduced has remained elusive. A number of recent studies on previously known ion channels along with the identification of novel MA ion channels have greatly transformed our understanding of touch and hearing in both vertebrates and invertebrates. Here, we present an updated review of eukaryotic ion channel families that have been implicated in mechanotransduction processes and evaluate the qualifications of the candidate genes according to specified criteria. We then discuss the proposed gating models for MA ion channels and highlight recent structural studies of mechanosensitive potassium channels. PMID:26402601

  12. C. elegans TRP channels

    PubMed Central

    Xiao, Rui; Xu, X.Z. Shawn

    2010-01-01

    TRP (transient receptor potential) channels represent a superfamily of cation channels found in all eukaryotes. The C. elegans genome encodes seventeen TRP channels covering all of the seven TRP subfamilies. Genetic analyses in C. elegans have implicated TRP channels in a wide spectrum of behavioral and physiological processes, ranging from sensory transduction (e.g. chemosensation, touch sensation, proprioception and osmosensation) to fertilization, drug dependence, organelle biogenesis, apoptosis, gene expression, and neurotransmitter/hormone release. Many C. elegans TRP channels share similar activation and regulatory mechanisms with their vertebrate counterparts. Studies in C. elegans have also revealed some previously unrecognized functions and regulatory mechanisms of TRP channels. C. elegans represents an excellent genetic model organism for the study of function and regulation of TRP channels in vivo. PMID:21290304

  13. MgtE From Rhizobium leguminosarum Is a Mg²⁺ Channel Essential for Growth at Low pH and N2 Fixation on Specific Plants.

    PubMed

    Hood, Graham; Karunakaran, Ramakrishnan; Downie, J Allan; Poole, Philip

    2015-12-01

    MgtE is predicted to be a Rhizobium leguminosarum channel and is essential for growth when both Mg²⁺ is limiting and the pH is low. N₂was only fixed at 8% of the rate of wild type when the crop legume Pisum sativum was inoculated with an mgtE mutant of R. leguminosarum and, although bacteroids were present, they were few in number and not fully developed. R. leguminosarum MgtE was also essential for N₂fixation on the native legume Vicia hirsuta but not when in symbiosis with Vicia faba. The importance of MgtE and the relevance of the contrasting phenotypes is discussed. PMID:26422403

  14. Specificity, affinity and efficacy of iota-conotoxin RXIA, an agonist of voltage-gated sodium channels NaV1.2, 1.6 and 1.7

    PubMed Central

    Fiedler, Brian; Zhang, Min-Min; Buczek, Oga; Azam, Layla; Bulaj, Grzegorz; Norton, Raymond S; Olivera, Baldomero M; Yoshikami, Doju

    2009-01-01

    The excitotoxic conopeptide ι-RXIA induces repetitive action potentials in frog motor axons and seizures upon intracranial injection into mice. We recently discovered that ι-RXIA shifts the voltage-dependence of activation of voltage-gated sodium channel NaV1.6 to a more hyperpolarized level. Here, we performed voltage-clamp experiments to examine its activity against rodent NaV1.1 through NaV1.7 co-expressed with the β1 subunit in Xenopus oocytes and NaV1.8 in dissociated mouse DRG neurons. The order of sensitivity to ι-RXIA was NaV1.6 > 1.2 > 1.7, and the remaining subtypes were insensitive. The time course of ι-RXIA-activity on NaV1.6 during exposure to different peptide concentrations were well fit by single-exponential curves that provided kobs. The plot of kobs versus [ι-RXIA] was linear, consistent with a bimolecular reaction with a Kd of ~3 μM, close to the steady-state EC50 of ~2 μM. ι-RXIA has an unusual residue, D-Phe, and the analog with an L-Phe instead, ι-RXIA[L-Phe44], had a two-fold lower affinity and two-fold faster off-rate than ι-RXIA on NaV1.6 and furthermore was inactive on NaV1.2. ι-RXIA induced repetitive action potentials in mouse sciatic nerve with conduction velocities of both A- and C-fibers, consistent with the presence of NaV1.6 at nodes of Ranvier as well as in unmyelinated axons. Sixteen peptides homologous to ι-RXIA have been identified from a single species of Conus, so these peptides represent a rich family of novel sodium channel-targeting ligands. PMID:18486102

  15. Herbal Compounds and Toxins Modulating TRP Channels

    PubMed Central

    Vriens, Joris; Nilius, Bernd; Vennekens, Rudi

    2008-01-01

    Although the benefits are sometimes obvious, traditional or herbal medicine is regarded with skepticism, because the mechanism through which plant compounds exert their powers are largely elusive. Recent studies have shown however that many of these plant compounds interact with specific ion channels and thereby modulate the sensing mechanism of the human body. Especially members of the Transient Receptor Potential (TRP) channels have drawn large attention lately as the receptors for plant-derived compounds such as capsaicin and menthol. TRP channels constitute a large and diverse family of channel proteins that can serve as versatile sensors that allow individual cells and entire organisms to detect changes in their environment. For this family, a striking number of empirical views have turned into mechanism-based actions of natural compounds. In this review we will give an overview of herbal compounds and toxins, which modulate TRP channels. PMID:19305789

  16. Modeling ion channels: Past, present, and future

    PubMed Central

    2014-01-01

    Ion channels are membrane-bound enzymes whose catalytic sites are ion-conducting pores that open and close (gate) in response to specific environmental stimuli. Ion channels are important contributors to cell signaling and homeostasis. Our current understanding of gating is the product of 60 plus years of voltage-clamp recording augmented by intervention in the form of environmental, chemical, and mutational perturbations. The need for good phenomenological models of gating has evolved in parallel with the sophistication of experimental technique. The goal of modeling is to develop realistic schemes that not only describe data, but also accurately reflect mechanisms of action. This review covers three areas that have contributed to the understanding of ion channels: traditional Eyring kinetic theory, molecular dynamics analysis, and statistical thermodynamics. Although the primary emphasis is on voltage-dependent channels, the methods discussed here are easily generalized to other stimuli and could be applied to any ion channel and indeed any macromolecule. PMID:24935742

  17. TRPC channels as signal transducers.

    PubMed

    Glazebrook, Patricia A; Schilling, William P; Kunze, Diana L

    2005-10-01

    The study of the TRPC cation channels as signal transducers in sensory neurons is in its infancy. Mechanoreceptors that monitor arterial pressure are prime candidates for the involvement of TRPC channels as either primary mechanical transducers or as modulators of the transduction process. Their activity patterns can be regulated by growth factors such as BDNF and by a variety of ligands that activate Gq-coupled receptors, mechanisms that have been shown in heterologous expression systems to activate TRPC channels. We investigated the distribution of TRPC1 and TRPC3-7 in nodose sensory neurons and in their peripheral axons that terminate as mechanosensitive receptors in the aortic arch of the rat. Using immunocytochemical techniques we identified these six TRPC proteins in the soma of the nodose neurons but only TRPC1 and TRPC3-5 were found to distribute to the peripheral axons and the mechanosensory terminals. TRPC1 and TRPC3 extended into the low threshold complex sensory endings with very strong labeling. In contrast, TRPC4 and TRPC5 were found primarily in major branches of the receptor but immunoreactivity was weak in the region where mechanotransduction is presumed to occur. Terminals arising from unmyelinated fibers also expressed TRPC1 and TRPC3-5 but not all fibers expressed all of the channels suggesting that specific TRPC protein may be aligned with previously described subclasses of the unmyelinated C-fibers. PMID:15971079

  18. Fading channel simulator

    DOEpatents

    Argo, Paul E.; Fitzgerald, T. Joseph

    1993-01-01

    Fading channel effects on a transmitted communication signal are simulated with both frequency and time variations using a channel scattering function to affect the transmitted signal. A conventional channel scattering function is converted to a series of channel realizations by multiplying the square root of the channel scattering function by a complex number of which the real and imaginary parts are each independent variables. The two-dimensional inverse-FFT of this complex-valued channel realization yields a matrix of channel coefficients that provide a complete frequency-time description of the channel. The transmitted radio signal is segmented to provide a series of transmitted signal and each segment is subject to FFT to generate a series of signal coefficient matrices. The channel coefficient matrices and signal coefficient matrices are then multiplied and subjected to inverse-FFT to output a signal representing the received affected radio signal. A variety of channel scattering functions can be used to characterize the response of a transmitter-receiver system to such atmospheric effects.

  19. P/Q-type calcium channel modulators

    PubMed Central

    Nimmrich, V; Gross, G

    2012-01-01

    P/Q-type calcium channels are high-voltage-gated calcium channels contributing to vesicle release at synaptic terminals. A number of neurological diseases have been attributed to malfunctioning of P/Q channels, including ataxia, migraine and Alzheimer's disease. To date, only two specific P/Q-type blockers are known: both are peptides deriving from the spider venom of Agelenopsis aperta, ω-agatoxins. Other peptidic calcium channel blockers with activity at P/Q channels are available, albeit with less selectivity. A number of low molecular weight compounds modulate P/Q-type currents with different characteristics, and some exhibit a peculiar bidirectional pattern of modulation. Interestingly, there are a number of therapeutics in clinical use, which also show P/Q channel activity. Because selectivity as well as the exact mode of action is different between all P/Q-type channel modulators, the interpretation of clinical and experimental data is complicated and needs a comprehensive understanding of their target profile. The situation is further complicated by the fact that information on potency varies vastly in the literature, which may be the result of different experimental systems, conditions or the splice variants of the P/Q channel. This review attempts to provide a comprehensive overview of the compounds available that affect the P/Q-type channel and should help with the interpretation of results of in vitro experiments and animal models. It also aims to explain some clinical observations by implementing current knowledge about P/Q channel modulation of therapeutically used non-selective drugs. Chances and challenges of the development of P/Q channel-selective molecules are discussed. PMID:22670568

  20. Voltage-Gated Calcium Channels in Nociception

    NASA Astrophysics Data System (ADS)

    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.

  1. The roles of K(+) channels in cancer.

    PubMed

    Pardo, Luis A; Stühmer, Walter

    2014-01-01

    Potassium channels are transmembrane proteins that selectively facilitate the flow of potassium ions down an electrochemical gradient. These molecules have been studied in great detail in the context of cell excitability, but their roles in less cell type-specific functions, such as cell proliferation, angiogenesis or cell migration, have only recently been assessed. Moreover, the importance of these channels for tumour biology has become evident. This, coupled with the fact that they are accessible proteins and that their pharmacology is well characterized, has increased the interest in investigating potassium channels as therapeutic targets in cancer patients. PMID:24336491

  2. Jamming in Vertical Channels

    NASA Astrophysics Data System (ADS)

    Baxter, G. William; Steel, Fiona

    2011-03-01

    We study jamming of low aspect-ratio cylindrical Delrin grains in a vertical channel. Grain heights are less than their diameter so the grains resemble antacid tablets, coins, or poker chips. These grains are allowed to fall through a vertical channel with a square cross section where the channel width is greater than the diameter of a grain and constant throughout the length of the channel with no obstructions or constrictions. Grains are sometimes observed to form jams, stable structures supported by the channel walls with no support beneath them. The probability of jam occurrence and the strength or robustness of a jam is effected by grain and channel sizes. We will present experimental measurements of the jamming probability and jam strength in this system and discuss the relationship of these results to other experiments and theories. Supported by an Undergraduate Research Grant from Penn State Erie, The Behrend College.

  3. Symmetrization for redundant channels

    NASA Technical Reports Server (NTRS)

    Tulplue, Bhalchandra R. (Inventor); Collins, Robert E. (Inventor)

    1988-01-01

    A plurality of redundant channels in a system each contain a global image of all the configuration data bases in each of the channels in the system. Each global image is updated periodically from each of the other channels via cross channel data links. The global images of the local configuration data bases in each channel are separately symmetrized using a voting process to generate a system signal configuration data base which is not written into by any other routine and is available for indicating the status of the system within each channel. Equalization may be imposed on a suspect signal and a number of chances for that signal to heal itself are provided before excluding it from future votes. Reconfiguration is accomplished upon detecting a channel which is deemed invalid. A reset function is provided which permits an externally generated reset signal to permit a previously excluded channel to be reincluded within the system. The updating of global images and/or the symmetrization process may be accomplished at substantially the same time within a synchronized time frame common to all channels.

  4. Phosphoinositides regulate ion channels

    PubMed Central

    Hille, Bertil; Dickson, Eamonn J.; Kruse, Martin; Vivas, Oscar; Suh, Byung-Chang

    2014-01-01

    Phosphoinositides serve as signature motifs for different cellular membranes and often are required for the function of membrane proteins. Here, we summarize clear evidence supporting the concept that many ion channels are regulated by membrane phosphoinositides. We describe tools used to test their dependence on phosphoinositides, especially phosphatidylinositol 4,5-bisphosphate, and consider mechanisms and biological meanings of phosphoinositide regulation of ion channels. This lipid regulation can underlie changes of channel activity and electrical excitability in response to receptors. Since different intracellular membranes have different lipid compositions, the activity of ion channels still in transit towards their final destination membrane may be suppressed until they reach an optimal lipid environment. PMID:25241941

  5. Oxidation of ion channels in the aging nervous system.

    PubMed

    Patel, Rahul; Sesti, Federico

    2016-05-15

    Ion channels are integral membrane proteins that allow passive diffusion of ions across membranes. In neurons and in other excitable cells, the harmonious coordination between the numerous types of ion channels shape and propagate electrical signals. Increased accumulation of reactive oxidative species (ROS), and subsequent oxidation of proteins, including ion channels, is a hallmark feature of aging and may contribute to cell failure as a result. In this review we discuss the effects of ROS on three major types of ion channels of the central nervous system, namely the potassium (K(+)), calcium (Ca(2+)) and sodium (Na(+)) channels. We examine two general mechanisms through which ROS affect ion channels: via direct oxidation of specific residues and via indirect interference of pathways that regulate the channels. The overall status of the present studies indicates that the interaction of ion channels with ROS is multimodal and pervasive in the central nervous system and likely constitutes a general mechanism of aging susceptibility. PMID:26947620

  6. Optical stimulation in mice lacking the TRPV1 channel

    NASA Astrophysics Data System (ADS)

    Suh, Eul; Izzo Matic, Agnella; Otting, Margarete; Walsh, Joseph T., Jr.; Richter, Claus-Peter

    2009-02-01

    Lasers can be used to stimulate neural tissue, including the sciatic nerve or auditory neurons. Wells and coworkers suggested that neural tissue is likely stimulated by heat.[1,2] Ion channels that can be activated by heat are the TRPV channels, a subfamily of the Transient Receptor Potential (TRP) ion channels. TRPV channels are nonselective cation channels found in sensory neurons involved in nociception. In addition to various chemicals, TRPV channels can also be thermally stimulated. The activation temperature for the different TRPV channels varies and is 43°C for TRPV1 and 39°C for TRPV3. By performing an immunohistochemical staining procedure on frozen 20 μm cochlear slices using a primary TRPV1 antibody, we observed specific immunostaining of the spiral ganglion cells. Here we show that in mice that lack the gene for the TRPV1 channel optical radiation cannot evoke action potentials on the auditory nerve.

  7. NMR and restrained molecular dynamics study of the three-dimensional solution structure of toxin FS2, a specific blocker of the L-type calcium channel, isolated from black mamba venom.

    PubMed

    Albrand, J P; Blackledge, M J; Pascaud, F; Hollecker, M; Marion, D

    1995-05-01

    The three-dimensional solution structure of toxin FS2, a 60-residue polypeptide isolated from the venom of black mamba snake (Dendroaspis polylepis polylepis), has been determined by nuclear magnetic resonance spectroscopy. Using 600 NOE constraints and 55 dihedral angle constraints, a set of 20 structures obtained from distance-geometry calculations was further refined by molecular dynamics calculations using a combined simulated annealing-restrained MD protocol. The resulting 20 conformers, taken to represent the solution structure, give an average rmsd of 1.2 A for their backbone atoms, relative to the average structure. The overall resulting three-fingered structure is similar to those already observed in several postsynaptic neurotoxins, cardiotoxins, and fasciculins, which all share with toxin FS2 the same network of four disulfide bridges. The overall concavity of the molecule, considered as a flat bottomed dish, is oriented toward the C-terminal loop of the molecule. This orientation is similar to that of fasciculins and cardiotoxins but opposite to that of neurotoxins. On the basis of the local rms displacements between the 20 conformers, the structure of the first loop appears to be less well defined in FS2 than in the previously reported neurotoxin structures, but fasciculin 1 shows a similar trend with particularly high temperature factors for this part of the X-ray structure. The concave side which presents most of the positively charged residues is quite similar in FS2 and fasciculin 1. The main difference is shown by the convex side of the third loop, mostly hydrophobic in FS2, in contrast to the pair of negatively charged aspartates in fasciculin 1. This difference could be one of the factors leading to the distinct pharmacological properties-L-type calcium channel blocker for FS2 and cholinesterase inhibitor for fasciculin--observed for these two subgroups of the "angusticeps-type" toxins. PMID:7727450

  8. Ion channels in cancer: future perspectives and clinical potential.

    PubMed

    Lang, Florian; Stournaras, Christos

    2014-03-19

    Ion transport across the cell membrane mediated by channels and carriers participate in the regulation of tumour cell survival, death and motility. Moreover, the altered regulation of channels and carriers is part of neoplastic transformation. Experimental modification of channel and transporter activity impacts tumour cell survival, proliferation, malignant progression, invasive behaviour or therapy resistance of tumour cells. A wide variety of distinct Ca(2+) permeable channels, K(+) channels, Na(+) channels and anion channels have been implicated in tumour growth and metastasis. Further experimental information is, however, needed to define the specific role of individual channel isoforms critically important for malignancy. Compelling experimental evidence supports the assumption that the pharmacological inhibition of ion channels or their regulators may be attractive targets to counteract tumour growth, prevent metastasis and overcome therapy resistance of tumour cells. This short review discusses the role of Ca(2+) permeable channels, K(+) channels, Na(+) channels and anion channels in tumour growth and metastasis and the therapeutic potential of respective inhibitors. PMID:24493756

  9. Novel perspectives in cancer therapy: Targeting ion channels.

    PubMed

    Arcangeli, Annarosa; Becchetti, Andrea

    2015-01-01

    By controlling ion fluxes at multiple time scales, ion channels shape rapid cell signals, such as action potential and synaptic transmission, as well as much slower processes, such as mitosis and cell migration. As is currently increasingly recognized, a variety of channel types are involved in cancer hallmarks, and regulate specific stages of neoplastic progression. Long-term in vitro work has established that inhibition of these ion channels impairs the growth of cancer cells. Recently, these studies have been followed up in vivo, hence revealing that ion channels constitute promising pharmacological targets in oncology. The channel proteins can be often accessed from the extracellular milieu, which allows use of lower drug doses and decrease untoward toxicity. However, because of the central physiological roles exerted by ion channels in excitable cells, other types of side effects may arise, the gravest of which is cardiac arrhythmia. A paradigmatic case is offered by Kv11.1 (hERG1) channels. HERG1 blockers attenuate the progression of both hematologic malignancies and solid tumors, but may also lead to the lengthening of the electrocardiographic QT interval, thus predisposing the patient to ventricular arrhythmias. These side effects can be avoided by specifically inhibiting the channel isoforms which are highly expressed in certain tumors, such as Kv11.1B and the neonatal forms of voltage-gated Na(+) channels. Preclinical studies are also being explored in breast and prostate cancer (targeting voltage-gated Na(+) channels), and gliomas (targeting CLC-3). Overall, the possible approaches to improve the efficacy and safety of ion channel targeting in oncology include: (1) the development of specific inhibitors for the channel subtypes expressed in specific tumors; (2) drug delivery into the tumor by using antibodies or nanotechnology-based approaches; (3) combination regimen therapy and (4) blocking specific conformational states of the ion channel. We believe

  10. Basaltic Lava Channels

    NASA Astrophysics Data System (ADS)

    Cashman, K. V.; Griffiths, R. W.; Kerr, R. C.

    2004-12-01

    In Hawaii, the mode of lava transport - through open channels or through insulating lava tubes - determines the thermal, rheological, and emplacement history of a lava flow. Most Hawaiian lavas are erupted at near-liquidus temperatures and are therefore crystal-poor; lava transport through open channels allows rapid cooling and consequent rapid increases in lava crystallinity. Solidified aa flows resulting from channelized flow are typically fine-grained throughout their thickness, indicating cooling of the entire flow thickness during transport. In contrast, transport of lava through insulating tubes permits flow over long distances with little cooling. Flows emerging from such tubes typically have pahoehoe flow surfaces with glassy crusts. Groundmass textures that coarsen from the flow rind to the interior reflect rates of post-emplacement, rather than syn-emplacement, cooling. To distinguish eruption conditions that result in lava channels from those that allow formation of lava tubes, we have performed a series of laboratory experiments involving injection of PEG 600 (a wax with a Newtonian rheology and freezing temperature of 19ºC) into cold water through both uniform and non-uniform sloping channels. In uniform channels, tube formation can be distinguished from open channel flow using a dimensionless parameter based on a solidification time scale, an advection time scale, and a Rayleigh number that describes convection by heat loss from crust-free shear zones. Theoretical analysis predicts that in the open channel regime, the width of the crust (dc) will vary with the channel width (W) as dc = W5/3. Crustal coverage of non-uniform channels in both laboratory experiments and field examples from Kilauea Volcano, Hawaii, is consistent with this prediction. However, experiments in non-uniform channels illustrate additional controls on the surface coverage of lava channels. Most important is crustal extension resulting from flow acceleration through constrictions

  11. Dysfunctional HCN ion channels in neurological diseases

    PubMed Central

    DiFrancesco, Jacopo C.; DiFrancesco, Dario

    2015-01-01

    Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are expressed as four different isoforms (HCN1-4) in the heart and in the central and peripheral nervous systems. HCN channels are activated by membrane hyperpolarization at voltages close to resting membrane potentials and carry the hyperpolarization-activated current, dubbed If (funny current) in heart and Ih in neurons. HCN channels contribute in several ways to neuronal activity and are responsible for many important cellular functions, including cellular excitability, generation, and modulation of rhythmic activity, dendritic integration, transmission of synaptic potentials, and plasticity phenomena. Because of their role, defective HCN channels are natural candidates in the search for potential causes of neurological disorders in humans. Several data, including growing evidence that some forms of epilepsy are associated with HCN mutations, support the notion of an involvement of dysfunctional HCN channels in different experimental models of the disease. Additionally, some anti-epileptic drugs are known to modify the activity of the Ih current. HCN channels are widely expressed in the peripheral nervous system and recent evidence has highlighted the importance of the HCN2 isoform in the transmission of pain. HCN channels are also present in the midbrain system, where they finely regulate the activity of dopaminergic neurons, and a potential role of these channels in the pathogenesis of Parkinson’s disease has recently emerged. The function of HCN channels is regulated by specific accessory proteins, which control the correct expression and modulation of the neuronal Ih current. Alteration of these proteins can severely interfere with the physiological channel function, potentially predisposing to pathological conditions. In this review we address the present knowledge of the association between HCN dysfunctions and neurological diseases, including clinical, genetic, and physiopathological

  12. The allocation of cargo to channel missions

    SciTech Connect

    Liu, Cheng; Harrison, G.

    1992-01-01

    Each month the armed services provide a forecast of tons of cargo by channel to MAC. The purpose of the Channels Allocation Algorithm is to allocate cargo requirements to specific Channel Missions. The objective of the allocation is algorithm is to minimize frequency and cargo requirements shortfall. The constraints on the allocation model include flying hours, channel frequencies, mission structure, mission operation days, and aircraft capacity. Cargo requirements shortfall is defined as the tonnage of cargo not moved from the airfields in the United States that are channel staging points to overseas locations. Channel frequencies are defined by the number of times a destination is served by an origin in one month. The mission structures are defined as sets of missions usually in the form of circuit. Mission operating days are determined by the operating day rules for the month, or they can be input by the user for an individual month. One of the assumptions in this model is that there is only one transshipment allowed between any origin and a destination if there is no mission that actually connects the stations. The transshipment stations are also restricted in that only certain stations can serve as transshipment stations. The Channels Allocation Algorithm consists of two linear programs that incorporate three objectives. The objectives are: (1) to minimize that number of frequency channels not met. (2) to minimize cargo shortfall, and (3) to minimize operating cost. The first linear program minimizes frequency channels not met, subject to the mission structure, number of times the mission operates, and total flying hours available. The second linear program minimizes the fleet operating cost cargo handling cost, and cargo shortfall, subject to frequency channels met by the first linear program, aircraft capacity, and total flying hours available. This document is comprised of viewgraphs.

  13. The allocation of cargo to channel missions

    SciTech Connect

    Liu, Cheng; Harrison, G.

    1992-06-01

    Each month the armed services provide a forecast of tons of cargo by channel to MAC. The purpose of the Channels Allocation Algorithm is to allocate cargo requirements to specific Channel Missions. The objective of the allocation is algorithm is to minimize frequency and cargo requirements shortfall. The constraints on the allocation model include flying hours, channel frequencies, mission structure, mission operation days, and aircraft capacity. Cargo requirements shortfall is defined as the tonnage of cargo not moved from the airfields in the United States that are channel staging points to overseas locations. Channel frequencies are defined by the number of times a destination is served by an origin in one month. The mission structures are defined as sets of missions usually in the form of circuit. Mission operating days are determined by the operating day rules for the month, or they can be input by the user for an individual month. One of the assumptions in this model is that there is only one transshipment allowed between any origin and a destination if there is no mission that actually connects the stations. The transshipment stations are also restricted in that only certain stations can serve as transshipment stations. The Channels Allocation Algorithm consists of two linear programs that incorporate three objectives. The objectives are: (1) to minimize that number of frequency channels not met. (2) to minimize cargo shortfall, and (3) to minimize operating cost. The first linear program minimizes frequency channels not met, subject to the mission structure, number of times the mission operates, and total flying hours available. The second linear program minimizes the fleet operating cost cargo handling cost, and cargo shortfall, subject to frequency channels met by the first linear program, aircraft capacity, and total flying hours available. This document is comprised of viewgraphs.

  14. A computer program for analyzing channel geometry

    USGS Publications Warehouse

    Regan, R.S.; Schaffranek, R.W.

    1985-01-01

    The Channel Geometry Analysis Program (CGAP) provides the capability to process, analyze, and format cross-sectional data for input to flow/transport simulation models or other computational programs. CGAP allows for a variety of cross-sectional data input formats through use of variable format specification. The program accepts data from various computer media and provides for modification of machine-stored parameter values. CGAP has been devised to provide a rapid and efficient means of computing and analyzing the physical properties of an open-channel reach defined by a sequence of cross sections. CGAP 's 16 options provide a wide range of methods by which to analyze and depict a channel reach and its individual cross-sectional properties. The primary function of the program is to compute the area, width, wetted perimeter, and hydraulic radius of cross sections at successive increments of water surface elevation (stage) from data that consist of coordinate pairs of cross-channel distances and land surface or channel bottom elevations. Longitudinal rates-of-change of cross-sectional properties are also computed, as are the mean properties of a channel reach. Output products include tabular lists of cross-sectional area, channel width, wetted perimeter, hydraulic radius, average depth, and cross-sectional symmetry computed as functions of stage; plots of cross sections; plots of cross-sectional area and (or) channel width as functions of stage; tabular lists of cross-sectional area and channel width computed as functions of stage for subdivisions of a cross section; plots of cross sections in isometric projection; and plots of cross-sectional area at a fixed stage as a function of longitudinal distance along an open-channel reach. A Command Procedure Language program and Job Control Language procedure exist to facilitate program execution on the U.S. Geological Survey Prime and Amdahl computer systems respectively. (Lantz-PTT)

  15. Ion channels in microbes

    PubMed Central

    Martinac, Boris; Saimi, Yoshiro; Kung, Ching

    2008-01-01

    Summary Studies of ion channels have for long been dominated by the animalcentric, if not anthropocentric view of physiology. The structures and activities of ion channels had, however, evolved long before the appearance of complex multicellular organisms on Earth. The diversity of ion channels existing in cellular membranes of prokaryotes is a good example. Though at first it may appear as a paradox that most of what we know about the structure of eukaryotic ion channels is based on the structure of bacterial channels, this should not be surprising given the evolutionary relatedness of all living organisms and suitability of microbial cells for structural studies of biological macromolecules in a laboratory environment. Genome sequences of the human as well as various microbial, plant and animal organisms unambiguously established the evolutionary links, whereas crystallographic studies of the structures of major types of ion channels published over the last decade clearly demonstrated the advantage of using microbes as experimental organisms. The purpose of this review is not only to provide an account of acquired knowledge on microbial ion channels but also to show that the study of microbes and their ion channels may also hold a key to solving unresolved molecular mysteries in the future. PMID:18923187

  16. RFI channels, 2

    NASA Technical Reports Server (NTRS)

    Mceliece, R. J.

    1981-01-01

    The cutoff parameters for a class of channel models exhibiting burst noise behavior were calculated and the performance of interleaved coding strategies was evaluated. It is concluded that, provided the channel memory is large enough and is properly exploited, interleaved coding is nearly optimal.

  17. Venus - Sinuous Channel

    NASA Technical Reports Server (NTRS)

    1991-01-01

    This full resolution radar mosaic from Magellan at 49 degrees south latitude, 273 degrees east longitude of an area with dimensions of 130 by 190 kilometers (81 by 118 miles), shows a 200 kilometer (124 mile) segment of a sinuous channel on Venus. The channel is approximately 2 kilometers (1.2 miles) wide. These channel-like features are common on the plains of Venus. In some places they appear to have been formed by lava which may have melted or thermally eroded a path over the plains' surface. Most are 1 to 3 kilometers (0.6 to 2 miles) wide. They resemble terrestrial rivers in some respects, with meanders, cutoff oxbows, and abandoned channel segments. However, Venus channels are not as tightly sinuous as terrestrial rivers. Most are partly buried by younger lava plains, making their sources difficult to identify. A few have vast radar-dark plains units associated with them, suggesting large flow volumes. These channels appear to be older than other channel types on Venus, as they are crossed by fractures and wrinkle ridges, and are often buried by other volcanic materials. In addition, they appear to run both upslope and downslope, suggesting that the plains were warped by regional tectonism after channel formation. Resolution of the Magellan data is about 120 meters (400 feet).

  18. Athermalized channeled spectropolarimeter enhancement.

    SciTech Connect

    Jones, Julia Craven; Way, Brandyn Michael; Mercier, Jeffrey Alan; Hunt, Jeffery P.

    2013-09-01

    Channeled spectropolarimetry can measure the complete polarization state of light as a function of wavelength. Typically, a channeled spectropolarimeter uses high order retarders made of uniaxial crystal to amplitude modulate the measured spectrum with the spectrally-dependent Stokes polarization information. A primary limitation of conventional channeled spectropolarimeters is related to the thermal variability of the retarders. Thermal variation often forces frequent system recalibration, particularly for field deployed systems. However, implementing thermally stable retarders, made of biaxial crystal, results in an athermal channeled spectropolarimeter that relieves the need for frequent recalibration. This report presents experimental results for an anthermalized channeled spectropolarimeter prototype produced using potassium titanyl phosphate. The results of this prototype are compared to the current thermal stabilization state of the art. Finally, the application of the technique to the thermal infrared is studied, and the athermalization concept is applied to an infrared imaging spectropolarimeter design.

  19. Generalized channeled polarimetry.

    PubMed

    Alenin, Andrey S; Tyo, J Scott

    2014-05-01

    Channeled polarimeters measure polarization by modulating the measured intensity in order to create polarization-dependent channels that can be demodulated to reveal the desired polarization information. A number of channeled systems have been described in the past, but their proposed designs often unintentionally sacrifice optimality for ease of algebraic reconstruction. To obtain more optimal systems, a generalized treatment of channeled polarimeters is required. This paper describes methods that enable handling of multi-domain modulations and reconstruction of polarization information using linear algebra. We make practical choices regarding use of either Fourier or direct channels to make these methods more immediately useful. Employing the introduced concepts to optimize existing systems often results in superficial system changes, like changing the order, orientation, thickness, or spacing of polarization elements. For the two examples we consider, we were able to reduce noise in the reconstruction to 34.1% and 57.9% of the original design values. PMID:24979633

  20. Jamming in Vertical Channels

    NASA Astrophysics Data System (ADS)

    Baxter, G. William; McCausland, Jeffrey; Steel, Fiona

    2010-03-01

    We experimentally study jamming of cylindrical grains in a vertical channel. The grains have a low aspect-ratio (height/diameter < 1) so their shape is like antacid tablets or poker chips. They are allowed to fall through a vertical channel with a square cross section. The channel width is greater than the diameter of a grain and constant throughout the length of the channel with no obstructions or constrictions. It is observed that grains sometimes jam in this apparatus. In a jam, grains form a stable structure from one side of the channel to the other with nothing beneath them. Jams may be strong enough to support additional grains above. The probability of a jam occurring is a function of the grain height and diameter. We will present experimental measurements of the jamming probability in this system and discuss the relationship of these results to other experiments and theories.

  1. Single-Molecule Ion Channel Conformational Dynamics in Living Cells

    NASA Astrophysics Data System (ADS)

    Lu, H. Peter

    2014-03-01

    Stochastic and inhomogeneous conformational changes regulate the function and dynamics of ion channels that are crucial for cell functions, neuronal signaling, and brain functions. Such complexity makes it difficult, if not impossible, to characterize ion channel dynamics using conventional electrical recording alone since that the measurement does not specifically interrogate the associated conformational changes but rather the consequences of the conformational changes. Recently, new technology developments on single-molecule spectroscopy, and especially, the combined approaches of using single ion channel patch-clamp electrical recording and single-molecule fluorescence imaging have provided us the capability of probing ion channel conformational changes simultaneously with the electrical single channel recording. By combining real-time single-molecule fluorescence imaging measurements with real-time single-channel electric current measurements in artificial lipid bilayers and in living cell membranes, we were able to probe single ion-channel-protein conformational changes simultaneously, and thus providing an understanding the dynamics and mechanism of ion-channel proteins at the molecular level. The function-regulating and site-specific conformational changes of ion channels are now measurable under physiological conditions in real-time, one molecule at a time. We will focus our discussion on the new development and results of real-time imaging of the dynamics of gramicidin, colicin, and NMDA receptor ion channels in lipid bilayers and living cells. Our results shed light on new perspectives of the intrinsic interplay of lipid membrane dynamics, solvation dynamics, and the ion channel functions.

  2. Cross-channel variability in benthic habitat

    USGS Publications Warehouse

    Vayssieres, Marc; Peterson, Heather

    2003-01-01

    The Interagency Ecological Program’s Environmental Monitoring Program (EMP) has monitored benthic invertebrates since the mid-1970s. A recent review of the EMP found that the spatial study design of the benthos monitoring element was in need of a thorough reexamination through intense special studies and extensive historic data analyses. This article reports the results of preliminary analyses of historical EMP data focusing on cross-channel variability. Specific questions are: (1) do benthic habitats and community assemblages vary between positions across a river channel? (2) Are benthic samples taken at a single channel position sufficiently representative of benthos assemblages across the channel to characterize long term changes in the benthos community of a particular section of a river?

  3. Fractional channel multichannel analyzer

    DOEpatents

    Brackenbush, L.W.; Anderson, G.A.

    1994-08-23

    A multichannel analyzer incorporating the features of the present invention obtains the effect of fractional channels thus greatly reducing the number of actual channels necessary to record complex line spectra. This is accomplished by using an analog-to-digital converter in the asynchronous mode, i.e., the gate pulse from the pulse height-to-pulse width converter is not synchronized with the signal from a clock oscillator. This saves power and reduces the number of components required on the board to achieve the effect of radically expanding the number of channels without changing the circuit board. 9 figs.

  4. Fractional channel multichannel analyzer

    DOEpatents

    Brackenbush, Larry W.; Anderson, Gordon A.

    1994-01-01

    A multichannel analyzer incorporating the features of the present invention obtains the effect of fractional channels thus greatly reducing the number of actual channels necessary to record complex line spectra. This is accomplished by using an analog-to-digital converter in the asynscronous mode, i.e., the gate pulse from the pulse height-to-pulse width converter is not synchronized with the signal from a clock oscillator. This saves power and reduces the number of components required on the board to achieve the effect of radically expanding the number of channels without changing the circuit board.

  5. Channel coding for satellite mobile channels

    NASA Astrophysics Data System (ADS)

    Wong, K. H. H.; Hanzo, L.; Steele, R.

    1989-09-01

    The deployment of channel coding and interleaving to enhance the bit-error performance of a satellite mobile radio channel is addressed for speech and data transmissions. Different convolutional codes (CC) using Viterbi decoding with soft decision are examined with interblock interleaving. Reed-Solomon (RS) codes with Berlekamp-Massey hard decision decoding or soft decision trellis decoding combined with block interleaving are also investigated. A concatenated arrangement employing RS and CC coding as the outer and inner coders, respectively, is used for transmissions via minimum shift keying over Gaussian and Rayleigh fading channels. For an interblock interleaving period of 2880 bits, a concatenated arrangement of an RS(48,36), over the Galois field GF(256) and punctured PCC(3,1,7) yielding an overall coding rate of 1/2, provides a coding gain of 42dB for a BER of 10 to the -6th, and an uncorrectable error detection probability of 1 - 10 to the -9th.

  6. A PLCγ1-Dependent, Force-Sensitive Signaling Network in the Myogenic Constriction of Cerebral Arteries

    PubMed Central

    Gonzales, Albert L.; Yang, Ying; Sullivan, Michelle N.; Sanders, Lindsey; Dabertrand, Fabrice; Hill-Eubanks, David C.; Nelson, Mark T.; Earley, Scott

    2014-01-01

    Maintaining constant blood flow in the face of fluctuations in blood pressure is a critical autoregulatory feature of cerebral arteries. An increase in pressure within the artery lumen causes the vessel to constrict through depolarization and contraction of the encircling smooth muscle cells. This pressure-sensing mechanism involves activation of two types of transient receptor potential (TRP) channels: TRPC6 and TRPM4. We provide evidence that the activation of the γ1 isoform of phospholipase C (PLCγ1) is critical for pressure sensing in cerebral arteries. Inositol 1,4,5-trisphosphate (IP3), generated by PLCγ1 in response to pressure, sensitized IP3 receptors (IP3Rs) to Ca2+ influx mediated by the mechanosensitive TRPC6 channel, synergistically increasing IP3R-mediated Ca2+ release to activate TRPM4 currents, leading to smooth muscle depolarization and constriction of isolated cerebral arteries. Proximity ligation assays demonstrated colocalization of PLCγ1 and TRPC6 with TRPM4, suggesting the presence of a force-sensitive, local signaling network comprising PLCγ1, TRPC6, TRPM4, and IP3Rs. Src tyrosine kinase activity was necessary for stretch-induced TRPM4 activation and myogenic constriction, consistent with the ability of Src to activate PLCγ isoforms. We conclude that contraction of cerebral artery smooth muscle cells requires the integration of pressure-sensing signaling pathways and their convergence on IP3Rs, which mediate localized Ca2+-dependent depolarization through the activation of TRPM4. PMID:24866019

  7. Photochemical control of endogenous ion channels and cellular excitability.

    PubMed

    Fortin, Doris L; Banghart, Matthew R; Dunn, Timothy W; Borges, Katharine; Wagenaar, Daniel A; Gaudry, Quentin; Karakossian, Movses H; Otis, Thomas S; Kristan, William B; Trauner, Dirk; Kramer, Richard H

    2008-04-01

    Light-activated ion channels provide a precise and noninvasive optical means for controlling action potential firing, but the genes encoding these channels must first be delivered and expressed in target cells. Here we describe a method for bestowing light sensitivity onto endogenous ion channels that does not rely on exogenous gene expression. The method uses a synthetic photoisomerizable small molecule, or photoswitchable affinity label (PAL), that specifically targets K+ channels. PALs contain a reactive electrophile, enabling covalent attachment of the photoswitch to naturally occurring nucleophiles in K+ channels. Ion flow through PAL-modified channels is turned on or off by photoisomerizing PAL with different wavelengths of light. We showed that PAL treatment confers light sensitivity onto endogenous K+ channels in isolated rat neurons and in intact neural structures from rat and leech, allowing rapid optical regulation of excitability without genetic modification. PMID:18311146

  8. Neuronal and Cardiovascular Potassium Channels as Therapeutic Drug Targets

    PubMed Central

    Humphries, Edward S. A.

    2015-01-01

    Potassium (K+) channels, with their diversity, often tissue-defined distribution, and critical role in controlling cellular excitability, have long held promise of being important drug targets for the treatment of dysrhythmias in the heart and abnormal neuronal activity within the brain. With the exception of drugs that target one particular class, ATP-sensitive K+ (KATP) channels, very few selective K+ channel activators or inhibitors are currently licensed for clinical use in cardiovascular and neurological disease. Here we review what a range of human genetic disorders have told us about the role of specific K+ channel subunits, explore the potential of activators and inhibitors of specific channel populations as a therapeutic strategy, and discuss possible reasons for the difficulty in designing clinically relevant K+ channel modulators. PMID:26303307

  9. P-Channel Lateral Double-Diffused Metal-Oxide-Semiconductor Field-Effect Transistor with Split N-Type Buried Layer for High Breakdown Voltage and Low Specific On-Resistance

    NASA Astrophysics Data System (ADS)

    Liaw, Chorng-Wei; Chang, Ching-Hung; Lin, Ming-Jang; King, Ya-Ching; Hsu, Charles Ching-Hsiang; Lin, Chrong Jung

    2007-07-01

    Many high voltage complementary metal-oxide-semiconductor (HV-CMOS) processes are modified from a standard 5 V CMOS process by adding an N-type heavily doped layer under the P-well of a HV-PMOS drain terminal to isolate a high voltage P-well from a grounded P-substrate. The limitation of breakdown voltage is dominated by P-well concentration and junction depth. For designing a certain breakdown voltage (\\mathit{BV}dss) for a HV-PMOS, the original 5 V CMOS P-well concentration should be decreased, which could degrade 5 V CMOS characteristics, such as NMOS punch through and latch-up immunity. In this study, we demonstrate a novel HV-PMOS based on a split N-type buried layer (NBL), which provides a high \\mathit{BV}dss in a HV-CMOS process. The newly proposed device with NBL split under the P-well of a drain electrode increases \\mathit{BV}dss without degrading specific on-resistance (Ron,sp) and any added process complexity. From this result, P-well concentration could be increased to improve both 5 V NMOS characteristics and HV-PMOS Ron,sp.

  10. TRPC4- and TRPC4-containing channels.

    PubMed

    Freichel, Marc; Tsvilovskyy, Volodymyr; Camacho-Londoño, Juan E

    2014-01-01

    TRPC4 proteins comprise six transmembrane domains, a putative pore-forming region, and an intracellularly located amino- and carboxy-terminus. Among eleven splice variants identified so far, TRPC4α and TRPC4β are the most abundantly expressed and functionally characterized. TRPC4 is expressed in various organs and cell types including the soma and dendrites of numerous types of neurons; the cardiovascular system including endothelial, smooth muscle, and cardiac cells; myometrial and skeletal muscle cells; kidney; and immune cells such as mast cells. Both recombinant and native TRPC4-containing channels differ tremendously in their permeability and other biophysical properties, pharmacological modulation, and mode of activation depending on the cellular environment. They vary from inwardly rectifying store-operated channels with a high Ca(2+) selectivity to non-store-operated channels predominantly carrying Na(+) and activated by Gαq- and/or Gαi-coupled receptors with a complex U-shaped current-voltage relationship. Thus, individual TRPC4-containing channels contribute to agonist-induced Ca(2+) entry directly or indirectly via depolarization and activation of voltage-gated Ca(2+) channels. The differences in channel properties may arise from variations in the composition of the channel complexes, in the specific regulatory pathways in the corresponding cell system, and/or in the expression pattern of interaction partners which comprise other TRPC proteins to form heteromultimeric channels. Additional interaction partners of TRPC4 that can mediate the activity of TRPC4-containing channels include (1) scaffolding proteins (e.g., NHERF) that may mediate interactions with signaling molecules in or in close vicinity to the plasma membrane such as Gα proteins or phospholipase C and with the cytoskeleton, (2) proteins in specific membrane microdomains (e.g., caveolin-1), or (3) proteins on cellular organelles (e.g., Stim1). The diversity of TRPC4-containing channels

  11. 28-Channel rotary transformer

    NASA Technical Reports Server (NTRS)

    Mclyman, W. T.

    1981-01-01

    Transformer transmits power and digital data across rotating interface. Array has many parallel data channels, each with potential l megabaud data rate. Ferrite-cored transformers are spaced along rotor; airgap between them reduces crosstalk.

  12. A Simple Water Channel

    ERIC Educational Resources Information Center

    White, A. S.

    1976-01-01

    Describes a simple water channel, for use with an overhead projector. It is run from a water tap and may be used for flow visualization experiments, including the effect of streamlining and elementary building aerodynamics. (MLH)

  13. Chondrocyte channel transcriptomics

    PubMed Central

    Lewis, Rebecca; May, Hannah; Mobasheri, Ali; Barrett-Jolley, Richard

    2013-01-01

    To date, a range of ion channels have been identified in chondrocytes using a number of different techniques, predominantly electrophysiological and/or biomolecular; each of these has its advantages and disadvantages. Here we aim to compare and contrast the data available from biophysical and microarray experiments. This letter analyses recent transcriptomics datasets from chondrocytes, accessible from the European Bioinformatics Institute (EBI). We discuss whether such bioinformatic analysis of microarray datasets can potentially accelerate identification and discovery of ion channels in chondrocytes. The ion channels which appear most frequently across these microarray datasets are discussed, along with their possible functions. We discuss whether functional or protein data exist which support the microarray data. A microarray experiment comparing gene expression in osteoarthritis and healthy cartilage is also discussed and we verify the differential expression of 2 of these genes, namely the genes encoding large calcium-activated potassium (BK) and aquaporin channels. PMID:23995703

  14. Proton channel models

    PubMed Central

    Pupo, Amaury; Baez-Nieto, David; Martínez, Agustín; Latorre, Ramón; González, Carlos

    2014-01-01

    Voltage-gated proton channels are integral membrane proteins with the capacity to permeate elementary particles in a voltage and pH dependent manner. These proteins have been found in several species and are involved in various physiological processes. Although their primary topology is known, lack of details regarding their structures in the open conformation has limited analyses toward a deeper understanding of the molecular determinants of their function and regulation. Consequently, the function-structure relationships have been inferred based on homology models. In the present work, we review the existing proton channel models, their assumptions, predictions and the experimental facts that support them. Modeling proton channels is not a trivial task due to the lack of a close homolog template. Hence, there are important differences between published models. This work attempts to critically review existing proton channel models toward the aim of contributing to a better understanding of the structural features of these proteins. PMID:24755912

  15. Channel in Kasei

    NASA Technical Reports Server (NTRS)

    2004-01-01

    14 November 2004 The Kasei Valles are a suite of very large, ancient outflow channels. This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows a portion of the youngest channel system in the Kasei Valles. Torrents of mud, rocks, and water carved this channel as flow was constricted through a narrow portion of the valley. Layers exposed by the erosion that created the channel can be seen in its walls. This 1.4 meters (5 feet) per pixel image is located near 21.1oN, 72.6oW. The picture covers an area approximately 3 km (1.9 mi) across. Sunlight illuminates the scene from the lower left.

  16. Ion Channels in Epithelial Cells

    NASA Astrophysics Data System (ADS)

    Palmer, Lawrence G.

    Ion channels in epithelial cells serve to move ions, and in some cases fluid, between compartments of the body. This function of the transfer of material is fundamentally different from that of the transfer of information, which is the main job of most channels in excitable cells. Nevertheless the basic construction of the channels is similar in many respects in the two tissue types. This chapter reviews the nature of channels in epithelia and discusses how their functions have evolved to accomplish the basic tasks for which they are responsible. I will focus on three channel types: epithelial Na+ channels, inward-rectifier K+ channels, and CFTR Cl- channels.

  17. Modulation of voltage-gated K+ channels by the sodium channel β1 subunit

    PubMed Central

    Nguyen, Hai M.; Miyazaki, Haruko; Hoshi, Naoto; Smith, Brian J.; Nukina, Nobuyuki; Goldin, Alan L.; Chandy, K. George

    2012-01-01

    Voltage-gated sodium (NaV) and potassium (KV) channels are critical components of neuronal action potential generation and propagation. Here, we report that NaVβ1 encoded by SCN1b, an integral subunit of NaV channels, coassembles with and modulates the biophysical properties of KV1 and KV7 channels, but not KV3 channels, in an isoform-specific manner. Distinct domains of NaVβ1 are involved in modulation of the different KV channels. Studies with channel chimeras demonstrate that NaVβ1-mediated changes in activation kinetics and voltage dependence of activation require interaction of NaVβ1 with the channel’s voltage-sensing domain, whereas changes in inactivation and deactivation require interaction with the channel’s pore domain. A molecular model based on docking studies shows NaVβ1 lying in the crevice between the voltage-sensing and pore domains of KV channels, making significant contacts with the S1 and S5 segments. Cross-modulation of NaV and KV channels by NaVβ1 may promote diversity and flexibility in the overall control of cellular excitability and signaling. PMID:23090990

  18. Chloride channels in stroke

    PubMed Central

    Zhang, Ya-ping; Zhang, Hao; Duan, Dayue Darrel

    2013-01-01

    Vascular remodeling of cerebral arterioles, including proliferation, migration, and apoptosis of vascular smooth muscle cells (VSMCs), is the major cause of changes in the cross-sectional area and diameter of the arteries and sudden interruption of blood flow or hemorrhage in the brain, ie, stroke. Accumulating evidence strongly supports an important role for chloride (Cl−) channels in vascular remodeling and stroke. At least three Cl− channel genes are expressed in VSMCs: 1) the TMEM16A (or Ano1), which may encode the calcium-activated Cl− channels (CACCs); 2) the CLC-3 Cl− channel and Cl−/H+ antiporter, which is closely related to the volume-regulated Cl− channels (VRCCs); and 3) the cystic fibrosis transmembrane conductance regulator (CFTR), which encodes the PKA- and PKC-activated Cl− channels. Activation of the CACCs by agonist-induced increase in intracellular Ca2+ causes membrane depolarization, vasoconstriction, and inhibition of VSMC proliferation. Activation of VRCCs by cell volume increase or membrane stretch promotes the production of reactive oxygen species, induces proliferation and inhibits apoptosis of VSMCs. Activation of CFTR inhibits oxidative stress and may prevent the development of hypertension. In addition, Cl− current mediated by gamma-aminobutyric acid (GABA) receptor has also been implicated a role in ischemic neuron death. This review focuses on the functional roles of Cl− channels in the development of stroke and provides a perspective on the future directions for research and the potential to develop Cl− channels as new targets for the prevention and treatment of stroke. PMID:23103617

  19. Bright ion channels and lipid bilayers.

    PubMed

    Szymański, Wiktor; Yilmaz, Duygu; Koçer, Armağan; Feringa, Ben L

    2013-12-17

    If we look at a simple organism such as a zebrafish under a microscope, we would see many cells working in harmony. If we zoomed in, we would observe each unit performing its own tasks in a special aqueous environment isolated from the other units by a lipid bilayer approximately 5 nm thick. These confined units are social: they communicate with one another by sensing and responding to the chemical changes in their environment through receptors and ion channels. These channels control the highly specific and selective passage of ions from one side of the cell to the other and are embedded in lipid bilayers. The movement of ions through ion channels supports excitation and electrical signaling in the nervous system. Ion channels have fascinated scientists not only because of their specificity and selectivity, but also for their functions, the serious consequences when they malfunction, and the other potential applications of these molecules. Light is a useful trigger to control and manipulate ion channels externally. With the many state-of-the-art optical technologies available, light offers a high degree of spatial and temporal control, millisecond precision, and noninvasive intervention and does not change the chemical environment of the system of interest. In this Account, we discuss research toward the dynamic control of lipid bilayer assembly and channel function, particularly the transport across the lipid bilayer-ion channel barrier of cells using light. We first summarize the manipulation of ion channel activity with light to modulate the channel's natural activity. Based on the type of photoswitch employed, we can achieve novel functionalities with these channels, and control neural activity. Then we discuss the recent developments in light-induced transport through lipid bilayers. We focus on three different approaches: the incorporation of photoswitchable copolymers into the lipids, the doping of the lipid bilayer with photosensitive amphiphiles and the

  20. Fracture channel waves

    SciTech Connect

    Nihei, K.T.; Yi, W.; Myer, L.R.; Cook, N.G.; Schoenberg, M.

    1999-03-01

    The properties of guided waves which propagate between two parallel fractures are examined. Plane wave analysis is used to obtain a dispersion equation for the velocities of fracture channel waves. Analysis of this equation demonstrates that parallel fractures form an elastic waveguide that supports two symmetric and two antisymmetric dispersive Rayleigh channel waves, each with particle motions and velocities that are sensitive to the normal and tangential stiffnesses of the fractures. These fracture channel waves degenerate to shear waves when the fracture stiffnesses are large, to Rayleigh waves and Rayleigh-Lamb plate waves when the fracture stiffnesses are low, and to fracture interface waves when the fractures are either very closely spaced or widely separated. For intermediate fracture stiffnesses typical of fractured rock masses, fracture channel waves are dispersive and exhibit moderate to strong localization of guided wave energy between the fractures. The existence of these waves is examined using laboratory acoustic measurements on a fractured marble plate. This experiment confirms the distinct particle motion of the fundamental antisymmetric fracture channel wave (A{sub 0} mode) and demonstrates the ease with which a fracture channel wave can be generated and detected. {copyright} 1999 American Geophysical Union

  1. Mechanosensitive channels in microbes.

    PubMed

    Kung, Ching; Martinac, Boris; Sukharev, Sergei

    2010-01-01

    All cells, including microbes, detect and respond to mechanical forces, of which osmotic pressure is most ancient and universal. Channel proteins have evolved such that they can be directly stretched open when the membrane is under turgor pressure. Osmotic downshock, as in rain, opens bacterial mechanosensitive (MS) channels to jettison osmolytes, relieving pressure and preventing cell lysis. The ion flux through individual channel proteins can be observed directly with a patch clamp. MS channels of large and small conductance (MscL and MscS, respectively) have been cloned, crystallized, and subjected to biophysical and genetic analyses in depth. They are now models to scrutinize how membrane forces direct protein conformational changes. Eukaryotic microbes have homologs from animal sensory channels of the TRP superfamily. The MS channel in yeast is also directly sensitive to membrane stretch. This review examines the key concept that proteins embedded in the lipid bilayer can respond to the changes in the mechanical environment the lipid bilayer provides. PMID:20825352

  2. Ion Channels as Drug Targets in Central Nervous System Disorders

    PubMed Central

    Waszkielewicz, A.M; Gunia, A; Szkaradek, N; Słoczyńska, K; Krupińska, S; Marona, H

    2013-01-01

    Ion channel targeted drugs have always been related with either the central nervous system (CNS), the peripheral nervous system, or the cardiovascular system. Within the CNS, basic indications of drugs are: sleep disorders, anxiety, epilepsy, pain, etc. However, traditional channel blockers have multiple adverse events, mainly due to low specificity of mechanism of action. Lately, novel ion channel subtypes have been discovered, which gives premises to drug discovery process led towards specific channel subtypes. An example is Na+ channels, whose subtypes 1.3 and 1.7-1.9 are responsible for pain, and 1.1 and 1.2 – for epilepsy. Moreover, new drug candidates have been recognized. This review is focusing on ion channels subtypes, which play a significant role in current drug discovery and development process. The knowledge on channel subtypes has developed rapidly, giving new nomenclatures of ion channels. For example, Ca2+ channels are not any more divided to T, L, N, P/Q, and R, but they are described as Cav1.1-Cav3.3, with even newer nomenclature α1A-α1I and α1S. Moreover, new channels such as P2X1-P2X7, as well as TRPA1-TRPV1 have been discovered, giving premises for new types of analgesic drugs. PMID:23409712

  3. 33 CFR 117.966 - Galveston Channel.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Texas § 117.966 Galveston Channel. The drawspan for..., (GIWW mile 356.1) at Galveston, Texas, must open on signal; except that, from 6:40 a.m. to 8:10 a.m.,...

  4. 33 CFR 117.966 - Galveston Channel.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Texas § 117.966 Galveston Channel. The drawspan for..., (GIWW mile 356.1) at Galveston, Texas, must open on signal; except that, from 6:40 a.m. to 8:10 a.m.,...

  5. 33 CFR 117.966 - Galveston Channel.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Texas § 117.966 Galveston Channel. The drawspan for..., (GIWW mile 356.1) at Galveston, Texas, must open on signal; except that, from 6:40 a.m. to 8:10 a.m.,...

  6. Morphodynamics of Floodplain Chute Channels

    NASA Astrophysics Data System (ADS)

    David, S. R.; Edmonds, D. A.

    2015-12-01

    Floodplain chute channel formation is a key process that can enable rivers to transition from single-thread to multi-thread planform geometries. Floodplain chute channels are usually incisional channels connecting topographic lows across point bars and in the floodplain. Surprisingly, it is still not clear what conditions promote chute channel formation and what governs their morphodynamic behavior. Towards this end we have initiated an empirical and theoretical study of floodplain chute channels in Indiana, USA. Using elevation models and satellite imagery we mapped 3064 km2 of floodplain in Indiana, and find that 37.3% of mapped floodplains in Indiana have extensive chute channel networks. These chute channel networks consist of two types of channel segments: meander cutoffs of the main channel and chute channels linking the cutoffs together. To understand how these chute channels link meander cutoffs together and eventually create floodplain channel networks we use Delft3D to explore floodplain morphodynamics. Our first modeling experiment starts from a generic floodplain prepopulated with meander cutoffs to test under what conditions chute channels form.We find that chute channel formation is optimized at an intermediate flood discharge. If the flood discharge is too large the meander cutoffs erosively diffuse, whereas if the floodwave is too small the cutoffs fill with sediment. A moderately sized floodwave reworks the sediment surrounding the topographic lows, enhancing the development of floodplain chute channels. Our second modeling experiments explore how floodplain chute channels evolve on the West Fork of the White River, Indiana, USA. We find that the floodplain chute channels are capable of conveying the entire 10 yr floodwave (Q=1330m3/s) leaving the inter-channel areas dry. Moreover, the chute channels can incise into the floodplain while the margins of channels are aggrading, creating levees. Our results suggest that under the right conditions

  7. The Enigmatic Cytoplasmic Regions of KCNH Channels

    PubMed Central

    Morais-Cabral, João H.; Robertson, Gail A.

    2014-01-01

    KCNH channels are expressed across a vast phylogenetic and evolutionary spectrum. In humans they function in a wide range of tissues and serve as biomarkers and targets for diseases such as cancer and cardiac arrhythmias. These channels share a general architecture with other voltage-gated ion channels but are distinguished by the presence of an N-terminal Per-Arnt-Sim (PAS) domain and a C-terminal domain with homology to cyclic nucleotide binding domains (referred to as the CNBh domain). Cytosolic regions outside these domains show little conservation between KCNH families but within a family are strongly conserved across species, likely reflecting variability that confers specificity to individual channel types. PAS and CNBh domains participate in channel gating, but at least twice in evolutionary history the PAS domain has been lost, and in one family it is omitted by alternate transcription to create a distinct channel subunit. In this focused review we present current knowledge of the structure and function of these cytosolic regions, discuss their evolution as modular domains, and provide our perspective on the important questions moving forward. PMID:25158096

  8. Potassium channels – multiplicity and challenges

    PubMed Central

    Jenkinson, Donald H

    2006-01-01

    The development of our knowledge of the function, structure and pharmacology of K+ channels is briefly outlined. This is the most diverse of all the ion channel families with at least 75 coding genes in mammals. Alternative splicing as well as variations in the channel subunits and accessory proteins that co-assemble to form the functional channel add to the multiplicity. Whereas diversity of this order suggests that it may be possible to develop new classes of drug, for example, for immunomodulation and some diseases of the central nervous system, the ubiquity of K+ channels imposes stringent requirements for selectivity. Animal toxins from the snake, bee and scorpion provide useful leads, though only in a few instances (e.g. with apamin) it has been possible to produce non-peptidic analogues of high potency. The scale of the resources needed to identify, and characterize fully, specific K+ channel as targets and then develop modulators with the required selectivity presents a challenge to both academic and applied pharmacologists. PMID:16402122

  9. The enigmatic cytoplasmic regions of KCNH channels.

    PubMed

    Morais-Cabral, João H; Robertson, Gail A

    2015-01-16

    KCNH channels are expressed across a vast phylogenetic and evolutionary spectrum. In humans, they function in a wide range of tissues and serve as biomarkers and targets for diseases such as cancer and cardiac arrhythmias. These channels share a general architecture with other voltage-gated ion channels but are distinguished by the presence of an N-terminal PAS (Per-Arnt-Sim) domain and a C-terminal domain with homology to cyclic nucleotide binding domains (referred to as the CNBh domain). Cytosolic regions outside these domains show little conservation between KCNH families but are strongly conserved across species within a family, likely reflecting variability that confers specificity to individual channel types. PAS and CNBh domains participate in channel gating, but at least twice in evolutionary history, the PAS domain has been lost and it is omitted by alternate transcription to create a distinct channel subunit in one family. In this focused review, we present current knowledge of the structure and function of these cytosolic regions, discuss their evolution as modular domains and provide our perspective on the important questions moving forward. PMID:25158096

  10. Optical Communications Channel Combiner

    NASA Technical Reports Server (NTRS)

    Quirk, Kevin J.; Quirk, Kevin J.; Nguyen, Danh H.; Nguyen, Huy

    2012-01-01

    NASA has identified deep-space optical communications links as an integral part of a unified space communication network in order to provide data rates in excess of 100 Mb/s. The distances and limited power inherent in a deep-space optical downlink necessitate the use of photon-counting detectors and a power-efficient modulation such as pulse position modulation (PPM). For the output of each photodetector, whether from a separate telescope or a portion of the detection area, a communication receiver estimates a log-likelihood ratio for each PPM slot. To realize the full effective aperture of these receivers, their outputs must be combined prior to information decoding. A channel combiner was developed to synchronize the log-likelihood ratio (LLR) sequences of multiple receivers, and then combines these into a single LLR sequence for information decoding. The channel combiner synchronizes the LLR sequences of up to three receivers and then combines these into a single LLR sequence for output. The channel combiner has three channel inputs, each of which takes as input a sequence of four-bit LLRs for each PPM slot in a codeword via a XAUI 10 Gb/s quad optical fiber interface. The cross-correlation between the channels LLR time series are calculated and used to synchronize the sequences prior to combining. The output of the channel combiner is a sequence of four-bit LLRs for each PPM slot in a codeword via a XAUI 10 Gb/s quad optical fiber interface. The unit is controlled through a 1 Gb/s Ethernet UDP/IP interface. A deep-space optical communication link has not yet been demonstrated. This ground-station channel combiner was developed to demonstrate this capability and is unique in its ability to process such a signal.

  11. 33 CFR 117.225 - Yellow Mill Channel.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 1 2013-07-01 2013-07-01 false Yellow Mill Channel. 117.225 Section 117.225 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Connecticut § 117.225 Yellow Mill Channel....

  12. 33 CFR 117.225 - Yellow Mill Channel.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Yellow Mill Channel. 117.225 Section 117.225 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY BRIDGES DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Connecticut § 117.225 Yellow Mill Channel....

  13. 33 CFR 117.598 - Eel Pond Channel.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 33 Navigation and Navigable Waters 1 2012-07-01 2012-07-01 false Eel Pond Channel. 117.598 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Massachusetts § 117.598 Eel Pond Channel. The following requirements apply to the draw of Eel Pond (Water Street) drawbridge at mile 0.0 at...

  14. 33 CFR 117.598 - Eel Pond Channel.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 33 Navigation and Navigable Waters 1 2013-07-01 2013-07-01 false Eel Pond Channel. 117.598 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Massachusetts § 117.598 Eel Pond Channel. The following requirements apply to the draw of Eel Pond (Water Street) drawbridge at mile 0.0 at...

  15. 33 CFR 117.598 - Eel Pond Channel.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 33 Navigation and Navigable Waters 1 2014-07-01 2014-07-01 false Eel Pond Channel. 117.598 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Massachusetts § 117.598 Eel Pond Channel. The following requirements apply to the draw of Eel Pond (Water Street) drawbridge at mile 0.0 at...

  16. 33 CFR 117.598 - Eel Pond Channel.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Eel Pond Channel. 117.598 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Massachusetts § 117.598 Eel Pond Channel. The following requirements apply to the draw of Eel Pond (Water Street) drawbridge at mile 0.0 at...

  17. 47 CFR 90.525 - Administration of interoperability channels.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... Frequencies in the 763-775 and 793-805 MHz Bands § 90.525 Administration of interoperability channels. (a) States are responsible for administration of the Interoperability channels in the 769-775 MHz and 799-805... in the 769-775 MHz and 799-805 MHz frequency bands without a specific authorization from...

  18. 33 CFR 117.225 - Yellow Mill Channel.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Yellow Mill Channel. 117.225... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Connecticut § 117.225 Yellow Mill Channel. The drawspan of the Stratford Avenue Bridge, mile 0.3 at Bridgeport, must open on signal if at least...

  19. 33 CFR 117.822 - Beaufort Channel, NC.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Beaufort Channel, NC. 117.822... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements North Carolina § 117.822 Beaufort Channel, NC. The draw of the US 70 bridge, mile 0.1, at Beaufort, shall open as follows: (a) From 6 a.m. to 10 p.m.,...

  20. 33 CFR 117.822 - Beaufort Channel, NC.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Beaufort Channel, NC. 117.822... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements North Carolina § 117.822 Beaufort Channel, NC. The draw of the US 70 bridge, mile 0.1, at Beaufort, shall open as follows: (a) From 6 a.m. to 10 p.m.,...

  1. 33 CFR 117.225 - Yellow Mill Channel.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 33 Navigation and Navigable Waters 1 2011-07-01 2011-07-01 false Yellow Mill Channel. 117.225... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Connecticut § 117.225 Yellow Mill Channel. The drawspan of the Stratford Avenue Bridge, mile 0.3 at Bridgeport, must open on signal if at least...

  2. 33 CFR 117.598 - Eel Pond Channel.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Eel Pond Channel. 117.598 Section... DRAWBRIDGE OPERATION REGULATIONS Specific Requirements Massachusetts § 117.598 Eel Pond Channel. The following requirements apply to the draw of Eel Pond (Water Street) drawbridge at mile 0.0 at...

  3. MEMS in microfluidic channels.

    SciTech Connect

    Ashby, Carol Iris Hill; Okandan, Murat; Michalske, Terry A.; Sounart, Thomas L.; Matzke, Carolyn M.

    2004-03-01

    Microelectromechanical systems (MEMS) comprise a new class of devices that include various forms of sensors and actuators. Recent studies have shown that microscale cantilever structures are able to detect a wide range of chemicals, biomolecules or even single bacterial cells. In this approach, cantilever deflection replaces optical fluorescence detection thereby eliminating complex chemical tagging steps that are difficult to achieve with chip-based architectures. A key challenge to utilizing this new detection scheme is the incorporation of functionalized MEMS structures within complex microfluidic channel architectures. The ability to accomplish this integration is currently limited by the processing approaches used to seal lids on pre-etched microfluidic channels. This report describes Sandia's first construction of MEMS instrumented microfluidic chips, which were fabricated by combining our leading capabilities in MEMS processing with our low-temperature photolithographic method for fabricating microfluidic channels. We have explored in-situ cantilevers and other similar passive MEMS devices as a new approach to directly sense fluid transport, and have successfully monitored local flow rates and viscosities within microfluidic channels. Actuated MEMS structures have also been incorporated into microfluidic channels, and the electrical requirements for actuation in liquids have been quantified with an elegant theory. Electrostatic actuation in water has been accomplished, and a novel technique for monitoring local electrical conductivities has been invented.

  4. TRP Channels and Analgesia

    PubMed Central

    Premkumar, Louis S.; Abooj, Mruvil

    2013-01-01

    Since cloning and characterizing the first nociceptive ion channel Transient Receptor Potential (TRP) Vanilloid 1 (TRPV1), other TRP channels involved in nociception have been cloned and characterized, which include TRP Vanilloid 2 (TRPV2), TRP Vanilloid 3 (TRPV3), TRP Vanilloid 4 (TRPV4), TRP Ankyrin 1 (TRPA1) and TRP Melastatin 8 (TRPM8), more recently TRP Canonical 1, 5, 6 (TRPC1, 5, 6), TRP Melastatin 2 (TRPM2) and TRP Melastatin 3 (TRPM3). These channels are predominantly expressed in C and Aδ nociceptors and transmit noxious thermal, mechanical and chemical sensitivities. TRP channels are modulated by pro-inflammatory mediators, neuropeptides and cytokines. Significant advances have been made targeting these receptors either by antagonists or agonists to treat painful conditions. In this review, we will discuss TRP channels as targets for next generation analgesics and the side effects that may ensue as a result of blocking/activating these receptors, because they are also involved in physiological functions such as release of vasoactive neuropeptides and regulation of vascular tone, maintenance of the body temperature, gastrointestinal motility, urinary bladder control etc. PMID:22910182

  5. Mitochondrial Ion Channels

    PubMed Central

    O’Rourke, Brian

    2009-01-01

    In work spanning more than a century, mitochondria have been recognized for their multifunctional roles in metabolism, energy transduction, ion transport, inheritance, signaling, and cell death. Foremost among these tasks is the continuous production of ATP through oxidative phosphorylation, which requires a large electrochemical driving force for protons across the mitochondrial inner membrane. This process requires a membrane with relatively low permeability to ions to minimize energy dissipation. However, a wealth of evidence now indicates that both selective and nonselective ion channels are present in the mitochondrial inner membrane, along with several known channels on the outer membrane. Some of these channels are active under physiological conditions, and others may be activated under pathophysiological conditions to act as the major determinants of cell life and death. This review summarizes research on mitochondrial ion channels and efforts to identify their molecular correlates. Except in a few cases, our understanding of the structure of mitochondrial ion channels is limited, indicating the need for focused discovery in this area. PMID:17059356

  6. MLKL forms cation channels

    PubMed Central

    Xia, Bingqing; Fang, Sui; Chen, Xueqin; Hu, Hong; Chen, Peiyuan; Wang, Huayi; Gao, Zhaobing

    2016-01-01

    The mixed lineage kinase domain-like (MLKL) protein is a key factor in tumor necrosis factor-induced necroptosis. Recent studies on necroptosis execution revealed a commitment role of MLKL in membrane disruption. However, our knowledge of how MLKL functions on membrane remains very limited. Here we demonstrate that MLKL forms cation channels that are permeable preferentially to Mg2+ rather than Ca2+ in the presence of Na+ and K+. Moreover, the N-terminal domain containing six helices (H1-H6) is sufficient to form channels. Using the substituted cysteine accessibility method, we further determine that helix H1, H2, H3, H5 and H6 are transmembrane segments, while H4 is located in the cytoplasm. Finally, MLKL-induced membrane depolarization and cell death exhibit a positive correlation to its channel activity. The Mg2+-preferred permeability and five transmembrane segment topology distinguish MLKL from previously identified Mg2+-permeable channels and thus establish MLKL as a novel class of cation channels. PMID:27033670

  7. Chaos in quantum channels

    NASA Astrophysics Data System (ADS)

    Hosur, Pavan; Qi, Xiao-Liang; Roberts, Daniel A.; Yoshida, Beni

    2016-02-01

    We study chaos and scrambling in unitary channels by considering their entanglement properties as states. Using out-of-time-order correlation functions to diagnose chaos, we characterize the ability of a channel to process quantum information. We show that the generic decay of such correlators implies that any input subsystem must have near vanishing mutual information with almost all partitions of the output. Additionally, we propose the negativity of the tripartite information of the channel as a general diagnostic of scrambling. This measures the delocalization of information and is closely related to the decay of out-of-time-order correlators. We back up our results with numerics in two non-integrable models and analytic results in a perfect tensor network model of chaotic time evolution. These results show that the butterfly effect in quantum systems implies the information-theoretic definition of scrambling.

  8. Trp channels and itch.

    PubMed

    Sun, Shuohao; Dong, Xinzhong

    2016-05-01

    Itch is a unique sensation associated with the scratch reflex. Although the scratch reflex plays a protective role in daily life by removing irritants, chronic itch remains a clinical challenge. Despite urgent clinical need, itch has received relatively little research attention and its mechanisms have remained poorly understood until recently. The goal of the present review is to summarize our current understanding of the mechanisms of acute as well as chronic itch and classifications of the primary itch populations in relationship to transient receptor potential (Trp) channels, which play pivotal roles in multiple somatosensations. The convergent involvement of Trp channels in diverse itch signaling pathways suggests that Trp channels may serve as promising targets for chronic itch treatments. PMID:26385480

  9. Dequantization Via Quantum Channels

    NASA Astrophysics Data System (ADS)

    Andersson, Andreas

    2016-08-01

    For a unital completely positive map {Φ} ("quantum channel") governing the time propagation of a quantum system, the Stinespring representation gives an enlarged system evolving unitarily. We argue that the Stinespring representations of each power {Φ^m} of the single map together encode the structure of the original quantum channel and provide an interaction-dependent model for the bath. The same bath model gives a "classical limit" at infinite time {mto∞} in the form of a noncommutative "manifold" determined by the channel. In this way, a simplified analysis of the system can be performed by making the large-m approximation. These constructions are based on a noncommutative generalization of Berezin quantization. The latter is shown to involve very fundamental aspects of quantum-information theory, which are thereby put in a completely new light.

  10. Channel plate for DNA sequencing

    DOEpatents

    Douthart, R.J.; Crowell, S.L.

    1998-01-13

    This invention is a channel plate that facilitates data compaction in DNA sequencing. The channel plate has a length, a width and a thickness, and further has a plurality of channels that are parallel. Each channel has a depth partially through the thickness of the channel plate. Additionally an interface edge permits electrical communication across an interface through a buffer to a deposition membrane surface. 15 figs.

  11. Channel plate for DNA sequencing

    DOEpatents

    Douthart, Richard J.; Crowell, Shannon L.

    1998-01-01

    This invention is a channel plate that facilitates data compaction in DNA sequencing. The channel plate has a length, a width and a thickness, and further has a plurality of channels that are parallel. Each channel has a depth partially through the thickness of the channel plate. Additionally an interface edge permits electrical communication across an interface through a buffer to a deposition membrane surface.

  12. The neutron channeling phenomenon.

    PubMed

    Khanouchi, A; Sabir, A; Boulkheir, M; Ichaoui, R; Ghassoun, J; Jehouani, A

    1997-01-01

    Shields, used for protection against radiation, are often pierced with vacuum channels for passing cables and other instruments for measurements. The neutron transmission through these shields is an unavoidable phenomenon. In this work we study and discuss the effect of channels on neutron transmission through shields. We consider an infinite homogeneous slab, with a fixed thickness (20 lambda, with lambda the mean free path of the neutron in the slab), which contains a vacuum channel. This slab is irradiated with an infinite source of neutrons on the left side and on the other side (right side) many detectors with windows equal to 2 lambda are placed in order to evaluate the neutron transmission probabilities (Khanouchi, A., Aboubekr, A., Ghassoun, J. and Jehouani, A. (1994) Rencontre Nationale des Jeunes Chercheurs en Physique. Casa Blanca Maroc; Khanouchi, A., Sabir, A., Ghassoun, J. and Jehouani, A. (1995) Premier Congré International des Intéractions Rayonnements Matière. Eljadida Maroc). The neutron history within the slab is simulated by the Monte Carlo method (Booth, T. E. and Hendricks, J. S. (1994) Nuclear Technology 5) and using the exponential biasing technique in order to improve the Monte Carlo calculation (Levitt, L. B. (1968) Nuclear Science and Engineering 31, 500-504; Jehouani, A., Ghassoun, J. and Aboubker, A. (1994) In Proceedings of the 6th International Symposium on Radiation Physics, Rabat, Morocco). Then different geometries of the vacuum channel have been studied. For each geometry we have determined the detector response and calculated the neutron transmission probability for different detector positions. This neutron transmission probability presents a peak for the detectors placed in front of the vacuum channel. This study allowed us to clearly identify the neutron channeling phenomenon. One application of our study is to detect vacuum defects in materials. PMID:9463884

  13. Fluoropolymer piezoelectrets with tubular channels: resonance behavior controlled by channel geometry

    NASA Astrophysics Data System (ADS)

    Pisani Altafim, Ruy Alberto; Corrêa Altafim, Ruy Alberto; Qiu, Xunlin; Raabe, Sebastian; Wirges, Werner; Basso, Heitor Cury; Gerhard, Reimund

    2012-06-01

    Ferro- or piezoelectrets are dielectric materials with two elastically very different macroscopic phases and electrically charged interfaces between them. One of the newer piezoelectret variants is a system of two fluoroethylenepropylene (FEP) films that are first laminated around a polytetrafluoroethylene (PTFE) template. Then, by removing the PTFE template, a two-layer FEP structure with open tubular channels is obtained. After electrical charging, the channels form easily deformable macroscopic electric dipoles whose changes under mechanical or electrical stress lead to significant direct or inverse piezoelectricity, respectively. Here, different PTFE templates are employed to generate channel geometries that vary in height or width. It is shown that the control of the channel geometry allows a direct adjustment of the resonance frequencies in the tubular-channel piezoelectrets. By combining several different channel widths in a single ferroelectret, it is possible to obtain multiple resonance peaks that may lead to a rather flat frequency-response region of the transducer material. A phenomenological relation between the resonance frequency and the geometrical parameters of a tubular channel is also presented. This relation may help to design piezoelectrets with a specific frequency response.

  14. Zeolites: Exploring Molecular Channels

    SciTech Connect

    Arslan, Ilke; Derewinski, Mirek

    2015-05-22

    Synthetic zeolites contain microscopic channels, sort of like a sponge. They have many uses, such as helping laundry detergent lather, absorbing liquid in kitty litter, and as catalysts to produce fuel. Of the hundreds of types of zeolites, only about 15 are used for catalysis. PNNL catalysis scientists Ilke Arslan and Mirek Derewinksi are studying these zeolites to understand what make them special. By exploring the mystery of these microscopic channels, their fundamental findings will help design better catalysts for applications such as biofuel production.

  15. Channel on Ascraeus Mons

    NASA Technical Reports Server (NTRS)

    2004-01-01

    6 May 2004 This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows a portion of a deep channel formed on the northern flank of the large volcano, Ascraeus Mons. Layers of volcanic rock are exposed in the channel walls, and the dark dots on the valley floor are boulders derived from erosion of these materials. The picture occurs near 14.5oN, 102.8oW, and is illuminated from the lower left. The picture covers an area about 3 km (1.9 mi) across.

  16. Alluvial channel hydraulics

    NASA Astrophysics Data System (ADS)

    Ackers, Peter

    1988-07-01

    The development and utilisation of water resources for irrigation, hydropower and public supply can be severely affected by sediment. Where there is a mature and well vegetated landscape, sediment problems may be relatively minor; but where slopes are steep and vegetation sparse, the yield of sediment from the catchment gives high concentrations in the rivers. In utilising these resources, for whatever purpose, an understanding of the hydraulics of alluvial channels is vital. The regime of any conveyance channel in alluvium depends on the interrelationships of sediment transport, channel resistance and bank stability. The regime concept was originally based on empirical relations obtained from observations from canal systems in the Indian subcontinent, and for many years was surrounded by a certain degree of mystique and much scepticism from academics. In more recent years the unabashed empiricism of the original method has been replaced by process-based methods, which have also served as broad confirmation of the classic regime formulae, including their extension to natural channels and meandering channels. The empirical approach to the hydraulics of alluvial channels has thus been updated by physically based formulae for sediment transport and resistance, though there remains some uncertainty about the third function to complete the definition of slope and geometry. Latest thoughts in this respect are that the channel seeks a natural optimum state. Physical modelling using scaled down representations of rivers and estuaries has been used for almost a century, but it requires the correct simulation of the relevant processes. The coming of a better understanding of the physics of sediment transport and the complexity of alluvial channel roughness leads to the conclusion that only in very restricted circumstances can scale models simulate closely the full-size condition. However, the quantification of these processes has been instrumental in the development of

  17. Transient receptor potential channel 1 (TRPC1) reduces calcium permeability in heteromeric channel complexes.

    PubMed

    Storch, Ursula; Forst, Anna-Lena; Philipp, Maximilian; Gudermann, Thomas; Mederos y Schnitzler, Michael

    2012-01-27

    Specific biological roles of the classical transient receptor potential channel 1 (TRPC1) are still largely elusive. To investigate the function of TRPC1 proteins in cell physiology, we studied heterologously expressed TRPC1 channels and found that recombinant TRPC1 subunits do not form functional homomeric channels. Instead, by electrophysiological analysis TRPC1 was shown to form functional heteromeric, receptor-operated channel complexes with TRPC3, -4, -5, -6, and -7 indicating that TRPC1 proteins can co-assemble with all members of the TRPC subfamily. In all TRPC1-containing heteromers, TRPC1 subunits significantly decreased calcium permeation. The exchange of select amino acids in the putative pore-forming region of TRPC1 further reduced calcium permeability, suggesting that TRPC1 subunits contribute to the channel pore. In immortalized immature gonadotropin-releasing hormone neurons endogenously expressing TRPC1, -2, -5, and -6, down-regulation of TRPC1 resulted in increased calcium permeability and elevated basal cytosolic calcium concentrations. We did not observe any involvement of TRPC1 in store-operated cation influx. Notably, TRPC1 suppressed the migration of gonadotropin-releasing hormone neurons without affecting cell proliferation. Conversely, in TRPC1 knockdown neurons, specific migratory properties like distance covered, locomotion speed, and directionality were increased. These findings suggest a novel regulatory mechanism relying on the expression of TRPC1 and the subsequent formation of heteromeric TRPC channel complexes with reduced calcium permeability, thereby fine-tuning neuronal migration. PMID:22157757

  18. Transient Receptor Potential Channel 1 (TRPC1) Reduces Calcium Permeability in Heteromeric Channel Complexes

    PubMed Central

    Storch, Ursula; Forst, Anna-Lena; Philipp, Maximilian; Gudermann, Thomas; Mederos y Schnitzler, Michael

    2012-01-01

    Specific biological roles of the classical transient receptor potential channel 1 (TRPC1) are still largely elusive. To investigate the function of TRPC1 proteins in cell physiology, we studied heterologously expressed TRPC1 channels and found that recombinant TRPC1 subunits do not form functional homomeric channels. Instead, by electrophysiological analysis TRPC1 was shown to form functional heteromeric, receptor-operated channel complexes with TRPC3, -4, -5, -6, and -7 indicating that TRPC1 proteins can co-assemble with all members of the TRPC subfamily. In all TRPC1-containing heteromers, TRPC1 subunits significantly decreased calcium permeation. The exchange of select amino acids in the putative pore-forming region of TRPC1 further reduced calcium permeability, suggesting that TRPC1 subunits contribute to the channel pore. In immortalized immature gonadotropin-releasing hormone neurons endogenously expressing TRPC1, -2, -5, and -6, down-regulation of TRPC1 resulted in increased calcium permeability and elevated basal cytosolic calcium concentrations. We did not observe any involvement of TRPC1 in store-operated cation influx. Notably, TRPC1 suppressed the migration of gonadotropin-releasing hormone neurons without affecting cell proliferation. Conversely, in TRPC1 knockdown neurons, specific migratory properties like distance covered, locomotion speed, and directionality were increased. These findings suggest a novel regulatory mechanism relying on the expression of TRPC1 and the subsequent formation of heteromeric TRPC channel complexes with reduced calcium permeability, thereby fine-tuning neuronal migration. PMID:22157757

  19. Conductive Channel for Energy Transmission

    NASA Astrophysics Data System (ADS)

    Apollonov, Victor V.

    2011-11-01

    For many years the attempts to create conductive channels of big length were taken in order to study the upper atmosphere and to settle special tasks, related to energy transmission. There upon the program of creation of "Impulsar" represents a great interest, as this program in a combination with high-voltage high repetition rate electrical source can be useful to solve the above mentioned problems (N. Tesla ideas for the days of high power lasers). The principle of conductive channel production can be shortly described as follows. The "Impulsar"—laser jet engine vehicle—propulsion take place under the influence of powerful high repetition rate pulse-periodic laser radiation. In the experiments the CO2—laser and solid state Nd:YAG laser systems had been used. Active impulse appears thanks to air breakdown (<30 km) or to the breakdown of ablated material on the board (>30 km), placed in the vicinity of the focusing mirror-acceptor of the breakdown waves. With each pulse of powerful laser the device rises up, leaving a bright and dense trace of products with high degree of ionization and metallization by conductive nano-particles due to ablation. Conductive dust plasma properties investigation in our experiments was produced by two very effective approaches: high power laser controlled ablation and by explosion of wire. Experimental and theoretical results of conductive canal modeling will be presented. The estimations show that with already experimentally demonstrated figures of specific thrust impulse the lower layers of the Ionosphere can be reached in several ten seconds that is enough to keep the high level of channel conductivity and stability with the help of high repetition rate high voltage generator. Some possible applications for new technology are highlighted.

  20. Oxidative Modulation of Voltage-Gated Potassium Channels

    PubMed Central

    Sahoo, Nirakar; Hoshi, Toshinori

    2014-01-01

    Abstract Significance: Voltage-gated K+ channels are a large family of K+-selective ion channel protein complexes that open on membrane depolarization. These K+ channels are expressed in diverse tissues and their function is vital for numerous physiological processes, in particular of neurons and muscle cells. Potentially reversible oxidative regulation of voltage-gated K+ channels by reactive species such as reactive oxygen species (ROS) represents a contributing mechanism of normal cellular plasticity and may play important roles in diverse pathologies including neurodegenerative diseases. Recent Advances: Studies using various protocols of oxidative modification, site-directed mutagenesis, and structural and kinetic modeling provide a broader phenomenology and emerging mechanistic insights. Critical Issues: Physicochemical mechanisms of the functional consequences of oxidative modifications of voltage-gated K+ channels are only beginning to be revealed. In vivo documentation of oxidative modifications of specific amino-acid residues of various voltage-gated K+ channel proteins, including the target specificity issue, is largely absent. Future Directions: High-resolution chemical and proteomic analysis of ion channel proteins with respect to oxidative modification combined with ongoing studies on channel structure and function will provide a better understanding of how the function of voltage-gated K+ channels is tuned by ROS and the corresponding reducing enzymes to meet cellular needs. Antioxid. Redox Signal. 21, 933–952. PMID:24040918

  1. Notch Activation of Ca(2+) Signaling in the Development of Hypoxic Pulmonary Vasoconstriction and Pulmonary Hypertension.

    PubMed

    Smith, Kimberly A; Voiriot, Guillaume; Tang, Haiyang; Fraidenburg, Dustin R; Song, Shanshan; Yamamura, Hisao; Yamamura, Aya; Guo, Qiang; Wan, Jun; Pohl, Nicole M; Tauseef, Mohammad; Bodmer, Rolf; Ocorr, Karen; Thistlethwaite, Patricia A; Haddad, Gabriel G; Powell, Frank L; Makino, Ayako; Mehta, Dolly; Yuan, Jason X-J

    2015-09-01

    Hypoxic pulmonary vasoconstriction (HPV) is an important physiological response that optimizes the ventilation/perfusion ratio. Chronic hypoxia causes vascular remodeling, which is central to the pathogenesis of hypoxia-induced pulmonary hypertension (HPH). We have previously shown that Notch3 is up-regulated in HPH and that activation of Notch signaling enhances store-operated Ca(2+) entry (SOCE), an important mechanism that contributes to pulmonary arterial smooth muscle cell (PASMC) proliferation and contraction. Here, we investigate the role of Notch signaling in HPV and hypoxia-induced enhancement of SOCE. We examined SOCE in human PASMCs exposed to hypoxia and pulmonary arterial pressure in mice using the isolated perfused/ventilated lung method. Wild-type and canonical transient receptor potential (TRPC) 6(-/-) mice were exposed to chronic hypoxia to induce HPH. Inhibition of Notch signaling with a γ-secretase inhibitor attenuates hypoxia-enhanced SOCE in PASMCs and hypoxia-induced increase in pulmonary arterial pressure. Our results demonstrate that hypoxia activates Notch signaling and up-regulates TRPC6 channels. Additionally, treatment with a Notch ligand can mimic hypoxic responses. Finally, inhibition of TRPC6, either pharmacologically or genetically, attenuates HPV, hypoxia-enhanced SOCE, and the development of HPH. These results demonstrate that hypoxia-induced activation of Notch signaling mediates HPV and the development of HPH via functional activation and up-regulation of TRPC6 channels. Understanding the molecular mechanisms that regulate cytosolic free Ca(2+) concentration and PASMC proliferation is critical to elucidation of the pathogenesis of HPH. Targeting Notch regulation of TRPC6 will be beneficial in the development of novel therapies for pulmonary hypertension associated with hypoxia. PMID:25569851

  2. Ion channels enable electrical communication within bacterial communities

    PubMed Central

    Prindle, Arthur; Liu, Jintao; Asally, Munehiro; Ly, San; Garcia-Ojalvo, Jordi; Süel, Gürol M.

    2016-01-01

    The study of bacterial ion channels has provided fundamental insights into the structural basis of neuronal signaling. However, the native role of ion channels in bacteria has remained elusive. Here we show that ion channels conduct long-range electrical signals within bacterial biofilm communities through spatially propagating waves of potassium. These waves result from a positive feedback loop, in which a metabolic trigger induces release of intracellular potassium, which in turn depolarizes neighboring cells. Propagating through the biofilm, this wave of depolarization coordinates metabolic states among cells in the interior and periphery of the biofilm. Deletion of the potassium channel abolishes this response. As predicted by a mathematical model, we further show that spatial propagation can be hindered by specific genetic perturbations to potassium channel gating. Together, these results demonstrate a function for ion channels in bacterial biofilms, and provide a prokaryotic paradigm for active, long-range electrical signaling in cellular communities. PMID:26503040

  3. Neurological perspectives on voltage-gated sodium channels

    PubMed Central

    Linley, John E.; Baker, Mark D.; Minett, Michael S.; Cregg, Roman; Werdehausen, Robert; Rugiero, François

    2012-01-01

    The activity of voltage-gated sodium channels has long been linked to disorders of neuronal excitability such as epilepsy and chronic pain. Recent genetic studies have now expanded the role of sodium channels in health and disease, to include autism, migraine, multiple sclerosis, cancer as well as muscle and immune system disorders. Transgenic mouse models have proved useful in understanding the physiological role of individual sodium channels, and there has been significant progress in the development of subtype selective inhibitors of sodium channels. This review will outline the functions and roles of specific sodium channels in electrical signalling and disease, focusing on neurological aspects. We also discuss recent advances in the development of selective sodium channel inhibitors. PMID:22961543

  4. Biophysical properties of the voltage gated proton channel HV1

    PubMed Central

    Musset, Boris; DeCoursey, Thomas

    2012-01-01

    The biophysical properties of the voltage gated proton channel (HV1) are the key elements of its physiological function. The voltage gated proton channel is a unique molecule that in contrast to all other ion channels is exclusively selective for protons. Alone among proton channels, it has voltage and time dependent gating like other “classical” ion channels. HV1 is furthermore a sensor for the pH in the cell and the surrounding media. Its voltage dependence is strictly coupled to the pH gradient across the membrane. This regulation restricts opening of the channel to specific voltages at any given pH gradient, therefore allowing HV1 to perform its physiological task in the tissue it is expressed in. For HV1 there is no known blocker. The most potent channel inhibitor is zinc (Zn2+) which prevents channel opening. An additional characteristic of HV1 is its strong temperature dependence of both gating and conductance. In contrast to single-file water filled pores like the gramicidin channel, HV1 exhibits pronounced deuterium effects and temperature effects on conduction, consistent with a different conduction mechanism than other ion channels. These properties may be explained by the recent identification of an aspartate in the pore of HV1 that is essential to its proton selectivity. PMID:23050239

  5. Channel Islands rare plants

    USGS Publications Warehouse

    McEachern, K.

    1999-01-01

    Database contains information on 65 rare plant taxa on six islands from archive searches and field surveys, including population location, size and extent 1920-1999, population and habitat conditions, census data, phenological information, associated species. USGS-BRD, Channel Islands Field Station, Ventura, CA.

  6. Channels of Propaganda.

    ERIC Educational Resources Information Center

    Sproule, J. Michael

    Defining propaganda as "efforts by special interests to win over the public covertly by infiltrating messages into various channels of public expression ordinarily viewed as politically neutral," this book argues that propaganda has become pervasive in American life. Pointing out that the 1990s society is inundated with propaganda from numerous…

  7. SK channels and calmodulin

    PubMed Central

    Adelman, John P

    2016-01-01

    Calcium ions are Nature's most widely used signaling mechanism, mediating communication between pathways at virtually every physiological level. Ion channels are no exception, as the activities of a wide range of ion channels are intricately shaped by fluctuations in intracellular Ca2+ levels. Mirroring the importance and the breadth of Ca2+ signaling, free Ca2+ levels are tightly controlled, and a myriad of Ca2+ binding proteins transduce Ca2+ signals, each with its own nuance, comprising a constantly changing symphony of metabolic activity. The founding member of Ca2+ binding proteins is calmodulin (CaM), a small, acidic, modular protein endowed with gymnastic-like flexibility and E-F hand motifs that chelate Ca2+ ions. In this review, I will trace the history that led to the realization that CaM serves as the Ca2+-gating cue for SK channels, the experiments that revealed that CaM is an intrinsic subunit of SK channels, and itself a target of regulation. PMID:25942650

  8. SK channels and calmodulin.

    PubMed

    Adelman, John P

    2016-01-01

    Calcium ions are Nature's most widely used signaling mechanism, mediating communication between pathways at virtually every physiological level. Ion channels are no exception, as the activities of a wide range of ion channels are intricately shaped by fluctuations in intracellular Ca(2+) levels. Mirroring the importance and the breadth of Ca(2+) signaling, free Ca(2+) levels are tightly controlled, and a myriad of Ca(2+) binding proteins transduce Ca(2+) signals, each with its own nuance, comprising a constantly changing symphony of metabolic activity. The founding member of Ca(2+) binding proteins is calmodulin (CaM), a small, acidic, modular protein endowed with gymnastic-like flexibility and E-F hand motifs that chelate Ca(2+) ions. In this review, I will trace the history that led to the realization that CaM serves as the Ca(2+)-gating cue for SK channels, the experiments that revealed that CaM is an intrinsic subunit of SK channels, and itself a target of regulation. PMID:25942650

  9. Chemistry in Microfluidic Channels

    ERIC Educational Resources Information Center

    Chia, Matthew C.; Sweeney, Christina M.; Odom, Teri W.

    2011-01-01

    General chemistry introduces principles such as acid-base chemistry, mixing, and precipitation that are usually demonstrated in bulk solutions. In this laboratory experiment, we describe how chemical reactions can be performed in a microfluidic channel to show advanced concepts such as laminar fluid flow and controlled precipitation. Three sets of…

  10. Keeping the Channels Clear.

    ERIC Educational Resources Information Center

    Weisberg, Jacob

    1996-01-01

    Institutional communication channels need to be clear so that administrators have the information necessary to make informed decisions whenever and wherever required. The secret is to treat the arrival of information--the good, the bad, and the neutral--in essentially the same way, and always thank the person who brings the news, regardless of its…

  11. Developments in relativistic channeling

    SciTech Connect

    Carrigan, R.A. Jr.

    1996-10-01

    The possibility of using channeling as a tool for high energy accelerator applications and particle physics has now been extensively investigated. Bent crystals have been used for accelerator extraction and for particle deflection. Applications as accelerating devices have been discussed but have not yet been tried. 61 refs., 1 fig.

  12. Cohering and decohering power of quantum channels

    NASA Astrophysics Data System (ADS)

    Mani, Azam; Karimipour, Vahid

    2015-09-01

    We introduce the concepts of cohering and decohering power of quantum channels. Using the axiomatic definition of the coherence measure, we show that the optimization required for calculations of these measures can be restricted to pure input states and hence greatly simplified. We then use two examples of this measure, one based on the skew information and the other based on the l1 norm; we find the cohering and decohering measures of a number of one-, two-, and n -qubit channels. Contrary to the view at first glance, it is seen that quantum channels can have cohering power. It is also shown that a specific property of a qubit unitary map is that it has equal cohering and decohering power in any basis. Finally, we derive simple relations between cohering and decohering powers of unitary qubit gates and their tensor products, results which have physically interesting implications.

  13. Majorization preservation of Gaussian bosonic channels

    NASA Astrophysics Data System (ADS)

    Jabbour, Michael G.; García-Patrón, Raúl; Cerf, Nicolas J.

    2016-07-01

    It is shown that phase-insensitive Gaussian bosonic channels are majorization-preserving over the set of passive states of the harmonic oscillator. This means that comparable passive states under majorization are transformed into equally comparable passive states by any phase-insensitive Gaussian bosonic channel. Our proof relies on a new preorder relation called Fock-majorization, which coincides with regular majorization for passive states but also induces another order relation in terms of mean boson number, thereby connecting the concepts of energy and disorder of a quantum state. The consequences of majorization preservation are discussed in the context of the broadcast communication capacity of Gaussian bosonic channels. Because most of our results are independent of the specific nature of the system under investigation, they could be generalized to other quantum systems and Hamiltonians, providing a new tool that may prove useful in quantum information theory and especially quantum thermodynamics.

  14. Channeling through Bent Crystals

    SciTech Connect

    Mack, Stephanie; /Ottawa U. /SLAC

    2012-09-07

    Bent crystals have demonstrated potential for use in beam collimation. A process called channeling is when accelerated particle beams are trapped by the nuclear potentials in the atomic planes within a crystal lattice. If the crystal is bent then the particles can follow the bending angle of the crystal. There are several different effects that are observed when particles travel through a bent crystal including dechanneling, volume capture, volume reflection and channeling. With a crystal placed at the edge of a particle beam, part of the fringe of the beam can be deflected away towards a detector or beam dump, thus helping collimate the beam. There is currently FORTRAN code by Igor Yazynin that has been used to model the passage of particles through a bent crystal. Using this code, the effects mentioned were explored for beam energy that would be seen at the Facility for Advanced Accelerator Experimental Tests (FACET) at a range of crystal orientations with respect to the incoming beam. After propagating 5 meters in vacuum space past the crystal the channeled particles were observed to separate from most of the beam with some noise due to dechanneled particles. Progressively smaller bending radii, with corresponding shorter crystal lengths, were compared and it was seen that multiple scattering decreases with the length of the crystal therefore allowing for cleaner detection of the channeled particles. The input beam was then modified and only a portion of the beam sent through the crystal. With the majority of the beam not affected by the crystal, most particles were not deflected and after propagation the channeled particles were seen to be deflected approximately 5mm. After a portion of the beam travels through the crystal, the entire beam was then sent through a quadrupole magnet, which increased the separation of the channeled particles from the remainder of the beam to a distance of around 20mm. A different code, which was developed at SLAC, was used to

  15. Analysis of aquifer mineralization by paleodrainage channels

    USGS Publications Warehouse

    Rubin, H.; Buddemeier, R.W.

    2003-01-01

    Mineralization of groundwater resources is a problem in south-central Kansas, due to the penetration of saline water from Permian bedrock formations into the overlying alluvial aquifer. One of the mechanisms involved in the mineralization involves small bedrock features of high permeability located in places occupied by streams and rivers in past geological eras. These geological features are termed 'paleodrainage channels'. The permeability of the overlying aquifer can be significantly smaller than that of the channel fill material. The comparatively fast migration of saline water through these channels of high permeability is associated with the transfer of minerals into the overlying freshwater aquifer. This study applies a set of boundary layer approaches to quantify the process of mineral transfer from the channels into the aquifer. The methods used in the present study provide quick estimation and evaluation of the dilution of the channel flow, as well as mineral concentration profile changes in the mineralized zone created in the overlying aquifer. More generally, the method can also be useful for the analysis and evaluation of various types of groundwater contamination in heterogeneous aquifers. The application of the method is exemplified by a complete set of calculations characterizing the possible mineralization process at a specific channel in south central Kansas. Sensitivity analyses are performed and provide information about the importance of the various parameters that affect the mineralization process. Some possible scenarios for the aquifer mineralization phenomena are described and evaluated. It is shown that the channel mineralization may create either several stream tubes of the aquifer with high mineral concentration, or many stream tubes mineralized to a lesser extent. Characteristics of these two patterns of aquifer mineralization are quantified and discussed. ?? 2003 Published by Elsevier Science B.V.

  16. Specification Reformulation During Specification Validation

    NASA Technical Reports Server (NTRS)

    Benner, Kevin M.

    1992-01-01

    The goal of the ARIES Simulation Component (ASC) is to uncover behavioral errors by 'running' a specification at the earliest possible points during the specification development process. The problems to be overcome are the obvious ones the specification may be large, incomplete, underconstrained, and/or uncompilable. This paper describes how specification reformulation is used to mitigate these problems. ASC begins by decomposing validation into specific validation questions. Next, the specification is reformulated to abstract out all those features unrelated to the identified validation question thus creating a new specialized specification. ASC relies on a precise statement of the validation question and a careful application of transformations so as to preserve the essential specification semantics in the resulting specialized specification. This technique is a win if the resulting specialized specification is small enough so the user my easily handle any remaining obstacles to execution. This paper will: (1) describe what a validation question is; (2) outline analysis techniques for identifying what concepts are and are not relevant to a validation question; and (3) identify and apply transformations which remove these less relevant concepts while preserving those which are relevant.

  17. Military specifications

    NASA Technical Reports Server (NTRS)

    Reynolds, Philip

    1987-01-01

    The current situation relative to the military specification is that there is not one specific model of turbulence which people are using. Particular disagreement exists on how turbulence levels will vary with qualitative analysis. It does not tie one down to specifics. When it comes to flying quality specifications, many feel that one should stay with the definitions of the Cooper-Harper rating scale but allow the levels to shift depending on the level of turbulence. There is a ride quality specification in the MIL-SPEC having to do with flight control systems design that is related to a turbulence model. This spec (MIL-F8785C) and others are discussed.

  18. Scorpion venom components that affect ion-channels function

    PubMed Central

    Quintero-Hernández, V.; Jiménez-Vargas, J.M.; Gurrola, G.B.; Valdivia, H.H.F.; Possani, L.D.

    2014-01-01

    SUMMARY The number and types of venom components that affect ion-channel function are reviewed. These are the most important venom components responsible for human intoxication, deserving medical attention, often requiring the use of specific anti-venoms. Special emphasis is given to peptides that recognize Na+-, K+- and Ca++-channels of excitable cells. Knowledge generated by direct isolation of peptides from venom and components deduced from cloned genes, whose amino acid sequences are deposited into databanks are now adays in the order of 1.5 thousands, out of an estimate biodiversity closed to 300,000. Here the diversity of components is briefly reviewed with mention to specific references. Structural characteristic are discussed with examples taken from published work. The principal mechanisms of action of the three different types of peptides are also reviewed. Na+-channel specific venom components usually are modifier of the open and closing kinetic mechanisms of the ion-channels, whereas peptides affecting K+-channels are normally pore blocking agents. The Ryanodine Ca++-channel specific peptides are known for causing sub-conducting stages of the channels conductance and some were shown to be able to internalize penetrating inside the muscle cells. PMID:23891887

  19. An intracellular anion channel critical for pigmentation

    PubMed Central

    Bellono, Nicholas W; Escobar, Iliana E; Lefkovith, Ariel J; Marks, Michael S; Oancea, Elena

    2014-01-01

    Intracellular ion channels are essential regulators of organellar and cellular function, yet the molecular identity and physiological role of many of these channels remains elusive. In particular, no ion channel has been characterized in melanosomes, organelles that produce and store the major mammalian pigment melanin. Defects in melanosome function cause albinism, characterized by vision and pigmentation deficits, impaired retinal development, and increased susceptibility to skin and eye cancers. The most common form of albinism is caused by mutations in oculocutaneous albinism II (OCA2), a melanosome-specific transmembrane protein with unknown function. Here we used direct patch-clamp of skin and eye melanosomes to identify a novel chloride-selective anion conductance mediated by OCA2 and required for melanin production. Expression of OCA2 increases organelle pH, suggesting that the chloride channel might regulate melanin synthesis by modulating melanosome pH. Thus, a melanosomal anion channel that requires OCA2 is essential for skin and eye pigmentation. DOI: http://dx.doi.org/10.7554/eLife.04543.001 PMID:25513726

  20. Are Fibre Channel SANs a Commodity?

    NASA Technical Reports Server (NTRS)

    Becker, Jeff; Jacob, Matt; Biegel, Bryan A. (Technical Monitor)

    2000-01-01

    The goal of this work is to evaluate the feasibility of putting together a Fibre Channel Storage Area Network with heterogeneous hardware running both open-source and commercial operating systems. Adherence to the Fibre Channel Specification is supposed to guarantee interoperability in such an environment. We also want to evaluate how difficult it might be to put together a SAN using open-source components. While all the commercial vendors provide Fibre Channel support, this comes at a cost, e.g., not only O/S and drivers, but usually an expensive support contract. The open-source model could lower the cost of building and maintaining a SAN. Of course, for this to be the case, the open-source platforms would have to provide the functionality to construct a SAN. We are assembling a Fibre Channel SAN from heterogeneous hardware (i386, alpha, sparc) running *BSD, Linux, Tru64, NT and Solaris operating systems. We are running several tests to investigate the level of Fibre Channel support provided by each OS. Our current testbed is specified in the table below. Currently, it only contains open-source platforms. We plan to add a PC running OpenBSD, as well as the following commercial systems: Sun Ultra 1/Solaris, DEC AlphaServer 4000/Tru64 Unix, Pentium Pro PC/Windows NT.

  1. An intracellular anion channel critical for pigmentation.

    PubMed

    Bellono, Nicholas W; Escobar, Iliana E; Lefkovith, Ariel J; Marks, Michael S; Oancea, Elena

    2014-01-01

    Intracellular ion channels are essential regulators of organellar and cellular function, yet the molecular identity and physiological role of many of these channels remains elusive. In particular, no ion channel has been characterized in melanosomes, organelles that produce and store the major mammalian pigment melanin. Defects in melanosome function cause albinism, characterized by vision and pigmentation deficits, impaired retinal development, and increased susceptibility to skin and eye cancers. The most common form of albinism is caused by mutations in oculocutaneous albinism II (OCA2), a melanosome-specific transmembrane protein with unknown function. Here we used direct patch-clamp of skin and eye melanosomes to identify a novel chloride-selective anion conductance mediated by OCA2 and required for melanin production. Expression of OCA2 increases organelle pH, suggesting that the chloride channel might regulate melanin synthesis by modulating melanosome pH. Thus, a melanosomal anion channel that requires OCA2 is essential for skin and eye pigmentation. PMID:25513726

  2. Quantum steganography with noisy quantum channels

    SciTech Connect

    Shaw, Bilal A.; Brun, Todd A.

    2011-02-15

    Steganography is the technique of hiding secret information by embedding it in a seemingly ''innocent'' message. We present protocols for hiding quantum information by disguising it as noise in a codeword of a quantum error-correcting code. The sender (Alice) swaps quantum information into the codeword and applies a random choice of unitary operation, drawing on a secret random key she shares with the receiver (Bob). Using the key, Bob can retrieve the information, but an eavesdropper (Eve) with the power to monitor the channel, but without the secret key, cannot distinguish the message from channel noise. We consider two types of protocols: one in which the hidden quantum information is stored locally in the codeword, and another in which it is embedded in the space of error syndromes. We analyze how difficult it is for Eve to detect the presence of secret messages, and estimate rates of steganographic communication and secret key consumption for specific protocols and examples of error channels. We consider both the case where there is no actual noise in the channel (so that all errors in the codeword result from the deliberate actions of Alice), and the case where the channel is noisy and not controlled by Alice and Bob.

  3. CLC chloride channels in Caenorhabditis elegans.

    PubMed

    Schriever, A M; Friedrich, T; Pusch, M; Jentsch, T J

    1999-11-26

    The genome of the nematode Caenorhabditis elegans encodes six putative chloride channels (CeCLC-1 through CeCLC-6) that represent all three known branches of the mammalian CLC gene family. Using promoter fragments to drive the expression of the green fluorescent protein, CeCLC-2, -3, and -4 expression was studied in transgenic C. elegans. CeCLC-4 was specifically expressed in the large H-shaped excretory cell, where it was co-expressed with CeCLC-3, which is also expressed in other cells, including neurons, muscles, and epithelial cells. Also, CeCLC-2 was expressed in several cells of the nervous system, intestinal cells, and vulval muscle cells. Similar to mammalian CLC proteins, only two nematode CLC channels elicited detectable plasma membrane currents in Xenopus oocytes. CeCLC-3 currents were inwardly rectifying and were activated by positive prepulses. Its complex gating behavior can be explained by two gates, at least one of which depends on extracellular anions. In this respect it resembles some mammalian chloride channels with which it also shares a preference of chloride over iodide. C. elegans thus provides new opportunities to understand common mechanisms underlying structure and function in CLC channels and will allow for a genetic dissection of chloride channels in this simple model organism. PMID:10567397

  4. Channeling of aluminum in silicon

    SciTech Connect

    Wilson, R.G.; Hopkins, C.G.

    1985-05-15

    A systematic study of channeling of aluminum in the silicon crystal is reported. Depth distributions measured by secondary ion mass spectrometry are reported for 40-, 75-, and 150-keV aluminum channeled in the <100> and <110> directions of silicon. The profile dependence on alignment angle is shown for 150-keV aluminum in the <110> of silicon. Aluminum has low electronic stopping in silicon and corresponding deep channeled profiles are observed for aligned implants and deep channeling tails are observed on random implants. The maximum channeling range for 150-keV Al in <100> silicon is about 2.8 ..mu..m and is about 6.4 ..mu..m in <110> silicon. Some ions will reach the maximum channeling range even for 2/sup 0/ misalignment. Many of the deep channeling tails and ''supertails'' reported in earlier literature can be explained by the normal channeling of aluminum in silicon.

  5. Fluid channeling system

    NASA Technical Reports Server (NTRS)

    Davis, Donald Y. (Inventor); Hitch, Bradley D. (Inventor)

    1994-01-01

    A fluid channeling system includes a fluid ejector, a heat exchanger, and a fluid pump disposed in series flow communication The ejector includes a primary inlet for receiving a primary fluid, and a secondary inlet for receiving a secondary fluid which is mixed with the primary fluid and discharged therefrom as ejector discharge. Heat is removed from the ejector discharge in the heat exchanger, and the heat exchanger discharge is compressed in the fluid pump and channeled to the ejector secondary inlet as the secondary fluid In an exemplary embodiment, the temperature of the primary fluid is greater than the maximum operating temperature of a fluid motor powering the fluid pump using a portion of the ejector discharge, with the secondary fluid being mixed with the primary fluid so that the ejector discharge temperature is equal to about the maximum operating temperature of the fluid motor.

  6. Geysering in boiling channels

    SciTech Connect

    Aritomi, Masanori; Takemoto, Takatoshi; Chiang, Jing-Hsien

    1995-09-01

    A concept of natural circulation BWRs such as the SBWR has been proposed and seems to be promising in that the primary cooling system can be simplified. The authors have been investigating thermo-hydraulic instabilities which may appear during the start-up in natural circulation BWRs. In our previous works, geysering was investigated in parallel boiling channels for both natural and forced circulations, and its driving mechanism and the effect of system pressure on geysering occurrence were made clear. In this paper, geysering is investigated in a vertical column and a U-shaped vertical column heated in the lower parts. It is clarified from the results that the occurrence mechanism of geysering and the dependence of system pressure on geysering occurrence coincide between parallel boiling channels in circulation systems and vertical columns in non-circulation systems.

  7. The human role in changing river channels

    NASA Astrophysics Data System (ADS)

    Gregory, K. J.

    2006-09-01

    Direct consequences of the human role, where human activity affects river channels through engineering works including channelization, dam construction, diversion and culverting, have been long recognised [Marsh, G.P., 1864. Man and Nature or Physical Geography as Modified by Human Action. Charles Scribner, New York; Thomas Jr., W.L., (ed.) 1956. Man's Role in Changing the Face of the Earth. Chicago, University of Chicago Press, Chicago.]. The less obvious indirect effects of point and reach changes occurring downstream and throughout the basin, however, are much more recently appreciated, dating from key contributions by Strahler [Strahler, A.N., 1956. The nature of induced erosion and aggradation. In W. L. Thomas (Ed.), Man's Role in Changing the Face of the Earth. University of Chicago Press, Chicago, 621-638.], Wolman [Wolman, M.G., 1967. A cycle of sedimentation and erosion in urban river channels. Geografiska Annaler 49A, 385-95.], Schumm [Schumm, S.A., 1969. River metamorphosis. Proceedings American Society of Civil Engineers, Journal Hydraulics Division 95, 255-73.], and Graf [Graf, W.L., 1977. The rate law in fluvial geomorphology. American Journal of Science, 277, 178-191.]. These are complemented by effects of alterations of land use, such as deforestation, intensive agriculture and incidence of fire, with the most extreme effects produced by building activity and urbanisation. Changing river channels are most evident in the channel cross-section where changes of size, shape and composition are now well-established, with up to tenfold increases or decreases illustrated by results from more than 200 world studies. In addition the overall channel planform, the network and the ecology have changed. Specific terms have become associated with changing river channels including enlargement, shrinkage and metamorphosis. Although the scope of adjustment has been established, it has not always been possible to predict what will happen in a particular location

  8. Athermal channeled spectropolarimeter

    SciTech Connect

    Jones, Julia Craven

    2015-12-08

    A temperature insensitive (athermal) channeled spectropolarimeter (CSP) is described. The athermal CSP includes a crystal retarder formed of a biaxial crystal. The crystal retarder has three crystal axes, wherein each axis has its own distinct index of refraction. The axes are oriented in a particular manner, causing an amplitude modulating carrier frequency induced by the crystal retarder to be thermally invariant. Accordingly, a calibration beam technique can be used over a relatively wide range of ambient temperatures, with a common calibration data set.

  9. Radar channel balancing with commutation

    SciTech Connect

    Doerry, Armin Walter

    2014-02-01

    When multiple channels are employed in a pulse-Doppler radar, achieving and maintaining balance between the channels is problematic. In some circumstances the channels may be commutated to achieve adequate balance. Commutation is the switching, trading, toggling, or multiplexing of the channels between signal paths. Commutation allows modulating the imbalance energy away from the balanced energy in Doppler, where it can be mitigated with filtering.

  10. The alpha channeling effect

    SciTech Connect

    Fisch, N. J.

    2015-12-10

    Alpha particles born through fusion reactions in a tokamak reactor tend to slow down on electrons, but that could take up to hundreds of milliseconds. Before that happens, the energy in these alpha particles can destabilize on collisionless timescales toroidal Alfven modes and other waves, in a way deleterious to energy confinement. However, it has been speculated that this energy might be instead be channeled into useful energy, so as to heat fuel ions or to drive current. Such a channeling needs to be catalyzed by waves Waves can produce diffusion in energy of the alpha particles in a way that is strictly coupled to diffusion in space. If these diffusion paths in energy-position space point from high energy in the center to low energy on the periphery, then alpha particles will be cooled while forced to the periphery. The energy from the alpha particles is absorbed by the wave. The amplified wave can then heat ions or drive current. This process or paradigm for extracting alpha particle energy collisionlessly has been called alpha channeling. While the effect is speculative, the upside potential for economical fusion is immense. The paradigm also operates more generally in other contexts of magnetically confined plasma.

  11. Ion channeling revisited.

    SciTech Connect

    Doyle, Barney Lee; Corona, Aldo; Nguyen, Anh

    2014-09-01

    A MS Excel program has been written that calculates accidental, or unintentional, ion channeling in cubic bcc, fcc and diamond lattice crystals or polycrystalline materials. This becomes an important issue when simulating the creation by energetic neutrons of point displacement damage and extended defects using beams of ions. All of the tables and graphs in the three Ion Beam Analysis Handbooks that previously had to be manually looked up and read from were programed into Excel in handy lookup tables, or parameterized, for the case of the graphs, using rather simple exponential functions with different powers of the argument. The program then offers an extremely convenient way to calculate axial and planar half-angles and minimum yield or dechanneling probabilities, effects on half-angles of amorphous overlayers, accidental channeling probabilities for randomly oriented crystals or crystal