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Sample records for calcium channels expressed

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

    Science.gov (United States)

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

    2009-05-01

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

  2. Calcium channel blocker overdose

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    ... 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 ...

  3. Calcium Channel Blockers

    Science.gov (United States)

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

  4. Calcium channel blocker poisoning

    Directory of Open Access Journals (Sweden)

    Miran Brvar

    2005-04-01

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

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

    DEFF Research Database (Denmark)

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

    2012-01-01

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

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

    DEFF Research Database (Denmark)

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

    2001-01-01

    The distribution of voltage-dependent calcium channels in kidney pre- and postglomerular resistance vessels was determined at the molecular and functional levels. Reverse transcription-polymerase chain reaction analysis of microdissected rat preglomerular vessels and cultured smooth muscle cells...... on vascular diameter in the afferent arteriole. We conclude that voltage-dependent L- and T-type calcium channels are expressed and of functional significance in renal cortical preglomerular vessels, in juxtamedullary efferent arterioles, and in outer medullary vasa recta, but not in cortical efferent...

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

    Czech Academy of Sciences Publication Activity Database

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

    2016-01-01

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

  8. Ryanodine receptors, a family of intracellular calcium ion channels, are expressed throughout early vertebrate development

    Directory of Open Access Journals (Sweden)

    Wu Houdini HT

    2011-12-01

    Full Text Available Abstract Background Calcium signals ([Ca2+]i direct many aspects of embryo development but their regulation is not well characterised. Ryanodine receptors (RyRs are a family of intracellular Ca2+ release channels that control the flux of Ca2+ from internal stores into the cytosol. RyRs are primarily known for their role in excitation-contraction coupling in adult striated muscle and ryr gene mutations are implicated in several human diseases. Current evidence suggests that RyRs do not have a major role to play prior to organogenesis but regulate tissue differentiation. Findings The sequences of the five zebrafish ryr genes were confirmed, their evolutionary relationship established and the primary sequences compared to other vertebrates, including humans. RyRs are differentially expressed in slow (ryr1a, fast (ryr3 and both types (ryr1b of developing skeletal muscle. There are two ryr2 genes (ryr2a and ryr2b which are expressed exclusively in developing CNS and cardiac tissue, respectively. In addition, ryr3 and ryr2a mRNA is detectable in the initial stages of development, prior to embryonic axis formation. Conclusions Our work reveals that zebrafish ryr genes are differentially expressed throughout the developing embryo from cleavage onwards. The data suggests that RyR-regulated Ca2+ signals are associated with several aspects of embryonic development, from organogenesis through to the differentiation of the musculoskeletal, cardiovascular and nervous system. These studies will facilitate further work to explore the developmental function of RyRs in each of these tissue types.

  9. Voltage-Gated Calcium Channels

    Science.gov (United States)

    Zamponi, Gerald Werner

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

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

    Science.gov (United States)

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

    2016-08-01

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

  11. A Crash Course in Calcium Channels.

    Science.gov (United States)

    Zamponi, Gerald W

    2017-12-20

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

  12. Glucose Regulates Cyclin D2 Expression in Quiescent and Replicating Pancreatic β-Cells Through Glycolysis and Calcium Channels

    Science.gov (United States)

    Salpeter, Seth J.; Klochendler, Agnes; Weinberg-Corem, Noa; Porat, Shay; Granot, Zvi; Shapiro, A. M. James; Magnuson, Mark A.; Eden, Amir; Grimsby, Joseph; Glaser, Benjamin

    2011-01-01

    Understanding the molecular triggers of pancreatic β-cell proliferation may facilitate the development of regenerative therapies for diabetes. Genetic studies have demonstrated an important role for cyclin D2 in β-cell proliferation and mass homeostasis, but its specific function in β-cell division and mechanism of regulation remain unclear. Here, we report that cyclin D2 is present at high levels in the nucleus of quiescent β-cells in vivo. The major regulator of cyclin D2 expression is glucose, acting via glycolysis and calcium channels in the β-cell to control cyclin D2 mRNA levels. Furthermore, cyclin D2 mRNA is down-regulated during S-G2-M phases of each β-cell division, via a mechanism that is also affected by glucose metabolism. Thus, glucose metabolism maintains high levels of nuclear cyclin D2 in quiescent β-cells and modulates the down-regulation of cyclin D2 in replicating β-cells. These data challenge the standard model for regulation of cyclin D2 during the cell division cycle and suggest cyclin D2 as a molecular link between glucose levels and β-cell replication. PMID:21521747

  13. Trafficking of neuronal calcium channels

    Czech Academy of Sciences Publication Activity Database

    Weiss, Norbert; Zamponi, G. W.

    2017-01-01

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

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

    DEFF Research Database (Denmark)

    Liu, Ying; Krueger, Katharina; Hovsepian, Anahit

    2011-01-01

    patients with chronic kidney disease and 19 age- and sex-matched healthy control subjects. TRPC3 channels were identified by immunoblotting using specific antibodies and TRPC3 protein was further confirmed by mass spectrometry. We observed a significant increase of TRPC3 channel protein expression...

  15. L-type calcium channels play a critical role in maintaining lens transparency by regulating phosphorylation of aquaporin-0 and myosin light chain and expression of connexins.

    Science.gov (United States)

    Maddala, Rupalatha; Nagendran, Tharkika; de Ridder, Gustaaf G; Schey, Kevin L; Rao, Ponugoti Vasantha

    2013-01-01

    Homeostasis of intracellular calcium is crucial for lens cytoarchitecture and transparency, however, the identity of specific channel proteins regulating calcium influx within the lens is not completely understood. Here we examined the expression and distribution profiles of L-type calcium channels (LTCCs) and explored their role in morphological integrity and transparency of the mouse lens, using cDNA microarray, RT-PCR, immunoblot, pharmacological inhibitors and immunofluorescence analyses. The results revealed that Ca (V) 1.2 and 1.3 channels are expressed and distributed in both the epithelium and cortical fiber cells in mouse lens. Inhibition of LTCCs with felodipine or nifedipine induces progressive cortical cataract formation with time, in association with decreased lens weight in ex-vivo mouse lenses. Histological analyses of felodipine treated lenses revealed extensive disorganization and swelling of cortical fiber cells resembling the phenotype reported for altered aquaporin-0 activity without detectable cytotoxic effects. Analysis of both soluble and membrane rich fractions from felodipine treated lenses by SDS-PAGE in conjunction with mass spectrometry and immunoblot analyses revealed decreases in β-B1-crystallin, Hsp-90, spectrin and filensin. Significantly, loss of transparency in the felodipine treated lenses was preceded by an increase in aquaporin-0 serine-235 phosphorylation and levels of connexin-50, together with decreases in myosin light chain phosphorylation and the levels of 14-3-3ε, a phosphoprotein-binding regulatory protein. Felodipine treatment led to a significant increase in gene expression of connexin-50 and 46 in the mouse lens. Additionally, felodipine inhibition of LTCCs in primary cultures of mouse lens epithelial cells resulted in decreased intracellular calcium, and decreased actin stress fibers and myosin light chain phosphorylation, without detectable cytotoxic response. Taken together, these observations reveal a crucial

  16. Calcium channel blockers and Alzheimer's disease★

    Science.gov (United States)

    Tan, Yi; Deng, Yulin; Qing, Hong

    2012-01-01

    Alzheimer's disease is characterized by two pathological hallmarks: amyloid plaques and neurofibrillary tangles. In addition, calcium homeostasis is disrupted in the course of human aging. Recent research shows that dense plaques can cause functional alteration of calcium signals in mice with Alzheimer's disease. Calcium channel blockers are effective therapeutics for treating Alzheimer's disease. This review provides an overview of the current research of calcium channel blockers involved in Alzheimer's disease therapy. PMID:25767489

  17. Calcium homeostasis modulator (CALHM) ion channels.

    Science.gov (United States)

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

    2016-03-01

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

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

    NARCIS (Netherlands)

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

    1993-01-01

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

  19. L-Type Calcium Channels Modulation by Estradiol.

    Science.gov (United States)

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

    2017-09-01

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

  20. Potassium and calcium channel gene expression in small arteries in porcine and rat models of diet-induced obesity (Poster)

    DEFF Research Database (Denmark)

    Jensen, Lars Jørn; Salomonsson, Max; Sørensen, Charlotte Mehlin

    2014-01-01

    Obesity is an increasing problem worldwide leading to cardiovascular morbidity. Only limited information exists on the transcriptional regulation of arterial K+ and Ca2+ channels in obesity. We quantified, by real-time PCR, mRNA expression of K+ channels and L-type Ca2+ channels (LTCC) in small...... mesenteric (MA), middle cerebral (MCA), and left coronary arteries (LCA) of lean vs. obese rats and minipigs. Male Sprague Dawley rats were fed a high-fat (FAT; N=5), high-fructose (FRUC; N=7), high-fat/high-fructose (FAT/FRUC; N=7) or standard diet (STD; N=7-11) for 28 Weeks. FAT and FAT/FRUC became obese...... increased in OB and OB+DIAB. BKca, IKca, SKca and/or LTCC mRNA was up-regulated in LCA from OB and OB+DIAB (n.s.). Expression of BKca mRNA was increased, whereas IKca mRNA decreased in MCA from OB (n.s.). SKca mRNA was decreased in MA from OB (n.s.). Diet-induced obesity in rats and minipigs lead to complex...

  1. Single channel recording of a mitochondrial calcium uniporter.

    Science.gov (United States)

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

    2018-01-29

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

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

    NARCIS (Netherlands)

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

    2017-01-01

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

  3. Anion-sensitive regions of L-type CaV1.2 calcium channels expressed in HEK293 cells.

    Directory of Open Access Journals (Sweden)

    Norbert Babai

    2010-01-01

    Full Text Available L-type calcium currents (I(Ca are influenced by changes in extracellular chloride, but sites of anion effects have not been identified. Our experiments showed that CaV1.2 currents expressed in HEK293 cells are strongly inhibited by replacing extracellular chloride with gluconate or perchlorate. Variance-mean analysis of I(Ca and cell-attached patch single channel recordings indicate that gluconate-induced inhibition is due to intracellular anion effects on Ca(2+ channel open probability, not conductance. Inhibition of CaV1.2 currents produced by replacing chloride with gluconate was reduced from approximately 75%-80% to approximately 50% by omitting beta subunits but unaffected by omitting alpha(2delta subunits. Similarly, gluconate inhibition was reduced to approximately 50% by deleting an alpha1 subunit N-terminal region of 15 residues critical for beta subunit interactions regulating open probability. Omitting beta subunits with this mutant alpha1 subunit did not further diminish inhibition. Gluconate inhibition was unchanged with expression of different beta subunits. Truncating the C terminus at AA1665 reduced gluconate inhibition from approximately 75%-80% to approximately 50% whereas truncating it at AA1700 had no effect. Neutralizing arginines at AA1696 and 1697 by replacement with glutamines reduced gluconate inhibition to approximately 60% indicating these residues are particularly important for anion effects. Expressing CaV1.2 channels that lacked both N and C termini reduced gluconate inhibition to approximately 25% consistent with additive interactions between the two tail regions. Our results suggest that modest changes in intracellular anion concentration can produce significant effects on CaV1.2 currents mediated by changes in channel open probability involving beta subunit interactions with the N terminus and a short C terminal region.

  4. Restoration of Motor Defects Caused by Loss of Drosophila TDP-43 by Expression of the Voltage-Gated Calcium Channel, Cacophony, in Central Neurons.

    Science.gov (United States)

    Lembke, Kayly M; Scudder, Charles; Morton, David B

    2017-09-27

    Defects in the RNA-binding protein, TDP-43, are known to cause a variety of neurodegenerative diseases, including amyotrophic lateral sclerosis and frontotemporal lobar dementia. A variety of experimental systems have shown that neurons are sensitive to TDP-43 expression levels, yet the specific functional defects resulting from TDP-43 dysregulation have not been well described. Using the Drosophila TDP-43 ortholog TBPH, we previously showed that TBPH-null animals display locomotion defects as third instar larvae. Furthermore, loss of TBPH caused a reduction in cacophony , a Type II voltage-gated calcium channel, expression and that genetically restoring cacophony in motor neurons in TBPH mutant animals was sufficient to rescue the locomotion defects. In the present study, we examined the relative contributions of neuromuscular junction physiology and the motor program to the locomotion defects and identified subsets of neurons that require cacophony expression to rescue the defects. At the neuromuscular junction, we showed mEPP amplitudes and frequency require TBPH. Cacophony expression in motor neurons rescued mEPP frequency but not mEPP amplitude. We also showed that TBPH mutants displayed reduced motor neuron bursting and coordination during crawling and restoring cacophony selectively in two pairs of cells located in the brain, the AVM001b/2b neurons, also rescued the locomotion and motor defects, but not the defects in neuromuscular junction physiology. These results suggest that the behavioral defects associated with loss of TBPH throughout the nervous system can be associated with defects in a small number of genes in a limited number of central neurons, rather than peripheral defects. SIGNIFICANCE STATEMENT TDP-43 dysfunction is a common feature in neurodegenerative diseases, including amyotrophic lateral sclerosis, frontotemporal lobar dementia, and Alzheimer's disease. Loss- and gain-of-function models have shown that neurons are sensitive to TDP-43

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

    DEFF Research Database (Denmark)

    Hansen, Pernille B L

    2015-01-01

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

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

    International Nuclear Information System (INIS)

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

    2012-01-01

    Highlights: ► Caveolin-1 expression is regulated by calcium signaling at the transcriptional level. ► An inhibitor of or siRNA to L-type calcium channel suppressed caveolin-1 expression. ► Cyclosporine A or an NFAT inhibitor markedly reduced caveolin-1 expression. ► Caveolin-1 regulation by calcium signaling is observed in several mouse cell lines. -- Abstract: Caveolin-1, an indispensable component of caveolae serving as a transformation suppressor protein, is highly expressed in poorly metastatic mouse osteosarcoma FBJ-S1 cells while highly metastatic FBJ-LL cells express low levels of caveolin-1. Calcium concentration is higher in FBJ-S1 cells than in FBJ-LL cells; therefore, we investigated the possibility that calcium signaling positively regulates caveolin-1 in mouse FBJ-S1 cells. When cells were treated with the calcium channel blocker nifedipine, cyclosporin A (a calcineurin inhibitor), or INCA-6 (a nuclear factor of activated T-cells [NFAT] inhibitor), caveolin-1 expression at the mRNA and protein levels decreased. RNA silencing of voltage-dependent L-type calcium channel subunit alpha-1C resulted in suppression of caveolin-1 expression. This novel caveolin-1 regulation pathway was also identified in mouse NIH 3T3 cells and Lewis lung carcinoma cells. These results indicate that caveolin-1 is positively regulated at the transcriptional level through a novel calcium signaling pathway mediated by L-type calcium channel/Ca 2+ /calcineurin/NFAT.

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

    DEFF Research Database (Denmark)

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

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Nawal Zabouri

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

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

    Science.gov (United States)

    Zabouri, Nawal; Haverkamp, Silke

    2013-01-01

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

  10. Discovery and Development of Calcium Channel Blockers

    Directory of Open Access Journals (Sweden)

    Théophile Godfraind

    2017-05-01

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

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

    Czech Academy of Sciences Publication Activity Database

    Weiss, Norbert

    2015-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Erin K Purcell

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

  13. Use of calcium channel blockers in hypertension.

    Science.gov (United States)

    Conlin, P R; Williams, G H

    1998-01-01

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

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

    Science.gov (United States)

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

    2014-04-03

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

  15. Reconstruction of Cell Surface Densities of Ion Pumps, Exchangers, and Channels from mRNA Expression, Conductance Kinetics, Whole-Cell Calcium, and Current-Clamp Voltage Recordings, with an Application to Human Uterine Smooth Muscle Cells.

    Directory of Open Access Journals (Sweden)

    Jolene Atia

    2016-04-01

    Full Text Available Uterine smooth muscle cells remain quiescent throughout most of gestation, only generating spontaneous action potentials immediately prior to, and during, labor. This study presents a method that combines transcriptomics with biophysical recordings to characterise the conductance repertoire of these cells, the 'conductance repertoire' being the total complement of ion channels and transporters expressed by an electrically active cell. Transcriptomic analysis provides a set of potential electrogenic entities, of which the conductance repertoire is a subset. Each entity within the conductance repertoire was modeled independently and its gating parameter values were fixed using the available biophysical data. The only remaining free parameters were the surface densities for each entity. We characterise the space of combinations of surface densities (density vectors consistent with experimentally observed membrane potential and calcium waveforms. This yields insights on the functional redundancy of the system as well as its behavioral versatility. Our approach couples high-throughput transcriptomic data with physiological behaviors in health and disease, and provides a formal method to link genotype to phenotype in excitable systems. We accurately predict current densities and chart functional redundancy. For example, we find that to evoke the observed voltage waveform, the BK channel is functionally redundant whereas hERG is essential. Furthermore, our analysis suggests that activation of calcium-activated chloride conductances by intracellular calcium release is the key factor underlying spontaneous depolarisations.

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

    International Nuclear Information System (INIS)

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

    2017-01-01

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

  17. Single molecule microscopy on Store-Operated Calcium channels

    International Nuclear Information System (INIS)

    Madl, J.

    2011-01-01

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

  18. 43. Calmodulin regulating calcium sensitivity of Na channels

    Directory of Open Access Journals (Sweden)

    R. Vegiraju

    2016-07-01

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

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

    NARCIS (Netherlands)

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

    1991-01-01

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

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

    DEFF Research Database (Denmark)

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

    2003-01-01

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

  1. Activation of purified calcium channels by stoichiometric protein phosphorylation

    Energy Technology Data Exchange (ETDEWEB)

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

    1989-09-01

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

  2. Activation of purified calcium channels by stoichiometric protein phosphorylation

    International Nuclear Information System (INIS)

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

    1989-01-01

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

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

    NARCIS (Netherlands)

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

    2003-01-01

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

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

    DEFF Research Database (Denmark)

    VanBavel, Ed; Sorop, Oana; Andreasen, Ditte

    2002-01-01

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

  5. The functions of store-operated calcium channels.

    Science.gov (United States)

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

    2017-06-01

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

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

    Science.gov (United States)

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

    1993-01-01

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

  7. TRPV5: an ingeniously controlled calcium channel.

    NARCIS (Netherlands)

    Groot, T. de; Bindels, R.J.M.; Hoenderop, J.G.J.

    2008-01-01

    Body Ca(2+) homeostasis is tightly controlled and slight disturbances in renal Ca(2+) reabsorption can lead to excessive urine Ca(2+) excretion and promote kidney stone formation. The epithelial Ca(2+) channel TRPV5 constitutes the rate-limiting step of active Ca(2+) reabsorption in the kidney.

  8. Nifedipine, a calcium channel blocker, inhibits advanced glycation end product (AGE)-elicited mesangial cell damage by suppressing AGE receptor (RAGE) expression via peroxisome proliferator-activated receptor-gamma activation

    International Nuclear Information System (INIS)

    Matsui, Takanori; Yamagishi, Sho-ichi; Takeuchi, Masayoshi; Ueda, Seiji; Fukami, Kei; Okuda, Seiya

    2009-01-01

    The interaction between advanced glycation end products (AGE) and their receptor RAGE mediates the progressive alteration in renal architecture and loss of renal function in diabetic nephropathy. Oxidative stress generation and inflammation also play a central role in diabetic nephropathy. This study investigated whether and how nifedipine, a calcium channel blocker (CCB), blocked the AGE-elicited mesangial cell damage in vitro. Nifedipine, but not amlodipine, a control CCB, down-regulated RAGE mRNA levels and subsequently reduced reactive oxygen species (ROS) generation in AGE-exposed mesangial cells. AGE increased mRNA levels of vascular cell adhesion molecule-1 (VCAM-1) and induced monocyte chemoattractant protein-1 (MCP-1) production in mesangial cells, both of which were prevented by the treatment with nifedipine, but not amlodipine. The beneficial effects of nifedipine on AGE-exposed mesangial cells were blocked by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-γ (PPAR-γ). Although nifedipine did not affect expression levels of PPAR-γ, it increased the PPAR-γ transcriptional activity in mesangial cells. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-inflammatory agent against AGE by suppressing RAGE expression in cultured mesangial cells via PPAR-γ activation.

  9. Altered expression of the voltage-gated calcium channel subunit alpha(2)delta-1: A comparison between two experimental models of epilepsy and a sensory nerve ligation model of neuropathic pain

    Czech Academy of Sciences Publication Activity Database

    Nieto-Rostro, M.; Sandhu, G.; Bauer, C. S.; Jiruška, Přemysl; Jefferys, J. G. R.; Dolphin, A. C.

    2014-01-01

    Roč. 283, Dec (2014), s. 124-137 ISSN 0306-4522 R&D Projects: GA MZd(CZ) NT14489 Institutional support: RVO:67985823 Keywords : calcium channel * dorsal root ganglion (DRG) * alpha2delta subunit * epilepsy * neuropathic pain * reactive gliosis Subject RIV: FH - Neurology Impact factor: 3.357, year: 2014

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

    Directory of Open Access Journals (Sweden)

    Weiping Zhang

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

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

    International Nuclear Information System (INIS)

    Curtis, B.M.

    1986-01-01

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

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

    DEFF Research Database (Denmark)

    Svenningsen, Per; Hansen, Pernille B L

    2016-01-01

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

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

    Science.gov (United States)

    Cheng, Pan; Tang, Wanyi; He, Hao

    2018-02-01

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

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

    Science.gov (United States)

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

    2013-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Xianjie Wen

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

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

    Science.gov (United States)

    Kaneko, Shuji

    2015-02-01

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

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

    Science.gov (United States)

    Koch, Peter; Herzig, Stefan; Matthes, Jan

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

  18. Crystal structure of the epithelial calcium channel TRPV6.

    Science.gov (United States)

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

    2016-06-23

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

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

    Science.gov (United States)

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

    2017-06-01

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

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

    DEFF Research Database (Denmark)

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

    2001-01-01

    The physiological significance and subcellular distribution of voltage dependent calcium channels was defined using calcium channel blockers to inhibit potassium induced rises in cytosolic calcium concentration in cultured mouse neocortical neurons. The cytosolic calcium concentration was measured...... channels were differentially distributed in somata, neurites and nerve terminals. omega-conotoxin MVIIC (omega-CgTx MVIIC) inhibited approximately 40% of the Ca(2+)-rise in both somata and neurites and 60% of the potassium induced [3H]GABA release, indicating that the Q-type channel is the quantitatively...... most important voltage dependent calcium channel in all parts of the neuron. After treatment with thapsigargin the increase in cytosolic calcium was halved, indicating that calcium release from thapsigargin sensitive intracellular calcium stores is an important component of the potassium induced rise...

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

    Science.gov (United States)

    Benhassine, Narimane; Berger, Thomas

    2009-03-01

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

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

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    Kang Zhang

    2017-05-01

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

  3. Calcium homeostasis and vitamin D metabolism and expression in strongly calcifying laying birds.

    Science.gov (United States)

    Bar, Arie

    2008-12-01

    Egg laying and shell calcification impose severe extra demands on ionic calcium (Ca2+) homeostasis; especially in birds characterized by their long clutches (series of eggs laid sequentially before a "pause day"). These demands induce vitamin D metabolism and expression. The metabolism of vitamin D is also altered indirectly, by other processes associated with increased demands for calcium, such as growth, bone formation and egg production. A series of intestinal, renal or bone proteins are consequently expressed in the target organs via mechanisms involving a vitamin D receptor. Some of these proteins (carbonic anhydrase, calbindin and calcium-ATPase) are also found in the uterus (eggshell gland) or are believed to be involved in calcium transport in the intestine or kidney (calcium channels). The present review deals with vitamin D metabolism and the expression of the above-mentioned proteins in birds, with special attention to the strongly calcifying laying bird.

  4. 6-OHDA induced calcium influx through N-type calcium channel alters membrane properties via PKA pathway in substantia nigra pars compacta dopaminergic neurons.

    Science.gov (United States)

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

    2014-07-11

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

  5. Voltage-gated calcium channels of Paramecium cilia.

    Science.gov (United States)

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

    2016-10-01

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

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

    Science.gov (United States)

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

    2018-06-01

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

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

    Science.gov (United States)

    Naranjo, David; Wen, Hua; Brehm, Paul

    2015-01-01

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

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

    International Nuclear Information System (INIS)

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

    2016-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Lu Ake Tzu-Hui

    2012-12-01

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

  10. Interaction of H2S with Calcium Permeable Channels and Transporters

    Directory of Open Access Journals (Sweden)

    Weihua Zhang

    2015-01-01

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

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

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    Bruce Lyeth

    2013-06-01

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

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

    Directory of Open Access Journals (Sweden)

    Adela Banciu

    2018-05-01

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

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

    Science.gov (United States)

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

    2018-05-09

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

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

    Directory of Open Access Journals (Sweden)

    David Ramírez

    2017-10-01

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

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

    Science.gov (United States)

    Tuluc, Petronel; Flucher, Bernhard E

    2011-12-01

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

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

    Science.gov (United States)

    Myoga, Michael H.; Regehr, Wade G.

    2011-01-01

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

  17. LRRK2 regulates voltage-gated calcium channel function.

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    Cade eBedford

    2016-05-01

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

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

    Science.gov (United States)

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

    2012-07-01

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

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

    Science.gov (United States)

    Sturgill, M G; Seibold, J R

    1998-11-01

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

  20. Do cysteine residues regulate transient receptor potential canonical type 6 (TRPC6) channel protein expression?

    DEFF Research Database (Denmark)

    Thilo, Florian; Liu, Ying; Krueger, Katharina

    2012-01-01

    The regulation of calcium influx through transient receptor potential canonical type 6 channel is mandatory for the activity of human monocytes. We submit the first evidence that cysteine residues of homocysteine or acetylcysteine affect TRPC6 expression in human monocytes. We observed that patie......The regulation of calcium influx through transient receptor potential canonical type 6 channel is mandatory for the activity of human monocytes. We submit the first evidence that cysteine residues of homocysteine or acetylcysteine affect TRPC6 expression in human monocytes. We observed...... that patients with chronic renal failure had significantly elevated homocysteine levels and TRPC6 mRNA expression levels in monocytes compared to control subjects. We further observed that administration of homocysteine or acetylcysteine significantly increased TRPC6 channel protein expression compared...... to control conditions. We therefore hypothesize that cysteine residues increase TRPC6 channel protein expression in humans....

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

    Science.gov (United States)

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

    2018-05-02

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

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

    Science.gov (United States)

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

    1991-06-01

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

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

    Directory of Open Access Journals (Sweden)

    CECILIA HIDALGO

    2000-01-01

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

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

    Science.gov (United States)

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

    2015-01-01

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

  5. Dental enamel cells express functional SOCE channels.

    Science.gov (United States)

    Nurbaeva, Meerim K; Eckstein, Miriam; Concepcion, Axel R; Smith, Charles E; Srikanth, Sonal; Paine, Michael L; Gwack, Yousang; Hubbard, Michael J; Feske, Stefan; Lacruz, Rodrigo S

    2015-10-30

    Dental enamel formation requires large quantities of Ca(2+) yet the mechanisms mediating Ca(2+) dynamics in enamel cells are unclear. Store-operated Ca(2+) entry (SOCE) channels are important Ca(2+) influx mechanisms in many cells. SOCE involves release of Ca(2+) from intracellular pools followed by Ca(2+) entry. The best-characterized SOCE channels are the Ca(2+) release-activated Ca(2+) (CRAC) channels. As patients with mutations in the CRAC channel genes STIM1 and ORAI1 show abnormal enamel mineralization, we hypothesized that CRAC channels might be an important Ca(2+) uptake mechanism in enamel cells. Investigating primary murine enamel cells, we found that key components of CRAC channels (ORAI1, ORAI2, ORAI3, STIM1, STIM2) were expressed and most abundant during the maturation stage of enamel development. Furthermore, inositol 1,4,5-trisphosphate receptor (IP3R) but not ryanodine receptor (RyR) expression was high in enamel cells suggesting that IP3Rs are the main ER Ca(2+) release mechanism. Passive depletion of ER Ca(2+) stores with thapsigargin resulted in a significant raise in [Ca(2+)]i consistent with SOCE. In cells pre-treated with the CRAC channel blocker Synta-66 Ca(2+) entry was significantly inhibited. These data demonstrate that enamel cells have SOCE mediated by CRAC channels and implicate them as a mechanism for Ca(2+) uptake in enamel formation.

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

    International Nuclear Information System (INIS)

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

    1989-01-01

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

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

    DEFF Research Database (Denmark)

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

    2015-01-01

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

  8. Channel properties of Nax expressed in neurons.

    Directory of Open Access Journals (Sweden)

    Masahito Matsumoto

    Full Text Available Nax is a sodium-concentration ([Na+]-sensitive Na channel with a gating threshold of ~150 mM for extracellular [Na+] ([Na+]o in vitro. We previously reported that Nax was preferentially expressed in the glial cells of sensory circumventricular organs including the subfornical organ, and was involved in [Na+] sensing for the control of salt-intake behavior. Although Nax was also suggested to be expressed in the neurons of some brain regions including the amygdala and cerebral cortex, the channel properties of Nax have not yet been adequately characterized in neurons. We herein verified that Nax was expressed in neurons in the lateral amygdala of mice using an antibody that was newly generated against mouse Nax. To investigate the channel properties of Nax expressed in neurons, we established an inducible cell line of Nax using the mouse neuroblastoma cell line, Neuro-2a, which is endogenously devoid of the expression of Nax. Functional analyses of this cell line revealed that the [Na+]-sensitivity of Nax in neuronal cells was similar to that expressed in glial cells. The cation selectivity sequence of the Nax channel in cations was revealed to be Na+ ≈ Li+ > Rb+ > Cs+ for the first time. Furthermore, we demonstrated that Nax bound to postsynaptic density protein 95 (PSD95 through its PSD95/Disc-large/ZO-1 (PDZ-binding motif at the C-terminus in neurons. The interaction between Nax and PSD95 may be involved in promoting the surface expression of Nax channels because the depletion of endogenous PSD95 resulted in a decrease in Nax at the plasma membrane. These results indicated, for the first time, that Nax functions as a [Na+]-sensitive Na channel in neurons as well as in glial cells.

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

    Science.gov (United States)

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

    2014-01-01

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

  10. Calcium electrotransfer for termination of transgene expression in muscle

    DEFF Research Database (Denmark)

    Hojman, Pernille; Spanggaard, Iben; Olsen, Caroline Holkman

    2011-01-01

    Gene electrotransfer is expanding in clinical use, thus we have searched for an emergency procedure to stop transgene expression in case of serious adverse events. Calcium is cytotoxic at high intracellular levels, so we tested effects of calcium electrotransfer on transgene expression in muscle....... A clinical grade calcium solution (20 μl, 168 mM) was injected into transfected mouse or rat tibialis cranialis muscle. Ca(2+) uptake was quantified using calcium 45 ((45)Ca), and voltage and time between injection and pulsation were varied. Extinction of transgene expression was investigated by using both...... voltage pulses of 1000 V/cm. Using these parameters, in vivo imaging showed that transgene expression significantly decreased 4 hr after Ca(2+) electrotransfer and was eliminated within 24 hr. Similarly, serum erythropoietin was reduced by 46% at 4 hr and to control levels at 2 days. Histological analyses...

  11. Calcium electrotransfer for termination of transgene expression in muscle

    DEFF Research Database (Denmark)

    Hojman, Pernille; Spanggaard, Iben; Olsen, Caroline Holkman

    2011-01-01

    in vivo imaging of infrared fluorescent "Katushka" and erythropoietin evaluated by ELISA and hemoglobin. Histology was performed. Electrotransfer of Katushka and erythropoietin yielded significant expression. Maximal calcium uptake occurred after injection of Ca(2+) before electropulsing using eight high...

  12. Activation of calcium-sensing receptor increases TRPC3 expression in rat cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Feng, Shan-Li [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Sun, Ming-Rui [Department of Pharmacology, Qiqihaer Medical College, Qiqihaer 160001 (China); Li, Ting-Ting; Yin, Xin [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China); Xu, Chang-Qing [Department of Pathophysiology, Harbin Medical University, Harbin 150086 (China); Sun, Yi-Hua, E-mail: syh200415@126.com [Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086 (China)

    2011-03-11

    Research highlights: {yields} Calcium-sensing receptor (CaR) activation stimulates TRP channels. {yields} CaR promoted transient receptor potential C3 (TRPC3) expression. {yields} Adult rat ventricular myocytes display capacitative calcium entry (CCE), which was operated by TRPCs. {yields} TRPC channels activation induced by CaR activator sustained the increased [Ca{sup 2+}]{sub i} to evoke cardiomyocytes apoptosis. -- Abstract: Transient receptor potential (TRP) channels are expressed in cardiomyocytes, which gate a type of influx of extracellular calcium, the capacitative calcium entry. TRP channels play a role in mediating Ca{sup 2+} overload in the heart. Calcium-sensing receptors (CaR) are also expressed in rat cardiac tissue and promote the apoptosis of cardiomyocytes by Ca{sup 2+} overload. However, data about the link between CaR and TRP channels in rat heart are few. In this study, reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting were used to examine the expression of the TRP canonical proteins TRPC1 and TRPC3 in adult and neonatal rat cardiomyocytes. Laser scan confocal microscopy was used to detect intracellular [Ca{sup 2+}]{sub i} levels in isolated adult rat ventricular myocytes. The results showed that, in adult rat cardiomyocytes, the depletion of Ca{sup 2+} stores in the endoplasmic/sarcoplasmic reticulum (ER/SR) by thapsigargin induced a transient increase in [Ca{sup 2+}]{sub i} in the absence of [Ca{sup 2+}]{sub o} and the subsequent restoration of [Ca{sup 2+}]{sub o} sustained the increased [Ca{sup 2+}]{sub i} for a few minutes, whereas, the persisting elevation of [Ca{sup 2+}]{sub i} was reduced in the presence of the TRPC inhibitor SKF96365. The stimulation of CaR by its activator gadolinium chloride (GdCl{sub 3}) or spermine also resulted in the same effect and the duration of [Ca{sup 2+}]{sub i} increase was also shortened in the absence of [Ca{sup 2+}]{sub o}. In adult and neonatal rat cardiomyocytes, GdCl{sub 3

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

    NARCIS (Netherlands)

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

    2009-01-01

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

  14. 5,6-EET potently inhibits T-type calcium channels

    DEFF Research Database (Denmark)

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

    2014-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2011-07-29

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2007-08-01

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

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

    Directory of Open Access Journals (Sweden)

    Shuyuan Liu

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

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

    Science.gov (United States)

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

    2016-01-01

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

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

    OpenAIRE

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

    2009-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

    Lazniewska, Joanna; Weiss, Norbert

    2017-01-01

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

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

    DEFF Research Database (Denmark)

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

    2007-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2016-06-15

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

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

    International Nuclear Information System (INIS)

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

    1990-01-01

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

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

    Science.gov (United States)

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

    2014-04-01

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

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

    International Nuclear Information System (INIS)

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

    1989-01-01

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

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

    Science.gov (United States)

    Kamaleddin, Mohammad Amin

    2018-02-01

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

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

    Science.gov (United States)

    Benhassine, Narimane; Berger, Thomas

    2005-02-01

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

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

    DEFF Research Database (Denmark)

    Hansen, P B L

    2013-01-01

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

  9. Transcriptional expression analysis of genes involved in regulation of calcium translocation and storage in finger millet (Eleusine coracana L. Gartn.).

    Science.gov (United States)

    Mirza, Neelofar; Taj, Gohar; Arora, Sandeep; Kumar, Anil

    2014-10-25

    Finger millet (Eleusine coracana) variably accumulates calcium in different tissues, due to differential expression of genes involved in uptake, translocation and accumulation of calcium. Ca(2+)/H(+) antiporter (CAX1), two pore channel (TPC1), CaM-stimulated type IIB Ca(2+) ATPase and two CaM dependent protein kinase (CaMK1 and 2) homologs were studied in finger millet. Two genotypes GP-45 and GP-1 (high and low calcium accumulating, respectively) were used to understand the role of these genes in differential calcium accumulation. For most of the genes higher expression was found in the high calcium accumulating genotype. CAX1 was strongly expressed in the late stages of spike development and could be responsible for accumulating high concentrations of calcium in seeds. TPC1 and Ca(2+) ATPase homologs recorded strong expression in the root, stem and developing spike and signify their role in calcium uptake and translocation, respectively. Calmodulin showed strong expression and a similar expression pattern to the type IIB ATPase in the developing spike only and indicating developing spike or even seed specific isoform of CaM affecting the activity of downstream target of calcium transportation. Interestingly, CaMK1 and CaMK2 had expression patterns similar to ATPase and TPC1 in various tissues raising a possibility of their respective regulation via CaM kinase. Expression pattern of 14-3-3 gene was observed to be similar to CAX1 gene in leaf and developing spike inferring a surprising possibility of CAX1 regulation through 14-3-3 protein. Our results provide a molecular insight for explaining the mechanism of calcium accumulation in finger millet. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Calcium regulates the expression of a Dictyostelium discoideum ...

    Indian Academy of Sciences (India)

    In a screen for calcium-regulated gene expression during growth and development of Dictyostelium discoideum we have identified an asparaginyl tRNA synthetase (ddAsnRS) gene, the second tRNA synthetase gene identified in this organism. The ddAsnRS gene shows many unique features. One, it is repressed by ...

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

    Science.gov (United States)

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

    2013-11-15

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

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

    Directory of Open Access Journals (Sweden)

    Jianjun Wu

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

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

    DEFF Research Database (Denmark)

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

    2011-01-01

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

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

    Science.gov (United States)

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

    2010-12-30

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

  15. Fragile X mental retardation protein controls ion channel expression and activity.

    Science.gov (United States)

    Ferron, Laurent

    2016-10-15

    Fragile X-associated disorders are a family of genetic conditions resulting from the partial or complete loss of fragile X mental retardation protein (FMRP). Among these disorders is fragile X syndrome, the most common cause of inherited intellectual disability and autism. FMRP is an RNA-binding protein involved in the control of local translation, which has pleiotropic effects, in particular on synaptic function. Analysis of the brain FMRP transcriptome has revealed hundreds of potential mRNA targets encoding postsynaptic and presynaptic proteins, including a number of ion channels. FMRP has been confirmed to bind voltage-gated potassium channels (K v 3.1 and K v 4.2) mRNAs and regulates their expression in somatodendritic compartments of neurons. Recent studies have uncovered a number of additional roles for FMRP besides RNA regulation. FMRP was shown to directly interact with, and modulate, a number of ion channel complexes. The sodium-activated potassium (Slack) channel was the first ion channel shown to directly interact with FMRP; this interaction alters the single-channel properties of the Slack channel. FMRP was also shown to interact with the auxiliary β4 subunit of the calcium-activated potassium (BK) channel; this interaction increases calcium-dependent activation of the BK channel. More recently, FMRP was shown to directly interact with the voltage-gated calcium channel, Ca v 2.2, and reduce its trafficking to the plasma membrane. Studies performed on animal models of fragile X syndrome have revealed links between modifications of ion channel activity and changes in neuronal excitability, suggesting that these modifications could contribute to the phenotypes observed in patients with fragile X-associated disorders. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

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

    DEFF Research Database (Denmark)

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

    2007-01-01

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

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

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

    International Nuclear Information System (INIS)

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

    1985-01-01

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

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

    Science.gov (United States)

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

    2013-09-01

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

  20. A Gate Hinge Controls the Epithelial Calcium Channel TRPV5

    OpenAIRE

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

    2017-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Masahiro Kawahara

    2011-01-01

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

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

    International Nuclear Information System (INIS)

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

    1991-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Mueller Rachel

    2008-04-01

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

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

    Science.gov (United States)

    Walker, Michael J A

    2015-11-15

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

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

    Science.gov (United States)

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

    2009-06-01

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

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

    Science.gov (United States)

    Kumari, Neema; Gaur, Himanshu; Bhargava, Anamika

    2018-02-01

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

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

    Science.gov (United States)

    Hansen, P B L

    2013-04-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2002-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

    Proft, Juliane; Weiss, Norbert

    2015-01-01

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

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

    NARCIS (Netherlands)

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

    2017-01-01

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

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

    International Nuclear Information System (INIS)

    Robinson, P.J.; Lovenberg, Walter

    1986-01-01

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

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

    Science.gov (United States)

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

    2017-08-01

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

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

    Science.gov (United States)

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

    2015-11-25

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

  14. Calcium-mediated signaling and calmodulin-dependent kinase regulate hepatocyte-inducible nitric oxide synthase expression.

    Science.gov (United States)

    Zhang, Baochun; Crankshaw, Will; Nesemeier, Ryan; Patel, Jay; Nweze, Ikenna; Lakshmanan, Jaganathan; Harbrecht, Brian G

    2015-02-01

    Induced nitric oxide synthase (iNOS) is induced in hepatocytes by shock and inflammatory stimuli. Excessive NO from iNOS mediates shock-induced hepatic injury and death, so understanding the regulation of iNOS will help elucidate the pathophysiology of septic shock. In vitro, cytokines induce iNOS expression through activation of signaling pathways including mitogen-activated protein kinases and nuclear factor κB. Cytokines also induce calcium (Ca(2+)) mobilization and activate calcium-mediated intracellular signaling pathways, typically through activation of calmodulin-dependent kinases (CaMK). Calcium regulates NO production in macrophages but the role of calcium and calcium-mediated signaling in hepatocyte iNOS expression has not been defined. Primary rat hepatocytes were isolated, cultured, and induced to produce NO with proinflammatory cytokines. Calcium mobilization and Ca(2+)-mediated signaling were altered with ionophore, Ca(2+) channel blockers, and inhibitors of CaMK. The Ca(2+) ionophore A23187 suppressed cytokine-stimulated NO production, whereas Ethylene glycol tetraacetic acid and nifedipine increased NO production, iNOS messenger RNA, and iNOS protein expression. Inhibition of CaMK with KN93 and CBD increased NO production but the calcineurin inhibitor FK 506 decreased iNOS expression. These data demonstrate that calcium-mediated signaling regulates hepatocyte iNOS expression and does so through a mechanism independent of calcineurin. Changes in intracellular calcium levels may regulate iNOS expression during hepatic inflammation induced by proinflammatory cytokines. Copyright © 2015 Elsevier Inc. All rights reserved.

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

    DEFF Research Database (Denmark)

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

    2012-01-01

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

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

    Science.gov (United States)

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

    2017-08-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2016-01-01

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

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

    OpenAIRE

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

    2015-01-01

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

  19. Neuronal calcium channel antagonists. Discrimination between calcium channel subtypes using omega-conotoxin from Conus magus venom

    International Nuclear Information System (INIS)

    Olivera, B.M.; Cruz, L.J.; de Santos, V.

    1987-01-01

    The omega-conotoxins from the venom of fish-hunting cone snails are probably the most useful of presently available ligands for neuronal Ca channels from vertebrates. Two of these peptide toxins, omega-conotoxins MVIIA and MVIIB from the venom of Conus magus, were purified. The amino acid sequences show significant differences from omega-conotoxins from Conus geographus. Total synthesis of omega-conotoxin MVIIA was achieved, and biologically active radiolabeled toxin was produced by iodination. Although omega-conotoxins from C. geographus (GVIA) and C. magus (MVIIA) appear to compete for the same sites in mammalian brain, in amphibian brain the high-affinity binding of omega-conotoxin MVIIA has narrower specificity. In this system, it is demonstrated that a combination of two omega-conotoxins can be used for biochemically defining receptor subtypes and suggested that these correspond to subtypes of neutronal Ca 2+ channels

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

    Science.gov (United States)

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

    2013-11-15

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

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

    Science.gov (United States)

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

    2017-12-01

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

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

    Directory of Open Access Journals (Sweden)

    Yunzhao R Ren

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

  3. Testosterone increases urinary calcium excretion and inhibits expression of renal calcium transport proteins.

    NARCIS (Netherlands)

    Hsu, Y.J.; Dimke, H.; Schoeber, J.P.H.; Hsu, S.C.; Lin, S.H.; Chu, P.; Hoenderop, J.G.J.; Bindels, R.J.M.

    2010-01-01

    Although gender differences in the renal handling of calcium have been reported, the overall contribution of androgens to these differences remains uncertain. We determined here whether testosterone affects active renal calcium reabsorption by regulating calcium transport proteins. Male mice had

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

    Science.gov (United States)

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

    2013-11-01

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

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

    Science.gov (United States)

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

    2016-10-01

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

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

    Science.gov (United States)

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

    1993-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Ji-Guang Wang

    2009-07-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1985-05-01

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

  9. Autosomal dominant hypercalciuria in a mouse model due to a mutation of the epithelial calcium channel, TRPV5.

    Directory of Open Access Journals (Sweden)

    Nellie Y Loh

    Full Text Available Hypercalciuria is a major cause of nephrolithiasis, and is a common and complex disorder involving genetic and environmental factors. Identification of genetic factors for monogenic forms of hypercalciuria is hampered by the limited availability of large families, and to facilitate such studies, we screened for hypercalciuria in mice from an N-ethyl-N-nitrosourea mutagenesis programme. We identified a mouse with autosomal dominant hypercalciuria (HCALC1. Linkage studies mapped the Hcalc1 locus to a 11.94 Mb region on chromosome 6 containing the transient receptor potential cation channel, subfamily V, members 5 (Trpv5 and 6 (Trpv6 genes. DNA sequence analysis of coding regions, intron-exon boundaries and promoters of Trpv5 and Trpv6 identified a novel T to C transition in codon 682 of TRPV5, mutating a conserved serine to a proline (S682P. Compared to wild-type littermates, heterozygous (Trpv5(682P/+ and homozygous (Trpv5(682P/682P mutant mice had hypercalciuria, polyuria, hyperphosphaturia and a more acidic urine, and ∼10% of males developed tubulointerstitial nephritis. Trpv5(682P/682P mice also had normal plasma parathyroid hormone but increased 1,25-dihydroxyvitamin D(3 concentrations without increased bone resorption, consistent with a renal defect for the hypercalciuria. Expression of the S682P mutation in human embryonic kidney cells revealed that TRPV5-S682P-expressing cells had a lower baseline intracellular calcium concentration than wild-type TRPV5-expressing cells, suggesting an altered calcium permeability. Immunohistological studies revealed a selective decrease in TRPV5-expression from the renal distal convoluted tubules of Trpv5(682P/+ and Trpv5(682P/682P mice consistent with a trafficking defect. In addition, Trpv5(682P/682P mice had a reduction in renal expression of the intracellular calcium-binding protein, calbindin-D(28K, consistent with a specific defect in TRPV5-mediated renal calcium reabsorption. Thus, our findings

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

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    Gorash ZM

    2007-01-01

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

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

    DEFF Research Database (Denmark)

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

    2011-01-01

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

  12. Stretch induced endothelin-1 secretion by adult rat astrocytes involves calcium influx via stretch-activated ion channels (SACs)

    International Nuclear Information System (INIS)

    Ostrow, Lyle W.; Suchyna, Thomas M.; Sachs, Frederick

    2011-01-01

    Highlights: → Endothelin-1 expression by adult rat astrocytes correlates with cell proliferation. → Stretch-induced ET-1 is inhibited by GsMtx-4, a specific inhibitor of Ca 2+ permeant SACs. → The less specific SAC inhibitor streptomycin also inhibits ET-1 secretion. → Stretch-induced ET-1 production depends on a calcium influx. → SAC pharmacology may provide a new class of therapeutic agents for CNS pathology. -- Abstract: The expression of endothelins (ETs) and ET-receptors is often upregulated in brain pathology. ET-1, a potent vasoconstrictor, also inhibits the expression of astrocyte glutamate transporters and is mitogenic for astrocytes, glioma cells, neurons, and brain capillary endothelia. We have previously shown that mechanical stress stimulates ET-1 production by adult rat astrocytes. We now show in adult astrocytes that ET-1 production is driven by calcium influx through stretch-activated ion channels (SACs) and the ET-1 production correlates with cell proliferation. Mechanical stimulation using biaxial stretch ( 2+ threshold. This coupling of mechanical stress to the astrocyte endothelin system through SACs has treatment implications, since all pathology deforms the surrounding parenchyma.

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

    Directory of Open Access Journals (Sweden)

    Lei Zhu

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

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

    Science.gov (United States)

    Gulothungan, G.; Malathi, R.

    2018-04-01

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

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

    Science.gov (United States)

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

    2015-01-12

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

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

    International Nuclear Information System (INIS)

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

    2006-01-01

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

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

    Science.gov (United States)

    Boda, Dezső; Henderson, Douglas; Gillespie, Dirk

    2013-08-07

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

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

    Science.gov (United States)

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

    2017-05-01

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

  19. Human neuronal stargazin-like proteins, γ2, γ3 and γ4; an investigation of their specific localization in human brain and their influence on CaV2.1 voltage-dependent calcium channels expressed in Xenopus oocytes.

    Directory of Open Access Journals (Sweden)

    Dolphin Annette C

    2003-09-01

    Full Text Available Abstract Background Stargazin (γ2 and the closely related γ3, and γ4 transmembrane proteins are part of a family of proteins that may act as both neuronal voltage-dependent calcium channel (VDCC γ subunits and transmembrane α-amino-3-hydroxy-5-methyl-4-isoxazoleproponinc (AMPA receptor regulatory proteins (TARPs. In this investigation, we examined the distribution patterns of the stargazin-like proteins γ2, γ3, and γ4 in the human central nervous system (CNS. In addition, we investigated whether human γ2 or γ4 could modulate the electrophysiological properties of a neuronal VDCC complex transiently expressed in Xenopus oocytes. Results The mRNA encoding human γ2 is highly expressed in cerebellum, cerebral cortex, hippocampus and thalamus, whereas γ3 is abundant in cerebral cortex and amygdala and γ4 in the basal ganglia. Immunohistochemical analysis of the cerebellum determined that both γ2 and γ4 are present in the molecular layer, particularly in Purkinje cell bodies and dendrites, but have an inverse expression pattern to one another in the dentate cerebellar nucleus. They are also detected in the interneurons of the granule cell layer though only γ2 is clearly detected in granule cells. The hippocampus stains for γ2 and γ4 throughout the layers of the every CA region and the dentate gyrus, whilst γ3 appears to be localized particularly to the pyramidal and granule cell bodies. When co-expressed in Xenopus oocytes with a CaV2.1/β4 VDCC complex, either in the absence or presence of an α2δ2 subunit, neither γ2 nor γ4 significantly modulated the VDCC peak current amplitude, voltage-dependence of activation or voltage-dependence of steady-state inactivation. Conclusion The human γ2, γ3 and γ4 stargazin-like proteins are detected only in the CNS and display differential distributions among brain regions and several cell types in found in the cerebellum and hippocampus. These distribution patterns closely resemble those

  20. High calcium concentration in bones promotes bone metastasis in renal cell carcinomas expressing calcium-sensing receptor.

    Science.gov (United States)

    Joeckel, Elke; Haber, Tobias; Prawitt, Dirk; Junker, Kerstin; Hampel, Christian; Thüroff, Joachim W; Roos, Frederik C; Brenner, Walburgis

    2014-02-28

    The prognosis for renal cell carcinoma (RCC) is related to a high rate of metastasis, including 30% of bone metastasis. Characteristic for bone tissue is a high concentration of calcium ions. In this study, we show a promoting effect of an enhanced extracellular calcium concentration on mechanisms of bone metastasis via the calcium-sensing receptor (CaSR) and its downstream signaling molecules. Our analyses were performed using 33 (11/category) matched specimens of normal and tumor tissue and 9 (3/category) primary cells derived from RCC patients of the 3 categories: non-metastasized, metastasized into the lung and metastasized into bones during a five-year period after nephrectomy. Expression of CaSR was determined by RT-PCR, Western blot analyses and flow cytometry, respectively. Cells were treated by calcium and the CaSR inhibitor NPS 2143. Cell migration was measured in a Boyden chamber with calcium (10 μM) as chemotaxin and proliferation by BrdU incorporation. The activity of intracellular signaling mediators was quantified by a phospho-kinase array and Western blot. The expression of CaSR was highest in specimens and cells of patients with bone metastases. Calcium treatment induced an increased migration (19-fold) and proliferation (2.3-fold) exclusively in RCC cells from patients with bone metastases. The CaSR inhibitor NPS 2143 elucidated the role of CaSR on the calcium-dependent effects. After treatment with calcium, the activity of AKT, PLCγ-1, p38α and JNK was clearly enhanced and PTEN expression was almost completely abolished in bone metastasizing RCC cells. Our results indicate a promoting effect of extracellular calcium on cell migration and proliferation of bone metastasizing RCC cells via highly expressed CaSR and its downstream signaling pathways. Consequently, CaSR may be regarded as a new prognostic marker predicting RCC bone metastasis.

  1. Calcium activated K⁺ channels in the electroreceptor of the skate confirmed by cloning. Details of subunits and splicing.

    Science.gov (United States)

    King, Benjamin L; Shi, Ling Fang; Kao, Peter; Clusin, William T

    2016-03-01

    Elasmobranchs detect small potentials using excitable cells of the ampulla of Lorenzini which have calcium-activated K(+) channels, first described in 1974. A distinctive feature of the outward current in voltage clamped ampullae is its apparent insensitivity to voltage. The sequence of a BK channel α isoform expressed in the ampulla of the skate was characterized. A signal peptide is present at the beginning of the gene. When compared to human isoform 1 (the canonical sequence), the largest difference was absence of a 59 amino acid region from the S8-S9 intra-cellular linker that contains the strex regulatory domain. The ampulla isoform was also compared with the isoform predicted in late skate embryos where strex was also absent. The BK voltage sensors were conserved in both skate isoforms. Differences between the skate and human BK channel included alternative splicing. Alternative splicing occurs at seven previously defined sites that are characteristic for BK channels in general and hair cells in particular. Skate BK sequences were highly similar to the Australian ghost shark and several other vertebrate species. Based on alignment of known BK sequences with the skate genome and transcriptome, there are at least two isoforms of Kcnma1α expressed in the skate. One of the β subunits (β4), which is known to decrease voltage sensitivity, was also identified in the skate genome and transcriptome and in the ampulla. These studies advance our knowledge of BK channels and suggest further studies in the ampulla and other excitable tissues. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Calcium Activated K+ Channels in The Electroreceptor of the Skate Confirmed by Cloning. Details of Subunits and Splicing

    Science.gov (United States)

    King, Benjamin L.; Shi, Ling Fang; Kao, Peter; Clusin, William T.

    2015-01-01

    Elasmobranchs detect small potentials using excitable cells of the ampulla of Lorenzini which have calcium-activated K+ channels, first described in l974. A distinctive feature of the outward current in voltage clamped ampullae is its apparent insensitivity to voltage. The sequence of a BK channel α isoform expressed in the ampulla of the skate was characterized. A signal peptide is present at the beginning of the gene. When compared to human isoform 1 (the canonical sequence), the largest difference was absence of a 59 amino acid region from the S8-S9 intracellular linker that contains the strex regulatory domain. The ampulla isoform was also compared with the isoform predicted˜ in late skate embryos where strex was also absent. The BK voltage sensors were conserved in both skate isoforms. Differences between the skate and human BK channel included alternative splicing. Alternative splicing occurs at seven previously defined sites that are characteristic for BK channels in general and hair cells in particular. Skate BK sequences were highly similar to the Australian ghost shark and several other vertebrate species. Based on alignment of known BK sequences with the skate genome and transcriptome, there are at least two isoforms of Kcnma1α expressed in the skate. One of the β subunits (β4), which is known to decrease voltage sensitivity, was also identified in the skate genome and transcriptome and in the ampulla. These studies advance our knowledge of BK channels and suggest further studies in the ampulla and other excitable tissues. PMID:26687710

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

    Science.gov (United States)

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

    2010-01-01

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

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

    Science.gov (United States)

    Fried, G; Liu, Y A

    1994-08-01

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

  5. Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling.

    Science.gov (United States)

    Szebényi, Kornélia; Füredi, András; Kolacsek, Orsolya; Pergel, Enikő; Bősze, Zsuzsanna; Bender, Balázs; Vajdovich, Péter; Tóvári, József; Homolya, László; Szakács, Gergely; Héja, László; Enyedi, Ágnes; Sarkadi, Balázs; Apáti, Ágota; Orbán, Tamás I

    2015-08-03

    In drug discovery, prediction of selectivity and toxicity require the evaluation of cellular calcium homeostasis. The rat is a preferred laboratory animal for pharmacology and toxicology studies, while currently no calcium indicator protein expressing rat model is available. We established a transgenic rat strain stably expressing the GCaMP2 fluorescent calcium sensor by a transposon-based methodology. Zygotes were co-injected with mRNA of transposase and a CAG-GCaMP2 expressing construct, and animals with one transgene copy were pre-selected by measuring fluorescence in blood cells. A homozygous rat strain was generated with high sensor protein expression in the heart, kidney, liver, and blood cells. No pathological alterations were found in these animals, and fluorescence measurements in cardiac tissue slices and primary cultures demonstrated the applicability of this system for studying calcium signaling. We show here that the GCaMP2 expressing rat cardiomyocytes allow the prediction of cardiotoxic drug side-effects, and provide evidence for the role of Na(+)/Ca(2+) exchanger and its beneficial pharmacological modulation in cardiac reperfusion. Our data indicate that drug-induced alterations and pathological processes can be followed by using this rat model, suggesting that transgenic rats expressing a calcium-sensitive protein provide a valuable system for pharmacological and toxicological studies.

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

    Science.gov (United States)

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

    2018-04-26

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

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

    Science.gov (United States)

    Findeisen, Felix; Minor, Daniel L

    2009-03-01

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

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

    Directory of Open Access Journals (Sweden)

    Mercè eIzquierdo-Serra

    2013-03-01

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

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

    Directory of Open Access Journals (Sweden)

    Milanović Tamara

    2015-01-01

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

  10. Enhanced expression of a calcium-dependent protein kinase

    Indian Academy of Sciences (India)

    Among the downstream targets of calcium in plants, calcium-dependent protein kinases (CDPKs) form an interesting class of kinases which are activated by calcium binding. They have been implicated in a diverse array of responses to hormonal and environmental stimuli. In order to dissect the role of CDPKs in the moss ...

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

    Directory of Open Access Journals (Sweden)

    Charalampos Seretis

    2012-01-01

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

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

    International Nuclear Information System (INIS)

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

    1991-01-01

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

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

    Science.gov (United States)

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

    2009-12-01

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

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

    DEFF Research Database (Denmark)

    Hansen, Pernille B L

    2015-01-01

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

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

    Science.gov (United States)

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

    2017-01-01

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

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

    Science.gov (United States)

    Grimsley, Calum Alex; Green, David Brian

    2016-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Vanessa Silva-Moraes

    2013-08-01

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

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

    Directory of Open Access Journals (Sweden)

    Lei Wen

    2006-04-01

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

  19. Calcium

    Science.gov (United States)

    ... You can get decent amounts of calcium from baked beans, navy beans, white beans, and others. Canned fish. You're in luck if you like sardines and canned salmon with bones. Almond milk. Working Calcium Into Your ...

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

    Czech Academy of Sciences Publication Activity Database

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

    2009-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Yu. V. Lukina

    2010-01-01

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

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

    DEFF Research Database (Denmark)

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

    2015-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Yuhuan Luo

    2018-06-01

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

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

    International Nuclear Information System (INIS)

    Lian Luyun; Myatt, Daniel; Kitmitto, Ashraf

    2007-01-01

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

  5. Orai3 channel is the 2-APB-induced endoplasmic reticulum calcium leak.

    Science.gov (United States)

    Leon-Aparicio, Daniel; Pacheco, Jonathan; Chavez-Reyes, Jesus; Galindo, Jose M; Valdes, Jesus; Vaca, Luis; Guerrero-Hernandez, Agustin

    2017-07-01

    We have studied in HeLa cells the molecular nature of the 2-APB induced ER Ca 2+ leak using synthetic Ca 2+ indicators that report changes in both the cytoplasmic ([Ca 2+ ] i ) and the luminal ER ([Ca 2+ ] ER ) Ca 2+ concentrations. We have tested the hypothesis that Orai channels participate in the 2-APB-induced ER Ca 2+ leak that was characterized in the companion paper. The expression of the dominant negative Orai1 E106A mutant, which has been reported to block the activity of all three types of Orai channels, inhibited the effect of 2-APB on the [Ca 2+ ] ER but did not decrease the ER Ca 2+ leak after thapsigargin (TG). Orai3 channel, but neither Orai1 nor Orai2, colocalizes with expressed IP 3 R and only Orai3 channel supported the 2-APB-induced ER Ca 2+ leak, while Orai1 and Orai2 inhibited this type of ER Ca 2+ leak. Decreasing the expression of Orai3 inhibited the 2-APB-induced ER Ca 2+ leak but did not modify the ER Ca 2+ leak revealed by inhibition of SERCA pumps with TG. However, reducing the expression of Orai3 channel resulted in larger [Ca 2+ ] i response after TG but only when the ER store had been overloaded with Ca 2+ by eliminating the acidic internal Ca 2+ store with bafilomycin. These data suggest that Orai3 channel does not participate in the TG-revealed ER Ca 2+ leak but forms an ER Ca 2+ leak channel that is limiting the overloading with Ca 2+ of the ER store. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

    Science.gov (United States)

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

    2018-05-10

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

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

    Directory of Open Access Journals (Sweden)

    Abhinaba Ghosh

    2017-12-01

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

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

    Science.gov (United States)

    Mehanna, Ahmed S; Kim, Jin Yung

    2005-07-01

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

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

    International Nuclear Information System (INIS)

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

    1990-01-01

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

  10. The transition from proliferation to differentiation in colorectal cancer is regulated by the calcium activated chloride channel A1.

    Directory of Open Access Journals (Sweden)

    Bo Yang

    Full Text Available Breaking the balance between proliferation and differentiation in animal cells can lead to cancer, but the mechanisms maintaining this balance remain largely undefined. The calcium activated chloride channel A1 (CLCA1 is a member of the calcium sensitive chloride conductance family of proteins and is expressed mainly in the colon, small intestine and appendix. We show that CLCA1 plays a functional role in differentiation and proliferation of Caco-2 cells and of intestinal tissue. Caco-2 cells spontaneously differentiate either in confluent culture or when treated with butyrate, a molecule present naturally in the diet. Here, we compared CLCA1 expressional levels between patients with and without colorectal cancer (CRC and determined the functional role of CLCA1 in differentiation and proliferation of Caco-2 cells. We showed that: 1 CLCA1 and CLCA4 expression were down-regulated significantly in CRC patients; 2 CLCA1 expression was up-regulated in Caco-2 cells induced to differentiate by confluent culture or by treatment with sodium butyrate (NaBT; 3 Knockdown of CLCA1 with siRNA significantly inhibited cell differentiation and promoted cell proliferation in Caco-2 confluent cultures, and 4 In Caco-2 3D culture, suppression of CLCA1 significantly increased cell proliferation and compromised NaBT-induced inhibition of proliferation. In conclusion, CLCA1 may contribute to promoting spontaneous differentiation and reducing proliferation of Caco-2 cells and may be a target of NaBT-induced inhibition of proliferation and therefore a potential diagnostic marker for CRC prognosis.

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

    Science.gov (United States)

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

    2014-01-01

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

  12. Interactions of waterborne and dietary cadmium on the expression of calcium transporters in the gills of rainbow trout: Influence of dietary calcium supplementation

    International Nuclear Information System (INIS)

    Galvez, Fernando; Franklin, Natasha M.; Tuttle, Ryan B.; Wood, Chris M.

    2007-01-01

    Recent studies have shown that dietary Ca 2+ supplementation strongly inhibits uptake of Ca 2+ and Cd at the fish gill. To better understand the influence of dietary Ca 2+ on branchial Ca 2+ transport, we examined the expression of two trout gill calcium transporters during waterborne and dietary Cd exposure, at two different levels of dietary Ca 2+ . Quantitative polymerase chain reaction (PCR) was used to monitor epithelial calcium channel (ECaC) and sodium-calcium exchange (NCX) mRNA levels following 7-28 days of exposure to these treatments. In brief, juvenile rainbow trout (Oncorhynchus mykiss) were exposed to control, 3 μg/L waterborne Cd, 500 mg/kg dietary Cd, or a combined 3 μg/L waterborne plus 500 mg/kg dietary Cd exposure, supplemented with either 20 mg/g or 60 mg/g dietary calcium (Ca 2+ ). Two-way analysis of variance was used to discern the main effects of Cd exposure and dietary Ca 2+ supplementation on ECaC and NCX mRNA levels. We found that dietary Ca 2+ supplementation decreased significantly ECaC mRNA expression on days 14 and 21. In comparison, NCX mRNA levels were not influenced by dietary Ca 2+ supplementation, but rather were significantly inhibited in the combined waterborne and dietary Cd exposure on day 7 alone. Statistical analysis found no interactive effects between Cd exposure and dietary Ca 2+ exposure at any time point, except for day 28.This study provides evidence of the importance of nutritional status on the transcriptional regulation of ion transport at the fish gill. We discuss the importance of diet and nutritional status to the development of new regulatory approaches, such as the biotic ligand model, which currently do not account for the significance of diet on metal bioavailability in aquatic organisms

  13. Blockade of T-type calcium channels prevents tonic-clonic seizures in a maximal electroshock seizure model.

    Science.gov (United States)

    Sakkaki, Sophie; Gangarossa, Giuseppe; Lerat, Benoit; Françon, Dominique; Forichon, Luc; Chemin, Jean; Valjent, Emmanuel; Lerner-Natoli, Mireille; Lory, Philippe

    2016-02-01

    T-type (Cav3) calcium channels play important roles in neuronal excitability, both in normal and pathological activities of the brain. In particular, they contribute to hyper-excitability disorders such as epilepsy. Here we have characterized the anticonvulsant properties of TTA-A2, a selective T-type channel blocker, in mouse. Using the maximal electroshock seizure (MES) as a model of tonic-clonic generalized seizures, we report that mice treated with TTA-A2 (0.3 mg/kg and higher doses) were significantly protected against tonic seizures. Although no major change in Local Field Potential (LFP) pattern was observed during the MES seizure, analysis of the late post-ictal period revealed a significant increase in the delta frequency power in animals treated with TTA-A2. Similar results were obtained for Cav3.1-/- mice, which were less prone to develop tonic seizures in the MES test, but not for Cav3.2-/- mice. Analysis of extracellular signal-regulated kinase 1/2 (ERK) phosphorylation and c-Fos expression revealed a rapid and elevated neuronal activation in the hippocampus following MES clonic seizures, which was unchanged in TTA-A2 treated animals. Overall, our data indicate that TTA-A2 is a potent anticonvulsant and that the Cav3.1 isoform plays a prominent role in mediating TTA-A2 tonic seizure protection. Copyright © 2015. Published by Elsevier Ltd.

  14. Interactions between β-catenin and the HSlo potassium channel regulates HSlo surface expression.

    Directory of Open Access Journals (Sweden)

    Shumin Bian

    Full Text Available The large conductance calcium-activated potassium channel alpha-subunit (Slo is widely distributed throughout the body and plays an important role in a number of diseases. Prior work has shown that Slo, through its S10 region, interacts with β-catenin, a key component of the cytoskeleton framework and the Wnt signaling pathway. However, the physiological significance of this interaction was not clear.Using a combination of proteomic and cell biology tools we show the existence of additional multiple binding sites in Slo, and explore in detail β-catenin interactions with the S10 region. We demonstrate that deletion of this region reduces Slo surface expression in HEK cells, which indicates that interaction with beta-catenin is important for Slo surface expression. This is confirmed by reduced expression of Slo in HEK cells and chicken (Gallus gallus domesticus leghorn white hair cells treated with siRNA to β-catenin. HSlo reciprocally co-immunoprecipitates with β-catenin, indicating a stable binding between these two proteins, with the S10 deletion mutant having reduced binding with β-catenin. We also observed that mutations of the two putative GSK phosphorylation sites within the S10 region affect both the surface expression of Slo and the channel's voltage and calcium sensitivities. Interestingly, expression of exogenous Slo in HEK cells inhibits β-catenin-dependent canonical Wnt signaling.These studies identify for the first time a central role for β-catenin in mediating Slo surface expression. Additionally we show that Slo overexpression can lead to downregulation of Wnt signaling.

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

    Science.gov (United States)

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

    2013-09-15

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

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

    Directory of Open Access Journals (Sweden)

    Keith P. Johnson

    2017-10-01

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

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

    Science.gov (United States)

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

    1993-03-01

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

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

    Directory of Open Access Journals (Sweden)

    Matthew Prideaux

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

  19. Calcium measurements in living filamentous fungi expressing codon-optimized aequorin

    NARCIS (Netherlands)

    Nelson, G.; Kozlova-Zwinderman, O.; Collis, A.J.; Knight, M.R.; Fincham, J.R.S.; Stanger, C.P.; Renwick, A.; Hessing, J.G.M.; Punt, P.J.; Hondel, C.A.M.J.J. van den; Read, N.D.

    2004-01-01

    Calcium signalling is little understood in filamentous fungi largely because easy and routine methods for calcium measurement in living hyphae have previously been unavailable. We have developed the recombinant aequorin method for this purpose. High levels of aequorin expression were obtained in

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

    International Nuclear Information System (INIS)

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

    1989-01-01

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

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

    Science.gov (United States)

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

    2015-12-01

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

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

    Directory of Open Access Journals (Sweden)

    See-Ziau Hoe

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    John P Cavaretta

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

  4. Differential intracellular calcium influx, nitric oxide production, ICAM-1 and IL8 expression in primary bovine endothelial cells exposed to nonesterified fatty acids.

    Science.gov (United States)

    Loaiza, Anitsi; Carretta, María D; Taubert, Anja; Hermosilla, Carlos; Hidalgo, María A; Burgos, Rafael A

    2016-02-25

    Nonesterified fatty acids (NEFAs) are involved in proinflammatory processes in cattle, including in the increased expression of adhesion molecules in endothelial cells. However, the mechanisms underlying these effects are still unknown. The aim of this study was to assess the effects of NEFAs on the intracellular calcium (Ca(2+) i) influx, nitric oxide production, and ICAM-1 and IL-8 expression in primary bovine umbilical vein endothelial cells (BUVECs). Myristic (MA), palmitic (PA), stearic (SA), oleic (OA) and linoleic acid (LA) rapidly increased Ca(2+) i. The calcium response to all tested NEFAs showed an extracellular calcium dependence and only the LA response was significantly inhibited until the intracellular calcium was chelated. The EC50 values for MA and LA were 125 μM and 37 μM, respectively, and the MA and LA effects were dependent on calcium release from the endoplasmic reticulum stores and on the L-type calcium channels. Only the calcium response to MA was significantly reduced by GW1100, a selective G-protein-coupled free fatty acid receptor (GPR40) antagonist. We also detected a functional FFAR1/GPR40 protein in BUVECs by using western blotting and the FFAR1/GPR40 agonist TAK-875. Only LA increased the cellular nitric oxide levels in a calcium-dependent manner. LA stimulation but not MA stimulation increased ICAM-1 and IL-8-expression in BUVECs. This effect was inhibited by GW1100, an antagonist of FFAR1/GPR40, but not by U-73122, a phospholipase C inhibitor. These findings strongly suggest that each individual NEFA stimulates endothelial cells in a different way, with clearly different effects on intracellular calcium mobilization, NO production, and IL-8 and ICAM-1 expression in primary BUVECs. These findings not only extend our understanding of NEFA-mediated diseases in ruminants, but also provide new insight into the different molecular mechanisms involved during endothelial cell activation by NEFAs.

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

    OpenAIRE

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

    2013-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2016-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

    Lacinová, L.; Weiss, Norbert

    2016-01-01

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

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

    Science.gov (United States)

    Doyle, Jefferson J; Doyle, Alexander J; Wilson, Nicole K; Habashi, Jennifer P; Bedja, Djahida; Whitworth, Ryan E; Lindsay, Mark E; Schoenhoff, Florian; Myers, Loretha; Huso, Nick; Bachir, Suha; Squires, Oliver; Rusholme, Benjamin; Ehsan, Hamid; Huso, David; Thomas, Craig J; Caulfield, Mark J; Van Eyk, Jennifer E; Judge, Daniel P; Dietz, Harry C

    2015-10-27

    Calcium channel blockers (CCBs) are prescribed to patients with Marfan syndrome for prophylaxis against aortic aneurysm progression, despite limited evidence for their efficacy and safety in the disorder. Unexpectedly, Marfan mice treated with CCBs show accelerated aneurysm expansion, rupture, and premature lethality. This effect is both extracellular signal-regulated kinase (ERK1/2) dependent and angiotensin-II type 1 receptor (AT1R) dependent. We have identified protein kinase C beta (PKCβ) as a critical mediator of this pathway and demonstrate that the PKCβ inhibitor enzastaurin, and the clinically available anti-hypertensive agent hydralazine, both normalize aortic growth in Marfan mice, in association with reduced PKCβ and ERK1/2 activation. Furthermore, patients with Marfan syndrome and other forms of inherited thoracic aortic aneurysm taking CCBs display increased risk of aortic dissection and need for aortic surgery, compared to patients on other antihypertensive agents.

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

    Science.gov (United States)

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

    2014-01-01

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

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

    Science.gov (United States)

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

    2017-12-21

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

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

    Directory of Open Access Journals (Sweden)

    Mandeep Kaur

    2011-01-01

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

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

    Science.gov (United States)

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

    2016-09-01

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

  13. The gene for the alpha 1 subunit of the skeletal muscle dihydropyridine-sensitive calcium channel (Cchl1a3) maps to mouse chromosome 1.

    Science.gov (United States)

    Chin, H; Krall, M; Kim, H L; Kozak, C A; Mock, B

    1992-12-01

    Cchl1a3 encodes the dihydropyridine-sensitive calcium channel alpha 1 subunit isoform predominantly expressed in skeletal muscle. mdg (muscular dysgenesis) has previously been implicated as a mutant allele of this gene. Hybridization of a rat brain cDNA probe for Cchl1a3 to Southern blots of DNAs from a panel of Chinese hamster x mouse somatic cell hybrids suggested that this gene maps to mouse Chromosome 1. Analysis of the progeny of an inbred strain cross-positioned Cchl1a3 1.3 cM proximal to the Pep-3 locus on Chr 1.

  14. STIM and Orai isoform expression in pregnant human myometrium: a potential role in calcium signaling during pregnancy.

    Directory of Open Access Journals (Sweden)

    Evonne eChin-Smith

    2014-05-01

    Full Text Available Store-operated calcium (Ca2+ entry (SOCE can be mediated by two novel proteins, STIM/Orai. We have previously demonstrated that members of the TRPC family, putative basal and store operated calcium entry channels, are present in human myometrium and regulated by labor associated stimuli IL-1β and mechanical stretch. Although STIM and Orai isoforms (1-3 have been reported in other smooth muscle cell types, there is little known about the expression or gestational regulation of STIM and Orai expression in human myometrium. Total RNA was isolated from lower segment human myometrial biopsies obtained at caesarean section from women at the time of preterm no labor (PTNL, preterm labor (PTL, term non-labor (TNL and term with labor (TL; primary cultured human uterine smooth muscle cells, and a human myometrial cell line (hTERT-HM. STIM1-2, and Orai1-3 mRNA expression was assessed by quantitative real-time PCR. All five genes were expressed in myometrial tissue and cultured cells. Orai2 was the most abundant Orai isoform in human myometrium. Expression of STIM1-2/Orai1-3 did not alter with the onset of labor. Orai1 mRNA expression in cultured cells was enhanced by IL-1β treatment. This novel report of STIM1-2 and Orai1-3 mRNA expression in pregnant human myometrium and Orai1 regulation by IL-1β indicates a potential role for these proteins in calcium signaling in human myometrium during pregnancy.

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

    DEFF Research Database (Denmark)

    Dalsgaard, Thomas; Kroigaard, Christel; Bek, Toke

    2009-01-01

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

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

    DEFF Research Database (Denmark)

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

    1992-01-01

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

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

    Science.gov (United States)

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

    2012-01-01

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

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

    NARCIS (Netherlands)

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

    2006-01-01

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

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

    Science.gov (United States)

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

    2003-06-01

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

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

    Science.gov (United States)

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

    2016-01-01

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

  1. Establishing homology between mitochondrial calcium uniporters, prokaryotic magnesium channels and chlamydial IncA proteins.

    Science.gov (United States)

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

    2014-08-01

    Mitochondrial calcium uniporters (MCUs) (TC no. 1.A.77) are oligomeric channel proteins found in the mitochondrial inner membrane. MCUs have two well-conserved transmembrane segments (TMSs), connected by a linker, similar to bacterial MCU homologues. These proteins and chlamydial IncA proteins (of unknown function; TC no. 9.B.159) are homologous to prokaryotic Mg(2+) transporters, AtpI and AtpZ, based on comparison scores of up to 14.5 sds. A phylogenetic tree containing all of these proteins showed that the AtpZ proteins cluster coherently as a subset within the large and diverse AtpI cluster, which branches separately from the MCUs and IncAs, both of which cluster coherently. The MCUs and AtpZs share the same two TMS topology, but the AtpIs have four TMSs, and IncAs can have either two (most frequent) or four (less frequent) TMSs. Binary alignments, comparison scores and motif analyses showed that TMSs 1 and 2 align with TMSs 3 and 4 of the AtpIs, suggesting that the four TMS AtpI proteins arose via an intragenic duplication event. These findings establish an evolutionary link interconnecting eukaryotic and prokaryotic Ca(2+) and Mg(2+) transporters with chlamydial IncAs, and lead us to suggest that all members of the MCU superfamily, including IncAs, function as divalent cation channels. © 2014 The Authors.

  2. Identification of cyclic nucleotide gated channels using regular expressions

    KAUST Repository

    Zelman, Alice K.

    2013-09-03

    Cyclic nucleotide-gated channels (CNGCs) are nonselective cation channels found in plants, animals, and some bacteria. They have a six-transmembrane/one- pore structure, a cytosolic cyclic nucleotide-binding domain, and a cytosolic calmodulin-binding domain. Despite their functional similarities, the plant CNGC family members appear to have different conserved amino acid motifs within corresponding functional domains than animal and bacterial CNGCs do. Here we describe the development and application of methods employing plant CNGC-specific sequence motifs as diagnostic tools to identify novel candidate channels in different plants. These methods are used to evaluate the validity of annotations of putative orthologs of CNGCs from plant genomes. The methods detail how to employ regular expressions of conserved amino acids in functional domains of annotated CNGCs and together with Web tools such as PHI-BLAST and ScanProsite to identify novel candidate CNGCs in species including Physcomitrella patens. © Springer Science+Business Media New York 2013.

  3. Decreased expression of Kv7 channels in Hirchsprung's disease.

    Science.gov (United States)

    O'Donnell, Anne-Marie; Coyle, David; Puri, Prem

    2017-07-01

    Voltage-dependent K + channels (Kv channels) participate in electrical rhythmicity and smooth muscle responses and are regulated by excitatory and inhibitory neurotransmitters. Kv channels also participate in the interstitial cell of Cajal (ICC) and smooth muscle cell (SMC) responses to neural inputs. The Kv family consists of 12 subfamilies, Kv1-Kv12, with five members of the Kv7 family identified to date: Kv7.1-Kv7.5. A recent study identified the potassium channel Kv7.5 as having a role in the excitability of ICC-IM in the mouse colon. We therefore designed this study to test the hypothesis that Kv7 channels are present in the normal human colon and are reduced in Hirschprung's disease (HSCR). HSCR tissue specimens were collected at the time of pull-through surgery (n=10), while normal control tissue specimens were obtained at the time of colostomy closure in patients with imperforate anus (n=10). Kv7.3-Kv7.5 immunohistochemistry was performed and visualized using confocal microscopy to assess their distribution. Western blot analysis was undertaken to determine Kv7.3-Kv7.5 protein quantification. Kv7.3 and Kv7.4-immunoreactivity was co-localized with neuron and ICC markers, while Kv7.5 was found to be expressed on both ICCs and SMCs. Western blot analysis revealed similar levels of Kv7.3 and Kv7.5 expression in the normal colon and HSCR colon, while Kv7.4 proteins were found to be markedly decreased in ganglionic specimens and decreased further in aganglionic specimens. A deficiency of Kv7.4 channels in the ganglionic and aganglionic bowel may place a role in colonic dysmotility in HSCR. Copyright © 2017 Elsevier Inc. All rights reserved.

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

    Directory of Open Access Journals (Sweden)

    Linn S Strandberg

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

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

    Science.gov (United States)

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

    2006-10-01

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

  6. NMDA receptors in mouse anterior piriform cortex initialize early odor preference learning and L-type calcium channels engage for long-term memory.

    Science.gov (United States)

    Mukherjee, Bandhan; Yuan, Qi

    2016-10-14

    The interactions of L-type calcium channels (LTCCs) and NMDA receptors (NMDARs) in memories are poorly understood. Here we investigated the specific roles of anterior piriform cortex (aPC) LTCCs and NMDARs in early odor preference memory in mice. Using calcium imaging in aPC slices, LTCC activation was shown to be dependent on NMDAR activation. Either D-APV (NMDAR antagonist) or nifedipine (LTCC antagonist) reduced somatic calcium transients in pyramidal cells evoked by lateral olfactory tract stimulation. However, nifedipine did not further reduce calcium in the presence of D-APV. In mice that underwent early odor preference training, blocking NMDARs in the aPC prevented short-term (3 hr) and long-term (24 hr) odor preference memory, and both memories were rescued when BayK-8644 (LTCC agonist) was co-infused. However, activating LTCCs in the absence of NMDARs resulted in loss of discrimination between the conditioned odor and a similar odor mixture at 3 hr. Elevated synaptic AMPAR expression at 3 hr was prevented by D-APV infusion but restored when LTCCs were directly activated, mirroring the behavioral outcomes. Blocking LTCCs prevented 24 hr memory and spared 3 hr memory. These results suggest that NMDARs mediate stimulus-specific encoding of odor memory while LTCCs mediate intracellular signaling leading to long-term memory.

  7. Voltage-gated proton channel is expressed on phagosomes

    International Nuclear Information System (INIS)

    Okochi, Yoshifumi; Sasaki, Mari; Iwasaki, Hirohide; Okamura, Yasushi

    2009-01-01

    Voltage-gated proton channel has been suggested to help NADPH oxidase activity during respiratory burst of phagocytes through its activities of compensating charge imbalance and regulation of pH. In phagocytes, robust production of reactive oxygen species occurs in closed membrane compartments, which are called phagosomes. However, direct evidence for the presence of voltage-gated proton channels in phagosome has been lacking. In this study, the expression of voltage-gated proton channels was studied by Western blot with the antibody specific to the voltage-sensor domain protein, VSOP/Hv1, that has recently been identified as the molecular correlate for the voltage-gated proton channel. Phagosomal membranes of neutrophils contain VSOP/Hv1 in accordance with subunits of NADPH oxidases, gp91, p22, p47 and p67. Superoxide anion production upon PMA activation was significantly reduced in neutrophils from VSOP/Hv1 knockout mice. These are consistent with the idea that voltage-gated proton channels help NADPH oxidase in phagocytes to produce reactive oxygen species.

  8. Does Increased Expression of the Plasma Membrane Calcium-ATPase Isoform 2 Confer Resistance to Apoptosis on Breast Cancer Cells?

    National Research Council Canada - National Science Library

    VanHouten, Joshua N

    2008-01-01

    The plasma membrane calcium ATPase isoform 2 (PMCA2) is highly expressed on the apical membrane of mammary epithelial cells during lactation, and is the predominant pump responsible for calcium transport into milk...

  9. Enhanced expression of a calcium-dependent protein kinase from ...

    Indian Academy of Sciences (India)

    Unknown

    low calcium medium; LNM, low nitrate medium; LPM, low phosphate medium; LSM, low sulphate medium; MMG, minimal medium with glucose; NR, nitrate reductase; ORF, open reading frame; PCR, polymerase chain reaction; SnRK, sucrose non fer- menting ..... amino acids in contrast to the usual number of 31 in other.

  10. Nuclear calcium signaling induces expression of the synaptic organizers Lrrtm1 and Lrrtm2.

    Science.gov (United States)

    Hayer, Stefanie N; Bading, Hilmar

    2015-02-27

    Calcium transients in the cell nucleus evoked by synaptic activity in hippocampal neurons function as a signaling end point in synapse-to-nucleus communication. As an important regulator of neuronal gene expression, nuclear calcium is involved in the conversion of synaptic stimuli into functional and structural changes of neurons. Here we identify two synaptic organizers, Lrrtm1 and Lrrtm2, as targets of nuclear calcium signaling. Expression of both Lrrtm1 and Lrrtm2 increased in a synaptic NMDA receptor- and nuclear calcium-dependent manner in hippocampal neurons within 2-4 h after the induction of action potential bursting. Induction of Lrrtm1 and Lrrtm2 occurred independently of the need for new protein synthesis and required calcium/calmodulin-dependent protein kinases and the nuclear calcium signaling target CREB-binding protein. Analysis of reporter gene constructs revealed a functional cAMP response element in the proximal promoter of Lrrtm2, indicating that at least Lrrtm2 is regulated by the classical nuclear Ca(2+)/calmodulin-dependent protein kinase IV-CREB/CREB-binding protein pathway. These results suggest that one mechanism by which nuclear calcium signaling controls neuronal network function is by regulating the expression of Lrrtm1 and Lrrtm2. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Nuclear Calcium Signaling Induces Expression of the Synaptic Organizers Lrrtm1 and Lrrtm2*

    Science.gov (United States)

    Hayer, Stefanie N.; Bading, Hilmar

    2015-01-01

    Calcium transients in the cell nucleus evoked by synaptic activity in hippocampal neurons function as a signaling end point in synapse-to-nucleus communication. As an important regulator of neuronal gene expression, nuclear calcium is involved in the conversion of synaptic stimuli into functional and structural changes of neurons. Here we identify two synaptic organizers, Lrrtm1 and Lrrtm2, as targets of nuclear calcium signaling. Expression of both Lrrtm1 and Lrrtm2 increased in a synaptic NMDA receptor- and nuclear calcium-dependent manner in hippocampal neurons within 2–4 h after the induction of action potential bursting. Induction of Lrrtm1 and Lrrtm2 occurred independently of the need for new protein synthesis and required calcium/calmodulin-dependent protein kinases and the nuclear calcium signaling target CREB-binding protein. Analysis of reporter gene constructs revealed a functional cAMP response element in the proximal promoter of Lrrtm2, indicating that at least Lrrtm2 is regulated by the classical nuclear Ca2+/calmodulin-dependent protein kinase IV-CREB/CREB-binding protein pathway. These results suggest that one mechanism by which nuclear calcium signaling controls neuronal network function is by regulating the expression of Lrrtm1 and Lrrtm2. PMID:25527504

  12. Eukaryotic translation initiation factor 3 subunit e controls intracellular calcium homeostasis by regulation of cav1.2 surface expression.

    Directory of Open Access Journals (Sweden)

    Pawel Buda

    Full Text Available Inappropriate surface expression of voltage-gated Ca(2+channels (CaV in pancreatic ß-cells may contribute to the development of type 2 diabetes. First, failure to increase intracellular Ca(2+ concentrations at the sites of exocytosis impedes insulin release. Furthermore, excessive Ca(2+ influx may trigger cytotoxic effects. The regulation of surface expression of CaV channels in the pancreatic β-cells remains unknown. Here, we used real-time 3D confocal and TIRFM imaging, immunocytochemistry, cellular fractionation, immunoprecipitation and electrophysiology to study trafficking of L-type CaV1.2 channels upon β-cell stimulation. We found decreased surface expression of CaV1.2 and a corresponding reduction in L-type whole-cell Ca(2+ currents in insulin-secreting INS-1 832/13 cells upon protracted (15-30 min stimulation. This internalization occurs by clathrin-dependent endocytosis and could be prevented by microtubule or dynamin inhibitors. eIF3e (Eukaryotic translation initiation factor 3 subunit E is part of the protein translation initiation complex, but its effect on translation are modest and effects in ion channel trafficking have been suggested. The factor interacted with CaV1.2 and regulated CaV1.2 traffic bidirectionally. eIF3e silencing impaired CaV1.2 internalization, which resulted in an increased intracellular Ca(2+ load upon stimulation. These findings provide a mechanism for regulation of L-type CaV channel surface expression with consequences for β-cell calcium homeostasis, which will affect pancreatic β-cell function and insulin production.

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

    Directory of Open Access Journals (Sweden)

    Gerhard P. Dahl

    2016-08-01

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

  14. Opposing Roles of Calcium and Intracellular ATP on Gating of the Purinergic P2X2 Receptor Channel

    Directory of Open Access Journals (Sweden)

    Milos B. Rokic

    2018-04-01

    Full Text Available P2X2 receptors (P2X2R exhibit a slow desensitization during the initial ATP application and a progressive, calcium-dependent increase in rates of desensitization during repetitive stimulation. This pattern is observed in whole-cell recordings from cells expressing recombinant and native P2X2R. However, desensitization is not observed in perforated-patched cells and in two-electrode voltage clamped oocytes. Addition of ATP, but not ATPγS or GTP, in the pipette solution also abolishes progressive desensitization, whereas intracellular injection of apyrase facilitates receptor desensitization. Experiments with injection of alkaline phosphatase or addition of staurosporine and ATP in the intracellular solution suggest a role for a phosphorylation-dephosphorylation in receptor desensitization. Mutation of residues that are potential phosphorylation sites identified a critical role of the S363 residue in the intracellular ATP action. These findings indicate that intracellular calcium and ATP have opposing effects on P2X2R gating: calcium allosterically facilitates receptor desensitization and ATP covalently prevents the action of calcium. Single cell measurements further revealed that intracellular calcium stays elevated after washout in P2X2R-expressing cells and the blockade of mitochondrial sodium/calcium exchanger lowers calcium concentrations during washout periods to basal levels, suggesting a role of mitochondria in this process. Therefore, the metabolic state of the cell can influence P2X2R gating.

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

    Science.gov (United States)

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

    2017-12-01

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

  16. Hyperthyroidism enhances 5-HT-induced contraction of the rat pulmonary artery: role of calcium-activated chloride channel activation.

    Science.gov (United States)

    Oriowo, Mabayoje A; Oommen, Elsie; Khan, Islam

    2011-11-01

    Experimentally-induced hyperthyroidism in rodents is associated with signs and symptoms of pulmonary hypertension. The main objective of the present study was to investigate the effect of thyroxine-induced pulmonary hypertension on the contractile response of the pulmonary artery to 5-HT and the possible underlying signaling pathway. 5-HT concentration-dependently contracted artery segments from control and thyroxine-treated rats with pD(2) values of 5.04 ± 0.19 and 5.34 ± 0.14, respectively. The maximum response was significantly greater in artery segments from thyroxine-treated rats. Neither BW 723C86 (5-HT(2B)-receptor agonist) nor CP 93129 (5-HT(1B)-receptor agonist) contracted ring segments of the pulmonary artery from control and thyroxine-treated rats at concentrations up to 10(-4)M. There was no significant difference in the level of expression of 5-HT(2A)-receptor protein between the two groups. Ketanserin (3 × 10(-8)M) produced a rightward shift of the concentration-response curve to 5-HT in both groups with equal potency (-logK(B) values were 8.1 ± 0.2 and 7.9 ± 0.1 in control and thyroxine-treated rats, respectively). Nifedipine (10(-6)M) inhibited 5-HT-induced contractions in artery segments from control and thyroxine-treated rats and was more effective against 5-HT-induced contraction in artery segments for thyroxine-treated rats. The calcium-activated chloride channel blocker, niflumic acid (10(-4)M) also inhibited 5-HT-induced contractions in artery segments from control and thyroxine-treated rats and was more effective against 5-HT-induced contraction in artery segments for thyroxine-treated rats. It was concluded that hyperthyroidism enhanced 5-HT-induced contractions of the rat pulmonary artery by a mechanism involving increased activity of calcium-activated chloride channels. Copyright © 2011 Elsevier B.V. All rights reserved.

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

    DEFF Research Database (Denmark)

    Kurtz, A; Skott, O; Chegini, S

    1990-01-01

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

  18. Endoplasmic reticulum calcium transport ATPase expression during differentiation of colon cancer and leukaemia cells

    International Nuclear Information System (INIS)

    Papp, Bela; Brouland, Jean-Philippe; Gelebart, Pascal; Kovacs, Tuende; Chomienne, Christine

    2004-01-01

    The calcium homeostasis of the endoplasmic reticulum (ER) is connected to a multitude of cell functions involved in intracellular signal transduction, control of proliferation, programmed cell death, or the synthesis of mature proteins. Calcium is accumulated in the ER by various biochemically distinct sarco/endoplasmic reticulum calcium transport ATPase isoenzymes (SERCA isoforms). Experimental data indicate that the SERCA composition of some carcinoma and leukaemia cell types undergoes significant changes during differentiation, and that this is accompanied by modifications of SERCA-dependent calcium accumulation in the ER. Because ER calcium homeostasis can also influence cell differentiation, we propose that the modulation of the expression of various SERCA isoforms, and in particular, the induction of the expression of SERCA3-type proteins, is an integral part of the differentiation program of some cancer and leukaemia cell types. The SERCA content of the ER may constitute a new parameter by which the calcium homeostatic characteristics of the organelle are adjusted. The cross-talk between ER calcium homeostasis and cell differentiation may have some implications for the better understanding of the signalling defects involved in the acquisition and maintenance of the malignant phenotype

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

    Science.gov (United States)

    Minor, Daniel L; Findeisen, Felix

    2010-01-01

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

  20. Sculpting ion channel functional expression with engineered ubiquitin ligases

    Science.gov (United States)

    Kanner, Scott A; Morgenstern, Travis

    2017-01-01

    The functional repertoire of surface ion channels is sustained by dynamic processes of trafficking, sorting, and degradation. Dysregulation of these processes underlies diverse ion channelopathies including cardiac arrhythmias and cystic fibrosis. Ubiquitination powerfully regulates multiple steps in the channel lifecycle, yet basic mechanistic understanding is confounded by promiscuity among E3 ligase/substrate interactions and ubiquitin code complexity. Here we targeted the catalytic domain of E3 ligase, CHIP, to YFP-tagged KCNQ1 ± KCNE1 subunits with a GFP-nanobody to selectively manipulate this channel complex in heterologous cells and adult rat cardiomyocytes. Engineered CHIP enhanced KCNQ1 ubiquitination, eliminated KCNQ1 surface-density, and abolished reconstituted K+ currents without affecting protein expression. A chemo-genetic variation enabling chemical control of ubiquitination revealed KCNQ1 surface-density declined with a ~ 3.5 hr t1/2 by impaired forward trafficking. The results illustrate utility of engineered E3 ligases to elucidate mechanisms underlying ubiquitin regulation of membrane proteins, and to achieve effective post-translational functional knockdown of ion channels. PMID:29256394

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

    Science.gov (United States)

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

    2018-05-01

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

  2. Calcium Homeostasis Modulator 1-Like Currents in Rat Fungiform Taste Cells Expressing Amiloride-Sensitive Sodium Currents.

    Science.gov (United States)

    Bigiani, Albertino

    2017-05-01

    Salt reception by taste cells is still the less understood transduction process occurring in taste buds, the peripheral sensory organs for the detection of food chemicals. Although there is evidence suggesting that the epithelial sodium channel (ENaC) works as sodium receptor, yet it is not clear how salt-detecting cells signal the relevant information to nerve endings. Taste cells responding to sweet, bitter, and umami substances release ATP as neurotransmitter through a nonvesicular mechanism. Three different channel proteins have been proposed as conduit for ATP secretion: pannexin channels, connexin hemichannels, and calcium homeostasis modulator 1 (CALHM1) channels. In heterologous expression systems, these channels mediate outwardly rectifying membrane currents with distinct biophysical and pharmacological properties. I therefore tested whether also salt-detecting taste cells were endowed with these currents. To this aim, I applied the patch-clamp techniques to single cells in isolated taste buds from rat fungiform papillae. Salt-detecting cells were functionally identified by exploiting the effect of amiloride, which induces a current response by shutting down ENaCs. I looked for the presence of outwardly rectifying currents by using appropriate voltage-clamp protocols and specific pharmacological tools. I found that indeed salt-detecting cells possessed these currents with properties consistent with the presence, at least in part, of CALHM1 channels. Unexpectedly, CALHM1-like currents in taste cells were potentiated by known blockers of pannexin, suggesting a possible inhibitory action of this protein on CALMH1. These findings indicate that communication between salt-detecting cells and nerve endings might involve ATP release by CALMH1 channels. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  3. Interactions of waterborne and dietary cadmium on the expression of calcium transporters in the gills of rainbow trout: Influence of dietary calcium supplementation

    Energy Technology Data Exchange (ETDEWEB)

    Galvez, Fernando [Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803 (United States); Department of Biology, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada)], E-mail: galvezf@lsu.edu; Franklin, Natasha M. [Department of Biology, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada); Tuttle, Ryan B. [Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803 (United States); Wood, Chris M. [Department of Biology, McMaster University, Hamilton, Ontario, L8S 4K1 (Canada)

    2007-08-30

    Recent studies have shown that dietary Ca{sup 2+} supplementation strongly inhibits uptake of Ca{sup 2+} and Cd at the fish gill. To better understand the influence of dietary Ca{sup 2+} on branchial Ca{sup 2+} transport, we examined the expression of two trout gill calcium transporters during waterborne and dietary Cd exposure, at two different levels of dietary Ca{sup 2+}. Quantitative polymerase chain reaction (PCR) was used to monitor epithelial calcium channel (ECaC) and sodium-calcium exchange (NCX) mRNA levels following 7-28 days of exposure to these treatments. In brief, juvenile rainbow trout (Oncorhynchus mykiss) were exposed to control, 3 {mu}g/L waterborne Cd, 500 mg/kg dietary Cd, or a combined 3 {mu}g/L waterborne plus 500 mg/kg dietary Cd exposure, supplemented with either 20 mg/g or 60 mg/g dietary calcium (Ca{sup 2+}). Two-way analysis of variance was used to discern the main effects of Cd exposure and dietary Ca{sup 2+} supplementation on ECaC and NCX mRNA levels. We found that dietary Ca{sup 2+} supplementation decreased significantly ECaC mRNA expression on days 14 and 21. In comparison, NCX mRNA levels were not influenced by dietary Ca{sup 2+} supplementation, but rather were significantly inhibited in the combined waterborne and dietary Cd exposure on day 7 alone. Statistical analysis found no interactive effects between Cd exposure and dietary Ca{sup 2+} exposure at any time point, except for day 28.This study provides evidence of the importance of nutritional status on the transcriptional regulation of ion transport at the fish gill. We discuss the importance of diet and nutritional status to the development of new regulatory approaches, such as the biotic ligand model, which currently do not account for the significance of diet on metal bioavailability in aquatic organisms.

  4. Antispasmodic and Antidiarrheal Activities of Valeriana hardwickii Wall. Rhizome Are Putatively Mediated through Calcium Channel Blockade.

    Science.gov (United States)

    Bashir, Samra; Memon, Raafia; Gilani, Anwar H

    2011-01-01

    Valeriana hardwickii is indigenous to Pakistan, Burma and Ceylon, where it is traditionally being used as an antispasmodic and antidiarrheal, besides its culinary use as spice. The aim of this paper was to provide pharmacological validation to these medicinal uses. The crude aqueous-methanolic extract of Valeriana hardwickii rhizome (Vh.Cr) was studied on isolated rabbit jejunum and castor oil-induced diarrhea in mice for spasmolytic and antidiarrheal properties, respectively. Vh.Cr caused concentration-dependent (0.01-1 mg/mL) relaxation of spontaneous contractions in isolated rabbit jejunum and inhibited K(+)-induced contractions (0.01-0.3 mg/mL), similar to verapamil, suggestive of calcium channel blockade (CCB). The CCB effect was confirmed when pretreatment of the jejunum preparations with Vh.Cr produced a concentration-dependent (0.03-0.1 mg/mL) rightward shift in the Ca(++) concentration-response curves, as caused by verapamil. Vh.Cr exhibited dose-dependent (100-300 mg/kg) protection against castor oil-induced diarrhea in mice. Loperamide, a standard antidiarrheal drug, similarly prevented the diarrhea. These data indicate the presence of CCB effect in the extract of Valeriana hardwickii rhizome, possibly mediating its antispasmodic and antidiarrheal activities and provide a scientific base for its traditional use in hyperactive gut disorders.

  5. Antispasmodic and Antidiarrheal Activities of Valeriana hardwickii Wall. Rhizome Are Putatively Mediated through Calcium Channel Blockade

    Directory of Open Access Journals (Sweden)

    Samra Bashir

    2011-01-01

    Full Text Available Valeriana hardwickii is indigenous to Pakistan, Burma and Ceylon, where it is traditionally being used as an antispasmodic and antidiarrheal, besides its culinary use as spice. The aim of this paper was to provide pharmacological validation to these medicinal uses. The crude aqueous-methanolic extract of Valeriana hardwickii rhizome (Vh.Cr was studied on isolated rabbit jejunum and castor oil-induced diarrhea in mice for spasmolytic and antidiarrheal properties, respectively. Vh.Cr caused concentration-dependent (0.01–1 mg/mL relaxation of spontaneous contractions in isolated rabbit jejunum and inhibited K+-induced contractions (0.01–0.3 mg/mL, similar to verapamil, suggestive of calcium channel blockade (CCB. The CCB effect was confirmed when pretreatment of the jejunum preparations with Vh.Cr produced a concentration-dependent (0.03–0.1 mg/mL rightward shift in the Ca++ concentration-response curves, as caused by verapamil. Vh.Cr exhibited dose-dependent (100–300 mg/kg protection against castor oil-induced diarrhea in mice. Loperamide, a standard antidiarrheal drug, similarly prevented the diarrhea. These data indicate the presence of CCB effect in the extract of Valeriana hardwickii rhizome, possibly mediating its antispasmodic and antidiarrheal activities and provide a scientific base for its traditional use in hyperactive gut disorders.

  6. Polyaniline-graphene oxide nanocomposite sensor for quantification of calcium channel blocker levamlodipine.

    Science.gov (United States)

    Jain, Rajeev; Sinha, Ankita; Khan, Ab Lateef

    2016-08-01

    A novel polyaniline-graphene oxide nanocomposite (PANI/GO/GCE) sensor has been fabricated for quantification of a calcium channel blocker drug levamlodipine (LAMP). Fabricated sensor has been characterized by electrochemical impedance spectroscopy, square wave and cyclic voltammetry, Raman spectroscopy and Fourier transform infrared (FTIR) spectroscopy. The developed PANI/GO/GCE sensor has excellent analytical performance towards electrocatalytic oxidation as compared to PANI/GCE, GO/GCE and bare GCE. Under optimized experimental conditions, the fabricated sensor exhibits a linear response for LAMP for its oxidation over a concentration range from 1.25μgmL(-1) to 13.25μgmL(-1) with correlation coefficient of 0.9950 (r(2)), detection limit of 1.07ngmL(-1) and quantification limit of 3.57ngmL(-1). The sensor shows an excellent performance for detecting LAMP with reproducibility of 2.78% relative standard deviation (RSD). The proposed method has been successfully applied for LAMP determination in pharmaceutical formulation with a recovery from 99.88% to 101.75%. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Regulation of the sperm calcium channel CatSper by endogenous steroids and plant triterpenoids.

    Science.gov (United States)

    Mannowetz, Nadja; Miller, Melissa R; Lishko, Polina V

    2017-05-30

    The calcium channel of sperm (CatSper) is essential for sperm hyperactivated motility and fertility. The steroid hormone progesterone activates CatSper of human sperm via binding to the serine hydrolase ABHD2. However, steroid specificity of ABHD2 has not been evaluated. Here, we explored whether steroid hormones to which human spermatozoa are exposed in the male and female genital tract influence CatSper activation via modulation of ABHD2. The results show that testosterone, estrogen, and hydrocortisone did not alter basal CatSper currents, whereas the neurosteroid pregnenolone sulfate exerted similar effects as progesterone, likely binding to the same site. However, physiological concentrations of testosterone and hydrocortisone inhibited CatSper activation by progesterone. Additionally, testosterone antagonized the effect of pregnenolone sulfate. We have also explored whether steroid-like molecules, such as the plant triterpenoids pristimerin and lupeol, affect sperm fertility. Interestingly, both compounds competed with progesterone and pregnenolone sulfate and significantly reduced CatSper activation by either steroid. Furthermore, pristimerin and lupeol considerably diminished hyperactivation of capacitated spermatozoa. These results indicate that ( i ) pregnenolone sulfate together with progesterone are the main steroids that activate CatSper and ( ii ) pristimerin and lupeol can act as contraceptive compounds by averting sperm hyperactivation, thus preventing fertilization.

  8. Mechanisms of the gabapentinoids and α 2 δ-1 calcium channel subunit in neuropathic pain.

    Science.gov (United States)

    Patel, Ryan; Dickenson, Anthony H

    2016-04-01

    The gabapentinoid drugs gabapentin and pregabalin are key front-line therapies for various neuropathies of peripheral and central origin. Originally designed as analogs of GABA, the gabapentinoids bind to the α 2 δ-1 and α 2 δ-2 auxiliary subunits of calcium channels, though only the former has been implicated in the development of neuropathy in animal models. Transgenic approaches also identify α 2 δ-1 as key in mediating the analgesic effects of gabapentinoids, however the precise molecular mechanisms remain unclear. Here we review the current understanding of the pathophysiological role of the α 2 δ-1 subunit, the mechanisms of analgesic action of gabapentinoid drugs and implications for efficacy in the clinic. Despite widespread use, the number needed to treat for gabapentin and pregabalin averages from 3 to 8 across neuropathies. The failure to treat large numbers of patients adequately necessitates a novel approach to treatment selection. Stratifying patients by sensory profiles may imply common underlying mechanisms, and a greater understanding of these mechanisms could lead to more direct targeting of gabapentinoids.

  9. Cross talk among calcium, hydrogen peroxide, and nitric oxide and activation of gene expression involving calmodulins and calcium-dependent protein kinases in Ulva compressa exposed to copper excess.

    Science.gov (United States)

    González, Alberto; Cabrera, M de Los Ángeles; Henríquez, M Josefa; Contreras, Rodrigo A; Morales, Bernardo; Moenne, Alejandra

    2012-03-01

    To analyze the copper-induced cross talk among calcium, nitric oxide (NO), and hydrogen peroxide (H(2)O(2)) and the calcium-dependent activation of gene expression, the marine alga Ulva compressa was treated with the inhibitors of calcium channels, ned-19, ryanodine, and xestospongin C, of chloroplasts and mitochondrial electron transport chains, 3-(3,4-dichlorophenyl)-1,1-dimethylurea and antimycin A, of pyruvate dehydrogenase, moniliformin, of calmodulins, N-(6-aminohexyl)-5-chloro-1-naphtalene sulfonamide, and of calcium-dependent protein kinases, staurosporine, as well as with the scavengers of NO, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, and of H(2)O(2), ascorbate, and exposed to a sublethal concentration of copper (10 μm) for 24 h. The level of NO increased at 2 and 12 h. The first peak was inhibited by ned-19 and 3-(2,3-dichlorophenyl)-1,1-dimethylurea and the second peak by ned-19 and antimycin A, indicating that NO synthesis is dependent on calcium release and occurs in organelles. The level of H(2)O(2) increased at 2, 3, and 12 h and was inhibited by ned-19, ryanodine, xestospongin C, and moniliformin, indicating that H(2)O(2) accumulation is dependent on calcium release and Krebs cycle activity. In addition, pyruvate dehydrogenase, 2-oxoxglutarate dehydrogenase, and isocitrate dehydrogenase activities of the Krebs cycle increased at 2, 3, 12, and/or 14 h, and these increases were inhibited in vitro by EGTA, a calcium chelating agent. Calcium release at 2, 3, and 12 h was inhibited by 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide and ascorbate, indicating activation by NO and H(2)O(2). In addition, the level of antioxidant protein gene transcripts decreased with N-(6-aminohexyl)-5-chloro-1-naphtalene sulfonamide and staurosporine. Thus, there is a copper-induced cross talk among calcium, H(2)O(2), and NO and a calcium-dependent activation of gene expression involving calmodulins and calcium-dependent protein

  10. Characterization of calcium signals in human induced pluripotent stem cell-derived dentate gyrus neuronal progenitors and mature neurons, stably expressing an advanced calcium indicator protein.

    Science.gov (United States)

    Vőfély, Gergő; Berecz, Tünde; Szabó, Eszter; Szebényi, Kornélia; Hathy, Edit; Orbán, Tamás I; Sarkadi, Balázs; Homolya, László; Marchetto, Maria C; Réthelyi, János M; Apáti, Ágota

    2018-04-01

    Pluripotent stem cell derived human neuronal progenitor cells (hPSC-NPCs) and their mature neuronal cell culture derivatives may efficiently be used for central nervous system (CNS) drug screening, including the investigation of ligand-induced calcium signalization. We have established hippocampal NPC cultures derived from human induced PSCs, which were previously generated by non-integrating Sendai virus reprogramming. Using established protocols these NPCs were differentiated into hippocampal dentate gyrus neurons. In order to study calcium signaling without the need of dye loading, we have stably expressed an advanced calcium indicator protein (GCaMP6fast) in the NPCs using the Sleeping Beauty transposon system. We observed no significant effects of the long-term GCaMP6 expression on NPC morphology, gene expression pattern or neural differentiation capacity. In order to compare the functional properties of GCaMP6-expressing neural cells and the corresponding parental cells loaded with calcium indicator dye Fluo-4, a detailed characterization of calcium signals was performed. We found that the calcium signals induced by ATP, glutamate, LPA, or proteases - were similar in these two systems. Moreover, the presence of the calcium indicator protein allowed for a sensitive, repeatable detection of changes in calcium signaling during the process of neurogenesis and neuronal maturation. Copyright © 2018 Elsevier Inc. All rights reserved.

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

    DEFF Research Database (Denmark)

    Stankevicius, Edgaras; Dalsgaard, Thomas; Kroigaard, Christel

    2011-01-01

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

  12. Novel CLCNKB mutations causing Bartter syndrome affect channel surface expression.

    Science.gov (United States)

    Keck, Mathilde; Andrini, Olga; Lahuna, Olivier; Burgos, Johanna; Cid, L Pablo; Sepúlveda, Francisco V; L'hoste, Sébastien; Blanchard, Anne; Vargas-Poussou, Rosa; Lourdel, Stéphane; Teulon, Jacques

    2013-09-01

    Mutations in the CLCNKB gene encoding the ClC-Kb Cl(-) channel cause Bartter syndrome, which is a salt-losing renal tubulopathy. Here, we investigate the functional consequences of seven mutations. When expressed in Xenopus laevis oocytes, four mutants carried no current (c.736G>C, p.Gly246Arg; c.1271G>A, p.Gly424Glu; c.1313G>A, p.Arg438His; c.1316T>C, p.Leu439Pro), whereas others displayed a 30%-60% reduction in conductance as compared with wild-type ClC-Kb (c.242T>C, p.Leu81Pro; c.274C>T, p.Arg92Trp; c.1052G>C, p.Arg351Pro). Anion selectivity and sensitivity to external Ca(2+) and H(+), typical of the ClC-Kb channel, were not modified in the partially active mutants. In oocytes, we found that all the mutations reduced surface expression with a profile similar to that observed for currents. In HEK293 cells, the currents in the mutants had similar profiles to those obtained in oocytes, except for p.Leu81Pro, which produced no current. Furthermore, p.Arg92Trp and p.Arg351Pro mutations did not modify the unit-conductance of closely related ClC-K1. Western blot analysis in HEK293 cells showed that ClC-Kb protein abundance was lower for the nonconducting mutants but similar to wild-type for other mutants. Overall, two classes of mutants can be distinguished: nonconducting mutants associated with low total protein expression, and partially conducting mutants with unaltered channel properties and ClC-Kb protein abundance. © 2013 WILEY PERIODICALS, INC.

  13. The effects of calcium channel blockade on agouti-induced obesity

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jung Han; Moustaid, N.; Zemel, M.B. [Univ. of Tennessee, Knoxville, TN (United States)] [and others

    1996-12-01

    We have previously observed that obese viable yellow (A{sup vy}/a) mice exhibit increased intracellular Ca{sup 2+} ([Ca{sup 2+}]i) and fatty acid synthase (FAS) gene expression; further, recombinant agouti protein increases in cultured adipocytes and these effects are inhibited by Ca{sup 2+} channel blockade. Accordingly, we determined the effect of Ca{sup 2+} channel blockade (nifedipine for 4 wk) on FAS and obesity in transgenic mice expressing the agouti gene in a ubiquitous manner. The transgenic mice initially were significantly heavier (30.5 {+-} 0.6 vs. 27.3 {+-} 0.3 g; P<0.001) and exhibited a 0.81{degrees}C lower initial core temperature (P<0.0005), an approximately twofold increase in fat pad weights (P=0.002), a sevenfold increase in adipose FAS activity (P=0.009), and a twofold increase in plasma insulin level (P<0.05) compared to control mice. Nifedipine treatment resulted in an 18% decrease in fat pad weights (P<0.007) and a 74% decrease in adipose FAS activity (P=0.03), normalized circulating insulin levels and insulin sensitivity (P,0.05), and transiently elevated core temperature in the transgenic mice, but was without effect in the control mice. These data suggest that agouti regulates FAS, fat storage, and possibly thermogenesis, at least partially, via a [Ca{sup 2+}]{sub i}-dependent mechanism, and that Ca{sup 2+} channel blockade may partially attenuate agouti-induced obesity. 42 refs., 4 figs., 1 tab.

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

    Science.gov (United States)

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

    2013-03-01

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

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

    OpenAIRE

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

    2008-01-01

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

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

    OpenAIRE

    Gulmez, Oyku

    2008-01-01

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

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

    OpenAIRE

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

    2008-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2006-01-01

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

  19. Expression profiling of colorectal cancer cells reveals inhibition of DNA replication licensing by extracellular calcium.

    Science.gov (United States)

    Aggarwal, Abhishek; Schulz, Herbert; Manhardt, Teresa; Bilban, Martin; Thakker, Rajesh V; Kallay, Enikö

    2017-06-01

    Colorectal cancer is one of the most common cancers in industrialised societies. Epidemiological studies, animal experiments, and randomized clinical trials have shown that dietary factors can influence all stages of colorectal carcinogenesis, from initiation through promotion to progression. Calcium is one of the factors with a chemoprophylactic effect in colorectal cancer. The aim of this study was to understand the molecular mechanisms of the anti-tumorigenic effects of extracellular calcium ([Ca 2+ ] o ) in colon cancer cells. Gene expression microarray analysis of colon cancer cells treated for 1, 4, and 24h with 2mM [Ca 2+ ] o identified significant changes in expression of 1571 probe sets (ANOVA, pcalcium-sensing receptor (CaSR), a G protein-coupled receptor is a mediator involved in this process. To test whether these results were physiologically relevant, we fed mice with a standard diet containing low (0.04%), intermediate (0.1%), or high (0.9%) levels of dietary calcium. The main molecules regulating replication licensing were inhibited also in vivo, in the colon of mice fed high calcium diet. We show that among the mechanisms behind the chemopreventive effect of [Ca 2+ ] o is inhibition of replication licensing, a process often deregulated in neoplastic transformation. Our data suggest that dietary calcium is effective in preventing replicative stress, one of the main drivers of cancer and this process is mediated by the calcium-sensing receptor. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

    Neely, Alan; Hidalgo, Patricia

    2014-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Alan eNeely

    2014-06-01

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

  2. Zinc oxide nanoparticles decrease the expression and activity of plasma membrane calcium ATPase, disrupt the intracellular calcium homeostasis in rat retinal ganglion cells.

    Science.gov (United States)

    Guo, Dadong; Bi, Hongsheng; Wang, Daoguang; Wu, Qiuxin

    2013-08-01

    Zinc oxide nanoparticle is one of the most important materials with diverse applications. However, it has been reported that zinc oxide nanoparticles are toxic to organisms, and that oxidative stress is often hypothesized to be an important factor in cytotoxicity mediated by zinc oxide nanoparticles. Nevertheless, the mechanism of toxicity of zinc oxide nanoparticles has not been completely understood. In this study, we investigated the cytotoxic effect of zinc oxide nanoparticles and the possible molecular mechanism involved in calcium homeostasis mediated by plasma membrane calcium ATPase in rat retinal ganglion cells. Real-time cell electronic sensing assay showed that zinc oxide nanoparticles could exert cytotoxic effect on rat retinal ganglion cells in a concentration-dependent manner; flow cytometric analysis indicated that zinc oxide nanoparticles could lead to cell damage by inducing the overproduction of reactive oxygen species. Furthermore, zinc oxide nanoparticles could also apparently decrease the expression level and their activity of plasma membrane calcium ATPase, which finally disrupt the intracellular calcium homeostasis and result in cell death. Taken together, zinc oxide nanoparticles could apparently decrease the plasma membrane calcium ATPase expression, inhibit their activity, cause the elevated intracellular calcium ion level and disrupt the intracellular calcium homeostasis. Further, the disrupted calcium homeostasis will trigger mitochondrial dysfunction, generate excessive reactive oxygen species, and finally initiate cell death. Thus, the disrupted calcium homeostasis is involved in the zinc oxide nanoparticle-induced rat retinal ganglion cell death. Copyright © 2013 Elsevier Ltd. All rights reserved.

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

    Science.gov (United States)

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

    2000-06-01

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

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

    Directory of Open Access Journals (Sweden)

    Andreas Totzeck

    2016-09-01

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

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

    Science.gov (United States)

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

    1990-09-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2017-01-01

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

  7. Mechanisms underlying the cardiac pacemaker: the role of SK4 calcium-activated potassium channels.

    Science.gov (United States)

    Weisbrod, David; Khun, Shiraz Haron; Bueno, Hanna; Peretz, Asher; Attali, Bernard

    2016-01-01

    The proper expression and function of the cardiac pacemaker is a critical feature of heart physiology. The sinoatrial node (SAN) in human right atrium generates an electrical stimulation approximately 70 times per minute, which propagates from a conductive network to the myocardium leading to chamber contractions during the systoles. Although the SAN and other nodal conductive structures were identified more than a century ago, the mechanisms involved in the generation of cardiac automaticity remain highly debated. In this short review, we survey the current data related to the development of the human cardiac conduction system and the various mechanisms that have been proposed to underlie the pacemaker activity. We also present the human embryonic stem cell-derived cardiomyocyte system, which is used as a model for studying the pacemaker. Finally, we describe our latest characterization of the previously unrecognized role of the SK4 Ca(2+)-activated K(+) channel conductance in pacemaker cells. By exquisitely balancing the inward currents during the diastolic depolarization, the SK4 channels appear to play a crucial role in human cardiac automaticity.

  8. The overview of channels, transporters, and calcium signaling molecules during amelogenesis.

    Science.gov (United States)

    Kim, Hee-Eun; Hong, Jeong Hee

    2018-05-20

    Enamel is a highly calcified tissue. Its formation requires a progressive and dynamic system for the regulation of electrolyte concentration by enamel epithelia. A critical function of enamel epithelial cells, ameloblasts, is the secretion and movement of electrolytes via various channels and transporters to develop the enamel tissue. Enamel formation generates protons, which need to be neutralised. Thus, ameloblasts possess a buffering system to sustain mineral accretion. Normal tooth formation involves stage-dependent net fluctuations in pH during amelogenesis. To date, all of our information about ion transporters in dental enamel tissue is based solely on immunostaining-expression techniques. This review critically evaluates the current understanding and recent discoveries and physiological role of ion channels and transporters, Mg 2+ transporters, and Ca 2+ regulatory proteins during amelogenesis in enamel formation. The ways in which ameloblasts modulate ions are discussed in the context of current research for developing a novel morphologic-functional model of enamel maturation. Copyright © 2018 Elsevier Ltd. All rights reserved.

  9. Troponin T3 regulates nuclear localization of the calcium channel Cavβ1a subunit in skeletal muscle

    International Nuclear Information System (INIS)

    Zhang, Tan; Taylor, Jackson; Jiang, Yang; Pereyra, Andrea S.; Messi, Maria Laura; Wang, Zhong-Min; Hereñú, Claudia; Delbono, Osvaldo

    2015-01-01

    The voltage-gated calcium channel (Ca v ) β 1a subunit (Ca v β 1a ) plays an important role in excitation–contraction coupling (ECC), a process in the myoplasm that leads to muscle-force generation. Recently, we discovered that the Ca v β 1a subunit travels to the nucleus of skeletal muscle cells where it helps to regulate gene transcription. To determine how it travels to the nucleus, we performed a yeast two-hybrid screening of the mouse fast skeletal muscle cDNA library and identified an interaction with troponin T3 (TnT3), which we subsequently confirmed by co-immunoprecipitation and co-localization assays in mouse skeletal muscle in vivo and in cultured C2C12 muscle cells. Interacting domains were mapped to the leucine zipper domain in TnT3 COOH-terminus (160–244 aa) and Ca v β 1a NH 2 -terminus (1–99 aa), respectively. The double fluorescence assay in C2C12 cells co-expressing TnT3/DsRed and Ca v β 1a /YFP shows that TnT3 facilitates Ca v β 1a nuclear recruitment, suggesting that the two proteins play a heretofore unknown role during early muscle differentiation in addition to their classical role in ECC regulation. - Highlights: • Previously, we demonstrated that Ca v β 1a is a gene transcription regulator. • Here, we show that TnT3 interacts with Ca v β 1a . • We mapped TnT3 and Ca v β 1a interaction domain. • TnT3 facilitates Ca v β 1a nuclear enrichment. • The two proteins play a heretofore unknown role during early muscle differentiation

  10. MANAGEMENT OF HYPERTENSION- INSIGHTS INTO REAL-WORLD CLINICAL PRACTICE FOR DIFFERENTIAL USAGE OF CALCIUM CHANNEL BLOCKERS (CCBS

    Directory of Open Access Journals (Sweden)

    Arup Dasbiswas

    2017-05-01

    Full Text Available BACKGROUND Calcium channel blockers (CCB like amlodipine, S (- amlodipine and cilnidipine, etc. have established place in the treatment of hypertension (HTN. As perceived by most of the physicians, they have comparative antihypertensive efficacy. However, available evidences suggest varied differences in incidence of pedal oedema. Aim- This survey was planned to understand real-world clinical practice pattern of Indian physicians for usage of various antihypertensive agents with emphasis on CCBs and whether differential incidence of oedema with CCBs is encountered in their clinical practice. MATERIALS AND METHODS Survey questionnaire consisting of 10 questions about preferred antihypertensive choice for different subsets of patients with HTN and efficacy and safety of S (- amlodipine was prepared and validated in small group of physicians. Overall, 494 general physicians and cardiologists practising in India were approached for seeking their opinion on usage of various CCBs. Statistical Analysis- Data were expressed in percentage. Design- Prospective, cross sectional, questionnaire-based survey. RESULTS Amongst various anti-hypertensive agents, majority of the physicians preferred CCB as their initial drug of choice for patients with HTN (53.8%, HTN with CKD (41.1%, elderly (55.3%, and young (30.8% patients. Though amlodipine was preferred by 75.7% physicians, pedal oedema was observed in >10% patients by 40.5% physicians. Most of the physicians rated S (- amlodipine to have better efficacy (79.4% and safety profile (88.3% with decreased incidence of pedal oedema than racemic Amlodipine. CONCLUSION Available evidences suggest comparative efficacy of S (- amlodipine and racemic amlodipine with varied differences in incidence of pedal oedema. However, our survey suggests better efficacy and safety of S (- amlodipine over racemic amlodipine as opined by most of the physicians of India. The survey findings need to be further evaluated in randomised

  11. L-type calcium channel CaV 1.2 in transgenic mice overexpressing human AbetaPP751 with the London (V717I) and Swedish (K670M/N671L) mutations.

    Science.gov (United States)

    Willis, Michael; Kaufmann, Walter A; Wietzorrek, Georg; Hutter-Paier, Birgit; Moosmang, Sven; Humpel, Christian; Hofmann, Franz; Windisch, Manfred; Knaus, Hans-Günther; Marksteiner, Josef

    2010-01-01

    Cumulative evidence indicates that amyloid-beta peptides exert some of their neurodegenerative effects through modulation of L-type voltage gated calcium channels, which play key roles in a diverse range of CNS functions. In this study we examined the expression of CaV1.2 L-type voltage gated calcium channels in transgenic mice overexpressing human AbetaPP751 with the London (V717I) and Swedish (K670M/N671L) mutations by immunohistochemistry in light and electron microscopy. In hippocampal layers of wild type and transgenic mice, CaV1.2 channels were predominantly localized to somato-dendritic domains of neurons, and to astrocytic profiles with an age-dependent increase in labeling density. In transgenic animals, CaV1.2-like immunoreactive clusters were found in neuronal profiles in association with amyloid-beta plaques. Both the number and density of these clusters depended upon age of animals and number of plaques. The most striking difference between wild type and transgenic mice was the age-dependent expression of CaV1.2 channels in reactive astrocytes. At the age of 6 month, CaV1.2 channels were rarely detected in reactive astrocytes of transgenic mice, but an incremental number of CaV1.2 expressing reactive astrocytes was found with increasing age of animals and number of amyloid-beta plaques. This study demonstrates that CaV1.2 channels are highly expressed in reactive astrocytes of 12-months of age transgenic mice, which might be a consequence of the increasing amyloid burden. Further studies should clarify which functional implications are associated with the higher availability of CaV1.2 channels in late stage Alzheimer's disease.

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

    Science.gov (United States)

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

    2011-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2005-10-15

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

  14. Voltage Gated Calcium Channel Activation by Backpropagating Action Potentials Downregulates NMDAR Function

    Directory of Open Access Journals (Sweden)

    Anne-Kathrin Theis

    2018-04-01

    Full Text Available The majority of excitatory synapses are located on dendritic spines of cortical glutamatergic neurons. In spines, compartmentalized Ca2+ signals transduce electrical activity into specific long-term biochemical and structural changes. Action potentials (APs propagate back into the dendritic tree and activate voltage gated Ca2+ channels (VGCCs. For spines, this global mode of spine Ca2+ signaling is a direct biochemical feedback of suprathreshold neuronal activity. We previously demonstrated that backpropagating action potentials (bAPs result in long-term enhancement of spine VGCCs. This activity-dependent VGCC plasticity results in a large interspine variability of VGCC Ca2+ influx. Here, we investigate how spine VGCCs affect glutamatergic synaptic transmission. We combined electrophysiology, two-photon Ca2+ imaging and two-photon glutamate uncaging in acute brain slices from rats. T- and R-type VGCCs were the dominant depolarization-associated Ca2+conductances in dendritic spines of excitatory layer 2 neurons and do not affect synaptic excitatory postsynaptic potentials (EPSPs measured at the soma. Using two-photon glutamate uncaging, we compared the properties of glutamatergic synapses of single spines that express different levels of VGCCs. While VGCCs contributed to EPSP mediated Ca2+ influx, the amount of EPSP mediated Ca2+ influx is not determined by spine VGCC expression. On a longer timescale, the activation of VGCCs by bAP bursts results in downregulation of spine NMDAR function.

  15. Characterization of [125I]omega-conotoxin binding to brain N calcium channels and (-)[3H] desmethoxyverapamil binding to novel calcium channels in osteoblast-like osteosarcoma cells

    International Nuclear Information System (INIS)

    Wagner, J.A.

    1987-01-01

    This dissertation provides molecular evidence for a diversity of Ca 2+ channels in neuronal and non-neuronal tissues. First, I demonstrated specific, reversible, saturable binding sites for omega [ 125 I]conotoxin GVIA (omega[ 125 I]CTX) in rat brain and rabbit sympathetic ganglion. Omega [ 125 I]CTX binding has a unique pharmacology, ion selectivity, and anatomical distribution in rat brain. Omega [ 125 I]CTX binding was solubilized, retaining an appropriate pharmacology and ion selectivity. Omega[ 125 I]CTX binding may be associated with a Ca 2+ channel because the K/sub D/ of omega [ 125 I]CTX is similar to the IC 50 of inhibition of depolarization-induced 45 Ca 2+ flux into rat brain synaptosomes. Specific (-)[ 3 H]desmethoxyverapamil ((-)[ 3 H]DMV) binding sites were demonstrated on osteoblast-like osteosarcoma cell membranes

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

    Science.gov (United States)

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

    2017-01-01

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

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

    Science.gov (United States)

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

    2013-09-09

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

  18. Scientific research related to calcium channel blockers poisoning: Bibliometric analysis in Scopus, 1968-2012.

    Science.gov (United States)

    Zyoud, S H; Al-Jabi, S W; Sweileh, W M; Waring, W S

    2015-11-01

    Calcium channel blockers (CCBs) were the most common agents associated with a significant morbidity and mortality rate. The main objective of this study was to examine the publication pattern related to CCBs poisoning at the global level using bibliometric analysis of articles published in SciVerse Scopus online database. Data were searched for documents that contained specific words regarding CCB poisoning as keywords in the title. No time period limitations were specified in the search regarding the starting year. The ending date of the search was 31 December 2012. The criteria were met by 713 publications from 53 countries. The largest number of articles associated with CCBs was from the United States (30%), followed by the United Kingdom (7.4%), Japan (6%), and Germany (5.6%). No data related to CCBs were published from 159 (75%) of 212 countries registered in World Bank online database. There was no correlation between the number of published articles in the country and its population size (r = 0.03, p > 0.926). United Kingdom and Australia were the leading countries in terms of number of CCBs publications per million inhabitants (0.83 and 0.82 articles per million inhabitants, respectively), followed by the United States (0.68). Countries with a large population, such as India, tended to rank relatively low (0.01 articles per million inhabitants). The total number of citations at the time of data analysis (23 October 2014) was 6462, with an average of 9.1 citations per document. The highest median (interquartile range) number of citations was 8 (8-18) for the United States, followed by 6 (1-21) for Australia, 5 (1-15) for the United Kingdom, and 5 (1-24) for Canada. The h-index of the retrieved documents was 37. Scientific production on CCBs poisoning is increasing; nonetheless, the international collaboration is still rare. The amount of CCBs-based research activity was low or not available in most countries. More regional epidemiological studies are

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

    International Nuclear Information System (INIS)

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

    2008-01-01

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

  20. Differential effect of T-type voltage-gated calcium channel disruption on renal plasma flow and glomerular filtration rate in vivo

    DEFF Research Database (Denmark)

    Thuesen, Anne D; Andersen, Henrik; Cardel, Majken

    2014-01-01

    Voltage-gated calcium channels (Cav) play an essential role in regulation of renal blood flow and GFR. Because T-type Cavs are differentially expressed in pre- and postglomerular vessels it was hypothesized that they impact renal blood flow and GFR differentially. The question was addressed by use...... of two T-type Cav knock-out mice strains. Continuous recordings of blood pressure and heart rate, and para-aminohippurate clearance (renal plasma flow) and inulin clearance (GFR) were performed in conscious, chronically catheterized, wild type and Cav 3.1-/- and Cav 3.2-/- mice. Contractility of afferent...... and efferent arterioles was determined in isolated perfused blood vessels. Efferent arterioles from Cav 3.2-/- mice constricted significantly more in response to a depolarization compared to Wt mice. GFR was increased in Cav 3.2-/- mice with no significant changes in renal plasma flow, heart rate and blood...

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

    Directory of Open Access Journals (Sweden)

    Joo Hyun Nam

    2016-07-01

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

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  3. Double-Nanodomain Coupling of Calcium Channels, Ryanodine Receptors, and BK Channels Controls the Generation of Burst Firing.

    Science.gov (United States)

    Irie, Tomohiko; Trussell, Laurence O

    2017-11-15

    Action potentials clustered into high-frequency bursts play distinct roles in neural computations. However, little is known about ionic currents that control the duration and probability of these bursts. We found that, in cartwheel inhibitory interneurons of the dorsal cochlear nucleus, the likelihood of bursts and the interval between their spikelets were controlled by Ca 2+ acting across two nanodomains, one between plasma membrane P/Q Ca 2+ channels and endoplasmic reticulum (ER) ryanodine receptors and another between ryanodine receptors and large-conductance, voltage- and Ca 2+ -activated K + (BK) channels. Each spike triggered Ca 2+ -induced Ca 2+ release (CICR) from the ER immediately beneath somatic, but not axonal or dendritic, plasma membrane. Moreover, immunolabeling demonstrated close apposition of ryanodine receptors and BK channels. Double-nanodomain coupling between somatic plasma membrane and hypolemmal ER cisterns provides a unique mechanism for rapid control of action potentials on the millisecond timescale. Copyright © 2017 Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

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

    2015-01-01

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

  5. Gene expression changes of single skeletal muscle fibers in response to modulation of the mitochondrial calcium uniporter (MCU

    Directory of Open Access Journals (Sweden)

    Francesco Chemello

    2015-09-01

    Full Text Available The mitochondrial calcium uniporter (MCU gene codifies for the inner mitochondrial membrane (IMM channel responsible for mitochondrial Ca2+ uptake. Cytosolic Ca2+ transients are involved in sarcomere contraction through cycles of release and storage in the sarcoplasmic reticulum. In addition cytosolic Ca2+ regulates various signaling cascades that eventually lead to gene expression reprogramming. Mitochondria are strategically placed in close contact with the ER/SR, thus cytosolic Ca2+ transients elicit large increases in the [Ca2+] of the mitochondrial matrix ([Ca2+]mt. Mitochondrial Ca2+ uptake regulates energy production and cell survival. In addition, we recently showed that MCU-dependent mitochondrial Ca2+ uptake controls skeletal muscle trophism. In the same report, we dissected the effects of MCU-dependent mitochondrial Ca2+ uptake on gene expression through microarray gene expression analysis upon modulation of MCU expression by in vivo AAV infection. Analyses were performed on single skeletal muscle fibers at two time points (7 and 14 days post-AAV injection. Raw and normalized data are available on the GEO database (http://www.ncbi.nlm.nih.gov/geo/ (GSE60931.

  6. Expression and distribution of voltage-gated ion channels in ferret sinoatrial node.

    Science.gov (United States)

    Brahmajothi, Mulugu V; Morales, Michael J; Campbell, Donald L; Steenbergen, Charles; Strauss, Harold C

    2010-10-01

    Spontaneous diastolic depolarization in the sinoatrial (SA) node enables it to serve as pacemaker of the heart. The variable cell morphology within the SA node predicts that ion channel expression would be heterogeneous and different from that in the atrium. To evaluate ion channel heterogeneity within the SA node, we used fluorescent in situ hybridization to examine ion channel expression in the ferret SA node region and atrial appendage. SA nodal cells were distinguished from surrounding cardiac myocytes by expression of the slow (SA node) and cardiac (surrounding tissue) forms of troponin I. Nerve cells in the sections were identified by detection of GAP-43 and cytoskeletal middle neurofilament. Transcript expression was characterized for the 4 hyperpolarization-activated cation channels, 6 voltage-gated Na(+) channels, 3 voltage-gated Ca(2+) channels, 24 voltage-gated K(+) channel α-subunits, and 3 ancillary subunits. To ensure that transcript expression was representative of protein expression, immunofluorescence was used to verify localization patterns of voltage-dependent K(+) channels. Colocalizations were performed to observe any preferential patterns. Some overlapping and nonoverlapping binding patterns were observed. Measurement of different cation channel transcripts showed heterogeneous expression with many different patterns of expression, attesting to the complexity of electrical activity in the SA node. This study provides insight into the possible role ion channel heterogeneity plays in SA node pacemaker activity.

  7. The alpha2-delta protein: an auxiliary subunit of voltage-dependent calcium channels as a recognized drug target.

    Science.gov (United States)

    Thorpe, Andrew J; Offord, James

    2010-07-01

    Currently, there are two drugs on the market, gabapentin (Neurontin) and pregabalin (Lyrica), that are proposed to exert their therapeutic effect through binding to the alpha2-delta subunit of voltage-sensitive calcium channels. This activity was unexpected, as the alpha2-delta subunit had previously been considered not to be a pharmacological target. In this review, the role of the alpha2-delta subunits is discussed and the mechanism of action of the alpha2-delta ligands in vitro and in vivo is summarized. Finally, new insights into the mechanism of drugs that bind to this protein are discussed.

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

    OpenAIRE

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

    2015-01-01

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

  9. Identification of cyclic nucleotide gated channels using regular expressions

    KAUST Repository

    Zelman, Alice K.; Dawe, Adam Sean; Berkowitz, Gerald A.

    2013-01-01

    Cyclic nucleotide-gated channels (CNGCs) are nonselective cation channels found in plants, animals, and some bacteria. They have a six-transmembrane/one- pore structure, a cytosolic cyclic nucleotide-binding domain, and a cytosolic calmodulin

  10. Inhibition of T-Type Voltage Sensitive Calcium Channel Reduces Load-Induced OA in Mice and Suppresses the Catabolic Effect of Bone Mechanical Stress on Chondrocytes.

    Directory of Open Access Journals (Sweden)

    Padma P Srinivasan

    Full Text Available Voltage-sensitive calcium channels (VSCC regulate cellular calcium influx, one of the earliest responses to mechanical stimulation in osteoblasts. Here, we postulate that T-type VSCCs play an essential role in bone mechanical response to load and participate in events leading to the pathology of load-induced OA. Repetitive mechanical insult was used to induce OA in Cav3.2 T-VSCC null and wild-type control mouse knees. Osteoblasts (MC3T3-E1 and chondrocytes were treated with a selective T-VSCC inhibitor and subjected to fluid shear stress to determine how blocking of T-VSCCs alters the expression profile of each cell type upon mechanical stimulation. Conditioned-media (CM obtained from static and sheared MC3T3-E1 was used to assess the effect of osteoblast-derived factors on the chondrocyte phenotype. T-VSCC null knees exhibited significantly lower focal articular cartilage damage than age-matched controls. In vitro inhibition of T-VSCC significantly reduced the expression of both early and late mechanoresponsive genes in osteoblasts but had no effect on gene expression in chondrocytes. Furthermore, treatment of chondrocytes with CM obtained from sheared osteoblasts induced expression of markers of hypertrophy in chondrocytes and this was nearly abolished when osteoblasts were pre-treated with the T-VSCC-specific inhibitor. These results indicate that T-VSCC plays a role in signaling events associated with induction of OA and is essential to the release of osteoblast-derived factors that promote an early OA phenotype in chondrocytes. Further, these findings suggest that local inhibition of T-VSCC may serve as a therapy for blocking load-induced bone formation that results in cartilage degeneration.

  11. TRPA1 expression levels and excitability brake by KV channels influence cold sensitivity of TRPA1-expressing neurons.

    Science.gov (United States)

    Memon, Tosifa; Chase, Kevin; Leavitt, Lee S; Olivera, Baldomero M; Teichert, Russell W

    2017-06-14

    The molecular sensor of innocuous (painless) cold sensation is well-established to be transient receptor potential cation channel, subfamily M, member 8 (TRPM8). However, the role of transient receptor potential cation channel, subfamily A, member 1 (TRPA1) in noxious (painful) cold sensation has been controversial. We find that TRPA1 channels contribute to the noxious cold sensitivity of mouse somatosensory neurons, independent of TRPM8 channels, and that TRPA1-expressing neurons are largely non-overlapping with TRPM8-expressing neurons in mouse dorsal-root ganglia (DRG). However, relatively few TRPA1-expressing neurons (e.g., responsive to allyl isothiocyanate or AITC, a selective TRPA1 agonist) respond overtly to cold temperature in vitro, unlike TRPM8-expressing neurons, which almost all respond to cold. Using somatosensory neurons from TRPM8-/- mice and subtype-selective blockers of TRPM8 and TRPA1 channels, we demonstrate that responses to cold temperatures from TRPA1-expressing neurons are mediated by TRPA1 channels. We also identify two factors that affect the cold-sensitivity of TRPA1-expressing neurons: (1) cold-sensitive AITC-sensitive neurons express relatively more TRPA1 transcripts than cold-insensitive AITC-sensitive neurons and (2) voltage-gated potassium (K V ) channels attenuate the cold-sensitivity of some TRPA1-expressing neurons. The combination of these two factors, combined with the relatively weak agonist-like activity of cold temperature on TRPA1 channels, partially explains why few TRPA1-expressing neurons respond to cold. Blocking K V channels also reveals another subclass of noxious cold-sensitive DRG neurons that do not express TRPM8 or TRPA1 channels. Altogether, the results of this study provide novel insights into the cold-sensitivity of different subclasses of somatosensory neurons. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

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

    Science.gov (United States)

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

    2013-01-01

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

  13. Single Nisoldipine-Sensitive Calcium Channels in Smooth Muscle Cells Isolated from Rabbit Mesenteric Artery

    Science.gov (United States)

    Worley, Jennings F.; Deitmer, Joachim W.; Nelson, Mark T.

    1986-08-01

    Single smooth muscle cells were enzymatically isolated from the rabbit mesenteric artery. At physiological levels of external Ca, these cells were relaxed and contracted on exposure to norepinephrine, caffeine, or high levels of potassium. The patch-clamp technique was used to measure unitary currents through single channels in the isolated cells. Single channels were selective for divalent cations and exhibited two conductance levels, 8 pS and 15 pS. Both types of channels were voltage-dependent, and channel activity occurred at potentials positive to -40 mV. The activity of both channel types was almost completely inhibited by 50 nM nisoldipine. These channels appear to be the pathways for voltage-dependent Ca influx in vascular smooth muscle and may be the targets of the clinically used dihydropyridines.

  14. Macrophage activation by a vanadyl-aspirin complex is dependent on L-type calcium channel and the generation of nitric oxide

    International Nuclear Information System (INIS)

    Molinuevo, Maria Silvina; Etcheverry, Susana Beatriz; Cortizo, Ana Maria

    2005-01-01

    Bone homeostasis is the result of a tight balance between bone resorption and bone formation where macrophage activation is believed to contribute to bone resorption. We have previously shown that a vanadyl(IV)-aspirin complex (VOAspi) regulates cell proliferation and differentiation of osteoblasts in culture. In this study, we assessed VOAspi and VO effects and their possible mechanism of action on a mouse macrophage cell line RAW 264.7. Both vanadium compounds inhibited cell proliferation in a dose-dependent manner. Nifedipine completely reversed the VOAspi-induced macrophage cytotoxicity, while it could not block the effect of VO. VOAspi also stimulated nitric oxide (NO) production, the oxidation of dihydrorhodamine 123 (DHR-123) and enhanced the expression of both constitutive and inducible isoforms of nitric oxide syntases (NOS). All these effects were abolished by nifedipine. Althogether our finding give evidence that VOAspi-induced macrophage cytotoxicity is dependent on L-type calcium channel and the generation of NO though the induction of eNOS and iNOS. Contrary, the parent compound VO exerted a cytotoxic effect by mechanisms independent of a calcium entry and the NO/NOS activation

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

    Directory of Open Access Journals (Sweden)

    Zhi-Ren Zhang

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

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

    Science.gov (United States)

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

    2014-07-01

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

  17. Expression of Water Channel Proteins in Mesembryanthemum crystallinum1

    Science.gov (United States)

    Kirch, Hans-Hubert; Vera-Estrella, Rosario; Golldack, Dortje; Quigley, Francoise; Michalowski, Christine B.; Barkla, Bronwyn J.; Bohnert, Hans J.

    2000-01-01

    We have characterized transcripts for nine major intrinsic proteins (MIPs), some of which function as water channels (aquaporins), from the ice plant Mesembryanthemum crystallinum. To determine the cellular distribution and expression of these MIPs, oligopeptide-based antibodies were generated against MIP-A, MIP-B, MIP-C, or MIP-F, which, according to sequence and functional characteristics, are located in the plasma membrane (PM) and tonoplast, respectively. MIPs were most abundant in cells involved in bulk water flow and solute flux. The tonoplast MIP-F was found in all cells, while signature cell types identified different PM-MIPs: MIP-A predominantly in phloem-associated cells, MIP-B in xylem parenchyma, and MIP-C in the epidermis and endodermis of immature roots. Membrane protein analysis confirmed MIP-F as tonoplast located. MIP-A and MIP-B were found in tonoplast fractions and also in fractions distinct from either the tonoplast or PM. MIP-C was most abundant but not exclusive to PM fractions, where it is expected based on its sequence signature. We suggest that within the cell, MIPs are mobile, which is similar to aquaporins cycling through animal endosomes. MIP cycling and the differential regulation of these proteins observed under conditions of salt stress may be fundamental for the control of tissue water flux. PMID:10806230

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

    Science.gov (United States)

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

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Alberto F Rubio-Guerra

    2009-11-01

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

  20. Expression, purification and functional reconstitution of slack sodium-activated potassium channels.

    Science.gov (United States)

    Yan, Yangyang; Yang, Youshan; Bian, Shumin; Sigworth, Fred J

    2012-11-01

    The slack (slo2.2) gene codes for a potassium-channel α-subunit of the 6TM voltage-gated channel family. Expression of slack results in Na(+)-activated potassium channel activity in various cell types. We describe the purification and reconstitution of Slack protein and show that the Slack α-subunit alone is sufficient for potassium channel activity activated by sodium ions as assayed in planar bilayer membranes and in membrane vesicles.

  1. Loss of ATP-Sensitive Potassium Channel Surface Expression in Heart Failure Underlies Dysregulation of Action Potential Duration and Myocardial Vulnerability to Injury.

    Directory of Open Access Journals (Sweden)

    Zhan Gao

    Full Text Available The search for new approaches to treatment and prevention of heart failure is a major challenge in medicine. The adenosine triphosphate-sensitive potassium (KATP channel has been long associated with the ability to preserve myocardial function and viability under stress. High surface expression of membrane KATP channels ensures a rapid energy-sparing reduction in action potential duration (APD in response to metabolic challenges, while cellular signaling that reduces surface KATP channel expression blunts APD shortening, thus sacrificing energetic efficiency in exchange for greater cellular calcium entry and increased contractile force. In healthy hearts, calcium/calmodulin-dependent protein kinase II (CaMKII phosphorylates the Kir6.2 KATP channel subunit initiating a cascade responsible for KATP channel endocytosis. Here, activation of CaMKII in a transaortic banding (TAB model of heart failure is coupled with a 35-40% reduction in surface expression of KATP channels compared to hearts from sham-operated mice. Linkage between KATP channel expression and CaMKII is verified in isolated cardiomyocytes in which activation of CaMKII results in downregulation of KATP channel current. Accordingly, shortening of monophasic APD is slowed in response to hypoxia or heart rate acceleration in failing compared to non-failing hearts, a phenomenon previously shown to result in significant increases in oxygen consumption. Even in the absence of coronary artery disease, failing myocardium can be further injured by ischemia due to a mismatch between metabolic supply and demand. Ischemia-reperfusion injury, following ischemic preconditioning, is diminished in hearts with CaMKII inhibition compared to wild-type hearts and this advantage is largely eliminated when myocardial KATP channel expression is absent, supporting that the myocardial protective benefit of CaMKII inhibition in heart failure may be substantially mediated by KATP channels. Recognition of Ca

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

    Directory of Open Access Journals (Sweden)

    Nooshin Masoudian

    2015-04-01

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

  3. Differential expression of BK channel isoforms and beta-subunits in rat neuro-vascular tissues

    DEFF Research Database (Denmark)

    Poulsen, Asser Nyander; Wulf, Helle; Hay-Schmidt, Anders

    2009-01-01

    We investigated the expression of splice variants and beta-subunits of the BK channel (big conductance Ca(2+)-activated K(+) channel, Slo1, MaxiK, K(Ca)1.1) in rat cerebral blood vessels, meninges, trigeminal ganglion among other tissues. An alpha-subunit splice variant X1(+24) was found expresse...

  4. Expression of the Sodium/Calcium/Potassium Exchanger, NCKX4, in Ameloblasts

    Science.gov (United States)

    Hu, Ping; Lacruz, Rodrigo S.; Smith, Charles E.; Smith, Susan M.; Kurtz, Ira; Paine, Michael L.

    2012-01-01

    Transcellular calcium transport is an essential activity in mineralized tissue formation, including dental hard tissues. In many organ systems, this activity is regulated by membrane-bound sodium/calcium (Na+/Ca2+) exchangers, which include the NCX and NCKX [sodium/calcium-potassium (Na+/Ca2+-K+ ) exchanger] proteins. During enamel maturation, when crystals expand in thickness, Ca2+ requirements vastly increase but exactly how Ca2+ traffics through ameloblasts remains uncertain. Previous studies have shown that several NCX proteins are expressed in ameloblasts, although no significant shifts in expression were observed during maturation which pointed to the possible identification of other Ca2+ membrane transporters. NCKX proteins are encoded by members of the solute carrier gene family, Slc24a, which include 6 different proteins (NCKX1–6). NCKX are bidirectional electrogenic transporters regulating Ca2+ transport in and out of cells dependent on the transmembrane ion gradient. In this study we show that all NCKX mRNAs are expressed in dental tissues. Real-time PCR indicates that of all the members of the NCKX group, NCKX4 is the most highly expressed gene transcript during the late stages of amelogenesis. In situ hybridization and immunolocalization analyses clearly establish that in the enamel organ, NCKX4 is expressed primarily by ameloblasts during the maturation stage. Further, during the mid-late maturation stages of amelogenesis, the expression of NCKX4 in ameloblasts is most prominent at the apical poles and at the lateral membranes proximal to the apical ends. These data suggest that NCKX4 might be an important regulator of Ca2+ transport during amelogenesis. PMID:22677781

  5. Channel formation in model membranes by the adenylate cyclase toxin of Bordetella pertussis: Effect if Calcium

    Czech Academy of Sciences Publication Activity Database

    Knapp, O.; Maier, E.; Polleichtner, G.; Mašín, Jiří; Šebo, Peter; Benz, R.

    2003-01-01

    Roč. 42, - (2003), s. 8077-8084 ISSN 0006-2960 R&D Projects: GA AV ČR IPP1050128 Institutional research plan: CEZ:AV0Z5020903 Keywords : bordetella pertussis * calcium Subject RIV: EE - Microbiology, Virology Impact factor: 3.922, year: 2003

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

    DEFF Research Database (Denmark)

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

    2013-01-01

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

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

    Science.gov (United States)

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

    2012-03-01

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

  8. Inhibition of Calcium-Activated Chloride Channel ANO1/TMEM16A Suppresses Tumor Growth and Invasion in Human Lung Cancer.

    Directory of Open Access Journals (Sweden)

    Linghan Jia

    Full Text Available Lung cancer or pulmonary carcinoma is primarily derived from epithelial cells that are thin and line on the alveolar surfaces of the lung for gas exchange. ANO1/TMEM16A, initially identified from airway epithelial cells, is a member of Ca2+-activated Cl- channels (CaCCs that function to regulate epithelial secretion and cell volume for maintenance of ion and tissue homeostasis. ANO1/TMEM16A has recently been shown to be highly expressed in several epithelium originated carcinomas. However, the role of ANO1 in lung cancer remains unknown. In this study, we show that inhibition of calcium-activated chloride channel ANO1/TMEM16A suppresses tumor growth and invasion in human lung cancer. ANO1 is upregulated in different human lung cancer cell lines. Knocking-down ANO1 by small hairpin RNAs inhibited proliferation, migration and invasion of GLC82 and NCI-H520 cancel cells evaluated by CCK-8, would-healing, transwell and 3D soft agar assays. ANO1 protein is overexpressed in 77.3% cases of human lung adenocarcinoma tissues detected by immunohistochemistry. Furthermore, the tumor growth in nude mice implanted with GLC82 cells was significantly suppressed by ANO1 silencing. Taken together, our findings provide evidence that ANO1 overexpression contributes to tumor growth and invasion of lung cancer; and suppressing ANO1 overexpression may have therapeutic potential in lung cancer therapy.

  9. Functional expression of T-type Ca2+ channels in spinal motoneurons of the adult turtle.

    Directory of Open Access Journals (Sweden)

    Martha Canto-Bustos

    Full Text Available Voltage-gated Ca2+ (CaV channels are transmembrane proteins comprising three subfamilies named CaV1, CaV2 and CaV3. The CaV3 channel subfamily groups the low-voltage activated Ca2+ channels (LVA or T-type a significant role in regulating neuronal excitability. CaV3 channel activity may lead to the generation of complex patterns of action potential firing such as the postinhibitory rebound (PIR. In the adult spinal cord, these channels have been found in dorsal horn interneurons where they control physiological events near the resting potential and participate in determining excitability. In motoneurons, CaV3 channels have been found during development, but their functional expression has not yet been reported in adult animals. Here, we show evidence for the presence of CaV3 channel-mediated PIR in motoneurons of the adult turtle spinal cord. Our results indicate that Ni2+ and NNC55-0396, two antagonists of CaV3 channel activity, inhibited PIR in the adult turtle spinal cord. Molecular biology and biochemical assays revealed the expression of the CaV3.1 channel isotype and its localization in motoneurons. Together, these results provide evidence for the expression of CaV3.1 channels in the spinal cord of adult animals and show also that these channels may contribute to determine the excitability of motoneurons.

  10. Ion channel expression patterns in glioblastoma stem cells with functional and therapeutic implications for malignancy.

    Directory of Open Access Journals (Sweden)

    Julia Pollak

    Full Text Available Ion channels and transporters have increasingly recognized roles in cancer progression through the regulation of cell proliferation, migration, and death. Glioblastoma stem-like cells (GSCs are a source of tumor formation and recurrence in glioblastoma multiforme, a highly aggressive brain cancer, suggesting that ion channel expression may be perturbed in this population. However, little is known about the expression and functional relevance of ion channels that may contribute to GSC malignancy. Using RNA sequencing, we assessed the enrichment of ion channels in GSC isolates and non-tumor neural cell types. We identified a unique set of GSC-enriched ion channels using differential expression analysis that is also associated with distinct gene mutation signatures. In support of potential clinical relevance, expression of selected GSC-enriched ion channels evaluated in human glioblastoma databases of The Cancer Genome Atlas and Ivy Glioblastoma Atlas Project correlated with patient survival times. Finally, genetic knockdown as well as pharmacological inhibition of individual or classes of GSC-enriched ion channels constrained growth of GSCs compared to normal neural stem cells. This first-in-kind global examination characterizes ion channels enriched in GSCs and explores their potential clinical relevance to glioblastoma molecular subtypes, gene mutations, survival outcomes, regional tumor expression, and experimental responses to loss-of-function. Together, the data support the potential biological and therapeutic impact of ion channels on GSC malignancy and provide strong rationale for further examination of their mechanistic and therapeutic importance.

  11. Ion channel expression patterns in glioblastoma stem cells with functional and therapeutic implications for malignancy.

    Science.gov (United States)

    Pollak, Julia; Rai, Karan G; Funk, Cory C; Arora, Sonali; Lee, Eunjee; Zhu, Jun; Price, Nathan D; Paddison, Patrick J; Ramirez, Jan-Marino; Rostomily, Robert C

    2017-01-01

    Ion channels and transporters have increasingly recognized roles in cancer progression through the regulation of cell proliferation, migration, and death. Glioblastoma stem-like cells (GSCs) are a source of tumor formation and recurrence in glioblastoma multiforme, a highly aggressive brain cancer, suggesting that ion channel expression may be perturbed in this population. However, little is known about the expression and functional relevance of ion channels that may contribute to GSC malignancy. Using RNA sequencing, we assessed the enrichment of ion channels in GSC isolates and non-tumor neural cell types. We identified a unique set of GSC-enriched ion channels using differential expression analysis that is also associated with distinct gene mutation signatures. In support of potential clinical relevance, expression of selected GSC-enriched ion channels evaluated in human glioblastoma databases of The Cancer Genome Atlas and Ivy Glioblastoma Atlas Project correlated with patient survival times. Finally, genetic knockdown as well as pharmacological inhibition of individual or classes of GSC-enriched ion channels constrained growth of GSCs compared to normal neural stem cells. This first-in-kind global examination characterizes ion channels enriched in GSCs and explores their potential clinical relevance to glioblastoma molecular subtypes, gene mutations, survival outcomes, regional tumor expression, and experimental responses to loss-of-function. Together, the data support the potential biological and therapeutic impact of ion channels on GSC malignancy and provide strong rationale for further examination of their mechanistic and therapeutic importance.

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

    Directory of Open Access Journals (Sweden)

    Mehrdad Roghani

    2006-03-01

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

  13. Voltage gated potassium channels expressed in Xenopus laevis(AMPHIBIA oocytes

    Directory of Open Access Journals (Sweden)

    Hedna Chaves

    2003-01-01

    Full Text Available Heterologous expression has been an important tool for structural and functionalcharacterization of proteins. The study of biophysical properties of ion channels,pumps and transporters has been possible thanks to their expression in Xenopuslaevisoocytes. Here we report the expression of two voltage gated channels, Kv1.1and Shaker, in X. laevisoocytes using a method for oocyte extraction, isolation, cul-ture, and microinjection adapted to the latitude and altitude conditions of Bogotá,Colombia.

  14. Expression of the calcium-binding proteins MRP8 and MRP14 in monocytes is regulated by a calcium-induced suppressor mechanism.

    OpenAIRE

    Roth, J; Goebeler, M; Wrocklage, V; van den Bos, C; Sorg, C

    1994-01-01

    MRP8 and MRP14 are two calcium-binding proteins of the S-100 family the expression of which is restricted to distinct stages of monocytic differentiation. Heteromeric MRP8/MRP14 complexes have been shown to represent their biologically active forms. However, it is not as yet clear whether biochemical modification of complexes, or regulation on the transcriptional level, are responsible for the control of MRP8/MRP14 expression. Employing Western-blot analysis and metabolic labelling we have de...

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

    Science.gov (United States)

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

    2009-08-01

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

  16. Calcium channel blockers inhibit retinal degeneration in the retinal-degeneration-B mutant of Drosophila.

    Science.gov (United States)

    Sahly, I; Bar Nachum, S; Suss-Toby, E; Rom, A; Peretz, A; Kleiman, J; Byk, T; Selinger, Z; Minke, B

    1992-01-01

    Light accelerates degeneration of photoreceptor cells of the retinal degeneration B (rdgB) mutant of Drosophila. During early stages of degeneration, light stimuli evoke spikes from photoreceptors of the mutant fly; no spikes can be recorded from photoreceptors of the wild-type fly. Production of spike potentials from mutant photoreceptors was blocked by diltiazem, verapamil hydrochloride, and cadmium. Little, if any, effect of the (-)-cis isomer or (+)-cis isomer of diltiazem on the light response was seen. Further, the (+)-cis isomer was approximately 50 times more effective than the (-)-cis isomer in blocking the Ca2+ spikes, indicating that diltiazem action on the rdgB eye is mediated by means of blocking voltage-sensitive Ca2+ channels, rather than by blocking the light-sensitive channels. Application of the Ca(2+)-channel blockers (+)-cis-diltiazem and verapamil hydrochloride to the eyes of rdgB flies over a 7-day period largely inhibited light-dependent degeneration of the photoreceptor cells. Pulse labeling with [32P]phosphate showed much greater incorporation into eye proteins of [32P]phosphate in rdgB flies than in wild-type flies. Retarding the light-induced photoreceptor degeneration in the mutant by Ca(2+)-channel blockers, thus, suggests that toxic increase in intracellular Ca2+ by means of voltage-gated Ca2+ channels, possibly secondary to excessive phosphorylation, leads to photoreceptor degeneration in the rdgB mutant. Images PMID:1309615

  17. Calcium channel blockers inhibit endogenous pyrogen fever in rats and rabbits.

    Science.gov (United States)

    Stitt, J T; Shimada, S G

    1991-09-01

    We have previously shown that febrile responses in both rats and rabbits are elicited by the intravenous injection of a semipurified endogenous pyrogen (EP) prepared from human monocytes. We are now presenting evidence that these febrile responses are mediated via activation of Ca2+ channels by EP. The febrile responses of male New Zealand White rabbits and Sprague-Dawley rats to a standard dose of EP were determined at their respective thermoneutral ambient temperatures. The animals were then treated with Ca2+ channel blocker verapamil (7.5 mg/kg iv) 30-60 min before the EP challenge. In every case the febrile response to EP was markedly attenuated after verapamil pretreatment, while administration of verapamil by itself had no detectable effect on body temperature. Another Ca2+ channel blocker, nifedipine (5 mg/kg iv), was shown to possess antipyretic activity in rats also. To localize where in the fever pathway these Ca2+ channel blockers were acting, we investigated the effect of verapamil at the same dose on fevers that were produced by microinjection of prostaglandin E (PGE) directly into the brain. These PGE fevers were unaffected by verapamil pretreatment, indicating that the antipyretic action of Ca2+ channel blockers occurs before the formation of PGE in response to EP stimulation. The most likely locus of action is the activation of the enzyme phospholipase A2, which regulates the production of arachidonic acid from cellular phospholipids in the prostanoid cascade.

  18. Neuronal MHC Class I Expression Is Regulated by Activity Driven Calcium Signaling.

    Directory of Open Access Journals (Sweden)

    Dan Lv

    Full Text Available MHC class I (MHC-I molecules are important components of the immune system. Recently MHC-I have been reported to also play important roles in brain development and synaptic plasticity. In this study, we examine the molecular mechanism(s underlying activity-dependent MHC-I expression using hippocampal neurons. Here we report that neuronal expression level of MHC-I is dynamically regulated during hippocampal development after birth in vivo. Kainic acid (KA treatment significantly increases the expression of MHC-I in cultured hippocampal neurons in vitro, suggesting that MHC-I expression is regulated by neuronal activity. In addition, KA stimulation decreased the expression of pre- and post-synaptic proteins. This down-regulation is prevented by addition of an MHC-I antibody to KA treated neurons. Further studies demonstrate that calcium-dependent protein kinase C (PKC is important in relaying KA simulation activation signals to up-regulated MHC-I expression. This signaling cascade relies on activation of the MAPK pathway, which leads to increased phosphorylation of CREB and NF-κB p65 while also enhancing the expression of IRF-1. Together, these results suggest that expression of MHC-I in hippocampal neurons is driven by Ca2+ regulated activation of the MAPK signaling transduction cascade.

  19. Cryo-electron microscopy structure of the lysosomal calcium-permeable channel TRPML3.

    Science.gov (United States)

    Hirschi, Marscha; Herzik, Mark A; Wie, Jinhong; Suo, Yang; Borschel, William F; Ren, Dejian; Lander, Gabriel C; Lee, Seok-Yong

    2017-10-19

    The modulation of ion channel activity by lipids is increasingly recognized as a fundamental component of cellular signalling. The transient receptor potential mucolipin (TRPML) channel family belongs to the TRP superfamily and is composed of three members: TRPML1-TRPML3. TRPMLs are the major Ca 2+ -permeable channels on late endosomes and lysosomes (LEL). They regulate the release of Ca 2+ from organelles, which is important for various physiological processes, including organelle trafficking and fusion. Loss-of-function mutations in the MCOLN1 gene, which encodes TRPML1, cause the neurodegenerative lysosomal storage disorder mucolipidosis type IV, and a gain-of-function mutation (Ala419Pro) in TRPML3 gives rise to the varitint-waddler (Va) mouse phenotype. Notably, TRPML channels are activated by the low-abundance and LEL-enriched signalling lipid phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)P 2 ), whereas other phosphoinositides such as PtdIns(4,5)P 2 , which is enriched in plasma membranes, inhibit TRPMLs. Conserved basic residues at the N terminus of the channel are important for activation by PtdIns(3,5)P 2 and inhibition by PtdIns(4,5)P 2 . However, owing to a lack of structural information, the mechanism by which TRPML channels recognize PtdIns(3,5)P 2 and increase their Ca 2+ conductance remains unclear. Here we present the cryo-electron microscopy (cryo-EM) structure of a full-length TRPML3 channel from the common marmoset (Callithrix jacchus) at an overall resolution of 2.9 Å. Our structure reveals not only the molecular basis of ion conduction but also the unique architecture of TRPMLs, wherein the voltage sensor-like domain is linked to the pore via a cytosolic domain that we term the mucolipin domain. Combined with functional studies, these data suggest that the mucolipin domain is responsible for PtdIns(3,5)P 2 binding and subsequent channel activation, and that it acts as a 'gating pulley' for lipid-dependent TRPML gating.

  20. Calcium-channel blockers do not alter the clinical efficacy of clopidogrel after myocardial infarction: a nationwide cohort study

    DEFF Research Database (Denmark)

    Olesen, Jonas B; Gislason, Gunnar H; Charlot, Mette G

    2011-01-01

    Objectives The purpose of this study was to determine the risk of adverse cardiovascular events associated with concomitant use of clopidogrel and calcium-channel blockers (CCBs) in patients with myocardial infarction (MI). Background CCBs inhibit a variety of cytochrome P-450 enzymes, some...... patients treated and not treated with clopidogrel, with a hazard ratio of 1.15 (95% confidence interval [CI]: 1.07 to 1.24) and 1.05 (95% CI: 1.01 to 1.11), respectively. The increased risk was independent of clopidogrel use; the hazard rate ratio was 1.08 (95% CI: 0.99 to 1.18). Analyses of all additional...... adverse end points and propensity score–matched models provided similar results. Conclusions The clinical efficacy of clopidogrel in patients with a recent MI is not modified by concomitant CCB treatment. This potential drug interaction is unlikely to have clinical significance....

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

    Science.gov (United States)

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

    2012-01-01

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

  2. Dose calcium channel blocker verapamil decrease urinary VMA levels in sympathoadrenal hyperactive patients with posttraumatic stress disorder?

    Institute of Scientific and Technical Information of China (English)

    Munawar Alam Ansari; Shahida PAhmed; Zahida Memon

    2008-01-01

    Objective:The majority of the patients with posttraumatic stress disorders (PTSD)embrace augmented urina-ry flow of Vanillylmandelic Acid (VMA)than normal subjects owing to superior sympathetic doings,which steer to cardiovascular catastrophe.Urinary flow of VMA was evaluated as sympathoadrenal bustle marker in patients with posttraumatic stress disorder.Calcium ion shows a noteworthy dependability in nervousness owing to its special effects on brain synaptosomes.So this study was conducted to explore the effects of Verapamil on sympathoadrenal motion in patients with PTSD.Methods:Placebo controlled clinical tryout was conducted. At first hundred (100)PTSD patients were chosen and enrolled in the study,from department of Psychological Medicine Dow University of Health Sciences,Karachi.Verapamil 120 mg/day was specified in divided doses to group-I (n =50)patients and group-II (n =50)patients received placebo therapy on a daily basis for nine weeks.Each and every patient was monitored weekly,all the way through extent of study.Results:Under-neath the posttraumatic stress disorder,urinary excretion of VMA was greater.Calcium channel blocker vera-pamil additionally abolished the embellished retort in urinary flow of VMA appreciably in patients with PTSD. Conclusion:Verapamil was experiential to be exceedingly effectual treatment.It reduces VMA levels in u-rine,and on the whole cardiovascular threat in PTSD patients.

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

    Directory of Open Access Journals (Sweden)

    Oyku Gulmez

    2008-12-01

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

  4. A voltage-gated calcium channel regulates lysosomal fusion with endosomes and autophagosomes and is required for neuronal homeostasis.

    Directory of Open Access Journals (Sweden)

    Xuejun Tian

    2015-03-01

    Full Text Available Autophagy helps deliver sequestered intracellular cargo to lysosomes for proteolytic degradation and thereby maintains cellular homeostasis by preventing accumulation of toxic substances in cells. In a forward mosaic screen in Drosophila designed to identify genes required for neuronal function and maintenance, we identified multiple cacophony (cac mutant alleles. They exhibit an age-dependent accumulation of autophagic vacuoles (AVs in photoreceptor terminals and eventually a degeneration of the terminals and surrounding glia. cac encodes an α1 subunit of a Drosophila voltage-gated calcium channel (VGCC that is required for synaptic vesicle fusion with the plasma membrane and neurotransmitter release. Here, we show that cac mutant photoreceptor terminals accumulate AV-lysosomal fusion intermediates, suggesting that Cac is necessary for the fusion of AVs with lysosomes, a poorly defined process. Loss of another subunit of the VGCC, α2δ or straightjacket (stj, causes phenotypes very similar to those caused by the loss of cac, indicating that the VGCC is required for AV-lysosomal fusion. The role of VGCC in AV-lysosomal fusion is evolutionarily conserved, as the loss of the mouse homologues, Cacna1a and Cacna2d2, also leads to autophagic defects in mice. Moreover, we find that CACNA1A is localized to the lysosomes and that loss of lysosomal Cacna1a in cerebellar cultured neurons leads to a failure of lysosomes to fuse with endosomes and autophagosomes. Finally, we show that the lysosomal CACNA1A but not the plasma-membrane resident CACNA1A is required for lysosomal fusion. In summary, we present a model in which the VGCC plays a role in autophagy by regulating the fusion of AVs with lysosomes through its calcium channel activity and hence functions in maintaining neuronal homeostasis.

  5. Brugada syndrome and calcium channel mutation in a patient with congenital deaf mutism

    Directory of Open Access Journals (Sweden)

    Uğur Canpolat

    2017-01-01

    Full Text Available To the best of our knowledge, for the first time in the literature, we described a congenitally deaf-mute patient with Brugada syndrome (BrS in whom a mutation in L-type Ca+2 channel [CACNA1C (Cav1.2α1] was identified.

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

    NARCIS (Netherlands)

    Majoie, H.J.; Baets, M.H.V. de; Renier, W.O.; Lang, B.; Vincent, A.

    2006-01-01

    OBJECTIVE: To determine the prevalence of antibodies to ion channels in patients with long standing epilepsy. BACKGROUND: Although the CNS is thought to be protected from circulating antibodies by the blood brain barrier, glutamate receptor antibodies have been reported in Rasmussen's encephalitis,

  7. Kokumi Substances, Enhancers of Basic Tastes, Induce Responses in Calcium-Sensing Receptor Expressing Taste Cells

    Science.gov (United States)

    Maruyama, Yutaka; Yasuda, Reiko; Kuroda, Motonaka; Eto, Yuzuru

    2012-01-01

    Recently, we reported that calcium-sensing receptor (CaSR) is a receptor for kokumi substances, which enhance the intensities of salty, sweet and umami tastes. Furthermore, we found that several γ-glutamyl peptides, which are CaSR agonists, are kokumi substances. In this study, we elucidated the receptor cells for kokumi substances, and their physiological properties. For this purpose, we used Calcium Green-1 loaded mouse taste cells in lingual tissue slices and confocal microscopy. Kokumi substances, applied focally around taste pores, induced an increase in the intracellular Ca2+ concentration ([Ca2+]i) in a subset of taste cells. These responses were inhibited by pretreatment with the CaSR inhibitor, NPS2143. However, the kokumi substance-induced responses did not require extracellular Ca2+. CaSR-expressing taste cells are a different subset of cells from the T1R3-expressing umami or sweet taste receptor cells. These observations indicate that CaSR-expressing taste cells are the primary detectors of kokumi substances, and that they are an independent population from the influenced basic taste receptor cells, at least in the case of sweet and umami. PMID:22511946

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

    Science.gov (United States)

    Gulati, Puja; Muthuraman, Arunachalam; Kaur, Parneet

    2015-04-01

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

  9. Differential Modulation of Rhythmic Brain Activity in Healthy Adults by a T-Type Calcium Channel Blocker: An MEG Study.

    Science.gov (United States)

    Walton, Kerry D; Maillet, Emeline L; Garcia, John; Cardozo, Timothy; Galatzer-Levy, Isaac; Llinás, Rodolfo R

    2017-01-01

    1-octanol is a therapeutic candidate for disorders involving the abnormal activation of the T-type calcium current since it blocks this current specifically. Such disorders include essential tremor and a group of neurological and psychiatric disorders resulting from thalamocortical dysrhythmia (TCD). For example, clinically, the observable phenotype in essential tremor is the tremor itself. The differential diagnostic of TCD is not based only on clinical signs and symptoms. Rather, TCD incorporates an electromagnetic biomarker, the presence of abnormal thalamocortical low frequency brain oscillations. The effect of 1-octanol on brain activity has not been tested. As a preliminary step to such a TCD study, we examined the short-term effects of a single dose of 1-octanol on resting brain activity in 32 healthy adults using magnetoencephalograpy. Visual inspection of baseline power spectra revealed that the subjects fell into those with strong low frequency activity (set 2, n = 11) and those without such activity, but dominated by an alpha peak (set 1, n = 22). Cross-validated linear discriminant analysis, using mean spectral density (MSD) in nine frequency bands as predictors, found overall that 82.5% of the subjects were classified as determined by visual inspection. The effect of 1-octanol on the MSD in narrow frequency bands differed between the two subject groups. In set 1 subjects the MSD increased in the 4.5-6.5Hz and 6.5-8.5 Hz bands. This was consistent with a widening of the alpha peak toward lower frequencies. In the set two subjects the MSD decrease in the 2.5-4.5 Hz and 4.5-6.5 Hz bands. This decreased power is consistent with the blocking effect of 1-octanol on T-type calcium channels. The subjects reported no adverse effects of the 1-octanol. Since stronger low frequency activity is characteristic of patients with TCD, 1-octanol and other T-type calcium channel blockers are good candidates for treatment of this group of disorders following a placebo

  10. Differential Modulation of Rhythmic Brain Activity in Healthy Adults by a T-Type Calcium Channel Blocker: An MEG Study

    Science.gov (United States)

    Walton, Kerry D.; Maillet, Emeline L.; Garcia, John; Cardozo, Timothy; Galatzer-Levy, Isaac; Llinás, Rodolfo R.

    2017-01-01

    1-octanol is a therapeutic candidate for disorders involving the abnormal activation of the T-type calcium current since it blocks this current specifically. Such disorders include essential tremor and a group of neurological and psychiatric disorders resulting from thalamocortical dysrhythmia (TCD). For example, clinically, the observable phenotype in essential tremor is the tremor itself. The differential diagnostic of TCD is not based only on clinical signs and symptoms. Rather, TCD incorporates an electromagnetic biomarker, the presence of abnormal thalamocortical low frequency brain oscillations. The effect of 1-octanol on brain activity has not been tested. As a preliminary step to such a TCD study, we examined the short-term effects of a single dose of 1-octanol on resting brain activity in 32 healthy adults using magnetoencephalograpy. Visual inspection of baseline power spectra revealed that the subjects fell into those with strong low frequency activity (set 2, n = 11) and those without such activity, but dominated by an alpha peak (set 1, n = 22). Cross-validated linear discriminant analysis, using mean spectral density (MSD) in nine frequency bands as predictors, found overall that 82.5% of the subjects were classified as determined by visual inspection. The effect of 1-octanol on the MSD in narrow frequency bands differed between the two subject groups. In set 1 subjects the MSD increased in the 4.5-6.5Hz and 6.5–8.5 Hz bands. This was consistent with a widening of the alpha peak toward lower frequencies. In the set two subjects the MSD decrease in the 2.5–4.5 Hz and 4.5–6.5 Hz bands. This decreased power is consistent with the blocking effect of 1-octanol on T-type calcium channels. The subjects reported no adverse effects of the 1-octanol. Since stronger low frequency activity is characteristic of patients with TCD, 1-octanol and other T-type calcium channel blockers are good candidates for treatment of this group of disorders following a

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

    Directory of Open Access Journals (Sweden)

    Niko Bekjarovski

    2013-09-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2006-01-01

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

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

    Science.gov (United States)

    Temme, Stephanie J.; Murphy, Geoffrey G.

    2017-01-01

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

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

    Science.gov (United States)

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

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

    DEFF Research Database (Denmark)

    Kroigaard, Christel; Dalsgaard, Thomas; Nielsen, Gorm

    2012-01-01

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

  16. Calcium channel alpha-2-delta-1 protein upregulation in dorsal spinal cord mediates spinal cord injury-induced neuropathic pain states.

    Science.gov (United States)

    Boroujerdi, Amin; Zeng, Jun; Sharp, Kelli; Kim, Donghyun; Steward, Oswald; Luo, Z David

    2011-03-01

    Spinal cord injury (SCI) commonly results in the development of neuropathic pain, which can dramatically impair the quality of life for SCI patients. SCI-induced neuropathic pain can be manifested as both tactile allodynia (a painful sensation to a non-noxious stimulus) and hyperalgesia (an enhanced sensation to a painful stimulus). The mechanisms underlying these pain states are poorly understood. Clinical studies have shown that gabapentin, a drug that binds to the voltage-gated calcium channel alpha-2-delta-1 subunit (Ca(v)α2δ-1) proteins is effective in the management of SCI-induced neuropathic pain. Accordingly, we hypothesized that tactile allodynia post SCI is mediated by an upregulation of Ca(v)α2δ-1 in dorsal spinal cord. To test this hypothesis, we examined whether SCI-induced dysregulation of spinal Ca(v)α2δ-1 plays a contributory role in below-level allodynia development in a rat spinal T9 contusion injury model. We found that Ca(v)α2δ-1 expression levels were significantly increased in L4-6 dorsal, but not ventral, spinal cord of SCI rats that correlated with tactile allodynia development in the hind paw plantar surface. Furthermore, both intrathecal gabapentin treatment and blocking SCI-induced Ca(v)α2δ-1 protein upregulation by intrathecal Ca(v)α2δ-1 antisense oligodeoxynucleotides could reverse tactile allodynia in SCI rats. These findings support that SCI-induced Ca(v)α2δ-1 upregulation in spinal dorsal horn is a key component in mediating below-level neuropathic pain states, and selectively targeting this pathway may provide effective pain relief for SCI patients. Spinal cord contusion injury caused increased calcium channel Ca(v)α2δ-1 subunit expression in dorsal spinal cord that contributes to neuropathic pain states. Copyright © 2010 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

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

    2010-01-01

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

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

    Science.gov (United States)

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

    2018-03-23

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

  19. A New Negative Allosteric Modulator AP14145 for the Study of Small Conductance Calcium-Activated Potassium Channels

    DEFF Research Database (Denmark)

    Simo Vicens, Rafel; Kirchhoff, Jeppe Egedal; Dolce, Bernardo

    2017-01-01

    ) prolongation in anaesthetised rats and a beam walk test was performed in mice to determine acute CNS related effects of the drug. Key results: AP14145 was found to be an equipotent negative allosteric modulator of KCa2.2 and KCa2.3 channels (IC50 = 1.1 ± 0.3 μM L-1). The presence of AP14145 (10 μM L-1......) increased the EC50 of Ca2+ on KCa2.3 from 0.36 ± 0.02 μM L-1 to 1.2 ± 0.1 μM L-1. The inhibitory effect strongly depended on two amino acids, S508 and A533. AP14145 concentration-dependently prolonged AERP in rats. Moreover, AP14145 (10 mg kg-1) did not trigger any apparent CNS effects in mice. Conclusion...... and implications: AP14145 is a negative allosteric modulator of KCa2.2 and KCa2.3 that shifts the calcium dependence of channel activation, an effect strongly dependent on two identified amino acids. AP14145 prolongs AERP in rats and does not trigger any acute CNS effects in mice. The understanding of how KCa2...

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2006-01-01

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

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

    International Nuclear Information System (INIS)

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

    2006-01-01

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

  2. Micromolar-Affinity Benzodiazepine Receptors Regulate Voltage-Sensitive Calcium Channels in Nerve Terminal Preparations

    Science.gov (United States)

    Taft, William C.; Delorenzo, Robert J.

    1984-05-01

    Benzodiazepines in micromolar concentrations significantly inhibit depolarization-sensitive Ca2+ uptake in intact nerve-terminal preparations. Benzodiazepine inhibition of Ca2+ uptake is concentration dependent and stereospecific. Micromolar-affinity benzodiazepine receptors have been identified and characterized in brain membrane and shown to be distinct from nanomolar-affinity benzodiazepine receptors. Evidence is presented that micromolar, and not nanomolar, benzodiazepine binding sites mediate benzodiazepine inhibition of Ca2+ uptake. Irreversible binding to micromolar benzodiazepine binding sites also irreversibly blocked depolarization-dependent Ca2+ uptake in synaptosomes, indicating that these compounds may represent a useful marker for identifying the molecular components of Ca2+ channels in brain. Characterization of benzodiazepine inhibition of Ca2+ uptake demonstrates that these drugs function as Ca2+ channel antagonists, because benzodiazepines effectively blocked voltage-sensitive Ca2+ uptake inhibited by Mn2+, Co2+, verapamil, nitrendipine, and nimodipine. These results indicate that micromolar benzodiazepine binding sites regulate voltage-sensitive Ca2+ channels in brain membrane and suggest that some of the neuronal stabilizing effects of micromolar benzodiazepine receptors may be mediated by the regulation of Ca2+ conductance.

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

    Science.gov (United States)

    Sukiasyan, N; Hultborn, H; Zhang, M

    2009-03-03

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

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

    Science.gov (United States)

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

    2010-08-15

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

  5. Corticolimbic expression of TRPC4 and TRPC5 channels in the rodent brain.

    Directory of Open Access Journals (Sweden)

    Melissa A Fowler

    2007-06-01

    Full Text Available The canonical transient receptor potential (TRPC channels are a family of non-selective cation channels that are activated by increases in intracellular Ca(2+ and G(q/phospholipase C-coupled receptors. We used quantitative real-time PCR, in situ hybridization, immunoblots and patch-clamp recording from several brain regions to examine the expression of the predominant TRPC channels in the rodent brain. Quantitative real-time PCR of the seven TRPC channels in the rodent brain revealed that TRPC4 and TRPC5 channels were the predominant TRPC subtypes in the adult rat brain. In situ hybridization histochemistry and immunoblotting further resolved a dense corticolimbic expression of the TRPC4 and TRPC5 channels. Total protein expression of HIP TRPC4 and 5 proteins increased throughout development and peaked late in adulthood (6-9 weeks. In adults, TRPC4 expression was high throughout the frontal cortex, lateral septum (LS, pyramidal cell layer of the hippocampus (HIP, dentate gyrus (DG, and ventral subiculum (vSUB. TRPC5 was highly expressed in the frontal cortex, pyramidal cell layer of the HIP, DG, and hypothalamus. Detailed examination of frontal cortical layer mRNA expression indicated TRPC4 mRNA is distributed throughout layers 2-6 of the prefrontal cortex (PFC, motor cortex (MCx, and somatosensory cortex (SCx. TRPC5 mRNA expression was concentrated specifically in the deep layers 5/6 and superficial layers 2/3 of the PFC and anterior cingulate. Patch-clamp recording indicated a strong metabotropic glutamate-activated cation current-mediated depolarization that was dependent on intracellular Ca(2+and inhibited by protein kinase C in brain regions associated with dense TRPC4 or 5 expression and absent in regions lacking TRPC4 and 5 expression. Overall, the dense corticolimbic expression pattern suggests that these Gq/PLC coupled nonselective cation channels may be involved in learning, memory, and goal-directed behaviors.

  6. Calcium-Sensing Receptor Tumor Expression and Lethal Prostate Cancer Progression.

    Science.gov (United States)

    Ahearn, Thomas U; Tchrakian, Nairi; Wilson, Kathryn M; Lis, Rosina; Nuttall, Elizabeth; Sesso, Howard D; Loda, Massimo; Giovannucci, Edward; Mucci, Lorelei A; Finn, Stephen; Shui, Irene M

    2016-06-01

    Prostate cancer metastases preferentially target bone, and the calcium-sensing receptor (CaSR) may play a role in promoting this metastatic progression. We evaluated the association of prostate tumor CaSR expression with lethal prostate cancer. A validated CaSR immunohistochemistry assay was performed on tumor tissue microarrays. Vitamin D receptor (VDR) expression and phosphatase and tensin homolog tumor status were previously assessed in a subset of cases by immunohistochemistry. Cox proportional hazards models adjusting for age and body mass index at diagnosis, Gleason grade, and pathological tumor node metastasis stage were used to estimate hazard ratios (HR) and 95% confidence intervals (CI) for the association of CaSR expression with lethal prostate cancer. The investigation was conducted in the Health Professionals Follow-up Study and Physicians' Health Study. We studied 1241 incident prostate cancer cases diagnosed between 1983 and 2009. Participants were followed up or cancer-specific mortality or development of metastatic disease. On average, men were followed up 13.6 years, during which there were 83 lethal events. High CaSR expression was associated with lethal prostate cancer independent of clinical and pathological variables (HR 2.0; 95% CI 1.2-3.3). Additionally, there was evidence of effect modification by VDR expression; CaSR was associated with lethal progression among men with low tumor VDR expression (HR 3.2; 95% CI 1.4-7.3) but not in cases with high tumor VDR expression (HR 0.8; 95% CI 0.2-3.0). Tumor CaSR expression is associated with an increased risk of lethal prostate cancer, particularly in tumors with low VDR expression. These results support further investigating the mechanism linking CaSR with metastases.

  7. Ectopic expression of the calcium-binding protein parvalbumin in mouse liver endothelial cells

    DEFF Research Database (Denmark)

    Castillo, M B; Berchtold, M W; Rülicke, T

    1997-01-01

    To elucidate the physiological role of the Ca2+ binding protein parvalbumin, we have generated transgenic mice carrying the full-length complementary DNA (cDNA) of rat parvalbumin under the control of the heavy-metal inducible metallothionein IIA promoter. Immunohistochemical and biochemical...... methods have been used to detect the presence of ectopic parvalbumin expression in different tissues. Here we show the expression of parvalbumin in endothelial cells lining the liver sinusoids in situ and after isolation in vitro. The hemodynamic effects of endothelin 1, a peptide hormone mediating potent...... vasoconstriction via calcium signalling, were investigated in the mouse liver perfused in situ. Vasoconstriction, thought to be mediated by the Ito cell, was not affected in the transgenic animals, whereas microvascular exchange, probed with the multiple indicator dilution technique, was markedly decreased...

  8. Potential role of melastatin-related transient receptor potential cation channel subfamily M gene expression in the pathogenesis of urinary bladder cancer.

    Science.gov (United States)

    Ceylan, Gülay Güleç; Önalan, Ebru Etem; Kuloğlu, Tuncay; Aydoğ, Gülten; Keleş, İbrahim; Tonyali, Şenol; Ceylan, Cavit

    2016-12-01

    Urinary bladder cancer is one of the most common malignancies of the urinary tract. Ion channels and calcium homeostasis are involved in almost all basic cellular mechanisms. The transient receptor potential cation channel subfamily M (TRPM) takes its name from the melastatin protein, which is classified as potential tumor suppressor. To the best of our knowledge, there have been no previous studies in the literature investigating the role of these ion channels in bladder cancer. The present study aimed to determine whether bladder cancer is associated with mRNA expression levels of TRPM ion channel genes, and whether there is the potential to conduct further studies to establish novel treatment modalities. The present study included a total of 47 subjects, of whom 40 were bladder cancer patients and 7 were controls. Following the histopathological evaluation for bladder carcinoma, the mRNA and protein expression of TRPM were examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry in tumor and normal tissues, in order to determine whether there is a difference in the expression of these channels in tumor and normal tissues. Immunoreactivity for TRPM2, TRPM4, TRPM7 and TRPM8 was observed in epithelial bladder cells in the two groups. RT-qPCR revealed a significant increase in TRPM7 expression in bladder cancer tissue compared to the controls (healthy bladder tissue), whereas no differences in TRPM2 or TRPM4 expression levels were observed. There were significant reductions in the expression levels of TRPM5 and TRPM8 in bladder cancer tissues. In the present study, the effects of TRP ion channels on the formation of bladder cancer was investigated. This study is instructive for TRPM2, TRPM4, TRPM5, TRPM7 and TRPM8 and their therapeutic role in bladder cancer. The results support the fact that these gens can be novel targets and can also be tested for during the treatment of bladder cancer.

  9. Calcium-Release Channels in Paramecium. Genomic Expansion, Differential Positioning and Partial Transcriptional Elimination

    Science.gov (United States)

    Ladenburger, Eva-Maria; Plattner, Helmut

    2011-01-01

    The release of Ca2+ from internal stores is a major source of signal Ca2+ in almost all cell types. The internal Ca2+ pools are activated via two main families of intracellular Ca2+-release channels, the ryanodine and the inositol 1,4,5-trisphosphate (InsP3) receptors. Among multicellular organisms these channel types are ubiquitous, whereas in most unicellular eukaryotes the identification of orthologs is impaired probably due to evolutionary sequence divergence. However, the ciliated protozoan Paramecium allowed us to prognosticate six groups, with a total of 34 genes, encoding proteins with characteristics typical of InsP3 and ryanodine receptors by BLAST search of the Paramecium database. We here report that these Ca2+-release channels may display all or only some of the characteristics of canonical InsP3 and ryanodine receptors. In all cases, prediction methods indicate the presence of six trans-membrane regions in the C-terminal domains, thus corresponding to canonical InsP3 receptors, while a sequence homologous to the InsP3-binding domain is present only in some types. Only two types have been analyzed in detail previously. We now show, by using antibodies and eventually by green fluorescent protein labeling, that the members of all six groups localize to distinct organelles known to participate in vesicle trafficking and, thus, may provide Ca2+ for local membrane-membrane interactions. Whole genome duplication can explain radiation within the six groups. Comparative and evolutionary evaluation suggests derivation from a common ancestor of canonical InsP3 and ryanodine receptors. With one group we could ascertain, to our knowledge for the first time, aberrant splicing in one thoroughly analyzed Paramecium gene. This yields truncated forms and, thus, may indicate a way to pseudogene formation. No comparable analysis is available for any other, free-living or parasitic/pathogenic protozoan. PMID:22102876

  10. Calcium-release channels in paramecium. Genomic expansion, differential positioning and partial transcriptional elimination.

    Directory of Open Access Journals (Sweden)

    Eva-Maria Ladenburger

    Full Text Available The release of Ca²⁺ from internal stores is a major source of signal Ca²⁺ in almost all cell types. The internal Ca²⁺ pools are activated via two main families of intracellular Ca²⁺-release channels, the ryanodine and the inositol 1,4,5-trisphosphate (InsP₃ receptors. Among multicellular organisms these channel types are ubiquitous, whereas in most unicellular eukaryotes the identification of orthologs is impaired probably due to evolutionary sequence divergence. However, the ciliated protozoan Paramecium allowed us to prognosticate six groups, with a total of 34 genes, encoding proteins with characteristics typical of InsP₃ and ryanodine receptors by BLAST search of the Paramecium database. We here report that these Ca²⁺-release channels may display all or only some of the characteristics of canonical InsP₃ and ryanodine receptors. In all cases, prediction methods indicate the presence of six trans-membrane regions in the C-terminal domains, thus corresponding to canonical InsP₃ receptors, while a sequence homologous to the InsP₃-binding domain is present only in some types. Only two types have been analyzed in detail previously. We now show, by using antibodies and eventually by green fluorescent protein labeling, that the members of all six groups localize to distinct organelles known to participate in vesicle trafficking and, thus, may provide Ca²⁺ for local membrane-membrane interactions. Whole genome duplication can explain radiation within the six groups. Comparative and evolutionary evaluation suggests derivation from a common ancestor of canonical InsP₃ and ryanodine receptors. With one group we could ascertain, to our knowledge for the first time, aberrant splicing in one thoroughly analyzed Paramecium gene. This yields truncated forms and, thus, may indicate a way to pseudogene formation. No comparable analysis is available for any other, free-living or parasitic/pathogenic protozoan.

  11. Inhibitory effects and mechanism of dihydroberberine on hERG channels expressed in HEK293 cells.

    Directory of Open Access Journals (Sweden)

    Dahai Yu

    Full Text Available The human ether-a-go-go-related gene (hERG potassium channel conducts rapid delayed rectifier potassium currents (IKr and contributes to phase III cardiac action potential repolarization. Drugs inhibit hERG channels by binding to aromatic residues in hERG helixes. Berberine (BBR has multiple actions, and its hydrogenated derivative dihydroberberine (DHB is a potential candidate for developing new drugs. Previous studies have demonstrated that BBR blocks hERG channels and prolongs action potential duration (APD. Our present study aimed to investigate the effects and mechanism of DHB on hERG channels. Protein expression and the hERG current were analyzed using western blotting and patch-clamp, respectively. DHB inhibited the hERG current concentration-dependently after instantaneous perfusion, accelerated channel inactivation by directly binding tyrosine (Tyr652 and phenylalanine (Phe656, and decreased mature (155-kDa and simultaneously increased immature (135-kDa hERG expression, respectively. This suggests disruption of forward trafficking of hERG channels. Besides, DHB remarkably reduced heat shock protein 90 (Hsp90 expression and its interaction with hERG, indicating that DHB disrupted hERG trafficking by impairing channel folding. Meanwhie, DHB enhanced the expression of cleaved activating transcription factor-6 (ATF-6, a biomarker of unfolded protein response (UPR. Expression of calnexin and calreticulin, chaperones activated by ATF-6 to facilitate channel folding, were also increased, which indicating UPR activation. Additionally, the degradation rate of mature 155-kDa hERG increased following DHB exposure. In conclusion, we demonstrated that DHB acutely blocked hERG channels by binding the aromatic Tyr652 and Phe656. DHB may decrease hERG plasma membrane expression through two pathways involving disruption of forward trafficking of immature hERG channels and enhanced degradation of mature hERG channels. Furthermore, forward trafficking was

  12. Substituted 2-Acylaminocycloalkylthiophene-3-carboxylic Acid Arylamides as Inhibitors of the Calcium-Activated Chloride Channel Transmembrane Protein 16A (TMEM16A).

    Science.gov (United States)

    Truong, Eric C; Phuan, Puay W; Reggi, Amanda L; Ferrera, Loretta; Galietta, Luis J V; Levy, Sarah E; Moises, Alannah C; Cil, Onur; Diez-Cecilia, Elena; Lee, Sujin; Verkman, Alan S; Anderson, Marc O

    2017-06-08

    Transmembrane protein 16A (TMEM16A), also called anoctamin 1 (ANO1), is a calcium-activated chloride channel expressed widely mammalian cells, including epithelia, vascular smooth muscle tissue, electrically excitable cells, and some tumors. TMEM16A inhibitors have been proposed for treatment of disorders of epithelial fluid and mucus secretion, hypertension, asthma, and possibly cancer. Herein we report, by screening, the discovery of 2-acylaminocycloalkylthiophene-3-carboxylic acid arylamides (AACTs) as inhibitors of TMEM16A and analysis of 48 synthesized analogs (10ab-10bw) of the original AACT compound (10aa). Structure-activity studies indicated the importance of benzene substituted as 2- or 4-methyl, or 4-fluoro, and defined the significance of thiophene substituents and size of the cycloalkylthiophene core. The most potent compound (10bm), which contains an unusual bromodifluoroacetamide at the thiophene 2-position, had IC 50 of ∼30 nM, ∼3.6-fold more potent than the most potent previously reported TMEM16A inhibitor 4 (Ani9), and >10-fold improved metabolic stability. Direct and reversible inhibition of TMEM16A by 10bm was demonstrated by patch-clamp analysis. AACTs may be useful as pharmacological tools to study TMEM16A function and as potential drug development candidates.

  13. High Glucose Represses hERG K+ Channel Expression through Trafficking Inhibition

    Directory of Open Access Journals (Sweden)

    Yuan-Qi Shi

    2015-08-01

    Full Text Available Background/Aims: Abnormal QT prolongation is the most prominent cardiac electrical disturbance in patients with diabetes mellitus (DM. It is well known that the human ether-ago-go-related gene (hERG controls the rapid delayed rectifier K+ current (IKr in cardiac cells. The expression of the hERG channel is severely down-regulated in diabetic hearts, and this down-regulation is a critical contributor to the slowing of repolarization and QT prolongation. However, the intracellular mechanisms underlying the diabetes-induced hERG deficiency remain unknown. Methods: The expression of the hERG channel was assessed via western blot analysis, and the hERG current was detected with a patch-clamp technique. Results: The results of our study revealed that the expression of the hERG protein and the hERG current were substantially decreased in high-glucose-treated hERG-HEK cells. Moreover, we demonstrated that the high-glucose-mediated damage to the hERG channel depended on the down-regulation of protein levels but not the alteration of channel kinetics. These discoveries indicated that high glucose likely disrupted hERG channel trafficking. From the western blot and immunoprecipitation analyses, we found that high glucose induced trafficking inhibition through an effect on the expression of Hsp90 and its interaction with hERG. Furthermore, the high-glucose-induced inhibition of hERG channel trafficking could activate the unfolded protein response (UPR by up-regulating the expression levels of activating transcription factor-6 (ATF-6 and the ER chaperone protein calnexin. In addition, we demonstrated that 100 nM insulin up-regulated the expression of the hERG channel and rescued the hERG channel repression caused by high glucose. Conclusion: The results of our study provide the first evidence of a high-glucose-induced hERG channel deficiency resulting from the inhibition of channel trafficking. Furthermore, insulin promotes the expression of the hERG channel

  14. A Unified Channel Charges Expression for Analytic MOSFET Modeling

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    Hugues Murray

    2012-01-01

    Full Text Available Based on a 1D Poissons equation resolution, we present an analytic model of inversion charges allowing calculation of the drain current and transconductance in the Metal Oxide Semiconductor Field Effect Transistor. The drain current and transconductance are described by analytical functions including mobility corrections and short channel effects (CLM, DIBL. The comparison with the Pao-Sah integral shows excellent accuracy of the model in all inversion modes from strong to weak inversion in submicronics MOSFET. All calculations are encoded with a simple C program and give instantaneous results that provide an efficient tool for microelectronics users.

  15. Synaptotagmin-7 is an asynchronous calcium sensor for synaptic transmission in neurons expressing SNAP-23.

    Directory of Open Access Journals (Sweden)

    Jens P Weber

    Full Text Available Synchronization of neurotransmitter release with the presynaptic action potential is essential for maintaining fidelity of information transfer in the central nervous system. However, synchronous release is frequently accompanied by an asynchronous release component that builds up during repetitive stimulation, and can even play a dominant role in some synapses. Here, we show that substitution of SNAP-23 for SNAP-25 in mouse autaptic glutamatergic hippocampal neurons results in asynchronous release and a higher frequency of spontaneous release events (mEPSCs. Use of neurons from double-knock-out (SNAP-25, synaptotagmin-7 mice in combination with viral transduction showed that SNAP-23-driven release is triggered by endogenous synaptotagmin-7. In the absence of synaptotagmin-7 release became even more asynchronous, and the spontaneous release rate increased even more, indicating that synaptotagmin-7 acts to synchronize release and suppress spontaneous release. However, compared to synaptotagmin-1, synaptotagmin-7 is a both leaky and asynchronous calcium sensor. In the presence of SNAP-25, consequences of the elimination of synaptotagmin-7 were small or absent, indicating that the protein pairs SNAP-25/synaptotagmin-1 and SNAP-23/synaptotagmin-7 might act as mutually exclusive calcium sensors. Expression of fusion proteins between pHluorin (pH-sensitive GFP and synaptotagmin-1 or -7 showed that vesicles that fuse using the SNAP-23/synaptotagmin-7 combination contained synaptotagmin-1, while synaptotagmin-7 barely displayed activity-dependent trafficking between vesicle and plasma membrane, implying that it acts as a plasma membrane calcium sensor. Overall, these findings support the idea of alternative syt∶SNARE combinations driving release with different kinetics and fidelity.

  16. An amplified promoter system for targeted expression of calcium indicator proteins in the cerebellar cortex

    Directory of Open Access Journals (Sweden)

    Bernd eKuhn

    2012-07-01

    Full Text Available Recording of identified neuronal network activity using genetically encoded calcium indicators (GECIs requires labeling that is cell type-specific and bright enough for the detection of functional signals. However, specificity and strong expression are often not achievable using the same promoter. Here we present a combinatorial approach for targeted expression and single-cell-level quantification in which a weak promoter is used to drive trans-amplification under a strong general promoter. We demonstrated this approach using recombinant adeno-associated viruses (rAAVs to deliver the sequence of the GECI D3cpv in the mouse cerebellar cortex. Direct expression under the human synapsin promoter (hSYN led to high levels of expression (50-100 µM in five interneuron types of the cerebellar cortex but not in Purkinje cells (PCs (≤10 μM, yielding sufficient contrast to allow functional signals to be recorded from somata and processes in awake animals using two-photon microscopy. When the hSYN promoter was used to drive expression of the tetracycline transactivator (tTA, a second rAAV containing the bidirectional TET promoter (Ptetbi could drive strong D3cpv expression in PCs (10-300 µM, enough to allow reliable complex spike detection in the dendritic arbor. An amplified approach should be of use in monitoring neural processing in selected cell types and boosting expression of optogenetic probes. Additionally, we overcome cell toxicity associated with rAAV injection and/or local GECI overexpression by combining the virus injection with systemic pre-injection of hyperosmotic D-mannitol, and by this double the time window for functional imaging.

  17. Expression of calcium binding protein S100 A7 (psoriasin) in laryngeal carcinoma.

    Science.gov (United States)

    Tiveron, Rogério Costa; de Freitas, Luiz Carlos Conti; Figueiredo, David L; Serafini, Luciano N; Mamede, Rui Celso Martins; Zago, Marco A

    2012-01-01

    Many studies have reported increased expression of S100 A7 (psoriasin) in neoplastic lesions. Among them are studies on breast carcinoma, bladder squamous cell carcinoma, skin tumors and oral cavity squamous cell carcinoma. The expression of S100 A7 has not been described for laryngeal cancer. This study aims to identify the expression of the calcium-binding protein S100 A7 and its correlation with squamous cell carcinomas of the larynx. Specimens from 63 patients were submitted to immunohistochemistry testing with antibody S100 A7. Results were classified and compared. The group with highly differentiated tumors had the highest treatment failure scores. Moderately differentiated tumors had higher treatment failure scores than poorly differentiated tumors. Higher scores were predominantly seen on stages I and II in moderately differentiated tumors, whereas score distribution was more homogeneous in advanced stage disease (III and IV). Regarding failure in treatment, the group scoring zero (3/4 complications: 75%) differed significantly from the remaining groups (13/59: 22%). S100 A7 marker was expressed in 93.7% of laryngeal cancer cases, with higher positive correlation rates in more differentiated tumors and significantly lower rates of treatment failure. Scores had no impact on survival rates.

  18. Broad neutralization of calcium-permeable amyloid pore channels with a chimeric Alzheimer/Parkinson peptide targeting brain gangliosides.

    Science.gov (United States)

    Di Scala, Coralie; Yahi, Nouara; Flores, Alessandra; Boutemeur, Sonia; Kourdougli, Nazim; Chahinian, Henri; Fantini, Jacques

    2016-02-01

    Growing evidence supports a role for brain gangliosides in the pathogenesis of neurodegenerative diseases including Alzheimer's and Parkinson's. Recently we deciphered the ganglioside-recognition code controlling specific ganglioside binding to Alzheimer's β-amyloid (Aβ1-42) peptide and Parkinson's disease-associated protein α-synuclein. Cracking this code allowed us to engineer a short chimeric Aβ/α-synuclein peptide that recognizes all brain gangliosides. Here we show that ganglioside-deprived neural cells do no longer sustain the formation of zinc-sensitive amyloid pore channels induced by either Aβ1-42 or α-synuclein, as assessed by single-cell Ca(2+) fluorescence microscopy. Thus, amyloid channel formation, now considered a key step in neurodegeneration, is a ganglioside-dependent process. Nanomolar concentrations of chimeric peptide competitively inhibited amyloid pore formation induced by Aβ1-42 or α-synuclein in cultured neural cells. Moreover, this peptide abrogated the intracellular calcium increases induced by Parkinson's-associated mutant forms of α-synuclein (A30P, E46K and A53T). The chimeric peptide also prevented the deleterious effects of Aβ1-42 on synaptic vesicle trafficking and decreased the Aβ1-42-induced impairment of spontaneous activity in rat hippocampal slices. Taken together, these data show that the chimeric peptide has broad anti-amyloid pore activity, suggesting that a common therapeutic strategy based on the prevention of amyloid-ganglioside interactions is a reachable goal for both Alzheimer's and Parkinson's diseases. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Site of action of calcium channel blockers in inhibiting endogenous pyrogen fever in rats.

    Science.gov (United States)

    Stitt, J T; Shimada, S G

    1991-09-01

    We have demonstrated that the Ca2+ channel blocker verapamil, administered intravenously, exerts an antipyretic effect on the febrile responses of rats to intravenously injected endogenous pyrogen (EP). We have also shown that the same intravenous dose of verapamil is ineffective in blocking fevers induced by the microinjection of exogenous prostaglandin E (PGE) into the organum vasculosum laminae terminalis (OVLT) of rats. Experiments were conducted to determine whether the site of this verapamil antipyresis was in the OVLT itself. The febrile responses of six male Sprague-Dawley rats to EP were determined at thermoneutrality. Verapamil (10 micrograms/rat) was microinjected directly into the OVLT, and the febrile responses to the EP dose were redetermined 15-30 min later. In every case the EP fevers were attenuated after verapamil pretreatment. Intra-OVLT injections of verapamil alone were without effect on body temperature. When the same dose of verapamil was injected into the OVLT 15 min before the injection of PGE into the same site, it had no effect on the ensuing PGE-induced fever. In view of the fact that less than 1/250th of the effective systemic dose of verapamil, when injected into the OVLT, was equally effective in blocking the EP fevers, we conclude that verapamil acts within the OVLT to block fever rather than peripherally. Furthermore, because verapamil administered into the OVLT does not block PGE fevers, it is unlikely that PGE produces fever by acting as a Ca2+ ionophore on hypothalamic neurons.

  20. Basic calcium phosphate crystal-induced Egr-1 expression stimulates mitogenesis in human fibroblasts

    International Nuclear Information System (INIS)

    Zeng, Xiao R.; Sun Yubo; Wenger, Leonor; Cheung, Herman S.

    2005-01-01

    Previously, we have reported that basic calcium phosphate (BCP) crystals stimulate mitogenesis and synthesis of matrix metalloproteinases in cultured human foreskin and synovial fibroblasts. However, the detailed mechanisms involved are still unclear. In the present study, using RT-PCR and Egr-1 promoter analysis we showed that BCP crystals could stimulate early growth response gene Egr-1 transcription through a PKCα-dependent p44/p42 MAPK pathway. Using a retrovirus gene expression system (Clontech) to overexpress Egr-1 in human fibroblast BJ-1 cells resulted in promotion of mitogenesis measured either by MTT cell proliferation analysis or by direct cell counting. The results demonstrate that Egr-1 may play a key role in mediating BCP crystal-induced synovial fibroblast mitogenesis

  1. THE ROLE OF S-AMLODIPINE IN ARTERIAL HYPERTENSION THERAPY WITH COMBINATION OF CALCIUM CHANNEL BLOCKERS AND BETA-BLOCKERS

    Directory of Open Access Journals (Sweden)

    M. A. Maksimova

    2013-01-01

    Full Text Available Aim. To study efficacy and safety of calcium channel blocker, S-amlodipine, in combination with β-blocker, atenolol, in patients with arterial hypertension (HT 1-2 degree com- pared to fixed combination of racemic amlodipine and atenolol.Material and methods. Patients (n=31, 7 men and 24 women with HT 1–2 degree were included into the study. The patients were randomized into two groups by the com- binations sequence. Treatment with each combination lasted 4 weeks. Office blood pressure (BP was assessed at baseline and at the end of the treatment periods, possible side effects were registered.Results. All patients completed the study. Both combination of S-amlodipine+atenolol and fixed combination of racemic amlodipine+atenolol reduced systolic (in average, -15.9 and -12.7 mm Hg, respectively and diastolic (in average, -7.3 and -5.3 mmHg, respectively BP significantly. Heart rate also decreased during therapy (in average, -3 and -4 bt/min, respectively. The differences between combinations BP and heart rate effects were not significant. 8 and 16 adverse events were registered during S-amlodipine+atenolol and racemic amlodipine+atenolol therapies, respectively Conclusion. Combination of S-amlodipine+atenolol, as well as combination of racemic amlodipine+atenolol are effective in the treatment of patients with HT 1-2 degree, however combination with S-amlodipine has less number of adverse events.

  2. Roselle Polyphenols Exert Potent Negative Inotropic Effects via Modulation of Intracellular Calcium Regulatory Channels in Isolated Rat Heart.

    Science.gov (United States)

    Lim, Yi-Cheng; Budin, Siti Balkis; Othman, Faizah; Latip, Jalifah; Zainalabidin, Satirah

    2017-07-01

    Roselle (Hibiscus sabdariffa Linn.) calyces have demonstrated propitious cardioprotective effects in animal and clinical studies; however, little is known about its action on cardiac mechanical function. This study was undertaken to investigate direct action of roselle polyphenols (RP) on cardiac function in Langendorff-perfused rat hearts. We utilized RP extract which consists of 12 flavonoids and seven phenolic acids (as shown by HPLC profiling) and has a safe concentration range between 125 and 500 μg/ml in this study. Direct perfusion of RP in concentration-dependent manner lowered systolic function of the heart as shown by lowered LVDP and dP/dt max , suggesting a negative inotropic effect. RP also reduced heart rate (negative chronotropic action) while simultaneously increasing maximal velocity of relaxation (positive lusitropic action). Conversely, RP perfusion increased coronary pressure, an indicator for improvement in coronary blood flow. Inotropic responses elicited by pharmacological agonists for L-type Ca 2+ channel [(±)-Bay K 8644], ryanodine receptor (4-chloro-m-cresol), β-adrenergic receptor (isoproterenol) and SERCA blocker (thapsigargin) were all abolished by RP. In conclusion, RP elicits negative inotropic, negative chronotropic and positive lusitropic responses by possibly modulating calcium entry, release and reuptake in the heart. Our findings have shown the potential use of RP as a therapeutic agent to treat conditions like arrhythmia.

  3. A Rare Case of Cerebellar Ataxia Due to Voltage-Gated Calcium Channel and Glutamic Acid Decarboxylase Autoantibodies.

    Science.gov (United States)

    Annunziata, Giuseppe; Lobo, Pamela; Carbuccia, Cristian

    2017-11-27

    BACKGROUND Autoimmune cerebellar ataxia can be paraneoplastic in nature or can occasionally present without evidence of an ongoing malignancy. The detection of specific autoantibodies has been statistically linked to different etiologies. CASE REPORT A 55-year-old African-American woman with hypertension and a past history of morbid obesity and uncontrolled diabetes status post gastric bypass four years prior to the visit (with significantly improved body mass index and hemoglobin A1c controlled at the time of the clinical encounter) presented to the office complaining of gradual onset of unsteadiness and recurrent falls for the past three years, as well as difficulties coordinating routine daily activities. The neurologic exam showed moderate dysarthria and ataxic gait with bilateral dysmetria and positive Romberg test. Routine laboratory test results were only remarkable for a mild elevation of erythrocyte sedimentation rate, and most laboratory and imaging tests for common causes of ataxia failed to demonstrate an etiology. Upon further workup, evidence of anti-voltage-gated calcium channel and anti-glutamic acid decarboxylase antibody was demonstrated. She was then treated with intravenous immunoglobulins with remarkable clinical improvement. CONCLUSIONS We present a case of antibody-mediated ataxia not associated with malignancy. While ataxia is rarely related to autoantibodies, in such cases it is critical to understand the etiology of this disabling condition in order to treat it correctly. Clinicians should be aware of the possible association with specific autoantibodies and the necessity to rule out an occult malignancy in such cases.

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

    Science.gov (United States)

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

    2012-01-01

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

  5. Mechanistic Exploration of Cancer Stem Cell Marker Voltage-Dependent Calcium Channel α2δ1 Subunit-mediated Chemotherapy Resistance in Small-Cell Lung Cancer.

    Science.gov (United States)

    Yu, Jiangyong; Wang, Shuhang; Zhao, Wei; Duan, Jianchun; Wang, Zhijie; Chen, Hanxiao; Tian, Yanhua; Wang, Di; Zhao, Jun; An, Tongtong; Bai, Hua; Wu, Meina; Wang, Jie

    2018-05-01

    Purpose: Chemoresistance in small-cell lung cancer (SCLC) is reportedly attributed to the existence of resistant cancer stem cells (CSC). Studies involving CSC-specific markers and related mechanisms in SCLC remain limited. This study explored the role of the voltage-dependent calcium channel α2δ1 subunit as a CSC marker in chemoresistance of SCLC, and explored the potential mechanisms of α2δ1-mediated chemoresistance and strategies of overcoming the resistance. Experimental Design: α2δ1-positive cells were identified and isolated from SCLC cell lines and patient-derived xenograft (PDX) models, and CSC-like properties were subsequently verified. Transcriptome sequencing and Western blotting were carried out to identify pathways involved in α2δ1-mediated chemoresistance in SCLC. In addition, possible interventions to overcome α2δ1-mediated chemoresistance were examined. Results: Different proportions of α2δ1 + cells were identified in SCLC cell lines and PDX models. α2δ1 + cells exhibited CSC-like properties (self-renewal, tumorigenic, differentiation potential, and high expression of genes related to CSCs and drug resistance). Chemotherapy induced the enrichment of α2δ1 + cells instead of CD133 + cells in PDXs, and an increased proportion of α2δ1 + cells corresponded to increased chemoresistance. Activation and overexpression of ERK in the α2δ1-positive H1048 cell line was identified at the protein level. mAb 1B50-1 was observed to improve the efficacy of chemotherapy and delay relapse as maintenance therapy in PDX models. Conclusions: SCLC cells expressing α2δ1 demonstrated CSC-like properties, and may contribute to chemoresistance. ERK may play a key role in α2δ1-mediated chemoresistance. mAb 1B50-1 may serve as a potential anti-SCLC drug. Clin Cancer Res; 24(9); 2148-58. ©2018 AACR . ©2018 American Association for Cancer Research.

  6. Ion channel gene expressions in infertile men: A case-control study

    Directory of Open Access Journals (Sweden)

    Serkan Carkci

    2017-12-01

    Full Text Available Background: Infertility is described as not receiving pregnancy despite unprotected and regular sexual intercourse in a 1 yr period. It is detected by 15% of the couples. Male and female factor in the etiology may be detected in similar rates. Objective: The present study aims to investigate ion channel gene expression in semen samples of infertile male compared with fertile men. Materials and Methods: A total of 150 men who applied to the urology clinic due to infertility were divided into five equal groups: asthenozoospermia, oligozoospermia, oligoasthenoteratozoospermia, teratozoospermia, and normozoospermia (control. All paticipants were evaluated with Cation Channel Spermia (CatSper 1, 2, 3, 4, Proton Voltage Gated Ion Channel1 (Hv1, Potassium Channel Subfamily U1 (KCNU1, and transmembrane protein (TMEM16A gene expression in semen samples. Results: “CatSper1, 4, HV1, KCNU1, and TMEM16A gene expression were detected higher in the oligozoospermia group compared to the controls. CatSper1, 2, 3, 4, KCNU1, and TMEM16A gene expression in the asthenozoospermia group and CatSper1, 2, 3, 4, KCNU1, and TMEM16A gene expression in the teratozoospermia group were detected lower compared to the controls. CatSper1, 4, HV1, and TMEM16A gen expression were higher in the oligoasthenoteratozoospermia men than the controls while CatSper3 gen expression was detected as lower.” Conclusion: It was detected that these ion channels have an effect on sperm progressive motility and morphology. It may be considered that mutations in these ion channels may result in infertility

  7. The effects of calcium on the expression of genes involved in ...

    African Journals Online (AJOL)

    Calcium regulation of the genes involved in ethylene biosynthesis and ethylene receptors in flower abscission zones (AZ) of wild-type tomato (Lycopersicon esculentum Mill.) was investigated in this study. Calcium treatment delayed abscission of pedicel explants. However, verapamil (VP, calcium inhibitor) treatments ...

  8. Cholesterol regulates HERG K+ channel activation by increasing phospholipase C β1 expression.

    Science.gov (United States)

    Chun, Yoon Sun; Oh, Hyun Geun; Park, Myoung Kyu; Cho, Hana; Chung, Sungkwon

    2013-01-01

    Human ether-a-go-go-related gene (HERG) K(+) channel underlies the rapidly activating delayed rectifier K(+) conductance (IKr) during normal cardiac repolarization. Also, it may regulate excitability in many neuronal cells. Recently, we showed that enrichment of cell membrane with cholesterol inhibits HERG channels by reducing the levels of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] due to the activation of phospholipase C (PLC). In this study, we further explored the effect of cholesterol enrichment on HERG channel kinetics. When membrane cholesterol level was mildly increased in human embryonic kidney (HEK) 293 cells expressing HERG channel, the inactivation and deactivation kinetics of HERG current were not affected, but the activation rate was significantly decelerated at all voltages tested. The application of PtdIns(4,5)P2 or inhibitor for PLC prevented the effect of cholesterol enrichment, while the presence of antibody against PtdIns(4,5)P2 in pipette solution mimicked the effect of cholesterol enrichment. These results indicate that the effect of cholesterol enrichment on HERG channel is due to the depletion of PtdIns(4,5)P2. We also found that cholesterol enrichment significantly increases the expression of β1 and β3 isoforms of PLC (PLCβ1, PLCβ3) in the membrane. Since the effects of cholesterol enrichment on HERG channel were prevented by inhibiting transcription or by inhibiting PLCβ1 expression, we conclude that increased PLCβ1 expression leads to the deceleration of HERG channel activation rate via downregulation of PtdIns(4,5)P2. These results confirm a crosstalk between two plasma membrane-enriched lipids, cholesterol and PtdIns(4,5)P2, in the regulation of HERG channels.

  9. Altered expression of two-pore domain potassium (K2P channels in cancer.

    Directory of Open Access Journals (Sweden)

    Sarah Williams

    Full Text Available Potassium channels have become a focus in cancer biology as they play roles in cell behaviours associated with cancer progression, including proliferation, migration and apoptosis. Two-pore domain (K2P potassium channels are background channels which enable the leak of potassium ions from cells. As these channels are open at rest they have a profound effect on cellular membrane potential and subsequently the electrical activity and behaviour of cells in which they are expressed. The K2P family of channels has 15 mammalian members and already 4 members of this family (K2P2.1, K2P3.1, K2P9.1, K2P5.1 have been implicated in cancer. Here we examine the expression of all 15 members of the K2P family of channels in a range of cancer types. This was achieved using the online cancer microarray database, Oncomine (www.oncomine.org. Each gene was examined across 20 cancer types, comparing mRNA expression in cancer to normal tissue. This analysis revealed all but 3 K2P family members (K2P4.1, K2P16.1, K2P18.1 show altered expression in cancer. Overexpression of K2P channels was observed in a range of cancers including breast, leukaemia and lung while more cancers (brain, colorectal, gastrointestinal, kidney, lung, melanoma, oesophageal showed underexpression of one or more channels. K2P1.1, K2P3.1, K2P12.1, were overexpressed in a range of cancers. While K2P1.1, K2P3.1, K2P5.1, K2P6.1, K2P7.1 and K2P10.1 showed significant underexpression across the cancer types examined. This analysis supports the view that specific K2P channels may play a role in cancer biology. Their altered expression together with their ability to impact the function of other ion channels and their sensitivity to environmental stimuli (pO2, pH, glucose, stretch makes understanding the role these channels play in cancer of key importance.

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

    Czech Academy of Sciences Publication Activity Database

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

    2007-01-01

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

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

    Science.gov (United States)

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

    2017-12-14

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

  12. Calcium waves.

    Science.gov (United States)

    Jaffe, Lionel F

    2008-04-12

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

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

    DEFF Research Database (Denmark)

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

    2016-01-01

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

  14. Letrozole induced low estrogen levels affected the expressions of duodenal and renal calcium-processing gene in laying hens.

    Science.gov (United States)

    Li, Qiao; Zhao, Xingkai; Wang, Shujie; Zhou, Zhenlei

    2018-01-01

    Estrogen regulates the calcium homeostasis in hens, but the mechanisms involved are still unclear fully. In this study, we investigated whether letrozole (LZ) induced low estrogen levels affected the calcium absorption and transport in layers. In the duodenum, we observed a significant decrease of mRNA expressions of Calbindin-28k (CaBP-28k) and plasma membrane Ca 2+ -ATPase (PMCA 1b) while CaBP-28k protein expression was declined in birds with LZ treatment, and the mRNA levels of duodenal transient receptor potential vanilloid 6 (TRPV6) and Na + /Ca 2+ exchanger 1 (NCX1) were not affected. Interestingly, we observed the different changes in the kidney. The renal mRNA expressions of TRPV6 and NCX1 were unregulated while the PMCA1b was down-regulated in low estrogen layers, however, the CaBP-28k gene and protein expressions were no changed in the kidney. Furthermore, it showed that the duodenal estradiol receptor 2 (ESR2) transcripts rather than parathyroid hormone 1 receptor (PTH1R) and calcitonin receptor (CALCR) played key roles to down-regulate calcium transport in LZ-treated birds. In conclusion, CaBP-28k, PMCA 1b and ESR2 genes in the duodenum may be primary targets for estrogen regulation in order to control calcium homeostasis in hens. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. A Single Nucleotide Polymorphism (rs4236480 in TRPV5 Calcium Channel Gene Is Associated with Stone Multiplicity in Calcium Nephrolithiasis Patients

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    Anas Khaleel

    2015-01-01

    Full Text Available Nephrolithiasis is characterized by calcification of stones in the kidneys from an unknown cause. Animal models demonstrated the functional roles of the transient receptor potential vanilloid member 5 (TRPV5 gene in calcium renal reabsorption and hypercalciuria. Therefore, TRPV5 was suggested to be involved in calcium homeostasis. However, whether genetic polymorphisms of TRPV5 are associated with kidney stone multiplicity or recurrence is unclear. In this study, 365 Taiwanese kidney-stone patients were recruited. Both biochemical data and DNA samples were collected. Genotyping was performed by a TaqMan allelic discrimination assay. We found that a TRPV5 polymorphism (rs4236480 was observed to be associated with stone multiplicity of calcium nephrolithiasis, as the risk of stone multiplicity was higher in patients with the TT+CT genotype than in patients with the CC genotype (p=0.0271. In summary, despite the complexity of nephrolithiasis and the potential association of numerous calcium homeostatic absorption/reabsorption factors, TRPV5 plays an important role in the pathogenesis of calcium nephrolithiasis.

  16. EXPRESSION OF CALCIUM-BINDING PROTEINS IN THE NEUROTROPHIN-3-DEPENDENT SUBPOPULATION OF RAT EMBRYONIC DORSAL-ROOT GANGLION-CELLS IN CULTURE

    NARCIS (Netherlands)

    COPRAY, JCVM; MANTINGHOTTER, IJ; BROUWER, N

    1994-01-01

    In this study we have examined the calcium-binding protein expression in rat embryonic (E16) dorsal root ganglia (DRG) neurons in vitro in the presence of neurotrophin-3 (NT-3). A comparison was made with the expression of calcium-binding proteins in DRG subpopulations that depended in vitro on

  17. Ether à go-go potassium channel expression in soft tissue sarcoma patients

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    Stühmer Walter

    2006-10-01

    Full Text Available Abstract Background The expression of the human Eag1 potassium channel (Kv10.1 is normally restricted to the adult brain, but it has been detected in both tumour cell lines and primary tumours. Our purpose was to determine the frequency of expression of Eag1 in soft tissue sarcoma and its potential clinical implications. Results We used specific monoclonal antibodies to determine the expression levels of Eag1 in soft tissue sarcomas from 210 patients by immunohistochemistry. Eag1 was expressed in 71% of all tumours, with frequencies ranging from 56% (liposarcoma to 82% (rhabdomyosarcoma. We detected differences in expression levels depending on the histological type, but no association was seen between expression of this protein and sex, age, grade or tumour size. Four cell lines derived from relevant sarcoma histological types (fibrosarcoma and rhabdomyosarcoma were tested for Eag1 expression by real-time RT-PCR. We found all four lines to be positive for Eag1. In these cell lines, blockage of Eag1 by RNA interference led to a decrease in proliferation. Conclusion Eag1 is aberrantly expressed in over 70% sarcomas. In sarcoma cell lines, inhibition of Eag1 expression and/or function leads to reduced proliferation. The high frequency of expression of Eag1 in primary tumours and the restriction of normal expression of the channel to the brain, suggests the application of this protein for diagnostic or therapeutic purposes.

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

    Science.gov (United States)

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

    2014-10-01

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

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

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    Melissa eGómez

    2015-03-01

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

  20. Expression of G-protein inwardly rectifying potassium channels (GIRKs in lung cancer cell lines

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    Schuller Hildegard M

    2005-08-01

    Full Text Available Abstract Background Previous data from our laboratory has indicated that there is a functional link between the β-adrenergic receptor signaling pathway and the G-protein inwardly rectifying potassium channel (GIRK1 in human breast cancer cell lines. We wanted to determine if GIRK channels were expressed in lung cancers and if a similar link exists in lung cancer. Methods GIRK1-4 expression and levels were determined by reverse transcription polymerase chain reaction (RT-PCR and real-time PCR. GIRK protein levels were determined by western blots and cell proliferation was determined by a 5-bromo-2'-deoxyuridine (BrdU assay. Results GIRK1 mRNA was expressed in three of six small cell lung cancer (SCLC cell lines, and either GIRK2, 3 or 4 mRNA expression was detected in all six SCLC cell lines. Treatment of NCI-H69 with β2-adrenergic antagonist ICI 118,551 (100 μM daily for seven days led to slight decreases of GIRK1 mRNA expression levels. Treatment of NCI-H69 with the β-adrenergic agonist isoproterenol (10 μM decreased growth rates in these cells. The GIRK inhibitor U50488H (2 μM also inhibited proliferation, and this decrease was potentiated by isoproterenol. In the SCLC cell lines that demonstrated GIRK1 mRNA expression, we also saw GIRK1 protein expression. We feel these may be important regulatory pathways since no expression of mRNA of the GIRK channels (1 & 2 was found in hamster pulmonary neuroendocrine cells, a suggested cell of origin for SCLC, nor was GIRK1 or 2 expression found in human small airway epithelial cells. GIRK (1,2,3,4 mRNA expression was also seen in A549 adenocarcinoma and NCI-H727 carcinoid cell lines. GIRK1 mRNA expression was not found in tissue samples from adenocarcinoma or squamous cancer patients, nor was it found in NCI-H322 or NCI-H441 adenocarcinoma cell lines. GIRK (1,3,4 mRNA expression was seen in three squamous cell lines, GIRK2 was only expressed in one squamous cell line. However, GIRK1 protein

  1. Reduced myocardial 18F-FDG uptake after calcium channel blocker administration. Initial observation for a potential new method to improve plaque detection

    International Nuclear Information System (INIS)

    Gaeta, Chiara; Flotats, Albert; Artigas, Carles; Deportos, Jordi; Geraldo, Llanos; Carrio, Ignasi; Fernandez, Yolanda; Pavia, Javier

    2011-01-01

    Physiological glucose uptake by the myocardium may hamper visualization of coronary atherosclerotic plaques in 18 F-FDG PET studies. Intracellular myocardial calcium relates to glucose influx. We assessed whether administration of a calcium channel blocker such as verapamil could decrease myocardial 18 F-FDG uptake in mice. Experiments were conducted on ten male C57BL/6JOlaHsd mice. The mice were studied by 18 F-FDG PET/CT under basal conditions and after a single administration of verapamil injected 1 h prior to 18 F-FDG administration at doses of 1 mg/kg (group A, n = 5) and 20 mg/kg (group B, n = 5). PET scanning was started 60 min after injection of 18 F-FDG employing a dedicated small-animal PET/CT system (ARGUS-CT). In each mouse, post-verapamil PET images were coregistered with the basal PET images. Volumetric regions of interest (VOI) were drawn on the basal study containing the myocardium of the whole left ventricle and quantitatively compared with the same VOI applied to the post-verapamil scan. The SUV mean was used to express the mean myocardial 18 F-FDG uptake. The relative coefficient of variation (RV) between the basal and post-verapamil conditions was calculated. Verapamil administration decreased myocardial 18 F-FDG uptake in all animals. The median (range) SUV mean values in group A were 2.6 (1.6-4.1) under basal conditions and 1.7 (1.1-2.9) after verapamil administration (p = 0.043), and in group B were 1.6 (1.3-2.0) under basal conditions and 1.0 (0.9-1.4) after verapamil administration (p = 0.043). The median (range) RV values were -31% (-5%, -50%) in group A, and -37% (-10%, -51%) in group B (p = 0.6). In this animal model there was a significant reduction in 18 F-FDG uptake in the myocardium following verapamil administration. This type of intervention could facilitate the definition of coronary atherosclerotic plaque inflammation on 18 F-FDG PET scans. (orig.)

  2. The effect of an L/N-type calcium channel blocker on intradialytic blood pressure in intradialytic hypertensive patients.

    Science.gov (United States)

    Ito, Takayasu; Fujimoto, Naoki; Ishikawa, Eiji; Dohi, Kaoru; Fujimoto, Mika; Murata, Tomohiro; Kiyohara, Michiyo; Takeuchi, Hideyuki; Koyabu, Sukenari; Nishimura, Hiroyuki; Takeuchi, Toshiaki; Ito, Masaaki

    2018-03-27

    Intradialytic hypertension (HTN), which is one of the poor prognostic markers in patients undergoing hemodialysis, may be associated with sympathetic overactivity. The L/N-type calcium channel blocker, cilnidipine, has been reported to suppress sympathetic nerves activity in vivo. Therefore, we hypothesized that cilnidipine could attenuate intradialytic systolic blood pressure (SBP) elevation. Fifty-one patients on chronic hemodialysis who had intradialytic-HTN (SBP elevation ≥10 mmHg during hemodialysis) and no fluid overload were prospectively randomized into two groups: control and cilnidipine groups. Cilnidipine group patients took cilnidipine (10 mg/day) for 12 weeks. The primary endpoint was the change in the intradialytic SBP elevation before and after the 12-week intervention. Before the intervention, no differences were observed in age, sex or pre-dialytic SBP (148.5 ± 12.9 vs. 148.3 ± 19.3 mmHg) between the two groups. Intradialytic SBP elevation was unchanged in the control group. Cilnidipine significantly lowered the post-dialytic SBP with an attenuation of the intradialytic SBP elevation from 12.0 ± 15.4 mmHg to 4.8 ± 10.1 mmHg. However, the observed difference in the intradialytic SBP elevation by cilnidipine did not reach statistical significance (group×time interaction effect p = 0.25). Cathecolamine levels were unaffected by the intervention in both groups. Cilnidipine lowers both the pre- and post-dialytic SBP and might attenuate intradialytic SBP elevation. Therefore, cilnidipine may be effective in lowering SBP during HD in patients with intradialytic-HTN.

  3. Calcium channel blockers, more than diuretics, enhance vascular protective effects of angiotensin receptor blockers in salt-loaded hypertensive rats.

    Directory of Open Access Journals (Sweden)

    Eiichiro Yamamoto

    Full Text Available The combination therapy of an angiotensin receptor blocker (ARB with a calcium channel blocker (CCB or with a diuretic is favorably recommended for the treatment of hypertension. However, the difference between these two combination therapies is unclear. The present work was undertaken to examine the possible difference between the two combination therapies in vascular protection. Salt-loaded stroke-prone spontaneously hypertensive rats (SHRSP were divided into 6 groups, and they were orally administered (1 vehicle, (2 olmesartan, an ARB, (3 azelnidipine, a CCB, (4 hydrochlorothiazide, a diuretic, (5 olmesartan combined with azelnidipine, or (6 olmesartan combined with hydrochlorothiazide. Olmesartan combined with either azelnidipine or hydrochlorothiazide ameliorated vascular endothelial dysfunction and remodeling in SHRSP more than did monotherapy with either agent. However, despite a comparable blood pressure lowering effect between the two treatments, azelnidipine enhanced the amelioration of vascular endothelial dysfunction and remodeling by olmesartan to a greater extent than did hydrochlorothiazide in salt-loaded SHRSP. The increased enhancement by azelnidipine of olmesartan-induced vascular protection than by hydrochlorothiazide was associated with a greater amelioration of vascular nicotinamide adenine dinucleotide phosphate (NADPH oxidase activation, superoxide, mitogen-activated protein kinase activation, and with a greater activation of the Akt/endothelial nitric oxide synthase (eNOS pathway. These results provided the first evidence that a CCB potentiates the vascular protective effects of an ARB in salt-sensitive hypertension, compared with a diuretic, and provided a novel rationale explaining the benefit of the combination therapy with an ARB and a CCB.

  4. Sigma-1 receptor agonist increases axon outgrowth of hippocampal neurons via voltage-gated calcium ions channels.

    Science.gov (United States)

    Li, Dong; Zhang, Shu-Zhuo; Yao, Yu-Hong; Xiang, Yun; Ma, Xiao-Yun; Wei, Xiao-Li; Yan, Hai-Tao; Liu, Xiao-Yan

    2017-12-01

    Sigma-1 receptors (Sig-1Rs) are unique endoplasmic reticulum proteins that have been implicated in both neurodegenerative and ischemic diseases, such as Alzheimer's disease and stroke. Accumulating evidence has suggested that Sig-1R plays a role in neuroprotection and axon outgrowth. The underlying mechanisms of Sig-1R-mediated neuroprotection have been well elucidated. However, the mechanisms underlying the effects of Sig-1R on axon outgrowth are not fully understood. To clarify this issue, we utilized immunofluorescence to compare the axon lengths of cultured naïve hippocampal neurons before and after the application of the Sig-1R agonist, SA4503. Then, electrophysiology and immunofluorescence were used to examine voltage-gated calcium ion channel (VGCCs) currents in the cell membranes and growth cones. We found that Sig-1R activation dramatically enhanced the axonal length of the naïve hippocampal neurons. Application of the Sig-1R antagonist NE100 and gene knockdown techniques both demonstrated the effects of Sig-1R. The growth-promoting effect of SA4503 was accompanied by the inhibition of voltage-gated Ca 2+ influx and was recapitulated by incubating the neurons with the L-type, N-type, and P/Q-type VGCC blockers, nimodipine, MVIIA and ω-agatoxin IVA, respectively. This effect was unrelated to glial cells. The application of SA4503 transformed the growth cone morphologies from complicated to simple, which favored axon outgrowth. Sig-1R activation can enhance axon outgrowth and may have a substantial influence on neurogenesis and neurodegenerative diseases. © 2017 John Wiley & Sons Ltd.

  5. Calcium channel blockers and breast cancer incidence: An updated systematic review and meta-analysis of the evidence.

    Science.gov (United States)

    Wright, Cameron M; Moorin, Rachael E; Chowdhury, Enayet K; Stricker, Bruno H; Reid, Christopher M; Saunders, Christobel M; Hughes, Jeffery D

    2017-10-01

    Controversy exists regarding the potential association between taking calcium channel blockers (CCBs) and the development of breast cancer. As a positive association would have important public health implications due to the widespread use of CCBs, this study aimed to incorporate new evidence to determine whether an association is likely to exist. We searched MEDLINE, EMBASE and the Cochrane Library to 28 June 2016 for relevant literature. References and citing articles were checked and authors contacted as necessary. Two authors independently selected articles and extracted data. Twenty-nine studies were reviewed; 26 were non-randomised studies (NRS). Meta-analysis of study data where adjustment for 'confounding by indication' was judged to be present suggests that an association, if any, is likely to be modest in magnitude (pooled odds/risk ratio 1.09 (95% confidence interval (CI) 1.03-1.15, I 2 =0%, 8 sub-studies; pooled hazard ratio 0.99 (95% CI 0.94-1.03, I 2 =35%, 9 sub-studies)). There are credible study data showing an increased relative risk with long-term use of CCBs, but the results of our meta-analysis and of meta-regression of log relative risk against minimum follow-up time are mixed. The current summative evidence does not support a clear association between taking CCBs and developing breast cancer. However, uncertainty remains, especially for long-term use and any association might not be uniform between different populations and/or breast cancer sub-types. We thus recommend further NRS in settings where CCB use is highly prevalent and population-based cancer, prescription and health-registries exist, to resolve this continuing uncertainty. PROSPERO, CRD42015026712. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Comprehensive behavioral analysis of voltage-gated calcium channel beta-anchoring and -regulatory protein knockout mice

    Science.gov (United States)

    Nakao, Akito; Miki, Takafumi; Shoji, Hirotaka; Nishi, Miyuki; Takeshima, Hiroshi; Miyakawa, Tsuyoshi; Mori, Yasuo

    2015-01-01

    Calcium (Ca2+) influx through voltage-gated Ca2+ channels (VGCCs) induces numerous intracellular events such as neuronal excitability, neurotransmitter release, synaptic plasticity, and gene regulation. It has been shown that genes related to Ca2+ signaling, such as the CACNA1C, CACNB2, and CACNA1I genes that encode VGCC subunits, are associated with schizophrenia and other psychiatric disorders. Recently, VGCC beta-anchoring and -regulatory protein (BARP) was identified as a novel regulator of VGCC activity via the interaction of VGCC β subunits. To examine the role of the BARP in higher brain functions, we generated BARP knockout (KO) mice and conducted a comprehensive battery of behavioral tests. BARP KO mice exhibited greatly reduced locomotor activity, as evidenced by decreased vertical activity, stereotypic counts in the open field test, and activity level in the home cage, and longer latency to complete a session in spontaneous T-maze alteration test, which reached “study-wide significance.” Acoustic startle response was also reduced in the mutants. Interestingly, they showed multiple behavioral phenotypes that are seemingly opposite to those seen in the mouse models of schizophrenia and its related disorders, including increased working memory, flexibility, prepulse inhibition, and social interaction, and decreased locomotor activity, though many of these phenotypes are statistically weak and require further replications. These results demonstrate that BARP is involved in the regulation of locomotor activity and, possibly, emotionality. The possibility was also suggested that BARP KO mice may serve as a unique tool for investigating the pathogenesis/pathophysiology of schizophrenia and related disorders. Further evaluation of the molecular and physiological phenotypes of the mutant mice would provide new insights into the role of BARP in higher brain functions. PMID:26136667

  7. Unidirectional photoreceptor-to-Müller glia coupling and unique K+ channel expression in Caiman retina.

    Directory of Open Access Journals (Sweden)

    Astrid Zayas-Santiago

    Full Text Available Müller cells, the principal glial cells of the vertebrate retina, are fundamental for the maintenance and function of neuronal cells. In most vertebrates, including humans, Müller cells abundantly express Kir4.1 inwardly rectifying potassium channels responsible for hyperpolarized membrane potential and for various vital functions such as potassium buffering and glutamate clearance; inter-species differences in Kir4.1 expression were, however, observed. Localization and function of potassium channels in Müller cells from the retina of crocodiles remain, hitherto, unknown.We studied retinae of the Spectacled caiman (Caiman crocodilus fuscus, endowed with both diurnal and nocturnal vision, by (i immunohistochemistry, (ii whole-cell voltage-clamp, and (iii fluorescent dye tracing to investigate K+ channel distribution and glia-to-neuron communications.Immunohistochemistry revealed that caiman Müller cells, similarly to other vertebrates, express vimentin, GFAP, S100β, and glutamine synthetase. In contrast, Kir4.1 channel protein was not found in Müller cells but was localized in photoreceptor cells. Instead, 2P-domain TASK-1 channels were expressed in Müller cells. Electrophysiological properties of enzymatically dissociated Müller cells without photoreceptors and isolated Müller cells with adhering photoreceptors were significantly different. This suggests ion coupling between Müller cells and photoreceptors in the caiman retina. Sulforhodamine-B injected into cones permeated to adhering Müller cells thus revealing a uni-directional dye coupling.Our data indicate that caiman Müller glial cells are unique among vertebrates studied so far by predominantly expressing TASK-1 rather than Kir4.1 K+ channels and by bi-directional ion and uni-directional dye coupling to photoreceptor cells. This coupling may play an important role in specific glia-neuron signaling pathways and in a new type of K+ buffering.

  8. Cholesterol Down-Regulates BK Channels Stably Expressed in HEK 293 Cells

    Science.gov (United States)

    Deng, Xiu-Ling; Sun, Hai-Ying; Li, Gui-Rong

    2013-01-01

    Cholesterol is one of the major lipid components of the plasma membrane in mammalian cells and is involved in the regulation of a number of ion channels. The present study investigates how large conductance Ca2+-activated K+ (BK) channels are regulated by membrane cholesterol in BK-HEK 293 cells expressing both the α-subunit hKCa1.1 and the auxiliary β1-subunit or in hKCa1.1-HEK 293 cells expressing only the α-subunit hKCa1.1 using approaches of electrophysiology, molecular biology, and immunocytochemistry. Membrane cholesterol was depleted in these cells with methyl-β-cyclodextrin (MβCD), and enriched with cholesterol-saturated MβCD (MβCD-cholesterol) or low-density lipoprotein (LDL). We found that BK current density was decreased by cholesterol enrichment in BK-HEK 293 cells, with a reduced expression of KCa1.1 protein, but not the β1-subunit protein. This effect was fully countered by the proteasome inhibitor lactacystin or the lysosome function inhibitor bafilomycin A1. Interestingly, in hKCa1.1-HEK 293 cells, the current density was not affected by cholesterol enrichment, but directly decreased by MβCD, suggesting that the down-regulation of BK channels by cholesterol depends on the auxiliary β1-subunit. The reduced KCa1.1 channel protein expression was also observed in cultured human coronary artery smooth muscle cells with cholesterol enrichment using MβCD-cholesterol or LDL. These results demonstrate the novel information that cholesterol down-regulates BK channels by reducing KCa1.1 protein expression via increasing the channel protein degradation, and the effect is dependent on the auxiliary β1-subunit. PMID:24260325

  9. Cloning, functional expression, and characterization of a PKA-activated gastric Cl- channel.

    Science.gov (United States)

    Malinowska, D H; Kupert, E Y; Bahinski, A; Sherry, A M; Cuppoletti, J

    1995-01-01

    cDNA encoding a Cl- channel was isolated from a rabbit gastric library, sequenced, and expressed in Xenopus oocytes. The predicted protein (898 amino acids, relative molecular mass 98,433 Da) was overall 93% similar to the rat brain ClC-2 Cl- channel. However, a 151-amino acid stretch toward the COOH-terminus was 74% similar to ClC-2 with six amino acids deleted. Two new potential protein kinase A (PKA) phosphorylation sites (also protein kinase C phosphorylation sites) were introduced. cRNA-injected Xenopus oocytes expressed a Cl- channel that was active at pHtrans 3 and had a linear current-voltage (I-V) curve and a slope conductance of 29 +/- 1 pS at 800 mM CsCl. A fivefold Cl- gradient caused a rightward shift in the I-V curve with a reversal potential of +30 +/- 3 mV, indicating anion selectivity. The selectivity was I- > Cl- > NO3-. The native and recombinant Cl- channel were both activated in vitro by PKA catalytic subunit and ATP. The electrophysiological and regulatory properties of the cloned and the native channel were similar. The cloned protein may be the Cl- channel involved in gastric HCl secretion.

  10. Calcium-binding protein expression in peritoneal endometriosis-associated nerve fibres.

    Science.gov (United States)

    Barcena de Arellano, M L; Münch, S; Arnold, J; Helbig, S; Schneider, A; Mechsner, S

    2013-11-01

    Recent studies demonstrated the potential involvement of nerve fibres in the chronic inflammatory process of endometriosis. We aimed to characterize nerve fibres in the proximal and distal areas of the peritoneal endometriotic lesions in order to understand the chronic inflammatory process in endometriosis. Peritoneal endometriotic lesions (proximal area) (n = 17), the matching unaffected peritoneum (distal area) and healthy peritoneum of patients without endometriosis (n = 15) were analysed with the neuronal markers PGP 9.5, calbindin, calretinin and parvalbumin. Peritoneal fluids of women with and without endometriosis were used for Western blot analysis and for the neuronal growth assay. The protein expression of neuronal PC-12 cells incubated with peritoneal fluids was analysed. The overall nerve fibre density was significantly reduced in the distal area of the lesion when compared with the proximal area or with healthy peritoneum. The density of calbindin-, calretinin- and parvalbumin-positive nerve fibres was significantly increased in the endometriosis group. Calretinin expression was elevated in the peritoneal fluid of women with symptomatic endometriosis when compared with women with asymptomatic endometriosis. Furthermore, PC-12 cells incubated with peritoneal fluid of women with endometriosis showed a higher proliferation rate and a stronger neurite outgrowth than the control group. PC-12 cells incubated in peritoneal fluids of women with endometriosis expressed less calretinin but more calbindin than the control group. Calcium-binding proteins seem to be increased in endometriosis-associated nerve fibres and might play an important role in the chronic inflammatory condition and the pain pathogenesis of endometriosis. © 2013 European Federation of International Association for the Study of Pain Chapters.

  11. EXPRESSION OF CALCIUM-DEPENDENT PROTEIN KINASE (CDPK GENES IN VITIS AMURENSIS UNDER ABIOTIC STRESS CONDITIONS

    Directory of Open Access Journals (Sweden)

    Dubrovina A.S.

    2012-08-01

    Full Text Available Abiotic stresses, such as extreme temperatures, soil salinity, or water deficit, are one of the major limiting factors of crop productivity worldwide. Examination of molecular and genetic mechanisms of abiotic stress tolerance in plants is of great interest to plant biologists. Calcium-dependent protein kinases (CDPKs, which are the most important Ca2+ sensors in plants, are known to play one of the key roles in plant adaptation to abiotic stress. CDPK is a multigene family of enzymes. Analysis of CDPK gene expression under various abiotic stress conditions would help identify those CDPKs that might play important roles in plant adaptation to abiotic stress. We focused on studying CDPK gene expression under osmotic, water deficit, and temperature stress conditions in a wild-growing grapevine Vitis amurensis Rurp., which is native to the Russian Far East and is known to possess high adaptive potential and high level of resistance against adverse environmental conditions. Healthy V. amurensis cuttings (excised young stems with one healthy leaf were used for the treatments. For the non-stress treatment, we placed the cuttings in distilled water for 12 h at room temperature. For the water-deficit stress, detached cuttings were laid on a paper towel for 12 h at room temperature. For osmotic stress treatments, the cuttings were placed in 0.4 М NaCl and 0.4 М mannitol solutions for 12 h at room temperature. To examine temperature stress tolerance, the V. amurensis cuttings were placed in a growth chamber at +10oC and +37oC for 12 h. The total expression of VaCDPK genes was examined by semiquantitative RT-PCR with degenerate primers designed to the CDPK kinase domain. The total level of CDPK gene expression increased under salt and decreased under low temperature stress conditions. We sequenced 300 clones of the amplified part of different CDPK transcripts obtained from the analyzed cDNA probes. Analysis of the cDNA sequences identified 8 different

  12. Chronic exposure of NG108-15 cells to amyloid beta peptide (A beta(1-42)) abolishes calcium influx via N-type calcium channels

    Czech Academy of Sciences Publication Activity Database

    Kašparová, Jana; Lisá, Věra; Tuček, Stanislav; Doležal, Vladimír

    2001-01-01

    Roč. 26, 8-9 (2001), s. 1079-1084 ISSN 0364-3190 R&D Projects: GA MZd NF5183 Institutional research plan: CEZ:AV0Z5011922 Keywords : amyloid beta peptide * Alzheimer's disease * calcium Subject RIV: FH - Neurology Impact factor: 1.638, year: 2001

  13. Calcium in plant cells

    Directory of Open Access Journals (Sweden)

    V. V. Schwartau

    2014-04-01

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

  14. Polyunsaturated fatty acids are potent openers of human M-channels expressed in Xenopus laevis oocytes

    DEFF Research Database (Denmark)

    Liin, Sara I; Karlsson, Urban; Bentzen, Bo Hjorth

    2016-01-01

    the threshold current to evoke action potentials in dorsal root ganglion neurons. The polyunsaturated fatty acids docosahexaenoic acid, α-linolenic acid, and eicosapentaenoic acid facilitated opening of the human M-channel, comprised of the heteromeric human KV 7.2/3 channel expressed in Xenopus oocytes......, by shifting the conductance-versus-voltage curve towards more negative voltages (by -7.4 to -11.3 mV by 70 μM). Uncharged docosahexaenoic acid methyl ester and monounsaturated oleic acid did not facilitate opening of the human KV 7.2/3 channel. CONCLUSIONS: These findings suggest that circulating...... polyunsaturated fatty acids, with a minimum requirement of multiple double bonds and a charged carboxyl group, dampen excitability by opening neuronal M-channels. Collectively, our data bring light to the molecular targets of polyunsaturated fatty acids and thus a possible mechanism by which polyunsaturated fatty...

  15. N-linked glycans are required on epithelial Na+ channel subunits for maturation and surface expression.

    Science.gov (United States)

    Kashlan, Ossama B; Kinlough, Carol L; Myerburg, Michael M; Shi, Shujie; Chen, Jingxin; Blobner, Brandon M; Buck, Teresa M; Brodsky, Jeffrey L; Hughey, Rebecca P; Kleyman, Thomas R

    2018-03-01

    Epithelial Na + channel (ENaC) subunits undergo N-linked glycosylation in the endoplasmic reticulum where they assemble into an αβγ complex. Six, 13, and 5 consensus sites (Asn-X-Ser/Thr) for N-glycosylation reside in the extracellular domains of the mouse α-, β-, and γ-subunits, respectively. Because the importance of ENaC N-linked glycans has not been fully addressed, we examined the effect of preventing N-glycosylation of specific subunits on channel function, expression, maturation, and folding. Heterologous expression in Xenopus oocytes or Fischer rat thyroid cells with αβγ-ENaC lacking N-linked glycans on a single subunit reduced ENaC activity as well as the inhibitory response to extracellular Na + . The lack of N-linked glycans on the β-subunit also precluded channel activation by trypsin. However, channel activation by shear stress was N-linked glycan independent, regardless of which subunit was modified. We also discovered that the lack of N-linked glycans on any one subunit reduced the total and surface levels of cognate subunits. The lack of N-linked glycans on the β-subunit had the largest effect on total levels, with the lack of N-linked glycans on the γ- and α-subunits having intermediate and modest effects, respectively. Finally, channels with wild-type β-subunits were more sensitive to limited trypsin proteolysis than channels lacking N-linked glycans on the β-subunit. Our results indicate that N-linked glycans on each subunit are required for proper folding, maturation, surface expression, and function of the channel.

  16. Distribution and expression of non-neuronal transient receptor potential (TRPV) ion channels in rosacea.

    Science.gov (United States)

    Sulk, Mathias; Seeliger, Stephan; Aubert, Jerome; Schwab, Verena D; Cevikbas, Ferda; Rivier, Michel; Nowak, Pawel; Voegel, Johannes J; Buddenkotte, Jörg; Steinhoff, Martin

    2012-04-01

    Rosacea is a frequent chronic inflammatory skin disease of unknown etiology. Because early rosacea reveals all characteristics of neurogenic inflammation, a central role of sensory nerves in its pathophysiology has been discussed. Neuroinflammatory mediators and their receptors involved in rosacea are poorly defined. Good candidates may be transient receptor potential (TRP) ion channels of vanilloid type (TRPV), which can be activated by many trigger factors of rosacea. Interestingly, TRPV2, TRPV3, and TRPV4 are expressed by both neuronal and non-neuronal cells. Here, we analyzed the expression and distribution of TRPV receptors in the various subtypes of rosacea on non-neuronal cells using immunohistochemistry, morphometry, double immunoflourescence, and quantitative real-time PCR (qRT-PCR) as compared with healthy skin and lupus erythematosus. Our results show that dermal immunolabeling of TRPV2 and TRPV3 and gene expression of TRPV1 is significantly increased in erythematotelangiectatic rosacea (ETR). Papulopustular rosacea (PPR) displayed an enhanced immunoreactivity for TRPV2, TRPV4, and also of TRPV2 gene expression. In phymatous rosacea (PhR)-affected skin, dermal immunostaining of TRPV3 and TRPV4 and gene expression of TRPV1 and TRPV3 was enhanced, whereas epidermal TRPV2 staining was decreased. Thus, dysregulation of TRPV channels also expressed by non-neuronal cells may be critically involved in the initiation and/or development of rosacea. TRP ion channels may be targets for the treatment of rosacea.

  17. Sequence genomic organization and expression of two channel catfish Ictalurus punctatus Ghrelin receptors

    Science.gov (United States)

    Two ghrelin receptor (GHS-R) genes were isolated from channel catfish tissue and a bacterial artificial chromosome (BAC) library. The two receptors were characterized by determining tissue distribution, ontogeny of receptor mRNA expression, and effects of exogenous homologous ghrelin administration ...

  18. L-type calcium channels play a crucial role in the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wen, Li [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Wang, Yu [Department of Oncology, Xijing Hospital, Fourth Military Medical University, Xi' an 710032 (China); Wang, Huan [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Kong, Lingmin [Department of Fundamental Medicine, Cell Engineering Research Centre, Fourth Military Medical University, Xi' an 710032 (China); Zhang, Liang [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Chen, Xin [Department of General Dentistry, The 174th Hospital of Chinese PLA, Xiamen 361003 (China); Ding, Yin, E-mail: dingyin@fmmu.edu.cn [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer We detect the functional Ca{sup 2+} currents and mRNA expression of VDCC{sub L} in rMSCs. Black-Right-Pointing-Pointer Blockage of VDCC{sub L} exert antiproliferative and apoptosis-inducing effects on rMSCs. Black-Right-Pointing-Pointer Inhibiting VDCC{sub L} can suppress the ability of rMSCs to differentiate into osteoblasts. Black-Right-Pointing-Pointer {alpha}1C of VDCC{sub L} may be a primary functional subunit in VDCC{sub L}-regulating rMSCs. -- Abstract: L-type voltage-dependent Ca{sup 2+} channels (VDCC{sub L}) play an important role in the maintenance of intracellular calcium homeostasis, and influence multiple cellular processes. They have been confirmed to contribute to the functional activities of osteoblasts. Recently, VDCC{sub L} expression was reported in mesenchymal stem cells (MSCs), but the role of VDCC{sub L} in MSCs is still undetermined. The aim of this study was to determine whether VDCC{sub L} may be regarded as a new regulator in the proliferation and osteogenic differentiation of rat MSC (rMSCs). In this study, we examined functional Ca{sup 2+} currents (I{sub Ca}) and mRNA expression of VDCC{sub L} in rMSCs, and then suppressed VDCC{sub L} using nifedipine (Nif), a VDCC{sub L} blocker, to investigate its role in rMSCs. The proliferation and osteogenic differentiation of MSCs were analyzed by MTT, flow cytometry, alkaline phosphatase (ALP), Alizarin Red S staining, RT-PCR, and real-time PCR assays. We found that Nif exerts antiproliferative and apoptosis-inducing effects on rMSCs. ALP activity and mineralized nodules were significantly decreased after Nif treatment. Moreover, the mRNA levels of the osteogenic markers, osteocalcin (OCN), bone sialoprotein (BSP), and runt-related transcription factor 2 (Runx2), were also down-regulated. In addition, we transfected {alpha}1C-siRNA into the cells to further confirm the role of VDCC{sub L} in rMSCs, and a similar effect on osteogenesis was found. These

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

    Directory of Open Access Journals (Sweden)

    Diana C. Pachajoa

    2017-09-01

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

  20. Rab4GTPase modulates CFTR function by impairing channel expression at plasma membrane

    International Nuclear Information System (INIS)

    Saxena, Sunil K.; Kaur, Simarna; George, Constantine

    2006-01-01

    Cystic fibrosis (CF), an autosomal recessive disorder, is caused by the disruption of biosynthesis or the function of a membrane cAMP-activated chloride channel, CFTR. CFTR regulatory mechanisms include recruitment of channel proteins to the cell surface from intracellular pools and by protein-protein interactions. Rab proteins are small GTPases involved in regulated trafficking controlling vesicle docking and fusion. Rab4 controls recycling events from endosome to the plasma membrane, fusion, and degradation. The colorectal cell line HT-29 natively expresses CFTR and responds to cAMP stimulation with an increase in CFTR-mediated currents. Rab4 over-expression in HT-29 cells inhibits both basal and cAMP-stimulated CFTR-mediated currents. GTPase-deficient Rab4Q67L and GDP locked Rab4S22N both inhibit channel activity, which appears characteristically different. Active status of Rab4 was confirmed by GTP overlay assay, while its expression was verified by Western blotting. The pull-down and immunoprecipitation experiments suggest that Rab4 physically interacts with CFTR through protein-protein interaction. Biotinylation with cell impermeant NHS-Sulfo-SS-Biotin implies that Rab4 impairs CFTR expression at cell surface. The enhanced cytosolic CFTR indicates that Rab4 expression restrains CFTR appearance at the cell membrane. The study suggests that Rab4 regulates the channel through multiple mechanisms that include protein-protein interaction, GTP/GDP exchange, and channel protein trafficking. We propose that Rab4 is a dynamic molecule with a significant role in CFTR function

  1. Comparative study the expression of calcium cycling genes in Bombay duck (Harpadon nehereus and beltfish (Trichiurus lepturus with different swimming activities

    Directory of Open Access Journals (Sweden)

    Hui Zhang

    2017-06-01

    Full Text Available The contraction and relaxation events of the muscle is mediated by the coordination of many important calcium cycling proteins of ryanodine receptor (RYR, troponin C (TNNC, parvalbumin (PVALB, sarcoendoplasmic reticulum calcium transport ATPase (SERCA and calsequestrin (CASQ. In higher vertebrates, the expression level of calcium cycling proteins are positively correlated to the muscle contraction/relaxation ability of the cell. In this study, we used RNAseq to explore the expression profile of calcium cycling genes between two marine fish of Bombay duck (Harpadon nehereus and beltfish (Trichiurus lepturus with poor and robust swimming activities, respectively. We have studied the hypothesis whether the expression level of calcium cycling proteins are also positive correlated to swimming ability in fish. We used Illumina sequencing technology (NextSeq500 to sequence, assemble and annotate the muscle transcriptome of Bombay duck for the first time. A total of 47,752,240 cleaned reads (deposited in NCBI SRA database with accession number of SRX1706379 were obtained from RNA sequencing and 26,288 unigenes (with N50 of 486 bp were obtained after de novo assembling with Trinity software. BLASTX against NR, GO, KEGG and eggNOG databases show 100%, 65%, 26%, 94% and 88% annotation rate, respectively. Comparison of the dominantly expressed unigenes in fish muscle shows calcium cycling gene expression in beltfish (SRX1674471 is 1.4- to 51.6-fold higher than Bombay duck. Among five calcium cycling genes, the fold change results are very significant in CASQ (51.6 fold and PVALB (9.1 fold and both of them are responsive for calcium binding to reduce free calcium concentration in the sarcoendoplasmic reticulum and cytoplasm. In conclusion, we confirmed that the high abundant expression rate of calcium cycling genes in robust swimming fish species. The current muscle transcriptome and identified calcium cycling gene data can provide more insights into the

  2. Cloning and functional expression of a plant voltage-dependent chloride channel.

    Science.gov (United States)

    Lurin, C; Geelen, D; Barbier-Brygoo, H; Guern, J; Maurel, C

    1996-01-01

    Plant cell membrane anion channels participate in basic physiological functions, such as cell volume regulation and signal transduction. However, nothing is known about their molecular structure. Using a polymerase chain reaction strategy, we have cloned a tobacco cDNA (CIC-Nt1) encoding a 780-amino acid protein with several putative transmembrane domains. CIC-Nt1 displays 24 to 32% amino acid identity with members of the animal voltage-dependent chloride channel (CIC) family, whose archetype is CIC-0 from the Torpedo marmorata electric organ. Injection of CIC-Nt1 complementary RNA into Xenopus oocytes elicited slowly activating inward currents upon membrane hyperpolarization more negative than -120 mV. These currents were carried mainly by anions, modulated by extracellular anions, and totally blocked by 10 mM extracellular calcium. The identification of CIC-Nt1 extends the CIC family to higher plants and provides a molecular probe for the study of voltage-dependent anion channels in plants. PMID:8624442

  3. Specific reduction of calcium-binding protein (28-kilodalton calbindin-D) gene expression in aging and neurodegenerative diseases

    International Nuclear Information System (INIS)

    Iacopino, A.M.; Christakos, S.

    1990-01-01

    The present studies establish that there are specific, significant decreases in the neuronal calcium-binding protein (28-kDa calbindin-D) gene expression in aging and in neurodegenerative diseases. The specificity of the changes observed in calbindin mRNA levels was tested by reprobing blots with calmodulin, cyclophilin, and B-actin cDNAs. Gross brain regions of the aging rat exhibited specific, significant decreases in calbindin·mRNA and protein levels in the cerebellum, corpus striatum, and brain-stem region but not in the cerebral cortex or hippocampus. Discrete areas of the aging human brain exhibited significant decreases in calbindin protein and mRNA in the cerebellum, corpus striatum, and nucleus basalis but not in the neocortex, hippocampus, amygdala, locus ceruleus, or nucleus raphe dorsalis. Comparison of diseased human brain tissue with age- and sex-matched controls yielded significant decreases calbindin protein and mRNA in the substantia nigra (Parkinson disease), in the corpus striatum (Huntington disease), in the nucleus basalis (Alzheimer disease), and in the hippocampus and nucleus raphe dorsalis (Parkinson, Huntington, and Alzheimer diseases) but not in the cerebellum, neocortex, amygdala, or locus ceruleus. These findings suggest that decreased calbindin gene expression may lead to a failure of calcium buffering or intraneuronal calcium homeostasis, which contributes to calcium-mediated cytotoxic events during aging and in the pathogenesis of neurodegenerative diseases

  4. Ca2+ Channel Re-localization to Plasma-Membrane Microdomains Strengthens Activation of Ca2+-Dependent Nuclear Gene Expression

    Directory of Open Access Journals (Sweden)

    Krishna Samanta

    2015-07-01

    Full Text Available In polarized cells or cells with complex geometry, clustering of plasma-membrane (PM ion channels is an effective mechanism for eliciting spatially restricted signals. However, channel clustering is also seen in cells with relatively simple topology, suggesting it fulfills a more fundamental role in cell biology than simply orchestrating compartmentalized responses. Here, we have compared the ability of store-operated Ca2+ release-activated Ca2+ (CRAC channels confined to PM microdomains with a similar number of dispersed CRAC channels to activate transcription factors, which subsequently increase nuclear gene expression. For similar levels of channel activity, we find that channel confinement is considerably more effective in stimulating gene expression. Our results identify a long-range signaling advantage to the tight evolutionary conservation of channel clustering and reveal that CRAC channel aggregation increases the strength, fidelity, and reliability of the general process of excitation-transcription coupling.

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

    Science.gov (United States)

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

    2009-04-01

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

  6. Differential expression of calcium/calmodulin-regulated SlSRs in response to abiotic and biotic stresses in tomato fruit.

    Science.gov (United States)

    Yang, Tianbao; Peng, Hui; Whitaker, Bruce D; Jurick, Wayne M

    2013-07-01

    Calcium has been shown to enhance stress tolerance, maintain firmness and reduce decay in fruits. Previously we reported that seven tomato SlSRs encode calcium/calmodulin-regulated proteins, and that their expressions are developmentally regulated during fruit development and ripening, and are also responsive to ethylene. To study their expressions in response to stresses encountered during postharvest handling, tomato fruit at the mature-green stage was subjected to chilling and wounding injuries, infected with Botrytis cinerea and treated with salicylic acid or methyl jasmonate. Gene expression studies revealed that the seven SlSRs differentially respond to different stress signals. SlSR2 was the only gene upregulated by all the treatments. SlSR4 acted as a late pathogen-induced gene; it was upregulated by salicylic acid and methyl jasmonate, but downregulated by cold treatment. SlSR3L was cold- and wound-responsive and was also induced by salicylic acid. SlSR1 and SlSR1L were repressed by cold, wounding and pathogen infection, but were upregulated by salicylic acid and methyl jasmonate. Overall, results of these expression studies indicate that individual SlSRs have distinct roles in responses to the specific stress signals, and SlSRs may act as a coordinator(s) connecting calcium-mediated signaling with other stress signal transduction pathways during fruit ripening and storage. © 2013 Scandinavian Plant Physiology Society.

  7. Pulsatile atheroprone shear stress affects the expression of transient receptor potential channels in human endothelial cells

    DEFF Research Database (Denmark)

    Thilo, Florian; Vorderwülbecke, Bernd J; Marki, Alex

    2012-01-01

    in comparison with endothelial cells grown under static conditions. There was a significant association between the expression of TRPC6 and tumor necrosis factor-α mRNA in human vascular tissue. No-flow and atheroprone flow conditions are equally characterized by an increase in the expression of tumor necrosis......The goal of the study was to assess whether pulsatile atheroprone shear stress modulates the expression of transient receptor potential (TRP) channels, TRPC3, TRPC6, TRPM7, and TRPV1 mRNA, in human umbilical vascular endothelial cells. Exposure of cultured vascular endothelial cells to defined...

  8. Differential gene expression of cardiac ion channels in human dilated cardiomyopathy.

    Directory of Open Access Journals (Sweden)

    Maria Micaela Molina-Navarro

    Full Text Available BACKGROUND: Dilated cardiomyopathy (DCM is characterized by idiopathic dilation and systolic contractile dysfunction of the cardiac chambers. The present work aimed to study the alterations in gene expression of ion channels involved in cardiomyocyte function. METHODS AND RESULTS: Microarray profiling using the Affymetrix Human Gene® 1.0 ST array was performed using 17 RNA samples, 12 from DCM patients undergoing cardiac transplantation and 5 control donors (CNT. The analysis focused on 7 cardiac ion channel genes, since this category has not been previously studied in human DCM. SCN2B was upregulated, while KCNJ5, KCNJ8, CLIC2, CLCN3, CACNB2, and CACNA1C were downregulated. The RT-qPCR (21 DCM and 8 CNT samples validated the gene expression of SCN2B (p < 0.0001, KCNJ5 (p < 0.05, KCNJ8 (p < 0.05, CLIC2 (p < 0.05, and CACNB2 (p < 0.05. Furthermore, we performed an IPA analysis and we found a functional relationship between the different ion channels studied in this work. CONCLUSION: This study shows a differential expression of ion channel genes involved in cardiac contraction in DCM that might partly underlie the changes in left ventricular function observed in these patients. These results could be the basis for new genetic therapeutic approaches.

  9. Heterologous Expression of Tulip Petal Plasma Membrane Aquaporins in Pichia pastoris for Water Channel Analysis▿

    Science.gov (United States)

    Azad, Abul Kalam; Sawa, Yoshihiro; Ishikawa, Takahiro; Shibata, Hitoshi

    2009-01-01

    Water channels formed by aquaporins (AQPs) play an important role in the control of water homeostasis in individual cells and in multicellular organisms. Plasma membrane intrinsic proteins (PIPs) constitute a subclass of plant AQPs. TgPIP2;1 and TgPIP2;2 from tulip petals are members of the PIP family. In this study, we overexpressed TgPIP2;1 and TgPIP2;2 in Pichia pastoris and monitored their water channel activity (WCA) either by an in vivo spheroplast-bursting assay performed after hypo-osmotic shock or by growth assay. Osmolarity, pH, and inhibitors of AQPs, protein kinases (PKs), and protein phosphatases (PPs) affect the WCA of heterologous AQPs in this expression system. The WCA of TgPIP2;2-expressing spheroplasts was affected by inhibitors of PKs and PPs, which indicates that the water channel of this homologue is regulated by phosphorylation in P. pastoris. From the results reported herein, we suggest that P. pastoris can be employed as a heterologous expression system to assay the WCA of PIPs and to monitor the AQP-mediated channel gating mechanism, and it can be developed to screen inhibitors/effectors of PIPs. PMID:19251885

  10. Heterologous expression of tulip petal plasma membrane aquaporins in Pichia pastoris for water channel analysis.

    Science.gov (United States)

    Azad, Abul Kalam; Sawa, Yoshihiro; Ishikawa, Takahiro; Shibata, Hitoshi

    2009-05-01

    Water channels formed by aquaporins (AQPs) play an important role in the control of water homeostasis in individual cells and in multicellular organisms. Plasma membrane intrinsic proteins (PIPs) constitute a subclass of plant AQPs. TgPIP2;1 and TgPIP2;2 from tulip petals are members of the PIP family. In this study, we overexpressed TgPIP2;1 and TgPIP2;2 in Pichia pastoris and monitored their water channel activity (WCA) either by an in vivo spheroplast-bursting assay performed after hypo-osmotic shock or by growth assay. Osmolarity, pH, and inhibitors of AQPs, protein kinases (PKs), and protein phosphatases (PPs) affect the WCA of heterologous AQPs in this expression system. The WCA of TgPIP2;2-expressing spheroplasts was affected by inhibitors of PKs and PPs, which indicates that the water channel of this homologue is regulated by phosphorylation in P. pastoris. From the results reported herein, we suggest that P. pastoris can be employed as a heterologous expression system to assay the WCA of PIPs and to monitor the AQP-mediated channel gating mechanism, and it can be developed to screen inhibitors/effectors of PIPs.

  11. TRPV4 calcium-permeable channel is a novel regulator of oxidized LDL-induced macrophage foam cell formation.

    Science.gov (United States)

    Goswami, Rishov; Merth, Michael; Sharma, Shweta; Alharbi, Mazen O; Aranda-Espinoza, Helim; Zhu, Xiaoping; Rahaman, Shaik O

    2017-09-01

    Cardiovascular disease is the number one cause of death in United States, and atherosclerosis, a chronic inflammatory arterial disease, is the most dominant underlying pathology. Macrophages are thought to orchestrate atherosclerosis by generating lipid-laden foam cells and by secreting inflammatory mediators. Emerging data support a role for a mechanical factor, e.g., matrix stiffness, in regulation of macrophage function, vascular elasticity, and atherogenesis. However, the identity of the plasma membrane mechanosensor and the mechanisms by which pro-atherogenic signals are transduced/maintained are unknown. We have obtained evidence that TRPV4, an ion channel in the transient receptor potential vanilloid family and a known mechanosensor, is the likely mediator of oxidized low-density lipoprotein (oxLDL)-dependent macrophage foam cell formation, a critical process in atherogenesis. Specifically, we found that: i) genetic ablation of TRPV4 or pharmacologic inhibition of TRPV4 activity by a specific antagonist blocked oxLDL-induced macrophage foam cell formation, and ii) TRPV4 deficiency prevented pathophysiological range matrix stiffness or scratch-induced exacerbation of oxLDL-induced foam cell formation. Mechanistically, we found that: i) plasma membrane localization of TRPV4 was sensitized to the increasing level of matrix stiffness, ii) lack of foam cell formation in TRPV4 null cells was not due to lack of expression of CD36, a major receptor for oxLDL, and iii) TRPV4 channel activity regulated oxLDL uptake but not its binding on macrophages. Altogether, these findings identify a novel role for TRPV4 in regulating macrophage foam cell formation by modulating uptake of oxLDL. These findings suggest that therapeutic targeting of TRPV4 may provide a selective approach to the treatment of atherosclerosis. Copyright © 2017. Published by Elsevier Inc.

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

    DEFF Research Database (Denmark)

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

    2015-01-01

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

  13. Adrenomedullin increases the short-circuit current in the rat prostate: Receptors, chloride channels, the effects of cAMP and calcium ions and implications on fluid secretion.

    Science.gov (United States)

    Liao, S B; Cheung, K H; Cheung, M P L; Wong, P F; O, W S; Tang, F

    2014-05-01

    In this study, we have investigated the effects of adrenomedullin on chloride and fluid secretion in the rat prostate. The presence of adrenomedullin (ADM) in rat prostate was confirmed using immunostaining, and the molecular species was determined using gel filtration chromatography coupled with an enzyme-linked assay for ADM. The effects of ADM on fluid secretion were studied by short-circuit current technique in a whole mount preparation of the prostate in an Ussing chamber. The results indicated that the ADM level was higher in the ventral than the dorso-lateral prostate and the major molecular species was the active peptide. ADM increased the short-circuit current through both the cAMP- and calcium-activated chloride channels in the ventral lobe, but only through the calcium-activated channels in the dorso-lateral lobe. These stimulatory effects were blocked by the calcitonin gene-related peptide (CGRP) receptor antagonist, hCGRP8-37. We conclude that ADM may regulate prostatic fluid secretion through the chloride channels, which may affect the composition of the seminal plasma bathing the spermatozoa and hence fertility. © 2014 American Society of Andrology and European Academy of Andrology.

  14. Novel insights into the distribution of cardiac HCN channels: an expression study in the mouse heart.

    Science.gov (United States)

    Herrmann, Stefan; Layh, Beate; Ludwig, Andreas

    2011-12-01

    HCN pacemaker channels (I(f) channels) are believed to contribute to important functions in the heart; thus these channels became an attractive target for generating transgenic mouse mutants to elucidate their role in physiological and pathophysiological cardiac conditions. A full understanding of cardiac I(f) and the interpretation of studies using HCN mouse mutants require detailed information about the expression profile of the individual HCN subunits. Here we investigate the cardiac expression pattern of the HCN isoforms at the mRNA as well as at the protein level. The specificity of antibodies used was strictly confirmed by the use of HCN1, HCN2 and HCN4 knockout animals. We find a low, but highly differential HCN expression profile outside the cardiac conduction pathway including left and right atria and ventricles. Additionally HCN distribution was investigated in tissue slices of the sinoatrial node, the atrioventricular node, the bundle of His and the bundle branches. The conduction system was marked by acetylcholine esterase staining. HCN4 was confirmed as the predominant isoform of the primary pacemaker followed by a distinct expression of HCN1. In contrast HCN2 shows only a confined expression to individual pacemaker cells. Immunolabeling of the AV-node reveals also a pronounced specificity for HCN1 and HCN4. Compared to the SN and AVN we found a low but selective expression of HCN4 as the only isoform in the atrioventricular bundle. However in the bundle branches HCN1, HCN4 and also HCN2 show a prominent and selective expression pattern. Our results display a characteristic distribution of individual HCN isoforms in several cardiac compartments and reveal that beside HCN4, HCN1 represents the isoform which is selectively expressed in most parts of the conduction system suggesting a substantial contribution of HCN1 to pacemaking. 2011 Elsevier Ltd. All rights reserved.

  15. Expression and distribution of transient receptor potential (TRP) channels in bladder epithelium.

    Science.gov (United States)

    Yu, Weiqun; Hill, Warren G; Apodaca, Gerard; Zeidel, Mark L

    2011-01-01

    The urothelium is proposed to be a sensory tissue that responds to mechanical stress by undergoing dynamic membrane trafficking and neurotransmitter release; however, the molecular basis of this function is poorly understood. Transient receptor potential (TRP) channels are ideal candidates to fulfill such a role as they can sense changes in temperature, osmolarity, and mechanical stimuli, and several are reported to be expressed in the bladder epithelium. However, their complete expression profile is unknown and their cellular localization is largely undefined. We analyzed expression of all 33 TRP family members in mouse bladder and urothelium by RT-PCR and found 22 specifically expressed in the urothelium. Of the latter, 10 were chosen for closer investigation based on their known mechanosensory or membrane trafficking functions in other cell types. Western blots confirmed urothelial expression of TRPC1, TRPC4, TRPV1, TRPV2, TRPV4, TRPM4, TRPM7, TRPML1, and polycystins 1 and 2 (PKD1 and PKD2) proteins. We further defined the cellular and subcellular localization of all 10 TRP channels. TRPV2 and TRPM4 were prominently localized to the umbrella cell apical membrane, while TRPC4 and TRPV4 were identified on their abluminal surfaces. TRPC1, TRPM7, and TRPML1 were localized to the cytoplasm, while PKD1 and PKD2 were expressed on the apical and basolateral membranes of umbrella cells as well as in the cytoplasm. The cellular location of TRPV1 in the bladder has been debated, but colocalization with neuronal marker calcitonin gene-related peptide indicated clearly that it is present on afferent neurons that extend into the urothelium, but may not be expressed by the urothelium itself. These findings are consistent with the hypothesis that the urothelium acts as a sentinel and by expressing multiple TRP channels it is likely it can detect and presumably respond to a diversity of external stimuli and suggest that it plays an important role in urothelial signal

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1991-06-01

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

  17. Enhanced pre-synaptic glutamate release in deep-dorsal horn contributes to calcium channel alpha-2-delta-1 protein-mediated spinal sensitization and behavioral hypersensitivity

    Directory of Open Access Journals (Sweden)

    Dickenson Anthony H

    2009-02-01

    Full Text Available Abstract Nerve injury-induced expression of the spinal calcium channel alpha-2-delta-1 subunit (Cavα2δ1 has been shown to mediate behavioral hypersensitivity through a yet identified mechanism. We examined if this neuroplasticity modulates behavioral hypersensitivity by regulating spinal glutamatergic neurotransmission in injury-free transgenic mice overexpressing the Cavα2δ1 proteins in neuronal tissues. The transgenic mice exhibited hypersensitivity to mechanical stimulation (allodynia similar to the spinal nerve ligation injury model. Intrathecally delivered antagonists for N-methyl-D-aspartate (NMDA and α-amino-3-hydroxyl-5-methylisoxazole-4-propionic acid (AMPA/kainate receptors, but not for the metabotropic glutamate receptors, caused a dose-dependent allodynia reversal in the transgenic mice without changing the behavioral sensitivity in wild-type mice. This suggests that elevated spinal Cavα2δ1 mediates allodynia through a pathway involving activation of selective glutamate receptors. To determine if this is mediated by enhanced spinal neuronal excitability or pre-synaptic glutamate release in deep-dorsal horn, we examined wide-dynamic-range (WDR neuron excitability with extracellular recording and glutamate-mediated excitatory postsynaptic currents with whole-cell patch recording in deep-dorsal horn of the Cavα2δ1 transgenic mice. Our data indicated that overexpression of Cavα2δ1 in neuronal tissues led to increased frequency, but not amplitude, of miniature excitatory post synaptic currents mediated mainly by AMPA/kainate receptors at physiological membrane potentials, and also by NMDA receptors upon depolarization, without changing the excitability of WDR neurons to high intensity stimulation. Together, these findings support a mechanism of Cavα2δ1-mediated spinal sensitization in which elevated Cavα2δ1 causes increased pre-synaptic glutamate release that leads to reduced excitation thresholds of post-synaptic dorsal

  18. Enhanced pre-synaptic glutamate release in deep-dorsal horn contributes to calcium channel alpha-2-delta-1 protein-mediated spinal sensitization and behavioral hypersensitivity

    Science.gov (United States)

    Nguyen, David; Deng, Ping; Matthews, Elizabeth A; Kim, Doo-Sik; Feng, Guoping; Dickenson, Anthony H; Xu, Zao C; Luo, Z David

    2009-01-01

    Nerve injury-induced expression of the spinal calcium channel alpha-2-delta-1 subunit (Cavα2δ1) has been shown to mediate behavioral hypersensitivity through a yet identified mechanism. We examined if this neuroplasticity modulates behavioral hypersensitivity by regulating spinal glutamatergic neurotransmission in injury-free transgenic mice overexpressing the Cavα2δ1 proteins in neuronal tissues. The transgenic mice exhibited hypersensitivity to mechanical stimulation (allodynia) similar to the spinal nerve ligation injury model. Intrathecally delivered antagonists for N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxyl-5-methylisoxazole-4-propionic acid (AMPA)/kainate receptors, but not for the metabotropic glutamate receptors, caused a dose-dependent allodynia reversal in the transgenic mice without changing the behavioral sensitivity in wild-type mice. This suggests that elevated spinal Cavα2δ1 mediates allodynia through a pathway involving activation of selective glutamate receptors. To determine if this is mediated by enhanced spinal neuronal excitability or pre-synaptic glutamate release in deep-dorsal horn, we examined wide-dynamic-range (WDR) neuron excitability with extracellular recording and glutamate-mediated excitatory postsynaptic currents with whole-cell patch recording in deep-dorsal horn of the Cavα2δ1 transgenic mice. Our data indicated that overexpression of Cavα2δ1 in neuronal tissues led to increased frequency, but not amplitude, of miniature excitatory post synaptic currents mediated mainly by AMPA/kainate receptors at physiological membrane potentials, and also by NMDA receptors upon depolarization, without changing the excitability of WDR neurons to high intensity stimulation. Together, these findings support a mechanism of Cavα2δ1-mediated spinal sensitization in which elevated Cavα2δ1 causes increased pre-synaptic glutamate release that leads to reduced excitation thresholds of post-synaptic dorsal horn neurons to innocuous

  19. Upregulation of calcium channel alpha-2-delta-1 subunit in dorsal horn contributes to spinal cord injury-induced tactile allodynia.

    Science.gov (United States)

    Kusuyama, Kazuki; Tachibana, Toshiya; Yamanaka, Hiroki; Okubo, Masamichi; Yoshiya, Shinichi; Noguchi, Koichi

    2018-01-31

    Spinal cord injury (SCI) commonly results not only in motor paralysis but also in the emergence of neuropathic pain (NeuP), both of which can impair the quality of life for patients with SCI. In the clinical field, it is well known that pregabalin, which binds to the voltage-gated calcium channel alpha-2-delta-1 (α 2 δ-1) subunit has therapeutic effects on NeuP after SCI. A previous study has demonstrated that SCI increased α 2 δ-1 in the L4-L6 dorsal spinal cord of SCI rats by Western blot analysis and that the increase of α 2 δ-1 was correlated with tactile allodynia of the hind paw. However, the detailed feature of an increase in α 2 δ-1 protein in the spinal dorsal horn and the mechanism of pregabalin effect on SCI-induced NeuP have not been fully examined. This study aimed to examine the detailed distribution of α 2 δ-1 expression in the lumbar spinal cord after thoracic SCI in rats and the correlation of the therapeutic effect of pregabalin in SCI rats. Male Sprague-Dawley rats underwent thoracic (T10) spinal cord contusion injury using the IH impactor device. Spinal cord injury rats received pregabalin (30 mg/kg) once a day for 2 weeks over a 4-week period after SCI. The mechanical threshold in the rat hind paw was measured over 4 weeks. Alpha-2-delta-1 expression in the lumbar spinal cord and in the dorsal root ganglion (DRG) was analyzed using immunohistochemistry and in situ hybridization histochemistry. A significant reduction of the withdrawal threshold of mechanical stimuli to the hind paw was observed for 2 weeks and continued at least 4 weeks after SCI. In the control rats, expression of α 2 δ-1 immunoreactivity was detected mainly in laminae I and II in the lumbar dorsal horn. Thoracic SCI significantly increased α 2 δ-1 immunoreactivity in laminae I and II in the lumbar dorsal horn 4 weeks after SCI; however, thoracic SCI did not affect the expression of α 2 δ-1 mRNA in the L4 and L5 DRGs. Meanwhile, the signal intensity of α 2

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

    Science.gov (United States)

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

    2018-01-01

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

  1. Comparison of the efficacy of dihydropyridine calcium channel blockers in African American patients with hypertension. ISHIB Investigators Group. International Society on Hypertension in Blacks.

    Science.gov (United States)

    Hall, W D; Reed, J W; Flack, J M; Yunis, C; Preisser, J

    1998-10-12

    Hypertension is a prevalent disease among African Americans, and successful treatment rates are low. Since calcium channel blockers are well-tolerated and efficacious in African Americans, we undertook this study to compare the efficacy, safety, and tolerability of 3 commonly prescribed calcium channel blockers: amlodipine besylate (Norvasc), nifedipine coat core (CC) (Adalat CC), and nifedipine gastrointestinal therapeutic system (GITS) (Procardia XL). One hundred ninety-two hypertensive patients across 10 study centers were randomly assigned to double-blind monotherapy with amlodipine besylate (5 mg/d), nifedipine CC (30 mg/d), or nifedipine GITS (30 mg/d) for 8 weeks. Patients not achieving therapeutic response after 4 weeks had their dose doubled for the next 4 weeks. The primary end point was a comparison of the average reduction (week 8 minus baseline) in 24-hour ambulatory diastolic blood pressure (DBP). Secondary end points included a comparison of average 24-hour ambulatory systolic blood pressure (SBP), office SBP or DBP reduction, responder rates, safety, and tolerability. One hundred sixty-three patients were evaluable for efficacy after 8 weeks. There was no significant difference in the average 24-hour ambulatory DBP (-8.5, -9.0, and -6.1 mm Hg, respectively) or SBP (-14.3, -15.7, and -11.8 mm Hg, respectively) reduction. Average office SBP and DBP were reduced to a comparable degree (19-22 mm Hg [P =.50] and 12-14 mm Hg [P =.51], respectively). Responder rates (DBP or = 10 mm Hg) were similar (P = .38). Discontinuation rates and adverse event frequency were distributed similarly across the 3 treatment groups. The efficacy, safety, and tolerability of the 3 dihydropyridine calcium channel blockers are equivalent in African Americans with stages 1 and 2 hypertension.

  2. Decreased calcium pump expression in human erythrocytes is connected to a minor haplotype in the ATP2B4 gene.

    Science.gov (United States)

    Zámbó, Boglárka; Várady, György; Padányi, Rita; Szabó, Edit; Németh, Adrienn; Langó, Tamás; Enyedi, Ágnes; Sarkadi, Balázs

    2017-07-01

    Plasma membrane Ca 2+ -ATPases are key calcium exporter proteins in most tissues, and PMCA4b is the main calcium transporter in the human red blood cells (RBCs). In order to assess the expression level of PMCA4b, we have developed a flow cytometry and specific antibody binding method to quantitatively detect this protein in the erythrocyte membrane. Interestingly, we found several healthy volunteers showing significantly reduced expression of RBC-PMCA4b. Western blot analysis of isolated RBC membranes confirmed this observation, and indicated that there are no compensatory alterations in other PMCA isoforms. In addition, reduced PMCA4b levels correlated with a lower calcium extrusion capacity in these erythrocytes. When exploring the potential genetic background of the reduced PMCA4b levels, we found no missense mutations in the ATP2B4 coding regions, while a formerly unrecognized minor haplotype in the predicted second promoter region closely correlated with lower erythrocyte PMCA4b protein levels. In recent GWA studies, SNPs in this ATP2B4 haplotype have been linked to reduced mean corpuscular hemoglobin concentrations (MCHC), and to protection against malaria infection. Our data suggest that an altered regulation of gene expression is responsible for the reduced RBC-PMCA4b levels that is probably linked to the development of human disease-related phenotypes. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Lipid nanocapsules containing the non-ionic surfactant Solutol HS15 inhibit the transport of calcium through hyperforin-activated channels in neuronal cells.

    Science.gov (United States)

    Chauvet, Sylvain; Barras, Alexandre; Boukherroub, Rabah; Bouron, Alexandre

    2015-12-01

    Hyperforin is described as a natural antidepressant inhibiting the reuptake of neurotransmitters and also activating cation channels. However the blood-brain barrier limits the access to the brain of this biomolecule. To circumvent this problem it was envisaged to encapsulate hyperforin into biomimetic lipid nano-carriers like lipid nanocapsules (LNCs). When testing the safety of 25 nm LNCs it appeared that they strongly blocked hyperforin-activated Ca2+ channels of cultured cortical neurons. This inhibition was due to one of their main component: solutol HS15 (polyoxyethylene-660-12-hydroxy stearate), a non-ionic soluble surfactant. Solutol HS15 rapidly depresses in a concentration-dependent manner the entry of Ca2+ through hyperforin-activated channels without influencing store-operated channels. This effect is mimicked by Brij58 but not by PEG600, indicating that the lipid chain of Solutol HS15 is important in determining its effects on the channels. The inhibition of the Ca2+ fluxes depends on the cellular cholesterol content; it is stronger after depleting cholesterol with methyl-β-cyclodextrin and is nearly absent on cells cultured in a cholesterol-rich medium. When chronically applied for 24 h, Solutol HS15 slightly up-regulates the entry of Ca2+ through hyperforin-activated channels. Similar observations were made when testing 25 nm lipid nanocapsules containing the surfactant Solutol HS15. Altogether, this study shows that Solutol HS15 perturbs in a cholesterol-dependent manner the activity of some neuronal channels. This is the first demonstration that LNCs containing this surfactant can influence cellular calcium signaling in the brain, a finding that can have important clinical implications. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Molecular and functional expression of high conductance Ca 2+ activated K+ channels in the eel intestinal epithelium

    DEFF Research Database (Denmark)

    Lionetto, Maria G; Rizzello, Antonia; Giordano, Maria E

    2008-01-01

    Several types of K(+) channels have been identified in epithelial cells. Among them high conductance Ca(2+)-activated K(+) channels (BK channels) are of relevant importance for their involvement in regulatory volume decrease (RVD) response following hypotonic stress. The aim of the present work...... was to investigate the functional and molecular expression of BK in the eel intestine, which is a useful experimental model for cell volume regulation research. In the present paper using rat BK channel-specific primer, a RT-PCR signal of 696 pb cDNA was detected in eel intestine, whole nucleotide sequence showed...... high similarity (83%) to the alpha subunit of BK channel family. BK channel protein expression was verified by immunoblotting and confocal microscopy, while the functional role of BK channels in epithelial ion transport mechanisms and cell volume regulation was examined by electrophysiological...

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

    Directory of Open Access Journals (Sweden)

    H SADRAEI

    2000-12-01

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

  6. Expression and activity of acid-sensing ion channels in the mouse anterior pituitary.

    Directory of Open Access Journals (Sweden)

    Jianyang Du

    Full Text Available Acid sensing ion channels (ASICs are proton-gated cation channels that are expressed in the nervous system and play an important role in fear learning and memory. The function of ASICs in the pituitary, an endocrine gland that contributes to emotions, is unknown. We sought to investigate which ASIC subunits were present in the pituitary and found mRNA expression for all ASIC isoforms, including ASIC1a, ASIC1b, ASIC2a, ASIC2b, ASIC3 and ASIC4. We also observed acid-evoked ASIC-like currents in isolated anterior pituitary cells that were absent in mice lacking ASIC1a. The biophysical properties and the responses to PcTx1, amiloride, Ca2+ and Zn2+ suggested that ASIC currents were mediated predominantly by heteromultimeric channels that contained ASIC1a and ASIC2a or ASIC2b. ASIC currents were also sensitive to FMRFamide (Phe-Met-Arg-Phe amide, suggesting that FMRFamide-like compounds might endogenously regulate pituitary ASICs. To determine whether ASICs might regulate pituitary cell function, we applied low pH and found that it increased the intracellular Ca2+ concentration. These data suggest that ASIC channels are present and functionally active in anterior pituitary cells and may therefore influence their function.

  7. Safety assessment of the calcium-binding protein, apoaequorin, expressed by Escherichia coli.

    Science.gov (United States)

    Moran, Daniel L; Tetteh, Afua O; Goodman, Richard E; Underwood, Mark Y

    2014-07-01

    Calcium-binding proteins are ubiquitous modulators of cellular activity and function. Cells possess numerous calcium-binding proteins that regulate calcium concentration in the cytosol by buffering excess free calcium ion. Disturbances in intracellular calcium homeostasis are at the heart of many age-related conditions making these proteins targets for therapeutic intervention. A calcium-binding protein, apoaequorin, has shown potential utility in a broad spectrum of applications for human health and well-being. Large-scale recombinant production of the protein has been successful; enabling further research and development and commercialization efforts. Previous work reported a 90-day subchronic toxicity test that demonstrated this protein has no toxicity by oral exposure in Sprague-Dawley rodents. The current study assesses the allergenic potential of the purified protein using bioinformatic analysis and simulated gastric digestion. The results from the bioinformatics searches with the apoaequorin sequence show the protein is not a known allergen and not likely to cross-react with known allergens. Apoaequorin is easily digested by pepsin, a characteristic commonly exhibited by many non-allergenic dietary proteins. From these data, there is no added concern of safety due to unusual stability of the protein by ingestion. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Molecular imaging of in vivo calcium ion expression in area postrema of total sleep deprived rats: Implications for cardiovascular regulation by TOF-SIMS analysis

    Science.gov (United States)

    Mai, Fu-Der; Chen, Li-You; Ling, Yong-Chien; Chen, Bo-Jung; Wu, Un-In; Chang, Hung-Ming

    2010-05-01

    Excessive calcium influx in chemosensitive neurons of area postrema (AP) is detrimental for sympathetic activation and participates in the disruption of cardiovascular activities. Since total sleep deprivation (TSD) is a stressful condition known to harm the cardiovascular function, the present study is aimed to determine whether the in vivo calcium expression in AP would significantly alter following TSD by the use of time-of-flight secondary ion mass spectrometry (TOF-SIMS) and calretinin (a specific calcium sensor protein in AP neurons) immunohistochemistry. The results indicated that in normal rats, the calcium intensity was estimated to be 0.5 × 10 5 at m/ z 40.08. However, following TSD, the intensity for calcium ions was greatly increased to 1.2 × 10 5. Molecular imaging revealed that after TSD, various strongly expressed calcium signals were distributed throughout AP with clear identified profiles instead of randomly scattered within this region in normal rats. Immunohistochemical staining corresponded well with ionic image in which a majority of calcium-enriched gathering co-localized with calretinin positive neurons. The functional significance of TSD-induced calcium augmentation was demonstrated by increased heart rate and mean arterial pressure, clinical markers for cardiovascular dysfunction. Considering AP-mediated sympathetic activation is important for cardiovascular regulation, exaggerated calcium influx in AP would render this neurocircuitry more vulnerable to over-excitation, which might serve as the underlying mechanism for the development of TSD-relevant cardiovascular deficiency.

  9. Spinal afferent neurons projecting to the rat lung and pleura express acid sensitive channels

    Directory of Open Access Journals (Sweden)

    Kummer Wolfgang

    2006-07-01

    Full Text Available Abstract Background The acid sensitive ion channels TRPV1 (transient receptor potential vanilloid receptor-1 and ASIC3 (acid sensing ion channel-3 respond to tissue acidification in the range that occurs during painful conditions such as inflammation and ischemia. Here, we investigated to which extent they are expressed by rat dorsal root ganglion neurons projecting to lung and pleura, respectively. Methods The tracer DiI was either injected into the left lung or applied to the costal pleura. Retrogradely labelled dorsal root ganglion neurons were subjected to triple-labelling immunohistochemistry using antisera against TRPV1, ASIC3 and neurofilament 68 (marker for myelinated neurons, and their soma diameter was measured. Results Whereas 22% of pulmonary spinal afferents contained neither channel-immunoreactivity, at least one is expressed by 97% of pleural afferents. TRPV1+/ASIC3- neurons with probably slow conduction velocity (small soma, neurofilament 68-negative were significantly more frequent among pleural (35% than pulmonary afferents (20%. TRPV1+/ASIC3+ neurons amounted to 14 and 10% respectively. TRPV1-/ASIC3+ neurons made up between 44% (lung and 48% (pleura of neurons, and half of them presumably conducted in the A-fibre range (larger soma, neurofilament 68-positive. Conclusion Rat pleural and pulmonary spinal afferents express at least two different acid-sensitive channels that make them suitable to monitor tissue acidification. Patterns of co-expression and structural markers define neuronal subgroups that can be inferred to subserve different functions and may initiate specific reflex responses. The higher prevalence of TRPV1+/ASIC3- neurons among pleural afferents probably reflects the high sensitivity of the parietal pleura to painful stimuli.

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

    Science.gov (United States)

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

    2015-03-01

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

  11. [Effect of hydrostatic pressure on intracellular free calcium concentration and transient receptor potential vanilloid expression in human bladder smooth muscle cells].

    Science.gov (United States)

    Han, Zhenwei; Wang, Kunjie; Chen, Lin; Wei, Tangqiang; Luo, Deyi; Li, Shengfu

    2012-04-01

    To explore the effect of hydrostatic pressure on intracellular free calcium concentration ([Ca2+]i) and the gene expression of transient receptor potential vanilloid (TRPV) in cultured human bladder smooth muscle cells (hb-SMCs), and to preliminarily probe into the possible molecular mechanism of hb-SMCs proliferation stimulated by hydrostatic pressure. The passage 6-7 hb-SMCs were loaded with Ca2+ indicator Fluo-3/AM. When the hb-SMCs were under 0 cm H2O (1cm H2O = 0.098 kPa) (group A) or 200 cm H2O hydrostatic pressure for 30 minutes (group B) and then removing the 200 cm H2O hydrostatic pressure (group C), the [Ca2+]i was measured respectively by inverted laser scanning confocal microscope. When the hb-SMCs were given the 200 cm H2O hydrostatic pressure for 0 hour, 2 hours, 6 hours, 12 hours, and 24 hours, the mRNA expressions of TRPV1, TRPV2, and TRPV4 were detected by RT-PCR technique. The [Ca2+]i of group A, group B, and group C were (100.808 +/- 1.724), (122.008 +/- 1.575), and (99.918 +/- 0.887) U, respectively; group B was significantly higher than groups A and C (P pressure (t = 0.919, P = 0.394). The TRPV1, TRPV2, and TRPV4 genes expressed in hb-SMCs under 200 cm H2O hydrostatic pressure at 0 hour, 2 hours, 6 hours, 12 hours, and 24 hours, but the expressions had no obvious changes with time. There was no significant difference in the expressions of TRPV1, TRPV2, and TRPV4 among 3 groups (P > 0.05). The [Ca2+]i of hb-SMCs increases significantly under high hydrostatic pressure. As possible genes in stretch-activated cation channel, the TRPV1, TRPV2, and TRPV4 express in hb-SMCs under 200 cm H2O hydrostatic pressure. It is possible that the mechanical pressure regulates the [Ca2+]i of hb-SMCs by opening the stretch-activated cation channel rather than up-regulating its expression.

  12. TRPV3, a thermosensitive channel is expressed in mouse distal colon epithelium

    International Nuclear Information System (INIS)

    Ueda, Takashi; Yamada, Takahiro; Ugawa, Shinya; Ishida, Yusuke; Shimada, Shoichi

    2009-01-01

    The thermo-transient receptor potential (thermoTRP) subfamily is composed of channels that are important in nociception and thermo-sensing. Here, we show a selective expression of TRPV3 channel in the distal colon throughout the gastrointestinal tract. Expression analyses clearly revealed that TRPV3 mRNA and proteins were expressed in the superficial epithelial cells of the distal colon, but not in those of the stomach, duodenum or proximal colon. In a subset of primary epithelial cells cultured from the distal colon, carvacrol, an agonist for TRPV3, elevated cytosolic Ca 2+ concentration in a concentration-dependent manner. This response was inhibited by ruthenium red, a TRPV channel antagonist. Organotypic culture supported that the carvacrol-responsive cells were present in superficial epithelial cells. Moreover, application of carvacrol evoked ATP release in primary colonic epithelial cells. We conclude that TRPV3 is present in absorptive cells in the distal colon and may be involved in a variety of cellular functions.

  13. The cell surface expressed nucleolin is a glycoprotein that triggers calcium entry into mammalian cells

    International Nuclear Information System (INIS)

    Losfeld, Marie-Estelle; Khoury, Diala El; Mariot, Pascal; Carpentier, Mathieu; Krust, Bernard; Briand, Jean-Paul; Mazurier, Joel; Hovanessian, Ara G.; Legrand, Dominique

    2009-01-01

    Nucleolin is an ubiquitous nucleolar phosphoprotein involved in fundamental aspects of transcription regulation, cell proliferation and growth. It has also been described as a shuttling molecule between nucleus, cytosol and the cell surface. Several studies have demonstrated that surface nucleolin serves as a receptor for various extracellular ligands implicated in cell proliferation, differentiation, adhesion, mitogenesis and angiogenesis. Previously, we reported that nucleolin in the extranuclear cell compartment is a glycoprotein containing N- and O-glycans. In the present study, we show that glycosylation is an essential requirement for surface nucleolin expression, since it is prevented when cells are cultured in the presence of tunicamycin, an inhibitor of N-glycosylation. Accordingly, surface but not nuclear nucleolin is radioactively labeled upon metabolic labeling of cells with [ 3 H]glucosamine. Besides its well-demonstrated role in the internalization of specific ligands, here we show that ligand binding to surface nucleolin could also induce Ca 2+ entry into cells. Indeed, by flow cytometry, microscopy and patch-clamp experiments, we show that the HB-19 pseudopeptide, which binds specifically surface nucleolin, triggers rapid and intense membrane Ca 2+ fluxes in various types of cells. The use of several drugs then indicated that Store-Operated Ca 2+ Entry (SOCE)-like channels are involved in the generation of these fluxes. Taken together, our findings suggest that binding of an extracellular ligand to surface nucleolin could be involved in the activation of signaling pathways by promoting Ca 2+ entry into cells

  14. Association of CD147 and Calcium Exporter PMCA4 Uncouples IL-2 Expression from Early TCR Signaling.

    Science.gov (United States)

    Supper, Verena; Schiller, Herbert B; Paster, Wolfgang; Forster, Florian; Boulègue, Cyril; Mitulovic, Goran; Leksa, Vladimir; Ohradanova-Repic, Anna; Machacek, Christian; Schatzlmaier, Philipp; Zlabinger, Gerhard J; Stockinger, Hannes

    2016-02-01

    The Ig superfamily member CD147 is upregulated following T cell activation and was shown to serve as a negative regulator of T cell proliferation. Thus, Abs targeting CD147 are being tested as new treatment strategies for cancer and autoimmune diseases. How CD147 mediates immunosuppression and whether association with other coreceptor complexes is needed have remained unknown. In the current study, we show that silencing of CD147 in human T cells increases IL-2 production without affecting the TCR proximal signaling components. We mapped the immunosuppressive moieties of CD147 to its transmembrane domain and Ig-like domain II. Using affinity purification combined with mass spectrometry, we determined the domain specificity of CD147 interaction partners and identified the calcium exporter plasma membrane calcium ATPase isoform 4 (PMCA4) as the interaction partner of the immunosuppressive moieties of CD147. CD147 does not control the proper membrane localization of PMCA4, but PMCA4 is essential for the CD147-dependent inhibition of IL-2 expression via a calcium-independent mechanism. In summary, our data show that CD147 interacts via its immunomodulatory domains with PMCA4 to bypass TCR proximal signaling and inhibit IL-2 expression. Copyright © 2016 by The American Association of Immunologists, Inc.

  15. Transcriptome Analysis of Calcium- and Hormone-Related Gene Expressions during Different Stages of Peanut Pod Development

    Science.gov (United States)

    Li, Yan; Meng, Jingjing; Yang, Sha; Guo, Feng; Zhang, Jialei; Geng, Yun; Cui, Li; Wan, Shubo; Li, Xinguo

    2017-01-01

    Peanut is one of the calciphilous plants. Calcium serves as a ubiquitous central hub in a large number of signaling pathways. In the field, free calcium ion (Ca2+)-deficient soil can result in unfilled pods. Four pod stages were analyzed to determine the relationship between Ca2+ excretion and pod development. Peanut shells showed Ca2+ excretion at all four stages; however, both the embryo of Stage 4 (S4) and the red skin of Stage 3 (S3) showed Ca2+ absorbance. These results showed that embryo and red skin of peanut need Ca2+ during development. In order to survey the relationship among calcium, hormone and seed development from gene perspective, we further analyzed the seed transcriptome at Stage 2 (S2), S3, and S4. About 70 million high quality clean reads were generated, which were assembled into 58,147 unigenes. By comparing these three stages, total 4,457 differentially expressed genes were identified. In these genes, 53 Ca2+ related genes, 40 auxin related genes, 15 gibberellin genes, 20 ethylene related genes, 2 abscisic acid related genes, and 7 cytokinin related genes were identified. Additionally, a part of them were validated by qRT-PCR. Most of their expressions changed during the pod development. Since some reports showed that Ca2+ signal transduction pathway is involved in hormone regulation pathway, these results implied that peanut seed development might be regulated by the collaboration of Ca2+ signal transduction pathway and hormone regulation pathway. PMID:28769950

  16. Transcriptome Analysis of Calcium- and Hormone-Related Gene Expressions during Different Stages of Peanut Pod Development

    Directory of Open Access Journals (Sweden)

    Yan Li

    2017-07-01

    Full Text Available Peanut is one of the calciphilous plants. Calcium serves as a ubiquitous central hub in a large number of signaling pathways. In the field, free calcium ion (Ca2+-deficient soil can result in unfilled pods. Four pod stages were analyzed to determine the relationship between Ca2+ excretion and pod development. Peanut shells showed Ca2+ excretion at all four stages; however, both the embryo of Stage 4 (S4 and the red skin of Stage 3 (S3 showed Ca2+ absorbance. These results showed that embryo and red skin of peanut need Ca2+ during development. In order to survey the relationship among calcium, hormone and seed development from gene perspective, we further analyzed the seed transcriptome at Stage 2 (S2, S3, and S4. About 70 million high quality clean reads were generated, which were assembled into 58,147 unigenes. By comparing these three stages, total 4,457 differentially expressed genes were identified. In these genes, 53 Ca2+ related genes, 40 auxin related genes, 15 gibberellin genes, 20 ethylene related genes, 2 abscisic acid related genes, and 7 cytokinin related genes were identified. Additionally, a part of them were validated by qRT-PCR. Most of their expressions changed during the pod development. Since some reports showed that Ca2+ signal transduction pathway is involved in hormone regulation pathway, these results implied that peanut seed development might be regulated by the collaboration of Ca2+ signal transduction pathway and hormone regulation pathway.

  17. Vitamin K3 inhibits mouse uterine contraction in vitro via interference with the calcium transfer and the potassium channels.

    Science.gov (United States)

    Zhang, Xian-Xia; Lu, Li-Min; Wang, Li

    2016-08-05

    Previous studies have demonstrated vitamin K3 had a great relief to smooth muscle spastic disorders, but no researches have yet pinpointed its possible anti-contractile activity in the uterus. Here, we evaluated the effect of vitamin K3 on myometrial contractility and explored the possible mechanisms of vitamin K3 action. Myograph apparatus were used to record the changes in contractility of isolated mouse uterine strips in a tissue bath. Uterine strips were exposed to vitamin K3 or vehicle. Vitamin K3 suppressed spontaneous contractions in a concentration dependent manner. It significantly decreased the contractile frequency induced by PGF2ɑ but not their amplitude (expect 58.0 μM). Prior incubation with vitamin K3 reduced the effectiveness of PGF2ɑ-induced contraction. The antispasmodic effect of vitamin K3 was also sensitive to potassium channel blockers, such as tetraethylammonium, 4-aminopyridine, iberiotoxin) but not to the nitric oxide related pathway blockers. High concentrations (29.0, 58.0 μM) of vitamin K3 weakened the Ca(2+) dose response and inhibited phase 1 contraction (intracellular stored calcium release). These dates suggest that vitamin K3 specifically suppresses myometrial contractility by affecting calcium and potassium channels; thus, this approach has potential therapy for uterine contractile activity related disorders. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. When lithium does not help: the use of anticonvulsants and calcium channel blockers in the treatment of bipolar disorder in the older person.

    Science.gov (United States)

    Masters, J C

    1996-01-01

    Although anticonvulsant agents and calcium channel blockers do not have any clear advantages over lithium, they do offer patients who cannot (or will not) take lithium another treatment option. It is not yet clear from the literature who will respond best to which drug or combination of drugs. The nurse should be supportive to the patients and family, in what may be a drawn out process, to find the best treatment. Optimism is justified because a lack of response to one drug is not indicative of nonresponse to other drugs. It is important to actively treat bipolar disorder because each episode of mania increases the risk of progression of the illness, with increasingly severe episodes occurring closer together. Bipolar disorder has high social costs (legal, financial, and relationship problems) that make improvements in treatment important for the patient and society. Anticonvulsant agents and calcium channel blockers may also be useful in treating depression. The number of people whose depressive symptoms respond is far less (25% to 30%) than the number who respond to the anti-manic effects, but this is an option when antidepressants and electroconvulsive therapy are not effective.

  19. Preventing effect of L-type calcium channel blockade on electrophysiological alterations in dentate gyrus granule cells induced by entorhinal amyloid pathology.

    Directory of Open Access Journals (Sweden)

    Hamid Gholami Pourbadie

    Full Text Available The entorhinal cortex (EC is one of the earliest affected brain regions in Alzheimer's disease (AD. EC-amyloid pathology induces synaptic failure in the dentate gyrus (DG with resultant behavioral impairment, but there is little known about its impact on neuronal properties in the DG. It is believed that calcium dyshomeostasis plays a pivotal role in the etiology of AD. Here, the effect of the EC amyloid pathogenesis on cellular properties of DG granule cells and also possible neuroprotective role of L-type calcium channel blockers (CCBs, nimodipine and isradipine, were investigated. The amyloid beta (Aβ 1-42 was injected bilaterally into the EC of male rats and one week later, electrophysiological properties of DG granule cells were assessed. Voltage clamp recording revealed appearance of giant sIPSC in combination with a decrease in sEPSC frequency which was partially reversed by CCBs in granule cells from Aβ treated rats. EC amyloid pathogenesis induced a significant reduction of input resistance (Rin accompanied by a profound decreased excitability in the DG granule cells. However, daily administration of CCBs, isradipine or nimodipine (i.c.v. for 6 days, almost preserved the normal excitability against Aβ. In conclusion, lower tendency to fire AP along with reduced Rin suggest that DG granule cells might undergo an alteration in the membrane ion channel activities which finally lead to the behavioral deficits observed in animal models and patients with early-stage Alzheimer's disease.

  20. Resveratrol Protects Against Ultraviolet A-Mediated Inhibition of the Phagocytic Function of Human Retinal Pigment Epithelial Cells Via Large-Conductance Calcium-Activated Potassium Channels

    Directory of Open Access Journals (Sweden)

    Shwu-Jiuan Sheu

    2009-07-01

    Full Text Available This study was undertaken to examine the protective effect of resveratrol on human retinal pigment epithelial (RPE cell phagocytosis against ultraviolet irradiation damage. Cultured RPE cells were exposed to ultraviolet A (UVA, 20 minutes irradiation, and treated with meclofenamic acid (30μM, 20 minutes, paxilline (100 μM, 20 minutes or resveratrol (10μM, 20 minutes. Meclofenamic acid and resveratrol were given after exposure to UVA. Pretreatment with meclofenamic acid, resveratrol or paxilline before UVA irradiation was also performed. Fluorescent latex beads were then fed for 4 hours and the phagocytotic function was assessed by flow cytometry. UVA irradiation inhibited the phagocytic function of human RPE cells. The large-conductance calcium-activated potassium channel activator meclofenamic acid ameliorated the damage caused by UVA irradiation. Pretreatment with resveratrol acid also provided protection against damage caused by UVA. Posttreatment with meclofenamic acid offered mild protection, whereas resveratrol did not. In conclusion, the red wine flavonoid resveratrol ameliorated UVA-mediated inhibition of human RPE phagocytosis. The underlying mechanism might involve the large-conductance calcium-activated potassium channels.

  1. Troponin T3 regulates nuclear localization of the calcium channel Ca{sub v}β{sub 1a} subunit in skeletal muscle

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Tan; Taylor, Jackson; Jiang, Yang [Department of Internal Medicine-Gerontology, Wake Forest School of Medicine, Winston-Salem, NC 27157 (United States); Pereyra, Andrea S. [Department of Histology, National University of La Plata, 1900 La Plata (Argentina); Messi, Maria Laura; Wang, Zhong-Min [Department of Internal Medicine-Gerontology, Wake Forest School of Medicine, Winston-Salem, NC 27157 (United States); Hereñú, Claudia [Department of Histology, National University of La Plata, 1900 La Plata (Argentina); Delbono, Osvaldo, E-mail: odelbono@wakehealth.edu [Department of Internal Medicine-Gerontology, Wake Forest School of Medicine, Winston-Salem, NC 27157 (United States); Neuroscience Program, Wake Forest School of Medicine, Winston-Salem, NC 27157 (United States)

    2015-08-15

    The voltage-gated calcium channel (Ca{sub v}) β{sub 1a} subunit (Ca{sub v}β{sub 1a}) plays an important role in excitation–contraction coupling (ECC), a process in the myoplasm that leads to muscle-force generation. Recently, we discovered that the Ca{sub v}β{sub 1a} subunit travels to the nucleus of skeletal muscle cells where it helps to regulate gene transcription. To determine how it travels to the nucleus, we performed a yeast two-hybrid screening of the mouse fast skeletal muscle cDNA library and identified an interaction with troponin T3 (TnT3), which we subsequently confirmed by co-immunoprecipitation and co-localization assays in mouse skeletal muscle in vivo and in cultured C2C12 muscle cells. Interacting domains were mapped to the leucine zipper domain in TnT3 COOH-terminus (160–244 aa) and Ca{sub v}β{sub 1a} NH{sub 2}-terminus (1–99 aa), respectively. The double fluorescence assay in C2C12 cells co-expressing TnT3/DsRed and Ca{sub v}β{sub 1a}/YFP shows that TnT3 facilitates Ca{sub v}β{sub 1a} nuclear recruitment, suggesting that the two proteins play a heretofore unknown role during early muscle differentiation in addition to their classical role in ECC regulation. - Highlights: • Previously, we demonstrated that Ca{sub v}β{sub 1a} is a gene transcription regulator. • Here, we show that TnT3 interacts with Ca{sub v}β{sub 1a}. • We mapped TnT3 and Ca{sub v}β{sub 1a} interaction domain. • TnT3 facilitates Ca{sub v}β{sub 1a} nuclear enrichment. • The two proteins play a heretofore unknown role during early muscle differentiation.

  2. Modulation of intracellular Ca2+ via L-type calcium channels in heart cells by the autoantibody directed against the second extracellular loop of the alpha1-adrenoceptors.

    Science.gov (United States)

    Bkaily, Ghassan; El-Bizri, Nesrine; Bui, Michel; Sukarieh, Rami; Jacques, Danielle; Fu, Michael L X

    2003-03-01

    The effects of methoxamine, a selective alpha1-adrenergic receptor agonist, and the autoantibody directed against the second extracellular loop of alpha1-adrenoceptors were studied on intracellular free Ca2+ levels using confocal microscopy and ionic currents using the whole-cell patch clamp technique in single cells of 10-day-old embryonic chick and 20-week-old fetal human hearts. We observed that like methoxamine, the autoantibody directed against the second extracellular loop of alpha1-adrenoreceptors significantly increased the L-type calcium current (I(Ca(L))) but had no effect on the T-type calcium current (I(Ca(T))), the delayed outward potassium current, or the fast sodium current. This effect of the autoantibody was prevented by a prestimulation of the receptors with methoxamine and vice versa. Moreover, treating the cells with prazosin, a selective alpha1-adrenergic receptor antagonist blocked the methoxamine and the autoantibody-induced increase in I(Ca(L)), respectively. In absence of prazosin, both methoxamine and the autoantibody showed a substantial enhancement in the frequency of cell contraction and that of the concomitant cytosolic and nuclear free Ca2+ variations. The subsequent addition of nifedipine, a specific L-type Ca2+ channel blocker, reversed not only the methoxamine or the autoantibody-induced effect but also completely abolished cell contraction. These results demonstrated that functional alpha1-adrenoceptors exist in both 10-day-old embryonic chick and 20-week-old human fetal hearts and that the autoantibody directed against the second extracellular loop of this type of receptors plays an important role in stimulating their activity via activation of L-type calcium channels. This loop seems to have a functional significance by being the target of alpha1-receptor agonists like methoxamine.

  3. A new functional role for mechanistic/mammalian target of rapamycin complex 1 (mTORC1 in the circadian regulation of L-type voltage-gated calcium channels in avian cone photoreceptors.

    Directory of Open Access Journals (Sweden)

    Cathy Chia-Yu Huang

    Full Text Available In the retina, the L-type voltage-gated calcium channels (L-VGCCs are responsible for neurotransmitter release from photoreceptors and are under circadian regulation. Both the current densities and protein expression of L-VGCCs are significantly higher at night than during the day. However, the underlying mechanisms of circadian regulation of L-VGCCs in the retina are not completely understood. In this study, we demonstrated that the mechanistic/mammalian target of rapamycin complex (mTORC signaling pathway participated in the circadian phase-dependent modulation of L-VGCCs. The activities of the mTOR cascade, from mTORC1 to its downstream targets, displayed circadian oscillations throughout the course of a day. Disruption of mTORC1 signaling dampened the L-VGCC current densities, as well as the protein expression of L-VGCCs at night. The decrease of L-VGCCs at night by mTORC1 inhibition was in part due to a reduction of L-VGCCα1 subunit translocation from the cytosol to the plasma membrane. Finally, we showed that mTORC1 was downstream of the phosphatidylionositol 3 kinase-protein kinase B (PI3K-AKT signaling pathway. Taken together, mTORC1 signaling played a role in the circadian regulation of L-VGCCs, in part through regulation of ion channel trafficking and translocation, which brings to light a new functional role for mTORC1: the modulation of ion channel activities.

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

    DEFF Research Database (Denmark)

    Andreasen, Ditte; Friis, Ulla G; Uhrenholt, Torben R

    2006-01-01

    Voltage-dependent Ca2+ channels Cav1.2 (L type) and Cav2.1 (P/Q type) are expressed in vascular smooth muscle cells (VSMCs) and are important for the contraction of renal resistance vessels. In the present study we examined whether native renal VSMCs coexpress L-, P-, and Q-type Ca2+ currents...... microscopy revealed expression of both channels in all of the smooth muscle cells. Whole-cell patch clamp on single preglomerular VSMCs from mice showed L-, P-, and Q-type currents. Blockade of the L-type currents by calciseptine (20 nmol/L) inhibited 35.6+/-3.9% of the voltage-dependent Ca2+ current......-type and P-type channels inhibited 58.0+/-11.8%, and simultaneous inhibition of L-, P-, and Q-type channels led to blockade (88.7+/-5.6%) of the Ca2+ current. We conclude that aortic and renal preglomerular smooth muscle cells express L-, P-, and Q-type voltage-dependent Ca2+ channels in the rat and mouse....

  5. Expression and function of K(V)2-containing channels in human urinary bladder smooth muscle.

    Science.gov (United States)

    Hristov, Kiril L; Chen, Muyan; Afeli, Serge A Y; Cheng, Qiuping; Rovner, Eric S; Petkov, Georgi V

    2012-06-01

    The functional role of the voltage-gated K(+) (K(V)) channels in human detrusor smooth muscle (DSM) is largely unexplored. Here, we provide molecular, electrophysiological, and functional evidence for the expression of K(V)2.1, K(V)2.2, and the electrically silent K(V)9.3 subunits in human DSM. Stromatoxin-1 (ScTx1), a selective inhibitor of K(V)2.1, K(V)2.2, and K(V)4.2 homotetrameric channels and of K(V)2.1/9.3 heterotetrameric channels, was used to examine the role of these channels in human DSM function. Human DSM tissues were obtained during open bladder surgeries from patients without a history of overactive bladder. Freshly isolated human DSM cells were studied using RT-PCR, immunocytochemistry, live-cell Ca(2+) imaging, and the perforated whole cell patch-clamp technique. Isometric DSM tension recordings of human DSM isolated strips were conducted using tissue baths. RT-PCR experiments showed mRNA expression of K(V)2.1, K(V)2.2, and K(V)9.3 (but not K(V)4.2) channel subunits in human isolated DSM cells. K(V)2.1 and K(V)2.2 protein expression was confirmed by Western blot analysis and immunocytochemistry. Perforated whole cell patch-clamp experiments revealed that ScTx1 (100 nM) inhibited the amplitude of the voltage step-induced K(V) current in freshly isolated human DSM cells. ScTx1 (100 nM) significantly increased the intracellular Ca(2+) level in DSM cells. In human DSM isolated strips, ScTx1 (100 nM) increased the spontaneous phasic contraction amplitude and muscle force, and enhanced the amplitude of the electrical field stimulation-induced contractions within the range of 3.5-30 Hz stimulation frequencies. These findings reveal that ScTx1-sensitive K(V)2-containing channels are key regulators of human DSM excitability and contractility and may represent new targets for pharmacological or genetic intervention for bladder dysfunction.

  6. Role of protein kinase A and class II phosphatidylinositol 3-kinase C2β in the downregulation of KCa3.1 channel synthesis and membrane surface expression by lyso-globotriaosylceramide

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Ju Yeon; Park, Seonghee, E-mail: sp@ewha.ac.kr

    2016-02-19

    The intermediate conductance calcium-activated potassium channel (KCa3.1) mediates proliferation of many cell types including fibroblasts, and is a molecular target for intervention in various cell proliferative diseases. Our previous study showed that reduction of KCa3.1 channel expression by lyso-globotriaosylceramide (lyso-Gb3) inhibits differentiation into myofibroblasts and collagen synthesis, which might lead to development of ascending thoracic aortic aneurysm secondary to Fabry disease. However, how lyso-Gb3 downregulates KCa3.1 channel expression is unknown. Therefore, we aimed to investigate the underlying mechanisms of lyso-Gb3-mediated KCa3.1 channel downregulation, focusing on the cAMP signaling pathway. We found that lyso-Gb3 increased the intracellular cAMP concentration by upregulation of adenylyl cyclase 6 and inhibited ERK 1/2 phosphorylation through the protein kinase A (PKA) pathway, leading to the inhibition of KCa3.1 channel synthesis, not the exchange protein directly activated by cAMP (Epac) pathway. Moreover, lyso-Gb3 suppressed expression of class II phosphatidylinositol 3-kinase C2β (PI3KC2β) by PKA activation, which reduces the production of phosphatidylinositol 3-phosphate [PI(3)P], and the reduced membrane surface expression of KCa3.1 channel was recovered by increasing the intracellular levels of PI(3)P. Consequently, our findings that lyso-Gb3 inhibited both KCa3.1 channel synthesis and surface expression by increasing intracellular cAMP, and controlled surface expression through changes in PI3KC2β-mediated PI(3)P production, suggest that modulation of PKA and PI3KC2β activity to control of KCa3.1 channel expression can be an alternative important target to attenuate ascending thoracic aortic aneurysms in Fabry disease. - Highlights: • Lyso-Gb3 causes elevation of intracellular cAMP. • Lyso-Gb3 inhibits the ERK 1/2 phosphorylation through PKA, thereby reducing KCa3.1 channel synthesis. • Lyso-Gb3 reduces PI3KC2

  7. Upregulation of SK3 and IK1 channels contributes to the enhanced endothelial calcium signaling and the preserved coronary relaxation in obese Zucker rats.

    Directory of Open Access Journals (Sweden)

    Belén Climent

    Full Text Available Endothelial small- and intermediate-conductance KCa channels, SK3 and IK1, are key mediators in the endothelium-derived hyperpolarization and relaxation of vascular smooth muscle and also in the modulation of endothelial Ca2+ signaling and nitric oxide (NO release. Obesity is associated with endothelial dysfunction and impaired relaxation, although how obesity influences endothelial SK3/IK1 function is unclear. Therefore we assessed whether the role of these channels in the coronary circulation is altered in obese animals.In coronary arteries mounted in microvascular myographs, selective blockade of SK3/IK1 channels unmasked an increased contribution of these channels to the ACh- and to the exogenous NO- induced relaxations in arteries of Obese Zucker Rats (OZR compared to Lean Zucker Rats (LZR. Relaxant responses induced by the SK3/IK1 channel activator NS309 were enhanced in OZR and NO- endothelium-dependent in LZR, whereas an additional endothelium-independent relaxant component was found in OZR. Fura2-AM fluorescence revealed a larger ACh-induced intracellular Ca2+ mobilization in the endothelium of coronary arteries from OZR, which was inhibited by blockade of SK3/IK1 channels in both LZR and OZR. Western blot analysis showed an increased expression of SK3/IK1 channels in coronary arteries of OZR and immunohistochemistry suggested that it takes place predominantly in the endothelial layer.Obesity may induce activation of adaptive vascular mechanisms to preserve the dilator function in coronary arteries. Increased function and expression of SK3/IK1 channels by influencing endothelial Ca2+ dynamics might contribute to the unaltered endothelium-dependent coronary relaxation in the early stages of obesity.

  8. Beyond Gap Junction Channel Function: the Expression of Cx43 Contributes to Aldosterone-Induced Mesangial Cell Proliferation via the ERK1/2 and PKC Pathways

    Directory of Open Access Journals (Sweden)

    Aiqing Zhang

    2015-06-01

    Full Text Available Aims: This study aimed to explore the precise mechanism and signaling pathways of mesangial cell (MC proliferation from a new point of view considering Connexin 43 (Cx43. Methods: MC proliferation was measured by the incorporation of 3H-thymidine (3H-TdR. Cx43 was over-expressed in MC cells using lipofectamine 2000, and the expression level was tested with reverse transcription-polymerase chain reaction (RT-PCR and Western blot analyses. The gap junction channel function was explored by Lucifer Yellow scrape loading and dye transfer (SLDT, and the intracellular calcium concentrations ([Ca2+]i were characterized by confocal microscopy on cells loaded with Fura-3/AM. Results: There was an inverse correlation between Cx43 expression and MC proliferation (P0.05. Our data also showed that the mineralcorticoid receptor (MR antagonist spironolactone, ERK1/2 inhibitor PD98059 and PKC inhibitor GF109203X could attenuate the down-regulation of Cx43 expression in Aldo-induced MC proliferation; however, the PI3K inhibitor LY294002 could block MC proliferation without affecting Cx43 expression at either the mRNA or protein level. In addition, Aldo promoted MC proliferation in parallel with increasing [Ca2+]i (PConclusions: Our study provides preliminary evidence that Cx43 is an important regulator of Aldo-promoted MC proliferation. Furthermore, reduced Cx43 expression promoted MC proliferation independent of the gap junction channel function, and this process might be mediated through the ERK1/2- and PKC-dependent pathways.

  9. T-type Ca(2+) channels and Autoregulation of Local Blood Flow

    DEFF Research Database (Denmark)

    Jensen, Lars Jørn; Nielsen, Morten Schak; Salomonsson, Max

    2017-01-01

    L-type voltage gated Ca(2+) channels are considered to be the primary source of calcium influx during the myogenic response. However, many vascular beds also express T-type voltage gated Ca(2+) channels. Recent studies suggest that these channels may also play a role in autoregulation. At low pre...

  10. The effects of calcium on the expression of genes involved in ...

    African Journals Online (AJOL)

    USER

    2010-06-14

    Jun 14, 2010 ... Following incubation, RNA was extracted from a 2 mm ... RNA from samples treated with ethylene was used to synthesize first-strand. cDNA using SuperScript II RNase H-Reverse Transcriptase. (Invitrogen .... Effects of calcium and VP on endogenous ethylene production in AZ of tomato pedicel explants.

  11. TREK-1 Channel Expression in Smooth Muscle as a Target for Regulating Murine Intestinal Contractility: Therapeutic Implications for Motility Disorders

    Directory of Open Access Journals (Sweden)

    Ruolin Ma

    2018-03-01