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Calcium Release (calcium + release)
Kinds of Calcium Release Selected AbstractsCalcium signaling in invertebrate glial cellsGLIA, Issue 7 2006Christian Lohr Abstract Calcium signaling studies in invertebrate glial cells have been performed mainly in the nervous systems of the medicinal leech (Hirudo medicinalis) and the sphinx moth Manduca sexta. The main advantages of studing glial cells in invertebrate nervous systems are the large size of invertebrate glial cells and their easy accessibility for optical and electrophysiological recordings. Glial cells in both insects and annelids express voltage-gated calcium channels and, in the case of leech glial cells, calcium-permeable neurotransmitter receptors, which allow calcium influx as one major source for cytosolic calcium transients. Calcium release from intracellular stores can be induced by metabotropic receptor activation in leech glial cells, but appears to play a minor role in calcium signaling. In glial cells of the antennal lobe of Manduca, voltage-gated calcium signaling changes during postembryonic development and is essential for the migration of the glial cells, a key step in axon guidance and in stabilization of the glomerular structures that are characteristic of primary olfactory centers. © 2006 Wiley-Liss, Inc. [source] Co-occurring increases of calcium and organellar reactive oxygen species determine differential activation of antioxidant and defense enzymes in Ulva compressa (Chlorophyta) exposed to copper excessPLANT CELL & ENVIRONMENT, Issue 10 2010ALBERTO GONZALEZ ABSTRACT In order to analyse copper-induced calcium release and (reactive oxygen species) ROS accumulation and their role in antioxidant and defense enzymes activation, the marine alga Ulva compressa was exposed to 10 µM copper for 7 d. The level of calcium, extracellular hydrogen peroxide (eHP), intracellular hydrogen peroxide (iHP) and superoxide anions (SA) as well as the activities of ascorbate peroxidase (AP), glutathione reductase (GR), glutathione-S-transferase (GST), phenylalanine ammonia lyase (PAL) and lipoxygenase (LOX) were determined. Calcium release showed a triphasic pattern with peaks at 2, 3 and 12 h. The second peak was coincident with increases in eHP and iHP and the third peak with the second increase of iHP. A delayed wave of SA occurred after day 3 and was not accompanied by calcium release. The accumulation of iHP and SA was mainly inhibited by organellar electron transport chains inhibitors (OETCI), whereas calcium release was inhibited by ryanodine. AP activation ceased almost completely after the use of OETCI. On the other hand, GR and GST activities were partially inhibited, whereas defense enzymes were not inhibited. In contrast, PAL and LOX were inhibited by ryanodine, whereas AP was not inhibited. Thus, copper stress induces calcium release and organellar ROS accumulation that determine the differential activation of antioxidant and defense enzymes. [source] Functional role of human NK cell receptor 2B4 (CD244) isoformsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 6 2009Stephen O. Mathew Abstract 2B4 (CD244), a member of the signaling lymphocyte-activation molecule (SLAM/CD150), is expressed on all NK cells, a subpopulation of T cells, monocytes and basophils. Human NK cells express two isoforms of 2B4, h2B4-A and h2B4-B that differ in a small portion of the extracellular domain. In the present investigation, we have studied the functions of h2B4-A and h2B4-B. Our study demonstrated that these two isoforms differ in their binding affinity for CD48, which results in differential cytotoxic activity as well as intracellular calcium release by NK cells upon target cell recognition. Analysis of the predicted 3-D structure of the two isoforms showed conformational differences that could account for their differences in binding affinity to CD48. h2B4-A was able to mediate natural cytotoxicity against CD48-expressing K562 target cells and induce intracellular calcium release, whereas h2B4-B showed no effects. NK-92MI, U937, THP-1, KU812, primary monocytes, basophils and NK cells showed expression of both h2B4-A and h2B4-B whereas YT and IL-2-activated NK cells did not show any h2B4-B expression. Stimulation of NK cells through 2B4 resulted in decreased mRNA levels of both h2B4-A and h2B4-B indicating that down-regulation of 2B4 isoforms may be an important factor in controlling NK cell activation during immune responses. [source] Lack of PSD-95 drives hippocampal neuronal cell death through activation of an ,CaMKII transduction pathwayEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 5 2002Fabrizio Gardoni Abstract The PSD-95 protein family organizes the glutamatergic postsynaptic density and it is involved in the regulation of the excitatory signal at central nervous system synapses. We show here that PSD-95 deficiency by means of antisense oligonucleotides induces significant neuronal cell death within 24 h both in primary hippocampal cultures and in organotypic hippocampal slices. On the other hand, cultured cortical neurons are spared by PSD-95 antisense toxicity until they reach a NR2A detectable protein level (24 days in vitro). The neurotoxic event is characterized by increased ,CaMKII association to NR2 regulatory subunits of NMDA receptor complex. As a direct consequence of ,CaMKII association, we found increased GluR1 delivery to cell surface in cultured hippocampal neurons paralleled by AMPA-dependent increase in [Na+]I levels. In addition, both CaMKII specific inhibitor KN-93 and AMPA receptor antagonists CNQX and NBQX rescued neuronal survival to control values. On the other hand, both the NMDA channel blocker MK-801 and Dantrolene, an inhibitor of calcium release from ryanodine-sensitive endoplasmic reticulum stores, failed to have any effect on neuronal survival in PSD-95 deficient neurons. Thus, our data provide clues that PSD-95 reduced expression in neurons is responsible for neuronal vulnerability mediated by direct activation of ,CaMKII transduction pathway in the postsynaptic compartment. [source] Antioxidant and anti-inflammatory activities of melanocortin peptidesEXPERIMENTAL DERMATOLOGY, Issue 9 2004J. W. Haycock ,-Melanocyte-stimulating hormone (,-MSH) has previously been identified as a potent anti-inflammatory agent in various tissues including the skin. It operates by binding to the melanocortin-1 receptor (MC-1R) which results in the elevation of cyclic AMP. ,-MSH opposes the action of several proinflammatory cytokines including tumour necrosis factor-, (TNF-,). We have shown that ,-MSH can inhibit TNF-,-stimulated activation of nuclear factor-,B (NF-,B) in human cultured melanocytes, melanoma cells, keratinocytes, fibroblasts, Schwann cells and olfactory ensheathing cells. It also inhibits TNF-,-stimulated upregulation of intercellular adhesion molecule-1 (ICAM-1) in many of these cells and can inhibit peroxide-stimulated activation of glutathione peroxidase, suggesting an antioxidant role. ,-MSH is also able to stimulate intracellular calcium release in keratinocytes and fibroblasts (which do not readily show detectible cyclic AMP elevation) but only in the presence of PIA (an adenosine agonist). The carboxyl terminal tripeptides KPV/KP-D-V are reported to be the minimal sequences necessary to convey anti-inflammatory potential, but evidence on how they act is not fully known. Stable transfection of Chinese hamster ovary cells with MC-1R suggests that the KPV peptides operate by this receptor, at least by elevating intracellular calcium. Elevation of cyclic AMP by these tripeptides has not been detected in any cell type studied; however, calcium elevation can inhibit TNF-,-stimulated NF-,B activity (as for cyclic AMP). In conclusion, the MSH peptides convey anti-inflammatory and antioxidant activity in many cell types in skin and nerve, by counteracting proinflammatory cytokine signalling. The KPV peptides appear to act functionally via the MC-1R and can also elevate intracellular calcium. [source] Actions of Arachidonic Acid on Contractions and Associated Electrical Activity in Guinea-Pig Isolated Ventricular MyocytesEXPERIMENTAL PHYSIOLOGY, Issue 4 2001M. A. Mamas The actions of arachidonic acid (AA) were investigated in guinea-pig isolated ventricular myocytes. Exposure of myocytes to 10 ,M AA reduced the amplitude of contractions and calcium transients accompanying action potentials at a frequency of 1 Hz. AA (10 ,M) also reduced the amplitude of calcium currents recorded under voltage-clamp conditions. The suppression of contraction by AA was not prevented by either 10 ,M trihydroindomethicin (to inhibit cyclo-oxygenase) or 10 ,M ETYA (5,8,11,14-eicosatetraynoic acid, to inhibit AA metabolising enzymes), showing that the actions of AA appeared not to be mediated by these metabolites. The reduction of contraction by 10 ,M AA was also not prevented by the protein kinase C inhibitor, Ro31-8220 (1 ,M), showing that this pathway appeared not to be required for the observed effect. Direct effects of AA may be involved. A further action of 10 ,M AA was to suppress spontaneous electrical activity induced by either the ,-adrenergic agonist isoprenaline or the Na+ pump inhibitor, ouabain. This effect of AA on spontaneous activity might be associated with the observed reduction of calcium entry through L-type calcium channels, although additional effects of AA on calcium release from the sarcoplasmic reticulum might also be involved. [source] Na,K-ATPase ,2 inhibition alters calcium responses in optic nerve astrocytesGLIA, Issue 3 2004April K. Hartford Abstract Experiments were conducted to test the effect of 1 ,M ouabain, an Na,K-ATPase inhibitor, on capacitative calcium entry (CCE) and calcium responses elicited by ATP in rat optic nerve astrocytes. In the rat, 1 ,M ouabain is sufficient to inhibit the ,2 Na,K-ATPase, but not the ,1. Immortalized astrocytes derived from Na,K-ATPase ,2 homozygous knockout (KO) mice and wild-type (WT) littermates were also used. Cytosolic calcium and sodium concentrations were measured using Fura-2 and SBFI, respectively. The magnitude of the increase in cytosolic calcium concentration during CCE was significantly greater in rat astrocytes exposed to 1 ,M ouabain. To measure calcium release from stores, cells were exposed to ATP in the absence of extracellular calcium. In astrocytes exposed to 1 ,M ouabain, a significantly greater calcium response to ATP was observed. 1 ,M ouabain was shown to inhibit ATP hydrolysis in membrane material containing Na,K-ATPase ,2 and ,1 isoforms (rat muscle) but not in membranes containing only Na,K-ATPase ,1 (rat kidney). In intact astrocytes, 1 ,M ouabain did not alter the cell-wide cytosolic sodium concentration. In mouse Na,K-ATPase ,2 KO astrocytes, the calcium increase during CCE was significantly higher than in WT cells, as was the magnitude of the calcium response to ATP. In KO astrocytes, but not WT, the cytosolic calcium increase during CCE was insensitive to 1 ,M ouabain. Taken together, the results suggest that selective inhibition of the Na,K-ATPase ,2 isoform has the potential to change calcium signaling and CCE. © 2003 Wiley-Liss, Inc. [source] Cyclooxygenase-2 Expression and Prostaglandin E2 Production in Response to Acidic pH Through OGR1 in a Human Osteoblastic Cell Line,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2008Hideaki Tomura Abstract Acidosis has been shown to induce depletion of bone calcium from the body. This calcium release process is thought to be partially cell mediated. In an organ culture of bone, acidic pH has been shown to induce cyclooxygenase-2 (COX-2) induction and prostaglandin E2 (PGE2) production, resulting in stimulation of bone calcium release. However, the molecular mechanisms whereby osteoblasts sense acidic circumstances and thereby induce COX-2 induction and PGE2 production remain unknown. In this study, we used a human osteoblastic cell line (NHOst) to characterize cellular activities, including inositol phosphate production, intracellular Ca2+ concentration ([Ca2+]i), PGE2 production, and COX-2 mRNA and protein expression, in response to extracellular acidification. Small interfering RNA (siRNA) specific to the OGR1 receptor and specific inhibitors for intracellular signaling pathways were used to characterize acidification-induced cellular activities. We found that extracellular acidic pH induced a transient increase in [Ca2+]i and inositol phosphate production in the cells. Acidification also induced COX-2 induction, resulting in PGE2 production. These proton-induced actions were markedly inhibited by siRNA targeted for the OGR1 receptor and the inhibitors for Gq/11 protein, phospholipase C, and protein kinase C. We conclude that the OGR1/Gq/11/phospholipase C/protein kinase C pathway regulates osteoblastic COX-2 induction and subsequent PGE2 production in response to acidic circumstances. [source] Modulation of cardiac ionic homeostasis by 3-iodothyronamineJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 9b 2009Sandra Ghelardoni Abstract 3-iodothyronamine (T1AM) is a novel endogenous relative of thyroid hormone, able to interact with trace amine-associated receptors, a class of plasma membrane G protein-coupled receptors, and to produce a negative inotropic and chronotropic effect. In the isolated rat heart 20,25 ,M T1AM decreased cardiac contractility, but oxygen consumption and glucose uptake were either unchanged or disproportionately high when compared to mechanical work. In adult rat cardiomyocytes acute exposure to 20 ,M T1AM decreased the amplitude and duration of the calcium transient. In patch clamped cardiomyocytes sarcolemmal calcium current density was unchanged while current facilitation by membrane depolarization was abolished consistent with reduced sarcoplasmic reticulum (SR) calcium release. In addition, T1AM decreased transient outward current (Ito) and IK1 background current. SR studies involving 20 ,M T1AM revealed a significant decrease in ryanodine binding due to reduced Bmax, no significant change in the rate constant of calcium-induced calcium release, a significant increase in calcium leak measured under conditions promoting channel closure, and no effect on oxalate-supported calcium uptake. Based on these observations we conclude T1AM affects calcium and potassium homeostasis and suggest its negative inotropic action is due to a diminished pool of SR calcium as a result of increased diastolic leak through the ryanodine receptor, while increased action potential duration is accounted for by inhibition of Ito and IK1 currents. [source] Substance P release evoked by capsaicin or potassium from rat cultured dorsal root ganglion neurons is conversely modulated with bradykininJOURNAL OF NEUROCHEMISTRY, Issue 5 2006He-Bin Tang Abstract To clarify the molecular mechanism of substance P (SP) release from dorsal root ganglion (DRG) neurons, we investigated the involvement of several intracellular effectors in the regulation of SP release evoked by capsaicin, potassium or/and bradykinin. Bradykinin-evoked SP release from cultured adult rat DRG neurons was attenuated by either the mitogen-activated protein kinase kinase (MEK) inhibitor (U0126) or cycloheximide. As the long-term exposure of DRG neurons to bradykinin (3 h) resulted in extracellular signal-regulated kinase (ERK) phosphorylation at an early stage and thereafter induced cyclooxygenase-2 (COX-2) protein expression, which both contribute to the SP release triggered by bradykinin B2 receptor. The long-term exposure of DRG neurons to bradykinin enhanced the SP release by capsaicin, but attenuated that by potassium. Interestingly, the inositol 1,4,5-triphosphate (IP3)-induced calcium release blocker [2-aminoethyl diphenylborinate (2-APB)] not only inhibited the potassium-evoked SP release, but also completely abolished the enhancement of capsaicin-induced SP release by bradykinin from cultured DRG neurons. Together, these findings suggest that the molecular mechanisms of SP release by bradykinin involve the activation of MEK, and also require the de novo protein synthesis of COX-2 in DRG neurons. The IP3 -dependent calcium release could be involved in the processes of the regulation by bradykinin of capsaicin-triggered SP release. [source] Bone resorption activity of osteolytic metastatic lung and breast cancersJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 6 2004Lih-Yuann Shih Abstract Production of bone resorption mediators and bone resorption activity were compared among osteolytic metastatic cancers, normal bone tissues, and soft tissue metastatic cancers to search for the possible factors leading to cancer-induced bone resorption. Twenty-five patients with untreated osteolytic metastatic breast or non-small cell lung cancers consisted of the study group. Normal bone tissues obtained from the same patient were used as internal controls; and tumor tissues from patients with soft tissue metastasis were used as external controls. Serum and urinary bone turnover markers were measured. Tissues harvested during surgery were subjected to tissue culture. The levels of prostaglandin E2 (PGE2), tumor necrosis factor-, (TNF-,), and interleukin-6 (IL-6) in the supernatant after 72 h of culture were measured. Bone resorption activity was measured by calcium release from cultured calvarias, and bone volume as well as osteoclast number in bone sections. Patients with osteolytic metastatic cancers showed significantly decreased serum osteocalcin, increased serum alkaline phosphatase, and urinary deoxypyridinoline levels. Osteolytic metastatic cancers produced significantly more PGE2 than both controls. Conditioned medium from osteolytic metastatic tumors showed significantly enhanced bone resorption activity, and indomethacin significantly reduced this activity. Levels of PGE2, and bone resorption activity increased in osteolytic tumor tissues than soft tissue metastatic tissues in the same patient indicated that the same tumor cells might respond differently to different microenvironments. Our observation showed that PGE2 was produced by osteolytic metastatic cancers and stimulated bone resorption in mice calvarias. PGE2 inhibitor may be applicable in reducing bone resorption by osteolytic metastatic cancers. © 2004 Orthopaedic Research Society. Published by Elsevier Ltd. All rights reserved. [source] Identification of genes related to mechanical stress in human periodontal ligament cells using microarray analysisJOURNAL OF PERIODONTAL RESEARCH, Issue 1 2007R. M. S. De Araujo Background and Objective:, Differential expression of genes in human periodontal ligament (PDL) under mechanical stress, such as orthodontic force, is thought to be involved in the remodeling of PDL cells and periodontal tissues. However, little is known about the genes expressed in PDL cells under mechanical stress. Material and Methods:, We employed microarray analysis to assess, in a comprehensive manner, the gene expression profiles in PDL cells compressed by a static force using an in vitro three-dimensional culture system. Six genes were selected and validated by quantitative real-time polymerase chain reaction analysis, consistent with the microarray data. Results:, The microarray data revealed that 108 of 30,000 genes tested were differentially expressed by mechanical force loading. Among them, 85 genes were up-regulated by mechanical stress, while 23 genes were down-regulated, judging by the thresholds of a two-fold increase/decrease compared with the controls. Thirty-two of the up-regulated and eight of the down-regulated genes, well-characterized in protein function, were involved in numerous biological processes including cell communication, cell signaling, cell cycle, stress response, and calcium release. However, several genes differentially expressed in our microarray data have not been well defined as stress-response molecules. Conclusion:, Our microarray is the first to show the gene profile in PDL cells caused by mechanical stress; however, further studies to clarify the physiological function of these molecules in PDL cells are required. [source] Acamprosate: Recent Findings and Future Research DirectionsALCOHOLISM, Issue 7 2008Karl Mann This article explores the mechanisms of action and the potential responder profile of acamprosate, a compound efficacious in relapse prevention of alcoholism. New evidence at the molecular and cellular level suggests that acamprosate attenuates hyper-glutamatergic states that occur during early abstinence and involves iono (NMDA)- and metabotrotropic (mGluR5) glutamate receptors along with augmented intracellular calcium release and electrophysiological changes. Thus mutant mice with enhanced glutamate levels exhibit higher alcohol consumption than wild type mice and respond better to acamprosate, demonstrating that acamprosate acts mainly on a hyper-glutamatergic system. This mode of action further suggests that acamprosate exhibits neuroprotective properties. In rats, cue-induced reinstatement behavior is significantly reduced by acamprosate treatment whereas cue-induced craving responses in alcohol-dependent patients seem not to be affected by this treatment. An ongoing study ("Project Predict") defines specific responder profiles for an individualized use of acamprosate and naltrexone. Neurophysiological as well as psychometric data are used to define 2 groups of patients: "reward cravers" and "relief cravers". While naltrexone should work better in the first group, acamprosate is hypothesized to be efficacious in the latter where withdrawal associated and/or cue induced hyper-glutamatergic states are thought to trigger relapse. Further research should target the definition of subgroups applying endophenotypic approaches, e.g. by detecting a hyperglutamatergic syndrome using MR spectroscopy. [source] Impaired inhibitory G-protein function contributes to increased calcium currents in rats with diabetic neuropathyJOURNAL OF THE PERIPHERAL NERVOUS SYSTEM, Issue 2 2002KE Hall There is a growing body of evidence that sensory neuropathy in diabetes is associated with abnormal calcium signaling in dorsal root ganglion (DRG) neurons. Enhanced influx of calcium via multiple high-threshold calcium currents is present in sensory neurons of several models of diabetes mellitus, including the spontaneously diabetic BioBred/Worchester (BB/W) rat and the chemical streptozotocin (STZ)-induced rat. We believe that abnormal calcium signaling in diabetes has pathologic significance as elevation of calcium influx and cytosolic calcium release has been implicated in other neurodegenerative conditions characterized by neuronal dysfunction and death. Using electrophysiologic and pharmacologic techniques, the present study provides evidence that significant impairment of G-protein-coupled modulation of calcium channel function may underlie the enhanced calcium entry in diabetes. N- and P-type voltage-activated, high-threshold calcium channels in DRGs are coupled to mu opiate receptors via inhibitory G(o)-type G proteins. The responsiveness of this receptor coupled model was tested in dorsal root ganglion (DRG) neurons from spontaneously-diabetic BB/W rats, and streptozotocin-induced (STZ) diabetic rats. Intracellular dialysis with GTPgammaS decreased calcium current amplitude in diabetic BB/W DRG neurons compared with those of age-matched, nondiabetic controls, suggesting that inhibitory G-protein activity was diminished in diabetes, resulting in larger calcium currents. Facilitation of calcium current density (I(DCa)) by large-amplitude depolarizing prepulses (proposed to transiently inactivate G proteins), was significantly less effective in neurons from BB/W and STZ-induced diabetic DRGs. Facilitation was enhanced by intracellular dialysis with GTPgammaS, decreased by pertussis toxin, and abolished by GDPbetaS within 5 min. Direct measurement of GTPase activity using opiate-mediated GTPgamma[(35)S] binding, confirmed that G-protein activity was significantly diminished in STZ-induced diabetic neurons compared with age-matched nondiabetic controls. Diabetes did not alter the level of expression of mu opiate receptors and G-protein alpha subunits. These studies indicate that impaired regulation of calcium channels by G proteins is an important mechanism contributing to enhanced calcium influx in diabetes. [source] Lipid rafts are required in G,i signaling downstream of the P2Y12 receptor during ADP-mediated platelet activationJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 5 2005T. M. QUINTON Summary., ADP is important in propagating hemostasis upon its secretion from activated platelets in response to other agonists. Lipid rafts are microdomains within the plasma membrane that are rich in cholesterol and sphingolipids, and have been implicated in the stimulatory mechanisms of platelet agonists. We sought to determine the importance of lipid rafts in ADP-mediated platelet activation via the G protein-coupled P2Y1 and P2Y12 receptors using lipid raft disruption by cholesterol depletion with methyl- , -cyclodextrin. Stimulation of cholesterol-depleted platelets with ADP resulted in a reduction in the extent of aggregation but no difference in the extent of shape change or intracellular calcium release. Furthermore, repletion of cholesterol to previously depleted membranes restored ADP-mediated platelet aggregation. In addition, P2Y12-mediated inhibition of cAMP formation was significantly decreased upon cholesterol depletion from platelets. Stimulation of cholesterol-depleted platelets with agonists that depend upon G,i activation for full activation displayed significant loss of aggregation and secretion, but showed restoration when simultaneously stimulated with the G,z -coupled agonist epinephrine. Finally, G,i preferentially localizes to lipid rafts as determined by sucrose density centrifugation. We conclude that G,i signaling downstream of P2Y12 activation, but not G,q or G,z signaling downstream of P2Y1 or ,2A activation, respectively, has a requirement for lipid rafts that is necessary for its function in ADP-mediated platelet activation. [source] Medicinal chemistry and therapeutic potential of muscarinic M3 antagonistsMEDICINAL RESEARCH REVIEWS, Issue 6 2009Ilaria Peretto Abstract Muscarinic acetylcholine receptors belong to the G-protein-coupled receptors family. Currently five different receptor subtypes have been identified and cloned. M3 receptor subtypes are coupled to Gq family proteins and increase phosphatidyl inositol hydrolysis and calcium release from internal stores. They are widely distributed both in the central nervous system and in the periphery. At the central level, M3 receptor subtypes are involved in modulation of neurotransmitter release, temperature homeostasis, and food intake, while in the periphery they induce smooth muscle contraction, gland secretion, indirect relaxation of vascular smooth muscle, and miosis. The main therapeutic applications of M3 antagonists include overactive bladder (OAB), chronic obstructive pulmonary disease (COPD), and pain-predominant irritable bowel syndrome (IBS). The introduction of selective M3 antagonists has not improved clinical efficacy compared with the old non-selective antimuscarinics but has reduced the rate of adverse events mediated by the blockade of cardiac M2 receptors (tachycardia) and central M1 receptors (cognitive impairment). Improved tolerability has been obtained also with controlled release or with inhaled formulations. However, there is still a need for safer M3 antagonists for the treatment of COPD and better-tolerated and more effective compounds for the therapy of OAB. New selective muscarinic M3 antagonists currently in early discovery and under development have been designed to address these issues. However, as M3 receptors are widely located in various tissues including salivary glands, gut smooth muscles, iris, and ciliary muscles, further clinical improvements may derive from the discovery and the development of new compounds with tissue rather than muscarinic receptor subtype selectivity. © 2009 Wiley Periodicals, Inc. Med Res Rev, 29, No. 6, 867,902, 2009 [source] Anti-thrombotic effect of milrinone is caused by inhibition of calcium release from the dense tubular system in human platelets,ACTA ANAESTHESIOLOGICA SCANDINAVICA, Issue 1 2003N. Hiramatsu Aim: Milrinone, a phosphodiesterase III inhibitor, exerts positive inotropic effects which induce an increase in the intracellular calcium concentration by raising the cyclic adenosine monophosphate level in cardiac muscle. Milrinone was also reported to inhibit platelet aggregation, however, its mechanism remains unknown. Therefore, we investigated the effects of milrinone on intracellular calcium mobilization when platelets were activated. Methods: Washed platelets, obtained from six healthy volunteers, were preincubated with milrinone (0.9 µM) for 1 min and then exposed to 0.015 iµ ml,1 thrombin for 5 min. The effect of milrinone on changes in the intracellular calcium level using a fluorescent dye, fura-2, was also observed. Calcium mobilizations via plasma membrane calcium channels and the dense tubular system were assessed differentially. Results: Milrinone (0.9 µM) significantly suppressed the aggregation ratios at 5 min compared with those in controls (86±5%) to 75±8%. The increase in the intracellular calcium concentration was also significantly suppressed (controls, 915±293 nM vs. 405±240 nM) when stimulated by thrombin. Milrinone also significantly inhibited the release of calcium from the dense tubular system (controls, 284±111 nM vs. 158±51 nM). Calcium influx through the plasma membrane was suppressed by milrinone 2.4 µM. Conclusion: Milrinone (0.9 µM) inhibited thrombin-induced platelet aggregation. This inhibitory effect was mainly mediated by suppressing calcium release from the dense tubular system. [source] Sarcoplasmic reticulum: The dynamic calcium governor of muscleMUSCLE AND NERVE, Issue 6 2006Ann E. Rossi MS Abstract The sarcoplasmic reticulum (SR) provides feedback control required to balance the processes of calcium storage, release, and reuptake in skeletal muscle. This balance is achieved through the concerted action of three major classes of SR calcium-regulatory proteins: (1) luminal calcium-binding proteins (calsequestrin, histidine-rich calcium-binding protein, junctate, and sarcalumenin) for calcium storage; (2) SR calcium release channels (type 1 ryanodine receptor or RyR1 and IP3 receptors) for calcium release; and (3) sarco(endo)plasmic reticulum Ca2+ -ATPase (SERCA) pumps for calcium reuptake. Proper calcium storage, release, and reuptake are essential for normal skeletal muscle function. We review SR structure and function during normal skeletal muscle activity, the proteins that orchestrate calcium storage, release, and reuptake, and how phenotypically distinct muscle diseases (e.g., malignant hyperthermia, central core disease, and Brody disease) can result from subtle alterations in the activity of several key components of the SR calcium-regulatory machinery. Muscle Nerve, 2006 [source] Effect of dantrolene in an in vivo and in vitro model of myocardial reperfusion injuryACTA ANAESTHESIOLOGICA SCANDINAVICA, Issue 2 2000B. Preckel Background: In skeletal muscle, dantrolene reduces free cytosolic calcium by inhibiting calcium release from the sarcoplasmic reticulum. A similar effect in ischemic-reperfused heart cells would protect myocardial tissue against reperfusion injury. We tested the hypothesis that dantrolene infusion during reperfusion protects the heart against reperfusion injury. Methods: Isovolumetric beating rat hearts were subjected to 30 min of ischemia followed by 60 min of reperfusion. Left ventricular (LV) developed pressure (LVDP) and creatine kinase release (CKR) were determined as indices of myocardial performance and cellular injury, respectively. In the treatment groups, dantrolene (25 (DAN25) or 100 (DAN100) ,mol l,1) was infused during the first 15 min of reperfusion; control hearts received the respective concentration of the vehicle (mannitol (CON25, CON100), each group n=7). To investigate the effects of dantrolene on reperfusion injury in vivo, 18 chloralose-anesthetized rabbits were subjected to 30 min occlusion and 180 min reperfusion of a major coronary artery. LV pressure (LVP), cardiac output (CO), and infarct size were determined. During the last 5 min of ischemia, nine rabbits received 10 mg kg,1 dantrolene intravenously (DAN). Another nine rabbits received the vehicle (dimethylsulfoxide) and served as controls (CON). Results: In isolated rat hearts, there was no recovery of LVDP in any group. Total CKR during 1 h of reperfusion was 845±76 (CON100) and 550±81 U g,1 dry mass (DAN100, P<0.05). In rabbits in vivo, hemodynamic baseline values were similar between groups (CON vs. DAN: LVP, 99±6 (mean±SEM) vs. 91±6mm Hg, P=0.29; CO, 252±26 vs. 275±23 ml min,1, P=0.53). During coronary artery occlusion, LVP and CO were reduced in both groups (CON: LVP, 89±3%; CO, 90±5% of baseline values) and LVP did not recover to baseline values during reperfusion (51±5% (CON) vs. 67±7% (DAN) of baseline, P=0.10). Infarct size was 41±4% of the area at risk in controls and 37±6% in dantrolene treated hearts (P=0.59). Conclusions: Dantrolene reduced CKR, indicating an attenuation of lethal cellular reperfusion injury in isolated rat hearts. However, in the rabbit in vivo, there was no effect on the extent of reperfusion injury after regional myocardial ischemia. [source] Co-occurring increases of calcium and organellar reactive oxygen species determine differential activation of antioxidant and defense enzymes in Ulva compressa (Chlorophyta) exposed to copper excessPLANT CELL & ENVIRONMENT, Issue 10 2010ALBERTO GONZALEZ ABSTRACT In order to analyse copper-induced calcium release and (reactive oxygen species) ROS accumulation and their role in antioxidant and defense enzymes activation, the marine alga Ulva compressa was exposed to 10 µM copper for 7 d. The level of calcium, extracellular hydrogen peroxide (eHP), intracellular hydrogen peroxide (iHP) and superoxide anions (SA) as well as the activities of ascorbate peroxidase (AP), glutathione reductase (GR), glutathione-S-transferase (GST), phenylalanine ammonia lyase (PAL) and lipoxygenase (LOX) were determined. Calcium release showed a triphasic pattern with peaks at 2, 3 and 12 h. The second peak was coincident with increases in eHP and iHP and the third peak with the second increase of iHP. A delayed wave of SA occurred after day 3 and was not accompanied by calcium release. The accumulation of iHP and SA was mainly inhibited by organellar electron transport chains inhibitors (OETCI), whereas calcium release was inhibited by ryanodine. AP activation ceased almost completely after the use of OETCI. On the other hand, GR and GST activities were partially inhibited, whereas defense enzymes were not inhibited. In contrast, PAL and LOX were inhibited by ryanodine, whereas AP was not inhibited. Thus, copper stress induces calcium release and organellar ROS accumulation that determine the differential activation of antioxidant and defense enzymes. [source] Proteomics reveals lowering oxygen alters cytoskeletal and endoplasmatic stress proteins in human endothelial cellsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 19 2009Louise Østergaard Abstract A proteomic approach was applied to explore the signalling pathways elicited by lowering O2 in endothelial cells. Endothelial cells isolated from native umbilical cords were subjected to 21, 5, or 1% O2 for 24,h. 2-D PAGE was performed and candidate proteins were identified using LC-MS/MS. Lowering of O2 from 21 to 5% induced upregulation of cofilin-1, cyclophilin A, tubulin and tubulin fragments, a fragment of glucose-regulated protein 78 (Grp78) and calmodulin. The upregulation of Grp78 suggested that ER stress proteins were altered and indeed Grp94 and caspase 12 expression were increased in cells exposed to 5% O2. The presence of ER stress is also supported by findings of blunted caffeine-evoked ER calcium release in cells exposed to 5 and 1% O2. Exposure to 1% O2 caused increases in cofilin-1, cyclophilin A, and caspase 12 as well as a decrease of ,-actin, but it did not alter the expression of calmodulin, tubulin, Grp78, and Grp94. Incubation with CoCl2, a stabilizer of the hypoxia-inducible factor, increased the expression of several of the proteins. The present investigations reveal that lowering O2, probably in part through hypoxia-inducible factor, alter the expression of a series of proteins mainly involved in cytoskeletal changes (e.g. cofilin-1, tubulin, and ,-actin) and in ER stress/apoptosis (e.g. Grp78/94, caspase 12, and cyclophilin A). [source] Calsequestrin, triadin and more: the molecules that modulate calcium release in cardiac and skeletal muscleTHE JOURNAL OF PHYSIOLOGY, Issue 13 2009Eduardo Ríos No abstract is available for this article. [source] Evolution and modulation of intracellular calcium release during long-lasting, depleting depolarization in mouse muscleTHE JOURNAL OF PHYSIOLOGY, Issue 19 2008Leandro Royer Intracellular calcium signals regulate multiple cellular functions. They depend on release of Ca2+ from cellular stores into the cytosol, a process that in many types of cells appears to be tightly controlled by changes in [Ca2+] within the store. In contrast with cardiac muscle, where depletion of Ca2+ in the sarcoplasmic reticulum is a crucial determinant of termination of Ca2+ release, in skeletal muscle there is no agreement regarding the sign, or even the existence of an effect of SR Ca2+ level on Ca2+ release. To address this issue we measured Ca2+ transients in mouse flexor digitorum brevis (FDB) skeletal muscle fibres under voltage clamp, using confocal microscopy and the Ca2+ monitor rhod-2. The evolution of Ca2+ release flux was quantified during long-lasting depolarizations that reduced severely the Ca2+ content of the SR. As in all previous determinations in mammals and non-mammals, release flux consisted of an early peak, relaxing to a lower level from which it continued to decay more slowly. Decay of flux in this second stage, which has been attributed largely to depletion of SR Ca2+, was studied in detail. A simple depletion mechanism without change in release permeability predicts an exponential decay with time. In contrast, flux decreased non-exponentially, to a finite, measurable level that could be maintained for the longest pulses applied (1.8 s). An algorithm on the flux record allowed us to define a quantitative index, the normalized flux rate of change (NFRC), which was shown to be proportional to the ratio of release permeability P and inversely proportional to Ca2+ buffering power B of the SR, thus quantifying the ,evacuability' or ability of the SR to empty its content. When P and B were constant, flux then decayed exponentially, and NFRC was equal to the exponential rate constant. Instead, in most cases NFRC increased during the pulse, from a minimum reached immediately after the early peak in flux, to a time between 200 and 250 ms, when the index was no longer defined. NFRC increased by 111% on average (in 27 images from 18 cells), reaching 300% in some cases. The increase may reflect an increase in P, a decrease in B, or both. On experimental and theoretical grounds, both changes are to be expected upon SR depletion. A variable evacuability helps maintain a constant Ca2+ output under conditions of diminishing store Ca2+ load. [source] AMPA-sst2 somatostatin receptor interaction in rat hypothalamus requires activation of nmda and/or metabotropic glutamate receptors and depends on intracellular calciumTHE JOURNAL OF PHYSIOLOGY, Issue 1 2003Stéphane Peineau Modulation of glutamatergic transmission by neuropeptides is an essential aspect of neuronal network activity. Activation of the hypothalamic somatostatin sst2 receptor subtype by octreotide decreases AMPA glutamate responses, indicating a central link between a neurohormonal and neuromodulatory peptide and the main hypothalamic fast excitatory neurotransmitter. In mediobasal hypothalamic slices, sst2 activation inhibits the AMPA component of glutamatergic synaptic responses but is ineffective when AMPA currents are pharmacologically isolated. In mediobasal hypothalamic cultures, the decrease of AMPA currents induced by octreotide requires a concomitant activation of sst2 receptors with either NMDA and/or metabotropic glutamate receptors. This modulation depends on changes in intracellular calcium concentration induced by calcium flux through NMDA receptors or calcium release from intracellular stores following metabotropic glutamate receptor activation. These results highlight an unusual regulatory mechanism in which the simultaneous activation of at least three different types of receptor is necessary to allow somatostatin-induced modulation of fast synaptic glutamatergic transmission in the hypothalamus. [source] Regulation of junctional and non-junctional sarcoplasmic reticulum calcium release in excitation-contraction coupling in cat atrial myocytesTHE JOURNAL OF PHYSIOLOGY, Issue 1 2003Katherine A. Sheehan We have characterized the dependence on membrane potential (Vm) and calcium current (ICa) of calcium-induced calcium release (CICR) from the junctional-SR (j-SR, in the subsarcolemmal (SS) space) and non-junctional-SR (nj-SR, in the central (CT) region of the cell) of cat atrial myocytes using whole-cell voltage-clamp together with spatially resolved laser-scanning confocal microscopy. Subsarcolemmal and central [Ca2+]i transient amplitudes and ICa had a bell-shaped dependence on Vm, but [Ca2+]i reached a maximum at more negative Vm (-10 to 0 mV) than ICa (+10 mV). Termination of ICa after a brief depolarization (2.5 to 22.5 ms) immediately interrupted only the SS [Ca2+]i transient, leaving the development of the CT [Ca2+]i transient unaffected. Block of SR function with 20 ,m ryanodine and 2 ,m thapsigargin, revealed that > 90 % of the control [Ca2+]i transient amplitude was attributable to active SR Ca2+ release through ryanodine receptors (RyRs). The gain of SR Ca2+ release was highest in the SS space at negative test potentials and was less pronounced in the CT region. Inhibition of Na+ -Ca2+ exchange resulted in prolonged and higher amplitude [Ca2+]i transients, elevated resting [Ca2+]i, accelerated propagation of CICR, decreased extrusion of Ca2+ and an increase in j-SR Ca2+ load. Increasing the cytosolic Ca2+ buffer capacity by internal perfusion with 1 mm EGTA limited SR Ca2+ release to the SS region, indicating that Ca2+ release from nj-SR is initiated by diffusion of Ca2+ from the cell periphery and propagating CICR. Junctional-SR Ca2+ release occurred at discrete sites whose order of activation and amplitude of release varied from beat to beat. In conclusion, during normal excitation-contraction coupling in cat atrial myocytes, only Ca2+ release from the j-SR is directly activated by Ca2+ entering via ICa. Elevation of SS [Ca2+]i is required to provide the cytosolic Ca2+ gradient needed to initiate regenerative and propagating CICR from nj-SR. [source] RGS4 Controls Renal Blood Flow and Inhibits Cyclosporine-Mediated NephrotoxicityAMERICAN JOURNAL OF TRANSPLANTATION, Issue 2 2010A. Siedlecki Calcineurin inhibitors (CNI) are powerful immunomodulatory agents that produce marked renal dysfunction due in part to endothelin-1-mediated reductions in renal blood flow. Ligand-stimulated Gq protein signaling promotes the contraction of smooth muscle cells via phospholipase C,-mediated stimulation of cytosolic calcium release. RGS4 is a GTPase activating protein that promotes the deactivation of Gq and Gi family members. To investigate the role of G protein-mediated signaling in the pathogenesis of CNI-mediated renal injury, we used mice deficient for RGS4 (rgs4,/,). Compared to congenic wild type control animals, rgs4,/, mice were intolerant of the CNI, cyclosporine (CyA), rapidly developing fatal renal failure. Rgs4,/, mice exhibited markedly reduced renal blood flow after CyA treatment when compared to congenic wild type control mice as measured by magnetic resonance imaging (MRI). Hypoperfusion was reversed by coadministration of CyA with the endothelin antagonist, bosentan. The MAPK/ERK pathway was activated by cyclosporine administration and was inhibited by cotreatment with bosentan. These results show that endothelin-1-mediated Gq protein signaling plays a key role in the pathogenesis of vasoconstrictive renal injury and that RGS4 antagonizes the deleterious effects of excess endothelin receptor activation in the kidney. [source] Melatonin Counteracts Alterations in Oxidative Metabolism and Cell Viability Induced by Intracellular Calcium Overload in Human Leucocytes: Changes with AgeBASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 1 2010Javier Espino In fact, the free radical theory of ageing proposes that deleterious actions of free radicals are responsible for the functional deterioration associated with ageing. Moreover, a close relationship exists between calcium homeostasis and oxidative stress. The current work was aimed at proving that intracellular calcium overload induced by N -formyl-methionyl-leucyl-phenylalanine (FMLP) and/or thapsigargin leads to oxidative stress. We additionally examined the effect of melatonin on the levels of reactive oxygen species (ROS) and cell viability in human leucocytes collected from young (20,30-year-old) and elderly (65,75-year-old) individuals under both basal and oxidative stress-induced conditions. Treatments with 10 nM FMLP and/or 1 ,M thapsigargin induced a transient increase in cytosolic free-calcium concentration ([Ca2 + ]c) in human leucocytes due to calcium release from internal stores, and led in turn to oxidative stress, as assessed by intracellular ROS measurement. Non-treated leucocytes from aged individuals exhibited higher ROS levels and lower rates of cell survival when compared to leucocytes from young individuals. Similar results were obtained in FMLP and/or thapsigargin-treated leucocytes from elderly individuals when compared to those from the young individuals. Melatonin treatment significantly reduced both hydrogen peroxide (H2O2) and superoxide anion levels, likely due to its free-radical scavenging properties, and enhanced leucocyte viability in both age groups. Therefore, melatonin may be a useful tool for the treatment of disease states and processes where an excessive production of oxidative damage occurs. [source] Model study of time-dependent muscle response to pulsed electrical stimulationBIOELECTROMAGNETICS, Issue 5 2010Ravindra P. Joshi Abstract A systems-level model analysis of neuromuscular response to external electrical stimulation is presented. Action potential (AP) generation, dynamics of voltage-based calcium release at the motor endplates controlled by the arrival of APs, and muscle force production are all comprehensively included. Numerical predictions exhibit trends that are qualitatively similar to measurements of muscle response in rats from a burst of cortical stimulation and a nanosecond impulse. Modulation of neural membrane conductances (including possible electroporation) that alters the neural impulse generation frequency is hypothesized as a possible mechanism leading to observed changes in muscle force production. Other possibilities such as calcium release at nerve end endings also exist. It is also proposed that multipulsing strategies and changing the electric field direction by using multielectrode systems would be useful. Bioelectromagnetics 31:361,370, 2010. © 2010 Wiley-Liss, Inc. [source] The developing relationship between receptor-operated and store-operated calcium channels in smooth muscleBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2002Ian McFadzean Contraction of smooth muscle is initiated, and to a lesser extent maintained, by a rise in the concentration of free calcium in the cell cytoplasm ([Ca2+]i). This activator calcium can originate from two intimately linked sources , the extracellular space and intracellular stores, most notably the sarcoplasmic reticulum. Smooth muscle contraction activated by excitatory neurotransmitters or hormones usually involves a combination of calcium release and calcium entry. The latter occurs through a variety of calcium permeable ion channels in the sarcolemma membrane. The best-characterized calcium entry pathway utilizes voltage-operated calcium channels (VOCCs). However, also present are several types of calcium-permeable channels which are non-voltage-gated, including the so-called receptor-operated calcium channels (ROCCs), activated by agonists acting on a range of G-protein-coupled receptors, and store-operated calcium channels (SOCCs), activated by depletion of the calcium stores within the sarcoplasmic reticulum. In this article we will review the electrophysiological, functional and pharmacological properties of ROCCs and SOCCs in smooth muscle and highlight emerging evidence that suggests that the two channel types may be closely related, being formed from proteins of the Transient Receptor Potential Channel (TRPC) family. British Journal of Pharmacology (2002) 135, 1,13; doi:10.1038/sj.bjp.0704468 [source] Respiratory syncytial virus and neutrophil activationCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 3 2005E. L. Bataki Summary Respiratory syncytial virus infects almost all children by 2 years of age. Neutrophils are the predominant airway leucocytes in RSV bronchiolitis and they are activated in the presence of infection. However it is not clear whether RSV can directly signal to activate neutrophil cytotoxic function. To investigate this we have used a preparation of RSV washed using a new centrifugal diafiltration method to rapidly remove inflammatory molecules produced by the epithelial cells used to propagate the RSV stock. Human neutrophils were isolated from peripheral blood and activated with either the unwashed crude RSV preparations or the purified intact RSV. Neutrophils were also challenged with purified RSV G-glycoprotein. The effect of challenging human neutrophils with these preparations of intact RSV, or the RSV G-glycoprotein, was assessed by measuring the cell surface expression of CD11b and CD18b, the phagocytic oxidative burst, and intracellular release of calcium pools. Neutrophils challenged with the washed RSV exhibited significantly lower activation of surface marker expression (P < 0·001) and oxidative burst (P < 0·001) than those challenged with unwashed virus or with virus free supernatant. There was no increase in intracellular calcium release on exposure to the washed RSV. Purified G glycoprotein did not stimulate neutrophils, whilst the use of a blocking antibody to the F protein did not prevent unwashed RSV from activating cytotoxic responses. These results suggest that neutrophils have no innate signalling system that recognizes RSV but they are activated at sites of RSV infection as a result of the cytokines and inflammatory molecules released by virally infected cells. [source] | ||||||||||||||||||||||||||||